Cancer Associated Fibroblasts express pro-inflammatory factors in human breast and ovarian tumors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Erez, Neta, E-mail: netaerez@post.tau.ac.il; Glanz, Sarah; Raz, Yael

Highlights: •CAFs in human breast and ovarian tumors express pro-inflammatory factors. •Expression of pro-inflammatory factors correlates with tumor invasiveness. •Expression of pro-inflammatory factors is associated with NF-κb activation in CAFs. -- Abstract: Inflammation has been established in recent years as a hallmark of cancer. Cancer Associated Fibroblasts (CAFs) support tumorigenesis by stimulating angiogenesis, cancer cell proliferation and invasion. We previously demonstrated that CAFs also mediate tumor-enhancing inflammation in a mouse model of skin carcinoma. Breast and ovarian carcinomas are amongst the leading causes of cancer-related mortality in women and cancer-related inflammation is linked with both these tumor types. However, themore » role of CAFs in mediating inflammation in these malignancies remains obscure. Here we show that CAFs in human breast and ovarian tumors express high levels of the pro-inflammatory factors IL-6, COX-2 and CXCL1, previously identified to be part of a CAF pro-inflammatory gene signature. Moreover, we show that both pro-inflammatory signaling by CAFs and leukocyte infiltration of tumors are enhanced in invasive ductal carcinoma as compared with ductal carcinoma in situ. The pro-inflammatory genes expressed by CAFs are known NF-κB targets and we show that NF-κB is up-regulated in breast and ovarian CAFs. Our data imply that CAFs mediate tumor-promoting inflammation in human breast and ovarian tumors and thus may be an attractive target for stromal-directed therapeutics.« less

Human amyloid β peptide and tau co-expression impairs behavior and causes specific gene expression changes in Caenorhabditis elegans.

PubMed

Wang, Chenyin; Saar, Valeria; Leung, Ka Lai; Chen, Liang; Wong, Garry

2018-01-01

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by the presence of extracellular amyloid plaques consisting of Amyloid-β peptide (Aβ) aggregates and neurofibrillary tangles formed by aggregation of hyperphosphorylated microtubule-associated protein tau. We generated a novel invertebrate model of AD by crossing Aβ1-42 (strain CL2355) with either pro-aggregating tau (strain BR5270) or anti-aggregating tau (strain BR5271) pan-neuronal expressing transgenic Caenorhabditis elegans. The lifespan and progeny viability of the double transgenic strains were significantly decreased compared with wild type N2 (P<0.0001). In addition, co-expression of these transgenes interfered with neurotransmitter signaling pathways, caused deficits in chemotaxis associative learning, increased protein aggregation visualized by Congo red staining, and increased neuronal loss. Global transcriptomic RNA-seq analysis revealed 248 up- and 805 down-regulated genes in N2 wild type versus Aβ1-42+pro-aggregating tau animals, compared to 293 up- and 295 down-regulated genes in N2 wild type versus Aβ1-42+anti-aggregating tau animals. Gene set enrichment analysis of Aβ1-42+pro-aggregating tau animals uncovered up-regulated annotation clusters UDP-glucuronosyltransferase (5 genes, P<4.2E-4), protein phosphorylation (5 genes, P<2.60E-02), and aging (5 genes, P<8.1E-2) while the down-regulated clusters included nematode cuticle collagen (36 genes, P<1.5E-21). RNA interference of 13 available top up-regulated genes in Aβ1-42+pro-aggregating tau animals revealed that F-box family genes and nep-4 could enhance life span deficits and chemotaxis deficits while Y39G8C.2 (TTBK2) could suppress these behaviors. Comparing the list of regulated genes from C. elegans to the top 60 genes related to human AD confirmed an overlap of 8 genes: patched homolog 1, PTCH1 (ptc-3), the Rab GTPase activating protein, TBC1D16 (tbc-16), the WD repeat and FYVE domain-containing protein 3, WDFY3 (wdfy-3), ADP-ribosylation factor guanine nucleotide exchange factor 2, ARFGEF2 (agef-1), Early B-cell Factor, EBF1 (unc-3), d-amino-acid oxidase, DAO (daao-1), glutamate receptor, metabotropic 1, GRM1 (mgl-2), prolyl 4-hydroxylase subunit alpha 2, P4HA2 (dpy-18 and phy-2). Taken together, our C. elegans double transgenic model provides insight on the fundamental neurobiologic processes underlying human AD and recapitulates selected transcriptomic changes observed in human AD brains. Copyright © 2017 Elsevier Inc. All rights reserved.

Characterization of the procera Tomato Mutant Shows Novel Functions of the SlDELLA Protein in the Control of Flower Morphology, Cell Division and Expansion, and the Auxin-Signaling Pathway during Fruit-Set and Development1[C][W

PubMed Central

Carrera, Esther; Ruiz-Rivero, Omar; Peres, Lazaro Eustaquio Pereira; Atares, Alejandro; Garcia-Martinez, Jose Luis

2012-01-01

procera (pro) is a tall tomato (Solanum lycopersicum) mutant carrying a point mutation in the GRAS region of the gene encoding SlDELLA, a repressor in the gibberellin (GA) signaling pathway. Consistent with the SlDELLA loss of function, pro plants display a GA-constitutive response phenotype, mimicking wild-type plants treated with GA3. The ovaries from both nonemasculated and emasculated pro flowers had very strong parthenocarpic capacity, associated with enhanced growth of preanthesis ovaries due to more and larger cells. pro parthenocarpy is facultative because seeded fruits were obtained by manual pollination. Most pro pistils had exserted stigmas, thus preventing self-pollination, similar to wild-type pistils treated with GA3 or auxins. However, Style2.1, a gene responsible for long styles in noncultivated tomato, may not control the enhanced style elongation of pro pistils, because its expression was not higher in pro styles and did not increase upon GA3 application. Interestingly, a high percentage of pro flowers had meristic alterations, with one additional petal, sepal, stamen, and carpel at each of the four whorls, respectively, thus unveiling a role of SlDELLA in flower organ development. Microarray analysis showed significant changes in the transcriptome of preanthesis pro ovaries compared with the wild type, indicating that the molecular mechanism underlying the parthenocarpic capacity of pro is complex and that it is mainly associated with changes in the expression of genes involved in GA and auxin pathways. Interestingly, it was found that GA activity modulates the expression of cell division and expansion genes and an auxin signaling gene (tomato AUXIN RESPONSE FACTOR7) during fruit-set. PMID:22942390

Co-transfection of decorin and interleukin-10 modulates pro-fibrotic extracellular matrix gene expression in human tenocyte culture

NASA Astrophysics Data System (ADS)

Abbah, Sunny A.; Thomas, Dilip; Browne, Shane; O'Brien, Timothy; Pandit, Abhay; Zeugolis, Dimitrios I.

2016-02-01

Extracellular matrix synthesis and remodelling are driven by increased activity of transforming growth factor beta 1 (TGF-β1). In tendon tissue repair, increased activity of TGF-β1 leads to progressive fibrosis. Decorin (DCN) and interleukin 10 (IL-10) antagonise pathological collagen synthesis by exerting a neutralising effect via downregulation of TGF-β1. Herein, we report that the delivery of DCN and IL-10 transgenes from a collagen hydrogel system supresses the constitutive expression of TGF-β1 and a range of pro-fibrotic extracellular matrix genes.

Analysis of Ly49 gene transcripts in mature NK cells supports a role for the Pro1 element in gene activation, not gene expression.

PubMed

McCullen, M V; Li, H; Cam, M; Sen, S K; McVicar, D W; Anderson, S K

2016-09-01

The variegated expression of murine Ly49 loci has been associated with the probabilistic behavior of an upstream promoter active in immature cells, the Pro1 element. However, recent data suggest that Pro1 may be active in mature natural killer (NK) cells and function as an enhancer element. To assess directly if Pro1 transcripts are present in mature Ly49-expressing NK cells, RNA-sequencing of the total transcript pool was performed on freshly isolated splenic NK cells sorted for expression of either Ly49G or Ly49I. No Pro1 transcripts were detected from the Ly49a, Ly49c or Ly49i genes in mature Ly49(+) NK cells that contained high levels of Pro2 transcripts. Low levels of Ly49g Pro1 transcripts were found in both Ly49G(+) and Ly49G(-) populations, consistent with the presence of a small population of mature NK cells undergoing Ly49g gene activation, as previously demonstrated by culture of splenic NK cells in interleukin-2. Ly49 gene reporter constructs containing Pro1 failed to show any enhancer activity of Pro1 on Pro2 in a mature Ly49-expressing cell line. Taken together, the results are consistent with Pro1 transcription having a role in gene activation in developing NK, and argue against a role for Pro1 in Ly49 gene transcription by mature NK cells.

Differential gene expression in porcine SK6 cells infected with wild-type and SAP domain-mutant foot-and-mouth disease virus.

PubMed

Ni, Zixin; Yang, Fan; Cao, Weijun; Zhang, Xiangle; Jin, Ye; Mao, Ruoqing; Du, Xiaoli; Li, Weiwei; Guo, Jianhong; Liu, Xiangtao; Zhu, Zixiang; Zheng, Haixue

2016-06-01

Foot-and-mouth disease virus (FMDV) is the causative agent of a highly contagious disease in livestock. The viral proteinase L(pro) of FMDV is involved in pathogenicity, and mutation of the L(pro) SAP domain reduces FMDV pathogenicity in pigs. To determine the gene expression profiles associated with decreased pathogenicity in porcine cells, we performed transcriptome analysis using next-generation sequencing technology and compared differentially expressed genes in SK6 cells infected with FMDV containing L(pro) with either a wild-type or mutated version of the SAP domain. This analysis yielded 1,853 genes that exhibited a ≥ 2-fold change in expression and was validated by real-time quantitative PCR detection of several differentially expressed genes. Many of the differentially expressed genes correlated with antiviral responses corresponded to genes associated with transcription factors, immune regulation, cytokine production, inflammatory response, and apoptosis. Alterations in gene expression profiles may be responsible for the variations in pathogenicity observed between the two FMDV variants. Our results provided genes of interest for the further study of antiviral pathways and pathogenic mechanisms related to FMDV L(pro).

Association of Pro12Ala Polymorphism of Peroxisome Proliferator-Activated Receptor gamma 2 (PPARγ2) Gene with Type 2 Diabetes Mellitus in Ethnic Kashmiri Population.

PubMed

Majid, Misbah; Masood, Akbar; Kadla, Showkat Ahmad; Hameed, Iqra; Ganai, Bashir A

2017-02-01

Type 2 diabetes mellitus (T2DM) is characterized by chronic hyperglycemia associated with insulin resistance and relative insulin deficiency. T2DM is believed to be attributable to the combined effect of genetic and environmental factors. Peroxisome proliferator-activated receptor gamma 2 (PPARγ2) is one of the main candidate genes that are implicated in T2DM. A common proline 12 alanine (Pro12Ala) polymorphism in PPARγ2 has been shown to be associated with T2DM. The aim of this work was to investigate the possible role of PPARγ2 gene polymorphism, as a genetic risk factor for T2DM. The study comprised 200 ethnic unrelated subjects (100 T2DM patients and 100 controls). PCR-RFLP technique was used for genotyping analysis. The frequency of the Pro allele was 79 and 91.5 % for controls and cases, respectively (P < 0.05; OR 3.2; 95 % CI 1.64-6.3). The Pro12Ala polymorphism was in Hardy-Weinberg equilibrium in both patients and controls (χ 2  = 0.13, P > 0.05). We found a significant association of Pro12Ala polymorphism of PPARγ2 gene with T2DM, however the genotypes showed statistically significant association only with few clinical parameters including body mass index, total cholesterol, and low-density lipoprotein (P < 0.05). The study signifies that Pro allele in PPARγ2 may be a genotypic risk factor that confers susceptibility to T2DM in ethnic Kashmiri population.

Combining laser microdissection and RNA-seq to chart the transcriptional landscape of fungal development

PubMed Central

2012-01-01

Background During sexual development, filamentous ascomycetes form complex, three-dimensional fruiting bodies for the protection and dispersal of sexual spores. Fruiting bodies contain a number of cell types not found in vegetative mycelium, and these morphological differences are thought to be mediated by changes in gene expression. However, little is known about the spatial distribution of gene expression in fungal development. Here, we used laser microdissection (LM) and RNA-seq to determine gene expression patterns in young fruiting bodies (protoperithecia) and non-reproductive mycelia of the ascomycete Sordaria macrospora. Results Quantitative analysis showed major differences in the gene expression patterns between protoperithecia and total mycelium. Among the genes strongly up-regulated in protoperithecia were the pheromone precursor genes ppg1 and ppg2. The up-regulation was confirmed by fluorescence microscopy of egfp expression under the control of ppg1 regulatory sequences. RNA-seq analysis of protoperithecia from the sterile mutant pro1 showed that many genes that are differentially regulated in these structures are under the genetic control of transcription factor PRO1. Conclusions We have generated transcriptional profiles of young fungal sexual structures using a combination of LM and RNA-seq. This allowed a high spatial resolution and sensitivity, and yielded a detailed picture of gene expression during development. Our data revealed significant differences in gene expression between protoperithecia and non-reproductive mycelia, and showed that the transcription factor PRO1 is involved in the regulation of many genes expressed specifically in sexual structures. The LM/RNA-seq approach will also be relevant to other eukaryotic systems in which multicellular development is investigated. PMID:23016559

Processing of the mother-cell sigma factor, sigma K, may depend on events occurring in the forespore during Bacillus subtilis development.

PubMed Central

Lu, S; Halberg, R; Kroos, L

1990-01-01

During sporulation of the Gram-positive bacterium Bacillus subtilis, transcription of genes encoding spore coat proteins in the mother-cell compartment of the sporangium is controlled by RNA polymerase containing the sigma subunit called sigma K. Based on comparison of the N-terminal amino acid sequence of sigma K with the nucleotide sequence of the gene encoding sigma K (sigK), the primary product of sigK was inferred to be a pro-protein (pro-sigma K) with 20 extra amino acids at the N terminus. Using antibodies generated against pro-sigma K, we have detected pro-sigma K beginning at the third hour of sporulation and sigma K beginning about 1 hr later. Even when pro-sigma K is expressed artificially during growth and throughout sporulation, sigma K appears at the normal time and expression of a sigma K-controlled gene occurs normally. These results suggest that pro-sigma K is an inactive precursor that is proteolytically processed to active sigma K in a developmentally regulated fashion. Mutations that block forespore gene expression block accumulation of sigma K but not accumulation of pro-sigma K, suggesting that pro-sigma K processing is a regulatory device that couples the programs of gene expression in the two compartments of the sporangium. We propose that this regulatory device ensures completion of forespore morphogenesis prior to the synthesis in the mother-cell of spore coat proteins that will encase the forespore. Images PMID:2124700

Stage-specific control of early B cell development by the transcription factor Ikaros

PubMed Central

Gültekin, Sinan; Dakic, Aleksandar; Axelsson, Elin; Minnich, Martina; Ebert, Anja; Werner, Barbara; Roth, Mareike; Cimmino, Luisa; Dickins, Ross A.; Zuber, Johannes; Jaritz, Markus; Busslinger, Meinrad

2018-01-01

Ikaros is an essential regulator of lymphopoiesis. Here, we studied the B-cell-specific function of Ikaros by conditional Ikzf1 inactivation in pro-B cells. B-cell development was arrested at an aberrant ‘pro-B’ cell stage characterized by increased cell adhesion and loss of pre-B cell receptor signaling. Ikaros was found to activate genes coding for pre-BCR signal transducers and to repress genes involved in the downregulation of pre-BCR signaling and upregulation of the integrin signaling pathway. Unexpectedly, derepression of Aiolos expression could not compensate for the loss of Ikaros in pro-B cells. Ikaros induced or suppressed active chromatin at regulatory elements of activated or repressed target genes. Notably, Ikaros binding and target gene expression was dynamically regulated at distinct stages of early B-lymphopoiesis. PMID:24509509

Fluoride exposure abates pro-inflammatory response and induces in vivo apoptosis rendering zebrafish (Danio rerio) susceptible to bacterial infections.

PubMed

Singh, Rashmi; Khatri, Preeti; Srivastava, Nidhi; Jain, Shruti; Brahmachari, Vani; Mukhopadhyay, Asish; Mazumder, Shibnath

2017-04-01

The present study describes the immunotoxic effect of chronic fluoride exposure on adult zebrafish (Danio rerio). Zebrafish were exposed to fluoride (71.12 mg/L; 1/10 LC 50 ) for 30 d and the expression of selected genes studied. We observed significant elevation in the detoxification pathway gene cyp1a suggesting chronic exposure to non-lethal concentration of fluoride is indeed toxic to fish. Fluoride mediated pro-oxidative stress is implicated with the downregulation in superoxide dismutase 1 and 2 (sod1/2) genes. Fluoride affected DNA repair machinery by abrogating the expression of the DNA repair gene rad51 and growth arrest and DNA damage inducible beta a gene gadd45ba. The upregulated expression of casp3a coupled with altered Bcl-2 associated X protein/B-cell lymphoma 2 ratio (baxa/bcl2a) clearly suggested chronic fluoride exposure induced the apoptotic cascade in zebrafish. Fluoride-exposed zebrafish when challenged with non-lethal dose of fish pathogen A. hydrophila revealed gross histopathology in spleen, bacterial persistence and significant mortality. We report that fluoride interferes with system-level output of pro-inflammatory cytokines tumour necrosis factor-α, interleukin-1β and interferon-γ, as a consequence, bacteria replicate efficiently causing significant fish mortality. We conclude, chronic fluoride exposure impairs the redox balance, affects DNA repair machinery with pro-apoptotic implications and suppresses pro-inflammatory cytokines expression abrogating host immunity to bacterial infections. Copyright © 2017 Elsevier Ltd. All rights reserved.

Gene expression changes in mononuclear cells in patients with metabolic syndrome after acute intake of phenol-rich virgin olive oil

PubMed Central

2010-01-01

Background Previous studies have shown that acute intake of high-phenol virgin olive oil reduces pro-inflammatory, pro-oxidant and pro-thrombotic markers compared with low phenols virgin olive oil, but it still remains unclear whether effects attributed to its phenolic fraction are exerted at transcriptional level in vivo. To achieve this goal, we aimed at identifying expression changes in genes which could be mediated by virgin olive oil phenol compounds in the human. Results Postprandial gene expression microarray analysis was performed on peripheral blood mononuclear cells during postprandial period. Two virgin olive oil-based breakfasts with high (398 ppm) and low (70 ppm) content of phenolic compounds were administered to 20 patients suffering from metabolic syndrome following a double-blinded, randomized, crossover design. To eliminate the potential effect that might exist in their usual dietary habits, all subjects followed a similar low-fat, carbohydrate rich diet during the study period. Microarray analysis identified 98 differentially expressed genes (79 underexpressed and 19 overexpressed) when comparing the intake of phenol-rich olive oil with low-phenol olive oil. Many of these genes seem linked to obesity, dyslipemia and type 2 diabetes mellitus. Among these, several genes seem involved in inflammatory processes mediated by transcription factor NF-κB, activator protein-1 transcription factor complex AP-1, cytokines, mitogen-activated protein kinases MAPKs or arachidonic acid pathways. Conclusion This study shows that intake of virgin olive oil based breakfast, which is rich in phenol compounds is able to repress in vivo expression of several pro-inflammatory genes, thereby switching activity of peripheral blood mononuclear cells to a less deleterious inflammatory profile. These results provide at least a partial molecular basis for reduced risk of cardiovascular disease observed in Mediterranean countries, where virgin olive oil represents a main source of dietary fat. Admittedly, other lifestyle factors are also likely to contribute to lowered risk of cardiovascular disease in this region. PMID:20406432

HC-Pro silencing suppressor significantly alters the gene expression profile in tobacco leaves and flowers

PubMed Central

2011-01-01

Background RNA silencing is used in plants as a major defence mechanism against invasive nucleic acids, such as viruses. Accordingly, plant viruses have evolved to produce counter defensive RNA-silencing suppressors (RSSs). These factors interfere in various ways with the RNA silencing machinery in cells, and thereby disturb the microRNA (miRNA) mediated endogene regulation and induce developmental and morphological changes in plants. In this study we have explored these effects using previously characterized transgenic tobacco plants which constitutively express (under CaMV 35S promoter) the helper component-proteinase (HC-Pro) derived from a potyviral genome. The transcript levels of leaves and flowers of these plants were analysed using microarray techniques (Tobacco 4 × 44 k, Agilent). Results Over expression of HC-Pro RSS induced clear phenotypic changes both in growth rate and in leaf and flower morphology of the tobacco plants. The expression of 748 and 332 genes was significantly changed in the leaves and flowers, respectively, in the HC-Pro expressing transgenic plants. Interestingly, these transcriptome alterations in the HC-Pro expressing tobacco plants were similar as those previously detected in plants infected with ssRNA-viruses. Particularly, many defense-related and hormone-responsive genes (e.g. ethylene responsive transcription factor 1, ERF1) were differentially regulated in these plants. Also the expression of several stress-related genes, and genes related to cell wall modifications, protein processing, transcriptional regulation and photosynthesis were strongly altered. Moreover, genes regulating circadian cycle and flowering time were significantly altered, which may have induced a late flowering phenotype in HC-Pro expressing plants. The results also suggest that photosynthetic oxygen evolution, sugar metabolism and energy levels were significantly changed in these transgenic plants. Transcript levels of S-adenosyl-L-methionine (SAM) were also decreased in these plants, apparently leading to decreased transmethylation capacity. The proteome analysis using 2D-PAGE indicated significantly altered proteome profile, which may have been both due to altered transcript levels, decreased translation, and increased proteosomal/protease activity. Conclusion Expression of the HC-Pro RSS mimics transcriptional changes previously shown to occur in plants infected with intact viruses (e.g. Tobacco etch virus, TEV). The results indicate that the HC-Pro RSS contributes a significant part of virus-plant interactions by changing the levels of multiple cellular RNAs and proteins. PMID:21507209

A novel pleiotropic effect of aspirin: Beneficial regulation of pro- and anti-inflammatory mechanisms in microglial cells.

PubMed

Kata, Diana; Földesi, Imre; Feher, Liliana Z; Hackler, Laszlo; Puskas, Laszlo G; Gulya, Karoly

2017-06-01

Aspirin, one of the most widely used non-steroidal anti-inflammatory drugs, has extensively studied effects on the cardiovascular system. To reveal further pleiotropic, beneficial effects of aspirin on a number of pro- and anti-inflammatory microglial mechanisms, we performed morphometric and functional studies relating to phagocytosis, pro- and anti-inflammatory cytokine production (IL-1β, tumor necrosis factor-α (TNF-α) and IL-10, respectively) and analyzed the expression of a number of inflammation-related genes, including those related to the above functions, in pure microglial cells. We examined the effects of aspirin (0.1mM and 1mM) in unchallenged (control) and bacterial lipopolysaccharide (LPS)-challenged secondary microglial cultures. Aspirin affected microglial morphology and functions in a dose-dependent manner as it inhibited LPS-elicited microglial activation by promoting ramification and the inhibition of phagocytosis in both concentrations. Remarkably, aspirin strongly reduced the pro-inflammatory IL-1β and TNF-α production, while it increased the anti-inflammatory IL-10 level in LPS-challenged cells. Moreover, aspirin differentially regulated the expression of a number of inflammation-related genes as it downregulated such pro-inflammatory genes as Nos2, Kng1, IL1β, Ptgs2 or Ccr1, while it upregulated some anti-inflammatory genes such as IL10, Csf2, Cxcl1, Ccl5 or Tgfb1. Thus, the use of aspirin could be beneficial for the prophylaxis of certain neurodegenerative disorders as it effectively ameliorates inflammation in the brain. Copyright © 2017 Elsevier Inc. All rights reserved.

Bee venom inhibits hepatic fibrosis through suppression of pro-fibrogenic cytokine expression.

PubMed

Kim, Soo-Jung; Park, Ji-Hyun; Kim, Kyung-Hyun; Lee, Woo-Ram; Chang, Young-Chae; Park, Kwan-Kyu; Lee, Kwang-Gill; Han, Sang-Mi; Yeo, Joo-Hong; Pak, Sok Cheon

2010-01-01

Bee venom (BV) has a long tradition of use for the control of pain and inflammation in various chronic diseases. Carbon tetrachloride (CCl4) is known to induce hepatotoxicity after being metabolized to the highly reactive trichloromethyl free radical and its peroxy radical. The purpose of the current study was to examine whether BV regulates the pro-inflammation and fibrosis related genes against a mouse model of hepatic fibrosis induced by CCl4 and ethanol-treated hepatocytes (ETH). Test mice were administered with CCl4 (2 ml/mg) and hepatocytes were treated with 25 mM ethanol. BV was added to the final concentration of 0.05-0.5 mg/kg and 1-100 ng/ml for in vivo and in vitro testing, respectively. Fibrotic livers and ETH were used for the measurement of hepatocyte necrosis, pro-inflammatory cytokines and fibrogenic genes. BV suppressed CCl4-induced hepatocyte necrosis markers of serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT). It also inhibited the secretion of interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha. Moreover, BV inhibited CCl4-induced expression of transforming growth factor (TGF)-beta1, alpha-smooth muscle actin (SMA) and fibronectin. Similarly, ETH exhibited significant suppression of IL-1beta, TNF-alpha, TGF-beta1 and fibronectin when cultured with BV. These results suggest that BV possesses anti-fibrogenic properties that are mediated by the suppression of pro-inflammatory cytokines and fibrogenic gene expression. BV has substantial therapeutic potential for the treatment of fibrotic diseases.

Prognostic Role of Host Cyclooxygenase and Cytokine Genotypes in a Caucasian Cohort of Patients with Gastric Adenocarcinoma

PubMed Central

García-González, María Asunción; Nicolás-Pérez, David; Lanas, Angel; Bujanda, Luis; Carrera, Patricia; Benito, Rafael; Strunk, Mark; Sopeña, Federico; Santolaria, Santos; Piazuelo, Elena; Jiménez, Pilar; Campo, Rafael; Espinel, Jesús; Manzano, Marisa; Geijo, Fernando; Pellisé, María; González-Huix, Ferrán; Espinós, Jorge; Zaballa, Manuel; Titó, Llúcia; Barranco, Luis; Pazo, Roberto; Quintero, Enrique

2012-01-01

Background Genetic factors influencing the prognosis of gastric adenocarcinoma (GAC) are not well known. Given the relevance of cytokines and other pro-inflammatory mediators in cancer progression and invasiveness, we aimed to assess the prognostic role of several functional cytokine and cyclooxygenase gene polymorphisms in patients with GAC. Methodology Genomic DNA from 380 Spanish Caucasian patients with primary GAC was genotyped for 23 polymorphisms in pro-inflammatory (IL1B, TNFA, LTA, IL6, IL12p40), anti-inflammatory (IL4, IL1RN, IL10, TGFB1) cytokine, and cyclooxygenase (PTGS1 and PTGS2) genes by PCR, RFLP and TaqMan assays. Clinical and histological information was collected prospectively. Survival curves were estimated by the Kaplan-Meier method and compared using the log rank test. Outcome was determined by analysis of Cox proportional hazards, adjusting for confounding factors. Results The median follow-up period and median overall survival (OS) time were 9.9 months (range 0.4–120.3) and 10.9 months (95% CI: 8.9–14.1), respectively. Multivariate analysis identified tumor stages III (HR, 3.23; 95% CI:2–5.22) and IV (HR, 5.5; 95% CI: 3.51–8.63) as independent factors associated with a significantly reduced OS, whereas surgical treatment (HR: 0.44; 95%CI: 0.3–0.6) was related to a better prognosis of the disease. Concerning genetic factors, none of the 23 polymorphisms evaluated in the current study did influence survival. Moreover, no gene-environment interactions on GAC prognosis were observed. Conclusions Our results show that, in our population, the panel of selected pro- and anti-inflammatory cytokine, and cyclooxygenase gene polymorphisms are not relevant in determining the prognosis of gastric adenocarcinoma. PMID:23029430

Tumour necrosis factor receptor trafficking dysfunction opens the TRAPS door to pro-inflammatory cytokine secretion

PubMed Central

Turner, Mark D.; Chaudhry, Anupama; Nedjai, Belinda

2011-01-01

Cytokines are secreted from macrophages and other cells of the immune system in response to pathogens. Additionally, in autoinflammatory diseases cytokine secretion occurs in the absence of pathogenic stimuli. In the case of TRAPS [TNFR (tumour necrosis factor receptor)-associated periodic syndrome], inflammatory episodes result from mutations in the TNFRSF1A gene that encodes TNFR1. This work remains controversial, however, with at least three distinct separate mechanisms of receptor dysfunction having been proposed. Central to these hypotheses are the NF-κB (nuclear factor κB) and MAPK (mitogen-activated protein kinase) families of transcriptional activators that are able to up-regulate expression of a number of genes, including pro-inflammatory cytokines. The present review examines each proposed mechanism of TNFR1 dysfunction, and addresses how these processes might ultimately impact upon cytokine secretion and disease pathophysiology. PMID:22115362

Impact of the Pro12Ala polymorphism of the PPARγ2 gene on diabetes and obesity in a highly consanguineous population

PubMed Central

Bener, Abdulbari; Zirie, M; Al-Hamaq, AOAA; Nawaz, Z; Samson, N; Mohammad, R

2015-01-01

Background: The peroxisome proliferator-activated receptors (PPARs) are members of the nuclear hormone receptor subfamily of transcription factors. It has been reported that they play important roles in obesity and the development of type 2 diabetes mellitus (T2DM). Materials and Methods: This case-control study was carried out among 764 Qatari patients with diabetes and 764 healthy subjects above 20 years of age at Primary Healthcare Clinics (PHCs) from January 2011 to December 2012. Face-to-face interviews were based on a questionnaire that included variables such as age, sex, sociodemographic status, body mass index (BMI) and other clinical parameters. The Pro12Ala in the PPARγ2 gene was detected on the LightCycler using two specific probes. Univariate and multivariate statistical analysis were performed. Results: The study revealed that in the diabetes group, Pro/(10.2% vs 9.4%; P = 0.606) and Ala/Ala (1.4% vs 0.9%; P = 0.343) were higher than in controls, whereas Pro/Pro (88.4% vs 89.7%;P = 0.413) was lower in diabetes patients, but no significant difference was observed among the genotype groups. In obese patients with diabetes, Pro/Pro (89% vs 89.9%;P = 0.792) and Pro/Ala (8.9% vs 10.1%;P = 0.671) were lower than in obese healthy subjects. No homozygous Ala/Ala was found in obese healthy subjects, whereas 6 Ala/Ala homozygotes were in obese diabetes group. But in diabetes group, obese patients had higher homozygous of Pro/Pro (89.3% vs 87.8%;P = 0.523) and Ala/Ala (1.8% vs 1.2%;P = 0.771) compared to non-obese patients. Conclusion: The current study did not reveal an association between the Pro12Ala polymorphism of the PPAR γ2 gene and type 2 diabetes (T2D) in Qatari's population. PMID:25593831

Generation and CRISPR/Cas9 editing of transformed progenitor B cells as a pseudo-physiological system to study DNA repair gene function in V(D)J recombination.

PubMed

Lenden Hasse, Hélène; Lescale, Chloé; Bianchi, Joy J; Yu, Wei; Bedora-Faure, Marie; Deriano, Ludovic

2017-12-01

Antigen receptor gene assembly is accomplished in developing lymphocytes by the V(D)J recombination reaction, which can be separated into two steps: DNA cleavage by the recombination-activating gene (RAG) nuclease and joining of DNA double strand breaks (DSBs) by components of the nonhomologous end joining (NHEJ) pathway. Deficiencies for NHEJ factors can result in immunodeficiency and a propensity to accumulate genomic instability, thus highlighting the importance of identifying all players in this process and deciphering their functions. Bcl2 transgenic v-Abl kinase-transformed pro-B cells provide a pseudo-physiological cellular system to study V(D)J recombination. Treatment of v-Abl/Bcl2 pro-B cells with the Abl kinase inhibitor Imatinib leads to G1 cell cycle arrest, the rapid induction of Rag1/2 gene expression and V(D)J recombination. In this system, the Bcl2 transgene alleviates Imatinib-induced apoptosis enabling the analysis of induced V(D)J recombination. Although powerful, the use of mouse models carrying the Bcl2 transgene for the generation of v-Abl pro-B cell lines is time and money consuming. Here, we describe a method for generating v-Abl/Bcl2 pro-B cell lines from wild type mice and for performing gene knock-out using episomal CRISPR/Cas9 targeting vectors. Using this approach, we generated distinct NHEJ-deficient pro-B cell lines and quantified V(D)J recombination levels in these cells. Furthermore, this methodology can be adapted to generate pro-B cell lines deficient for any gene suspected to play a role in V(D)J recombination, and more generally DSB repair. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

An engineered promoter driving expression of a microbial avirulence gene confers recognition of TAL effectors and reduces growth of diverse Xanthomonas strains in citrus.

PubMed

Shantharaj, Deepak; Römer, Patrick; Figueiredo, Jose F L; Minsavage, Gerald V; Krönauer, Christina; Stall, Robert E; Moore, Gloria A; Fisher, Latanya C; Hu, Yang; Horvath, Diana M; Lahaye, Thomas; Jones, Jeffrey B

2017-09-01

Xanthomonas citri ssp. citri (X. citri), causal agent of citrus canker, uses transcription activator-like effectors (TALEs) as major pathogenicity factors. TALEs, which are delivered into plant cells through the type III secretion system (T3SS), interact with effector binding elements (EBEs) in host genomes to activate the expression of downstream susceptibility genes to promote disease. Predictably, TALEs bind EBEs in host promoters via known combinations of TALE amino acids to DNA bases, known as the TALE code. We introduced 14 EBEs, matching distinct X. citri TALEs, into the promoter of the pepper Bs3 gene (ProBs3 1EBE ), and fused this engineered promoter with multiple EBEs (ProBs3 14EBE ) to either the β-glucuronidase (GUS) reporter gene or the coding sequence (cds) of the pepper gene, Bs3. TALE-induced expression of the Bs3 cds in citrus leaves resulted in no visible hypersensitive response (HR). Therefore, we utilized a different approach in which ProBs3 1EBE and ProBs3 14EBE were fused to the Xanthomonas gene, avrGf1, which encodes a bacterial effector that elicits an HR in grapefruit and sweet orange. We demonstrated, in transient assays, that activation of ProBs3 14EBE by X. citri TALEs is T3SS dependent, and that the expression of AvrGf1 triggers HR and correlates with reduced bacterial growth. We further demonstrated that all tested virulent X. citri strains from diverse geographical locations activate ProBs3 14EBE . TALEs are essential for the virulence of X. citri strains and, because the engineered promoter traps are activated by multiple TALEs, this concept has the potential to confer broad-spectrum, durable resistance to citrus canker in stably transformed plants. © 2016 BSPP AND JOHN WILEY & SONS LTD.

Ultrafine particles from diesel vehicle emissions at different driving cycles induce differential vascular pro-inflammatory responses: Implication of chemical components and NF-κB signaling

PubMed Central

2010-01-01

Background Epidemiological evidence supports the association between exposure to ambient particulate matter (PM) and cardiovascular diseases. Chronic exposure to ultrafine particles (UFP; Dp <100 nm) is reported to promote atherosclerosis in ApoE knockout mice. Atherogenesis-prone factors induce endothelial dysfunction that contributes to the initiation and progression of atherosclerosis. We previously demonstrated that UFP induced oxidative stress via c-Jun N-terminal Kinases (JNK) activation in endothelial cells. In this study, we investigated pro-inflammatory responses of human aortic endothelial cells (HAEC) exposed to UFP emitted from a diesel truck under an idling mode (UFP1) and an urban dynamometer driving schedule (UFP2), respectively. We hypothesize that UFP1 and UFP2 with distinct chemical compositions induce differential pro-inflammatory responses in endothelial cells. Results UFP2 contained a higher level of redox active organic compounds and metals on a per PM mass basis than UFP1. While both UFP1 and UFP2 induced superoxide production and up-regulated stress response genes such as heme oxygenease-1 (HO-1), OKL38, and tissue factor (TF), only UFP2 induced the expression of pro-inflammatory genes such as IL-8 (2.8 ± 0.3-fold), MCP-1 (3.9 ± 0.4-fold), and VCAM (6.5 ± 1.1-fold) (n = 3, P < 0.05). UFP2-exposed HAEC also bound to a higher number of monocytes than UFP1-exposed HAEC (Control = 70 ± 7.5, UFP1 = 106.7 ± 12.5, UFP2 = 137.0 ± 8.0, n = 3, P < 0.05). Adenovirus NF-κB Luciferase reporter assays revealed that UFP2, but not UFP1, significantly induced NF-κB activities. NF-κB inhibitor, CAY10512, significantly abrogated UFP2-induced pro-inflammatory gene expression and monocyte binding. Conclusion While UFP1 induced higher level of oxidative stress and stress response gene expression, only UFP2, with higher levels of redox active organic compounds and metals, induced pro-inflammatory responses via NF-κB signaling. Thus, UFP with distinct chemical compositions caused differential response patterns in endothelial cells. PMID:20307321

Glycosylphosphatidylinositol Anchor Modification Machinery Deficiency Is Responsible for the Formation of Pro-Prion Protein (PrP) in BxPC-3 Protein and Increases Cancer Cell Motility*

PubMed Central

Yang, Liheng; Gao, Zhenxing; Hu, Lipeng; Wu, Guiru; Yang, Xiaowen; Zhang, Lihua; Zhu, Ying; Wong, Boon-Seng; Xin, Wei; Sy, Man-Sun; Li, Chaoyang

2016-01-01

The normal cellular prion protein (PrP) is a glycosylphosphatidylinositol (GPI)-anchored cell surface glycoprotein. However, in pancreatic ductal adenocarcinoma cell lines, such as BxPC-3, PrP exists as a pro-PrP retaining its glycosylphosphatidylinositol (GPI) peptide signaling sequence. Here, we report the identification of another pancreatic ductal adenocarcinoma cell line, AsPC-1, which expresses a mature GPI-anchored PrP. Comparison of the 24 genes involved in the GPI anchor modification pathway between AsPC-1 and BxPC-3 revealed 15 of the 24 genes, including PGAP1 and PIG-F, were down-regulated in the latter cells. We also identified six missense mutations in DPM2, PIG-C, PIG-N, and PIG-P alongside eight silent mutations. When BxPC-3 cells were fused with Chinese hamster ovary (CHO) cells, which lack endogenous PrP, pro-PrP was successfully converted into mature GPI-anchored PrP. Expression of the individual gene, such as PGAP1, PIG-F, or PIG-C, into BxPC-3 cells does not result in phosphoinositide-specific phospholipase C sensitivity of PrP. However, when PIG-F but not PIG-P is expressed in PGAP1-expressing BxPC-3 cells, PrP on the surface of the cells becomes phosphoinositide-specific phospholipase C-sensitive. Thus, low expression of PIG-F and PGAP1 is the major factor contributing to the accumulation of pro-PrP. More importantly, BxPC-3 cells expressing GPI-anchored PrP migrate much slower than BxPC-3 cells bearing pro-PrP. In addition, GPI-anchored PrP-bearing AsPC-1 cells also migrate slower than pro-PrP bearing BxPC-3 cells, although both cells express filamin A. “Knocking out” PRNP in BxPC-3 cell drastically reduces its migration. Collectively, these results show that multiple gene irregularity in BxPC-3 cells is responsible for the formation of pro-PrP, and binding of pro-PrP to filamin A contributes to enhanced tumor cell motility. PMID:26683373

Increased megakaryocytic proliferation, pro-platelet deposition and expression of fibrosis-associated factors in children with chronic myeloid leukaemia with bone marrow fibrosis.

PubMed

Hussein, K; Stucki-Koch, A; Göhring, G; Kreipe, H; Suttorp, M

2017-07-01

Paediatric chronic myeloid leukaemia (ped-CML) is rare and ped-CML with fibre accumulation in the bone marrow (MF) is thought to be even rarer. In adults (ad-CML), fibrosis represents an adverse prognostic factor. So far, the pro-fibrotic changes in the bone marrow microenvironment have not been investigated in detail in ped-CML. From a total of 66 ped-CML in chronic phase, biopsies were analysable and 10 had MF1/2 (MF1, n=8/10; MF2, n=2/10). We randomly selected 16 ped-CML and 16 ad-CML cases with and without fibrosis (each n=8) as well as 18 non-neoplastic controls. Bone marrow samples were analysed with a real-time PCR-based assay (including 127 genes for paediatric cases) and by immunohistochemistry. We found increased expression of megakaryocytic genes in ped-CML. The number of megakaryocytes and pro-platelets are increased in CML patients, but the most significant increase was noted for ped-CML-MF1/2. Anti-fibrotic MMP9 expression was lower in children than in adults. Cell mobilisation-related CXCL12 was decreased in young and adult patients with CML but not the corresponding receptor CXCR4. In summary, fibre accumulation in ped-CML-MF1/2 is associated with increased megakaryocytic proliferation and increased interstitial pro-platelet deposition. Deregulated expression of matrix-modulating factors shifts the bone marrow microenvironment towards fibrosis.

Biological properties of human skeletal myoblasts genetically modified to simultaneously overexpress the pro-angiogenic factors vascular endothelial growth factor-A and fibroblast growth factor-4.

PubMed

Zimna, A; Janeczek, A; Rozwadowska, N; Fraczek, M; Kucharzewska, P; Rucinski, M; Mietkiewski, T; Kurpisz, M

2014-04-01

Myocardial infarction results in cardiomyocyte loss and may eventually lead to cardiac failure. Skeletal myoblast transplantation into the scar area may compensate for this observed cell loss by strengthening the weakened myocardium and inducing myogenesis. Moreover, skeletal myoblasts may serve as potential transgene carriers for the myocardium (i.e., delivering pro-angiogenic factors, which may potentially improve blood perfusion in infarcted heart). We examined the influence of the simultaneous overexpression of two potent pro-angiogenic factors, fibroblast growth factor-4 (FGF-4) and vascular endothelial growth factor (VEGF), on human primary myoblast proliferation, cell cycle, resistance to hypoxic stress conditions and myogenic gene expression, as well as the induction of pro-angiogenic activities. We used a bicistronic plasmid vector encoding two factors introduced via an efficient myoblast electroporation method. The levels of overexpressed proteins were assessed, and their functionality at capillary formation was evaluated. This combined approach led to a high level of non-viral transient overexpression of both pro-angiogenic proteins, which proved to be potent regulators of blood vessel development assayed in capillary formation tests. We demonstrated in in vitro conditions that the transfection of human skeletal myoblasts with both FGF-4 and VEGF did not affect their basic biological properties such as the cell cycle, proliferation or expression of myogenic lineage-specific genes, and the modified cells adapted to oxidative stress conditions. Overall, the results obtained suggest that the applied combined approach with the use of two pro-angiogenic genes overexpressed in skeletal muscle stem cells may be an interesting alternative for the effective therapy of myocardial infarction in animal models and/or prospective clinical trials.

Spi-C has opposing effects to PU.1 on gene expression in progenitor B cells.

PubMed

Schweitzer, Brock L; Huang, Kelly J; Kamath, Meghana B; Emelyanov, Alexander V; Birshtein, Barbara K; DeKoter, Rodney P

2006-08-15

The Ets transcription factor Spi-C, expressed in B cells and macrophages, is closely related to PU.1 and has the ability to recognize the same DNA consensus sequence. However, the function of Spi-C has yet to be determined. The purpose of this study is to further examine Spi-C activity in B cell development. First, using retroviral vectors to infect PU.1(-/-) fetal liver progenitors, Spi-C was found to be inefficient at inducing cytokine-dependent proliferation and differentiation of progenitor B (pro-B) cells or macrophages relative to PU.1 or Spi-B. Next, Spi-C was ectopically expressed in fetal liver-derived, IL-7-dependent pro-B cell lines. Wild-type (WT) pro-B cells ectopically expressing Spi-C (WT-Spi-C) have several phenotypic characteristics of pre-B cells such as increased CD25 and decreased c-Kit surface expression. In addition, WT-Spi-C pro-B cells express increased levels of IgH sterile transcripts and reduced levels of expression and transcription of the FcgammaRIIb gene. Gel-shift analysis suggests that Spi-C, ectopically expressed in pro-B cells, can bind PU.1 consensus sites in the IgH intronic enhancer and FcgammaRIIb promoter. Transient transfection analysis demonstrated that PU.1 functions to repress the IgH intronic enhancer and activate the FcgammaRIIb promoter, while Spi-C opposes these activities. WT-Spi-C pro-B cells have reduced levels of dimethylation on lysine 9 of histone H3 within the IgH 3' regulatory region, indicating that Spi-C can contribute to removal of repressive features in the IgH locus. Overall, these studies suggest that Spi-C may promote B cell differentiation by modulating the activity of PU.1-dependent genes.

RAAS gene polymorphisms influence progression of pediatric hypertrophic cardiomyopathy.

PubMed

Kaufman, Beth D; Auerbach, Scott; Reddy, Sushma; Manlhiot, Cedric; Deng, Liyong; Prakash, Ashwin; Printz, Beth F; Gruber, Dorota; Papavassiliou, Dimitrios P; Hsu, Daphne T; Sehnert, Amy J; Chung, Wendy K; Mital, Seema

2007-12-01

Hypertrophic Cardiomyopathy (HCM) is a disease with variable rate of progression. Young age is an independent risk factor for poor outcome in HCM. The influence of renin-angiotensin-aldosterone (RAAS) genotype on the progression of HCM in children is unknown. Children with HCM (n = 65) were enrolled prospectively across two centers (2001-2005). All subjects were genotyped for five RAAS gene polymorphisms previously associated with LV hypertrophy (pro-LVH): AGT M235T, ACE DD, CMA-1903 A/G, AGTR1 1666 A/C and CYP11B2-344 C/T. Linear regression models, based on maximum likelihood estimates, were created to assess the independent effect of RAAS genotype on LV hypertrophy (LVH). Forty-six subjects were homozygous for <2 and 19 were homozygous for > or =2 pro-LVH RAAS polymorphisms. Mean age at presentation was 9.6 +/- 6 years. Forty children had follow-up echocardiograms after a median of 1.5 years. Indexed LV mass (LVMI) and LV mass z-scores were higher at presentation and follow-up in subjects with > or =2 pro-LVH genotypes compared to those with <2 (P < 0.05). Subjects with > or =2 pro-LVH genotypes also demonstrated a greater increase in septal thickness (IVST) and in LV outflow tract (LVOT) obstruction on follow-up (P < 0.05). On multivariate analysis, a higher number of pro-LVH genotypes was associated with a larger effect size (P < 0.05). Pro-LVH RAAS gene polymorphisms are associated with progressive septal hypertrophy and LVOT obstruction in children with HCM. Identification of RAAS modifier genes may help to risk-stratify patients with HCM.

Recurrent pregnancy failure is associated with a polymorphism in the p53 tumour suppressor gene.

PubMed

Pietrowski, Detlef; Bettendorf, Hertha; Riener, Eva-Katrin; Keck, Christoph; Hefler, Lukas A; Huber, Johannes C; Tempfer, Clemens

2005-04-01

The p53 tumour suppressor gene is a well-known factor regulating apoptosis in a wide variety of cells and tissues. Alterations in the p53 gene are among the most common genetic changes in human cancers. In addition, recent data provide evidence that p53 plays a critical role in mediating pregnancy by regulating steroid hormone activation. In idiopathic recurrent miscarriages (IRM), causes and associations are much debated as the exact pathophysiological mechanisms are unknown. In this study, we assess whether an established polymorphism in the p53 gene is associated with the occurrence of IRM. Genotyping was performed by PCR-based amplification of the p53 Arg and Pro variants at codon 72 in 175 cases of IRM and 143 controls. We observed a statistically significant association between carriage of the Pro allele and the occurrence of IRM (P = 0.03, odds ratio 1.49, confidence interval 1.04-2.14). Distribution of genotypes was in Hardy-Weinberg equilibrium. Our results indicate an over-representation of the Pro allele of the p53 gene in women with IRM, giving support to the theory that p53 has a potential role during pregnancy.

AMP-activated Protein Kinase Mediates Apoptosis in Response to Bioenergetic Stress through Activation of the Pro-apoptotic Bcl-2 Homology Domain-3-only Protein BMF*

PubMed Central

Kilbride, Seán M.; Farrelly, Angela M.; Bonner, Caroline; Ward, Manus W.; Nyhan, Kristine C.; Concannon, Caoimhín G.; Wollheim, Claes B.; Byrne, Maria M.; Prehn, Jochen H. M.

2010-01-01

Heterozygous loss-of-function mutations in the hepatocyte nuclear factor 1A (HNF1A) gene result in the pathogenesis of maturity-onset diabetes-of-the-young type 3, (HNF1A-MODY). This disorder is characterized by a primary defect in metabolism-secretion coupling and decreased beta cell mass, attributed to excessive beta cell apoptosis. Here, we investigated the link between energy stress and apoptosis activation following HNF1A inactivation. This study employed single cell fluorescent microscopy, flow cytometry, gene expression analysis, and gene silencing to study the effects of overexpression of dominant-negative (DN)-HNF1A expression on cellular bioenergetics and apoptosis in INS-1 cells. Induction of DN-HNF1A expression led to reduced ATP levels and diminished the bioenergetic response to glucose. This was coupled with activation of the bioenergetic stress sensor AMP-activated protein kinase (AMPK), which preceded the onset of apoptosis. Pharmacological activation of AMPK using aminoimidazole carboxamide ribonucleotide (AICAR) was sufficient to induce apoptosis in naive cells. Conversely, inhibition of AMPK with compound C or AMPKα gene silencing protected against DN-HNF1A-induced apoptosis. Interestingly, AMPK mediated the induction of the pro-apoptotic Bcl-2 homology domain-3-only protein Bmf (Bcl-2-modifying factor). Bmf expression was also elevated in islets of DN-HNF1A transgenic mice. Furthermore, knockdown of Bmf expression in INS-1 cells using siRNA was sufficient to protect against DN-HNF1A-induced apoptosis. Our study suggests that overexpression of DN-HNF1A induces bioenergetic stress and activation of AMPK. This in turn mediates the transcriptional activation of the pro-apoptotic Bcl-2-homology protein BMF, coupling prolonged energy stress to apoptosis activation. PMID:20841353

Collagen-derived dipeptide prolyl-hydroxyproline promotes differentiation of MC3T3-E1 osteoblastic cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kimira, Yoshifumi, E-mail: kimira@josai.ac.jp; Ogura, Kana; Taniuchi, Yuri

Highlights: • Pro-Hyp did not affect MC3T3-E1 cell proliferation and matrix mineralization. • Pro-Hyp significantly increased alkaline phosphatase activity. • Pro-Hyp significantly upregulated gene expression of Runx2, Osterix, and Col1α1. - Abstract: Prolyl-hydroxyproline (Pro-Hyp) is one of the major constituents of collagen-derived dipeptides. The objective of this study was to investigate the effects of Pro-Hyp on the proliferation and differentiation of MC3T3-E1 osteoblastic cells. Addition of Pro-Hyp did not affect MC3T3-E1 cell proliferation and matrix mineralization but alkaline phosphatase activity was significantly increased. Furthermore, cells treated with Pro-Hyp significantly upregulated gene expression of Runx2, Osterix, and Col1α1. These results indicatemore » that Pro-Hyp promotes osteoblast differentiation. This study demonstrates for the first time that Pro-Hyp has a positive effect on osteoblast differentiation with upregulation of Runx2, Osterix, and Collα1 gene expression.« less

The B-cell identity factor Pax5 regulates distinct transcriptional programmes in early and late B lymphopoiesis

PubMed Central

Revilla-i-Domingo, Roger; Bilic, Ivan; Vilagos, Bojan; Tagoh, Hiromi; Ebert, Anja; Tamir, Ido M; Smeenk, Leonie; Trupke, Johanna; Sommer, Andreas; Jaritz, Markus; Busslinger, Meinrad

2012-01-01

Pax5 controls the identity and development of B cells by repressing lineage-inappropriate genes and activating B-cell-specific genes. Here, we used genome-wide approaches to identify Pax5 target genes in pro-B and mature B cells. In these cell types, Pax5 bound to 40% of the cis-regulatory elements defined by mapping DNase I hypersensitive (DHS) sites, transcription start sites and histone modifications. Although Pax5 bound to 8000 target genes, it regulated only 4% of them in pro-B and mature B cells by inducing enhancers at activated genes and eliminating DHS sites at repressed genes. Pax5-regulated genes in pro-B cells account for 23% of all expression changes occurring between common lymphoid progenitors and committed pro-B cells, which identifies Pax5 as an important regulator of this developmental transition. Regulated Pax5 target genes minimally overlap in pro-B and mature B cells, which reflects massive expression changes between these cell types. Hence, Pax5 controls B-cell identity and function by regulating distinct target genes in early and late B lymphopoiesis. PMID:22669466

Mutation Screening of the Krüppel-like Factor 1 Gene in Individuals With Increased Fetal Hemoglobin Referred for Hemoglobinopathy Investigation in South of Iran.

PubMed

Hamid, Mohammad; Ershadi Oskouei, Sanaz; Shariati, Gholamreza; Babaei, Esmaeil; Galehdari, Hamid; Saberi, Alihossein; Sedaghat, Alireza

2018-04-01

Any mutation in the Krüppel-like factor 1 (KLF1) gene may interfere with its proper related function in the erythropoiesis process and lead to alterations in proper activation of its downstream protein through globin switching, which results in an increase in fetal hemoglobin (HbF). This study aimed to investigate whether KLF1 mutation can associate with high level of HbF in individuals with increased fetal hemoglobin referred for screening of hemoglobinopathies in south of Iran. The human KLF1 gene was amplified via the polymerase chain reaction procedure, and sequencing was used to determine any mutation in these patients. Moreover, XmnI polymorphisms in the position of -158 of γ-globin gene promoter were analyzed in all patients by polymerase chain reaction restriction fragment length polymorphism. Analysis of sequencing revealed a missense mutation in the KLF1 gene, p.Ser102Pro (c.304T>C), which was detectable in 10 of 23 cases with elevated HbF level. This mutation was only detected in individuals who had a HbF level between 3.1% and 25.6%. Statistical analysis showed that the frequency of C allele is significantly correlated with a high level of HbF (P<0.05). The allele frequency of positive result of XmnI polymorphism in individuals with increased HbF level was also significant, which showed an association with increased HbF level (P<0.05). To the best of our knowledge, this is the first report of p.Ser102Pro (c.304T>C) in the KLF1 gene in β-thalassemia patients with increased level of fetal hemoglobin. According to statistical results of p.Ser102Pro mutation and XmnI polymorphism, it has been strongly suggested that both polymorphisms have an association with increased HbF samples. These nucleotide changes alone may not be the only elements raising the level of HbF, and other regulatory and modifying factors also play a role in HbF production.

Molecular risk assessment of BIG 1-98 participants by expression profiling using RNA from archival tissue

PubMed Central

2010-01-01

Background The purpose of the work reported here is to test reliable molecular profiles using routinely processed formalin-fixed paraffin-embedded (FFPE) tissues from participants of the clinical trial BIG 1-98 with a median follow-up of 60 months. Methods RNA from fresh frozen (FF) and FFPE tumor samples of 82 patients were used for quality control, and independent FFPE tissues of 342 postmenopausal participants of BIG 1-98 with ER-positive cancer were analyzed by measuring prospectively selected genes and computing scores representing the functions of the estrogen receptor (eight genes, ER_8), the progesterone receptor (five genes, PGR_5), Her2 (two genes, HER2_2), and proliferation (ten genes, PRO_10) by quantitative reverse transcription PCR (qRT-PCR) on TaqMan Low Density Arrays. Molecular scores were computed for each category and ER_8, PGR_5, HER2_2, and PRO_10 scores were combined into a RISK_25 score. Results Pearson correlation coefficients between FF- and FFPE-derived scores were at least 0.94 and high concordance was observed between molecular scores and immunohistochemical data. The HER2_2, PGR_5, PRO_10 and RISK_25 scores were significant predictors of disease free-survival (DFS) in univariate Cox proportional hazard regression. PRO_10 and RISK_25 scores predicted DFS in patients with histological grade II breast cancer and in lymph node positive disease. The PRO_10 and PGR_5 scores were independent predictors of DFS in multivariate Cox regression models incorporating clinical risk indicators; PRO_10 outperformed Ki-67 labeling index in multivariate Cox proportional hazard analyses. Conclusions Scores representing the endocrine responsiveness and proliferation status of breast cancers were developed from gene expression analyses based on RNA derived from FFPE tissues. The validation of the molecular scores with tumor samples of participants of the BIG 1-98 trial demonstrates that such scores can serve as independent prognostic factors to estimate disease free survival (DFS) in postmenopausal patients with estrogen receptor positive breast cancer. Trial Registration Current Controlled Trials: NCT00004205 PMID:20144231

Toll-like receptor-mediated inhibition of Gas6 and ProS expression facilitates inflammatory cytokine production in mouse macrophages

PubMed Central

Deng, Tingting; Zhang, Yue; Chen, Qiaoyuan; Yan, Keqin; Han, Daishu

2012-01-01

Activation of Toll-like receptors (TLRs) triggers rapid inflammatory cytokine production in various cell types. The exogenous product of growth-arrest-specific gene 6 (Gas6) and Protein S (ProS) inhibit the TLR-triggered inflammatory responses through the activation of Tyro3, Axl and Mer (TAM) receptors. However, regulation of the Gas6/ProS-TAM system remains largely unknown. In the current study, mouse macrophages are shown to constitutively express Gas6 and ProS, which synergistically suppress the basal and TLR-triggered production of inflammatory cytokines, including those of tumour necrosis factor-α, interleukin-6 and interleukin-1β, by the macrophages in an autocrine manner. Notably, TLR signalling markedly decreases Gas6 and ProS expression in macrophages through the activation of the nuclear factor-κB. Further, the down-regulation of Gas6 and ProS by TLR signalling facilitates the TLR-mediated inflammatory cytokine production in mouse macrophages. These results describe a self-regulatory mechanism of TLR signalling through the suppression of Gas6 and ProS expression. PMID:22043818

LuFLA1PRO and LuBGAL1PRO promote gene expression in the phloem fibres of flax (Linum usitatissimum).

PubMed

Hobson, Neil; Deyholos, Michael K

2013-04-01

Cell type-specific promoters were identified that drive gene expression in an industrially important product. To identify flax (Linum usitatissimum) gene promoters, we analyzed the genomic regions upstream of a fasciclin-like arabinogalactan protein (LuFLA1) and a beta-galactosidase (LuBGAL1). Both of these genes encode transcripts that have been found to be highly enriched in tissues bearing phloem fibres. Using a beta-glucuronidase (GUS) reporter construct, we found that a 908-bp genomic sequence upstream of LuFLA1 (LuFLA1PRO) directed GUS expression with high specificity to phloem fibres undergoing secondary cell wall development. The DNA sequence upstream of LuBGAL1 (LuBGAL1PRO) likewise produced GUS staining in phloem fibres with developing secondary walls, as well as in tissues of developing flowers and seed bolls. These data provide further evidence of a specific role for LuFLA1 in phloem fibre development, and demonstrate the utility of LuFLA1PRO and LuBGAL1PRO as tools for biotechnology and further investigations of phloem fibre development.

Genome-Wide Association Mapping Combined with Reverse Genetics Identifies New Effectors of Low Water Potential-Induced Proline Accumulation in Arabidopsis1[W][OPEN

PubMed Central

Verslues, Paul E.; Lasky, Jesse R.; Juenger, Thomas E.; Liu, Tzu-Wen; Kumar, M. Nagaraj

2014-01-01

Arabidopsis (Arabidopsis thaliana) exhibits natural genetic variation in drought response, including varying levels of proline (Pro) accumulation under low water potential. As Pro accumulation is potentially important for stress tolerance and cellular redox control, we conducted a genome-wide association (GWAS) study of low water potential-induced Pro accumulation using a panel of natural accessions and publicly available single-nucleotide polymorphism (SNP) data sets. Candidate genomic regions were prioritized for subsequent study using metrics considering both the strength and spatial clustering of the association signal. These analyses found many candidate regions likely containing gene(s) influencing Pro accumulation. Reverse genetic analysis of several candidates identified new Pro effector genes, including thioredoxins and several genes encoding Universal Stress Protein A domain proteins. These new Pro effector genes further link Pro accumulation to cellular redox and energy status. Additional new Pro effector genes found include the mitochondrial protease LON1, ribosomal protein RPL24A, protein phosphatase 2A subunit A3, a MADS box protein, and a nucleoside triphosphate hydrolase. Several of these new Pro effector genes were from regions with multiple SNPs, each having moderate association with Pro accumulation. This pattern supports the use of summary approaches that incorporate clusters of SNP associations in addition to consideration of individual SNP probability values. Further GWAS-guided reverse genetics promises to find additional effectors of Pro accumulation. The combination of GWAS and reverse genetics to efficiently identify new effector genes may be especially applicable for traits difficult to analyze by other genetic screening methods. PMID:24218491

The Pro-inflammatory Effects of Glucocorticoids in the Brain

PubMed Central

Duque, Erica de Almeida; Munhoz, Carolina Demarchi

2016-01-01

Glucocorticoids are a class of steroid hormones derived from cholesterol. Their actions are mediated by the glucocorticoid and mineralocorticoid receptors, members of the superfamily of nuclear receptors, which, once bound to their ligands, act as transcription factors that can directly modulate gene expression. Through protein–protein interactions with other transcription factors, they can also regulate the activity of many genes in a composite or tethering way. Rapid non-genomic signaling was also demonstrated since glucocorticoids can act through membrane receptors and activate signal transduction pathways, such as protein kinases cascades, to modulate other transcriptions factors and activate or repress various target genes. By all these different mechanisms, glucocorticoids regulate numerous important functions in a large variety of cells, not only in the peripheral organs but also in the central nervous system during development and adulthood. In general, glucocorticoids are considered anti-inflammatory and protective agents due to their ability to inhibit gene expression of pro-inflammatory mediators and other possible damaging molecules. Nonetheless, recent studies have uncovered situations in which these hormones can act as pro-inflammatory agents depending on the dose, chronicity of exposure, and the structure/organ analyzed. In this review, we will provide an overview of the conditions under which these phenomena occur, a discussion that will serve as a basis for exploring the mechanistic foundation of glucocorticoids pro-inflammatory gene regulation in the brain. PMID:27445981

Functional polymorphisms in the sigma1 receptor gene associated with alcoholism.

PubMed

Miyatake, Ryosuke; Furukawa, Aizo; Matsushita, Sachio; Higuchi, Susumu; Suwaki, Hiroshi

2004-01-01

Sigma1 receptors are involved in the pathogenesis of drug abuse. Two polymorphisms (GC-241-240TT and Gln2Pro) in the sigma1 receptor gene (SIGMAR1) have been identified. To investigate the role of SIGMAR1 in conveying susceptibility to alcoholism, we performed a functional analysis of polymorphisms in the SIGMAR1 and a case-control study. We initially screened for polymorphisms in the 5'-upstream region. The effects of the polymorphisms on transcriptional activity were determined using a gene reporter assay. The distribution of SIGMAR1 polymorphisms was analyzed in 307 alcoholic and 302 control subjects. A novel T-485A polymorphism was identified. The transcriptional activity of the A-485 allele and the TT-241-240 allele was significantly reduced compared with that of the T-485 allele and the GC-241-240 allele. The frequencies of the A-485 allele (chi2=5.575, df=1, p=.0205) and the TT-241-240/Pro2 haplotype (chi2=21.464, df=1, p<.0001) were significantly higher in control subjects compared with alcoholic subjects. The T-485A and the GC-241-240TT may be functional polymorphisms, and the A-485 allele and TT-241-240/Pro2 haplotype are possible protective factors for the development of alcoholism.

Glucocorticoid Receptor-Mediated Repression of Pro-Inflammatory Genes in Rheumatoid Arthritis

DTIC Science & Technology

2015-10-01

1 AWARD NUMBER: W81XWH-14-1-0314 TITLE: Glucocorticoid Receptor-Mediated Repression of Pro-Inflammatory Genes in Rheumatoid Arthritis ...19 Sep 2015 4. TITLE AND SUBTITLE Glucocorticoid Receptor-Mediated Repression of Pro- Inflammatory Genes in Rheumatoid Arthritis 5a. CONTRACT NUMBER...SUBJECT TERMS Rheumatoid arthritis , inflammation and autoimmunity, macrophages, glucocorticoid receptor, transcriptional regulation, coactivators and

Cyclo(His-Pro) promotes cytoprotection by activating Nrf2-mediated up-regulation of antioxidant defence

PubMed Central

Minelli, Alba; Conte, Carmela; Grottelli, Silvia; Bellezza, Maria; Cacciatore, Ivana; Bolaños, Juan P

2009-01-01

Hystidyl-proline [cyclo(His-Pro)] is an endogenous cyclic dipeptide produced by the cleavage of thyrotropin releasing hormone. Previous studies have shown that cyclo(His-Pro) protects against oxidative stress, although the underlying mechanism has remained elusive. Here, we addressed this issue and found that cyclo(His-Pro) triggered nuclear accumulation of NF-E2-related factor-2 (Nrf2), a transcription factor that up-regulates antioxidant-/electrophile-responsive element (ARE-EpRE)-related genes, in PC12 cells. Cyclo(His-Pro) attenuated reactive oxygen species production, and prevented glutathione depletion caused by glutamate, rotenone, paraquat and β-amyloid treatment. Moreover, real-time PCR analyses revealed that cyclo(His-Pro) induced the expression of a number of ARE-related genes and protected cells against hydrogen peroxide-mediated apoptotic death. Furthermore, these effects were abolished by RNA interference-mediated Nrf2 knockdown. Finally, pharmacological inhibition of p-38 MAPK partially prevented both cyclo(His-Pro)-mediated Nrf2 activation and cellular protection. These results suggest that the signalling mechanism responsible for the cytoprotective actions of cyclo(His-Pro) would involve p-38 MAPK activation leading to Nrf2-mediated up-regulation of antioxidant cellular defence. PMID:18373731

Comparative Analysis of the Regulation of Expression and Structures of Two Evolutionarily Divergent Genes for Δ1-Pyrroline-5-Carboxylate Synthetase from Tomato1

PubMed Central

Fujita, Tomomichi; Maggio, Albino; Garcia-Rios, Mario; Bressan, Ray A.; Csonka, Laszlo N.

1998-01-01

We isolated two tomato (Lycopersicon esculentum) cDNA clones, tomPRO1 and tomPRO2, specifying Δ1-pyrroline-5-carboxylate synthetase (P5CS), the first enzyme of proline (Pro) biosynthesis. tomPRO1 is unusual because it resembles prokaryotic polycistronic operons (M.G. García-Ríos, T. Fujita, P.C. LaRosa, R.D. Locy, J.M. Clithero, R.A. Bressan, L.N. Csonka [1997] Proc Natl Acad Sci USA 94: 8249–8254), whereas tomPRO2 encodes a full-length P5CS. We analyzed the accumulation of Pro and the tomPRO1 and tomPRO2 messages in response to NaCl stress and developmental signals. Treatment with 200 mm NaCl resulted in a >60-fold increase in Pro levels in roots and leaves. However, there was a <3-fold increase in the accumulation of the tomPRO2 message and no detectable induction in the level of the tomPRO1 message in response to NaCl stress. Although pollen contained approximately 100-fold higher levels of Pro than other plant tissues, there was no detectable increase in the level of either message in pollen. We conclude that transcriptional regulation of these genes for P5CS is probably not important for the osmotic or pollen-specific regulation of Pro synthesis in tomato. Using restriction fragment-length polymorphism mapping, we determined the locations of tomPRO1 and tomPRO2 loci in the tomato nuclear genome. Sequence comparison suggested that tomPRO1 is similar to prokaryotic P5CS loci, whereas tomPRO2 is closely related to other eukaryotic P5CS genes. PMID:9765552

Characterization of CpSte11, a MAPKKK gene of Cryphonectria parasitica, and initial evidence of its involvement in the pheromone response pathway.

PubMed

Park, Jin-Ah; Kim, Jung-Mi; Park, Seung-Moon; Kim, Dae-Hyuk

2012-04-01

The gene CpSte11 of Cryphonectria parasitica, which encodes a yeast Ste11 homologue, was cloned and characterized. Gene replacement analysis revealed a high frequency of CpSte11 null mutants. When compared with the wild-type parent strain, CpSte11 null mutants showed no difference in terms of growth rate or pigmentation. However, CpSte11 null mutants showed a marked decrease in both the number and size of stromal pustules on chestnut twigs. The virulence test showed that, in comparison with those of the wild-type and virus-infected hypovirulent strains, CpSte11 null mutants produced necrotic areas of intermediate size. Disruption of the CpSte11 gene also resulted in defects in female fertility. Down-regulation of transcripts for the mating pheromone precursor gene, Mf2/2, and mating response transcription factors, such as cpst12 and pro1, was observed in CpSte11 null mutants. The down-regulation of Mf2/2, cpst12 and pro1 was also observed in the mutant phenotype of Cpmk2, a mating response Fus3-like mitogen-activated protein kinase (MAPK) gene, but not in the mutant of Cpmk1, a high-osmolarity glycerol Hog1-like MAPK gene. These results indicate that the cloned CpSte11 gene is functionally involved in the mating response pathway and acts through downstream targets, including Cpmk2, cpst12, pro1 and Mf2/2. However, the characteristics of the CpSte11 null mutant were fully phenocopied only in the cpst12 null mutant, but not in other studied null mutants of components of the putative mating response pathway. © 2011 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2011 BSPP AND BLACKWELL PUBLISHING LTD.

Effects of maternal dietary olive oil on pathways involved in diabetic embryopathy.

PubMed

Higa, Romina; Roberti, Sabrina Lorena; Musikant, Daniel; Mazzucco, María Belén; White, Verónica; Jawerbaum, Alicia

2014-11-01

Maternal diabetes induces a pro-oxidant/pro-inflammatory intrauterine environment related to the induction of congenital anomalies. Peroxisome proliferator activated receptors (PPARs) are transcription factors that regulate antioxidant and anti-inflammatory pathways. We investigated whether maternal diets supplemented with olive oil, enriched in oleic acid, a PPAR agonist, can regulate the expression of PPAR system genes, levels of lipoperoxidation and activity of matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) in embryos and decidua from diabetic rats. The embryos and decidua from diabetic rats showed reduced expression of PPARs and increased concentration of lipoperoxidation, MMPs and TIMPs, whereas the maternal treatments enriched in olive oil increased PPARδ in embryos and PPARγ and PPARγ-coactivator-1α expression in decidua, and increased TIMPs concentrations and decreased lipoperoxidation and MMPs activity in both tissues. Thus, maternal diets enriched in olive oil can regulate embryonic and decidual PPAR system genes expression and reduce the pro-oxidant/pro-inflammatory environment during rat early organogenesis. Copyright © 2014 Elsevier Inc. All rights reserved.

Vibrio cholerae ToxR downregulates virulence factor production in response to cyclo(Phe-Pro).

PubMed

Bina, X Renee; Taylor, Dawn L; Vikram, Amit; Ante, Vanessa M; Bina, James E

2013-08-27

Vibrio cholerae is an aquatic organism that causes the severe acute diarrheal disease cholera. The ability of V. cholerae to cause disease is dependent upon the production of two critical virulence determinants, cholera toxin (CT) and the toxin-coregulated pilus (TCP). The expression of the genes that encode for CT and TCP production is under the control of a hierarchical regulatory system called the ToxR regulon, which functions to activate virulence gene expression in response to in vivo stimuli. Cyclic dipeptides have been found to be produced by numerous bacteria, yet their biological function remains unknown. V. cholerae has been shown to produce cyclo(Phe-Pro). Previous studies in our laboratory demonstrated that cyclo(Phe-Pro) inhibited V. cholerae virulence factor production. For this study, we report on the mechanism by which cyclo(Phe-Pro) inhibited virulence factor production. We have demonstrated that exogenous cyclo(Phe-Pro) activated the expression of leuO, a LysR-family regulator that had not been previously associated with V. cholerae virulence. Increased leuO expression repressed aphA transcription, which resulted in downregulation of the ToxR regulon and attenuated CT and TCP production. The cyclo(Phe-Pro)-dependent induction of leuO expression was found to be dependent upon the virulence regulator ToxR. Cyclo(Phe-Pro) did not affect toxR transcription or ToxR protein levels but appeared to enhance the ToxR-dependent transcription of leuO. These results have identified leuO as a new component of the ToxR regulon and demonstrate for the first time that ToxR is capable of downregulating virulence gene expression in response to an environmental cue. The ToxR regulon has been a focus of cholera research for more than three decades. During this time, a model has emerged wherein ToxR functions to activate the expression of Vibrio cholerae virulence factors upon host entry. V. cholerae and other enteric bacteria produce cyclo(Phe-Pro), a cyclic dipeptide that we identified as an inhibitor of V. cholerae virulence factor production. This finding suggested that cyclo(Phe-Pro) was a negative effector of virulence factor production and represented a molecule that could potentially be exploited for therapeutic development. In this work, we investigated the mechanism by which cyclo(Phe-Pro) inhibited virulence factor production. We found that cyclo(Phe-Pro) signaled through ToxR to activate the expression of leuO, a new virulence regulator that functioned to repress virulence factor production. Our results have identified a new arm of the ToxR regulon and suggest that ToxR may play a broader role in pathogenesis than previously known.

Adjunctive Dexamethasone Affects the Expression of Genes Related to Inflammation, Neurogenesis and Apoptosis in Infant Rat Pneumococcal Meningitis

PubMed Central

Blaser, Cornelia; Wittwer, Matthias; Grandgirard, Denis; Leib, Stephen L.

2011-01-01

Streptococcus pneumoniae is the most common pathogen causing non-epidemic bacterial meningitis worldwide. The immune response and inflammatory processes contribute to the pathophysiology. Hence, the anti-inflammatory dexamethasone is advocated as adjuvant treatment although its clinical efficacy remains a question at issue. In experimental models of pneumococcal meningitis, dexamethasone increased neuronal damage in the dentate gyrus. Here, we investigated expressional changes in the hippocampus and cortex at 72 h after infection when dexamethasone was given to infant rats with pneumococcal meningitis. Nursing Wistar rats were intracisternally infected with Streptococcus pneumoniae to induce experimental meningitis or were sham-infected with pyrogen-free saline. Besides antibiotics, animals were either treated with dexamethasone or saline. Expressional changes were assessed by the use of GeneChip® Rat Exon 1.0 ST Arrays and quantitative real-time PCR. Protein levels of brain-derived neurotrophic factor, cytokines and chemokines were evaluated in immunoassays using Luminex xMAP® technology. In infected animals, 213 and 264 genes were significantly regulated by dexamethasone in the hippocampus and cortex respectively. Separately for the cortex and the hippocampus, Gene Ontology analysis identified clusters of biological processes which were assigned to the predefined categories “inflammation”, “growth”, “apoptosis” and others. Dexamethasone affected the expression of genes and protein levels of chemokines reflecting diminished activation of microglia. Dexamethasone-induced changes of genes related to apoptosis suggest the downregulation of the Akt-survival pathway and the induction of caspase-independent apoptosis. Signalling of pro-neurogenic pathways such as transforming growth factor pathway was reduced by dexamethasone resulting in a lack of pro-survival triggers. The anti-inflammatory properties of dexamethasone were observed on gene and protein level in experimental pneumococcal meningitis. Further dexamethasone-induced expressional changes reflect an increase of pro-apoptotic signals and a decrease of pro-neurogenic processes. The findings may help to identify potential mechanisms leading to apoptosis by dexamethasone in experimental pneumococcal meningitis. PMID:21412436

St. John's wort extract and hyperforin inhibit multiple phosphorylation steps of cytokine signaling and prevent inflammatory and apoptotic gene induction in pancreatic β cells.

PubMed

Novelli, Michela; Menegazzi, Marta; Beffy, Pascale; Porozov, Svetlana; Gregorelli, Alex; Giacopelli, Daniela; De Tata, Vincenzo; Masiello, Pellegrino

2016-12-01

The extract of the herbaceous plant St. John's wort (SJW) and its phloroglucinol component hyperforin (HPF) were previously shown to inhibit cytokine-induced STAT-1 and NF-κB activation and prevent damage in pancreatic β cells. To further clarify the mechanisms underlying their protective effects, we evaluated the phosphorylation state of various factors of cytokine signaling pathways and the expression of target genes involved in β-cell function, inflammatory response and apoptosis induction. In the INS-1E β-cell line, exposed to a cytokine mixture with/without SJW extract (2-5μg/ml) or HPF (1-5μM), protein phosphorylation was assessed by western blotting and expression of target genes by real-time quantitative PCR. SJW and HPF markedly inhibited, in a dose-dependent manner (from 60 to 100%), cytokine-induced activating phosphorylations of STAT-1, NF-κB p65 subunit and IKK (NF-κB inhibitory subunit IκBα kinase). MAPK and Akt pathways were also modulated by the vegetal compounds through hindrance of p38 MAPK, ERK1/2, JNK and Akt phosphorylations, each reduced by at least 65% up to 100% at the higher dose. Consistently, SJW and HPF a) abolished cytokine-induced mRNA expression of pro-inflammatory genes; b) avoided down-regulation of relevant β-cell functional/differentiation genes; c) corrected cytokine-driven imbalance between pro- and anti-apoptotic factors, by fully preventing up-regulation of pro-apoptotic genes and preserving expression or function of anti-apoptotic Bcl-2 family members; d) protected INS-1E cells against cytokine-induced apoptosis. In conclusion, SJW extract and HPF exert their protective effects through simultaneous inhibition of multiple phosphorylation steps along various cytokine signaling pathways and consequent restriction of inflammatory and apoptotic gene expression. Thus, they have a promising therapeutic potential for the prevention or limitation of immune-mediated β-cell dysfunction and damage leading to type 1 diabetes. Copyright © 2016 Elsevier Ltd. All rights reserved.

Lung cancer risk associated with Thr495Pro polymorphism of GHR in Chinese population.

PubMed

Cao, Guochun; Lu, Hongna; Feng, Jifeng; Shu, Jian; Zheng, Datong; Hou, Yayi

2008-04-01

The incidence of lung cancer has been increasing over recent decades. Previous studies showed that polymorphisms of the genes involved in carcinogen-detoxication, DNA repair and cell cycle control comprise risk factors for lung cancer. Recent observations revealed that the growth hormone receptor (GHR) might play important roles in carcinogenesis and Rudd et al. found that the Thr495Pro polymorphism of GHR was strongly associated with lung cancer risk in Caucasians living in the UK (OR = 12.98, P = 0.0019, 95% CI: 1.77-infinity). To test whether this variant of GHR would modify the risk of lung cancer in Chinese population, we compared the polymorphism between 778 lung cancer patients and 781 healthy control subjects. Our results indicate that the frequency of 495Thr (2.8%) allele in cases was significantly higher than in controls (OR = 2.04, P = 0.006, 95% CI: 1.21-3.42) which indicated this allele might be a risk factor for lung cancer. Further analyses revealed Thr495Pro variant was associated with lung cancer in the subpopulation with higher risk for lung cancer: male subpopulation, still-smokers subpopulation and the subpopulation with familial history of cancer. In different histological types of lung cancer, Thr495Pro SNP was significantly associated with small cell and squamous cell lung cancer, but not with adenocarcinoma, which suggested a potential interaction between this polymorphism and metabolic pathways related to smoking. The potential gene-environment interaction on lung cancer risk was evaluated using MDR software. A significant redundant interaction between Thr495Pro polymorphism and smoking dose and familial history of cancer was identified and the combination of genetic factors and smoking status or familial history of cancer barely increased the cancer risk prediction accuracy. In conclusion, our results suggested that the Thr495Pro polymorphism of GHR was associated with the risk of lung cancer in a redundant interaction with smoking and familial history of cancer.

Butyrate modulating effects on pro-inflammatory pathways in human intestinal epithelial cells.

PubMed

Elce, A; Amato, F; Zarrilli, F; Calignano, A; Troncone, R; Castaldo, G; Canani, R B

2017-10-13

Butyrate acts as energy source for intestinal epithelial cells and as key mediator of several immune processes, modulating gene expression mainly through histone deacetylation inhibition. Thanks to these effects, butyrate has been proposed for the treatment of many intestinal diseases. Aim of this study was to investigate the effect of butyrate on the expression of a large series of target genes encoding proteins involved in pro-inflammatory pathways. We performed quantitative real-time-PCR analysis of the expression of 86 genes encoding proteins bearing to pro-inflammatory pathways, before and after butyrate exposure, in primary epithelial cells derived from human small intestine and colon. Butyrate significantly down-regulated the expression of genes involved in inflammatory response, among which nuclear factor kappa beta, interferon-gamma, Toll like 2 receptor and tumour necrosis factor-alpha. Further confirmations of these data, including studies at protein level, would support the use of butyrate as effective therapeutic strategy in intestinal inflammatory disorders.

A family with the Arg103Pro mutation in the NEUROD1 gene detected by next-generation sequencing - Clinical characteristics of mutation carriers.

PubMed

Szopa, Magdalena; Ludwig-Galezowska, Agnieszka H; Radkowski, Piotr; Skupien, Jan; Machlowska, Julita; Klupa, Tomasz; Wolkow, Pawel; Borowiec, Maciej; Mlynarski, Wojciech; Malecki, Maciej T

2016-02-01

Until now only a few families with early onset autosomal diabetes due to the NEUROD1 gene mutations have been identified. Moreover, only some of them meet strict MODY (maturity-onset diabetes of the young) criteria. Next-generation sequencing (NGS) provides an opportunity to detect more pathogenic mutations in this gene. Here, we evaluated the segregation of the Arg103Pro mutation in the NEUROD1 gene in a pedigree in which it was detected, and described the clinical characteristics of the mutation carriers. We included 156 diabetic probands of MODY families, among them 52 patients earlier tested for GCK-MODY and/or HNF1A-MODY by Sanger sequencing with negative results. Genetic testing was performed by targeted NGS sequencing using a panel of 28 monogenic diabetes genes. As detected by NGS, one patient had the missense Arg103Pro (CGC/CCC) mutation in the gene NEUROD1 changing the amino-acid structure of the DNA binding domain of this transcription factor. We confirmed this sequence difference by Sanger sequencing. This family had previously been tested with negative results for HNF1A gene mutations. 17 additional members of this family were invited for further testing. We confirmed the presence of the mutation in 11 subjects. Seven adult mutation carriers (all but one) from three generations had been already diagnosed with diabetes. There were 3 individuals with the Arg103Pro mutation diagnosed before the age of 30 years in the family. The range of age of the four unaffected mutation carriers (3 minors and 1 adult) was 3-48 years. Interestingly, one mutation carrier had a history of transient neonatal hypoglycemia, of which the clinical course resembled episodes typical for HNF4A-MODY. We report a family with autosomal dominant diabetes related to a new NEUROD1 mutation, one of very few meeting MODY criteria. The use of the NGS method will facilitate identification of more families with rare forms of MODY. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

β3 Integrin Haplotype Influences Gene Regulation and Plasma von Willebrand Factor Activity

PubMed Central

Payne, Katie E; Bray, Paul F; Grant, Peter J; Carter, Angela M

2008-01-01

The Leu33Pro polymorphism of the gene encoding β3 integrin (ITGB3) is associated with acute coronary syndromes and influences platelet aggregation. Three common promoter polymorphisms have also been identified. The aims of this study were to (1) investigate the influence of the ITGB3 −400C/A, −425A/C and −468G/A promoter polymorphisms on reporter gene expression and nuclear protein binding and (2) determine genotype and haplotype associations with platelet αIIbβ3 receptor density. Promoter haplotypes were introduced into an ITGB3 promoter-pGL3 construct by site directed mutagenesis and luciferase reporter gene expression analysed in HEL and HMEC-1 cells. Binding of nuclear proteins was assessed by electrophoretic mobility shift assay. The association of ITGB3 haplotype with platelet αIIbβ3 receptor density was determined in 223 subjects. Species conserved motifs were identified in the ITGB3 promoter in the vicinity of the 3 polymorphisms. The GAA, GCC, AAC, AAA and ACC constructs induced ~50% increased luciferase expression relative to the GAC construct in both cell types. Haplotype analysis including Leu33Pro indicated 5 common haplotypes; no associations between ITGB3 haplotypes and receptor density were found. However, the GCC-Pro33 haplotype was associated with significantly higher vWF activity (128.6 [112.1–145.1]%) compared with all other haplotypes (107.1 [101.2–113.0]%, p=0.02). In conclusion, the GCC-Pro33 haplotype was associated with increased vWF activity but not with platelet αIIbβ3 receptor density, which may indicate ITGB3 haplotype influences endothelial function. PMID:18045606

The cachectic mediator proteolysis inducing factor activates NF-kappaB and STAT3 in human Kupffer cells and monocytes.

PubMed

Watchorn, Tammy M; Dowidar, Nabil; Dejong, Cornelis H C; Waddell, Ian D; Garden, O James; Ross, James A

2005-10-01

A novel proteoglycan, proteolysis inducing factor (PIF), is capable of inducing muscle proteolysis during the process of cancer cachexia, and of inducing an acute phase response in human hepatocytes. We investigated whether PIF is able to activate pro-inflammatory pathways in human Kupffer cells, the resident macrophages of the liver, and in monocytes, resulting in the production of pro-inflammatory cytokines. Normal liver tissue was obtained from patients undergoing partial hepatectomy and Kupffer cells were isolated. Monocytes were isolated from peripheral blood. Following exposure to native PIF, pro-inflammatory cytokine production from Kupffer cells and monocytes was measured and the NF-kappaB and STAT3 transcriptional pathways were investigated using electrophoretic mobility shift assays. We demonstrate that PIF is able to activate the transcription factor NF-kappaB and NF-kappaB-inducible genes in human Kupffer cells, and in monocytes, resulting in the production of pro-inflammatory cytokines such as TNF-alpha, IL-8 and IL-6. PIF enhances the expression of the cell surface molecules LFA-1 and CD14 on macrophages. PIF also activates the transcription factor STAT3 in Kupffer cells. The pro-inflammatory effects of PIF, mediated via NF-kappaB and STAT3, are important in macrophage behaviour and may contribute to the inflammatory pro-cachectic process in the liver.

Association of mRNA expression of TP53 and the TP53 codon 72 Arg/Pro gene polymorphism with colorectal cancer risk in Asian population: a bioinformatics analysis and meta-analysis.

PubMed

Dong, Zhiyong; Zheng, Longzhi; Liu, Weimin; Wang, Cunchuan

2018-01-01

The relationship between TP53 codon 72 Pro/Arg gene polymorphism and colorectal cancer risk in Asians is still controversial, and this bioinformatics analysis and meta-analysis was performed to assess the associations. The association studies were identified from PubMed, and eligible reports were included. RevMan 5.3.1 software, Oncolnc, cBioPortal, and Oncomine online tools were used for statistical analysis. A random/fixed effects model was used in meta-analysis. The data were reported as risk ratios or mean differences with corresponding 95% CI. We confirmed that TP53 was associated with colorectal cancer, the alteration frequency of TP53 was 53% mutation and 7% deep deletion, and TP53 mRNA expression was different in different types of colorectal cancer based on The Cancer Genome Atlas database. Then, 18 studies were included that examine the association of TP53 codon 72 gene polymorphism with colorectal cancer risk in Asians. The meta-analysis indicated that TP53 Pro allele and Pro/Pro genotype were associated with colorectal cancer risk in Asian population, but Arg/Arg genotype was not (Pro allele: odds ratios [OR]=1.20, 95% CI: 1.06 to 1.35, P =0.003; Pro/Pro genotype: OR=1.39, 95% CI: 1.15 to 1.69, P =0.0007; Arg/Arg genotype: OR=0.86, 95% CI: 0.74 to 1.00, P =0.05). Interestingly, in the meta-analysis of the controls from the population-based studies, we found that TP53 codon 72 Pro/Arg gene polymorphism was associated with colorectal cancer risk (Pro allele: OR=1.33, 95% CI: 1.15 to 1.55, P =0.0002; Pro/Pro genotype: OR=1.61, 95% CI: 1.28 to 2.02, P <0.0001; Arg/Arg genotype: OR=0.77, 95% CI: 0.63 to 0.93, P =0.009). TP53 was associated with colorectal cancer, but the different value levels of mRNA expression were not associated with survival rate of colon and rectal cancer. TP53 Pro allele and Pro/Pro genotype were associated with colorectal cancer risk in Asians.

Association of mRNA expression of TP53 and the TP53 codon 72 Arg/Pro gene polymorphism with colorectal cancer risk in Asian population: a bioinformatics analysis and meta-analysis

PubMed Central

Liu, Weimin; Wang, Cunchuan

2018-01-01

Background The relationship between TP53 codon 72 Pro/Arg gene polymorphism and colorectal cancer risk in Asians is still controversial, and this bioinformatics analysis and meta-analysis was performed to assess the associations. Methods The association studies were identified from PubMed, and eligible reports were included. RevMan 5.3.1 software, Oncolnc, cBioPortal, and Oncomine online tools were used for statistical analysis. A random/fixed effects model was used in meta-analysis. The data were reported as risk ratios or mean differences with corresponding 95% CI. Results We confirmed that TP53 was associated with colorectal cancer, the alteration frequency of TP53 was 53% mutation and 7% deep deletion, and TP53 mRNA expression was different in different types of colorectal cancer based on The Cancer Genome Atlas database. Then, 18 studies were included that examine the association of TP53 codon 72 gene polymorphism with colorectal cancer risk in Asians. The meta-analysis indicated that TP53 Pro allele and Pro/Pro genotype were associated with colorectal cancer risk in Asian population, but Arg/Arg genotype was not (Pro allele: odds ratios [OR]=1.20, 95% CI: 1.06 to 1.35, P=0.003; Pro/Pro genotype: OR=1.39, 95% CI: 1.15 to 1.69, P=0.0007; Arg/Arg genotype: OR=0.86, 95% CI: 0.74 to 1.00, P=0.05). Interestingly, in the meta-analysis of the controls from the population-based studies, we found that TP53 codon 72 Pro/Arg gene polymorphism was associated with colorectal cancer risk (Pro allele: OR=1.33, 95% CI: 1.15 to 1.55, P=0.0002; Pro/Pro genotype: OR=1.61, 95% CI: 1.28 to 2.02, P<0.0001; Arg/Arg genotype: OR=0.77, 95% CI: 0.63 to 0.93, P=0.009). Conclusion TP53 was associated with colorectal cancer, but the different value levels of mRNA expression were not associated with survival rate of colon and rectal cancer. TP53 Pro allele and Pro/Pro genotype were associated with colorectal cancer risk in Asians. PMID:29872345

Proline Coordination with Fatty Acid Synthesis and Redox Metabolism of Chloroplast and Mitochondria1[OPEN

PubMed Central

Shinde, Suhas; Villamor, Joji Grace; Lin, Wendar; Verslues, Paul E.

2016-01-01

Proline (Pro) accumulation is one of the most prominent changes in plant metabolism during drought and low water potential; however, the regulation and function of Pro metabolism remain unclear. We used a combination of forward genetic screening based on a Proline Dehydrogenase1 (PDH1) promoter-luciferase reporter (PDH1pro:LUC2) and RNA sequencing of the Pro synthesis mutant p5cs1-4 to identify multiple loci affecting Pro accumulation in Arabidopsis (Arabidopsis thaliana). Two mutants having high PDH1pro:LUC2 expression and increased Pro accumulation at low water potential were found to be alleles of Cytochrome P450, Family 86, Subfamily A, Polypeptide2 (CYP86A2) and Long Chain Acyl Synthetase2 (LACS2), which catalyze two successive steps in very-long-chain fatty acid (VLCFA) synthesis. Reverse genetic experiments found additional VLCFA and lipid metabolism-related mutants with increased Pro accumulation. Altered cellular redox status is a key factor in the coordination of Pro and VLCFA metabolism. The NADPH oxidase inhibitor diphenyleneiodonium (DPI) induced high levels of Pro accumulation and strongly repressed PDH1pro:LUC2 expression. cyp86a2 and lacs2 mutants were hypersensitive to diphenyleneiodonium but could be reverted to wild-type Pro and PDH1pro:LUC2 expression by reactive oxygen species scavengers. The coordination of Pro and redox metabolism also was indicated by the altered expression of chloroplast and mitochondria electron transport genes in p5cs1-4. These results show that Pro metabolism is both influenced by and influences cellular redox status via previously unknown coordination with several metabolic pathways. In particular, Pro and VLCFA synthesis share dual roles to help buffer cellular redox status while producing products useful for stress resistance, namely the compatible solute Pro and cuticle lipids. PMID:27512016

Altered Cytokine Gene Expression in Peripheral Blood Monocytes across the Menstrual Cycle in Primary Dysmenorrhea: A Case-Control Study

PubMed Central

Ma, Hongyue; Hong, Min; Duan, Jinao; Liu, Pei; Fan, Xinsheng; Shang, Erxin; Su, Shulan; Guo, Jianming; Qian, Dawei; Tang, Yuping

2013-01-01

Primary dysmenorrhea is one of the most common gynecological complaints in young women, but potential peripheral immunologic features underlying this condition remain undefined. In this paper, we compared 84 common cytokine gene expression profiles of peripheral blood mononuclear cells (PBMCs) from six primary dysmenorrheic young women and three unaffected controls on the seventh day before (secretory phase), and the first (menstrual phase) and the fifth (regenerative phase) days of menstruation, using a real-time PCR array assay combined with pattern recognition and gene function annotation methods. Comparisons between dysmenorrhea and normal control groups identified 11 (nine increased and two decreased), 14 (five increased and nine decreased), and 15 (seven increased and eight decreased) genes with ≥2-fold difference in expression (P<0.05) in the three phases of menstruation, respectively. In the menstrual phase, genes encoding pro-inflammatory cytokines (IL1B, TNF, IL6, and IL8) were up-regulated, and genes encoding TGF-β superfamily members (BMP4, BMP6, GDF5, GDF11, LEFTY2, NODAL, and MSTN) were down-regulated. Functional annotation revealed an excessive inflammatory response and insufficient TGF-β superfamily member signals with anti-inflammatory consequences, which may directly contribute to menstrual pain. In the secretory and regenerative phases, increased expression of pro-inflammatory cytokines and decreased expression of growth factors were also observed. These factors may be involved in the regulation of decidualization, endometrium breakdown and repair, and indirectly exacerbate primary dysmenorrhea. This first study of cytokine gene expression profiles in PBMCs from young primary dysmenorrheic women demonstrates a shift in the balance between expression patterns of pro-inflammatory cytokines and TGF-β superfamily members across the whole menstrual cycle, underlying the peripheral immunologic features of primary dysmenorrhea. PMID:23390521

Pro12Ala PPAR γ2 gene polymorphism in PCOS women: the role of compounds regulating satiety.

PubMed

Bidzińska-Speichert, Bożena; Lenarcik, Agnieszka; Tworowska-Bardzińska, Urszula; Slęzak, Ryszard; Bednarek-Tupikowska, Grażyna; Milewicz, Andrzej

2012-03-01

Five to ten percent of women of reproductive age suffer from polycystic ovary syndrome (PCOS). Leptin, NPY, galanin, cholecystokinin (CCK) are involved in the regulation of eating behavior. PPARγ are receptors that are probably involved in hyperandrogenism. This study was designed to assess associations between the Pro12Ala PPARγ2 gene polymorphism and satiety factors in PCOS. Fifty-four PCOS women and 51 healthy women were studied. Leptin, NPY, galanin, CCK levels, and genetic studies to detect Pro12Ala PPARγ2 gene polymorphism were assessed. The leptin levels in the PCOS women carrying Pro12Ala genotype were higher than in those with Pro12Pro and Ala12Ala. The PCOS women had higher leptin and NPY levels and lower galanin levels. Obese PCOS patients had lower CCK levels. In the PCOS women, a single Ala allele may have a protective role as far as hyperleptinemia is concerned. The PCOS women may reveal a disrupted central leptin/NPY feedback loop with some shifts in food intake.

KLF5 regulates infection- and inflammation-induced pro-labour mediators in human myometrium.

PubMed

Lappas, Martha

2015-05-01

The transcription factor Kruppel-like factor 5 (KLF5) has been shown to associate with nuclear factor kappa B (NFκB) to regulate genes involved in inflammation. However, there are no studies on the expression and regulation of KLF5 in the processes of human labour and delivery. Thus, the aims of this study were to determine the effect of i) human labour on KLF5 expression in both foetal membranes and myometrium; ii) the pro-inflammatory cytokine interleukin 1 beta (IL1β), bacterial product flagellin and the viral dsRNA analogue poly(I:C) on KLF5 expression and iii) KLF5 knockdown by siRNA in human myometrial primary cells on pro-inflammatory and pro-labour mediators. In foetal membranes, there was no effect of term or preterm labour on KLF5 expression. In myometrium, the term labour was associated with an increase in nuclear KLF5 protein expression. Moreover, KLF5 expression was also increased in myometrial cells treated with IL1β, flagellin or poly(IC), likely factors contributing to preterm birth. KLF5 silencing in myometrial cells significantly decreased IL1β-induced cytokine expression (IL6 and IL8 mRNA expression and release), COX2 mRNA expression, and subsequent release of prostaglandins PGE2 and PGF2 α. KLF5 silencing also significantly reduced flagellin- and poly(I:C)-induced IL6 and IL8 mRNA expression. Lastly, IL1β-, flagellin- and poly(I:C)-stimulated NFκB transcriptional activity was significantly suppressed in KLF5-knockout myometrial cells. In conclusion, this study describes novel data in which KLF5 is increased in labouring myometrium, and KLF5 silencing decreased inflammation- and infection-induced pro-labour mediators. © 2015 Society for Reproduction and Fertility.

Maternal obesity is associated with ovarian inflammation and up-regulation of early growth response factor 1

USDA-ARS?s Scientific Manuscript database

Obesity impairs reproductive functions through multiple mechanisms, possibly through disruption of ovarian function. We hypothesized that increased adiposity will lead to a pro-inflammatory gene signature and up-regulation of Egr-1 protein in ovaries from obese (OB, n=7) compared to lean (LN, n=10) ...

The transcription factor MEF2C mediates cardiomyocyte hypertrophy induced by IGF-1 signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Munoz, Juan Pablo; Collao, Andres; Chiong, Mario

2009-10-09

Myocyte enhancer factor 2C (MEF2C) plays an important role in cardiovascular development and is a key transcription factor for cardiac hypertrophy. Here, we describe MEF2C regulation by insulin-like growth factor-1 (IGF-1) and its role in IGF-1-induced cardiac hypertrophy. We found that IGF-1 addition to cultured rat cardiomyocytes activated MEF2C, as evidenced by its increased nuclear localization and DNA binding activity. IGF-1 stimulated MEF2 dependent-gene transcription in a time-dependent manner, as indicated by increased MEF2 promoter-driven reporter gene activity; IGF-1 also induced p38-MAPK phosphorylation, while an inhibitor of p38-MAPK decreased both effects. Additionally, inhibitors of phosphatidylinositol 3-kinase and calcineurin prevented IGF-1-inducedmore » MEF2 transcriptional activity. Via MEF2C-dependent signaling, IGF-1 also stimulated transcription of atrial natriuretic factor and skeletal {alpha}-actin but not of fos-lux reporter genes. These novel data suggest that MEF2C activation by IGF-1 mediates the pro-hypertrophic effects of IGF-1 on cardiac gene expression.« less

Proline Coordination with Fatty Acid Synthesis and Redox Metabolism of Chloroplast and Mitochondria.

PubMed

Shinde, Suhas; Villamor, Joji Grace; Lin, Wendar; Sharma, Sandeep; Verslues, Paul E

2016-10-01

Proline (Pro) accumulation is one of the most prominent changes in plant metabolism during drought and low water potential; however, the regulation and function of Pro metabolism remain unclear. We used a combination of forward genetic screening based on a Proline Dehydrogenase1 (PDH1) promoter-luciferase reporter (PDH1 pro :LUC2) and RNA sequencing of the Pro synthesis mutant p5cs1-4 to identify multiple loci affecting Pro accumulation in Arabidopsis (Arabidopsis thaliana). Two mutants having high PDH1 pro :LUC2 expression and increased Pro accumulation at low water potential were found to be alleles of Cytochrome P450, Family 86, Subfamily A, Polypeptide2 (CYP86A2) and Long Chain Acyl Synthetase2 (LACS2), which catalyze two successive steps in very-long-chain fatty acid (VLCFA) synthesis. Reverse genetic experiments found additional VLCFA and lipid metabolism-related mutants with increased Pro accumulation. Altered cellular redox status is a key factor in the coordination of Pro and VLCFA metabolism. The NADPH oxidase inhibitor diphenyleneiodonium (DPI) induced high levels of Pro accumulation and strongly repressed PDH1 pro :LUC2 expression. cyp86a2 and lacs2 mutants were hypersensitive to diphenyleneiodonium but could be reverted to wild-type Pro and PDH1 pro :LUC2 expression by reactive oxygen species scavengers. The coordination of Pro and redox metabolism also was indicated by the altered expression of chloroplast and mitochondria electron transport genes in p5cs1-4 These results show that Pro metabolism is both influenced by and influences cellular redox status via previously unknown coordination with several metabolic pathways. In particular, Pro and VLCFA synthesis share dual roles to help buffer cellular redox status while producing products useful for stress resistance, namely the compatible solute Pro and cuticle lipids. © 2016 American Society of Plant Biologists. All Rights Reserved.

Branched-chain amino acids prevent hepatic fibrosis and development of hepatocellular carcinoma in a non-alcoholic steatohepatitis mouse model

PubMed Central

Takegoshi, Kai; Honda, Masao; Okada, Hikari; Takabatake, Riuta; Matsuzawa-Nagata, Naoto; Campbell, Jean S.; Nishikawa, Masashi; Shimakami, Tetsuro; Shirasaki, Takayoshi; Sakai, Yoshio; Yamashita, Taro; Takamura, Toshinari; Tanaka, Takuji; Kaneko, Shuichi

2017-01-01

Oral supplementation with branched-chain amino acids (BCAA; leucine, isoleucine, and valine) in patients with liver cirrhosis potentially suppresses the incidence of hepatocellular carcinoma (HCC) and improves event-free survival. However, the detailed mechanisms of BCAA action have not been fully elucidated. BCAA were administered to atherogenic and high-fat (Ath+HF) diet-induced nonalcoholic steatohepatitis (NASH) model mice. Liver histology, tumor incidence, and gene expression profiles were evaluated. Ath+HF diet mice developed hepatic tumors at a high frequency at 68 weeks. BCAA supplementation significantly improved hepatic steatosis, inflammation, fibrosis, and tumors in Ath+HF mice at 68 weeks. GeneChip analysis demonstrated the significant resolution of pro-fibrotic gene expression by BCAA supplementation. The anti-fibrotic effect of BCAA was confirmed further using platelet-derived growth factor C transgenic mice, which develop hepatic fibrosis and tumors. In vitro, BCAA restored the transforming growth factor (TGF)-β1-stimulated expression of pro-fibrotic genes in hepatic stellate cells (HSC). In hepatocytes, BCAA restored TGF-β1-induced apoptosis, lipogenesis, and Wnt/β-Catenin signaling, and inhibited the transformation of WB-F344 rat liver epithelial stem-like cells. BCAA repressed the promoter activity of TGFβ1R1 by inhibiting the expression of the transcription factor NFY and histone acetyltransferase p300. Interestingly, the inhibitory effect of BCAA on TGF-β1 signaling was mTORC1 activity-dependent, suggesting the presence of negative feedback regulation from mTORC1 to TGF-β1 signaling. Thus, BCAA induce an anti-fibrotic effect in HSC, prevent apoptosis in hepatocytes, and decrease the incidence of HCC; therefore, BCAA supplementation would be beneficial for patients with advanced liver fibrosis with a high risk of HCC. PMID:28212548

Branched-chain amino acids prevent hepatic fibrosis and development of hepatocellular carcinoma in a non-alcoholic steatohepatitis mouse model.

PubMed

Takegoshi, Kai; Honda, Masao; Okada, Hikari; Takabatake, Riuta; Matsuzawa-Nagata, Naoto; Campbell, Jean S; Nishikawa, Masashi; Shimakami, Tetsuro; Shirasaki, Takayoshi; Sakai, Yoshio; Yamashita, Taro; Takamura, Toshinari; Tanaka, Takuji; Kaneko, Shuichi

2017-03-14

Oral supplementation with branched-chain amino acids (BCAA; leucine, isoleucine, and valine) in patients with liver cirrhosis potentially suppresses the incidence of hepatocellular carcinoma (HCC) and improves event-free survival. However, the detailed mechanisms of BCAA action have not been fully elucidated. BCAA were administered to atherogenic and high-fat (Ath+HF) diet-induced nonalcoholic steatohepatitis (NASH) model mice. Liver histology, tumor incidence, and gene expression profiles were evaluated. Ath+HF diet mice developed hepatic tumors at a high frequency at 68 weeks. BCAA supplementation significantly improved hepatic steatosis, inflammation, fibrosis, and tumors in Ath+HF mice at 68 weeks. GeneChip analysis demonstrated the significant resolution of pro-fibrotic gene expression by BCAA supplementation. The anti-fibrotic effect of BCAA was confirmed further using platelet-derived growth factor C transgenic mice, which develop hepatic fibrosis and tumors. In vitro, BCAA restored the transforming growth factor (TGF)-β1-stimulated expression of pro-fibrotic genes in hepatic stellate cells (HSC). In hepatocytes, BCAA restored TGF-β1-induced apoptosis, lipogenesis, and Wnt/β-Catenin signaling, and inhibited the transformation of WB-F344 rat liver epithelial stem-like cells. BCAA repressed the promoter activity of TGFβ1R1 by inhibiting the expression of the transcription factor NFY and histone acetyltransferase p300. Interestingly, the inhibitory effect of BCAA on TGF-β1 signaling was mTORC1 activity-dependent, suggesting the presence of negative feedback regulation from mTORC1 to TGF-β1 signaling. Thus, BCAA induce an anti-fibrotic effect in HSC, prevent apoptosis in hepatocytes, and decrease the incidence of HCC; therefore, BCAA supplementation would be beneficial for patients with advanced liver fibrosis with a high risk of HCC.

Irradiation induces regionally specific alterations in pro-inflammatory environments in rat brain

PubMed Central

Lee, Won Hee; Sonntag, William E.; Mitschelen, Matthew; Yan, Han; Lee, Yong Woo

2010-01-01

Purpose Pro-inflammatory environments in the brain have been implicated in the onset and progression of neurological disorders. In the present study, we investigate the hypothesis that brain irradiation induces regionally specific alterations in cytokine gene and protein expression. Materials and methods Four month old F344 × BN rats received either whole brain irradiation with a single dose of 10 Gy γ-rays or sham-irradiation, and were maintained for 4, 8, and 24 h following irradiation. The mRNA and protein expression levels of pro-inflammatory mediators were analysed by real-time reverse transcriptase-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and immunofluorescence staining. To elucidate the molecular mechanisms of irradiation-induced brain inflammation, effects of irradiation on the DNA-binding activity of pro-inflammatory transcription factors were also examined. Results A significant and marked up-regulation of mRNA and protein expression of pro-inflammatory mediators, including tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and monocyte chemoattractant protein-1 (MCP-1), was observed in hippocampal and cortical regions isolated from irradiated brain. Cytokine expression was regionally specific since TNF-α levels were significantly elevated in cortex compared to hippocampus (57% greater) and IL-1β levels were elevated in hippocampus compared to cortical samples (126% greater). Increases in cytokine levels also were observed after irradiation of mouse BV-2 microglial cells. A series of electrophoretic mobility shift assays (EMSA) demonstrated that irradiation significantly increased activation of activator protein-1 (AP-1), nuclear factor-κB (NF-κB), and cAMP response element-binding protein (CREB). Conclusion The present study demonstrated that whole brain irradiation induces regionally specific pro-inflammatory environments through activation of AP-1, NF-κB, and CREB and overexpression of TNF-α, IL-1β, and MCP-1 in rat brain and may contribute to unique pathways for the radiation-induced impairments in tissue function. PMID:20148699

Optimization of culture conditions for stem cells derived from human anterior cruciate ligament and bone marrow.

PubMed

Cheng, Ming-Te; Liu, Chien-Lin; Chen, Tain-Hsiung; Lee, Oscar K

2014-01-01

Tissue engineering with stem cells is a fascinating approach for treating anterior cruciate ligament (ACL) injuries. In our previous study, stem cells isolated from the human anterior cruciate ligament were shown to possess extensive proliferation and differentiation capabilities when treated with specific growth factors. However, optimal culture conditions and the usefulness of fetal bovine serum (FBS) as a growth factor in in vitro culture systems are yet to be determined. In this study, we compared the effects of different culture media containing combinations of various concentrations of FBS and the growth factors basic fibroblastic growth factor (bFGF) and transforming growth factor-β1 (TGF-β1) on the proliferation and differentiation of ligament-derived stem cells (LSCs) and bone marrow mesenchymal stem cells (BMSCs). We found that α-MEM plus 10% FBS and bFGF was able to maintain both LSCs and BMSCs in a relatively undifferentiated state but with lower major extracellular matrix (ECM) component gene expression and protein production, which is beneficial for stem cell expansion. However, the differentiation and proliferation potentials of LSCs and BMSCs were increased when cultured in MesenPRO, a commercially available stem cell medium containing 2% FBS. MesenPRO in conjunction with TGF-β1 had the greatest ability to induce the differentiation of BMSCs and LSCs to ligament fibroblasts, which was evidenced by the highest ligamentous ECM gene expression and protein production. These results indicate that culture media and growth factors play a very important role in the success of tissue engineering. With α-MEM plus 10% FBS and bFGF, rapid proliferation of stem cells can be achieved. In this study, MesenPRO was able to promote differentiation of both LSCs and BMSCs to ligament fibroblasts. Differentiation was further increased by TGF-β1. With increasing understanding of the effects of different culture media and growth factors, manipulation of stem cells in the desired direction for ligament tissue engineering can be achieved.

Molecular dissection of prethymic progenitor entry into the T lymphocyte developmental pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fung, Elizabeth-sharon

2008-01-01

Notch signaling activates T lineage differentiation from hemopoietic progenitors, but relatively few regulators that initiate this program have been identified, e.g., GATA3 and T cell factor-I (TCF-1) (gene name Tcli). To identify additional regulators of T cell specification, a cDNA libnlrY from mouse Pro-T cells was screened for genes that are specifically up-regulated in intrathymic T cell precursors as compared with myeloid progenitors. Over 90 genes of interest were identified, and 35 of 44 tested were confirmed to be more highly expressed in T lineage precursors relative to precursors of B and/or myeloid lineage. To a remarkable extent, however, expressionmore » of these T lineage-enriched genes, including zinc finger transcription factor, helicase, and signaling adaptor genes, was also shared by stem cells (Lin{sup -}Sca-1{sup +}Kit{sup +}CD27{sup -}) and multipotent progenitors (Lin{sup -}Sca-l{sup +}Kit{sup +}CD27{sup +}), although down-regulated in other lineages. Thus, a major fraction of these early T lineage genes are a regulatory legacy from stem cells. The few genes sharply up-regulated between multipotent progenitors and Pro-T cell stages included those encoding transcription factors Bclllb, TCF-I (Tcli), and HEBalt, Notch target Deltexl, Deltex3L, Fkbp5, Eval, and Tmem13l. Like GATA3 and Deltexl, Bclllb, Fkbp5, and Eval were dependent on Notch/Delta signaling for induction in fetal liver precursors, but only BcIlI band HEBalt were up-regulated between the first two stages of intrathymic T cell development (double negative I and double negative 2) corresponding to T lineage specification. Bclllb was uniquely T lineage restricted and induced by NotchlDelta signaling specifically upon entry into the T lineage differentiation pathway.« less

Gene expression profile of the fibrotic response in the peritoneal cavity.

PubMed

Le, S J; Gongora, M; Zhang, B; Grimmond, S; Campbell, G R; Campbell, J H; Rolfe, B E

2010-01-01

The cellular response to materials implanted in the peritoneal cavity has been utilised to produce tissue for grafting to hollow smooth muscle organs (blood vessels, bladder, uterus and vas deferens). To gain insight into the regulatory mechanisms involved in encapsulation of a foreign object, and subsequent differentiation of encapsulating cells, the present study used microarray technology and real-time RT-PCR to identify the temporal changes in gene expression associated with tissue development. Immunohistochemical analysis showed that 3-7 days post-implantation of foreign objects (cubes of boiled egg white) into rats, they were encapsulated by tissue comprised primarily of haemopoietic (CD45(+)) cells, mainly macrophages (CD68(+), CCR1(+)). By day 14, tissue capsule cells no longer expressed CD68, but were positive for myofibroblast markers alpha-smooth muscle (SM) actin and SM22. In accordance with these results, gene expression data showed that early capsule (days 3-7) development was dominated by the expression of monocyte/macrophage-specific genes (CD14, CSF-1, CSF-1R, MCP-1) and pro-inflammatory mediators such as transforming growth factor (TGF-beta). As tissue capsule development progressed (days 14-21), myofibroblast-associated and pro-fibrotic genes (associated with TGF-beta and Wnt/beta-catenin signalling pathways, including Wnt 4, TGFbetaRII, connective tissue growth factor (CTGF), SMADs-1, -2, -4 and collagen-1 subunits) were significantly up-regulated. The up-regulation of genes associated with Cardiovascular and Skeletal and Muscular System Development at later time-points suggests the capacity of cells within the tissue capsule for further differentiation to smooth muscle, and possibly other cell types. The identification of key regulatory pathways and molecules associated with the fibrotic response to implanted materials has important applications not only for optimising tissue engineering strategies, but also to control deleterious fibrotic responses.

Gene-gene, gene-environment, gene-nutrient interactions and single nucleotide polymorphisms of inflammatory cytokines.

PubMed

Nadeem, Amina; Mumtaz, Sadaf; Naveed, Abdul Khaliq; Aslam, Muhammad; Siddiqui, Arif; Lodhi, Ghulam Mustafa; Ahmad, Tausif

2015-05-15

Inflammation plays a significant role in the etiology of type 2 diabetes mellitus (T2DM). The rise in the pro-inflammatory cytokines is the essential step in glucotoxicity and lipotoxicity induced mitochondrial injury, oxidative stress and beta cell apoptosis in T2DM. Among the recognized markers are interleukin (IL)-6, IL-1, IL-10, IL-18, tissue necrosis factor-alpha (TNF-α), C-reactive protein, resistin, adiponectin, tissue plasminogen activator, fibrinogen and heptoglobins. Diabetes mellitus has firm genetic and very strong environmental influence; exhibiting a polygenic mode of inheritance. Many single nucleotide polymorphisms (SNPs) in various genes including those of pro and anti-inflammatory cytokines have been reported as a risk for T2DM. Not all the SNPs have been confirmed by unifying results in different studies and wide variations have been reported in various ethnic groups. The inter-ethnic variations can be explained by the fact that gene expression may be regulated by gene-gene, gene-environment and gene-nutrient interactions. This review highlights the impact of these interactions on determining the role of single nucleotide polymorphism of IL-6, TNF-α, resistin and adiponectin in pathogenesis of T2DM.

Col1A1 Production and Apoptotic Resistance in TGF-β1-Induced Epithelial-to-Mesenchymal Transition-Like Phenotype of 603B Cells

PubMed Central

Liu, Jun; Eischeid, Alex N.; Chen, Xian-Ming

2012-01-01

Recent studies have suggested that proliferating cholangiocytes have an important role in the induction of fibrosis, either directly via epithelial-to-mesenchymal transition (EMT), or indirectly via activation of other liver cell types. Transforming growth factor beta 1 (TGF-β1), a critical fibrotic cytokine for hepatic fibrosis, is a potent EMT inducer. This study aimed to clarify the potential contributions of TGF-β1-induced EMT-like cholangiocyte phenotype to collagen production and cell survival of cholangiocytes in vitro. Mouse cholangiocytes (603B cells) were treated with TGF-β1 and EMT-like phenotype alterations were monitored by morphological changes and expression of EMT-associated genes. Alterations in Col1A1 gene, Col1A1-associated miR-29s, and pro-apoptotic genes were measured in TGF-β1-treated 603B cells. Snail1 knockdown was achieved using shRNA to evaluate the contribution of EMT-associated changes to Col1A1 production and cell survival. We found TGF-β1 treatment induced partial EMT-like phenotype transition in 603B cells in a Snail1-dependent manner. TGF-β1 also stimulated collagen α1(I) expression in 603B cells. However, this induction was not parallel to the EMT-like alterations and independent of Snail1 or miR-29 expression. Cells undergoing EMT-like changes showed a modest down-regulation of multiple pro-apoptotic genes and displayed resistance to TNF-α-induced apoptosis. TGF-β1-induced apoptosis resistance was attenuated in Snail1 knockdown 603B cells. TGF-β1-induced Col1A1 production seems to be independent of EMT-like transition and miR-29 expression. Nevertheless, TGF-β1-induced EMT may contribute to the increased survival capacity of cholangiocytes via modulating the expression of pro-apoptotic genes. PMID:23236489

Effects of Pro12Ala polymorphism in peroxisome proliferator-activated receptor-γ2 gene on metabolic syndrome risk: a meta-analysis.

PubMed

Zhang, Ruyi; Wang, Jiao; Yang, Rui; Sun, Jia; Chen, Rongping; Luo, Haizhao; Liu, Duan; Cai, Dehong

2014-02-01

Associations between peroxisome proliferator-activated receptor γ2 (PPARγ2) gene polymorphism and metabolic syndrome risk remained controversial and ambiguous. Thus, we performed a meta-analysis to assess the association between Pro12Ala polymorphism in PPARγ2 gene and metabolic syndrome susceptibility. An electronic literature search was conducted on Medline, OVID, Cochrane Library database, and the China National Knowledge Internet up to March 2013. Odds ratios (ORs) with 95% confidence intervals (CIs) were used to calculate the strength of association in the fixed or random effects model. Ten studies involving a total of 4456 cases and 10343 controls were included in this meta-analysis. No statistical evidence of association was found between Pro12Ala polymorphism and metabolic syndrome risk in all genetic models (homozygote model: OR=0.83, 95% CI=0.62-1.12; heterozygote model: OR=1.04, 95% CI=0.94-1.14; dominant model: OR=1.02, 95% CI=0.93-1.12; recessive model: OR=0.83, 95% CI=0.62-1.11). No statistical evidence of significant association was observed when stratified by ethnicity, definition of metabolic syndrome, source of control groups and quality score of the selected articles. All in all, the results did not support a major role of the Pro12Ala variant of the PPARγ2 gene in metabolic syndrome risk. This meta-analysis suggested that the effect of Pro12Ala polymorphism in PPARγ2 gene may not be related to metabolic syndrome as an entity. However, Pro12Ala may affect the single component of metabolic syndrome. A large, well designed study is required to more adequately assess the role for Pro12Ala polymorphism on metabolic syndrome. © 2013 Elsevier B.V. All rights reserved.

Cross-species microarray hybridization to identify developmentally regulated genes in the filamentous fungus Sordaria macrospora.

PubMed

Nowrousian, Minou; Ringelberg, Carol; Dunlap, Jay C; Loros, Jennifer J; Kück, Ulrich

2005-04-01

The filamentous fungus Sordaria macrospora forms complex three-dimensional fruiting bodies that protect the developing ascospores and ensure their proper discharge. Several regulatory genes essential for fruiting body development were previously isolated by complementation of the sterile mutants pro1, pro11 and pro22. To establish the genetic relationships between these genes and to identify downstream targets, we have conducted cross-species microarray hybridizations using cDNA arrays derived from the closely related fungus Neurospora crassa and RNA probes prepared from wild-type S. macrospora and the three developmental mutants. Of the 1,420 genes which gave a signal with the probes from all the strains used, 172 (12%) were regulated differently in at least one of the three mutants compared to the wild type, and 17 (1.2%) were regulated differently in all three mutant strains. Microarray data were verified by Northern analysis or quantitative real time PCR. Among the genes that are up- or down-regulated in the mutant strains are genes encoding the pheromone precursors, enzymes involved in melanin biosynthesis and a lectin-like protein. Analysis of gene expression in double mutants revealed a complex network of interaction between the pro gene products.

In vitro non-viral murine pro-neurotrophin 3 gene transfer into rat bone marrow stromal cells.

PubMed

Darabi, Shahram; Tiraihi, Taki; Delshad, AliReza; Sadeghizadeh, Majid; Khalil, Wisam; Taheri, Taher

2017-04-15

Neurotrophin 3 (NT-3) is an important factor for promoting prenatal neural development, as well as regeneration, axogenesis and plasticity in postnatal life. Therapy with NT-3 was reported to improve the condition of patients suffering from degenerative diseases and traumatic injuries, however, the disadvantage of NT-3 protein delivery is its short half-life, thus our alternative approach is the use of NT-3 gene therapy. In this study, the bone marrow stromal cells (BMSCs) were isolated from adult rats, cultured for 4 passages and transfected with either pEGFP-N1 or a constructed vector containing murine proNT-3 (pSecTag2/HygroB-murine proNT-3) using Lipofectamine 2000 followed by Hygromycin B (200mg/kg). The transfection efficiency of the transiently transfected BMSCs was evaluated using the green fluorescence protein containing vector (pEGFP-N1). A quantitative evaluation of the NT-3 expression of mRNA using real time qRT-PCR shows that there was double fold increase in NT-3 gene expression compared with non-transfected BMSCs, also, the culture supernatant yielded double fold increase in NT-3 using ELISA technique, the data were supported by immunoblotting technique. This suggests that the use of this transfection technique can be useful for gene therapy in different neurological disorders with neurodegenerative or traumatic origins. Copyright © 2017 Elsevier B.V. All rights reserved.

Gene Expression Profiles of Sporadic Canine Hemangiosarcoma Are Uniquely Associated with Breed

PubMed Central

Tamburini, Beth A.; Trapp, Susan; Phang, Tzu Lip; Schappa, Jill T.; Hunter, Lawrence E.; Modiano, Jaime F.

2009-01-01

The role an individual's genetic background plays on phenotype and biological behavior of sporadic tumors remains incompletely understood. We showed previously that lymphomas from Golden Retrievers harbor defined, recurrent chromosomal aberrations that occur less frequently in lymphomas from other dog breeds, suggesting spontaneous canine tumors provide suitable models to define how heritable traits influence cancer genotypes. Here, we report a complementary approach using gene expression profiling in a naturally occurring endothelial sarcoma of dogs (hemangiosarcoma). Naturally occurring hemangiosarcomas of Golden Retrievers clustered separately from those of non-Golden Retrievers, with contributions from transcription factors, survival factors, and from pro-inflammatory and angiogenic genes, and which were exclusively present in hemangiosarcoma and not in other tumors or normal cells (i.e., they were not due simply to variation in these genes among breeds). Vascular Endothelial Growth Factor Receptor 1 (VEGFR1) was among genes preferentially enriched within known pathways derived from gene set enrichment analysis when characterizing tumors from Golden Retrievers versus other breeds. Heightened VEGFR1 expression in these tumors also was apparent at the protein level and targeted inhibition of VEGFR1 increased proliferation of hemangiosarcoma cells derived from tumors of Golden Retrievers, but not from other breeds. Our results suggest heritable factors mold gene expression phenotypes, and consequently biological behavior in sporadic, naturally occurring tumors. PMID:19461996

Genome-wide binding of transcription factor ZEB1 in triple-negative breast cancer cells.

PubMed

Maturi, Varun; Enroth, Stefan; Heldin, Carl-Henrik; Moustakas, Aristidis

2018-05-10

Zinc finger E-box binding homeobox 1 (ZEB1) is a transcriptional regulator involved in embryonic development and cancer progression. ZEB1 induces epithelial-mesenchymal transition (EMT). Triple-negative human breast cancers express high ZEB1 mRNA levels and exhibit features of EMT. In the human triple-negative breast cancer cell model Hs578T, ZEB1 associates with almost 2,000 genes, representing many cellular functions, including cell polarity regulation (DLG2 and FAT3). By introducing a CRISPR-Cas9-mediated 30 bp deletion into the ZEB1 second exon, we observed reduced migratory and anchorage-independent growth capacity of these tumor cells. Transcriptomic analysis of control and ZEB1 knockout cells, revealed 1,372 differentially expressed genes. The TIMP metallopeptidase inhibitor 3 and the teneurin transmembrane protein 2 genes showed increased expression upon loss of ZEB1, possibly mediating pro-tumorigenic actions of ZEB1. This work provides a resource for regulators of cancer progression that function under the transcriptional control of ZEB1. The data confirm that removing a single EMT transcription factor, such as ZEB1, is not sufficient for reverting the triple-negative mesenchymal breast cancer cells into more differentiated, epithelial-like clones, but can reduce tumorigenic potential, suggesting that not all pro-tumorigenic actions of ZEB1 are linked to the EMT. © 2018 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.

Dexamethasone inhibits inflammatory response via down regulation of AP-1 transcription factor in human lung epithelial cells.

PubMed

Patil, Rajeshwari H; Naveen Kumar, M; Kiran Kumar, K M; Nagesh, Rashmi; Kavya, K; Babu, R L; Ramesh, Govindarajan T; Chidananda Sharma, S

2018-03-01

The production of inflammatory mediators by epithelial cells in inflammatory lung diseases may represent an important target for the anti-inflammatory effects of glucocorticoids. Activator protein-1 is a major activator of inflammatory genes and has been proposed as a target for inhibition by glucocorticoids. We have used human pulmonary type-II A549 cells to examine the effect of dexamethasone on the phorbol ester (PMA)/Lipopolysaccharide (LPS) induced pro-inflammatory cytokines and AP-1 factors. A549 cells were treated with and without PMA or LPS or dexamethasone and the cell viability and nitric oxide production was measured by MTT assay and Griess reagent respectively. Expression of pro-inflammatory cytokines and AP-1 factors mRNA were measured using semi quantitative RT-PCR. The PMA/LPS treated cells show significant 2-3 fold increase in the mRNA levels of pro-inflammatory cytokines (IL-1β, IL-2, IL-6, IL-8 and TNF-α), cyclo‑oxygenase-2 (COX-2) and specific AP-1 factors (c-Jun, c-Fos and Jun-D). Whereas, pretreatment of cells with dexamethasone significantly inhibited the LPS induced nitric oxide production and PMA/LPS induced mRNAs expression of above pro-inflammatory cytokines, COX-2 and AP-1 factors. Cells treated with dexamethasone alone at both the concentrations inhibit the mRNAs expression of IL-1β, IL-6 and TNF-α compared to control. Our study reveals that dexamethasone decreased the mRNAs expression of c-Jun and c-Fos available for AP-1 formation suggested that AP-1 is the probable key transcription factor involved in the anti-inflammatory activity of dexamethasone. This may be an important molecular mechanism of steroid action in asthma and other chronic inflammatory lung diseases which may be useful for treatment of lung inflammatory diseases. Copyright © 2017 Elsevier B.V. All rights reserved.

Notch/Delta signaling constrains reengineering of pro-T cells by PU.1

PubMed Central

Franco, Christopher B.; Scripture-Adams, Deirdre D.; Proekt, Irina; Taghon, Tom; Weiss, Angela H.; Yui, Mary A.; Adams, Stephanie L.; Diamond, Rochelle A.; Rothenberg, Ellen V.

2006-01-01

PU.1 is essential for early stages of mouse T cell development but antagonizes it if expressed constitutively. Two separable mechanisms are involved: attenuation and diversion. Dysregulated PU.1 expression inhibits pro-T cell survival, proliferation, and passage through β-selection by blocking essential T cell transcription factors, signaling molecules, and Rag gene expression, which expression of a rearranged T cell antigen receptor transgene cannot rescue. However, Bcl2 transgenic cells are protected from this attenuation and may even undergo β-selection, as shown by PU.1 transduction of defined subsets of Bcl2 transgenic fetal thymocytes with differentiation in OP9-DL1 and OP9 control cultures. The outcome of PU.1 expression in these cells depends on Notch/Delta signaling. PU.1 can efficiently divert thymocytes toward a myeloid-like state with multigene regulatory changes, but Notch/Delta signaling vetoes diversion. Gene expression analysis distinguishes sets of critical T lineage regulatory genes with different combinatorial responses to PU.1 and Notch/Delta signals, suggesting particular importance for inhibition of E proteins, Myb, and/or Gfi1 (growth factor independence 1) in diversion. However, Notch signaling only protects against diversion of cells that have undergone T lineage specification after Thy-1 and CD25 up-regulation. The results imply that in T cell precursors, Notch/Delta signaling normally acts to modulate and channel PU.1 transcriptional activities during the stages from T lineage specification until commitment. PMID:16880393

Erdosteine protects HEI-OC1 auditory cells from cisplatin toxicity through suppression of inflammatory cytokines and induction of Nrf2 target proteins

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Se-Jin; Park, Channy; Lee, Joon No

Cisplatin has many adverse effects, which are a major limitation to its use, including ototoxicity, neurotoxicity, and nephrotoxicity. This study aims to elucidate the protective mechanisms of erdosteine against cisplatin in HEI-OC1 cells. Pretreatment with erdosteine protects HEI-OC1 cells from cisplatin-medicated apoptosis, which is characterized by increase in nuclear fragmentation, DNA laddering, sub-G{sub 0}/G{sub 1} phase, H2AX phosphorylation, PARP cleavage, and caspase-3 activity. Erdosteine significantly suppressed the production of reactive nitrogen/oxygen species and pro-inflammatory cytokines such as tumor necrosis factor-α, interleukin (IL)-1β, and IL-6 in cisplatin-treated cells. Studies using pharmacologic inhibitors demonstrated that phosphatidylinositol-3-kinases (PI3K) and protein kinase B (Akt)more » have protective roles in the action of erdosteine against cisplatin in HEI-OC1 cells. In addition, pretreatment with erdosteine clearly suppressed the phosphorylation of p53 (Ser15) and expression of p53-upregulated modulator of apoptosis. Erdosteine markedly induces expression of NF-E2-related factor 2 (Nrf2), which may contribute to the increase in expression of glutathione redox genes γ-L-glutamate-L-cysteine-ligase catalytic and γ-L-glutamate-L-cysteine-ligase modifier subunits, as well as in the antioxidant genes HO-1 and SOD2 in cisplatin-treated HEI-OC1 cells. Furthermore, the increase in expression of phosphorylated p53 induced by cisplatin is markedly attenuated by pretreatment with erdosteine in the mitochondrial fraction. This increased expression may inhibit the cytosolic expression of the apoptosis-inducing factor, cytochrome c, and Bax/Bcl-xL ratio. Thus, our results suggest that treatment with erdosteine is significantly attenuated cisplatin-induced damage through the activation of Nrf2-dependent antioxidant genes, inhibition of pro-inflammatory cytokines, activation of the PI3K/Akt signaling, and mitochondrial-related inhibition of pro-apoptotic protein expression in HEI-OC1 auditory cells. - Highlights: • Erdosteine significantly attenuated cisplatin-induced cytotoxicity in HEI-OC1 cells. • Erdosteine markedly increased the activation of PI3K/Akt pathway. • Erdosteine induces the expression of Nrf2 and its target genes. • Erdosteine significantly attenuated the accumulation of p53 in mitochondria. • Erdosteine suppressed the release of AIF and cytochrome c from mitochondria.« less

MicroRNA-29 regulates myocardial microvascular endothelial cells proliferation and migration in association with IGF1 in type 2 diabetes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Zhenjie; Jiang, Runxia; Yue, Qingcai

Background: In our study, we investigated the expression and function of microRNA-29 in myocardial microvascular endothelial cells (MMEVC) in type 2 diabetic Goto-Kakizaki (GK) rats. Methods: MiR-29 gene expression was compared, by qRT-PCR between diabetic GK rat MMEVC and non-diabetic Wistar rat MMEVC. MiR-29 was downregulated in GK MMEVC and its effect on angiogenic properties of proliferation and migration was examined. Potential downstream target gene of miR-29, insulin growth factor 1 (IGF1), was assessed by dual-luciferase reporter assay, qRT-PCR and western blot in GK MMEVC. IGF1 was also downregulated by siRNA in miR-29-downregulated GK MMEVC. Its effect on miR-29-associated angiogenicmore » regulation on MMEVC proliferation and migration was further investigated. Results: MiR-29 was substantially upregulated in GK MMEVC than in Wistar MMEVC. Transfection of synthetic miR-29 inhibitor successfully downregulate endogenous miR-29 in GK MMEVC, and subsequently promoted angiogenesis by increasing cell proliferation and migration. IGF1 was confirmed to be downstream target gene of miR-29 in GK MMEVC, with its gene and protein expressions both upregulated in miR-29-downregualted GK MMEVC. Conversely, siRNA-mediated IGF1 downregulation reversed the pro-angiogenic effect of miR-29 downregulation in GK MMEVC, as it decreased cell proliferation and migration. Conclusion: Our study suggests that miR-29 downregulation, through its inverse regulation on downstream target of IGF1 gene, is a pro-angiogenic factor in MMEVC in type 2 diabetic rats.« less

Association of pro-ghrelin and GHS-R1A gene polymorphisms and haplotypes with heavy alcohol use and body mass.

PubMed

Landgren, Sara; Jerlhag, Elisabet; Zetterberg, Henrik; Gonzalez-Quintela, Arturo; Campos, Joaquin; Olofsson, Ulrica; Nilsson, Staffan; Blennow, Kaj; Engel, Jörgen A

2008-12-01

Ghrelin, an orexigenic peptide, acts on growth hormone secretagogue receptors (GHS-R1A), expressed in the hypothalamus as well as in important reward nodes such as the ventral tegmental area. Interestingly, ghrelin has been found to activate an important part of the reward systems, i.e., the cholinergic-dopaminergic reward link. Additionally, the rewarding and neurochemical properties of alcohol are, at least in part, mediated via this reward link. There is comorbidity between alcohol dependence and eating disorders. Thus, plasma levels of ghrelin are altered in patients with addictive behaviors such as alcohol and nicotine dependence and in binge eating disorder. This overlap prompted as to investigate the pro-ghrelin and GHS-R1A genes in a haplotype analysis of heavy alcohol-using individuals. A total of 417 Spanish individuals (abstainers, moderate, and heavy alcohol drinkers) were investigated in a haplotype analysis of the pro-ghrelin and GHS-R1A genes. Tag SNPs were chosen using HapMap data and the Tagger and Haploview softwares. These SNPs were then genotyped using TaqMan Allelic Discrimination. SNP rs2232165 of the GHS-R1A gene was associated with heavy alcohol consumption and SNP rs2948694 of the same gene as well as haplotypes of both the pro-ghrelin and the GHS-R1A genes were associated with body mass in heavy alcohol consuming individuals. The present findings are the first to disclose an association between the pro-ghrelin and GHS-R1A genes and heavy alcohol use, further strengthening the role of the ghrelin system in addictive behaviors and brain reward.

NSAID-activated gene 1 mediates pro-inflammatory signaling activation and paclitaxel chemoresistance in type I human epithelial ovarian cancer stem-like cells.

PubMed

Kim, Ki-Hyung; Park, Seong-Hwan; Do, Kee Hun; Kim, Juil; Choi, Kyung Un; Moon, Yuseok

2016-11-01

Epithelial ovarian cancer (EOC) remains the most lethal gynecologic malignancy in developed countries. Chronic endogenous sterile pro-inflammatory responses are strongly linked to EOC progression and chemoresistance to anti-cancer therapeutics. In the present study, the activity of epithelial NF-κB, a key pro-inflammatory transcription factor, was enhanced with the progress of EOC. This result was mechanistically linked with an increased expression of NSAID-Activated Gene 1 (NAG-1) in MyD88-positive type I EOC stem-like cells, compared with that in MyD88-negative type II EOC cells. Elevated NAG-1 as a potent biomarker of poor prognosis in the ovarian cancer was positively associated with the levels of NF-κB activation, chemokines and stemness markers in type I EOC cells. In terms of signal transduction, NAG-1-activated SMAD-linked and non-canonical TGFβ-activated kinase 1 (TAK-1)-activated pathways contributed to NF-κB activation and the subsequent induction of some chemokines and cancer stemness markers. In addition to effects on NF-κB-dependent gene regulation, NAG-1 was involved in expression of EGF receptor and subsequent activation of EGF receptor-linked signaling. The present study also provided evidences for links between NAG-1-linked signaling and chemoresistance in ovarian cancer cells. NAG-1 and pro-inflammatory NF-κB were positively associated with resistance to paclitaxel in MyD88-positive type I EOC cells. Mechanistically, this chemoresistance occurred due to enhanced activation of the SMAD-4- and non-SMAD-TAK-1-linked pathways. All of the present data suggested NAG-1 protein as a crucial mediator of EOC progression and resistance to the standard first-line chemotherapy against EOC, particularly in MyD88-positive ovarian cancer stem-like cells.

Therapeutic potential of GW501516 and the role of Peroxisome proliferator-activated receptor β/δ and B-cell lymphoma 6 in inflammatory signaling in human pancreatic cancer cells.

PubMed

Smith, Russell W; Coleman, Jeffrey D; Thompson, Jerry T; Vanden Heuvel, John P

2016-12-01

Peroxisome proliferator-activated receptor β/δ (PPARβ/δ) is a member of the nuclear receptor superfamily and a ligand-activated transcription factor that is involved in the regulation of the inflammatory response via activation of anti-inflammatory target genes and ligand-induced disassociation with the transcriptional repressor B-cell lymphoma 6 (BCL6). Chronic pancreatitis is considered to be a significant etiological factor for pancreatic cancer development, and a better understanding of the underlying mechanisms of the transition between inflammation and carcinogenesis would help further elucidate chemopreventative options. The aim of this study was to determine the role of PPARβ/δ and BCL6 in human pancreatic cancer of ductal origin, as well as the therapeutic potential of PPARβ/δ agonist, GW501516. Over-expression of PPARβ/δ inhibited basal and TNFα-induced Nfkb luciferase activity. GW501516-activated PPARβ/δ suppressed TNFα-induced Nfkb reporter activity. RNAi knockdown of Pparb attenuated the GW501516 effect on Nfkb luciferase, while knockdown of Bcl6 enhanced TNFα-induced Nfkb activity. PPARβ/δ activation induced expression of several anti-inflammatory genes in a dose-dependent manner, and GW501516 inhibited Mcp1 promoter-driven luciferase in a BCL6-dependent manner. Several pro-inflammatory genes were suppressed in a BCL6-dependent manner. Conditioned media from GW501516-treated pancreatic cancer cells suppressed pro-inflammatory expression in THP-1 macrophages as well as reduced invasiveness across a basement membrane. These results demonstrate that PPARβ/δ and BCL6 regulate anti-inflammatory signaling in human pancreatic cancer cells by inhibiting NFκB and pro-inflammatory gene expression, and via induction of anti-inflammatory target genes. Activation of PPARβ/δ may be a useful target in pancreatic cancer therapeutics.

Expression of a partially deleted gene of human type II procollagen (COL2A1) in transgenic mice produces a chondrodysplasia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vandenberg, P.; Khillan, J.S.; Prockop, D.J.

A minigene version of the human gene for type II procollagen (COL2AI) was prepared that lacked a large central region containing 12 of the 52 exons and therefore 291 of the 1523 codons of the gene. The construct was modeled after sporadic in-frame deletions of collagen genes that cause synthesis of shortened pro{alpha} chains that associate with normal pro{alpha} chains and thereby cause degradation of the shortened and normal pro{alpha} chains through a process called procollagen suicide. The gene construct was used to prepare five lines of transgenic mice expressing the minigene. A large proportion of the mice expressing themore » minigene developed a phenotype of a chondrodysplasia with dwarfism, short and thick limbs, a short snout, a cranial bulge, a cleft palate, and delayed mineralization of bone. A number of mice died shortly after birth. Microscopic examination of cartilage revealed decreased density and organization of collagen fibrils. In cultured chondrocytes from the transgenic mice, the minigene was expressed as shortened pro{alpha}1(II) chains that were disulfide-linked to normal mouse pro{alpha}1(II) chains. Therefore, the phenotype is probably explained by depletion of the endogenous mouse type II procollagen through the phenomenon of procollagen suicide.« less

Simultaneous Overexpression of Functional Human HO-1, E5NT and ENTPD1 Protects Murine Fibroblasts against TNF-α-Induced Injury In Vitro

PubMed Central

Cinti, Alessandro; De Giorgi, Marco; Chisci, Elisa; Arena, Claudia; Galimberti, Gloria; Farina, Laura; Bugarin, Cristina; Rivolta, Ilaria; Gaipa, Giuseppe; Smolenski, Ryszard Tom; Cerrito, Maria Grazia; Lavitrano, Marialuisa; Giovannoni, Roberto

2015-01-01

Several biomedical applications, such as xenotransplantation, require multiple genes simultaneously expressed in eukaryotic cells. Advances in genetic engineering technologies have led to the development of efficient polycistronic vectors based on the use of the 2A self-processing oligopeptide. The aim of this work was to evaluate the protective effects of the simultaneous expression of a novel combination of anti-inflammatory human genes, ENTPD1, E5NT and HO-1, in eukaryotic cells. We produced an F2A system-based multicistronic construct to express three human proteins in NIH3T3 cells exposed to an inflammatory stimulus represented by tumor necrosis factor alpha (TNF-α), a pro-inflammatory cytokine which plays an important role during inflammation, cell proliferation, differentiation and apoptosis and in the inflammatory response during ischemia/reperfusion injury in several organ transplantation settings. The protective effects against TNF-α-induced cytotoxicity and cell death, mediated by HO-1, ENTPD1 and E5NT genes were better observed in cells expressing the combination of genes as compared to cells expressing each single gene and the effect was further improved by administrating enzymatic substrates of the human genes to the cells. Moreover, a gene expression analyses demonstrated that the expression of the three genes has a role in modulating key regulators of TNF-α signalling pathway, namely Nemo and Tnfaip3, that promoted pro-survival phenotype in TNF-α injured cells. These results could provide new insights in the research of protective mechanisms in transplantation settings. PMID:26513260

Cognitive-behavioral stress management reverses anxiety-related leukocyte transcriptional dynamics

PubMed Central

Antoni, Michael H.; Lutgendorf, Susan K.; Blomberg, Bonnie; Carver, Charles S.; Lechner, Suzanne; Diaz, Alain; Stagl, Jamie; Arevalo, Jesusa M.G.; Cole, Steven W.

2011-01-01

Background Chronic threat and anxiety are associated with pro-inflammatory transcriptional profiles in circulating leukocytes, but the causal direction of that relationship has not been established. This study tested whether a Cognitive-Behavioral Stress Management (CBSM) intervention targeting negative affect and cognition might counteract anxiety-related transcriptional alterations in people confronting a major medical threat. Methods 199 women undergoing primary treatment of Stage 0–III breast cancer were randomized to a 10-week CBSM protocol or an active control condition. 79 provided peripheral blood leukocyte samples for genome-wide transcriptional profiling and bioinformatic analyses at baseline, 6-, and 12-month follow-ups. Results Baseline negative affect was associated with > 50% differential expression of 201 leukocyte transcripts, including up-regulated expression of pro-inflammatory and metastasis-related genes. CBSM altered leukocyte expression of 91 genes by > 50% at follow-up (Group × Time interaction), including down-regulation of pro-inflammatory and metastasis-related genes and up-regulation of Type I interferon response genes. Promoter-based bioinformatic analyses implicated decreased activity of NF-κB/Rel and GATA family transcription factors and increased activity of Interferon Response Factors and the Glucocorticoid Receptor (GR) as potential mediators of CBSM-induced transcriptional alterations. Conclusions In early stage breast cancer patients, a 10-week CBSM intervention can reverse anxiety-related up-regulation of pro-inflammatory gene expression in circulating leukocytes. These findings clarify the molecular signaling pathways by which behavioral interventions can influence physical health and alter peripheral inflammatory processes that may reciprocally affect brain affective and cognitive processes. PMID:22088795

Identification of an epigenetic signature of early mouse liver regeneration that is disrupted by Zn-HDAC inhibition.

PubMed

Huang, Jiansheng; Schriefer, Andrew E; Yang, Wei; Cliften, Paul F; Rudnick, David A

2014-11-01

Liver regeneration has been well studied with hope of discovering strategies to improve liver disease outcomes. Nevertheless, the signals that initiate such regeneration remain incompletely defined, and translation of mechanism-based pro-regenerative interventions into new treatments for hepatic diseases has not yet been achieved. We previously reported the isoform-specific regulation and essential function of zinc-dependent histone deacetylases (Zn-HDACs) during mouse liver regeneration. Those data suggest that epigenetically regulated anti-proliferative genes are deacetylated and transcriptionally suppressed by Zn-HDAC activity or that pro-regenerative factors are acetylated and induced by such activity in response to partial hepatectomy (PH). To investigate these possibilities, we conducted genome-wide interrogation of the liver histone acetylome during early PH-induced liver regeneration in mice using acetyL-histone chromatin immunoprecipitation and next generation DNA sequencing. We also compared the findings of that study to those seen during the impaired regenerative response that occurs with Zn-HDAC inhibition. The results reveal an epigenetic signature of early liver regeneration that includes both hyperacetylation of pro-regenerative factors and deacetylation of anti-proliferative and pro-apoptotic genes. Our data also show that administration of an anti-regenerative regimen of the Zn-HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) not only disrupts gene-specific pro-regenerative changes in liver histone deacetylation but also reverses PH-induced effects on histone hyperacetylation. Taken together, these studies offer new insight into and suggest novel hypotheses about the epigenetic mechanisms that regulate liver regeneration.

Induction of Interferon-Stimulated Genes by IRF3 Promotes Replication of Toxoplasma gondii

PubMed Central

Majumdar, Tanmay; Chattopadhyay, Saurabh; Ozhegov, Evgeny; Dhar, Jayeeta; Goswami, Ramansu; Sen, Ganes C.; Barik, Sailen

2015-01-01

Innate immunity is the first line of defense against microbial insult. The transcription factor, IRF3, is needed by mammalian cells to mount innate immune responses against many microbes, especially viruses. IRF3 remains inactive in the cytoplasm of uninfected cells; upon virus infection, it gets phosphorylated and then translocates to the nucleus, where it binds to the promoters of antiviral genes and induces their expression. Such genes include type I interferons (IFNs) as well as Interferon Stimulated Genes (ISGs). IRF3-/- cells support enhanced replication of many viruses and therefore, the corresponding mice are highly susceptible to viral pathogenesis. Here, we provide evidence for an unexpected pro-microbial role of IRF3: the replication of the protozoan parasite, Toxoplasma gondii, was significantly impaired in IRF3-/- cells. In exploring whether the transcriptional activity of IRF3 was important for its pro-parasitic function, we found that ISGs induced by parasite-activated IRF3 were indeed essential, whereas type I interferons were not important. To delineate the signaling pathway that activates IRF3 in response to parasite infection, we used genetically modified human and mouse cells. The pro-parasitic signaling pathway, which we termed PISA (Parasite-IRF3 Signaling Activation), activated IRF3 without any involvement of the Toll-like receptor or RIG-I-like receptor pathways, thereby ruling out a role of parasite-derived RNA species in activating PISA. Instead, PISA needed the presence of cGAS, STING, TBK1 and IRF3, indicating the necessity of DNA-triggered signaling. To evaluate the physiological significance of our in vitro findings, IRF3-/- mice were challenged with parasite infection and their morbidity and mortality were measured. Unlike WT mice, the IRF3-/- mice did not support replication of the parasite and were resistant to pathogenesis caused by it. Our results revealed a new paradigm in which the antiviral host factor, IRF3, plays a cell-intrinsic pro-parasitic role. PMID:25811886

Induction of interferon-stimulated genes by IRF3 promotes replication of Toxoplasma gondii.

PubMed

Majumdar, Tanmay; Chattopadhyay, Saurabh; Ozhegov, Evgeny; Dhar, Jayeeta; Goswami, Ramansu; Sen, Ganes C; Barik, Sailen

2015-03-01

Innate immunity is the first line of defense against microbial insult. The transcription factor, IRF3, is needed by mammalian cells to mount innate immune responses against many microbes, especially viruses. IRF3 remains inactive in the cytoplasm of uninfected cells; upon virus infection, it gets phosphorylated and then translocates to the nucleus, where it binds to the promoters of antiviral genes and induces their expression. Such genes include type I interferons (IFNs) as well as Interferon Stimulated Genes (ISGs). IRF3-/- cells support enhanced replication of many viruses and therefore, the corresponding mice are highly susceptible to viral pathogenesis. Here, we provide evidence for an unexpected pro-microbial role of IRF3: the replication of the protozoan parasite, Toxoplasma gondii, was significantly impaired in IRF3-/- cells. In exploring whether the transcriptional activity of IRF3 was important for its pro-parasitic function, we found that ISGs induced by parasite-activated IRF3 were indeed essential, whereas type I interferons were not important. To delineate the signaling pathway that activates IRF3 in response to parasite infection, we used genetically modified human and mouse cells. The pro-parasitic signaling pathway, which we termed PISA (Parasite-IRF3 Signaling Activation), activated IRF3 without any involvement of the Toll-like receptor or RIG-I-like receptor pathways, thereby ruling out a role of parasite-derived RNA species in activating PISA. Instead, PISA needed the presence of cGAS, STING, TBK1 and IRF3, indicating the necessity of DNA-triggered signaling. To evaluate the physiological significance of our in vitro findings, IRF3-/- mice were challenged with parasite infection and their morbidity and mortality were measured. Unlike WT mice, the IRF3-/- mice did not support replication of the parasite and were resistant to pathogenesis caused by it. Our results revealed a new paradigm in which the antiviral host factor, IRF3, plays a cell-intrinsic pro-parasitic role.

Mindfulness-Based Stress Reduction training reduces loneliness and pro-inflammatory gene expression in older adults: a small randomized controlled trial.

PubMed

Creswell, J David; Irwin, Michael R; Burklund, Lisa J; Lieberman, Matthew D; Arevalo, Jesusa M G; Ma, Jeffrey; Breen, Elizabeth Crabb; Cole, Steven W

2012-10-01

Lonely older adults have increased expression of pro-inflammatory genes as well as increased risk for morbidity and mortality. Previous behavioral treatments have attempted to reduce loneliness and its concomitant health risks, but have had limited success. The present study tested whether the 8-week Mindfulness-Based Stress Reduction (MBSR) program (compared to a Wait-List control group) reduces loneliness and downregulates loneliness-related pro-inflammatory gene expression in older adults (N = 40). Consistent with study predictions, mixed effect linear models indicated that the MBSR program reduced loneliness, compared to small increases in loneliness in the control group (treatment condition × time interaction: F(1,35) = 7.86, p = .008). Moreover, at baseline, there was an association between reported loneliness and upregulated pro-inflammatory NF-κB-related gene expression in circulating leukocytes, and MBSR downregulated this NF-κB-associated gene expression profile at post-treatment. Finally, there was a trend for MBSR to reduce C Reactive Protein (treatment condition × time interaction: (F(1,33) = 3.39, p = .075). This work provides an initial indication that MBSR may be a novel treatment approach for reducing loneliness and related pro-inflammatory gene expression in older adults. Copyright © 2012 Elsevier Inc. All rights reserved.

Mindfulness-Based Stress Reduction Training Reduces Loneliness and Pro-Inflammatory Gene Expression in Older Adults: A Small Randomized Controlled Trial

PubMed Central

Creswell, J. David; Irwin, Michael R.; Burklund, Lisa J.; Lieberman, Matthew D.; Arevalo, Jesusa M. G.; Ma, Jeffrey; Breen, Elizabeth Crabb; Cole, Steven W.

2013-01-01

Lonely older adults have increased expression of pro-inflammatory genes as well as increased risk for morbidity and mortality. Previous behavioral treatments have attempted to reduce loneliness and its concomitant health risks, but have had limited success. The present study tested whether the 8-week Mindfulness-Based Stress Reduction (MBSR) program (compared to a Wait-List control group) reduces loneliness and downregulates loneliness-related pro-inflammatory gene expression in older adults (N=40). Consistent with study predictions, mixed effect linear models indicated that the MBSR program reduced loneliness, compared to small increases in loneliness in the control group (treatment condition × time interaction: F(1,35)=7.86, p=.008). Moreover, at baseline, there was an association between reported loneliness and upregulated pro-inflammatory NF-κB-related gene expression in circulating leukocytes, and MBSR downregulated this NF-κB-associated gene expression profile at post-treatment. Finally, there was a trend for MBSR to reduce C Reactive Protein (treatment condition × time interaction: (F(1,33)=3.39, p=.075). This work provides an initial indication that MBSR may be a novel treatment approach for reducing loneliness and related pro-inflammatory gene expression in older adults. PMID:22820409

c-Myb Coordinates Survival and the Expression of Genes That Are Critical for the Pre-BCR Checkpoint.

PubMed

Fahl, Shawn P; Daamen, Andrea R; Crittenden, Rowena B; Bender, Timothy P

2018-05-15

The c-Myb transcription factor is required for adult hematopoiesis, yet little is known about c-Myb function during lineage-specific differentiation due to the embryonic lethality of Myb -null mutations. We previously used tissue-specific inactivation of the murine Myb locus to demonstrate that c-Myb is required for differentiation to the pro-B cell stage, survival during the pro-B cell stage, and the pro-B to pre-B cell transition during B lymphopoiesis. However, few downstream mediators of c-Myb-regulated function have been identified. We demonstrate that c-Myb regulates the intrinsic survival of CD19 + pro-B cells in the absence of IL-7 by repressing expression of the proapoptotic proteins Bmf and Bim and that levels of Bmf and Bim mRNA are further repressed by IL-7 signaling in pro-B cells. c-Myb regulates two crucial components of the IL-7 signaling pathway: the IL-7Rα-chain and the negative regulator SOCS3 in CD19 + pro-B cells. Bypassing IL-7R signaling through constitutive activation of Stat5b largely rescues survival of c-Myb-deficient pro-B cells, whereas constitutively active Akt is much less effective. However, rescue of pro-B cell survival is not sufficient to rescue proliferation of pro-B cells or the pro-B to small pre-B cell transition, and we further demonstrate that c-Myb-deficient large pre-B cells are hypoproliferative. Analysis of genes crucial for the pre-BCR checkpoint demonstrates that, in addition to IL-7Rα, the genes encoding λ5, cyclin D3, and CXCR4 are downregulated in the absence of c-Myb, and λ5 is a direct c-Myb target. Thus, c-Myb coordinates survival with the expression of genes that are required during the pre-BCR checkpoint. Copyright © 2018 by The American Association of Immunologists, Inc.

Brain-derived neurotrophic factor (BDNF) and its precursor (proBDNF) in genetically defined fear-induced aggression.

PubMed

Ilchibaeva, Tatiana V; Kondaurova, Elena M; Tsybko, Anton S; Kozhemyakina, Rimma V; Popova, Nina K; Naumenko, Vladimir S

2015-09-01

The brain-derived neurotrophic factor (BDNF), its precursor (proBDNF) and BDNF mRNA levels were studied in the brain of wild rats selectively bred for more than 70 generations for either high level or for the lack of affective aggressiveness towards man. Significant increase of BDNF mRNA level in the frontal cortex and increase of BDNF level in the hippocampus of aggressive rats was revealed. In the midbrain and hippocampus of aggressive rats proBDNF level was increased, whereas BDNF/proBDNF ratio was reduced suggesting the prevalence and increased influence of proBDNF in highly aggressive rats. In the frontal cortex, proBDNF level in aggressive rats was decreased. Thus, considerable structure-specific differences in BDNF and proBDNF levels as well as in BDNF gene expression between highly aggressive and nonaggressive rats were shown. The data suggested the implication of BDNF and its precursor proBDNF in the mechanism of aggressiveness and in the creation of either aggressive or nonaggressive phenotype. Copyright © 2015 Elsevier B.V. All rights reserved.

Transforming growth factor beta-1 (TGF-β1) gene single nucleotide polymorphisms (SNPs) and susceptibility to pre-eclampsia in Iranian women: A case-control study.

PubMed

Khani, Masood; Amani, Davar; Taheripanah, Robabeh; Sanadgol, Nima; Feizollahzadeh, Sadegh; Rahmani, Zahra

2015-10-01

Pre-eclampsia (PE) is a disorder of pregnancy characterized by high blood pressure and proteinuria. Transforming growth factor beta-1 (TGF-β1) is an important replicated PE candidate gene, and few studies have evaluated the direct association of TGF-β polymorphisms and risk to PE. The aim of this study was to investigate the association between three SNPs of TGF-β1 and serum level of this cytokine in PE patients and controls. In this study the polymorphisms of the TGF-β1 gene at the coding region, and positions 29T→C (Leu 10 Pro), 74G→C (Arg 25 Pro) and 788C→T (Thr 263 Ile) were studied in 123 PE and 120 normal subjects using PCR-restriction fragment length polymorphism PCR-(RFLP) and amplification refractory mutation system (ARMS)-PCR methods. Moreover, serum TGF-β1 was determined by enzyme-linked immunosorbent assay (ELISA) technique. At positions 74G→C and 29T→C the genotypes and allele frequencies showed no significant differences between PE patients and normal controls (P=0.3 and P=0.5 respectively). While in the case of position 788C→T both genotypes and allele frequencies were significantly different between PE patients and controls (P=0.02). Haplotype analysis on three polymorphic sites showed no significant differences between PE and control individuals (P=0.8). TGC and CGC haplotypes were the most frequent in both studied groups. The mean serum TGF-β1 level was significantly higher (62.73ng/ml) in PE patients compared with pregnant (47.01ng/ml) and non-pregnant (40.68ng/ml) control groups (P=0.0001). The results of this study suggest that TGF-β1 gene 788C→T polymorphism is an important factor mediating the casual pathway of preeclampsia. Copyright © 2015 International Society for the Study of Hypertension in Pregnancy. Published by Elsevier B.V. All rights reserved.

The opiorphin gene (ProL1) and its homologues function in erectile physiology.

PubMed

Tong, Yuehong; Tar, Moses; Melman, Arnold; Davies, Kelvin

2008-09-01

To determine if ProL1, a member of the opiorphin family of genes, can modulate erectile physiology, as it encodes a peptide which acts as a neutral endopeptidase inhibitor, other examples of which (Vcsa1, hSMR3A) modulate erectile physiology. We cloned members of the opiorphin family of genes into the same mammalian expression backbone (pVAX); 100 microg of these plasmids (pVAX-Vcsa1, -hSMR3A, -hSMR3B and -ProL1) were injected intracorporally into retired breeder rats and the affect on erectile physiology assessed visually, by histology and by measuring the intracavernous pressure (ICP) and blood pressure (BP). As a positive control, rats were treated with pVAX-hSlo (expressing the MaxiK potassium channel) and as a negative control the empty backbone plasmid was injected (pVAX). We also compared the level of expression of ProL1 in corporal tissue of patients not reporting erectile dysfunction (ED), ED associated with diabetes and ED not caused by diabetes. Gene transfer of plasmids expressing all members of the opiorphin family had a similar and significant effect on erectile physiology. At the concentration used in these experiments (100 microg) they resulted in higher resting ICP, and histological and visual analysis showed evidence of a priapic-like condition. After electrostimulation of the cavernous nerve, rats had significantly better ICP/BP than the negative control (pVAX). Gene transfer of pVAX-hSlo increased the ICP/BP ratio to a similar extent to the opiorphin homologues, but with no evidence for a priapic-like condition. Corpora cavernosa tissue samples obtained from men with ED, regardless of underlying causes, had significant down-regulation of both hSMR3A and ProL1. All members of the human opiorphin family of genes can potentially modulate erectile physiology. Both hSMR3 and ProL1 are down-regulated in the corpora of men with ED, and therefore both genes can potentially act as markers of ED.

The opiorphin gene (ProL1) and its homologues function in erectile physiology

PubMed Central

Tong, Yuehong; Tar, Moses; Melman, Arnold; Davies, Kelvin

2010-01-01

OBJECTIVE To determine if ProL1, a member of the opiorphin family of genes, can modulate erectile physiology, as it encodes a peptide which acts as a neutral endopeptidase inhibitor, other examples of which (Vcsa1, hSMR3A) modulate erectile physiology. MATERIALS AND METHODS We cloned members of the opiorphin family of genes into the same mammalian expression backbone (pVAX); 100 μg of these plasmids (pVAX-Vcsa1, -hSMR3A, -hSMR3B and -ProL1) were injected intracorporally into retired breeder rats and the affect on erectile physiology assessed visually, by histology and by measuring the intracavernous pressure (ICP) and blood pressure (BP). As a positive control, rats were treated with pVAX-hSlo (expressing the MaxiK potassium channel) and as a negative control the empty backbone plasmid was injected (pVAX). We also compared the level of expression of ProL1 in corporal tissue of patients not reporting erectile dysfunction (ED), ED associated with diabetes and ED not caused by diabetes. RESULTS Gene transfer of plasmids expressing all members of the opiorphin family had a similar and significant effect on erectile physiology. At the concentration used in these experiments (100 μg) they resulted in higher resting ICP, and histological and visual analysis showed evidence of a priapiclike condition. After electrostimulation of the cavernous nerve, rats had significantly better ICP/BP than the negative control (pVAX). Gene transfer of pVAX-hSlo increased the ICP/BP ratio to a similar extent to the opiorphin homologues, but with no evidence for a priapic-like condition. Corpora cavernosa tissue samples obtained from men with ED, regardless of underlying causes, had significant down-regulation of both hSMR3A and ProL1. CONCLUSION All members of the human opiorphin family of genes can potentially modulate erectile physiology. Both hSMR3 and ProL1 are down-regulated in the corpora of men with ED, and therefore both genes can potentially act as markers of ED. PMID:18410445

AKT activation drives the nuclear localization of CSE1L and a pro-oncogenic transcriptional activation in ovarian cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lorenzato, Annalisa; Biolatti, Marta; Institute for Cancer Research at Candiolo, Candiolo, Torino

The human homolog of the yeast cse1 gene (CSE1L) is over-expressed in ovarian cancer. CSE1L forms complex with Ran and importin-α and has roles in nucleocytoplasmic traffic and gene expression. CSE1L accumulated in the nucleus of ovarian cancer cell lines, while it was localized also in the cytoplasm of other cancer cell lines. Nuclear localization depended on AKT, which was constitutively active in ovarian cancer cells, as the CSE1L protein translocated to the cytoplasm when AKT was inactivated. Moreover, the expression of a constitutively active AKT forced the translocation of CSE1L from the cytoplasm to the nucleus in other cancermore » cells. Nuclear accrual of CSE1L was associated to the nuclear accumulation of the phosphorylated Ran Binding protein 3 (RanBP3), which depended on AKT as well. Also in samples of human ovarian cancer, AKT activation was associated to nuclear accumulation of CSE1L and phosphorylation of RanBP3. Expression profiling of ovarian cancer cells after CSE1L silencing showed that CSE1L was required for the expression of genes promoting invasion and metastasis. In agreement, CSE1L silencing impaired motility and invasiveness of ovarian cancer cells. Altogether these data show that in ovarian cancer cells activated AKT by affecting RanBP3 phosphorylation determines the nuclear accumulation of CSE1L and likely the nuclear concentration of transcription factors conveying pro-oncogenic signals. - highlights: • CSE1L is a key player in nucleocytoplasmic traffic by forming complex with Ran. • AKT phosphorylates RanBP3 that regulates the nucleocytoplasmic gradient of Ran. • The activated oncogenic AKT drives the nuclear accumulation of CSE1L. • CSE1L in the nucleus up-regulates genes conveying pro-oncogenic signals. • CSE1L might contribute to tumor progression driven by the activated oncogenic AKT.« less

Differential regulation of gene expression of neurotensin and prohormone convertases PC1 and PC2 in the bovine ocular ciliary epithelium: possible implications on neurotensin processing.

PubMed

Ortego, Javier; Wollmann, Guido; Coca-Prados, Miguel

2002-11-15

Prohormone convertases PC1 and PC2 are enzymes involved in the intracellular processing of pro-neurotensin/neuromedin N (pro-NT/NN) through the regulated secretory pathway. In this study, we present evidence of the differential gene expression of pro-NT/NN, pro-PC1 and pro-PC2 in two cell lines established from the neuroendocrine ocular ciliary epithelium. Dexamethasone and forskolin were found to synergistically up-regulate NT/NN mRNA expression in both cell types. The pigmented cells released NT, and this release was enhanced by agents that induced its biosynthesis. In contrast, nonpigmented cells exhibited a significantly reduced neurotensin secretion in response to inducers, leading to an accumulation of the peptide. PC1 and PC2 mRNA expression was induced in a cell-specific manner by the same agents that enhanced pro-NT/NN biosynthesis. These results demonstrate cell-specific processing of pro-NT/NN by the ciliary epithelium. Copyright 2002 Elsevier Science Ireland Ltd.

Insufficiency of pro-heparin-binding epidermal growth factor-like growth factor shedding enhances hypoxic cell death in H9c2 cardiomyoblasts via the activation of caspase-3 and c-Jun N-terminal kinase.

PubMed

Uetani, Teruyoshi; Nakayama, Hironao; Okayama, Hideki; Okura, Takafumi; Higaki, Jitsuo; Inoue, Hirofumi; Higashiyama, Shigeki

2009-05-01

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a cardiogenic and cardiohypertrophic growth factor. ProHB-EGF, a product of the Hb-egf gene and the precursor of HB-EGF, is anchored to the plasma membrane. Its ectodomain region is shed by a disintegrin and metalloproteases (ADAMs) when activated by various stimulations. It has been reported that an uncleavable mutant of Hb-egf, uc-Hb-egf, produces uc-proHB-EGF, which is not cleaved by ADAMs and causes dilation of the heart in knock-in mice. This suggests that the shedding of proHB-EGF is essential for the development and survival of cardiomyocytes: however, the molecular mechanism involved has remained unclear. In this study, we investigated the relationship between uc-proHB-EGF expression and cardiomyocyte survival. Human uc-proHB-EGF was adenovirally introduced into the rat cardiomyoblast cell line H9c2, and the cells were cultured under normoxic and hypoxic conditions. Uc-proHB-EGF-expressing H9c2 cells underwent apoptosis under normoxic conditions, which distinctly increased under hypoxic conditions. Furthermore, we observed an increased Caspase-3 activity, reactive oxygen species accumulation, and an increased c-Jun N-terminal kinase (JNK) activity in the uc-proHB-EGF-expressing H9c2 cells. Treatment of the uc-proHB-EGF transfectants with inhibitors of Caspase-3, reactive oxygen species, and JNK, namely, Z-VAD-fmk, N-acetylcysteine, and SP600125, respectively, significantly reduced hypoxic cell death. These data indicate that insufficiency of proHB-EGF shedding under hypoxic stress leads to cardiomyocyte apoptosis via Caspase-3- and JNK-dependent pathways.

Signatures of positive selection in the cis-regulatory sequences of the human oxytocin receptor (OXTR) and arginine vasopressin receptor 1a (AVPR1A) genes.

PubMed

Schaschl, Helmut; Huber, Susanne; Schaefer, Katrin; Windhager, Sonja; Wallner, Bernard; Fieder, Martin

2015-05-13

The evolutionary highly conserved neurohypophyseal hormones oxytocin and arginine vasopressin play key roles in regulating social cognition and behaviours. The effects of these two peptides are meditated by their specific receptors, which are encoded by the oxytocin receptor (OXTR) and arginine vasopressin receptor 1a genes (AVPR1A), respectively. In several species, polymorphisms in these genes have been linked to various behavioural traits. Little, however, is known about whether positive selection acts on sequence variants in genes influencing variation in human behaviours. We identified, in both neuroreceptor genes, signatures of balancing selection in the cis-regulative acting sequences such as transcription factor binding and enhancer sequences, as well as in a transcriptional repressor sequence motif. Additionally, in the intron 3 of the OXTR gene, the SNP rs59190448 appears to be under positive directional selection. For rs59190448, only one phenotypical association is known so far, but it is in high LD' (>0.8) with loci of known association; i.e., variants associated with key pro-social behaviours and mental disorders in humans. Only for one SNP on the OXTR gene (rs59190448) was a sign of positive directional selection detected with all three methods of selection detection. For rs59190448, however, only one phenotypical association is known, but rs59190448 is in high LD' (>0.8), with variants associated with important pro-social behaviours and mental disorders in humans. We also detected various signatures of balancing selection on both neuroreceptor genes.

A novel proteolytic processing of prolysyl oxidase

PubMed Central

Atsawasuwan, Phimon; Mochida, Yoshiyuki; Katafuchi, Michitsuna; Tokutomi, Kentaro; Mocanu, Viorel; Parker, Carol E.; Yamauchi, Mitsuo

2012-01-01

Lysyl oxidase (LOX) is an amine oxidase that is critical for the stability of connective tissues. The secreted proLOX is enzymatically quiescent and is activated through proteolytic cleavage between residue Gly162 and Asp163 (residue numbers according to the mouse LOX) by bone morphogenetic protein (BMP)-1 gene products. Here we report a novel processing of proLOX identified in vitro and in vivo. Two forms of mature LOX were identified and characterized by their immunoreactivity to specific antibodies, amine oxidase activity and mass spectrometry. One form was identified as a well characterized BMP-1 processed LOX protein. Another was found to be a truncated form of LOX (tLOX) resulting from the cleavage at the carboxy terminus of Arg192. The tLOX still appeared to retain amine oxidase activity. The results from the proLOX gene deletion and mutation experiments indicated that the processing occurs independent of the cleavage of proLOX by BMP-1 gene products and likely requires the presence of LOX propeptide. These results indicate that proLOX could be processed by two different mechanisms producing two forms of active LOX. PMID:21591931

A novel proteolytic processing of prolysyl oxidase.

PubMed

Atsawasuwan, Phimon; Mochida, Yoshiyuki; Katafuchi, Michitsuna; Tokutomi, Kentaro; Mocanu, Viorel; Parker, Carol E; Yamauchi, Mitsuo

2011-01-01

Lysyl oxidase (LOX) is an amine oxidase that is critical for the stability of connective tissues. The secreted proLOX is enzymatically quiescent and is activated through proteolytic cleavage between residues Gly(162) and Asp(163) (residue numbers according to the mouse LOX) by bone morphogenetic protein (BMP)-1 gene products. Here we report a novel processing of proLOX identified in vitro and in vivo. Two forms of mature LOX were identified and characterized by their immunoreactivity to specific antibodies, amine oxidase activity, and mass spectrometry. One form was identified as a well-characterized BMP-1 processed LOX protein. Another was found to be a truncated form of LOX resulting from the cleavage at the carboxy terminus of Arg(192). The truncated form of LOX still appeared to retain amine oxidase activity. The results from the proLOX gene deletion and mutation experiments indicated that the processing occurs independent of the cleavage of proLOX by BMP-1 gene products and likely requires the presence of LOX propeptide. These results indicate that proLOX could be processed by two different mechanisms producing two forms of active LOX.

Hormetic modulation of hepatic insulin sensitivity by advanced glycation end products.

PubMed

Fabre, Nelly T; Thieme, Karina; Silva, Karolline S; Catanozi, Sérgio; Cavaleiro, Ana Mercedes; Pinto, Danilo A C; Okamoto, Maristela M; Morais, Mychel Raony P T; Falquetto, Bárbara; Zorn, Telma M; Machado, Ubiratan F; Passarelli, Marisa; Correa-Giannella, Maria Lúcia

2017-05-15

Because of the paucity of information regarding metabolic effects of advanced glycation end products (AGEs) on liver, we evaluated effects of AGEs chronic administration in (1) insulin sensitivity; (2) hepatic expression of genes involved in AGEs, glucose and fat metabolism, oxidative stress and inflammation and; (3) hepatic morphology and glycogen content. Rats received intraperitoneally albumin modified (AlbAGE) or not by advanced glycation for 12 weeks. AlbAGE induced whole-body insulin resistance concomitantly with increased hepatic insulin sensitivity, evidenced by activation of AKT, inactivation of GSK3, increased hepatic glycogen content, and decreased expression of gluconeogenesis genes. Additionally there was reduction in hepatic fat content, in expression of lipogenic, pro-inflamatory and pro-oxidative genes and increase in reactive oxygen species and in nuclear expression of NRF2, a transcription factor essential to cytoprotective response. Although considered toxic, AGEs become protective when administered chronically, stimulating AKT signaling, which is involved in cellular defense and insulin sensitivity. Copyright © 2017 Elsevier B.V. All rights reserved.

Cardiac and Carotid Markers Link With Accelerated Brain Atrophy: The AGES-Reykjavik Study (Age, Gene/Environment Susceptibility-Reykjavik).

PubMed

Sabayan, Behnam; van Buchem, Mark A; Sigurdsson, Sigurdur; Zhang, Qian; Meirelles, Osorio; Harris, Tamara B; Gudnason, Vilmundur; Arai, Andrew E; Launer, Lenore J

2016-11-01

Pathologies in the heart-brain axis might, independently or in combination, accelerate the process of brain parenchymal loss. We aimed to investigate the association of serum N-terminal brain natriuretic peptide (NT-proBNP), as a marker of cardiac dysfunction, and carotid intima media thickness (CIMT), as a marker of carotid atherosclerosis burden, with structural brain changes. In the longitudinal population-based AGES-Reykjavik study (Age, Gene/Environment Susceptibility-Reykjavik), we included 2430 subjects (mean age, 74.6 years; 41.4% men) with baseline data on NT-proBNP and CITM (assessed by ultrasound imaging). Participants underwent a high-resolution brain magnetic resonance imaging at baseline and 5 years later to assess total brain (TBV), gray matter, and white matter volumes. Each unit higher log-transformed NT-proBNP was associated with 3.6 mL (95% confidence interval [CI], -6.0 to -1.1) decline in TBV and 3.5 mL (95% CI, -5.7 to -1.3) decline in gray matter volume. Likewise, each millimeter higher CIMT was associated with 10.8 mL (95% CI, -17.3 to -4.2) decline in TBV and 8.6 mL (95% CI, -14.4 to -2.8) decline in gray matter volume. There was no association between NT-proBNP and CIMT and changes in white matter volume. Compared with participants with low NT-proBNP and CIMT, participants with both high NT-proBNP and CIMT had 3.8 mL (95% CI, -6.0 to -1.6) greater decline in their TBV and 4 mL (95% CI, -6.0 to -2.0) greater decline in GMW. These associations were independent of sociodemographic and cardiovascular factors. Older subjects with both cardiac dysfunction and carotid atherosclerosis are at an increased risk for brain parenchymal loss. Accumulated pathologies in the heart-brain axis might accelerate brain atrophy. © 2016 American Heart Association, Inc.

NFAT5-Regulated Macrophage Polarization Supports the Proinflammatory Function of Macrophages and T Lymphocytes.

PubMed

Tellechea, Mónica; Buxadé, Maria; Tejedor, Sonia; Aramburu, Jose; López-Rodríguez, Cristina

2018-01-01

Macrophages are exquisite sensors of tissue homeostasis that can rapidly switch between pro- and anti-inflammatory or regulatory modes to respond to perturbations in their microenvironment. This functional plasticity involves a precise orchestration of gene expression patterns whose transcriptional regulators have not been fully characterized. We had previously identified the transcription factor NFAT5 as an activator of TLR-induced responses, and in this study we explore its contribution to macrophage functions in different polarization settings. We found that both in classically and alternatively polarized macrophages, NFAT5 enhanced functions associated with a proinflammatory profile such as bactericidal capacity and the ability to promote Th1 polarization over Th2 responses. In this regard, NFAT5 upregulated the Th1-stimulatory cytokine IL-12 in classically activated macrophages, whereas in alternatively polarized ones it enhanced the expression of the pro-Th1 mediators Fizz-1 and arginase 1, indicating that it could promote proinflammatory readiness by regulating independent genes in differently polarized macrophages. Finally, adoptive transfer assays in vivo revealed a reduced antitumor capacity in NFAT5-deficient macrophages against syngeneic Lewis lung carcinoma and ID8 ovarian carcinoma cells, a defect that in the ID8 model was associated with a reduced accumulation of effector CD8 T cells at the tumor site. Altogether, detailed analysis of the effect of NFAT5 in pro- and anti-inflammatory macrophages uncovered its ability to regulate distinct genes under both polarization modes and revealed its predominant role in promoting proinflammatory macrophage functions. Copyright © 2017 by The American Association of Immunologists, Inc.

TNF-α Gene Knockout in Triple Negative Breast Cancer Cell Line Induces Apoptosis

PubMed Central

Pileczki, Valentina; Braicu, Cornelia; Gherman, Claudia D.; Berindan-Neagoe, Ioana

2013-01-01

Tumor necrosis factor alpha (TNF-α) is a pro-inflammatory cytokine involved in the promotion and progression of cancer, including triple negative breast cancer cells. Thus, there is significant interest in understanding the molecular signaling pathways that connect TNF-α with the survival of tumor cells. In our experiments, we used as an in vitro model for triple negative breast cancer the cell line Hs578T. The purpose of this study is to determine the gene expression profiling of apoptotic signaling networks after blocking TNF-α formation by using specially designed siRNA molecules to target TNF-α messenger RNA. Knockdown of TNF-α gene was associated with cell proliferation inhibition and apoptosis, as observed by monitoring the cell index using the xCELLigence RTCA System and flow cytometry. PCR array technology was used to examine the transcript levels of 84 genes involved in apoptosis. 15 genes were found to be relevant after comparing the treated group with the untreated one of which 3 were down-regulated and 12 up-regulated. The down-regulated genes are all involved in cell survival, whereas the up-regulated ones are involved in and interact with pro-apoptotic pathways. The results described here indicate that the direct target of TNF-α in the Hs578T breast cancer cell line increases the level of certain pro-apoptotic factors that modulate different cellular networks that direct the cells towards death. PMID:23263670

Effects of the nuclear localization of the N(pro) protein of classical swine fever virus on its virulence in pigs.

PubMed

Li, Yongfeng; Shen, Liang; Sun, Yuan; Wang, Xiao; Li, Chao; Huang, Junhua; Chen, Jianing; Li, Lianfeng; Zhao, Bibo; Luo, Yuzi; Li, Su; Qiu, Hua-Ji

2014-12-05

The N(pro) protein of classical swine fever virus (CSFV) is localized in the cytoplasm and nucleus. However, it is unknown whether the nuclear localization of N(pro) correlates with the virulence of CSFV in the host. Previously, we showed that the N(pro) protein fused with interferon regulatory factor 3 (IRF3) was present only in the cytoplasm. Here, we generated and evaluated a recombinant CSFV vSM-IRF3 harboring the IRF3 gene inserted into the N(pro) gene of the highly virulent CSFV Shimen strain. Compared to the even nuclear and cytoplasmic distribution of the enhanced green fluorescent protein (EGFP)-N(pro) fusion expressed by the recombinant CSFV EGFP-CSFV, vSM-IRF3 expressed an IRF3-N(pro) fusion protein that only was localized in the cytoplasm. vSM-IRF3 was markedly attenuated in vitro and in vivo, and the inoculated pigs were completely protected from lethal CSFV challenge, whereas the parental virus as well as EGFP-CSFV exhibited a typical virulent phenotype. Taken together, the nuclear localization of N(pro) plays a significant role in the CSFV replication and virulence. Copyright © 2014 Elsevier B.V. All rights reserved.

Identification of an epigenetic signature of early mouse liver regeneration that is disrupted by Zn-HDAC inhibition

PubMed Central

Huang, Jiansheng; Schriefer, Andrew E; Yang, Wei; Cliften, Paul F; Rudnick, David A

2014-01-01

Liver regeneration has been well studied with hope of discovering strategies to improve liver disease outcomes. Nevertheless, the signals that initiate such regeneration remain incompletely defined, and translation of mechanism-based pro-regenerative interventions into new treatments for hepatic diseases has not yet been achieved. We previously reported the isoform-specific regulation and essential function of zinc-dependent histone deacetylases (Zn-HDACs) during mouse liver regeneration. Those data suggest that epigenetically regulated anti-proliferative genes are deacetylated and transcriptionally suppressed by Zn-HDAC activity or that pro-regenerative factors are acetylated and induced by such activity in response to partial hepatectomy (PH). To investigate these possibilities, we conducted genome-wide interrogation of the liver histone acetylome during early PH-induced liver regeneration in mice using acetyL-histone chromatin immunoprecipitation and next generation DNA sequencing. We also compared the findings of that study to those seen during the impaired regenerative response that occurs with Zn-HDAC inhibition. The results reveal an epigenetic signature of early liver regeneration that includes both hyperacetylation of pro-regenerative factors and deacetylation of anti-proliferative and pro-apoptotic genes. Our data also show that administration of an anti-regenerative regimen of the Zn-HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) not only disrupts gene-specific pro-regenerative changes in liver histone deacetylation but also reverses PH-induced effects on histone hyperacetylation. Taken together, these studies offer new insight into and suggest novel hypotheses about the epigenetic mechanisms that regulate liver regeneration. PMID:25482284

The Lipopolysaccharide and β-1,3-Glucan Binding Protein Gene Is Upregulated in White Spot Virus-Infected Shrimp (Penaeus stylirostris)

PubMed Central

Roux, Michelle M.; Pain, Arnab; Klimpel, Kurt R.; Dhar, Arun K.

2002-01-01

Pattern recognition proteins such as lipopolysaccharide and β-1,3-glucan binding protein (LGBP) play an important role in the innate immune response of crustaceans and insects. Random sequencing of cDNA clones from a hepatopancreas cDNA library of white spot virus (WSV)-infected shrimp provided a partial cDNA (PsEST-289) that showed similarity to the LGBP gene of crayfish and insects. Subsequently full-length cDNA was cloned by the 5′-RACE (rapid amplification of cDNA ends) technique and sequenced. The shrimp LGBP gene is 1,352 bases in length and is capable of encoding a polypeptide of 376 amino acids that showed significant similarity to homologous genes from crayfish, insects, earthworms, and sea urchins. Analysis of the shrimp LGBP deduced amino acid sequence identified conserved features of this gene family including a potential recognition motif for β-(1→3) linkage of polysaccharides and putative RGD cell adhesion sites. It is known that LGBP gene expression is upregulated in bacterial and fungal infection and that the binding of lipopolysaccharide and β-1,3-glucan to LGBP activates the prophenoloxidase (proPO) cascade. The temporal expression of LGBP and proPO genes in healthy and WSV-challenged Penaeus stylirostris shrimp was measured by real-time quantitative reverse transcription-PCR, and we showed that LGBP gene expression in shrimp was upregulated as the WSV infection progressed. Interestingly, the proPO expression was upregulated initially after infection followed by a downregulation as the viral infection progressed. The downward trend in the expression of proPO coincided with the detection of WSV in the infected shrimp. Our data suggest that shrimp LGBP is an inducible acute-phase protein that may play a critical role in shrimp-WSV interaction and that the WSV infection regulates the activation and/or activity of the proPO cascade in a novel way. PMID:12072514

The AP-1 Transcription Factor c-Jun Promotes Arthritis by Regulating Cyclooxygenase-2 and Arginase-1 Expression in Macrophages.

PubMed

Hannemann, Nicole; Jordan, Jutta; Paul, Sushmita; Reid, Stephen; Baenkler, Hanns-Wolf; Sonnewald, Sophia; Bäuerle, Tobias; Vera, Julio; Schett, Georg; Bozec, Aline

2017-05-01

Activation of proinflammatory macrophages is associated with the inflammatory state of rheumatoid arthritis. Their polarization and activation are controlled by transcription factors such as NF-κB and the AP-1 transcription factor member c-Fos. Surprisingly, little is known about the role of the AP-1 transcription factor c-Jun in macrophage activation. In this study, we show that mRNA and protein levels of c-Jun are increased in macrophages following pro- or anti-inflammatory stimulations. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment cluster analyses of microarray data using wild-type and c-Jun-deleted macrophages highlight the central function of c-Jun in macrophages, in particular for immune responses, IL production, and hypoxia pathways. Mice deficient for c-Jun in macrophages show an amelioration of inflammation and bone destruction in the serum-induced arthritis model. In vivo and in vitro gene profiling, together with chromatin immunoprecipitation analysis of macrophages, revealed direct activation of the proinflammatory factor cyclooxygenase-2 and indirect inhibition of the anti-inflammatory factor arginase-1 by c-Jun. Thus, c-Jun regulates the activation state of macrophages and promotes arthritis via differentially regulating cyclooxygenase-2 and arginase-1 levels. Copyright © 2017 by The American Association of Immunologists, Inc.

TrSDB: a proteome database of transcription factors

PubMed Central

Hermoso, Antoni; Aguilar, Daniel; Aviles, Francesc X.; Querol, Enrique

2004-01-01

TrSDB—TranScout Database—(http://ibb.uab.es/trsdb) is a proteome database of eukaryotic transcription factors based upon predicted motifs by TranScout and data sources such as InterPro and Gene Ontology Annotation. Nine eukaryotic proteomes are included in the current version. Extensive and diverse information for each database entry, different analyses considering TranScout classification and similarity relationships are offered for research on transcription factors or gene expression. PMID:14681387

Tight regulation of a timed nuclear import wave of EKLF by PKCθ and FOE during Pro-E to Baso-E transition.

PubMed

Shyu, Yu-Chiau; Lee, Tung-Liang; Chen, Xin; Hsu, Pang-Hung; Wen, Shau-Ching; Liaw, Yi-Wei; Lu, Chi-Huan; Hsu, Po-Yen; Lu, Mu-Jie; Hwang, JauLang; Tsai, Ming-Daw; Hwang, Ming-Jing; Chen, Jim-Ray; Shen, Che-Kun James

2014-02-24

Erythropoiesis is a highly regulated process during which BFU-E are differentiated into RBCs through CFU-E, Pro-E, PolyCh-E, OrthoCh-E, and reticulocyte stages. Uniquely, most erythroid-specific genes are activated during the Pro-E to Baso-E transition. We show that a wave of nuclear import of the erythroid-specific transcription factor EKLF occurs during the Pro-E to Baso-E transition. We further demonstrate that this wave results from a series of finely tuned events, including timed activation of PKCθ, phosphorylation of EKLF at S68 by P-PKCθ(S676), and sumoylation of EKLF at K74. The latter EKLF modifications modulate its interactions with a cytoplasmic ankyrin-repeat-protein FOE and importinβ1, respectively. The role of FOE in the control of EKLF nuclear import is further supported by analysis of the subcellular distribution patterns of EKLF in FOE-knockout mice. This study reveals the regulatory mechanisms of the nuclear import of EKLF, which may also be utilized in the nuclear import of other factors. Copyright © 2014 Elsevier Inc. All rights reserved.

Molecular inflammation as an underlying mechanism of the aging process and age-related diseases.

PubMed

Chung, H Y; Lee, E K; Choi, Y J; Kim, J M; Kim, D H; Zou, Y; Kim, C H; Lee, J; Kim, H S; Kim, N D; Jung, J H; Yu, B P

2011-07-01

Aging is a biological process characterized by time-dependent functional declines that are influenced by changes in redox status and by oxidative stress-induced inflammatory reactions. An organism's pro-inflammatory status may underlie the aging process and age-related diseases. In this review, we explore the molecular basis of low-grade, unresolved, subclinical inflammation as a major risk factor for exacerbating the aging process and age-related diseases. We focus on the redox-sensitive transcription factors, NF-κB and FOXO, which play essential roles in the expression of pro-inflammatory mediators and anti-oxidant enzymes, respectively. Major players in molecular inflammation are discussed with respect to the age-related up-regulation of pro-inflammatory cytokines and adhesion molecules, cyclo-oxygenase-2, lipoxygenase, and inducible nitric oxide synthase. The molecular inflammation hypothesis proposed by our laboratory is briefly described to give further molecular insights into the intricate interplay among redox balance, pro-inflammatory gene activation, and chronic age-related inflammatory diseases. The final section discusses calorie restriction as an aging-retarding intervention that also exhibits extraordinarily effective anti-inflammatory activity by modulating GSH redox, NF-κB, SIRT1, PPARs, and FOXOs.

Common non-synonymous polymorphisms in the BRCA1 Associated RING Domain (BARD1) gene are associated with breast cancer susceptibility: a case-control analysis.

PubMed

Huo, Xiang; Hu, Zhibin; Zhai, Xiangjun; Wang, Yan; Wang, Shui; Wang, Xuechen; Qin, Jianwei; Chen, Wenseng; Jin, Guangfu; Liu, Jiyong; Gao, Jun; Wei, Qingyi; Wang, Xinru; Shen, Hongbing

2007-05-01

The BRCA1 Associated RING Domain (BARD1) gene has been identified as a high penetrance gene for breast cancer, whose germline and somatic mutations were reported in both non-BRCA1/2 hereditary site-specific and sporadic breast cancer cases. BARD1 plays a crucial role in tumor repression, along with its heterodimeric partner BRCA1. In the current study, we tested the hypothesis that common non-synonymous polymorphisms in BARD1 are associated with breast cancer susceptibility in a case-control study of 507 patients with incident breast cancer and 539 frequency-matched cancer-free controls in Chinese women. We genotyped all three common (minor allele frequency (MAF)>0.10) non-synonymous polymorphisms (Pro24Ser, Arg378Ser, and Val507Met) in BARD1. We found that the BARD1 Pro24Ser variant genotypes (24Pro/Ser and 24Ser/Ser) and Arg378Ser variant homozygote 378Ser/Ser were associated with a significantly decreased breast cancer risk, compared with their wild-type homozygotes, respectively. Furthermore, a significant locus-locus interaction was evident between Pro24Ser and Arg378Ser (P(int )= 0.032). Among the 378Ser variant allele carriers, the 24Pro/Pro wild-type homozygote was associated with a significantly increased breast cancer risk (adjusted OR=1.81, 95% CI=1.11-2.95), but the subjects having 24Pro/Ser or Ser/Ser variant genotypes had a significantly decreased risk (adjusted OR=0.74, 95% CI=0.56-0.99). In stratified analysis, this locus-locus interaction was more evident among subjects without family cancer history, those with positive estrogen receptor (ER) and individuals with negative progesterone receptor (PR). These findings indicate that the potentially functional polymorphisms Pro24Ser and Arg378Ser in BARD1 may jointly contribute to the susceptibility of breast cancer.

Characterization of Arginase Expression in Glioma-Associated Microglia and Macrophages

PubMed Central

Zhang, Leying; Gao, Hang; Song, Yanyan; Ren, Hui; Ouyang, Mao; Wu, Xiwei; D’Apuzzo, Massimo; Badie, Behnam

2016-01-01

Microglia (MG) and macrophages (MPs) represent a significant component of the inflammatory response to gliomas. When activated, MG/MP release a variety of pro-inflammatory cytokines, however, they also secrete anti-inflammatory factors that limit their cytotoxic function. The balance between pro and anti-inflammatory functions dictates their antitumor activity. To evaluate potential variations in MG and MP function in gliomas, we isolated these cells (and other Gr1+ cells) from intracranial GL261 murine gliomas by FACS and evaluated their gene expression profiles by microarray analysis. As expected, arginase 1 (Arg1, M2 marker) was highly expressed by tumor-associated Gr1+, MG and MP. However, in contrast to MP and Gr1+ cells that expressed Arg1 shortly after tumor trafficking, Arg1 expression in MG was delayed and occurred in larger tumors. Interestingly, depletion of MPs in tumors did not prevent MG polarization, suggesting direct influence of tumor-specific factors on MG Arg1 upregulation. Finally, Arg1 expression was confirmed in human GBM samples, but most Arg1+ cells were neutrophils and not MPs. These findings confirm variations in tumor MG and MP polarization states and its dependency on tumor microenvironmental factors. PMID:27936099

Characterization of Arginase Expression in Glioma-Associated Microglia and Macrophages.

PubMed

Zhang, Ian; Alizadeh, Darya; Liang, Junling; Zhang, Leying; Gao, Hang; Song, Yanyan; Ren, Hui; Ouyang, Mao; Wu, Xiwei; D'Apuzzo, Massimo; Badie, Behnam

2016-01-01

Microglia (MG) and macrophages (MPs) represent a significant component of the inflammatory response to gliomas. When activated, MG/MP release a variety of pro-inflammatory cytokines, however, they also secrete anti-inflammatory factors that limit their cytotoxic function. The balance between pro and anti-inflammatory functions dictates their antitumor activity. To evaluate potential variations in MG and MP function in gliomas, we isolated these cells (and other Gr1+ cells) from intracranial GL261 murine gliomas by FACS and evaluated their gene expression profiles by microarray analysis. As expected, arginase 1 (Arg1, M2 marker) was highly expressed by tumor-associated Gr1+, MG and MP. However, in contrast to MP and Gr1+ cells that expressed Arg1 shortly after tumor trafficking, Arg1 expression in MG was delayed and occurred in larger tumors. Interestingly, depletion of MPs in tumors did not prevent MG polarization, suggesting direct influence of tumor-specific factors on MG Arg1 upregulation. Finally, Arg1 expression was confirmed in human GBM samples, but most Arg1+ cells were neutrophils and not MPs. These findings confirm variations in tumor MG and MP polarization states and its dependency on tumor microenvironmental factors.

Aging is associated with altered inflammatory, arachidonic acid cascade and synaptic markers, influenced by epigenetic modifications, in the human frontal cortex

PubMed Central

Keleshian, Vasken L.; Modi, Hiren R.; Rapoport, Stanley I.; Rao, Jagadeesh S.

2013-01-01

Aging is a risk factor for Alzheimer’s disease (AD) and is associated with cognitive decline. However, underlying molecular mechanisms of brain aging are not clear. Recent studies suggest epigenetic influences on gene expression in AD, since DNA methylation levels influence protein and mRNA expression in postmortem AD brain. We hypothesized that some of these changes occur with normal aging. To test this hypothesis, we measured markers of the arachidonic acid (AA) cascade, neuroinflammation, pro- and anti-apoptosis factors, and gene specific epigenetic modifications in postmortem frontal cortex from nine middle-aged (41 ± 1 (SEM) years) and ten aged subjects (70 ± 3 years). The aged compared with middle-aged brain showed elevated levels of neuroinflammatory and AA cascade markers, altered pro and anti-apoptosis factors and loss of synaptophysin. Some of these changes correlated with promoter hypermethylation of BDNF, CREB, and synaptophysin and hypomethylation of BAX. These molecular alterations in aging are different from or more subtle than changes associated with AD pathology. The degree to which they are related to changes in cognition or behavior during normal aging remains to be evaluated. PMID:23336521

Uncovering DELLA-Independent Gibberellin Responses by Characterizing New Tomato procera Mutants

PubMed Central

Livne, Sivan; Lor, Vai S.; Nir, Ido; Eliaz, Natanella; Aharoni, Asaph; Olszewski, Neil E.; Eshed, Yuval; Weiss, David

2015-01-01

Gibberellin (GA) regulates plant development primarily by triggering the degradation/deactivation of the DELLA proteins. However, it remains unclear whether all GA responses are regulated by DELLAs. Tomato (Solanum lycopersicum) has a single DELLA gene named PROCERA (PRO), and its recessive pro allele exhibits constitutive GA activity but retains responsiveness to external GA. In the loss-of-function mutant proΔGRAS, all examined GA developmental responses were considerably enhanced relative to pro and a defect in seed desiccation tolerance was uncovered. As pro, but not proΔGRAS, elongation was promoted by GA treatment, pro may retain residual DELLA activity. In agreement with homeostatic feedback regulation of the GA biosynthetic pathway, we found that GA20oxidase1 expression was suppressed in proΔGRAS and was not affected by exogenous GA3. In contrast, expression of GA2oxidase4 was not affected by the elevated GA signaling in proΔGRAS but was strongly induced by exogenous GA3. Since a similar response was found in Arabidopsis thaliana plants with impaired activity of all five DELLA genes, we suggest that homeostatic GA responses are regulated by both DELLA-dependent and -independent pathways. Transcriptome analysis of GA-treated proΔGRAS leaves suggests that 5% of all GA-regulated genes in tomato are DELLA independent. PMID:26036254

P53 gene codon 72 polymorphism in patients with oral squamous cell carcinoma in the population of northern Iran.

PubMed

Sina, Mahmud; Pedram, Mehrdad; Ghojazadeh, Morteza; Kochaki, Ahmad; Aghbali, Amirala

2014-11-01

Squamous cell carcinoma is the most common cancer of the oral cavity, and several etiologic factors are involved in its developing. Single nucleotide polymorphism (SNP) of the P53 gene codon 72 (P53c72) changes the structure of the protein and affects its activity. The prevalence of P53c72 different genotypes, which seems to vary with race and geographic location, has shown a strong correlation with many types of human cancers. The aim of this study was to investigate the correlation between P53c72polymorphism and risk of oral squamous cell carcinoma (OSCC) in the heavily populated Gilan Province in northern Iran. This case-control study was done on 55 paraffin-embedded samples from OSCC patients and 100 samples of non-dysplastic oral cavity lesions. The P53c72 genotypes were determined using the ARMS-PCR method. SPSS-15 software was used for statistical analysis. There were no significant statistical differences found between the prevalence of different P53c72 genotypes in the OSCC group vs. the control. However, the Pro/Pro genotype in OSCC samples showed a strong correlation with age, as 70% of such patients were below 50 years old. Interestingly, a large portion (40%) of the patients with the Pro/Pro genotype had the tumor in the lip area. Although P53c72 polymorphism does not appear to be a predisposing factor for OSCC in the population of Northern Iran, the Pro/Pro genotype could be considered as a risk factor for OSCC in adults below 50 years old and the anatomical location of the tumor.

The transcription repressor, ZEB1, cooperates with CtBP2 and HDAC1 to suppress IL-2 gene activation in T cells.

PubMed

Wang, Jun; Lee, Seungsoo; Teh, Charis En-Yi; Bunting, Karen; Ma, Lina; Shannon, M Frances

2009-03-01

Activation of T cells leads to the induction of many cytokine genes that are required for appropriate immune responses, including IL-2, a key cytokine for T cell proliferation and homeostasis. The activating transcription factors such as nuclear factor of activated T cells, nuclear factor kappaB/Rel and activated protein-1 family members that regulate inducible IL-2 gene expression have been well documented. However, negative regulation of the IL-2 gene is less studied. Here we examine the role of zinc finger E-box-binding protein (ZEB) 1, a homeodomain/Zn finger transcription factor, as a repressor of IL-2 gene transcription. We show here that ZEB1 is expressed in non-stimulated and stimulated T cells and using chromatin immunoprecipitation assays we show that ZEB1 binds to the IL-2 promoter. Over-expression of ZEB1 can repress IL-2 promoter activity, as well as endogenous IL-2 mRNA production in EL-4 T cells, and this repression is dependent on the ZEB-binding site at -100. ZEB1 cooperates with the co-repressor C-terminal-binding protein (CtBP) 2 and with histone deacetylase 1 to repress the IL-2 promoter and this cooperation depends on the ZEB-binding site in the promoter as well as the Pro-X-Asp-Leu-Ser protein-protein interaction domain in CtBP2. Thus, ZEB1 may function to recruit a repressor complex to the IL-2 promoter.

Ursolic Acid Inhibits Na+/K+-ATPase Activity and Prevents TNF-α-Induced Gene Expression by Blocking Amino Acid Transport and Cellular Protein Synthesis

PubMed Central

Yokomichi, Tomonobu; Morimoto, Kyoko; Oshima, Nana; Yamada, Yuriko; Fu, Liwei; Taketani, Shigeru; Ando, Masayoshi; Kataoka, Takao

2011-01-01

Pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-α, induce the expression of a wide variety of genes, including intercellular adhesion molecule-1 (ICAM-1). Ursolic acid (3β-hydroxy-urs-12-en-28-oic acid) was identified to inhibit the cell-surface ICAM-1 expression induced by pro-inflammatory cytokines in human lung carcinoma A549 cells. Ursolic acid was found to inhibit the TNF-α-induced ICAM-1 protein expression almost completely, whereas the TNF-α-induced ICAM-1 mRNA expression and NF-κB signaling pathway were decreased only partially by ursolic acid. In line with these findings, ursolic acid prevented cellular protein synthesis as well as amino acid uptake, but did not obviously affect nucleoside uptake and the subsequent DNA/RNA syntheses. This inhibitory profile of ursolic acid was similar to that of the Na+/K+-ATPase inhibitor, ouabain, but not the translation inhibitor, cycloheximide. Consistent with this notion, ursolic acid was found to inhibit the catalytic activity of Na+/K+-ATPase. Thus, our present study reveals a novel molecular mechanism in which ursolic acid inhibits Na+/K+-ATPase activity and prevents the TNF-α-induced gene expression by blocking amino acid transport and cellular protein synthesis. PMID:24970122

A Rosa canina WUSCHEL-related homeobox gene, RcWOX1, is involved in auxin-induced rhizoid formation.

PubMed

Gao, Bin; Wen, Chao; Fan, Lusheng; Kou, Yaping; Ma, Nan; Zhao, Liangjun

2014-12-01

Homeobox (HB) proteins are important transcription factors that regulate the developmental decisions of eukaryotes. WUSCHEL-related homeobox (WOX) transcription factors, known as a plant-specific HB family, play a key role in plant developmental processes. Our previous work has indicated that rhizoids are induced by auxin in rose (Rosa spp.), which acts as critical part of an efficient plant regeneration system. However, the function of WOX genes in auxin-induced rhizoid formation remains unclear. Here, we isolated and characterized a WUSCHEL-related homeobox gene from Rosa canina, RcWOX1, containing a typical homeodomain with 65 amino acid residues. Real-time reverse transcription PCR (qRT-PCR) analysis revealed that RcWOX1 was expressed in the whole process of callus formation and in the early stage of rhizoid formation. Moreover, its expression was induced by auxin treatment. In Arabidopsis transgenic lines expressing the RcWOX1pro::GUS and 35S::GFP-RcWOX1, RcWOX1 was specifically expressed in roots and localized to the nucleus. Overexpression of RcWOX1 in Arabidopsis increased lateral root density and induced upregulation of PIN1 and PIN7 genes. Therefore, we postulated that RcWOX1 is a functional transcription factor that plays an essential role in auxin-induced rhizoid formation.

Heterochromatin protein 1 gamma and IκB kinase alpha interdependence during tumour necrosis factor gene transcription elongation in activated macrophages.

PubMed

Thorne, James L; Ouboussad, Lylia; Lefevre, Pascal F

2012-09-01

IκB kinase α (IKKα) is part of the cytoplasmic IKK complex regulating nuclear factor-κB (NF-κB) release and translocation into the nucleus in response to pro-inflammatory signals. IKKα can also be recruited directly to the promoter of NF-κB-dependent genes by NF-κB where it phosphorylates histone H3 at serine 10, triggering recruitment of the bromodomain-containing protein 4 and the positive transcription elongation factor b. Herein, we report that IKKα travels with the elongating form of ribonucleic acid polymerase II together with heterochromatin protein 1 gamma (HP1γ) at NF-κB-dependent genes in activated macrophages. IKKα binds to and phosphorylates HP1γ, which in turn controls IKKα binding to chromatin and phosphorylation of the histone variant H3.3 at serine 31 within transcribing regions. Downstream of transcription end sites, IKKα accumulates with its inhibitor the CUE-domain containing protein 2, suggesting a link between IKKα inactivation and transcription termination.

Novel XLRS1 gene mutations cause X-linked juvenile retinoschisis in Chinese families.

PubMed

Ma, Xiang; Li, Xiaoxin; Wang, Lihua

2008-01-01

To investigate various XLRS1 (RS1) gene mutations in Chinese families with X-linked juvenile retinoschisis (XLRS or RS). Genomic DNA was isolated from leukocytes of 29 male patients with X-linked juvenile retinoschisis, 38 female carriers, and 100 normal controls. All 6 exons of the RS1 gene were amplified by polymerase chain reaction, and the RS1 gene mutations were determined by direct sequencing. Eleven different RS1 mutations in 12 families were identified in the 29 male patients. The mutations comprised eight missense, two frameshift, and one splice donor site mutation. Four of these mutations, one frameshift mutation (26 del T) in exon 1, one frameshift mutation (488 del G) in exon 5, Asp145His and Arg156Gly in exon 5, have not been previously described. One novel non-disease-related polymorphism, 576C to T (Pro192Pro) in exon 6, was also found. Six recurrent mutations, Ser73Pro and Arg102Gln mutations in exon 4 and Arg200Cys, Arg209His, Arg213Gln, and Cys223Arg mutations in exon 6, were also identified in this study. RS1 gene mutations caused X-linked juvenile retinoschisis in these Chinese families.

Polymorphisms of glutathione peroxidase 1 (GPX1 Pro198Leu) and catalase (CAT C-262T) in women with spontaneous abortion.

PubMed

Sabet, Eliza Eskafi; Salehi, Zivar; Khodayari, Siamak; Zarafshan, Samin Sabouhi; Zahiri, Ziba

2014-10-01

About 10%-15% of conceptions are lost spontaneously prior to 20 weeks. Apart from the clinical problems, genetic variations have also been proposed as a susceptibility factor to miscarriage. Glutathione peroxidase 1 (GPX1) and catalase (CAT) encode two antioxidant enzymes that detoxify H2O2 and protect the cells from oxidative damage. A functional polymorphism at codon 198 of the GPX1 gene causes a C/T substitution in exon 2, which encodes for either proline or leucine (Pro198Leu). The CAT gene has a polymorphic site in the promoter region at position -262 (C-262T) which alters the expression and enzyme blood levels, leading to some pathological clinical conditions. In this study, we evaluated the association of these two polymorphisms with the risk of spontaneous abortion. Genomic DNA from 105 cases with spontaneous abortion and 90 healthy women were genotyped using allele-specific PCR (AS-PCR) and polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP). The genetic distributions for GPX1 did not differ significantly between cases and controls (p = 0.680). However, C-262T polymorphism was significantly associated with the risk of the disease (OR, 5.50; 95% CI, 1.43-21.09; p = 0.012). In conclusion, this study indicates that CAT -262T/T genotype confers less susceptibility to spontaneous abortion, while GPX1 Pro198Leu polymorphism may not be correlated with the disease.

Sleep deprivation attenuates endotoxin-induced cytokine gene expression independent of day length and circulating cortisol in male Siberian hamsters (Phodopus sungorus).

PubMed

Ashley, Noah T; Walton, James C; Haim, Achikam; Zhang, Ning; Prince, Laura A; Fruchey, Allison M; Lieberman, Rebecca A; Weil, Zachary M; Magalang, Ulysses J; Nelson, Randy J

2013-07-15

Sleep is restorative, whereas reduced sleep leads to negative health outcomes, such as increased susceptibility to disease. Sleep deprivation tends to attenuate inflammatory responses triggered by infection or exposure to endotoxin, such as bacterial lipopolysaccharide (LPS). Previous studies have demonstrated that Siberian hamsters (Phodopus sungorus), photoperiodic rodents, attenuate LPS-induced fever, sickness behavior and upstream pro-inflammatory gene expression when adapted to short day lengths. Here, we tested whether manipulation of photoperiod alters the suppressive effects of sleep deprivation upon cytokine gene expression after LPS challenge. Male Siberian hamsters were adapted to long (16 h:8 h light:dark) or short (8 h:16 h light:dark) photoperiods for >10 weeks, and were deprived of sleep for 24 h using the multiple platform method or remained in their home cage. Hamsters received an intraperitoneal injection of LPS or saline (control) 18 h after starting the protocol, and were killed 6 h later. LPS increased liver and hypothalamic interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF) gene expression compared with vehicle. Among LPS-challenged hamsters, sleep deprivation reduced IL-1 mRNA levels in liver and hypothalamus, but not TNF. IL-1 attenuation was independent of circulating baseline cortisol, which did not increase after sleep deprivation. Conversely, photoperiod altered baseline cortisol, but not pro-inflammatory gene expression in sleep-deprived hamsters. These results suggest that neither photoperiod nor glucocorticoids influence the suppressive effect of sleep deprivation upon LPS-induced inflammation.

Characteristic Changes in Decidual Gene Expression Signature in Spontaneous Term Parturition

PubMed Central

El-Azzamy, Haidy; Balogh, Andrea; Romero, Roberto; Xu, Yi; LaJeunesse, Christopher; Plazyo, Olesya; Xu, Zhonghui; Price, Theodore G.; Dong, Zhong; Tarca, Adi L.; Papp, Zoltan; Hassan, Sonia S.; Chaiworapongsa, Tinnakorn; Kim, Chong Jai; Gomez-Lopez, Nardhy; Than, Nandor Gabor

2017-01-01

Background The decidua has been implicated in the “terminal pathway” of human term parturition, which is characterized by the activation of pro-inflammatory pathways in gestational tissues. However, the transcriptomic changes in the decidua leading to terminal pathway activation have not been systematically explored. This study aimed to compare the decidual expression of developmental signaling and inflammation-related genes before and after spontaneous term labor in order to reveal their involvement in this process. Methods Chorioamniotic membranes were obtained from normal pregnant women who delivered at term with spontaneous labor (TIL, n = 14) or without labor (TNL, n = 15). Decidual cells were isolated from snap-frozen chorioamniotic membranes with laser microdissection. The expression of 46 genes involved in decidual development, sex steroid and prostaglandin signaling, as well as pro- and anti-inflammatory pathways, was analyzed using high-throughput quantitative real-time polymerase chain reaction (qRT-PCR). Chorioamniotic membrane sections were immunostained and then semi-quantified for five proteins, and immunoassays for three chemokines were performed on maternal plasma samples. Results The genes with the highest expression in the decidua at term gestation included insulin-like growth factor-binding protein 1 (IGFBP1), galectin-1 (LGALS1), and progestogen-associated endometrial protein (PAEP); the expression of estrogen receptor 1 (ESR1), homeobox A11 (HOXA11), interleukin 1β (IL1B), IL8, progesterone receptor membrane component 2 (PGRMC2), and prostaglandin E synthase (PTGES) was higher in TIL than in TNL cases; the expression of chemokine C-C motif ligand 2 (CCL2), CCL5, LGALS1, LGALS3, and PAEP was lower in TIL than in TNL cases; immunostaining confirmed qRT-PCR data for IL-8, CCL2, galectin-1, galectin-3, and PAEP; and no correlations between the decidual gene expression and the maternal plasma protein concentrations of CCL2, CCL5, and IL-8 were found. Conclusions Our data suggests that with the initiation of parturition, the decidual expression of anti-inflammatory mediators decreases, while the expression of pro-inflammatory mediators and steroid receptors increases. This shift may affect downstream signaling pathways that can lead to parturition. PMID:28226203

The Anti-Inflammatory Effect of Algae-Derived Lipid Extracts on Lipopolysaccharide (LPS)-Stimulated Human THP-1 Macrophages

PubMed Central

Robertson, Ruairi C.; Guihéneuf, Freddy; Bahar, Bojlul; Schmid, Matthias; Stengel, Dagmar B.; Fitzgerald, Gerald F.; Ross, R. Paul; Stanton, Catherine

2015-01-01

Algae contain a number of anti-inflammatory bioactive compounds such as omega-3 polyunsaturated fatty acids (n-3 PUFA) and chlorophyll a, hence as dietary ingredients, their extracts may be effective in chronic inflammation-linked metabolic diseases such as cardiovascular disease. In this study, anti-inflammatory potential of lipid extracts from three red seaweeds (Porphyra dioica, Palmaria palmata and Chondrus crispus) and one microalga (Pavlova lutheri) were assessed in lipopolysaccharide (LPS)-stimulated human THP-1 macrophages. Extracts contained 34%–42% total fatty acids as n-3 PUFA and 5%–7% crude extract as pigments, including chlorophyll a, β-carotene and fucoxanthin. Pretreatment of the THP-1 cells with lipid extract from P. palmata inhibited production of the pro-inflammatory cytokines interleukin (IL)-6 (p < 0.05) and IL-8 (p < 0.05) while that of P. lutheri inhibited IL-6 (p < 0.01) production. Quantitative gene expression analysis of a panel of 92 genes linked to inflammatory signaling pathway revealed down-regulation of the expression of 14 pro-inflammatory genes (TLR1, TLR2, TLR4, TLR8, TRAF5, TRAF6, TNFSF18, IL6R, IL23, CCR1, CCR4, CCL17, STAT3, MAP3K1) by the lipid extracts. The lipid extracts effectively inhibited the LPS-induced pro-inflammatory signaling pathways mediated via toll-like receptors, chemokines and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling molecules. These results suggest that lipid extracts from P. lutheri, P. palmata, P. dioica and C. crispus can inhibit LPS-induced inflammatory pathways in human macrophages. Therefore, algal lipid extracts should be further explored as anti-inflammatory ingredients for chronic inflammation-linked metabolic diseases. PMID:26308008

Adenoviral mediated interferon-alpha 2b gene therapy suppresses the pro-angiogenic effect of vascular endothelial growth factor in superficial bladder cancer.

PubMed

Adam, Liana; Black, Peter C; Kassouf, Wassim; Eve, Beryl; McConkey, David; Munsell, Mark F; Benedict, William F; Dinney, Colin P N

2007-05-01

Intravesical adenovirus mediated interferon-alpha gene transfer has a potent therapeutic effect against superficial human bladder carcinoma xenografts growing in the bladder of athymic nude mice. We determined whether the inhibition of angiogenesis might contribute to the antitumor effect. We treated several human urothelial carcinoma cells with adenovirus mediated interferon-alpha 2b and monitored its effects on the production of angiogenic factors using real-time reverse-transcription polymerase chain reaction, Western blotting, and immunohistochemical analysis and a gel shift based transcription factor array. To assess the role of adenovirus mediated interferon 2b in angiogenic activity we used in vitro invasion assays and evaluated the anti-angiogenic effects of adenovirus mediated interferon gene therapy in an orthotopic murine model of human superficial bladder cancer. In adenovirus mediated interferon-alpha infected 253J B-V cells vascular endothelial growth factor was decreased and anti-angiogenic interferon-gamma inducible protein 10 was up-regulated. In contrast, the addition of as much as 100,000 IU recombinant interferon had no apparent effect on vascular endothelial growth factor production. Conditioned medium derived from adenovirus mediated interferon 2b infected 253J B-V cells greatly decreased the invasive potential of human endothelial cells and down-regulated their matrix metalloproteinase 2 expression compared to controls. Furthermore, adenovirus mediated interferon 2b blocked pro-angiogenic nuclear signals, such as the transcription factors activating protein-1 and 2, stimulating protein-1, nuclear factor kappaB and c-myb. In vivo experiments revealed significant vascular endothelial growth factor down-regulation and decreased tumor vessel density in the adenovirus mediated interferon 2b treated group compared to controls. Treatment with adenovirus mediated interferon 2b increases the angiostatic activity of the bladder cancer microenvironment. This inhibition may prove beneficial for treating superficial bladder cancer with adenovirus mediated interferon-alpha and hopefully contribute to a decreased recurrence rate of this neoplasm.

Inflammatory and age-related pathologies in mice with ectopic expression of human PARP-1.

PubMed

Mangerich, Aswin; Herbach, Nadja; Hanf, Benjamin; Fischbach, Arthur; Popp, Oliver; Moreno-Villanueva, María; Bruns, Oliver T; Bürkle, Alexander

2010-06-01

Poly(ADP-ribose) polymerase-1 (PARP-1) is a sensor for DNA strand breaks and some unusual DNA structures and catalyzes poly(ADP-ribosyl)ation of nuclear proteins with NAD(+) serving as substrate. PARP-1 is involved in the regulation of genomic integrity, transcription, inflammation, and cell death. Due to its versatile role, PARP-1 is discussed both as a longevity factor and as an aging-promoting factor. Recently, we generated a mouse model with ectopic integration of full-length hPARP-1 [Mangerich, A., Scherthan, H., Diefenbach, J., Kloz, U., van der Hoeven, F., Beneke, S. and Bürkle, A., 2009. A caveat in mouse genetic engineering: ectopic gene targeting in ES cells by bidirectional extension of the homology arms of a gene replacement vector carrying human PARP-1. Transgenic Res. 18, 261-279]. Here, we show that hPARP-1 mice exhibit impaired survival rates accompanied by reduced hair growth and premature development of several inflammation and age-associated pathologies, such as adiposity, kyphosis, nephropathy, dermatitis, pneumonitis, cardiomyopathy, hepatitis, and anemia. Moreover, mutant male mice showed impaired glucose tolerance, yet without developing manifest diabetes. Overall tumor burden was comparable in wild-type and hPARP-1 mice, but tumor spectrum was shifted in mutant mice, showing lower incidence of sarcomas, but increased incidence of carcinomas. Furthermore, DNA repair was delayed in splenocytes of hPARP-1 mice, and gene expression of pro-inflammatory cytokines was dysregulated. Our results suggest that in hPARP-1 mice impaired DNA repair, accompanied by a continuous low-level increase in pro-inflammatory stimuli, causes development of chronic diseases leading to impaired survival. (c) 2010 Elsevier Ireland Ltd. All rights reserved.

The pro-Forms of Insulin-Like Growth Factor I (IGF-I) Are Predominant in Skeletal Muscle and Alter IGF-I Receptor Activation

PubMed Central

Durzyńska, Julia; Philippou, Anastassios; Brisson, Becky K.; Nguyen-McCarty, Michelle

2013-01-01

IGF-I is a key regulator of muscle development and growth. The pre-pro-peptide produced by the Igf1gene undergoes several posttranslational processing steps to result in a secreted mature protein, which is thought to be the obligate ligand for the IGF-I receptor (IGF-IR). The goals of this study were to determine what forms of IGF-I exist in skeletal muscle, and whether the mature IGF-I protein was the only form able to activate the IGF-IR. We measured the proportion of IGF-I species in murine skeletal muscle and found that the predominant forms were nonglycosylated pro-IGF-I and glycosylated pro-IGF-I, which retained the C-terminal E peptide extension, instead of mature IGF-I. These forms were validated using samples subjected to viral expression of IGF-I combined with furin and glycosidase digestion. To determine whether the larger molecular weight IGF-I forms were also ligands for the IGF-IR, we generated each specific form through transient transfection of 3T3 cells and used the enriched media to perform kinase receptor activation assays. Compared with mature IGF-I, nonglycosylated pro-IGF-I had similar ability to activate the IGF-IR, whereas glycosylation of pro-IGF-I significantly reduced receptor activation. Thus, it is important to understand not only the quantity, but also the proportion of IGF-I forms produced, to evaluate the true biological activity of this growth factor. PMID:23407451

The BCL-2 family protein Bid is critical for pro-inflammatory signaling in astrocytes.

PubMed

König, Hans-Georg; Coughlan, Karen S; Kinsella, Sinéad; Breen, Bridget A; Prehn, Jochen H M

2014-10-01

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of motoneurons in the spinal cord, brainstem and motor cortex. Mutations in the superoxide dismutase 1 (SOD1) gene represent a frequent genetic determinant and recapitulate a disease phenotype similar to ALS when expressed in mice. Previous studies using SOD1(G93A) transgenic mice have suggested a paracrine mechanism of neuronal loss, in which cytokines and other toxic factors released from astroglia or microglia trigger motoneuron degeneration. Several pro-inflammatory cytokines activate death receptors and may downstream from this activate the Bcl-2 family protein, Bid. We here sought to investigate the role of Bid in astrocyte activation and non-cell autonomous motoneuron degeneration. We found that spinal cord Bid protein levels increased significantly during disease progression in SOD1(G93A) mice. Subsequent experiments in vitro indicated that Bid was expressed at relatively low levels in motoneurons, but was enriched in astrocytes and microglia. Bid was strongly induced in astrocytes in response to pro-inflammatory cytokines or exposure to lipopolysaccharide. Experiments in bid-deficient astrocytes or astrocytes treated with a small molecule Bid inhibitor demonstrated that Bid was required for the efficient activation of transcription factor nuclear factor-κB in response to these pro-inflammatory stimuli. Finally, we found that conditioned medium from wild-type astrocytes, but not from bid-deficient astrocytes, was toxic when applied to primary motoneuron cultures. Collectively, our data demonstrate a new role for the Bcl-2 family protein Bid as a mediator of astrocyte activation during neuroinflammation, and suggest that Bid activation may contribute to non-cell autonomous motoneuron degeneration in ALS. Copyright © 2014 Elsevier Inc. All rights reserved.

Differential Effects of HNF-1α Mutations Associated with Familial Young-Onset Diabetes on Target Gene Regulation

PubMed Central

Galán, Maria; García-Herrero, Carmen-Maria; Azriel, Sharona; Gargallo, Manuel; Durán, Maria; Gorgojo, Juan-Jose; Andía, Victor-Manuel; Navas, Maria-Angeles

2011-01-01

Hepatocyte nuclear factor 1-α (HNF-1α) is a homeodomain transcription factor expressed in a variety of tissues (including liver and pancreas) that regulates a wide range of genes. Heterozygous mutations in the gene encoding HNF-1α (HNF1A) cause familial young-onset diabetes, also known as maturity-onset diabetes of the young, type 3 (MODY3). The variability of the MODY3 clinical phenotype can be due to environmental and genetic factors as well as to the type and position of mutations. Thus, functional characterization of HNF1A mutations might provide insight into the molecular defects explaining the variability of the MODY3 phenotype. We have functionally characterized six HNF1A mutations identified in diabetic patients: two novel ones, p.Glu235Gly and c-57-64delCACGCGGT;c-55G>C; and four previously described, p.Val133Met, p.Thr196Ala, p.Arg271Trp and p.Pro379Arg. The effects of mutations on transcriptional activity have been measured by reporter assays on a subset of HNF-1α target promoters in Cos7 and Min6 cells. Target DNA binding affinities have been quantified by electrophoretic mobility shift assay using bacterially expressed glutathione-S-transferase (GST)-HNF-1α fusion proteins and nuclear extracts of transfected Cos7 cells. Our functional studies revealed that mutation c-57-64delCACGCGGT;c-55G>C reduces HNF1A promoter activity in Min6 cells and that missense mutations have variable effects. Mutation p.Arg271Trp impairs HNF-1α activity in all conditions tested, whereas mutations p.Val133Met, p.Glu235Gly and p.Pro379Arg exert differential effects depending on the target promoter. In contrast, substitution p.Thr196Ala does not appear to alter HNF-1α function. Our results suggest that HNF1A mutations may have differential effects on the regulation of specific target genes, which could contribute to the variability of the MODY3 clinical phenotype. PMID:21170474

MeHG Stimulates Antiapoptotic Signaling in Stem Cells

DTIC Science & Technology

2010-07-01

hormone signaling. Hypothyroid rats display increased caspase 3 activity and increased levels of pro-apoptotic Bcl2 members and decreased Bcl2 family...2006) Increased pro-nerve growth factor and p75 neurotrophin receptor levels in developing hypothyroid rat cerebral cortex are associated with enhanced...Tiwari M and Godbole MM (2003) Hypothyroidism alters the expression of Bcl-2 family genes to induce enhanced apoptosis in the developing

Variable transcription of pro- and anti-inflammatory cytokines in phocine lymphocytes following canine distemper virus infection.

PubMed

Seibel, H; Siebert, U; Rosenberger, T; Baumgärtner, W

2014-10-15

Canine distemper virus (CDV) is a highly contagious viral pathogen. Domesticated dogs are the main reservoir of CDV. Although phocine distemper virus was responsible for the recent epidemics in seals in the North and Baltic Seas, most devastating epidemics in seals were also caused by CDV. To further study the pathogenesis of CDV infection in seals, it was the aim of the present study to investigate the mechanisms of CDV induced immunosuppression in seals by analyzing the gene transcription of different pro- and anti-inflammatory cytokines in Concanavalin A (Con A) stimulated and non-stimulated phocine lymphocytes in vitro following infection with the CDV Onderstepoort (CDV-OND) strain. Phocine lymphocytes were isolated via density gradient centrifugation. The addition of 1 μg/ml Con A and virus was either performed simultaneously or lymphocytes were stimulated for 48 h with Con A prior to virus infection. Gene transcription of interleukin (IL)-6, IL-12 and tumor necrosis factor alpha (TNFα) as pro-inflammatory cytokines and IL-4, IL-10 and transforming growth factor beta (TGFβ) as anti-inflammatory cytokines were determined by using RT-qPCR. CDV-OND infection caused an initial increase of pro-inflammatory phocine cytokines mRNA 24h after infection, followed by a decrease in gene transcription after 48 h. A strong increase in the transcription of IL-4 and TGFβ was detected after 48 h when virus and mitogen were added simultaneously. An increased IL-10 production occurred only when stimulation and infection were performed simultaneously. Furthermore, an inhibition of IL-12 on IL-4 was noticed in phocine lymphocytes which were stimulated for 48 h prior to infection. In summary, the duration of the stimulation or the lymphocytes seem to have an important influence on the cytokine transcription and indicates that the outcome of CDV infection is dependent on various factors that might sensitize lymphocytes or make them more susceptible or reactive to CDV infection. Copyright © 2014 Elsevier B.V. All rights reserved.

Anti-adipogenic effects of KD025 (SLx-2119), a ROCK2-specific inhibitor, in 3T3-L1 cells.

PubMed

Diep, Duy Trong Vien; Hong, Kyungki; Khun, Triyeng; Zheng, Mei; Ul-Haq, Asad; Jun, Hee-Sook; Kim, Young-Bum; Chun, Kwang-Hoon

2018-02-06

Adipose tissue is a specialized organ that synthesizes and stores fat. During adipogenesis, Rho and Rho-associated kinase (ROCK) 2 are inactivated, which enhances the expression of pro-adipogenic genes and induces the loss of actin stress fibers. Furthermore, pan ROCK inhibitors enhance adipogenesis in 3T3-L1 cells. Here, we show that KD025 (formerly known as SLx-2119), a ROCK2-specific inhibitor, suppresses adipogenesis in 3T3-L1 cells partially through a ROCK2-independent mechanism. KD025 downregulated the expression of key adipogenic transcription factors PPARγ and C/EBPα during adipogenesis in addition to lipogenic factors FABP4 and Glut4. Interestingly, adipogenesis was blocked by KD025 during days 1~3 of differentiation; after differentiation terminated, lipid accumulation was unaffected. Clonal expansion occurred normally in KD025-treated cells. These results suggest that KD025 could function during the intermediate stage after clonal expansion. Data from depletion of ROCKs showed that KD025 suppressed cell differentiation partially independent of ROCK's activity. Furthermore, no further loss of actin stress fibers emerged in KD025-treated cells during and after differentiation compared to control cells. These results indicate that in contrast to the pro-adipogenic effect of pan-inhibitors, KD025 suppresses adipogenesis in 3T3-L1 cells by regulating key pro-adipogenic factors. This outcome further implies that KD025 could be a potential anti-adipogenic/obesity agent.

Discovery and Complete Genome Sequence of a Bacteriophage from an Obligate Intracellular Symbiont of a Cellulolytic Protist in the Termite Gut

PubMed Central

Pramono, Ajeng K.; Kuwahara, Hirokazu; Itoh, Takehiko; Toyoda, Atsushi; Yamada, Akinori; Hongoh, Yuichi

2017-01-01

Termites depend nutritionally on their gut microbes, and protistan, bacterial, and archaeal gut communities have been extensively studied. However, limited information is available on viruses in the termite gut. We herein report the complete genome sequence (99,517 bp) of a phage obtained during a genome analysis of “Candidatus Azobacteroides pseudotrichonymphae” phylotype ProJPt-1, which is an obligate intracellular symbiont of the cellulolytic protist Pseudotrichonympha sp. in the gut of the termite Prorhinotermes japonicus. The genome of the phage, designated ProJPt-Bp1, was circular or circularly permuted, and was not integrated into the two circular chromosomes or five circular plasmids composing the host ProJPt-1 genome. The phage was putatively affiliated with the order Caudovirales based on sequence similarities with several phage-related genes; however, most of the 52 protein-coding sequences had no significant homology to sequences in the databases. The phage genome contained a tRNA-Gln (CAG) gene, which showed the highest sequence similarity to the tRNA-Gln (CAA) gene of the host “Ca. A. pseudotrichonymphae” phylotype ProJPt-1. Since the host genome lacked a tRNA-Gln (CAG) gene, the phage tRNA gene may compensate for differences in codon usage bias between the phage and host genomes. The phage genome also contained a non-coding region with high nucleotide sequence similarity to a region in one of the host plasmids. No other phage-related sequences were found in the host ProJPt-1 genome. To the best of our knowledge, this is the first report of a phage from an obligate, mutualistic endosymbiont permanently associated with eukaryotic cells. PMID:28321010

Fatigue and gene expression in human leukocytes: Increased NF-κB and decreased glucocorticoid signaling in breast cancer survivors with persistent fatigue

PubMed Central

Bower, Julienne E.; Ganz, Patricia A.; Irwin, Michael R.; Arevalo, Jesusa M.G.; Cole, Steve W.

2013-01-01

Fatigue is highly prevalent in the general population and is one of the most common side effects of cancer treatment. There is growing evidence that pro-inflammatory cytokines play a role in cancer-related fatigue, although the molecular mechanisms for chronic inflammation and fatigue have not been determined. The current study utilized genome-wide expression microarrays to identify differences in gene expression and associated alterations in transcriptional activity in leukocytes from breast cancer survivors with persistent fatigue (n = 11) and non-fatigued controls (n = 10). We focused on transcription of inflammation-related genes, particularly those responsive to the pro-inflammatory NF-κB transcription control pathway. Further, given the role of glucocorticoids as key regulators of inflammatory processes, we examined transcription of glucocorticoid-responsive genes indicative of potential glucocorticoid receptor (GR) desensitization. Plasma levels of cortisol were also assessed. Consistent with hypotheses, results showed increased expression of transcripts with response elements for NF-κB, and reduced expression of transcripts with response elements for glucocorticoids (p < .05) in fatigued breast cancer survivors. No differences in plasma levels of cortisol were observed. These data indicate that increased activity of pro-inflammatory transcription factors may contribute to persistent cancer-related fatigue and provide insight into potential mechanisms for tonic increases in NF-κB activity, specifically decreased expression of GR anti-inflammatory transcription factors. PMID:20854893

Small Molecule-Induced Complement Factor D (Adipsin) Promotes Lipid Accumulation and Adipocyte Differentiation

PubMed Central

Jang, Byung-Hyun; Chang, Seo-Hyuk; Yun, Ui Jeong; Park, Ki-Moon; Waki, Hironori; Li, Dean Y.; Tontonoz, Peter; Park, Kye Won

2016-01-01

Adipocytes are differentiated by various transcriptional cascades integrated on the master regulator, Pparγ. To discover new genes involved in adipocyte differentiation, preadipocytes were treated with three newly identified pro-adipogenic small molecules and GW7845 (a Pparγ agonist) for 24 hours and transcriptional profiling was analyzed. Four genes, Peroxisome proliferator-activated receptor γ (Pparγ), human complement factor D homolog (Cfd), Chemokine (C-C motif) ligand 9 (Ccl9), and GIPC PDZ Domain Containing Family Member 2 (Gipc2) were induced by at least two different small molecules but not by GW7845. Cfd and Ccl9 expressions were specific to adipocytes and they were altered in obese mice. Small hairpin RNA (shRNA) mediated knockdown of Cfd in preadipocytes inhibited lipid accumulation and expression of adipocyte markers during adipocyte differentiation. Overexpression of Cfd promoted adipocyte differentiation, increased C3a production, and led to induction of C3a receptor (C3aR) target gene expression. Similarly, treatments with C3a or C3aR agonist (C4494) also promoted adipogenesis. C3aR knockdown suppressed adipogenesis and impaired the pro-adipogenic effects of Cfd, further suggesting the necessity for C3aR signaling in Cfd-mediated pro-adipogenic axis. Together, these data show the action of Cfd in adipogenesis and underscore the application of small molecules to identify genes in adipocytes. PMID:27611793

Evaluation of human epidermal growth factor receptor 2 (HER2) single nucleotide polymorphisms (SNPs) in normal and breast tumor tissues and their link with breast cancer prognostic factors.

PubMed

Furrer, Daniela; Lemieux, Julie; Côté, Marc-André; Provencher, Louise; Laflamme, Christian; Barabé, Frédéric; Jacob, Simon; Michaud, Annick; Diorio, Caroline

2016-12-01

Amplification of the human epidermal growth factor receptor 2 (HER2) gene is associated with worse prognosis and decreased overall survival in breast cancer patients. The HER2 gene contains several polymorphisms; two of the best-characterized HER2 polymorphisms are Ile655Val and Ala1170Pro. The aim of this study was to evaluate the association between these two HER2 polymorphisms in normal breast and breast cancer tissues and known breast cancer prognostic factors in a retrospective cohort study of 73 women with non-metastatic HER2-positive breast cancer. HER2 polymorphisms were assessed in breast cancer tissue and normal breast tissue using TaqMan assay. Ala1170Pro polymorphism in normal breast tissue was associated with age at diagnosis (p = 0.007), tumor size (p = 0.004) and lymphovascular invasion (p = 0.06). Similar significant associations in cancer tissues were observed. No association between the Ile655Val polymorphism and prognostic factors were observed. However, we found significant differences in the distribution of Ile655Val (p = 0.03) and Ala1170Pro (p = 0.01) genotypes between normal breast and breast tumor tissues. This study demonstrates that only the Ala1170Pro polymorphism is associated with prognostic factors in HER2-positive breast cancer patients. Moreover, our results suggest that both HER2 polymorphisms could play a significant role in carcinogenesis in non-metastatic HER2-positive breast cancer women. Copyright © 2016 Elsevier Ltd. All rights reserved.

Effects of inhalational anaesthetics in experimental allergic asthma.

PubMed

Burburan, S M; Silva, J D; Abreu, S C; Samary, C S; Guimarães, I H L; Xisto, D G; Morales, M M; Rocco, P R M

2014-06-01

We evaluated whether isoflurane, halothane and sevoflurane attenuate the inflammatory response and improve lung morphofunction in experimental asthma. Fifty-six BALB/c mice were sensitised and challenged with ovalbumin and anaesthetised with isoflurane, halothane, sevoflurane or pentobarbital sodium for one hour. Lung mechanics and histology were evaluated. Gene expression of pro-inflammatory (tumour necrosis factor-α), pro-fibrogenic (transforming growth factor-β) and pro-angiogenic (vascular endothelial growth factor) mediators, as well as oxidative process modulators, were analysed. These modulators included nuclear factor erythroid-2 related factor 2, sirtuin, catalase and glutathione peroxidase. Isoflurane, halothane and sevoflurane reduced airway resistance, static lung elastance and atelectasis when compared with pentobarbital sodium. Sevoflurane minimised bronchoconstriction and cell infiltration, and decreased tumour necrosis factor-α, transforming growth factor-β, vascular endothelial growth factor, sirtuin, catalase and glutathione peroxidase, while increasing nuclear factor erythroid-2-related factor 2 expression. Sevoflurane down-regulated inflammatory, fibrogenic and angiogenic mediators, and modulated oxidant-antioxidant imbalance, improving lung function in this model of asthma. © 2014 The Association of Anaesthetists of Great Britain and Ireland.

Threonine-4 of mammalian RNA polymerase II CTD is targeted by Polo-like kinase 3 and required for transcriptional elongation

PubMed Central

Hintermair, Corinna; Heidemann, Martin; Koch, Frederic; Descostes, Nicolas; Gut, Marta; Gut, Ivo; Fenouil, Romain; Ferrier, Pierre; Flatley, Andrew; Kremmer, Elisabeth; Chapman, Rob D; Andrau, Jean-Christophe; Eick, Dirk

2012-01-01

Eukaryotic RNA polymerase II (Pol II) has evolved an array of heptad repeats with the consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7 at the carboxy-terminal domain (CTD) of the large subunit (Rpb1). Differential phosphorylation of Ser2, Ser5, and Ser7 in the 5′ and 3′ regions of genes coordinates the binding of transcription and RNA processing factors to the initiating and elongating polymerase complexes. Here, we report phosphorylation of Thr4 by Polo-like kinase 3 in mammalian cells. ChIPseq analyses indicate an increase of Thr4-P levels in the 3′ region of genes occurring subsequently to an increase of Ser2-P levels. A Thr4/Ala mutant of Pol II displays a lethal phenotype. This mutant reveals a global defect in RNA elongation, while initiation is largely unaffected. Since Thr4 replacement mutants are viable in yeast we conclude that this amino acid has evolved an essential function(s) in the CTD of Pol II for gene transcription in mammalian cells. PMID:22549466

TGF{beta}1 polymorphisms and late clinical radiosensitivity in patients treated for gynecologic tumors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ruyck, Kim de; Van Eijkeren, Marc; Claes, Kathleen

2006-07-15

Purpose: To investigate the association between six transforming growth factor {beta}1 gene (TGF{beta}1) polymorphisms (-1.552delAGG, -800G>A, -509C>T, Leu10Pro, Arg25Pro, Thr263Ile) and the occurrence of late normal tissue reactions after gynecologic radiotherapy (RT). Methods and Materials: Seventy-eight women with cervical or endometrial cancer and 140 control individuals were included in the study. According to the Common Terminology Criteria for Adverse Events version 3.0 (CTCAEv3.0) scale, 25 patients showed late adverse RT reactions (CTC2+), of whom 11 had severe complications (CTC3+). Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), single base extension and genotyping assays were performed to examine the polymorphic sites inmore » TGF{beta}1. Results: Homozygous variant -1.552delAGG, -509TT, and 10Pro genotypes were associated with the risk of developing late severe RT reactions. Triple (variant) homozygous patients had a 3.6 times increased risk to develop severe RT reactions (p = 0.26). Neither the -800A allele, nor the 25Pro allele or the 263Ile allele were associated with clinical radiosensitivity. There was perfect linkage disequilibrium (LD) between the -1.552delAGG and the -509C>T polymorphisms, and tight LD between the -1.552/-509 and the Leu10Pro polymorphisms. Haplotype analysis revealed two major haplotypes but could not distinguish radiosensitive from nonradiosensitive patients. Conclusions: The present study shows that homozygous variant TGF{beta}1 -1.552delAGG, -509TT, and 10Pro genotypes may be associated with severe clinical radiosensitivity after gynecologic RT.« less

E1(-)E4(+) adenoviral gene transfer vectors function as a "pro-life" signal to promote survival of primary human endothelial cells.

PubMed

Ramalingam, R; Rafii, S; Worgall, S; Brough, D E; Crystal, R G

1999-05-01

Although endothelial cells are quiescent and long-lived in vivo, when they are removed from blood vessels and cultured in vitro they die within days to weeks. In studies of the interaction of E1(-)E4(+) replication-deficient adenovirus (Ad) vectors and human endothelium, the cells remained quiescent and were viable for prolonged periods. Evaluation of these cultures showed that E1(-)E4(+) Ad vectors provide an "antiapoptotic" signal that, in association with an increase in the ratio of Bcl2 to Bax levels, induces the endothelial cells to enter a state of "suspended animation," remaining viable for at least 30 days, even in the absence of serum and growth factors. Although the mechanisms initiating these events are unclear, the antiapoptoic signal requires the presence of E4 genes in the vector genome, suggesting that one or more E4 open reading frames of subgroup C Ad initiate a "pro-life" program that modifies cultured endothelial cells to survive for prolonged periods.

Mutations in the LHX2 gene are not a frequent cause of micro/anophthalmia

PubMed Central

Desmaison, Annaïck; Vigouroux, Adeline; Rieubland, Claudine; Peres, Christine; Calvas, Patrick

2010-01-01

Purpose Microphthalmia and anophthalmia are at the severe end of the spectrum of abnormalities in ocular development. A few genes (orthodenticle homeobox 2 [OTX2], retina and anterior neural fold homeobox [RAX], SRY-box 2 [SOX2], CEH10 homeodomain-containing homolog [CHX10], and growth differentiation factor 6 [GDF6]) have been implicated mainly in isolated micro/anophthalmia but causative mutations of these genes explain less than a quarter of these developmental defects. The essential role of the LIM homeobox 2 (LHX2) transcription factor in early eye development has recently been documented. We postulated that mutations in this gene could lead to micro/anophthalmia, and thus performed molecular screening of its sequence in patients having micro/anophthalmia. Methods Seventy patients having non-syndromic forms of colobomatous microphthalmia (n=25), isolated microphthalmia (n=18), or anophthalmia (n=17), and syndromic forms of micro/anophthalmia (n=10) were included in this study after negative molecular screening for OTX2, RAX, SOX2, and CHX10 mutations. Mutation screening of LHX2 was performed by direct sequencing of the coding sequences and intron/exon boundaries. Results Two heterozygous variants of unknown significance (c.128C>G [p.Pro43Arg]; c.776C>A [p.Pro259Gln]) were identified in LHX2 among the 70 patients. These variations were not identified in a panel of 100 control patients of mixed origins. The variation c.776C>A (p.Pro259Gln) was considered as non pathogenic by in silico analysis, while the variation c.128C>G (p.Pro43Arg) considered as deleterious by in silico analysis and was inherited from the asymptomatic father. Conclusions Mutations in LHX2 do not represent a frequent cause of micro/anophthalmia. PMID:21203406

Mutations in the LHX2 gene are not a frequent cause of micro/anophthalmia.

PubMed

Desmaison, Annaïck; Vigouroux, Adeline; Rieubland, Claudine; Peres, Christine; Calvas, Patrick; Chassaing, Nicolas

2010-12-18

Microphthalmia and anophthalmia are at the severe end of the spectrum of abnormalities in ocular development. A few genes (orthodenticle homeobox 2 [OTX2], retina and anterior neural fold homeobox [RAX], SRY-box 2 [SOX2], CEH10 homeodomain-containing homolog [CHX10], and growth differentiation factor 6 [GDF6]) have been implicated mainly in isolated micro/anophthalmia but causative mutations of these genes explain less than a quarter of these developmental defects. The essential role of the LIM homeobox 2 (LHX2) transcription factor in early eye development has recently been documented. We postulated that mutations in this gene could lead to micro/anophthalmia, and thus performed molecular screening of its sequence in patients having micro/anophthalmia. Seventy patients having non-syndromic forms of colobomatous microphthalmia (n=25), isolated microphthalmia (n=18), or anophthalmia (n=17), and syndromic forms of micro/anophthalmia (n=10) were included in this study after negative molecular screening for OTX2, RAX, SOX2, and CHX10 mutations. Mutation screening of LHX2 was performed by direct sequencing of the coding sequences and intron/exon boundaries. Two heterozygous variants of unknown significance (c.128C>G [p.Pro43Arg]; c.776C>A [p.Pro259Gln]) were identified in LHX2 among the 70 patients. These variations were not identified in a panel of 100 control patients of mixed origins. The variation c.776C>A (p.Pro259Gln) was considered as non pathogenic by in silico analysis, while the variation c.128C>G (p.Pro43Arg) considered as deleterious by in silico analysis and was inherited from the asymptomatic father. Mutations in LHX2 do not represent a frequent cause of micro/anophthalmia.

No association of the neuropeptide Y (Leu7Pro) and ghrelin gene (Arg51Gln, Leu72Met, Gln90Leu) single nucleotide polymorphisms with eating disorders.

PubMed

Kindler, Jochen; Bailer, Ursula; de Zwaan, Martina; Fuchs, Karoline; Leisch, Friedrich; Grün, Bettina; Strnad, Alexandra; Stojanovic, Mirjana; Windisch, Julia; Lennkh-Wolfsberg, Claudia; El-Giamal, Nadja; Sieghart, Werner; Kasper, Siegfried; Aschauer, Harald

2011-06-01

Genetic factors likely contribute to the biological vulnerability of eating disorders. Case-control association study on one neuropeptide Y gene (Leu7Pro) polymorphism and three ghrelin gene (Arg51Gln, Leu72Met and Gln90Leu) polymorphisms. 114 eating disorder patients (46 with anorexia nervosa, 30 with bulimia nervosa, 38 with binge eating disorder) and 164 healthy controls were genotyped. No differences were detected between patients and controls for any of the four polymorphisms in allele frequency and genotype distribution (P > 0.05). Allele frequencies and genotypes had no significant influence on body mass index (P > 0.05) in eating disorder patients. Positive findings of former case-control studies of associations between ghrelin gene polymorphisms and eating disorders could not be replicated. Neuropeptide Y gene polymorphisms have not been investigated in eating disorders before.

A Sordaria macrospora mutant lacking the leu1 gene shows a developmental arrest during fruiting body formation.

PubMed

Kück, Ulrich

2005-10-01

Developmental mutants with defects in fruiting body formation are excellent resources for the identification of genetic components that control cellular differentiation processes in filamentous fungi. The mutant pro4 of the ascomycete Sordaria macrospora is characterized by a developmental arrest during the sexual life cycle. This mutant generates only pre-fruiting bodies (protoperithecia), and is unable to form ascospores. Besides being sterile, pro4 is auxotrophic for leucine. Ascospore analysis revealed that the two phenotypes are genetically linked. After isolation of the wild-type leu1 gene from S. macrospora, complementation experiments demonstrated that the gene was able to restore both prototrophy and fertility in pro4. To investigate the control of leu1 expression, other genes involved in leucine biosynthesis specifically and in the general control of amino acid biosynthesis ("cross-pathway control") have been analysed using Northern hybridization and quantitative RT-PCR. These analyses demonstrated that genes of leucine biosynthesis are transcribed at higher levels under conditions of amino acid starvation. In addition, the expression data for the cpc1 and cpc2 genes indicate that cross-pathway control is superimposed on leucine-specific regulation of fruiting body development in the leu1 mutant. This was further substantiated by growth experiments in which the wild-type strain was found to show a sterile phenotype when grown on a medium containing the amino acid analogue 5-methyl-tryptophan. Taken together, these data show that pro4 represents a novel mutant type in S. macrospora, in which amino acid starvation acts as a signal that interrupts the development of the fruiting body.

Notch regulates BMP responsiveness and lateral branching in vessel networks via SMAD6

PubMed Central

Mouillesseaux, Kevin P.; Wiley, David S.; Saunders, Lauren M.; Wylie, Lyndsay A.; Kushner, Erich J.; Chong, Diana C.; Citrin, Kathryn M.; Barber, Andrew T.; Park, Youngsook; Kim, Jun-Dae; Samsa, Leigh Ann; Kim, Jongmin; Liu, Jiandong; Jin, Suk-Won; Bautch, Victoria L.

2016-01-01

Functional blood vessel growth depends on generation of distinct but coordinated responses from endothelial cells. Bone morphogenetic proteins (BMP), part of the TGFβ superfamily, bind receptors to induce phosphorylation and nuclear translocation of SMAD transcription factors (R-SMAD1/5/8) and regulate vessel growth. However, SMAD1/5/8 signalling results in both pro- and anti-angiogenic outputs, highlighting a poor understanding of the complexities of BMP signalling in the vasculature. Here we show that BMP6 and BMP2 ligands are pro-angiogenic in vitro and in vivo, and that lateral vessel branching requires threshold levels of R-SMAD phosphorylation. Endothelial cell responsiveness to these pro-angiogenic BMP ligands is regulated by Notch status and Notch sets responsiveness by regulating a cell-intrinsic BMP inhibitor, SMAD6, which affects BMP responses upstream of target gene expression. Thus, we reveal a paradigm for Notch-dependent regulation of angiogenesis: Notch regulates SMAD6 expression to affect BMP responsiveness of endothelial cells and new vessel branch formation. PMID:27834400

Tumor necrosis factor-alpha activates signal transduction in hypothalamus and modulates the expression of pro-inflammatory proteins and orexigenic/anorexigenic neurotransmitters.

PubMed

Amaral, Maria E; Barbuio, Raquel; Milanski, Marciane; Romanatto, Talita; Barbosa, Helena C; Nadruz, Wilson; Bertolo, Manoel B; Boschero, Antonio C; Saad, Mario J A; Franchini, Kleber G; Velloso, Licio A

2006-07-01

Tumor necrosis factor-alpha (TNF-alpha) is known to participate in the wastage syndrome that accompanies cancer and severe infectious diseases. More recently, a role for TNF-alpha in the pathogenesis of type 2 diabetes mellitus and obesity has been shown. Much of the regulatory action exerted by TNF-alpha upon the control of energy stores depends on its action on the hypothalamus. In this study, we show that TNF-alpha activates canonical pro-inflammatory signal transduction pathways in the hypothalamus of rats. These signaling events lead to the transcriptional activation of an early responsive gene and to the induction of expression of cytokines and a cytokine responsive protein such as interleukin-1beta, interleukin-6, interleukin-10 and suppressor of cytokine signalling-3, respectively. In addition, TNF-alpha induces the expression of neurotransmitters involved in the control of feeding and thermogenesis. Thus, TNF-alpha may act directly in the hypothalamus inducing a pro-inflammatory response and the modulation of expression of neurotransmitters involved in energy homeostasis.

The effect of newly synthesized progesterone derivatives on apoptotic and angiogenic pathway in MCF-7 breast cancer cells.

PubMed

Yahya, Shaymaa M M; Abdelhamid, Abdou O; Abd-Elhalim, Mervat M; Elsayed, Ghada H; Eskander, Emad F

2017-10-01

Due to its high potency and selectivity, anticancer agents consisting of combined molecules have gained great interests. The current study introduces newly synthesized progesterone derivatives of promising anticancer effect. Moreover, the pro-apoptotic and anti-angiogenic effects of these compounds were studied extensively. Several thiazole, pyridine, pyrazole, thiazolopyridine and pyrazolopyridine progesterone derivatives were synthesized. The structure of the novel progesterone derivatives was elucidated and confirmed using the analytical and spectral data. This novel derivatives were tested for their cytotoxic effect against human breast cancer cells (MCF-7) using neutral red uptake assay. Tested compounds showed anticancer activity against MCF-7 cancer cell line in the descending order of 7>2>3>8>6>9>4. The expression levels of Bcl-2, survivin, CCND1, CDC2, P53 and P21, VEGF, Hif-1α, MMP-2, MMP-9, Ang-1, Ang-2, and FGF-1 genes were investigated using QRT-PCR (Quantitative real time-polymerase chain reaction). The study clarified that compounds 2, 3, 4, 6, 7, 8 and 9 showed significant pro-apoptotic effect through the down regulation of Bcl-2., besides, survivin and CCND1 expression levels were down regulated by compounds 3, 4, 6, 7, 8, 9. However, Compound 4 may exert this pro-apoptotic effect through the up-regulation of P53 gene expression. On the other hand, the anti-angiogenic effect of these newly synthesized derivatives was due to their down regulation of VEGF, Ang-2, MMP-9 and FGF-1; and the up-regulation of HIF-1α and ang-1. This study recommended promising pro-apoptotic and anti-angiogenic anticancer agents acting through the regulation of key regulators of apoptosis, cell cycle genes, and pro-angiogenic genes. Copyright © 2017 Elsevier Inc. All rights reserved.

Calcitriol enhances fat synthesis factors and calpain activity in co-cultured cells.

PubMed

Choi, Hyuck; Myung, Kyuho

2014-08-01

We have conducted an in vitro experiment to determine whether calcitriol can act as a fat synthesizer and/or meat tenderizer when skeletal muscle cells, adipose tissue, and macrophages are co-cultured. When co-cultured, pro-inflammatory cytokine tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) expression increased, whereas decreased anti-inflammatory cytokine (IL-10 and IL-15) expression decreased in both C2C12 and 3T3-L1 cells. Calcitriol increased reactive oxygen species (ROS) production in the media. While adiponectin gene expression decreased, leptin, resistin, CCAAT-enhancer-binding protein-beta (C/EBP-β), and peroxisome proliferator-activated receptor gamma (PPAR-γ) gene expression was significantly (P < 0.047) increased with calcitriol in 3T3-L1 cells co-cultured with two different cell types. Inducible nitric oxide synthase (iNOS) protein levels were also stimulated in the C2C12 and 3T3-L1 cells, but arginase l was attenuated by calcitriol. Cacitriol highly amplified (P = 0.008) µ-calpain gene expression in co-cultured C2C12 cells. The results showed an overall increase in pro-inflammatory cytokines and a decrease in anti-inflammatory cytokines of C2C12 and 3T3-L1 cells with calcitriol in co-culture systems. µ-Calpain protein was also augmented in differentiated C2C12 cells with calcitriol. These findings suggest that calcitriol can be used as not only fat synthesizer, but meat tenderizer, in meat-producing animals. © 2014 International Federation for Cell Biology.

Analysis of PGC-1{alpha} variants Gly482Ser and Thr612Met concerning their PPAR{gamma}2-coactivation function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nitz, Inke; Ewert, Agnes; Klapper, Maja

2007-02-09

Peroxisome proliferator-activated receptor-{gamma} coactivator-1{alpha} (PGC-1{alpha}) is a cofactor involved in adaptive thermogenesis, fatty acid oxidation, and gluconeogenesis. Dysfunctions of this protein are likely to contribute to the development of obesity and the metabolic syndrome. This is in part but not definitely confirmed by results of population studies. The aim of this study was to investigate if common genetic variants rs8192678 (Gly482Ser) and rs3736265 (Thr612Met) in the PGC-1{alpha} gene lead to a functional consequence in cofactor activity using peroxisome proliferator-activated receptor-{gamma} 2 (PPAR{gamma}2) as interacting transcription factor. Reporter gene assays in HepG2 cells with wildtype and mutant proteins of both PGC1{alpha}more » and PPAR{gamma}2 (Pro12Ala, rs1801282) using the acyl-CoA-binding protein (ACBP) promoter showed no difference in coactivator activity. This is First study implicating that the Gly482Ser and Thr612Met polymorphisms in PGC-1{alpha} and Pro12Ala polymorphism in PPAR{gamma}2 do not affect the functional integrity of these proteins.« less

Gene-specific mechanisms direct glucocorticoid-receptor-driven repression of inflammatory response genes in macrophages

PubMed Central

Sacta, Maria A; Tharmalingam, Bowranigan; Coppo, Maddalena; Rollins, David A; Deochand, Dinesh K; Benjamin, Bradley; Yu, Li; Zhang, Bin; Hu, Xiaoyu; Li, Rong; Chinenov, Yurii

2018-01-01

The glucocorticoid receptor (GR) potently represses macrophage-elicited inflammation, however, the underlying mechanisms remain obscure. Our genome-wide analysis in mouse macrophages reveals that pro-inflammatory paused genes, activated via global negative elongation factor (NELF) dissociation and RNA Polymerase (Pol)2 release from early elongation arrest, and non-paused genes, induced by de novo Pol2 recruitment, are equally susceptible to acute glucocorticoid repression. Moreover, in both cases the dominant mechanism involves rapid GR tethering to p65 at NF-kB-binding sites. Yet, specifically at paused genes, GR activation triggers widespread promoter accumulation of NELF, with myeloid cell-specific NELF deletion conferring glucocorticoid resistance. Conversely, at non-paused genes, GR attenuates the recruitment of p300 and histone acetylation, leading to a failure to assemble BRD4 and Mediator at promoters and enhancers, ultimately blocking Pol2 initiation. Thus, GR displays no preference for a specific pro-inflammatory gene class; however, it effects repression by targeting distinct temporal events and components of transcriptional machinery. PMID:29424686

Macrophage Depletion Impairs Skeletal Muscle Regeneration: the Roles of Pro-fibrotic Factors, Inflammation, and Oxidative Stress.

PubMed

Xiao, Weihua; Liu, Yu; Chen, Peijie

2016-12-01

Muscle contusion is one of the most common muscle injuries in sports medicine. Macrophages play complex roles in the regeneration of skeletal muscle. However, the roles of macrophages, especially the mechanisms involved, in the regeneration of muscle contusion are still not fully understood. We hypothesize that the depletion of macrophages impairs skeletal muscle regeneration and that pro-fibrotic factors, inflammation, and oxidative stress may be involved in the process. To test these hypotheses, we constructed a muscle contusion injury and a macrophage depletion model and followed it up with morphological and gene expression analyses. The data showed that fibrotic scars were formed in the muscle of contusion injury, and they deteriorated in the mice of macrophage depletion. Furthermore, the sizes of regenerating myofibers were significantly reduced by macrophage depletion. Pro-fibrotic factors, inflammatory cytokines, chemokines, and oxidative stress-related enzymes increased significantly after muscle injury. Moreover, the expression of these factors was delayed by macrophage depletion. Most of them were still significantly higher in the later stage of regeneration. These results suggest that macrophage depletion impairs skeletal muscle regeneration and that pro-fibrotic factors, inflammation, and oxidative stress may play important roles in the process.

Apple cider vinegar boosted immunomodulatory and health promoting effects of Lactobacillus casei in common carp (Cyprinus carpio).

PubMed

Safari, Roghieh; Hoseinifar, Seyed Hossein; Nejadmoghadam, Shabnam; Khalili, Mohsen

2017-08-01

The present study was performed to investigate the immunomodulatory and health promoting effects of combined or singular administration of apple cider vinegar (ACV) and Lactobacillus casei in common carp (Cyprinus carpio) diet. An 8-week feeding trial was designed with following treatments: Control (basal diet), Pro (contains 10 7  CFU g -1 L. casei), LACV (contains 1% ACV), HACV (contains 2% ACV), Pro + LACV (contains 10 7  CFU g -1 L. casei plus 1% ACV) and Pro + HACV (contains 10 7  CFU g -1 L. casei plus 2% ACV). Evaluation of skin mucus revealed notable increase of total Ig level and lysozyme activity in Pro + LACV and Pro + HACV treatments compared other groups (P < 0.05). Similarly, serum total Ig and lysozyme activity in HACV, Pro + LACV and Pro + HACV fed carps was remarkably higher than other groups (P < 0.05). However, regarding serum alternative complement (ACH50) activity significant difference was observed just between Pro + HACV and control treatment (P < 0.05). The highest expression of immune related (LYZ, TNF-alpha, IL1b, IL8) and antioxidant enzymes genes (GSR, GST) were observed in carps fed Pro + HACV and Pro + LACV. The expression of GH gene expression in Pro, LACV and HACV treatments was significantly higher than those in control group (P < 0.05). The highest expression level of GH and IGF1 was observed in fish fed combined Pro and ACV (P < 0.05). These results indicated that co-administration of ACV boosted immunomodulatory and health promoting effects of L. casei and can be considered as a promising immunostimulants in early stage of common carp culture. Copyright © 2017 Elsevier Ltd. All rights reserved.

The nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activator dh404 protects against diabetes-induced endothelial dysfunction.

PubMed

Sharma, Arpeeta; Rizky, Luddwi; Stefanovic, Nada; Tate, Mitchel; Ritchie, Rebecca H; Ward, Keith W; de Haan, Judy B

2017-03-03

Vascular dysfunction is a pivotal event in the development of diabetes-associated vascular disease. Increased inflammation and oxidative stress are major contributors to vascular dysfunction. Nrf2, a master regulator of several anti-oxidant genes and a suppressor of inflammatory NF-κB, has potential as a target to combat oxidative stress and inflammation. The aim of this study was to investigate the effects of a novel Nrf2 activator, the bardoxolone methyl derivative dh404, on endothelial function in vitro and in vivo. dh404 at 3 mg/kg was administered to male Akita mice, an established diabetic mouse model of insulin insufficiency and hyperglycemia, from 6 weeks of age. At 26 weeks of age, vascular reactivity was assessed by wire myography, pro-inflammatory expression was assessed in the aortas by qRT-PCR and immunohistochemistry, and systemic and vascular oxidative stress measurements were determined. Additionally, studies in human aortic endothelial cells (HAECs) derived from normal and diabetic patients in the presence or absence of dh404 included assessment of pro-inflammatory genes by qRT-PCR and western blotting. Oxidative stress was assessed by three methods; L-012, DCFDA and amplex red. Static adhesion assays were performed to determine the leukocyte-endothelial interaction in the presence or absence of dh404. Dh404 significantly attenuated endothelial dysfunction in diabetic Akita mice characterized by reduced contraction in response to phenylephrine and the downregulation of inflammatory genes (VCAM-1, ICAM-1, p65, IL-1β) and pro-oxidant genes (Nox1 and Nox2). Furthermore, reduced systemic and vascular oxidative stress levels were observed in diabetic Akita mice. dh404 exhibited cytoprotective effects in diabetic HAECs in vitro, reflected by significant upregulation of Nrf2-responsive genes, NAD(P)H quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1), reduction of oxidative stress markers (O 2 ·- and H 2 O 2 ), inhibition of inflammatory genes (VCAM-1 and the p65 subunit of NF-κB) and attenuation of leukocyte-endothelial interactions (P < 0.05 for all in vitro and in vivo parameters; one or two-way ANOVA as appropriate with post hoc testing). These studies demonstrate that upregulation of Nrf2 by dh404 represents a novel therapeutic strategy to limit diabetes-associated vascular injury.

Influence of candidate polymorphisms on the dipeptidyl peptidase IV and μ-opioid receptor genes expression in aspect of the β-casomorphin-7 modulation functions in autism.

PubMed

Cieślińska, Anna; Sienkiewicz-Szłapka, Edyta; Wasilewska, Jolanta; Fiedorowicz, Ewa; Chwała, Barbara; Moszyńska-Dumara, Małgorzata; Cieśliński, Tomasz; Bukało, Marta; Kostyra, Elżbieta

2015-03-01

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder with population prevalence of approximately 60-70 per 10,000. Data shows that both opioid system function enhancement and opiate administration can result in autistic-like symptoms. Cow milk opioid peptides, including β-casomorphin-7 (BCM7, Tyr-Pro-Phe-Pro-Gly-Pro-Ile), affect the μ-opioid receptor (MOR) and are subjected to degradation resulting from the proline dipeptidyl peptidase IV (DPPIV, EC 3.4.14.5) enzyme activity. The presence of MOR and DPPIV activity are crucial factors determining biological activity of BCM7 in the human body. Our study examined the effect of β-casomorphin-7 on the MOR and DPPIV genes expression according to specific point mutations in these genes. In addition, we investigated frequency of A118G SNP in the MOR gene and rs7608798 of the DPPIV (A/G) gene in healthy and autistic children. Our research indicated correlation in DPPIV gene expression under the influence of BCM7 and hydrolyzed milk between healthy and ASD-affected children with genotype GG (P<0.0001). We also observed increased MOR gene expression in healthy children with genotype AG at polymorphic site A118G under influence of BCM7 and hydrolyzed milk. The G allele frequency was 0.09 in MOR gene and 0.68 in the DPPIV gene. But our results suggest no association between presence of the alleles G and A at position rs7608798 in DPPIV gene nor alleles A and G at position A118G of the MOR and increased incidence of ASD. Our studies emphasize the compulsion for genetic analysis in correlation with genetic factors affecting development and enhancement of autism symptoms. Copyright © 2015 Elsevier Inc. All rights reserved.

Polymorphism of the p53 tumor suppressor gene is associated with susceptibility to uterine leiomyoma.

PubMed

Denschlag, Dominik; Bettendorf, Herta; Watermann, Dirk; Keck, Christoph; Tempfer, Clemens; Pietrowski, Detlef

2005-07-01

To evaluate the association between the presence of uterine leiomyoma and two single nuclear polymorphisms of the p53 tumor suppressor and the angiopoietin-2 (ANGPT2) genes. Prospective case control study. Academic research institution. One hundred thirty-two women with clinically and surgically diagnosed uterine leiomyomas and 280 controls. Peripheral venous puncture. Genotyping was performed by polymerase chain reaction-based amplification of the Arg and Pro variants at codon 72 of the p53 gene and by restriction fragment length polymorphism analysis of the G/G and G/A alleles in exon 4 of the ANGPT2 gene. Comparing women with uterine leiomyomas and controls, no statistically significant difference with respect to allele frequency and genotype distribution were ascertained for the ANGPT2 polymorphism (P=.2 and P=.5, respectively). However, for the p53 tumor suppressor gene polymorphism, statistically significant differences in terms of a higher Pro allele frequency and a higher prevalence of the Pro/Pro genotype among women with uterine leiomyoma (32.0% vs. 16.0%, respectively, and 21.3% vs. 4.7%, respectively) were ascertained (P=.001, OR 1.74; 95% CI 1.24-2.45, P=.001; OR 3.84, 95% CI 1.81-8.14; respectively). Carriage of the p53 polymorphism at codon 72 predicts the susceptibility to leiomyoma in a Caucasian population and may contribute to the pathogenesis of uterine leiomyoma.

Twist promotes tumor metastasis in basal-like breast cancer by transcriptionally upregulating ROR1.

PubMed

Cao, Jingying; Wang, Xin; Dai, Tao; Wu, Yuanzhong; Zhang, Meifang; Cao, Renxian; Zhang, Ruhua; Wang, Gang; Jiang, Rou; Zhou, Binhua P; Shi, Jian; Kang, Tiebang

2018-01-01

Rationale: Twist is a key transcription factor for induction of epithelial-mesenchymal transition (EMT), which promotes cell migration, invasion, and cancer metastasis, confers cancer cells with stem cell-like characteristics, and provides therapeutic resistance. However, the functional roles and targeted genes of Twist in EMT and cancer progression remain elusive. Methods: The potential targeted genes of Twist were identified from the global transcriptomes of T47D/Twist cells by microarray analysis. EMT phenotype was detected by western blotting and immunofluorescence of marker proteins. The dual-luciferase reporter and chromatin immunoprecipitation assays were employed to observe the direct transcriptional induction of ROR1 by Twist. A lung metastasis model was used to study the pro-metastatic role of Twist and ROR1 by injecting MDA-MB-231 cells into tail vein of nude mice. Bio-informatics analysis was utilized to measure the metastasis-free survival of breast cancer patients. Results: Twist protein was proved to directly activate the transcription of ROR1 gene, a receptor of Wnt5a in non-canonical WNT signaling pathway. Silencing of ROR1 inhibited EMT process, cell migration, invasion, and cancer metastasis of basal-like breast cancer (BLBC) cells. Knockdown of ROR1 also ameliorated the pro-metastatic effect of Twist. Furthermore, analyses of clinical specimens indicated that high expression of both ROR1 and Twist tightly correlates with poor metastasis-free survival of breast cancer patients. Conclusion: ROR1 is a targeted gene of Twist. Twist/ROR1 signaling is critical for invasion and metastasis of BLBC cells.

Induction of activation of the antioxidant response element and stabilization of Nrf2 by 3-(3-pyridylmethylidene)-2-indolinone (PMID) confers protection against oxidative stress-induced cell death

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yao, Jia-Wei; Beijing Institute of Radiation Medicine, Beijing 100850; Liu, Jing

2012-03-01

The antioxidant response elements (ARE) are a cis-acting enhancer sequence located in regulatory regions of antioxidant and detoxifying genes. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a member of the Cap ‘n’ Collar family of transcription factors that binds to the ARE and regulates the transcription of specific ARE-containing genes. Under oxidative stress, Nrf2/ARE induction is fundamental to defense against reactive oxygen species (ROS) and serves as a key factor in the protection against toxic xenobiotics. 3-(3-Pyridylmethylidene)-2-Indolinone (PMID) is a derivative of 2-indolinone compounds which act as protein kinase inhibitors and show anti-tumor activity. However, the role of PMID inmore » the oxidative stress remains unknown. In the present study, we showed that PMID induced the activation of ARE-mediated transcription, increased the DNA-binding activity of Nrf2 and then up-regulated the expression of antioxidant genes such as HO-1, SOD, and NQO1. The level of Nrf2 protein was increased in cells treated with PMID by a post-transcriptional mechanism. Under CHX treatment, the stability of Nrf2 protein was enhanced by PMID with decreased turnover rate. We showed that PMID reduced the ubiquitination of Nrf2 and disrupted the Cullin3 (Cul3)-Keap1 interaction. Furthermore, cells treated with PMID showed resistance to cytotoxicity by H{sub 2}O{sub 2} and pro-oxidant 6-OHDA. PMID also up-regulated the antioxidant level in BALB/c mice. Taken together, the compound PMID induces the ARE-mediated gene expression through stabilization of Nrf2 protein and activation of Nrf2/ARE pathway and protects against oxidative stress-mediated cell death. -- Highlights: ► PMID up-regulates ARE-mediated antioxidant gene expression in vitro and in vivo. ► PMID enhances the stabilization of Nrf2 protein, decreasing Nrf2 turnover rate. ► PMID disrupted the Cullin3 (Cul3)-Keap1 interaction. ► PMID protects against cell death induced by H{sub 2}O{sub 2} and pro-oxidant 6-OHDA.« less

Distribution and genotype frequency of the C1431T and pro12ala polymorphisms of the peroxisome proliferator activator receptor gamma gene in an Iranian population

PubMed Central

Rooki, Hassan; Haerian, Monir-Sadat; Azimzadeh, Pedram; Ebrahimi, Mahmoud; Mirhafez, Reza; Ferns, Gordon; Ghayour-Mobarhan, Majid; Zali, Mohammad-Reza

2013-01-01

BACKGROUND: Peroxisome proliferator activator receptor gamma (PPARγ) is a nuclear transcription factor regulating multiple genes involved in cell growth, differentiation, carbohydrate and lipid metabolism and energy production. Several genetic variations in the PPARγ gene have been identified to be associated with diabetes, obesity, dyslipidemia, insulin resistance, metabolic syndrome and coronary artery disease. The present study was designed to explore the distribution of two common single nucleotide polymorphisms of the PPARγ gene (C1431T and Pro12Ala) in an Iranian population. MATERIALS AND METHODS: Genotype frequencies for these two polymorphisms were compared for 160 healthy Iranian individuals with reports from other populations. The Genotyping was performed using real-time polymerase chain reaction. RESULTS: The genotype distribution of the C1431T PPARγ polymorphism was 0.869 for the CC genotype, 0.119 for the CT genotype and 0.013 for uncommon TT genotype. Allelic frequencies were 0.93 for C and 0.07 for T allele respectively. For the Pro12Ala polymorphism of PPARγ gene, genotypic distributions and allelic frequencies were, 0.813 for CC, 0.181 for CG and 0.06 for GG and 0.903 for C and 0.097 for G respectively. Allelic and genotypic frequencies for both polymorphisms of PPARγ gene were in Hardy-Weinberg equilibrium. CONCLUSIONS: Iran is a country with an ethnically diverse population and a comparison of allelic and genotypic frequencies of PPARγ C1431T and Pro12Ala polymorphisms between our population and others showed significant differences. PMID:24497707

Anti-inflammatory action of high molecular weight Mytilus edulis hydrolysates fraction in LPS-induced RAW264.7 macrophage via NF-κB and MAPK pathways.

PubMed

Kim, Young-Sang; Ahn, Chang-Bum; Je, Jae-Young

2016-07-01

Anti-inflammatory Mytilus edulis hydrolysates (MEHs) were prepared by peptic hydrolysis and MEH was further fractionated into three fractions based on molecular weight, namely >5kDa, 1-5kDa, and <1kDa. The >5kDa peptide fraction exerted the highest nitric oxide (NO) inhibitory activity and inhibited prostaglandin E2 (PGE2) secretion in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. Pretreatment with the >5kDa peptide fraction markedly inhibited LPS-stimulated inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein and gene expressions. Stimulation by LPS induced the production of pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and -1β (IL-1β), whereas co-treatment with the >5kDa peptide fraction suppressed pro-inflammatory cytokine production. The >5kDa peptide fraction inhibited the translocation of NF-κB (nuclear factor-kappa B) through the prevention of IκBα (inhibitory factor kappa B alpha) phosphorylation and degradation and also inhibited the MAPK signaling pathway in LPS-stimulated RAW264.7 macrophages. Copyright © 2016 Elsevier Ltd. All rights reserved.

Suppression of pro-inflammatory and pro-survival biomarkers in oral cancer patients consuming a black raspberry phytochemical-rich troche

PubMed Central

Knobloch, Thomas J.; Uhrig, Lana K.; Pearl, Dennis K.; Casto, Bruce C.; Warner, Blake M.; Clinton, Steven K.; Sardo-Molmenti, Christine L.; Ferguson, Jeanette M.; Daly, Brett T.; Riedl, Kenneth; Schwartz, Steven J.; Vodovotz, Yael; Buchta, Anthony J.; Schuller, David E.; Ozer, Enver; Agrawal, Amit; Weghorst, Christopher M.

2016-01-01

Black raspberries (BRBs) demonstrate potent inhibition of aerodigestive tract carcinogenesis in animal models. However, translational clinical trials evaluating the ability of BRB phytochemicals to impact molecular biomarkers in the oral mucosa remain limited. The present phase 0 study addresses a fundamental question for oral cancer food-based prevention: Do BRB phytochemicals successfully reach the targeted oral tissues and reduce pro-inflammatory and anti-apoptotic gene expression profiles? Patients with biopsy-confirmed oral squamous cell carcinomas (OSCCs) administered oral troches containing freeze-dried BRB powder from the time of enrollment to the date of curative intent surgery (13.9 ± 1.27 days). Transcriptional biomarkers were evaluated in patient-matched OSCCs and non-involved high at-risk mucosa (HARM) for BRB-associated changes. Significant expression differences between baseline OSCC and HARM tissues were confirmed using a panel of genes commonly deregulated during oral carcinogenesis. Following BRB troche administration, the expression of pro-survival genes (AURKA, BIRC5, EGFR) and pro-inflammatory genes (NFKB1, PTGS2) were significantly reduced. There were no BRB-associated Grade 3–4 toxicities or adverse events and 79.2% (N = 30) of patients successfully completed the study with high levels of compliance (97.2%). The BRB phytochemicals cyanidin-3-rutinoside and cyanidin-3-xylosylrutinoside were detected in all OSCC tissues analyzed, demonstrating that bioactive components were successfully reaching targeted OSCC tissues. We confirmed that hallmark anti-apoptotic and pro-inflammatory molecular biomarkers were over-expressed in OSCCs and that their gene expression was significantly reduced following BRB troche administration. Since these molecular biomarkers are fundamental to oral carcinogenesis and are modifiable, they may represent emerging biomarkers of molecular efficacy for BRB-mediated oral cancer chemoprevention. PMID:26701664

Fe2+ chelator proferrorosamine A: a gene cluster of Erwinia rhapontici P45 involved in its synthesis and its impact on growth of Erwinia amylovora CFBP1430.

PubMed

Born, Yannick; Remus-Emsermann, Mitja N P; Bieri, Marco; Kamber, Tim; Piel, Jörn; Pelludat, Cosima

2016-02-01

Proferrorosamine A (proFRA) is an iron (Fe2+) chelator produced by the opportunistic plant pathogen Erwinia rhapontici P45. To identify genes involved in proFRA synthesis, transposon mutagenesis was performed. The identified 9.3 kb gene cluster, comprising seven genes, designated rosA-rosG, encodes proteins that are involved in proFRA synthesis. Based on gene homologies, a biosynthetic pathway model for proFRA is proposed. To obtain a better understanding of the effect of proFRA on non-proFRA producing bacteria, E. rhapontici P45 was co-cultured with Erwinia amylovora CFBP1430, a fire-blight-causing plant pathogen. E. rhapontici P45, but not corresponding proFRA-negative mutants, led to a pink coloration of E. amylovora CFBP1430 colonies on King's B agar, indicating accumulation of the proFRA-iron complex ferrorosamine, and growth inhibition in vitro. By saturating proFRA-containing extracts with Fe2+, the inhibitory effect was neutralized, suggesting that the iron-chelating capability of proFRA is responsible for the growth inhibition of E. amylovora CFBP1430.

[The Effects of Chronic Alcoholization on the Expression of Brain-Derived Neurotrophic Factor and Its Receptors in the Brains of Mice Genetically Predisposed to Depressive-Like Behavior].

PubMed

Bazovkina, D V; Kondaurova, E M; Tsybko, A S; Kovetskaya, A I; Ilchibaeva, T V; Naumenko, V S

2017-01-01

Brain-derived neurotropic factor (BDNF) plays an important role in mechanisms of depression. Precursor protein of this factor (proBDNF) can initiate apoptosis in the brain, while the mature form of BDNF is involved in neurogenesis. It is known that chronic alcoholization leads to the activation of apoptotic processes, neurodegeneration, brain injury, and cognitive dysfunction. In this work, we have studied the influence of long-term ethanol exposure on the proBDNF and BDNF protein levels, as well as on the expression of genes that encode these proteins in the brain structures of ASC mice with genetic predisposition to depressive-like behavior and in mice from parental nondepressive CBA strain. It was shown that chronic alcoholization results in a reduction of the BDNF level in the hippocampus and an increase in the amount of TrkB and p75 receptors in the frontal cortex of nondepressive CBA mice. At the same time, the long-term alcoholization of depressive ASC mice results in an increase of the proBDNF level in the frontal cortex and a reduction in the p75 protein level in the hippocampus. It has also been shown that, in depressive ASC mice, proBDNF and BDNF levels are significantly lower in the hippocampus and the frontal cortex compared with nondepressive CBA strain. However, no significant differences in the expression of genes encoding the studied proteins were observed. Thus, changes in the expression patterns of proBDNF, BDNF, and their receptors under the influence of alcoholization in the depressive ASC strain and nondepressive CBA strain mice are different.

Genome-Wide Analyses and Functional Classification of Proline Repeat-Rich Proteins: Potential Role of eIF5A in Eukaryotic Evolution

PubMed Central

Mandal, Ajeet; Mandal, Swati; Park, Myung Hee

2014-01-01

The eukaryotic translation factor, eIF5A has been recently reported as a sequence-specific elongation factor that facilitates peptide bond formation at consecutive prolines in Saccharomyces cerevisiae, as its ortholog elongation factor P (EF-P) does in bacteria. We have searched the genome databases of 35 representative organisms from six kingdoms of life for PPP (Pro-Pro-Pro) and/or PPG (Pro-Pro-Gly)-encoding genes whose expression is expected to depend on eIF5A. We have made detailed analyses of proteome data of 5 selected species, Escherichia coli, Saccharomyces cerevisiae, Drosophila melanogaster, Mus musculus and Homo sapiens. The PPP and PPG motifs are low in the prokaryotic proteomes. However, their frequencies markedly increase with the biological complexity of eukaryotic organisms, and are higher in newly derived proteins than in those orthologous proteins commonly shared in all species. Ontology classifications of S. cerevisiae and human genes encoding the highest level of polyprolines reveal their strong association with several specific biological processes, including actin/cytoskeletal associated functions, RNA splicing/turnover, DNA binding/transcription and cell signaling. Previously reported phenotypic defects in actin polarity and mRNA decay of eIF5A mutant strains are consistent with the proposed role for eIF5A in the translation of the polyproline-containing proteins. Of all the amino acid tandem repeats (≥3 amino acids), only the proline repeat frequency correlates with functional complexity of the five organisms examined. Taken together, these findings suggest the importance of proline repeat-rich proteins and a potential role for eIF5A and its hypusine modification pathway in the course of eukaryotic evolution. PMID:25364902

Differential regulations of fibronectin and laminin in Smad2 activation in vascular endothelial cells in response to disturbed flow.

PubMed

Yang, Tung-Lin; Lee, Pei-Ling; Lee, Ding-Yu; Wang, Wei-Li; Wei, Shu-Yi; Lee, Chih-I; Chiu, Jeng-Jiann

2018-01-02

Atherosclerosis occurs in arterial curvatures and branches, where the flow is disturbed with low and oscillatory shear stress (OSS). The remodeling and alterations of extracellular matrices (ECMs) and their composition is the critical step in atherogenesis. In this study, we investigated the effects of different ECM proteins on the regulation of mechanotransduction in vascular endothelial cells (ECs) in response to OSS. Through the experiments ranging from in vitro cell culture studies on effects of OSS on molecular signaling to in vivo examinations on clinical specimens from patients with coronary artery disease (CAD), we elucidated the roles of integrins and different ECMs, i.e., fibronectin (FN) and laminin (LM), in transforming growth factor (TGF)-β receptor (TβR)-mediated Smad2 activation and nuclear factor-κB (NF-κB) signaling in ECs in response to OSS and hence atherogenesis. OSS at 0.5±12 dynes/cm 2 induces sustained increases in the association of types I and II TβRs with β1 and β3 integrins in ECs grown on FN, but it only transient increases in ECs grown on LM. OSS induces a sustained activation of Smad2 in ECs on FN, but only a transient activation of Smad2 in ECs on LM. OSS-activation of Smad2 in ECs on FN regulates downstream NF-κB signaling and pro-inflammatory gene expression through the activation of β1 integrin and its association with TβRs. In contrast, OSS induces transient activations of β1 and β3 integrins in ECs on LM, which associate with type I TβR to regulate Smad2 phosphorylation, resulting in transient induction of NF-κB and pro-inflammatory gene expression. In vivo investigations on diseased human coronary arteries from CAD patients revealed that Smad2 is highly activated in ECs of atherosclerotic lesions, which is accompanied by the concomitant increase of FN rather than LM in the EC layer and neointimal region of atherosclerotic lesions. Our findings provide new insights into the mechanisms of how OSS regulates Smad2 signaling and pro-inflammatory genes through the complex signaling networks of integrins, TβRs, and ECMs, thus illustrating the molecular basis of regional pro-inflammatory activation within disturbed flow regions in the arterial tree.

Gene-diet-related factors of hyperglycaemia in postmenopausal women.

PubMed

Grygiel-Górniak, Bogna; Kaczmarek, Elżbieta; Mosor, Maria; Przysławski, Juliusz; Nowak, Jerzy

2018-05-01

As ageing and increased body fat are the signs of insulin resistance, we have studied whether the presence of Pro12Ala and C1431T of peroxisome proliferator-activated receptor gamma 2 gene and Trp64Arg of beta 3-adrenergic receptor gene may predispose to the hyperglycaemia development in postmenopausal women, who have never undergone hypoglycaemic treatment. The distributions of selected allele and genotype frequencies were determined by the PCR-RFLP method in normo- and hyperglycaemic, who have never been diagnosed and treated for diabetes mellitus were measured. The amount of body fat and lean body mass (LBM) were assessed by the bioimpedance method and nutritional habits by 7-day dietary recall. There were no differences between the distribution of genotypes and the allele frequencies of the Pro12Ala, C1431T and Trp64Arg polymorphisms in normo- and hyperglycaemic women. Hyperglycaemic women were characterized by visceral obesity, hypertension, higher serum insulin and triglycerides, higher intake of fat and lower consumption of complex carbohydrates and B vitamins. Normoglycaemic women with Pro12Pro polymorphism acquired higher energy from dietary fat (p < 0.0276) and lower energy from carbohydrates (p < 0.0480) than normoglycaemic Ala12 carriers. Subjects with Pro12Pro polymorphism and LBM > 58% of total body mass or with Trp64Trp and normal triglycerides have higher chance of normoglycaemia. Genotyping for Pro12Ala and Trp64Arg polymorphism in postmenopausal women may have the clinical benefit of predicting hyperglycaemia, thereby contributing to the prevention of diabetes mellitus development in the future. However, not only the genetic background but also the dietary habits (intake of fat, carbohydrates and B vitamins) determine the risk of hyperglycaemia.

PPARγ gene polymorphism, C-reactive protein level, BMI and periodontitis in post-menopausal Japanese women.

PubMed

Wang, Yangming; Sugita, Noriko; Yoshihara, Akihiro; Iwasaki, Masanori; Miyazaki, Hideo; Nakamura, Kazutoshi; Yoshie, Hiromasa

2016-03-01

Several studies have reported inconsistent results regarding the association between the PPARγPro12Ala polymorphism and obesity. Obese individuals had higher C-reactive protein (CRP) levels compared with those of normal weight, and PPARγ activation could significantly reduce serum high-sensitive CRP level. We have previously suggested that the Pro12Ala polymorphism represents a susceptibility factor for periodontitis, which is a known risk factor for increased CRP level. The aim was to investigate associations between PPARγ gene polymorphism, serum CRP level, BMI and/or periodontitis among post-menopausal Japanese women. The final sample in this study comprised 359 post-menopausal Japanese women. Periodontal parameters, including PD, CAL and BOP, were measured per tooth. PPARγPro12Ala genotype was determined by PCR-RFLP. Hs-CRP value was measured by a latex nephelometry assay. No significant differences in age, BMI or periodontal parameters were found between the genotypes. The percentages of sites with PD ≥ 4 mm were significantly higher among the hsCRP ≥ 1 mg/l group than the hsCRP < 1 mg/l group (p = 0.003). Positive correlations were found between serum hsCRP levels and the percentages of sites with PD ≥ 4 mm (p = 0.043) in PPARγ Ala allele carriers, and BMI (p = 0.033) in non-carriers. After adjustment for model covariates, BMI was significantly associated with serum hsCRP level. The PPARγPro12Ala polymorphism was not independently associated with periodontitis, serum CRP level or BMI in post-menopausal Japanese women. However, serum hsCRP level correlated with periodontitis in Ala allele carriers, and with BMI in non-carriers. © 2014 John Wiley & Sons A/S and The Gerodontology Society. Published by John Wiley & Sons Ltd.

Chemopreventive effects of a curcumin-like diarylpentanoid [2,6-bis(2,5-dimethoxybenzylidene)cyclohexanone] in cellular targets of rheumatoid arthritis in vitro.

PubMed

Lee, Ka-Heng; Abas, Faridah; Mohamed Alitheen, Noorjahan Banu; Shaari, Khozirah; Lajis, Nordin Haji; Israf, Daud Ahmad; Syahida, Ahmad

2015-07-01

Synovial fibroblast has emerged as a potential cellular target in progressive joint destruction in rheumatoid arthritis development. In this study, BDMC33 (2,6-bis[2,5-dimethoxybenzylidene]cyclohexanone), a curcumin analogue with enhanced anti-inflammatory activity has been synthesized and the potency of BDMC33 on molecular and cellular basis of synovial fibroblasts (SF) were evaluated in vitro. Synovial fibroblast cells (HIG-82) were cultured in vitro and induced by phorbol-12-myristate acetate (PMA) to stimulate the expression of matrix metalloproteinase (MMPs) and pro-inflammatory cytokines. The protective effects of BDMC33 were evaluated toward MMP activities, pro-inflammatory cytokine expression and nuclear factor kappa-B (NF-κB) activation by using various bioassay methods, including zymography, Western blotting, reverse transcription polymerase chain reaction, immunofluorescense microscopy and electrophoretic mobility shift assay. The results showed that BDMC33 significantly inhibited the pro-gelatinase B (pro-MMP-9) and collagenase activities via suppression of MMP-1 in activated SF. In addition, BDMC33 strongly suppressed MMP-3 gene expression as well as inhibited COX-2 and IL-6 pro-inflammatory gene expression. We also demonstrated that BDMC33 abolished the p65 NF-κB nuclear translocation and NF-κB DNA binding activity in PMA-stimulated SF. BDMC33 represents an effective chemopreventive agent and could be used as a promising lead compound for further development of rheumatoid arthritis therapeutic intervention. © 2014 Asia Pacific League of Associations for Rheumatology and Wiley Publishing Asia Pty Ltd.

Explore the Features of Brain-Derived Neurotrophic Factor in Mood Disorders

PubMed Central

Yeh, Fan-Chi; Kao, Chung-Feng; Kuo, Po-Hsiu

2015-01-01

Objectives Brain-derived neurotrophic factor (BDNF) plays important roles in neuronal survival and differentiation; however, the effects of BDNF on mood disorders remain unclear. We investigated BDNF from the perspective of various aspects of systems biology, including its molecular evolution, genomic studies, protein functions, and pathway analysis. Methods We conducted analyses examining sequences, multiple alignments, phylogenetic trees and positive selection across 12 species and several human populations. We summarized the results of previous genomic and functional studies of pro-BDNF and mature-BDNF (m-BDNF) found in a literature review. We identified proteins that interact with BDNF and performed pathway-based analysis using large genome-wide association (GWA) datasets obtained for mood disorders. Results BDNF is encoded by a highly conserved gene. The chordate BDNF genes exhibit an average of 75% identity with the human gene, while vertebrate orthologues are 85.9%-100% identical to human BDNF. No signs of recent positive selection were found. Associations between BDNF and mood disorders were not significant in most of the genomic studies (e.g., linkage, association, gene expression, GWA), while relationships between serum/plasma BDNF level and mood disorders were consistently reported. Pro-BDNF is important in the response to stress; the literature review suggests the necessity of studying both pro- and m-BDNF with regard to mood disorders. In addition to conventional pathway analysis, we further considered proteins that interact with BDNF (I-Genes) and identified several biological pathways involved with BDNF or I-Genes to be significantly associated with mood disorders. Conclusions Systematically examining the features and biological pathways of BDNF may provide opportunities to deepen our understanding of the mechanisms underlying mood disorders. PMID:26091093

Extracellular HSP27 acts as a signaling molecule to activate NF-κB in macrophages.

PubMed

Salari, Samira; Seibert, Tara; Chen, Yong-Xiang; Hu, Tieqiang; Shi, Chunhua; Zhao, Xiaoling; Cuerrier, Charles M; Raizman, Joshua E; O'Brien, Edward R

2013-01-01

Heat shock protein 27 (HSP27) shows attenuated expression in human coronary arteries as the extent of atherosclerosis progresses. In mice, overexpression of HSP27 reduces atherogenesis, yet the precise mechanism(s) are incompletely understood. Inflammation plays a central role in atherogenesis, and of particular interest is the balance of pro- and anti-inflammatory factors produced by macrophages. As nuclear factor-kappa B (NF-κB) is a key immune signaling modulator in atherogenesis, and macrophages are known to secrete HSP27, we sought to determine if recombinant HSP27 (rHSP27) alters NF-κB signaling in macrophages. Treatment of THP-1 macrophages with rHSP27 resulted in the degradation of an inhibitor of NF-κB, IκBα, nuclear translocation of the NF-κB p65 subunit, and increased NF-κB transcriptional activity. Treatment of THP-1 macrophages with rHSP27 yielded increased expression of a variety of genes, including the pro-inflammatory factors, IL-1β, and TNF-α. However, rHSP27 also increased the expression of the anti-inflammatory factors IL-10 and GM-CSF both at the mRNA and protein levels. Our study suggests that in macrophages, activation of NF-κB signaling by rHSP27 is associated with upregulated expression and secretion of key pro- and anti-inflammatory cytokines. Moreover, we surmise that it is the balance in expression of these mediators and antagonists of inflammation, and hence atherogenesis, that yields a favorable net effect of HSP27 on the vessel wall.

Effects of Almond- and Olive Oil-Based Docosahexaenoic- and Vitamin E-Enriched Beverage Dietary Supplementation on Inflammation Associated to Exercise and Age

PubMed Central

Capó, Xavier; Martorell, Miquel; Sureda, Antoni; Riera, Joan; Drobnic, Franchek; Tur, Josep Antoni; Pons, Antoni

2016-01-01

n-3-polyunsaturated fatty acids and polyphenols are potential key factors for the treatment and prevention of chronic inflammation associated to ageing and non-communicable diseases. The aim was to analyse effects of an almond and olive oil beverage enriched with α-tocopherol and docosahexaenoic, exercise and age on inflammatory plasma markers, and immune gene expression in peripheral blood mononuclear cells (PBMCs). Five young and five senior athletes who were supplemented for five weeks with a functional beverage performed a stress test under controlled conditions before and after beverage supplementation. Blood samples were taken immediately before and 1 h after each test. Plasma, erythrocytes and PBMCs were isolated. Beverage supplementation increased plasmatic Tumour Necrosis Factor α (TNFα) levels depending on age and exercise. Exercise increased plasma non esterified fatty acids (NEFAs), soluble Intercellular adhesion molecule 3 (sICAM3) and soluble L-selectin (sL-Selectin), and this increase was attenuated by the supplementation. Exercise increased PGE2 plasma levels in supplemented young and in senior placebo athletes. Exercise increased NFkβ-activated levels in PBMCs, which are primed to a pro-inflammatory response increasing pro-inflammatory genes expression after the exercise mainly in the young group after the supplementation. The functional beverage supplementation to young athletes enhances a pro-inflammatory circulating environment in response to the exercise that was less evident in the senior group. PMID:27735833

Effects of Almond- and Olive Oil-Based Docosahexaenoic- and Vitamin E-Enriched Beverage Dietary Supplementation on Inflammation Associated to Exercise and Age.

PubMed

Capó, Xavier; Martorell, Miquel; Sureda, Antoni; Riera, Joan; Drobnic, Franchek; Tur, Josep Antoni; Pons, Antoni

2016-10-09

n -3-polyunsaturated fatty acids and polyphenols are potential key factors for the treatment and prevention of chronic inflammation associated to ageing and non-communicable diseases. The aim was to analyse effects of an almond and olive oil beverage enriched with α-tocopherol and docosahexaenoic, exercise and age on inflammatory plasma markers, and immune gene expression in peripheral blood mononuclear cells (PBMCs). Five young and five senior athletes who were supplemented for five weeks with a functional beverage performed a stress test under controlled conditions before and after beverage supplementation. Blood samples were taken immediately before and 1 h after each test. Plasma, erythrocytes and PBMCs were isolated. Beverage supplementation increased plasmatic Tumour Necrosis Factor α (TNFα) levels depending on age and exercise. Exercise increased plasma non esterified fatty acids (NEFAs), soluble Intercellular adhesion molecule 3 (sICAM3) and soluble L-selectin (sL-Selectin), and this increase was attenuated by the supplementation. Exercise increased PGE2 plasma levels in supplemented young and in senior placebo athletes. Exercise increased NFkβ-activated levels in PBMCs, which are primed to a pro-inflammatory response increasing pro-inflammatory genes expression after the exercise mainly in the young group after the supplementation. The functional beverage supplementation to young athletes enhances a pro-inflammatory circulating environment in response to the exercise that was less evident in the senior group.

Gene-gene interaction between PPAR gamma 2 and ADR beta 3 increases obesity risk in children and adolescents.

PubMed

Ochoa, M C; Marti, A; Azcona, C; Chueca, M; Oyarzábal, M; Pelach, R; Patiño, A; Moreno-Aliaga, M J; Martínez-González, M A; Martínez, J A

2004-11-01

Multiple genes are likely to be involved in obesity and these genes may interact with environmental factors to influence obesity risk. Our aim was to explore the synergistic contribution of the two polymorphisms: Pro12Ala of the PPAR gamma 2 gene and Trp64Arg of the ADR beta 3 gene to obesity risk in a Spanish children and adolescent population. We designed a sex- and age-matched case-control study. Participants were 185 obese and 185 control children (aged 5-18 y) from the Navarra region, recruited through Departments of Pediatrics (Hospital Virgen del Camino, Navarra University Clinic and several Primary Health Centers). The obesity criterion (case definition) was BMI above the 97th percentile according to Spanish BMI reference data for age and gender. Anthropometric parameters were measured by standard protocols. The genotype was assessed by PCR-RFLP after digestion with BstUI for PPAR gamma 2 mutation and BstNI for ADR beta 3 variants. Face-to-face interviews were conducted to assess the physical activity. Using a validated physical activity questionnaire, we computed an activity metabolic equivalent index (METs h/week), which represents the physical exercise during the week for each participant. Statistical analysis was performed by conditional logistic regression, taking into account the matching between cases and controls. Carriers of the polymorphism Pro12Ala of the PPAR gamma 2 gene had a significantly higher obesity risk than noncarriers (odds ratio (OR)=2.18, 95% CI=1.09-4.36) when we adjusted for sex, age and physical activity. Moreover, the risk of obesity was higher (OR=2.59, 95% CI=1.17-5.34) when family history of obesity was also taken into account in the model. The OR for obesity linked to both polymorphisms (PPAR gamma 2 and ADR beta 3) was 5.30 (95% CI=1.08-25.97) when we adjusted for sex, age and physical activity. After adjustment for family history of obesity, the OR for carriers of both polymorphisms was 19.5 (95% CI=2.43-146.8). A synergistic effect between polymorphism Pro12Ala of the PPAR gamma 2 gene and Trp64Arg of the ADR beta 3 gene for obesity risk was found in a case-control study including children and adolescents.

Fatigue and gene expression in human leukocytes: increased NF-κB and decreased glucocorticoid signaling in breast cancer survivors with persistent fatigue.

PubMed

Bower, Julienne E; Ganz, Patricia A; Irwin, Michael R; Arevalo, Jesusa M G; Cole, Steve W

2011-01-01

Fatigue is highly prevalent in the general population and is one of the most common side effects of cancer treatment. There is growing evidence that pro-inflammatory cytokines play a role in cancer-related fatigue, although the molecular mechanisms for chronic inflammation and fatigue have not been determined. The current study utilized genome-wide expression microarrays to identify differences in gene expression and associated alterations in transcriptional activity in leukocytes from breast cancer survivors with persistent fatigue (n=11) and non-fatigued controls (n=10). We focused on transcription of inflammation-related genes, particularly those responsive to the pro-inflammatory NF-κB transcription control pathway. Further, given the role of glucocorticoids as key regulators of inflammatory processes, we examined transcription of glucocorticoid-responsive genes indicative of potential glucocorticoid receptor (GR) desensitization. Plasma levels of cortisol were also assessed. Consistent with hypotheses, results showed increased expression of transcripts with response elements for NF-κB, and reduced expression of transcripts with response elements for glucocorticoids (p<.05) in fatigued breast cancer survivors. No differences in plasma levels of cortisol were observed. These data indicate that increased activity of pro-inflammatory transcription factors may contribute to persistent cancer-related fatigue and provide insight into potential mechanisms for tonic increases in NF-κB activity, specifically decreased expression of GR anti-inflammatory transcription factors. Copyright © 2010 Elsevier Inc. All rights reserved.

Mutation spectrum of NDP, FZD4 and TSPAN12 genes in Indian patients with retinopathy of prematurity.

PubMed

Rathi, Sonika; Jalali, Subhadra; Musada, Ganeswara Rao; Patnaik, Satish; Balakrishnan, Divya; Hussain, Anjli; Kaur, Inderjeet

2018-02-01

Retinopathy of prematurity (ROP) is a vasoproliferative eye disease in preterm infants. Based on its phenotypic similarities with familial exudative vitreo retinopathy (FEVR), the present study was conducted to screen the Norrin signalling pathway genes (already been implicated in FEVR) for understanding their involvement among Indian patients with ROP. The study cohort consisted of patients with ROP (n=246) and controls (n=300) that included full term (n=110) and preterm babies devoid of ROP (n=190). Screening of the NDP, FZD4, TSPAN12 genes were accomplished by resequencing the entire coding and untranslated regions (UTR). The genotype data of the patients with ROP were analysed in the background of their clinical manifestations and further analysed in conjunction with other available data on these genes worldwide. Two novel variants in intron 1 (IVS1 +16A>G) and 3'UTR (c.5 22T>C) along with a previously reported change in the 5'UTR (c.395_409del14bp) were observed in the NDP gene in three patients with ROP. Screening of the FZD4 revealed four heterozygous variants, p.(Pro33Ser), p.(Pro168Ser), p.(Ile192Ile) and p.(Ile360Val), a compound heterozygous (p.(Pro33Ser)/p.(Pro168Ser)) and a 3'UTR (c*G>T) variants in the study cohort. Variants p.(Pro33Ser) and p.(Pro168Ser) were found to be significantly associated with ROP. A heterozygous variant p.(Leu119Arg) in TSPAN12 gene was observed in a patient with threshold ROP. However, a formal genotype-phenotype correlation could not be established due to the low frequencies of the variant alleles in these genes. This is a first study that revealed association of few variants in Norrin signalling genes among Indian patients with ROP that warrants further detailed investigation worldwide. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Purinergic receptor ligands stimulate pro-opiomelanocortin gene expression in AtT-20 pituitary corticotroph cells.

PubMed

Zhao, L-F; Iwasaki, Y; Oki, Y; Tsugita, M; Taguchi, T; Nishiyama, M; Takao, T; Kambayashi, M; Hashimoto, K

2006-04-01

Although recent studies have suggested that purinergic receptors are expressed in the anterior pituitary gland, their involvement in the regulation of pituitary hormone gene expression is not completely understood. In the present study, we examined the expression of purinergic receptors and the effects of purinergic receptor ligands on pro-opiomelanocortin (POMC) gene expression, in AtT20 mouse corticotroph cells. We identified the expression of most of the purinergic receptor subtypes (A1, A2, P2X1, 3-7, P2Y1, 2, 4) mRNAs, analysed by the reverse transcriptase-polymerase chain reaction. We also found that adenosine and ATP, two representative and endogenous agonists of A1-3 and P2X/P2Y receptors, respectively, stimulated the 5'-promoter activity of the POMC gene in a dose- and time-related manner. When these ligands were simultaneously used with corticotrophin-releasing hormone (CRH), effects that were more than additive were observed, suggesting an enhancing role of these compounds in CRH-mediated adrenocorticotrophic hormone (ACTH) synthesis. These ligands also stimulated the expression of transcription factors involved in the regulation of the POMC gene, but did not enhance ACTH secretion. Finally, the positive effect of adenosine as well as CRH was completely inhibited by the protein kinase A inhibitor H89, whereas that of ATP was not influenced, indicating that different intracellular signalling pathways mediate these effects. Altogether, our results suggest a stimulatory role for these purinergic receptor ligands in the regulation of POMC gene expression in corticotroph cells. Because adenosine and ATP are known to be produced within the pituitary gland, it is possible they may be acting in an autocrine/paracrine fashion.

The AP-1 transcription factor FOSL1 causes melanocyte reprogramming and transformation.

PubMed

Maurus, K; Hufnagel, A; Geiger, F; Graf, S; Berking, C; Heinemann, A; Paschen, A; Kneitz, S; Stigloher, C; Geissinger, E; Otto, C; Bosserhoff, A; Schartl, M; Meierjohann, S

2017-09-07

The MAPK pathway is activated in the majority of melanomas and is the target of therapeutic approaches. Under normal conditions, it initiates the so-called immediate early response, which encompasses the transient transcription of several genes belonging to the AP-1 transcription factor family. Under pathological conditions, such as continuous MAPK pathway overactivation due to oncogenic alterations occurring in melanoma, these genes are constitutively expressed. The consequences of a permanent expression of these genes are largely unknown. Here, we show that FOSL1 is the main immediate early AP-1 member induced by melanoma oncogenes. We first examined its role in established melanoma cells. We found that FOSL1 is involved in melanoma cell migration as well as cell proliferation and anoikis-independent growth, which is mediated by the gene product of its target gene HMGA1, encoding a multipotent chromatin modifier. As FOSL1 expression is increased in patient melanoma samples compared to nevi, we investigated the effect of enhanced FOSL1 expression on melanocytes. Intriguingly, we found that FOSL1 acts oncogenic and transforms melanocytes, enabling subcutaneous tumor growth in vivo. During the process of transformation, FOSL1 reprogrammed the melanocytes and downregulated MITF in a HMGA1-dependent manner. At the same time, AXL was upregulated, leading to a shift in the MITF/AXL balance. Furthermore, FOSL1 re-enforced pro-tumorigenic transcription factors MYC, E2F3 and AP-1. Together, this led to the enhancement of several growth-promoting processes, such as ribosome biogenesis, cellular detachment and pyrimidine metabolism. Overall, we demonstrate that FOSL1 is a novel reprogramming factor for melanocytes with potent tumor transformation potential.

PPARγ2Pro12Ala Polymorphism and Human Health

PubMed Central

He, Weimin

2009-01-01

The nuclear hormone receptor peroxisome proliferator activated receptor gamma (PPARγ) is an important transcription factor regulating adipocyte differentiation, lipid and glucose homeostasis, and insulin sensitivity. Numerous genetic mutations of PPARγ have been identified and these mutations positively or negatively regulate insulin sensitivity. Among these, a relatively common polymorphism of PPARγ, Pro12Ala of PPARγ2, the isoform expressed only in adipose tissue has been shown to be associated with lower body mass index, enhanced insulin sensitivity, and resistance to the risk of type 2 diabetes in human subjects carrying this mutation. Subsequent studies in different ethnic populations, however, have revealed conflicting results, suggesting a complex interaction between the PPARγ2 Pro12Ala polymorphism and environmental factors such as the ratio of dietary unsaturated fatty acids to saturated fatty acids and/or between the PPARγ2 Pro12Ala polymorphism and genetic factors such as polymorphic mutations in other genes. In addition, this polymorphic mutation in PPARγ2 is associated with other aspects of human diseases, including cancers, polycystic ovary syndrome, Alzheimer disease and aging. This review will highlight findings from recent studies. PMID:19390629

The role of FGF2 in migration and tubulogenesis of endothelial progenitor cells in relation to pro-angiogenic growth factor production.

PubMed

Litwin, Monika; Radwańska, Agata; Paprocka, Maria; Kieda, Claudine; Dobosz, Tadeusz; Witkiewicz, Wojciech; Baczyńska, Dagmara

2015-12-01

In recent years, special attention has been paid to finding new pro-angiogenic factors which could be used in gene therapy of vascular diseases such as critical limb ischaemia (CLI). Angiogenesis, the formation of new blood vessels, is a complex process dependent on different cytokines, matrix proteins, growth factors and other pro- or anti-angiogenic stimuli. Numerous lines of evidence suggest that key mediators of angiogenesis, vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) together with fibroblast growth factor2 (FGF2) are involved in regulation of the normal and pathological process of angiogenesis. However, less information is available on the complex interactions between these and other angiogenic factors. The aim of this study was to characterise the effect of fibroblast growth factor2 on biological properties of human endothelial progenitor cells with respect to the expression level of other regulatory cytokines. Ectopic expression of FGF2 in EP cells stimulates their pro-angiogenic behaviour, leading to increased proliferation, migration and tube formation abilities. Moreover, we show that the expression profile of VEGF and other pro-angiogenic cytokines, such as HGF, MCP2, and interleukins, is affected differently by FGF2 in EPC. In conclusion, we provide evidence that FGF2 directly affects not only the biological properties of EP cells but also the expression pattern and secretion of numerous chemocytokines. Our results suggest that FGF2 could be applied in therapeutic approaches for CLI and other ischaemic diseases of the vascular system in vivo.

Novel mutation identified in severe early-onset tumor necrosis factor receptor-associated periodic syndrome: a case report.

PubMed

Radhakrishna, Suhas M; Grimm, Amy; Broderick, Lori

2017-04-20

Tumor Necrosis Factor Receptor-Associated Periodic Syndrome (TRAPS) is the second most common heritable autoinflammatory disease, typically presenting in pre-school aged children with fever episodes lasting 1-3 weeks. Systemic symptoms can include rash, myalgia, ocular inflammation, and serositis. Here we report an unusual presentation of TRAPS in a 7 month old girl who presented with only persistent fever. She was initially diagnosed with incomplete Kawasaki Disease and received IVIG and infliximab; however, her fevers quickly recurred. Subsequent testing revealed a urinary tract infection, but she did not improve despite appropriate therapy. As fever continued, she developed significant abdominal distension with imaging concerning for appendicitis, followed by hyperthermia and hemodynamic instability. Given her protracted clinical course and maternal history of a poorly defined inflammatory condition, an autoinflammatory disease was considered. Therapy with anakinra was initiated, resulting in rapid resolution of fever and normalization of inflammatory markers. She was found to have a previously unreported mutation, Thr90Pro, in the TNFRSF1A gene associated with TRAPS. This novel mutation was also confirmed in the patient's mother and maternal uncle. This report reviews a severe case of TRAPS in infancy associated with a novel mutation, Thr90Pro, in the TNFRSF1A gene, and emphasizes that autoinflammatory disease should be considered in the differential of infants with fever of unknown origin.

Joint acute and endocrine disruptive toxicities of malathion, cypermethrin and prochloraz to embryo-larval zebrafish, Danio rerio.

PubMed

Guo, Dongmei; Wang, Yanhua; Qian, Yongzhong; Chen, Chen; Jiao, Bining; Cai, Leiming; Wang, Qiang

2017-01-01

It remains a daunting challenge to determine ecotoxicological risks of exposure to mixtures of endocrine disrupting chemicals (EDCs) in environmental toxicology. In the present study, we investigated acute and endocrine disruptive toxicities of cypermethrin (CPM), malathion (MAL), prochloraz (PRO) and their binary mixtures of MAL + CPM and MAL + PRO to the early life stages of zebrafish. In the acute lethal toxicity test, three pesticides exhibited different levels of toxicity to zebrafish larvae, and the order of toxicity was as follows: CPM > PRO > MAL. The binary mixture of MAL + CPM displayed a synergistic effect on zebrafish larvae after exposure for 24, 48, 72 and 96 h. However, binary mixture of MAL + PRO showed an antagonistic effect. To evaluate the estrogenic effect, the expression of genes in the hypothalamic-pituitary-gonadal axis was assessed after zebrafish embryos were exposed to CPM, MAL, PRO and their binary mixtures from blastula stage (1 h post-fertilization, 1 hpf) to 14 dpf (14 d post-fertilization). Our data indicated that the transcription patterns of many key genes (vtg1, vtg2, era, erβ1, erβ2, cyp19a1a and cyp19a1b) were affected in hatched zebrafish after exposure to CPM, MAL and PRO. Moreover, following exposure to binary mixtures of 1000 μg/L MAL +4 μg/L CPM and 1000 μg/L MAL +900 μg/L PRO, the gene expressions were significantly changed compared with the individual pesticides. Our data provided a better understanding of bidirectional interactions of toxic response induced by these pesticides. Copyright © 2016 Elsevier Ltd. All rights reserved.

MYCN controls an alternative RNA splicing program in high-risk metastatic neuroblastoma

PubMed Central

Zhang, Shile; Wei, Jun S.; Li, Samuel Q.; Badgett, Tom C.; Song, Young K.; Agarwal, Saurabh; Coarfa, Cristian; Tolman, Catherine; Hurd, Laura; Liao, Hongling; He, Jianbin; Wen, Xinyu; Liu, Zhihui; Thiele, Carol J.; Westermann, Frank; Asgharzadeh, Shahab; Seeger, Robert C.; Maris, John M.; Auvil, Jamie M Guidry; Smith, Malcolm A; Kolaczyk, Eric D; Shohet, Jason; Khan, Javed

2016-01-01

The molecular mechanisms underlying the aggressive behavior of MYCN driven neuroblastoma (NBL) is under intense investigation; however, little is known about the impact of this family of transcription factors on the splicing program. Here we used high-throughput RNA sequencing to systematically study the expression of RNA isoforms in stage 4 MYCN-amplified NBL, an aggressive subtype of metastatic NBL. We show that MYCN-amplified NBL tumors display a distinct gene splicing pattern affecting multiple cancer hallmark functions. Six splicing factors displayed unique differential expression patterns in MYCN-amplified tumors and cell lines, and the binding motifs for some of these splicing factors are significantly enriched in differentially-spliced genes. Direct binding of MYCN to promoter regions of the splicing factors PTBP1 and HNRNPA1 detected by ChIP-seq demonstrates MYCN controls the splicing pattern by direct regulation of the expression of these key splicing factors. Furthermore, high expression of PTBP1 and HNRNPA1 was significantly associated with poor overall survival of stage4 NBL patients (p≤0.05). Knocking down PTBP1, HNRNPA1 and their downstream target PKM2, an isoform of pro-tumor-growth, result in repressed growth of NBL cells. Therefore, our study reveals a novel role of MYCN in controlling global splicing program through regulation of splicing factors in addition to its well-known role in the transcription program. These findings suggest a therapeutically potential to target the key splicing factors or gene isoforms in high-risk NBL with MYCN-amplification. PMID:26683771

Bone Morphogenetic Protein 3 Controls Insulin Gene Expression and Is Down-regulated in INS-1 Cells Inducibly Expressing a Hepatocyte Nuclear Factor 1A–Maturity-onset Diabetes of the Young Mutation*

PubMed Central

Bonner, Caroline; Farrelly, Angela M.; Concannon, Caoimhín G.; Dussmann, Heiko; Baquié, Mathurin; Virard, Isabelle; Wobser, Hella; Kögel, Donat; Wollheim, Claes B.; Rupnik, Marjan; Byrne, Maria M.; König, Hans-Georg; Prehn, Jochen H. M.

2011-01-01

Inactivating mutations in the transcription factor hepatocyte nuclear factor (HNF) 1A cause HNF1A–maturity-onset diabetes of the young (HNF1A-MODY), the most common monogenic form of diabetes. To examine HNF1A-MODY-induced defects in gene expression, we performed a microarray analysis of the transcriptome of rat INS-1 cells inducibly expressing the common hot spot HNF1A frameshift mutation, Pro291fsinsC-HNF1A. Real-time quantitative PCR (qPCR), Western blotting, immunohistochemistry, reporter assays, and chromatin immunoprecipitation (ChIP) were used to validate alterations in gene expression and to explore biological activities of target genes. Twenty-four hours after induction of the mutant HNF1A protein, we identified a prominent down-regulation of the bone morphogenetic protein 3 gene (Bmp-3) mRNA expression. Reporter assays, qPCR, and Western blot analysis validated these results. In contrast, inducible expression of wild-type HNF1A led to a time-dependent increase in Bmp-3 mRNA and protein levels. Moreover, reduced protein levels of BMP-3 and insulin were detected in islets of transgenic HNF1A-MODY mice. Interestingly, treatment of naïve INS-1 cells or murine organotypic islet cultures with recombinant human BMP-3 potently increased their insulin levels and restored the decrease in SMAD2 phosphorylation and insulin gene expression induced by the HNF1A frameshift mutation. Our study suggests a critical link between HNF1A-MODY-induced alterations in Bmp-3 expression and insulin gene levels in INS-1 cells and indicates that the reduced expression of growth factors involved in tissue differentiation may play an important role in the pathophysiology of HNF1A-MODY. PMID:21628466

Platelet lysate-based pro-angiogenic nanocoatings.

PubMed

Oliveira, Sara M; Pirraco, Rogério P; Marques, Alexandra P; Santo, Vítor E; Gomes, Manuela E; Reis, Rui L; Mano, João F

2016-03-01

Human platelet lysate (PL) is a cost-effective and human source of autologous multiple and potent pro-angiogenic factors, such as vascular endothelial growth factor A (VEGF A), fibroblast growth factor b (FGF b) and angiopoietin-1. Nanocoatings previously characterized were prepared by layer-by-layer assembling incorporating PL with marine-origin polysaccharides and were shown to activate human umbilical vein endothelial cells (HUVECs). Within 20 h of incubation, the more sulfated coatings induced the HUVECS to the form tube-like structures accompanied by an increased expression of angiogenic-associated genes, such as angiopoietin-1 and VEGF A. This may be a cost-effective approach to modify 2D/3D constructs to instruct angiogenic cells towards the formation of neo-vascularization, driven by multiple and synergistic stimulations from the PL combined with sulfated polysaccharides. The presence, or fast induction, of a stable and mature vasculature inside 3D constructs is crucial for new tissue formation and its viability. This has been one of the major tissue engineering challenges, limiting the dimensions of efficient tissue constructs. Many approaches based on cells, growth factors, 3D bioprinting and channel incorporation have been proposed. Herein, we explored a versatile technique, layer-by-layer assembling in combination with platelet lysate (PL), that is a cost-effective source of many potent pro-angiogenic proteins and growth factors. Results suggest that the combination of PL with sulfated polyelectrolytes might be used to introduce interfaces onto 2D/3D constructs with potential to induce the formation of cell-based tubular structures. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A novel role for GSK3 in the regulation of the processes of human labour.

PubMed

Lim, Ratana; Lappas, Martha

2015-02-01

Preterm birth remains the largest single cause of neonatal death and morbidity. Infection and/or inflammation are strongly associated with preterm delivery. Glycogen synthase kinase 3 (GSK3) is known to be a crucial mediator of inflammation homeostasis. The aims of this study were to determine the effect of spontaneous human labour in foetal membranes and myometrium on GSK3α/β expression, and the effect of inhibition of GSK3α/β on pro-labour mediators in foetal membranes and myometrium stimulated with Toll-like receptor (TLR) ligands and pro-inflammatory cytokines. Term and preterm labour in foetal membranes was associated with significantly decreased serine phosphorylated GSK3α and β expression, and thus increased GSK3 activity. There was no effect of term labour on serine phosphorylated GSK3β expression in myometrium. The specific GSK3α/β inhibitor CHIR99021 significantly decreased lipopolysaccharide (ligand to TLR4)-stimulated pro-inflammatory cytokine gene expression and release; COX2 gene expression and prostaglandin release; and MMP9 gene expression and pro MMP9 release in foetal membranes and/or myometrium. CHIR99021 also decreased FSL1 (TLR2 ligand) and flagellin (TLR5 ligand)-induced pro-inflammatory cytokine gene expression and release and COX2 mRNA expression and prostaglandin release. GSK3β siRNA knockdown in primary myometrial cells was associated with a significant decrease in IL1β and TNFα-induced pro-inflammatory cytokine and prostaglandin release. In conclusion, GSK3α/β activity is increased in foetal membranes after term and preterm labour. Pharmacological blockade of the kinase GSK3 markedly reduced pro-inflammatory and pro-labour mediators in human foetal membranes and myometrium, providing a possible therapeutics for the management of preterm labour. © 2015 Society for Reproduction and Fertility.

Acute stress alters transcript expression pattern and reduces processing of proBDNF to mature BDNF in Dicentrarchus labrax

PubMed Central

2010-01-01

Background Stress involves alterations of brain functioning that may precipitate to mood disorders. The neurotrophin Brain Derived Neurotrophic Factor (BDNF) has recently been involved in stress-induced adaptation. BDNF is a key regulator of neuronal plasticity and adaptive processes. Regulation of BDNF is complex and may reflect not only stress-specific mechanisms but also hormonal and emotional responses. For this reason we used, as an animal model of stress, a fish whose brain organization is very similar to that of higher vertebrates, but is generally considered free of emotional reactions. Results We provide a comprehensive characterization of BDNF gene in the Dicentrarchus labrax and its transcriptional, translational and post-translational regulation following acute stress. While total BDNF mRNA levels are unchanged, BDNF transcripts 1c and 1d resulted down regulated after acute stress. Acute stress induces also a significant increase in proBDNF levels and reduction in mature BDNF suggesting altered regulation of proBDNF proteolytic processing. Notably, we provide here the first evidence that fishes possess a simplified proteolytic regulation of BDNF since the pro28Kda form, generated by the SKI-1 protease in mammals, is absent in fishes because the cleavage site has first emerged in reptilians. Finally, we show that the proBDNF/totBDNF ratio is a highly predictive novel quantitative biomarker to detect stress in fishes with sensitivity = 100%, specificity = 87%, and Negative Predictive Value = 100%. Conclusion The high predictivity of proBDNF/totBDNF ratio for stress in lower vertebrates indicates that processing of BDNF is a central mechanism in adaptation to stress and predicts that a similar regulation of pro/mature BDNF has likely been conserved throughout evolution of vertebrates from fish to man. PMID:20074340

Acute stress alters transcript expression pattern and reduces processing of proBDNF to mature BDNF in Dicentrarchus labrax.

PubMed

Tognoli, Chiara; Rossi, Federica; Di Cola, Francesco; Baj, Gabriele; Tongiorgi, Enrico; Terova, Genciana; Saroglia, Marco; Bernardini, Giovanni; Gornati, Rosalba

2010-01-14

Stress involves alterations of brain functioning that may precipitate to mood disorders. The neurotrophin Brain Derived Neurotrophic Factor (BDNF) has recently been involved in stress-induced adaptation. BDNF is a key regulator of neuronal plasticity and adaptive processes. Regulation of BDNF is complex and may reflect not only stress-specific mechanisms but also hormonal and emotional responses. For this reason we used, as an animal model of stress, a fish whose brain organization is very similar to that of higher vertebrates, but is generally considered free of emotional reactions. We provide a comprehensive characterization of BDNF gene in the Dicentrarchus labrax and its transcriptional, translational and post-translational regulation following acute stress. While total BDNF mRNA levels are unchanged, BDNF transcripts 1c and 1d resulted down regulated after acute stress. Acute stress induces also a significant increase in proBDNF levels and reduction in mature BDNF suggesting altered regulation of proBDNF proteolytic processing. Notably, we provide here the first evidence that fishes possess a simplified proteolytic regulation of BDNF since the pro28Kda form, generated by the SKI-1 protease in mammals, is absent in fishes because the cleavage site has first emerged in reptilians. Finally, we show that the proBDNF/totBDNF ratio is a highly predictive novel quantitative biomarker to detect stress in fishes with sensitivity = 100%, specificity = 87%, and Negative Predictive Value = 100%. The high predictivity of proBDNF/totBDNF ratio for stress in lower vertebrates indicates that processing of BDNF is a central mechanism in adaptation to stress and predicts that a similar regulation of pro/mature BDNF has likely been conserved throughout evolution of vertebrates from fish to man.

Association of cytokine gene polymorphisms in CWP and its severity in Turkish coal workers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ates, I.; Suzen, H.S.; Yucesoy, B.

2008-10-15

Cytokines appear to play a key role in some inflammatory reactions affecting the interactions among pro- and anti-inflammatory mechanisms that result in several diseases such as coal workers' pneumoconiosis (CWP). In this study, to determine the cytokine gene profiles of Turkish coal miners, we performed genotyping analysis to investigate the polymorphisms of CWP-related pro-inflammatory (TNFA, IL1A, IL1B, and IL6) and anti-inflammatory cytokines (IL-1RN and TGFB1). Genotyping was carried out by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. TNFA (-238) gene polymorphism principally affected CWP development and severity (OR=3.47: 95% CI, 1.12-10.77 and OR=4.30: 95% CI, 1.25-14.74, respectively) and alsomore » risk of CWP (OR=3.79: 95% CI, 1.37-10.46). The TNFA (-308) variant was associated with a risk for the CWP severity (OR = 2.84: 95% CI, 1.08-7.39). A protective effect of IL6 was found on the development (OR = 0.48: 95% CI, 0.21-0.93) and severity of CWP (OR = 0.37: 95% CI, 0.15-0.91). We suggest that TNFA (-238) variant may be a risk factor in both development and the severity, of CWP while TNFA (-308) variant seems to be important only in disease severity On the other hand, IL6 variant may have a protective effect on the development and disease severity.« less

Development of a bioassay to screen for chemicals mimicking the anti-aging effects of calorie restriction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chiba, Takuya, E-mail: takuya@nagasaki-u.ac.jp; Tsuchiya, Tomoshi; Komatsu, Toshimitsu

2010-10-15

Research highlights: {yields} We identified four sequence motifs lying upstream of putative pro-longevity genes. {yields} One of these motifs binds to HNF-4{alpha}. {yields} HNF-4{alpha}/PGC-1{alpha} could up-regulate the transcription of a reporter gene linked to this motif. {yields} The reporter system described here could be used to screen candidate anti-aging molecules. -- Abstract: Suppression of the growth hormone/insulin-like growth factor-I pathway in Ames dwarf (DF) mice, and caloric restriction (CR) in normal mice extends lifespan and delays the onset of age-related disorders. In combination, these interventions have an additive effect on lifespan in Ames DF mice. Therefore, common signaling pathways regulatedmore » by DF and CR could have additive effects on longevity. In this study, we tried to identity the signaling mechanism and develop a system to assess pro-longevity status in cells and mice. We previously identified genes up-regulated in the liver of DF and CR mice by DNA microarray analysis. Motif analysis of the upstream sequences of those genes revealed four major consensus sequence motifs, which have been named dwarfism and calorie restriction-responsive elements (DFCR-REs). One of the synthesized sequences bound to hepatocyte nuclear factor-4{alpha} (HNF-4{alpha}), an important transcription factor involved in liver metabolism. Furthermore, using this sequence information, we developed a highly sensitive bioassay to identify chemicals mimicking the anti-aging effects of CR. When the reporter construct, containing an element upstream of a secreted alkaline phosphatase (SEAP) gene, was co-transfected with HNF-4{alpha} and its regulator peroxisome proliferator-activated receptor (PPAR) {gamma} coactivator-1{alpha} (PGC-1{alpha}), SEAP activity was increased compared with untransfected controls. Moreover, transient transgenic mice established using this construct showed increased SEAP activity in CR mice compared with ad libitum-fed mice. These data suggest that because of its rapidity, ease of use, and specificity, our bioassay will be more useful than the systems currently employed to screen for CR mimetics, which mimic the beneficial effects of CR. Our system will be particularly useful for high-throughput screening of natural and synthetic candidate molecules.« less

Molecular cloning and expression analysis of the gene encoding proline dehydrogenase from Jatropha curcas L.

PubMed

Wang, Haibo; Ao, Pingxing; Yang, Shuanglong; Zou, Zhurong; Wang, Shasha; Gong, Ming

2015-03-01

Proline dehydrogenase (ProDH) (EC 1.5.99.8) is a key enzyme in the catabolism of proline. The enzyme JcProDH and its complementary DNA (cDNA) were isolated from Jatropha curcas L., an important woody oil plant used as a raw material for biodiesels. It has been classified as a member of the Pro_dh superfamily based on multiple sequence alignment, phylogenetic characterization, and its role in proline catabolism. Its cDNA is 1674 bp in length with a complete open reading frame of 1485 bp, which encodes a polypeptide chain of 494 amino acids with a predicted molecular mass of 54 kD and a pI of 8.27. Phylogenetic analysis indicated that JcProDH showed high similarity with ProDH from other plants. Reverse transcription PCR (RT-PCR) analysis revealed that JcProDH was especially abundant in the seeds and flowers but scarcely present in the stems, roots, and leaves. In addition, the expression of JcProDH increased in leaves experiencing environmental stress such as cold (5 °C), heat (42 °C), salt (300 mM), and drought (30 % PEG6000). The JcProDH protein was successfully expressed in the yeast strain INVSc1 and showed high enzyme activity in proline catabolism. This result confirmed that the JcProDH gene negatively participated in the stress response.

Human native lipoprotein-induced de novo DNA methylation is associated with repression of inflammatory genes in THP-1 macrophages.

PubMed

Rangel-Salazar, Rubén; Wickström-Lindholm, Marie; Aguilar-Salinas, Carlos A; Alvarado-Caudillo, Yolanda; Døssing, Kristina B V; Esteller, Manel; Labourier, Emmanuel; Lund, Gertrud; Nielsen, Finn C; Rodríguez-Ríos, Dalia; Solís-Martínez, Martha O; Wrobel, Katarzyna; Wrobel, Kazimierz; Zaina, Silvio

2011-11-25

We previously showed that a VLDL- and LDL-rich mix of human native lipoproteins induces a set of repressive epigenetic marks, i.e. de novo DNA methylation, histone 4 hypoacetylation and histone 4 lysine 20 (H4K20) hypermethylation in THP-1 macrophages. Here, we: 1) ask what gene expression changes accompany these epigenetic responses; 2) test the involvement of candidate factors mediating the latter. We exploited genome expression arrays to identify target genes for lipoprotein-induced silencing, in addition to RNAi and expression studies to test the involvement of candidate mediating factors. The study was conducted in human THP-1 macrophages. Native lipoprotein-induced de novo DNA methylation was associated with a general repression of various critical genes for macrophage function, including pro-inflammatory genes. Lipoproteins showed differential effects on epigenetic marks, as de novo DNA methylation was induced by VLDL and to a lesser extent by LDL, but not by HDL, and VLDL induced H4K20 hypermethylation, while HDL caused H4 deacetylation. The analysis of candidate factors mediating VLDL-induced DNA hypermethylation revealed that this response was: 1) surprisingly, mediated exclusively by the canonical maintenance DNA methyltransferase DNMT1, and 2) independent of the Dicer/micro-RNA pathway. Our work provides novel insights into epigenetic gene regulation by native lipoproteins. Furthermore, we provide an example of DNMT1 acting as a de novo DNA methyltransferase independently of canonical de novo enzymes, and show proof of principle that de novo DNA methylation can occur independently of a functional Dicer/micro-RNA pathway in mammals.

Corrected and Republished from: BCL11A Is a Critical Component of a Transcriptional Network That Activates Recombinase Activating Gene Expression and V(D)J Recombination

PubMed Central

Lee, Baeck-Seung; Lee, Bum-Kyu; Iyer, Vishwanath R.; Sleckman, Barry P.; Shaffer, Arthur L.; Ippolito, Gregory C.

2017-01-01

ABSTRACT Recombination activating gene 1 (RAG1) and RAG2 are critical enzymes for initiating variable-diversity-joining [V(D)J] segment recombination, an essential process for antigen receptor expression and lymphocyte development. The BCL11A transcription factor is required for B cell and plasmacytoid dendritic cell (pDC) development, but its molecular function(s) in early B cell fate specification and commitment is unknown. We show here that the major B cell isoform, BCL11A-XL, binds directly to the RAG1 promoter as well as directly to regulatory regions of transcription factors previously implicated in both B cell and pDC development to activate RAG1 and RAG2 gene transcription in pro- and pre-B cells. We employed BCL11A overexpression with recombination substrates to demonstrate direct consequences of BCL11A/RAG modulation on V(D)J recombination. We conclude that BCL11A is a critical component of a transcriptional network that regulates B cell fate by controlling V(D)J recombination. PMID:29038163

Factors of transforming growth factor beta signalling are co-regulated in human hepatocellular carcinoma.

PubMed

Longerich, Thomas; Breuhahn, Kai; Odenthal, Margarete; Petmecky, Katharina; Schirmacher, Peter

2004-12-01

Transforming growth factor beta (TGFbeta) is a central mitoinhibitory factor for epithelial cells, and alterations of TGFbeta signalling have been demonstrated in many different human cancers. We have analysed human hepatocellular carcinomas (HCCs) for potential pro-tumourigenic alterations in regard to expression of Smad4 and mutations and expression changes of the pro-oncogenic transcriptional co-repressors Ski and SnoN, as well as mRNA levels of matrix metalloproteinase-2 (MMP2), which is transcriptionally regulated by TGFbeta. Smad4 mRNA was detected in all HCCs; while, using immunohistology, loss of Smad4 expression was found in 10% of HCCs. Neither mutations in the transformation-relevant sequences nor significant pro-tumourigenic expression changes of the Ski and SnoN genes were detected. In HCC cell lines, expression of both genes was regulated, potentially involving phosphorylation. Ski showed a distinct nuclear speckled pattern, indicating recruitment to active transcription complexes. MMP2 mRNA levels were increased in 19% of HCCs, whereas MMP2 mRNA was not detectable in HCC cell lines, suggesting that MMP2 was derived only from tumour stroma cells. Transcript levels of Smad4, Ski, SnoN and MMP2 correlated well. These data argue against a significant role of Ski and SnoN in human hepatocarcinogenesis and suggest that, in the majority of HCCs, the analysed factors are co-regulated by an upstream mechanism, potentially by TGFbeta itself.

Cytokine gene signatures in neural tissue of horses with equine protozoal myeloencephalitis or equine herpes type 1 myeloencephalopathy.

PubMed

Pusterla, N; Wilson, W D; Conrad, P A; Barr, B C; Ferraro, G L; Daft, B M; Leutenegger, C M

2006-09-09

This study was designed to determine the relative levels of gene transcription of selected pathogens and cytokines in the brain and spinal cord of 12 horses with equine protozoal myeloencephalitis (EPM), 11 with equine herpesvirus type 1 (EHV-1) myeloencephalopathy, and 12 healthy control horses by applying a real time pcr to the formalin-fixed and paraffin-embedded tissues. Total rna was extracted from each tissue, transcribed to complementary dna (cDNA) and assayed for Sarcocystis neurona, Neospora hughesi, EHV-1, equine GAPDH (housekeeping gene), tumour necrosis factor (TNF)-alpha, interferon (IFN)-gamma, interleukin (IL)-1beta, IL-2, IL-4, IL-6, IL-8, IL-10 AND IL-12 p40. S neurona cdna was detected in the neural tissue from all 12 horses with EPM, and two of them also had amplifiable cDNA of N hughesi. The relative levels of transcription of protozoal cdna ranged from 1 to 461 times baseline (mean 123). All the horses with ehv-1 myeloencephalopathy had positive viral signals by PCR with relative levels of transcription ranging from 1 to 1618 times baseline (mean 275). All the control horses tested negative for S neurona, N hughesi and EHV-1 cdna. The cytokine profiles of each disease indicated a balance between pro- and anti-inflammatory markers. In the horses with epm the pro-inflammatory Th1 cytokines (IL-8, TNF-alpha and IFN-gamma) were commonly expressed but the anti-inflammatory Th2 cytokines (IL-4, IL-6 AND IL-10) were absent or rare. In the horses with ehv-1 the proinflammatory cytokine IL-8 was commonly expressed, but IL-10 and IFN-gamma were not, and TNF-alpha was rare. Tissue from the control horses expressed only the gene GAPDH.

HNF1B variants associate with promoter methylation and regulate gene networks activated in prostate and ovarian cancer

PubMed Central

Ross-Adams, Helen; Ball, Stephen; Lawrenson, Kate; Halim, Silvia; Russell, Roslin; Wells, Claire; Strand, Siri H.; Ørntoft, Torben F.; Larson, Melissa; Armasu, Sebastian; Massie, Charles E.; Asim, Mohammad; Mortensen, Martin M.; Borre, Michael; Woodfine, Kathryn; Warren, Anne Y.; Lamb, Alastair D.; Kay, Jonathan; Whitaker, Hayley; Ramos-Montoya, Antonio; Murrell, Adele; Sørensen, Karina D.; Fridley, Brooke L.; Goode, Ellen L.; Gayther, Simon A.; Masters, John

2016-01-01

Two independent regions within HNF1B are consistently identified in prostate and ovarian cancer genome-wide association studies (GWAS); their functional roles are unclear. We link prostate cancer (PC) risk SNPs rs11649743 and rs3760511 with elevated HNF1B gene expression and allele-specific epigenetic silencing, and outline a mechanism by which common risk variants could effect functional changes that increase disease risk: functional assays suggest that HNF1B is a pro-differentiation factor that suppresses epithelial-to-mesenchymal transition (EMT) in unmethylated, healthy tissues. This tumor-suppressor activity is lost when HNF1B is silenced by promoter methylation in the progression to PC. Epigenetic inactivation of HNF1B in ovarian cancer also associates with known risk SNPs, with a similar impact on EMT. This represents one of the first comprehensive studies into the pleiotropic role of a GWAS-associated transcription factor across distinct cancer types, and is the first to describe a conserved role for a multi-cancer genetic risk factor. PMID:27732966

Airborne nitro-PAHs induce Nrf2/ARE defense system against oxidative stress and promote inflammatory process by activating PI3K/Akt pathway in A549 cells.

PubMed

Shang, Yu; Zhou, Qian; Wang, Tiantian; Jiang, Yuting; Zhong, Yufang; Qian, Guangren; Zhu, Tong; Qiu, Xinghua; An, Jing

2017-10-01

Ambient particulate matter (PM) is a worldwide health issue of concern. However, limited information is available regarding the toxic contributions of the nitro-derivatives of polycyclic aromatic hydrocarbons (nitro-PAHs). This study intend to examine whether 1-nitropyrene (1-NP) and 3-nitrofluoranthene (3-NF) could activate the nuclear factor-erythroid 2-related factor 2/antioxidant response element (Nrf2/ARE) antioxidant defense system, and whether the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway participates in regulating pro-inflammatory responses in A549 cells. Firstly, 1-NP and 3-NF concentration-dependently induced cellular apoptosis, reactive oxygen species (ROS) generation, DNA damage, S phase cell cycle arrest and differential expression of related cytokine genes. Secondly, 1-NP and 3-NF activated the Nrf2/ARE defense system, as evidenced by increased protein expression levels and nuclear translocation of transcription factor Nrf2, elevated Nrf2/ARE binding activity, up-regulated expression of the target gene heme oxygenase-1 (HO-1). Significantly increased protein expression of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and phosphorylation level of Akt indicated that the PI3K/Akt pathway was activated during pro-inflammatory process. Further, both PI3K inhibitor (LY294002) and Akt inhibitor (MK-2206) reversed the elevated TNF-α expression to control level. Our results suggested that Nrf2/ARE pathway activation might cause an initiation step in cellular protection against oxidative stress caused by nitro-PAHs, and the PI3K/Akt pathway participated in regulating inflammatory responses. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hydrolyzed fish proteins modulates both inflammatory and antioxidant gene expression as well as protein expression in a co culture model of liver and head kidney cells isolated from Atlantic salmon (Salmo salar).

PubMed

Holen, Elisabeth; He, Juyun; Araujo, Pedro; Seliussen, Jørgen; Espe, Marit

2016-07-01

Hydrolyzed fish proteins (H-pro) contain high concentrations of free amino acids and low molecular peptides that potentially may benefit fish health. The following study aimed to test whether the water-soluble phase of H-pro could attenuate lipopolysaccharide (LPS) provoked inflammation in liver cells and head kidney cells isolated from Atlantic salmon. Cells were grown as mono cultures or co cultures to assess possible crosstalk between immune cells and metabolic cells during treatments. Cells were added media with or without H-pro for 2 days before LPS exposure and harvested 24 h post LPS exposure. Respective cells without H-pro and LPS were used as controls. H-pro alone could affect expression of proteins directly as H-pro increased catalase protein expression in head kidney- and liver cells, regardless of culturing methods and LPS treatment. Leukotriene B4 (LTB4) production was also increased by H-pro in head kidney cells co cultured with liver cells. H-pro increased LPS induced interleukin 1β (IL-1β) transcription in liver cells co cultured with head kidney cells. All cultures of head kidney cells showed a significant increase in IL-1β transcription when treated with H-pro + LPS. H-pro decreased caspase-3 transcription in liver cells cultured co cultured with head kidney cells. Peroxisome proliferator activated receptor α (PPAR α) was upregulated, regardless of treatment, in liver cells co cultured with head kidney cells clearly showing that culturing method alone affected gene transcription. H-pro alone and together with LPS as an inflammation inducer, affect both antioxidant and inflammatory responses. Copyright © 2016 Elsevier Ltd. All rights reserved.

Interactive effects of inflammatory cytokine and abundant low-molecular-weight PAHs on inhibition of gap junctional intercellular communication, disruption of cell proliferation control, and the AhR-dependent transcription.

PubMed

Kabátková, Markéta; Svobodová, Jana; Pěnčíková, Kateřina; Mohatad, Dilshad Shaik; Šmerdová, Lenka; Kozubík, Alois; Machala, Miroslav; Vondráček, Jan

2015-01-05

Polycyclic aromatic hydrocarbons (PAHs) with lower molecular weight exhibit lesser genotoxicity and carcinogenicity than highly carcinogenic PAHs with a higher number of benzene rings. Nevertheless, they elicit specific effects linked with tumor promotion, such as acute inhibition of gap junctional intercellular communication (GJIC). Although inflammatory reaction may alter bioactivation and toxicity of carcinogenic PAHs, little is known about the impact of pro-inflammatory cytokines on toxic effects of the low-molecular-weight PAHs. Here, we investigated the impact of a pro-inflammatory cytokine, tumor necrosis factor-α (TNF-α), on the effects associated with tumor promotion and with induction of the aryl hydrocarbon receptor (AhR)-dependent gene expression in rat liver epithelial cells. We found that a prolonged incubation with TNF-α induced a down-regulation of GJIC, associated with reduced expression of connexin 43 (Cx43), a major connexin isoform found in liver epithelial cells. The Cx43 down-regulation was partly mediated by the activity of the mitogen-activated protein (MAP) p38 kinase. Independently of GJIC modulation, or p38 activation, TNF-α potentiated the AhR-dependent proliferative effect of a model low-molecular-weight PAH, fluoranthene, on contact-inhibited cells. In contrast, this pro-inflammatory cytokine repressed the fluoranthene-induced expression of a majority of model AhR gene targets, such as Cyp1a1, Ahrr or Tiparp. The results of the present study indicate that inflammatory reaction may differentially modulate various toxic effects of low-molecular-weight PAHs; the exposure to pro-inflammatory cytokines may both strengthen (inhibition of GJIC, disruption of contact inhibition) and repress (expression of a majority of AhR-dependent genes) their impact on toxic endpoints associated with carcinogenesis. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

The cAMP receptor protein CRP can function as an osmoregulator of transcription in Escherichia coli

PubMed Central

Landis, Lenore; Xu, Jimin; Johnson, Reid C.

1999-01-01

Transcription of the P1 promoter of the Escherichia coli proP gene, which encodes a transporter of osmoprotectants, is strongly induced by a shift to hyperosmotic media. Unlike most other osmotically regulated promoters, the induction occurs for a brief period of time, corresponding to the replacement of intracellular K+ glutamate with osmoprotecting compounds. This burst of proP transcription is correlated with the osmolarity-dependent binding of the cAMP receptor protein CRP to a site within the proP P1 promoter. We show that CRP–cAMP functions as an osmotically sensitive repressor of proP P1 transcription in vitro. Binding of CRP to the proP promoter in vivo is transiently destabilized after a hyperosmotic shift with kinetics that correspond to the derepression of transcription, whereas Fis and Lac repressor binding is not osmotically sensitive. Similar osmotic regulation of proP P1 transcription by the CRP* mutant implies that binding of cAMP is not responsible for the unusual osmotic sensitivity of CRP activity. Osmotic regulation of CRP activity is not limited to proP. Activation of the lac promoter by CRP is also transiently inhibited after an osmotic upshift, as is the binding of CRP to the galΔ4 P1 promoter. These findings suggest that CRP functions in certain contexts to regulate gene expression in response to osmotic changes, in addition to its role in catabolite control. PMID:10601034

The cAMP receptor protein CRP can function as an osmoregulator of transcription in Escherichia coli.

PubMed

Landis, L; Xu, J; Johnson, R C

1999-12-01

Transcription of the P1 promoter of the Escherichia coli proP gene, which encodes a transporter of osmoprotectants, is strongly induced by a shift to hyperosmotic media. Unlike most other osmotically regulated promoters, the induction occurs for a brief period of time, corresponding to the replacement of intracellular K(+) glutamate with osmoprotecting compounds. This burst of proP transcription is correlated with the osmolarity-dependent binding of the cAMP receptor protein CRP to a site within the proP P1 promoter. We show that CRP-cAMP functions as an osmotically sensitive repressor of proP P1 transcription in vitro. Binding of CRP to the proP promoter in vivo is transiently destabilized after a hyperosmotic shift with kinetics that correspond to the derepression of transcription, whereas Fis and Lac repressor binding is not osmotically sensitive. Similar osmotic regulation of proP P1 transcription by the CRP* mutant implies that binding of cAMP is not responsible for the unusual osmotic sensitivity of CRP activity. Osmotic regulation of CRP activity is not limited to proP. Activation of the lac promoter by CRP is also transiently inhibited after an osmotic upshift, as is the binding of CRP to the galdelta4P1 promoter. These findings suggest that CRP functions in certain contexts to regulate gene expression in response to osmotic changes, in addition to its role in catabolite control.

[Expression of plant antimicrobial peptide pro-SmAMP2 gene increases resistance of transgenic potato plants to Alternaria and Fusarium pathogens].

PubMed

Vetchinkina, E M; Komakhina, V V; Vysotskii, D A; Zaitsev, D V; Smirnov, A N; Babakov, A V; Komakhin, R A

2016-09-01

The chickweed (Stellaria media L.) pro-SmAMP2 gene encodes the hevein-like peptides that have in vitro antimicrobial activity against certain harmful microorganisms. These peptides play an important role in protecting the chickweed plants from infection, and the pro-SmAMP2 gene was previously used to protect transgenic tobacco and Arabidopsis plants from phytopathogens. In this study, the pro-SmAMP2 gene under control of viral CaMV35S promoter or under control of its own pro-SmAMP2 promoter was transformed into cultivated potato plants of two cultivars, differing in the resistance to Alternaria: Yubiley Zhukova (resistant) and Skoroplodny (susceptible). With the help of quantitative real-time PCR, it was demonstrated that transgenic potato plants expressed the pro-SmAMP2 gene under control of both promoters at the level comparable to or exceeding the level of the potato actin gene. Assessment of the immune status of the transformants demonstrated that expression of antimicrobial peptide pro-SmAMP2 gene was able to increase the resistance to a complex of Alternaria sp. and Fusarium sp. phytopathogens only in potato plants of the Yubiley Zhukova cultivar. The possible role of the pro-SmAMP2 products in protecting potatoes from Alternaria sp. and Fusarium sp. is discussed.

Osteosarcoma cells promote the production of pro-tumor cytokines in mesenchymal stem cells by inhibiting their osteogenic differentiation through the TGF-β/Smad2/3 pathway.

PubMed

Tu, Bing; Peng, Zhao-Xiang; Fan, Qi-Ming; Du, Lin; Yan, Wei; Tang, Ting-Ting

2014-01-01

Mesenchymal stem cells (MSCs) are among the most important components of the osteosarcoma microenvironment and are reported to promote tumor progression. However, the means by which osteosarcoma cells modulate MSC behavior remains unclear. The aim of this study was to determine the effects of osteosarcoma cells on both the production of pro-tumor cytokines by mesenchymal stem cells (MSCs) and the osteogenic differentiation of MSCs. High level of transforming growth factor-β (TGF-β) was detected in three osteosarcoma cell lines. Conditioned media (CM) from the osteosarcoma cell lines Saos-2 and U2-OS were used to stimulate the cultured MSCs. We found that osteosarcoma cells promoted the production of IL-6 and VEGF in MSCs by inhibiting their osteogenic differentiation. Furthermore, TGF-β in tumor CM was proved to be an important factor. The TGF-β neutralizing antibody antagonized the effects induced by osteosarcoma CM. The inhibition of Smad2/3 by siRNA significantly decreased the production of IL-6 and VEGF in MSCs and induced their osteogenic differentiation. We also found that Smad2/3 enhanced the expression of β-catenin in MSCs by decreasing the level of Dickkopf-1 (DKK1). Although the inhibition of β-catenin did not affect the production of IL-6 or VEGF, or the gene expression of the early osteogenic markers Runx2 and ALP, it did enhance the gene expression of osteocalcin. Taken together, our data indicate that osteosarcoma cells secrete TGF-β to maintain the stemness of MSCs and promote the production of pro-tumor cytokines by these cells. © 2013 Published by Elsevier Inc.

Mycobacterium avium subspecies induce differential expression of pro-inflammatory mediators in a murine macrophage model: evidence for enhanced pathogenicity of Mycobacterium avium subspecies paratuberculosis.

PubMed

Basler, Tina; Geffers, Robert; Weiss, Siegfried; Valentin-Weigand, Peter; Goethe, Ralph

2008-01-01

Mycobacterium avium subspecies (ssp.) paratuberculosis (MAP) is the etiological agent of paratuberculosis, a chronic, non-treatable granulomatous enteritis of ruminants. MAP is the only mycobacterium affecting the intestinal tract, which is of interest since it is presently the most favoured pathogen linked to Crohn's disease (CD) in humans due to its frequent detection in CD tissues. MAP is genetically closely related to other M. avium ssp. such as M. avium ssp. avium (MAA) and M. avium ssp. hominissuis (MAH) which can cause mycobacteriosis in animals and immunocompromised humans. We have recently shown that murine macrophage cell lines represent suitable systems to analyse M. avium ssp. patho-mechanisms and could show that MAP, but not MAA, specifically inhibited the antigen-specific stimulatory capacity for CD4(+) T-cells. In the present study, we compared gene expression profiles of murine RAW264.7 macrophages in response to infections with MAP or MAA using murine high-density oligonucleotide Affymetrix microarrays. A comparison of MAP and MAA infection revealed 17 differentially expressed genes. They were expressed at a much lower level in MAP-infected macrophages than in MAA-infected macrophages. Among these were the genes for IL-1beta, IL-1alpha, CXCL2, PTGS2 (COX2), lipocalin (LCN2) and TNF, which are important pro-inflammatory factors. The microarray data were confirmed for selected genes by quantitative real-time reverse transcription PCR and, by protein array analyses and ELISA. Similar to MAA, infection with MAH also showed robust induction of IL-1beta, CXCL2, COX2, LCN2 and TNF. Taken together, our results from M. avium ssp.-infected murine macrophages provide evidence that MAP in contrast to MAA and MAH specifically suppresses the pro-inflammatory defence mechanisms of infected macrophages.

Do gene polymorphism in IL-1β, TNF-α and IL-6 influence therapeutic response in patients with drug refractory epilepsy?

PubMed

Tiwari, Prabhakar; Dwivedi, Rekha; Mansoori, Nasim; Alam, Rizwan; Chauhan, Ugam Kumari; Tripathi, Manjari; Mukhopadhyay, Asok Kumar

2012-09-01

Pro-inflammatory cytokines may play an important pathophysiological role in patients with epilepsy. To understand the role of genes encoding pro-inflammatory cytokines in epilepsy, this study aimed to evaluate the polymorphisms of the promoter regions of IL-1β-511C>T (rs16944), TNF-α-308G>A (rs1800629) and IL-6-174G>C (rs1800795) genes and to look into the interaction between these genes in influencing seizure susceptibility, seizure frequency and response to therapy. The comparative frequency of polymorphism was determined in rs16944, rs1800629 and rs1800795 using PCR-RFLP in a group of 120 persons with epilepsy (PWE) and 110 ethnically matched healthy subjects of comparable age and sex in the North Indian population. Alleles and genotypes of rs16944, rs1800629 and rs1800795 were not found to influence the odds ratio of having susceptibility to epilepsy. Also gene-gene interaction of possible nine combinations of these genes did not show any positive association with epilepsy. The genotype and allelic frequency of rs1800795 showed a significant association (p<0.05) in seizure frequency (number of seizures/6-months) and drug refractory epilepsy. However, the genotype and allelic frequency of rs16944 and rs1800629 were not found to have such effect. This study demonstrates that the rs16944, rs1800629 and rs1800795 polymorphism does not act as a strong susceptibility factor for epilepsy in North Indian population. The genotypic association of rs1800795 with seizure frequency and drug-refractory epilepsy raises the issue that a specific set of polymorphic genes can influence seizures and therapeutic response in epilepsy. Copyright © 2012 Elsevier B.V. All rights reserved.

Pseudomonas aeruginosa infection alters the macrophage phenotype switching process during wound healing in diabetic mice.

PubMed

Chen, Sinuo; Li, Renren; Cheng, Chun; Xu, Jing-Ying; Jin, Caixia; Gao, Furong; Wang, Juan; Zhang, Jieping; Zhang, Jingfa; Wang, Hong; Lu, Lixia; Xu, Guo-Tong; Tian, Haibin

2018-03-07

Macrophages play critical roles in wound healing process. They switch from "classically activated" (M1) phenotype in the early inflammatory phase to "alternatively activated" (M2) phenotype in the later healing phase. However, the dynamic process of macrophage phenotype switching in diabetic wounds burdened with bacteria is unclear. In this report, Pseudomonas aeruginosa, frequently detected in diabetic foot ulcers, was inoculated into cutaneous wounds of db/db diabetic mice to mimic bacterium-infected diabetic wound healing. We observed that P. aeruginosa infection impaired diabetic wound healing and quickly promoted the expression of pro-inflammatory genes (M1 macrophage markers) tumor necrosis factor-α (tnf-α), interleukin-1β (il-1β) and il-6 in wounds. The expression of markers of M2 macrophages, including il-10, arginase-1, and ym1 were also upregulated. In addition, similar gene expression patterns were observed in macrophages isolated directly from wounds. Immunostaining showed that P. aeruginosa infection increased both the ratios of M1 and M2 macrophages in wounds compared with that in control groups, which was further confirmed by in vitro culturing macrophages with P. aeruginosa and skin fibroblast conditioned medium. However, the ratios of the expression levels of pro-inflammatory genes to anti-inflammatory gene il-10 was increased markedly in P. aeruginosa infected wounds and macrophages compared with that in control groups, and P. aeruginosa prolonged the presence of M1 macrophages in the wounds. These data demonstrated that P. aeruginosa in diabetic wounds activates a mixed M1/M2 macrophage phenotype with an excessive activation of M1 phenotype or relatively inadequate activation of M2 phenotype. © 2018 International Federation for Cell Biology.

The Role of Myoepithelial Maspin in Breast Carcinoma Progression Diagnosis and Screening

DTIC Science & Technology

2003-08-01

expression pro- xenografts, the tumors were essentially avascular and their filing (gene chip analysis) to see whether the different myo- 116 S.H. Barskv...factor; aFGF, acidic fibroblast growth factor; TFGa, transforming growth factor a; TGFP3. transforming growth factor 03; TNFa, tumor necrosis factor a...angiogenesis but containing bound angiogenic inhibitors. These myoepithelial xenografts exhibit only minimal hypoxia but extensive necrosis compared with their

Genetic organization and regulation of a meta cleavage pathway for catechols produced from catabolism of toluene, benzene, phenol, and cresols by Pseudomonas pickettii PKO1.

PubMed Central

Kukor, J J; Olsen, R H

1991-01-01

Plasmid pRO1957 contains a 26.5-kb BamHI restriction endonuclease-cleaved DNA fragment cloned from the chromosome of Pseudomonas pickettii PKO1 that allows P. aeruginosa PAO1c to grow on toluene, benzene, phenol, or m-cresol as the sole carbon source. The genes encoding enzymes for meta cleavage of catechol or 3-methylcatechol, derived from catabolism of these substrates, were subcloned from pRO1957 and were shown to be organized into a single operon with the promoter proximal to tbuE. Deletion and analysis of subclones demonstrated that the order of genes in the meta cleavage operon was tbuEFGKIHJ, which encoded catechol 2,3-dioxygenase, 2-hydroxymuconate semialdehyde hydrolase, 2-hydroxymuconate semialdehyde dehydrogenase, 4-hydroxy-2-oxovalerate aldolase, 4-oxalocrotonate decarboxylase, 4-oxalocrotonate isomerase, and 2-hydroxypent-2,4-dienoate hydratase, respectively. The regulatory gene for the tbuEFGKIHJ operon, designated tbuS, was subcloned into vector plasmid pRO2317 from pRO1957 as a 1.3-kb PstI fragment, designated pRO2345. When tbuS was not present, meta pathway enzyme expression was partially derepressed, but these activity levels could not be fully induced. However, when tbuS was present in trans with tbuEFGKIHJ, meta pathway enzymes were repressed in the absence of an effector and were fully induced when an effector was present. This behavior suggests that the gene product of tbuS acts as both a repressor and an activator. Phenol and m-cresol were inducers of meta pathway enzymatic activity. Catechol, 3-methylcatechol, 4-methylcatechol, o-cresol, and p-cresol were not inducers but could be metabolized by cells previously induced by phenol or m-cresol. PMID:1856161

Effects of extending the length of pro-oestrus in an oestradiol- and progesterone-based oestrus synchronisation program on ovarian function, uterine environment and pregnancy establishment in beef heifers.

PubMed

de la Mata, J J; Núñez-Olivera, R; Cuadro, F; Bosolasco, D; de Brun, V; Meikle, A; Bó, G A; Menchaca, A

2018-05-21

The aim of the present study was to investigate the effects of a strategy for extending pro-oestrus (the interval between luteolysis and ovulation) in an oestrus synchronisation protocol (named J-Synch) in beef heifers on follicular growth, sexual steroid concentrations, the oestrogen receptor ERα and progesterone receptors (PR) in the uterus, insulin-like growth factor (IGF) 1 and pregnancy rates. In Experiment 1, heifers treated with the new J-Synch protocol had a longer pro-oestrus period than those treated with the conventional protocol (mean (±s.e.m.) 93.7±12.9 vs 65.0±13.7h respectively; P<0.05). The rate of dominant follicle growth from the time of progesterone device removal to ovulation was greater in heifers in the J-Synch than conventional group (P<0.05). Luteal area and serum progesterone concentrations were greater in the J-Synch Group (P<0.05) for the 12 days after ovulation. Progesterone receptor (PGR) staining on Day 6 after ovulation in the uterine stroma was lower in the J-Synch than conventional group (P<0.05), and the expression of PR gene (PGR) and IGF1 gene tended to be lower in J-Synch-treated heifers (P<0.1). In Experiment 2 (n=2349), the pregnancy rate 30-35 days after fixed-time AI (FTAI) was greater for heifers in the J-Synch than conventional group (56.1% vs 50.7% respectively). In conclusion, our strategy for extending pro-oestrus (i.e. the J-Synch protocol) significantly improves pregnancy establishment in beef heifers. This improvement was related to an increased rate of growth of the dominant ovulatory follicle, greater progesterone concentrations during the ensuing luteal phase and different uterine patterns of PGR and IGF1, which may have favoured embryo development and pregnancy establishment.

Prevalence of the Pro12Ala missense mutation in the PPARG2 gene in Kuwaiti patients with primary knee osteoarthritis

PubMed Central

Al-Jarallah, Khaled F.; Shehab, Diaa K.; Haider, Mohammad Z.

2011-01-01

BACKGROUND AND OBJECTIVES: Peroxisome proliferator–activated receptors (PPARs) play an important role in a number of cellular and metabolic functions. This study was carried out to determine the prevalence of a missense mutation (Pro12Ala) in the PPARG2 gene in Kuwaiti Arab patients with primary knee osteoarthritis (OA) and healthy controls with the aim of identifying a possible association. DESIGN AND SETTING: A prospective cross-sectional study carried out at three major teaching hospitals (referral centers) in the country over a one-year period. PATIENTS AND METHODS: The prevalence of PPARG2 gene Pro12Ala missense mutation was determined in 104 Kuwaiti Arab patients with primary knee OA and 111 ethnically matched healthy controls. The prevalence of this Pro12Ala missense mutation was also determined in clinical subgroups of OA patients divided on the basis of age at onset, function and radiologic grading. RESULTS The Pro-Pro genotype of the PPARG2 gene Pro12Ala missense mutation was detected in 95/104 (91.3%) cases compared to 111/111 (100%) in the control subjects. The heterozygous Pro-Ala genotype was detected in 9/104 (8.7%) of the OA patients, while it was not detected in any of the controls. The Ala-Ala genotype was not detected in any of the OA patients or the controls. No significant differences were detected in the PPARG2 gene Pro12Ala genotypes in the subgroups of patients classified on the basis of age at onset, functional assessment using Lequesne’s functional index, and radiological grading using Kellgren-Lawrence (K-L) grading. CONCLUSIONS This study found no significant association between the PPARG2 gene Pro12Ala missense mutation and knee OA. However, the presence of the Pro-Pro genotype of the PPARG2 gene mutation has a protective effect against development of OA. PMID:21245597

GRBase, a new gene regulation data base available by anonymous ftp.

PubMed Central

Collier, B; Danielsen, M

1994-01-01

The Gene Regulation Database (GRBase) is a compendium of information on the structure and function of proteins involved in the control of gene expression in eukaryotes. These proteins include transcription factors, proteins involved in signal transduction, and receptors. The database can be obtained by FTP in Filemaker Pro, text, and postscript formats. The database will be expanded in the coming year to include reviews on families of proteins involved in gene regulation and to allow online searching. PMID:7937071

TCF21 and the environmental sensor aryl-hydrocarbon receptor cooperate to activate a pro-inflammatory gene expression program in coronary artery smooth muscle cells

PubMed Central

Nguyen, Trieu; Iyer, Dharini; Liu, Boxiang; Wang, Ting; Sazonova, Olga; Matic, Ljubica Perisic; Maegdefessel, Lars; Quertermous, Thomas

2017-01-01

Both environmental factors and genetic loci have been associated with coronary artery disease (CAD), however gene-gene and gene-environment interactions that might identify molecular mechanisms of risk are not easily studied by human genetic approaches. We have previously identified the transcription factor TCF21 as the causal CAD gene at 6q23.2 and characterized its downstream transcriptional network that is enriched for CAD GWAS genes. Here we investigate the hypothesis that TCF21 interacts with a downstream target gene, the aryl hydrocarbon receptor (AHR), a ligand-activated transcription factor that mediates the cellular response to environmental contaminants, including dioxin and polycyclic aromatic hydrocarbons (e.g., tobacco smoke). Perturbation of TCF21 expression in human coronary artery smooth muscle cells (HCASMC) revealed that TCF21 promotes expression of AHR, its heterodimerization partner ARNT, and cooperates with these factors to upregulate a number of inflammatory downstream disease related genes including IL1A, MMP1, and CYP1A1. TCF21 was shown to bind in AHR, ARNT and downstream target gene loci, and co-localization was noted for AHR-ARNT and TCF21 binding sites genome-wide in regions of HCASMC open chromatin. These regions of co-localization were found to be enriched for GWAS signals associated with cardio-metabolic as well as chronic inflammatory disease phenotypes. Finally, we show that similar to TCF21, AHR gene expression is increased in atherosclerotic lesions in mice in vivo using laser capture microdissection, and AHR protein is localized in human carotid atherosclerotic lesions where it is associated with protein kinases with a critical role in innate immune response. These data suggest that TCF21 can cooperate with AHR to activate an inflammatory gene expression program that is exacerbated by environmental stimuli, and may contribute to the overall risk for CAD. PMID:28481916

Pro-inflammatory cytokines and oxidative stress/antioxidant parameters characterize the bio-humoral profile of early cachexia in lung cancer patients.

PubMed

Fortunati, Nicoletta; Manti, Roberta; Birocco, Nadia; Pugliese, Mariateresa; Brignardello, Enrico; Ciuffreda, Libero; Catalano, Maria G; Aragno, Manuela; Boccuzzi, Giuseppe

2007-12-01

Cancer-related cachexia, that is present in about 50% of cancer patients and accounts for 20% of all cancer deaths, is clinically characterized by progressive weight loss, anorexia, metabolic alterations, asthenia, depletion of lipid stores and severe loss of skeletal muscle proteins. The main biochemical and molecular alterations that are responsible for the syndrome are prematurely present in the progress of the disease and the identification of the early stages of cachexia can be useful in targetting patients who will benefit from early treatment. The aim of the present study was to delineate the bio-humoral profile of a group of lung cancer patients either non-cachectic or cachectic by evaluating serum pro-inflammatory cytokines and oxidative stress/antioxidant parameters (both recognized to be involved in cachexia pathogenesis) and pro-inflammatory cytokine gene expression in PBMC (Peripheral blood mononuclear cells) of cancer patients. All serum pro-inflammatory cytokines and oxidative stress/antioxidant parameters significantly increased in neoplastic patients, but only TNF-alpha, ROS, GSH and vitamin E showed a significantly greater increase in cachectic patients. Pro-inflammatory cytokine gene expression mirrored serum level behaviour except for IL-6 that was increased in serum but not as gene expression, suggesting its provenience from tumour tissue. Our data support that the simultaneous determination of ROS, GSH, vitamin E, together with TNF-alpha allows the identification of a lung cancer patient developing cancer-related cachexia. This bio-humoral profile should be used for the early diagnosis and follow-up of the syndrome. Moreover, the evaluation of gene expression in patient PBMC was helpful in differentiating tumour vs host factors, therefore being useful in the study of pathogenetic mechanisms in neoplastic cachectic patients.

Globular adiponectin induces a pro-inflammatory response in human astrocytic cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wan, Zhongxiao; Mah, Dorrian; Simtchouk, Svetlana

Highlights: • Adiponectin receptors are expressed in human astrocytes. • Globular adiponectin induces secretion of IL-6 and MCP-1 from cultured astrocytes. • Adiponectin may play a pro-inflammatory role in astrocytes. - Abstract: Neuroinflammation, mediated in part by activated brain astrocytes, plays a critical role in the development of neurodegenerative disorders, including Alzheimer’s disease (AD). Adiponectin is the most abundant adipokine secreted from adipose tissue and has been reported to exert both anti- and pro-inflammatory effects in peripheral tissues; however, the effects of adiponectin on astrocytes remain unknown. Shifts in peripheral concentrations of adipokines, including adiponectin, could contribute to the observedmore » link between midlife adiposity and increased AD risk. The aim of the present study was to characterize the effects of globular adiponectin (gAd) on pro-inflammatory cytokine mRNA expression and secretion in human U373 MG astrocytic cells and to explore the potential involvement of nuclear factor (NF)-κB, p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase (ERK)1/2, c-Jun N-terminal kinase (JNK) and phosphatidylinositide 3-kinases (PI3 K) signaling pathways in these processes. We demonstrated expression of adiponectin receptor 1 (adipoR1) and adipoR2 in U373 MG cells and primary human astrocytes. gAd induced secretion of interleukin (IL)-6 and monocyte chemoattractant protein (MCP)-1, and gene expression of IL-6, MCP-1, IL-1β and IL-8 in U373 MG cells. Using specific inhibitors, we found that NF-κB, p38MAPK and ERK1/2 pathways are involved in gAd-induced induction of cytokines with ERK1/2 contributing the most. These findings provide evidence that gAd may induce a pro-inflammatory phenotype in human astrocytes.« less

CRM1 protein-mediated regulation of nuclear clusterin (nCLU), an ionizing radiation-stimulated, Bax-dependent pro-death factor.

PubMed

Leskov, Konstantin S; Araki, Shinako; Lavik, John-Paul; Gomez, Jose A; Gama, Vivian; Gonos, Efstathios S; Trougakos, Ioannis P; Matsuyama, Shigemi; Boothman, David A

2011-11-18

Expression of the clusterin (CLU) gene results in the synthesis of a conventional secretory isoform set (pre- and mature secretory clusterin proteins, psCLU/sCLU), as well as another set of intracellular isoforms, appearing in the cytoplasm (pre-nuclear CLU, pnCLU) and in the nucleus as an ∼55-kDa mature nuclear clusterin (nCLU) form. These two isoform sets have opposing cell functions: pro-survival and pro-death, respectively. Although much is known about the regulation and function of sCLU as a pro-survival factor, the regulation and function of endogenous nCLU in cell death are relatively unexplored. Here, we show that depletion of endogenous nCLU protein using siRNA specific to its truncated mRNA increased clonogenic survival of ionizing radiation (IR)-exposed cells. nCLU-mediated apoptosis was Bax-dependent, and lethality correlated with accumulation of mature nCLU protein. nCLU accumulation was regulated by CRM1 because binding between CRM1 and nCLU proteins was significantly diminished by leptomycin B (LMB), and nuclear levels of nCLU protein were significantly enhanced by LMB and IR co-treatment. Moreover, LMB treatment significantly enhanced IR-induced nCLU-mediated cell death responses. Importantly, bax(-/-) and bax(-/-)/bak(-/-) double knock-out cells were resistant to nCLU-mediated cell death, whereas bak(-/-) or wild-type bax(+/+)/bak(+/+) cells were hypersensitive. The regulation of nCLU by CRM1 nuclear export/import may explain recent clinical results showing that highly malignant tumors have lost the ability to accumulate nCLU levels, thereby avoiding growth inhibition and cell death.

Functional Specialization and Evolution of Leader Proteinases in the Family Closteroviridae

PubMed Central

Peng, Chih-Wen; Peremyslov, Valera V.; Mushegian, Arcady R.; Dawson, William O.; Dolja, Valerian V.

2001-01-01

Members of the Closteroviridae and Potyviridae families of the plant positive-strand RNA viruses encode one or two papain-like leader proteinases. In addition to a C-terminal proteolytic domain, each of these proteinases possesses a nonproteolytic N-terminal domain. We compared functions of the several leader proteinases using a gene swapping approach. The leader proteinase (L-Pro) of Beet yellows virus (BYV; a closterovirus) was replaced with L1 or L2 proteinases of Citrus tristeza virus (CTV; another closterovirus), P-Pro proteinase of Lettuce infectious yellows virus (LIYV; a crinivirus), and HC-Pro proteinase of Tobacco etch virus (a potyvirus). Each foreign proteinase efficiently processed the chimeric BYV polyprotein in vitro. However, only L1 and P-Pro, not L2 and HC-Pro, were able to rescue the amplification of the chimeric BYV variants. The combined expression of L1 and L2 resulted in an increased RNA accumulation compared to that of the parental BYV. Remarkably, this L1-L2 chimera exhibited reduced invasiveness and inability to move from cell to cell. Similar analyses of the BYV hybrids, in which only the papain-like domain of L-Pro was replaced with those derived from L1, L2, P-Pro, and HC-Pro, also revealed functional specialization of these domains. In subcellular-localization experiments, distinct patterns were observed for the leader proteinases of BYV, CTV, and LIYV. Taken together, these results demonstrated that, in addition to a common proteolytic activity, the leader proteinases of closteroviruses possess specialized functions in virus RNA amplification, virus invasion, and cell-to-cell movement. The phylogenetic analysis suggested that functionally distinct L1 and L2 of CTV originated by a gene duplication event. PMID:11711606

Functional specialization and evolution of leader proteinases in the family Closteroviridae.

PubMed

Peng, C W; Peremyslov, V V; Mushegian, A R; Dawson, W O; Dolja, V V

2001-12-01

Members of the Closteroviridae and Potyviridae families of the plant positive-strand RNA viruses encode one or two papain-like leader proteinases. In addition to a C-terminal proteolytic domain, each of these proteinases possesses a nonproteolytic N-terminal domain. We compared functions of the several leader proteinases using a gene swapping approach. The leader proteinase (L-Pro) of Beet yellows virus (BYV; a closterovirus) was replaced with L1 or L2 proteinases of Citrus tristeza virus (CTV; another closterovirus), P-Pro proteinase of Lettuce infectious yellows virus (LIYV; a crinivirus), and HC-Pro proteinase of Tobacco etch virus (a potyvirus). Each foreign proteinase efficiently processed the chimeric BYV polyprotein in vitro. However, only L1 and P-Pro, not L2 and HC-Pro, were able to rescue the amplification of the chimeric BYV variants. The combined expression of L1 and L2 resulted in an increased RNA accumulation compared to that of the parental BYV. Remarkably, this L1-L2 chimera exhibited reduced invasiveness and inability to move from cell to cell. Similar analyses of the BYV hybrids, in which only the papain-like domain of L-Pro was replaced with those derived from L1, L2, P-Pro, and HC-Pro, also revealed functional specialization of these domains. In subcellular-localization experiments, distinct patterns were observed for the leader proteinases of BYV, CTV, and LIYV. Taken together, these results demonstrated that, in addition to a common proteolytic activity, the leader proteinases of closteroviruses possess specialized functions in virus RNA amplification, virus invasion, and cell-to-cell movement. The phylogenetic analysis suggested that functionally distinct L1 and L2 of CTV originated by a gene duplication event.

ProDiGe: Prioritization Of Disease Genes with multitask machine learning from positive and unlabeled examples

PubMed Central

2011-01-01

Background Elucidating the genetic basis of human diseases is a central goal of genetics and molecular biology. While traditional linkage analysis and modern high-throughput techniques often provide long lists of tens or hundreds of disease gene candidates, the identification of disease genes among the candidates remains time-consuming and expensive. Efficient computational methods are therefore needed to prioritize genes within the list of candidates, by exploiting the wealth of information available about the genes in various databases. Results We propose ProDiGe, a novel algorithm for Prioritization of Disease Genes. ProDiGe implements a novel machine learning strategy based on learning from positive and unlabeled examples, which allows to integrate various sources of information about the genes, to share information about known disease genes across diseases, and to perform genome-wide searches for new disease genes. Experiments on real data show that ProDiGe outperforms state-of-the-art methods for the prioritization of genes in human diseases. Conclusions ProDiGe implements a new machine learning paradigm for gene prioritization, which could help the identification of new disease genes. It is freely available at http://cbio.ensmp.fr/prodige. PMID:21977986

LentiPro26: novel stable cell lines for constitutive lentiviral vector production.

PubMed

Tomás, H A; Rodrigues, A F; Carrondo, M J T; Coroadinha, A S

2018-03-27

Lentiviral vectors (LVs) are excellent tools to promote gene transfer and stable gene expression. Their potential has been already demonstrated in gene therapy clinical trials for the treatment of diverse disorders. For large scale LV production, a stable producer system is desirable since it allows scalable and cost-effective viral productions, with increased reproducibility and safety. However, the development of stable systems has been challenging and time-consuming, being the selection of cells presenting high expression levels of Gag-Pro-Pol polyprotein and the cytotoxicity associated with some viral components, the main limitations. Hereby is described the establishment of a new LV producer cell line using a mutated less active viral protease to overcome potential cytotoxic limitations. The stable transfection of bicistronic expression cassettes with re-initiation of the translation mechanism enabled the generation of LentiPro26 packaging populations supporting high titers. Additionally, by skipping intermediate clone screening steps and performing only one final clone screening, it was possible to save time and generate LentiPro26-A59 cell line, that constitutively produces titers above 10 6 TU.mL -1 .day -1 , in less than six months. This work constitutes a step forward towards the development of improved LV producer cell lines, aiming to efficiently supply the clinical expanding gene therapy applications.

Invivo monitoring of fetoplacental vegfr2 gene activity in a murine pregnancy model using a vegfr2 -luc reporter gene and bioluminescent imaging

USDA-ARS?s Scientific Manuscript database

Vascular endothelial growth factor receptor-2 (VEGFR2) plays a pivotal role in angiogenesis by eliciting vascular endothelial cell growth when bound to VEGF, a powerful pro-angiogenic ligand. While Vegf and Vegfr2 are expressed throughout gestation, the latter third of gestation in mice is character...

AKT activation drives the nuclear localization of CSE1L and a pro-oncogenic transcriptional activation in ovarian cancer cells.

PubMed

Lorenzato, Annalisa; Biolatti, Marta; Delogu, Giuseppe; Capobianco, Giampiero; Farace, Cristiano; Dessole, Salvatore; Cossu, Antonio; Tanda, Francesco; Madeddu, Roberto; Olivero, Martina; Di Renzo, Maria Flavia

2013-10-15

The human homolog of the yeast cse1 gene (CSE1L) is over-expressed in ovarian cancer. CSE1L forms complex with Ran and importin-α and has roles in nucleocytoplasmic traffic and gene expression. CSE1L accumulated in the nucleus of ovarian cancer cell lines, while it was localized also in the cytoplasm of other cancer cell lines. Nuclear localization depended on AKT, which was constitutively active in ovarian cancer cells, as the CSE1L protein translocated to the cytoplasm when AKT was inactivated. Moreover, the expression of a constitutively active AKT forced the translocation of CSE1L from the cytoplasm to the nucleus in other cancer cells. Nuclear accrual of CSE1L was associated to the nuclear accumulation of the phosphorylated Ran Binding protein 3 (RanBP3), which depended on AKT as well. Also in samples of human ovarian cancer, AKT activation was associated to nuclear accumulation of CSE1L and phosphorylation of RanBP3. Expression profiling of ovarian cancer cells after CSE1L silencing showed that CSE1L was required for the expression of genes promoting invasion and metastasis. In agreement, CSE1L silencing impaired motility and invasiveness of ovarian cancer cells. Altogether these data show that in ovarian cancer cells activated AKT by affecting RanBP3 phosphorylation determines the nuclear accumulation of CSE1L and likely the nuclear concentration of transcription factors conveying pro-oncogenic signals. © 2013 Elsevier Inc. All rights reserved.

Reduction in (pro-)inflammatory responses of lung cells exposed in vitro to diesel exhaust treated with a non-catalyzed diesel particle filter

NASA Astrophysics Data System (ADS)

Steiner, Sandro; Czerwinski, Jan; Comte, Pierre; Müller, Loretta L.; Heeb, Norbert V.; Mayer, Andreas; Petri-Fink, Alke; Rothen-Rutishauser, Barbara

2013-12-01

Increasingly stringent regulation of particulate matter emissions from diesel vehicles has led to the widespread use of diesel particle filters (DPFs), the effect of which on exhaust toxicity is so far poorly understood. We exposed a cellular model of the human respiratory epithelium at the air-liquid interface to non-catalyzed wall-flow DPF-filtered diesel exhaust and compared the resulting biological responses to the ones observed upon exposure to unfiltered exhaust. Filtered diesel exhaust acted highly oxidative, even though to a lesser extent than unfiltered exhaust (quantification of total reduced glutathione), and both exhaust types triggered comparable responses to oxidative stress (measurement of heme-oxygenase 1 (HMOX1) and superoxide-dismutase (SOD1) gene expression). Further, diesel exhaust filtration significantly reduced pro-inflammatory responses (measurement of tumor necrosis factor (TNF) and interleukin-8 (IL-8) gene expression and quantification of the secretion of their gene products TNF-α and IL-8). Because inflammatory processes are central to the onset of adverse respiratory health effects caused by diesel exhaust inhalation, our results imply that DPFs may make a valuable contribution to the detoxification of diesel vehicle emissions. The induction of significant oxidative stress by filtered diesel exhaust however, also implies that the non-particulate exhaust components also need to be considered for lung cell risk assessment.

Plant viral synergism: the potyviral genome encodes a broad-range pathogenicity enhancer that transactivates replication of heterologous viruses.

PubMed Central

Pruss, G; Ge, X; Shi, X M; Carrington, J C; Bowman Vance, V

1997-01-01

Synergistic viral diseases of higher plants are caused by the interaction of two independent viruses in the same host and are characterized by dramatic increases in symptoms and in accumulation of one of the coinfecting viruses. In potato virus X (PVX)/potyviral synergism, increased pathogenicity and accumulation of PVX are mediated by the expression of potyviral 5' proximal sequences encoding P1, the helper component proteinase (HC-Pro), and a fraction of P3. Here, we report that the same potyviral sequence (termed P1/HC-Pro) enhances the pathogenicity and accumulation of two other heterologous viruses: cucumber mosaic virus and tobacco mosaic virus. In the case of PVX-potyviral synergism, we show that the expression of the HC-Pro gene product, but not the RNA sequence itself, is sufficient to induce the increase in PVX pathogenicity and that both P1 and P3 coding sequences are dispensable for this aspect of the synergistic interaction. In protoplasts, expression of the potyviral P1/HC-Pro region prolongs the accumulation of PVX (-) strand RNA and transactivates expression of a reporter gene from a PVX subgenomic promoter. Unlike the synergistic enhancement of PVX pathogenicity, which requires only expression of HC-Pro, the enhancement of PVX (-) strand RNA accumulation in protoplasts is significantly greater when the entire P1/HC-Pro sequence is expressed. These results indicate that the potyviral P1/HC-Pro region affects a step in disease development that is common to a broad range of virus infections and suggest a mechanism involving transactivation of viral replication. PMID:9212462

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, De-Ju; Xu, Yan-Ming; Du, Ji-Ying

Highlights: • Normal human bronchial epithelial cells (BEAS-2B) were dosed with cadmium (Cd). • A low level (2 μM) of Cd treatment for 36 h elicited negligible cytotoxicity. • High levels (20 or 30 μM) of Cd treatment for 36 h induced cell death. • High levels of Cd can upregulate the protein levels of eIF5A1 and NF-κB p65. • We suggest that eIF5A1 level is possibly modulated by NF-κB. - Abstract: Cadmium (Cd) and Cd compounds are widely-distributed in the environment and well-known carcinogens. Here, we report that in CdCl{sub 2}-exposed human bronchial epithelial cells (BEAS-2B), the level ofmore » p53 is dramatically decreased in a time- and dose-dependent manner, suggesting that the observed Cd-induced cytotoxicity is not likely due to the pro-apoptotic function of p53. Therefore, this prompted us to further study the responsive pro-apoptotic factors by proteomic approaches. Interestingly, we identified that high levels (20 or 30 μM) of Cd can significantly upregulate the protein levels of eukaryotic translation initiation factor 5A1 (eIF5A1) and redox-sensitive transcription factor NF-κB p65. Moreover, there is an enhanced NF-κB nuclear translocation as well as chromatin-binding in Cd-treated BEAS-2B cells. We also show that small interfering RNA-specific knockdown of eIF5A1 in Cd-exposed cells attenuated the Cd cytotoxicity, indicating the potential role of eIF5A1 in Cd cytotoxicity. As eIF5A1 is reported to be related with cell apoptosis but little is known about its transcriptional control, we hypothesize that NF-κB might likely modulate eIF5A1 gene expression. Notably, by bioinformatic analysis, several potential NF-κB binding sites on the upstream promoter region of eIF5A1 gene can be found. Subsequent chromatin immunoprecipitation assay revealed that indeed there is enhanced NF-κB binding on eIF5A1 promoter region of Cd-treated BEAS-2B cells. Taken together, our findings suggest for the first time a regulatory mechanism for the pro-apoptotic protein eIF5A1 in which its level is possibly modulated by NF-κB in human lung cells.« less

No association between the sigma receptor type 1 gene and schizophrenia: results of analysis and meta-analysis of case-control studies

PubMed Central

Uchida, Naohiko; Ujike, Hiroshi; Nakata, Kenji; Takaki, Manabu; Nomura, Akira; Katsu, Takeshi; Tanaka, Yuji; Imamura, Takaki; Sakai, Ayumu; Kuroda, Shigetoshi

2003-01-01

Background Several lines of evidence have supported possible roles of the sigma receptors in the etiology of schizophrenia and mechanisms of antipsychotic efficacy. An association study provided genetic evidence that the sigma receptor type 1 gene (SIGMAR1) was a possible susceptibility factor for schizophrenia, however, it was not replicated by a subsequent study. It is necessary to evaluate further the possibility that the SIGMAR1 gene is associated with susceptibility to schizophrenia. Methods A case-control association study between two polymorphisms of the SIGMAR1 gene, G-241T/C-240T and Gln2Pro, and schizophrenia in Japanese population, and meta-analysis including present and previous studies. Results There was no significant association of any allele or genotype of the polymorphisms with schizophrenia. Neither significant association was observed with hebephrenic or paranoid subtype of schizophrenia. Furthermore, a meta-analysis including the present and previous studies comprising 779 controls and 636 schizophrenics also revealed no significant association between the SIGMAR1 gene and schizophrenia. Conclusion In view of this evidence, it is likely that the SIGMAR1 gene does not confer susceptibility to schizophrenia. PMID:14567761

No association between the sigma receptor type 1 gene and schizophrenia: results of analysis and meta-analysis of case-control studies.

PubMed

Uchida, Naohiko; Ujike, Hiroshi; Nakata, Kenji; Takaki, Manabu; Nomura, Akira; Katsu, Takeshi; Tanaka, Yuji; Imamura, Takaki; Sakai, Ayumu; Kuroda, Shigetoshi

2003-10-21

Several lines of evidence have supported possible roles of the sigma receptors in the etiology of schizophrenia and mechanisms of antipsychotic efficacy. An association study provided genetic evidence that the sigma receptor type 1 gene (SIGMAR1) was a possible susceptibility factor for schizophrenia, however, it was not replicated by a subsequent study. It is necessary to evaluate further the possibility that the SIGMAR1 gene is associated with susceptibility to schizophrenia. A case-control association study between two polymorphisms of the SIGMAR1 gene, G-241T/C-240T and Gln2Pro, and schizophrenia in Japanese population, and meta-analysis including present and previous studies. There was no significant association of any allele or genotype of the polymorphisms with schizophrenia. Neither significant association was observed with hebephrenic or paranoid subtype of schizophrenia. Furthermore, a meta-analysis including the present and previous studies comprising 779 controls and 636 schizophrenics also revealed no significant association between the SIGMAR1 gene and schizophrenia. In view of this evidence, it is likely that the SIGMAR1 gene does not confer susceptibility to schizophrenia.

Human adipocytes are highly sensitive to intermittent hypoxia induced NF-kappaB activity and subsequent inflammatory gene expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Taylor, Cormac T.; Kent, Brian D.; Crinion, Sophie J.

Highlights: • Intermittent hypoxia (IH) leads to NF-κB activation in human primary adipocytes. • Adipocytes bear higher pro-inflammatory potential than other human primary cells. • IH leads to upregulation of multiple pro-inflammatory genes in human adipocytes. - Abstract: Introduction: Intermittent hypoxia (IH)-induced activation of pro-inflammatory pathways is a major contributing factor to the cardiovascular pathophysiology associated with obstructive sleep apnea (OSA). Obesity is commonly associated with OSA although it remains unknown whether adipose tissue is a major source of inflammatory mediators in response to IH. The aim of this study was to test the hypothesis that IH leads to augmentedmore » inflammatory responses in human adipocytes when compared to cells of non-adipocyte lineages. Methods and results: Human primary subcutaneous and visceral adipocytes, human primary microvascular pulmonary endothelial cells (HUMEC-L) and human primary small airway epithelial cells (SAEC) were exposed to 0, 6 or 12 cycles of IH or stimulated with tumor necrosis factor (TNF)-α. IH led to a robust increase in NF-κB DNA-binding activity in adipocytes compared with normoxic controls regardless of whether the source of adipocytes was visceral or subcutaneous. Notably, the NF-κB response of adipocytes to both IH and TNF-α was significantly greater than that in HUMEC-L and SAEC. Western blotting confirmed enhanced nuclear translocation of p65 in adipocytes in response to IH, accompanied by phosphorylation of I-κB. Parallel to p65 activation, we observed a significant increase in secretion of the adipokines interleukin (IL)-8, IL-6 and TNF-α with IH in adipocytes accompanied by significant upregulation of mRNA expression. PCR-array suggested profound influence of IH on pro-inflammatory gene expression in adipocytes. Conclusion: Human adipocytes demonstrate strong sensitivity to inflammatory gene expression in response to acute IH and hence, adipose tissue may be a key source of inflammatory mediators in OSA.« less

Serum and gene expression profile of cytokines in first-episode psychosis.

PubMed

Di Nicola, Marco; Cattaneo, Annamaria; Hepgul, Nilay; Di Forti, Marta; Aitchison, Katherine J; Janiri, Luigi; Murray, Robin M; Dazzan, Paola; Pariante, Carmine M; Mondelli, Valeria

2013-07-01

An inflammatory syndrome has been previously reported in chronic schizophrenia. The aims of this study were to investigate: (1) serum levels and leukocyte gene expression of cytokines in patients with first-episode psychosis and controls; and (2) possible causes of abnormal cytokine levels in first-episode psychosis, testing their association with psychosocial stressors, current nicotine and cannabis use, and duration of antipsychotic treatment. We recruited 24 first-episode psychosis patients and 24 healthy controls matched for age, gender, ethnicity and body mass index. Serum interleukin(IL)-1α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, Tumour Necrosis Factor- α (TNF-α), Interferon- γ (IFN-γ), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), and monocyte chemotactic protein-1 (MCP-1) were analysed in all subjects. Leukocyte gene expression analyses were conducted only for those cytokines that were different between-groups in the serum analyses. Patients had significantly higher serum levels of IL-1α (effect size d=0.6, p=0.03), IL-1β (d=0.4, p=0.01), IL-8 (d=0.6, p=0.01) and TNF-α (d=0.7, p=0.05) and a trend for higher IL-6 serum levels (d=0.3, p=0.09) when compared with controls. Leukocyte m-RNA levels of IL-1α (d=0.6, p=0.04), IL-6 (d=0.7, p=0.01) and TNF-α (d=1.6, p<0.001), but not IL-1β and IL-8, were also significantly higher in patients. A history of childhood trauma was associated with higher TNF-α serum levels (p=0.01), while more recent stressful life-events were associated with higher TNF-α mRNA levels in leukocytes (p=0.002). In conclusion, first-episode psychosis is characterised by a pro-inflammatory state supported, at least in part, by activation of leukocytes. Past and recent stressors contribute to this pro-inflammatory state. Copyright © 2012 Elsevier Inc. All rights reserved.

The Transcription Factor p53 Influences Microglial Activation Phenotype

PubMed Central

Jayadev, Suman; Nesser, Nicole K.; Hopkins, Stephanie; Myers, Scott J.; Case, Amanda; Lee, Rona J.; Seaburg, Luke A.; Uo, Takuma; Murphy, Sean P.; Morrison, Richard S.; Garden, Gwenn A.

2011-01-01

Several neurodegenerative diseases are influenced by the innate immune response in the central nervous system (CNS). Microglia, have pro-inflammatory and subsequently neurotoxic actions as well as anti-inflammatory functions that promote recovery and repair. Very little is known about the transcriptional control of these specific microglial behaviors. We have previously shown that in HIV associated neurocognitive disorders (HAND), the transcription factor p53 accumulates in microglia and that microglial p53 expression is required for the in vitro neurotoxicity of the HIV coat glycoprotein gp120. These findings suggested a novel function for p53 in regulating microglial activation. Here we report that in the absence of p53, microglia demonstrate a blunted response to interferon-γ, failing to increase expression of genes associated with classical macrophage activation or secrete pro-inflammatory cytokines. Microarray analysis of global gene expression profiles revealed increased expression of genes associated with anti-inflammatory functions, phagocytosis and tissue repair in p53 knockout (p53−/−) microglia compared with those cultured from strain matched p53 expressing (p53+/+) mice. We further observed that p53−/− microglia demonstrate increased phagocytic activity in vitro and expression of markers for alternative macrophage activation both in vitro and in vivo. In HAND brain tissue, the alternative activation marker CD163 was expressed in a separate subset of microglia than those demonstrating p53 accumulation. These data suggest that p53 influences microglial behavior, supporting the adoption of a pro-inflammatory phenotype, while p53 deficiency promotes phagocytosis and gene expression associated with alternative activation and anti-inflammatory functions. PMID:21598312

The transcriptional PPARβ/δ network in human macrophages defines a unique agonist-induced activation state

PubMed Central

Adhikary, Till; Wortmann, Annika; Schumann, Tim; Finkernagel, Florian; Lieber, Sonja; Roth, Katrin; Toth, Philipp M.; Diederich, Wibke E.; Nist, Andrea; Stiewe, Thorsten; Kleinesudeik, Lara; Reinartz, Silke; Müller-Brüsselbach, Sabine; Müller, Rolf

2015-01-01

Peroxisome proliferator-activated receptor β/δ (PPARβ/δ) is a lipid ligand-inducible transcription factor with established metabolic functions, whereas its anti-inflammatory function is poorly understood. To address this issue, we determined the global PPARβ/δ-regulated signaling network in human monocyte-derived macrophages. Besides cell type-independent, canonical target genes with metabolic and immune regulatory functions we identified a large number of inflammation-associated NFκB and STAT1 target genes that are repressed by agonists. Accordingly, PPARβ/δ agonists inhibited the expression of multiple pro-inflammatory mediators and induced an anti-inflammatory, IL-4-like morphological phenotype. Surprisingly, bioinformatic analyses also identified immune stimulatory effects. Consistent with this prediction, PPARβ/δ agonists enhanced macrophage survival under hypoxic stress and stimulated CD8+ T cell activation, concomitantly with the repression of immune suppressive target genes and their encoded products CD274 (PD-1 ligand), CD32B (inhibitory Fcγ receptor IIB) and indoleamine 2,3-dioxygenase 1 (IDO-1), as well as a diminished release of the immune suppressive IDO-1 metabolite kynurenine. Comparison with published data revealed a significant overlap of the PPARβ/δ transcriptome with coexpression modules characteristic of both anti-inflammatory and pro-inflammatory cytokines. Our findings indicate that PPARβ/δ agonists induce a unique macrophage activation state with strong anti-inflammatory but also specific immune stimulatory components, pointing to a context-dependent function of PPARβ/δ in immune regulation. PMID:25934804

Atf3 mutant mice show reduced axon regeneration and impaired regeneration-associated gene induction after peripheral nerve injury

PubMed Central

Gey, Manuel; Wanner, Renate; Schilling, Corinna; Pedro, Maria T.; Sinske, Daniela

2016-01-01

Axon injury in the peripheral nervous system (PNS) induces a regeneration-associated gene (RAG) response. Atf3 (activating transcription factor 3) is such a RAG and ATF3's transcriptional activity might induce ‘effector’ RAGs (e.g. small proline rich protein 1a (Sprr1a), Galanin (Gal), growth-associated protein 43 (Gap43)) facilitating peripheral axon regeneration. We provide a first analysis of Atf3 mouse mutants in peripheral nerve regeneration. In Atf3 mutant mice, facial nerve regeneration and neurite outgrowth of adult ATF3-deficient primary dorsal root ganglia neurons was decreased. Using genome-wide transcriptomics, we identified a neuropeptide-encoding RAG cluster (vasoactive intestinal peptide (Vip), Ngf, Grp, Gal, Pacap) regulated by ATF3. Exogenous administration of neuropeptides enhanced neurite growth of Atf3 mutant mice suggesting that these molecules might be effector RAGs of ATF3's pro-regenerative function. In addition to the induction of growth-promoting molecules, we present data that ATF3 suppresses growth-inhibiting molecules such as chemokine (C-C motif) ligand 2. In summary, we show a pro-regenerative ATF3 function during PNS nerve regeneration involving transcriptional activation of a neuropeptide-encoding RAG cluster. ATF3 is a general injury-inducible factor, therefore ATF3-mediated mechanisms identified herein might apply to other cell and injury types. PMID:27581653

Angiogenesis and Vascular Architecture in Pheochromocytomas

PubMed Central

Favier, Judith; Plouin, Pierre-François; Corvol, Pierre; Gasc, Jean-Marie

2002-01-01

Angiogenesis is a critical step in tumor growth and metastatic invasion. We here report the study of the vascular status of 10 benign and 9 malignant pheochromocytomas. We examined the vascular architecture after immunostaining endothelial cells (CD34) and vascular smooth muscle cells (α-actin) and identified a vascular pattern characteristic of malignant lesions. To define a gene expression profile indicative of the invasive phenotype, we studied by in situ hybridization the expression of genes encoding several pro- and anti-angiogenic factors [hypoxia-inducible factor (HIF-1α), EPAS1, vascular endothelial growth factor (VEGF), VEGF receptors, angiopoietins and their receptor Tie2, five genes of the endothelin system, and thrombospondin 1]. A semiquantitative evaluation of the labeling revealed an induction of genes encoding EPAS1, VEGF, VEGFR-1, VEGFR-2, endothelin receptor, type B (ETB) and endothelin receptor, type A (ETA) in malignant pheochromocytomas as compared to benign tumors. These differences were observed in tumor cells, in endothelial cells, or in both. Quantification by real-time reverse-transcriptase polymerase chain reaction showed an increase of EPAS1, VEGF, and ETB transcripts of 4.5-, 3.5-, and 10-fold, respectively, in malignant versus benign tumors. Furthermore, we observed a strong correlation between the expression of EPAS1 and VEGF in tumoral tissue and between EPAS1 and ETB in endothelial cells. Altogether, our observations show that analysis of angiogenesis provides promising new criteria for the diagnosis of malignant pheochromocytomas. PMID:12368197

Ionizing radiation stimulates expression of pro-osteoclastogenic genes in marrow and skeletal tissue

DOE PAGES

Alwood, Joshua S.; Shahnazari, Mohammad; Chicana, Betsabel; ...

2015-03-03

Exposure to ionizing radiation can cause rapid mineral loss and increase bone-resorbing osteoclasts within metabolically active, cancellous bone tissue leading to structural deficits. To better understand mechanisms involved in rapid, radiation-induced bone loss, we determined the influence of total body irradiation on expression of select cytokines known both to stimulate osteoclastogenesis and contribute to inflammatory bone disease. Adult (16 week), male C57BL/6J mice were exposed to either 2 Gy gamma rays ( 137Cs, 0.8 Gy/min) or heavy ions ( 56Fe, 600MeV, 0.50–1.1 Gy/min); this dose corresponds to either a single fraction of radiotherapy (typical total dose is ≥10 Gy) ormore » accumulates over long-duration interplanetary missions. Serum, marrow, and mineralized tissue were harvested 4 h—7 days later. Gamma irradiation caused a prompt (2.6-fold within 4 h) and persistent (peaking at 4.1-fold within 1 day) rise in the expression of the obligate osteoclastogenic cytokine, receptor activator of nuclear factor kappa-B ligand ( Rankl), within marrow cells over controls. Similarly, Rankl expression peaked in marrow cells within 3 days of iron exposure (9.2-fold). Changes in Rankl expression induced by gamma irradiation preceded and overlapped with a rise in expression of other pro-osteoclastic cytokines in marrow (eg, monocyte chemotactic protein-1 increased by 11.9-fold, and tumor necrosis factor-alpha increased by 1.7-fold over controls). The ratio, Rankl/ Opg, in marrow increased by 1.8-fold, a net pro-resorption balance. In the marrow, expression of the antioxidant transcription factor, Nfe2l2, strongly correlated with expression levels of Nfatc1, Csf1, Tnf, and Rankl. Radiation exposure increased a serum marker of bone resorption (tartrate-resistant acid phosphatase) and led to cancellous bone loss (16% decrement after 1 week). Finally, we conclude that total body irradiation (gamma or heavy-ion) caused temporal elevations in the concentrations of specific genes expressed within marrow and mineralized tissue related to bone resorption, including select cytokines that lead to osteoclastogenesis and elevated resorption; this is likely to account for rapid and progressive deterioration of cancellous microarchitecture following exposure to ionizing radiation.« less

Ionizing radiation stimulates expression of pro-osteoclastogenic genes in marrow and skeletal tissue

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alwood, Joshua S.; Shahnazari, Mohammad; Chicana, Betsabel

Exposure to ionizing radiation can cause rapid mineral loss and increase bone-resorbing osteoclasts within metabolically active, cancellous bone tissue leading to structural deficits. To better understand mechanisms involved in rapid, radiation-induced bone loss, we determined the influence of total body irradiation on expression of select cytokines known both to stimulate osteoclastogenesis and contribute to inflammatory bone disease. Adult (16 week), male C57BL/6J mice were exposed to either 2 Gy gamma rays ( 137Cs, 0.8 Gy/min) or heavy ions ( 56Fe, 600MeV, 0.50–1.1 Gy/min); this dose corresponds to either a single fraction of radiotherapy (typical total dose is ≥10 Gy) ormore » accumulates over long-duration interplanetary missions. Serum, marrow, and mineralized tissue were harvested 4 h—7 days later. Gamma irradiation caused a prompt (2.6-fold within 4 h) and persistent (peaking at 4.1-fold within 1 day) rise in the expression of the obligate osteoclastogenic cytokine, receptor activator of nuclear factor kappa-B ligand ( Rankl), within marrow cells over controls. Similarly, Rankl expression peaked in marrow cells within 3 days of iron exposure (9.2-fold). Changes in Rankl expression induced by gamma irradiation preceded and overlapped with a rise in expression of other pro-osteoclastic cytokines in marrow (eg, monocyte chemotactic protein-1 increased by 11.9-fold, and tumor necrosis factor-alpha increased by 1.7-fold over controls). The ratio, Rankl/ Opg, in marrow increased by 1.8-fold, a net pro-resorption balance. In the marrow, expression of the antioxidant transcription factor, Nfe2l2, strongly correlated with expression levels of Nfatc1, Csf1, Tnf, and Rankl. Radiation exposure increased a serum marker of bone resorption (tartrate-resistant acid phosphatase) and led to cancellous bone loss (16% decrement after 1 week). Finally, we conclude that total body irradiation (gamma or heavy-ion) caused temporal elevations in the concentrations of specific genes expressed within marrow and mineralized tissue related to bone resorption, including select cytokines that lead to osteoclastogenesis and elevated resorption; this is likely to account for rapid and progressive deterioration of cancellous microarchitecture following exposure to ionizing radiation.« less

IL-1β-specific recruitment of GCN5 histone acetyltransferase induces the release of PAF1 from chromatin for the de-repression of inflammatory response genes.

PubMed

Kim, Nari; Sun, Hwa-Young; Youn, Min-Young; Yoo, Joo-Yeon

2013-04-01

To determine the functional specificity of inflammation, it is critical to orchestrate the timely activation and repression of inflammatory responses. Here, we explored the PAF1 (RNA polymerase II associated factor)-mediated signal- and locus-specific repression of genes induced through the pro-inflammatory cytokine interleukin (IL)-1β. Using microarray analysis, we identified the PAF1 target genes whose expression was further enhanced by PAF1 knockdown in IL-1β-stimulated HepG2 hepatocarcinomas. PAF1 bound near the transcription start sites of target genes and dissociated on stimulation. In PAF1-deficient cells, more elongating RNA polymerase II and acetylated histones were observed, although IL-1β-mediated activation and recruitment of nuclear factor κB (NF-κB) were not altered. Under basal conditions, PAF1 blocked histone acetyltransferase general control non-depressible 5 (GCN5)-mediated acetylation on H3K9 and H4K5 residues. On IL-1β stimulation, activated GCN5 discharged PAF1 from chromatin, allowing productive transcription to occur. PAF1 bound to histones but not to acetylated histones, and the chromatin-binding domain of PAF1 was essential for target gene repression. Moreover, IL-1β-induced cell migration was similarly controlled through counteraction between PAF1 and GCN5. These results suggest that the IL-1β signal-specific exchange of PAF1 and GCN5 on the target locus limits inappropriate gene induction and facilitates the timely activation of inflammatory responses.

Genetic Contributions to Disparities in Preterm Birth

PubMed Central

Anum, Emmanuel A.; Springel, Edward H.; Shriver, Mark D.; Strauss, Jerome F.

2008-01-01

Ethnic disparity in preterm delivery between African Americans and European Americans has existed for decades, and is likely the consequence of multiple factors, including socioeconomic status, access to care, environment, and genetics. This review summarizes existing information on genetic variation and its association with preterm birth in African Americans. Candidate gene-based association studies, in which investigators have evaluated particular genes selected primarily because of their potential roles in the process of normal and pathological parturition, provide evidence that genetic contributions from both mother and fetus account for some of the disparity in preterm births. To date, most attention has been focused on genetic variation in pro- and anti-inflammatory cytokine genes and their respective receptors. These genes, particularly the pro-inflammatory cytokine genes and their receptors, are linked to matrix metabolism since these cytokines increase expression of matrix degrading metalloproteinases. However, the role that genetic variants that are different between populations play in preterm birth cannot yet be quantified. Future studies based on genome wide association or admixture mapping may reveal other genes that contribute to disparity in prematurity. PMID:18787421

MYCN controls an alternative RNA splicing program in high-risk metastatic neuroblastoma.

PubMed

Zhang, Shile; Wei, Jun S; Li, Samuel Q; Badgett, Tom C; Song, Young K; Agarwal, Saurabh; Coarfa, Cristian; Tolman, Catherine; Hurd, Laura; Liao, Hongling; He, Jianbin; Wen, Xinyu; Liu, Zhihui; Thiele, Carol J; Westermann, Frank; Asgharzadeh, Shahab; Seeger, Robert C; Maris, John M; Guidry Auvil, Jamie M; Smith, Malcolm A; Kolaczyk, Eric D; Shohet, Jason; Khan, Javed

2016-02-28

The molecular mechanisms underlying the aggressive behavior of MYCN driven neuroblastoma (NBL) is under intense investigation; however, little is known about the impact of this family of transcription factors on the splicing program. Here we used high-throughput RNA sequencing to systematically study the expression of RNA isoforms in stage 4 MYCN-amplified NBL, an aggressive subtype of metastatic NBL. We show that MYCN-amplified NBL tumors display a distinct gene splicing pattern affecting multiple cancer hallmark functions. Six splicing factors displayed unique differential expression patterns in MYCN-amplified tumors and cell lines, and the binding motifs for some of these splicing factors are significantly enriched in differentially-spliced genes. Direct binding of MYCN to promoter regions of the splicing factors PTBP1 and HNRNPA1 detected by ChIP-seq demonstrates that MYCN controls the splicing pattern by direct regulation of the expression of these key splicing factors. Furthermore, high expression of PTBP1 and HNRNPA1 was significantly associated with poor overall survival of stage4 NBL patients (p ≤ 0.05). Knocking down PTBP1, HNRNPA1 and their downstream target PKM2, an isoform of pro-tumor-growth, result in repressed growth of NBL cells. Therefore, our study reveals a novel role of MYCN in controlling global splicing program through regulation of splicing factors in addition to its well-known role in the transcription program. These findings suggest a therapeutically potential to target the key splicing factors or gene isoforms in high-risk NBL with MYCN-amplification. Published by Elsevier Ireland Ltd.

The dipeptide Pro-Asp promotes IGF-1 secretion and expression in hepatocytes by enhancing JAK2/STAT5 signaling pathway.

PubMed

Wang, Songbo; Wang, Guoqing; Zhang, Mengyuan; Zhuang, Lu; Wan, Xiaojuan; Xu, Jingren; Wang, Lina; Zhu, Xiaotong; Gao, Ping; Xi, Qianyun; Zhang, Yongliang; Shu, Gang; Jiang, Qingyan

2016-11-15

It has been implicated that IGF-1 secretion can be regulated by dietary protein. However, whether the dipeptides, one of digested products of dietary protein, have influence on IGF-1 secretion remain largely unknown. Our study aimed to investigate the effects of the dipeptide Pro-Asp on IGF-1 secretion and expression in hepatocytes and to explore the possible underlying mechanisms. Our findings demonstrated that Pro-Asp promoted the secretion and gene expression of IGF-1 in HepG2 cells and primary porcine hepatocytes. Meanwhile, Pro-Asp activated the ERK and Akt signaling pathways, downstream of IGF-1. In addition, Pro-Asp enhanced GH-mediated JAK2/STAT5 signaling pathway, while inhibition of JAK2/STAT5 blocked the promotive effect of Pro-Asp on IGF-1 secretion and expression. Moreover, acute injection of Pro-Asp stimulated IGF-1 expression and activated JAK2/STAT5 signaling pathway in mice liver. Together, these results suggested that the dipeptide Pro-Asp promoted IGF-1 secretion and expression in hepatocytes by enhancing GH-mediated JAK2/STAT5 signaling pathway. Copyright © 2016. Published by Elsevier Ireland Ltd.

KLF6 contributes to myeloid cell plasticity in the pathogenesis of intestinal inflammation

PubMed Central

Goodman, Wendy A.; Omenetti, Sara; Date, Dipali; Di Martino, Luca; De Salvo, Carlo; Kim, Gun-Dong; Chowdhry, Saleem; Bamias, Giorgos; Cominelli, Fabio; Pizarro, Theresa T.; Mahabeleshwar, Ganapati H.

2016-01-01

Inflammatory bowel disease (IBD) is associated with dysregulated macrophage responses, such that quiescent macrophages acquire a pro-inflammatory activation state and contribute to chronic intestinal inflammation. The transcriptional events governing macrophage activation and gene expression in the context of chronic inflammation such as IBD remain incompletely understood. Here, we identify Kruppel-like transcription factor-6 (KLF6) as a critical regulator of pathogenic myeloid cell activation in human and experimental IBD. We found that KLF6 was significantly upregulated in myeloid cells and intestinal tissue from IBD patients and experimental models of IBD, particularly in actively inflamed regions of the colon. Using complementary gain- and loss-of-function studies, we observed that KLF6 promotes pro-inflammatory gene expression through enhancement of NFκB signaling, while simultaneously suppressing anti-inflammatory gene expression through repression of STAT3 signaling. To study the in vivo role of myeloid KLF6, we treated myeloid-specific KLF6-knockout mice (Mac-KLF6-KO) with dextran sulfate-sodium (DSS) and found that Mac-KLF6-KO mice were protected against chemically-induced colitis; this highlights the central role of myeloid KLF6 in promoting intestinal inflammation. Collectively, our results point to a novel gene regulatory program underlying pathogenic, pro-inflammatory macrophage activation in the setting of chronic intestinal inflammation. PMID:26838049

Molecular and cellular characterization of the tomato pollen profilin, LePro1

USDA-ARS?s Scientific Manuscript database

Profilin is an actin-binding protein involved in the dynamic turnover and restructuring of the actin cytoskeleton in all eukaryotic cells. We previously cloned a profilin gene, designated as LePro1 from tomato pollen. To investigate its biological role, in the present study, We investigated the tem...

Foxm1 transcription factor is required for lung fibrosis and epithelial-to-mesenchymal transition

PubMed Central

Balli, David; Ustiyan, Vladimir; Zhang, Yufang; Wang, I-Ching; Masino, Alex J; Ren, Xiaomeng; Whitsett, Jeffrey A; Kalinichenko, Vladimir V; Kalin, Tanya V

2013-01-01

Alveolar epithelial cells (AECs) participate in the pathogenesis of pulmonary fibrosis, producing pro-inflammatory mediators and undergoing epithelial-to-mesenchymal transition (EMT). Herein, we demonstrated the critical role of Forkhead Box M1 (Foxm1) transcription factor in radiation-induced pulmonary fibrosis. Foxm1 was induced in AECs following lung irradiation. Transgenic expression of an activated Foxm1 transcript in AECs enhanced radiation-induced pneumonitis and pulmonary fibrosis, and increased the expression of IL-1β, Ccl2, Cxcl5, Snail1, Zeb1, Zeb2 and Foxf1. Conditional deletion of Foxm1 from respiratory epithelial cells decreased radiation-induced pulmonary fibrosis and prevented the increase in EMT-associated gene expression. siRNA-mediated inhibition of Foxm1 prevented TGF-β-induced EMT in vitro. Foxm1 bound to and increased promoter activity of the Snail1 gene, a critical transcriptional regulator of EMT. Expression of Snail1 restored TGF-β-induced loss of E-cadherin in Foxm1-deficient cells in vitro. Lineage-tracing studies demonstrated that Foxm1 increased EMT during radiation-induced pulmonary fibrosis in vivo. Foxm1 is required for radiation-induced pulmonary fibrosis by enhancing the expression of genes critical for lung inflammation and EMT. PMID:23288041

Acute Toxicity of Radiochemotherapy in Rectal Cancer Patients: A Risk Particularly for Carriers of the TGFB1 Pro25 variant

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schirmer, Markus Anton; Mergler, Caroline Patricia Nadine; Rave-Fraenk, Margret

Purpose: Transforming growth factor-beta1 is related to adverse events in radiochemotherapy. We investigated TGFB1 genetic variability in relation to quality of life-impairing acute organ toxicity (QAOT) of neoadjuvant radiochemotherapy under clinical trial conditions. Methods and Materials: Two independent patient cohorts (n = 88 and n = 75) diagnosed with International Union Against Cancer stage II/III rectal cancer received neoadjuvant radiation doses of 50.4 Gy combined with 5-fluorouracil-based chemotherapy. Toxicity was monitored according to Common Terminology Criteria for Adverse Events. QAOT was defined as a CTCAE grade {>=}2 for at least one case of enteritis, proctitis, cystitis, or dermatitis. Nine germlinemore » polymorphisms covering the common genetic diversity in the TGFB1 gene were genotyped. Results: In both cohorts, all patients carrying the TGFB1 Pro25 variant experienced QAOT (positive predictive value of 100%, adjusted p = 0.0006). In a multivariate logistic regression model, gender, age, body mass index, type of chemotherapy, or disease state had no significant impact on QAOT. Conclusion: The TGFB1 Pro25 variant could be a relevant marker for individual treatment stratification and carriers may benefit from adaptive clinical care or specific radiation techniques.« less

[Agrobacterium-mediated sunflower transformation (Helianthus annuus L.) in vitro and in Planta using strain of LBA4404 harboring binary vector pBi2E with dsRNA-suppressor proline dehydrogenase gene].

PubMed

Tishchenko, E N; Komisarenko, A G; Mikhal'skaia, S I; Sergeeva, L E; Adamenko, N I; Morgun, B V; Kochetov, A V

2014-01-01

To estimate the efficiency of proline dehydrogenase gene suppression towards increasing of sunflower (Helianthus annuus L.) tolerance level to water deficit and salinity, we employed strain LBA4404 harboring pBi2E with double-stranded RNA-suppressor, which were prepared on basis arabidopsis ProDH1 gene. The techniques of Agrobacterium-mediated transformation in vitro and in planta during fertilization sunflower have been proposed. There was shown the genotype-depended integration of T-DNA in sunflower genome. PCR-analysis showed that ProDH1 presents in genome of inbred lines transformed in planta, as well as in T1- and T2-generations. In trans-genic regenerants the essential accumulation of free L-proline during early stages of in vitro cultivation under normal conditions was shown. There was established the essential accumulation of free proline in transgenic regenerants during cultivation under lethal stress pressure (0.4 M mannitol and 2.0% sea water salts) and its decline upon the recovery period. These data are declared about effectiveness of suppression of sunflower ProDH and gene participation in processes connected with osmotolerance.

RNA-Seq analysis reveals new evidence for inflammation-related changes in aged kidney

PubMed Central

Park, Daeui; Kim, Byoung-Chul; Kim, Chul-Hong; Choi, Yeon Ja; Jeong, Hyoung Oh; Kim, Mi Eun; Lee, Jun Sik; Park, Min Hi; Chung, Ki Wung; Kim, Dae Hyun; Lee, Jaewon; Im, Dong-Soon; Yoon, Seokjoo; Lee, Sunghoon; Yu, Byung Pal; Bhak, Jong; Chung, Hae Young

2016-01-01

Age-related dysregulated inflammation plays an essential role as a major risk factor underlying the pathophysiological aging process. To better understand how inflammatory processes are related to aging at the molecular level, we sequenced the transcriptome of young and aged rat kidney using RNA-Seq to detect known genes, novel genes, and alternative splicing events that are differentially expressed. By comparing young (6 months of age) and old (25 months of age) rats, we detected 722 up-regulated genes and 111 down-regulated genes. In the aged rats, we found 32 novel genes and 107 alternatively spliced genes. Notably, 6.6% of the up-regulated genes were related to inflammation (P < 2.2 × 10−16, Fisher exact t-test); 15.6% were novel genes with functional protein domains (P = 1.4 × 10−5); and 6.5% were genes showing alternative splicing events (P = 3.3 × 10−4). Based on the results of pathway analysis, we detected the involvement of inflammation-related pathways such as cytokines (P = 4.4 × 10−16), which were found up-regulated in the aged rats. Furthermore, an up-regulated inflammatory gene analysis identified the involvement of transcription factors, such as STAT4, EGR1, and FOSL1, which regulate cancer as well as inflammation in aging processes. Thus, RNA changes in these pathways support their involvement in the pro-inflammatory status during aging. We propose that whole RNA-Seq is a useful tool to identify novel genes and alternative splicing events by documenting broadly implicated inflammation-related genes involved in aging processes. PMID:27153548

Irisin is a pro-myogenic factor that induces skeletal muscle hypertrophy and rescues denervation-induced atrophy.

PubMed

Reza, Musarrat Maisha; Subramaniyam, Nathiya; Sim, Chu Ming; Ge, Xiaojia; Sathiakumar, Durgalakshmi; McFarlane, Craig; Sharma, Mridula; Kambadur, Ravi

2017-10-24

Exercise induces expression of the myokine irisin, which is known to promote browning of white adipose tissue and has been shown to mediate beneficial effects following exercise. Here we show that irisin induces expression of a number of pro-myogenic and exercise response genes in myotubes. Irisin increases myogenic differentiation and myoblast fusion via activation of IL6 signaling. Injection of irisin in mice induces significant hypertrophy and enhances grip strength of uninjured muscle. Following skeletal muscle injury, irisin injection improves regeneration and induces hypertrophy. The effects of irisin on hypertrophy are due to activation of satellite cells and enhanced protein synthesis. In addition, irisin injection rescues loss of skeletal muscle mass following denervation by enhancing satellite cell activation and reducing protein degradation. These data suggest that irisin functions as a pro-myogenic factor in mice.

Dual transcriptome sequencing reveals resistance of TLR4 ligand-activated bone marrow-derived macrophages to inflammation mediated by the BET inhibitor JQ1

PubMed Central

Das, Amitabh; Chai, Jin Choul; Yang, Chul-su; Lee, Young Seek; Das, Nando Dulal; Jung, Kyoung Hwa; Chai, Young Gyu

2015-01-01

Persistent macrophage activation is associated with the expression of various pro-inflammatory genes, cytokines and chemokines, which may initiate or amplify inflammatory disorders. A novel synthetic BET inhibitor, JQ1, was proven to exert immunosuppressive activities in macrophages. However, a genome-wide search for JQ1 molecular targets has not been undertaken. The present study aimed at evaluating the anti-inflammatory function and underlying genes that are targeted by JQ1 in LPS-stimulated primary bone marrow-derived macrophages (BMDMs) using global transcriptomic RNA sequencing and quantitative real-time PCR. Among the annotated genes, transcriptional sequencing of BMDMs that were treated with JQ1 revealed a selective effect on LPS-induced gene expression in which the induction of cytokines/chemokines, interferon-stimulated genes, and prominent (transcription factors) TFs was suppressed. Additionally, we found that JQ1 reduced the expression of previously unidentified genes that are important in inflammation. Importantly, these inflammatory genes were not affected by JQ1 treatment alone. Furthermore, we confirmed that JQ1 reduced cytokines/chemokines in the supernatants of LPS treated BMDMs. Moreover, the biological pathways and gene ontology of the differentially expressed genes were determined in the JQ1 treatment of BMDMs. These unprecedented results suggest that the BET inhibitor JQ1 is a candidate for the prevention or therapeutic treatment of inflammatory disorders. PMID:26582142

Edible blue-green algae reduce the production of pro-inflammatory cytokines by inhibiting NF-κB pathway in macrophages and splenocytes

PubMed Central

Ku, Chai Siah; Pham, Tho X.; Park, Youngki; Kim, Bohkyung; Shin, Min; Kang, Insoo; Lee, Jiyoung

2013-01-01

Background Chronic inflammation contributes to the development of pathological disorders including insulin resistance and atherosclerosis. Identification of anti-inflammatory natural products can prevent the inflammatory diseases. Methods Anti-inflammatory effects of blue-green algae (BGA), i.e., Nostoc commune var. Sphaeroides Kützing (NO) and Spirulina Platensis (SP), were compared in RAW 264.7 and mouse bone marrow-derived macrophages (BMM) as well as splenocytes from apolipoprotein E knockout (apoE−/−) mice fed BGA. Results When macrophages pretreated with 100 μg/ml NO lipid extract (NOE) or SP lipid extract (SPE) were activated by lipopolysaccharide (LPS), expression and secretion of pro-inflammatory cytokines, such as tumor necrosis factor α (TNFα), interleukin 1β (IL-1β), and IL-6, were significantly repressed. NOE and SPE also significantly repressed the expression of TNFα and IL-1β in BMM. LPS-induced secretion of IL-6 was lower in splenocytes from apoE−/− fed an atherogenic diet containing 5% NO or SP for 12 weeks. In RAW 264.7 macrophages, NOE and SPE markedly decreased nuclear translocation of NF-κB. The degree of repression of pro-inflammatory gene expression by algal extracts was much stronger than that of SN50, an inhibitor of NF-κB nuclear translocation. Trichostatin A, a pan histone deacetylase inhibitor, increased basal expression of IL-1β and attenuated the repression of the gene expression by SPE. SPE significantly down-regulated mRNA abundance of 11 HDAC isoforms, consequently increasing acetylated histone 3 levels. Conclusion NOE and SPE repress pro-inflammatory cytokine expression and secretion in macrophages and splenocytes via inhibition of NF-κB pathway. Histone acetylation state is likely involved in the inhibition. General significance This study underscores natural products can exert anti-inflammatory effects by epigenetic modifications such as histone acetylation. PMID:23357040

Expression of genes associated with immunity in the endometrium of cattle with disparate postpartum uterine disease and fertility

PubMed Central

Herath, Shan; Lilly, Sonia T; Santos, Natalia R; Gilbert, Robert O; Goetze, Leopold; Bryant, Clare E; White, John O; Cronin, James; Sheldon, I Martin

2009-01-01

Background Contamination of the uterine lumen with bacteria is ubiquitous in cattle after parturition. Some animals develop endometritis and have reduced fertility but others have no uterine disease and readily conceive. The present study tested the hypothesis that postpartum cattle that develop persistent endometritis and infertility are unable to limit the inflammatory response to uterine bacterial infection. Methods Endometrial biopsies were collected several times during the postpartum period from animals that were subsequently infertile with persistent endometritis (n = 4) or had no clinical disease and conceived to first insemination (n = 4). Quantitative PCR was used to determine the expression of candidate genes in the endometrial biopsies, including the Toll-like receptor (TLR 1 to 10) family of innate immune receptors, inflammatory mediators and their cognate receptors. Selected proteins were examined by immunohistochemistry. Results The expression of genes encoding pro-inflammatory mediators such as interleukins (IL1A, IL1B and IL6), and nitric oxide synthase 2 (NOS2) were higher during the first week post partum than subsequently. During the first week post partum, there was higher gene expression in infertile than fertile animals of TLR4, the receptor for bacterial lipopolysaccharide, and the pro-inflammatory cytokines IL1A and IL1B, and their receptor IL1R2. The expression of genes encoding other Toll-like receptors, transforming growth factor beta receptor 1 (TGFBR1) or prostaglandin E2 receptors (PTGER2 and PTGER4) did not differ significantly between the animal groups. Gene expression did not differ significantly between infertile and fertile animals after the first week postpartum. However, there were higher ratios of IL1A or IL1B mRNA to the anti-inflammatory cytokine IL10, during the first week post partum in the infertile than fertile animals, and the protein products of these genes were mainly localised to the epithelium of the endometrium. Conclusion Cattle may maintain fertility by limiting the inflammatory response to postpartum bacterial infection in the endometrium during the first week after parturition. PMID:19476661

c-Kit modifies the inflammatory status of smooth muscle cells

PubMed Central

Song, Lei; Martinez, Laisel; Zigmond, Zachary M.; Hernandez, Diana R.; Lassance-Soares, Roberta M.; Selman, Guillermo

2017-01-01

Background c-Kit is a receptor tyrosine kinase present in multiple cell types, including vascular smooth muscle cells (SMC). However, little is known about how c-Kit influences SMC biology and vascular pathogenesis. Methods High-throughput microarray assays and in silico pathway analysis were used to identify differentially expressed genes between primary c-Kit deficient (KitW/W–v) and control (Kit+/+) SMC. Quantitative real-time RT-PCR and functional assays further confirmed the differences in gene expression and pro-inflammatory pathway regulation between both SMC populations. Results The microarray analysis revealed elevated NF-κB gene expression secondary to the loss of c-Kit that affects both the canonical and alternative NF-κB pathways. Upon stimulation with an oxidized phospholipid as pro-inflammatory agent, c-Kit deficient SMC displayed enhanced NF-κB transcriptional activity, higher phosphorylated/total p65 ratio, and increased protein expression of NF-κB regulated pro-inflammatory mediators with respect to cells from control mice. The pro-inflammatory phenotype of mutant cells was ameliorated after restoring c-Kit activity using lentiviral transduction. Functional assays further demonstrated that c-Kit suppresses NF-κB activity in SMC in a TGFβ-activated kinase 1 (TAK1) and Nemo-like kinase (NLK) dependent manner. Discussion Our study suggests a novel mechanism by which c-Kit suppresses NF-κB regulated pathways in SMC to prevent their pro-inflammatory transformation. PMID:28626608

c-Kit modifies the inflammatory status of smooth muscle cells.

PubMed

Song, Lei; Martinez, Laisel; Zigmond, Zachary M; Hernandez, Diana R; Lassance-Soares, Roberta M; Selman, Guillermo; Vazquez-Padron, Roberto I

2017-01-01

c-Kit is a receptor tyrosine kinase present in multiple cell types, including vascular smooth muscle cells (SMC). However, little is known about how c-Kit influences SMC biology and vascular pathogenesis. High-throughput microarray assays and in silico pathway analysis were used to identify differentially expressed genes between primary c-Kit deficient (Kit W/W-v ) and control (Kit +/+ ) SMC. Quantitative real-time RT-PCR and functional assays further confirmed the differences in gene expression and pro-inflammatory pathway regulation between both SMC populations. The microarray analysis revealed elevated NF-κB gene expression secondary to the loss of c-Kit that affects both the canonical and alternative NF-κB pathways. Upon stimulation with an oxidized phospholipid as pro-inflammatory agent, c-Kit deficient SMC displayed enhanced NF-κB transcriptional activity, higher phosphorylated/total p65 ratio, and increased protein expression of NF-κB regulated pro-inflammatory mediators with respect to cells from control mice. The pro-inflammatory phenotype of mutant cells was ameliorated after restoring c-Kit activity using lentiviral transduction. Functional assays further demonstrated that c-Kit suppresses NF-κB activity in SMC in a TGFβ-activated kinase 1 (TAK1) and Nemo-like kinase (NLK) dependent manner. Our study suggests a novel mechanism by which c-Kit suppresses NF-κB regulated pathways in SMC to prevent their pro-inflammatory transformation.

TGFbeta1 (Leu10Pro), p53 (Arg72Pro) can predict for increased risk for breast cancer in south Indian women and TGFbeta1 Pro (Leu10Pro) allele predicts response to neo-adjuvant chemo-radiotherapy.

PubMed

Rajkumar, Thangarajan; Samson, Mani; Rama, Ranganathan; Sridevi, Veluswami; Mahji, Urmila; Swaminathan, Rajaraman; Nancy, Nirmala K

2008-11-01

The breast cancer incidence has been increasing in the south Indian women. A case (n=250)-control (n=500) study was undertaken to investigate the role of Single Nucleotide Polymorphisms (SNP's) in GSTM1 (Present/Null); GSTP1 (Ile105Val), p53 (Arg72Pro), TGFbeta1 (Leu10Pro), c-erbB2 (Ile655Val), and GSTT1 (Null/Present) in breast cancer. In addition, the value of the SNP's in predicting primary tumor's pathologic response following neo-adjuvant chemo-radiotherapy was assessed. Genotyping was done using PCR (GSTM1, GSTT1), Taqman Allelic discrimination assay (GSTP1, c-erbB2) and PCR-CTPP (p53 and TGFbeta1). None of the gene SNP's studied were associated with a statistically significant increased risk for the breast cancer. However, combined analysis of the SNP's showed that p53 (Arg/Arg and Arg/Pro) with TGFbeta1 (Pro/Pro and Leu/Pro) were associated with greater than 2 fold increased risk for breast cancer in Univariate (P=0.01) and Multivariate (P=0.003) analysis. There was no statistically significant association for the GST family members with the breast cancer risk. TGFbeta1 (Pro/Pro) allele was found to predict complete pathologic response in the primary tumour following neo-adjuvant chemo-radiotherapy (OR=6.53 and 10.53 in Univariate and Multivariate analysis respectively) (P=0.004) and was independent of stage. This study suggests that SNP's can help predict breast cancer risk in south Indian women and that TGFbeta1 (Pro/Pro) allele is associated with a better pCR in the primary tumour.

Inhibitory Effects of Bangladeshi Medicinal Plant Extracts on Interactions between Transcription Factors and Target DNA Sequences

PubMed Central

Lampronti, Ilaria; Khan, Mahmud T.H.; Borgatti, Monica; Bianchi, Nicoletta

2008-01-01

Several transcription factors (TFs) play crucial roles in governing the expression of different genes involved in the immune response, embryo or cell lineage development, cell apoptosis, cell cycle progression, oncogenesis, repair and fibrosis processes and inflammation. As far as inflammation, TFs playing pivotal roles are nuclear factor kappa B (NF-kB), activator protein (AP-1), signal transducer and activator of transcription (STATs), cAMP response element binding protein (CREB) and GATA-1 factors. All these TFs regulate the expression of pro-inflammatory cytokines and are involved in the pathogenesis of a number of human disorders, particularly those with an inflammatory component. Since several medicinal plants can be employed to produce extracts exhibiting biological effects and because alteration of gene transcription represents a very interesting approach to control the expression of selected genes, this study sought to verify the ability of several extracts derived from Bangladeshi medicinal plants in interfering with molecular interactions between different TFs and specific DNA sequences. We first analyzed the antiproliferative activity of 19 medicinal plants on different human cell lines, including erythroleukemia K562, B lymphoid Raji and T lymphoid Jurkat cell lines. Secondly, we employed the electrophoretic mobility shift assay as a suitable technique for a fast screening of plant extracts altering the binding between NF-kB, AP-1, GATA-1, STAT-3, CREB and the relative target DNA elements. PMID:18830455

Association between the Peroxisome proliferator-activated receptor-gamma Pro12Ala Variant and Haplotype and Pancreatic Cancer in a High-Risk Cohort of Smokers: A Pilot Study

PubMed Central

Fesinmeyer, Megan Dann; Stanford, Janet L.; Brentnall, Teresa A.; Mandelson, Margaret T.; Farin, Federico M.; Srinouanprachanh, Sengkeo; Afsharinejad, Zahra; Goodman, Gary E.; Barnett, Matt J.; Austin, Melissa A.

2010-01-01

Objectives The Pro12Ala variant in the proliferator-activated receptor-gamma (PPARG) gene has been associated with diabetes and several cancers. This pilot study tested for the association between Pro12Ala and pancreatic cancer risk in a high-risk sample of smokers. Methods A nested case-control study was conducted in 83 incident cases of pancreatic cancer and 166 matched controls originally recruited into a cohort chemoprevention study of lung cancer. Associations between Pro12Ala and pancreatic cancer risk were measured using conditional logistic regression. Results Carriers of the G allele (Ala) of the Pro12Ala variant had a borderline increased relative risk of pancreatic cancer compared to homozygous carriers of the C allele (Pro), with an odds ratio (OR) of 1.79 (95% confidence interval (CI): 0.96–3.33, p-value: 0.06). Among subjects randomized to high-dose vitamin A, the OR was 2.80 (95: CI: 1.16–6.74, p-value 0.02) versus 1.20 (95% CI: 0.45–3.23, p-value 0.71) in the placebo group. A haplotype including Pro12Ala was also significantly associated with pancreatic cancer risk in all subjects and in subjects randomized to vitamin A. Conclusions This analysis presents the first evidence that PPARG may be associated with pancreatic cancer risk, and this candidate gene should be investigated in future, larger studies. PMID:19436234

A double mutation in AGXT gene in families with primary hyperoxaluria type 1.

PubMed

Kanoun, Houda; Jarraya, Faiçal; Hadj Salem, Ikhlass; Mahfoudh, Hichem; Chaabouni, Yosr; Makni, Fatma; Hachicha, Jamil; Fakhfakh, Faiza

2013-12-01

Primary hyperoxaluria type 1 (PH1) is a severe autosomal recessive inherited disorder of glyoxylate metabolism caused by mutations in the AGXT gene on chromosome 2q37.3 that encodes the hepatic peroxisomal enzyme alanine:glyoxylate aminotransferase. These mutations are found throughout the entire gene and cause a wide spectrum of clinical severity. Rare in Europe, PH1 is responsible for 13% of the end stage renal failure in the Tunisian child. In the present work, we identified the double mutation c.32C>T (Pro11Leu) and c.731T>C (p.Ile244Thr) in AGXT gene in five unrelated Tunisian families with PH1 disease. Our results provide evidence regarding the potential involvement of c.32C>T, originally described as common polymorphism, on the resulting phenotype. We also reported an extreme intrafamilial heterogeneity in clinical presentation of PH1. Despite the same genetic background, the outcome of the affected members differs widely. The significant phenotypic heterogeneity observed within a same family, with a same genotype, suggests the existence of relevant modifier factors. © 2013.

Transcriptional up-regulation of nitric oxide synthase II by nuclear factor-kappaB at rostral ventrolateral medulla in a rat mevinphos intoxication model of brain stem death.

PubMed

Chan, Julie Y H; Wu, Carol H Y; Tsai, Ching-Yi; Cheng, Hsiao-Lei; Dai, Kuang-Yu; Chan, Samuel H H; Chang, Alice Y W

2007-06-15

As the origin of a 'life-and-death' signal that reflects central cardiovascular regulatory failure during brain stem death, the rostral ventrolateral medulla (RVLM) is a suitable neural substrate for mechanistic delineation of this vital phenomenon. Using a clinically relevant animal model that employed the organophosphate pesticide mevinphos (Mev) as the experimental insult, we evaluated the hypothesis that transcriptional up-regulation of nitric oxide synthase I or II (NOS I or II) gene expression by nuclear factor-kappaB (NF-kappaB) on activation of muscarinic receptors in the RVLM underlies brain stem death. In Sprague-Dawley rats maintained under propofol anaesthesia, co-microinjection of muscarinic M2R (methoctramine) or M4R (tropicamide), but not M1R (pirenzepine) or M3R (4-diphenylacetoxy-N-dimethylpiperidinium) antagonist significantly reduced the enhanced NOS I-protein kinase G signalling ('pro-life' phase) or augmented NOS II-peroxynitrite cascade ('pro-death' phase) in ventrolateral medulla, blunted the biphasic increase and decrease in baroreceptor reflex-mediated sympathetic vasomotor tone that reflect the transition from life to death, and diminished the elevated DNA binding activity or nucleus-bound translocation of NF-kappaB in RVLM neurons induced by microinjection of Mev into the bilateral RVLM. However, NF-kappaB inhibitors (diethyldithiocarbamate or pyrrolidine dithiocarbamate) or double-stranded kappaB decoy DNA preferentially antagonized the augmented NOS II-peroxynitrite cascade and the associated cardiovascular depression exhibited during the 'pro-death' phase. We conclude that transcriptional up-regulation of NOS II gene expression by activation of NF-kappaB on selective stimulation of muscarinic M2 or M4 subtype receptors in the RVLM underlies the elicited cardiovascular depression during the 'pro-death' phase in our Mev intoxication model of brain stem death.

GPER signalling in both cancer-associated fibroblasts and breast cancer cells mediates a feedforward IL1β/IL1R1 response

PubMed Central

De Marco, Paola; Lappano, Rosamaria; Francesco, Ernestina Marianna De; Cirillo, Francesca; Pupo, Marco; Avino, Silvia; Vivacqua, Adele; Abonante, Sergio; Picard, Didier; Maggiolini, Marcello

2016-01-01

Cancer-associated fibroblasts (CAFs) contribute to the malignant aggressiveness through secreted factors like IL1β, which may drive pro-tumorigenic inflammatory phenotypes mainly acting via the cognate receptor named IL1R1. Here, we demonstrate that signalling mediated by the G protein estrogen receptor (GPER) triggers IL1β and IL1R1 expression in CAFs and breast cancer cells, respectively. Thereby, ligand-activation of GPER generates a feedforward loop coupling IL1β induction by CAFs to IL1R1 expression by cancer cells, promoting the up-regulation of IL1β/IL1R1 target genes such as PTGES, COX2, RAGE and ABCG2. This regulatory interaction between the two cell types induces migration and invasive features in breast cancer cells including fibroblastoid cytoarchitecture and F-actin reorganization. A better understanding of the mechanisms involved in the regulation of pro-inflammatory cytokines by GPER-integrated estrogen signals may be useful to target these stroma-cancer interactions. PMID:27072893

c-Rel Deficiency Increases Caspase-4 Expression and Leads to ER Stress and Necrosis in EBV-Transformed Cells

PubMed Central

Valentín-Acevedo, Aníbal; Sinquett, Frank L.; Covey, Lori R.

2011-01-01

LMP1-mediated activation of nuclear factor of kappaB (NF-κB) is critical for the ligand independent proliferation and cell survival of in vitro EBV-transformed lymphoblastoid cell lines (LCLs). Previous experiments revealed that a majority of LMP1-dependent responses are regulated by NF-κB. However, the extent that individual NF-κB family members are required for these responses, in particular, c-Rel, whose expression is restricted to mature hematopoietic cells, remains unclear. Here we report that low c-Rel expression in LCLs derived from a patient with hyper-IgM syndrome (Pt1), resulted in defects in proliferation and cell survival. In contrast to studies that associated loss of NF-κB with increased apoptosis, Pt1 LCLs failed to initiate apoptosis and alternatively underwent autophagy and necrotic cell death. Whereas the proliferation defect appeared linked to a c-Rel-associated decrease in c-myc expression, identified pro-survival and pro-apoptotic targets were expressed at or near control levels consistent with the absence of apoptosis. Ultrastructural examination of Pt1 LCLs revealed a high level of cellular and ER stress that was further supported by gene expression profiling showing the upregulation of several genes involved in stress and inflammation. Apoptosis-independent cell death was accompanied by increased expression of the inflammatory marker, caspase-4. Using gene overexpression and siRNA knockdown we demonstrated that levels of c-Rel directly modulated expression of caspase-4 as well as other ER stress genes. Overall, these findings reveal the importance of c-Rel in maintaining LCL viability and that decreased expression results in ER stress and a default response leading to necrotic cell death. PMID:21984918

Association of P53 gene polymorphism with gastric cancer in Northern Iran as a high-risk region.

PubMed

Hedayatizadeh-Omran, Akbar; Alizadeh-Navaei, Reza; Janbabaei, Ghasem; Omrani-Nava, Versa; Hasheminasab, Yahya; Amjadi, Omolbanin; Tehrani, Mohsen

2018-05-01

Gastric cancer has the fourth highest morbidity rate of all cancers worldwide. Genetic factors including alterations in oncogenes and tumor suppressor genes serve an important role in gastric cancer development and progression. The P53 gene acts as a tumor suppressor gene by regulating the cell cycle, DNA transcription and repair, apoptosis, senescence and genome stability. In addition to somatic P53 mutations in cancer development, germline polymorphisms are also involved in different malignancies. The polymorphism of P53 at codon 72 (Arg72Pro) is established as a common variant that increases susceptibility to various cancers. The present case-control study was conducted to evaluate the possible association between this P53 polymorphism and gastric cancer in the Iranian population. A total of 59 patients with gastric cancer and 59 healthy controls were enrolled in the present study. Genomic DNA was extracted from peripheral blood mononuclear cells and genotype analysis was performed using a polymerase chain reaction-based restriction fragment length polymorphism assay. Genotype frequencies did not differ significantly between the patients and controls (P=0.4); the frequencies of the three genotypes Arg/Arg, Arg/Pro and Pro/Pro in gastric cancer patients were 28.8, 49.2 and 22.0%, and in controls were 37.3, 49.2 and 13.6%. Additionally, there were no differences in genotype frequencies based on tumor location, histological differentiation or tumor stage. Based on these findings, it may be concluded that the P53 codon 72 polymorphism does not contribute to gastric cancer susceptibility in Northern Iran.

Role of Brain-Derived Neurotrophic Factor in Beneficial Effects of Repetitive Transcranial Magnetic Stimulation for Upper Limb Hemiparesis after Stroke.

PubMed

Niimi, Masachika; Hashimoto, Kenji; Kakuda, Wataru; Miyano, Satoshi; Momosaki, Ryo; Ishima, Tamaki; Abo, Masahiro

2016-01-01

Repetitive transcranial magnetic stimulation (rTMS) can improve upper limb hemiparesis after stroke but the mechanism underlying its efficacy remains elusive. rTMS seems to alter brain-derived neurotrophic factor (BDNF) and such effect is influenced by BDNF gene polymorphism. To investigate the molecular effects of rTMS on serum levels of BDNF, its precursor proBDNF and matrix metalloproteinase-9 (MMP-9) in poststroke patients with upper limb hemiparesis. Poststroke patients with upper limb hemiparesis were studied. Sixty-two patients underwent rehabilitation plus rTMS combination therapy and 33 patients underwent rehabilitation monotherapy without rTMS for 14 days at our hospital. One Hz rTMS was applied over the motor representation of the first dorsal interosseous muscle on the non-lesional hemisphere. Fugl-Meyer Assessment and Wolf Motor Function (WMFT) were used to evaluate motor function on the affected upper limb before and after intervention. Blood samples were collected for analysis of BDNF polymorphism and measurement of BDNF, proBDNF and MMP-9 levels. Two-week combination therapy increased BDNF and MMP-9 serum levels, but not serum proBDNF. Serum BDNF and MMP-9 levels did not correlate with motor function improvement, though baseline serum proBDNF levels correlated negatively and significantly with improvement in WMFT (ρ = -0.422, p = 0.002). The outcome of rTMS therapy was not altered by BDNF gene polymorphism. The combination therapy of rehabilitation plus low-frequency rTMS seems to improve motor function in the affected limb, by activating BDNF processing. BDNF and its precursor proBDNF could be potentially suitable biomarkers for poststroke motor recovery.

Oncogenic fusion proteins adopt the insulin-like growth factor signaling pathway.

PubMed

Werner, Haim; Meisel-Sharon, Shilhav; Bruchim, Ilan

2018-02-19

The insulin-like growth factor-1 receptor (IGF1R) has been identified as a potent anti-apoptotic, pro-survival tyrosine kinase-containing receptor. Overexpression of the IGF1R gene constitutes a typical feature of most human cancers. Consistent with these biological roles, cells expressing high levels of IGF1R are expected not to die, a quintessential feature of cancer cells. Tumor specific chromosomal translocations that disrupt the architecture of transcription factors are a common theme in carcinogenesis. Increasing evidence gathered over the past fifteen years demonstrate that this type of genomic rearrangements is common not only among pediatric and hematological malignancies, as classically thought, but may also provide a molecular and cytogenetic foundation for an ever-increasing portion of adult epithelial tumors. In this review article we provide evidence that the mechanism of action of oncogenic fusion proteins associated with both pediatric and adult malignancies involves transactivation of the IGF1R gene, with ensuing increases in IGF1R levels and ligand-mediated receptor phosphorylation. Disrupted transcription factors adopt the IGF1R signaling pathway and elicit their oncogenic activities via activation of this critical regulatory network. Combined targeting of oncogenic fusion proteins along with the IGF1R may constitute a promising therapeutic approach.

Estrogen Modulates Specific Life and Death Signals Induced by LH and hCG in Human Primary Granulosa Cells In Vitro.

PubMed

Casarini, Livio; Riccetti, Laura; De Pascali, Francesco; Gilioli, Lisa; Marino, Marco; Vecchi, Eugenia; Morini, Daria; Nicoli, Alessia; La Sala, Giovanni Battista; Simoni, Manuela

2017-04-28

Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) are glycoprotein hormones used for assisted reproduction acting on the same receptor (LHCGR) and mediating different intracellular signaling. We evaluated the pro- and anti-apoptotic effect of 100 pM LH or hCG, in the presence or in the absence of 200 pg/mL 17β-estradiol, in long-term, serum-starved human primary granulosa cells (hGLC) and a transfected granulosa cell line overexpressing LHCGR (hGL5/LHCGR). To this purpose, phospho-extracellular-regulated kinase 1/2 (pERK1/2), protein kinase B (pAKT), cAMP-responsive element binding protein (pCREB) activation and procaspase 3 cleavage were evaluated over three days by Western blotting, along with the expression of target genes by real-time PCR and cell viability by colorimetric assay. We found that LH induced predominant pERK1/2 and pAKT activation STARD1 , CCND2 and anti-apoptotic XIAP gene expression, while hCG mediated more potent CREB phosphorylation, expression of CYP19A1 and procaspase 3 cleavage than LH. Cell treatment by LH is accompanied by increased (serum-starved) cell viability, while hCG decreased the number of viable cells. The hCG-specific, pro-apoptotic effect was blocked by a physiological dose of 17β-estradiol, resulting in pAKT activation, lack of procaspase 3 cleavage and increased cell viability. These results confirm that relatively high levels of steroidogenic pathway activation are linked to pro-apoptotic signals in vitro, which may be counteracted by other factors, i.e., estrogens.

P5CDH affects the pathways contributing to Pro synthesis after ProDH activation by biotic and abiotic stress conditions

PubMed Central

Rizzi, Yanina S.; Monteoliva, Mariela I.; Fabro, Georgina; Grosso, Carola L.; Laróvere, Laura E.; Alvarez, María E.

2015-01-01

Plants facing adverse conditions usually alter proline (Pro) metabolism, generating changes that help restore the cellular homeostasis. These organisms synthesize Pro from glutamate (Glu) or ornithine (Orn) by two-step reactions that share Δ1 pyrroline-5-carboxylate (P5C) as intermediate. In the catabolic process, Pro is converted back to Glu using a different pathway that involves Pro dehydrogenase (ProDH), P5C dehydrogenase (P5CDH), and P5C as intermediate. Little is known about the coordination of the catabolic and biosynthetic routes under stress. To address this issue, we analyzed how P5CDH affects the activation of Pro synthesis, in Arabidopsis tissues that increase ProDH activity by transient exposure to exogenous Pro, or infection with Pseudomonas syringae pv. tomato. Wild-type (Col-0) and p5cdh mutant plants subjected to these treatments were used to monitor the Pro, Glu, and Orn levels, as well as the expression of genes from Pro metabolism. Col-0 and p5cdh tissues consecutively activated ProDH and Pro biosynthetic genes under both conditions. However, they manifested a different coordination between these routes. When external Pro supply was interrupted, wild-type leaves degraded Pro to basal levels at which point Pro synthesis, mainly via Glu, became activated. Under the same condition, p5cdh leaves sustained ProDH induction without reducing the Pro content but rather increasing it, apparently by stimulating the Orn pathway. In response to pathogen infection, both genotypes showed similar trends. While Col-0 plants seemed to induce both Pro biosynthetic routes, p5cdh mutant plants may primarily activate the Orn route. Our study contributes to the functional characterization of P5CDH in biotic and abiotic stress conditions, by revealing its capacity to modulate the fate of P5C, and prevalence of Orn or Glu as Pro precursors in tissues that initially consumed Pro. PMID:26284090

Altered processing of the neurotensin/neuromedin N precursor in PC2 knock down mice: a biochemical and immunohistochemical study.

PubMed

Villeneuve, Pierre; Feliciangeli, Sylvain; Croissandeau, Gilles; Seidah, Nabil G; Mbikay, Majambu; Kitabgi, Patrick; Beaudet, Alain

2002-08-01

Neurotensin (NT) and neuromedin N (NN) are generated by endoproteolytic cleavage of a common precursor molecule, pro-NT/NN. To gain insight into the role of prohormone convertases PC1, PC2, and PC7 in this process, we investigated the maturation of pro-NT/NN in the brain of PC7 (PC7-/-), PC2 (PC2-/-), and/or PC1 (PC1+/- and PC2-/-; PC1+/-) knock down mice. Inactivation of the PC7 gene was without effect, suggesting that this convertase is not involved in the processing of pro-NT/NN. By contrast, there was a 15% decrease in NT and a 50% decrease in NN levels, as measured by radioimmunoassay, in whole brain extracts from PC2 null as compared with wild type mice. Using immunohistochemistry, we found that this decrease in pro-NT/NN maturation products was uneven and that it was most pronounced in the medial preoptic area, lateral hypothalamus, and paraventricular hypothalamic nuclei. These results suggest that PC2 plays a critical role in the processing of pro-NT/NN in mouse brain and that its deficiency may be compensated to a regionally variable extent by other convertases. Previous data have suggested that PC1 might be subserving this role. However, there was no change in the maturation of pro-NT/NN in the brain of mice in which the PC1 gene had been partially inactivated, implying that complete PC1 knock down may be required for loss of function.

Sphingosine-1-Phosphate Signaling and Metabolism Gene Signature in Pediatric Inflammatory Bowel Disease: A Matched-case Control Pilot Study

PubMed Central

Suh, Jung H.; Degagné, Émilie; Gleghorn, Elizabeth E.; Setty, Mala; Rodriguez, Alexis; Park, K. T.; Verstraete, Sofia G.; Heyman, Melvin B.; Patel, Ashish S.; Irek, Melissa; Gildengorin, Ginny L.; Hubbard, Neil E.; Borowsky, Alexander D.; Saba, Julie D.

2018-01-01

Goal The aim of this study was to investigate gene expression levels of proteins involved in sphingosine-1-phosphate (S1P) metabolism and signaling in a pediatric inflammatory bowel disease (IBD) patient population. Background IBD is a debilitating disease affecting 0.4% of the US population. The incidence of IBD in childhood is rising. Identifying effective targeted therapies that can be used safely in young patients and developing tools for selecting specific candidates for targeted therapies are important goals. Clinical IBD trials now underway target S1PR1, a receptor for the pro-inflammatory sphingolipid S1P. However, circulating and tissue sphingolipid levels and S1P-related gene expression have not been characterized in pediatric IBD. Methods Pediatric IBD patients and controls were recruited in a four-site study. Patients received a clinical score using PUCAI or PCDAI evaluation. Colon biopsies were collected during endoscopy. Gene expression was measured by qRT-PCR. Plasma and gut tissue sphingolipids were measured by LC-MS/MS. Results Genes of S1P synthesis (SPHK1, SPHK2), degradation (SGPL1), and signaling (S1PR1, S1PR2, and S1PR4) were significantly upregulated in colon biopsies of IBD patients with moderate/severe symptoms compared with controls or patients in remission. Tissue ceramide, dihydroceramide, and ceramide-1-phosphate (C1P) levels were significantly elevated in IBD patients compared with controls. Conclusions A signature of elevated S1P-related gene expression in colon tissues of pediatric IBD patients correlates with active disease and normalizes in remission. Biopsied gut tissue from symptomatic IBD patients contains high levels of pro-apoptotic and pro-inflammatory sphingolipids. A combined analysis of gut tissue sphingolipid profiles with this S1P-related gene signature may be useful for monitoring response to conventional therapy. PMID:29788359

Isothiocyanate-enriched moringa seed extract alleviates ulcerative colitis symptoms in mice

PubMed Central

Wu, Alex G.; Jaja-Chimedza, Asha; Graf, Brittany L.; Waterman, Carrie; Verzi, Michael P.; Raskin, Ilya

2017-01-01

Moringa (Moringa oleifera Lam.) seed extract (MSE) has anti-inflammatory and antioxidant activities. We investigated the effects of MSE enriched in moringa isothiocyanate-1 (MIC-1), its putative bioactive, on ulcerative colitis (UC) and its anti-inflammatory/antioxidant mechanism likely mediated through Nrf2-signaling pathway. Dextran sulfate sodium (DSS)-induced acute (n = 8/group; 3% DSS for 5 d) and chronic (n = 6/group; cyclic rotations of 2.5% DSS/water for 30 d) UC was induced in mice that were assigned to 4 experimental groups: healthy control (water/vehicle), disease control (DSS/vehicle), MSE treatment (DSS/MSE), or 5-aminosalicyic acid (5-ASA) treatment (positive control; DSS/5-ASA). Following UC induction, water (vehicle), 150 mg/kg MSE, or 50 mg/kg 5-ASA were orally administered for 1 or 2 wks. Disease activity index (DAI), spleen/colon sizes, and colonic histopathology were measured. From colon and/or fecal samples, pro-inflammatory biomarkers, tight-junction proteins, and Nrf2-mediated enzymes were analyzed at protein and/or gene expression levels. Compared to disease control, MSE decreased DAI scores, and showed an increase in colon lengths and decrease in colon weight/length ratios in both UC models. MSE also reduced colonic inflammation/damage and histopathological scores (modestly) in acute UC. MSE decreased colonic secretions of pro-inflammatory keratinocyte-derived cytokine (KC), tumor necrosis factor (TNF)-α, nitric oxide (NO), and myeloperoxidase (MPO) in acute and chronic UC; reduced fecal lipocalin-2 in acute UC; downregulated gene expression of pro-inflammatory interleukin (IL)-1, IL-6, TNF-α, and inducible nitric oxide synthase (iNOS) in acute UC; upregulated expression of claudin-1 and ZO-1 in acute and chronic UC; and upregulated GSTP1, an Nrf2-mediated phase II detoxifying enzyme, in chronic UC. MSE was effective in mitigating UC symptoms and reducing UC-induced colonic pathologies, likely by suppressing pro-inflammatory biomarkers and increasing tight-junction proteins. This effect is consistent with Nrf2-mediated anti-inflammatory/antioxidant signaling pathway documented for other isothiocyanates similar to MIC-1. Therefore, MSE, enriched with MIC-1, may be useful in prevention and treatment of UC. PMID:28922365

Isothiocyanate-enriched moringa seed extract alleviates ulcerative colitis symptoms in mice.

PubMed

Kim, Youjin; Wu, Alex G; Jaja-Chimedza, Asha; Graf, Brittany L; Waterman, Carrie; Verzi, Michael P; Raskin, Ilya

2017-01-01

Moringa (Moringa oleifera Lam.) seed extract (MSE) has anti-inflammatory and antioxidant activities. We investigated the effects of MSE enriched in moringa isothiocyanate-1 (MIC-1), its putative bioactive, on ulcerative colitis (UC) and its anti-inflammatory/antioxidant mechanism likely mediated through Nrf2-signaling pathway. Dextran sulfate sodium (DSS)-induced acute (n = 8/group; 3% DSS for 5 d) and chronic (n = 6/group; cyclic rotations of 2.5% DSS/water for 30 d) UC was induced in mice that were assigned to 4 experimental groups: healthy control (water/vehicle), disease control (DSS/vehicle), MSE treatment (DSS/MSE), or 5-aminosalicyic acid (5-ASA) treatment (positive control; DSS/5-ASA). Following UC induction, water (vehicle), 150 mg/kg MSE, or 50 mg/kg 5-ASA were orally administered for 1 or 2 wks. Disease activity index (DAI), spleen/colon sizes, and colonic histopathology were measured. From colon and/or fecal samples, pro-inflammatory biomarkers, tight-junction proteins, and Nrf2-mediated enzymes were analyzed at protein and/or gene expression levels. Compared to disease control, MSE decreased DAI scores, and showed an increase in colon lengths and decrease in colon weight/length ratios in both UC models. MSE also reduced colonic inflammation/damage and histopathological scores (modestly) in acute UC. MSE decreased colonic secretions of pro-inflammatory keratinocyte-derived cytokine (KC), tumor necrosis factor (TNF)-α, nitric oxide (NO), and myeloperoxidase (MPO) in acute and chronic UC; reduced fecal lipocalin-2 in acute UC; downregulated gene expression of pro-inflammatory interleukin (IL)-1, IL-6, TNF-α, and inducible nitric oxide synthase (iNOS) in acute UC; upregulated expression of claudin-1 and ZO-1 in acute and chronic UC; and upregulated GSTP1, an Nrf2-mediated phase II detoxifying enzyme, in chronic UC. MSE was effective in mitigating UC symptoms and reducing UC-induced colonic pathologies, likely by suppressing pro-inflammatory biomarkers and increasing tight-junction proteins. This effect is consistent with Nrf2-mediated anti-inflammatory/antioxidant signaling pathway documented for other isothiocyanates similar to MIC-1. Therefore, MSE, enriched with MIC-1, may be useful in prevention and treatment of UC.

Rapid Activation of Bone Morphogenic Protein 9 by Receptor-mediated Displacement of Pro-domains*

PubMed Central

Kienast, Yvonne; Jucknischke, Ute; Scheiblich, Stefan; Thier, Martina; de Wouters, Mariana; Haas, Alexander; Lehmann, Christian; Brand, Verena; Bernicke, Dirk; Honold, Konrad; Lorenz, Stefan

2016-01-01

By non-covalent association after proteolytic cleavage, the pro-domains modulate the activities of the mature growth factor domains across the transforming growth factor-β family. In the case of bone morphogenic protein 9 (BMP9), however, the pro-domains do not inhibit the bioactivity of the growth factor, and the BMP9·pro-domain complexes have equivalent biological activities as the BMP9 mature ligand dimers. By using real-time surface plasmon resonance, we could demonstrate that either binding of pro-domain-complexed BMP9 to type I receptor activin receptor-like kinase 1 (ALK1), type II receptors, co-receptor endoglin, or to mature BMP9 domain targeting antibodies leads to immediate and complete displacement of the pro-domains from the complex. Vice versa, pro-domain binding by an anti-pro-domain antibody results in release of the mature BMP9 growth factor. Based on these findings, we adjusted ELISA assays to measure the protein levels of different BMP9 variants. Although mature BMP9 and inactive precursor BMP9 protein were directly detectable by ELISA, BMP9·pro-domain complex could only be measured indirectly as dissociated fragments due to displacement of mature growth factor and pro-domains after antibody binding. Our studies provide a model in which BMP9 can be readily activated upon getting into contact with its receptors. This increases the understanding of the underlying biology of BMP9 activation and also provides guidance for ELISA development for the detection of circulating BMP9 variants. PMID:26677222

Anti-inflammatory effects of conjugated linoleic acid isomers and essential fatty acids in bovine mammary epithelial cells.

PubMed

Dipasquale, D; Basiricò, L; Morera, P; Primi, R; Tröscher, A; Bernabucci, U

2018-01-09

Fatty acids are important modulators of inflammatory responses, in particular, n-3 and n-6 essential fatty acids and CLA have received particular attention for their ability to modulate inflammation. The objectives of this study were to compare the effects of CLA and essential fatty acids on the expression of pro and anti- inflammatory cytokines and their protective efficacy against inflammatory status in mammary gland by an in vitro model based on bovine mammary epithelial cells (BME-UV1). Bovine mammary epithelial cells were treated with complete medium containing either 50 µM of cis-9, trans-11 CLA (c9,t11 CLA) or trans-10, cis-12 CLA (t10,c12 CLA) or (α)-linolenic acid (aLnA) or (γ)-linolenic acid (gLnA) or linoleic acid (LA). After 48 h by fatty acids administration the cells were treated for 3 h with 20 µM of lipopolysaccharide (LPS) to induce inflammatory stimulus. Reactive oxygen species (ROS) production after treatments was assessed to verify and to compare the potential protection of different fatty acids against LPS-induced oxidative stress. The messenger RNA abundance of bovine pro and anti-inflammatory cytokines (tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6) and interleukine-10 (IL-10)) and peroxisome proliferator receptor-α/γ (PPARγ/α) were determined in BME-UV1 by real-time PCR. The results showed that cells treated with fatty acids and LPS increased ROS production compared with control cells. Among treatments, cells treated with c9,t11 CLA and t10,c12 CLA isomers revealed significant lower levels of ROS production compared with other fatty acids. All fatty acids reduced the gene expression of pro- and anti-inflammatory cytokines. Among fatty acids, t10,c12 CLA, LA and gLnA showed an homogeneous reduction of the three pro-inflammatory cytokines and this may correspond to more balanced and efficient physiological activity and may trigger a better protective effect. The PPARγ gene expression was significantly greater in cells treated with t10,c12 CLA, aLnA and LA, whereas the PPARα gene expression levels were significantly lower in cells treated with all different fatty acids, compared with the control. These results suggest that fatty acids inhibited the transcription of pro-inflammatory cytokines by the upregulation of PPARγ expression.

Improvements in glucose homeostasis in response to regular exercise are influenced by the PPARG Pro12Ala variant: results from the HERITAGE Family Study.

PubMed

Ruchat, S-M; Rankinen, T; Weisnagel, S J; Rice, T; Rao, D C; Bergman, R N; Bouchard, C; Pérusse, L

2010-04-01

Exercise training improves glucose homeostasis, but large inter-individual differences are reported, suggesting a role of genetic factors. We investigated whether variants either confirmed or newly identified as diabetes susceptibility variants through genome-wide association studies (GWAS) modulate changes in phenotypes derived from an IVGTT in response to an endurance training programme. We analysed eight polymorphisms in seven type 2 diabetes genes (CDKAL1 rs7756992; CDKN2A and CDKN2B rs10811661 and rs564398; HHEX rs7923837; IGF2BP2 rs4402960; KCNJ11 rs5215; PPARG rs1801282; and TCF7L2 rs7903146) in a maximum of 481 sedentary, non-diabetic white individuals, who participated in a 20-week endurance training programme. Associations were tested between the variants and changes in IVGTT-derived phenotypes. The only evidence of association with training response was found with PPARG rs1801282 (Pro12Ala). We observed that Ala carriers experienced greater increase in overall glucose tolerance (Deltaglucose disappearance index Ala/Ala 0.22 +/- 0.22, Pro/Ala 0.14 +/- 0.06, Pro/Pro 0.004 +/- 0.03; p = 0.0008), glucose effectiveness (Ala/Ala 0.28 +/- 0.41, Pro/Ala 0.44 +/- 0.14, Pro/Pro 0.09 +/- 0.06; p = 0.004), acute insulin response to glucose (Ala/Ala 64.21 +/- 37.73, Pro/Ala -11.92 +/- 40.30, Pro/Pro -46.30 +/- 14.70; p = 0.03) and disposition index (Ala/Ala 551.8 +/- 448.5, Pro/Ala 534.6 +/- 218.3, Pro/Pro -7.44 +/- 88.18; p = 0.003). Compared with Pro/Pro individuals, PPARG Ala carriers experienced greater improvements in glucose and insulin metabolism in response to regular endurance training. However, we did not find evidence of association between type 2 diabetes susceptibility variants recently identified through GWAS and glucose homeostasis response to exercise. Our results extend those of previous studies showing that Ala carriers appear to be more responsive to beneficial health effects of lifestyle interventions.

Ionizing Radiation Stimulates Expression of Pro-Osteoclastogenic Genes in Marrow and Skeletal Tissue

NASA Technical Reports Server (NTRS)

Alwood, J. S.; Shahnazari, M.; Chicana, B.; Schreurs, A. S.; Kumar, A.; Bartolini, A.; Shirazi-Fard, Y.; Globus, R. K.

2015-01-01

Exposure to ionizing radiation can cause rapid mineral loss and increase bone-resorbing osteoclasts within metabolically-active, cancellous-bone tissue leading to structural deficits. To better understand mechanisms involved in rapid, radiation-induced bone loss, we determined the influence of total-body irradiation on expression of select cytokines known both to stimulate osteoclastogenesis and contribute to inflammatory bone disease. Adult (16wk), male C57BL/6J mice were exposed to either 2Gy gamma rays (137Cs, 0.8Gy/min) or heavy ions (56Fe, 600MeV, 0.50-1.1Gy/min); this dose corresponds to either a single fraction of radiotherapy (typical total dose is =10Gy) or accumulates over long-duration, interplanetary missions. Serum, marrow, and mineralized tissue were harvested 4hrs-7d later. Gamma irradiation caused a prompt (2.6-fold within 4hrs) and persistent (peaking at 4.1-fold within 1d) rise in the expression of the obligate osteoclastogenic cytokine, receptor activator of nuclear factor kappaB-ligand (Rankl) within marrow cells over controls. Similarly, Rankl expression peaked in marrow cells within 3d of iron exposure (9.2-fold). Changes in Rankl expression induced by gamma irradiation preceded and overlapped with a rise in expression of other pro-osteoclastic cytokines in marrow (e.g., monocyte chemotactic protein-1 increased 11.9-fold, tumor necrosis factor-alpha increased 1.7- fold over controls). Marrow expression of the RANKL decoy receptor, osteoprotegerin (Opg), also rose after irradiation (11.3-fold). The ratio Rankl/Opg in marrow was increased 1.8-fold, a net pro-resorption balance. As expected, radiation increased a serum marker of resorption (tartrate resistant acid phosphatase) and led to cancellous bone loss (16% decrease in bone volume/total volume) through reduced trabecular struts. We conclude that total-body irradiation (gamma or heavy-ion) caused temporal, concerted regulation of gene expression within marrow and mineralized tissue for select cytokines which are responsible for osteoclastogenesis and elevated resorption; this is likely to account for rapid and progressive 52 deterioration of cancellous microarchitecture following exposure to ionizing radiation.

Association of PPARγ2 gene variant Pro12Ala polymorphism with hypertension and obesity in the aboriginal Qatari population known for being consanguineous

PubMed Central

Bener, Abdulbari; Darwish, Sarah; Al-Hamaq, Abdulla OAA; Mohammad, Ramzi M; Yousafzai, Mohammad T

2013-01-01

Aim The aim of this study was to investigate the association of the Pro12Ala polymorphism of the human peroxisome proliferator-activated receptor gamma 2 (PPARγ2) gene with hypertension and obesity in a highly consanguineous aboriginal Qatari population. Design A cross-sectional survey conducted from January 2011–December 2012. Setting Primary health care clinics. Subjects A random sample of 1,528 Qatari male and female population older than 20 years of age. Materials and methods Data on age, sex, income, level of education, occupation status, body mass index, and blood pressure and lipid profile were obtained. The Pro12Ala in the PPARγ2 gene was detected on the LightCycler® using two specific probes: (Sensor [G] 5′-CTC CTA TTG ACG CAG AAA GCG-FL and PPAR Anchor 5′ LC Red 640- TCC TTC ACT GAT ACA CTG TCT GCA AAC ATA TC-PH). Univariate and multivariate logistic regression were performed. Result Out of a total 1,528 participants, 220 were diagnosed with essential hypertension, and 420 were obese. Participants with consanguinity were significantly higher among hypertensive than normotensive (41.9% versus 30.8%; P=0.001). Altogether, more than three-fourths (89%) of the participants had a wild genotype (Pro12Pro), 9.8% were heterozygous with Pro12Ala, and only 1.2% was homozygous with the Ala12Ala genotype. The frequency of the Pro allele was 94.5% in normotensive versus 90.5% in hypertensive, while the distribution of the Ala allele was 5.5% in normotensive versus 9.5% in the hypertensive group (P=0.001). The odds of hypertension were 1.7 times higher among the participants with the Ala allele as compared to those with the Pro, while adjusting for other potential confounders (adjusted odds ratio 1.69; 95% confidence interval 1.12–2.55; P=0.012). There was no association between the PPARγ2Ala allele and obesity (P=0.740). Conclusion The current study revealed an association between the PPARγ2Ala allele and hypertension in Qatar’s population. On the other hand, this study found no association between the Ala allele and obesity. PMID:24187509

A HIF-LIMD1 negative feedback mechanism mitigates the pro-tumorigenic effects of hypoxia.

PubMed

Foxler, Daniel E; Bridge, Katherine S; Foster, John G; Grevitt, Paul; Curry, Sean; Shah, Kunal M; Davidson, Kathryn M; Nagano, Ai; Gadaleta, Emanuela; Rhys, Hefin I; Kennedy, Paul T; Hermida, Miguel A; Chang, Ting-Yu; Shaw, Peter E; Reynolds, Louise E; McKay, Tristan R; Wang, Hsei-Wei; Ribeiro, Paulo S; Plevin, Michael J; Lagos, Dimitris; Lemoine, Nicholas R; Rajan, Prabhakar; Graham, Trevor A; Chelala, Claude; Hodivala-Dilke, Kairbaan M; Spendlove, Ian; Sharp, Tyson V

2018-06-21

The adaptive cellular response to low oxygen tensions is mediated by the hypoxia-inducible factors (HIFs), a family of heterodimeric transcription factors composed of HIF-α and HIF-β subunits. Prolonged HIF expression is a key contributor to cellular transformation, tumorigenesis and metastasis. As such, HIF degradation under hypoxic conditions is an essential homeostatic and tumour-suppressive mechanism. LIMD1 complexes with PHD2 and VHL in physiological oxygen levels (normoxia) to facilitate proteasomal degradation of the HIF-α subunit. Here, we identify LIMD1 as a HIF-1 target gene, which mediates a previously uncharacterised, negative regulatory feedback mechanism for hypoxic HIF-α degradation by modulating PHD2-LIMD1-VHL complex formation. Hypoxic induction of LIMD1 expression results in increased HIF-α protein degradation, inhibiting HIF-1 target gene expression, tumour growth and vascularisation. Furthermore, we report that copy number variation at the LIMD1 locus occurs in 47.1% of lung adenocarcinoma patients, correlates with enhanced expression of a HIF target gene signature and is a negative prognostic indicator. Taken together, our data open a new field of research into the aetiology, diagnosis and prognosis of LIMD1 -negative lung cancers. © 2018 The Authors. Published under the terms of the CC BY 4.0 license.

Lactoferricin enhances BMP7-stimulated anabolic pathways in intervertebral disc cells.

PubMed

Ellman, Michael B; Kim, Jaesung; An, Howard S; Chen, Di; Kc, Ranjan; Li, Xin; Xiao, Guozhi; Yan, Dongyao; Suh, Joon; van Wjnen, Andre J; Wang, James H-C; Kim, Su-Gwan; Im, Hee-Jeong

2013-07-25

Bone-morphogenetic protein-7 (BMP7) is a well-known anabolic and anti-catabolic growth factor on intervertebral disc (IVD) matrix and cell homeostasis. Similarly, Lactoferricin B (LfcinB) has recently been shown to have pro-anabolic, anti-catabolic, anti-oxidative and/or anti-inflammatory effects in bovine disc cells in vitro. In this study, we investigated the potential benefits of using combined peptide therapy with LfcinB and BMP7 for intervertebral disc matrix repair and to understand cellular and signaling mechanisms controlled by these factors. We studied the effects of BMP7 and LfcinB as individual treatments and combined therapy on bovine nucleus pulposus (NP) cells by assessing proteoglycan (PG) accumulation and synthesis, and the gene expression of matrix protein aggrecan and transcription factor SOX-9. We also analyzed the role of Noggin, a BMP antagonist, in IVD tissue and examined its effect after stimulation with LfcinB. To understand the molecular mechanisms by which LfcinB synergizes with BMP7, we investigated the ERK-SP1 axis as a downstream intracellular signaling regulator involved in BMP7 and LfcinB-mediated activities. Treatment of bovine NP cells cultured in alginate with LfcinB plus BMP7 synergistically stimulates PG synthesis and accumulation in part by upregulation of aggrecan gene expression. The synergism results from LfcinB-mediated activation of Sp1 and SMAD signaling pathways by (i) phosphorylation of SMAD 1/5/8; (ii) downregulation of SMAD inhibitory factors [i.e., noggin and SMAD6 (inhibitory SMAD)]; and (iii) upregulation of SMAD4 (universal co-SMAD). These data indicate that LfcinB-suppression of Noggin may eliminate the negative feedback of BMP7, thereby maximizing biological activity of BMP7 and ultimately shifting homeostasis to a pro-anabolic state in disc cells. We propose that combination growth factor therapy using BMP7 and LfcinB may be beneficial for treatment of disc degeneration. Copyright © 2013 Elsevier B.V. All rights reserved.

DDE and PCB 153 independently induce aryl hydrocarbon receptor (AhR) expression in peripheral blood mononuclear cells.

PubMed

Gaspar-Ramírez, Octavio; Pérez-Vázquez, Francisco J; Salgado-Bustamante, Mariana; González-Amaro, Roberto; Hernandez-Castro, Berenice; Pérez-Maldonado, Ivan N

2015-01-01

Recent studies have demonstrated that compounds inducing pro-inflammatory cytokines enhance AhR expression. The aim of this study was 2-fold: (1) to determine if two pro-inflammatory compounds, dichlorodiphenyldichloroethylene (DDE) and 2,2',4,4',5,5'-hexa-chlorobiphenyl (PCB 153), independently affect AhR gene expression in peripheral blood mononuclear cells (PBMC); and (2) if affected, to determine whether the mechanism involved was due to AhR activation or to a pro-inflammatory effect of the chemicals. PBMC isolated from healthy individuals were incubated in the presence of DDE (10 µg/ml) and PCB 153 (20 ng/ml) over time and AhR and CYP1A1 expression was assessed with a real-time PCR technique. The results indicated there was over-expression of the AhR mRNA in PBMC when the cells were treated with DDE and PCB 153. No changes in expression levels of CYP1A1 mRNA were found. Importantly, when the cells were exposed to DDE and PCB 153 in the presence of an antagonist of tumor necrosis factor (TNF)-α, the over-expression of AhR was abolished; as expected, the expression of CYP1A1 was unaffected. In conclusion, these studies demonstrated for the first time an increment of AhR expression "in vitro" in PBMC treated with two pro-inflammatory environmental pollutants, DDE and PCB153. Moreover, the over-expression of AhR was dependent of TNFα induced by DDE and PCB 153 and was independent of AhR activation.

Mitogen- and Stress-Activated Kinase 1 (MSK1) Regulates Cigarette Smoke-Induced Histone Modifications on NF-κB-dependent Genes

PubMed Central

Sundar, Isaac K.; Chung, Sangwoon; Hwang, Jae-woong; Lapek, John D.; Bulger, Michael; Friedman, Alan E.; Yao, Hongwei; Davie, James R.; Rahman, Irfan

2012-01-01

Cigarette smoke (CS) causes sustained lung inflammation, which is an important event in the pathogenesis of chronic obstructive pulmonary disease (COPD). We have previously reported that IKKα (I kappaB kinase alpha) plays a key role in CS-induced pro-inflammatory gene transcription by chromatin modifications; however, the underlying role of downstream signaling kinase is not known. Mitogen- and stress-activated kinase 1 (MSK1) serves as a specific downstream NF-κB RelA/p65 kinase, mediating transcriptional activation of NF-κB-dependent pro-inflammatory genes. The role of MSK1 in nuclear signaling and chromatin modifications is not known, particularly in response to environmental stimuli. We hypothesized that MSK1 regulates chromatin modifications of pro-inflammatory gene promoters in response to CS. Here, we report that CS extract activates MSK1 in human lung epithelial (H292 and BEAS-2B) cell lines, human primary small airway epithelial cells (SAEC), and in mouse lung, resulting in phosphorylation of nuclear MSK1 (Thr581), phospho-acetylation of RelA/p65 at Ser276 and Lys310 respectively. This event was associated with phospho-acetylation of histone H3 (Ser10/Lys9) and acetylation of histone H4 (Lys12). MSK1 N- and C-terminal kinase-dead mutants, MSK1 siRNA-mediated knock-down in transiently transfected H292 cells, and MSK1 stable knock-down mouse embryonic fibroblasts significantly reduced CS extract-induced MSK1, NF-κB RelA/p65 activation, and posttranslational modifications of histones. CS extract/CS promotes the direct interaction of MSK1 with RelA/p65 and p300 in epithelial cells and in mouse lung. Furthermore, CS-mediated recruitment of MSK1 and its substrates to the promoters of NF-κB-dependent pro-inflammatory genes leads to transcriptional activation, as determined by chromatin immunoprecipitation. Thus, MSK1 is an important downstream kinase involved in CS-induced NF-κB activation and chromatin modifications, which have implications in pathogenesis of COPD. PMID:22312446

Validation of the LupusPRO version 1.8: an update to a disease-specific patient-reported outcome tool for systemic lupus erythematosus.

PubMed

Azizoddin, D R; Weinberg, S; Gandhi, N; Arora, S; Block, J A; Sequeira, W; Jolly, M

2018-04-01

Objectives LupusPRO has shown good measurement properties as a disease-specific patient-reported outcome tool in systemic lupus erythematosus (SLE). For the purpose of clinical trials, the version 1.7 (v1.7) domain of Pain-Vitality was separated into distinct Pain, Vitality and Sleep domains in v1.8, and the psychometric properties examined. Methods A total of 131 consecutive SLE patients were self-administered surveys assessing fatigue (FACIT, SF-36), pain (Pain Inventory, SF-36), insomnia (Insomnia Severity Index), emotional health (PHQ-9, SF-36) and quality of life (SF-36, LupusPRO) at routine care visits. Internal consistency reliability (ICR) for each domain was obtained using Cronbach's alpha. The convergent construct validity of LupusPRO domains with corresponding SF-36 domains or tools were tested using Spearman correlation. Varimax rotations were conducted to assess factor structures of the LupusPRO v1.8. Results Mean (SD) age was 40.04 (14.10) years. Scores from the LupusPRO-Sleep domain strongly correlated with insomnia scores, while LupusPRO-Vitality correlated strongly with fatigue (FACIT) and SF-36 vitality. The LupusPRO-Pain domain correlated strongly with pain (SF36 Bodily-Pain, Pain Inventory) scores. Similarly, the LupusPRO domains of Physical and Emotional Health had significant correlations with corresponding SF-36 domains. The ICR for HRQoL and non-HRQoL were 0.96 and 0.81. LupusPRO (domains HRQoL and QoL) scores correlated with disease activity. Principal component analysis included seven factor loadings presenting for the HRQOL subscales (combined Sleep, Vitality, and Pain), and three factors for the NHRQoL (Combined Coping and Social Support). Conclusions LupusPRO v1.8 (including its Sleep, Vitality, and Pain domains) has acceptable reliability and validity. Use of LupusPRO as an outcome measure in clinical trials would facilitate responsiveness assessment.

Anti-apoptotic A1 is not essential for lymphoma development in Eµ-Myc mice but helps sustain transplanted Eµ-Myc tumour cells.

PubMed

Mensink, Mark; Anstee, Natasha S; Robati, Mikara; Schenk, Robyn L; Herold, Marco J; Cory, Suzanne; Vandenberg, Cassandra J

2018-03-01

The transcription factor c-MYC regulates a multiplicity of genes involved in cellular growth, proliferation, metabolism and DNA damage response and its overexpression is a hallmark of many tumours. Since MYC promotes apoptosis under conditions of stress, such as limited availability of nutrients or cytokines, MYC-driven cells are very much dependent on signals that inhibit cell death. Stress signals trigger apoptosis via the pathway regulated by opposing fractions of the BCL-2 protein family and previous genetic studies have shown that the development of B lymphoid tumours in Eµ-Myc mice is critically dependent on expression of pro-survival BCL-2 relatives MCL-1, BCL-W and, to a lesser extent, BCL-X L , but not BCL-2 itself, and that sustained growth of these lymphomas is dependent on MCL-1. Using recently developed mice that lack expression of all three functional pro-survival A1 genes, we show here that the kinetics of lymphoma development in Eµ-Myc mice and the competitive repopulation capacity of Eµ-Myc haemopoietic stem and progenitor cells is unaffected by the absence of A1. However, conditional loss of a single remaining functional A1 gene from transplanted A1-a -/- A1-b fl/fl A1-c -/- Eµ-Myc lymphomas slowed their expansion, significantly extending the life of the transplant recipients. Thus, A1 contributes to the survival of malignant Eµ-Myc-driven B lymphoid cells. These results strengthen the case for BFL-1, the human homologue of A1, being a valid target for drug development for MYC-driven tumours.

Sulforaphane Inhibits Lipopolysaccharide-Induced Inflammation, Cytotoxicity, Oxidative Stress, and miR-155 Expression and Switches to Mox Phenotype through Activating Extracellular Signal-Regulated Kinase 1/2-Nuclear Factor Erythroid 2-Related Factor 2/Antioxidant Response Element Pathway in Murine Microglial Cells.

PubMed

Eren, Erden; Tufekci, Kemal Ugur; Isci, Kamer Burak; Tastan, Bora; Genc, Kursad; Genc, Sermin

2018-01-01

Sulforaphane (SFN) is a natural product with cytoprotective, anti-inflammatory, and antioxidant effects. In this study, we evaluated the mechanisms of its effects on lipopolysaccharide (LPS)-induced cell death, inflammation, oxidative stress, and polarization in murine microglia. We found that SFN protects N9 microglial cells upon LPS-induced cell death and suppresses LPS-induced levels of secreted pro-inflammatory cytokines, tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. SFN is also a potent inducer of redox sensitive transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), which is responsible for the transcription of antioxidant, cytoprotective, and anti-inflammatory genes. SFN induced translocation of Nrf2 to the nucleus via extracellular signal-regulated kinase 1/2 (ERK1/2) pathway activation. siRNA-mediated knockdown study showed that the effects of SFN on LPS-induced reactive oxygen species, reactive nitrogen species, and pro-inflammatory cytokine production and cell death are partly Nrf2 dependent. Mox phenotype is a novel microglial phenotype that has roles in oxidative stress responses. Our results suggested that SFN induced the Mox phenotype in murine microglia through Nrf2 pathway. SFN also alleviated LPS-induced expression of inflammatory microRNA, miR-155. Finally, SFN inhibits microglia-mediated neurotoxicity as demonstrated by conditioned medium and co-culture experiments. In conclusion, SFN exerts protective effects on microglia and modulates the microglial activation state.

Sulforaphane Inhibits Lipopolysaccharide-Induced Inflammation, Cytotoxicity, Oxidative Stress, and miR-155 Expression and Switches to Mox Phenotype through Activating Extracellular Signal-Regulated Kinase 1/2–Nuclear Factor Erythroid 2-Related Factor 2/Antioxidant Response Element Pathway in Murine Microglial Cells

PubMed Central

Eren, Erden; Tufekci, Kemal Ugur; Isci, Kamer Burak; Tastan, Bora; Genc, Kursad; Genc, Sermin

2018-01-01

Sulforaphane (SFN) is a natural product with cytoprotective, anti-inflammatory, and antioxidant effects. In this study, we evaluated the mechanisms of its effects on lipopolysaccharide (LPS)-induced cell death, inflammation, oxidative stress, and polarization in murine microglia. We found that SFN protects N9 microglial cells upon LPS-induced cell death and suppresses LPS-induced levels of secreted pro-inflammatory cytokines, tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. SFN is also a potent inducer of redox sensitive transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), which is responsible for the transcription of antioxidant, cytoprotective, and anti-inflammatory genes. SFN induced translocation of Nrf2 to the nucleus via extracellular signal-regulated kinase 1/2 (ERK1/2) pathway activation. siRNA-mediated knockdown study showed that the effects of SFN on LPS-induced reactive oxygen species, reactive nitrogen species, and pro-inflammatory cytokine production and cell death are partly Nrf2 dependent. Mox phenotype is a novel microglial phenotype that has roles in oxidative stress responses. Our results suggested that SFN induced the Mox phenotype in murine microglia through Nrf2 pathway. SFN also alleviated LPS-induced expression of inflammatory microRNA, miR-155. Finally, SFN inhibits microglia-mediated neurotoxicity as demonstrated by conditioned medium and co-culture experiments. In conclusion, SFN exerts protective effects on microglia and modulates the microglial activation state. PMID:29410668

Evaluation of glucose metabolism and reproductive hormones in polycystic ovary syndrome on the basis of peroxisome proliferator-activated receptor (PPAR)-gamma2 Pro12Ala genotype.

PubMed

Tok, E C; Aktas, A; Ertunc, D; Erdal, E M; Dilek, S

2005-06-01

Peroxisome proliferator-activated receptor (PPAR)-gamma2 Pro12Ala polymorphism has been suggested as a protective factor for polycystic ovary syndrome (PCOS). In this study, we aimed to investigate metabolic features and reproductive hormones in women with PCOS and compare these features with control women on the basis of Pro12Ala genotype. This study involved 60 randomly selected women with PCOS and 60 controls. Main outcome measures were anthropometric measures, variables of glucose metabolism and reproductive hormones. All the patients were genotyped for Pro12Ala variant of PPAR-gamma2 gene. Patients with Pro12Ala polymorphism were more obese in both groups. Furthermore, they had lower fasting insulin levels, were less insulin-resistant and were less glucose-intolerant as demonstrated by 2 h glucose concentrations. However, there was no difference in reproductive hormone levels on the basis of Pro12Ala genotype. Both control women and women with PCOS had significant differences in glucose metabolism on the basis of PPAR-gamma2 Pro12Ala polymorphism. Pro12Ala variant may break the process that leads to PCOS in susceptible women, instead of being a direct causal relationship between Pro12Ala polymorphism and PCOS.

A Violaxanthin Deepoxidase Interacts with a Viral Suppressor of RNA Silencing to Inhibit Virus Amplification1[OPEN

PubMed Central

Chen, Ling; Yan, Zhaoling; Cheng, Yuqin; Jiao, Zhiyuan; Sun, Biao

2017-01-01

RNA silencing plays a critical role against viral infection. To counteract this antiviral silencing, viruses have evolved various RNA silencing suppressors. Meanwhile, plants have evolved counter-counter defense strategies against RNA silencing suppression (RSS). In this study, the violaxanthin deepoxidase protein of maize (Zea mays), ZmVDE, was shown to interact specifically with the helper component-proteinase (HC-Pro; a viral RNA silencing suppressor) of Sugarcane mosaic virus (SCMV) via its mature protein region by yeast two-hybrid assay, which was confirmed by coimmunoprecipitation in Nicotiana benthamiana cells. It was demonstrated that amino acids 101 to 460 in HC-Pro and the amino acid glutamine-292 in ZmVDE mature protein were essential for this interaction. The mRNA levels of ZmVDE were down-regulated 75% to 65% at an early stage of SCMV infection. Interestingly, ZmVDE, which normally localized in the chloroplasts and cytoplasm, could relocalize to HC-Pro-containing aggregate bodies in the presence of HC-Pro alone or SCMV infection. In addition, ZmVDE could attenuate the RSS activity of HC-Pro in a specific protein interaction-dependent manner. Subsequently, transient silencing of the ZmVDE gene facilitated SCMV RNA and coat protein accumulation. Taken together, our results suggest that ZmVDE interacts with SCMV HC-Pro and attenuates its RSS activity, contributing to decreased SCMV accumulation. This study demonstrates that a host factor can be involved in secondary defense responses against viral infection in monocot plants. PMID:29021224

Proline with or without hydroxyproline influences collagen concentration and regulates prolyl 4-hydroxylase α (I) gene expression in juvenile turbo ( Scophthalmus maximus L.)

NASA Astrophysics Data System (ADS)

Zhang, Kaikai; Mai, Kangsen; Xu, Wei; Zhou, Huihui; Liufu, Zhiguo; Zhang, Yanjiao; Peng, Mo; Ai, Qinghui

2015-06-01

This study was conducted to investigate the effect of dietary proline (Pro), and Pro and hydroxyproline (Hyp) in combination on the growth performance, total Hyp and collagen concentrations of tissues, and prolyl 4-hydroxylase α(I) (P4H α(I)) gene expression in juvenile turbot feeding high plant protein diets. A diet containing 50% crude protein and 12% crude lipid was formulated as the basal and control, on which other two protein and lipid contents identical experimental diets were formulated by supplementing the basal with either 0.75% Pro (Pro-0.75) or 0.75% Pro and 0.75% Hyp (Pro+Hyp). Four groups of fish in indoor seawater recirculating systems, 35 individuals each, were fed twice a day to apparent satiation for 10 weeks. The results showed that dietary Pro and Hyp supplementation had no significant effect on growth performance and feed utilization of juvenile turbot (P > 0.05). Total Hyp and collagen concentrations in muscle were significantly increased when dietary Pro and Hyp increased (P <0.05), and fish fed diet Pro+Hyp showed significantly higher free Hyp content in plasma than those fed other diets (P <0.05). The expression of P4H a(I) gene in liver and muscle was significantly up regulated in fish fed diet Pro-0.75 in comparison with control (P <0.05); however the gene was significantly down regulated in fish fed diet Pro+Hyp in muscle in comparison with fish fed diet Pro-0.75 (P <0.05). It can be concluded that supplement of crystal L-Pro and L-Hyp to high plant protein diets did not show positive effects on growth performance of juvenile turbot, but enhanced total collagen concentrations in muscle.

Curcumin mediated suppression of nuclear factor-κB promotes chondrogenic differentiation of mesenchymal stem cells in a high-density co-culture microenvironment.

PubMed

Buhrmann, Constanze; Mobasheri, Ali; Matis, Ulrike; Shakibaei, Mehdi

2010-01-01

Osteoarthritis (OA) and rheumatoid arthritis (RA) are characterised by joint inflammation and cartilage degradation. Although mesenchymal stem cell (MSC)-like progenitors are resident in the superficial zone of articular cartilage, damaged tissue does not possess the capacity for regeneration. The high levels of pro-inflammatory cytokines present in OA/RA joints may impede the chondrogenic differentiation of these progenitors. Interleukin (IL)-1β activates the transcription factor nuclear factor-κB (NF-κB), which in turn activates proteins involved in matrix degradation, inflammation and apoptosis. Curcumin is a phytochemical capable of inhibiting IL-1β-induced activation of NF-κB and expression of apoptotic and pro-inflammatory genes in chondrocytes. Therefore, the aim of the present study was to evaluate the influence of curcumin on IL-1β-induced NF-κB signalling pathway in MSCs during chondrogenic differentiation. MSCs were either cultured in a ratio of 1:1 with primary chondrocytes in high-density culture or cultured alone in monolayer with/without curcumin and/or IL-1β. We demonstrate that although curcumin alone does not have chondrogenic effects on MSCs, it inhibits IL-1β-induced activation of NF-κB, activation of caspase-3 and cyclooxygenase-2 in MSCs time and concentration dependently, as it does in chondrocytes. In IL-1β stimulated co-cultures, four-hour pre-treatment with curcumin significantly enhanced the production of collagen type II, cartilage specific proteoglycans (CSPGs), β1-integrin, as well as activating MAPKinase signaling and suppressing caspase-3 and cyclooxygenase-2. Curcumin treatment may help establish a microenvironment in which the effects of pro-inflammatory cytokines are antagonized, thus facilitating chondrogenesis of MSC-like progenitor cells in vivo. This strategy may support the regeneration of articular cartilage.

Human IGF-I propeptide A promotes articular chondrocyte biosynthesis and employs glycosylation-dependent heparin binding.

PubMed

Shi, Shuiliang; Kelly, Brian J; Wang, Congrong; Klingler, Ken; Chan, Albert; Eckert, George J; Trippel, Stephen B

2018-03-01

Insulin-like growth factor I (IGF-I) is a key regulator of chondrogenesis, but its therapeutic application to articular cartilage damage is limited by rapid elimination from the repair site. The human IGF-I gene gives rise to three IGF-I propeptides (proIGF-IA, proIGF-IB and proIGF-IC) that are cleaved to create mature IGF-I. In this study, we elucidate the processing of IGF-I precursors by articular chondrocytes, and test the hypotheses that proIGF-I isoforms bind to heparin and regulate articular chondrocyte biosynthesis. Human IGF-I propeptides and mutants were overexpressed in bovine articular chondrocytes. IGF-I products were characterized by ELISA, western blot and FPLC using a heparin column. The biosynthetic activity of IGF-I products on articular chondrocytes was assayed for DNA and glycosaminoglycan that the cells produced. Secreted IGF-I propeptides stimulated articular chondrocyte biosynthetic activity to the same degree as mature IGF-I. Of the three IGF-I propeptides, only one, proIGF-IA, strongly bound to heparin. Interestingly, heparin binding of proIGF-IA depended on N-glycosylation at Asn92 in the EA peptide. To our knowledge, this is the first demonstration that N-glycosylation determines the binding of a heparin-binding protein to heparin. The biosynthetic and heparin binding abilities of proIGF-IA, coupled with its generation of IGF-I, suggest that proIGF-IA may have therapeutic value for articular cartilage repair. These data identify human pro-insulin-like growth factor IA as a bifunctional protein. Its combined ability to bind heparin and augment chondrocyte biosynthesis makes it a promising therapeutic agent for cartilage damage due to trauma and osteoarthritis. Copyright © 2017 Elsevier B.V. All rights reserved.

Overexpression of the Arabidopsis CBF3 Transcriptional Activator Mimics Multiple Biochemical Changes Associated with Cold Acclimation1

PubMed Central

Gilmour, Sarah J.; Sebolt, Audrey M.; Salazar, Maite P.; Everard, John D.; Thomashow, Michael F.

2000-01-01

We further investigated the role of the Arabidopsis CBF regulatory genes in cold acclimation, the process whereby certain plants increase in freezing tolerance upon exposure to low temperature. The CBF genes, which are rapidly induced in response to low temperature, encode transcriptional activators that control the expression of genes containing the C-repeat/dehydration responsive element DNA regulatory element in their promoters. Constitutive expression of either CBF1 or CBF3 (also known as DREB1b and DREB1a, respectively) in transgenic Arabidopsis plants has been shown to induce the expression of target COR (cold-regulated) genes and to enhance freezing tolerance in nonacclimated plants. Here we demonstrate that overexpression of CBF3 in Arabidopsis also increases the freezing tolerance of cold-acclimated plants. Moreover, we show that it results in multiple biochemical changes associated with cold acclimation: CBF3-expressing plants had elevated levels of proline (Pro) and total soluble sugars, including sucrose, raffinose, glucose, and fructose. Plants overexpressing CBF3 also had elevated P5CS transcript levels suggesting that the increase in Pro levels resulted, at least in part, from increased expression of the key Pro biosynthetic enzyme Δ1-pyrroline-5-carboxylate synthase. These results lead us to propose that CBF3 integrates the activation of multiple components of the cold acclimation response. PMID:11115899

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jin, Sun Hee; Choi, Dalwoong; Chun, Young-Jin

Keratinocytes are the major cellular components of human epidermis and play a key role in the modulating cutaneous inflammation and toxic responses. In human chronic skin diseases, the common skin inflammatory phenotypes like skin barrier disruption and epidermal hyperplasia are manifested in epidermal keratinocytes by interactions with T helper (Th) cells. To find a common gene expression signature of human keratinocytes in chronic skin diseases, we performed a whole genome microarray analysis on normal human epidermal keratinocytes (NHKs) treated with IFNγ, IL-4, IL-17A or IL-22, major cytokines from Th1, Th2, Th17 or Th22 cells, respectively. The microarray results showed thatmore » the four genes, IL-24, PDZK1IP1, H19 and filaggrin, had common expression profiles in NHKs exposed to Th cell cytokines. In addition, the acute phase pro-inflammatory cytokines, IL-1β, IL-6 and TNFα, also change the gene transcriptional profile of IL-24, PDZK1IP1, H19, and filaggrin in NHKs as those of Th cytokines. Therefore, the signature gene set, consisting of IL-24, PDZK1IP1, H19, and filaggrin, provides essential insights for understanding the process of cutaneous inflammation and toxic responses. We demonstrate that environmental toxic stressors, such as chemical irritants and ultraviolet irradiation stimulate the production of IL-24 in NHKs. IL-24 stimulates the JAK1-STAT3 and MAPK pathways in NHKs, and promotes the secretion of pro-inflammatory mediators IL-8, PGE2, and MMP-1. These results suggest that keratinocyte-derived IL-24 participates in the positive feedback regulation of epidermal inflammation in response to both endogenous and environmental toxic stressors. - Highlights: • Cutaneous inflammatory gene signature consists of PDZK1IP1, IL-24, H19 and filaggrin. • Pro-inflammatory cytokines increase IL-24 production in human keratinocytes. • Environmental toxic stressors increase IL-24 production in human keratinocytes. • IL-24 stimulates human keratinocytes to produce pro-inflammatory mediators. • IL-24 activates STAT3 and MAPK signaling pathways in human keratinocytes.« less

Effects of BDNF polymorphisms on antidepressant action.

PubMed

Tsai, Shih-Jen; Hong, Chen-Jee; Liou, Ying-Jay

2010-12-01

Evidence suggests that the down-regulation of the signaling pathway involving brain-derived neurotrophic factor (BDNF), a molecular element known to regulate neuronal plasticity and survival, plays an important role in the pathogenesis of major depression. The restoration of BDNF activity induced by antidepressant treatment has been implicated in the antidepressant therapeutic mechanism. Because there is variability among patients with major depressive disorder in terms of response to antidepressant treatment and since genetic factors may contribute to this inter-individual variability in antidepressant response, pharmacogenetic studies have tested the associations between genetic polymorphisms in candidate genes related to antidepressant therapeutic action. In human BDNF gene, there is a common functional polymorphism (Val66Met) in the pro-region of BDNF, which affects the intracellular trafficking of proBDNF. Because of the potentially important role of BDNF in the antidepressant mechanism, many pharmacogenetic studies have tested the association between this polymorphism and the antidepressant therapeutic response, but they have produced inconsistent results. A recent meta-analysis of eight studies, which included data from 1,115 subjects, suggested that the Val/Met carriers have increased antidepressant response in comparison to Val/Val homozygotes, particularly in the Asian population. The positive molecular heterosis effect (subjects heterozygous for a specific genetic polymorphism show a significantly greater effect) is compatible with animal studies showing that, although BDNF exerts an antidepressant effect, too much BDNF may have a detrimental effect on mood. Several recommendations are proposed for future antidepressant pharmacogenetic studies of BDNF, including the consideration of multiple polymorphisms and a haplotype approach, gene-gene interaction, a single antidepressant regimen, controlling for age and gender interactions, and pharmacogenetic effects on specific depressive symptom-clusters.

The Hippo pathway mediates inhibition of vascular smooth muscle cell proliferation by cAMP.

PubMed

Kimura, Tomomi E; Duggirala, Aparna; Smith, Madeleine C; White, Stephen; Sala-Newby, Graciela B; Newby, Andrew C; Bond, Mark

2016-01-01

Inhibition of vascular smooth muscle cell (VSMC) proliferation by intracellular cAMP prevents excessive neointima formation and hence angioplasty restenosis and vein-graft failure. These protective effects are mediated via actin-cytoskeleton remodelling and subsequent regulation of gene expression by mechanisms that are incompletely understood. Here we investigated the role of components of the growth-regulatory Hippo pathway, specifically the transcription factor TEAD and its co-factors YAP and TAZ in VSMC. Elevation of cAMP using forskolin, dibutyryl-cAMP or the physiological agonists, Cicaprost or adenosine, significantly increased phosphorylation and nuclear export YAP and TAZ and inhibited TEAD-luciferase report gene activity. Similar effects were obtained by inhibiting RhoA activity with C3-transferase, its downstream kinase, ROCK, with Y27632, or actin-polymerisation with Latrunculin-B. Conversely, expression of constitutively-active RhoA reversed the inhibitory effects of forskolin on TEAD-luciferase. Forskolin significantly inhibited the mRNA expression of the pro-mitogenic genes, CCN1, CTGF, c-MYC and TGFB2 and this was reversed by expression of constitutively-active YAP or TAZ phospho-mutants. Inhibition of YAP and TAZ function with RNAi or Verteporfin significantly reduced VSMC proliferation. Furthermore, the anti-mitogenic effects of forskolin were reversed by overexpression of constitutively-active YAP or TAZ. Taken together, these data demonstrate that cAMP-induced actin-cytoskeleton remodelling inhibits YAP/TAZ-TEAD dependent expression of pro-mitogenic genes in VSMC. This mechanism contributes novel insight into the anti-mitogenic effects of cAMP in VSMC and suggests a new target for intervention. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

The effect of hydrophilic titanium surface modification on macrophage inflammatory cytokine gene expression.

PubMed

Hamlet, Stephen; Alfarsi, Mohammed; George, Roy; Ivanovski, Saso

2012-05-01

Chemical modification of microrough titanium dental implants to produce a hydrophilic surface with increased wettability and improved surface energy has been demonstrated clinically to achieve superior bone wound healing and osseointegration compared to that achieved with a microrough titanium surface alone. As the recruitment of the necessary osseoinductive precursors involved in bone wound healing and osseointegration to the wound site is facilitated by the action of cytokines, this study sought to determine the in vitro effect of hydrophilic surface modification on the expression of pro-inflammatory cytokines from adherent macrophages. The surface topography and composition of the titanium surfaces was characterized by scanning electron microscopy and X-ray photoelectron spectroscopy. Macrophage attachment and proliferation was assessed using an MTT assay. The expression of 84 pro-inflammatory cytokines and chemokines by adherent RAW 264.7 cells, a murine leukaemic monocyte cell line, was assessed by PCR array after 24 h culture on either smooth polished, sand-blasted acid-etched (SLA) or hydrophilic-modified SLA (SLActive) titanium surfaces. Following 24 h culture on titanium, surface microroughness activated pro-inflammatory cytokine gene transcription in RAW 264.7 cells. Although there was no significant difference in the degree of cellular attachment or proliferation of RAW 264.7 cells to the different titanium surfaces, by 24 h the hydrophilic surface elicited a gene expression profile with significant down-regulation of the key pro-inflammatory cytokines Tnfα, IL-1α, IL-1β and the chemokine Ccl-2. Down-regulation of the expression of pro-inflammatory cytokine genes may thus modulate the inflammatory response and may facilitate the enhanced bone wound healing and osseointegration observed clinically using implants with a microrough hydrophilic surface. © 2011 John Wiley & Sons A/S.

Integrative analysis identifies targetable CREB1/FoxA1 transcriptional co-regulation as a predictor of prostate cancer recurrence.

PubMed

Sunkel, Benjamin; Wu, Dayong; Chen, Zhong; Wang, Chiou-Miin; Liu, Xiangtao; Ye, Zhenqing; Horning, Aaron M; Liu, Joseph; Mahalingam, Devalingam; Lopez-Nicora, Horacio; Lin, Chun-Lin; Goodfellow, Paul J; Clinton, Steven K; Jin, Victor X; Chen, Chun-Liang; Huang, Tim H-M; Wang, Qianben

2016-05-19

Identifying prostate cancer-driving transcription factors (TFs) in addition to the androgen receptor promises to improve our ability to effectively diagnose and treat this disease. We employed an integrative genomics analysis of master TFs CREB1 and FoxA1 in androgen-dependent prostate cancer (ADPC) and castration-resistant prostate cancer (CRPC) cell lines, primary prostate cancer tissues and circulating tumor cells (CTCs) to investigate their role in defining prostate cancer gene expression profiles. Combining genome-wide binding site and gene expression profiles we define CREB1 as a critical driver of pro-survival, cell cycle and metabolic transcription programs. We show that CREB1 and FoxA1 co-localize and mutually influence each other's binding to define disease-driving transcription profiles associated with advanced prostate cancer. Gene expression analysis in human prostate cancer samples found that CREB1/FoxA1 target gene panels predict prostate cancer recurrence. Finally, we showed that this signaling pathway is sensitive to compounds that inhibit the transcription co-regulatory factor MED1. These findings not only reveal a novel, global transcriptional co-regulatory function of CREB1 and FoxA1, but also suggest CREB1/FoxA1 signaling is a targetable driver of prostate cancer progression and serves as a biomarker of poor clinical outcomes. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Isolation of baker's yeast mutants with proline accumulation that showed enhanced tolerance to baking-associated stresses.

PubMed

Tsolmonbaatar, Ariunzaya; Hashida, Keisuke; Sugimoto, Yukiko; Watanabe, Daisuke; Furukawa, Shuhei; Takagi, Hiroshi

2016-12-05

During bread-making processes, yeast cells are exposed to baking-associated stresses such as freeze-thaw, air-drying, and high-sucrose concentrations. Previously, we reported that self-cloning diploid baker's yeast strains that accumulate proline retained higher-level fermentation abilities in both frozen and sweet doughs than the wild-type strain. Although self-cloning yeasts do not have to be treated as genetically modified yeasts, the conventional methods for breeding baker's yeasts are more acceptable to consumers than the use of self-cloning yeasts. In this study, we isolated mutants resistant to the proline analogue azetidine-2-carboxylate (AZC) derived from diploid baker's yeast of Saccharomyces cerevisiae. Some of the mutants accumulated a greater amount of intracellular proline, and among them, 5 mutants showed higher cell viability than that observed in the parent wild-type strain under freezing or high-sucrose stress conditions. Two of them carried novel mutations in the PRO1 gene encoding the Pro247Ser or Glu415Lys variant of γ-glutamyl kinase (GK), which is a key enzyme in proline biosynthesis in S. cerevisiae. Interestingly, we found that these mutations resulted in AZC resistance of yeast cells and desensitization to proline feedback inhibition of GK, leading to intracellular proline accumulation. Moreover, baker's yeast cells expressing the PRO1 P247S and PRO1 E415K gene were more tolerant to freezing stress than cells expressing the wild-type PRO1 gene. The approach described here could be a practical method for the breeding of proline-accumulating baker's yeasts with higher tolerance to baking-associated stresses. Copyright © 2016 Elsevier B.V. All rights reserved.

Pattern Recognition Protein Binds to Lipopolysaccharide and β-1,3-Glucan and Activates Shrimp Prophenoloxidase System*

PubMed Central

Amparyup, Piti; Sutthangkul, Jantiwan; Charoensapsri, Walaiporn; Tassanakajon, Anchalee

2012-01-01

The prophenoloxidase (proPO) system is activated upon recognition of pathogens by pattern recognition proteins (PRPs), including a lipopolysaccharide- and β-1,3-glucan-binding protein (LGBP). However, shrimp LGBPs that are involved in the proPO system have yet to be clarified. Here, we focus on characterizing the role of a Penaeus monodon LGBP (PmLGBP) in the proPO system. We found that PmLGBP transcripts are expressed primarily in the hemocytes and are increased at 24 h after pathogenic bacterium Vibrio harveyi challenge. The binding studies carried out using ELISA indicated that recombinant (r)PmLGBP binds to β-1,3-glucan and LPS with a dissociation constant of 6.86 × 10−7 m and 3.55 × 10−7 m, respectively. Furthermore, we found that rPmLGBP could enhance the phenoloxidase (PO) activity of hemocyte suspensions in the presence of LPS or β-1,3-glucan. Using dsRNA interference-mediated gene silencing assay, we further demonstrated that knockdown of PmLGBP in shrimp in vivo significantly decreased the PmLGBP transcript level but had no effect on the expression of the other immune genes tested, including shrimp antimicrobial peptides (AMPs). However, suppression of proPO expression down-regulated PmLGBP, proPO-activating enzyme (PmPPAE2), and AMPs (penaeidin and crustin). Such PmLGBP down-regulated shrimp showed significantly decreased total PO activity. We conclude that PmLGBP functions as a pattern recognition protein for LPS and β-1,3-glucan in the shrimp proPO activating system. PMID:22235126

Effects of pre- and pro-sequence of thaumatin on the secretion by Pichia pastoris.

PubMed

Ide, Nobuyuki; Masuda, Tetsuya; Kitabatake, Naofumi

2007-11-23

Thaumatin is a 22-kDa sweet-tasting protein containing eight disulfide bonds. When thaumatin is expressed in Pichia pastoris using the thaumatin cDNA fused with both the alpha-factor signal sequence and the Kex2 protease cleavage site from Saccharomyces cerevisiae, the N-terminal sequence of the secreted thaumatin molecule is not processed correctly. To examine the role of the thaumatin cDNA-encoded N-terminal pre-sequence and C-terminal pro-sequence on the processing of thaumatin and efficiency of thaumatin production in P. pastoris, four expression plasmids with different pre-sequence and pro-sequence were constructed and transformed into P. pastoris. The transformants containing pre-thaumatin gene that has the native plant signal, secreted thaumatin molecules in the medium. The N-terminal amino acid sequence of the secreted thaumatin molecule was processed correctly. The production yield of thaumatin was not affected by the C-terminal pro-sequence, and the pro-sequence was not processed in P. pastoris, indicating that pro-sequence is not necessary for thaumatin synthesis.

Endothelial pro-atherosclerotic response to extracellular diabetic-like environment: possible role of thioredoxin-interacting protein.

PubMed

Zitman-Gal, Tali; Green, Janice; Pasmanik-Chor, Metsada; Oron-Karni, Varda; Bernheim, Jacques

2010-07-01

BACKGROUND. High blood and tissue concentrations of glucose and advanced glycation end-products (AGEs) are thought to play an important role in the development of vascular diabetic complications. Therefore, the impact of extracellular AGEs and different glucose concentrations was evaluated by studying the gene expressions and the underlying cellular pathways involved in the development of inflammatory pro-atherosclerotic processes observed in cultured endothelial cells. METHODS. Fresh human umbilical vein cord endothelial cells (HUVEC) were treated in the presence of elevated extracellular glucose concentrations (5.5-28 mmol/l) with and without AGE-human serum albumin (HSA). Affymetrix GeneChip(R) Human Gene 1.0 ST arrays were used for gene expression analysis (total 20 chips). Genes of interest were further validated using real-time PCR and western blot techniques. RESULTS. Microarray analysis revealed significant changes in some gene expressions in the presence of the different stimuli, suggesting that different pathways are involved. Six genes were selected for validation as follows: thioredoxin-interacting protein (TXNIP), thioredoxin (TXN), nuclear factor of kappa B (NF-kappaB), interleukin 6 (IL6), interleukin 8 (IL8) and receptor of advanced glycation end-products (RAGE). Interestingly, it was found that the association of AGEs together with the highest pathophysiological concentration of glucose (28 mmol/l) diminished the expression of these specific genes, excluding TXN. CONCLUSIONS. In the present model that mimics a diabetic environment, the relatively short-term experimental conditions used showed an unexpected blunting action of AGEs in the presence of the highest glucose concentration (28 mmol/l). The interactive cellular pathways involved in these processes should be further investigated.

[Detection and prenatal diagnosis for RS1 gene mutations in two Chinese families with X-linked juvenile retinoschisis].

PubMed

Chu, Yan; Fang, Dong; Hou, Qiao-fang; Wang, Li-ya; Guo, Xi-rang; Wang, Ying-tai; Liao, Shi-xiu

2013-04-01

To identify potential mutations of retinoschisis 1 (RS1) gene responsible for X-linked retinoschisis (XLRS) in two Chinese families. The 6 exons and flanking intronic regions were analyzed with PCR and direct sequencing. Two RS1 mutations were identified in the two families, which included 1 frameshift mutation (c.573delG, p.Pro192fs) and 1 missense mutation (c.626G>A, p.Arg209His). Two RS1 mutations have been identified, among which Pro192fs mutation is discovered for the first time in Chinese population. Above results may enrich our understanding of the clinical manifestations of XLRS and facilitated early diagnosis and genetic counseling for the disease.

Inorganic arsenic represses interleukin-17A expression in human activated Th17 lymphocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morzadec, Claudie; Macoch, Mélinda; Robineau, Marc

2012-08-01

Trivalent inorganic arsenic [As(III)] is an efficient anticancer agent used to treat patients suffering from acute promyelocytic leukemia. Recently, experimental studies have clearly demonstrated that this metalloid can also cure lymphoproliferative and/or pro-inflammatory syndromes in different murine models of chronic immune-mediated diseases. T helper (Th) 1 and Th17 lymphocytes play a central role in development of these diseases, in mice and humans, especially by secreting the potent pro-inflammatory cytokine interferon-γ and IL-17A, respectively. As(III) impairs basic functions of human T cells but its ability to modulate secretion of pro-inflammatory cytokines by differentiated Th lymphocytes is unknown. In the present study,more » we demonstrate that As(III), used at concentrations clinically achievable in plasma of patients, has no effect on the secretion of interferon-γ from Th1 cells but almost totally blocks the expression and the release of IL-17A from human Th17 lymphocytes co-stimulated for five days with anti-CD3 and anti-CD28 antibodies, in the presence of differentiating cytokines. In addition, As(III) specifically reduces mRNA levels of the retinoic-related orphan receptor (ROR)C gene which encodes RORγt, a key transcription factor controlling optimal IL-17 expression in fully differentiated Th17 cells. The metalloid also blocks initial expression of IL-17 gene induced by the co-stimulation, probably in part by impairing activation of the JNK/c-Jun pathway. In conclusion, our results demonstrate that As(III) represses expression of the major pro-inflammatory cytokine IL-17A produced by human Th17 lymphocytes, thus strengthening the idea that As(III) may be useful to treat inflammatory immune-mediated diseases in humans. -- Highlights: ► Arsenic inhibits secretion of IL-17A from human naïve and memory Th17 lymphocytes. ► Arsenic represses early expression of IL-17A gene in human activated T lymphocytes. ► Arsenic interferes with activation of the JNK/c-Jun pathway in human T lymphocytes.« less

Association between the Pro12Ala polymorphism of the peroxisome proliferator-activated receptor gamma gene and strength athlete status.

PubMed

Maciejewska-Karlowska, Agnieszka; Sawczuk, Marek; Cieszczyk, Pawel; Zarebska, Aleksandra; Sawczyn, Stanislaw

2013-01-01

The 12Ala allele of the Peroxisome Proliferator-Activated Receptor gamma gene (PPARG) Pro12Ala polymorphism produces a decreased binding affinity of the PPARγ2 protein, resulting in low activation of the target genes. The 12Ala allele carriers display a significantly improved insulin sensitivity that may result in better glucose utilisation in working skeletal muscles. We hypothesise that the PPARG 12Ala allele could be associated with strength athlete status in Polish athletes. The genotype distribution of PPARG Pro12Ala was examined in 660 Polish athletes. The athletes were stratified into four subgroups: endurance, strength-endurance, sprint-strength and strength. Control samples were prepared from 684 unrelated sedentary volunteers. A χ(2) test was used to compare the PPARG Pro12Ala allele and genotype frequencies between the different groups of athletes and control subjects. Bonferroni's correction for multiple testing was applied. A statistically significant higher frequency of PPARG 12Ala alleles was observed in the subgroup of strength athletes performing short-term and very intense exertion characterised by predominant anaerobic energy production (13.2% vs. 7.5% in controls; P = 0.0007). The PPARG 12Ala allele may be a relevant genetic factor favouring strength abilities in professional athletes, especially in terms of insulin-dependent metabolism, a shift of the energy balance towards glucose utilisation and the development of a favourable weight-to-strength ratio.

Minnelide: A Novel Therapeutic That Promotes Apoptosis in Non-Small Cell Lung Carcinoma In Vivo

PubMed Central

Rousalova, Ilona; Banerjee, Sulagna; Sangwan, Veena; Evenson, Kristen; McCauley, Joel A.; Kratzke, Robert; Vickers, Selwyn M.; Saluja, Ashok; D’Cunha, Jonathan

2013-01-01

Background Minnelide, a pro-drug of triptolide, has recently emerged as a potent anticancer agent. The precise mechanisms of its cytotoxic effects remain unclear. Methods Cell viability was studied using CCK8 assay. Cell proliferation was measured real-time on cultured cells using Electric Cell Substrate Impedence Sensing (ECIS). Apoptosis was assayed by Caspase activity on cultured lung cancer cells and TUNEL staining on tissue sections. Expression of pro-survival and anti-apoptotic genes (HSP70, BIRC5, BIRC4, BIRC2, UACA, APAF-1) was estimated by qRTPCR. Effect of Minnelide on proliferative cells in the tissue was estimated by Ki-67 staining of animal tissue sections. Results In this study, we investigated in vitro and in vivo antitumor effects of triptolide/Minnelide in non-small cell lung carcinoma (NSCLC). Triptolide/Minnelide exhibited anti-proliferative effects and induced apoptosis in NSCLC cell lines and NSCLC mouse models. Triptolide/Minnelide significantly down-regulated the expression of pro-survival and anti-apoptotic genes (HSP70, BIRC5, BIRC4, BIRC2, UACA) and up-regulated pro-apoptotic APAF-1 gene, in part, via attenuating the NF-κB signaling activity. Conclusion In conclusion, our results provide supporting mechanistic evidence for Minnelide as a potential in NSCLC. PMID:24143232

Minnelide: a novel therapeutic that promotes apoptosis in non-small cell lung carcinoma in vivo.

PubMed

Rousalova, Ilona; Banerjee, Sulagna; Sangwan, Veena; Evenson, Kristen; McCauley, Joel A; Kratzke, Robert; Vickers, Selwyn M; Saluja, Ashok; D'Cunha, Jonathan

2013-01-01

Minnelide, a pro-drug of triptolide, has recently emerged as a potent anticancer agent. The precise mechanisms of its cytotoxic effects remain unclear. Cell viability was studied using CCK8 assay. Cell proliferation was measured real-time on cultured cells using Electric Cell Substrate Impedence Sensing (ECIS). Apoptosis was assayed by Caspase activity on cultured lung cancer cells and TUNEL staining on tissue sections. Expression of pro-survival and anti-apoptotic genes (HSP70, BIRC5, BIRC4, BIRC2, UACA, APAF-1) was estimated by qRTPCR. Effect of Minnelide on proliferative cells in the tissue was estimated by Ki-67 staining of animal tissue sections. In this study, we investigated in vitro and in vivo antitumor effects of triptolide/Minnelide in non-small cell lung carcinoma (NSCLC). Triptolide/Minnelide exhibited anti-proliferative effects and induced apoptosis in NSCLC cell lines and NSCLC mouse models. Triptolide/Minnelide significantly down-regulated the expression of pro-survival and anti-apoptotic genes (HSP70, BIRC5, BIRC4, BIRC2, UACA) and up-regulated pro-apoptotic APAF-1 gene, in part, via attenuating the NF-κB signaling activity. In conclusion, our results provide supporting mechanistic evidence for Minnelide as a potential in NSCLC.

Wnt interaction and extracellular release of prominin-1/CD133 in human malignant melanoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rappa, Germana; College of Pharmacy, Roseman University of Health Sciences, Henderson, NV 89104; Mercapide, Javier

2013-04-01

Prominin-1 (CD133) is the first identified gene of a novel class of pentaspan membrane glycoproteins. It is expressed by various epithelial and non-epithelial cells, and notably by stem and cancer stem cells. In non-cancerous cells such as neuro-epithelial and hematopoietic stem cells, prominin-1 is selectively concentrated in plasma membrane protrusions, and released into the extracellular milieu in association with small vesicles. Previously, we demonstrated that prominin-1 contributes to melanoma cells pro-metastatic properties and suggested that it may constitute a molecular target to prevent prominin-1-expressing melanomas from colonizing and growing in lymph nodes and distant organs. Here, we report that threemore » distinct pools of prominin-1 co-exist in cultures of human FEMX-I metastatic melanoma. Morphologically, in addition to the plasma membrane localization, prominin-1 is found within the intracellular compartments, (e.g., Golgi apparatus) and in association with extracellular membrane vesicles. The latter prominin-1–positive structures appeared in three sizes (small, ≤40 nm; intermediates ∼40–80 nm, and large, >80 nm). Functionally, the down-regulation of prominin-1 in FEMX-I cells resulted in a significant reduction of number of lipid droplets as observed by coherent anti-Stokes Raman scattering image analysis and Oil red O staining, and surprisingly in a decrease in the nuclear localization of beta-catenin, a surrogate marker of Wnt activation. Moreover, the T-cell factor/lymphoid enhancer factor (TCF/LEF) promoter activity was 2 to 4 times higher in parental than in prominin-1-knockdown cells. Collectively, our results point to Wnt signaling and/or release of prominin-1–containing membrane vesicles as mediators of the pro-metastatic activity of prominin-1 in FEMX-I melanoma. - Highlights: ► First report of release of prominin-1–containing microvesicles from cancer cells. ► Pro-metastatic role of prominin-1–containing microvesicles in FEMX-I melanoma. ► Down-regulation of prominin-1 results in decreased nuclear localization of β-catenin. ► Wnt signaling as mediator of the pro-metastatic activity of prominin-1.« less

Epigenetic synergies between biotin and folate in the regulation of pro-inflammatory cytokines and repeats

PubMed Central

Xue, Jing; Zempleni, Janos

2013-01-01

The protein biotin ligase, holocarboxylase synthetase (HLCS), is a chromatin protein that interacts physically with the DNA methyltransferase DNMT1, the methylated cytosine binding protein MeCP2, and the histone H3 K9-methyltransferase EHMT1, all of which participate in folate-dependent gene repression. Here we tested the hypothesis that biotin and folate synergize in the repression of pro-inflammatory cytokines and long-terminal repeats (LTRs), mediated by interactions between HLCS and other chromatin proteins. Biotin and folate supplementation could compensate for each other’s deficiency in the repression of LTRs in Jurkat and U937 cells. For example, when biotin-deficient Jurkat cells were supplemented with folate, the expression of LTRs decreased by >70%. Epigenetic synergies were more complex in the regulation of cytokines compared with LTRs. For example, the abundance of TNF-α was 100% greater in folate- and biotin-supplemented U937 cells compared with biotin-deficient and folate-supplemented cells. The NF-κB inhibitor curcumin abrogated the effects of folate and biotin in cytokine regulation, suggesting that transcription factor signaling adds an extra layer of complexity to the regulation of cytokine genes by epigenetic phenomena. We conclude that biotin and folate synergize in the repression of LTRs and that these interactions are probably mediated by HLCS-dependent epigenetic mechanisms. In contrast, synergies between biotin and folate in the regulation of cytokines need to be interpreted in the context of transcription factor signaling. PMID:24007195

Evaluation of inflammation-related genes polymorphisms in Mexican with Alzheimer’s disease: a pilot study

PubMed Central

Toral-Rios, Danira; Franco-Bocanegra, Diana; Rosas-Carrasco, Oscar; Mena-Barranco, Francisco; Carvajal-García, Rosa; Meraz-Ríos, Marco Antonio; Campos-Peña, Victoria

2015-01-01

Amyloid peptide is able to promote the activation of microglia and astrocytes in Alzheimer’s disease (AD), and this stimulates the production of pro-inflammatory cytokines. Inflammation contributes to the process of neurodegeneration and therefore is a key factor in the development of AD. Some of the most important proteins involved in AD inflammation are: clusterin (CLU), complement receptor 1 (CR1), C reactive protein (CRP), tumor necrosis factor α (TNF-α), the interleukins 1α (IL-1α), 6 (IL-6), 10 (IL-10) and cyclooxygenase 2 (COX-2). In particular, COX-2 is encoded by the prostaglandin-endoperoxide synthase 2 gene (PTGS2). Since variations in the genes that encode these proteins may modify gene expression or function, it is important to investigate whether these variations may change the developing AD. The aim of this study was to determine whether the presence of polymorphisms in the genes encoding the aforementioned proteins is associated in Mexican patients with AD. Fourteen polymorphisms were genotyped in 96 subjects with AD and 100 controls; the differences in allele, genotype and haplotype frequencies were analyzed. Additionally, an ancestry analysis was conducted to exclude differences in genetic ancestry among groups as a confounding factor in the study. Significant differences in frequencies between AD and controls were found for the single-nucleotide polymorphism (SNP) rs20417 within the PTGS2 gene. Ancestry analysis revealed no significant differences in the ancestry of the compared groups, and the association was significant even after adjustment for ancestry and correction for multiple testing, which strengthens the validity of the results. We conclude that this polymorphism plays an important role in the development of the AD pathology and further studies are required, including their proteins. PMID:26041990

Association of Circulating Biomarkers (Adrenomedullin, TNFR1, and NT-proBNP) With Renal Function Decline in Patients With Type 2 Diabetes: A French Prospective Cohort.

PubMed

Saulnier, Pierre-Jean; Gand, Elise; Velho, Gilberto; Mohammedi, Kamel; Zaoui, Philippe; Fraty, Mathilde; Halimi, Jean Michel; Roussel, Ronan; Ragot, Stéphanie; Hadjadj, Samy

2017-03-01

We explored the prognostic value of three circulating candidate biomarkers-midregional-proadrenomedullin (MR-proADM), soluble tumor necrosis factor receptor 1 (sTNFR1), and N-terminal prohormone brain natriuretic peptide (NT-proBNP)-for change in renal function in patients with type 2 diabetes. Outcomes were defined as renal function loss (RFL), ≥40% decline of estimated glomerular filtration rate (eGFR) from baseline, and rapid renal function decline (RRFD), absolute annual eGFR slope <-5 mL/min/year. We used a proportional hazard model for RFL and a logistic model for RRFD. Adjustments were performed for established risk factors (age, sex, diabetes duration, HbA 1c , blood pressure, baseline eGFR, and urinary albumin-to-creatinine ratio [uACR]). C-statistics were used to assess the incremental predictive value of the biomarkers to these risk factors. Among 1,135 participants (mean eGFR 76 mL/min, median uACR 2.6 mg/mmol, and median GFR slope -1.6 mL/min/year), RFL occurred in 397, RRFD developed in 233, and 292 died during follow-up. Each biomarker predicted RFL and RRFD. When combined, MR-proADM, sTNFR1, and NT-proBNP predicted RFL independently from the established risk factors (adjusted hazard ratio 1.59 [95% CI 1.34-1.89], P < 0.0001; 1.33 [1.14-1.55], P = 0.0003; and 1.22 [1.07-1.40], P = 0.004, respectively) and RRFD (adjusted odds ratio 1.56 [95% CI 1.7-2.09], P = 0.003; 1.72 [1.33-2.22], P < 0.0001; and 1.28 [1.03-1.59], P = 0.02, respectively). The combination of the three biomarkers yielded the highest discrimination (difference in C-statistic = 0.054, P < 0.0001; 0.067, P < 0.0001 for RFL; and 0.027, P < 0.0001 for RRFD). In addition to established risk factors, MR-proADM, sTNFR1, and NT-proBNP improve risk prediction of loss of renal function in patients with type 2 diabetes. © 2017 by the American Diabetes Association.

Characterization of lentiviral vector production using microwell suspension cultures of HEK293T-derived producer cells.

PubMed

Guy, Heather M; McCloskey, Laura; Lye, Gary J; Mitrophanous, Kyriacos A; Mukhopadhyay, Tarit K

2013-04-01

ProSavin(®) is a lentiviral vector (LV)-based gene therapy for Parkinson's disease. ProSavin(®) is currently in a Phase I/II clinical trial using material that was generated by transient transfection of adherent human embryonic kidney (HEK)293T cells. For future large-scale productions of ProSavin(®), we have previously reported the development and characterization of two inducible producer cell lines, termed PS5.8 and PS46.2. PS46.2 has been successfully adapted to grow in suspension cultures. The present study describes the creation of a small-scale (<2 ml) microwell-based experimental platform for the parallel investigation of ProSavin(®) production using suspension-adapted PS46.2. This is combined with statistical design of experiments (DoE) techniques to enable rapid characterization of the process conditions that impact cell growth and LV production. The effects of postinduction period, microwell liquid fill volume, and concentration of inducer (doxycycline) on ProSavin(®) titer and the particle:infectivity (P:I) ratio was investigated using three rounds of DoE, in order to identify appropriate factor ranges and optimize production conditions. We identified an optimal "harvest window" between approximately 26-46 hr within which maximal titers of around 6×10(4) transducing units (TU)/ml were obtained (an approximately 30-fold improvement compared to starting microwell conditions), providing that the fill volume was maintained at or below 1 ml and the doxycycline concentration was at least 1.0 μg/ml. Insights from the microwell studies were subsequently used to rapidly establish operating conditions for ProSavin(®) production in a 0.5-L wave bioreactor culture. The information presented herein thus aids the design and evaluation of scalable production processes for LVs.

Hypothyroidism alters the expression of Bcl-2 family genes to induce enhanced apoptosis in the developing cerebellum.

PubMed

Singh, R; Upadhyay, G; Kumar, S; Kapoor, A; Kumar, A; Tiwari, M; Godbole, M M

2003-01-01

Thyroid hormone (TH) deficiency results in delayed proliferation and migration of cerebellar granule cells. Although extensive cell loss during the development of the cerebellum under hypothyroid conditions is known, its nature and its mechanism are poorly understood. Bcl-2 family gene expression is known to determine the fate of cells to undergo apoptosis. We evaluated the effect of hypothyroidism on Bcl-2 family gene expression in the developing rat cerebellum. Electrophoresis and Western blotting were used to analyze DNA fragmentation and expression of DNA fragmentation factor (DFF-45), Bcl-2, Bcl-xL and Bax genes respectively. In the hypothyroid condition, extensive DNA fragmentation and enhanced cleavage of DFF-45 were seen throughout development (postnatal day 0 to day 24) and adulthood whereas they were absent in the euthyroid state. The anti-apoptotic genes Bcl-2 and Bcl-xL were down-regulated and the pro-apoptotic gene Bax was expressed at higher levels compared with the euthyroid state. These results suggest that normal levels of TH prevent cerebellar apoptosis to a large extent, whereas hypothyroidism not only increases the extent but also the duration of apoptosis by down-regulating the anti-apoptotic genes and maintaining a high level of the pro-apoptotic gene Bax.

Pro-apoptotic gene regulation in the Caribbean fruit fly, Anastrepha suspensa

USDA-ARS?s Scientific Manuscript database

Transcriptional activation of pro-apoptotic genes in response to cytotoxic stimuli is a conserved feature of the cell death pathway proposed for metazoans. However, understanding the extent of this conservation in insects, as well as other organisms, has been limited by the lack of known pro-apoptot...

UV Light Potentiates STING (Stimulator of Interferon Genes)-dependent Innate Immune Signaling through Deregulation of ULK1 (Unc51-like Kinase 1)*

PubMed Central

Kemp, Michael G.; Lindsey-Boltz, Laura A.; Sancar, Aziz

2015-01-01

The mechanism by which ultraviolet (UV) wavelengths of sunlight trigger or exacerbate the symptoms of the autoimmune disorder lupus erythematosus is not known but may involve a role for the innate immune system. Here we show that UV radiation potentiates STING (stimulator of interferon genes)-dependent activation of the immune signaling transcription factor interferon regulatory factor 3 (IRF3) in response to cytosolic DNA and cyclic dinucleotides in keratinocytes and other human cells. Furthermore, we find that modulation of this innate immune response also occurs with UV-mimetic chemical carcinogens and in a manner that is independent of DNA repair and several DNA damage and cell stress response signaling pathways. Rather, we find that the stimulation of STING-dependent IRF3 activation by UV is due to apoptotic signaling-dependent disruption of ULK1 (Unc51-like kinase 1), a pro-autophagic protein that negatively regulates STING. Thus, deregulation of ULK1 signaling by UV-induced DNA damage may contribute to the negative effects of sunlight UV exposure in patients with autoimmune disorders. PMID:25792739

Anti-inflammatory effects of ursodeoxycholic acid by lipopolysaccharide-stimulated inflammatory responses in RAW 264.7 macrophages

PubMed Central

Ko, Wan-Kyu; Lee, Soo-Hong; Kim, Sung Jun; Jo, Min-Jae; Kumar, Hemant; Han, In-Bo; Sohn, Seil

2017-01-01

Purpose The aim of this study was to investigate the anti-inflammatory effects of Ursodeoxycholic acid (UDCA) in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. Methods We induced an inflammatory process in RAW 264.7 macrophages using LPS. The anti-inflammatory effects of UDCA on LPS-stimulated RAW 264.7 macrophages were analyzed using nitric oxide (NO). Pro-inflammatory and anti-inflammatory cytokines were analyzed by quantitative real time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). The phosphorylations of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 in mitogen-activated protein kinase (MAPK) signaling pathways and nuclear factor kappa-light polypeptide gene enhancer in B-cells inhibitor, alpha (IκBα) signaling pathways were evaluated by western blot assays. Results UDCA decreased the LPS-stimulated release of the inflammatory mediator NO. UDCA also decreased the pro-inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin 1-α (IL-1α), interleukin 1-β (IL-1β), and interleukin 6 (IL-6) in mRNA and protein levels. In addition, UDCA increased an anti-inflammatory cytokine interleukin 10 (IL-10) in the LPS-stimulated RAW 264.7 macrophages. UDCA inhibited the expression of inflammatory transcription factor nuclear factor kappa B (NF-κB) in LPS-stimulated RAW 264.7 macrophages. Furthermore, UDCA suppressed the phosphorylation of ERK, JNK, and p38 signals related to inflammatory pathways. In addition, the phosphorylation of IκBα, the inhibitor of NF-κB, also inhibited by UDCA. Conclusion UDCA inhibits the pro-inflammatory responses by LPS in RAW 264.7 macrophages. UDCA also suppresses the phosphorylation by LPS on ERK, JNK, and p38 in MAPKs and NF-κB pathway. These results suggest that UDCA can serve as a useful anti-inflammatory drug. PMID:28665991

Shrimp serine proteinase homologues PmMasSPH-1 and -2 play a role in the activation of the prophenoloxidase system.

PubMed

Jearaphunt, Miti; Amparyup, Piti; Sangsuriya, Pakkakul; Charoensapsri, Walaiporn; Senapin, Saengchan; Tassanakajon, Anchalee

2015-01-01

Melanization mediated by the prophenoloxidase (proPO) activating system is a rapid immune response used by invertebrates against intruding pathogens. Several masquerade-like and serine proteinase homologues (SPHs) have been demonstrated to play an essential role in proPO activation in insects and crustaceans. In a previous study, we characterized the masquerade-like SPH, PmMasSPH1, in the black tiger shrimp Penaeus monodon as a multifunctional immune protein based on its recognition and antimicrobial activity against the Gram-negative bacteria Vibrio harveyi. In the present study, we identify a novel SPH, known as PmMasSPH2, composed of an N-terminal clip domain and a C-terminal SP-like domain that share high similarity to those of other insect and crustacean SPHs. We demonstrate that gene silencing of PmMasSPH1 and PmMasSPH2 significantly reduces PO activity, resulting in a high number of V. harveyi in the hemolymph. Interestingly, knockdown of PmMasSPH1 suppressed not only its gene transcript but also other immune-related genes in the proPO system (e.g., PmPPAE2) and antimicrobial peptides (e.g., PenmonPEN3, PenmonPEN5, crustinPm1 and Crus-likePm). The PmMasSPH1 and PmMasSPH2 also show binding activity to peptidoglycan (PGN) of Gram-positive bacteria. Using a yeast two-hybrid analysis and co-immunoprecipitation, we demonstrate that PmMasSPH1 specifically interacted with the final proteinase of the proPO cascade, PmPPAE2. Furthermore, the presence of both PmMasSPH1 and PmPPAE2 enhances PGN-induced PO activity in vitro. Taken together, these results suggest the importance of PmMasSPHs in the activation of the shrimp proPO system.

Shrimp Serine Proteinase Homologues PmMasSPH-1 and -2 Play a Role in the Activation of the Prophenoloxidase System

PubMed Central

Jearaphunt, Miti; Amparyup, Piti; Sangsuriya, Pakkakul; Charoensapsri, Walaiporn; Senapin, Saengchan; Tassanakajon, Anchalee

2015-01-01

Melanization mediated by the prophenoloxidase (proPO) activating system is a rapid immune response used by invertebrates against intruding pathogens. Several masquerade-like and serine proteinase homologues (SPHs) have been demonstrated to play an essential role in proPO activation in insects and crustaceans. In a previous study, we characterized the masquerade-like SPH, PmMasSPH1, in the black tiger shrimp Penaeus monodon as a multifunctional immune protein based on its recognition and antimicrobial activity against the Gram-negative bacteria Vibrio harveyi. In the present study, we identify a novel SPH, known as PmMasSPH2, composed of an N-terminal clip domain and a C-terminal SP-like domain that share high similarity to those of other insect and crustacean SPHs. We demonstrate that gene silencing of PmMasSPH1 and PmMasSPH2 significantly reduces PO activity, resulting in a high number of V. harveyi in the hemolymph. Interestingly, knockdown of PmMasSPH1 suppressed not only its gene transcript but also other immune-related genes in the proPO system (e.g., PmPPAE2) and antimicrobial peptides (e.g., PenmonPEN3, PenmonPEN5, crustinPm1 and Crus-likePm). The PmMasSPH1 and PmMasSPH2 also show binding activity to peptidoglycan (PGN) of Gram-positive bacteria. Using a yeast two-hybrid analysis and co-immunoprecipitation, we demonstrate that PmMasSPH1 specifically interacted with the final proteinase of the proPO cascade, PmPPAE2. Furthermore, the presence of both PmMasSPH1 and PmPPAE2 enhances PGN-induced PO activity in vitro. Taken together, these results suggest the importance of PmMasSPHs in the activation of the shrimp proPO system. PMID:25803442

The SIGMAR1 gene is associated with a risk of schizophrenia and activation of the prefrontal cortex.

PubMed

Ohi, Kazutaka; Hashimoto, Ryota; Yasuda, Yuka; Fukumoto, Motoyuki; Yamamori, Hidenaga; Umeda-Yano, Satomi; Kamino, Kouzin; Ikezawa, Koji; Azechi, Michiyo; Iwase, Masao; Kazui, Hiroaki; Kasai, Kiyoto; Takeda, Masatoshi

2011-07-01

Several studies have identified the possible involvement of sigma non-opioid intracellular receptor 1 (SIGMAR1) in the pathogenesis of schizophrenia. The Gln2Pro polymorphism in the SIGMAR1 gene has been extensively examined for an association with schizophrenia. However, findings across multiple studies have been inconsistent. We performed a meta-analysis of the association between the functional Gln2Pro polymorphism and schizophrenia using combined samples (1254 patients with schizophrenia and 1574 healthy controls) from previously published studies and our own additional samples (478 patients and 631 controls). We then used near-infrared spectroscopy to analyze the effects of the Gln2Pro genotype, a schizophrenia diagnosis and the interaction between genotype and diagnosis on activation of the prefrontal cortex (PFC) during a verbal fluency task (127 patients and 216 controls). The meta-analysis provided evidence of an association between Gln2Pro and schizophrenia without heterogeneity across studies (odds ratio=1.12, p=0.047). Consistent with previous studies, patients with schizophrenia showed lower bilateral activation of the PFC when compared to controls (p<0.05). We provide evidence that Pro carriers, who are more common among patients with schizophrenia, have significantly lower activation of the right PFC compared to subjects with the Gln/Gln genotype (p=0.013). These data suggest that the SIGMAR1 polymorphism is associated with an increased risk of schizophrenia and differential activation of the PFC. Copyright © 2011 Elsevier Inc. All rights reserved.

NT-proBNP is associated with coronary heart disease risk in healthy older women but fails to enhance prediction beyond established risk factors: results from the British Women's Heart and Health Study.

PubMed

Sattar, Naveed; Welsh, Paul; Sarwar, Nadeem; Danesh, John; Di Angelantonio, Emanuele; Gudnason, Vilmundur; Davey Smith, George; Ebrahim, Shah; Lawlor, Debbie A

2010-03-01

Limited evidence suggests NT-proBNP improves prediction of coronary heart disease (CHD) events but further data are needed, especially in people without pre-existing CHD and in women. We measured NT-proBNP in serum from 162 women with incident CHD events and 1226 controls (60-79 years) in a case-control study nested within the prospective British Women's Heart and Health Study. All cases and controls were free from CHD at baseline. We related NT-proBNP to CHD event risk, and determined to what extent NT-proBNP enhanced CHD risk prediction beyond established risk factors. The odds ratio for CHD per 1 standard deviation increase in log(e)NT-proBNP was 1.37 (95% CI: 1.13-1.68) in analyses adjusted for established CHD risk factors, social class, CRP and insulin. However, addition of log(e)NT-proBNP did not improve the discrimination of a prediction model including age, social class, smoking, physical activity, lipids, fasting glucose, waist:hip ratio, hypertension, statin and aspirin use, nor a standard Framingham risk score model; area under the receiver operator curve for the former model increased from 0.676 to 0.687 on inclusion of NT-proBNP (p=0.3). Furthermore, adding NT-proBNP did not improve calibration of a prediction model containing established risk factors, nor did inclusion more appropriately re-classify participants in relation to their final outcome. Findings were similar (independent associations, but no prediction improvement) for fasting insulin and CRP. These results caution against use of NT-proBNP for CHD risk prediction in healthy women and suggest a need for larger studies in both genders to resolve outstanding uncertainties.

Oleuropein potentiates anti-tumor activity of cisplatin against HepG2 through affecting proNGF/NGF balance.

PubMed

Sherif, Iman O; Al-Gayyar, Mohammed M H

2018-04-01

Oleuropein is considered as a new chemotherapeutic agent in human hepatocellular carcinoma (HCC) while, its exact underlying molecular mechanism still not yet explored. In addition, cisplatin is a standard anticancer drug against solid tumors with toxic side effects. Therefore, we conducted this study to assess antitumor activity of oleuropein either alone or in combination with cisplatin against HepG2, human HCC cell lines, via targeting pro-NGF/NGF signaling pathway. HepG2 cells were treated with cisplatin (20, 50, 100 μM) and oleuropein (100, 200, 300 and 400 μM) as well as some of the cells were treated with 50 μM cisplatin and different concentrations of oleuropein. Gene expressions of nerve growth factor (NGF), matrix metalloproteinase-7 (MMP-7) and caspase-3 were evaluated by real time-PCR. In addition, protein levels of NGF and pro-form of NGF (pro-NGF) were measured by ELISA while, nitric oxide (NO) content was determined colorimetrically. Cisplatin treatment showed a significant elevation of NO content and pro-NGF protein level with a marked reduction of NGF protein level in addition to the upregulation of caspase-3 along with downregulation of MMP-7 gene expressions in a dose-dependent manner. However, the combination of 50 μM cisplatin and 200 μM oleuropein showed the most potent effect on the molecular level when compared with oleuropein or cisplatin alone. Our results showed for the first time that the anti-tumor activity of oleuropein against HCC could be attributed to influencing the pro-NGF/NGF balance via affecting MMP-7 activity without affecting the gene expression of NGF. Concurrent treatment with both oleuropein and cisplatin could lead to more effective chemotherapeutic combination against HCC. Copyright © 2018 Elsevier Inc. All rights reserved.

TNF-alpha and IL-6 inhibit apolipoprotein A-IV production induced by linoleic acid in human intestinal Caco2 cells.

PubMed

Li, Xiaoming; Xu, Min; Liu, Min; Ji, Yong; Li, Zongfang

2015-01-01

Apolipoprotein A-IV (apoA-IV) is a protein mainly synthesized by enterocytes in the intestine. Its gene expression is suppressed during fasting and stimulated during active fat absorption. Chronic feeding of a high-fat (HF) diet abolishes the differential expression between fasting and fat-feeding and therefore may contribute to diet-induced obesity since apoA-IV is a potent satiety factor. It is well established that the circulating pro-inflammatory cytokines TNF-α and IL-6 are increased by HF feeding. To determine whether pro-inflammatory cytokines are involved in the diminished response of apoA-IV gene expression to fat-feeding, different concentrations of linoleic acid (LA), an important dietary fatty acid, was used to stimulate apoA-IV expression in human intestinal Caco2 cells. Cells were pre-treated with or without human recombinant TNF-α, IL-6 or their combination before the addition of LA. Real-time PCR and ELISA were used to detect and quantify RNA transcripts and proteins of apoA-IV and the cytokines. LA stimulated gene and protein expression of apoA-IV in a dose and time dependent manner. Pre-treatment with the cytokines for 72 h significantly inhibited the increased expression of apoA-IV gene and protein induced by LA. Furthermore, the cytokines, especially TNF-α, also positively up-regulate the cytokine themselves in Caco2 cells. Our data indicate that the pro-inflammatory cytokines may be responsible for the reduced apoA-IV production in response to fat feeding. Because of apoA-IV's role in satiety, we propose the inhibitory effect of circulating pro-inflammatory cytokines on apoA-IV production contributes to diet-induced obesity.

HULC long noncoding RNA silencing suppresses angiogenesis by regulating ESM-1 via the PI3K/Akt/mTOR signaling pathway in human gliomas.

PubMed

Zhu, Yu; Zhang, Xuebin; Qi, Lisha; Cai, Ying; Yang, Ping; Xuan, Geng; Jiang, Yuan

2016-03-22

Tumor angiogenesis plays a critical role in the tumor progression. Highly upregulated in liver cancer (HULC) is a long noncoding RNA (lncRNA) that acts as an oncogene in gliomas. We found that HULC, vascular endothelial growth factor (VEGF), and ESM-1 (endothelial cell specific molecule 1) expression and microvessel density were positively correlated with grade dependency in glioma patient tissues, and that HULC silencing suppressed angiogenesis by inhibiting glioma cells proliferation and invasion. This process induced anoikis and blocked the cell cycle at G1/S phase via the PI3K/Akt/mTOR signaling pathway, thus regulating the tumor-related genes involved in the above biological behavior in human glioma U87MG and U251 cells. However, these effects were reversed by ESM-1 overexpression, suggesting a mediating role of ESM-1 in the pro-angiogenesis effect of HULC. Our results define the mechanism of the pro-angiogenesis activity of HULC, which shows potential for application as a therapeutic target in glioma.

Anti-Inflammatory Effect of Apigenin on LPS-Induced Pro-Inflammatory Mediators and AP-1 Factors in Human Lung Epithelial Cells.

PubMed

Patil, Rajeshwari H; Babu, R L; Naveen Kumar, M; Kiran Kumar, K M; Hegde, Shubha M; Nagesh, Rashmi; Ramesh, Govindarajan T; Sharma, S Chidananda

2016-02-01

Apigenin is one of the plant flavonoids present in fruits and vegetables, acting as an important nutraceutical component. It is recognized as a potential antioxidant, antimicrobial, and anti-inflammatory molecule. In the present study, the mechanism of anti-inflammatory action of apigenin on lipopolysaccharide (LPS)-induced pro-inflammatory cytokines and activator protein-1 (AP-1) factors in human lung A549 cells was investigated. The anti-inflammatory activity of apigenin on LPS-induced inflammation was determined by analyzing the expression of pro-inflammatory cytokines, nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and different AP-1 factors. Apigenin significantly inhibited the LPS-induced expression of iNOS, COX-2, expression of pro-inflammatory cytokines (IL-1β, IL-2, IL-6, IL-8, and TNF-α), and AP-1 proteins (c-Jun, c-Fos, and JunB) including nitric oxide production. Study confirms the anti-inflammatory effect of apigenin by inhibiting the expression of inflammatory mediators and AP-1 factors involved in the inflammation and its importance in the treatment of lung inflammatory diseases.

Extremely low-frequency electromagnetic field exposure enhances inflammatory response and inhibits effect of antioxidant in RAW 264.7 cells.

PubMed

Kim, Soo Jeong; Jang, Ye Won; Hyung, Kyung Eun; Lee, Da Kyoung; Hyun, Kee Hyeob; Jeong, Seung Hwarn; Min, Kyung Hoon; Kang, Wonku; Jeong, Ji Hoon; Park, So-Young; Hwang, Kwang Woo

2017-07-01

In recent years, there has been a dramatic increase in the number and variety of electronic devices that emit electromagnetic waves. Because people live and work in close proximity to these pieces of electrical equipment, there is growing concern surrounding the destruction of homeostasis by electromagnetic field exposure. In the present study, the effects of 60 Hz 0.8 mT extremely low-frequency electromagnetic fields (ELF-EMF) on a macrophage cell line (RAW 264.7) were examined. Under defined ELF-EMF exposure conditions, the production of nitric oxide and pro-inflammatory cytokines, TNF-α, IL-1β, and IL-6, were increased in RAW 264.7 cells and the expression of those genes was also upregulated. However, cell proliferation was not altered. Translocation of NF-κB (nuclear factor kappa B), molecules that act downstream of the pro-inflammatory cytokines, were increased to the nucleus under ELF-EMF exposure conditions. In addition, we found that ELF-EMF exposure elevated activation of nuclear factor of activated T cells (NFAT) 2, as well as positively affected the influx of calcium. Furthermore, with both the presence of a potent antioxidant (Resveratrol) and downregulation of the antioxidant-related gene Prx-1 (Peroxiredoxin-1), ELF-EMF was associated with higher inflammatory responses of macrophages. These results suggest that an ELF-EMF amplifies inflammatory responses through enhanced macrophage activation and can decrease the effectiveness of antioxidants. Bioelectromagnetics. 38:374-385, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Flavocoxid, a dual inhibitor of cyclooxygenase and 5-lipoxygenase, blunts pro-inflammatory phenotype activation in endotoxin-stimulated macrophages.

PubMed

Altavilla, D; Squadrito, F; Bitto, A; Polito, F; Burnett, B P; Di Stefano, V; Minutoli, L

2009-08-01

The flavonoids, baicalin and catechin, from Scutellaria baicalensis and Acacia catechu, respectively, have been used for various clinical applications. Flavocoxid is a mixed extract containing baicalin and catechin, and acts as a dual inhibitor of cyclooxygenase (COX) and 5-lipoxygenase (LOX) enzymes. The anti-inflammatory activity, measured by protein and gene expression of inflammatory markers, of flavocoxid in rat peritoneal macrophages stimulated with Salmonella enteritidis lipopolysaccharide (LPS) was investigated. LPS-stimulated (1 microg.mL(-1)) peritoneal rat macrophages were co-incubated with different concentrations of flavocoxid (32-128 microg.mL(-1)) or RPMI medium for different incubation times. Inducible COX-2, 5-LOX, inducible nitric oxide synthase (iNOS) and inhibitory protein kappaB-alpha (IkappaB-alpha) levels were evaluated by Western blot analysis. Nuclear factor kappaB (NF-kappaB) binding activity was investigated by electrophoretic mobility shift assay. Tumour necrosis factor-alpha (TNF-alpha) gene and protein expression were measured by real-time polymerase chain reaction and enzyme-linked immunosorbent assay respectively. Finally, malondialdehyde (MDA) and nitrite levels in macrophage supernatants were evaluated. LPS stimulation induced a pro-inflammatory phenotype in rat peritoneal macrophages. Flavocoxid (128 microg.mL(-1)) significantly inhibited COX-2 (LPS = 18 +/- 2.1; flavocoxid = 3.8 +/- 0.9 integrated intensity), 5-LOX (LPS = 20 +/- 3.8; flavocoxid = 3.1 +/- 0.8 integrated intensity) and iNOS expression (LPS = 15 +/- 1.1; flavocoxid = 4.1 +/- 0.4 integrated intensity), but did not modify COX-1 expression. PGE(2) and LTB(4) levels in culture supernatants were consequently decreased. Flavocoxid also prevented the loss of IkappaB-alpha protein (LPS = 1.9 +/- 0.2; flavocoxid = 7.2 +/- 1.6 integrated intensity), blunted increased NF-kappaB binding activity (LPS = 9.2 +/- 2; flavocoxid = 2.4 +/- 0.7 integrated intensity) and the enhanced TNF-alpha mRNA levels (LPS = 8 +/- 0.9; flavocoxid = 1.9 +/- 0.8 n-fold/beta-actin) induced by LPS. Finally, flavocoxid decreased MDA, TNF and nitrite levels from LPS-stimulated macrophages. Flavocoxid might be useful as a potential anti-inflammatory agent, acting at the level of gene and protein expression.

Capture of microRNA-bound mRNAs identifies the tumor suppressor miR-34a as a regulator of growth factor signaling.

PubMed

Lal, Ashish; Thomas, Marshall P; Altschuler, Gabriel; Navarro, Francisco; O'Day, Elizabeth; Li, Xiao Ling; Concepcion, Carla; Han, Yoon-Chi; Thiery, Jerome; Rajani, Danielle K; Deutsch, Aaron; Hofmann, Oliver; Ventura, Andrea; Hide, Winston; Lieberman, Judy

2011-11-01

A simple biochemical method to isolate mRNAs pulled down with a transfected, biotinylated microRNA was used to identify direct target genes of miR-34a, a tumor suppressor gene. The method reidentified most of the known miR-34a regulated genes expressed in K562 and HCT116 cancer cell lines. Transcripts for 982 genes were enriched in the pull-down with miR-34a in both cell lines. Despite this large number, validation experiments suggested that ~90% of the genes identified in both cell lines can be directly regulated by miR-34a. Thus miR-34a is capable of regulating hundreds of genes. The transcripts pulled down with miR-34a were highly enriched for their roles in growth factor signaling and cell cycle progression. These genes form a dense network of interacting gene products that regulate multiple signal transduction pathways that orchestrate the proliferative response to external growth stimuli. Multiple candidate miR-34a-regulated genes participate in RAS-RAF-MAPK signaling. Ectopic miR-34a expression reduced basal ERK and AKT phosphorylation and enhanced sensitivity to serum growth factor withdrawal, while cells genetically deficient in miR-34a were less sensitive. Fourteen new direct targets of miR-34a were experimentally validated, including genes that participate in growth factor signaling (ARAF and PIK3R2) as well as genes that regulate cell cycle progression at various phases of the cell cycle (cyclins D3 and G2, MCM2 and MCM5, PLK1 and SMAD4). Thus miR-34a tempers the proliferative and pro-survival effect of growth factor stimulation by interfering with growth factor signal transduction and downstream pathways required for cell division.

Genome wide gene expression regulation by HIP1 Protein Interactor, HIPPI: prediction and validation.

PubMed

Datta, Moumita; Choudhury, Ananyo; Lahiri, Ansuman; Bhattacharyya, Nitai P

2011-09-26

HIP1 Protein Interactor (HIPPI) is a pro-apoptotic protein that induces Caspase8 mediated apoptosis in cell. We have shown earlier that HIPPI could interact with a specific 9 bp sequence motif, defined as the HIPPI binding site (HBS), present in the upstream promoter of Caspase1 gene and regulate its expression. We also have shown that HIPPI, without any known nuclear localization signal, could be transported to the nucleus by HIP1, a NLS containing nucleo-cytoplasmic shuttling protein. Thus our present work aims at the investigation of the role of HIPPI as a global transcription regulator. We carried out genome wide search for the presence of HBS in the upstream sequences of genes. Our result suggests that HBS was predominantly located within 2 Kb upstream from transcription start site. Transcription factors like CREBP1, TBP, OCT1, EVI1 and P53 half site were significantly enriched in the 100 bp vicinity of HBS indicating that they might co-operate with HIPPI for transcription regulation. To illustrate the role of HIPPI on transcriptome, we performed gene expression profiling by microarray. Exogenous expression of HIPPI in HeLa cells resulted in up-regulation of 580 genes (p < 0.05) while 457 genes were down-regulated. Several transcription factors including CBP, REST, C/EBP beta were altered by HIPPI in this study. HIPPI also interacted with P53 in the protein level. This interaction occurred exclusively in the nuclear compartment and was absent in cells where HIP1 was knocked down. HIPPI-P53 interaction was necessary for HIPPI mediated up-regulation of Caspase1 gene. Finally, we analyzed published microarray data obtained with post mortem brains of Huntington's disease (HD) patients to investigate the possible involvement of HIPPI in HD pathogenesis. We observed that along with the transcription factors like CREB, P300, SREBP1, Sp1 etc. which are already known to be involved in HD, HIPPI binding site was also significantly over-represented in the upstream sequences of genes altered in HD. Taken together, the results suggest that HIPPI could act as an important transcription regulator in cell regulating a vast array of genes, particularly transcription factors and at least, in part, play a role in transcription deregulation observed in HD.

Genome wide gene expression regulation by HIP1 Protein Interactor, HIPPI: Prediction and validation

PubMed Central

2011-01-01

Background HIP1 Protein Interactor (HIPPI) is a pro-apoptotic protein that induces Caspase8 mediated apoptosis in cell. We have shown earlier that HIPPI could interact with a specific 9 bp sequence motif, defined as the HIPPI binding site (HBS), present in the upstream promoter of Caspase1 gene and regulate its expression. We also have shown that HIPPI, without any known nuclear localization signal, could be transported to the nucleus by HIP1, a NLS containing nucleo-cytoplasmic shuttling protein. Thus our present work aims at the investigation of the role of HIPPI as a global transcription regulator. Results We carried out genome wide search for the presence of HBS in the upstream sequences of genes. Our result suggests that HBS was predominantly located within 2 Kb upstream from transcription start site. Transcription factors like CREBP1, TBP, OCT1, EVI1 and P53 half site were significantly enriched in the 100 bp vicinity of HBS indicating that they might co-operate with HIPPI for transcription regulation. To illustrate the role of HIPPI on transcriptome, we performed gene expression profiling by microarray. Exogenous expression of HIPPI in HeLa cells resulted in up-regulation of 580 genes (p < 0.05) while 457 genes were down-regulated. Several transcription factors including CBP, REST, C/EBP beta were altered by HIPPI in this study. HIPPI also interacted with P53 in the protein level. This interaction occurred exclusively in the nuclear compartment and was absent in cells where HIP1 was knocked down. HIPPI-P53 interaction was necessary for HIPPI mediated up-regulation of Caspase1 gene. Finally, we analyzed published microarray data obtained with post mortem brains of Huntington's disease (HD) patients to investigate the possible involvement of HIPPI in HD pathogenesis. We observed that along with the transcription factors like CREB, P300, SREBP1, Sp1 etc. which are already known to be involved in HD, HIPPI binding site was also significantly over-represented in the upstream sequences of genes altered in HD. Conclusions Taken together, the results suggest that HIPPI could act as an important transcription regulator in cell regulating a vast array of genes, particularly transcription factors and at least, in part, play a role in transcription deregulation observed in HD. PMID:21943362

Aberrant expression of NKL homeobox gene HLX in Hodgkin lymphoma.

PubMed

Nagel, Stefan; Pommerenke, Claudia; Meyer, Corinna; Kaufmann, Maren; MacLeod, Roderick A F; Drexler, Hans G

2018-03-06

NKL homeobox genes are basic regulators of cell and tissue differentiation, many acting as oncogenes in T-cell leukemia. Recently, we described an hematopoietic NKL-code comprising six particular NKL homeobox genes expressed in hematopoietic stem cells and lymphoid progenitors, unmasking their physiological roles in the development of these cell types. Hodgkin lymphoma (HL) is a B-cell malignancy showing aberrant activity of several developmental genes resulting in disturbed B-cell differentiation. To examine potential concordances in abnormal lymphoid differentiation of T- and B-cell malignancies we analyzed the expression of the hematopoietic NKL-code associated genes in HL, comprising HHEX, HLX, MSX1, NKX2-3, NKX3-1 and NKX6-3. Our approach revealed aberrant HLX activity in 8 % of classical HL patients and additionally in HL cell line L-540. Accordingly, to identify upstream regulators and downstream target genes of HLX we used L-540 cells as a model and performed chromosome and genome analyses, comparative expression profiling and functional assays via knockdown and overexpression experiments therein. These investigations excluded chromosomal rearrangements of the HLX locus at 1q41 and demonstrated that STAT3 operated directly as transcriptional activator of the HLX gene. Moreover, subcellular analyses showed highly enriched STAT3 protein in the nucleus of L-540 cells which underwent cytoplasmic translocation by repressing deacetylation. Finally, HLX inhibited transcription of B-cell differentiation factors MSX1, BCL11A and SPIB and of pro-apoptotic factor BCL2L11/BIM, thereby suppressing Etoposide-induced cell death. Collectively, we propose that aberrantly expressed NKL homeobox gene HLX is part of a pathological gene network in HL, driving deregulated B-cell differentiation and survival.

Increased expression of pro-angiogenic factors and vascularization in thyroid hyperfunctioning adenomas with and without TSH receptor activating mutations.

PubMed

Celano, Marilena; Sponziello, Marialuisa; Tallini, Giovanni; Maggisano, Valentina; Bruno, Rocco; Dima, Mariavittoria; Di Oto, Enrico; Redler, Adriano; Durante, Cosimo; Sacco, Rosario; Filetti, Sebastiano; Russo, Diego

2013-02-01

Autonomously functioning thyroid nodules (AFTN) are known to receive an increased blood influx necessary to sustain their high rate of growth and hormone production. Here, we investigated the expression of hematic and lymphatic vases in a series of 20 AFTN compared with the contralateral non-tumor tissues of the same patients, and the transcript levels of proteins involved in the control of vascular proliferation, including the vascular endothelial growth factor (VEGF) and platelet-derived growth factors (PDGF) and their receptors and the endothelial nitric oxide synthase (eNOS). In parallel, the expression of the differentiation markers sodium/iodide symporter (NIS), thyroperoxidase (TPO), thyroglobulin (Tg), and TSH receptor (TSHR) was also investigated. The data were further analyzed comparing subgroups of tumors with or without mutations in the TSHR gene. Analysis by means of CD31 and D2-40 immunostaining showed in AFTN an increased number of hematic, but not lymphatic, vessels in parallel with an enhanced proliferation rate shown by increased Ki67 staining. Quantitative RT-PCR analysis revealed an increase of VEGF, VEGFR1 and 2, PDGF-A, PDGF-B, and eNOS expression in tumor versus normal tissues. Also, higher transcript levels of NIS, TPO, and Tg were detected. Comparison of the two subgroups of samples revealed only few differences in the expression of the genes examined. In conclusion, these data demonstrate an increased expression of angiogenesis-related factors associated with an enhanced proliferation of hematic, but not lymphatic, vessels in AFTNs. In this context, the presence of TSHR mutations may only slightly influence the expression of pro-angiogenic growth factors.

Screening for Novel Germline Rare Mutations Associated with Aggressive Prostate Cancer

DTIC Science & Technology

2015-12-01

amino acid substitution from Threonine (Thr) to Proline (Pro); while rs61756080 results to the amino acid substitution from Glycine (Gly) to Glutamic...Single nucleotide polymorphisms 20 in the gene encoding Krüppel-like factor 7 are associated with type 2 diabetes . Diabetologia. 2005 Jul;48(7

Knowledge-guided gene prioritization reveals new insights into the mechanisms of chemoresistance.

PubMed

Emad, Amin; Cairns, Junmei; Kalari, Krishna R; Wang, Liewei; Sinha, Saurabh

2017-08-11

Identification of genes whose basal mRNA expression predicts the sensitivity of tumor cells to cytotoxic treatments can play an important role in individualized cancer medicine. It enables detailed characterization of the mechanism of action of drugs. Furthermore, screening the expression of these genes in the tumor tissue may suggest the best course of chemotherapy or a combination of drugs to overcome drug resistance. We developed a computational method called ProGENI to identify genes most associated with the variation of drug response across different individuals, based on gene expression data. In contrast to existing methods, ProGENI also utilizes prior knowledge of protein-protein and genetic interactions, using random walk techniques. Analysis of two relatively new and large datasets including gene expression data on hundreds of cell lines and their cytotoxic responses to a large compendium of drugs reveals a significant improvement in prediction of drug sensitivity using genes identified by ProGENI compared to other methods. Our siRNA knockdown experiments on ProGENI-identified genes confirmed the role of many new genes in sensitivity to three chemotherapy drugs: cisplatin, docetaxel, and doxorubicin. Based on such experiments and extensive literature survey, we demonstrate that about 73% of our top predicted genes modulate drug response in selected cancer cell lines. In addition, global analysis of genes associated with groups of drugs uncovered pathways of cytotoxic response shared by each group. Our results suggest that knowledge-guided prioritization of genes using ProGENI gives new insight into mechanisms of drug resistance and identifies genes that may be targeted to overcome this phenomenon.

A novel NFIA-NFκB feed-forward loop contributes to glioblastoma cell survival

PubMed Central

Lee, JunSung; Hoxha, Edlira

2017-01-01

Abstract Background. The nuclear factor I-A (NFIA) transcription factor promotes glioma growth and inhibits apoptosis in glioblastoma (GBM) cells. Here we report that the NFIA pro-survival effect in GBM is mediated in part via a novel NFIA–nuclear factor-kappaB (NFκB) p65 feed-forward loop. Methods. We examined effects of gain- and loss-of-function manipulations of NFIA and NFκB p65 on each other’s transcription, cell growth, apoptosis and sensitivity to chemotherapy in patient-derived GBM cells and established GBM cell lines. Results. NFIA enhanced apoptosis evasion by activating NFκB p65 and its downstream anti-apoptotic factors tumor necrosis factor receptor-associated factor 1 (TRAF1) and cellular inhibitor of apoptosis proteins (cIAPs). Induction of NFκB by NFIA was required to protect cells from apoptosis, and inhibition of NFκB effectively reversed the NFIA anti-apoptotic effect. Conversely, NFIA knockdown decreased expression of NFκB and anti-apoptotic genes TRAF1 and cIAPs, and increased baseline apoptosis. NFIA positively regulated NFκB transcription and NFκB protein level. Interestingly, NFκB also activated the NFIA promoter and increased NFIA level, and knockdown of NFIA was sufficient to attenuate the NFκB pro-survival effect, suggesting a reciprocal regulation between NFIA and NFκB in governing GBM cell survival. Supporting this, NFIA and NFκB expression levels were highly correlated in human GBM and patient-derived GBM cells. Conclusions. These data define a previously unknown NFIA-NFκB feed-forward regulation that may contribute to GBM cell survival. PMID:27994064

Phenylbutyrate Attenuates the Expression of Bcl-XL, DNA-PK, Caveolin-1, and VEGF in Prostate Cancer Cells1

PubMed Central

Goh, Meidee; Chen, Feng; Paulsen, Michelle T; Yeager, Ann M; Dyer, Erica S; Ljungman, Mats

2001-01-01

Abstract Phenylbutyrate (PB) is a histone deacetylase inhibitor that has been shown to induce differentiation and apoptosis in various cancer cell lines. Although these effects are most likely due to modulation of gene expression, the specific genes and gene products responsible for the effects of PB are not well characterized. In this study, we used cDNA expression arrays and Western blot to assess the effect that PB has on the expression of various cancer and apoptosis-regulatory gene products. We show that PB attenuates the expression of the apoptosis antagonist Bcl-XL, the double-strand break repair protein DNA-dependent protein kinase, the prostate progression marker caveolin -1, and the pro-angiogenic vascular endothelial growth factor. Furthermore, PB was found to act in synergy with ionizing radiation to induce apoptosis in prostate cancer cells. Taken together, our results point to the possibility that PB may be an effective anti-prostate cancer agent when used in combination with radiation or chemotherapy and for the inhibition of cancer progression. PMID:11571633

HOGA1 Gene Mutations of Primary Hyperoxaluria Type 3 in Tunisian Patients.

PubMed

M'dimegh, Saoussen; Aquaviva-Bourdain, Cécile; Omezzine, Asma; Souche, Geneviéve; M'barek, Ibtihel; Abidi, Kamel; Gargah, Tahar; Abroug, Saoussen; Bouslama, Ali

2017-05-01

Primary hyperoxaluria type 3 (PH3) is due to mutations in the recently identified 4-hydroxy-2-oxoglutarate aldolase (HOGA1) gene. PH3 might be the least severe form with a milder phenotype with good preservation of kidney function in most patients. The aim of this study was to report three PH3 cases carrying mutations in HOGA1. Genetic analysis of HOGA1 was performed in patients with a high clinical suspicion of PH after sequencing of AGXT and GRHPR genes, which was negative. Also, a complete AGXT/GRHPR MLPA was performed in these patients in order to detect large deletions/insertions. Two different HOGA1 gene mutations were identified: the p.Pro190Leu in a homozygous state and the p.Gly287Val in two patients in homozygous and heterozygous carriers. The median age at onset of clinical symptoms was 3.93 years. Most of the patients had a positive family history for recurrent urolithiasis. The p.Pro190Leu mutation was reported with impaired renal function at follow-up; however, the p.Gly287Val was presented with normal renal function. All patients were presented with urolithiasis, but only one had a nephrocalcinosis. This study expanded the number of PH3 patients from 63 to 66 cases. The p.Pro190Leu and the p.Gly287Val mutations found in this study can provide a first-line investigation in Tunisian PH1 patients. © 2016 Wiley Periodicals, Inc.

PPARgamma gene polymorphism is associated with exercise-mediated changes of insulin resistance in healthy men.

PubMed

Kahara, Toshio; Takamura, Toshinari; Hayakawa, Tetsuo; Nagai, Yukihiro; Yamaguchi, Hiromi; Katsuki, Tatsuo; Katsuki, Ken-Ichi; Katsuki, Michio; Kobayashi, Ken-Ichi

2003-02-01

Exercise training improves insulin sensitivity, but individual responses vary greatly. Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a regulator of adipose cell differentiation and plays an important role in systemic insulin action. We investigated whether PPARgamma gene polymorphism affects insulin resistance in response to exercise in Japanese healthy men. The exercise program at an individual intensity of 50% of the maximal heart rate was performed for 20 to 60 min/d, and 2 to 3 days per week to attain a level of physical activity of 700 kcal/wk. The program was conducted for 3 months without any dietary intervention, and the clinical and metabolic characteristics were examined before and after the exercise program. Body mass index (BMI) did not change significantly after the exercise program, whereas percentage of body fat (% body fat), fasting plasma glucose (FPG), and serum leptin levels decreased significantly. Pro12Ala polymorphism in PPARgamma gene was performed on genomic DNA isolated from human leukocytes and examined with polymerase chain reaction (PCR) and subsequent restriction enzyme analysis using BstU-I. In this study, the Ala allele did not correlate with fasting immunoreactive insulin (IRI) and homeostasis model assessment-insulin resistance index (HOMA-R) at baseline, but did so with the changes in IRI and HOMA-R after exercise (DeltaIRI, Pro/Pro 0.55 +/- 3.49 microU/mL v Pro/Ala -2.83 +/- 1.47 microU/mL, P <.05; DeltaHOMA-R, Pro/Pro 0.09 +/- 0.86 v Pro/Ala -0.61 +/- 0.32, P <.05). This result suggests that the Ala allele is associated with improvement in insulin resistance after exercise. We conclude that PPARgamma gene polymorphism may be a reliable indicator of whether exercise will have a beneficial effect as part of the treatment of insulin resistance syndrome. Copyright 2003, Elsevier Science (USA). All rights reserved.

Imbalance in the pro-hepatocyte growth factor activation system in bleomycin-induced lung fibrosis in mice.

PubMed

Phin, Sophie; Marchand-Adam, Sylvain; Fabre, Aurélie; Marchal-Somme, Joëlle; Bantsimba-Malanda, Claudie; Kataoka, Hiroaki; Soler, Paul; Crestani, Bruno

2010-03-01

Hepatocyte growth factor (HGF) is a growth factor for alveolar epithelial cells. Activation of pro-HGF to HGF is regulated by the HGF activator (HGFA), a serine protease, and a specific inhibitor (HGFA inhibitor-1, HAI-1). An imbalance in the HGFA/HAI-1 system might contribute to lung fibrosis. Pro-HGF activation capacity from bronchoalveolar lavage (BAL) fluid was evaluated 3, 7, and 14 days after the intratracheal bleomycin injection (Bleo) in mice with or without thrombin. BAL fluid from naïve mice was used as control. HGFA and HAI-1 mRNA were evaluated by QPCR in the whole lung or by Western blot in BAL fluid. BAL fluid from control mice and Bleo mice activated pro-HGF in vitro at a similar degree. Thrombin accelerated proHGF activation by Bleo BAL on Day 3 and Day 7, but not on Day 14, or in control BAL. Incubation of pro-HGF with BAL from Bleo Day 3 and Day 7 mice increased phosphorylation of HGFR on A549 cells. Thrombin-induced pro-HGF activation was inhibited by an anti-HGFA antibody and accelerated by an anti-HAI-1 antibody. Active HGFA was not detected in control BAL and was strongly induced in Bleo BAL. HGFA concentrations were higher on Day 3 and Day 7 than on Day 14. HAI-1 was detected at low levels in control BAL and increased strongly by Day 3 with stable concentrations until Day 14. By demonstrating an imbalance between HGFA and HAI-1 expression in BAL fluid, our results highlight a defective thrombin-dependent proHGF activation system at the fibrotic phase of bleomycin-induced pulmonary fibrosis.

The CDK inhibitor p21 is a novel target gene of ATF4 and contributes to cell survival under ER stress.

PubMed

Inoue, Yasumichi; Kawachi, Shiori; Ohkubo, Tsubasa; Nagasaka, Mai; Ito, Shogo; Fukuura, Keishi; Itoh, Yuka; Ohoka, Nobumichi; Morishita, Daisuke; Hayashi, Hidetoshi

2017-11-01

Activating transcription factor 4 (ATF4) is well known for its role in the endoplasmic reticulum (ER) stress response. ATF4 also transcriptionally induces multiple effectors that determine cell fate depending on cellular context. In addition, ATF4 can communicate both pro-apoptotic and pro-survival signals. How ATF4 mediates its prosurvival roles, however, requires further investigation. Here, we report that the CDK inhibitor p21 is a novel target gene of ATF4. We identified two ATF4-responsive elements, one of which directly binds ATF4, within the first intron of the p21 gene. Importantly, overexpression of p21 enhances cell survival following ER stress induction, while p21 knockdown increases cell death. These results suggest that p21 induction plays a vital role in the cellular response to ER stress and indicate that p21 is a prosurvival effector of ATF4. © 2017 Federation of European Biochemical Societies.

A novel polymorphism in the PAI-1 gene promoter enhances gene expression. A novel pro-thrombotic risk factor?

PubMed

Liguori, Renato; Quaranta, Sandro; Di Fiore, Rosanna; Elce, Ausilia; Castaldo, Giuseppe; Amato, Felice

2014-12-01

Plasminogen activator inhibitor-1 (PAI-1) is the major physiological inhibitor of tissue-type plasminogen activator in plasma and the most important regulator of the fibrinolytic pathway. The 4G/5G polymorphism (rs1799889) in the PAI-1 promoter is associated with altered PAI-1 transcription. We have identified a new 4G/5G allele, in which a T is inserted near the 4G tract or replaces a G in the 5G tract, forming a T plus 4G (T4G) region. This new variant was first identified in two women, one had experienced juvenile myocardial infarction, the other repeated miscarriage; both had increased PAI-1 plasma activity. In view of the important influence of this promoter region on PAI-1 protein plasma level, we performed in vitro evaluation of the effects of the T4G variant on the transcription activity of the PAI-1 gene promoter. In silico prediction analysis showed that presence of the T4G allele disrupts the E-Box region upstream of the T4G variant, altering the affinity of the target sequence for E-Box binding factors like upstream stimulatory factor-1 (USF-1). Basal T4G promoter activity was 50% higher compared to 4G and 5G variants, but it was less stimulated by USF-1 overexpression. We also analyzed the effects of IL-1β and IL-6 on the PAI-1 promoter activity of our three constructs and showed that the T4G variant was less affected by IL-1β than the other variants. These findings indicate that the T4G variant may be a novel risk factor for thrombotic events. Copyright © 2014 Elsevier Ltd. All rights reserved.

Hepatic Leukemia Factor Promotes Resistance To Cell Death: Implications For Therapeutics and Chronotherapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Waters, Katrina M.; Sontag, Ryan L.; Weber, Thomas J.

Physiological variation related to circadian rhythms and aberrant gene expression patterns are believed to modulate therapeutic efficacy, but the precise molecular determinants remain unclear. Here we examine the regulation of cell death by hepatic leukemia factor (HLF), which is an output regulator of circadian rhythms and is aberrantly expressed in human cancers, using an ectopic expression strategy in JB6 mouse epidermal cells and human keratinocytes. Ectopic HLF expression inhibited cell death in both JB6 cells and human keratinocytes, as induced by serum-starvation, tumor necrosis factor alpha and ionizing radiation. Microarray analysis indicates that HLF regulates a complex multi-gene transcriptional programmore » encompassing upregulation of anti-apoptotic genes, downregulation of pro-apoptotic genes, and many additional changes that are consistent with an anti-death program. Collectively, our results demonstrate that ectopic expression of HLF, an established transcription factor that cycles with circadian rhythms, can recapitulate many features associated with circadian-dependent physiological variation.« less

Oxidized Guanine Base Lesions Function in 8-Oxoguanine DNA Glycosylase-1-mediated Epigenetic Regulation of Nuclear Factor κB-driven Gene Expression*

PubMed Central

Pan, Lang; Hao, Wenjing; Ba, Xueqing

2016-01-01

A large percentage of redox-responsive gene promoters contain evolutionarily conserved guanine-rich clusters; guanines are the bases most susceptible to oxidative modification(s). Consequently, 7,8-dihydro-8-oxoguanine (8-oxoG) is one of the most abundant base lesions in promoters and is primarily repaired via the 8-oxoguanine DNA glycosylase-1 (OOG1)-initiated base excision repair pathway. In view of a prompt cellular response to oxidative challenge, we hypothesized that the 8-oxoG lesion and the cognate repair protein OGG1 are utilized in transcriptional gene activation. Here, we document TNFα-induced enrichment of both 8-oxoG and OGG1 in promoters of pro-inflammatory genes, which precedes interaction of NF-κB with its DNA-binding motif. OGG1 bound to 8-oxoG upstream from the NF-κB motif increased its DNA occupancy by promoting an on-rate of both homodimeric and heterodimeric forms of NF-κB. OGG1 depletion decreased both NF-κB binding and gene expression, whereas Nei-like glycosylase-1 and -2 had a marginal effect. These results are the first to document a novel paradigm wherein the DNA repair protein OGG1 bound to its substrate is coupled to DNA occupancy of NF-κB and functions in epigenetic regulation of gene expression. PMID:27756845

Eight novel F13A1 gene missense mutations in patients with mild FXIII deficiency: in silico analysis suggests changes in FXIII-A subunit structure/function.

PubMed

Biswas, Arijit; Ivaskevicius, Vytautas; Thomas, Anne; Varvenne, Michael; Brand, Brigitte; Rott, Hannelore; Haussels, Iris; Ruehl, Heiko; Scholz, Ute; Klamroth, Robert; Oldenburg, Johannes

2014-10-01

Mild FXIII deficiency is an under-diagnosed disorder because the carriers of this deficiency are often asymptomatic and reveal a phenotype only under special circumstances like surgery or induced trauma. Mutational reports from this type of deficiency have been rare. In this study, we present the phenotypic and genotypic data of nine patients showing mild FXIII-A deficiency caused by eight novel heterozygous missense mutations (Pro166Leu, Arg171Gln, His342Tyr, Gln415Arg, Leu529Pro, Gln601Lys, Arg703Gln and Arg715Gly) in the F13A1 gene. None of these variants were seen in 200 healthy controls. In silico structural analysis of the local wild-type protein structures (activated and non-activated) from X-ray crystallographic models downloaded from the protein databank identified potential structural/functional effects for the identified mutations. The missense mutations in the core domain are suggested to be directly influencing the catalytic triad. Mutations on other domains might influence other critical factors such as activation peptide cleavage or the barrel domain integrity. In vitro expression and subsequent biochemical studies in the future will be able to confirm the pathophysiological mechanisms proposed for the mutations in this article.

The Multi-Biomarker Approach for Heart Failure in Patients with Hypertension

PubMed Central

Bielecka-Dabrowa, Agata; Gluba-Brzózka, Anna; Michalska-Kasiczak, Marta; Misztal, Małgorzata; Rysz, Jacek; Banach, Maciej

2015-01-01

We assessed the predictive ability of selected biomarkers using N-terminal pro-brain natriuretic peptide (NT-proBNP) as the benchmark and tried to establish a multi-biomarker approach to heart failure (HF) in hypertensive patients. In 120 hypertensive patients with or without overt heart failure, the incremental predictive value of the following biomarkers was investigated: Collagen III N-terminal propeptide (PIIINP), cystatin C (CysC), lipocalin-2/NGAL, syndecan-4, tumor necrosis factor-α (TNF-α), interleukin 1 receptor type I (IL1R1), galectin-3, cardiotrophin-1 (CT-1), transforming growth factor β (TGF-β) and N-terminal pro-brain natriuretic peptide (NT-proBNP). The highest discriminative value for HF was observed for NT-proBNP (area under the receiver operating characteristic curve (AUC) = 0.873) and TGF-β (AUC = 0.878). On the basis of ROC curve analysis we found that CT-1 > 152 pg/mL, TGF-β < 7.7 ng/mL, syndecan > 2.3 ng/mL, NT-proBNP > 332.5 pg/mL, CysC > 1 mg/L and NGAL > 39.9 ng/mL were significant predictors of overt HF. There was only a small improvement in predictive ability of the multi-biomarker panel including the four biomarkers with the best performance in the detection of HF—NT-proBNP, TGF-β, CT-1, CysC—compared to the panel with NT-proBNP, TGF-β and CT-1 only. Biomarkers with different pathophysiological backgrounds (NT-proBNP, TGF-β, CT-1, CysC) give additive prognostic value for incident HF in hypertensive patients compared to NT-proBNP alone. PMID:25984599

Effects of dietary inulin and mannan oligosaccharide on immune related genes expression and disease resistance of Pacific white shrimp, Litopenaeus vannamei.

PubMed

Li, Yun; Liu, Hong; Dai, Xilin; Li, Jingjing; Ding, Fujiang

2018-05-01

The effects of inulin and mannan oligosaccharide (MOS) at different doses (2.5, 4 and 10 mg/g) in singular or combined diet on growth rate, immune related genes expression, and resistance to white spot syndrome virus (WSSV) and Vibrio alginolyticus in Pacific white shrimp (Litopenaeus vannamei) were investigated. At the end of 28-day singular feeding experiment, the highest values of specific growth rate (SGR) and the expression of toll-like receptor1, 2 and 3 (TLR1, 2, 3), signal transducer and activator of transcription (STAT), crustin, anti-lipopolysaccharide factor (ALF) as well as prophenoloxidase (proPO) were observed in shrimp individually fed with 5 mg/g dietary inulin or MOS, respectively. Compared with individual treatments, diet containing combined prebiotics (5 mg/g inulin and MOS) significantly improved the expression of TLRs, STAT, proPO, crustin and ALF in L. vannamei after four-week feeding. Additionally, Pacific white shrimp fed with combined dietary prebiotics showed significantly higher expression of immune related genes and lower cumulative mortality in WSSV and Vibrio alginolyticus challenges, compared to the singular feeding groups and control. These results in the present study demonstrated that the combined supplementation of inulin (5 mg/g) and MOS (5 mg/g) remarkably enhanced innate immune response and pathogen resistance of shrimp, and should be considered as a promising immunostimulatory additive for the culture of Pacific white shrimp. Copyright © 2018 Elsevier Ltd. All rights reserved.

A new pro-migratory activity on human myogenic precursor cells for a synthetic peptide within the E domain of the mechano growth factor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mills, Philippe; Lafreniere, Jean-Francois; Benabdallah, Basma Fattouma

2007-02-01

Duchenne muscular dystrophy (DMD) is an inherited disease that leads to progressive muscle wasting. Myogenic precursor cell transplantation is an approach that can introduce the normal dystrophin gene in the muscle fibers of the patients. Unfortunately, these myogenic precursor cells do not migrate well in the muscle and thus many injections have to be done to enable a good graft success. Recent reports have shown that there is extensive splicing of the IGF-1 gene in muscles. The MGF isoform contains a C-terminal 24 amino acids peptide in the E domain (MGF-Ct24E) that has intrinsic properties. It can promote the proliferationmore » while delaying the differentiation of C{sub 2}C{sub 12} cells. Here, we demonstrated that this synthetic peptide is a motogenic factor for human precursor myogenic cells in vitro and in vivo. Indeed, MGF-Ct24E peptide can modulate members of the fibrinolytic and metalloproteinase systems, which are implicated in the migration of myogenic cells. MGF-Ct24E peptide enhances the expression of u-PA, u-PAR and MMP-7 while reducing PAI-1 activity. Moreover, it has no effect on the gelatinases MMP-2 and -9. Those combined effects can favour cell migration. Finally, we present some results suggesting that the MGF-Ct24E peptide induces these cell responses through a mechanism that does not involve the IGF-1 receptor. Thus, this MGF-Ct24E peptide has a new pro-migratory activity on human myogenic precursor cells that may be helpful in the treatment of DMD. Those results reinforce the possibility that the IGF-1Ec isoform may produce an E domain peptide that can act as a cytokine.« less

Lack of association between the Pro12Ala polymorphism of the PPAR-gamma 2 gene and type 2 diabetes mellitus in the Qatari consanguineous population.

PubMed

Badii, Ramin; Bener, Abdulbari; Zirie, Mahmoud; Al-Rikabi, Ammar; Simsek, Mehmet; Al-Hamaq, Abdulla O A A; Ghoussaini, Maya; Froguel, Philippe; Wareham, Nick J

2008-03-01

Peroxisome proliferators-activated receptor gamma (PPAR gamma) is a nuclear hormone receptor that serves as a master regulator for adipocytes-specific genes contributing to adipocytes differentiation, insulin sensitivity and lipid metabolism. The substitution of proline to alanine at codon 12 of the PPAR gamma 2 gene (Pro12Ala polymorphism) is most widely studied, and the associations with diabetes, obesity, and other clinical parameters have been reported and discussed in several ethnic groups. Among native Qatar ethnicity, however, there is no report about this polymorphism. The aim of this study was to estimate the allele frequency of the Pro12Ala polymorphism of PPAR gamma 2 gene among Qatari population and investigate the association between this polymorphism and obesity or type 2 diabetes. This is a matched case-control study. It was carried out among diabetic patients and healthy subjects at the Primary Healthcare Clinics, and the survey was conducted from February 2003 to March 2006 in Qatari male and female nationals aged 35 to 60 years. The study was based on matched age, sex, and ethnicity of 400 cases (with diabetes) and 450 controls (without diabetes). Face-to-face interviews were based on a questionnaire that included variables such as age, sex, sociodemographic status, body mass index (BMI), and obesity. Their health status was assessed by medical conditions, family history, and blood pressure measurements. The allele frequency of Pro12Ala polymorphism in PPAR gamma 2 gene among Qataris is lower than that in many Caucasian ethnic groups. No association is seen between the Pro12Ala and type 2 Diabetes (0.055 vs 0.059, OR = 1.1311, P = 0.669). Nearly half of the diabetic type 2 patients (48.5%) were obese (BMI > 30) compared to nondiabetic subjects (29.8%) (P < 0.001). In this study, no association is seen between the Pro12Ala polymorphism in PPAR gamma 2 gene and the type 2 diabetes in Qatar.

Neisseria meningitidis elicits a pro-inflammatory response involving IκBζ in a human blood-cerebrospinal fluid barrier model.

PubMed

Borkowski, Julia; Li, Li; Steinmann, Ulrike; Quednau, Natascha; Stump-Guthier, Carolin; Weiss, Christel; Findeisen, Peter; Gretz, Norbert; Ishikawa, Hiroshi; Tenenbaum, Tobias; Schroten, Horst; Schwerk, Christian

2014-09-13

The human-specific, Gram-negative bacterium Neisseria meningitidis (Nm) is a leading cause of bacterial meningitis worldwide. The blood-cerebrospinal fluid barrier (BCSFB), which is constituted by the epithelial cells of the choroid plexus (CP), has been suggested as one of the potential entry sites of Nm into the CSF and can contribute to the inflammatory response during infectious diseases of the brain. Toll-like receptors (TLRs) are involved in mediating signal transduction caused by the pathogens. Using a recently established in vitro model of the human BCSFB based on human malignant CP papilloma (HIBCPP) cells we investigated the cellular response of HIBCPP cells challenged with the meningitis-causing Nm strain, MC58, employing transcriptome and RT-PCR analysis, cytokine bead array, and enzyme-linked immunosorbent assay (ELISA). In comparison, we analyzed the answer to the closely related unencapsulated carrier isolate Nm α14. The presence of TLRs in HIBCPP and their role during signal transduction caused by Nm was studied by RT-PCR and the use of specific agonists and mutant bacteria. We observed a stronger transcriptional response after infection with strain MC58, in particular with its capsule-deficient mutant MC58siaD-, which correlated with bacterial invasion levels. Expression evaluation and Gene Set Enrichment Analysis pointed to a NFκB-mediated pro-inflammatory immune response involving up-regulation of the transcription factor IκBζ. Infected cells secreted significant levels of pro-inflammatory chemokines and cytokines, including, among others, IL8, CXCL1-3, and the IκBζ target gene product IL6. The expression profile of pattern recognition receptors in HIBCPP cells and the response to specific agonists indicates that TLR2/TLR6, rather than TLR4 or TLR2/TLR1, is involved in the cellular reaction following Nm infection. Our data show that Nm can initiate a pro-inflammatory response in human CP epithelial cells probably involving TLR2/TLR6 signaling and the transcriptional regulator IκBζ.

Phospholipase C-{delta}{sub 1} regulates interleukin-1{beta} and tumor necrosis factor-{alpha} mRNA expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chung, Eric; Jakinovich, Paul; Bae, Aekyung

Phospholipase C-{delta}{sub 1} (PLC{delta}{sub 1}) is a widely expressed highly active PLC isoform, modulated by Ca{sup 2+} that appears to operate downstream from receptor signaling and has been linked to regulation of cytokine production. Here we investigated whether PLC{delta}{sub 1} modulated expression of the pro-inflammatory cytokines interleukin-1{beta} (IL-1{beta}), tumor necrosis factor-{alpha} (TNF-{alpha}) and interleukin-6 (IL-6) in rat C6 glioma cells. Expression of PLC{delta}{sub 1} was specifically suppressed by small interfering RNA (siRNA) and the effects on cytokine mRNA expression, stimulated by the Toll-like receptor (TLR) agonist, lipopolysaccharide (LPS), were examined. Real-time polymerase chain reaction (RT-PCR) results showed that PLC{delta}{sub 1}more » knockdown enhanced expression IL-1{beta} and tumor necrosis factor-{alpha} (TNF-{alpha}) mRNA by at least 100 fold after 4 h of LPS stimulation compared to control siRNA treatment. PLC{delta}{sub 1} knock down caused persistently high Nf{kappa}b levels at 4 h of LPS stimulation compared to control siRNA-treated cells. PLC{delta}{sub 1} knockdown was also associated with elevated nuclear levels of c-Jun after 30 min of LPS stimulation, but did not affect LPS-stimulated p38 or p42/44 MAPK phosphorylation, normally associated with TLR activation of cytokine gene expression; rather, enhanced protein kinase C (PKC) phosphorylation of cellular proteins was observed in the absence of LPS stimulation. An inhibitor of PKC, bisindolylmaleimide II (BIM), reversed phosphorylation, prevented elevation of nuclear c-Jun levels, and inhibited LPS-induced increases of IL-1{beta} and TNF-{alpha} mRNA's induced by PLC{delta}{sub 1} knockdown. Our results show that loss of PLC{delta}{sub 1} enhances PKC/c-Jun signaling and up-modulates pro-inflammatory cytokine gene transcription in concert with the TLR-stimulated p38MAPK/Nf{kappa}b pathway. Our findings are consistent with the idea that PLC{delta}{sub 1} is a suppressor of PKC activity.« less

Parsing protection and risk for problem behavior versus pro-social behavior among US and Chinese adolescents.

PubMed

Jessor, Richard; Turbin, Mark S

2014-07-01

This study investigates the different roles played by protective factors and risk factors-and by particular protective and risk factors-when the concern is with accounting for adolescent problem behavior than when the concern is with accounting for adolescent pro-social behavior. The protective and risk factor literature on adolescent problem behavior reveals considerable conceptual and operational ambiguity; an aim of the present study was to advance understanding in this domain of inquiry by providing a systematic conceptualization of protection and risk and of their measurement. Within the systematic framework of Problem Behavior Theory, four protective and four risk factors are assessed in a cross-national study of both problem behavior and pro-social behavior involving large adolescent samples in China (N = 1,368) and the US (N = 1,087), in grades 9, 10, and 11; females 56 %, US; 50 %, China. The findings reveal quite different roles for protection and risk, and for particular protective and risk factors, when the outcome criterion is problem behavior than when it is pro-social behavior. The protective factor, Controls Protection, which engages rule and regulations and sanctions in the adolescent's ecology, emerges as most important in influencing problem behavior, but it plays a relatively minor role in relationship to pro-social behavior. By contrast, Models Protection, the presence of pro-social models in the adolescent's ecology, and Support Protection, the presence of interest and care in that same ecology, have no significant relationship to problem behavior variation, but they are both the major predictors of variation in pro-social behavior. The findings are robust across the samples from the two very diverse societies. These results suggest that greater attention be given to protection in problem behavior research and that a more nuanced perspective is needed about the roles that particular protective and risk factors play in reducing problem behavior and in promoting pro-social behavior.

Association of the gene expression variation of tumor necrosis factor-α and expressions changes of dopamine receptor genes in progression of diabetic severe foot ulcers

PubMed Central

Vaseghi, Hajar; Pornour, Majid; Djavid, Gholamreza Esmaeeli; Rigi, Garshasb; Ganji, Shahla Mohammad; Novin, Leila

2017-01-01

Objective(s): Regulation of pro-inflammatory factors such as TNF-α which are secreted by the immune cells through induction of their several receptors including dopamine receptors (especially DRD2 and DRD3) is one of the noticeable problems in diabetic severe foot ulcer healing. This study was conducted to evaluate the alteration of TNF-αin plasma as well as DRD2 and DRD3 changes in PBMCs of diabetics with severe foot ulcers. Materials and Methods: Peripheral blood samples were collected from 31 subjects with ulcers, 29 without ulcers, and 25 healthy individuals. Total mRNA was extracted from PBMCs for the study of DRD2, DRD3, and TNF-α gene expression variations. Expression patterns of these genes were evaluated by real-time PCR. Consequently, concentration of TNF-α was investigated in plasma. Results: Significant decrease in gene expression and plasma concentration of TNF-α in PBMCs was observed in both patient groups at P<0.05. These diminutions are correlated to the decrease in the expression of both DRD2 and DRD3 in PBMCs of both patient groups. Also, the same relationship is present between expressions of two new DRD3 transcripts with TNF-α downturn. Conclusion: We concluded that DRD2 and DRD3 expression alteration and presence of new DRD3 transcripts can be effective in reduction of TNF-α expression as a pro-inflammatory factor. Performing complementary studies, may explain that variations in DRD2 and DRD3 are prognostic and effective markers attributed to the development of diabetes severe foot ulcers. PMID:29299198

Anti-inflammatory and apoptotic effects of the polyphenol curcumin on human fibroblast-like synoviocytes.

PubMed

Kloesch, Burkhard; Becker, Tatjana; Dietersdorfer, Elisabeth; Kiener, Hans; Steiner, Guenter

2013-02-01

It has recently been reported that the polyphenol curcumin has pronounced anti-carcinogenic, anti-inflammatory and pro-apoptotic properties. This study investigated possible anti-inflammatory and apoptotic effects of curcumin on the human synovial fibroblast cell line MH7A, and on fibroblast-like synoviocytes (FLS) derived from patients with rheumatoid arthritis (RA). MH7A cells and RA-FLS were stimulated either with interleukin (IL)-1β or phorbol 12-myristate 13 acetate (PMA), and treated simultaneously or sequentially with increasing concentrations of curcumin. Release of interleukin (IL)-6 and vascular endothelial growth factor (VEGF)-A was quantified by enzyme-linked immunosorbent assays (ELISAs). In MH7A cells, modulation of the transcription factor nuclear factor kappa-B (NF-κB) and mitogen-activated protein kinases (MAPKs) such as p38 and extracellular-signal regulated kinase (ERK1/2) were analysed by a reporter gene assay and Western blot, respectively. Pro-apoptotic events were monitored by Annexin-V/7-AAD based assay. Cleavage of pro-caspase-3 and -7 was checked with specific antibodies. Curcumin effectively blocked IL-1β and PMA-induced IL-6 expression both in MH7A cells and RA-FLS. VEGF-A expression could only be detected in RA-FLS and was induced by PMA, but not by IL-1β. Furthermore, curcumin inhibited activation of NF-κB and induced dephosphorylation of ERK1/2. Treatment of FLS with high concentrations of curcumin was associated with a decrease in cell viability and induction of apoptosis. The natural compound curcumin represents strong anti-inflammatory properties and induces apoptosis in FLS. This study provides an insight into possible molecular mechanisms of this substance and suggests it as a natural remedy for the treatment of chronic inflammatory diseases like RA. Copyright © 2013 Elsevier B.V. All rights reserved.

Pro-Inflammatory Flagellin Proteins of Prevalent Motile Commensal Bacteria Are Variably Abundant in the Intestinal Microbiome of Elderly Humans

PubMed Central

Neville, B. Anne; Sheridan, Paul O.; Harris, Hugh M. B.; Coughlan, Simone; Flint, Harry J.; Duncan, Sylvia H.; Jeffery, Ian B.; Claesson, Marcus J.; Ross, R. Paul; Scott, Karen P.; O'Toole, Paul W.

2013-01-01

Some Eubacterium and Roseburia species are among the most prevalent motile bacteria present in the intestinal microbiota of healthy adults. These flagellate species contribute “cell motility” category genes to the intestinal microbiome and flagellin proteins to the intestinal proteome. We reviewed and revised the annotation of motility genes in the genomes of six Eubacterium and Roseburia species that occur in the human intestinal microbiota and examined their respective locus organization by comparative genomics. Motility gene order was generally conserved across these loci. Five of these species harbored multiple genes for predicted flagellins. Flagellin proteins were isolated from R. inulinivorans strain A2-194 and from E. rectale strains A1-86 and M104/1. The amino-termini sequences of the R. inulinivorans and E. rectale A1-86 proteins were almost identical. These protein preparations stimulated secretion of interleukin-8 (IL-8) from human intestinal epithelial cell lines, suggesting that these flagellins were pro-inflammatory. Flagellins from the other four species were predicted to be pro-inflammatory on the basis of alignment to the consensus sequence of pro-inflammatory flagellins from the β- and γ- proteobacteria. Many fliC genes were deduced to be under the control of σ28. The relative abundance of the target Eubacterium and Roseburia species varied across shotgun metagenomes from 27 elderly individuals. Genes involved in the flagellum biogenesis pathways of these species were variably abundant in these metagenomes, suggesting that the current depth of coverage used for metagenomic sequencing (3.13–4.79 Gb total sequence in our study) insufficiently captures the functional diversity of genomes present at low (≤1%) relative abundance. E. rectale and R. inulinivorans thus appear to synthesize complex flagella composed of flagellin proteins that stimulate IL-8 production. A greater depth of sequencing, improved evenness of sequencing and improved metagenome assembly from short reads will be required to facilitate in silico analyses of complete complex biochemical pathways for low-abundance target species from shotgun metagenomes. PMID:23935906

A novel IMPDH1 mutation (Arg231Pro) in a family with a severe form of autosomal dominant retinitis pigmentosa.

PubMed

Grover, Sandeep; Fishman, Gerald A; Stone, Edwin M

2004-10-01

To define ophthalmic findings in a family with autosomal dominant retinitis pigmentosa and a novel IMPDH1 gene mutation. Genetic and observational family study. Sixteen affected members of a family with autosomal dominant retinitis pigmentosa. Ophthalmic examination, including best-corrected visual acuity (VA), slit-lamp biomicroscopy, direct and indirect ophthalmoscopy, Goldmann kinetic perimetry, and electroretinography were performed. Deoxyribonucleic acid single-strand conformation polymorphism (SSCP) analysis was done. Abnormal polymerase chain reaction products identified by SSCP analysis were sequenced bidirectionally. All affected patients had the onset of night blindness within the first decade of life. Ocular findings were characterized by diffuse retinal pigmentary degenerative changes, marked restriction of peripheral visual fields, severe loss of VA, nondetectable electroretinography amplitudes, and a high frequency of posterior subcapsular lens opacities. Affected members were observed to harbor a novel IMPDH1 gene mutation. A novel IMPDH1 gene mutation (Arg231Pro) was associated with a severe form of autosomal dominant retinitis pigmentosa. Families affected with a severe form of this genetic subtype should be investigated for a mutation in the IMPDH1 gene.

The influence of occupational chronic lead exposure on the levels of selected pro-inflammatory cytokines and angiogenic factors.

PubMed

Machoń-Grecka, A; Dobrakowski, M; Boroń, M; Lisowska, G; Kasperczyk, A; Kasperczyk, S

2017-05-01

The aim of the study was to determine the effect of occupational exposure to lead on the blood levels of pro-inflammatory cytokines and selected factors that influence angiogenesis. The study population was divided into two groups. The first group consisted of 56 male workers chronically exposed to lead. The second group (control) was comprised of 24 male administrative workers. The serum levels of interleukin 1β (IL-1β), interleukin 6 (IL-6), and tumor necrosis factor α (TNF-α) were significantly higher in the group of workers chronically exposed to lead compared to control values by 38%, 68%, and 57%, respectively. Similarly, the values of soluble vascular endothelial growth factor receptor-1 (sVEGFR-1) and fibroblast growth factor-basic (FGF-basic) were higher by 19% and 63%, respectively. In the group of workers chronically exposed to lead, there were positive correlations between the levels of pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) and angiogenic factors (VEGF, FGF-basic, sVEGFR-1, and soluble angiopoietin receptor). In the control group, there were no correlations between the levels of the abovementioned parameters. Results of the present study indicate that chronic occupational lead exposure promotes inflammatory processes via induction of pro-inflammatory cytokines, modulates angiogenesis, and elicits interdependencies between the immune response and angiogenic factors.

Identification of the transcription factor ZEB1 as a central component of the adipogenic gene regulatory network

PubMed Central

Gubelmann, Carine; Schwalie, Petra C; Raghav, Sunil K; Röder, Eva; Delessa, Tenagne; Kiehlmann, Elke; Waszak, Sebastian M; Corsinotti, Andrea; Udin, Gilles; Holcombe, Wiebke; Rudofsky, Gottfried; Trono, Didier; Wolfrum, Christian; Deplancke, Bart

2014-01-01

Adipose tissue is a key determinant of whole body metabolism and energy homeostasis. Unraveling the regulatory mechanisms underlying adipogenesis is therefore highly relevant from a biomedical perspective. Our current understanding of fat cell differentiation is centered on the transcriptional cascades driven by the C/EBP protein family and the master regulator PPARγ. To elucidate further components of the adipogenic gene regulatory network, we performed a large-scale transcription factor (TF) screen overexpressing 734 TFs in mouse pre-adipocytes and probed their effect on differentiation. We identified 22 novel pro-adipogenic TFs and characterized the top ranking TF, ZEB1, as being essential for adipogenesis both in vitro and in vivo. Moreover, its expression levels correlate with fat cell differentiation potential in humans. Genomic profiling further revealed that this TF directly targets and controls the expression of most early and late adipogenic regulators, identifying ZEB1 as a central transcriptional component of fat cell differentiation. DOI: http://dx.doi.org/10.7554/eLife.03346.001 PMID:25163748

l-Proline Accumulation and Freeze Tolerance in Saccharomyces cerevisiae Are Caused by a Mutation in the PRO1 Gene Encoding γ-Glutamyl Kinase

PubMed Central

Morita, Yuko; Nakamori, Shigeru; Takagi, Hiroshi

2003-01-01

We previously isolated a mutant which showed a high tolerance to freezing that correlated with higher levels of intracellular l-proline derived from l-proline analogue-resistant mutants. The mutation responsible for the analogue resistance and l-proline accumulation was a single nuclear dominant mutation. By introducing the mutant-derived genomic library into a non-l-proline-utilizing strain, the mutant was found to carry an allele of the wild-type PRO1 gene encoding γ-glutamyl kinase, which resulted in a single amino acid replacement; Asp (GAC) at position 154 was replaced by Asn (AAC). Interestingly, the allele of PRO1 was shown to enhance the activities of γ-glutamyl kinase and γ-glutamyl phosphate reductase, both of which catalyze the first two steps of l-proline synthesis from l-glutamate and which together may form a complex in vivo. When cultured in liquid minimal medium, yeast cells expressing the mutated γ-glutamyl kinase were found to accumulate intracellular l-proline and showed a prominent increase in cell viability after freezing at −20°C compared to the viability of cells harboring the wild-type PRO1 gene. These results suggest that the altered γ-glutamyl kinase results in stabilization of the complex or has an indirect effect on γ-glutamyl phosphate reductase activity, which leads to an increase in l-proline production in Saccharomyces cerevisiae. The approach described in this paper could be a practical method for breeding novel freeze-tolerant yeast strains. PMID:12513997

Hypoxia, hypoxia-inducible factors and fibrogenesis in chronic liver diseases.

PubMed

Cannito, Stefania; Paternostro, Claudia; Busletta, Chiara; Bocca, Claudia; Colombatto, Sebastiano; Miglietta, Antonella; Novo, Erica; Parola, Maurizio

2014-01-01

Fibrogenic progression of chronic liver diseases (CLDs) towards the end-point of cirrhosis is currently regarded, whatever the aetiology, as a dynamic and highly integrated cellular response to chronic liver injury. Liver fibrogenesis (i.e., the process) is sustained by hepatic populations of highly proliferative, pro-fibrogenic and contractile myofibroblast-like cells (MFs) that mainly originate from hepatic stellate cells (HSC) or, to a less extent, from portal fibroblasts or bone marrow-derived cells. As is well known, liver fibrosis (i.e., the result) is accompanied by perpetuation of liver injury, chronic hepatitis and persisting activation of tissue repair mechanisms, leading eventually to excess deposition of extracellular matrix (ECM) components. In this scenario, hypoxic areas represent a very common and major feature of fibrotic and cirrhotic liver during the progression of CLDs. Cells exposed to hypoxia respond by means of heterodimeric hypoxia-inducible factors (HIFs) that translocate into the nucleus and binds to a specific core sequence defined hypoxia-responsive element (HRE), present in the promoter on several genes which are considered as hypoxia-regulated target genes. HIFs transcription factors can activate a complex genetic program designed to sustain several changes necessary to efficiently counteract the decrease in oxygen tension. Accordingly, hypoxia, through up-regulation of angiogenesis, is currently believed to significantly contribute to fibrogenic progression of CLDs, mostly by affecting the pro-fibrogenic and pro-angiogenic behaviour of hepatic MFs. In addition, experimental and clinical evidence generated in the last decade also indicates that angiogenesis and fibrogenesis in CLDs may also be sustained by HIF-dependent but hypoxia-independent mediators.

Manduca sexta proprophenoloxidase activating proteinase-3 (PAP3) stimulates melanization by activating proPAP3, proSPHs, and proPOs

PubMed Central

Wang, Yang; Lu, Zhiqiang; Jiang, Haobo

2014-01-01

Melanization participates in various insect physiological processes including antimicrobial immune responses. Phenoloxidase (PO), a critical component of the enzyme system catalyzing melanin formation, is produced as an inactive precursor prophenoloxidase (proPO) and becomes active via specific proteolytic cleavage by proPO activating proteinase (PAP). In Manduca sexta, three PAPs can activate proPOs in the presence of two serine proteinase homologs (SPH1 and SPH2). While the hemolymph proteinases (HPs) that generate the active PAPs are known, it is unclear how the proSPHs (especially proSPH1) are activated. In this study, we isolated from plasma of bar-stage M. sexta larvae an Ile-Glu-Ala-Arg-p-nitroanilide hydrolyzing enzyme that cleaved the proSPHs. This proteinase, PAP3, generated active SPH1 and SPH2, which function as cofactors for PAP3 in proPO activation. Cleavage of the purified recombinant proSPHs by PAP3 yielded 38 kDa bands similar in mobility to the SPHs formed in vivo. Surprisingly, PAP3 also can activate proPAP3 to stimulate melanization in a direct positive feedback loop. The enhanced proPO activation concurred with the cleavage activation of proHP6, proHP8, proPAP1, proPAP3, proSPH1, proSPH2, proPOs, but not proHP14 or proHP21. These results indicate that PAP3, like PAP1, is a key factor of the self-reinforcing mechanism in the proPO activation system, which is linked to other immune responses in M. sexta. PMID:24768974

Phenoxyacetic acid degradation by the 2,4-dichlorophenoxyacetic acid (TFD) pathway of plasmid pJP4: mapping and characterization of the TFD regulatory gene, tfdR.

PubMed Central

Harker, A R; Olsen, R H; Seidler, R J

1989-01-01

Plasmid pJP4 enables Alcaligenes eutrophus JMP134 to degrade 3-chlorobenzoate and 2,4-dichlorophenoxyacetic acid (TFD). Plasmid pRO101 is a derivative of pJP4 obtained by insertion of Tn1721 into a nonessential region of pJP4. Plasmid pRO101 was transferred by conjugation to several Pseudomonas strains and to A. eutrophus AEO106, a cured isolate of JMP134. AEO106(pRO101) and some Pseudomonas transconjugants grew on TFD. Transconjugants with a chromosomally encoded phenol hydroxylase also degraded phenoxyacetic acid (PAA) in the presence of an inducer of the TFD pathway, namely, TFD or 3-chlorobenzoate. A mutant of one such phenol-degrading strain, Pseudomonas putida PPO300(pRO101), grew on PAA as the sole carbon source in the absence of inducer. This isolate carried a mutant plasmid, designated pRO103, derived from pRO101 through the deletion of a 3.9-kilobase DNA fragment. Plasmid pRO103 constitutively expressed the TFD pathway, and this allowed the metabolism of PAA in the absence of the inducer, TFD. Complementation of pRO103 in trans by a DNA fragment corresponding to the fragment deleted in pRO101 indicates that a negative control-regulatory gene (tfdR) is located on the BamHI E fragment of pRO101. Other subcloning experiments resulted in the cloning of the tfdA monooxygenase gene on a 3.5-kilobase fragment derived from pRO101. This subclone, in the absence of other pRO101 DNA, constitutively expressed the tfdA gene and allowed PPO300 to grow on PAA. Preliminary evidence suggests that the monooxygenase activity encoded by this DNA fragment is feedback-inhibited by phenols. Images PMID:2914848

Upregulation of LYAR induces neuroblastoma cell proliferation and survival.

PubMed

Sun, Yuting; Atmadibrata, Bernard; Yu, Denise; Wong, Matthew; Liu, Bing; Ho, Nicholas; Ling, Dora; Tee, Andrew E; Wang, Jenny; Mungrue, Imran N; Liu, Pei Y; Liu, Tao

2017-09-01

The N-Myc oncoprotein induces neuroblastoma by regulating gene transcription and consequently causing cell proliferation. Paradoxically, N-Myc is well known to induce apoptosis by upregulating pro-apoptosis genes, and it is not clear how N-Myc overexpressing neuroblastoma cells escape N-Myc-mediated apoptosis. The nuclear zinc finger protein LYAR has recently been shown to modulate gene expression by forming a protein complex with the protein arginine methyltransferase PRMT5. Here we showed that N-Myc upregulated LYAR gene expression by binding to its gene promoter. Genome-wide differential gene expression studies revealed that knocking down LYAR considerably upregulated the expression of oxidative stress genes including CHAC1, which depletes intracellular glutathione and induces oxidative stress. Although knocking down LYAR expression with siRNAs induced oxidative stress, neuroblastoma cell growth inhibition and apoptosis, co-treatment with the glutathione supplement N-acetyl-l-cysteine or co-transfection with CHAC1 siRNAs blocked the effect of LYAR siRNAs. Importantly, high levels of LYAR gene expression in human neuroblastoma tissues predicted poor event-free and overall survival in neuroblastoma patients, independent of the best current markers for poor prognosis. Taken together, our data suggest that LYAR induces proliferation and promotes survival of neuroblastoma cells by repressing the expression of oxidative stress genes such as CHAC1 and suppressing oxidative stress, and identify LYAR as a novel co-factor in N-Myc oncogenesis.

The role of hypoxia inducible factor-1α in the increased MMP-2 and MMP-9 production by human monocytes exposed to nickel nanoparticles.

PubMed

Wan, Rong; Mo, Yiqun; Chien, Sufan; Li, Yihua; Li, Yixin; Tollerud, David J; Zhang, Qunwei

2011-12-01

Nickel is an important economic commodity, but it can cause skin sensitization and may cause lung diseases such as lung fibrosis, pneumonitis, bronchial asthma and lung cancer. With development of nanotechnology, nano-sized nickel (Nano-Ni) and nano-sized titanium dioxide (Nano-TiO₂) particles have been developed and produced for many years with new formulations and surface properties to meet novel demands. Our previous studies have shown that Nano-Ni instilled into rat lungs caused a greater inflammatory response as compared with standard-sized nickel (5 μm) at equivalent mass concentrations. Nano-Ni caused a persistent high level of inflammation in lungs even at low doses. Recently, several studies have shown that nanoparticles can translocate from the lungs to the circulatory system. To evaluate the potential systemic effects of metal nanoparticles, we compared the effects of Nano-Ni and Nano-TiO₂ on matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9) gene expression and activity. Our results showed that exposure of human monocyte U937 to Nano-Ni caused dose- and time- dependent increase in MMP-2 and MMP-9 mRNA expression and pro-MMP-2 and pro-MMP-9 activity, but Nano-TiO₂ did not. Nano-Ni also caused dose- and time- related increase in tissue inhibitor of metalloproteinases 1 (TIMP-1), but Nano-TiO₂ did not. To determine the potential mechanisms involved, we measured the expression of hypoxia inducible factor 1α (HIF-1α) in U937 cells exposed to Nano-Ni and Nano-TiO₂. Our results showed that exposure to Nano-Ni caused HIF-1α accumulation in the nucleus. Furthermore, pre-treatment of U937 cells with heat shock protein 90 (Hsp90) inhibitor, 17-(Allylamino)-17-demethoxygeldanamycin (17-AAG), prior to exposure to Nano-Ni significantly abolished Nano-Ni-induced MMP-2 and MMP-9 mRNA upregulation and increased pro-MMP-2 and pro-MMP-9 activity. Our results suggest that HIF-1α accumulation may be involved in the increased MMP-2 and MMP-9 production in U937 cells exposed to Nano-Ni.

Frequency of the AGT Pro11Leu polymorphism in humans: Does diet matter?

PubMed

Ségurel, Laure; Lafosse, Sophie; Heyer, Evelyne; Vitalis, Renaud

2010-01-01

The Pro11Leu substitution in the AGXT gene, which causes primary hyperoxaluria type 1, is found with high frequency in some human populations (e.g., 5-20% in Caucasians). It has been suggested that this detrimental mutation could have been positively selected in populations with a meat-rich diet. In order to test this hypothesis, we investigated the occurrence of Pro11Leu in both herder and agriculturalist populations from Central Asia. We found a lower frequency of this detrimental mutation in herders, whose diet is more meat-rich, as compared to agriculturalists, which therefore challenges the universality of the previous claim. Furthermore, when combining our original data with previously published results, we could show that the worldwide genetic differentiation measured at the Pro11Leu polymorphism does not depart from neutrality. Hence, the distribution of the variation observed in the AGXT gene could be due to demographic history, rather than local adaptation to diet.

Kcnh1 Voltage-gated Potassium Channels Are Essential for Early Zebrafish Development*

PubMed Central

Stengel, Rayk; Rivera-Milla, Eric; Sahoo, Nirakar; Ebert, Christina; Bollig, Frank; Heinemann, Stefan H.; Schönherr, Roland; Englert, Christoph

2012-01-01

The Kcnh1 gene encodes a voltage-gated potassium channel highly expressed in neurons and involved in tumor cell proliferation, yet its physiological roles remain unclear. We have used the zebrafish as a model to analyze Kcnh1 function in vitro and in vivo. We found that the kcnh1 gene is duplicated in teleost fish (i.e. kcnh1a and kcnh1b) and that both genes are maternally expressed during early development. In adult zebrafish, kcnh1a and kcnh1b have distinct expression patterns but share expression in brain and testis. Heterologous expression of both genes in Xenopus oocytes revealed a strong conservation of characteristic functional properties between human and fish channels, including a unique sensitivity to intracellular Ca2+/calmodulin and modulation of voltage-dependent gating by extracellular Mg2+. Using a morpholino antisense approach, we demonstrate a strong kcnh1 loss-of-function phenotype in developing zebrafish, characterized by growth retardation, delayed hindbrain formation, and embryonic lethality. This late phenotype was preceded by transcriptional up-regulation of known cell-cycle inhibitors (p21, p27, cdh2) and down-regulation of pro-proliferative factors, including cyclin D1, at 70% epiboly. These results reveal an unanticipated basic activity of kcnh1 that is crucial for early embryonic development and patterning. PMID:22927438

Analysis of the Effects of Five Factors Relevant to In Vitro Chondrogenesis of Human Mesenchymal Stem Cells Using Factorial Design and High Throughput mRNA-Profiling

PubMed Central

Jakobsen, Rune B.; Østrup, Esben; Zhang, Xiaolan; Mikkelsen, Tarjei S.; Brinchmann, Jan E.

2014-01-01

The in vitro process of chondrogenic differentiation of mesenchymal stem cells for tissue engineering has been shown to require three-dimensional culture along with the addition of differentiation factors to the culture medium. In general, this leads to a phenotype lacking some of the cardinal features of native articular chondrocytes and their extracellular matrix. The factors used vary, but regularly include members of the transforming growth factor β superfamily and dexamethasone, sometimes in conjunction with fibroblast growth factor 2 and insulin-like growth factor 1, however the use of soluble factors to induce chondrogenesis has largely been studied on a single factor basis. In the present study we combined a factorial quality-by-design experiment with high-throughput mRNA profiling of a customized chondrogenesis related gene set as a tool to study in vitro chondrogenesis of human bone marrow derived mesenchymal stem cells in alginate. 48 different conditions of transforming growth factor β 1, 2 and 3, bone morphogenetic protein 2, 4 and 6, dexamethasone, insulin-like growth factor 1, fibroblast growth factor 2 and cell seeding density were included in the experiment. The analysis revealed that the best of the tested differentiation cocktails included transforming growth factor β 1 and dexamethasone. Dexamethasone acted in synergy with transforming growth factor β 1 by increasing many chondrogenic markers while directly downregulating expression of the pro-osteogenic gene osteocalcin. However, all factors beneficial to the expression of desirable hyaline cartilage markers also induced undesirable molecules, indicating that perfect chondrogenic differentiation is not achievable with the current differentiation protocols. PMID:24816923

Resolvin D1 protects the liver from ischemia/reperfusion injury by enhancing M2 macrophage polarization and efferocytosis.

PubMed

Kang, Jung-Woo; Lee, Sun-Mee

2016-09-01

Resolution of inflammation is an active process involving a novel category of lipid factors known as specialized pro-resolving lipid mediators, which includes Resolvin D1 (RvD1). While accumulating evidence suggests that RvD1 counteracts proinflammatory signaling and promotes resolution, the specific cellular targets and mechanisms of action of RvD1 remain largely unknown. In the present study, we investigated the role and molecular mechanisms of RvD1 in ischemia/reperfusion (IR)-induced sterile liver inflammation. Male C57BL/6 mice underwent 70% hepatic ischemia for 60min, followed by reperfusion. RvD1 (5, 10, and 15μg/kg, i.p.) was administered to the mice 1h before ischemia and then immediately prior to reperfusion. RvD1 attenuated IR-induced hepatocellular damage and the proinflammatory response. In purified Kupffer cells (KCs) from mice exposed to IR, the levels of M1 marker genes (Nos2a and Cd40) increased, while those of M2 marker genes (Arg1, Cd206, and Mst1r) decreased, demonstrating a proinflammatory shift. RvD1 markedly attenuated these changes. Depletion of KCs by liposome clodronate abrogated the effects of RvD1 on proinflammatory mediators and macrophage polarization. In addition, RvD1 attenuated increases in myeloperoxidase activity and Cxcl1 and Cxcl2 mRNA expression. RvD1 markedly augmented the efferocytic activity of KCs, as indicated by increases in F4/80(+)Gr-1(+) cells in the liver. However, antagonist pretreatment or gene silencing of the RvD1 receptor, ALX/FPR2, abrogated the anti-inflammatory and pro-resolving actions of RvD1. These data indicate that RvD1 ameliorates IR-induced liver injury, and this protection is associated with enhancement of M2 polarization and efferocytosis via ALX/FPR2 activation. Copyright © 2016 Elsevier B.V. All rights reserved.

Urokinase–urokinase receptor interaction mediates an inhibitory signal for HIV-1 replication

PubMed Central

Alfano, Massimo; Sidenius, Nicolai; Panzeri, Barbara; Blasi, Francesco; Poli, Guido

2002-01-01

Elevated levels of soluble urokinase-type plasminogen activator (uPA) receptor, CD87/u-PAR, predict survival in individuals infected with HIV-1. Here, we report that pro-uPA (or uPA) inhibits HIV-1 expression in U937-derived chronically infected promonocytic U1 cells stimulated with phorbol 12-myristate 13-acetate (PMA) or tumor necrosis factor-α (TNF-α). However, pro-uPA did not inhibit PMA or TNF-α-dependent activation of nuclear factor-kB or activation protein-1 in U1 cells. Cell-associated HIV protein synthesis also was not decreased by pro-uPA, although the release of virion-associated reverse transcriptase activity was substantially inhibited, suggesting a functional analogy between pro-uPA and the antiviral effects of IFNs. Indeed, cell disruption reversed the inhibitory effect of pro-uPA on activated U1 cells, and ultrastructural analysis confirmed that virions were preferentially retained within cell vacuoles in pro-uPA treated cells. Neither expression of endogenous IFNs nor activation of the IFN-inducible Janus kinase/signal transducer and activator of transcription pathway were induced by pro-uPA. Pro-uPA also inhibited acute HIV replication in monocyte-derived macrophages and activated peripheral blood mononuclear cells, although with great inter-donor variability. However, pro-uPA inhibited HIV replication in acutely infected promonocytic U937 cells and in ex vivo cultures of lymphoid tissue infected in vitro. Because these effects occurred at concentrations substantially lower than those affecting thrombolysis, pro-uPA may represent a previously uncharacterized class of antiviral agents mimicking IFNs in their inhibitory effects on HIV expression and replication. PMID:12084931

NT-proBNP is superior to BNP for predicting first cardiovascular events in the general population: the Heinz Nixdorf Recall Study.

PubMed

Kara, Kaffer; Lehmann, Nils; Neumann, Till; Kälsch, Hagen; Möhlenkamp, Stefan; Dykun, Iryna; Broecker-Preuss, Martina; Pundt, Noreen; Moebus, Susanne; Jöckel, Karl-Heinz; Erbel, Raimund; Mahabadi, Amir A

2015-03-15

B-type natriuretic peptide (BNP) as well as N-terminal-proBNP (NT-proBNP) are associated with cardiac events in the general population. Yet, data from the general population comparing both peptides for their prognostic value is lacking. Participants from the population-based Heinz-Nixdorf-Recall-study without cardiovascular diseases were included. Associations of BNP and NT-proBNP with incident cardiovascular events (incident myocardial infarction, stroke, or cardiovascular death) were assessed using Cox regression; prognostic value was addressed using Harrell's c statistic. From overall 3589 subjects (mean age: 59.3 ± 7.7 yrs, 52.5% female), 235 subjects developed a cardiovascular event during 8.9 ± 2.2 yrs of follow-up. In regression analysis both natriuretic peptides were associated with incident cardiovascular events, independent of traditional risk factors (hazard ratio (HR) per unit increase on log-scale (95% CI): NT-proBNP: 1.60 (1.39; 1.84); BNP: 1.37 (1.19; 1.58), p<0.0001 respectively). Specifically looking at subjects <60 yrs only NT-proBNP, was linked with events (HR (95% CI): 1.59 (1.19; 2.13) for NT-proBNP, p=0.0019; HR: 1.25 (0.94; 1.65) for BNP, p=0.12, after adjustment for age and gender). Similar results were observed for females (HR (95% CI) 1.65 (1.28; 2.12), p=0.0001 for NT-proBNP, and 1.24 (0.96; 1.61), p=0.10 for BNP after adjustment for age). Adding NT-proBNP/BNP to traditional risk factors increased the prognostic value, with effects being stronger for NT-proBNP (Harrell's c, 0.724 to 0.741, p=0.034) as compared to BNP (0.724 to 0.732, p=0.20). Both, NT-proBNP and BNP are associated with future cardiovascular events in the general population. However, when both are available, NT-proBNP seems to be superior due to its higher prognostic value, especially in younger subjects and females. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

The pVHL172 isoform is not a tumor suppressor and up-regulates a subset of pro-tumorigenic genes including TGFB1 and MMP13

PubMed Central

Hascoet, Pauline; Chesnel, Franck; Jouan, Florence; Goff, Cathy Le; Couturier, Anne; Darrigrand, Eric; Mahe, Fabrice; Rioux-Leclercq, Nathalie; Goff, Xavier Le; Arlot-Bonnemains, Yannick

2017-01-01

The von Hippel-Lindau (VHL) tumor suppressor gene is often deleted or mutated in ccRCC (clear cell renal cell carcinoma) producing a non-functional protein. The gene encodes two mRNA, and three protein isoforms (pVHL213, pVHL160 and pVHL172). The pVHL protein is part of an E3 ligase complex involved in the ubiquitination and proteasomal degradation of different proteins, particularly hypoxia inducible factors (HIF) that drive the transcription of genes involved in the regulation of cell proliferation, angiogenesis or extracellular matrix remodelling. Other non-canonical (HIF-independent) pVHL functions have been described. A recent work reported the expression of the uncharacterized protein isoform pVHL172 which is translated from the variant 2 by alternative splicing of the exon 2. This splice variant is sometimes enriched in the ccRCCs and the protein has been identified in the respective samples of ccRCCs and different renal cell lines. Functional studies on pVHL have only concerned the pVHL213 and pVHL160 isoforms, but no function was assigned to pVHL172. Here we show that pVHL172 stable expression in renal cancer cells does not regulate the level of HIF, exacerbates tumorigenicity when 786-O-pVHL172 cells were xenografted in mice. The pVHL172-induced tumors developed a sarcomatoid phenotype. Moreover, pVHL172 expression was shown to up regulate a subset of pro-tumorigenic genes including TGFB1, MMP1 and MMP13. In summary we identified that pVHL172 is not a tumor suppressor. Furthermore our findings suggest an antagonistic function of this pVHL isoform in the HIF-independent aggressiveness of renal tumors compared to pVHL213. PMID:29100286

Expression of Essential B Cell Development Genes in Horses with Common Variable Immunodeficiency

PubMed Central

Tallmadge, R.L.; Such, K.A.; Miller, K.C.; Matychak, M.B.; Felippe, M.J.B.

2012-01-01

Common variable immunodeficiency (CVID) is a heterogeneous disorder of B cell differentiation or function with inadequate antibody production. Our laboratory studies a natural form of CVID in horses characterized by late-onset B cell lymphopenia due to impaired B cell production in the bone marrow. This study was undertaken to assess the status of B cell differentiation in the bone marrow of CVID-affected horses by measuring the expression of genes essential for early B cell commitment and development. Standard RT-PCR revealed that most of the transcription factors and key signaling molecules that directly regulate B cell differentiation in the bone marrow and precede PAX5 are expressed in the affected horses. Yet, the expression of PAX5 and relevant target genes was variable. Quantitative RT-PCR analysis confirmed that the mRNA expression of E2A, PAX5, CD19, and IGHD was significantly reduced in equine CVID patients when compared to healthy horses (p < 0.05). In addition, the PAX5/EBF1 and PAX5/B220 ratios were significantly reduced in CVID patients (p < 0.01). Immunohistochemical analysis confirmed the absence of PAX5-BSAP expression in the bone marrow of affected horses. Our data suggest that B cell development seems to be impaired at the transition between pre-pro-B cells and pro-B cells in equine CVID patients. PMID:22464097

Gene Expression and Structural Skeletal Responses to Long-Duration Simulated Microgravity in Rats

NASA Technical Reports Server (NTRS)

Shirazi-Fard, Yasaman; Rael, Victoria E.; Torres, Samantha; Steczina, Sonette; Bryant, Sheenah; Tahimic, Candice; Globus, Ruth K.

2017-01-01

In this study, we aim to examine skeletal responses to simulated long-duration spaceflight (90 days) and weight-bearing recovery on bone loss using the ground-based hindlimb unloading (HU) model in adolescent (3-month old) male rats. We hypothesized that simulated microgravity leads to the temporal regulation of oxidative defense genes and pro-bone resorption factors, where there is a progression and eventual plateau; furthermore, early transient changes in these pathways precede skeletal adaptations.

Transcriptional analysis of different stress response genes in Escherichia coli strains subjected to sodium chloride and lactic acid stress.

PubMed

Peng, Silvio; Stephan, Roger; Hummerjohann, Jörg; Tasara, Taurai

2014-12-01

Survival of Escherichia coli in food depends on its ability to adapt against encountered stress typically involving induction of stress response genes. In this study, the transcriptional induction of selected acid (cadA, speF) and salt (kdpA, proP, proW, otsA, betA) stress response genes was investigated among five E. coli strains, including three Shiga toxin-producing strains, exposed to sodium chloride or lactic acid stress. Transcriptional induction upon lactic acid stress exposure was similar in all but one E. coli strain, which lacked the lysine decarboxylase gene cadA. In response to sodium chloride stress exposure, proW and otsA were similarly induced, while significant differences were observed between the E. coli strains in induction of kdpA, proP and betA. The kdpA and betA genes were significantly induced in four and three strains, respectively, whereas one strain did not induce these genes. The proP gene was only induced in two E. coli strains. Interestingly, transcriptional induction differences in response to sodium chloride stress exposure were associated with survival phenotypes observed for the E. coli strains in cheese as the E. coli strain lacking significant induction in three salt stress response genes investigated also survived poorly compared to the other E. coli strains in cheese. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

NUDT2 Disruption Elevates Diadenosine Tetraphosphate (Ap4A) and Down-Regulates Immune Response and Cancer Promotion Genes

PubMed Central

Marriott, Andrew S.; Vasieva, Olga; Fang, Yongxiang; Copeland, Nikki A.; McLennan, Alexander G.; Jones, Nigel J.

2016-01-01

Regulation of gene expression is one of several roles proposed for the stress-induced nucleotide diadenosine tetraphosphate (Ap4A). We have examined this directly by a comparative RNA-Seq analysis of KBM-7 chronic myelogenous leukemia cells and KBM-7 cells in which the NUDT2 Ap4A hydrolase gene had been disrupted (NuKO cells), causing a 175-fold increase in intracellular Ap4A. 6,288 differentially expressed genes were identified with P < 0.05. Of these, 980 were up-regulated and 705 down-regulated in NuKO cells with a fold-change ≥ 2. Ingenuity® Pathway Analysis (IPA®) was used to assign these genes to known canonical pathways and functional networks. Pathways associated with interferon responses, pattern recognition receptors and inflammation scored highly in the down-regulated set of genes while functions associated with MHC class II antigens were prominent among the up-regulated genes, which otherwise showed little organization into major functional gene sets. Tryptophan catabolism was also strongly down-regulated as were numerous genes known to be involved in tumor promotion in other systems, with roles in the epithelial-mesenchymal transition, proliferation, invasion and metastasis. Conversely, some pro-apoptotic genes were up-regulated. Major upstream factors predicted by IPA® for gene down-regulation included NFκB, STAT1/2, IRF3/4 and SP1 but no major factors controlling gene up-regulation were identified. Potential mechanisms for gene regulation mediated by Ap4A and/or NUDT2 disruption include binding of Ap4A to the HINT1 co-repressor, autocrine activation of purinoceptors by Ap4A, chromatin remodeling, effects of NUDT2 loss on transcript stability, and inhibition of ATP-dependent regulatory factors such as protein kinases by Ap4A. Existing evidence favors the last of these as the most probable mechanism. Regardless, our results suggest that the NUDT2 protein could be a novel cancer chemotherapeutic target, with its inhibition potentially exerting strong anti-tumor effects via multiple pathways involving metastasis, invasion, immunosuppression and apoptosis. PMID:27144453

NUDT2 Disruption Elevates Diadenosine Tetraphosphate (Ap4A) and Down-Regulates Immune Response and Cancer Promotion Genes.

PubMed

Marriott, Andrew S; Vasieva, Olga; Fang, Yongxiang; Copeland, Nikki A; McLennan, Alexander G; Jones, Nigel J

2016-01-01

Regulation of gene expression is one of several roles proposed for the stress-induced nucleotide diadenosine tetraphosphate (Ap4A). We have examined this directly by a comparative RNA-Seq analysis of KBM-7 chronic myelogenous leukemia cells and KBM-7 cells in which the NUDT2 Ap4A hydrolase gene had been disrupted (NuKO cells), causing a 175-fold increase in intracellular Ap4A. 6,288 differentially expressed genes were identified with P < 0.05. Of these, 980 were up-regulated and 705 down-regulated in NuKO cells with a fold-change ≥ 2. Ingenuity® Pathway Analysis (IPA®) was used to assign these genes to known canonical pathways and functional networks. Pathways associated with interferon responses, pattern recognition receptors and inflammation scored highly in the down-regulated set of genes while functions associated with MHC class II antigens were prominent among the up-regulated genes, which otherwise showed little organization into major functional gene sets. Tryptophan catabolism was also strongly down-regulated as were numerous genes known to be involved in tumor promotion in other systems, with roles in the epithelial-mesenchymal transition, proliferation, invasion and metastasis. Conversely, some pro-apoptotic genes were up-regulated. Major upstream factors predicted by IPA® for gene down-regulation included NFκB, STAT1/2, IRF3/4 and SP1 but no major factors controlling gene up-regulation were identified. Potential mechanisms for gene regulation mediated by Ap4A and/or NUDT2 disruption include binding of Ap4A to the HINT1 co-repressor, autocrine activation of purinoceptors by Ap4A, chromatin remodeling, effects of NUDT2 loss on transcript stability, and inhibition of ATP-dependent regulatory factors such as protein kinases by Ap4A. Existing evidence favors the last of these as the most probable mechanism. Regardless, our results suggest that the NUDT2 protein could be a novel cancer chemotherapeutic target, with its inhibition potentially exerting strong anti-tumor effects via multiple pathways involving metastasis, invasion, immunosuppression and apoptosis.

Epigenetic Regulation of Inflammatory Gene Expression in Macrophages by Selenium

PubMed Central

Narayan, Vivek; Ravindra, Kodihalli C.; Liao, Chang; Kaushal, Naveen; Carlson, Bradley A.; Prabhu, K. Sandeep

2014-01-01

Acetylation of histone and non-histone proteins by histone acetyltransferases plays a pivotal role in the expression of pro-inflammatory genes. Given the importance of dietary selenium in mitigating inflammation, we hypothesized that selenium supplementation may regulate inflammatory gene expression at the epigenetic level. The effect of selenium towards histone acetylation was examined in both in vitro and in vivo models of inflammation by chromatin immunoprecipitation (ChIP) assays and immunoblotting. Our results indicated that selenium supplementation, as selenite, decreased acetylation of histone H4 at K12 and K16 in COX-2 and TNF promoters, and of the p65 subunit of the redox sensitive transcription factor NFκB in primary and immortalized macrophages. On the other hand, selenomethionine had a much weaker effect. Selenite treatment of HIV-1 infected human monocytes also significantly decreased the acetylation of H4 at K12 and K16 on the HIV-1 promoter, supporting the downregulation of proviral expression by selenium. A similar decrease in histone acetylation was also seen in the colonic extracts of mice treated with dextran sodium sulfate that correlated well with the levels of selenium in the diet. Bone marrow-derived macrophages from Trspfl/flCreLysM mice that lack expression of selenoproteins in macrophages confirmed the important role of selenoproteins in the inhibition of histone H4 acetylation. Our studies suggest that the ability of selenoproteins to skew the metabolism of arachidonic acid to contribute, in part, to their ability to inhibit histone acetylation. In summary, our studies suggest a new role for selenoproteins in the epigenetic modulation of pro-inflammatory genes. PMID:25458528

Betulinic acid inhibits colon cancer cell and tumor growth and induces proteasome-dependent and -independent downregulation of specificity proteins (Sp) transcription factors

PubMed Central

2011-01-01

Background Betulinic acid (BA) inhibits growth of several cancer cell lines and tumors and the effects of BA have been attributed to its mitochondriotoxicity and inhibition of multiple pro-oncogenic factors. Previous studies show that BA induces proteasome-dependent degradation of specificity protein (Sp) transcription factors Sp1, Sp3 and Sp4 in prostate cancer cells and this study focused on the mechanism of action of BA in colon cancer cells. Methods The effects of BA on colon cancer cell proliferation and apoptosis and tumor growth in vivo were determined using standardized assays. The effects of BA on Sp proteins and Sp-regulated gene products were analyzed by western blots, and real time PCR was used to determine microRNA-27a (miR-27a) and ZBTB10 mRNA expression. Results BA inhibited growth and induced apoptosis in RKO and SW480 colon cancer cells and inhibited tumor growth in athymic nude mice bearing RKO cells as xenograft. BA also decreased expression of Sp1, Sp3 and Sp4 transcription factors which are overexpressed in colon cancer cells and decreased levels of several Sp-regulated genes including survivin, vascular endothelial growth factor, p65 sub-unit of NFκB, epidermal growth factor receptor, cyclin D1, and pituitary tumor transforming gene-1. The mechanism of action of BA was dependent on cell context, since BA induced proteasome-dependent and proteasome-independent downregulation of Sp1, Sp3 and Sp4 in SW480 and RKO cells, respectively. In RKO cells, the mechanism of BA-induced repression of Sp1, Sp3 and Sp4 was due to induction of reactive oxygen species (ROS), ROS-mediated repression of microRNA-27a, and induction of the Sp repressor gene ZBTB10. Conclusions These results suggest that the anticancer activity of BA in colon cancer cells is due, in part, to downregulation of Sp1, Sp3 and Sp4 transcription factors; however, the mechanism of this response is cell context-dependent. PMID:21864401

Mechanistic insights into the link between visfatin gene C-1535T polymorphism and coronary artery disease: an in vitro study.

PubMed

Wang, Yong-Sheng; Gao, Wei; Li, Hong-Fen; Wang, Ze-Mu; Zhu, Jun; Zhao, Huan; Yan, Jian-Jun; Jia, En-Zhi; Yang, Zhi-Jian; Wang, Lian-Sheng

2012-04-01

Visfatin, a pro-inflammatory cytokine predominantly released from leucocytes, is correlated with coronary artery disease (CAD). We have previously reported that the -1535C>T polymorphism (rs1330082), which located on the promoter region of visfatin, was associated with decreased risk of CAD. Here, we investigated the underlying mechanism by which this polymorphism affects the genetic susceptibility to CAD. The difference of the promoter activities between -1535T variant and -1535C allele was tested by luciferase reporter gene assay. The difference of transcription factor binding activities between T and C allele was evaluated by electrophoretic mobility shift assay. In reporter gene assay, we showed that the T variant had a significantly reduced transcriptional activity compared with the C allele. The T-variant significantly attenuated the promoter binding affinity to nuclear transcription factors and this effect became much obvious after treatment with TNF-α. Moreover, competition experiment revealed that the retarded complex formed by T-1535- or C-1535-probe binding to nuclear extracts was nearly completely inhibited by unlabeled activator protein-1 (AP-1) specific probe, indicating that AP-1 might be the target nuclear effector. Taken together, our data provided potential mechanistic link between the visfatin -1535C>T polymorphism and reduced CAD risk.

N-terminal pro-brain natriuretic peptide and abnormal brain aging: The AGES-Reykjavik Study.

PubMed

Sabayan, Behnam; van Buchem, Mark A; de Craen, Anton J M; Sigurdsson, Sigurdur; Zhang, Qian; Harris, Tamara B; Gudnason, Vilmundur; Arai, Andrew E; Launer, Lenore J

2015-09-01

To investigate the independent association of serum N-terminal fragment of the prohormone natriuretic peptide (NT-proBNP) with structural and functional features of abnormal brain aging in older individuals. In this cross-sectional study based on the Age, Gene/Environment Susceptibility (AGES)-Reykjavik Study, we included 4,029 older community-dwelling individuals (born 1907 to 1935) with a measured serum level of NT-proBNP. Outcomes included parenchymal brain volumes estimated from brain MRI, cognitive function measured by tests of memory, processing speed, and executive functioning, and presence of depressive symptoms measured using the Geriatric Depression Scale. In a substudy, cardiac output of 857 participants was assessed using cardiac MRI. In multivariate analyses, adjusted for sociodemographic and cardiovascular factors, higher levels of NT-proBNP were independently associated with lower total (p < 0.001), gray matter (p < 0.001), and white matter (p = 0.001) brain volumes. Likewise, in multivariate analyses, higher levels of NT-proBNP were associated with worse scores in memory (p = 0.005), processing speed (p = 0.001), executive functioning (p < 0.001), and more depressive symptoms (p = 0.002). In the substudy, the associations of higher NT-proBNP with lower brain parenchymal volumes, impaired executive function and processing speed, and higher depressive symptoms were independent of the level of cardiac output. Higher serum levels of NT-proBNP, independent of cardiovascular risk factors and a measure of cardiac function, are linked with alterations in brain structure and function. Roles of natriuretic peptides in the process of brain aging need to be further elucidated. © 2015 American Academy of Neurology.

Specialized Transducing Phages Derived from Salmonella Phage P22

PubMed Central

Hoppe, Ingrid; Roth, John

1974-01-01

Salmonella phage P22 has been used in the construction of three sorts of specialized transducing phage: P22 proAB, P22 proABlac and P22 argF. The bacterial genes carried are derived from E. coli K12. Since E. coli and Salmonella chromosomes recombine very poorly, E. coli genes cannot be transduced into Salmonella recipients by P22's generalized transduction mechanism. Therefore, stable inheritance of E. coli material provides a means of detecting specialized transduction. Formation of these phages was possible because the P22 prophage recognizes an attachment site in the E. coli F' prolac episome. Salmonella strains carrying the F' prolac episome can be lysogenized by P22 so as to leave the prophage inserted into the E. coli material of the F' factor. Improper prophage excision can then lead to formation of P22 specialized phages carrying E. coli genetic material. PMID:4599252

Identification of the IGF-1 processing product human Ec/rodent Eb peptide in various tissues: Evidence for its differential regulation after exercise-induced muscle damage in humans.

PubMed

Vassilakos, George; Philippou, Anastassios; Koutsilieris, Michael

2017-02-01

Insulin-like growth factor-1 (IGF-1) is a pleiotropic factor expressed in various tissues and plays a critical role in skeletal muscle physiology. Alternative splicing of the IGF-1 gene gives rise to different precursor polypeptides (isoforms) which could undergo post-translational cleavage, generating the common mature IGF-1 peptide and different carboxyl terminal extension (E-) peptides, with the fate of the latter being, so far, unknown. The objective if this study was to identify the IGF-1Ec forms or processing product(s), other than mature IGF-1, generated in different human and rodent tissues and particularly in human skeletal muscle after exercise-induced damage. Protein lysates from a wide range of human and rodent tissues were immunoblotted with a rabbit anti-human Ec polyclonal antibody raised against the last 24 amino acids of the C-terminal of the Ec peptide. This antibody can recognize the Ec peptide, both as part of IGF-1Ec and alone, and also the corresponding rodent forms, due to the high homology that the human Ec shares with the rodent Eb. We were able to confirm, for the first time, that the human Ec peptide and its rodent homologous Eb peptide are produced simultaneously with their precursor protein (pro-IGF-1Ec/Eb) in vivo, in a wide range of tissues (e.g. muscle, liver, heart). Proprotein convertase furin digestion of human muscle and liver protein lysates confirmed that the higher molecular form, pro-IGF-1Ec, can be cleaved to produce the free Ec peptide. Furthermore, initial evidence is provided that Ec peptide is differentially regulated during the process of muscle regeneration after exercise-induced damage in humans. The findings of this study possibly imply that the post-translational modification of the IGF-1Ec pro-peptide may regulate the bioavailability and activity of the processing product(s). Copyright © 2016. Published by Elsevier Ltd.

Regulation of the angiopoietin-2 gene by hCG in ovarian cancer cell line OVCAR-3.

PubMed

Pietrowski, D; Wiehle, P; Sator, M; Just, A; Keck, C

2010-05-01

Angiogenesis is a crucial step in growing tissues including many tumors. It is regulated by pro- and antiangiogenic factors including the family of angiopoietins and their corresponding receptors. In previous work we have shown that in human ovarian cells the expression of angiopoietin 2 (ANG2) is regulated by human chorionic gonadotropin (hCG). To better understand the mechanisms of hCG-dependent regulation of the ANG2-gene we have now investigated upstream regulatory active elements of the ANG2-promoter in the ovarian carcinoma cell line OVCAR-3. We cloned several ANG2-promoter-fragments of different lengths into a luciferase reporter-gene-vector and analyzed the corresponding ANG2 expression before and after hCG stimulation. We identified regions of the ANG2-promoter between 1 048 bp and 613 bp upstream of the transcriptional start site where hCG-dependent pathways promote a significant downregulation of gene expression. By sequence analysis of this area we found several potential binding sites for transcription factors that are involved in regulation of ANG2-expression, vascular development and ovarian function. These encompass the forkhead family transcription factors FOXC2 and FOXO1 as well as the CCAAT/enhancer binding protein family (C/EBP). In conclusion, we have demonstrated that the regulation of ANG2-expression in ovarian cancer cells is hCG-dependent and we suggest that forkhead transcription factor and C/EBP-dependent pathways are involved in the regulation of ANG2-expression in ovarian cancer cells. Georg Thieme Verlag KG Stuttgart-New York.

The Effects of Exercise on Brain Inflammation

DTIC Science & Technology

2010-06-01

macrophages and lymphocytes release pro -inflammatory cytokines such as interleukin-1β and tumor necrosis factor alpha that induce the subsequent...inflammatory cascade. These pro -inflammatory cytokines are responsible for weakening the BBB so that effector cells from the periphery can enter the CNS...restrictions on Cover and Notice Page: Distribution A – Approved for Public Release DISCUSSION IL-1β Interleukin-1β (IL-1β) is a pro -inflammatory cytokine

PICT-1 triggers a pro-death autophagy through inhibiting rRNA transcription and AKT/mTOR/p70S6K signaling pathway.

PubMed

Chen, Hongbo; Duo, Yanhong; Hu, Bo; Wang, Zhiwei; Zhang, Fang; Tsai, Hsiangi; Zhang, Jianping; Zhou, Lanzhen; Wang, Lijun; Wang, Xinyu; Huang, Laiqiang

2016-11-29

PICT-1 was originally identified as a tumor suppressor. Here, we found that PICT-1 overexpression triggered pro-death autophagy without nucleolar disruption or p53 accumulation in U251 and MCF7 cells. Truncated PICT-1 fragments 181-346 and 1-346, which partly or totally lack nucleolar localization, showed weaker autophagy-inducing effects than full-length PICT-1 and a well-defined nucleolar mutant (181-479). Furthermore, PICT-1 partly localizes to the nucleolar fibrillar center (FC) and directly binds to ribosomal DNA (rDNA) gene loci, where it interacts with upstream binding factor (UBF). Overexpression of PICT-1 or the 181-479 mutant, but not the 1-346 or 181-346 mutants, markedly inhibited the phosphorylation of UBF and the recruitment of rRNA polymerase I (Pol I) to the rDNA promoter in response to serum stimulation, thereby suppressing rRNA transcription, suggesting that rRNA transcription inhibition might be an important contributor to PICT-1-induced autophagy. This is supported by the finding that CX-5461, a specific Pol I inhibitor, also induced autophagy. In addition, both CX-5461 and PICT-1, but not the 1-346 or 181-346 mutants, significantly suppressed the activation of the Akt/mTOR/p70S6K signaling pathway. Our data show that PICT-1 triggers pro-death autophagy through inhibition of rRNA transcription and the inactivation of AKT/mTOR/p70S6K pathway, independent of nucleolar disruption and p53 activation.

Evaluation of Seegene Allplex Respiratory Panel 1 kit for the detection of influenza virus and human respiratory syncytial virus.

PubMed

Gimferrer, Laura; Andrés, Cristina; Rando, Ariadna; Piñana, Maria; Codina, Maria Gema; Martin, Maria Del Carmen; Fuentes, Francisco; Rubio, Susana; Alcubilla, Pilar; Pumarola, Tomàs; Antón, Andrés

2018-05-25

Influenza (FLUV) and human respiratory syncytial (HRSV) viruses are etiological agents of respiratory infections that cause a significant morbidity and mortality worldwide. A rapid and accurate diagnosis of these respiratory viruses is essential for an appropriate patient management. Molecular tests are the best detection option due to their high sensitivity and specificity. Seegene's Allplex™ Respiratory Panel 1 (Allplex RP1) is a real-time one-step RT-PCR assay for the simultaneous detection of FLUAV, FLUBV, HRSV-A and HRSV-B. In addition, it allows the determination of FLUAV subtype (H1, H3 and H1pdm09). This study aims to evaluate Allplex RP1 as a rapid molecular test for the detection of FLUAV, FLUBV, HRSV-A and HRSV-B viruses. The Allplex RP1 assay will be compared with other two commercial molecular assays, Prodesse ProFlu+ and ProFAST+ (Hologic, Madison, WI, USA), and GeneXpert Flu/RSV XC (Cepheid, USA). Allplex RP1, ProFlu+ and GeneXpert tests showed 95%, 91% and 96% of accuracy; and 94%, 88% and 95% of sensitivity, respectively. Moreover, Allplex RP1 showed a FLUAV subtype sensitivity of 91% and 88% for FLUAV-H1pdm09 and FLUAV-H3 respectively, and ProFAST+ assay showed sensitivities of 100% for both targets. The three assays showed a 100% of specificity and PPV, while the NPV were 84%, 73% and 86% for Allplex RP1, Prodesse and GeneXpert, respectively. In this study, Seegene's Allplex RP1 assay showed to be highly sensitive, specific, and suitable for detection of FLUV and HRSV, including FLUAV subtyping. In addition, it is also a hands-on-time saving assay due to the automated nucleic acid extraction and PCR setup. Copyright © 2018. Published by Elsevier B.V.

Salt stress-induced proline transporters and salt stress-repressed broad specificity amino acid permeases identified by suppression of a yeast amino acid permease-targeting mutant.

PubMed Central

Rentsch, D; Hirner, B; Schmelzer, E; Frommer, W B

1996-01-01

A yeast mutant lacking SHR3, a protein specifically required for correct targeting of plasma membrane amino acid permeases, was used to study the targeting of plant transporters and as a tool to isolate new SHR3-independent amino acid transporters. For this purpose, an shr3 mutant was transformed with an Arabidopsis cDNA library. Thirty-four clones were capable of growth under selective conditions, but none showed homology with SHR3. However, genes encoding eight different amino acid transporters belonging to three different transporter families were isolated. Five of these are members of the general amino acid permease (AAP) gene family, one is a member of the NTR family, encoding an oligopeptide transporter, and two belong to a new class of transporter genes. A functional analysis of the latter two genes revealed that they encode specific proline transporters (ProT) that are distantly related to the AAP gene family. ProT1 was found to be expressed in all organs, but highest levels were found in roots, stems, and flowers. Expression in flowers was highest in the floral stalk phloem that enters the carpels and was downregulated after fertilization, indicating a specific role in supplying the ovules with proline. ProT2 transcripts were found ubiquitously throughout the plant, but expression was strongly induced under water or salt stress, implying that ProT2 plays an important role in nitrogen distribution during water stress, unlike members of the AAP gene family whose expression was repressed under the same conditions. These results corroborate the general finding that under water stress, amino acid export is impaired whereas proline export is increased. PMID:8776904

A prodrug of green tea polyphenol (-)-epigallocatechin-3-gallate (Pro-EGCG) serves as a novel angiogenesis inhibitor in endometrial cancer.

PubMed

Wang, Jianzhang; Man, Gene Chi Wai; Chan, Tak Hang; Kwong, Joseph; Wang, Chi Chiu

2018-01-01

Anti-angiogenesis effect of a prodrug of green tea polyphenol (-)-epigallocatechin-3-gallate (Pro-EGCG) in malignant tumors is not well studied. Here, we investigated how the treatment with Pro-EGCG inhibited tumor angiogenesis in endometrial cancer. Tumor xenografts of human endometrial cancer were established and subjected to microarray analysis after Pro-EGCG treatment. First, we showed Pro-EGCG inhibited tumor angiogenesis in xenograft models through down-regulation of vascular endothelial growth factor A (VEGFA) and hypoxia inducible factor 1 alpha (HIF1α) in tumor cells and chemokine (C-X-C motif) ligand 12 (CXCL12) in host stroma by immunohistochemical staining. Next, we investigated how HIF1α/VEGFA was down-regulated and how the reduction of CXCL12 inhibited tumor angiogenesis. We found that VEGFA secretion from endometrial cancer cells was decreased by Pro-EGCG treatment through inhibiting PI3K/AKT/mTOR/HIF1α pathway. Furthermore, the down-regulation of CXCL12 in stromal cells by Pro-EGCG treatment restricted migration and differentiation of macrophages thereby inhibited infiltration of VEGFA-expressing tumor-associated macrophages (TAMs). Taken together, we demonstrated that treatment with Pro-EGCG not only decreases cancer cell-secreted VEGFA but also inhibits TAM-secreted VEGFA in endometrial cancer. These findings demonstrate that Pro-EGCG is a novel angiogenesis inhibitor for endometrial cancer. Copyright © 2017 Elsevier B.V. All rights reserved.

Structure of genes and an insertion element in the methane producing archaebacterium Methanobrevibacter smithii.

PubMed

Hamilton, P T; Reeve, J N

1985-01-01

DNA fragments cloned from the methanogenic archaebacterium Methanobrevibacter smithii which complement mutations in the purE and proC genes of E. coli have been sequenced. Sequence analyses, transposon mutagenesis and expression in E. coli minicells indicate that purE and proC complementations result from the synthesis of M. smithii polypeptides with molecular weights of 36,697 and 27,836 respectively. The encoding genes appear to be located in operons. The M. smithii genome contains 69% A/T basepairs (bp) which is reflected in unusual codon usages and intergenic regions containing approximately 85% A/T bp. An insertion element, designated ISM1, was found within the cloned M. smithii DNA located adjacent to the proC complementing region. ISM1 is 1381 bp in length, has 29 bp terminal inverted repeat sequences and contains one major ORF encoded in 87% of the ISM1 sequence. ISM1 is mobile, present in approximately 10 copies per genome and integration duplicates 8 bp at the site of insertion. The duplicated sequences show homology with sequences within the 29 bp terminal repeat sequence of ISM1. Comparison of our data with sequences from halophilic archaebacteria suggests that 5'GAANTTTCA and 5'TTTTAATATAAA may be consensus promoter sequences for archaebacteria. These sequences closely resemble the consensus sequences which precede Drosophila heat-shock genes (Pelham 1982; Davidson et al. 1983). Methanogens appear to employ the eubacterial system of mRNA: 16SrRNA hybridization to ensure initiation of translation; the consensus ribosome binding sequence is 5'AGGTGA.

Discovery of the type VII ESX-1 secretion needle?

PubMed

Ates, Louis S; Brosch, Roland

2017-01-01

Mycobacterium tuberculosis, the etiological agent of human tuberculosis, harbours five ESAT-6/type VII secretion (ESX/T7S) systems. The first esx gene clusters were identified during the genome-sequencing project of M. tuberculosis H37Rv. Follow-up studies revealed additional genes playing important roles in ESX/T7S systems. Among the latter genes, one can find those that encode Pro-Glu (PE) and Pro-Pro-Glu (PPE) proteins as well as a gene cluster that is encoded >260 kb upstream of the esx-1 locus and encodes ESX-1 secretion-associated proteins EspA (Rv3616c), EspC (Rv3615c) and EspD (Rv3614c). The espACD cluster has been suggested to have an important function in ESX-1 secretion since EspA-EspC and EsxA-EsxB are mutually co-dependent on each other for secretion. However, the molecular mechanism of this co-dependence and interaction between the substrates remained unknown. In this issue of Molecular Microbiology, Lou and colleagues show that EspC forms high-molecular weight polymerization complexes that resemble selected components of type II, III and/or IV secretion systems of Gram-negative bacteria. Indeed, EspC-multimeric complexes form filamentous structures that could well represent a secretion needle of ESX-1 type VII secretion systems. This exciting observation opens new avenues for research to discover and characterize ESX/T7S components and elucidates the co-dependence of EsxA/B secretion with EspA/C. © 2016 John Wiley & Sons Ltd.

Heterozygous Mutations in BMP6 Pro-peptide Lead to Inappropriate Hepcidin Synthesis and Moderate Iron Overload in Humans.

PubMed

Daher, Raed; Kannengiesser, Caroline; Houamel, Dounia; Lefebvre, Thibaud; Bardou-Jacquet, Edouard; Ducrot, Nicolas; de Kerguenec, Caroline; Jouanolle, Anne-Marie; Robreau, Anne-Marie; Oudin, Claire; Le Gac, Gerald; Moulouel, Boualem; Loustaud-Ratti, Veronique; Bedossa, Pierre; Valla, Dominique; Gouya, Laurent; Beaumont, Carole; Brissot, Pierre; Puy, Hervé; Karim, Zoubida; Tchernitchko, Dimitri

2016-03-01

Hereditary hemochromatosis is a heterogeneous group of genetic disorders characterized by parenchymal iron overload. It is caused by defective expression of liver hepcidin, the main regulator of iron homeostasis. Iron stimulates the gene encoding hepcidin (HAMP) via the bone morphogenetic protein (BMP)6 signaling to SMAD. Although several genetic factors have been found to cause late-onset hemochromatosis, many patients have unexplained signs of iron overload. We investigated BMP6 function in these individuals. We sequenced the BMP6 gene in 70 consecutive patients with a moderate increase in serum ferritin and liver iron levels who did not carry genetic variants associated with hemochromatosis. We searched for BMP6 mutations in relatives of 5 probands and in 200 healthy individuals (controls), as well as in 2 other independent cohorts of hyperferritinemia patients. We measured serum levels of hepcidin by liquid chromatography-tandem mass spectrometry and analyzed BMP6 in liver biopsy specimens from patients by immunohistochemistry. The functions of mutant and normal BMP6 were assessed in transfected cells using immunofluorescence, real-time quantitative polymerase chain reaction, and immunoblot analyses. We identified 3 heterozygous missense mutations in BMP6 (p.Pro95Ser, p.Leu96Pro, and p.Gln113Glu) in 6 unrelated patients with unexplained iron overload (9% of our cohort). These mutations were detected in less than 1% of controls. p.Leu96Pro also was found in 2 patients from the additional cohorts. Family studies indicated dominant transmission. Serum levels of hepcidin were inappropriately low in patients. A low level of BMP6, compared with controls, was found in a biopsy specimen from 1 patient. In cell lines, the mutated residues in the BMP6 propeptide resulted in defective secretion of BMP6; reduced signaling via SMAD1, SMAD5, and SMAD8; and loss of hepcidin production. We identified 3 heterozygous missense mutations in BMP6 in patients with unexplained iron overload. These mutations lead to loss of signaling to SMAD proteins and reduced hepcidin production. These mutations might increase susceptibility to mild-to-moderate late-onset iron overload. Copyright © 2016 AGA Institute. Published by Elsevier Inc. All rights reserved.

N-terminal pro–brain natriuretic peptide and abnormal brain aging

PubMed Central

Sabayan, Behnam; van Buchem, Mark A.; de Craen, Anton J.M.; Sigurdsson, Sigurdur; Zhang, Qian; Harris, Tamara B.; Gudnason, Vilmundur; Arai, Andrew E.

2015-01-01

Objective: To investigate the independent association of serum N-terminal fragment of the prohormone natriuretic peptide (NT-proBNP) with structural and functional features of abnormal brain aging in older individuals. Methods: In this cross-sectional study based on the Age, Gene/Environment Susceptibility (AGES)–Reykjavik Study, we included 4,029 older community-dwelling individuals (born 1907 to 1935) with a measured serum level of NT-proBNP. Outcomes included parenchymal brain volumes estimated from brain MRI, cognitive function measured by tests of memory, processing speed, and executive functioning, and presence of depressive symptoms measured using the Geriatric Depression Scale. In a substudy, cardiac output of 857 participants was assessed using cardiac MRI. Results: In multivariate analyses, adjusted for sociodemographic and cardiovascular factors, higher levels of NT-proBNP were independently associated with lower total (p < 0.001), gray matter (p < 0.001), and white matter (p = 0.001) brain volumes. Likewise, in multivariate analyses, higher levels of NT-proBNP were associated with worse scores in memory (p = 0.005), processing speed (p = 0.001), executive functioning (p < 0.001), and more depressive symptoms (p = 0.002). In the substudy, the associations of higher NT-proBNP with lower brain parenchymal volumes, impaired executive function and processing speed, and higher depressive symptoms were independent of the level of cardiac output. Conclusions: Higher serum levels of NT-proBNP, independent of cardiovascular risk factors and a measure of cardiac function, are linked with alterations in brain structure and function. Roles of natriuretic peptides in the process of brain aging need to be further elucidated. PMID:26231259

Growth Factor FGF2 Cooperates with Interleukin-17 to Repair Intestinal Epithelial Damage.

PubMed

Song, Xinyang; Dai, Dai; He, Xiao; Zhu, Shu; Yao, Yikun; Gao, Hanchao; Wang, Jingjing; Qu, Fangfang; Qiu, Ju; Wang, Honglin; Li, Xiaoxia; Shen, Nan; Qian, Youcun

2015-09-15

The intestinal epithelial barrier plays a critical role in the mucosal immunity. However, it remains largely unknown how the epithelial barrier is maintained after damage. Here we show that growth factor FGF2 synergized with interleukin-17 (IL-17) to induce genes for repairing of damaged epithelium. FGF2 or IL-17 deficiency resulted in impaired epithelial proliferation, increased pro-inflammatory microbiota outgrowth, and consequently worse pathology in a DSS-induced colitis model. The dysregulated microbiota in the model induced transforming growth factor beta 1 (TGFβ1) expression, which in turn induced FGF2 expression mainly in regulatory T cells. Act1, an essential adaptor in IL-17 signaling, suppressed FGF2-induced ERK activation through binding to adaptor molecule GRB2 to interfere with its association with guanine nucleotide exchange factor SOS1. Act1 preferentially bound to IL-17 receptor complex, releasing its suppressive effect on FGF2 signaling. Thus, microbiota-driven FGF2 and IL-17 cooperate to repair the damaged intestinal epithelium through Act1-mediated direct signaling cross-talk. Copyright © 2015 Elsevier Inc. All rights reserved.

Helicobacter pylori virulence genes and host genetic polymorphisms as risk factors for peptic ulcer disease.

PubMed

Miftahussurur, Muhammad; Yamaoka, Yoshio

2015-01-01

Helicobacter pylori infection plays an important role in the pathogenesis of peptic ulcer disease (PUD). Several factors have been proposed as possible H. pylori virulence determinants; for example, bacterial adhesins and gastric inflammation factors are associated with an increased risk of PUD. However, differences in bacterial virulence factors alone cannot explain the opposite ends of the PUD disease spectrum, that is duodenal and gastric ulcers; presumably, both bacterial and host factors contribute to the differential response. Carriers of the high-producer alleles of the pro-inflammatory cytokines IL-1B, IL-6, IL-8, IL-10, and TNF-α who also carry low-producer allele of anti-inflammatory cytokines have severe gastric mucosal inflammation, whereas carriers of the alternative alleles have mild inflammation. Recent reports have suggested that the PSCA and CYP2C19 ultra-rapid metabolizer genotypes are also associated with PUD.

Early-onset ocular ochronosis in a girl with alkaptonuria (AKU) and a novel mutation in homogentisate 1,2-dioxygenase (HGD).

PubMed

Gucev, Z S; Slaveska, N; Laban, N; Danilovski, D; Tasic, V; Pop-Jordanova, N; Zatkova, A

2011-01-01

Alkaptonuria (AKU) is a disorder of phenylalanine/tyrosine metabolism due to a defect in the enzyme homogentisate 1,2-dioxygenase (HGD). This recessive disease is caused by mutations in the HGD gene. We report a 14-year-old girl who was referred after presenting black urine. Careful examination revealed ochronosis of the conjunctiva. There was no affection of the cardiac valves. Elevated excretion of homogentisic acid in urine was found. Sequence analysis of the HGD gene from genomic DNA revealed that the patient is a compound heterozygote with a previously described mutation (c.473C>T, p.Pro158Leu), and a novel one (c.821C>T, p.Pro274Leu). Her mother is heterozygous for the novel mutation, while the brother is heterozygous for the previously described mutation. In summary, we describe an alkaptonuric patient with ocular ochronosis and a novel HGD mutation, c.821C>T, p.Pro274Leu.

Association between p53 polymorphism at codon 72 and recurrent spontaneous abortion.

PubMed

Zhang, Ying; Wu, Yuan-Yuan; Qiao, Fu-Yuan; Zeng, Wan-Jiang

2016-06-01

p53 gene plays an important role in apoptosis, which is necessary for successful invasion of trophoblast cells. The change from an arginine (Arg) to a proline (Pro) at codon 72 can influence the biological activity of p53, which predisposes to an increased risk of recurrent spontaneous abortion (RSA). In order to investigate the association between p53 polymorphism at codon 72 and RSA, we conducted this meta-analysis. Pubmed, Embase and Web of science were used to identify the eligible studies. Odds ratio (OR) with 95% confidence interval (CI) was used to evaluate the strength of the association. Six studies containing 937 cases of RSA and 830 controls were included, and there was one study deviated from Hardy-Weinberg equilibrium (HWE). There was a significant association between p53 polymorphism at codon 72 and RSA in recessive model (Pro/Pro vs. Pro/Arg+Arg/Arg; OR=1.60, 95% CI: 1.14-2.24) and co-dominant model (Pro/Pro vs. Arg/Arg; OR=1.47, 95% CI: 1.02-2.12) whether the study that was deviated from HWE was eliminated or not. A significant association was observed in allelic model (Pro vs. Arg; OR=1.28, 95% CI: 1.04-1.57) after exclusion of the study that was deviated from HWE. No association was noted in recessive model (Pro/Pro+Pro/Arg vs. Arg/Arg; OR=1.05, 95% CI: 0.86-1.30) and co-dominant model (Pro/Arg vs. Arg/Arg; OR=0.96, 95% CI: 0.77-1.19). Subgroup analysis by ethnicity also indicated a significant association between p53 polymorphism at codon 72 and RSA in Caucasian group. No heterogeneity and publication bias were found. Our meta-analysis implied that p53 polymorphism at codon 72 carries high maternal risk of RSA.

Non-ribosomal Peptide Synthases from Pseudomonas aeruginosa Play a Role in Cyclodipeptide Biosynthesis, Quorum-Sensing Regulation, and Root Development in a Plant Host.

PubMed

González, Omar; Ortíz-Castro, Randy; Díaz-Pérez, César; Díaz-Pérez, Alma L; Magaña-Dueñas, Viridiana; López-Bucio, José; Campos-García, Jesús

2017-04-01

Diverse molecules mediate cross-kingdom communication between bacteria and their eukaryotic partners and determine pathogenic or symbiotic relationships. N-acyl-L-homoserine lactone-dependent quorum-sensing signaling represses the biosynthesis of bacterial cyclodipeptides (CDPs) that act as auxin signal mimics in the host plant Arabidopsis thaliana. In this work, we performed bioinformatics, biochemical, and plant growth analyses to identify non-ribosomal peptide synthase (NRPS) proteins of Pseudomonas aeruginosa, which are involved in CDP synthesis. A reverse genetics strategy allowed the identification of the genes encoding putative multi-modular-NRPS (MM-NRPS). Mutations in these genes affected the synthesis of the CDPs cyclo(L-Pro-L-Val), cyclo(L-Pro-L-Leu), and cyclo(L-Pro-L-Tyr), while showing wild-type-like levels of virulence factors, such as violacein, elastase, and pyocyanin. When analyzing the bioactivity of purified, naturally produced CDPs, it was found that cyclo(L-Pro-L-Tyr) and cyclo(L-Pro-L-Val) were capable of antagonizing quorum-sensing-LasR (QS-LasR)-dependent signaling in a contrasting manner in the cell-free supernatants of the selected NRPS mutants, which showed QS induction. Using a bacteria-plant interaction system, we further show that the pvdJ, ambB, and pchE P. aeruginosa mutants failed to repress primary root growth, but improved root branching in A. thaliana seedlings. These results indicated that the CDP production in P. aeruginosa depended on the functional MM-NRPS, which influences quorum-sensing of bacteria and plays a role in root architecture remodeling.

ISSLS PRIZE IN BASIC SCIENCE 2018: Growth differentiation factor-6 attenuated pro-inflammatory molecular changes in the rabbit anular-puncture model and degenerated disc-induced pain generation in the rat xenograft radiculopathy model.

PubMed

Miyazaki, Shingo; Diwan, Ashish D; Kato, Kenji; Cheng, Kevin; Bae, Won C; Sun, Yang; Yamada, Junichi; Muehleman, Carol; Lenz, Mary E; Inoue, Nozomu; Sah, Robert L; Kawakami, Mamoru; Masuda, Koichi

2018-04-01

To elucidate the effects of growth differentiation factor-6 (GDF6) on: (i) gene expression of inflammatory/pain-related molecules and structural integrity in the rabbit intervertebral disc (IVD) degeneration model, and (ii) sensory dysfunction and changes in pain-marker expression in dorsal nerve ganglia (DRGs) in the rat xenograft radiculopathy model. Forty-six adolescent rabbits received anular-puncture in two non-consecutive lumbar IVDs. Four weeks later, phosphate-buffered saline (PBS) or GDF6 (1, 10 or 100 µg) was injected into the nucleus pulposus (NP) of punctured discs and followed for 4 weeks for gene expression analysis and 12 weeks for structural analyses. For pain assessment, eight rabbits were sacrificed at 4 weeks post-injection and NP tissues of injected discs were transplanted onto L5 DRGs of 16 nude rats to examine mechanical allodynia. The rat DRGs were analyzed immunohistochemically. In GDF6-treated rabbit NPs, gene expressions of interleukin-6, tumor necrosis factor-α, vascular endothelial growth factor, prostaglandin-endoperoxide synthase 2, and nerve growth factor were significantly lower than those in the PBS group. GDF6 injections resulted in partial restoration of disc height and improvement of MRI disc degeneration grades with statistical significance in rabbit structural analyses. Allodynia induced by xenograft transplantation of rabbit degenerated NPs onto rat DRGs was significantly reduced by GDF6 injection. Staining intensities for ionized calcium-binding adaptor molecule-1 and calcitonin gene-related peptide in rat DRGs of the GDF6 group were significantly lower than those of the PBS group. GDF6 injection may change the pathological status of degenerative discs and attenuate degenerated IVD-induced pain.

Identification of the APC/C co-factor FZR1 as a novel therapeutic target for multiple myeloma.

PubMed

Crawford, Lisa J; Anderson, Gordon; Johnston, Cliona K; Irvine, Alexandra E

2016-10-25

Multiple Myeloma (MM) is a haematological neoplasm characterised by the clonal proliferation of malignant plasma cells in the bone marrow. The success of proteasome inhibitors in the treatment of MM has highlighted the importance of the ubiquitin proteasome system (UPS) in the pathogenesis of this disease. In this study, we analysed gene expression of UPS components to identify novel therapeutic targets within this pathway in MM. Here we demonstrate how this approach identified previously validated and novel therapeutic targets. In addition we show that FZR1 (Fzr), a cofactor of the multi-subunit E3 ligase complex anaphase-promoting complex/cyclosome (APC/C), represents a novel therapeutic target in myeloma. The APC/C associates independently with two cofactors, Fzr and Cdc20, to control cell cycle progression. We found high levels of FZR1 in MM primary cells and cell lines and demonstrate that expression is further increased on adhesion to bone marrow stromal cells (BMSCs). Specific knockdown of either FZR1 or CDC20 reduced viability and induced growth arrest of MM cell lines, and resulted in accumulation of APC/CFzr substrate Topoisomerase IIα (TOPIIα) or APC/CCdc20 substrate Cyclin B. Similar effects were observed following treatment with proTAME, an inhibitor of both APC/CFzr and APC/CCdc20. Combinations of proTAME with topoisomerase inhibitors, etoposide and doxorubicin, significantly increased cell death in MM cell lines and primary cells, particularly if TOPIIα levels were first increased through pre-treatment with proTAME. Similarly, combinations of proTAME with the microtubule inhibitor vincristine resulted in enhanced cell death. This study demonstrates the potential of targeting the APC/C and its cofactors as a therapeutic approach in MM.

Codon usage analysis of photolyase encoding genes of cyanobacteria inhabiting diverse habitats.

PubMed

Rajneesh; Pathak, Jainendra; Kannaujiya, Vinod K; Singh, Shailendra P; Sinha, Rajeshwar P

2017-07-01

Nucleotide and amino acid compositions were studied to determine the genomic and structural relationship of photolyase gene in freshwater, marine and hot spring cyanobacteria. Among three habitats, photolyase encoding genes from hot spring cyanobacteria were found to have highest GC content. The genomic GC content was found to influence the codon usage and amino acid variability in photolyases. The third position of codon was found to have more effect on amino acid variability in photolyases than the first and second positions of codon. The variation of amino acids Ala, Asp, Glu, Gly, His, Leu, Pro, Gln, Arg and Val in photolyases of three different habitats was found to be controlled by first position of codon (G1C1). However, second position (G2C2) of codon regulates variation of Ala, Cys, Gly, Pro, Arg, Ser, Thr and Tyr contents in photolyases. Third position (G3C3) of codon controls incorporation of amino acids such as Ala, Phe, Gly, Leu, Gln, Pro, Arg, Ser, Thr and Tyr in photolyases from three habitats. Photolyase encoding genes of hot spring cyanobacteria have 85% codons with G or C at third position, whereas marine and freshwater cyanobacteria showed 82 and 60% codons, respectively, with G or C at third position. Principal component analysis (PCA) showed that GC content has a profound effect in separating the genes along the first major axis according to their RSCU (relative synonymous codon usage) values, and neutrality analysis indicated that mutational pressure has resulted in codon bias in photolyase genes of cyanobacteria.

Redox-linked effects of green tea on DNA damage and repair, and influence of microsatellite polymorphism in HMOX-1: results of a human intervention trial.

PubMed

Choi, Siu-Wai; Yeung, Vincent T F; Collins, Andrew R; Benzie, Iris F F

2015-01-01

Green tea has many reported health benefits, including genoprotective and antioxidant effects, but green tea has pro-oxidant activity in vitro. A tea-induced pro-oxidant shift that triggers cytoprotective adaptations has been postulated, but human data are lacking. We investigated effects on oxidation-induced DNA damage and redox-linked cytoprotective factors, including 8-oxoguanine glycosylase (hOGG1) and heme oxygenase 1 (HMOX-1) in lymphocytes in a randomised, placebo-controlled, cross-over supplementation trial. hOGG1 catalyses the first step in base excision repair; increased HMOX-1 is a sign of cytoprotective response to pro-oxidant change. The influence of microsatellite polymorphisms in the HMOX-1 promoter region was also explored. Higher numbers of GT repeats [GT(n)] in this region reportedly diminish response to pro-oxidant change. Green tea [2 × 150 ml of 1% w/v tea/day (or water as control)] was taken for 12 weeks by 43 Type 2 diabetes subjects {20 with short [S/S; GT(n) < 25] and 23 with long [L/L; GT(n) ≥ 25]}. Fasting venous blood was collected before and after each treatment. The formamidopyrimidine DNA glycosylase-assisted comet assay was used to measure DNA damage in lymphocytes. For measuring hOGG1 activity, we used photo-damaged HeLa cells incubated with lymphocyte extracts from test subjects, in combination with the comet assay. Lymphocyte HMOX-1 and hOGG1 protein concentrations and expression (mRNA) of redox-sensitive genes, including HMOX-1 and hOGG1, were also investigated. Results showed significantly (P < 0.01) lower (~15%) DNA damage, higher (~50%) hOGG1 activity and higher (~40%) HMOX-1 protein concentration after tea. No changes in mRNA expression were seen. Baseline HMOX-1 protein and hOGG1 activity were higher (P < 0.05) in the S/S group, but tea-associated responses were similar in both GT(n) groups. Green tea is clearly associated with lowered DNA damage, increased hOGG1 activity and higher HMOX-1 protein levels. Further study is needed to confirm a cause and effect relationship and to establish if these effects are mediated by post-translational changes in proteins or by increased gene expression. © The Author 2014. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

NHR-49/HNF4 integrates regulation of fatty acid metabolism with a protective transcriptional response to oxidative stress and fasting.

PubMed

Goh, Grace Y S; Winter, Johnathan J; Bhanshali, Forum; Doering, Kelsie R S; Lai, Regina; Lee, Kayoung; Veal, Elizabeth A; Taubert, Stefan

2018-06-01

Endogenous and exogenous stresses elicit transcriptional responses that limit damage and promote cell/organismal survival. Like its mammalian counterparts, hepatocyte nuclear factor 4 (HNF4) and peroxisome proliferator-activated receptor α (PPARα), Caenorhabditis elegans NHR-49 is a well-established regulator of lipid metabolism. Here, we reveal that NHR-49 is essential to activate a transcriptional response common to organic peroxide and fasting, which includes the pro-longevity gene fmo-2/flavin-containing monooxygenase. These NHR-49-dependent, stress-responsive genes are also upregulated in long-lived glp-1/notch receptor mutants, with two of them making critical contributions to the oxidative stress resistance of wild-type and long-lived glp-1 mutants worms. Similar to its role in lipid metabolism, NHR-49 requires the mediator subunit mdt-15 to promote stress-induced gene expression. However, NHR-49 acts independently from the transcription factor hlh-30/TFEB that also promotes fmo-2 expression. We show that activation of the p38 MAPK, PMK-1, which is important for adaptation to a variety of stresses, is also important for peroxide-induced expression of a subset of NHR-49-dependent genes that includes fmo-2. However, organic peroxide increases NHR-49 protein levels, by a posttranscriptional mechanism that does not require PMK-1 activation. Together, these findings establish a new role for the HNF4/PPARα-related NHR-49 as a stress-activated regulator of cytoprotective gene expression. © 2018 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

In vitro infectivity and differential gene expression of Leishmania infantum metacyclic promastigotes: negative selection with peanut agglutinin in culture versus isolation from the stomodeal valve of Phlebotomus perniciosus.

PubMed

Alcolea, Pedro J; Alonso, Ana; Degayón, María A; Moreno-Paz, Mercedes; Jiménez, Maribel; Molina, Ricardo; Larraga, Vicente

2016-05-20

Leishmania infantum is the protozoan parasite responsible for zoonotic visceral leishmaniasis in the Mediterranean basin. A recent outbreak in humans has been reported in this area. The life cycle of the parasite is digenetic. The promastigote stage develops within the gut of phlebotomine sand flies, whereas amastigotes survive and multiply within phagolysosomes of mammalian host phagocytes. The major vector of L. infantum in Spain is Phlebotomus perniciosus. The axenic culture model of promastigotes is generally used because it is able to mimic the conditions of the natural environment (i.e. the sand fly vector gut). However, infectivity decreases with culture passages and infection of laboratory animals is frequently required. Enrichment of the stationary phase population in highly infective metacyclic promastigotes is achieved by negative selection with peanut agglutinin (PNA), which is possible only in certain Leishmania species such as L. major and L. infantum. In this study, in vitro infectivity and differential gene expression of cultured PNA-negative promastigotes (Pro-PNA(-)) and metacyclic promastigotes isolated from the sand fly anterior thoracic midgut (Pro-Pper) have been compared. In vitro infectivity is about 30 % higher in terms of rate of infected cells and number of amastigotes per infected cell in Pro-Pper than in Pro-PNA(-). This finding is in agreement with up-regulation of a leishmanolysin gene (gp63) and genes involved in biosynthesis of glycosylinositolphospholipids (GIPL), lipophosphoglycan (LPG) and proteophosphoglycan (PPG) in Pro-Pper. In addition, differences between Pro-Pper and Pro-PNA(-) in genes involved in important cellular processes (e.g. signaling and regulation of gene expression) have been found. Pro-Pper are significantly more infective than peanut lectin non-agglutinating ones. Therefore, negative selection with PNA is an appropriate method for isolating metacyclic promastigotes in stationary phase of axenic culture but it does not allow reaching the in vitro infectivity levels of Pro-Pper. Indeed, GIPL, LPG and PPG biosynthetic genes together with a gp63 gene are up-regulated in Pro-Pper and interestingly, the correlation coefficient between both transcriptomes in terms of transcript abundance is R (2) = 0.68. This means that the correlation is sufficiently high to consider that both samples are physiologically comparable (i.e. the experiment was correctly designed and performed) and sufficiently low to conclude that important differences in transcript abundance have been found. Therefore, the implications of axenic culture should be evaluated case-by-case in each experimental design even when the stationary phase population in culture is enriched in metacyclic promastigotes by negative selection with PNA.

Identification of a novel homozygous mutation Arg459Pro in SYNJ1 gene of an Indian family with autosomal recessive juvenile Parkinsonism.

PubMed

Kirola, Laxmi; Behari, Madhuri; Shishir, Chandan; Thelma, B K

2016-10-01

A novel homozygous missense mutation (c.773G > A, p.Arg258Gln) in Synaptojanin 1 (SYNJ1, 21q22.2) has recently been reported in two Italian and one Iranian consanguineous families with autosomal recessive juvenile Parkinsonism (ARJP). Contribution of this synaptic gene related to Parkinsonism phenotypes in other populations still remains unidentified. An ARJP family with two affected siblings characterized by frequent tremor with bradykinesia and rigidity was recruited in this study. Both siblings showed intense dyskinesia and dystonia on administration of Syndopa. The family was analyzed for both mutations and exon dosage variations in PARKIN, PINK1 and DJ1. Further, whole exome sequencing was performed in two affected and one unaffected sibling in the family. We identified a novel homozygous mutation (c.1376C > G, p.Arg459Pro) in SYNJ1 segregating in this family. This p.Arg459Pro mutation was not observed in 285 additional Parkinson disease (PD) samples (32 familial, 81 early onset and 172 late onset) screened by PCR-Sanger-sequencing. It was also absent in dbSNP, 1000 Genomes, ExAC, NHLBI-ESP database and in >250 ethnically matched exomes available in our laboratory. The arginine residue is highly conserved across species and predicted to be damaging by several in silico tools. As with the previous mutation p.Arg258Gln, p.Arg459Pro is also present in Sac 1 domain of SYNJ1 wherein p.Arg258Gln mutation has already been described to impair the phosphatase activity. We report another novel mutation in SYNJ1 of an Indian consanguineous ARJP family. Finding an additional mutation in this gene further supports the involvement of SYNJ1 in PD pathogenesis across different ethnicities. Copyright © 2016 Elsevier Ltd. All rights reserved.

Structure and activation of pro-activin A

PubMed Central

Wang, Xuelu; Fischer, Gerhard; Hyvönen, Marko

2016-01-01

Activins are growth factors with multiple roles in the development and homeostasis. Like all TGF-β family of growth factors, activins are synthesized as large precursors from which mature dimeric growth factors are released proteolytically. Here we have studied the activation of activin A and determined crystal structures of the unprocessed precursor and of the cleaved pro-mature complex. Replacing the natural furin cleavage site with a HRV 3C protease site, we show how the protein gains its bioactivity after proteolysis and is as active as the isolated mature domain. The complex remains associated in conditions used for biochemical analysis with a dissociation constant of 5 nM, but the pro-domain can be actively displaced from the complex by follistatin. Our high-resolution structures of pro-activin A share features seen in the pro-TGF-β1 and pro-BMP-9 structures, but reveal a new oligomeric arrangement, with a domain-swapped, cross-armed conformation for the protomers in the dimeric protein. PMID:27373274

Osteoarthritis year in review 2015: biology.

PubMed

Malfait, A M

2016-01-01

This review highlights a selection of recently published literature in the area of osteoarthritis biology. Major themes transpiring from a PubMed search covering the year between the 2014 and the 2015 Osteoarthritis Research Society International (OARSI) World Congress are explored. Inflammation emerged as a significant theme, revealing complex pathways that drive dramatic changes in cartilage homeostasis and in the synovium. Highlights include a homeostatic role for CXC chemokines in cartilage, identification of the zinc-ZIP8-MTF1 axis as an essential regulator of cartilage catabolism, and the discovery that a small aggrecan fragment can have catabolic and pro-inflammatory effects through Toll-like receptor 2. Synovitis can promote joint damage, partly through alarmins such as S100A8. Synovitis and synovial expression of the pro-algesic neurotrophin, Nerve Growth Factor, are associated with pain. Increasingly, researchers are considering specific pathogenic pathways that may operate in distinct subsets of osteoarthritis associated with distinct risk factors, including obesity, age, and joint injury. In obesity, the contribution of metabolic factors and diet is under intense investigation. The role of autophagy and oxidative stress in age-related osteoarthritis has been further explored. This approach may open avenues for targeted treatment of distinct phenotypes of osteoarthritis. Finally, a small selection of novel analgesic targets in the periphery is briefly discussed, including calcitonin gene-related peptide and the neuronal sodium voltage-gated channels, Nav1.7 and Nav1.8. Copyright © 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Lipophilic components of diesel exhaust particles induce pro-inflammatory responses in human endothelial cells through AhR dependent pathway(s).

PubMed

Brinchmann, Bendik C; Skuland, Tonje; Rambøl, Mia H; Szoke, Krisztina; Brinchmann, Jan E; Gutleb, Arno C; Moschini, Elisa; Kubátová, Alena; Kukowski, Klara; Le Ferrec, Eric; Lagadic-Gossmann, Dominique; Schwarze, Per E; Låg, Marit; Refsnes, Magne; Øvrevik, Johan; Holme, Jørn A

2018-05-11

Exposure to traffic-derived particulate matter (PM), such as diesel exhaust particles (DEP), is a leading environmental cause of cardiovascular disease (CVD), and may contribute to endothelial dysfunction and development of atherosclerosis. It is still debated how DEP and other inhaled PM can contribute to CVD. However, organic chemicals (OC) adhered to the particle surface, are considered central to many of the biological effects. In the present study, we have explored the ability of OC from DEP to reach the endothelium and trigger pro-inflammatory reactions, a central step on the path to atherosclerosis. Exposure-relevant concentrations of DEP (0.12 μg/cm 2 ) applied on the epithelial side of an alveolar 3D tri-culture, rapidly induced pro-inflammatory and aryl hydrocarbon receptor (AhR)-regulated genes in the basolateral endothelial cells. These effects seem to be due to soluble lipophilic constituents rather than particle translocation. Extractable organic material of DEP (DEP-EOM) was next fractionated with increasing polarity, chemically characterized, and examined for direct effects on pro-inflammatory and AhR-regulated genes in human microvascular endothelial (HMEC-1) cells and primary human endothelial cells (PHEC) from four healthy donors. Exposure-relevant concentrations of lipophilic DEP-EOM (0.15 μg/cm 2 ) induced low to moderate increases in IL-1α, IL-1β, COX2 and MMP-1 gene expression, and the MMP-1 secretion was increased. By contrast, the more polar EOM had negligible effects, even at higher concentrations. Use of pharmacological inhibitors indicated that AhR and protease-activated receptor-2 (PAR-2) were central in regulation of EOM-induced gene expression. Some effects also seemed to be attributed to redox-responses, at least at the highest exposure concentrations tested. Although the most lipophilic EOM, that contained the majority of PAHs and aliphatics, had the clearest low-concentration effects, there was no straight-forward link between chemical composition and biological effects. Lipophilic and semi-lipophilic chemicals seemed to detach from DEP, translocate through alveolar epithelial cells and trigger pro-inflammatory reactions in endothelial cells at exposure-relevant concentrations. These effects appeared to be triggered by AhR agonists, and involve PAR-2 signaling.

Meta-analysis of association between the Pro12Ala polymorphism of the peroxisome proliferator-activated receptor-γ2 gene and diabetic retinopathy in Caucasians and Asians.

PubMed

Ma, Jinlan; Li, Yan; Zhou, Fang; Xu, Xiaoyi; Guo, Gang; Qu, Yi

2012-01-01

The Pro12Ala polymorphism of the peroxisome proliferator-activated receptor-γ2 (PPARγ2) gene is reported to be associated with diabetes. However, the gene's association with diabetic retinopathy (DR) in type 2 diabetes mellitus (T2DM) has been investigated in numerous epidemiologic studies with controversial results. This meta-analysis aimed to collectively assess the association of the Pro12Ala polymorphism with DR in T2DM. An electronic literature search was conducted on PubMed, ISI Web of Knowledge, EMBASE, and the China National Knowledge Internet. A dominant model [(Pro/Ala +Ala/Ala) versus Pro/Pro] was used to ensure adequate statistical power. Crude odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using the fixed effect model. Potential sources of heterogeneity and bias were explored. This meta-analysis included genotype data from 2,720 cases with DR and 2,450 controls free of DR from eight eligible publications. The results showed the Ala allele had a protective effect on DR in T2DM (OR=0.81; 95% CI: 0.68-0.98, p=0.03). There was no significant evidence against homogeneity (I(2)=46%, P(heterogeneity)=0.07). The sensitivity analysis showed a robust association of the Pro12Ala polymorphism with DR in T2DM after a study involving Caucasians that presented a big effect on heterogeneity (OR=0.75; 95% CI: 0.62-0.91, p=0.003) was excluded. Possible ethnic differences in the association of the Pro12Ala single nucleotide polymorphism and DR were demonstrated; a significant association was illustrated in the Caucasian subgroup (OR=0.74; 95% CI: 0.59-0.94, p=0.01) but was not found in the Asian subgroup (OR=0.77; 95% CI: 0.55-1.07, p=0.12). No publication bias was observed. This meta-analysis suggested a significant association exists between the Pro12Ala polymorphism and DR in T2DM with ethnic differences. The Ala allele had a significant protective effect against DR in T2DM.

The significance of p53 codon 72 polymorphism for the development of cervical adenocarcinomas

PubMed Central

Andersson, S; Rylander, E; Strand, A; Sällström, J; Wilander, E

2001-01-01

Infection with the human papillomavirus is an important co-factor in the development of cervical carcinomas. Accordingly, HPV DNA is recognised in most of these tumours. Polymorphism of the p53 gene, codon 72, is also considered a risk factor in the development of cervical carcinoma. However, this finding is contradicted by several observers. In the present investigation, 111 cases of adenocarcinoma of the cervix collected through the Swedish Cancer Registry and 188 controls (females with normal cytology at organised gynaecological screening) were analysed with regard to p53, codon 72, polymorphism using a PCR- and SSCP-based technique. In the controls, 9% showed pro/pro, 44% pro/arg and 47% arg/arg, whereas in the invasive adenocarcinomas, the corresponding figures were 0%, 29% and 71%, respectively. The difference was statistically significant (P = 0.001). HPV DNA was identified in 86 tumours (HPV 18 in 48, HPV 16 in 31 and HPV of unknown type in 7 cases) and 25 tumours were HPV negative. The p53, codon 72, genotypes observed in HPV-positive and HPV-negative cervical adenocarcinomas were not statistically different (P = 0.690). The results indicate that women homozygotic for arg/arg in codon 72 of the p53 gene are at an increased risk for the development of cervical adenocarcinomas. However, this genetic disposition seems to be unrelated to the HPV infection. © 2001 Cancer Research Campaign  http://www.bjcancer.com PMID:11710828

Osteopontin ablation ameliorates muscular dystrophy by shifting macrophages to a pro-regenerative phenotype

PubMed Central

Capote, Joana; Martinez, Leonel; Vetrone, Sylvia; Barton, Elisabeth R.; Sweeney, H. Lee; Miceli, M. Carrie

2016-01-01

In the degenerative disease Duchenne muscular dystrophy, inflammatory cells enter muscles in response to repetitive muscle damage. Immune factors are required for muscle regeneration, but chronic inflammation creates a profibrotic milieu that exacerbates disease progression. Osteopontin (OPN) is an immunomodulator highly expressed in dystrophic muscles. Ablation of OPN correlates with reduced fibrosis and improved muscle strength as well as reduced natural killer T (NKT) cell counts. Here, we demonstrate that the improved dystrophic phenotype observed with OPN ablation does not result from reductions in NKT cells. OPN ablation skews macrophage polarization toward a pro-regenerative phenotype by reducing M1 and M2a and increasing M2c subsets. These changes are associated with increased expression of pro-regenerative factors insulin-like growth factor 1, leukemia inhibitory factor, and urokinase-type plasminogen activator. Furthermore, altered macrophage polarization correlated with increases in muscle weight and muscle fiber diameter, resulting in long-term improvements in muscle strength and function in mdx mice. These findings suggest that OPN ablation promotes muscle repair via macrophage secretion of pro-myogenic growth factors. PMID:27091452

TNFα- and IKKβ-mediated TANK/I-TRAF phosphorylation: implications for interaction with NEMO/IKKγ and NF-κB activation

PubMed Central

Bonif, Marianne; Meuwis, Marie-Alice; Close, Pierre; Benoit, Valérie; Heyninck, Karen; Chapelle, Jean-Paul; Bours, Vincent; Merville, Marie-Paule; Piette, Jacques; Beyaert, Rudi; Chariot, Alain

2005-01-01

Pro-inflammatory cytokines trigger signalling cascades leading to NF-κB (nuclear factor-κB)-dependent gene expression through IKK [IκB (inhibitory κB) kinase]-dependent phosphorylation and subsequent degradation of the IκB proteins and via induced phosphorylation of p65. These signalling pathways rely on sequentially activated kinases which are assembled by essential and non-enzymatic scaffold proteins into functional complexes. Here, we show that the pro-inflammatory cytokine TNFα (tumour necrosis factor α) promotes TANK [TRAF (TNF receptor-associated factor) family member associated NF-κB activator] recruitment to the IKK complex via a newly characterized C-terminal zinc finger. Moreover, we show that TANK is phosphorylated by IKKβ upon TNFα stimulation and that this modification negatively regulates TANK binding to NEMO (NF-κB essential modulator). Interestingly, reduced TANK expression by RNA interference attenuates TNFα-mediated induction of a subset of NF-κB target genes through decreased p65 transactivation potential. Therefore the scaffold protein TANK is required for the cellular response to TNFα by connecting upstream signalling molecules to the IKKs and p65, and its subsequent IKKβ-mediated phosphorylation may be a mechanism to terminate the TANK-dependent wave of NF-κB activation. PMID:16336209

Query-based biclustering of gene expression data using Probabilistic Relational Models.

PubMed

Zhao, Hui; Cloots, Lore; Van den Bulcke, Tim; Wu, Yan; De Smet, Riet; Storms, Valerie; Meysman, Pieter; Engelen, Kristof; Marchal, Kathleen

2011-02-15

With the availability of large scale expression compendia it is now possible to view own findings in the light of what is already available and retrieve genes with an expression profile similar to a set of genes of interest (i.e., a query or seed set) for a subset of conditions. To that end, a query-based strategy is needed that maximally exploits the coexpression behaviour of the seed genes to guide the biclustering, but that at the same time is robust against the presence of noisy genes in the seed set as seed genes are often assumed, but not guaranteed to be coexpressed in the queried compendium. Therefore, we developed ProBic, a query-based biclustering strategy based on Probabilistic Relational Models (PRMs) that exploits the use of prior distributions to extract the information contained within the seed set. We applied ProBic on a large scale Escherichia coli compendium to extend partially described regulons with potentially novel members. We compared ProBic's performance with previously published query-based biclustering algorithms, namely ISA and QDB, from the perspective of bicluster expression quality, robustness of the outcome against noisy seed sets and biological relevance.This comparison learns that ProBic is able to retrieve biologically relevant, high quality biclusters that retain their seed genes and that it is particularly strong in handling noisy seeds. ProBic is a query-based biclustering algorithm developed in a flexible framework, designed to detect biologically relevant, high quality biclusters that retain relevant seed genes even in the presence of noise or when dealing with low quality seed sets.

Cardiac stress and inflammatory markers as predictors of heart failure in patients with type 2 diabetes: the ADVANCE trial

PubMed Central

Ohkuma, Toshiaki; Jun, Min; Woodward, Mark; Zoungas, Sophia; Cooper, Mark E.; Grobbee, Diederick E.; Hamet, Pavel; Mancia, Giuseppe; Williams, Bryan; Welsh, Paul; Sattar, Naveed; Shaw, Jonathan E.; Rahimi, Kazem; Chalmers, John

2018-01-01

Objective This study examined the individual and combined impact of N-terminal pro-B-type natriuretic peptide (NT-proBNP), high-sensitivity cardiac troponin T (hs-cTnT), interleukin-6 (IL-6), and high-sensitivity C-reactive protein (hs-CRP) on the prediction of heart failure incidence or progression in patients with type 2 diabetes. Research Design and Methods A nested case-cohort study was conducted in 3,098 participants with type 2 diabetes in the Action in Diabetes and Vascular Disease: Preterax and Diamicron Modified Release Controlled Evaluation (ADVANCE) trial. Results A higher value of each biomarker was significantly associated with a higher risk of heart failure incidence or progression, after adjustment for major risk factors. The hazard ratios per 1-SD increase were 3.06 (95% CI 2.37-3.96) for NT-proBNP, 1.50 (1.27-1.77) for hs-cTnT, 1.48 (1.27-1.72) for IL-6, and 1.32 (1.12-1.55) for hs-CRP. Addition of NT-proBNP to the model including conventional risk factors meaningfully improved 5-year risk predictive performance (c-statistic 0.8162 to 0.8800; continuous net reclassification improvement [NRI] 73.1%; categorical NRI [<5%, 5-10%, >10% 5-year risk] 24.2%). In contrast, addition of hs-cTnT, IL-6 or hs-CRP did not improve the prediction metrics consistently either in combination or when added to NT-proBNP. Conclusions Only NT-proBNP, strongly and consistently improved prediction of heart failure in patients with type 2 diabetes beyond a wide range of clinical risk factors and biomarkers. PMID:28684396

Cardiac Stress and Inflammatory Markers as Predictors of Heart Failure in Patients With Type 2 Diabetes: The ADVANCE Trial.

PubMed

Ohkuma, Toshiaki; Jun, Min; Woodward, Mark; Zoungas, Sophia; Cooper, Mark E; Grobbee, Diederick E; Hamet, Pavel; Mancia, Giuseppe; Williams, Bryan; Welsh, Paul; Sattar, Naveed; Shaw, Jonathan E; Rahimi, Kazem; Chalmers, John

2017-09-01

This study examined the individual and combined effect of N-terminal pro-B-type natriuretic peptide (NT-proBNP), high-sensitivity cardiac troponin T (hs-cTnT), interleukin-6 (IL-6), and hs-CRP on the prediction of heart failure incidence or progression in patients with type 2 diabetes. A nested case-cohort study was conducted in 3,098 participants with type 2 diabetes in the Action in Diabetes and Vascular Disease: Preterax and Diamicron Modified Release Controlled Evaluation (ADVANCE) trial. A higher value of each biomarker was significantly associated with a higher risk of heart failure incidence or progression, after adjustment for major risk factors. The hazard ratios per 1-SD increase were 3.06 (95% CI 2.37, 3.96) for NT-proBNP, 1.50 (1.27, 1.77) for hs-cTnT, 1.48 (1.27, 1.72) for IL-6, and 1.32 (1.12, 1.55) for hs-CRP. The addition of NT-proBNP to the model including conventional risk factors meaningfully improved 5-year risk-predictive performance (C statistic 0.8162 to 0.8800; continuous net reclassification improvement [NRI] 73.1%; categorical NRI [<5%, 5-10%, >10% 5-year risk] 24.2%). In contrast, the addition of hs-cTnT, IL-6, or hs-CRP did not improve the prediction metrics consistently in combination or when added to NT-proBNP. Only NT-proBNP strongly and consistently improved the prediction of heart failure in patients with type 2 diabetes beyond a wide range of clinical risk factors and biomarkers. © 2017 by the American Diabetes Association.

MicroRNA-9 regulates the expression of peroxisome proliferator-activated receptor δ in human monocytes during the inflammatory response

PubMed Central

THULIN, PETRA; WEI, TIANLING; WERNGREN, OLIVERA; CHEUNG, LOUISA; FISHER, RACHEL M.; GRANDÉR, DAN; CORCORAN, MARTIN; EHRENBORG, EWA

2013-01-01

PPARδ is involved in the inflammatory response and its expression is induced by cytokines, however, limited knowledge has been produced regarding its regulation. Since recent findings have shown that microRNAs, which are small non-coding RNAs that regulate gene expression, are involved in the immune response, we set out to investigate whether PPARδ can be regulated by microRNAs expressed in monocytes. Bioinformatic analysis identified a putative miR-9 target site within the 3′-UTR of PPARδ that was subsequently verified to be functional using reporter constructs. Primary human monocytes stimulated with LPS showed a downregulation of PPARδ and its target genes after 4 h while the expression of miR-9 was induced. Analysis of pro-inflammatory (M1) and anti-inflammatory (M2) macrophages showed that human PPARδ mRNA as well as miR-9 expression was higher in M1 compared to M2 macrophages. Furthermore, treatment with the PPARδ agonist, GW501516, induced the expression of PPARδ target genes in the pro-inflammatory M1 macrophages while no change was observed in the anti-inflammatory M2 macrophages. Taken together, these data suggest that PPARδ is regulated by miR-9 in monocytes and that activation of PPARδ may be of importance in M1 pro-inflammatory but not in M2 anti-inflammatory macrophages in humans. PMID:23525285

Secretory leukocyte protease inhibitor gene deletion alters bleomycin-induced lung injury, but not development of pulmonary fibrosis.

PubMed

Habgood, Anthony N; Tatler, Amanda L; Porte, Joanne; Wahl, Sharon M; Laurent, Geoffrey J; John, Alison E; Johnson, Simon R; Jenkins, Gisli

2016-06-01

Idiopathic pulmonary fibrosis is a progressive, fatal disease with limited treatment options. Protease-mediated transforming growth factor-β (TGF-β) activation has been proposed as a pathogenic mechanism of lung fibrosis. Protease activity in the lung is tightly regulated by protease inhibitors, particularly secretory leukocyte protease inhibitor (SLPI). The bleomycin model of lung fibrosis was used to determine the effect of increased protease activity in the lungs of Slpi(-/-) mice following injury. Slpi(-/-), and wild-type, mice received oropharyngeal administration of bleomycin (30 IU) and the development of pulmonary fibrosis was assessed. Pro and active forms of matrix metalloproteinase (MMP)-2 and MMP-9 were measured. Lung fibrosis was determined by collagen subtype-specific gene expression, hydroxyproline concentration, and histological assessment. Alveolar TGF-β activation was measured using bronchoalveolar lavage cell pSmad2 levels and global TGF-β activity was assessed by pSmad2 immunohistochemistry. The active-MMP-9 to pro-MMP-9 ratio was significantly increased in Slpi(-/-) animals compared with wild-type animals, demonstrating enhanced metalloproteinase activity. Wild-type animals showed an increase in TGF-β activation following bleomycin, with a progressive and sustained increase in collagen type I, alpha 1 (Col1α1), III, alpha 1(Col3α1), IV, alpha 1(Col4α1) mRNA expression, and a significant increase in total lung collagen 28 days post bleomycin. In contrast Slpi(-/-) mice showed no significant increase of alveolar TGF-β activity following bleomycin, above their already elevated levels, although global TGF-β activity did increase. Slpi(-/-) mice had impaired collagen gene expression but animals demonstrated minimal reduction in lung fibrosis compared with wild-type animals. These data suggest that enhanced proteolysis does not further enhance TGF-β activation, and inhibits sustained Col1α1, Col3α1, and Col4α1 gene expression following lung injury. However, these changes do not prevent the development of lung fibrosis. Overall, these data suggest that the absence of Slpi does not markedly modify the development of lung fibrosis following bleomycin-induced lung injury.

The Genome and Development-Dependent Transcriptomes of Pyronema confluens: A Window into Fungal Evolution

PubMed Central

Traeger, Stefanie; Altegoer, Florian; Freitag, Michael; Gabaldon, Toni; Kempken, Frank; Kumar, Abhishek; Marcet-Houben, Marina; Pöggeler, Stefanie; Stajich, Jason E.; Nowrousian, Minou

2013-01-01

Fungi are a large group of eukaryotes found in nearly all ecosystems. More than 250 fungal genomes have already been sequenced, greatly improving our understanding of fungal evolution, physiology, and development. However, for the Pezizomycetes, an early-diverging lineage of filamentous ascomycetes, there is so far only one genome available, namely that of the black truffle, Tuber melanosporum, a mycorrhizal species with unusual subterranean fruiting bodies. To help close the sequence gap among basal filamentous ascomycetes, and to allow conclusions about the evolution of fungal development, we sequenced the genome and assayed transcriptomes during development of Pyronema confluens, a saprobic Pezizomycete with a typical apothecium as fruiting body. With a size of 50 Mb and ∼13,400 protein-coding genes, the genome is more characteristic of higher filamentous ascomycetes than the large, repeat-rich truffle genome; however, some typical features are different in the P. confluens lineage, e.g. the genomic environment of the mating type genes that is conserved in higher filamentous ascomycetes, but only partly conserved in P. confluens. On the other hand, P. confluens has a full complement of fungal photoreceptors, and expression studies indicate that light perception might be similar to distantly related ascomycetes and, thus, represent a basic feature of filamentous ascomycetes. Analysis of spliced RNA-seq sequence reads allowed the detection of natural antisense transcripts for 281 genes. The P. confluens genome contains an unusually high number of predicted orphan genes, many of which are upregulated during sexual development, consistent with the idea of rapid evolution of sex-associated genes. Comparative transcriptomics identified the transcription factor gene pro44 that is upregulated during development in P. confluens and the Sordariomycete Sordaria macrospora. The P. confluens pro44 gene (PCON_06721) was used to complement the S. macrospora pro44 deletion mutant, showing functional conservation of this developmental regulator. PMID:24068976

Effect of polymorphisms in candidate genes on carcass and meat quality traits in double muscled Piemontese cattle.

PubMed

Ribeca, C; Bonfatti, V; Cecchinato, A; Albera, A; Gallo, L; Carnier, P

2014-03-01

The aim of this study was to investigate the association between 10 candidate genes and carcass weight and conformation, carcass daily gain, and meat quality (pH, color, cooking loss, drip loss and shear force) in 990 double-muscled Piemontese young bulls. Animals were genotyped at each of the following genes: growth hormone, growth hormone receptor, pro-opiomelanocortin, pro-opiomelanocortin class 1 homeobox 1, melanocortin-4 receptor, corticotrophin-releasing hormone, diacylglycerol O-acyltransferase-1, thyroglobulin, carboxypeptidase E and gamma-3 regulatory subunit of the AMP-activated protein kinase. All the investigated SNPs had additive effects which were relevant for at least one of the traits. Relevant associations between the investigated SNPs and carcass weight, carcass daily gain and carcass conformation were detected, whereas associations of SNPs with meat quality were moderate. Results confirmed some of previously reported associations, but diverged for others. Validation in other cattle breeds is required to use these SNPs in gene-assisted selection programs for enhancement of carcass traits and meat quality. Copyright © 2013 Elsevier Ltd. All rights reserved.

New associations: INFG and TGFB1 genes and the inhibitor development in severe haemophilia A.

PubMed

de Alencar, J B; Macedo, L C; de Barros, M F; Rodrigues, C; Shinzato, A H; Pelissari, C B; Machado, J; Sell, A M; Visentainer, J E L

2015-07-01

The development of factor VIII (FVIII) inhibitor is the main complication of replacement therapy in patients with haemophilia A (HA). A ratio of 5-7% of individuals HA develops antibodies (inhibitors) against the FVIII infused during the treatment, thereby reducing their pro-coagulant activity. The immunomodulatory cytokine genes have been related to the risk of development of alloantibodies in several studies, mainly in HA with severe form. We investigated the polymorphisms in regulatory regions of cytokine genes (IL1A, IL1B, IL1R, IL1RA, IL4RA, IL12, INFG, TGFB1, TNF, IL2, IL4, IL6, IL10) that could influence the risk of developing inhibitors in patients with severe HA. The genotyping of cytokine genes of 117 patients with HA was performed by polymerase chain reaction with sequence-specific primers (PCR-SSP) using the protocol recommended by the manufacturer (Invitrogen kit Cytokines(®) , Canoga Park, USA) RESULTS: From the cohort of 117 patients with severe HA, 35 developed inhibitors. There was a higher frequency of +874 T allele in INFG and of +869 TT and TG/TG in TGFB1 genes on patients with inhibitors. This suggests that polymorphisms in INFG and in TGFB1 genes are related to risk of developing inhibitor, and could contribute to a genetic profile of the individual HA for the risk of inhibitors development to FVIII. © 2015 John Wiley & Sons Ltd.

Niclosamide, an anti-helminthic molecule, downregulates the retroviral oncoprotein Tax and pro-survival Bcl-2 proteins in HTLV-1-transformed T lymphocytes

PubMed Central

Chen, Li; Liu, Xin; Belani, Chandra; Cheng, Hua

2015-01-01

Adult T cell leukemia and lymphoma (ATL) is a highly aggressive form of hematological malignancy and is caused by chronic infection of human T cell leukemia virus type 1 (HTLV-1). The viral genome encodes an oncogenic protein, Tax, which plays a key role in transactivating viral gene transcription and in deregulating cellular oncogenic signaling to promote survival, proliferation and transformation of virally infected T cells. Hence, Tax is a desirable therapeutic target, particularly at early stage of HTLV-1-mediated oncogenesis. We here show that niclosamide, an anti-helminthic molecule, induced apoptosis of HTLV-1-transformed T cells. Niclosamide facilitated degradation of the Tax protein in proteasome. Consistent with niclosamide-mediated Tax degradation, this compound inhibited activities of MAPK/ERK1/2 and IκB kinases. In addition, niclosamide downregulated Stat3 and pro-survival Bcl-2 family members such as Mcl-1 and repressed the viral gene transcription of HTLV-1 through induction of Tax degradation. Since Tax, Stat3 and Mcl-1 are crucial molecules for promoting survival and growth of HTLV-1-transformed T cells, our findings demonstrate a novel mechanism of niclosamide in inducing Tax degradation and downregulating various cellular pro-survival molecules, thereby promoting apoptosis of HTLV-1-associated leukemia cells. PMID:26116531

A comparative genome analysis of PME and PMEI families reveals the evolution of pectin metabolism in plant cell walls.

PubMed

Wang, Maojun; Yuan, Daojun; Gao, Wenhui; Li, Yang; Tan, Jiafu; Zhang, Xianlong

2013-01-01

Pectins are fundamental polysaccharides in the plant primary cell wall. Pectins are synthesized and secreted to cell walls as highly methyl-esterified polymers and then demethyl-esterified by pectin methylesterases (PMEs), which are spatially regulated by pectin methylesterase inhibitors (PMEIs). Although PME and PMEI genes are pivotal in plant cell wall formation, few studies have focused on the evolutionary patterns of the PME and PMEI gene families. In this study, the gene origin, evolution, and expression diversity of these two families were systematically analyzed using 11 representative species, including algae, bryophytes, lycophytes and flowering land plants. The results show that 1) for the two subfamilies (PME and proPME) of PME, the origin of the PME subfamily is consistent with the appearance of pectins in early charophyte cell walls, 2) Whole genome duplication (WGD) and tandem duplication contribute to the expansion of proPME and PMEI families in land plants, 3) Evidence of selection pressure shows that the proPME and PMEI families have rapidly evolved, particularly the PMEI family in vascular plants, and 4) Comparative expression profile analysis of the two families indicates that the eudicot Arabidopsis and monocot rice have different expression patterns. In addition, the gene structure and sequence analyses show that the origin of the PMEI domain may be derived from the neofunctionalization of the pro domain after WGD. This study will advance the evolutionary understanding of the PME and PMEI families and plant cell wall development.

Niclosamide, an anti-helminthic molecule, downregulates the retroviral oncoprotein Tax and pro-survival Bcl-2 proteins in HTLV-1-transformed T lymphocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xiang, Di; Yuan, Yunsheng; Engineering Research Center of Cell and Therapeutic Antibody, Ministry of Education, School of Pharmacy, Shanghai Jiao Tong University, Shanghai

Adult T cell leukemia and lymphoma (ATL) is a highly aggressive form of hematological malignancy and is caused by chronic infection of human T cell leukemia virus type 1 (HTLV-1). The viral genome encodes an oncogenic protein, Tax, which plays a key role in transactivating viral gene transcription and in deregulating cellular oncogenic signaling to promote survival, proliferation and transformation of virally infected T cells. Hence, Tax is a desirable therapeutic target, particularly at early stage of HTLV-1-mediated oncogenesis. We here show that niclosamide, an anti-helminthic molecule, induced apoptosis of HTLV-1-transformed T cells. Niclosamide facilitated degradation of the Tax proteinmore » in proteasome. Consistent with niclosamide-mediated Tax degradation, this compound inhibited activities of MAPK/ERK1/2 and IκB kinases. In addition, niclosamide downregulated Stat3 and pro-survival Bcl-2 family members such as Mcl-1 and repressed the viral gene transcription of HTLV-1 through induction of Tax degradation. Since Tax, Stat3 and Mcl-1 are crucial molecules for promoting survival and growth of HTLV-1-transformed T cells, our findings demonstrate a novel mechanism of niclosamide in inducing Tax degradation and downregulating various cellular pro-survival molecules, thereby promoting apoptosis of HTLV-1-associated leukemia cells. - Highlights: • Niclosamide is a promising therapeutic candidate for adult T cell leukemia. • Niclosamide employs a novel mechanism through proteasomal degradation of Tax. • Niclosamide downregulates certain cellular pro-survival molecules.« less

Exploring the effect and mechanism of Hibiscus sabdariffa on urinary tract infection and experimental renal inflammation.

PubMed

Chou, Shun-Ting; Lo, Hsin-Yi; Li, Chia-Cheng; Cheng, Lu-Chen; Chou, Pei-Chi; Lee, Yu-Chen; Ho, Tin-Yun; Hsiang, Chien-Yun

2016-12-24

Hibiscus sabdariffa Linn., also known as roselle, is used in folk medicine as an anti-inflammatory agent. Urinary tract infection (UTI) is a common problem in long-term care facilities. However, effects of roselle on UTI and renal inflammation remained to be analyzed. Here we surveyed the effect of roselle drink on the prevention of UTI in long-term care facilities and analyzed the anti-inflammatory potential of roselle on lipopolysaccharide (LPS)-induced renal inflammation in mice. Survey questionnaires and clinical observation were applied to evaluate the use of roselle and the incidence of UTI in long-term care facilities. Mice were administrated roselle orally for 7 consecutive days and then challenged with LPS. Anti-renal inflammatory effects of roselle were analyzed by microarray and immunohistochemical staining. Clinical observation showed that taking roselle drink in residents with urinary catheters reduced the incidence of UTI in long-term care facilities. Renal inflammation is a key event of UTI. Roselle suppressed LPS-induced nuclear factor-κB (NF-κB) activation in cells and LPS-induced interleukin-1β production in mice a dose-dependent manner. Immunohistochemical staining showed that roselle inhibited LPS-induced NF-κB activation and inflammatory cell infiltration in kidney. Gene expression profiling further showed that roselle suppressed the expression of pro-inflammatory cytokine genes and enzyme genes involved in the production of prostaglandin and nitric oxide. In addition, NF-κB was the main transcription factor involved in the regulation of roselle-regulated gene expression in kidney. This is the first report applying clinical observation-guided transcriptomic study to explore the application and mechanism of roselle on UTI. Our findings suggested that roselle drink ameliorated LPS-induced renal inflammation via downregulation of cytokine network, pro-inflammatory product production, and NF-κB pathway. Moreover, this report suggested the potential benefit of roselle drink on UTI. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Flavocoxid, a dual inhibitor of cyclooxygenase and 5-lipoxygenase, blunts pro-inflammatory phenotype activation in endotoxin-stimulated macrophages

PubMed Central

Altavilla, D; Squadrito, F; Bitto, A; Polito, F; Burnett, BP; Di Stefano, V; Minutoli, L

2009-01-01

Background and purpose: The flavonoids, baicalin and catechin, from Scutellaria baicalensis and Acacia catechu, respectively, have been used for various clinical applications. Flavocoxid is a mixed extract containing baicalin and catechin, and acts as a dual inhibitor of cyclooxygenase (COX) and 5-lipoxygenase (LOX) enzymes. The anti-inflammatory activity, measured by protein and gene expression of inflammatory markers, of flavocoxid in rat peritoneal macrophages stimulated with Salmonella enteritidis lipopolysaccharide (LPS) was investigated. Experimental approach: LPS-stimulated (1 µg·mL−1) peritoneal rat macrophages were co-incubated with different concentrations of flavocoxid (32–128 µg·mL−1) or RPMI medium for different incubation times. Inducible COX-2, 5-LOX, inducible nitric oxide synthase (iNOS) and inhibitory protein κB-α (IκB-α) levels were evaluated by Western blot analysis. Nuclear factor κB (NF-κB) binding activity was investigated by electrophoretic mobility shift assay. Tumour necrosis factor-α (TNF-α) gene and protein expression were measured by real-time polymerase chain reaction and enzyme-linked immunosorbent assay respectively. Finally, malondialdehyde (MDA) and nitrite levels in macrophage supernatants were evaluated. Key results: LPS stimulation induced a pro-inflammatory phenotype in rat peritoneal macrophages. Flavocoxid (128 µg·mL−1) significantly inhibited COX-2 (LPS = 18 ± 2.1; flavocoxid = 3.8 ± 0.9 integrated intensity), 5-LOX (LPS = 20 ± 3.8; flavocoxid = 3.1 ± 0.8 integrated intensity) and iNOS expression (LPS = 15 ± 1.1; flavocoxid = 4.1 ± 0.4 integrated intensity), but did not modify COX-1 expression. PGE2 and LTB4 levels in culture supernatants were consequently decreased. Flavocoxid also prevented the loss of IκB-α protein (LPS = 1.9 ± 0.2; flavocoxid = 7.2 ± 1.6 integrated intensity), blunted increased NF-κB binding activity (LPS = 9.2 ± 2; flavocoxid = 2.4 ± 0.7 integrated intensity) and the enhanced TNF-α mRNA levels (LPS = 8 ± 0.9; flavocoxid = 1.9 ± 0.8 n-fold/β-actin) induced by LPS. Finally, flavocoxid decreased MDA, TNF and nitrite levels from LPS-stimulated macrophages. Conclusion and implications: Flavocoxid might be useful as a potential anti-inflammatory agent, acting at the level of gene and protein expression. PMID:19681869

ProP Is Required for the Survival of Desiccated Salmonella enterica Serovar Typhimurium Cells on a Stainless Steel Surface

PubMed Central

Finn, Sarah; Händler, Kristian; Condell, Orla; Colgan, Aoife; Cooney, Shane; McClure, Peter; Amézquita, Aléjandro; Hinton, Jay C. D.

2013-01-01

Consumers trust commercial food production to be safe, and it is important to strive to improve food safety at every level. Several outbreaks of food-borne disease have been caused by Salmonella strains associated with dried food. Currently we do not know the mechanisms used by Salmonella enterica serovar Typhimurium to survive in desiccated environments. The aim of this study was to discover the responses of S. Typhimurium ST4/74 at the transcriptional level to desiccation on a stainless steel surface and to subsequent rehydration. Bacterial cells were dried onto the same steel surfaces used during the production of dry foods, and RNA was recovered for transcriptomic analysis. Subsequently, dried cells were rehydrated and were again used for transcriptomic analysis. A total of 266 genes were differentially expressed under desiccation stress compared with a static broth culture. The osmoprotectant transporters proP, proU, and osmU (STM1491 to STM1494) were highly upregulated by drying. Deletion of any one of these transport systems resulted in a reduction in the long-term viability of S. Typhimurium on a stainless steel food contact surface. The proP gene was critical for survival; proP deletion mutants could not survive desiccation for long periods and were undetectable after 4 weeks. Following rehydration, 138 genes were differentially expressed, with upregulation observed for genes such as proP, proU, and the phosphate transport genes (pstACS). In time, this knowledge should prove valuable for understanding the underlying mechanisms involved in pathogen survival and should lead to improved methods for control to ensure the safety of intermediate- and low-moisture foods. PMID:23666329

The Frustrated Host Response to Legionella pneumophila Is Bypassed by MyD88-Dependent Translation of Pro-inflammatory Cytokines

PubMed Central

Asrat, Seblewongel; Dugan, Aisling S.; Isberg, Ralph R.

2014-01-01

Many pathogens, particularly those that require their host for survival, have devised mechanisms to subvert the host immune response in order to survive and replicate intracellularly. Legionella pneumophila, the causative agent of Legionnaires' disease, promotes intracellular growth by translocating proteins into its host cytosol through its type IV protein secretion machinery. At least 5 of the bacterial translocated effectors interfere with the function of host cell elongation factors, blocking translation and causing the induction of a unique host cell transcriptional profile. In addition, L. pneumophila also interferes with translation initiation, by preventing cap-dependent translation in host cells. We demonstrate here that protein translation inhibition by L. pneumophila leads to a frustrated host MAP kinase response, where genes involved in the pathway are transcribed but fail to be translated due to the bacterium-induced protein synthesis inhibition. Surprisingly, few pro-inflammatory cytokines, such as IL-1α and IL-1β, bypass this inhibition and get synthesized in the presence of Legionella effectors. We show that the selective synthesis of these genes requires MyD88 signaling and takes place in both infected cells that harbor bacteria and neighboring bystander cells. Our findings offer a perspective of how host cells are able to cope with pathogen-encoded activities that disrupt normal cellular process and initiate a successful inflammatory response. PMID:25058342

EFFECT OF HYPOXIA ON THE EXPRESSION OF GENES THAT ENCODE SOME IGFBP AND CCN PROTEINS IN U87 GLIOMA CELLS DEPENDS ON IRE1 SIGNALING.

PubMed

Minchenko, O H; Kharkova, A P; Minchenko, D O; Karbovskyi, L L

2015-01-01

We have studied hypoxic regulation of the expression of different insulin-like growth factor binding protein genes in U87 glioma cells in relation to inhibition of IRE1 (inositol requiring enzyme-1), a central mediator of endoplasmic reticulum stress, which controls cell proliferation and tumor growth. We have demonstrated that hypoxia leads to up-regulation of the expression of IGFBP6, IGFBP7, IGFBP10/CYR61, WISP1, and WISP2 genes and down-regulation--of IGFBP9/NOV gene at the mRNA level in control glioma cells, being more signifcant changes for IGFBP10/CYR61 and WISP2 genes. At the same time, inhibition of IRE1 modifies the effect of hypoxia on the expression of all studied genes: eliminates sensitivity to hypoxia the expression of IGFBP7 and IGFBP9/NOV genes, suppresses effect of hypoxia on IGFBP6, IGFBP10/CYR61, and WISP2 genes, and slightly enhances hypoxic regulation of WISP1 gene expression in glioma cells. We have also demonstrated that the expression of all studied genes in glioma cells is regulated by IRE1 signaling enzyme upon normoxic condition, because inhibition of IRE1 significantly up-regulates IGFBP7, IGFBP10/CYR61, WISP1, and WISP2 genes and down-regulates IGFBP6 and IGFBP9/NOV genes as compared to control glioma cells. The present study demonstrates that hypoxia, which contributes to tumor growth, affects all studied IGFBP and WISP gene expressions and that inhibition of IRE1 preferentially abolishes or suppresses the hypoxic regulation of these gene expressions and thus possibly contributes to slower glioma growth. Moreover, inhibition of IRE1, which correlates with suppression of cell proliferation and glioma growth, is down-regulated expression of pro-proliferative IGFBP genes, attesting to the fact that endoplasmic reticulum stress is a necessary component of malignant tumor growth.

Hypoxia-inducible factor-1 (HIF-1) promotes its degradation by induction of HIF-α-prolyl-4-hydroxylases

PubMed Central

2004-01-01

An important regulator involved in oxygen-dependent gene expression is the transcription factor HIF (hypoxia-inducible factor), which is composed of an oxygen-sensitive α-subunit (HIF-1α or HIF-2α) and a constitutively expressed β-subunit. In normoxia, HIF-1α is destabilized by post-translational hydroxylation of Pro-564 and Pro-402 by a family of oxygen-sensitive dioxygenases. The three HIF-modifying human enzymes have been termed prolyl hydroxylase domain containing proteins (PHD1, PHD2 and PHD3). Prolyl hydroxylation leads to pVHL (von-Hippel-Lindau protein)-dependent ubiquitination and rapid proteasomal degradation of HIF-1α. In the present study, we report that human PHD2 and PHD3 are induced by hypoxia in primary and transformed cell lines. In the human osteosarcoma cell line, U2OS, selective suppression of HIF-1α expression by RNA interference resulted in a complete loss of hypoxic induction of PHD2 and PHD3. Induction of PHD2 by hypoxia was lost in pVHL-deficient RCC4 cells. These results suggest that hypoxic induction of PHD2 and PHD3 is critically dependent on HIF-α. Using a VHL capture assay, we demonstrate that HIF-α prolyl-4-hydroxylase capacity of cytoplasmic and nuclear protein extracts was enhanced by prolonged exposure to hypoxia. Degradation of HIF-1α after reoxygenation was accelerated, which demonstrates functional relevance of the present results. We propose a direct, negative regulatory mechanism, which limits accumulation of HIF-1α in hypoxia and leads to accelerated degradation on reoxygenation after long-term hypoxia. PMID:15104534

MAPK signaling promotes axonal degeneration by speeding the turnover of the axonal maintenance factor NMNAT2

PubMed Central

Walker, Lauren J; Summers, Daniel W; Sasaki, Yo; Brace, EJ; Milbrandt, Jeffrey; DiAntonio, Aaron

2017-01-01

Injury-induced (Wallerian) axonal degeneration is regulated via the opposing actions of pro-degenerative factors such as SARM1 and a MAPK signal and pro-survival factors, the most important of which is the NAD+ biosynthetic enzyme NMNAT2 that inhibits activation of the SARM1 pathway. Here we investigate the mechanism by which MAPK signaling facilitates axonal degeneration. We show that MAPK signaling promotes the turnover of the axonal survival factor NMNAT2 in cultured mammalian neurons as well as the Drosophila ortholog dNMNAT in motoneurons. The increased levels of NMNAT2 are required for the axonal protection caused by loss of MAPK signaling. Regulation of NMNAT2 by MAPK signaling does not require SARM1, and so cannot be downstream of SARM1. Hence, pro-degenerative MAPK signaling functions upstream of SARM1 by limiting the levels of the essential axonal survival factor NMNAT2 to promote injury-dependent SARM1 activation. These findings are consistent with a linear molecular pathway for the axonal degeneration program. DOI: http://dx.doi.org/10.7554/eLife.22540.001 PMID:28095293

Role of the epidermal growth factor receptor in signaling strain-dependent activation of the brain natriuretic peptide gene.

PubMed

Anderson, Hope D I; Wang, Feng; Gardner, David G

2004-03-05

The epidermal growth factor receptor (EGFR) and ectoshedding of heparin-binding epidermal growth factor (HBEGF), an EGFR ligand, have been linked to the development of cardiac myocyte hypertrophy. However, the precise role that the liganded EGFR plays in the transcriptional activation of the gene program that accompanies hypertrophy remains undefined. Utilizing the human (h) BNP gene as a model of hypertrophy-dependent gene activation, we show that activation of the EGFR plays an important role in mediating mechanical strain-dependent stimulation of the hBNP promoter. Strain promotes endothelin (ET) generation through NAD(P)H oxidase-dependent production of reactive oxygen species. ET in turn induces metalloproteinase-mediated cleavage of pro-HBEGF and ectoshedding of HBEGF, which activates the EGFR and stimulates hBNP promoter activity. HBEGF also stimulates other phenotypic markers of hypertrophy including protein synthesis and sarcomeric assembly. The antioxidant N-acetylcysteine or the NAD(P)H oxidase inhibitor, apocynin, inhibited strain-dependent activation of the ET-1 promoter, HBEGF shedding, and hBNP promoter activation. The metalloproteinase inhibitor, GM-6001, prevented the induction of HBEGF ectoshedding and the hBNP promoter response to strain, suggesting a critical role for the metalloproteinase-dependent cleavage event in signaling the strain response. These findings suggest that metalloproteinase activity as an essential step in this pathway may prove to be a relevant therapeutic target in the management of cardiac hypertrophy.

Curcuma longa polyphenols improve insulin-mediated lipid accumulation and attenuate proinflammatory response of 3T3-L1 adipose cells during oxidative stress through regulation of key adipokines and antioxidant enzymes.

PubMed

Septembre-Malaterre, Axelle; Le Sage, Fanny; Hatia, Sarah; Catan, Aurélie; Janci, Laurent; Gonthier, Marie-Paule

2016-07-08

Plant polyphenols may exert beneficial action against obesity-related oxidative stress and inflammation which promote insulin resistance. This study evaluated the effect of polyphenols extracted from French Curcuma longa on 3T3-L1 adipose cells exposed to H2 O2 -mediated oxidative stress. We found that Curcuma longa extract exhibited high amounts of curcuminoids identified as curcumin, demethoxycurcumin, and bisdemethoxycurcumin, which exerted free radical-scavenging activities. Curcuma longa polyphenols improved insulin-mediated lipid accumulation and upregulated peroxisome proliferator-activated receptor-gamma gene expression and adiponectin secretion which decreased in H2 O2 -treated cells. Curcuminoids attenuated H2 O2 -enhanced production of pro-inflammatory molecules such as interleukin-6, tumor necrosis factor-alpha, monocyte chemoattractant protein-1, and nuclear factor κappa B. Moreover, they reduced intracellular levels of reactive oxygen species elevated by H2 O2 and modulated the expression of genes encoding superoxide dismutase and catalase antioxidant enzymes. Collectively, these findings highlight that Curcuma longa polyphenols protect adipose cells against oxidative stress and may improve obesity-related metabolic disorders. © 2016 BioFactors, 42(4):418-430, 2016. © 2016 International Union of Biochemistry and Molecular Biology.

Immunohistochemical profile of cytokines and growth factors expressed in vestibular schwannoma and in normal vestibular nerve tissue.

PubMed

Taurone, Samanta; Bianchi, Enrica; Attanasio, Giuseppe; Di Gioia, Cira; Ierinó, Rocco; Carubbi, Cecilia; Galli, Daniela; Pastore, Francesco Saverio; Giangaspero, Felice; Filipo, Roberto; Zanza, Christian; Artico, Marco

2015-07-01

Vestibular schwannomas, also known as acoustic neuromas, are benign tumors, which originate from myelin-forming Schwann cells. They develop in the vestibular branch of the eighth cranial nerve in the internal auditory canal or cerebellopontine angle. The clinical progression of the condition involves slow and progressive growth, eventually resulting in brainstem compression. The objective of the present study was to investigate the expression level and the localization of the pro-inflammatory cytokines, transforming growth factor-β1 (TGF-β1) interleukin (IL)-1β, IL-6 and tumor necrosis factor-α (TNF-α), as well as the adhesion molecules, intracellular adhesion molecule-1 and vascular endothelial growth factor (VEGF), in order to determine whether these factors are involved in the transformation and development of human vestibular schwannoma. The present study investigated whether changes in inflammation are involved in tumor growth and if so, the mechanisms underlying this process. The results of the current study demonstrated that pro-inflammatory cytokines, including TGF-β1, IL-1β and IL-6 exhibited increased expression in human vestibular schwannoma tissue compared with normal vestibular nerve samples. TNF-α was weakly expressed in Schwann cells, confirming that a lower level of this cytokine is involved in the proliferation of Schwann cells. Neoplastic Schwann cells produce pro-inflammatory cytokines that may act in an autocrine manner, stimulating cellular proliferation. In addition, the increased expression of VEGF in vestibular schwannoma compared with that in normal vestibular nerve tissue, suggests that this factor may induce neoplastic growth via the promotion of angiogenesis. The present findings suggest that inflammation may promote angiogenesis and consequently contribute to tumor progression. In conclusion, the results of the present study indicated that VEGF and pro-inflammatory cytokines may be potential therapeutic targets in vestibular schwannoma. Further studies are necessary to confirm the involvement of these factors in the growth of neoplasms and to develop inhibitors of pro-inflammatory cytokines as a potential treatment option in the future.

Study on the antiulcer effects of Veronicastrum axillare on gastric ulcer in rats induced by ethanol based on tumor necrosis factor-α (TNF-α) and endothelin-1 (ET-1).

PubMed

Du, Yong; Zhao, Weichun; Lu, Leilei; Zheng, Jiayan; Hu, Xishi; Yu, Zhehan; Zhu, Lixin

2013-12-01

To assess whether Veronicastrum axillare (V. axillare) can ameliorate ethanol-induced gastric mucosal lesions in rats, reduce the production of pro-inflammatory cytokines, suppress apoptosis and improve local microcirculation disturbances. Totally 48 male Sprague-Dawley rats were randomly divided into six groups, eight rats in each group. Rats in the normal group and the model group were administered with 0.9% normal saline respectively. Rats in the positive group and ranitidine group were administered with 0.18% ranitidine suspension by intragastric administration respectively. Those in the high dose V. axillare group, the medium dose V. axillare group and the low dose V. axillare group were administrated with V. axillare at the daily dose of 2.8 g/kg, 1.4 g/kg and 0.7 g/kg by intragastric administration. Gastric mucosal lesions were produced by intragastric administration of absolute ethanol. Water extract of V. axillare was successively injected for 14 d and last day was injected 1 h before ethanol administration. Gastric mucosal ulcer index and ulcer inhibitory rate were counted by improved Guth methods. The tissue sections were made for pathological histology analysis. Also, we measured the concentrations of tumor necrosis factor-α (TNF-α) and endothelin-1 (ET-1) in gastric mucosal, as an index of the pro-inflammatory cytokines, apoptosis and local microcirculation. Besides, the mRNA contents of TNF-α and ET-1 were measured to verify effects on gene expression by real-time fluorescent quantitative PCR. Water extract of V. axillare significantly ameliorated the gastric mucosal lesions induced by ethanol administration (P<0.01). Pro-inflammatory cytokines, TNF-α and ET-1 were increased after ethanol administration and significantly reduced by water extract of V. axillare. The expressions of TNF-α and ET-1 mRNA were also be inhibited by water extract of V. axillare. Current evidences show water extract of V. axillare is effective for defending against ethanol-induced gastric mucosal lesions, significantly inhibiting the production of pro-inflammatory cytokines and the expressions of TNF-α and ET-1 mRNA, which may be useful for inhibiting apoptosis and improving local microcirculation. Copyright © 2013 Asian Pacific Tropical Biomedical Magazine. Published by Elsevier B.V. All rights reserved.

Brain-derived neurotrophic factor and its clinical implications

PubMed Central

Bathina, Siresha

2015-01-01

Brain-derived neurotrophic factor (BDNF) plays an important role in neuronal survival and growth, serves as a neurotransmitter modulator, and participates in neuronal plasticity, which is essential for learning and memory. It is widely expressed in the CNS, gut and other tissues. BDNF binds to its high affinity receptor TrkB (tyrosine kinase B) and activates signal transduction cascades (IRS1/2, PI3K, Akt), crucial for CREB and CBP production, that encode proteins involved in β cell survival. BDNF and insulin-like growth factor-1 have similar downstream signaling mechanisms incorporating both p-CAMK and MAPK that increase the expression of pro-survival genes. Brain-derived neurotrophic factor regulates glucose and energy metabolism and prevents exhaustion of β cells. Decreased levels of BDNF are associated with neurodegenerative diseases with neuronal loss, such as Parkinson's disease, Alzheimer's disease, multiple sclerosis and Huntington's disease. Thus, BDNF may be useful in the prevention and management of several diseases including diabetes mellitus. PMID:26788077

An inflammation-responsive transcription factor in the pathophysiology of osteoarthritis.

PubMed

Ray, Alpana; Ray, Bimal K

2008-01-01

A number of risk factors including biomechanical stress on the articular cartilage imposed by joint overloading due to obesity, repetitive damage of the joint tissues by injury of the menisci and ligaments, and abnormal joint alignment play a significant role in the onset of osteoarthritis (OA). Genetic predisposition can also lead to the formation of defective cartilage matrix because of abnormal gene expression in the cartilage-specific cells. Another important biochemical event in OA is the consequence of inflammation. It has been shown that synovial inflammation triggers the synthesis of biological stimuli such as cytokines and growth factors which subsequently reach the chondrocyte cells of the articular cartilage activating inflammatory events in the chondrocytes leading to cartilage destruction. In addition to cartilage degradation, hypertrophy of the subchondral bone and osteophyte formation at the joint margins also takes place in OA. Both processes involve abnormal expression of a number of genes including matrix metalloproteinases (MMPs) for cartilage degradation and those associated with bone formation during osteophyte development. To address how diverse groups of genes are activated in OA chondrocyte, we have studied their induction mechanism. We present evidence for abundant expression of an inflammation-responsive transcription factor, SAF-1, in moderate to severely damaged OA cartilage tissues. In contrast, cells in normal cartilage matrix contain very low level of SAF-1 protein. SAF-1 is identified as a major regulator of increased synthesis of MMP-1 and -9 and pro-angiogenic factor, vascular endothelial growth factor (VEGF). While VEGF by stimulating angiogenesis plays a key role in new bone formation in osteophyte, increase of MMP-1 and -9 is instrumental for cartilage erosion in the pathogenesis of OA. Increased expression in degenerated cartilage matrix and in the osteophytes indicate for a key regulatory role of SAF-1 in directing catabolic matrix degrading and anabolic matrix regenerating activities.

p75 Neurotrophin Receptor Signaling Activates Sterol Regulatory Element-binding Protein-2 in Hepatocyte Cells via p38 Mitogen-activated Protein Kinase and Caspase-3.

PubMed

Pham, Dan Duc; Do, Hai Thi; Bruelle, Céline; Kukkonen, Jyrki P; Eriksson, Ove; Mogollón, Isabel; Korhonen, Laura T; Arumäe, Urmas; Lindholm, Dan

2016-05-13

Nerve growth factor (NGF) influences the survival and differentiation of a specific population of neurons during development, but its role in non-neuronal cells has been less studied. We observed here that NGF and its pro-form, pro-NGF, are elevated in fatty livers from leptin-deficient mice compared with controls, concomitant with an increase in low density lipoprotein receptors (LDLRs). Stimulation of mouse primary hepatocytes with NGF or pro-NGF increased LDLR expression through the p75 neurotrophin receptor (p75NTR). Studies using Huh7 human hepatocyte cells showed that the neurotrophins activate the sterol regulatory element-binding protein-2 (SREBP2) that regulates genes involved in lipid metabolism. The mechanisms for this were related to stimulation of p38 mitogen-activated protein kinase (p38 MAPK) and activation of caspase-3 and SREBP2 cleavage following NGF and pro-NGF stimulations. Cell fractionation experiments showed that caspase-3 activity was increased particularly in the membrane fraction that harbors SREBP2 and caspase-2. Experiments showed further that caspase-2 interacts with pro-caspase-3 and that p38 MAPK reduced this interaction and caused caspase-3 activation. Because of the increased caspase-3 activity, the cells did not undergo cell death following p75NTR stimulation, possibly due to concomitant activation of nuclear factor-κB (NF-κB) pathway by the neurotrophins. These results identify a novel signaling pathway triggered by ligand-activated p75NTR that via p38 MAPK and caspase-3 mediate the activation of SREBP2. This pathway may regulate LDLRs and lipid uptake particularly after injury or during tissue inflammation accompanied by an increased production of growth factors, including NGF and pro-NGF. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

The dipteran parasitoid Exorista bombycis induces pro- and anti-oxidative reactions in the silkworm Bombyx mori: Enzymatic and genetic analysis.

PubMed

Makwana, Pooja; Pradeep, Appukuttan Nair R; Hungund, Shambhavi P; Ponnuvel, Kangayam M; Trivedy, Kanika

2017-02-01

Hymenopteran parasitoids inject various factors including polydnaviruses along with their eggs into their host insects that suppress host immunity reactions to the eggs and larvae. Less is known about the mechanisms evolved in dipteran parasitoids that suppress host immunity. Here we report that the dipteran, Exorista bombycis, parasitization leads to pro-oxidative reactions and activation of anti-oxidative enzymes in the silkworm Bombyx mori larva. We recorded increased activity of oxidase, superoxide dismutase, thioredoxin peroxidase, catalase, glutathione-S-transferase (GST), and peroxidases in the hemolymph plasma, hemocytes, and fat body collected from B. mori after E. bombycis parasitization. Microarray and qPCR showed differential expression of genes encoding pro- and anti-oxidant enzymes in the hemocytes. The significance of this work lies in increased understanding of dipteran parasitoid biology. © 2017 Wiley Periodicals, Inc.

Lsd1 regulates skeletal muscle regeneration and directs the fate of satellite cells.

PubMed

Tosic, Milica; Allen, Anita; Willmann, Dominica; Lepper, Christoph; Kim, Johnny; Duteil, Delphine; Schüle, Roland

2018-01-25

Satellite cells are muscle stem cells required for muscle regeneration upon damage. Of note, satellite cells are bipotent and have the capacity to differentiate not only into skeletal myocytes, but also into brown adipocytes. Epigenetic mechanisms regulating fate decision and differentiation of satellite cells during muscle regeneration are not yet fully understood. Here, we show that elevated levels of lysine-specific demethylase 1 (Kdm1a, also known as Lsd1) have a beneficial effect on muscle regeneration and recovery after injury, since Lsd1 directly regulates key myogenic transcription factor genes. Importantly, selective Lsd1 ablation or inhibition in Pax7-positive satellite cells, not only delays muscle regeneration, but changes cell fate towards brown adipocytes. Lsd1 prevents brown adipocyte differentiation of satellite cells by repressing expression of the novel pro-adipogenic transcription factor Glis1. Together, downregulation of Glis1 and upregulation of the muscle-specific transcription program ensure physiological muscle regeneration.

Autophagy promotes apoptosis of mesenchymal stem cells under inflammatory microenvironment.

PubMed

Dang, Shipeng; Yu, Zhi-Ming; Zhang, Chang-Ying; Zheng, Jie; Li, Ku-Lin; Wu, Ying; Qian, Ling-Ling; Yang, Zhen-Yu; Li, Xiao-Rong; Zhang, Yanyun; Wang, Ru-Xing

2015-12-15

Mesenchymal stem cells (MSCs) have been widely applied to treat various inflammatory diseases. Inflammatory cytokines can induce both apoptosis and autophagy in MSCs. However, whether autophagy plays a pro- or con-apoptosis effect on MSCs in an inflammatory microenvironment has not been clarified. We inhibited autophagy by constructing MSCs with lentivirus containing small hairpin RNA to knockdown Beclin-1 and applied these MSCs to a model of sepsis to evaluate therapeutic effect of MSCs. Here we show that inhibition of autophagy in MSCs increases the survival rate of septic mice more than control MSCs, and autophagy promotes apoptosis of MSCs during application to septic mice. Further study demonstrated that autophagy aggravated tumor necrosis factor alpha plus interferon gamma-induced apoptosis of MSCs. Mechanically, autophagy inhibits the expression of the pro-survival gene Bcl-2 via suppressing reactive oxygen species/mitogen-activated protein kinase 1/3 pathway. Our findings indicate that an inflammatory microenvironment-induced autophagy promotes apoptosis of MSCs. Therefore, modulation of autophagy in MSCs would provide a novel approach to improve MSC survival during immunotherapy.

What are the local and systemic biologic reactions and mediators to wear debris, and what host factors determine or modulate the biologic response to wear particles?

PubMed

Tuan, Rocky S; Lee, Francis Young-In; T Konttinen, Yrjö; Wilkinson, J Mark; Smith, Robert Lane

2008-01-01

New clinical and basic science data on the cellular and molecular mechanisms by which wear particles stimulate the host inflammatory response have provided deeper insight into the pathophysiology of periprosthetic bone loss. Interactions among wear particles, macrophages, osteoblasts, bone marrow-derived mesenchymal stem cells, fibroblasts, endothelial cells, and T cells contribute to the production of pro-inflammatory and pro-osteoclastogenic cytokines such as TNF-alpha, RANKL, M-SCF, PGE2, IL-1, IL-6, and IL-8. These cytokines not only promote osteoclastogenesis but interfere with osteogenesis led by osteoprogenitor cells. Recent studies indicate that genetic variations in TNF-alpha, IL-1, and FRZB can result in subtle changes in gene function, giving rise to altered susceptibility or severity for periprosthetic inflammation and bone loss. Continuing research on the biologic effects and mechanisms of action of wear particles will provide a rational basis for the development of novel and effective ways of diagnosis, prevention, and treatment of periprosthetic inflammatory bone loss.

Age-related regulation of genes: slow homeostatic changes and age-dimension technology

NASA Astrophysics Data System (ADS)

Kurachi, Kotoku; Zhang, Kezhong; Huo, Jeffrey; Ameri, Afshin; Kuwahara, Mitsuhiro; Fontaine, Jean-Marc; Yamamoto, Kei; Kurachi, Sumiko

2002-11-01

Through systematic studies of pro- and anti-blood coagulation factors, we have determined molecular mechanisms involving two genetic elements, age-related stability element (ASE), GAGGAAG and age-related increase element (AIE), a unique stretch of dinucleotide repeats (AIE). ASE and AIE are essential for age-related patterns of stable and increased gene expression patterns, respectively. Such age-related gene regulatory mechanisms are also critical for explaining homeostasis in various physiological reactions as well as slow homeostatic changes in them. The age-related increase expression of the human factor IX (hFIX) gene requires the presence of both ASE and AIE, which apparently function additively. The anti-coagulant factor protein C (hPC) gene uses an ASE (CAGGAG) to produce age-related stable expression. Both ASE sequences (G/CAGAAG) share consensus sequence of the transcriptional factor PEA-3 element. No other similar sequences, including another PEA-3 consensus sequence, GAGGATG, function in conferring age-related gene regulation. The age-regulatory mechanisms involving ASE and AIE apparently function universally with different genes and across different animal species. These findings have led us to develop a new field of research and applications, which we named “age-dimension technology (ADT)”. ADT has exciting potential for modifying age-related expression of genes as well as associated physiological processes, and developing novel, more effective prophylaxis or treatments for age-related diseases.

Identification of Novel Inherited Genetic Markers for Aggressive PCa in European and African Americans Using Whole Genome Sequencing

DTIC Science & Technology

2014-10-01

rs115393439 leads to the amino acid substitution from Threonine (Thr) to Proline (Pro); while rs61756080 results to the amino acid substitution from...gene encoding Krüppel-like factor 7 are associated with type 2 diabetes . Diabetologia. 2005 Jul;48(7):1315-22. Epub 2005 Jun 4. Karlsson R, Aly M

IDENTIFICATION AND MOLECULAR CHARACTERIZATION OF TWO SERINE PROTEASES AND THEIR POTENTIAL INVOLVEMENT IN PROPHENOLOXIDASE ACTIVATION IN Plutella xylostella.

PubMed

Gao, Gang; Xu, Xiao-Xia; Yu, Jing; Li, Lin-Miao; Ju, Wen-Yan; Jin, Feng-Liang; Freed, Shoaib

2016-09-01

The proteolytic activation of prophenoloxidase (proPO) is a humoral defense mechanism in insects and crustaceans. Phenoloxidase (PO) is produced as an inactive precursor namely, proPO and is activated via specific proteolytic cleavage by proPO-activating proteinase. The current research reports two novel serine proteinase genes (PxSP1-768 bp and PxSP2-816 bp) from Plutella xylostella, encoding 255 and 271 amino acid residues, respectively. Tissue distribution analyses by semiquantitative reverse transcription-PCR (RT-PCR) revealed the resultant genes to be primarily expressed in the hemocytes, while quantitative-RT-PCR (qRT-PCR) assay showed that transcription level of PxSP1 and PxSP2 increased significantly after injection of the fungal pathogen Beauveria bassiana. Purified recombinant fusion proteins of PxSP2 and PxSP1 were injected to New Zealand white rabbits and polyclonal antibodies were generated with the titers of 1:12,800. After silencing the expression of PxSP2 by RNAi, the PO activity decreased significantly. The results show that PxSP2 is involved in prophenoloxidase activation in P. xylostella. © 2016 Wiley Periodicals, Inc.

Clinical and Mutational Analysis of the GCDH Gene in Malaysian Patients with Glutaric Aciduria Type 1.

PubMed

Abdul Wahab, Siti Aishah; Yakob, Yusnita; Abdul Azize, Nor Azimah; Md Yunus, Zabedah; Huey Yin, Leong; Mohd Khalid, Mohd Khairul Nizam; Lock Hock, Ngu

Glutaric aciduria type 1 (GA1) is an autosomal recessive metabolic disorder caused by deficiency of glutaryl-CoA dehydrogenase enzyme encoded by the GCDH gene. In this study, we presented the clinical and molecular findings of seven GA1 patients in Malaysia. All the patients were symptomatic from infancy and diagnosed clinically from large excretion of glutaric and 3-hydroxyglutaric acids. Bidirectional sequencing of the GCDH gene revealed ten mutations, three of which were novel (Gln76Pro, Glu131Val, and Gly390Trp). The spectrum of mutations included eight missense mutations, a nonsense mutation, and a splice site mutation. Two mutations (Gln76Pro and Arg386Gln) were homozygous in two patients with parental consanguinity. All mutations were predicted to be disease causing by MutationTaster2. In conclusion, this is the first report of both clinical and molecular aspects of GA1 in Malaysian patients. Despite the lack of genotype and phenotype correlation, early diagnosis and timely treatment remained the most important determinant of patient outcome.

Clinical and Mutational Analysis of the GCDH Gene in Malaysian Patients with Glutaric Aciduria Type 1

PubMed Central

Yakob, Yusnita; Abdul Azize, Nor Azimah; Md Yunus, Zabedah; Huey Yin, Leong; Mohd Khalid, Mohd Khairul Nizam; Lock Hock, Ngu

2016-01-01

Glutaric aciduria type 1 (GA1) is an autosomal recessive metabolic disorder caused by deficiency of glutaryl-CoA dehydrogenase enzyme encoded by the GCDH gene. In this study, we presented the clinical and molecular findings of seven GA1 patients in Malaysia. All the patients were symptomatic from infancy and diagnosed clinically from large excretion of glutaric and 3-hydroxyglutaric acids. Bidirectional sequencing of the GCDH gene revealed ten mutations, three of which were novel (Gln76Pro, Glu131Val, and Gly390Trp). The spectrum of mutations included eight missense mutations, a nonsense mutation, and a splice site mutation. Two mutations (Gln76Pro and Arg386Gln) were homozygous in two patients with parental consanguinity. All mutations were predicted to be disease causing by MutationTaster2. In conclusion, this is the first report of both clinical and molecular aspects of GA1 in Malaysian patients. Despite the lack of genotype and phenotype correlation, early diagnosis and timely treatment remained the most important determinant of patient outcome. PMID:27672653

The BOS1 gene encodes an essential 27-kD putative membrane protein that is required for vesicular transport from the ER to the Golgi complex in yeast

PubMed Central

1991-01-01

We recently described the identification of BOS1 (Newman, A., J. Shim, and S. Ferro-Novick. 1990. Mol. Cell. Biol. 10:3405-3414.). BOS1 is a gene that in multiple copy suppresses the growth and secretion defect of bet1 and sec22, two mutants that disrupt transport from the ER to the Golgi complex in yeast. The ability of BOS1 to specifically suppress mutants blocked at a particular stage of the secretory pathway suggested that this gene encodes a protein that functions in this process. The experiments presented in this study support this hypothesis. Specifically, the BOS1 gene was found to be essential for cellular growth. Furthermore, cells depleted of the Bos1 protein fail to transport pro-alpha-factor and carboxypeptidase Y (CPY) to the Golgi apparatus. This defect in export leads to the accumulation of an extensive network of ER and small vesicles. DNA sequence analysis predicts that Bos1 is a 27-kD protein containing a putative membrane- spanning domain. This prediction is supported by differential centrifugation experiments. Thus, Bos1 appears to be a membrane protein that functions in conjunction with Bet1 and Sec22 to facilitate the transport of proteins at a step subsequent to translocation into the ER but before entry into the Golgi apparatus. PMID:2007627

Mycobacterium bovis BCG promotes tumor cell survival from tumor necrosis factor-α-induced apoptosis.

PubMed

Holla, Sahana; Ghorpade, Devram Sampat; Singh, Vikas; Bansal, Kushagra; Balaji, Kithiganahalli Narayanaswamy

2014-09-11

Increased incidence of lung cancer among pulmonary tuberculosis patients suggests mycobacteria-induced tumorigenic response in the host. The alveolar epithelial cells, candidate cells that form lung adenocarcinoma, constitute a niche for mycobacterial replication and infection. We thus explored the possible mechanism of M. bovis Bacillus Calmette-Guérin (BCG)-assisted tumorigenicity in type II epithelial cells, human lung adenocarcinoma A549 and other cancer cells. Cancer cell lines originating from lung, colon, bladder, liver, breast, skin and cervix were treated with tumor necrosis factor (TNF)-α in presence or absence of BCG infection. p53, COP1 and sonic hedgehog (SHH) signaling markers were determined by immunoblotting and luciferase assays, and quantitative real time PCR was done for p53-responsive pro-apoptotic genes and SHH signaling markers. MTT assays and Annexin V staining were utilized to study apoptosis. Gain- and loss-of-function approaches were used to investigate the role for SHH and COP1 signaling during apoptosis. A549 xenografted mice were used to validate the contribution of BCG during TNF-α treatment. Here, we show that BCG inhibits TNF-α-mediated apoptosis in A549 cells via downregulation of p53 expression. Substantiating this observation, BCG rescued A549 xenografts from TNF-α-mediated tumor clearance in nude mice. Furthermore, activation of SHH signaling by BCG induced the expression of an E3 ubiquitin ligase, COP1. SHH-driven COP1 targeted p53, thereby facilitating downregulation of p53-responsive pro-apoptotic genes and inhibition of apoptosis. Similar effects of BCG could be shown for HCT116, T24, MNT-1, HepG2 and HELA cells but not for HCT116 p53(-/-) and MDA-MB-231 cells. Our results not only highlight possible explanations for the coexistence of pulmonary tuberculosis and lung cancer but also address probable reasons for failure of BCG immunotherapy of cancers.

Effect of age on the sensitivity of the rat thyroid gland to ionizing radiation

PubMed Central

Matsuu-Matsuyama, Mutsumi; Shichijo, Kazuko; Okaichi, Kumio; Kurashige, Tomomi; Kondo, Hisayoshi; Miura, Shiro; Nakashima, Masahiro

2015-01-01

Exposure to ionizing radiation during childhood is a well-known risk factor for thyroid cancer. Our study evaluated the effect of age on the radiosensitivity of rat thyroid glands. Four-week-old (4W), 7 -week-old (7W), and 8-month-old (8M) male Wistar rats were exposed to 8 Gy of whole-body X-ray irradiation. Thyroids were removed 3–72 h after irradiation, and non-irradiated thyroids served as controls. Ki67-positivity and p53 binding protein 1 (53BP1) focus formation (a DNA damage response) were evaluated via immunohistochemistry. Amounts of proteins involved in DNA damage response (p53, p53 phosphorylated at serine 15, p21), apoptosis (cleaved caspase-3), and autophagy (LC3, p62) were determined via western blotting. mRNA levels of 84 key autophagy-related genes were quantified using polymerase chain reaction arrays. Ki67-positive cells in 4W (with high proliferative activity) and 7W thyroids significantly decreased in number post-irradiation. The number of 53BP1 foci and amount of p53 phosphorylated at serine 15 increased 3 h after irradiation, regardless of age. No increase in apoptosis or in the levels of p53, p21 or cleaved caspase-3 was detected for any ages. Levels of LC3-II and p62 increased in irradiated 4W but not 8M thyroids, whereas expression of several autophagy-related genes was higher in 4W than 8M irradiated thyroids. Irradiation increased the expression of genes encoding pro-apoptotic proteins in both 4W and 8M thyroids. In summary, no apoptosis or p53 accumulation was noted, despite the expression of some pro-apoptotic genes in immature and adult thyroids. Irradiation induced autophagy in immature, but not in adult, rat thyroids. PMID:25691451

GPR120 in adipocytes has differential roles in the production of pro-inflammatory adipocytokines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hasan, Arif Ul, E-mail: ahasan@med.kagawa-u.ac.jp; Department of Pharmacology, Faculty of Medicine, Kagawa University, 1750-1, Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0793; Ohmori, Koji

How nutritional excess leads to inflammatory responses in metabolic syndrome is not well characterized. Here, we evaluated the effects of ω-3 polyunsaturated fatty acid specific G-protein coupled receptor 120 (GPR120) activation on inflammatory pathways in adipocytes, and the influence of this process on macrophage migration. Using 3T3-L1 adipocytes, we found that agonizing GPR120 using its synthetic ligand, GSK137647, attenuated both basal and lipopolysaccharide-induced production of interleukin-6 (IL-6) and C-C motif chemokine ligand 2 (CCL2). Moreover, the intervention reduced the phosphorylation of nuclear factor kappa B inhibitor alpha (IκBα) and nuclear translocation of nuclear factor kappa-B p65 subunit (p65). Furthermore, themore » silencing of GPR120 itself reduced IL-6 and CCL2 mRNA expression. Inhibition of protein kinase C (PKC) augmented the down-regulatory effect of GSK137647 on IL-6 and CCL2 mRNA. Using a luciferase assay to measure promoter activity of the IL-6 gene in mouse embryonic fibroblasts, we demonstrated that exogenous transfection of GPR120 alone reduced the promoter activity, which was augmented by GSK137647. Inhibition of PKC further reduced the promoter activity. Nevertheless, RAW 264.7 macrophages grown in conditioned medium collected from GSK137647-treated adipocytes attenuated the expressions of matrix metalloproteinases-9 and -3, and tissue inhibitor of metalloproteinase-1. Conditioned medium also inhibited the lipopolysaccharide-induced migration of these macrophages. Taken together, these findings provide critical evidence that although GPR120 is associated with a PKC-mediated pro-inflammatory pathway, the direct inhibitory effects of GPR120 on the nuclear factor kappa B pathway are anti-inflammatory. Moreover, GPR120 activity can attenuate the adipocyte-mediated enhanced production of extracellular matrix-modulating factors in macrophages and can reduce their migration by a paracrine mechanism. - Highlights: • Agonizing GPR120 differentially regulates the pro-inflammatory adipocytokines. • Agonizing GPR120 in adipocytes attenuates NF-κB mediated IL-6 and CCL2 production. • Agonizing GPR120 concomitantly triggers a PKC mediated pro-inflammatory pathway. • However, the resulted effect in adipocytes remains anti-inflammatory. • Agonizing GPR120 in adipocytes reduces macrophage migration in a paracrine manner.« less

Hepatic leukemia factor promotes resistance to cell death: Implications for therapeutics and chronotherapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Waters, Katrina M.; Sontag, Ryan L.; Weber, Thomas J., E-mail: Thomas.Weber@pnl.gov

Physiological variation related to circadian rhythms and aberrant gene expression patterns are believed to modulate therapeutic efficacy, but the precise molecular determinants remain unclear. Here we examine the regulation of cell death by hepatic leukemia factor (HLF), which is an output regulator of circadian rhythms and is aberrantly expressed in human cancers, using an ectopic expression strategy in JB6 mouse epidermal cells and human keratinocytes. Ectopic HLF expression inhibited cell death in both JB6 cells and human keratinocytes, as induced by serum-starvation, tumor necrosis factor alpha and ionizing radiation. Microarray analysis indicates that HLF regulates a complex multi-gene transcriptional programmore » encompassing upregulation of anti-apoptotic genes, downregulation of pro-apoptotic genes, and many additional changes that are consistent with an anti-death program. Collectively, our results demonstrate that ectopic expression of HLF, an established transcription factor that cycles with circadian rhythms, can recapitulate many features associated with circadian-dependent physiological variation. - Highlights: ► Circadian-dependent physiological variation impacts therapeutic efficacy. ► Hepatic leukemia factor inhibits cell death and is a candidate circadian factor. ► Hepatic leukemia factor anti-death program is conserved in murine and human cells. ► Transcriptomics indicates the anti-death program results from a systems response.« less

Suppression of severe achondroplasia with developmental delay and acanthosis nigricans by the p.Thr651Pro mutation.

PubMed

Manickam, Kandamurugu; Donoghue, Daniel J; Meyer, April N; Snyder, Pamela J; Prior, Thomas W

2014-01-01

Severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN) is an extremely rare severe skeletal dysplasia characterized by significant developmental delay, brain structural abnormalities, hearing loss, and acanthosis nigricans. The disorder is the result of a single missense mutation at codon 650 (p.Lys650Met) in the fibroblast growth factor receptor 3 gene (FGFR3). We describe a child who initially presented with a mild achondroplasia or hypochondroplasia like phenotype. Molecular analysis of the FGFR3 gene showed the common SADDAN mutation and a second novel mutation at codon 651 (p.Thr651Pro). Both mutations were shown to occur on the same allele (cis) and de novo. Transient transfection studies with FGFR3 double mutant constructs show that the p.Thr651Pro mutation causes a dramatic decrease in constitutive receptor kinase activity than that observed by the p.Lys650Met mutation. Our data suggest that the molecular effect by the p.Thr651Pro is to elicit a conformational change that decreases the FGFR3 tyrosine kinase activity, which is constitutively activated by the SADDAN mutation. Due to the inheritance of both a gain-of-function and a loss-of-function mutation, we conclude that a reduction of constitutive activation caused the milder skeletal phenotype. Although the occurrence of double mutations are expected to be rare, the presence of other FGFR3 modifiers may be responsible for some of the clinically discrepant skeletal dysplasia cases. © 2013 Wiley Periodicals, Inc.

DUSP5 functions as a feedback regulator of TNFα-induced ERK1/2 dephosphorylation and inflammatory gene expression in adipocytes.

PubMed

Habibian, Justine S; Jefic, Mitra; Bagchi, Rushita A; Lane, Robert H; McKnight, Robert A; McKinsey, Timothy A; Morrison, Ron F; Ferguson, Bradley S

2017-10-10

Adipose tissue inflammation is a central pathological element that regulates obesity-mediated insulin resistance and type II diabetes. Evidence demonstrates that extracellular signal-regulated kinase (ERK 1/2) activation (i.e. phosphorylation) links tumor necrosis factor α (TNFα) to pro-inflammatory gene expression in the nucleus. Dual specificity phosphatases (DUSPs) inactivate ERK 1/2 through dephosphorylation and can thus inhibit inflammatory gene expression. We report that DUSP5, an ERK1/2 phosphatase, was induced in epididymal white adipose tissue (WAT) in response to diet-induced obesity. Moreover, DUSP5 mRNA expression increased during obesity development concomitant to increases in TNFα expression. Consistent with in vivo findings, DUSP5 mRNA expression increased in adipocytes in response to TNFα, parallel with ERK1/2 dephosphorylation. Genetic loss of DUSP5 exacerbated TNFα-mediated ERK 1/2 signaling in 3T3-L1 adipocytes and in adipose tissue of mice. Furthermore, inhibition of ERK 1/2 and c-Jun N terminal kinase (JNK) signaling attenuated TNFα-induced DUSP5 expression. These data suggest that DUSP5 functions in the feedback inhibition of ERK1/2 signaling in response to TNFα, which resulted in increased inflammatory gene expression. Thus, DUSP5 potentially acts as an endogenous regulator of adipose tissue inflammation; although its role in obesity-mediated inflammation and insulin signaling remains unclear.

Plant polyphenols differentially modulate inflammatory responses of human keratinocytes by interfering with activation of transcription factors NF{kappa}B and AhR and EGFR-ERK pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Potapovich, Alla I.; Biology Department, Belarus State University, Skorina Prosp. 10, Minsk 220050; Lulli, Daniela

Molecular mechanisms underlying modulation of inflammatory responses in primary human keratinocytes by plant polyphenols (PPs), namely the glycosylated phenylpropanoid verbascoside, the stilbenoid resveratrol and its glycoside polydatin, and the flavonoid quercetin and its glycoside rutin were evaluated. As non-lethal stimuli, the prototypic ligand for epidermal growth factor receptor (EGFR) transforming growth factor alpha (TGFalpha), the combination of tumor necrosis factor (TNFalpha) and interferon (IFNgamma) (T/I), UVA + UVB irradiation, and bacterial lipopolysaccharide (LPS) were used. We demonstrated differential modulation of inflammatory responses in keratinocytes at signal transduction, gene transcription, and protein synthesis levels as a function of PP chemical structure,more » the pro-inflammatory trigger used, and PP interaction with intracellular detoxifying systems. The PPs remarkably inhibited constitutive, LPS- and T/I-induced but not TGFalpha-induced ERK phosphorylation. They also suppressed NFkappaB activation by LPS and T/I. Verbascoside and quercetin invariably impaired EGFR phosphorylation and UV-associated aryl hydrocarbon receptor (AhR)-mediated signaling, while rutin, polydatin and resveratrol did not affect EGFR phosphorylation and further activated AhR machinery in UV-exposed keratinocytes. In general, PPs down-regulated gene expression of pro-inflammatory cytokines/enzymes, except significant up-regulation of IL-8 observed under stimulation with TGFalpha. Both spontaneous and T/I-induced release of IL-8 and IP-10 was suppressed, although 50 {mu}M resveratrol and polydatin up-regulated IL-8. At this concentration, resveratrol activated both gene expression and de novo synthesis of IL-8 and AhR-mediated mechanisms were involved. We conclude that PPs differentially modulate the inflammatory response of human keratinocytes through distinct signal transduction pathways, including AhR and EGFR. - Graphical abstract: Display Omitted Highlights: > Effects of plant polyphenols on inflammatory responses in human keratinocytes. > Inflammatory stimuli used: TGFalpha, TNFalpha+IFNgamma, UVA+UVB, and LPS. > Inflammatory pathways connected with NFB, ERK1/2, EGFR, and AhR were investigated. > Plant polyphenols, flavonoids, stilbenoids, and phenylpropanoids, were studied. > Modulation of inflammation depends on phenolic core structure and glycosylation.« less

L-Proline uptake in Saccharomyces cerevisiae mitochondria can contribute to bioenergetics during nutrient stress as alternative mitochondrial fuel.

PubMed

Pallotta, Maria Luigia

2014-01-01

L-Proline (pyrrolidine-2-carboxylic acid) is a distinctive metabolite both biochemically and biotechnologically and is currently recognized to have a cardinal role in gene expression and cellular signaling pathways in stress response. Proline-fueled mitochondrial metabolism involves the oxidative conversion of L-Proline to L-Glutamate in two enzymatic steps by means of Put1p and Put2p that help Saccharomyces cerevisiae to respond to changes in the nutritional environment by initiating the breakdown of L-Proline as a source for nitrogen, carbon, and energy. Compartmentalization of L-Proline catabolic pathway implies that extensive L-Proline transport must take place between the cytosol where its biogenesis via Pro1p, Pro2p, Pro3p occurs and mitochondria. L-Proline uptake in S. cerevisiae purified and active mitochondria was investigated by swelling experiments, oxygen uptake and fluorimetric measurement of a membrane potential generation (ΔΨ). Our results strongly suggest that L-Proline uptake occurs via a carried-mediated process as demonstrated by saturation kinetics and experiments with N-ethylmaleimide, a pharmacological compound that is a cysteine-modifying reagent in hydrophobic protein domains and that inhibited mitochondrial transport. Plasticity of S. cerevisiae cell biochemistry according to background fluctuations is an important factor of adaptation to stress. Thus L-Proline → Glutamate route feeds Krebs cycle providing energy and anaplerotic carbon for yeast survival.

The Level of Testosterone, Vitamin D, and Irregular Menstruation More Important than Omega-3 in Non-Symptomatic Women Will Define the Fate of Multiple Scleroses in Future.

PubMed

Tavakol, Shima; Shakibapour, Sahar; Bidgoli, Sepideh Arbabi

2018-01-01

Multiple sclerosis is one of the most salient degenerative disorders of CNS with dysregulated immune process that resulted in axonal damage and demyelination. In the present investigation, the serum level of testosterone was assessed in women who were struggling with multiple sclerosis (MS). Also, the level of omega-3, vitamin D, and the irregular menstruation in women 5 years before the onset MS symptoms were surveyed. Although the levels of omega-3 and vitamin D in women MS patients were non-significant and significantly less than the healthy ones, they were significantly less in the whole population of MS patients. However, the MS patients more experienced more irregular menstruation some years before the onset of MS with the low level of testosterone. Based on the presented findings, it might be said that the vitamin D intake has significant protective role in women and men MS patients unlike the omega-3 that had significant protective role just in men. However, vitamin D metabolism encoding genes of CYP27B1 and CYP24A1 and predicting MS risk gene of HLA-DRB1*15:01 define its fate as well. Besides, vitamin D intake, through the proliferation decrement of pro-inflammatory cells, decreases of pro-inflammatory markers (IL-6, TNF-α, INF-γ) and auto-immune pathways have potential role in recovery of irregular menstruation in women with the low level of testosterone as a red warning factor of MS development. The low level of testosterone and vitamin D consumption increase the neural damage and pro-inflammatory pathways in MS patients, and the difference among the investigations is related to the long-standing history of MS that influences severity of damage to the neural cells and biomolecules and complicate its recovery.

Glycogen synthase kinase 3 regulates expression of nuclear factor-erythroid-2 related transcription factor-1 (Nrf1) and inhibits pro-survival function of Nrf1

PubMed Central

Biswas, Madhurima; Kwong, Erick K.; Park, Eujean; Nagra, Parminder; Chan, Jefferson Y.

2013-01-01

Nuclear factor E2-related factor-1 (Nrf1) is a basic leucine zipper transcription factor that is known to regulate antioxidant and cytoprotective gene expression. It was recently shown that Nrf1 is regulated by SCF-Fbw7 ubiquitin ligase. However our knowledge of upstream signals that targets Nrf1 for degradation by the UPS is not known. We report here that Nrf1 expression is negatively regulated by glycogen synthase kinase 3 (GSK3) in Fbw7-dependent manner. We show that GSK3 interacts with Nrf1 and phosphorylates the Cdc4 phosphodegron domain (CPD) in Nrf1. Mutation of serine residue in the CPD of Nrf1 to alanine (S350A), blocks Nrf1 from phosphorylation by GSK3, and stabilizes Nrf1. Knockdown of Nrf1 and expression of a constitutively active form of GSK3 results in increased apoptosis in neuronal cells in response to ER stress, while expression of the GSK3 phosphorylation resistant S350A–Nrfl attenuates apoptotic cell death. Together these data suggest that GSK3 regulates Nrf1 expression and cell survival function in response to stress activation. PMID:23623971

Leprosy Drug Resistance Surveillance in Colombia: The Experience of a Sentinel Country

PubMed Central

Beltrán-Alzate, Camilo; López Díaz, Fernando; Romero-Montoya, Marcela; Sakamuri, Rama; Li, Wei; Kimura, Miyako; Brennan, Patrick

2016-01-01

An active search for Mycobacterium leprae drug resistance was carried out, 243 multibacillary patients from endemic regions of Colombia were included from 2004 to 2013 in a surveillance program. This program was a World Health Organization initiative for drug resistance surveillance in leprosy, where Colombia is a sentinel country. M. leprae DNA from slit skin smear and/or skin biopsy samples was amplified and sequenced to identify mutations in the drug resistance determining region (DRDR) in rpoB, folP1, gyrA, and gyrB, the genes responsible for rifampicin, dapsone and ofloxacin drug-resistance, respectively. Three isolates exhibited mutations in the DRDR rpoB gene (Asp441Tyr, Ser456Leu, Ser458Met), two in the DRDR folP1 gene (Thr53Ala, Pro55Leu), and one isolate exhibited mutations in both DRDR rpoB (Ser456Met) and DRDR folP1 (Pro55Leu), suggesting multidrug resistance. One isolate had a double mutation in folP1 (Thr53Ala and Thr88Pro). Also, we detected mutations outside of DRDR that required in vivo evaluation of their association or not with drug resistance: rpoB Arg505Trp, folP1 Asp91His, Arg94Trp, and Thr88Pro, and gyrA Ala107Leu. Seventy percent of M. leprae mutations were related to drug resistance and were isolated from relapsed patients; the likelihood of relapse was significantly associated with the presence of confirmed resistance mutations (OR range 20.1–88.7, p < 0.05). Five of these relapsed patients received dapsone monotherapy as a primary treatment. In summary, the current study calls attention to M. leprae resistance in Colombia, especially the significant association between confirmed resistance mutations and relapse in leprosy patients. A high frequency of DRDR mutations for rifampicin was seen in a region where dapsone monotherapy was used extensively. PMID:27706165

A Transcriptional Regulatory Role for the Membrane Type-1 Matrix Metalloproteinase in Carcinogen-Induced Inflammasome Gene Expression.

PubMed

Sheehy, Samuel; Annabi, Borhane

2017-01-01

Signal-transducing functions driven by the cytoplasmic domain of membrane type-1 matrix metalloproteinase (MT1-MMP) are believed to regulate many inflammation-associated cancer cell functions including migration, proliferation, and survival. Aside from upregulation of the inflammation biomarker cyclooxygenase-2 (COX-2) expression, MT1-MMP's role in relaying intracellular signals triggered by extracellular pro-inflammatory cues remains poorly understood. Here, we triggered inflammation in HT1080 fibrosarcoma cells with phorbol-12-myristate-13-acetate (PMA), an inducer of COX-2 and of MT1-MMP. To assess the global transcriptional regulatory role that MT1-MMP may exert on inflammation biomarkers, we combined gene array screens with a transient MT1-MMP gene silencing strategy. Expression of MT1-MMP was found to exert both stimulatory and repressive transcriptional control of several inflammasome-related biomarkers such as interleukin (IL)-1B, IL-6, IL-12A, and IL-33, as well as of transcription factors such as EGR1, ELK1, and ETS1/2 in PMA-treated cells. Among the signal-transducing pathways explored, the silencing of MT1-MMP prevented PMA from phosphorylating extracellular signal-regulated kinase, inhibitor of κB, and p105 nuclear factor κB (NF-κB) intermediates. We also found a signaling axis linking MT1-MMP to MMP-9 transcriptional regulation. Altogether, our data indicate a significant involvement of MT1-MMP in the transcriptional regulation of inflammatory biomarkers consolidating its contribution to signal transduction functions in addition to its classical hydrolytic activity.

The Role of Neurotrophins in Major Depressive Disorder.

PubMed

Jiang, Cheng; Salton, Stephen R

2013-03-01

Neurotrophins and other growth factors have been advanced as critical modulators of depressive behavior. Support for this model is based on analyses of knockout and transgenic mouse models, human genetic studies, and screens for gene products that are regulated by depressive behavior and/or antidepressants. Even subtle alteration in the regulated secretion of brain-derived neurotrophic factor (BDNF), for example, due to a single nucleotide polymorphism (SNP)-encoded Val-Met substitution in proBDNF that affects processing and sorting, impacts behavior and cognition. Alterations in growth factor expression result in changes in neurogenesis as well as structural changes in neuronal cytoarchitecture, including effects on dendritic length and spine density, in the hippocampus, nucleus accumbens, and prefrontal cortex. These changes have the potential to impact the plasticity and stability of synapses in the CNS, and the complex brain circuitry that regulates behavior. Here we review the role that neurotrophins play in the modulation of depressive behavior, and the downstream signaling targets they regulate that potentially mediate these behavioral pro-depressant and antidepressant effects.

The Role of Neurotrophins in Major Depressive Disorder

PubMed Central

Jiang, Cheng; Salton, Stephen R.

2013-01-01

Neurotrophins and other growth factors have been advanced as critical modulators of depressive behavior. Support for this model is based on analyses of knockout and transgenic mouse models, human genetic studies, and screens for gene products that are regulated by depressive behavior and/or antidepressants. Even subtle alteration in the regulated secretion of brain-derived neurotrophic factor (BDNF), for example, due to a single nucleotide polymorphism (SNP)-encoded Val-Met substitution in proBDNF that affects processing and sorting, impacts behavior and cognition. Alterations in growth factor expression result in changes in neurogenesis as well as structural changes in neuronal cytoarchitecture, including effects on dendritic length and spine density, in the hippocampus, nucleus accumbens, and prefrontal cortex. These changes have the potential to impact the plasticity and stability of synapses in the CNS, and the complex brain circuitry that regulates behavior. Here we review the role that neurotrophins play in the modulation of depressive behavior, and the downstream signaling targets they regulate that potentially mediate these behavioral pro-depressant and antidepressant effects. PMID:23691270

Overexpression of cellular repressor of E1A-stimulated genes inhibits TNF-{alpha}-induced apoptosis via NF-{kappa}B in mesenchymal stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Peng, Cheng-Fei; Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang; Han, Ya-Ling, E-mail: hanyaling53@gmail.com

2011-03-25

Research highlights: {yields} CREG protected MSCs from tumor necrosis factor-{alpha} (TNF-{alpha}) induced apoptosis. {yields} CREG inhibits the phosphorylation of I{kappa}B{alpha} and prevents the activation of NF-{kappa}B. {yields} CREG inhibits NF-{kappa}B nuclear translocation and pro-apoptosis protein transcription. {yields} CREG anti-apoptotic effect involves inhibition of the death receptor pathway. {yields} p53 is downregulated by CREG via NF-{kappa}B pathway under TNF-{alpha} stimulation. -- Abstract: Bone marrow-derived mesenchymal stem cells (MSCs) show great potential for therapeutic repair after myocardial infarction. However, poor viability of transplanted MSCs in the ischemic heart has limited their use. Cellular repressor of E1A-stimulated genes (CREG) has been identified asmore » a potent inhibitor of apoptosis. This study therefore aimed to determine if rat bone marrow MSCs transfected with CREG-were able to effectively resist apoptosis induced by inflammatory mediators, and to demonstrate the mechanism of CREG action. Apoptosis was determined by flow cytometric and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling assays. The pathways mediating these apoptotic effects were investigated by Western blotting. Overexpression of CREG markedly protected MSCs from tumor necrosis factor-{alpha} (TNF-{alpha}) induced apoptosis by 50% after 10 h, through inhibition of the death-receptor-mediated apoptotic pathway, leading to attenuation of caspase-8 and caspase-3. Moreover, CREG resisted the serine phosphorylation of I{kappa}B{alpha} and prevented the nuclear translocation of the transcription factor nuclear factor-{kappa}B (NF-{kappa}B) under TNF-{alpha} stimulation. Treatment of cells with the NF-{kappa}B inhibitor pyrrolidine dithiocarbamate (PDTC) significantly increased the transcription of pro-apoptosis proteins (p53 and Fas) by NF-{kappa}B, and attenuated the anti-apoptotic effects of CREG on MSCs. The results of this study indicate that CREG acts as a novel and potent survival factor in MSCs, and may therefore be a useful therapeutic adjunct for transplanting MSCs into the damaged heart after myocardial infarction.« less

l-lysine production by Bacillus methanolicus: Genome-based mutational analysis and l-lysine secretion engineering.

PubMed

Nærdal, Ingemar; Netzer, Roman; Irla, Marta; Krog, Anne; Heggeset, Tonje Marita Bjerkan; Wendisch, Volker F; Brautaset, Trygve

2017-02-20

Bacillus methanolicus is a methylotrophic bacterium with an increasing interest in academic research and for biotechnological applications. This bacterium was previously applied for methanol-based production of l-glutamate, l-lysine and the five-carbon diamine cadaverine by wild type, classical mutant and recombinant strains. The genomes of two different l-lysine secreting B. methanolicus classical mutant strains, NOA2#13A52-8A66 and M168-20, were sequenced. We focused on mutational mapping in genes present in l-lysine and other relevant amino acid biosynthetic pathways, as well as in the primary cell metabolism important for precursor supply. In addition to mutations in the aspartate pathway genes dapG, lysA and hom-1, new mutational target genes like alr, proA, proB1, leuC, odhA and pdhD were identified. Surprisingly, no mutations were found in the putative l-lysine transporter gene lysE MGA3 . Inspection of the wild type B. methanolicus strain PB1 genome sequence identified two homologous putative l-lysine transporter genes, lysE PB1 and lysE2 PB1 . The biological role of these putative l-lysine transporter genes, together with the heterologous l-lysine exporter gene lysE Cg from Corynebacterium glutamicum, were therefore investigated. Our results demonstrated that the titer of secreted l-lysine in B. methanolicus was significantly increased by overexpression of lysE Cg while overexpression of lysE MGA3 , lysE PB1 and lysE2 PB1 had no measurable effect. Copyright © 2017 Elsevier B.V. All rights reserved.

Association of the Pro12Ala polymorphism in peroxisome proliferator-activated receptor gamma2 with decreased basic metabolic rate in women with polycystic ovary syndrome.

PubMed

Koika, Vasiliki; Marioli, Dimitra J; Saltamavros, Alexandros D; Vervita, Vasiliki; Koufogiannis, Kleanthis D; Adonakis, George; Decavalas, George; Georgopoulos, Neoklis A

2009-08-01

The peroxisome proliferator-activated receptor (PPAR)gamma is a transcription factor involved in glucose homeostasis and energy metabolism. A missense mutation at codon 12 in the PPARgamma2 has been associated with increased body mass index (BMI) and attenuated insulin resistance (IR) in polycystic ovary syndrome (PCOS). We have recently shown a decreased basic metabolic rate (BMR) in PCOS. The aim of the present study is to determine the prevalence of the Pro12Ala polymorphism of the PPARgamma2 gene and its associations with indices of IR and BMR in lean and slightly overweight PCOS women. Case-control association study involving 156 PCOS women with biochemical hyperandrogenism, chronic anovulation and polycystic ovarian morphology in ultrasound and 56 unrelated healthy controls. Hormonal determinations were performed by electrochemiluminescence quantitation or RIA. BMR was measured by indirect calorimetry. All subjects were genotyped by a PCR-restriction fragment length polymorphism assay. Genotype frequencies of the Pro12Ala polymorphism in PPARgamma2 did not differ among PCOS women and control subjects. The presence of Pro12Ala polymorphism of PPARgamma2 was associated with lower BMR (P=0.04). This finding was valid in our subgroup of lean PCOS (BMI<25 kg/m(2)), in which the Ala variant was also associated with higher total testosterone values. The Pro12Ala polymorphism in the PPARgamma2 gene is associated with decreased BMR in women with PCOS and biochemical hyperandrogenemia. These young women are therefore at risk to increase their body weight and should restrict their energy intake by diet and enhance their energy expenditure by exercise.

N-terminal pro-brain-type natriuretic peptide (NT-pro-BNP) and mortality risk in early inflammatory polyarthritis: results from the Norfolk Arthritis Registry (NOAR)

PubMed Central

Mirjafari, Hoda; Welsh, Paul; Verstappen, Suzanne M M; Wilson, Paddy; Marshall, Tarnya; Edlin, Helena; Bunn, Diane; Chipping, Jacqueline; Lunt, Mark; Symmons, Deborah P M; Sattar, Naveed; Bruce, Ian N

2014-01-01

Background We measured N-terminal pro-brain natriuretic peptide (NT-pro-BNP), a marker of cardiac dysfunction, in an inception cohort with early inflammatory polyarthritis (IP) and assessed its association with disease phenotype, cardiovascular disease (CVD), all-cause and CVD related mortality. Methods Subjects with early IP were recruited to the Norfolk Arthritis Register from January 2000 to December 2008 and followed up to death or until March 2010 including any data from the national death register. The associations of baseline NT-pro-BNP with IP related factors and CVD were assessed by linear regression. Cox proportional hazards models examined the independent association of baseline NT-pro-BNP with all-cause and CVD mortality. Results We studied 960 early IP subjects; 163 (17%) had prior CVD. 373 (39%) patients had a baseline NT-pro-BNP levels ≥100 pg/ml. NT-pro-BNP was associated with age, female gender, HAQ score, CRP, current smoking, history of hypertension, prior CVD and the presence of carotid plaque. 92 (10%) IP subjects died including 31 (3%) from CVD. In an age and gender adjusted analysis, having a raised NT-pro-BNP level (≥100 pg/ml) was associated with both all-cause and CVD mortality (adjusted HR (95% CI) 2.36 (1.42 to 3.94) and 3.40 (1.28 to 9.03), respectively). These findings were robust to adjustment for conventional CVD risk factors and prevalent CVD. Conclusions In early IP patients, elevated NT-pro-BNP is related to HAQ and CRP and predicts all-cause and CVD mortality independently of conventional CVD risk factors. Further study is required to identify whether NT-pro-BNP may be clinically useful in targeting intensive interventions to IP patients at greatest risk of CVD. PMID:23511225

Novel compound heterozygous DPH1 mutations in a patient with the unique clinical features of airway obstruction and external genital abnormalities.

PubMed

Nakajima, Junya; Oana, Shingo; Sakaguchi, Tomohiro; Nakashima, Mitsuko; Numabe, Hironao; Kawashima, Hisashi; Matsumoto, Naomichi; Miyake, Noriko

2018-04-01

The diphthamide biosynthesis 1 (DPH1) gene encodes one of the essential components of the enzyme catalyzing the first step of diphthamide formation on eukaryotic elongation factor 2 (EEF2). Diphthamide is the posttranslationally modified histidine residue on EEF2 that promotes protein chain elongation in the ribosome. DPH1 defects result in a failure of protein synthesis involving EEF2, leading to growth defects, embryonic lethality, and cell death. In humans, DPH1 mutations cause developmental delay with a short stature, dysmorphic features, and sparse hair, and are inherited in an autosomal recessive manner (MIM#616901). To date, only two homozygous missense mutations in DPH1 (c.17T>A, p.Met6Lys and c.701T>C, p.Leu234Pro) have been reported. We used WES to identify novel compound heterozygous mutations in DPH1 (c.289delG, p.Glu97Lysfs*8 and c.491T>C, p.Leu164Pro) in a patient from a nonconsanguineous family presenting with intellectual disability, a short stature, craniofacial abnormalities, and external genital abnormalities. The clinical phenotype of all patients with DPH1 mutations, including the current patient, revealed core features, although the external genital anomaly was newly recognized in our case.

Regulation of LPS-induced mRNA expression of pro-inflammatory cytokines via alteration of NF-κB activity in mouse peritoneal macrophages exposed to fluoride.

PubMed

Tian, Yuhu; Huo, Meijun; Li, Guangsheng; Li, Yanyan; Wang, Jundong

2016-10-01

F toxicity to immune system, especially to macrophage, has been studied a lot recently. Nuclear factor-kappa B (NF-κB), as a transcription factor, plays a central role in immune and inflammatory responses via the regulation of downstream gene expression. Recent studies indicated that fluoride effect on inflammatory cytokine secretion, however, the molecular mechanism was less understood. In our study, peritoneal macrophages (PMs) were divided several groups and were administrated sodium fluoride (NaF, 50, 100, 200, 400, 800 μM) and/or lipopolysaccharide (LPS, 30 ng/mg). The mRNA expression of p65, inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) in macrophages exposed to fluoride was determined by quantitative real-time RT-PCR respectively. The translocation of NF-κB from cytoplasm to nucleus, which in a way reflects NF-κB activity, was demonstrated by Immunofluorescence and ELISA. Our results showed that fluoride had a dose-dependent effect on NF-κB activity, which coincided with LPS-induced mRNA expression of its downstream genes, iNOS and IL-1β. Fluoride alone causes no effect on gene expression. However, the mRNA expression of TNF-α showed non-NF-κB-dependent manner. Therefore, we come to the conclusion that fluoride can regulate LPS-induced mRNA expression of iNOS and IL-1β via NF-κB pathway in mouse peritoneal macrophages. Copyright © 2016 Elsevier Ltd. All rights reserved.

Regulation of apoptosis by peroxisome proliferators.

PubMed

Roberts, Ruth A; Michel, Cecile; Coyle, Beth; Freathy, Caroline; Cain, Kelvin; Boitier, Eric

2004-04-01

Peroxisome proliferators (PPs) constitute a large and chemically diverse family of non-genotoxic rodent hepatocarcinogens that activate the PP-activated receptor alpha (PPARalpha). In order to investigate the hypothesis that PPs elicit their carcinogenic effects through the suppression of apoptosis, we established an in vitro assay for apoptosis using both primary rat hepatocytes and the FaO rat hepatoma cell line. Apoptosis was induced by transforming growth factor beta1 (TGFbeta1), the physiological negative regulator of liver growth. In this system, PPs could suppress both spontaneous and TGFbeta1-induced apoptosis. In order to understand the mechanisms of this regulation of apoptosis, we conducted microarray analysis followed by pathway-specific gene clustering in TGFbeta1-treated cells. After treatment, 76 genes were up-regulated and 185 were down-regulated more than 1.5-fold. Cluster analysis of up-regulated genes revealed three clusters, A-C. Cluster A (4h) was associated with 12% apoptosis and consisted of genes mainly of the cytoskeleton and extracellular matrix such as troponin and the proteoglycan SDC4. In cluster B (8h; 25% apoptosis), there were many pro- and anti-apoptotic genes such as XIAP, BAK1 and BAD, whereas at 16h (40% apoptosis) the regulated genes were mainly those of the cellular stress pathways such as the genes implicated in the activation of the transcription factor NFkappab. Genes found down-regulated in response to TGFbeta1 were mainly those associated with oxidative stress and several genes implicated in glutathione production and maintenance. Thus, TGFbeta1 may induce apoptosis via a down regulation of oxidant defence leading to the generation of reactive oxygen species. The ability of PPs to impact on these apoptosis pathways remains to be determined. To approach this question, we have developed a technique using laser capture microdissection of livers treated with the PP, clofibric acid coupled with gene expression array analysis. Results show that some of the key steps of the LCM process had an impact on the gene profiles generated. However, this did not preclude accurate determination of a PP-specific molecular signature. Thus, the choice of appropriate controls will ensure that meaningful gene expression analyses can be performed on tissue microdissected from the foci generated in clofibric acid treated livers. These data will allow the identification of specific genes that are regulated by PPs leading to changes in apoptosis and ultimately to tumours.

Interactions of the PPARγ2 polymorphism with fat intake affecting energy metabolism and nutritional outcomes in obese women.

PubMed

Rosado, Eliane L; Bressan, Josefina; Martínez, J Alfredo; Marques-Lopes, Iva

2010-01-01

To determine the influence of the Pro12Ala polymorphism of the PPARγ2 gene and the dietary lipid intake on energy metabolism and nutritional outcomes in obese women after an acute fat load or following a low-calorie diet for 10 weeks. Sixty obese women (aged 30-46 years) participated in the study and were assigned to 2 groups depending on the genotype: Pro12Pro and Pro12Ala/Ala12Ala carriers. At baseline and after 2 nutritional (short- or long-term) interventions, measurement of anthropometrical and body composition (bioelectrical impedance) variables, dietary assessments, energy metabolism (indirect calorimetry) measurements as well as biochemical and molecular (PPARγ2 genotype) analyses were performed. All women received a high-fat test meal to determine the postprandial metabolism (short term) and an energy-restricted diet for 10 weeks (long term). The frequencies of the Pro12Pro and Pro12Ala/Ala12Ala genotypes were 83.33 and 16.67%, respectively, and reached Hardy-Weinberg equilibrium. Interestingly, the postprandial energy expenditure after the fat load was higher in subjects carrying the Ala allele. At baseline, the habitual monounsaturated fatty acid (MUFA) intake inversely correlated with fat oxidation and body mass index in the obese Pro12Ala/Ala12Ala carriers, while a lower PUFA intake (%) in the long-term trial was associated with an increase in the respiratory quotient only in Ala carriers but not in the Pro12Pro genotyped group. The Pro12Ala polymorphism in the PPARγ2 gene influenced energy metabolism in the assayed short- and long-term situations since the response to both nutritional interventions differed according to the genotype. The results suggest that fat oxidation and energy expenditure may be lower in Pro12Pro carriers compared to Pro12Ala/Ala12Ala genotypes, while in obese women with Pro12Ala/Ala12Ala polymorphisms in the PPARγ2 gene fat oxidation was negatively correlated with the MUFA and PUFA (%) intake. Copyright © 2010 S. Karger AG, Basel.

N-terminal pro-B-type natriuretic peptide and the prediction of primary cardiovascular events: results from 15-year follow-up of WOSCOPS

PubMed Central

Welsh, Paul; Doolin, Orla; Willeit, Peter; Packard, Chris; Macfarlane, Peter; Cobbe, Stuart; Gudnason, Vilmundur; Di Angelantonio, Emanuele; Ford, Ian; Sattar, Naveed

2013-01-01

Aims To test whether N-terminal pro-B-type natriuretic peptide (NT-proBNP) was independently associated with, and improved the prediction of, cardiovascular disease (CVD) in a primary prevention cohort. Methods and results In the West of Scotland Coronary Prevention Study (WOSCOPS), a cohort of middle-aged men with hypercholesterolaemia at a moderate risk of CVD, we related the baseline NT-proBNP (geometric mean 28 pg/mL) in 4801 men to the risk of CVD over 15 years during which 1690 experienced CVD events. Taking into account the competing risk of non-CVD death, NT-proBNP was associated with an increased risk of all CVD [HR: 1.17 (95% CI: 1.11–1.23) per standard deviation increase in log NT-proBNP] after adjustment for classical and clinical cardiovascular risk factors plus C-reactive protein. N-terminal pro-B-type natriuretic peptide was more strongly related to the risk of fatal [HR: 1.34 (95% CI: 1.19–1.52)] than non-fatal CVD [HR: 1.17 (95% CI: 1.10–1.24)] (P= 0.022). The addition of NT-proBNP to traditional risk factors improved the C-index (+0.013; P < 0.001). The continuous net reclassification index improved with the addition of NT-proBNP by 19.8% (95% CI: 13.6–25.9%) compared with 9.8% (95% CI: 4.2–15.6%) with the addition of C-reactive protein. N-terminal pro-B-type natriuretic peptide correctly reclassified 14.7% of events, whereas C-reactive protein correctly reclassified 3.4% of events. Results were similar in the 4128 men without evidence of angina, nitrate prescription, minor ECG abnormalities, or prior cerebrovascular disease. Conclusion N-terminal pro-B-type natriuretic peptide predicts CVD events in men without clinical evidence of CHD, angina, or history of stroke, and appears related more strongly to the risk for fatal events. N-terminal pro-B-type natriuretic peptide also provides moderate risk discrimination, in excess of that provided by the measurement of C-reactive protein. Clinical trial registration WOSCOPS was carried out and completed prior to the requirement for clinical trial registration. PMID:22942340

Increased sorbitol levels in the hypertrophic ligamentum flavum of diabetic patients with lumbar spinal canal stenosis.

PubMed

Luo, Jiaquan; Huang, Lu; Chen, Zhuo; Zeng, Zhaoxun; Miyamoto, Takeshi; Wu, Hao; Zhang, Zhongzu; Pan, Zhimin; Fujita, Nobuyuki; Hikata, Tomohiro; Iwanami, Akio; Tsuji, Takashi; Ishii, Ken; Nakamura, Masaya; Matsumoto, Morio; Watanabe, Kota; Cao, Kai

2017-05-01

The pathomechanism of the ligamentum flavum (LF) hypertrophy in diabetic patients with lumbar spinal canal stenosis (LSCS) remains unclear. A cross-sectional study was undertaken to investigate the mechanism of LF hypertrophy in these patients. Twenty-four diabetic and 20 normoglycemic patients with LSCS were enrolled in the study. The structure of the LF in the study subjects was evaluated using histological and immunohistochemical methods, and the levels of sorbitol, pro-inflammatory cytokines, and the fibrogenic factor, TGF-β1, in the LF were analyzed. In vitro experiments were performed using NIH3T3 fibroblasts to evaluate the effect of high-glucose conditions and an aldose reductase inhibitor on the cellular production of sorbitol, pro-inflammatory factors, and TGF-β1. We found that the LF of diabetic patients exhibited significantly higher levels of sorbitol and pro-inflammatory cytokines, TGF-β1 and of CD68-positive staining than that of the normoglycemic subjects. The diabetic LF was significantly thicker than that of the controls, and showed evidence of degeneration. The high glucose-cultured fibroblasts exhibited significantly higher levels of sorbitol, pro-inflammatory factors, and TGF-β1 compared to the low glucose-cultured cells, and these levels were dose-dependently reduced by treatment with the aldose reductase inhibitor. Taken together, our data suggests that increased sorbitol levels in the LF of diabetic patients results in increased production of pro-inflammatory and fibrogenic factor, which contribute to LF hypertrophy, and could increase the susceptibility of diabetic patients to LSCS. Furthermore, aldose reductase inhibition effectively reduced the levels of sorbitol and sorbitol-induced pro-inflammatory factor expression in high glucose-cultured fibroblasts. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1058-1066, 2017. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

The genetic component of human longevity: New insights from the analysis of pathway-based SNP-SNP interactions.

PubMed

Dato, Serena; Soerensen, Mette; De Rango, Francesco; Rose, Giuseppina; Christensen, Kaare; Christiansen, Lene; Passarino, Giuseppe

2018-06-01

In human longevity studies, single nucleotide polymorphism (SNP) analysis identified a large number of genetic variants with small effects, yet not easily replicable in different populations. New insights may come from the combined analysis of different SNPs, especially when grouped by metabolic pathway. We applied this approach to study the joint effect on longevity of SNPs belonging to three candidate pathways, the insulin/insulin-like growth factor signalling (IIS), DNA repair and pro/antioxidant. We analysed data from 1,058 tagging SNPs in 140 genes, collected in 1825 subjects (1,089 unrelated nonagenarians from the Danish 1905 Birth Cohort Study and 736 Danish controls aged 46-55 years) for evaluating synergic interactions by SNPsyn. Synergies were further tested by the multidimensional reduction (MDR) approach, both intra- and interpathways. The best combinations (FDR<0.0001) resulted those encompassing IGF1R-rs12437963 and PTPN1-rs6067484, TP53-rs2078486 and ERCC2-rs50871, TXNRD1-rs17202060 and TP53-rs2078486, the latter two supporting a central role of TP53 in mediating the concerted activation of the DNA repair and pro-antioxidant pathways in human longevity. Results were consistently replicated with both approaches, as well as a significant effect on longevity was found for the GHSR gene, which also interacts with partners belonging to both IIS and DNA repair pathways (PAPPA, PTPN1, PARK7, MRE11A). The combination GHSR-MREA11, positively associated with longevity by MDR, was further found influencing longitudinal survival in nonagenarian females (p = .026). Results here presented highlight the validity of SNP-SNP interactions analyses for investigating the genetics of human longevity, confirming previously identified markers but also pointing to novel genes as central nodes of additional networks involved in human longevity. © 2018 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

Ablation of the Regulatory IE1 Protein of Murine Cytomegalovirus Alters In Vivo Pro-inflammatory TNF-alpha Production during Acute Infection

PubMed Central

Wilhelmi, Vanessa; Lisnic, Vanda Juranic; Hsieh, Wei Yuan; Blanc, Mathieu; Livingston, Andrew; Busche, Andreas; Tekotte, Hille; Messerle, Martin; Auer, Manfred; Fraser, Iain; Jonjic, Stipan; Angulo, Ana; Reddehase, Matthias J.; Ghazal, Peter

2012-01-01

Little is known about the role of viral genes in modulating host cytokine responses. Here we report a new functional role of the viral encoded IE1 protein of the murine cytomegalovirus in sculpting the inflammatory response in an acute infection. In time course experiments of infected primary macrophages (MΦs) measuring cytokine production levels, genetic ablation of the immediate-early 1 (ie1) gene results in a significant increase in TNFα production. Intracellular staining for cytokine production and viral early gene expression shows that TNFα production is highly associated with the productively infected MΦ population of cells. The ie1- dependent phenotype of enhanced MΦ TNFα production occurs at both protein and RNA levels. Noticeably, we show in a series of in vivo infection experiments that in multiple organs the presence of ie1 potently inhibits the pro-inflammatory cytokine response. From these experiments, levels of TNFα, and to a lesser extent IFNβ, but not the anti-inflammatory cytokine IL10, are moderated in the presence of ie1. The ie1- mediated inhibition of TNFα production has a similar quantitative phenotype profile in infection of susceptible (BALB/c) and resistant (C57BL/6) mouse strains as well as in a severe immuno-ablative model of infection. In vitro experiments with infected macrophages reveal that deletion of ie1 results in increased sensitivity of viral replication to TNFα inhibition. However, in vivo infection studies show that genetic ablation of TNFα or TNFRp55 receptor is not sufficient to rescue the restricted replication phenotype of the ie1 mutant virus. These results provide, for the first time, evidence for a role of IE1 as a regulator of the pro-inflammatory response and demonstrate a specific pathogen gene capable of moderating the host production of TNFα in vivo. PMID:22952450

Effects of probiotics on methionine choline deficient diet-induced steatohepatitis in rats.

PubMed

Karahan, Nermin; Işler, Mehmet; Koyu, Ahmet; Karahan, Aynur G; Başyığıt Kiliç, Gülden; Cırış, Ibrahim Metin; Sütçü, Recep; Onaran, Ibrahim; Cam, Hakan; Keskın, Muharrem

2012-04-01

Intestinal bacteria induce endogenous signals that play a pathogenic role in hepatic insulin resistance and non-alcoholic fatty liver disease. Probiotics could modulate the gut flora and could influence the gut-liver axis. We aimed to investigate the preventive effect of two probiotic mixtures on the methionine choline-deficient diet-induced non-alcoholic steatohepatitis model in rats. Two studies, short-term (2 weeks) and long-term (6 weeks), were carried out using 60 male Wistar rats. The 2-week study included six groups. Rats were fed with methionine choline-deficient diet or pair-fed control diet and were given a placebo or one of two probiotic mixtures (Pro-1 and Pro-2) by orogastric gavage. In the 6-week study, rats were allocated into four groups and were fed with methionine choline-deficient diet or pair-fed control diet and given a placebo or Pro-2. At the end of the 2- and 6-week periods, blood samples were obtained, the animals were sacrificed, and liver tissues were removed. Serum alanine aminotransferase activity was determined; histologic and immunohistochemical analysis was performed for steatosis, inflammation, protein expression of tumor necrosis factor-α, and apoptosis markers. In both studies, methionine choline-deficient diet caused an elevation of serum alanine aminotransferase activity, which was slightly reduced by Pro-1 and Pro-2. In the 2- and 6-week studies, feeding with methionine choline-deficient diet resulted in steatosis and inflammation, but not fibrosis, in all rats. In the 2-week study, in rats fed with methionine choline-deficient diet and given Pro-1, steatosis and inflammation were present in 2 of 6 rats. In rats fed with methionine choline-deficient diet and given Pro-2, steatosis was detected in 3 of 6 rats, while inflammation was present in 2 of 6 rats. In the 6-week study, in rats fed with methionine choline-deficient diet and given Pro-2, steatosis and inflammation were present in 3 of 6 rat livers. In both the 2- and 6-week studies, methionine choline-deficient diet resulted in tumor necrosis factor-α, proapoptotic Bax, caspase 3, caspase 8, and anti-apoptotic Bcl-2 expression in all rat livers. Pro-1 and Pro-2 treatment influenced protein expression involved in apoptosis and tumor necrosis factor-α in varying degrees. Pro-1 and Pro-2 decrease methionine choline-deficient diet-induced steatohepatitis in rats. The preventive effect of probiotics may be due, in part, to modulation of apoptosis and their anti-inflammatory activity.

Tumor p38MAPK signaling enhances breast carcinoma vascularization and growth by promoting expression and deposition of pro-tumorigenic factors.

PubMed

Limoge, Michelle; Safina, Alfiya; Truskinovsky, Alexander M; Aljahdali, Ieman; Zonneville, Justin; Gruevski, Aleksandar; Arteaga, Carlos L; Bakin, Andrei V

2017-09-22

The breast carcinoma microenvironment strikingly influences cancer progression and response to therapy. Various cell types in the carcinoma microenvironment show significant activity of p38 mitogen-activated protein kinase (MAPK), although the role of p38MAPK in breast cancer progression is still poorly understood. The present study examined the contribution of tumor p38MAPK to breast carcinoma microenvironment and metastatic capacity. Inactivation of p38MAPK signaling in metastatic breast carcinoma cells was achieved by forced expression of the kinase-inactive mutant of p38/MAPK14 (a dominant-negative p38, dn-p38). Disruption of tumor p38MAPK signaling reduced growth and metastases of breast carcinoma xenografts. Importantly, dn-p38 markedly decreased tumor blood-vessel density and lumen sizes. Mechanistic studies revealed that p38 controls expression of pro-angiogenic extracellular factors such as matrix protein Fibronectin and cytokines VEGFA, IL8, and HBEGF. Tumor-associated fibroblasts enhanced tumor growth and vasculature as well as increased expression of the pro-angiogenic factors. These effects were blunted by dn-p38. Metadata analysis showed elevated expression of p38 target genes in breast cancers and this was an unfavorable marker of disease recurrence and poor-outcome. Thus, our study demonstrates that tumor p38MAPK signaling promotes breast carcinoma growth, invasive and metastatic capacities. Importantly, p38 enhances carcinoma vascularization by facilitating expression and deposition of pro-angiogenic factors. These results argue that p38MAPK is a valuable target for anticancer therapy affecting tumor vasculature. Anti-p38 drugs may provide new therapeutic strategies against breast cancer, including metastatic disease.

Tumor p38MAPK signaling enhances breast carcinoma vascularization and growth by promoting expression and deposition of pro-tumorigenic factors

PubMed Central

Limoge, Michelle; Safina, Alfiya; Truskinovsky, Alexander M.; Aljahdali, Ieman; Zonneville, Justin; Gruevski, Aleksandar; Arteaga, Carlos L.; Bakin, Andrei V.

2017-01-01

The breast carcinoma microenvironment strikingly influences cancer progression and response to therapy. Various cell types in the carcinoma microenvironment show significant activity of p38 mitogen-activated protein kinase (MAPK), although the role of p38MAPK in breast cancer progression is still poorly understood. The present study examined the contribution of tumor p38MAPK to breast carcinoma microenvironment and metastatic capacity. Inactivation of p38MAPK signaling in metastatic breast carcinoma cells was achieved by forced expression of the kinase-inactive mutant of p38/MAPK14 (a dominant-negative p38, dn-p38). Disruption of tumor p38MAPK signaling reduced growth and metastases of breast carcinoma xenografts. Importantly, dn-p38 markedly decreased tumor blood-vessel density and lumen sizes. Mechanistic studies revealed that p38 controls expression of pro-angiogenic extracellular factors such as matrix protein Fibronectin and cytokines VEGFA, IL8, and HBEGF. Tumor-associated fibroblasts enhanced tumor growth and vasculature as well as increased expression of the pro-angiogenic factors. These effects were blunted by dn-p38. Metadata analysis showed elevated expression of p38 target genes in breast cancers and this was an unfavorable marker of disease recurrence and poor-outcome. Thus, our study demonstrates that tumor p38MAPK signaling promotes breast carcinoma growth, invasive and metastatic capacities. Importantly, p38 enhances carcinoma vascularization by facilitating expression and deposition of pro-angiogenic factors. These results argue that p38MAPK is a valuable target for anticancer therapy affecting tumor vasculature. Anti-p38 drugs may provide new therapeutic strategies against breast cancer, including metastatic disease. PMID:28977919

Bioaerosols from a food waste composting plant affect human airway epithelial cell remodeling genes.

PubMed

Chang, Min-Wei; Lee, Chung-Ru; Hung, Hsueh-Fen; Teng, Kuo-Sheng; Huang, Hsin; Chuang, Chun-Yu

2013-12-24

The composting procedure in food waste plants generates airborne bioaerosols that have the potential to damage human airway epithelial cells. Persistent inflammation and repair responses induce airway remodeling and damage to the respiratory system. This study elucidated the expression changes of airway remodeling genes in human lung mucoepidermoid NCI-H292 cells exposed to bioaerosols from a composting plant. Different types of microorganisms were detectable in the composting plant, using the agar culture method. Real-time polymerase chain reaction was used to quantify the level of Aspergillus fumigatus and the profile of remodeling genes. The real-time PCR results indicated that the amount of A. fumigatus in the composting hall was less than 10(2) conidia. The endotoxins in the field bioaerosols were determined using a limulus amebocyte lysate test. The endotoxin levels depended on the type of particulate matter (PM), with coarse particles (2.5-10 μm) having higher endotoxin levels than did fine particles (0.5-2.5 μm). After exposure to the conditioned medium of field bioaerosol samples, NCI-H292 cells showed increased pro-inflammatory interleukin (IL)-6 release and activated epidermal growth factor receptor (EGFR), transforming growth factor (TGF)-β1 and cyclin-dependent kinase inhibitor 1 (p21 WAF1/CIP1) gene expression, but not of matrix metallopeptidase (MMP)-9. Airborne endotoxin levels were higher inside the composting hall than they were in other areas, and they were associated with PM. This suggested that airborne bioaerosols in the composting plant contained endotoxins and microorganisms besides A. fumigatus that cause the inflammatory cytokine secretion and augment the expression of remodeling genes in NCI-H292 cells. It is thus necessary to monitor potentially hazardous materials from bioaerosols in food composting plants, which could affect the health of workers.

Pro-inflammatory effects of metals in persons and animals exposed to tobacco smoke.

PubMed

Milnerowicz, Halina; Ściskalska, Milena; Dul, Magdalena

2015-01-01

Metals present in tobacco smoke have the ability to cause a pro-oxidant/antioxidant imbalance through the direct generation of free radicals in accordance with the Fenton or Haber-Weiss reaction and redox properties. Metals can also interact with antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase) and small molecular antioxidants (glutathione) through binding to SH groups or by replacement of metals ions in the catalytic center of enzymes. Excessive free radicals production can induce an inflammatory response. The aim of this study was to review the information on the induction of inflammation by metals present in tobacco smoke such as lead (Pb), cadmium (Cd), arsenic (As), aluminum (Al), nickel (Ni) and mercury (Hg). In cellular immune response, it was demonstrated that radicals induced by metals can disrupt the transcription signaling pathway mediated by the mitogen-activated protein kinase (induced by Pb), NLRP3-ASC-caspase 1 (induced by Ni), tyrosine kinase Src (induced by As) and the nuclear factor κB (induced by Pb, Ni, Hg). The result of this is a gene transcription for early inflammatory cytokines, such as Interleukine 1β, Interleukine 6, and Tumor necrosis factor α). These cytokines can cause leukocytes recruitment and secretions of other pro-inflammatory cytokines and chemokines, which intensifies the inflammatory response. Some metals, such as cadmium (Cd), can activate an inflammatory response through tissue damage induction mediated by free radicals, which also results in leukocytes recruitment and cytokines secretions. Inflammation generated by metals can be reduced by metallothionein, which has the ability to scavenge free radicals and bind toxic metals through the release of Zn and oxidation of SH groups. Copyright © 2014 Elsevier GmbH. All rights reserved.

Impact of Sub-Inhibitory Concentrations of Amoxicillin on Streptococcus suis Capsule Gene Expression and Inflammatory Potential.

PubMed

Haas, Bruno; Grenier, Daniel

2016-04-19

Streptococcus suis is an important swine pathogen and emerging zoonotic agent worldwide causing meningitis, endocarditis, arthritis and septicemia. Among the 29 serotypes identified to date, serotype 2 is mostly isolated from diseased pigs. Although several virulence mechanisms have been characterized in S. suis, the pathogenesis of S. suis infections remains only partially understood. This study focuses on the response of S. suis P1/7 to sub-inhibitory concentrations of amoxicillin. First, capsule expression was monitored by qRT-PCR when S. suis was cultivated in the presence of amoxicillin. Then, the pro-inflammatory potential of S. suis P1/7 culture supernatants or whole cells conditioned with amoxicillin was evaluated by monitoring the activation of the NF-κB pathway in monocytes and quantifying pro-inflammatory cytokines secreted by macrophages. It was found that amoxicillin decreased capsule expression in S. suis. Moreover, conditioning the bacterium with sub-inhibitory concentrations of amoxicillin caused an increased activation of the NF-κB pathway in monocytes following exposure to bacterial culture supernatants and to a lesser extent to whole bacterial cells. This was associated with an increased secretion of pro-inflammatory cytokines (CXCL8, IL-6, IL-1β) by macrophages. This study identified a new mechanism by which S. suis may increase its inflammatory potential in the presence of sub-inhibitory concentrations of amoxicillin, a cell wall-active antibiotic, thus challenging its use for preventive treatments or as growth factor.

Sequestosome-1 (SQSTM1) sequence variants in ALS cases in the UK: prevalence and coexistence of SQSTM1 mutations in ALS kindred with PDB.

PubMed

Kwok, Chun T; Morris, Alex; de Belleroche, Jacqueline S

2014-04-01

Mutations in the SQSTM1 gene have been reported to be associated with amyotrophic lateral sclerosis (ALS). We sought to determine the frequency of these mutations in a UK familial ALS (FALS) cohort. Sequences of all eight exons of the SQSTM1 gene were analysed in index cases from 61 different FALS kindred lacking known FALS mutations. Six exonic variants c.463G>A, p.(Glu155Lys), c.822G>C, p.(Glu274Asp), c.888G>T, p.(=), c.954C>T, p.(=), c.1038G>A, p.(=) and c.1175C>T, p.(Pro392Leu) were identified in five FALS index cases, three of which were non-synonymous and three were synonymous. One index case harboured three variants (c.822G>C, c.888G>T and c.954C>T), and a second index case harboured two variants (c.822G>C and c.954C>T). Only the p.(Pro392Leu) and p.(Glu155Lys) mutations were predicted to be pathogenic. In one p.(Pro392Leu) kindred, the carrier developed both ALS and Paget's disease of bone (PDB), and, in the p.(Glu155Lys) kindred, the father of the proband developed PDB. All p.(Pro392Leu) carriers were heterozygous for a previously reported founder haplotype for PDB, where this mutation has an established causal effect. The frequency of the p.(Pro392Leu) mutation in this UK FALS cohort was 2.3% and 0.97% overall including three previously screened FALS cohorts. Our results confirm the presence of the p.(Pro392Leu) SQSTM1 mutation in FALS. This mutation is the most common SQSTM1 mutation found in ALS to date, and a likely pathogenicity is supported by having an established causal role in PDB. The occurrence of the same mutation in ALS and PDB is indicative of a common pathogenic pathway that converges on protein homeostasis.

Elevated Plasma C-Terminal Endothelin-1 Precursor Fragment Concentrations Are Associated with Less Anxiety in Patients with Cardiovascular Risk Factors. Results from the Observational DIAST-CHF Study.

PubMed

Meyer, Thomas; Chavanon, Mira-Lynn; Herrrmann-Lingen, Christoph; Roggenthien, Maren; Nolte, Kathleen; Pieske, Burkert; Wachter, Rolf; Edelmann, Frank

2015-01-01

The role of endothelin-1 (ET-1) in the neurobiology of anxiety is unknown, therefore, we assessed in the observational multicenter DIAST-CHF study whether the C-terminal ET-1 precursor fragment (CT-proET-1) is linked to anxiety. Plasma concentrations of CT-proET-1 were measured in a total of 1,410 patients presenting with cardiovascular risk factors (mean age 66.91±8.2 years, 49.3% males, mean left ventricular ejection fraction 60.0±8.2%) who had completed the Hospital Anxiety and Depression Scale (HADS) questionnaire. Among the total study cohort (n = 1,410), there were 118 subjects (8.4%) with an HADS anxiety score above the cut-off level of 11 suggestive of clinically relevant anxiety. Plasma CT-proET-1 levels were significantly lower in the group of anxious patients as compared to non-anxious patients (p = 0.013). In regression models adjusted for sex, age, systolic blood pressure, and diameters of left atrium and ventricle, plasma CT-proET-1 was again linked to anxiety (Exp(β) = 0.247, 95%-confidence interval [95%-CI] = 0.067-0.914, p = 0.036). Given the high prevalence of depressive disorders in anxious patients, we additionally included the HADS depression score as an independent variable in the models and found that CT-proET-1 remained a significant predictor of anxiety, independent of comorbid depression (Exp(β) = 0.114, 95%-CI = 0.023-0.566, p = 0.008). Our data from a population-based study in outpatients with cardiovascular risk factors revealed that circulating CT-proET-1 levels are negatively associated with anxiety. Further investigations are required to clarify the putative anxiolytic effect of ET-1 or its precursor molecules in humans and to decipher its mechanistic pathways.

Nuclear translocation of Nrf2 and expression of antioxidant defence genes in THP-1 cells exposed to carbon nanotubes.

PubMed

Brown, David M; Donaldson, Kenneth; Stone, Vicki

2010-06-01

Carbon nanotubes have a wide range of applications in various industries and their use is likely to rise in the future. Currently, a major concern is that with the increasing use and production of these materials, there may be increased health risks to exposed workers. Long (> 15 microm) straight nanotubes may undergo frustrated phagocytosis which is likely to result in reduced clearance. We examine here the effects of multiwalled carbon nanotubes of different sizes on monocytic THP-1 cells, with regard to their ability to stimulate increased expression of the HO-1 and GST genes and their ability to produce nuclear translocation of the transcription factor, Nrf2, as well as the release of several pro-inflammatory cytokines and mediators of inflammation. Our results suggest that long (50 microm) carbon nanotubes (62.5 microg/ml for 4 hours) produce increased nuclear translocation of Nrf2 and increased HO-1 gene expression compared with shorter entangled nanotubes. There was no increased gene expression for GST. The long nanotubes (NT1) caused increased release of the proinflammatory cytokine IL-1beta, an effect which was diminished by the antioxidant trolox, suggesting a role of oxidative stress in the upregulation of this cytokine. Tentatively, our study suggests that long carbon nanotubes may exert their effect in THP-1 cells in part via an oxidative stress mechanism.

Suppressors of systemin signaling identify genes in the tomato wound response pathway.

PubMed Central

Howe, G A; Ryan, C A

1999-01-01

In tomato plants, systemic induction of defense genes in response to herbivory or mechanical wounding is regulated by an 18-amino-acid peptide signal called systemin. Transgenic plants that overexpress prosystemin, the systemin precursor, from a 35S::prosystemin (35S::prosys) transgene exhibit constitutive expression of wound-inducible defense proteins including proteinase inhibitors and polyphenol oxidase. To study further the role of (pro)systemin in the wound response pathway, we isolated and characterized mutations that suppress 35S::prosys-mediated phenotypes. Ten recessive, extragenic suppressors were identified. Two of these define new alleles of def-1, a previously identified mutation that blocks both wound- and systemin-induced gene expression and renders plants susceptible to herbivory. The remaining mutants defined four loci designated Spr-1, Spr-2, Spr-3, and Spr-4 (for Suppressed in 35S::prosystemin-mediated responses). spr-3 and spr-4 mutants were not significantly affected in their response to either systemin or mechanical wounding. In contrast, spr-1 and spr-2 plants lacked systemic wound responses and were insensitive to systemin. These results confirm the function of (pro)systemin in the transduction of systemic wound signals and further establish that wounding, systemin, and 35S::prosys induce defensive gene expression through a common signaling pathway defined by at least three genes (Def-1, Spr-1, and Spr-2). PMID:10545469

Differential expression of IGFBPs in Laron syndrome-derived lymphoblastoid cell lines: Potential correlation with reduced cancer incidence.

PubMed

Somri, Lina; Sarfstein, Rive; Lapkina-Gendler, Lena; Nagaraj, Karthik; Laron, Zvi; Bach, Leon A; Werner, Haim

2018-04-01

Laron syndrome (LS), or primary growth hormone (GH) insensitivity, is a growth disorder that results from mutation of the GH-receptor (GHR) gene leading to congenital insulin-like growth factor-1 (IGF-1) deficiency. Recent epidemiological studies have shown that LS patients are protected from cancer development. Genome-wide profiling identified genes and signaling pathways that are differentially represented in LS patients, and that may contribute to cancer protection. The present study was aimed at evaluating the hypothesis that IGF binding proteins (IGFBPs) are differentially expressed in LS, most probably as a result of low circulating levels of IGF-1. Furthermore, we postulated that IGFBPs might be differentially regulated by oxidative stress in this condition and, therefore, may contribute to cancer evasion. Our results show that IGFBP-3, which is predominantly protective, was highly expressed in LS-derived lymphoblastoid cells in comparison to control cells from the same ethnic group. On the other hand, levels of IGFBP-2, -4, -5, and -6 were diminished in LS patients, as demonstrated by RQ-PCR, Western immunoblots and confocal immunofluorescence. In addition, our data provide evidence for a pattern of IGFBP response to H 2 O 2 treatment that might be associated with distinct expression of apoptosis markers (BCL2, pro-caspase-9, pro-caspase-3) in LS. In summary, differential expression of specific IGFBPs in LS might be correlated with cellular mechanisms underlying cancer protection and, probably, additional phenotypes due to congenital IGF-1 deficiency. Copyright © 2017 Elsevier Ltd. All rights reserved.

Niclosamide, an anti-helminthic molecule, downregulates the retroviral oncoprotein Tax and pro-survival Bcl-2 proteins in HTLV-1-transformed T lymphocytes.

PubMed

Xiang, Di; Yuan, Yunsheng; Chen, Li; Liu, Xin; Belani, Chandra; Cheng, Hua

2015-08-14

Adult T cell leukemia and lymphoma (ATL) is a highly aggressive form of hematological malignancy and is caused by chronic infection of human T cell leukemia virus type 1 (HTLV-1). The viral genome encodes an oncogenic protein, Tax, which plays a key role in transactivating viral gene transcription and in deregulating cellular oncogenic signaling to promote survival, proliferation and transformation of virally infected T cells. Hence, Tax is a desirable therapeutic target, particularly at early stage of HTLV-1-mediated oncogenesis. We here show that niclosamide, an anti-helminthic molecule, induced apoptosis of HTLV-1-transformed T cells. Niclosamide facilitated degradation of the Tax protein in proteasome. Consistent with niclosamide-mediated Tax degradation, this compound inhibited activities of MAPK/ERK1/2 and IκB kinases. In addition, niclosamide downregulated Stat3 and pro-survival Bcl-2 family members such as Mcl-1 and repressed the viral gene transcription of HTLV-1 through induction of Tax degradation. Since Tax, Stat3 and Mcl-1 are crucial molecules for promoting survival and growth of HTLV-1-transformed T cells, our findings demonstrate a novel mechanism of niclosamide in inducing Tax degradation and downregulating various cellular pro-survival molecules, thereby promoting apoptosis of HTLV-1-associated leukemia cells. Copyright © 2015 Elsevier Inc. All rights reserved.

Functional anatomy of complement factor H.

PubMed

Makou, Elisavet; Herbert, Andrew P; Barlow, Paul N

2013-06-11

Factor H (FH) is a soluble regulator of the proteolytic cascade at the core of the evolutionarily ancient vertebrate complement system. Although FH consists of a single chain of similar protein modules, it has a demanding job description. Its chief role is to prevent complement-mediated injury to healthy host cells and tissues. This entails recognition of molecular patterns on host surfaces combined with control of one of nature's most dangerous examples of a positive-feedback loop. In this way, FH modulates, where and when needed, an amplification process that otherwise exponentially escalates the production of the pro-inflammatory, pro-phagocytic, and pro-cytolytic cleavage products of complement proteins C3 and C5. Mutations and single-nucleotide polymorphisms in the FH gene and autoantibodies against FH predispose individuals to diseases, including age-related macular degeneration, dense-deposit disease, and atypical hemolytic uremic syndrome. Moreover, deletions or variations of genes for FH-related proteins also influence the risk of disease. Numerous pathogens hijack FH and use it for self-defense. As reviewed herein, a molecular understanding of FH function is emerging. While its functional oligomeric status remains uncertain, progress has been achieved in characterizing its three-dimensional architecture and, to a lesser extent, its intermodular flexibility. Models are proposed, based on the reconciliation of older data with a wealth of recent evidence, in which a latent circulating form of FH is activated by its principal target, C3b tethered to a self-surface. Such models suggest hypotheses linking sequence variations to pathophysiology, but improved, more quantitative, functional assays and rigorous data analysis are required to test these ideas.

Proteomic analysis of mouse thymoma EL4 cells treated with bis(tri-n-butyltin)oxide (TBTO).

PubMed

Osman, Ahmed M; van Kol, Sandra; Peijnenburg, Ad; Blokland, Marco; Pennings, Jeroen L A; Kleinjans, Jos C S; van Loveren, Henk

2009-09-01

Here, we report the results of proteomic analysis of the mouse thymoma EL4 cell line exposed to bis(tri-n-butylin)oxide (TBTO), an immunotoxic organotin compound. The objective of the work was to examine whether TBTO affects the expression of proteins in this cell line and to compare the differentially expressed proteins with the corresponding mRNA expression data. The identified proteins were quantified using a label-free quantitative method based on counting the observed peptides as an index of protein abundance. The calculation of the ratio of peptides obtained from exposed and control samples allowed us to evaluate the effect of TBTO on protein expression and to compare these results to those obtained in gene expression profiling studies. Correlation of some of the differentially expressed proteins and their corresponding mRNAs was observed. The analysis of the protein ratios revealed that 12 proteins were significantly affected. These proteins included cytoskeleton proteins myosin-9, spectrin beta 2 and plectin 8. The first two proteins were down-regulated 3-fold, whereas the third was up-regulated 2-fold. Ras-related Rab1, a GTP binding protein and T-complex protein-1 subunit alpha, a chaperonin, were decreased 2- and 3.6-fold, respectively. The ribosomal S10 and eukaryotic translation factor (eIf4G1), which are involved in protein synthesis, were down-regulated 2.6- and 3.7-fold, respectively. Also, proteins involved in splicing of pre-mRNA and in transcription, splicing factor arginine/serine-rich 2 and chromodomain-helicase-DNA binding protein 4 (Chd4), were decreased 2.6- and 4.5 times, respectively. Nuclear RNA helicase II was reduced 2.8-fold. Finally, prothymosin-alpha (ProTalpha), an essential protein for cell proliferation, and a protein similar to ProTalpha, (with a molecular weight and a pI (3.54) comparable to that of ProTalpha) were also down-regulated 6-and 8-fold, respectively. We propose that the observed down-regulation of the expression level of ProTalpha in the TBTO-exposed cells could account for the previously reported anti-proliferative effect of TBTO.

Peroxisome proliferator-activated receptor-α agonist, Wy 14,643, improves metabolic indices, steatosis and ballooning in diabetic mice with non-alcoholic steatohepatitis.

PubMed

Larter, Claire Z; Yeh, Matthew M; Van Rooyen, Derrick M; Brooling, John; Ghatora, Kamaljit; Farrell, Geoffrey C

2012-02-01

Lipid accumulation precedes hepatocellular injury and liver inflammation in non-alcoholic steatohepatitis (NASH). The peroxisome proliferator-activated receptor (PPAR)α regulates hepatic lipid disposal. We studied whether pharmacological stimulation of PPARα reverses NASH associated with metabolic syndrome in high-fat (HF)-fed foz/foz obese/diabetic mice. Female foz/foz mice and wildtype (WT) littermates were fed HF diet for 16 weeks to initiate NASH then treated with Wy 14,643 (Wy) for 10 days or 20 days. Liver disease was assessed by histology, serum alanine aminotransferase, genes (real-time polymerase chain reaction) and proteins (Western blot, enzyme-linked immunosorbent assay) of interest and pro-inflammatory signaling pathways were determined. In diabetic foz/foz mice, NASH was associated with elevated serum MCP1 and hepatic activation of nuclear factor (NF)-κB and c-Jun N-terminal kinase, but not oxidative or endoplasmic reticulum stress. Wy treatment decreased steatosis and injury, although induction of PPARα-responsive fatty acid oxidation genes was proportionally less than in WT. The PPARα agonist lowered serum insulin, corrected hyperglycemia, and suppressed the carbohydrate-dependent lipogenic transcription factor, carbohydrate response element binding protein. Steatosis resolution was associated with suppression of NF-κB and JNK activation and decreased hepatic macrophages and neutrophils. Despite this, histology inflammation score remained high, associated with serum monocyte chemoattractant protein (MCP)1 elevation, a pro-inflammatory chemokine related to higher adipose, not liver MCP1 mRNA expression. Pharmacological activation of PPARα improves metabolic milieu, steatosis, ballooning, and combats NF-κB and JNK activation, neutrophil and F4/80 macrophage recruitment in diabetes-related NASH. However, persistent liver inflammation with high serum MCP1 due to unsuppressed adipose inflammation may limit PPARα agonists' efficacy as therapy for NASH. © 2011 Journal of Gastroenterology and Hepatology Foundation and Blackwell Publishing Asia Pty Ltd.

Uropathogenic E. coli Induce Different Immune Response in Testicular and Peritoneal Macrophages: Implications for Testicular Immune Privilege

PubMed Central

Bhushan, Sudhanshu; Hossain, Hamid; Lu, Yongning; Geisler, Andreas; Tchatalbachev, Svetlin; Mikulski, Zbigniew; Schuler, Gerhard; Klug, Jörg; Pilatz, Adrian; Wagenlehner, Florian; Chakraborty, Trinad; Meinhardt, Andreas

2011-01-01

Infertility affects one in seven couples and ascending bacterial infections of the male genitourinary tract by Escherichia coli are an important cause of male factor infertility. Thus understanding mechanisms by which immunocompetent cells such as testicular macrophages (TM) respond to infection and how bacterial pathogens manipulate defense pathways is of importance. Whole genome expression profiling of TM and peritoneal macrophages (PM) infected with uropathogenic E. coli (UPEC) revealed major differences in regulated genes. However, a multitude of genes implicated in calcium signaling pathways was a common feature which indicated a role of calcium-dependent nuclear factor of activated T cells (NFAT) signaling. UPEC-dependent NFAT activation was confirmed in both cultured TM and in TM in an in vivo UPEC infectious rat orchitis model. Elevated expression of NFATC2-regulated anti-inflammatory cytokines was found in TM (IL-4, IL-13) and PM (IL-3, IL-4, IL-13). NFATC2 is activated by rapid influx of calcium, an activity delineated to the pore forming toxin alpha-hemolysin by bacterial mutant analysis. Alpha-hemolysin suppressed IL-6 and TNF-α cytokine release from PM and caused differential activation of MAP kinase and AP-1 signaling pathways in TM and PM leading to reciprocal expression of key pro-inflammatory cytokines in PM (IL-1α, IL-1β, IL-6 downregulated) and TM (IL-1β, IL-6 upregulated). In addition, unlike PM, LPS-treated TM were refractory to NFκB activation shown by the absence of degradation of IκBα and lack of pro-inflammatory cytokine secretion (IL-6, TNF-α). Taken together, these results suggest a mechanism to the conundrum by which TM initiate immune responses to bacteria, while maintaining testicular immune privilege with its ability to tolerate neo-autoantigens expressed on developing spermatogenic cells. PMID:22164293

Coordinated Gene Expression of Neuroinflammatory and Cell Signaling Markers in Dorsolateral Prefrontal Cortex during Human Brain Development and Aging

PubMed Central

Primiani, Christopher T.; Ryan, Veronica H.; Rao, Jagadeesh S.; Cam, Margaret C.; Ahn, Kwangmi; Modi, Hiren R.; Rapoport, Stanley I.

2014-01-01

Background Age changes in expression of inflammatory, synaptic, and neurotrophic genes are not well characterized during human brain development and senescence. Knowing these changes may elucidate structural, metabolic, and functional brain processes over the lifespan, as well vulnerability to neurodevelopmental or neurodegenerative diseases. Hypothesis Expression levels of inflammatory, synaptic, and neurotrophic genes in the human brain are coordinated over the lifespan and underlie changes in phenotypic networks or cascades. Methods We used a large-scale microarray dataset from human prefrontal cortex, BrainCloud, to quantify age changes over the lifespan, divided into Development (0 to 21 years, 87 brains) and Aging (22 to 78 years, 144 brains) intervals, in transcription levels of 39 genes. Results Gene expression levels followed different trajectories over the lifespan. Many changes were intercorrelated within three similar groups or clusters of genes during both Development and Aging, despite different roles of the gene products in the two intervals. During Development, changes were related to reported neuronal loss, dendritic growth and pruning, and microglial events; TLR4, IL1R1, NFKB1, MOBP, PLA2G4A, and PTGS2 expression increased in the first years of life, while expression of synaptic genes GAP43 and DBN1 decreased, before reaching plateaus. During Aging, expression was upregulated for potentially pro-inflammatory genes such as NFKB1, TRAF6, TLR4, IL1R1, TSPO, and GFAP, but downregulated for neurotrophic and synaptic integrity genes such as BDNF, NGF, PDGFA, SYN, and DBN1. Conclusions Coordinated changes in gene transcription cascades underlie changes in synaptic, neurotrophic, and inflammatory phenotypic networks during brain Development and Aging. Early postnatal expression changes relate to neuronal, glial, and myelin growth and synaptic pruning events, while late Aging is associated with pro-inflammatory and synaptic loss changes. Thus, comparable transcriptional regulatory networks that operate throughout the lifespan underlie different phenotypic processes during Aging compared to Development. PMID:25329999

Coordinated gene expression of neuroinflammatory and cell signaling markers in dorsolateral prefrontal cortex during human brain development and aging.

PubMed

Primiani, Christopher T; Ryan, Veronica H; Rao, Jagadeesh S; Cam, Margaret C; Ahn, Kwangmi; Modi, Hiren R; Rapoport, Stanley I

2014-01-01

Age changes in expression of inflammatory, synaptic, and neurotrophic genes are not well characterized during human brain development and senescence. Knowing these changes may elucidate structural, metabolic, and functional brain processes over the lifespan, as well vulnerability to neurodevelopmental or neurodegenerative diseases. Expression levels of inflammatory, synaptic, and neurotrophic genes in the human brain are coordinated over the lifespan and underlie changes in phenotypic networks or cascades. We used a large-scale microarray dataset from human prefrontal cortex, BrainCloud, to quantify age changes over the lifespan, divided into Development (0 to 21 years, 87 brains) and Aging (22 to 78 years, 144 brains) intervals, in transcription levels of 39 genes. Gene expression levels followed different trajectories over the lifespan. Many changes were intercorrelated within three similar groups or clusters of genes during both Development and Aging, despite different roles of the gene products in the two intervals. During Development, changes were related to reported neuronal loss, dendritic growth and pruning, and microglial events; TLR4, IL1R1, NFKB1, MOBP, PLA2G4A, and PTGS2 expression increased in the first years of life, while expression of synaptic genes GAP43 and DBN1 decreased, before reaching plateaus. During Aging, expression was upregulated for potentially pro-inflammatory genes such as NFKB1, TRAF6, TLR4, IL1R1, TSPO, and GFAP, but downregulated for neurotrophic and synaptic integrity genes such as BDNF, NGF, PDGFA, SYN, and DBN1. Coordinated changes in gene transcription cascades underlie changes in synaptic, neurotrophic, and inflammatory phenotypic networks during brain Development and Aging. Early postnatal expression changes relate to neuronal, glial, and myelin growth and synaptic pruning events, while late Aging is associated with pro-inflammatory and synaptic loss changes. Thus, comparable transcriptional regulatory networks that operate throughout the lifespan underlie different phenotypic processes during Aging compared to Development.

Transcriptional profiling of human bronchial epithelial cell BEAS-2B exposed to diesel and biomass ultrafine particles.

PubMed

Grilli, Andrea; Bengalli, Rossella; Longhin, Eleonora; Capasso, Laura; Proverbio, Maria Carla; Forcato, Mattia; Bicciato, Silvio; Gualtieri, Maurizio; Battaglia, Cristina; Camatini, Marina

2018-04-27

Emissions from diesel vehicles and biomass burning are the principal sources of primary ultrafine particles (UFP). The exposure to UFP has been associated to cardiovascular and pulmonary diseases, including lung cancer. Although many aspects of the toxicology of ambient particulate matter (PM) have been unraveled, the molecular mechanisms activated in human cells by the exposure to UFP are still poorly understood. Here, we present an RNA-seq time-course experiment (five time point after single dose exposure) used to investigate the differential and temporal changes induced in the gene expression of human bronchial epithelial cells (BEAS-2B) by the exposure to UFP generated from diesel and biomass combustion. A combination of different bioinformatics tools (EdgeR, next-maSigPro and reactome FI app-Cytoscape and prioritization strategies) facilitated the analyses the temporal transcriptional pattern, functional gene set enrichment and gene networks related to cellular response to UFP particles. The bioinformatics analysis of transcriptional data reveals that the two different UFP induce, since the earliest time points, different transcriptional dynamics resulting in the activation of specific genes. The functional enrichment of differentially expressed genes indicates that the exposure to diesel UFP induces the activation of genes involved in TNFα signaling via NF-kB and inflammatory response, and hypoxia. Conversely, the exposure to ultrafine particles from biomass determines less distinct modifications of the gene expression profiles. Diesel UFP exposure induces the secretion of biomarkers associated to inflammation (CCXL2, EPGN, GREM1, IL1A, IL1B, IL6, IL24, EREG, VEGF) and transcription factors (as NFE2L2, MAFF, HES1, FOSL1, TGIF1) relevant for cardiovascular and lung disease. By means of network reconstruction, four genes (STAT3, HIF1a, NFKB1, KRAS) have emerged as major regulators of transcriptional response of bronchial epithelial cells exposed to diesel exhaust. Overall, this work highlights modifications of the transcriptional landscape in human bronchial cells exposed to UFP and sheds new lights on possible mechanisms by means of which UFP acts as a carcinogen and harmful factor for human health.

Association of the Genetic Polymorphisms in Transcription Factor 7-Like 2 and Peroxisome Proliferator-Activated Receptors-γ2 with Type 2 Diabetes Mellitus and Its Interaction with Obesity Status in Emirati Population

PubMed Central

Al-Safar, Habiba; Hassoun, Ahmed; Almazrouei, Shaikha; Kamal, Wala; Afandi, Bachar; Rais, Naushad

2015-01-01

Background. Transcription factor 7-like 2 gene (TCF7L2) and peroxisome proliferator-activated receptors-γ2 (PPAR-γ2) have a profound effect on the incidence of type 2 diabetes mellitus (T2DM) and had previously been found to be associated with T2DM risk in various ppopulations. However, studies in the Arab population are inconsistent. We conducted a case control study to confirm the association of variants rs10885409 of TCF7L2 and Pro12Ala (rs1801282) of PPAR-γ2 with risk of T2DM and related complications in Emirati population of Arab origin. We also investigated the interaction of these associations with obesity status. Methods. DNA was extracted from the saliva samples of 272 T2DM patients and 216 nondiabetic Emiratis. Genotyping for rs10885409 (TCF7L2) and rs1801282 (PPAR-γ2 P12A) variants was accomplished with a TaqMan assay. The subgroups were constituted according to obesity status. Results. In the nonobese group, the rs10885409 C allele in the recessive model was significantly associated with the incidence of T2DM (OR 1.975 [95% CI 1.127–3.461], P = 0.017), but this association was not observed in the obese group or when BMI was not considered. PPAR-γ2 risk allele Pro12 frequency (0.96) was similar in the groups tested and more than 90% population was homozygous for this allele. Conclusions. Our case-control study is the first of its kind in Emiratis which establishes TCF7L2 rs10885409 C allele as a T2DM risk factor in Emiratis and this association is modulated by obesity status. We also confirmed that Pro12Ala mutation in PPAR-γ2 is not associated with T2DM risk in this population. PMID:26273662