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Sample records for cia2 coordinately up-regulates

  1. Coordinately up-regulated genes in ovarian cancer.

    PubMed

    Hough, C D; Cho, K R; Zonderman, A B; Schwartz, D R; Morin, P J

    2001-05-15

    A better understanding of the molecular circuitry in normal ovarian tissues and in ovarian cancer will likely provide new targets for diagnosis and therapy. Recently, much has been learned about the genes expressed in ovarian cancer through studies with cDNA arrays and serial analysis of gene expression. However, these methods do not allow highly quantitative analysis of gene expression on a large number of specimens. Here, we have used quantitative real-time RT-PCR in a panel of 39 microdissected ovarian carcinomas of various subtypes to systematically analyze the expression of 13 genes, many of which were previously identified as up-regulated in a subset of ovarian cancers by serial analyses of gene expression. The genes analyzed are glutathione peroxidase 3 (GPX3), apolipoprotein J/clusterin, insulin-like growth factor-binding protein 2, epithelial cell adhesion molecule/GA733-2, Kop protease inhibitor, matrix gla protein, tissue inhibitor of metalloproteinase 3, folate receptor 1, S100A2, signal transducer and activator of transcription 1, secretory leukocyte protease inhibitor, apolipoprotein E, and ceruloplasmin. All of the genes were found overexpressed, some at extremely high levels, in the vast majority of ovarian carcinomas irrespective of the subtype. Interestingly, GPX3 was found at much higher levels in tumors with clear cell histology and may represent a biomarker for this subtype. Some of the genes studied here may thus represent targets for early detection ovarian cancer. The gene expression patterns were not associated with age at diagnosis, stage, or K-ras mutation status in ovarian cancer. We find that several genes are coordinately regulated in ovarian cancer, likely representing the fact that many genes are activated as part of common signaling pathways or that extensive cross-talk exists between several pathways in ovarian cancer. A statistical analysis shows that genes commonly up-regulated in ovarian cancer may result from the aberrant

  2. Coordinate up-regulation of low-density lipoprotein receptor and cyclo-oxygenase-2 gene expression in human colorectal cells and in colorectal adenocarcinoma biopsies

    NASA Technical Reports Server (NTRS)

    Lum, D. F.; McQuaid, K. R.; Gilbertson, V. L.; Hughes-Fulford, M.

    1999-01-01

    Many colorectal cancers have high levels of cyclo-oxygenase 2 (COX-2), an enzyme that metabolizes the essential fatty acids into prostaglandins. Since the low-density lipoprotein receptor (LDLr) is involved in the uptake of essential fatty acids, we studied the effect of LDL on growth and gene regulation in colorectal cancer cells. DiFi cells grown in lipoprotein-deficient sera (LPDS) grew more slowly than cells with LDL. LDLr antibody caused significant inhibition of tumor cell growth but did not affect controls. In addition, LDL uptake did not change in the presence of excess LDL, suggesting that ldlr mRNA lacks normal feedback regulation in some colorectal cancers. Analysis of the ldlr mRNA showed that excess LDL in the medium did not cause down-regulation of the message even after 24 hr. The second portion of the study examined the mRNA expression of ldlr and its co-regulation with cox-2 in normal and tumor specimens from patients with colorectal adenocarcinomas. The ratio of tumor:paired normal mucosa of mRNA expression of ldlr and of cox-2 was measured in specimens taken during colonoscopy. ldlr and cox-2 transcripts were apparent in 11 of 11 carcinomas. There was significant coordinate up-regulation both of ldlr and of cox-2 in 6 of 11 (55%) tumors compared with normal colonic mucosa. There was no up-regulation of cox-2 without concomitant up-regulation of ldlr. These data suggest that the LDLr is abnormally regulated in some colorectal tumors and may play a role in the up-regulation of cox-2. Copyright 1999 Wiley-Liss, Inc.

  3. Ephrin-A1 Is Up-Regulated by Hypoxia in Cancer Cells and Promotes Angiogenesis of HUVECs through a Coordinated Cross-Talk with eNOS

    PubMed Central

    Song, Kai; Shang, Zheng-Jun

    2013-01-01

    Hypoxia, ephrin-A1 and endothelial nitric oxide synthase (eNOS) have been proved to play critical roles in tumor angiogenesis. However, how ephrin-A1 is regulated by hypoxia and whether ephrin-A1 cooperates with eNOS in modulation of angiogenesis remain to be addressed in details. Here we demonstrated that both ephrin-A1 in squamous cell carcinoma cells (SCC-9) and especially soluble ephrin-A1 in the supernatants were up-regulated under hypoxic condition. An increased nitric oxide (NO) production in human umbilical vein endothelial cells (HUVECs) was observed in ephrin-A1-induced angiogenesis which was reversed after co-culture with eNOS specific inhibitor, N-nitro-L-arginine methyl ester hydrochloride (L-NAME). Western blot analysis confirmed that both phosphorylation of AktSer473 and eNOSSer1177 were up-regulated in ephrin-A1-stimulated HUVECs, with the total eNOS expression unchanged. The specific inhibitor of phosphatidylinositol 3-kinase (PI3K), LY294002, significantly down-regulated ephrin-A1-induced expression of phosphorylated AktSer473 as well as phosphorylation of eNOSSer1177. These results revealed a possible novel mechanism whereby ephrin-A1 is regulated in tumor microenvironment and promotes angiogenesis through a coordinated cross-talk with PI3K/Akt-dependent eNOS activation which may relate to normal vascular development and tumor neovascularization. PMID:24040255

  4. Coordinated action of IgE and a B-cell-stimulatory factor on the CD23 receptor molecule up-regulates B-lymphocyte growth.

    PubMed Central

    Guy, G R; Gordon, J

    1987-01-01

    The CD23 (BLAST-2) antigen, recently identified as the low-affinity IgE receptor of B lymphocytes, has also been implicated as the focus for growth-promoting signals delivered to activated B cells by a low molecular weight B-cell growth factor (BCGF). Here we show that IgE and BCGF can coordinate B-lymphocyte growth through their opposing effects on the CD23 molecule. While the activation of purified quiescent B cells with phorbol 12-myristate 13-acetate led to the induction of 45-kDa CD23 at the surface membrane, the inclusion of IgE increased CD23 expression by a factor of approximately equal to 5. The addition of BCGF resulted in the rapid release of a 35-kDa form of CD23 from the cell surface. This shed molecule is associated with autocrine growth factor activity. Substantially more of this material was generated by BCGF acting on cells that had been stimulated in the presence of IgE. The combined effects of IgE and BCGF on DNA synthesis in activated B cells were more than additive. IgE similarly augmented the stimulatory capacity of a CD23 antibody that mimics the biological actions of BCGF. Binding of the anti-receptor antibody to its 45-kDa target at the B-cell surface also prompted the release of the 35-kDa soluble species. These results demonstrate a pleiotropy in the CD23 molecule with regard to both ligand binding and the subsequent behavior of the receptor. The ability of this single receptor to orchestrate a B-lymphocyte response through a variety of ligands and its role in normal and transformed autocrine growth are discussed. Images PMID:2957693

  5. Sucrose prevents up-regulation of senescence-associated genes in carnation petals.

    PubMed

    Hoeberichts, Frank A; van Doorn, Wouter G; Vorst, Oscar; Hall, Robert D; van Wordragen, Monique F

    2007-01-01

    cDNA microarrays were used to characterize senescence-associated gene expression in petals of cut carnation (Dianthus caryophyllus) flowers, sampled from anthesis to the first senescence symptoms. The population of PCR fragments spotted on these microarrays was enriched for flower-specific and senescence-specific genes, using subtractive hybridization. About 90% of the transcripts showed a large increase in quantity, approximately 25% transiently, and about 65% throughout the 7 d experiment. Treatment with silver thiosulphate (STS), which blocks the ethylene receptor and prevented the normal senescence symptoms, prevented the up-regulation of almost all of these genes. Sucrose treatment also considerably delayed visible senescence. Its effect on gene expression was very similar to that of STS, suggesting that soluble sugars act as a repressor of ethylene signal transduction. Two fragments that encoded a carnation EIN3-like (EIL) protein were isolated, some of which are key transcription factors that control ethylene response genes. One of these (Dc-EIL3) was up-regulated during senescence. Its up-regulation was delayed by STS and prevented by sucrose. Sucrose, therefore, seems to repress ethylene signalling, in part, by preventing up-regulation of Dc-EIL3. Some other transcription factors displayed an early increase in transcript abundance: a MYB-like DNA binding protein, a MYC protein, a MADS-box factor, and a zinc finger protein. Genes suggesting a role in senescence of hormones other than ethylene encoded an Aux/IAA protein, which regulate transcription of auxin-induced genes, and a cytokinin oxidase/dehydrogenase, which degrades cytokinin. Taken together, the results suggest a master switch during senescence, controlling the co-ordinated up-regulation of numerous ethylene response genes. Dc-EIL3 might be (part of) this master switch.

  6. Nutraceutical up-regulation of serotonin paradoxically induces compulsive behavior

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The role of diet in either the etiology or treatment of complex mental disorder is highly controversial in psychiatry. However, physiological mechanisms by which diet can influence brain chemistry – particularly that of serotonin – are well established. Here we show that dietary up-regulation of br...

  7. FRZB up-regulated in hepatocellular carcinoma bone metastasis

    PubMed Central

    Huang, Jia; Hu, Wenhao; Lin, Xiangjin; Wang, Xuanwei; Jin, Ketao

    2015-01-01

    The clinical relevance of frizzled-related protein (FRZB) in hepatocellular carcinoma (HCC) bone metastasis remains uncertain. The aim of this study was to assess the clinical relationship of FRZB in patients with HCC bone metastasis after surgical resection. FRZB expression was evaluated by immunohistochemistry in formalin-fixed paraffin embedded (FFPE) HCC and paired bone metastasis tissues from 13 patients that underwent surgical resection. The clinical characteristics of 13 HCC patients with synchronous or metachronous bone metastasis received surgery were retrospectively reviewed. We found that FRZB was positive in 9 HCC tissues (69.2%) and in 11 paired bone metastatic tissues (84.6%) among these 13 paired samples. The expression of FRZB in the bone metastases was noticeably higher than that in the paired HCC tissues. The expression of FRZB was up-regulated in 10 (76.9%) paired bone metastases tissues. FRZB expression was up-regulated in HCC bone metastasis tissue, which suggested that FRZB might play a key role in the HCC bone metastasis. PMID:26722540

  8. A plant gene up-regulated at rust infection sites.

    PubMed

    Ayliffe, Michael A; Roberts, James K; Mitchell, Heidi J; Zhang, Ren; Lawrence, Gregory J; Ellis, Jeffrey G; Pryor, Tony J

    2002-05-01

    Expression of the fis1 gene from flax (Linum usitatissimum) is induced by a compatible rust (Melampsora lini) infection. Infection of transgenic plants containing a beta-glucuronidase (GUS) reporter gene under the control of the fis1 promoter showed that induction is highly localized to those leaf mesophyll cells within and immediately surrounding rust infection sites. The level of induction reflects the extent of fungal growth. In a strong resistance reaction, such as the hypersensitive fleck mediated by the L6 resistance gene, there is very little fungal growth and a microscopic level of GUS expression. Partially resistant flax leaves show levels of GUS expression that were intermediate to the level observed in the fully susceptible infection. Sequence and deletion analysis using both transient Agrobacterium tumefaciens expression and stable transformation assays have shown that the rust-inducible fis1 promoter is contained within a 580-bp fragment. Homologs of fis1 were identified in expressed sequence tag databases of a range of plant species including dicots, monocots, and a gymnosperm. Homologous genes isolated from maize (Zea mays; mis1), barley (Hordeum vulgare; bis1), wheat (Triticum aestivum; wis1), and Arabidopsis encode proteins that are highly similar (76%-82%) to the FIS1 protein. The Arabidopsis homologue has been reported to encode a delta1-pyrroline-5-carboxylate dehydrogenase that is involved in the catabolism of proline to glutamate. RNA-blot analysis showed that mis1 in maize and the bis1 homolog in barley are both up-regulated by a compatible infection with the corresponding species-specific rust. The rust-induced genes homologous to fis1 are present in many plants. The promoters of these genes have potential roles for the engineering of synthetic rust resistance genes by targeting transgene expression to the sites of rust infection.

  9. Mycoplasma arthritidis mitogen up-regulates human NK cell activity.

    PubMed Central

    D'Orazio, J A; Cole, B C; Stein-Streilein, J

    1996-01-01

    While the effects of superantigens on T lymphocytes are well characterized, how superantigens interact with other immune cells is less clear. This report examines the effects of Mycoplasma arthritidis mitogen (MAM) on human natural killer (NK) cell activity. Incubation of peripheral blood mononuclear cells (PBMC) with MAM for 16 to 20 h augmented NK cytotoxicity (against K562) in a dose-dependent manner (P < or = 0.05). Superantigen-dependent cellular cytotoxicity, an activity of superantigen-activated cytotoxic T cells, was not involved in lysis of K562 cells because the erythroleukemic tumor target cells expressed no class II major histocompatibility complex by fluorescence-activated cell sorter analysis. Kinetic experiments showed that the largest increase in NK activity induced by MAM occurred within 48 h. Incubation with MAM caused a portion of NK cells to become adherent to tissue culture flasks, a quality associated with activation, and augmented NK activity was found in both adherent and nonadherent subpopulations. Experiments using cytokine-specific neutralizing antibodies showed that interleukin-2 contributed to enhancement of the NK activity observed in superantigen-stimulated PBMC. Interestingly, MAM was able to augment NK lysis of highly purified NK (CD56+) cells in the absence of other immune cells in 9 of 12 blood specimens, with the augmented lytic activity ranging from 110 to 170% of unstimulated NK activity. In summary, data presented in this report show for the first time that MAM affects human NK cells directly by increasing their lytic capacity and indirectly in PBMC as a consequence of cytokines produced by T cells. Results of this work suggest that, in vivo, one consequence of interaction with superantigen-secreting microorganisms may be up-regulation of NK lytic activity. These findings may have clinical application as a means of generating augmented NK effector cells useful in the immunotherapy of parasitic infections or neoplasms. PMID

  10. Functional inactivation of CXC chemokine receptor 4-mediated responses through SOCS3 up-regulation.

    PubMed

    Soriano, Silvia F; Hernanz-Falcón, Patricia; Rodríguez-Frade, José Miguel; De Ana, Ana Martín; Garzón, Ruth; Carvalho-Pinto, Carla; Vila-Coro, Antonio J; Zaballos, Angel; Balomenos, Dimitrios; Martínez-A, Carlos; Mellado, Mario

    2002-08-01

    Hematopoietic cell growth, differentiation, and chemotactic responses require coordinated action between cytokines and chemokines. Cytokines promote receptor oligomerization, followed by Janus kinase (JAK) kinase activation, signal transducers and transactivators of transcription (STAT) nuclear translocation, and transcription of cytokine-responsive genes. These include genes that encode a family of negative regulators of cytokine signaling, the suppressors of cytokine signaling (SOCS) proteins. After binding their specific receptors, chemokines trigger receptor dimerization and activate the JAK/STAT pathway. We show that SOCS3 overexpression or up-regulation, stimulated by a cytokine such as growth hormone, impairs the response to CXCL12, measured by Ca(2+) flux and chemotaxis in vitro and in vivo. This effect is mediated by SOCS3 binding to the CXC chemokine receptor 4 receptor, blocking JAK/STAT and Galpha(i) pathways, without interfering with cell surface chemokine receptor expression. The data provide clear evidence for signaling cross-talk between cytokine and chemokine responses in building a functional immune system.

  11. Dysfunctional chloroplasts up-regulate the expression of mitochondrial genes in Arabidopsis seedlings.

    PubMed

    Liao, Jo-Chien; Hsieh, Wei-Yu; Tseng, Ching-Chih; Hsieh, Ming-Hsiun

    2016-02-01

    Chloroplasts and mitochondria play important roles in maintaining metabolic and energy homeostasis in the plant cell. The interactions between these two organelles, especially photosynthesis and respiration, have been intensively studied. Still, little is known about the regulation of mitochondrial gene expression by chloroplasts and vice versa. The gene expression machineries in chloroplasts and mitochondria rely heavily on the nuclear genome. Thus, the interactions between nucleus and these organelles, including anterograde and retrograde regulation, have been actively investigated in the last two decades. Norflurazon (NF) and lincomycin (Lin) are two commonly used inhibitors to study chloroplast-to-nucleus retrograde signaling in plants. We used NF and Lin to block the development and functions of chloroplasts and examined their effects on mitochondrial gene expression, RNA editing and splicing. The editing of most mitochondrial transcripts was not affected, but the editing extents of nad4-107, nad6-103, and ccmFc-1172 decreased slightly in NF- and Lin-treated seedlings. While the splicing of mitochondrial transcripts was not significantly affected, steady-state mRNA levels of several mitochondrial genes increased significantly in NF- and Lin-treated seedlings. Moreover, Lin seemed to have more profound effects than NF on the expression of mitochondrial genes, indicating that signals derived from these two inhibitors might be distinct. NF and Lin also significantly induced the expression of nuclear genes encoding subunits of mitochondrial electron transport chain complexes. Thus, dysfunctional chloroplasts may coordinately up-regulate the expression of nuclear and mitochondrial genes encoding subunits of respiratory complexes.

  12. Parallel up-regulation of the profilin gene family following independent domestication of diploid and allopolyploid cotton (Gossypium).

    PubMed

    Bao, Ying; Hu, Guanjing; Flagel, Lex E; Salmon, Armel; Bezanilla, Magdalena; Paterson, Andrew H; Wang, Zining; Wendel, Jonathan F

    2011-12-27

    Cotton is remarkable among our major crops in that four species were independently domesticated, two allopolyploids and two diploids. In each case thousands of years of human selection transformed sparsely flowering, perennial shrubs into highly productive crops with seeds bearing the vastly elongated and abundant single-celled hairs that comprise modern cotton fiber. The genetic underpinnings of these transformations are largely unknown, but comparative gene expression profiling experiments have demonstrated up-regulation of profilin accompanying domestication in all three species for which wild forms are known. Profilins are actin monomer binding proteins that are important in cytoskeletal dynamics and in cotton fiber elongation. We show that Gossypium diploids contain six profilin genes (GPRF1-GPRF6), located on four different chromosomes (eight chromosomes in the allopolyploid). All but one profilin (GPRF6) are expressed during cotton fiber development, and both homeologs of GPRF1-GPRF5 are expressed in fibers of the allopolyploids. Remarkably, quantitative RT-PCR and RNAseq data demonstrate that GPRF1-GPRF5 are all up-regulated, in parallel, in the three independently domesticated cottons in comparison with their wild counterparts. This result was additionally supported by iTRAQ proteomic data. In the allopolyploids, there This usage of novel should be fine, since it refers to a novel evolutionary process, not a novel discovery has been novel recruitment of the sixth profilin gene (GPRF6) as a result of domestication. This parallel up-regulation of an entire gene family in multiple species in response to strong directional selection is without precedent and suggests unwitting selection on one or more upstream transcription factors or other proteins that coordinately exercise control over profilin expression. PMID:22160709

  13. Up-Regulation of Antioxidant Proteins in the Plasma Proteome during Saturation Diving: Unique Coincidence under Hypobaric Hypoxia

    PubMed Central

    Domoto, Hideharu; Iwaya, Keiichi; Ikomi, Fumitaka; Matsuo, Hirotaka; Tadano, Yutaka; Fujii, Shigenori; Tachi, Kazuyoshi; Itoh, Yoshiyuki; Sato, Michiya; Inoue, Kimitoshi; Shinomiya, Nariyoshi

    2016-01-01

    Saturation diving (SD) is one of the safest techniques for tolerating hyperbaric conditions for long durations. However, the changes in the human plasma protein profile that occur during SD are unknown. To identify differential protein expression during or after SD, 65 blood samples from 15 healthy Japanese men trained in SD were analyzed by two-dimensional fluorescence difference gel electrophoresis. The expression of two proteins, one 32.4 kDa with an isoelectric point (pI) of 5.8 and the other 44.8 kDa with pI 4.0, were elevated during SD to 60, 100, and 200 meters sea water (msw). The expression of these proteins returned to pre-diving level when the SD training was completed. The two proteins were identified using in-gel digestion and mass spectrometric analysis; the 32.4 kDa protein was transthyretin and the 44.8 kDa protein was alpha-1-acid glycoprotein 1. Oxidation was detected at methionine 13 of transthyretin and at methionine 129 of alpha-1-acid glycoprotein 1 by tandem mass spectrometry. Moreover, haptoglobin was up-regulated during the decompression phase of 200 msw. These plasma proteins up-regulated during SD have a common function as anti-oxidants. This suggests that by coordinating their biological effects, these proteins activate a defense mechanism to counteract the effects of hyperbaric-hyperoxic conditions during SD. PMID:27741252

  14. Up-regulation in the expression of renin gene by the influence of aluminium.

    PubMed

    Ezomo, Ojeiru F; Matsushima, Fumiko; Meshitsuka, Shunsuke

    2009-11-01

    The excretion of aluminium in urine was significantly increased after intake of analgesics containing aluminium, confirming increased absorption and hence exposure to aluminium with such medication. The effect of aluminium on the kidney was further investigated by study of gene expression in mice. After a single dose of aluminium, an up-regulation of renin gene was found by DNA sequencing of the products of differential display analysis. The up-regulation of renin was confirmed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting experiments in the dose dependent treatments and the time course observation after aluminium citrate injection. The up-regulation of the renin expression by aluminium is a strong indication of the influence of aluminium on the renin-angiotensin-aldosterone-system, resulting in possible induction of essential hypertension.

  15. Up-regulation of emotional responses to reward-predicting stimuli: an ERP study.

    PubMed

    Langeslag, Sandra J E; van Strien, Jan W

    2013-09-01

    Altered reward processing is a hallmark symptom of many psychiatric disorders. It has recently been shown that people are capable of down-regulating reward processing. Here, we examined whether people are capable of up-regulating emotional responses to reward-predicting stimuli. Participants passively viewed colored squares that predicted a reward or no reward, and up- or down-regulated their emotional responses to these reward-predicting stimuli by focusing on the reward meaning or the color of the squares respectively. The amplitude of the late positive potential (LPP) was taken as an objective index of regulation success. The LPP in response to reward-predicting squares was enhanced by up-regulation, suggesting that up-regulation of emotional responses to reward-predicting stimuli using a cognitive strategy is feasible. These results are highly relevant for the treatment of disorders characterized by diminished motivation, and for reward-based decision making in daily life. PMID:23770414

  16. Mu opioid receptor up-regulation and participation in excitability of hippocampal pyramidal cell electrophysiology

    SciTech Connect

    Moudy, A.M.

    1988-01-01

    Chronic administration of opiate antagonists to rats results in up-regulation of their brain opioid receptors. Using subcellular fractionation techniques, brain opioid receptors were resolved into two membrane populations, one associated with synaptic plasma membranes (SPM) and the other enriched in smooth endoplasmic reticulum and Golgi (microsomes). This study addressed in part the question of whether an antagonist induces up-regulation uniformly in these two populations. Rats were administered naltrexone by subcutaneously implanted osmotic minipumps. Forebrain mu receptor levels were determined by homologous displacement of ({sup 3}H)D-ala{sup 2}-mePhe{sup 4}-gly-ol{sup 5}-enkephalin (DAGO) followed by computer estimation of binding parameters. Receptor levels in crude membranes rose 77% after treatment. Microsomes displayed a 92% increase, a two-fold greater change than in SPMs (51%). These results establish that naltrexone induces up-regulation of both membrane populations; and that microsomal and SPM receptors represent discrete populations of intracellular and cell surface sites, respectively. Binding experiments on isolated hippocampi also demonstrated up-regulation (71%) of mu receptors. To demonstrate up-regulation of opioid receptors electrophysiologically, hippocampal slices were prepared from rats which had been chronically treated with naltrexone. After superfusion with DAGO, these slices showed a 42% greater population spike output than controls in response to the same EPSP input. Hippocampi from animals treated for two weeks showed an additional increase in sensitivity. The results support a disinhibitory role for opioids in pyramidal cell hyper-excitability. More importantly, they demonstrate a significant physiological correlate to opioid receptor up-regulation.

  17. Neuropilin 1 Receptor Is Up-Regulated in Dysplastic Epithelium and Oral Squamous Cell Carcinoma.

    PubMed

    Shahrabi-Farahani, Shokoufeh; Gallottini, Marina; Martins, Fabiana; Li, Erik; Mudge, Dayna R; Nakayama, Hironao; Hida, Kyoko; Panigrahy, Dipak; D'Amore, Patricia A; Bielenberg, Diane R

    2016-04-01

    Neuropilins are receptors for disparate ligands, including proangiogenic factors such as vascular endothelial growth factor and inhibitory class 3 semaphorin (SEMA3) family members. Differentiated cells in skin epithelium and cutaneous squamous cell carcinoma highly express the neuropilin-1 (NRP1) receptor. We examined the expression of NRP1 in human and mouse oral mucosa. NRP1 was significantly up-regulated in oral epithelial dysplasia and oral squamous cell carcinoma (OSCC). NRP1 receptor localized to the outer suprabasal epithelial layers in normal tongue, an expression pattern similar to the normal skin epidermis. However, dysplastic tongue epithelium and OSCC up-regulated NRP1 in basal and proliferating epithelial layers, a profile unseen in cutaneous squamous cell carcinoma. NRP1 up-regulation is observed in a mouse carcinogen-induced OSCC model and in human tongue OSCC biopsies. Human OSCC cell lines express NRP1 protein in vitro and in mouse tongue xenografts. Sites of capillary infiltration into orthotopic OSCC tumors correlate with high NRP1 expression. HSC3 xenografts, which express the highest NRP1 levels of the cell lines examined, showed massive intratumoral lymphangiogenesis. SEMA3A inhibited OSCC cell migration, suggesting that the NRP1 receptor was bioactive in OSCC. In conclusion, NRP1 is regulated in the oral epithelium and is selectively up-regulated during epithelial dysplasia. NRP1 may function as a reservoir to sequester proangiogenic ligands within the neoplastic compartment, thereby recruiting neovessels toward tumor cells. PMID:26877262

  18. Executive functions and the down-regulation and up-regulation of emotion

    PubMed Central

    Gyurak, Anett; Goodkind, Madeleine S.; Kramer, Joel H.; Miller, Bruce L.; Levenson, Robert W.

    2011-01-01

    This study examined the relationship between individual differences in executive functions (EF; assessed by measures of working memory, Stroop, trail making, and verbal fluency) and ability to down-regulate and up-regulate responses to emotionally evocative film clips. To ensure a wide range of EF, 48 participants with diverse neurodegenerative disorders and 21 older neurologically normal aging participants were included. Participants were exposed to three different movie clips that were designed to elicit a mix of disgust and amusement. While watching the films they were either instructed to watch, down-regulate, and up-regulate their visible emotional responses. Heart-rate and facial behaviors were monitored throughout. Emotion regulatory ability was operationalized as changes in heart-rate and facial behavior in the down- and up-regulation conditions, controlling for responses in the watch condition. Results indicated that higher verbal fluency scores were related to greater ability to regulate emotion in both the down-regulation and up-regulation conditions. This finding remained significant even after controlling for age and general cognitive functioning. No relationships were found between emotion regulation and the other EF measures. We believe these results derive from differences among EF measures, with verbal fluency performance best capturing the complex sequence of controlled planning, activation, and monitoring required for successful emotion regulation. These findings contribute to our understanding of emotion-cognition interaction, suggesting a link between emotion-regulatory abilities and individual differences in complex executive functions. PMID:21432634

  19. Impaired up-regulation of type II corticosteroid receptors in hippocampus of aged rats.

    PubMed

    Eldridge, J C; Fleenor, D G; Kerr, D S; Landfield, P W

    1989-01-30

    Several recent investigations have reported a decline of rat hippocampal corticosteroid-binding receptors (CSRs) with aging. This decline has been proposed to be an initial cause (through disinhibition) of the elevated adrenal steroid secretion that apparently occurs with aging; however, it could instead be an effect of corticoid elevation (through down-regulation). In order to assess the effects of age on CSR biosynthetic capacity in the absence of down-regulatory influences of endogenous corticoids, as well as to study aging changes in CSR plasticity, we examined the up-regulation of hippocampal CSR that follows adrenalectomy (ADX). The rat hippocampus contains at least two types of CSR binding and differential analysis of types I and II CSR was accomplished by selective displacement of [3H]corticosterone with RU-28362, a specific type II agonist. In young (3 months old) Fischer-344 rat hippocampus, up-regulation of type II binding above 2-day ADX baseline was present by 3-7 days and increased still further by 8-10 days post-ADX; type I CSR density did not change significantly between 1 and 10 days post-ADX. However, in aged (24-26 months old) rats, type II CSR up-regulation did not occur over the 10 day post-ADX period. Thus, the age-related impairment of type II up-regulation may reflect an intrinsic deficit in CSR biosynthesis or lability that is independent of the acute endogenous adrenal steroid environment.

  20. Cotton Benzoquinone Reductase: Up-regulation During Early Cotton Fiber Developement

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Benzoquinone reductase (BR; EC 1.6.5.7) is an enzyme that catalyzes the bivalent redox reactions of quinones without the production of free radical intermediates. Using 2-D PAGE comparisons, two proteins were found to be up-regulated in wild-type cotton ovules during the fiber initiation stage but ...

  1. CD84 is markedly up-regulated in Kawasaki disease arteriopathy

    PubMed Central

    Reindel, R; Bischof, J; Kim, K-Y A; Orenstein, J M; Soares, M B; Baker, S C; Shulman, S T; Perlman, E J; Lingen, M W; Pink, A J; Trevenen, C; Rowley, A H

    2014-01-01

    The major goals of Kawasaki disease (KD) therapy are to reduce inflammation and prevent thrombosis in the coronary arteries (CA), but some children do not respond to currently available non-specific therapies. New treatments have been difficult to develop because the molecular pathogenesis is unknown. In order to identify dysregulated gene expression in KD CA, we performed high-throughput RNA sequencing on KD and control CA, validated potentially dysregulated genes by real-time reverse transcription–polymerase chain reaction (RT–PCR) and localized protein expression by immunohistochemistry. Signalling lymphocyte activation molecule CD84 was up-regulated 16-fold (P < 0·01) in acute KD CA (within 2 months of onset) and 32-fold (P < 0·01) in chronic CA (5 months to years after onset). CD84 was localized to inflammatory cells in KD tissues. Genes associated with cellular proliferation, motility and survival were also up-regulated in KD CA, and immune activation molecules MX2 and SP140 were up-regulated in chronic KD. CD84, which facilitates immune responses and stabilizes platelet aggregates, is markedly up-regulated in KD CA in patients with acute and chronic arterial disease. We provide the first molecular evidence of dysregulated inflammatory responses persisting for months to years in CA significantly damaged by KD. PMID:24635044

  2. Leptin up-regulates the lactogenic effect of prolactin in the bovine mammary gland in vitro.

    PubMed

    Feuermann, Y; Shamay, A; Mabjeesh, S J

    2008-11-01

    The ability of leptin to up-regulate prolactin action in the mammary gland is well established. We examined the effect of leptin and prolactin on traits associated with lactation. Leptin and prolactin enhanced proliferation (thymidine incorporation) of the mammary gland cells, elevated the cells' proliferation in a dose-responsive manner, and synergized to elevate the expression of amino acid metabolism via a 90% increase in aminopeptidase N expression. Leptin and prolactin decreased apoptosis (decreased caspase-3 expression by 60%) in the same manner. Leptin enhanced the effect of prolactin on all of these processes in bovine mammary explants. Leptin and prolactin regulated mTOR (mammalian target of rapamycin) by increasing expression by 66%, which is one of the signal-transduction junctions involved in the regulation of proliferation, apoptosis, and protein synthesis. These findings support the hypothesis that leptin up-regulates prolactin action in the bovine mammary gland. PMID:18946122

  3. Up-regulation of cyclooxygenase-2 by product-prostaglandin E2

    NASA Technical Reports Server (NTRS)

    Tjandrawinata, R. R.; Hughes-Fulford, M.

    1997-01-01

    The development of prostate cancer has been linked to high level of dietary fat intake. Our laboratory investigates the connection between cancer cell growth and fatty acid products. Studying human prostatic carcinoma PC-3 cells, we found that prostaglandin E2 (PGE2) increased cell growth and up-regulated the gene expression of its own synthesizing enzyme, cyclooxygenase-2 (COX-2). PGE2 increased COX-2 mRNA expression dose-dependently with the highest levels of stimulation seen at the 3-hour period following PGE2 addition. The NSAID flurbiprofen (5 microM), in the presence of exogenous PGE2, inhibited the up-regulation of COX-2 mRNA and cell growth. These data suggest that the levels of local intracellular PGE2 play a major role in the growth of prostate cancer cells through an activation of COX-2 gene expression.

  4. Eurycomanone induce apoptosis in HepG2 cells via up-regulation of p53

    PubMed Central

    Zakaria, Yusmazura; Rahmat, Asmah; Pihie, Azimahtol Hawariah Lope; Abdullah, Noor Rain; Houghton, Peter J

    2009-01-01

    Background Eurycomanone is a cytotoxic compound found in Eurycoma longifolia Jack. Previous studies had noted the cytotoxic effect against various cancer cell lines. The aim of this study is to investigate the cytotoxicity against human hepato carcinoma cell in vitro and the mode of action. The cytotoxicity of eurycomanone was evaluated using MTT assay and the mode of cell death was detected by Hoechst 33258 nuclear staining and flow cytometry with Annexin-V/propidium iodide double staining. The protein expression Bax, Bcl-2, p53 and cytochrome C were studied by flow cytometry using a spesific antibody conjugated fluorescent dye to confirm the up-regulation of p53 and Bax in cancer cells. Results The findings suggested that eurycomanone was cytotoxic on cancerous liver cell, HepG2 and less toxic on normal cells Chang's liver and WLR-68. Furthermore, various methods proved that apoptosis was the mode of death in eurycomanone-treated HepG2 cells. The characteristics of apoptosis including chromatin condensation, DNA fragmentation and apoptotic bodies were found following eurycomanone treatment. This study also found that apoptotic process triggered by eurycomanone involved the up-regulation of p53 tumor suppressor protein. The up-regulation of p53 was followed by the increasing of pro-apoptotic Bax and decreasing of anti-apoptotic Bcl-2. The increased of cytochrome C levels in cytosol also results in induction of apoptosis. Conclusion The data suggest that eurycomanone was cytotoxic on HepG2 cells by inducing apoptosis through the up-regulation of p53 and Bax, and down-regulation of Bcl-2. PMID:19508737

  5. High Glucose Up-regulates ADAM17 through HIF-1α in Mesangial Cells.

    PubMed

    Li, Renzhong; Uttarwar, Lalita; Gao, Bo; Charbonneau, Martine; Shi, Yixuan; Chan, John S D; Dubois, Claire M; Krepinsky, Joan C

    2015-08-28

    We previously showed that ADAM17 mediates high glucose-induced matrix production by kidney mesangial cells. ADAM17 expression is increased in diabetic kidneys, suggesting that its up-regulation may augment high glucose profibrotic responses. We thus studied the effects of high glucose on ADAM17 gene regulation. Primary rat mesangial cells were treated with high glucose (30 mm) or mannitol as osmotic control. High glucose dose-dependently increased ADAM17 promoter activity, transcript, and protein levels. This correlated with augmented ADAM17 activity after 24 h versus 1 h of high glucose. We tested involvement of transcription factors shown in other settings to regulate ADAM17 transcription. Promoter activation was not affected by NF-κB or Sp1 inhibitors, but was blocked by hypoxia-inducible factor-1α (HIF-1α) inhibition or down-regulation. This also prevented ADAM17 transcript and protein increases. HIF-1α activation by high glucose was shown by its increased nuclear translocation and activation of the HIF-responsive hypoxia-response element (HRE)-luciferase reporter construct. Assessment of ADAM17 promoter deletion constructs coupled with mutation analysis and ChIP studies identified HIF-1α binding to its consensus element at -607 as critical for the high glucose response. Finally, inhibitors of epidermal growth factor receptor (EGFR) and downstream PI3K/Akt, or ADAM17 itself, prevented high glucose-induced HIF-1α activation and ADAM17 up-regulation. Thus, high glucose induces ADAM17 transcriptional up-regulation in mesangial cells, which is associated with augmentation of its activity. This is mediated by HIF-1α and requires EGFR/ADAM17 signaling, demonstrating the potentiation by ADAM17 of its own up-regulation. ADAM17 inhibition thus provides a potential novel therapeutic strategy for the treatment of diabetic nephropathy.

  6. Selective CB2 up-regulation in women affected by endometrial inflammation

    PubMed Central

    Iuvone, Teresa; De Filippis, Daniele; Di Spiezio Sardo, Attilio; D'Amico, Alessandra; Simonetti, Sara; Sparice, Stefania; Esposito, Giuseppe; Bifulco, Giuseppe; Insabato, Luigi; Nappi, Carmine; Guida, Maurizio

    2008-01-01

    Abstract Endometritis is defined as an inflammation of the endometrial mucosa of the uterus. In endometritis large amounts of toxic mediators, including nitric oxide (NO) are released by inflammatory cells. As a consequence of nitric oxide-dependent injury, the cells respond by triggering protective mechanisms, by changing the endo-cannabinoid system (ECS) which comprises both CB1 and CB2 cannabinoid receptors and their endogenous ligands. The aim of our study was to seek out evidence for the presence of cannabinoid receptors in inflammatory endometrial tissue as well as for their potential role in endometrial inflammation. Our results showed a selective up-regulation of both transcription and expression of CB2 receptors in biopsies from women affected by endometrial inflammation compared to healthy women. The experiments with the nitric oxide-donor S-Nitroso-L-Glutathione (GSNO) suggest that such a selective up-regulation may be related to the nitric oxide release occurring during endometrial inflammation. In addition, we demonstrated an increase in chymase expression, a marker of mast cells, in biopsies of women affected by endometritis. Therefore our results support the hypothesis that the up-regulation of CB2 occurs mainly on mast cells and that it might tend to sensitize these cells to the anti-inflammatory effect exerted by endogenous cannabinoids by binding their receptor and thus preventing the mast cell degranulation and the release of pro-inflammatory mediators. In conclusion, we believe that the selective CB2 up-regulation might play a role as a novel prognostic factor in endometrial inflammation. PMID:18419603

  7. Rapid systemic up-regulation of genes after heat-wounding and electrical stimulation

    NASA Technical Reports Server (NTRS)

    Davies, E.; Vian, A.; Vian, C.; Stankovic, B.

    1997-01-01

    When one leaf of a tomato plant is electrically-stimulated or heat-wounded, proteinase inhibitor genes are rapidly up-regulated in distant leaves. The identity of the systemic wound signal(s) is not yet known, but major candidates include hormones transmitted via the phloem or the xylem, the electrically-stimulated self-propagating electrical signal in the phloem (the action potential, AP), or the heat-wound-induced surge in hydraulic pressure in the xylem evoking a local change in membrane potential in adjacent living cells (the variation potential, VP). In order to discriminate between these signals we have adopted two approaches. The first approach involves applying stimuli that evoke known signals and determining whether these signals have similar effects on the "model" transcripts for proteinase inhibitors (pin) and calmodulin (cal). Here we show that a heat wound almost invariably evokes a VP, while an electrical stimulation occasionally evokes an AP, and both of these signals induce accumulation of transcripts encoding proteinase inhibitors. The second approach involves identifying the array of genes turned on by heat-wounding. To this end, we have constructed a subtractive library for heat-wounded tissue, isolated over 800 putatively up-regulated clones, and shown that all but two of the fifty that we have analyzed by Northern hybridization are, indeed, up-regulated. Here we show the early kinetics of up-regulation of three of these transcripts in the terminal (4th) leaf in response to heat-wounding the 3rd leaf, about 5 cm away. Even though these transcripts show somewhat different time courses of induction, with one peaking at 30 min, another at 15 min, and another at 5 min after flaming of a distant leaf, they all exhibit a similar pattern, i.e., a transient period of transcript accumulation preceding a period of transcript decrease, followed by a second period of transcript accumulation.

  8. HO-1 up-regulation: a key point in high-risk neuroblastoma resistance to bortezomib.

    PubMed

    Furfaro, Anna Lisa; Piras, Sabrina; Passalacqua, Mario; Domenicotti, Cinzia; Parodi, Alessia; Fenoglio, Daniela; Pronzato, Maria Adelaide; Marinari, Umberto Maria; Moretta, Lorenzo; Traverso, Nicola; Nitti, Mariapaola

    2014-04-01

    High-risk neuroblastoma (NB) is characterized by the development of chemoresistance, and bortezomib (BTZ), a selective inhibitor of proteasome, has been proposed in order to overcome drug resistance. Considering the involvement of the nuclear factor-erythroid-derived 2-like 2 (Nrf2) and heme oxygenase-1 (HO-1) in the antioxidant and detoxifying ability of cancer cells, in this study we have investigated their role in differently aggressive NB cell lines treated with BTZ, focusing on the modulation of HO-1 to improve sensitivity to therapy. We have shown that MYCN amplified HTLA-230 cells were slightly sensitive to BTZ treatment, due to the activation of Nrf2 that led to an impressive up-regulation of HO-1. BTZ-treated HTLA-230 cells down-regulated p53 and up-regulated p21, favoring cell survival. The inhibition of HO-1 activity obtained by Zinc (II) protoprophyrin IX (ZnPPIX) was able to significantly increase the pro-apoptotic effect of BTZ in a p53- and p21-independent way. However, MYCN non-amplified SH-SY5Y cells showed a greater sensitivity to BTZ in relation to their inability to up-regulate HO-1. Therefore, we have shown that HO-1 inhibition improves the sensitivity of aggressive NB to proteasome inhibition-based therapy, suggesting that HO-1 up-regulation can be used as a marker of chemoresistance in NB. These results open up a new scenario in developing a combined therapy to overcome chemoresistance in high-risk neuroblastoma.

  9. Perlecan up-regulation of FRNK suppresses smooth muscle cell proliferation via inhibition of FAK signaling.

    PubMed

    Walker, Heather A; Whitelock, John M; Garl, Pamela J; Nemenoff, Raphael A; Stenmark, Kurt R; Weiser-Evans, Mary C M

    2003-05-01

    We previously reported that fully assembled basement membranes are nonpermissive to smooth muscle cell (SMC) replication and that perlecan (PN), a basement membrane heparan sulfate proteoglycan, is a dominant effector of this response. We report here that SMC adhesion to basement membranes, and perlecan in particular, up-regulate the expression of focal adhesion kinase-related nonkinase (FRNK), a SMC-specific endogenous inhibitor of FAK, which subsequently suppresses FAK-mediated, ERK1/2-dependent growth signals. Up-regulation of FRNK by perlecan is actively and continuously regulated. Relative to the matrix proteins studied, the effects are unique to perlecan, because plating of SMCs on several other basement membrane proteins is associated with low levels of FRNK and corresponding high levels of FAK and ERK1/2 phosphorylation and SMC growth. Perlecan supports SMC adhesion, although there is reduced cell spreading compared with fibronectin (FN), laminin (LN), or collagen type IV (IV). Despite the reduction in cell spreading, we report that perlecan-induced up-regulation of FRNK is independent of cell shape changes. Growth inhibition by perlecan was rescued by overexpressing a constitutively active FAK construct, but overexpressing kinase-inactivated mutant FAK or FRNK attenuated fibronectin-stimulated growth. These data indicate that perlecan functions as an endogenously produced inhibitor of SMC growth at least in part through the active regulation of FRNK expression. FRNK, in turn, may control SMC growth by downregulating FAK-dependent signaling events.

  10. Hyaluronan up-regulation is linked to renal dysfunction and hearing loss induced by silver nanoparticles.

    PubMed

    Feng, Hao; Pyykkö, Ilmari; Zou, Jing

    2015-10-01

    Increased application of silver nanoparticles (AgNPs) has raised concerns on their potential adverse effects on human health. However, the precise toxicological mechanisms are not known in detail. The current study hypothesized that AgNPs induced glycosaminoglycan accumulation in the basement membrane that associated with the up-regulation of its component hyaluronic acid, known as a hydrophilic molecule of binding and retaining water, and caused toxicities in the kidney and cochlea. Rats administered AgNPs through either intravenous or intratympanic injection were observed at different time points after exposure. The concentrations of creatinine and urea in the serum were elevated remarkably, and proteins leaked into the urine were increased. A significant hearing loss over a broad range of frequencies was indicated. AgNP exposure induced glycosaminoglycan accumulation and hyaluronic acid up-regulation in the basement membrane. Abundant apoptotic cell death was demonstrated in the AgNP-exposed organs. Our results suggested that glycosaminoglycan accumulation associated with the up-regulation of hyaluronic acid was involved in the toxicities of kidney and cochlea caused by AgNPs.

  11. Identification and analysis of Phytophthora cactorum genes up-regulated during cyst germination and strawberry infection.

    PubMed

    Chen, Xiaoren; Klemsdal, Sonja Sletner; Brurberg, May Bente

    2011-10-01

    The oomycete Phytophthora cactorum can cause economically important diseases on numerous host plants worldwide, such as crown rot on strawberry. To explore the molecular mechanisms underlying the pathogenicity of P. cactorum on strawberry, transcriptional analysis of P. cactorum during strawberry infection and cyst germination was performed by applying suppression subtractive hybridization (SSH) and effector-specific differential display (ESDD) techniques. Two SSH cDNA libraries were generated, enriched for P. cactorum genes expressed during infection or during cyst germination, respectively, and 137 unique differentially expressed genes were identified. To specifically select RxLR effector genes from P. cactorum, ESDD was performed using RxLR and EER motif-based degenerate primers. Eight RxLR effector candidate genes as well as 67 other genes were identified out of 124 selected fragments. The expression levels of 20 putatively up-regulated genes were further analyzed using real-time RT-PCR, showing that, indeed 19 of these 20 genes were up-regulated during at least one of the studied developmental stages or during strawberry crown invasion, relative to the mycelium. This study provides a first overview of P. cactorum genes that are up-regulated immediately prior to or during strawberry infection and also provides a novel method for selecting RxLR effector genes from the unsequenced genome of P. cactorum.

  12. Placental fractalkine is up-regulated in severe early onset preeclampsia

    PubMed Central

    Siwetz, Monika; Dieber-Rotheneder, Martina; Cervar-Zivkovic, Mila; Kummer, Daniel; Kremshofer, Julia; Weiss, Gregor; Herse, Florian; Huppertz, Berthold; Gauster, Martin

    2015-01-01

    The pathogenesis of preeclampsia includes the release of placental factors into the maternal circulation inducing an inflammatory environment in the mother. One of the factors may be the pro-inflammatory chemokine fractalkine, which is expressed in the syncytiotrophoblast of human placenta, from where it is released into the maternal circulation by constitutive shedding. We examined whether placental fractalkine is up-regulated in severe early onset preeclampsia and whether the pro-inflammatory cytokines tumor necrosis factor (TNF)-α and interleukin-6 are able to increase the expression of fractalkine. Gene expression analysis, ELISA, and immunohistochemistry consistently showed increased fractalkine expression in placentas from severe early onset preeclampsia, compared to gestational age-matched controls. Expression of the metalloproteinases ADAM10 and ADAM17, which convert transmembrane fractalkine into the soluble form, was significantly increased in these cases. Incubation of first trimester placental explants with TNF-α provoked a significant increase in fractalkine expression and release of the soluble form, whereas interleukin-6 had no effect. TNF-α-mediated up-regulation of placental fractalkine was reversed in the presence of the Aspirin-derivative salicylate, which impaired activation of NF-κB p65 in TNF-α-treated explants. Based on data from placental explants we suggest that increased maternal TNF-α may up-regulate the expression and release of placental fractalkine, which in turn may contribute to an exaggerated systemic inflammatory response in preeclampsia. PMID:25769431

  13. A20 is up-regulated in primary mouse hepatocytes subjected to hypoxia and reperfusion.

    PubMed

    Sun, Jiao; Sun, Luning; Zhang, Ning; Lu, Xiaomei; Zhang, Haipeng

    2012-12-01

    Hepatic ischemia reperfusion-induced injury is a major medical concern, and it is important to characterize the adaptive mechanisms of hepatocytes to hypoxia and reoxygenation to sustain liver function. In this study, we reported a proteomic analysis of ischemia reperfusion-induced global responses in primary hepatocytes. The primary hepatocytes were isolated from mice and exposed to oxygen to mimic ischemia reperfusion. Total proteins were extracted from the cells and analyzed by two-dimensional gel electrophoresis followed by matrix-assisted laser desorption time-of-flight mass spectrometry. Zinc finger protein A20, mercaptopyruvate sulfur transferase, apolipoprotein E precursor and carbamoyl-phosphate synthase mitochondrial precursor were identified as differentially expressed in differently exposed groups. Reverse transcriptase polymerase chain reaction and Western blot analysis validated that A20 was significantly up-regulated in the hepatocytes subjected to hypoxia and reperfusion. In addition, the expression of peroxisome proliferator-activated receptor α, an A20 target, was up-regulated in the hepatocytes subjected to hypoxia and reperfusion. Our results on A20 provide new insight into the mechanism underlying the adaptation of hepatocytes to hypoxia and reperfusion. Because of its role in the up-regulation of peroxisome proliferator-activated receptor α expression to protect hepatocytes from reperfusion-induced apoptosis, A20 is a potential target for the prevention and therapy of liver injury after ischemia reperfusion.

  14. Placental fractalkine is up-regulated in severe early-onset preeclampsia.

    PubMed

    Siwetz, Monika; Dieber-Rotheneder, Martina; Cervar-Zivkovic, Mila; Kummer, Daniel; Kremshofer, Julia; Weiss, Gregor; Herse, Florian; Huppertz, Berthold; Gauster, Martin

    2015-05-01

    The pathogenesis of preeclampsia (PE) includes the release of placental factors into the maternal circulation, inducing an inflammatory environment in the mother. One of the factors may be the proinflammatory chemokine fractalkine, which is expressed in the syncytiotrophoblast of human placenta, from where it is released into the maternal circulation by constitutive shedding. We examined whether placental fractalkine is up-regulated in severe early-onset PE and whether the proinflammatory cytokines tumor necrosis factor (TNF)-α and IL-6 are able to increase the expression of fractalkine. Gene expression analysis, enzyme-linked immunosorbent assay, and immunohistochemistry consistently showed increased fractalkine expression in placentas from severe early-onset PE, compared to gestational age-matched controls. Expression of a disintegrin and metalloproteinases (ADAMs) 10 and 17, which convert transmembrane fractalkine into the soluble form, was significantly increased in these cases. Incubation of first-trimester placental explants with TNF-α provoked a significant increase in fractalkine expression and release of the soluble form, whereas IL-6 had no effect. TNF-α-mediated up-regulation of placental fractalkine was reversed in the presence of the aspirin-derivative salicylate, which impaired activation of NF-κB p65 in TNF-α-treated explants. On the basis of data from placental explants, we suggest that increased maternal TNF-α may up-regulate the expression and release of placental fractalkine, which, in turn, may contribute to an exaggerated systemic inflammatory response in PE. PMID:25769431

  15. Up-regulation of miR-98 and unraveling regulatory mechanisms in gestational diabetes mellitus

    PubMed Central

    Cao, Jing-Li; Zhang, Lu; Li, Jian; Tian, Shi; Lv, Xiao-Dan; Wang, Xue-Qin; Su, Xing; Li, Ying; Hu, Yi; Ma, Xu; Xia, Hong-Fei

    2016-01-01

    MiR-98 expression was up-regulated in kidney in response to early diabetic nephropathy in mouse and down-regulated in muscle in type 2 diabetes in human. However, the expression prolife and functional role of miR-98 in human gestational diabetes mellitus (GDM) remained unclear. Here, we investigated its expression and function in placental tissues from GDM patients and the possible molecular mechanisms. The results showed that miR-98 was up-regulated in placentas from GDM patients compared with normal placentas. MiR-98 over-expression increased global DNA methylational level and miR-98 knockdown reduced global DNA methylational level. Further investigation revealed that miR-98 could inhibit Mecp2 expression by binding the 3′-untranslated region (UTR) of methyl CpG binding protein 2 (Mecp2), and then led to the expression dysregulation of canonical transient receptor potential 3 (Trpc3), a glucose uptake related gene. More importantly, in vivo analysis found that the expression level of Mecp2 and Trpc3 in placental tissues from GDM patients, relative to the increase of miR-98, was diminished, especially for GDM patients over the age of 35 years. Collectively, up-regulation of miR-98 in the placental tissues of human GDM is linked to the global DNA methylation via targeting Mecp2, which may imply a novel regulatory mechanism in GDM. PMID:27573367

  16. Protein kinase C mediates up-regulation of tetrodotoxin-resistant, persistent Na+ current in rat and mouse sensory neurones.

    PubMed

    Baker, Mark D

    2005-09-15

    The tetrodotoxin-resistant (TTX-r) persistent Na(+) current, attributed to Na(V)1.9, was recorded in small (< 25 mum apparent diameter) dorsal root ganglion (DRG) neurones cultured from P21 rats and from adult wild-type and Na(V)1.8 null mice. In conventional whole-cell recordings intracellular GTP-gamma-S caused current up-regulation, an effect inhibited by the PKC pseudosubstrate inhibitor, PKC19-36. The current amplitude was also up-regulated by 25 microM intracellular 1-oleoyl-2-acetyl-sn-glycerol (OAG) consistent with PKC involvement. In perforated-patch recordings, phorbol 12-myristate 13-acetate (PMA) up-regulated the current, whereas membrane-permeant activators of protein kinase A (PKA) were without effect. PGE(2) did not acutely up-regulate the current. Conversely, both PGE(2) and PKA activation up-regulated the major TTX-r Na(+) current, Na(V)1.8. Extracellular ATP up-regulated the persistent current with an average apparent K(d) near 13 microM, possibly consistent with P2Y receptor activation. Numerical simulation of the up-regulation qualitatively reproduced changes in sensory neurone firing properties. The activation of PKC appears to be a necessary step in the GTP-dependent up-regulation of persistent Na(+) current. PMID:16002450

  17. N-acetylcysteine inhibits the up-regulation of mitochondrial biogenesis genes in livers from rats fed ethanol chronically

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: Chronic ethanol (EtOH) administration to experimental animals induces hepatic oxidative stress and up-regulates mitochondrial biogenesis. The mechanisms by which chronic EtOH up-regulates mitochondrial biogenesis have not been fully explored. In this work, we hypothesized that oxidative ...

  18. Mutations in BALB Mitochondrial DNA Induce CCL20 Up-regulation Promoting Tumorigenic Phenotypes

    PubMed Central

    Sligh, James; Janda, Jaroslav; Jandova, Jana

    2014-01-01

    mtDNA mutations are common in human cancers and are thought to contribute to the process of neoplasia. We examined the role of mtDNA mutations in skin cancer by generating fibroblast cybrids harboring a mutation in the gene encoding the mitochondrial tRNA for arginine. This somatic mutation (9821insA) was previously reported in UV-induced hyperkeratotic skin tumors in hairless mice and confers specific tumorigenic phenotypes to mutant cybrids. Microarray analysis revealed and RT-PCR along with Western blot analysis confirmed the up-regulation of CCL20 and its receptor CCR6 in mtBALB haplotype containing the mt-Tr 9821insA allele compared to wild type mtB6 haplotype. Based on reported role of CCL20 in cancer progression we examined whether the hyper-proliferation and enhanced motility of mtBALB haplotype would be associated with CCL20 levels. Treatment of both genotypes with recombinant CCL20 (rmCCL20) resulted in enhanced growth and motility of mtB6 cybrids. Furthermore, the acquired somatic alteration increased the in vivo tumor growth of mtBALB cybrids through the up-regulation of CCL20 since neutralizing antibody significantly decreased in vivo tumor growth of these cells; and tumors from anti-CCL20 treated mice injected with mtBALB cybrids showed significantly decreased CCL20 levels. When rmCCL20 or mtBALB cybrids were used as chemotactic stimuli, mtB6 cybrids showed increased motility while anti-CCL20 antibody decreased the migration and in vivo tumor growth of mtBALB cybrids. Moreover, the inhibitors of MAPK signaling and NF-κB activation inhibited CCL20 expression in mtBALB cybrids and decreased their migratory capabilities. Thus, acquired mtDNA mutations may promote tumorigenic phenotypes through up-regulation of chemokine CCL20. PMID:25177208

  19. Nitric oxide up-regulates endothelial expression of angiotensin II type 2 receptors.

    PubMed

    Dao, Vu Thao-Vi; Medini, Sawsan; Bisha, Marion; Balz, Vera; Suvorava, Tatsiana; Bas, Murat; Kojda, Georg

    2016-07-15

    Increasing vascular NO levels following up-regulation of endothelial nitric oxide synthase (eNOS) is considered beneficial in cardiovascular disease. Whether such beneficial effects exerted by increased NO-levels include the vascular renin-angiotensin system remains elucidated. Exposure of endothelial cells originated from porcine aorta, mouse brain and human umbilical veins to different NO-donors showed that expression of the angiotensin-II-type-2-receptor (AT2) mRNA and protein is up-regulated by activation of soluble guanylyl cyclase, protein kinase G and p38 mitogen-activated protein kinase without changing AT2 mRNA stability. In mice, endothelial-specific overexpression of eNOS stimulated, while chronic treatment with the NOS-blocker l-nitroarginine inhibited AT2 expression. The NO-induced AT2 up-regulation was associated with a profound inhibition of angiotensin-converting enzyme (ACE)-activity. In endothelial cells this reduction of ACE-activity was reversed by either the AT2 antagonist PD 123119 or by inhibition of transcription with actinomycin D. Furthermore, in C57Bl/6 mice an acute i.v. bolus of l-nitroarginine did not change AT2-expression and ACE-activity suggesting that inhibition of ACE-activity by endogenous NO is crucially dependent on AT2 protein level. Likewise, three weeks of either voluntary or forced exercise training increased AT2 expression and reduced ACE-activity in C57Bl/6 but not in mice lacking eNOS suggesting significance of this signaling interaction for vascular physiology. Finally, aortic AT2 expression is about 5 times greater in female as compared to male C57Bl/6 and at the same time aortic ACE activity is reduced in females by more than 50%. Together these findings imply that endothelial NO regulates AT2 expression and that AT2 may regulate ACE-activity. PMID:27235748

  20. Urban air pollution produces up-regulation of myocardial inflammatory genes and dark chocolate provides cardioprotection.

    PubMed

    Villarreal-Calderon, Rodolfo; Reed, William; Palacios-Moreno, Juan; Keefe, Sheyla; Herritt, Lou; Brooks, Diane; Torres-Jardón, Ricardo; Calderón-Garcidueñas, Lilian

    2012-05-01

    Air pollution is a serious environmental problem. Elderly subjects show increased cardiac morbidity and mortality associated with air pollution exposure. Mexico City (MC) residents are chronically exposed to high concentrations of fine particulate matter (PM(2.5)) and PM-associated lipopolysaccharides (PM-LPS). To test the hypothesis that chronic exposure to urban pollution produces myocardial inflammation, female Balb-c mice age 4 weeks were exposed for 16 months to two distinctly different polluted areas within MC: southwest (SW) and northwest (NW). SW mice were given either no treatment or chocolate 2g/9.5 mg polyphenols/3 times per week. Results were compared to mice kept in clean air. Key inflammatory mediator genes: cyclooxygenase-2 (COX-2), interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α), and the LPS receptor CD14 (cluster of differentiation antigen 14) were measured by real-time polymerase chain reaction. Also explored were target NFκB (nuclear factor κB), oxidative stress and antioxidant defense genes. TNF-α, IL-6, and COX-2 were significantly increased in both NW and SWMC mice (p=0.0001). CD14 was up-regulated in SW mice in keeping with the high exposures to particulate matter associated endotoxin. Chocolate administration resulted in a significant down-regulation of TNF-α (p<0.0001), IL-6 (p=0.01), and IL-1β (p=0.02). The up-regulation of antioxidant enzymes and the down-regulation of potent oxidases, toll-like receptors, and pro-apoptotic signaling genes completed the protective profile. Exposure to air pollution produces up-regulation of inflammatory myocardial genes and endotoxin plays a key role in the inflammatory response. Regular consumption of dark chocolate may reduce myocardial inflammation and have cardioprotective properties in the setting of air pollution exposures. PMID:20932730

  1. Serum withdrawal up-regulates human SIRT1 gene expression in a p53-dependent manner.

    PubMed

    Shang, Linshan; Zhou, Haibin; Xia, Yu; Wang, Hui; Gao, Guimin; Chen, Bingxi; Liu, Qiji; Shao, Changshun; Gong, Yaoqin

    2009-10-01

    SIRT1, a nicotinamide adenine dinucleotide (NAD(+))-dependent histone/protein deacetylase, has been extensively studied recently for its critical role in the regulation of physiology, calorie restriction and aging. Studies on laboratory mice showed that expression of SIRT1 can be induced by starvation in a p53-dependent manner and requires the p53-binding sites present in the Sirt1 promoter. However, it remains to be determined whether these findings based on rodents apply to human beings. In this paper, we characterized a putative p53-binding element in the human SIRT1 promoter that might be required for the up-regulation of SIRT1 in response to nutritional stress. The p53-binding site in the promoter of human SIRT1 is more deviant from the consensus sequence than the corresponding sequence in the mouse Sirt1. There is a C to A change at the second half site in human SIRT1, thus disrupting the core-binding element CWWG in the canonical RRRCWWGYYY. To test whether such sequence change would affect its binding with p53 and the SIRT1 expression under stress, we studied various human cell lines with different p53 status and cells with ectopic expression of functionally distinct p53. We found that serum withdrawal also up-regulates human SIRT1 gene expression in a p53-dependent manner and that the p53-binding element in SIRT1 is required for the up-regulation. Thus, the mechanism responsible for the regulation of SIRT1 expression by p53 is conserved between mice and human beings.

  2. Up-regulation of nicotinic acetylcholine receptors in menthol cigarette smokers

    PubMed Central

    Brody, Arthur L; Mukhin, Alexey G; La Charite, Jaime; Ta, Karen; Farahi, Judah; Sugar, Catherine A.; Mamoun, Michael S.; Vellios, Evan; Archie, Meena; Kozman, Maggie; Phuong, Jonathan; Arlorio, Franca; Mandelkern, Mark A.

    2013-01-01

    One-third of smokers primarily use menthol cigarettes and usage of these cigarettes leads to elevated serum nicotine levels and more difficulty quitting in standard treatment programmes. Previous brain imaging studies demonstrate that smoking (without regard to cigarette type) leads to up-regulation of β2*-containing nicotinic acetylcholine receptors (nAChRs). We sought to determine if menthol cigarette usage results in greater nAChR up-regulation than non-menthol cigarette usage. Altogether, 114 participants (22 menthol cigarette smokers, 41 non-menthol cigarette smokers and 51 non-smokers) underwent positron emission tomography scanning using the α4β2* nAChR radioligand 2-[18F]fluoro-A-85380 (2-FA). In comparing menthol to non-menthol cigarette smokers, an overall test of 2-FA total volume of distribution values revealed a significant between-group difference, resulting from menthol smokers having 9–28% higher α4β2* nAChR densities than non-menthol smokers across regions. In comparing the entire group of smokers to non-smokers, an overall test revealed a significant between-group difference, resulting from smokers having higher α4β2* nAChR levels in all regions studied (36–42%) other than thalamus (3%). Study results demonstrate that menthol smokers have greater up-regulation of nAChRs than non-menthol smokers. This difference is presumably related to higher nicotine exposure in menthol smokers, although other mechanisms for menthol influencing receptor density are possible. These results provide additional information about the severity of menthol cigarette use and may help explain why these smokers have more trouble quitting in standard treatment programmes. PMID:23171716

  3. Up-regulation of gamma-glutamyl transpeptidase (GGT) activity in growth perturbed C6 astrocytes.

    PubMed

    Mares, V; Malík, R; Lisá, V; Sedo, A

    2005-05-20

    Activity of gamma-glutamyl transpeptidase (GGT) was studied in astrocyte-like C6 glial cells modulated in growth and maturation by different concentration of serum and dibutyryl cyclic AMP (Db-cAMP) supplement in culture medium. After reduction of serum concentration from 10% to 0.1%, the number of GGT positive cells determined histochemically increased 3.1 times and the GGT activity/mg protein in whole cell lysates was 5.1 times higher. In cultures with 0.1% serum + Db-cAMP, the histochemically and biochemically assayed GGT activity exceeded 5.1 and 7.9 times the values measured in control 10% serum cultures, respectively. The up-regulation of GGT was accompanied by an inhibition of proliferation, enhanced differentiation and hypertrophy of cells. In addition, the process of metabolic perturbation and/or cellular stress was revealed in these cultures by the (i) growth-support release followed by shrinkage and death of a small number of cells and (ii) higher oxidation of 2'7'dichlorofluorescein diacetate to its fluorescent form in the adherent/viable cells. The observed up-regulation of GGT is considered to primarily reflect increased metabolism of glutathione and/or the maintenance of the redox potential in cells stressed by sub-optimal concentration of serum and Db-cAMP supplement. The concomitant cellular hypertrophy and differentiation and their relationship to increased activity of GGT await further investigation. The study suggests that up-regulation of GGT can contribute to adaptation of astrocytic cells to metabolic and/or oxidative perturbances occurring under various pathological conditions, including radiation- and drug-induced toxicity. PMID:15893589

  4. Agonist- and antagonist-induced up-regulation of surface 5-HT3A receptors

    PubMed Central

    Morton, Russell A; Baptista-Hon, Daniel T; Hales, Tim G; Lovinger, David M

    2015-01-01

    Background and Purpose The 5-HT3 receptor is a member of the pentameric ligand-gated ion channel family and is pharmacologically targeted to treat irritable bowel syndrome and nausea/emesis. Furthermore, many antidepressants elevate extracellular concentrations of 5-HT. This study investigates the functional consequences of exposure of recombinant 5-HT3A receptors to agonists and antagonists. Experimental Approach We used HEK cells stably expressing recombinant 5-HT3A receptors and the ND7/23 (mouse neuroblastoma/dorsal root ganglion hybrid) cell line, which expresses endogenous 5-HT3 receptors. Surface expression of recombinant 5-HT3A receptors, modified to contain the bungarotoxin (BTX) binding sequence, was quantified using fluorescence microscopy to image BTX-conjugated fluorophores. Whole cell voltage-clamp electrophysiology was used to measure the density of current mediated by 5-HT3A receptors. Key Results 5-HT3A receptors were up-regulated by the prolonged presence of agonists (5-HT and m-chlorophenylbiguanide) and antagonists (MDL-72222 and morphine). The up-regulation of 5-HT3A receptors by 5-HT and MDL-72222 was time- and concentration-dependent but was independent of newly translated receptors. The phenomenon was observed for recombinant rodent and human 5-HT3A receptors and for endogenous 5-HT3 receptors in neuronal ND7/23 cells. Conclusions and Implications Up-regulation of 5-HT3A receptors, following exposure to either agonists or antagonists suggests that this phenomenon may occur in response to different therapeutic agents. Medications that elevate 5-HT levels, such as the antidepressant inhibitors of 5-HT reuptake and antiemetic inhibitors of 5-HT3 receptor function, may both raise receptor expression. However, this will require further investigation in vivo. PMID:25989383

  5. GATA-4 promotes myocardial transdifferentiation of mesenchymal stromal cells via up-regulating IGFBP-4

    PubMed Central

    LI, HONGXIA; ZUO, SHI; PASHA, ZEESHAN; YU, BIN; HE, ZHISONG; WANG, YIGANG; YANG, XIANGJUN; ASHRAF, MUHAMMAD; XU, MEIFENG

    2012-01-01

    Background aims GATA-4 is a cardiac transcription factor and plays an important role in cell lineage differentiation during development. We investigated whether overexpression of GATA-4 increases adult mesenchymal stromal cell (MSC) transdifferentiation into a cardiac phenotype in vitro. Methods MSC were harvested from rat bone marrow (BM) and transduced with GATA-4 (MSCGATA-4) using a murine stem cell virus (pMSCV) retroviral expression system. Gene expression in MSCGATA-4 was analyzed using quantitative reverse transcription–polymerase chain reaction (RT-PCR) and Western blotting. Native cardiomyocytes (CM) were isolated from ventricles of neonatal rats. Myocardial transdifferentiation of MSC was determined by immunostaining and electrophysiologic recording. The transdifferentiation rate was calculated directly from flow cytometery. Results The expression of cardiac genes, including brain natriuretic peptide (BNP), Islet-1 and α-sarcomeric actinin (α-SA), was up-regulated in MSCGATA-4 compared with control cells that were transfected with Green Fluorescent Protein (GFP) only (MSCNull). At the same time, insulin-like growth factor-binding protein (IGFBP)-4 was significantly up-regulated in MSCGATA-4. A synchronous beating of MSC with native CM was detected and an action potential was recorded. Some GFP + cells were positive for α-SA staining after MSC were co-cultured with native CM for 7 days. The transdifferentiation rate was significantly higher in MSCGATA-4. Functional studies indicated that the differentiation potential of MSCGATA-4 was decreased by knockdown of IGFBP-4. Conclusions Overexpression of GATA-4 significantly increases MSC differentiation into a myocardial phenotype, which might be associated with the up-regulation of IGFBP-4. PMID:21846294

  6. Erbb2 up-regulation of ADAM12 expression accelerates skin cancer progression.

    PubMed

    Rao, Velidi H; Vogel, Kristen; Yanagida, Jodi K; Marwaha, Nitin; Kandel, Amrit; Trempus, Carol; Repertinger, Susan K; Hansen, Laura A

    2015-10-01

    Solar ultraviolet (UV) radiation can cause severe damage to the skin and is the primary cause of most skin cancer. UV radiation causes DNA damage leading to mutations and also activates the Erbb2/HER2 receptor through indirect mechanisms involving reactive oxygen species. We hypothesized that Erbb2 activation accelerates the malignant progression of UV-induced skin cancer. Following the induction of benign squamous papillomas by UV exposure of v-ras(Ha) transgenic Tg.AC mice, mice were treated topically with the Erbb2 inhibitor AG825 and tumor progression monitored. AG825 treatment reduced tumor volume, increased tumor regression, and delayed the development of malignant squamous cell carcinoma (SCC). Progression to malignancy was associated with increased Erbb2 and ADAM12 (A Disintegin And Metalloproteinase 12) transcripts and protein, while inhibition of Erbb2 blocked the increase in ADAM12 message upon malignant progression. Similarly, human SCC and SCC cell lines had increased ADAM12 protein and transcripts when compared to normal controls. To determine whether Erbb2 up-regulation of ADAM12 contributed to malignant progression of skin cancer, Erbb2 expression was modulated in cultured SCC cells using forced over-expression or siRNA targeting, demonstrating up-regulation of ADAM12 by Erbb2. Furthermore, ADAM12 transfection or siRNA targeting revealed that ADAM12 increased both the migration and invasion of cutaneous SCC cells. Collectively, these results suggest Erbb2 up-regulation of ADAM12 as a novel mechanism contributing to the malignant progression of UV-induced skin cancer. Inhibition of Erbb2/HER2 reduced tumor burden, increased tumor regression, and delayed the progression of benign skin tumors to malignant SCC in UV-exposed mice. Inhibition of Erbb2 suppressed the increase in metalloproteinase ADAM12 expression in skin tumors, which in turn increased migration and tumor cell invasiveness.

  7. Urban Air Pollution Produces Up-Regulation of Myocardial Inflammatory Genes and Dark Chocolate Provides Cardioprotection

    PubMed Central

    Villarreal-Calderon, Rodolfo; Reed, William; Palacios-Moreno, Juan; Keefe, Sheyla; Herritt, Lou; Brooks, Diane; Torres-Jardón, Ricardo; Calderón-Garcidueñas, Lilian

    2010-01-01

    Air pollution is a serious environmental problem. Elderly subjects show increased cardiac morbidity and mortality associated with air pollution exposure. Mexico City (MC) residents are chronically exposed to high concentrations of fine particulate matter (PM2.5) and PM-associated lipopolysaccharides (PM-LPS). To test the hypothesis that chronic exposure to urban pollution produces myocardial inflammation, female Balb-c mice age 4 weeks were exposed for 16 months to two distinctly different polluted areas within MC: Southwest (SW) and Northwest (NW). SW mice were given either no treatment or chocolate 2g/9.5 mg polyphenols/3 times per week. Results were compared to mice kept in clean air. Key inflammatory mediator genes: cyclooxygenase-2 (COX-2), interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α), and the LPS receptor CD14 (cluster of differentiation antigen 14) were measured by real time polymerase chain reaction. Also explored were target NFκB (Nuclear Factor κ B), oxidative stress and antioxidant defense genes. TNF-α, IL-6, and COX-2 were significantly increased in both NW and SWMC mice (p=0.0001). CD14 was up-regulated in SW mice in keeping with the high exposures to particulate matter associated endotoxin. Chocolate administration resulted in a significant down-regulation of TNF-α (p<0.0001), IL-6 (p=0.01), and IL-1β (p=0.02). The up-regulation of antioxidant enzymes and the down-regulation of potent oxidases, toll-like receptors, and pro-apoptotic signaling genes completed the protective profile. Exposure to air pollution produces up-regulation of inflammatory myocardial genes and endotoxin plays a key role in the inflammatory response. Regular consumption of dark chocolate may reduce myocardial inflammation and have cardioprotective properties in the setting of air pollution exposures. PMID:20932730

  8. The Natural Antimicrobial Enzyme Lysozyme is Up-Regulated in Gastrointestinal Inflammatory Conditions.

    PubMed

    Rubio, Carlos A

    2014-01-01

    The cells that line the mucosa of the human gastrointestinal tract (GI, that is, oral cavity, oesophagus, stomach, small intestine, large intestine, and rectum) are constantly challenged by adverse micro-environmental factors, such as different pH, enzymes, and bacterial flora. With exception of the oral cavity, these microenvironments also contain remnant cocktails of secreted enzymes and bacteria from upper organs along the tract. The density of the GI bacteria varies, from 103/mL near the gastric outlet, to 1010/mL at the ileocecal valve, to 1011 to 1012/mL in the colon. The total microbial population (ca. 1014) exceeds the total number of cells in the tract. It is, therefore, remarkable that despite the prima facie inauspicious mixture of harmful secretions and bacteria, the normal GI mucosa retains a healthy state of cell renewal. To counteract the hostile microenvironment, the GI epithelia react by speeding cell exfoliation (the GI mucosa has a turnover time of two to three days), by increasing peristalsis, by eliminating bacteria through secretion of plasma cell-immunoglobulins and by increasing production of natural antibacterial compounds, such as defensin-5 and lysozyme. Only recently, lysozyme was found up-regulated in Barrett's oesophagitis, chronic gastritis, gluten-induced atrophic duodenitis (coeliac disease), collagenous colitis, lymphocytic colitis, and Crohn's colitis. This up-regulation is a response directed to the special types of bacteria recently detected in these diseases. The aim of lysozyme up-regulation is to protect individual mucosal segments to chronic inflammation. The molecular mechanisms connected to the crosstalk between the intraluminal bacterial flora and the production of lysozyme released by the GI mucosae, are discussed. Bacterial resistance continues to exhaust our supply of commercial antibiotics. The potential use of lysozyme to treat infectious diseases is receiving much attention. PMID:25437608

  9. Up-regulation of the adrenomedullin system mediates hypotension and hypoaldosteronism induced by simulated microgravity.

    PubMed

    Andreis, Paola G; Rossi, Gian Paolo; Bova, Sergio; Neri, Giuliano; Nussdorfer, Gastone G; Mazzocchi, Giuseppina

    2004-04-01

    We recently demonstrated that prolonged simulated microgravity (SMG) induced hypotension and hypoaldosteronism in rats, and gathered preliminary evidence for an involvement of circulating adrenomedullin (AM). Thus, we aimed to investigate whether short-term SMG elicits the same effects, and whether up-regulation of adrenal AM system plays a relevant role. Rats were exposed for 8 days to SMG in the form of hindlimb unweighting, and then, along with control animals, were given an intraperitoneal injection of AM22-52 and/or angiotensin-II (Ang-II) (100 nmoles/kg) or the saline vehicle. Systolic blood pressure (SBP) was measured by tail-cuff sphygmomanometry. The adrenal expression of AM was assayed by semiquantitative RT-PCR. The plasma concentrations of aldosterone (PAC) and AM, and adrenal AM content were measured by RIA. Short-term SMG induced significant decreases in SBP and PAC. Conversely, both the plasma and adrenal levels of AM, and adrenal AM mRNA were enhanced in SMG-exposed animals. The SMG-induced hypotension and hypoaldosteronism were reversed by AM22-52, an AM-receptor antagonist, thereby demonstrating a causal link between these effects and the up-regulation of AM system. SMG hampered SBP and PAC responses to Ang-II; the co-administration of AM22-52 restored these responses. These findings accord well with the known ability of AM to counteract the effects of Ang-II on both blood vessels and adrenocortical cells. Taken together, our findings allow us to conclude that up-regulation of the adrenal AM system i) occurs early and takes part in the adaptative changes occurring during SMG conditions; and ii) may account for both hypotension and hypoaldosteronism on returning to the normogravitational environment.

  10. Nicotine induces chromatin changes and c-Jun up-regulation in HL-60 leukemia cells.

    PubMed

    Landais, Emilie; El-Khoury, Victoria; Prevost, Alain; Dufer, Jean; Liautaud-Roger, Françoise

    2005-12-01

    Although nicotine has been implicated as a potential factor in the pathogenesis of human cancer, its mechanisms of action regarding cancer development remain largely unknown. HL-60 cells were used to investigate the effects of a short-term treatment with nicotine at concentrations found in the blood of smokers. The findings show that nicotine induces chromatin decondensation, histone H3 acetylation and up-regulation of the c-Jun transcription factor mRNA. This increase is inhibited by mecamylamine, a nicotinic receptor antagonist, suggesting that nicotine alters cellular function directly via nicotinic acetylcholine receptors and may then play a role in cell physiology and tumor promotion.

  11. Utrophin Up-Regulation by an Artificial Transcription Factor in Transgenic Mice

    PubMed Central

    Mattei, Elisabetta; Corbi, Nicoletta; Di Certo, Maria Grazia; Strimpakos, Georgios; Severini, Cinzia; Onori, Annalisa; Desantis, Agata; Libri, Valentina; Buontempo, Serena; Floridi, Aristide; Fanciulli, Maurizio; Baban, Dilair; Davies, Kay E.; Passananti, Claudio

    2007-01-01

    Duchenne Muscular Dystrophy (DMD) is a severe muscle degenerative disease, due to absence of dystrophin. There is currently no effective treatment for DMD. Our aim is to up-regulate the expression level of the dystrophin related gene utrophin in DMD, complementing in this way the lack of dystrophin functions. To this end we designed and engineered several synthetic zinc finger based transcription factors. In particular, we have previously shown that the artificial three zinc finger protein named Jazz, fused with the appropriate effector domain, is able to drive the transcription of a test gene from the utrophin promoter “A”. Here we report on the characterization of Vp16-Jazz-transgenic mice that specifically over-express the utrophin gene at the muscular level. A Chromatin Immunoprecipitation assay (ChIP) demonstrated the effective access/binding of the Jazz protein to active chromatin in mouse muscle and Vp16-Jazz was shown to be able to up-regulate endogenous utrophin gene expression by immunohistochemistry, western blot analyses and real-time PCR. To our knowledge, this is the first example of a transgenic mouse expressing an artificial gene coding for a zinc finger based transcription factor. The achievement of Vp16-Jazz transgenic mice validates the strategy of transcriptional targeting of endogenous genes and could represent an exclusive animal model for use in drug discovery and therapeutics. PMID:17712422

  12. Honey constituents up-regulate detoxification and immunity genes in the western honey bee Apis mellifera

    PubMed Central

    Mao, Wenfu; Schuler, Mary A.; Berenbaum, May R.

    2013-01-01

    As a managed pollinator, the honey bee Apis mellifera is critical to the American agricultural enterprise. Recent colony losses have thus raised concerns; possible explanations for bee decline include nutritional deficiencies and exposures to pesticides and pathogens. We determined that constituents found in honey, including p-coumaric acid, pinocembrin, and pinobanksin 5-methyl ether, specifically induce detoxification genes. These inducers are primarily found not in nectar but in pollen in the case of p-coumaric acid (a monomer of sporopollenin, the principal constituent of pollen cell walls) and propolis, a resinous material gathered and processed by bees to line wax cells. RNA-seq analysis (massively parallel RNA sequencing) revealed that p-coumaric acid specifically up-regulates all classes of detoxification genes as well as select antimicrobial peptide genes. This up-regulation has functional significance in that that adding p-coumaric acid to a diet of sucrose increases midgut metabolism of coumaphos, a widely used in-hive acaricide, by ∼60%. As a major component of pollen grains, p-coumaric acid is ubiquitous in the natural diet of honey bees and may function as a nutraceutical regulating immune and detoxification processes. The widespread apicultural use of honey substitutes, including high-fructose corn syrup, may thus compromise the ability of honey bees to cope with pesticides and pathogens and contribute to colony losses. PMID:23630255

  13. Water-soluble genistin glycoside isoflavones up-regulate antioxidant metallothionein expression and scavenge free radicals.

    PubMed

    Chung, Mi Ja; Kang, Ah-Young; Lee, Kyung Min; Oh, Eunji; Jun, Hee-Jin; Kim, Sang-Yeon; Auh, Joong Hyuck; Moon, Tae-Wha; Lee, Sung-Joon; Park, Kwan-Hwa

    2006-05-31

    Genistin has antioxidant activities; however, its insolubility in water often limits its biological availability in vivo. Using a novel transglycosylation process, the solubility of genistin glycosides was increased 1000 to 10000-fold, but it was not known whether these modified genistin glycosides maintained antioxidant activity. We found that both genistin and its glycosides similarly up-regulated the transcription of several metallothionein (MT) antioxidant genes (MT1A, MT2A, MT1E, and MT1X), as well as the glucose 6-phosphate dehydrogenase (G6PD) gene in HepG2 cells. This gene induction was mediated by the sequestration of zinc in the cytosol, which up-regulated the metal-responsive transcription factor-1 (MTF-1) that induced MT gene expression. Although not as effective as ascorbic acid, genistin glycosides possessed slightly greater reducing power than genistin. We concluded that genistin and genistin glycosides have a direct antioxidant effect and an indirect antioxidant effect, perhaps via induction of MT by activity of MTF-1.

  14. Tubular up-regulation of clusterin mRNA in murine lupus-like nephritis.

    PubMed Central

    Moll, S.; Menoud, P. A.; French, L.; Sappino, A. P.; Pastore, Y.; Schifferli, J. A.; Izui, S.

    1998-01-01

    Clusterin, a widely distributed glycoprotein, is detected in most tissues and in numerous physiological fluids. In the kidney, this protein is constitutively expressed in tubular epithelial cells, and its expression is enhanced following tubular injuries. In addition, clusterin has been detected in glomerular immune deposits of glomerulonephritis. The present study was designed to define the sites of clusterin mRNA accumulation in murine lupus-like nephritis in comparison with murine tubulopathies. In lupus-like nephritis, a significant increase of clusterin mRNA abundance was demonstrated. This up-regulation was localized exclusively in tubular epithelial cells exhibiting tubulointerstitial alterations, whereas no clusterin mRNA was detectable in diseased glomeruli, excluding an active synthesis of clusterin by glomerular cells. A similar tubular increase of clusterin mRNA abundance was observed in myeloma-like cast nephropathy induced by IgG3 monoclonal cryoglobulins and even in the absence of any detectable histological alterations in a model of septic shock induced by the injection of bacterial lipopolysaccharides. Our results suggest that tubular epithelial cells are the only sites of clusterin mRNA accumulation during the course of lupus-like nephritis and that the tubular up-regulation of clusterin gene expression may reflect the cellular response to various types of tubular injuries. Images Figure 1 Figure 3 Figure 4 Figure 5 Figure 7 PMID:9546356

  15. Low-Dose Cancer Risk Modeling Must Recognize Up-Regulation Of Protection

    PubMed Central

    Feinendegen, Ludwig E.; Pollycove, Myron; Neumann, Ronald D.

    2009-01-01

    Ionizing radiation primarily perturbs the basic molecular level proportional to dose, with potential damage propagation to higher levels: cells, tissues, organs, and whole body. There are three types of defenses against damage propagation. These operate deterministically and below a certain impact threshold there is no propagation. Physical-static defenses precede metabolic-dynamic defenses acting immediately: scavenging of toxins; - molecular repair, especially of DNA; - removal of damaged cells either by apoptosis, necrosis, phagocytosis, cell differentiation-senescence, or by immune responses, - followed by replacement of lost elements. Another metabolic-dynamic defense arises delayed by up-regulating immediately operating defense mechanisms. Some of these adaptive protections may last beyond a year and all create temporary protection against renewed potentially toxic impacts also from non-radiogenic endogenous sources. Adaptive protections have a maximum after single tissue absorbed doses around 100 to 200 mSv and disappear with higher doses. Low dose rates initiate maximum protection likely at lower cell doses delivered repetitively at certain time intervals. Adaptive protection preventing only about 2 – 3 % of endogenous life-time cancer risk would fully balance a calculated induced cancer risk at about 100 mSv, in agreement with epidemiological data and concordant with an hormetic effect. Low-dose-risk modeling must recognize up-regulation of protection. PMID:20585440

  16. N-glycoprotein analysis discovers new up-regulated glycoproteins in colorectal cancer tissue.

    PubMed

    Nicastri, Annalisa; Gaspari, Marco; Sacco, Rosario; Elia, Laura; Gabriele, Caterina; Romano, Roberto; Rizzuto, Antonia; Cuda, Giovanni

    2014-11-01

    Colorectal cancer is one of the leading causes of death due to cancer worldwide. Therefore, the identification of high-specificity and -sensitivity biomarkers for the early detection of colorectal cancer is urgently needed. Post-translational modifications, such as glycosylation, are known to play an important role in cancer progression. In the present work, we used a quantitative proteomic technique based on (18)O stable isotope labeling to identify differentially expressed N-linked glycoproteins in colorectal cancer tissue samples compared with healthy colorectal tissue from 19 patients undergoing colorectal cancer surgery. We identified 54 up-regulated glycoproteins in colorectal cancer samples, therefore potentially involved in the biological processes of tumorigenesis. In particular, nine of these (PLOD2, DPEP1, SE1L1, CD82, PAR1, PLOD3, S12A2, LAMP3, OLFM4) were found to be up-regulated in the great majority of the cohort, and, interestingly, the association with colorectal cancer of four (PLOD2, S12A2, PLOD3, CD82) has not been hitherto described.

  17. Up-regulation of vitamin B1 homeostasis genes in breast cancer.

    PubMed

    Zastre, Jason A; Hanberry, Bradley S; Sweet, Rebecca L; McGinnis, A Cary; Venuti, Kristen R; Bartlett, Michael G; Govindarajan, Rajgopal

    2013-09-01

    An increased carbon flux and exploitation of metabolic pathways for the rapid generation of biosynthetic precursors is a common phenotype observed in breast cancer. To support this metabolic phenotype, cancer cells adaptively regulate the expression of glycolytic enzymes and nutrient transporters. However, activity of several enzymes involved in glucose metabolism requires an adequate supply of cofactors. In particular, vitamin B1 (thiamine) is utilized as an essential cofactor for metabolic enzymes that intersect at critical junctions within the glycolytic network. Intracellular availability of thiamine is facilitated by the activity of thiamine transporters and thiamine pyrophosphokinase-1 (TPK-1). Therefore, the objective of this study was to establish if the cellular determinants regulating thiamine homeostasis differ between breast cancer and normal breast epithelia. Employing cDNA arrays of breast cancer and normal breast epithelial tissues, SLC19A2, SLC25A19 and TPK-1 were found to be significantly up-regulated. Similarly, up-regulation was also observed in breast cancer cell lines compared to human mammary epithelial cells. Thiamine transport assays and quantitation of intracellular thiamine and thiamine pyrophosphate established a significantly greater extent of thiamine transport and free thiamine levels in breast cancer cell lines compared to human mammary epithelial cells. Overall, these findings demonstrate an adaptive response by breast cancer cells to increase cellular availability of thiamine.

  18. Honey constituents up-regulate detoxification and immunity genes in the western honey bee Apis mellifera.

    PubMed

    Mao, Wenfu; Schuler, Mary A; Berenbaum, May R

    2013-05-28

    As a managed pollinator, the honey bee Apis mellifera is critical to the American agricultural enterprise. Recent colony losses have thus raised concerns; possible explanations for bee decline include nutritional deficiencies and exposures to pesticides and pathogens. We determined that constituents found in honey, including p-coumaric acid, pinocembrin, and pinobanksin 5-methyl ether, specifically induce detoxification genes. These inducers are primarily found not in nectar but in pollen in the case of p-coumaric acid (a monomer of sporopollenin, the principal constituent of pollen cell walls) and propolis, a resinous material gathered and processed by bees to line wax cells. RNA-seq analysis (massively parallel RNA sequencing) revealed that p-coumaric acid specifically up-regulates all classes of detoxification genes as well as select antimicrobial peptide genes. This up-regulation has functional significance in that that adding p-coumaric acid to a diet of sucrose increases midgut metabolism of coumaphos, a widely used in-hive acaricide, by ∼60%. As a major component of pollen grains, p-coumaric acid is ubiquitous in the natural diet of honey bees and may function as a nutraceutical regulating immune and detoxification processes. The widespread apicultural use of honey substitutes, including high-fructose corn syrup, may thus compromise the ability of honey bees to cope with pesticides and pathogens and contribute to colony losses.

  19. Laughter up-regulates the genes related to NK cell activity in diabetes.

    PubMed

    Hayashi, Takashi; Tsujii, Satoru; Iburi, Tadao; Tamanaha, Tamiko; Yamagami, Keiko; Ishibashi, Rieko; Hori, Miyo; Sakamoto, Shigeko; Ishii, Hitoshi; Murakami, Kazuo

    2007-12-01

    To elucidate the sustainable effects of laughter on gene expression, we recruited type 2 diabetic patients who were in-patient for receiving self-management education and examined time-dependent regulation for gene expression by laughter. Two-day experiment was performed. On one day, the patients watched comic video and laughed together with hospital staffs. On the other day, they participated in an inpatient diabetes educational program. Blood samples were collected before and 1.5, 4 h after watching comic video or spending lecture time, and changes in gene expression were comprehensively analyzed by microarray technique. Of the 41,000 genes analyzed, the laughter relatively up-regulated 39 genes, among which, 27 genes were relatively increased in the expression for all the observation period after watching comic video. By functional classification of these genes, 14 genes were found to be related to natural killer cell activity. No genes were included that are directly involved in blood glucose regulation, though successive suppression of postprandial blood glucose levels was observed. These results suggest that the laughter influences the expression of many genes classified into immune responses, and may contribute to amelioration of postprandial blood glucose elevation through a modulation of NK cell activity caused by up-regulation of relating genes.

  20. Up-regulation of NF45 correlates with Schwann cell proliferation after sciatic nerve crush.

    PubMed

    Wang, Youhua; Zhou, Shiran; Xu, Hua; Yan, Shixian; Xu, Dawei; Zhang, Yi

    2015-05-01

    Nuclear factor (NF)45 (also known as interleukin enhancer-binding factor (ILF)2), is a transcription factor that interacts with NF90 to regulate gene expression. It has long been implicated in the regulation of cell proliferation. However, the role of NF45 in the process of peripheral nervous system regeneration after injury remains poorly understood. Herein, we investigated the spatiotemporal expression of NF45 in a rat sciatic nerve crush model. We detected the up-regulated expression of NF45 in Schwann cell after sciatic nerve crush. What's more, the expression of the cell proliferation marker proliferating cell nuclear antigen (PCNA) exhibited a similar tendency with that of NF45. In cell cultures, we observed increased expression of NF45 during the process of TNF-α-induced Schwann cell proliferation, whereas the protein level of p21 was down-regulated. Interference of NF45 led to enhanced expression of p21 and also impaired proliferation of Schwan cells. Taken together, our data implicated that NF45 was up-regulated in the sciatic nerve after crush, which was associated with proliferation of Schwann cell.

  1. Up-regulated expression of Ran reveals its potential role to deltamethrin stress in Kc cells.

    PubMed

    Liu, Wei; Xu, Qin; Chi, Qingping; Hu, Junli; Li, Fengliang; Cheng, Luogen

    2016-05-25

    The GTP-binding nuclear protein Ran has mostly been reported to be an essential player in nuclear transport, chromosome alignment, microtubule dynamics, centrosome duplication, kinetochore attachment of microtubules, nuclear-envelope dynamics, and phagocytosis. However, until now, there has been no report showing the involvement of Ran in DM stress. In this paper, two-dimensional electrophoresis analysis showed that the expression level of Ran in Kc cells in response to DM was higher than that in the control group. In addition, quantitative analysis using real-time PCR revealed that the expression of Ran was obviously up-regulated at various concentrations of DM. Western blot analysis showed that Ran was up-regulated 2.27-fold over the control at 48h. Because we still could not pinpoint whether Ran was actually involved in DM stress reaction, to further verify the role of Ran in stress reaction, RNA interference and cell transfection were utilized. Overexpression of Ran in cells conferred a degree of protection against DM after 72h. Furthermore, interference with Ran significantly decrease cell viability. All of the above findings strongly imply that Ran may participate in the development of stress reaction to DM. Therefore, investigating the possible role of Ran in DM stress will broaden our limited knowledge regarding DM stress inducible genes. PMID:26924245

  2. Honey constituents up-regulate detoxification and immunity genes in the western honey bee Apis mellifera.

    PubMed

    Mao, Wenfu; Schuler, Mary A; Berenbaum, May R

    2013-05-28

    As a managed pollinator, the honey bee Apis mellifera is critical to the American agricultural enterprise. Recent colony losses have thus raised concerns; possible explanations for bee decline include nutritional deficiencies and exposures to pesticides and pathogens. We determined that constituents found in honey, including p-coumaric acid, pinocembrin, and pinobanksin 5-methyl ether, specifically induce detoxification genes. These inducers are primarily found not in nectar but in pollen in the case of p-coumaric acid (a monomer of sporopollenin, the principal constituent of pollen cell walls) and propolis, a resinous material gathered and processed by bees to line wax cells. RNA-seq analysis (massively parallel RNA sequencing) revealed that p-coumaric acid specifically up-regulates all classes of detoxification genes as well as select antimicrobial peptide genes. This up-regulation has functional significance in that that adding p-coumaric acid to a diet of sucrose increases midgut metabolism of coumaphos, a widely used in-hive acaricide, by ∼60%. As a major component of pollen grains, p-coumaric acid is ubiquitous in the natural diet of honey bees and may function as a nutraceutical regulating immune and detoxification processes. The widespread apicultural use of honey substitutes, including high-fructose corn syrup, may thus compromise the ability of honey bees to cope with pesticides and pathogens and contribute to colony losses. PMID:23630255

  3. Utrophin up-regulation by an artificial transcription factor in transgenic mice.

    PubMed

    Mattei, Elisabetta; Corbi, Nicoletta; Di Certo, Maria Grazia; Strimpakos, Georgios; Severini, Cinzia; Onori, Annalisa; Desantis, Agata; Libri, Valentina; Buontempo, Serena; Floridi, Aristide; Fanciulli, Maurizio; Baban, Dilair; Davies, Kay E; Passananti, Claudio

    2007-08-22

    Duchenne Muscular Dystrophy (DMD) is a severe muscle degenerative disease, due to absence of dystrophin. There is currently no effective treatment for DMD. Our aim is to up-regulate the expression level of the dystrophin related gene utrophin in DMD, complementing in this way the lack of dystrophin functions. To this end we designed and engineered several synthetic zinc finger based transcription factors. In particular, we have previously shown that the artificial three zinc finger protein named Jazz, fused with the appropriate effector domain, is able to drive the transcription of a test gene from the utrophin promoter "A". Here we report on the characterization of Vp16-Jazz-transgenic mice that specifically over-express the utrophin gene at the muscular level. A Chromatin Immunoprecipitation assay (ChIP) demonstrated the effective access/binding of the Jazz protein to active chromatin in mouse muscle and Vp16-Jazz was shown to be able to up-regulate endogenous utrophin gene expression by immunohistochemistry, western blot analyses and real-time PCR. To our knowledge, this is the first example of a transgenic mouse expressing an artificial gene coding for a zinc finger based transcription factor. The achievement of Vp16-Jazz transgenic mice validates the strategy of transcriptional targeting of endogenous genes and could represent an exclusive animal model for use in drug discovery and therapeutics.

  4. Celecoxib Up Regulates the Expression of Drug Efflux Transporter ABCG2 in Breast Cancer Cell Lines

    PubMed Central

    Kalalinia, Fatemeh; Elahian, Fatemeh; Mosaffa, Fatemeh; Behravan, Javad

    2014-01-01

    Elevated expression of the drug efflux transporter ABCG2 seems to correlate with multidrug resistance of cancer cells. Specific COX-2 inhibitor celecoxib has been shown to enhance the sensitivity of cancer cells to anticancer drugs. To clarify whether ABCG2 inhibition is involved in the sensitizing effect of celecoxib, we investigated whether the expression of ABCG2 in breast cancer cell lines, could be modulated by celecoxib. The expression of the multidrug resistant gene (ABCG2) at mRNA and protein level was detected by real-time quantitative reverse transcription-polymerase chain reaction and flow cytometry analysis, respectively. Among three human breast cancer cell lines ABCG2 and COX-2 were highly expressed in MCF7-MX and MDA-MB-231 cells, respectively. The COX-2 inhibitor celecoxib up-regulated the expression of ABCG2 mRNA in MCF-7 and MCF7-MX cells, which was accompanied by increased ABCG2 protein expression. While celecoxib was able to block the 12-O-tetradecanoylphorbol-13-acetate (TPA)-mediated increase in COX-2 expression in MDA-MB-231 cells, it increased the expression of ABCG2 up to 4.27 times to the control level at mRNA level and with less intensity at protein level. Our findings provide evidence that celecoxib up-regulates ABCG2 expression in human breast cancer cells and proposed that ABCG2 is not involved in chemosensitizing effects of celecoxib. PMID:25587329

  5. Transcriptional up-regulation of the human androgen receptor by androgen in bone cells.

    PubMed

    Wiren, K M; Zhang, X; Chang, C; Keenan, E; Orwoll, E S

    1997-06-01

    Androgen regulation of androgen receptor (AR) expression has been observed in a variety of tissues, generally as inhibition, and is thought to attenuate cellular responses to androgen. AR is expressed in osteoblasts, the bone-forming cell, suggesting direct actions of androgens on bone. Here we characterized the effect of androgen exposure on AR gene expression in human osteoblastic SaOS-2 and U-2 OS cells. Treatment of osteoblastic cells with the nonaromatizable androgen 5alpha-dihydrotestosterone increased AR steady state messenger RNA levels in a time- and dose-dependent fashion. Reporter assays with 2.3 kilobases of the proximal 5'-flanking region of the human AR promoter linked to the chloramphenicol acetyltransferase gene in transfected cultures showed that up-regulation of AR promoter activity by androgen was time and dose dependent. Treatment with other steroid hormones, including progesterone, 17beta-estradiol, and dexamethasone, was without effect. The antiandrogen hydroxyflutamide completely antagonized androgen up-regulation. Thus, in contrast to many other androgen target tissues, androgen exposure increases steady state AR messenger RNA levels in osteoblasts. This regulation occurs at least partially at the level of transcription, is mediated by the 5'-promoter region of the AR gene, and is dependent on functional AR. These results suggest that physiological concentrations of androgens have significant effects on AR expression in skeletal tissue. PMID:9165014

  6. HBx sensitizes hepatocellular carcinoma cells to lapatinib by up-regulating ErbB3

    PubMed Central

    Yen, Chia-Jui; Chen, Wen-Shu; Huang, Wei-Chien

    2016-01-01

    Poor prognosis of hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC) involves HBV X protein (HBx)-induced tumor progression. HBx also contributes to chemo-resistance via inducing the expressions of anti-apoptosis and multiple drug resistance genes. However, the impact of HBx expression on the therapeutic efficacy of various receptor tyrosine kinase inhibitors remains unknown. In this study, our data showed that HBx overexpression did not alter the cellular sensitivity of HCC cell lines to sorafenib but unexpectedly enhanced the cell death induced by EGFR family inhibitors, including gefitinib, erlotinib, and lapatinib due to ErbB3 up-regulation. Mechanistically, HBx transcriptionally up-regulates ErbB3 expression in a NF-κB dependent manner. In addition, HBx also physically interacts with ErbB2 and ErbB3 proteins and enhances the formation of ErbB2/ErbB3 heterodimeric complex. The cell viability of HBx-overexpressing cells was decreased by silencing ErbB3 expression, further revealing the pivotal role of ErbB3 in HBx-mediated cell survival. Our data suggest that HBx shifts the oncogenic addiction of HCC cells to ErbB2/ErbB3 signaling pathway via inducing ErbB3 expression and thereby enhances their sensitivity to EGFR/ErbB2 inhibitors. PMID:26595522

  7. Up-regulated Smad5 mediates apoptosis of gastric epithelial cells induced by Helicobacter pylori infection.

    PubMed

    Nagasako, Tomokazu; Sugiyama, Toshiro; Mizushima, Takuji; Miura, Yosuke; Kato, Mototsugu; Asaka, Masahiro

    2003-02-14

    The gastric pathogen Helicobacter pylori activates epithelial cell signaling pathways, and its infection induces changes in the expression of several genes in infected human gastric tissues. Recent studies have indicated that the ability of H. pylori to regulate epithelial cell responses depends on the presence of an intact cag pathogenicity island (cagPAI). We investigated altered mRNA expression of gastric epithelial cells after infection with H. pylori, both cagPAI-positive and cagPAI-negative strains, by cDNA microarray, reverse transcription PCR, and Northern blot analysis. Our results indicated that cagPAI-positive H. pylori strains (ATCC 43504 and clinical isolated strains) significantly activated Smad5 mRNA expression of human gastric epithelial cells (AGS, KATOIII, MKN28, and MKN45). We further examined whether the up-regulated Smad5 was related to apoptosis of gastric epithelial cells induced by H. pylori. Smad5 RNA interference completely inhibited H. pylori-induced apoptosis. These results suggest that Smad5 is up-regulated in gastric epithelial cells through the presence of cagPAI of H. pylori and that Smad5 mediates apoptosis of gastric epithelial cells induced by H. pylori infection. PMID:12473652

  8. Tubular up-regulation of clusterin mRNA in murine lupus-like nephritis.

    PubMed

    Moll, S; Menoud, P A; French, L; Sappino, A P; Pastore, Y; Schifferli, J A; Izui, S

    1998-04-01

    Clusterin, a widely distributed glycoprotein, is detected in most tissues and in numerous physiological fluids. In the kidney, this protein is constitutively expressed in tubular epithelial cells, and its expression is enhanced following tubular injuries. In addition, clusterin has been detected in glomerular immune deposits of glomerulonephritis. The present study was designed to define the sites of clusterin mRNA accumulation in murine lupus-like nephritis in comparison with murine tubulopathies. In lupus-like nephritis, a significant increase of clusterin mRNA abundance was demonstrated. This up-regulation was localized exclusively in tubular epithelial cells exhibiting tubulointerstitial alterations, whereas no clusterin mRNA was detectable in diseased glomeruli, excluding an active synthesis of clusterin by glomerular cells. A similar tubular increase of clusterin mRNA abundance was observed in myeloma-like cast nephropathy induced by IgG3 monoclonal cryoglobulins and even in the absence of any detectable histological alterations in a model of septic shock induced by the injection of bacterial lipopolysaccharides. Our results suggest that tubular epithelial cells are the only sites of clusterin mRNA accumulation during the course of lupus-like nephritis and that the tubular up-regulation of clusterin gene expression may reflect the cellular response to various types of tubular injuries.

  9. Midazolam inhibits the hypoxia-induced up-regulation of erythropoietin in the central nervous system.

    PubMed

    Matsuyama, Tomonori; Tanaka, Tomoharu; Tatsumi, Kenichiro; Daijo, Hiroki; Kai, Shinichi; Harada, Hiroshi; Fukuda, Kazuhiko

    2015-08-15

    Erythropoietin (EPO), a regulator of red blood cell production, is endogenously expressed in the central nervous system. It is mainly produced by astrocytes under hypoxic conditions and has proven to have neuroprotective and neurotrophic effects. In the present study, we investigated the effect of midazolam on EPO expression in primary cultured astrocytes and the mouse brain. Midazolam was administered to 6-week-old BALB/c male mice under hypoxic conditions and pregnant C57BL/6N mice under normoxic conditions. Primary cultured astrocytes were also treated with midazolam under hypoxic conditions. The expression of EPO mRNA in mice brains and cultured astrocytes was studied. In addition, the expression of hypoxia-inducible factor (HIF), known as the main regulator of EPO, was evaluated. Midazolam significantly reduced the hypoxia-induced up-regulation of EPO in BALB/c mice brains and primary cultured astrocytes and suppressed EPO expression in the fetal brain. Midazolam did not affect the total amount of HIF proteins but significantly inhibited the nuclear expression of HIF-1α and HIF-2α proteins. These results demonstrated the suppressive effects of midazolam on the hypoxia-induced up-regulation of EPO both in vivo and in vitro. PMID:26001375

  10. Transforming growth factor-β1 up-regulates connexin43 expression in human granulosa cells

    PubMed Central

    Chen, Yu-Ching; Chang, Hsun-Ming; Cheng, Jung-Chien; Tsai, Horng-Der; Wu, Cheng-Hsuan; Leung, Peter C.K.

    2015-01-01

    STUDY QUESTION Does transforming growth factor-β1 (TGF-β1) up-regulate connexin43 (Cx43) to promote cell–cell communication in human granulosa cells? SUMMARY ANSWER TGF-β1 up-regulates Cx43 and increases gap junction intercellular communication activities (GJIC) in human granulosa cells, and this effect occurs via the activin receptor-like kinase (ALK)5-mediated Sma- and Mad-related protein (SMAD)2/3-SMAD4-dependent pathway. WHAT IS KNOWN ALREADY TGF-β1 and its receptors are expressed in human granulosa cells, and follicular fluid contains TGF-β1 protein. In human granulosa cells, Cx43 gap junctions play an important role in the development of follicles and oocytes. STUDY DESIGN, SIZE, DURATION This is an experimental study which was performed over a 1-year period. PARTICIPANTS/MATERIALS, SETTING, METHODS Immortalized human granulosa cells (SVOG cells) and primary human granulosa-lutein cells obtained from women undergoing IVF in an academic research center were used as the study models. Cx43 mRNA and protein expression levels were examined after exposure of SVOG cells to recombinant human TGF-β1. An activin/TGF-β type I receptor inhibitor, SB431542, and small interfering RNAs targeting ALK4, ALK5, SMAD2, SMAD3 and SMAD4 were used to verify the specificity of the effects and to investigate the molecular mechanisms. Real-time-quantitative PCR and western blot analysis were used to detect the specific mRNA and protein levels, respectively. GJIC between SVOG cells were evaluated using a scrape loading and dye transfer assay. Results were analyzed by one-way analysis of variance. MAIN RESULTS AND THE ROLE OF CHANCE TGF-β1 treatment increased phosphorylation of SMAD2/3 (P < 0.0001) and up-regulated Cx43 mRNA and protein levels (P < 0.001) in SVOG cells and these stimulatory effects were abolished by the TGF-β type I receptor inhibitor SB431542. In addition, the up-regulatory effect of TGF-β1 on Cx43 expression (mRNA and protein) was confirmed in primary

  11. Up-regulation of CLDN1 in gastric cancer is correlated with reduced survival

    PubMed Central

    2013-01-01

    Background The genetic changes in gastric adenocarcinoma are extremely complex and reliable tumor markers have not yet been identified. There are also remarkable geographical differences in the distribution of this disease. Our aim was to identify the most differentially regulated genes in 20 gastric adenocarcinomas from a Norwegian selection, compared to matched normal mucosa, and we have related our findings to prognosis, survival and chronic Helicobacter pylori infection. Methods Biopsies from gastric adenocarcinomas and adjacent normal gastric mucosa were obtained from 20 patients immediately following surgical resection of the tumor. Whole genome, cDNA microarray analysis was performed on the RNA isolated from the sample pairs to compare the gene expression profiles between the tumor against matched mucosa. The samples were microscopically examined to classify gastritis. The presence of H. pylori was examined using microscopy and immunohistochemistry. Results 130 genes showed differential regulation above a predefined cut-off level. Interleukin-8 (IL-8) and Claudin-1 (CLDN1) were the most consistently up-regulated genes in the tumors. Very high CLDN1 expression in the tumor was identified as an independent and significant predictor gene of reduced post-operative survival. There were distinctly different expression profiles between the tumor group and the control mucosa group, and the histological subsets of mixed type, diffuse type and intestinal type cancer demonstrated further sub-clustering. Up-regulated genes were mapped to cell-adhesion, collagen-related processes and angiogenesis, whereas normal intestinal functions such as digestion and excretion were associated with down-regulated genes. We relate the current findings to our previous study on the gene response of gastric epithelial cells to H. pylori infection. Conclusions CLDN1 was highly up-regulated in gastric cancer, and CLDN1 expression was independently associated with a poor post

  12. Physiological effects of major up-regulated Alnus glutinosa peptides on Frankia sp. ACN14a.

    PubMed

    Carro, Lorena; Pujic, Petar; Alloisio, Nicole; Fournier, Pascale; Boubakri, Hasna; Poly, Franck; Rey, Marjolaine; Heddi, Abdelaziz; Normand, Philippe

    2016-07-01

    Alnus glutinosa has been shown previously to synthesize, in response to nodulation by Frankia sp. ACN14a, an array of peptides called Alnus symbiotic up-regulated peptides (ASUPs). In a previous study one peptide (Ag5) was shown to bind to Frankia nitrogen-fixing vesicles and to modify their porosity. Here we analyse four other ASUPs, alongside Ag5, to determine whether they have different physiological effects on in vitro grown Frankia sp. ACN14a. The five studied peptides were shown to have different effects on nitrogen fixation, respiration, growth, the release of ions and amino acids, as well as on cell clumping and cell lysis. The mRNA abundance for all five peptides was quantified in symbiotic nodules and one (Ag11) was found to be more abundant in the meristem part of the nodule. These findings point to some peptides having complementary effects on Frankia cells. PMID:27082768

  13. Up-regulation of vimentin expression during regeneration in the adult fish brain.

    PubMed

    Clint, Sorcha C; Zupanc, Günther K H

    2002-03-01

    In contrast to mammals, the brains of teleost fish exhibit an enormous regenerative capacity following injury. Here, we have examined the potential role of vimentin in this wound healing. Fifteen days after application of a mechanical lesion to the corpus cerebelli in the teleost fish Apteronotus leptoryhnchus, the areal density of vimentin-positive fibres increased significantly at the lesion site and in the remaining ipsilateral molecular layer. This density remained elevated throughout the time period of up to 100 days examined. Based on this spatio-temporal pattern of vimentin up-regulation we propose that this intermediate filament protein is involved in the survival, differentiation, and/or dendritic growth of the new cells that replace damaged cells in the injury zone.

  14. Fetal nicotine exposure produces postnatal up-regulation of adenylate cyclase activity in peripheral tissues

    SciTech Connect

    Slotkin, T.A.; Navarro, H.A.; McCook, E.C.; Seidler, F.J. )

    1990-01-01

    Gestational exposure to nicotine has been shown to affect development of noradrenergic activity in both the central and peripheral nervous systems. In the current study, pregnant rats received nicotine infusions of 6 mg/kg/day throughout gestation, administered by osmotic minipump implants. After birth, offspring of the nicotine-infused dams exhibited marked increases in basal adenylate cyclase activity in membranes prepared from kidney and heart, as well as supersensitivity to stimulation by either a {beta}-adrenergic agonist, isoproterenol, or by forskolin. The altered responses were not accompanied by up-regulation of {beta}-adrenergic receptors: in fact, ({sup 125}I)pindolol binding was significantly decreased in the nicotine group. These results indicate that fetal nicotine exposure affects enzymes involved in membrane receptor signal transduction, leading to altered responsiveness independently of changes at the receptor level.

  15. Water deprivation up-regulates urine osmolality and renal aquaporin 2 in Mongolian gerbils (Meriones unguiculatus).

    PubMed

    Xu, Meng-Meng; Wang, De-Hua

    2016-04-01

    To better understand how desert rodents adapt to water scarcity, we examined urine osmolality, renal distribution and expression of aquaporins (AQPs) in Mongolian gerbils (Meriones unguiculatus) during 7 days of water deprivation (WD). Urine osmolality of the gerbils during WD averaged 7503 mOsm kg(-1). Renal distributions of AQP1, AQP2, and AQP3 were similar to that described in other rodents. After the 7 day WD, renal AQP2 was up-regulated, while resting metabolic rate and total evaporative water loss decreased by 43% and 36%, respectively. Our data demonstrated that Mongolian gerbils showed high urine concentration, renal AQPs expression and body water conservation to cope with limited water availability, which may be critical for their survival during dry seasons in cold deserts. PMID:26806059

  16. Maggot debridement therapy promotes diabetic foot wound healing by up-regulating endothelial cell activity.

    PubMed

    Sun, Xinjuan; Chen, Jin'an; Zhang, Jie; Wang, Wei; Sun, Jinshan; Wang, Aiping

    2016-03-01

    To determine the role of maggot debridement therapy (MDT) on diabetic foot wound healing, we compared growth related factors in wounds before and after treatment. Furthermore, we utilized human umbilical vein endothelial cells (HUVECs) to explore responses to maggot excretions/secretions on markers of angiogenesis and proliferation. The results showed that there was neo-granulation and angiogenesis in diabetic foot wounds after MDT. Moreover, significant elevation in CD34 and CD68 levels was also observed in treated wounds. In vitro, ES increased HUVEC proliferation, improved tube formation, and increased expression of vascular endothelial growth factor receptor 2 in a dose dependent manner. These results demonstrate that MDT and maggot ES can promote diabetic foot wound healing by up-regulating endothelial cell activity.

  17. A novel transcript is up-regulated by fasting in the hypothalamus and enhances insulin signaling

    PubMed Central

    Chai, Biaoxin; Li, Ji-Yao; Fritze, Danielle; Zhang, Weizhen; Xia, Zefeng; Mulholland, Michael W

    2015-01-01

    A transcript of unknown function, regulated by fasting and feeding, was identified by microarray analysis. The transcript is up-regulated in the fasting state. An 1168 base-pair cDNA was cloned from rat hypothalamus and sequenced. This sequence is consistent with adipogenesis downregulating transcript 3 (AGD3) (also known as human OCC-1) mRNA. A protein sequence identical to AGD3 was determined by mass spectrometry. In rat brain, AGD3 mRNA is distributed in arcuate nucleus, ventromedial hypothalamus, amygdaloid nuclei, paraventricular nucleus (PVN), hippocampus, and somatic cortex. Double in situ hybridization showed that AGD3 mRNA is co-localized with pro-opiomelanocortin and neuropeptide Y in arcuate nucleus neurons. AGD3 binds with insulin receptor substrate 4 and increases insulin-stimulated phospho-AKT and regulates AMPK and mTOR downstream target S6 kinase phosphorylation. PMID:22935015

  18. A novel transcript is up-regulated by fasting in the hypothalamus and enhances insulin signalling.

    PubMed

    Chai, B; Li, J-Y; Fritze, D; Zhang, W; Xia, Z; Mulholland, M W

    2013-03-01

    A transcript of unknown function, regulated by fasting and feeding, was identified by microarray analysis. The transcript is up-regulated in the fasting state. An 1168-bp cDNA was cloned from rat hypothalamus and sequenced. This sequence is consistent with adipogenesis down-regulating transcript 3 (AGD3) (also known as human OCC-1) mRNA. A protein sequence identical to AGD3 was determined by mass spectrometry. In the rat brain, AGD3 mRNA is distributed in the arcuate nucleus, ventromedial hypothalamus, amygdaloid nuclei, hippocampus, and somatic cortex. Double in situ hybridisation showed that AGD3 mRNA is co-localised with pro-opiomelanocortin and neuropeptide Y in arcuate nucleus neurones. AGD3 binds with insulin receptor substrate 4 and increases insulin-stimulated phospho-Akt and regulates AMP-activated protein kinase and mammalian target of rapamycin downstream target S6 kinase phosphorylation.

  19. Exercise-induced up-regulation of MMP-1 and IL-8 genes in endurance horses

    PubMed Central

    Cappelli, Katia; Felicetti, Michela; Capomaccio, Stefano; Pieramati, Camillo; Silvestrelli, Maurizio; Verini-Supplizi, Andrea

    2009-01-01

    Background The stress response is a critical factor in the training of equine athletes; it is important for performance and for protection of the animal against physio-pathological disorders. In this study, the molecular mechanisms involved in the response to acute and strenuous exercise were investigated using peripheral blood mononuclear cells (PBMCs). Results Quantitative real-time PCR (qRT-PCR) was used to detect modifications in transcription levels of the genes for matrix metalloproteinase-1 (MMP-1) and interleukin 8 (IL-8), which were derived from previous genome-wide expression analysis. Significant up-regulation of these two genes was found in 10 horses that had completed a race of 90–120 km in a time-course experimental design. Conclusion These results suggest that MMP-1 and IL-8 are both involved in the exercise-induced stress response, and this represents a starting point from which to understand the adaptive responses to this phenomenon. PMID:19552796

  20. Axl receptor tyrosine kinase is up-regulated in metformin resistant prostate cancer cells

    PubMed Central

    Bansal, Nitu; Mishra, Prasun J.; Stein, Mark; DiPaola, Robert S.; Bertino, Joseph R.

    2015-01-01

    Recent epidemiological studies showed that metformin, a widely used anti-diabetic drug might prevent certain cancers. Metformin also has an anti-proliferative effect in preclinical studies of both hematologic malignancies as well as solid cancers and clinical studies testing metformin as an anti-cancer drug are in progress. However, all cancer types do not respond to metformin with the same effectiveness or acquire resistance. To understand the mechanism of acquired resistance and possibly its mechanism of action as an anti-proliferative agent, we developed metformin resistant LNCaP prostate cancer cells. Metformin resistant LNCaP cells had an increased proliferation rate, increased migration and invasion ability as compared to the parental cells, and expressed markers of epithelial-mesenchymal transition (EMT). A detailed gene expression microarray comparing the resistant cells to the wild type cells revealed that Edil2, Ereg, Axl, Anax2, CD44 and Anax3 were the top up-regulated genes and calbindin 2 and TPTE (transmembrane phosphatase with tensin homology) and IGF1R were down regulated. We focused on Axl, a receptor tyrosine kinase that has been shown to be up regulated in several drug resistance cancers. Here, we show that the metformin resistant cell line as well as castrate resistant cell lines that over express Axl were more resistant to metformin, as well as to taxotere compared to androgen sensitive LNCaP and CWR22 cells that do not overexpress Axl. Forced overexpression of Axl in LNCaP cells decreased metformin and taxotere sensitivity and knockdown of Axl in resistant cells increased sensitivity to these drugs. Inhibition of Axl activity by R428, a small molecule Axl kinase inhibitor, sensitized metformin resistant cells that overexpressed Axl to metformin. Inhibitors of Axl may enhance tumor responses to metformin and other chemotherapy in cancers that over express Axl. PMID:26036314

  1. Curcumin Inhibits Prostate Cancer Bone Metastasis by Up-Regulating Bone Morphogenic Protein-7 in Vivo

    PubMed Central

    Dorai, Thambi; Diouri, Janane; O'Shea, Orla; Doty, Stephen B.

    2014-01-01

    A number of studies have focused on the beneficial properties of Curcumin (diferuloyl methane, used in South Asian cuisine and traditional medicine) such as the chemoprevention of cancer. Recent studies have also indicated that this material has significant benefits for the treatment of cancer and is currently undergoing several clinical trials. We have been interested in the application of this compound as a therapeutic agent for advanced prostate cancer, particularly the skeletal complications in this malignancy. Our earlier work indicated that this compound could inhibit the osteomimetic properties which occur in castration resistant prostate cancer cells, by interfering with the common denominators between these cancer cells and the bone cells in the metastatic tumor microenvironment, namely the osteoblasts and the osteoclast. We predicted that curcumin could break the vicious cycle of reciprocal stimulation that results in uncontrolled osteolysis in the bony matrix. In this work, we have evaluated the potential of this compound in inhibiting the bone metastasis of hormone refractory prostate cancer cells in an established animal model. Our results strongly suggest that curcumin modulates the TGF-β signaling that occurs due to bone matrix degradation by up-regulating the metastasis inhibitory bone morphogenic protein-7 (BMP- 7). This enhancement of BMP-7 in the context of TGF-βin the tumor microenvironment is shown to enhance the mesenchymal-to-epithelial transition. Most importantly, we show that as a result of BMP-7 up-regulation, a novel brown/beige adipogenic differentiation program is also up-regu- lated which plays a role in the inhibition of bone metastasis. Our results suggest that curcumin may subvert the TGF-βsignaling to an alternative adipogenic differentiation program in addition to the previously established interference with the osteomimetic properties, thus inhibiting the bone metastatic processes in a chemopreventive as well as therapeutic

  2. Nicotine activates and up-regulates nicotinic acetylcholine receptors in bronchial epithelial cells.

    PubMed

    Fu, Xiao Wen; Lindstrom, Jon; Spindel, Eliot R

    2009-07-01

    Prenatal nicotine exposure impairs normal lung development and leads to diminished pulmonary function after birth. Previous work from our laboratory has demonstrated that nicotine alters lung development by affecting a nonneuronal cholinergic autocrine loop that is expressed in lung. Bronchial epithelial cells (BECs) express choline acetyltransferase, the choline high-affinity transporter and nicotinic acetylcholine (ACh) receptor (nAChR) subunits. We now demonstrate through a combination of morphological and electrophysiological techniques that nicotine affects this autocrine loop by up-regulating and activating cholinergic signaling. RT-PCR showed the expression of alpha 3, alpha 4, alpha 7, alpha 9, alpha 10, beta2, and beta 4 nAChR mRNAs in rhesus monkey lung and cultured BECs. The expression of alpha 7, alpha 4, and beta2 nAChR was confirmed by immunofluorescence in the cultured BECs and lung. The electrophysiological characteristics of nAChR in BECs were determined using whole-cell patch-clamp on cultured BECs. Both ACh and nicotine evoked an inward current, with a rapid desensitizing current. Nicotine induced inward currents in a concentration-dependent manner, with an EC(50) of 26.7 microM. Nicotine-induced currents were reversibly blocked by the nicotinic antagonists, mecamylamine, dihydro-beta-erythroidine, and methyllcaconitine. Incubation of BECs with 1 microM nicotine for 48 hours enhanced nicotine-induced currents by roughly 26%. The protein tyrosine phosphorylation inhibitor, genistein, increased nicotine-induced currents by 58% and enhanced methyllcaconitine-sensitive currents (alpha 7 nAChR activities) 2.3-fold, whereas the protein tyrosine phosphatase inhibitor, pervanadate, decreased the effects of nicotine. These results demonstrate that chronic nicotine exposure up-regulates nAChR activity in developing lung, and that nAChR activity can be further modified by tyrosine phosphorylation.

  3. Curcumin Inhibits Prostate Cancer Bone Metastasis by Up-Regulating Bone Morphogenic Protein-7 in Vivo.

    PubMed

    Dorai, Thambi; Diouri, Janane; O'Shea, Orla; Doty, Stephen B

    2014-04-01

    A number of studies have focused on the beneficial properties of Curcumin (diferuloyl methane, used in South Asian cuisine and traditional medicine) such as the chemoprevention of cancer. Recent studies have also indicated that this material has significant benefits for the treatment of cancer and is currently undergoing several clinical trials. We have been interested in the application of this compound as a therapeutic agent for advanced prostate cancer, particularly the skeletal complications in this malignancy. Our earlier work indicated that this compound could inhibit the osteomimetic properties which occur in castration resistant prostate cancer cells, by interfering with the common denominators between these cancer cells and the bone cells in the metastatic tumor microenvironment, namely the osteoblasts and the osteoclast. We predicted that curcumin could break the vicious cycle of reciprocal stimulation that results in uncontrolled osteolysis in the bony matrix. In this work, we have evaluated the potential of this compound in inhibiting the bone metastasis of hormone refractory prostate cancer cells in an established animal model. Our results strongly suggest that curcumin modulates the TGF-β signaling that occurs due to bone matrix degradation by up-regulating the metastasis inhibitory bone morphogenic protein-7 (BMP- 7). This enhancement of BMP-7 in the context of TGF-βin the tumor microenvironment is shown to enhance the mesenchymal-to-epithelial transition. Most importantly, we show that as a result of BMP-7 up-regulation, a novel brown/beige adipogenic differentiation program is also up-regu- lated which plays a role in the inhibition of bone metastasis. Our results suggest that curcumin may subvert the TGF-βsignaling to an alternative adipogenic differentiation program in addition to the previously established interference with the osteomimetic properties, thus inhibiting the bone metastatic processes in a chemopreventive as well as therapeutic

  4. Up-regulation of GLT-1 severely impairs LTD at mossy fibre--CA3 synapses.

    PubMed

    Omrani, Azar; Melone, Marcello; Bellesi, Michele; Safiulina, Victoria; Aida, Tomomi; Tanaka, Kohishi; Cherubini, Enrico; Conti, Fiorenzo

    2009-10-01

    Glutamate transporters are responsible for clearing synaptically released glutamate from the extracellular space. By this action, they maintain low levels of ambient glutamate, thus preventing excitotoxic damage, and contribute to shaping synaptic currents. We show that up-regulation of the glutamate transporter GLT-1 by ceftriaxone severely impaired mGluR-dependent long-term depression (LTD), induced at rat mossy fibre (MF)-CA3 synapses by repetitive stimulation of afferent fibres. This effect involved GLT-1, since LTD was rescued by the selective GLT-1 antagonist dihydrokainate (DHK). DHK per se produced a modest decrease in fEPSP amplitude that rapidly regained control levels after DHK wash out. Moreover, the degree of fEPSP inhibition induced by the low-affinity glutamate receptor antagonist gamma-DGG was similar during basal synaptic transmission but not during LTD, indicating that in ceftriaxone-treated rats LTD induction did not alter synaptic glutamate transient concentration. Furthermore, ceftriaxone-induced GLT-1 up-regulation significantly reduced the magnitude of LTP at MF-CA3 synapses but not at Schaffer collateral-CA1 synapses. Postembedding immunogold studies in rats showed an increased density of gold particles coding for GLT-1a in astrocytic processes and in mossy fibre terminals; in the latter, gold particles were located near and within the active zones. In both CEF-treated and untreated GLT-1 KO mice used for verifying the specificity of immunostaining, the density of gold particles in MF terminals was comparable to background levels. The enhanced expression of GLT-1 at release sites may prevent activation of presynaptic receptors, thus revealing a novel mechanism by which GLT-1 regulates synaptic plasticity in the hippocampus.

  5. Low-Level Laser Irradiation Stimulates Tenocyte Migration with Up-Regulation of Dynamin II Expression

    PubMed Central

    Tsai, Wen-Chung; Hsu, Chih-Chin; Pang, Jong-Hwei S.; Lin, Miao-Sui; Chen, Ying-Hsun; Liang, Fang-Chen

    2012-01-01

    Low-level laser therapy (LLLT) is commonly used to treat sports-related tendinopathy or tendon injury. Tendon healing requires tenocyte migration to the repair site, followed by proliferation and synthesis of the extracellular matrix. This study was designed to determine the effect of laser on tenocyte migration. Furthermore, the correlation between this effect and expression of dynamin 2, a positive regulator of cell motility, was also investigated. Tenocytes intrinsic to rat Achilles tendon were treated with low-level laser (660 nm with energy density at 1.0, 1.5, and 2.0 J/cm2). Tenocyte migration was evaluated by an in vitro wound healing model and by transwell filter migration assay. The messenger RNA (mRNA) and protein expressions of dynamin 2 were determined by reverse transcription/real-time polymerase chain reaction (real-time PCR) and Western blot analysis respectively. Immunofluorescence staining was used to evaluate the dynamin 2 expression in tenocytes. Tenocytes with or without laser irradiation was treated with dynasore, a dynamin competitor and then underwent transwell filter migration assay. In vitro wound model revealed that more tenocytes with laser irradiation migrated across the wound border to the cell-free zone. Transwell filter migration assay confirmed that tenocyte migration was enhanced dose-dependently by laser. Real-time PCR and Western-blot analysis demonstrated that mRNA and protein expressions of dynamin 2 were up-regulated by laser irradiation dose-dependently. Confocal microscopy showed that laser enhanced the expression of dynamin 2 in cytoplasm of tenocytes. The stimulation effect of laser on tenocytes migration was suppressed by dynasore. In conclusion, low-level laser irradiation stimulates tenocyte migration in a process that is mediated by up-regulation of dynamin 2, which can be suppressed by dynasore. PMID:22666495

  6. NOD1 receptor is up-regulated in diabetic human and murine myocardium.

    PubMed

    Prieto, Patricia; Vallejo-Cremades, María Teresa; Benito, Gemma; González-Peramato, Pilar; Francés, Daniel; Agra, Noelia; Terrón, Verónica; Gónzalez-Ramos, Silvia; Delgado, Carmen; Ruiz-Gayo, Mariano; Pacheco, Ivette; Velasco-Martín, Juan P; Regadera, Javier; Martín-Sanz, Paloma; López-Collazo, Eduardo; Boscá, Lisardo; Fernández-Velasco, María

    2014-12-01

    Type 2 diabetes has a complex pathology that involves a chronic inflammatory state. Emerging evidence suggests a link between the innate immune system receptor NOD1 (nucleotide-binding and oligomerization domain 1) and the pathogenesis of diabetes, in monocytes and hepatic and adipose tissues. The aim of the present study was to assess the role of NOD1 in the progression of diabetic cardiomyopathy. We have measured NOD1 protein in cardiac tissue from Type 2 diabetic (db) mice. Heart and isolated cardiomyocytes from db mice revealed a significant increase in NOD1, together with an up-regulation of nuclear factor κB (NF-κB) and increased apoptosis. Heart tissue also exhibited an enhanced expression of pro-inflammatory cytokines. Selective NOD1 activation with C12-γ-D-glutamyl-m-diaminopimelic acid (iEDAP) resulted in an increased NF-κB activation and apoptosis, demonstrating the involvement of NOD1 both in wild-type and db mice. Moreover, HL-1 cardiomyocytes exposed to elevated concentrations of glucose plus palmitate displayed an enhanced NF-κB activity and apoptotic profile, which was prevented by silencing of NOD1 expression. To address this issue in human pathology, NOD1 expression was evaluated in myocardium obtained from patients with Type 2 diabetes (T2DMH) and from normoglycaemic individuals without cardiovascular histories (NH). We have found that NOD1 was expressed in both NH and T2DMH; however, NOD1 expression was significantly pronounced in T2DMH. Furthermore, both the pro-inflammatory cytokine tumour necrosis factor α (TNF-α) and the apoptosis mediator caspase-3 were up-regulated in T2DMH samples. Taken together, our results define an active role for NOD1 in the heightened inflammatory environment associated with both experimental and human diabetic cardiac disease.

  7. Low-level laser irradiation stimulates tenocyte migration with up-regulation of dynamin II expression.

    PubMed

    Tsai, Wen-Chung; Hsu, Chih-Chin; Pang, Jong-Hwei S; Lin, Miao-Sui; Chen, Ying-Hsun; Liang, Fang-Chen

    2012-01-01

    Low-level laser therapy (LLLT) is commonly used to treat sports-related tendinopathy or tendon injury. Tendon healing requires tenocyte migration to the repair site, followed by proliferation and synthesis of the extracellular matrix. This study was designed to determine the effect of laser on tenocyte migration. Furthermore, the correlation between this effect and expression of dynamin 2, a positive regulator of cell motility, was also investigated. Tenocytes intrinsic to rat Achilles tendon were treated with low-level laser (660 nm with energy density at 1.0, 1.5, and 2.0 J/cm(2)). Tenocyte migration was evaluated by an in vitro wound healing model and by transwell filter migration assay. The messenger RNA (mRNA) and protein expressions of dynamin 2 were determined by reverse transcription/real-time polymerase chain reaction (real-time PCR) and Western blot analysis respectively. Immunofluorescence staining was used to evaluate the dynamin 2 expression in tenocytes. Tenocytes with or without laser irradiation was treated with dynasore, a dynamin competitor and then underwent transwell filter migration assay. In vitro wound model revealed that more tenocytes with laser irradiation migrated across the wound border to the cell-free zone. Transwell filter migration assay confirmed that tenocyte migration was enhanced dose-dependently by laser. Real-time PCR and Western-blot analysis demonstrated that mRNA and protein expressions of dynamin 2 were up-regulated by laser irradiation dose-dependently. Confocal microscopy showed that laser enhanced the expression of dynamin 2 in cytoplasm of tenocytes. The stimulation effect of laser on tenocytes migration was suppressed by dynasore. In conclusion, low-level laser irradiation stimulates tenocyte migration in a process that is mediated by up-regulation of dynamin 2, which can be suppressed by dynasore.

  8. Perforin and granzyme B. Cytolytic proteins up-regulated during rejection of rat small intestine allografts.

    PubMed

    McDiarmid, S V; Farmer, D G; Kuniyoshi, J S; Robert, M; Khadavi, A; Shaked, A; Busuttil, R W

    1995-03-15

    Perforin and granzyme B are 2 cytolytic proteins specific to activated killer cells, particularly CTL. We have studied the mRNA expression of these 2 proteins by a reverse transcriptase polymerase chain reaction method in a unidirectional model of rat small intestine transplant rejection. The allograft group consisted of Lewis x Brown Norway F1 donors into Lewis recipients. The isograft controls were Lewis donors into Lewis recipients. Grafts were placed heterotopically and no immunosuppression was given. Five animals in each group were killed at postoperative days (POD) 3, 5, 7, 8, 9, 10, 12, and 14. mRNA was extracted and a semiquantitative reverse transcriptase polymerase chain reaction was performed. For the semiquantitative analysis, we compared scintillation counts from excised bands. Results were expressed as a percent activity compared with beta-actin. From the same tissue samples, a histologic evaluation was made and rejection was graded according to severity. The isograft controls showed no evidence of histologic rejection and a very low expression of mRNA for perforin and granzyme B from POD 3-14. In contrast, the allograft group began to show histologic evidence of mild rejection on POD 5. By day 7, rejection was moderately severe and associated with a significant up-regulation of perforin and granzyme B in the allografts compared with the controls (P < 0.01), which persisted through POD 14. Peak expression for perforin and granzyme B was on POD 10 and 8, respectively. We conclude that the up-regulation of perforin and granzyme B in rat small intestine transplant allografts is a useful marker of clinically important rejection. PMID:7886805

  9. Novel durum wheat genes up-regulated in response to a combination of heat and drought stress.

    PubMed

    Rampino, Patrizia; Mita, Giovanni; Fasano, Pasqua; Borrelli, Grazia Maria; Aprile, Alessio; Dalessandro, Giuseppe; De Bellis, Luigi; Perrotta, Carla

    2012-07-01

    We report the effect of heat, drought and combined stress on the expression of a group of genes that are up-regulated under these conditions in durum wheat (Triticum turgidum subsp. durum) plants. Modulation of gene expression was studied by cDNA-AFLP performed on RNAs extracted from flag leaves. By this approach, we identified several novel durum wheat genes whose expression is modulated under different stress conditions. We focused on a group of hitherto undescribed up-regulated genes in durum wheat, among these, 7 are up-regulated by heat, 8 by drought stress, 15 by combined heat and drought stress, 4 are up-regulated by both heat and combined stress, and 3 by both drought and combined stress. The functional characterization of these genes will provide new data that could help the developing of strategies aimed at improving durum wheat tolerance to field stress.

  10. Insecticide-Mediated Up-Regulation of Cytochrome P450 Genes in the Red Flour Beetle (Tribolium castaneum)

    PubMed Central

    Liang, Xiao; Xiao, Da; He, Yanping; Yao, Jianxiu; Zhu, Guonian; Zhu, Kun Yan

    2015-01-01

    Some cytochrome P450 (CYP) genes are known for their rapid up-regulation in response to insecticide exposures in insects. To date, however, limited information is available with respect to the relationships among the insecticide type, insecticide concentration, exposure duration and the up-regulated CYP genes. In this study, we examined the transcriptional response of eight selected CYP genes, including CYP4G7, CYP4Q4, CYP4BR3, CYP12H1, CYP6BK11, CYP9D4, CYP9Z5 and CYP345A1, to each of four insecticides in the red flour beetle, Tribolium castaneum. Reverse transcription quantitative PCR (RT-qPCR) revealed that CYP4G7 and CYP345A1 can be significantly up-regulated by cypermethrin (1.97- and 2.06-fold, respectively), permethrin (2.00- and 2.03-fold) and lambda-cyhalothrin (1.73- and 1.81-fold), whereas CYP4BR3 and CYP345A1 can be significantly up-regulated by imidacloprid (1.99- and 1.83-fold) when 20-day larvae were exposed to each of these insecticides at the concentration of LC20 for 24 h. Our studies also showed that similar levels of up-regulation can be achieved for CYP4G7, CYP4BR3 and CYP345A1 by cypermethrin, permethrin, lambda-cyhalothrin or imidacloprid with approximately one fourth of LC20 in 6 h. Our study demonstrated that up-regulation of these CYP genes was rapid and only required low concentrations of insecticides, and the up-regulation not only depended on the CYP genes but also the type of insecticides. Our results along with those from previous studies also indicated that there were no specific patterns for predicting the up-regulation of specific CYP gene families based on the insecticide classification. PMID:25607733

  11. Insecticide-mediated up-regulation of cytochrome P450 genes in the red flour beetle (Tribolium castaneum).

    PubMed

    Liang, Xiao; Xiao, Da; He, Yanping; Yao, Jianxiu; Zhu, Guonian; Zhu, Kun Yan

    2015-01-19

    Some cytochrome P450 (CYP) genes are known for their rapid up-regulation in response to insecticide exposures in insects. To date, however, limited information is available with respect to the relationships among the insecticide type, insecticide concentration, exposure duration and the up-regulated CYP genes. In this study, we examined the transcriptional response of eight selected CYP genes, including CYP4G7, CYP4Q4, CYP4BR3, CYP12H1, CYP6BK11, CYP9D4, CYP9Z5 and CYP345A1, to each of four insecticides in the red flour beetle, Tribolium castaneum. Reverse transcription quantitative PCR (RT-qPCR) revealed that CYP4G7 and CYP345A1 can be significantly up-regulated by cypermethrin (1.97- and 2.06-fold, respectively), permethrin (2.00- and 2.03-fold) and lambda-cyhalothrin (1.73- and 1.81-fold), whereas CYP4BR3 and CYP345A1 can be significantly up-regulated by imidacloprid (1.99- and 1.83-fold) when 20-day larvae were exposed to each of these insecticides at the concentration of LC20 for 24 h. Our studies also showed that similar levels of up-regulation can be achieved for CYP4G7, CYP4BR3 and CYP345A1 by cypermethrin, permethrin, lambda-cyhalothrin or imidacloprid with approximately one fourth of LC20 in 6 h. Our study demonstrated that up-regulation of these CYP genes was rapid and only required low concentrations of insecticides, and the up-regulation not only depended on the CYP genes but also the type of insecticides. Our results along with those from previous studies also indicated that there were no specific patterns for predicting the up-regulation of specific CYP gene families based on the insecticide classification.

  12. Uncoupling protein-2 up-regulation and enhanced cyanide toxicity are mediated by PPAR{alpha} activation and oxidative stress

    SciTech Connect

    Zhang, X.; Li, L.; Prabhakaran, K.; Zhang, L.; Leavesley, H.B.; Borowitz, J.L.; Isom, G.E.

    2007-08-15

    Uncoupling protein 2 (UCP-2) is an inner mitochondrial membrane proton carrier that modulates mitochondrial membrane potential ({delta}{psi}{sub m}) and uncouples oxidative phosphorylation. We have shown that up-regulation of UCP-2 by Wy14,643, a selective peroxisome proliferator-activated receptor-{alpha} (PPAR{alpha}) agonist, enhances cyanide cytotoxicity. The pathway by which Wy14,643 up-regulates UCP-2 was determined in a dopaminergic cell line (N27 cells). Since dopaminergic mesencephalic cells are a primary brain target of cyanide, the N27 immortalized mesencephalic cell was used in this study. Wy14,643 produced a concentration- and time-dependent up-regulation of UCP-2 that was linked to enhanced cyanide-induced cell death. MK886 (PPAR{alpha} antagonist) or PPAR{alpha} knock-down by RNA interference (RNAi) inhibited PPAR{alpha} activity as shown by the peroxisome proliferator response element-luciferase reporter assay, but only partially decreased up-regulation of UCP-2. The role of oxidative stress as an alternative pathway to UCP-2 up-regulation was determined. Wy14,643 induced a rapid surge of ROS generation and loading cells with glutathione ethyl ester (GSH-EE) or pre-treatment with vitamin E attenuated up-regulation of UCP-2. On the other hand, RNAi knockdown of PPAR{alpha} did not alter ROS generation, suggesting a PPAR{alpha}-independent component to the response. Co-treatment with PPAR{alpha}-RNAi and GSH-EE blocked both the up-regulation of UCP-2 by Wy14,643 and the cyanide-induced cell death. It was concluded that a PPAR{alpha}-mediated pathway and an oxidative stress pathway independent of PPAR{alpha} mediate the up-regulation of UCP-2 and subsequent increased vulnerability to cyanide-induced cytotoxicity.

  13. NFAT5 Is Up-Regulated by Hypoxia: Possible Implications in Preeclampsia and Intrauterine Growth Restriction.

    PubMed

    Dobierzewska, Aneta; Palominos, Macarena; Irarrazabal, Carlos E; Sanchez, Marianela; Lozano, Mauricio; Perez-Sepulveda, Alejandra; Monteiro, Lara J; Burmeister, Yara; Figueroa-Diesel, Horacio; Rice, Gregory E; Illanes, Sebastian E

    2015-07-01

    During gestation, low oxygen environment is a major determinant of early placentation process, while persistent placental hypoxia leads to pregnancy-related complications such as preeclampsia (PE) and intrauterine growth restriction (IUGR). PE affects 5%-8% of all pregnancies worldwide and is a cause of maternal and fetal morbidity and mortality. During placental development, persistent hypoxia due to poor trophoblast invasion and reduced uteroplacental perfusion leads to maternal endothelial dysfunction and clinical manifestation of PE. Here we hypothesized that nuclear factor of activated T cells-5 (NFAT5), a well-known osmosensitive renal factor and recently characterized hypoxia-inducible protein, is also activated in vivo in placentas of PE and IUGR complications as well as in the in vitro model of trophoblast hypoxia. In JAR cells, low oxygen tension (1% O2) induced NFAT5 mRNA and increased its nuclear abundance, peaking at 16 h. This increase did not occur in parallel with the earlier HIF1A induction. Real-time PCR and Western blot analysis confirmed up-regulation of NFAT5 mRNA and NFAT5 nuclear content in human preeclamptic placentas and in rabbit placentas of an experimentally induced IUGR model, as compared with the control groups. In vitro lambda protein phosphatase (lambda PPase) treatment revealed that increased abundance of NFAT5 protein in nuclei of either JAR cells (16 h of hypoxia) or PE and IUGR placentas is at least partially due to NFAT5 phosphorylation. NFAT5 downstream targets aldose reductase (AR) and sodium-myo-inositol cotransporter (SMIT; official symbol SLC5A3) were not significantly up-regulated either in JAR cells exposed to hypoxia or in placentas of PE- and IUGR-complicated pregnancies, suggesting that hypoxia-dependent activation of NFAT5 serves as a separate function to its tonicity-dependent stimulation. In conclusion, we propose that NFAT5 may serve as a novel marker of placental hypoxia and ischemia independently of HIF1A. PMID

  14. Up-regulation of MicroRNA 146b is Associated with Myelofibrosis in Myeloproliferative Neoplasms.

    PubMed

    Ha, Jung-Sook; Jung, Hye-Ra

    2015-01-01

    In this study, our goal was to evaluate whether the expressions of microRNA (miR)-150, miR-146b, miR-31 and miR-95 demonstrate primary myelofibrosis (PMF) specificity, associations with fibrosis grade, hematologic phenotypes, or myeloproliferative neoplasm (MPN)-associated mutations. A total of 51 formalin-fixed and paraffin-embedded bone marrow MPN samples, including 15 polycythemia vera (PV), 26 essential thrombocythemia (ET), and 10 PMF, and 24 normal controls were included. The expression of microRNA (miRNA) was detected by quantitative real-time polymerase chain reaction using miRNA specific primers. RNU6-2 was analyzed for all samples as endogenous control for relative quantification. Information for fibrosis, hematologic parameters, Janus kinase 2 (JAK2) V617F, and calreticulin (CALR) mutations was obtained from medical records. Significant increment of miR-146b was detected in PMF compared to normal controls (P=0.008). Moreover, expression of miR-146b tended to increase according to increment of fibrosis grade, and patients with myelofibrosis (MF) grade 3 showed significantly higher expression than patients with MF 0 to 2 (P=0.022, 0.001 and 0.013, respectively) or normal controls (P<0.001). The expression of miR-31 also showed tendency to increase following fibrosis and miR-150 showed up-regulated expression in ET (P=0.015) compared to normal control. There was no relationship between miRNA expression and hematologic indices except miR-95 showed negative correlation with platelet count (P=0.024). There was no significant correlation between miRNA expression and JAK2 V617F or CALR mutation. Up-regulation of miR-146b could be used as a fibrosis-indicating marker and might be helpful in the study of fibrotic mechanism in MPN, as well as other fibrotic diseases. PMID:26116595

  15. G-protein Receptor Kinase 5 Regulates the Cannabinoid Receptor 2-induced Up-regulation of Serotonin 2A Receptors*

    PubMed Central

    Franklin, Jade M.; Carrasco, Gonzalo A.

    2013-01-01

    We have recently reported that cannabinoid agonists can up-regulate and enhance the activity of serotonin 2A (5-HT2A) receptors in the prefrontal cortex (PFCx). Increased expression and activity of cortical 5-HT2A receptors has been associated with neuropsychiatric disorders, such as anxiety and schizophrenia. Here we report that repeated CP55940 exposure selectively up-regulates GRK5 proteins in rat PFCx and in a neuronal cell culture model. We sought to examine the mechanism underlying the regulation of GRK5 and to identify the role of GRK5 in the cannabinoid agonist-induced up-regulation and enhanced activity of 5-HT2A receptors. Interestingly, we found that cannabinoid agonist-induced up-regulation of GRK5 involves CB2 receptors, β-arrestin 2, and ERK1/2 signaling because treatment with CB2 shRNA lentiviral particles, β-arrestin 2 shRNA lentiviral particles, or ERK1/2 inhibitor prevented the cannabinoid agonist-induced up-regulation of GRK5. Most importantly, we found that GRK5 shRNA lentiviral particle treatment prevented the cannabinoid agonist-induced up-regulation and enhanced 5-HT2A receptor-mediated calcium release. Repeated cannabinoid exposure was also associated with enhanced phosphorylation of CB2 receptors and increased interaction between β-arrestin 2 and ERK1/2. These latter phenomena were also significantly inhibited by GRK5 shRNA lentiviral treatment. Our results suggest that sustained activation of CB2 receptors, which up-regulates 5-HT2A receptor signaling, enhances GRK5 expression; the phosphorylation of CB2 receptors; and the β-arrestin 2/ERK interactions. These data could provide a rationale for some of the adverse effects associated with repeated cannabinoid agonist exposure. PMID:23592773

  16. A few shared up-regulated genes may influence conidia to yeast transformation in dimorphic fungal pathogens.

    PubMed

    Kirkland, Theo N

    2016-08-01

    The small number of fungi that commonly cause disease in normal people share the capacity to grow as mycelia in the soil at 25°C and as yeast (or spherules) in mammals at 37°C. This remarkable conversion has long been a topic of interest in medical mycology. The conidia to yeast conversion has been studied by transcription profiling in several fungal species, including Histoplasma capsulatum, Paracoccidioides brasiliensis, Coccidioides spp., Blastomyces dermatitidis, and Talaromyces marneffei One limitation of transcriptional profiling is determining which genes are involved in the process of conversion to yeast as opposed to a result of conversion to yeast. If there are genes that are up-regulated in the yeast phase of more than one dimorphic, pathogenic fungus they might be required for conversion to yeast (or spherules). To address this issue, 24 up-regulated genes common to Coccidioides spp spherules and H. capsulatum yeasts were identified. Four homologs of these genes were also found in P. brasiliensis, B. dermatitidis or T. marneffei genes that were up-regulated in yeast. 4-hydroxyphenylpurvate dioxygenase, a gene involved in tyrosine metabolism and melanin synthesis that has been shown to be required for yeast conversion, is conserved and up-regulated in yeast in all five species. Another up-regulated gene that is conserved in all five species is a MFS sugar porter. These results suggest that a minority of up-regulated yeast (or spherule) genes are conserved across species and raises the possibility that conserved up-regulated genes may be of special interest for differentiation of mycelium into yeast.

  17. A few shared up-regulated genes may influence conidia to yeast transformation in dimorphic fungal pathogens.

    PubMed

    Kirkland, Theo N

    2016-08-01

    The small number of fungi that commonly cause disease in normal people share the capacity to grow as mycelia in the soil at 25°C and as yeast (or spherules) in mammals at 37°C. This remarkable conversion has long been a topic of interest in medical mycology. The conidia to yeast conversion has been studied by transcription profiling in several fungal species, including Histoplasma capsulatum, Paracoccidioides brasiliensis, Coccidioides spp., Blastomyces dermatitidis, and Talaromyces marneffei One limitation of transcriptional profiling is determining which genes are involved in the process of conversion to yeast as opposed to a result of conversion to yeast. If there are genes that are up-regulated in the yeast phase of more than one dimorphic, pathogenic fungus they might be required for conversion to yeast (or spherules). To address this issue, 24 up-regulated genes common to Coccidioides spp spherules and H. capsulatum yeasts were identified. Four homologs of these genes were also found in P. brasiliensis, B. dermatitidis or T. marneffei genes that were up-regulated in yeast. 4-hydroxyphenylpurvate dioxygenase, a gene involved in tyrosine metabolism and melanin synthesis that has been shown to be required for yeast conversion, is conserved and up-regulated in yeast in all five species. Another up-regulated gene that is conserved in all five species is a MFS sugar porter. These results suggest that a minority of up-regulated yeast (or spherule) genes are conserved across species and raises the possibility that conserved up-regulated genes may be of special interest for differentiation of mycelium into yeast. PMID:27118798

  18. Compassion-based emotion regulation up-regulates experienced positive affect and associated neural networks

    PubMed Central

    Singer, Tania

    2015-01-01

    Emotion regulation research has primarily focused on techniques that attenuate or modulate the impact of emotional stimuli. Recent evidence suggests that this mode regulation can be problematic in the context of regulation of emotion elicited by the suffering of others, resulting in reduced emotional connectedness. Here, we investigated the effects of an alternative emotion regulation technique based on the up-regulation of positive affect via Compassion-meditation on experiential and neural affective responses to depictions of individuals in distress, and compared these with the established emotion regulation strategy of Reappraisal. Using fMRI, we scanned 15 expert practitioners of Compassion-meditation either passively viewing, or using Compassion-meditation or Reappraisal to modulate their emotional reactions to film clips depicting people in distress. Both strategies effectively, but differentially regulated experienced affect, with Compassion primarily increasing positive and Reappraisal primarily decreasing negative affect. Imaging results showed that Compassion, relative to both passive-viewing and Reappraisal increased activation in regions involved in affiliation, positive affect and reward processing including ventral striatum and medial orbitfrontal cortex. This network was shown to be active prior to stimulus presentation, suggesting that the regulatory mechanism of Compassion is the stimulus-independent endogenous generation of positive affect. PMID:25698699

  19. PPM1D controls nucleolar formation by up-regulating phosphorylation of nucleophosmin.

    PubMed

    Kozakai, Yuuki; Kamada, Rui; Furuta, Junya; Kiyota, Yuhei; Chuman, Yoshiro; Sakaguchi, Kazuyasu

    2016-01-01

    An increase of nucleolar number and size has made nucleoli essential markers for cytology and tumour development. However, the underlying basis for their structural integrity and abundance remains unclear. Protein phosphatase PPM1D was found to be up-regulated in different carcinomas including breast cancers. Here, we demonstrate for the first time that PPM1D regulates nucleolar formation via inducing an increased phosphorylation of the nucleolar protein NPM. We show that PPM1D overexpression induces an increase in the nucleolar number regardless of p53 status. We also demonstrated that specific sequential phosphorylation of NPM is important for nucleolar formation and that PPM1D is a novel upstream regulator of this phosphorylation pathway. These results enhance our understanding of the molecular mechanisms that govern nucleoli formation by demonstrating that PPM1D regulates nucleolar formation by regulating NPM phosphorylation status through a novel signalling pathway, PPM1D-CDC25C-CDK1-PLK1. PMID:27619510

  20. Early up-regulation of chemokine expression in fulminant hepatic failure.

    PubMed

    Leifeld, Ludger; Dumoulin, Franz-Ludwig; Purr, Ingvill; Janberg, Katrin; Trautwein, Christian; Wolff, Martin; Manns, Michael Peter; Sauerbruch, Tilman; Spengler, Ulrich

    2003-03-01

    CC-chemokines recruit and activate macrophages and T lymphocytes, the major components of inflammatory infiltrates in fulminant hepatic failure (FHF). To analyse the role of CC-chemokines in the pathogenesis of FHF, this study examined serum levels and intrahepatic expression of MCP-1, MIP-1alpha, MIP-1beta, and RANTES in the livers and sera of patients with FHF and controls by ELISA, immunohistochemistry, and competitive RT-PCR. Serum levels and intrahepatic expression of all chemokines studied in FHF exceeded the levels in chronic liver diseases and normal controls. Distinct patterns of expression of each chemokine were noted on Kupffer cells, sinusoidal endothelial cells, hepatocytes, lymphocytes, and bile ducts. Intrahepatic chemokine expression correlated closely with the extent of infiltration by macrophages and T lymphocytes (r = 0.65-0.95, p < 0.001). The functional relationship between intrahepatic chemokine release and infiltration was confirmed in chemotaxis assays by inhibiting chemotaxis induced by homogenates of liver tissue obtained from FHF patients with neutralizing MCP-1, MIP-1alpha, MIP-1beta, and RANTES antibodies. The time course of CC-chemokine release was studied in the concanavalin A and the galactosamine/LPS mouse models of FHF. In both models, intrahepatic chemokine up-regulation occurred as an early event prior to hepatic infiltration and liver damage. The data indicate that an abundant intrahepatic release of CC-chemokines is an early and pivotal step in the pathogenesis of FHF.

  1. Hypoxia Up-Regulates Galectin-3 in Mammary Tumor Progression and Metastasis.

    PubMed

    de Oliveira, Joana T; Ribeiro, Cláudia; Barros, Rita; Gomes, Catarina; de Matos, Augusto J; Reis, Celso A; Rutteman, Gerard R; Gärtner, Fátima

    2015-01-01

    The tumor microenvironment encompasses several stressful conditions for cancer cells such as hypoxia, oxidative stress and pH alterations. Galectin-3, a well-studied member of the beta-galactoside-binding animal family of lectins has been implicated in multiple steps of metastasis as cell-cell and cell-ECM adhesion, promotion of angiogenesis, cell proliferation and resistance to apoptosis. However, both its aberrantly up- and down-regulated expression was observed in several types of cancer. Thus, the mechanisms that regulate galectin-3 expression in neoplastic settings are not clear. In order to demonstrate the putative role of hypoxia in regulating galectin-3 expression in canine mammary tumors (CMT), in vitro and in vivo studies were performed. In malignant CMT cells, hypoxia was observed to induce expression of galectin-3, a phenomenon that was almost completely prevented by catalase treatment of CMT-U27 cells. Increased galectin-3 expression was confirmed at the mRNA level. Under hypoxic conditions the expression of galectin-3 shifts from a predominant nuclear location to cytoplasmic and membrane expressions. In in vivo studies, galectin-3 was overexpressed in hypoxic areas of primary tumors and well-established metastases. Tumor hypoxia thus up-regulates the expression of galectin-3, which may in turn increase tumor aggressiveness. PMID:26222311

  2. Compassion-based emotion regulation up-regulates experienced positive affect and associated neural networks.

    PubMed

    Engen, Haakon G; Singer, Tania

    2015-09-01

    Emotion regulation research has primarily focused on techniques that attenuate or modulate the impact of emotional stimuli. Recent evidence suggests that this mode regulation can be problematic in the context of regulation of emotion elicited by the suffering of others, resulting in reduced emotional connectedness. Here, we investigated the effects of an alternative emotion regulation technique based on the up-regulation of positive affect via Compassion-meditation on experiential and neural affective responses to depictions of individuals in distress, and compared these with the established emotion regulation strategy of Reappraisal. Using fMRI, we scanned 15 expert practitioners of Compassion-meditation either passively viewing, or using Compassion-meditation or Reappraisal to modulate their emotional reactions to film clips depicting people in distress. Both strategies effectively, but differentially regulated experienced affect, with Compassion primarily increasing positive and Reappraisal primarily decreasing negative affect. Imaging results showed that Compassion, relative to both passive-viewing and Reappraisal increased activation in regions involved in affiliation, positive affect and reward processing including ventral striatum and medial orbitfrontal cortex. This network was shown to be active prior to stimulus presentation, suggesting that the regulatory mechanism of Compassion is the stimulus-independent endogenous generation of positive affect. PMID:25698699

  3. Compassion-based emotion regulation up-regulates experienced positive affect and associated neural networks.

    PubMed

    Engen, Haakon G; Singer, Tania

    2015-09-01

    Emotion regulation research has primarily focused on techniques that attenuate or modulate the impact of emotional stimuli. Recent evidence suggests that this mode regulation can be problematic in the context of regulation of emotion elicited by the suffering of others, resulting in reduced emotional connectedness. Here, we investigated the effects of an alternative emotion regulation technique based on the up-regulation of positive affect via Compassion-meditation on experiential and neural affective responses to depictions of individuals in distress, and compared these with the established emotion regulation strategy of Reappraisal. Using fMRI, we scanned 15 expert practitioners of Compassion-meditation either passively viewing, or using Compassion-meditation or Reappraisal to modulate their emotional reactions to film clips depicting people in distress. Both strategies effectively, but differentially regulated experienced affect, with Compassion primarily increasing positive and Reappraisal primarily decreasing negative affect. Imaging results showed that Compassion, relative to both passive-viewing and Reappraisal increased activation in regions involved in affiliation, positive affect and reward processing including ventral striatum and medial orbitfrontal cortex. This network was shown to be active prior to stimulus presentation, suggesting that the regulatory mechanism of Compassion is the stimulus-independent endogenous generation of positive affect.

  4. Centenarians, but not octogenarians, up-regulate the expression of microRNAs.

    PubMed

    Serna, Eva; Gambini, Juan; Borras, Consuelo; Abdelaziz, Kheira M; Mohammed, Kheira; Belenguer, Angel; Sanchis, Paula; Avellana, Juan A; Rodriguez-Mañas, Leocadio; Viña, Jose

    2012-01-01

    Centenarians exhibit extreme longevity and a remarkable compression of morbidity. They have a unique capacity to maintain homeostatic mechanisms. Since small non-coding RNAs (including microRNAs) are implicated in the regulation of gene expression, we hypothesised that longevity of centenarians may reflect alterations in small non-coding RNA expression. We report the first comparison of microRNAs expression profiles in mononuclear cells from centenarians, octogenarians and young individuals resident near Valencia, Spain. Principal Component Analysis of the expression of 15,644 mature microRNAs and, 2,334 snoRNAs and scaRNAs in centenarians revealed a significant overlap with profiles in young individuals but not with octogenarians and a significant up-regulation of 7 small non-coding RNAs in centenarians compared to young persons and notably 102 small non-coding RNAs when compared with octogenarians. We suggest that the small non-coding RNAs signature in centenarians may provide insights into the underlying molecular mechanisms endowing centenarians with extreme longevity.

  5. Caffeine Induces the Stress Response and Up-Regulates Heat Shock Proteins in Caenorhabditis elegans.

    PubMed

    Al-Amin, Mohammad; Kawasaki, Ichiro; Gong, Joomi; Shim, Yhong-Hee

    2016-02-01

    Caffeine has both positive and negative effects on physiological functions in a dose-dependent manner. C. elegans has been used as an animal model to investigate the effects of caffeine on development. Caffeine treatment at a high dose (30 mM) showed detrimental effects and caused early larval arrest. We performed a comparative proteomic analysis to investigate the mode of action of high-dose caffeine treatment in C. elegans and found that the stress response proteins, heat shock protein (HSP)-4 (endoplasmic reticulum [ER] chaperone), HSP-6 (mitochondrial chaperone), and HSP-16 (cytosolic chaperone), were induced and their expression was regulated at the transcriptional level. These findings suggest that high-dose caffeine intake causes a strong stress response and activates all three stress-response pathways in the worms, including the ER-, mitochondrial-, and cytosolic pathways. RNA interference of each hsp gene or in triple combination retarded growth. In addition, caffeine treatment stimulated a food-avoidance behavior (aversion phenotype), which was enhanced by RNAi depletion of the hsp-4 gene. Therefore, up-regulation of hsp genes after caffeine treatment appeared to be the major responses to alleviate stress and protect against developmental arrest.

  6. CUL4B promotes replication licensing by up-regulating the CDK2-CDC6 cascade.

    PubMed

    Zou, Yongxin; Mi, Jun; Wang, Wenxing; Lu, Juanjuan; Zhao, Wei; Liu, Zhaojian; Hu, Huili; Yang, Yang; Gao, Xiaoxing; Jiang, Baichun; Shao, Changshun; Gong, Yaoqin

    2013-03-18

    Cullin-RING ubiquitin ligases (CRLs) participate in the regulation of diverse cellular processes including cell cycle progression. Mutations in the X-linked CUL4B, a member of the cullin family, cause mental retardation and other developmental abnormalities in humans. Cells that are deficient in CUL4B are severely selected against in vivo in heterozygotes. Here we report a role of CUL4B in the regulation of replication licensing. Strikingly, CDC6, the licensing factor in replication, was positively regulated by CUL4B and contributed to the loading of MCM2 to chromatin. The positive regulation of CDC6 by CUL4B depends on CDK2, which phosphorylates CDC6, protecting it from APC(CDH1)-mediated degradation. Thus, aside being required for cell cycle reentry from quiescence, CDK2 also contributes to pre-replication complex assembly in G1 phase of cycling cells. Interestingly, the up-regulation of CDK2 by CUL4B is achieved via the repression of miR-372 and miR-373, which target CDK2. Our findings thus establish a CUL4B-CDK2-CDC6 cascade in the regulation of DNA replication licensing.

  7. MicroRNA-276 promotes egg-hatching synchrony by up-regulating brm in locusts.

    PubMed

    He, Jing; Chen, Qianquan; Wei, Yuanyuan; Jiang, Feng; Yang, Meiling; Hao, Shuguang; Guo, Xiaojiao; Chen, Dahua; Kang, Le

    2016-01-19

    Developmental synchrony, the basis of uniform swarming, migration, and sexual maturation, is an important strategy for social animals to adapt to variable environments. However, the molecular mechanisms underlying developmental synchrony are largely unexplored. The migratory locust exhibits polyphenism between gregarious and solitarious individuals, with the former displaying more synchronous sexual maturation and migration than the latter. Here, we found that the egg-hatching time of gregarious locusts was more uniform compared with solitarious locusts and that microRNA-276 (miR-276) was expressed significantly higher in both ovaries and eggs of gregarious locusts than in solitarious locusts. Interestingly, inhibiting miR-276 in gregarious females and overexpressing it in solitarious females, respectively, caused more heterochronic and synchronous hatching of progeny eggs. Moreover, miR-276 directly targeted a transcription coactivator gene, brahma (brm), resulting in its up-regulation. Knockdown of brm not only resulted in asynchronous egg hatching in gregarious locusts but also impaired the miR-276-induced synchronous egg hatching in solitarious locusts. Mechanistically, miR-276 mediated brm activation in a manner that depended on the secondary structure of brm, namely, a stem-loop around the binding site of miR-276. Collectively, our results unravel a mechanism by which miR-276 enhances brm expression to promote developmental synchrony and provide insight into regulation of developmental homeostasis and population sustaining that are closely related to biological synchrony. PMID:26729868

  8. Up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver

    PubMed Central

    Zhang, Shuai; Li, Tao-Sheng; Soyama, Akihiko; Tanaka, Takayuki; Yan, Chen; Sakai, Yusuke; Hidaka, Masaaki; Kinoshita, Ayaka; Natsuda, Koji; Fujii, Mio; Kugiyama, Tota; Baimakhanov, Zhassulan; Kuroki, Tamotsu; Gu, Weili; Eguchi, Susumu

    2016-01-01

    Although the healthy liver is known to have high regenerative potential, poor liver regeneration under pathological conditions remains a substantial problem. We investigated the key molecules that impair the regeneration of cholestatic liver. C57BL/6 mice were randomly subjected to partial hepatectomy and bile duct ligation (PH+BDL group, n = 16), partial hepatectomy only (PH group, n = 16), or sham operation (Sham group, n = 16). The liver sizes and histological findings were similar in the PH and sham groups 14 days after operation. However, compared with those in the sham group, the livers in mice in the PH+BDL group had a smaller size, a lower cell proliferative activity, and more fibrotic tissue 14 days after the operation, suggesting the insufficient regeneration of the cholestatic liver. Pathway-focused array analysis showed that many genes were up- or down-regulated over 1.5-fold in both PH+BDL and PH groups at 1, 3, 7, and 14 days after treatment. Interestingly, more genes that were functionally related to the extracellular matrix and inflammatory chemokines were found in the PH+BDL group than in the PH group at 7 and 14 days after treatment. Our data suggest that up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver. PMID:27226149

  9. Hypoxia Induces Autophagy through Translational Up-Regulation of Lysosomal Proteins in Human Colon Cancer Cells

    PubMed Central

    Lai, Ming-Chih; Chang, Chiao-May; Sun, H. Sunny

    2016-01-01

    Hypoxia occurs in a wide variety of physiological and pathological conditions, including tumorigenesis. Tumor cells have to adapt to hypoxia by altering their gene expression and protein synthesis. Here, we showed that hypoxia inhibits translation through activation of PERK and inactivation of mTOR in human colon cancer HCT116 cells. Prolonged hypoxia (1% O2, 16 h) dramatically inhibits general translation in HCT116 cells, yet selected mRNAs remain efficiently translated under such a condition. Using microarray analysis of polysome- associated mRNAs, we identified a large number of hypoxia-regulated genes at the translational level. Efficiently translated mRNAs during hypoxia were validated by polysome profiling and quantitative real-time RT-PCR. Pathway enrichment analysis showed that many of the up-regulated genes are involved in lysosome, glycan and lipid metabolism, antigen presentation, cell adhesion, and remodeling of the extracellular matrix and cytoskeleton. The majority of down-regulated genes are involved in apoptosis, ubiquitin-mediated proteolysis, and oxidative phosphorylation. Further investigation showed that hypoxia induces lysosomal autophagy and mitochondrial dysfunction through translational regulation in HCT116 cells. The abundance of several translation factors and the mTOR kinase activity are involved in hypoxia-induced mitochondrial autophagy in HCT116 cells. Our studies highlight the importance of translational regulation for tumor cell adaptation to hypoxia. PMID:27078027

  10. Two-Dimensional Magnesium Phosphate Nanosheets Form Highly Thixotropic Gels That Up-Regulate Bone Formation.

    PubMed

    Laurenti, Marco; Al Subaie, Ahmed; Abdallah, Mohamed-Nur; Cortes, Arthur R G; Ackerman, Jerome L; Vali, Hojatollah; Basu, Kaustuv; Zhang, Yu Ling; Murshed, Monzur; Strandman, Satu; Zhu, Julian; Makhoul, Nicholas; Barralet, Jake E; Tamimi, Faleh

    2016-08-10

    Hydrogels composed of two-dimensional (2D) nanomaterials have become an important alternative to replace traditional inorganic scaffolds for tissue engineering. Here, we describe a novel nanocrystalline material with 2D morphology that was synthesized by tuning the crystallization of the sodium-magnesium-phosphate system. We discovered that the sodium ion can regulate the precipitation of magnesium phosphate by interacting with the crystal's surface causing a preferential crystal growth that results in 2D morphology. The 2D nanomaterial gave rise to a physical hydrogel that presented extreme thixotropy, injectability, biocompatibility, bioresorption, and long-term stability. The nanocrystalline material was characterized in vitro and in vivo and we discovered that it presented unique biological properties. Magnesium phosphate nanosheets accelerated bone healing and osseointegration by enhancing collagen formation, osteoblasts differentiation, and osteoclasts proliferation through up-regulation of COL1A1, RunX2, ALP, OCN, and OPN. In summary, the 2D magnesium phosphate nanosheets could bring a paradigm shift in the field of minimally invasive orthopedic and craniofacial interventions because it is the only material available that can be injected through high gauge needles into bone defects in order to accelerate bone healing and osseointegration. PMID:27280476

  11. MicroRNA-276 promotes egg-hatching synchrony by up-regulating brm in locusts

    PubMed Central

    He, Jing; Chen, Qianquan; Wei, Yuanyuan; Jiang, Feng; Yang, Meiling; Hao, Shuguang; Guo, Xiaojiao; Chen, Dahua; Kang, Le

    2016-01-01

    Developmental synchrony, the basis of uniform swarming, migration, and sexual maturation, is an important strategy for social animals to adapt to variable environments. However, the molecular mechanisms underlying developmental synchrony are largely unexplored. The migratory locust exhibits polyphenism between gregarious and solitarious individuals, with the former displaying more synchronous sexual maturation and migration than the latter. Here, we found that the egg-hatching time of gregarious locusts was more uniform compared with solitarious locusts and that microRNA-276 (miR-276) was expressed significantly higher in both ovaries and eggs of gregarious locusts than in solitarious locusts. Interestingly, inhibiting miR-276 in gregarious females and overexpressing it in solitarious females, respectively, caused more heterochronic and synchronous hatching of progeny eggs. Moreover, miR-276 directly targeted a transcription coactivator gene, brahma (brm), resulting in its up-regulation. Knockdown of brm not only resulted in asynchronous egg hatching in gregarious locusts but also impaired the miR-276–induced synchronous egg hatching in solitarious locusts. Mechanistically, miR-276 mediated brm activation in a manner that depended on the secondary structure of brm, namely, a stem-loop around the binding site of miR-276. Collectively, our results unravel a mechanism by which miR-276 enhances brm expression to promote developmental synchrony and provide insight into regulation of developmental homeostasis and population sustaining that are closely related to biological synchrony. PMID:26729868

  12. PPM1D controls nucleolar formation by up-regulating phosphorylation of nucleophosmin

    PubMed Central

    Kozakai, Yuuki; Kamada, Rui; Furuta, Junya; Kiyota, Yuhei; Chuman, Yoshiro; Sakaguchi, Kazuyasu

    2016-01-01

    An increase of nucleolar number and size has made nucleoli essential markers for cytology and tumour development. However, the underlying basis for their structural integrity and abundance remains unclear. Protein phosphatase PPM1D was found to be up-regulated in different carcinomas including breast cancers. Here, we demonstrate for the first time that PPM1D regulates nucleolar formation via inducing an increased phosphorylation of the nucleolar protein NPM. We show that PPM1D overexpression induces an increase in the nucleolar number regardless of p53 status. We also demonstrated that specific sequential phosphorylation of NPM is important for nucleolar formation and that PPM1D is a novel upstream regulator of this phosphorylation pathway. These results enhance our understanding of the molecular mechanisms that govern nucleoli formation by demonstrating that PPM1D regulates nucleolar formation by regulating NPM phosphorylation status through a novel signalling pathway, PPM1D-CDC25C-CDK1-PLK1. PMID:27619510

  13. Amyotrophic lateral sclerosis and denervation alter sphingolipids and up-regulate glucosylceramide synthase

    PubMed Central

    Henriques, Alexandre; Croixmarie, Vincent; Priestman, David A.; Rosenbohm, Angela; Dirrig-Grosch, Sylvie; D'Ambra, Eleonora; Huebecker, Mylene; Hussain, Ghulam; Boursier-Neyret, Claire; Echaniz-Laguna, Andoni; Ludolph, Albert C.; Platt, Frances M.; Walther, Bernard; Spedding, Michael; Loeffler, Jean-Philippe; Gonzalez De Aguilar, Jose-Luis

    2015-01-01

    Amyotrophic lateral sclerosis (ALS) is a fatal adult-onset disease characterized by upper and lower motor neuron degeneration, muscle wasting and paralysis. Growing evidence suggests a link between changes in lipid metabolism and ALS. Here, we used UPLC/TOF-MS to survey the lipidome in SOD1(G86R) mice, a model of ALS. Significant changes in lipid expression were evident in spinal cord and skeletal muscle before overt neuropathology. In silico analysis also revealed appreciable changes in sphingolipids including ceramides and glucosylceramides (GlcCer). HPLC analysis showed increased amounts of GlcCer and downstream glycosphingolipids (GSLs) in SOD1(G86R) muscle compared with wild-type littermates. Glucosylceramide synthase (GCS), the enzyme responsible for GlcCer biosynthesis, was up-regulated in muscle of SOD1(G86R) mice and ALS patients, and in muscle of wild-type mice after surgically induced denervation. Conversely, inhibition of GCS in wild-type mice, following transient peripheral nerve injury, reversed the overexpression of genes in muscle involved in oxidative metabolism and delayed motor recovery. GCS inhibition in SOD1(G86R) mice also affected the expression of metabolic genes and induced a loss of muscle strength and morphological deterioration of the motor endplates. These findings suggest that GSLs may play a critical role in ALS muscle pathology and could lead to the identification of new therapeutic targets. PMID:26483191

  14. Hypoxia Up-Regulates Galectin-3 in Mammary Tumor Progression and Metastasis

    PubMed Central

    Barros, Rita; Gomes, Catarina; de Matos, Augusto J.; Reis, Celso A.; Rutteman, Gerard R.; Gärtner, Fátima

    2015-01-01

    The tumor microenvironment encompasses several stressful conditions for cancer cells such as hypoxia, oxidative stress and pH alterations. Galectin-3, a well-studied member of the beta-galactoside-binding animal family of lectins has been implicated in multiple steps of metastasis as cell-cell and cell-ECM adhesion, promotion of angiogenesis, cell proliferation and resistance to apoptosis. However, both its aberrantly up- and down-regulated expression was observed in several types of cancer. Thus, the mechanisms that regulate galectin-3 expression in neoplastic settings are not clear. In order to demonstrate the putative role of hypoxia in regulating galectin-3 expression in canine mammary tumors (CMT), in vitro and in vivo studies were performed. In malignant CMT cells, hypoxia was observed to induce expression of galectin-3, a phenomenon that was almost completely prevented by catalase treatment of CMT-U27 cells. Increased galectin-3 expression was confirmed at the mRNA level. Under hypoxic conditions the expression of galectin-3 shifts from a predominant nuclear location to cytoplasmic and membrane expressions. In in vivo studies, galectin-3 was overexpressed in hypoxic areas of primary tumors and well-established metastases. Tumor hypoxia thus up-regulates the expression of galectin-3, which may in turn increase tumor aggressiveness. PMID:26222311

  15. Up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver.

    PubMed

    Zhang, Shuai; Li, Tao-Sheng; Soyama, Akihiko; Tanaka, Takayuki; Yan, Chen; Sakai, Yusuke; Hidaka, Masaaki; Kinoshita, Ayaka; Natsuda, Koji; Fujii, Mio; Kugiyama, Tota; Baimakhanov, Zhassulan; Kuroki, Tamotsu; Gu, Weili; Eguchi, Susumu

    2016-01-01

    Although the healthy liver is known to have high regenerative potential, poor liver regeneration under pathological conditions remains a substantial problem. We investigated the key molecules that impair the regeneration of cholestatic liver. C57BL/6 mice were randomly subjected to partial hepatectomy and bile duct ligation (PH+BDL group, n = 16), partial hepatectomy only (PH group, n = 16), or sham operation (Sham group, n = 16). The liver sizes and histological findings were similar in the PH and sham groups 14 days after operation. However, compared with those in the sham group, the livers in mice in the PH+BDL group had a smaller size, a lower cell proliferative activity, and more fibrotic tissue 14 days after the operation, suggesting the insufficient regeneration of the cholestatic liver. Pathway-focused array analysis showed that many genes were up- or down-regulated over 1.5-fold in both PH+BDL and PH groups at 1, 3, 7, and 14 days after treatment. Interestingly, more genes that were functionally related to the extracellular matrix and inflammatory chemokines were found in the PH+BDL group than in the PH group at 7 and 14 days after treatment. Our data suggest that up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver. PMID:27226149

  16. Up-regulation of steroid biosynthesis by retinoid signaling: Implications for aging

    PubMed Central

    Manna, Pulak R.; Stetson, Cloyce L.; Daugherty, Carol; Shimizu, Ikue; Syapin, Peter J.; Garrel, Ghislaine; Cohen-Tannoudji, Joelle; Huhtaniemi, Ilpo; Slominski, Andrzej T.; Pruitt, Kevin; Stocco, Douglas M.

    2015-01-01

    Retinoids (vitamin A and its derivatives) are critical for a spectrum of developmental and physiological processes, in which steroid hormones also play indispensable roles. The StAR protein predominantly regulates steroid biosynthesis in steroidogenic tissues. We reported that regulation of retinoid, especially atRA and 9-cis RA, responsive StAR transcription is largely mediated by an LXR-RXR/RAR heterodimeric motif in the mouse StAR promoter. Herein we demonstrate that retinoids are capable of enhancing StAR protein, P-StAR, and steroid production, in granulosa, adrenocortical, glial, and epidermal cells. Whereas transient expression of RARα and RXRα enhanced 9-cis RA-treated StAR gene transcription, silencing of RXRα with siRNA, decreased StAR and steroid levels. An oligonucleotide probe encompassing an LXR-RXR/RAR motif bound to adrenocortical and epidermal keratinocyte NEs in EMSAs. ChIP studies revealed association of RARα and RXRα with the StAR proximal promoter. Further studies demonstrated that StAR mRNA levels decreased in diseased and elderly men and women skin tissues and that atRA could restore steroidogenesis in epidermal keratinocytes of aged individuals. These findings provide novel insights into the relevance of retinoid signaling in the up-regulation of steroid biosynthesis in various target tissues, and indicate that retinoid therapy may have important implications in age-related complications and diseases. PMID:26303142

  17. Top-down and bottom-up regulation of macroalgal community structure on a Kenyan reef

    NASA Astrophysics Data System (ADS)

    Mörk, Erik; Sjöö, Gustaf Lilliesköld; Kautsky, Nils; McClanahan, Tim R.

    2009-09-01

    Top-down and bottom-up regulation in the form of grazing by herbivores and nutrient availability are important factors governing macroalgal communities in the coral reef ecosystem. Today, anthropogenic activities, such as over-harvesting of herbivorous fish and sea urchins and increased nutrient loading, are altering the interaction of these two structuring forces. The present study was conducted in Kenya and investigates the relative importance of herbivory and nutrient loading on macroalgal community dynamics, by looking at alterations in macroalgal functional groups, species diversity ( H') and biomass within experimental quadrats. The experiment was conducted in situ for 42 days during the dry season. Cages excluding large herbivorous fish and sea urchins were used in the study and nutrient addition was conducted using coated, slow-release fertilizer (nitrogen and phosphorous) at a site where herbivory is generally low and nutrient levels are relatively high for the region. Nutrient addition increased tissue nutrient content in the algae, and fertilized quadrats had 24% higher species diversity. Herbivore exclusion resulted in a 77% increase in algal biomass, mainly attributable to a >1000% increase in corticated forms. These results are in accordance with similar studies in other regions, but are unique in that they indicate that, even when prevailing nutrient levels are relatively high and herbivore pressure is relatively low, continued anthropogenic disturbance results in further ecological responses and increased reef degradation.

  18. MiR-21 is up-regulated in psoriasis and suppresses T cell apoptosis.

    PubMed

    Meisgen, Florian; Xu, Ning; Wei, Tianling; Janson, Peter C; Obad, Susanna; Broom, Oliver; Nagy, Nikoletta; Kauppinen, Sakari; Kemény, Lajos; Ståhle, Mona; Pivarcsi, Andor; Sonkoly, Enikö

    2012-04-01

    MicroRNAs are short non-coding RNAs that regulate gene expression. Previously, in a genome-wide screen, we found deregulation of microRNA expression in psoriasis skin. MicroRNA-21 (miR-21) is one of the microRNAs significantly up-regulated in psoriasis skin lesions. To identify the cell type responsible for the increased miR-21 level, we compared expression of miR-21 in epidermal cells and dermal T cells between psoriasis and healthy skin and found elevated levels of miR-21 in psoriasis in both cell types. In cultured T cells, expression of miR-21 increased markedly upon activation. To explore the function of miR-21 in primary human T helper cells, we inhibited miR-21 using a tiny seed-targeting LNA-anti-miR. Specific inhibition of miR-21 increased the apoptosis rate of activated T cells. Our results suggest that miR-21 suppresses apoptosis in activated T cells, and thus, overexpression of miR-21 may contribute to T cell-derived psoriatic skin inflammation.

  19. Up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver.

    PubMed

    Zhang, Shuai; Li, Tao-Sheng; Soyama, Akihiko; Tanaka, Takayuki; Yan, Chen; Sakai, Yusuke; Hidaka, Masaaki; Kinoshita, Ayaka; Natsuda, Koji; Fujii, Mio; Kugiyama, Tota; Baimakhanov, Zhassulan; Kuroki, Tamotsu; Gu, Weili; Eguchi, Susumu

    2016-01-01

    Although the healthy liver is known to have high regenerative potential, poor liver regeneration under pathological conditions remains a substantial problem. We investigated the key molecules that impair the regeneration of cholestatic liver. C57BL/6 mice were randomly subjected to partial hepatectomy and bile duct ligation (PH+BDL group, n = 16), partial hepatectomy only (PH group, n = 16), or sham operation (Sham group, n = 16). The liver sizes and histological findings were similar in the PH and sham groups 14 days after operation. However, compared with those in the sham group, the livers in mice in the PH+BDL group had a smaller size, a lower cell proliferative activity, and more fibrotic tissue 14 days after the operation, suggesting the insufficient regeneration of the cholestatic liver. Pathway-focused array analysis showed that many genes were up- or down-regulated over 1.5-fold in both PH+BDL and PH groups at 1, 3, 7, and 14 days after treatment. Interestingly, more genes that were functionally related to the extracellular matrix and inflammatory chemokines were found in the PH+BDL group than in the PH group at 7 and 14 days after treatment. Our data suggest that up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver.

  20. The yeast PNC1 longevity gene is up-regulated by mRNA mistranslation.

    PubMed

    Silva, Raquel M; Duarte, Iven C N; Paredes, João A; Lima-Costa, Tatiana; Perrot, Michel; Boucherie, Hélian; Goodfellow, Brian J; Gomes, Ana C; Mateus, Denisa D; Moura, Gabriela R; Santos, Manuel A S

    2009-01-01

    Translation fidelity is critical for protein synthesis and to ensure correct cell functioning. Mutations in the protein synthesis machinery or environmental factors that increase synthesis of mistranslated proteins result in cell death and degeneration and are associated with neurodegenerative diseases, cancer and with an increasing number of mitochondrial disorders. Remarkably, mRNA mistranslation plays critical roles in the evolution of the genetic code, can be beneficial under stress conditions in yeast and in Escherichia coli and is an important source of peptides for MHC class I complex in dendritic cells. Despite this, its biology has been overlooked over the years due to technical difficulties in its detection and quantification. In order to shed new light on the biological relevance of mistranslation we have generated codon misreading in Saccharomyces cerevisiae using drugs and tRNA engineering methodologies. Surprisingly, such mistranslation up-regulated the longevity gene PNC1. Similar results were also obtained in cells grown in the presence of amino acid analogues that promote protein misfolding. The overall data showed that PNC1 is a biomarker of mRNA mistranslation and protein misfolding and that PNC1-GFP fusions can be used to monitor these two important biological phenomena in vivo in an easy manner, thus opening new avenues to understand their biological relevance.

  1. HPV E1 up-regulates replication-related biochemistries of AAV Rep78.

    PubMed

    Bandyopadhyay, Sarmistha; Cao, Maohua; Liu, Yong; Hermonat, Paul L

    2010-06-20

    Human papillomavirus type 16 (HPV) E1 protein provides helper function for the adeno-associated virus type 2 (AAV) life cycle. E1 is the replication protein of HPV, analogous to AAV Rep78, but without the endonuclease/covalent attachment activity of Rep78. Previously we have shown that E1 and Rep78 interact in vitro. Here we investigated E1's effects on Rep78 interaction with AAV's inverted terminal repeat (ITR) DNA in vitro, using purified Rep78 and E1 proteins from bacteria. E1 enhanced Rep78-ITR binding, ATPase activity, Rep78-ITR-covalent linkage and Rep78-ITR-endonuclease activity (central to AAV replication). These enhancements occurred in a dose-dependent manner whenever assayed. However, overall Rep78-plus-E1 helicase activity was lower than Rep78's helicase activity. These data suggest that E1's broad-based helper function for the AAV life cycle (AAV DNA, mRNA, and protein levels are up-regulated by E1) is likely through its ability to enhance Rep78's critical replication-required biochemistries on ITR DNA.

  2. Bovine herpesvirus type 1 (BHV-1) up-regulates telomerase activity in MDBK cells.

    PubMed

    Pagnini, U; De Martino, L; Montagnaro, S; Diodato, A; Longo, M; Pacelli, F; Pisanelli, G; Iovane, G

    2006-03-31

    The proliferative capacity of mammalian cells is regulated by telomerase, an enzyme uniquely specialised for telomeric DNA synthesis. The critical role of telomerase activation in tumor progression and maintenance has been well established in studies of cancer and of oncogenic transformation in cell culture. Experimental data suggest that telomerase activation has an important role in normal somatic cells, and that failure to activate sufficient telomerase also promotes disease. Evidence regarding the role of telomerase in the pathogenesis of several viruses including human immunodeficiency virus has led to an increased interest in the role of telomerase activity in other virus infections. In this research we evaluated the telomerase modulating activity of Bovine herpesvirus 1 (BHV-1) in MDBK cells. MDBK cells were infected at different multiplicity of infection with BHV-1 Cooper strain and telomerase activity at different times post-infection was measured by the TRAP assay. Our data indicate that BHV-1 significantly up-regulates telomerase activity at 3 and 6h post-infection decreasing after the 24h post-infection. Our data, showed that the effect was mediated by an immediate-early or early viral gene, and use of the protein translation inhibitor cycloheximide confirmed that an immediate early gene is primarily responsible.

  3. Impaired endothelial shear stress induces podosome assembly via VEGF up-regulation.

    PubMed

    Fey, Theres; Schubert, Kai Michael; Schneider, Holger; Fein, Evelyn; Kleinert, Eike; Pohl, Ulrich; Dendorfer, Andreas

    2016-08-01

    Podosomes are dynamic cytoskeletal membrane structures with local adhesive and proteolytic activity. They are critically involved in angiogenesis and vascular adaptive growth. Here, we studied in HUVECs and murine small vessels whether shear stress controls podosome assembly and local proteolytic activity. Podosomes were characterized by immunohistochemistry, and their proteolytic activity was assessed as degradation imprints in fluorescent gelatin that was used as growth substrate. Compared with controls (10 dyn/cm(2)), the number of podosomes formed per time was doubled when cells were exposed to low shear stress (0.3 dyn/cm(2)) or even increased 5-fold under static conditions. This was a result of an enhanced expression of VEGF after reduction of shear stress. Consequently, enhanced podosome formation could be prevented by a VEGF receptor antagonist as well by interruption of VEGF signaling via inhibition of PI3K, Src, or p38. Increase of podosome assembly went along with significantly augmented cell motility. In vivo experiments in mouse arteries confirmed increased endothelial podosome numbers when shear stress was abolished by vessel occlusion. We conclude that shear stress, by reducing VEGF release, inhibits podosome assembly. Hence, endothelial cell-mediated matrix proteolysis and migratory activity are inhibited, thereby stabilizing the structure of the vessel wall.-Fey, T., Schubert, K. M., Schneider, H., Fein, E., Kleinert, E., Pohl, U., Dendorfer, A. Impaired endothelial shear stress induces podosome assembly via VEGF up-regulation.

  4. Rck1 up-regulates pseudohyphal growth by activating the Ras2 and MAP kinase pathways independently in Saccharomyces cerevisiae.

    PubMed

    Chang, Miwha; Kang, Chang-Min; Park, Yong-Sung; Yun, Cheol-Won

    2014-02-21

    Previously, we reported that Rck1 regulates Hog1 and Slt2 activities and affects MAP kinase activity in Saccharomyces cerevisiae. Recently, we found that Rck1 up-regulates phospho-Kss1 and phospho-Fus3. Kss1 has been known as a component in the pseudohyphal growth pathway, and we attempted to identify the function of Rck1 in pseudohyphal growth. Rck1 up-regulated Ras2 at the protein level, not the transcriptional level. Additionally, FLO11 transcription was up-regulated by RCK1 over-expression. RCK1 expression was up-regulated during growth on SLAD+1% butanol medium. On nitrogen starvation agar plates, RCK1 over-expression induced pseudohyphal growth of colonies, and cells over-expressing RCK1 showed a filamentous morphology when grown in SLAD medium. Furthermore, 1-butanol greatly induced filamentous growth when RCK1 was over-expressed. Moreover, invasive growth was activated in haploid cells when RCK1 was over-expressed. The growth defect of cells observed on 1-butanol medium was recovered when RCK1 was over-expressed. Interestingly, Ras2 and phospho-Kss1 were up-regulated by Rck1 independently. Together, these results suggest that Rck1 promotes pseudohyphal growth by activating Ras2 and Kss1 via independent pathways in S. cerevisiae. PMID:24491552

  5. Molecular characterization of Quercus suber MYB1, a transcription factor up-regulated in cork tissues.

    PubMed

    Almeida, Tânia; Menéndez, Esther; Capote, Tiago; Ribeiro, Teresa; Santos, Conceição; Gonçalves, Sónia

    2013-01-15

    The molecular processes associated with cork development in Quercus suber L. are poorly understood. A previous molecular approach identified a list of genes potentially important for cork formation and differentiation, providing a new basis for further molecular studies. This report is the first molecular characterization of one of these candidate genes, QsMYB1, coding for an R2R3-MYB transcription factor. The R2R3-MYB gene sub-family has been described as being involved in the phenylpropanoid and lignin pathways, both involved in cork biosynthesis. The results showed that the expression of QsMYB1 is putatively mediated by an alternative splicing (AS) mechanism that originates two different transcripts (QsMYB1.1 and QsMYB1.2), differing only in the 5'-untranslated region, due to retention of the first intron in one of the variants. Moreover, within the retained intron, a simple sequence repeat (SSR) was identified. The upstream regulatory region of QsMYB1 was extended by a genome walking approach, which allowed the identification of the putative gene promoter region. The relative expression pattern of QsMYB1 transcripts determined by reverse transcription quantitative polymerase chain reaction (RT-qPCR) revealed that both transcripts were up-regulated in cork tissues; the detected expression was several times higher in newly formed cork harvested from trees producing virgin, second or reproduction cork when compared with wood. Moreover, the expression analysis of QsMYB1 in several Q. suber organs showed very low expression in young branches and roots, whereas in leaves, immature acorns or male flowers, no expression was detected. These preliminary results suggest that QsMYB1 may be related to secondary growth and, in particular, with the cork biosynthesis process with a possible alternative splicing mechanism associated with its regulatory function.

  6. The Wnt Inhibitor Sclerostin Is Up-regulated by Mechanical Unloading in Osteocytes in Vitro.

    PubMed

    Spatz, Jordan M; Wein, Marc N; Gooi, Jonathan H; Qu, Yili; Garr, Jenna L; Liu, Shawn; Barry, Kevin J; Uda, Yuhei; Lai, Forest; Dedic, Christopher; Balcells-Camps, Mercedes; Kronenberg, Henry M; Babij, Philip; Pajevic, Paola Divieti

    2015-07-01

    Although bone responds to its mechanical environment, the cellular and molecular mechanisms underlying the response of the skeleton to mechanical unloading are not completely understood. Osteocytes are the most abundant but least understood cells in bones and are thought to be responsible for sensing stresses and strains in bone. Sclerostin, a product of the SOST gene, is produced postnatally primarily by osteocytes and is a negative regulator of bone formation. Recent studies show that SOST is mechanically regulated at both the mRNA and protein levels. During prolonged bed rest and immobilization, circulating sclerostin increases both in humans and in animal models, and its increase is associated with a decrease in parathyroid hormone. To investigate whether SOST/sclerostin up-regulation in mechanical unloading is a cell-autonomous response or a hormonal response to decreased parathyroid hormone levels, we subjected osteocytes to an in vitro unloading environment achieved by the NASA rotating wall vessel system. To perform these studies, we generated a novel osteocytic cell line (Ocy454) that produces high levels of SOST/sclerostin at early time points and in the absence of differentiation factors. Importantly, these osteocytes recapitulated the in vivo response to mechanical unloading with increased expression of SOST (3.4 ± 1.9-fold, p < 0.001), sclerostin (4.7 ± 0.1-fold, p < 0.001), and the receptor activator of nuclear factor κΒ ligand (RANKL)/osteoprotegerin (OPG) (2.5 ± 0.7-fold, p < 0.001) ratio. These data demonstrate for the first time a cell-autonomous increase in SOST/sclerostin and RANKL/OPG ratio in the setting of unloading. Thus, targeted osteocyte therapies could hold promise as novel osteoporosis and disuse-induced bone loss treatments by directly modulating the mechanosensing cells in bone.

  7. Up-regulation of IL-12 in monocytes: a fundamental defect in common variable immunodeficiency.

    PubMed

    Cambronero, R; Sewell, W A; North, M E; Webster, A D; Farrant, J

    2000-01-01

    We show that LPS-stimulated circulating CD14-positive monocytes from patients with common variable immunodeficiency (CVID) express a higher proportion of intracellular IL-12-positive cells than monocytes from patients with X-linked agammaglobulinemia or normal subjects. We used four-color flow cytometry and measured IL-12 with an Ab to the p40 subunit following stimulation with LPS. The raised IL-12 is associated with an increased frequency of IFN-gamma-positive T cells, but not of IFN-gamma-positive CD56+ NK cells. These increases in frequency of cytokine-positive cells are due to a decrease in the absolute numbers of circulating monocytes and T cells that are negative for IL-12 and IFN-gamma, respectively. The increased frequency of IL-12-positive monocytes appears to be selective because TNF-alpha was not increased, and is thus unlikely to reflect a general activation. Chronic infection is also unlikely to explain our data since cells from X-linked agammaglobulinemia patients with a similar Ig deficiency do not show these changes. Our data suggest a fundamental abnormality in the IL-12/IFN-gamma circuit in CVID, with up-regulation of IL-12 being the "primary" factor. This imbalance is likely to skew the immune response away from Ab production and also explains the failure of CVID T cells to make Ag-specific memory cells and the chronic inflammatory and granulomatous complications that are a feature of CVID. This disease appears to be a rare example of a polarized Th1-type response and may in part be due to a genetic defect in the control of IL-12 production.

  8. Up-regulation of the interferon-related genes in BRCA2 knockout epithelial cells

    PubMed Central

    Xu, Hong; Xian, Jian; Vire, Emmanuelle; McKinney, Steven; Wong, Jason; Wei, Vivien; Tong, Rebecca; Kouzarides, Tony; Caldas, Carlos; Aparicio, Samuel

    2016-01-01

    BRCA2 mutations are significantly associated with early onset breast cancer, and the tumour suppressing function of BRCA2 has been attributed to its involvement in homologous recombination [1]-mediated DNA repair. In order to identify additional functions of BRCA2, we generated BRCA2-knockout HCT116 human colorectal carcinoma cells. Using genome-wide microarray analyses, we have discovered a link between the loss of BRCA2 and the up-regulation of a subset of interferon (IFN)-related genes, including APOBEC3F and APOBEC3G. The over-expression of IFN-related genes was confirmed in different human BRCA2−/− and mouse Brca2−/− tumour cell lines, and was independent of either senescence or apoptosis. In isogenic wild type BRCA2 cells, we observed over-expression of IFN-related genes after treatment with DNA-damaging agents, and following ionizing radiation. Cells with endogenous DNA damage because of defective BRCA1 or RAD51 also exhibited over-expression of IFN-related genes. Transcriptional activity of the IFN-stimulated response element (ISRE) was increased in BRCA2 knockout cells, and the expression of BRCA2 greatly decreased IFN-α stimulated ISRE reporter activity, suggesting that BRCA2 directly represses the expression of IFN-related genes through the ISRE. Finally, the colony forming capacity of BRCA2 knockout cells was significantly reduced in the presence of either IFN-β or IFN-γ, suggesting that IFNs may have potential as therapeutic agents in cancer cells with BRCA2 mutations. PMID:25043256

  9. Up-regulation of niacinamide in intervertebral disc aggrecan in vitro.

    PubMed

    Xiong, Xiaoqian; Yang, Shuhua; Shao, Zengwu; Liu, Xin; Zhan, Zirui; Duan, Deyu

    2006-01-01

    The regulatory effects of niacinamide (Nia) on intervertebral disc (IVD) aggrecan in vitro was investigated. Chiba's 10 ng/mL interleukin-1 (IL-1)-induced rabbit IVD degeneration model in vitro was established. 0.5, 0.25 and 0.05 mg/mL Nia was added to normal and degenerated IVDs for intervention. On the first and second week after intervention, safranin O-fast green staining intensity and glycosaminoglycan (GS) content were measured. The expression of aggrecan core protein was detected by RT-PCR. The results showed: (1) After treatment with 0.5 mg/mL Nia for one week, the GS content in nucleus pulposus (NP) was increased by 44.8% as compared with control group (P < 0 01); The GS content in IL-1 induction groups was increased with the increase of Nia concentrations: After treatment with 0.5 mg/mL for one week, the GS content in NP was increased by 68.3% as compared with control group (P < 0.01). After two weeks, GS content in NP and fibrous rings was still higher than in control group at the same period (P < 0.01) and untreated group (P < 0.01). (2) Safranin O-fast green staining revealed that with the increase of Nia concentrations, staining density in NP and fibrous rings was increased and histological structure damage to IVDs by IL-1beta was alleviated. (3) RT-PCR showed that the expression of core protein gene in IL-1beta-induced degenerated IVDS was increased with the increase of Nia concentrations. It was concluded that under conditions in vitro, Nia could up-regulate the expression of aggrecan in IVDs and protect IVDs from IL-1beta-induced degeneration at least partially, which offers a potential choice for IVD degeneration clinical therapy.

  10. Neonatal maternal separation up-regulates protein signalling for cell survival in rat hypothalamus.

    PubMed

    Irles, Claudine; Nava-Kopp, Alicia T; Morán, Julio; Zhang, Limei

    2014-05-01

    We have previously reported that in response to early life stress, such as maternal hyperthyroidism and maternal separation (MS), the rat hypothalamic vasopressinergic system becomes up-regulated, showing enlarged nuclear volume and cell number, with stress hyperresponsivity and high anxiety during adulthood. The detailed signaling pathways involving cell death/survival, modified by adverse experiences in this developmental window remains unknown. Here, we report the effects of MS on cellular density and time-dependent fluctuations of the expression of pro- and anti-apoptotic factors during the development of the hypothalamus. Neonatal male rats were exposed to 3 h-daily MS from postnatal days 2 to 15 (PND 2-15). Cellular density was assessed in the hypothalamus at PND 21 using methylene blue staining, and neuronal nuclear specific protein and glial fibrillary acidic protein immunostaining at PND 36. Expression of factors related to apoptosis and cell survival in the hypothalamus was examined at PND 1, 3, 6, 9, 12, 15, 20 and 43 by Western blot. Rats subjected to MS exhibited greater cell-density and increased neuronal density in all hypothalamic regions assessed. The time course of protein expression in the postnatal brain showed: (1) decreased expression of active caspase 3; (2) increased Bcl-2/Bax ratio; (3) increased activation of ERK1/2, Akt and inactivation of Bad; PND 15 and PND 20 were the most prominent time-points. These data indicate that MS can induce hypothalamic structural reorganization by promoting survival, suppressing cell death pathways, increasing cellular density which may alter the contribution of these modified regions to homeostasis.

  11. Stat3 promotes invasion of esophageal squamous cell carcinoma through up-regulation of MMP2.

    PubMed

    Xuan, Xaioyan; Li, Shanshan; Lou, Xi; Zheng, Xianzhao; Li, Yunyun; Wang, Feng; Gao, Yuan; Zhang, Hongyan; He, Hongliu; Zeng, Qingru

    2015-05-01

    Stat3 alters the expression of its downstream genes and is associated with tumor invasion and metastasis in several human cancers. Its role in esophageal squamous cell carcinoma (ESCC) has not been well characterized. We examined the tumor sections of 100 cases of ESCC by immunohistochemistry and observed significant overexpression of Stat3 in the cytoplasm of 89% of ESCC cells and of phosphorylated Stat3 (p-Stat3) in the nuclei of 71% of ESCC when compare with normal esophageal mucosa (72%, p = 0.02; and 31%, p = 0.001). Overexpression of Stat3 and p-Stat3 positively correlated with that of matrix metalloproteinase-2 (MMP2), a known regulator for cell migration, in 65% of ESCC while only 26% shown in benign esophageal mucosa. To further investigate the association of Stat3 with tumor metastasis in vitro, invasion of EC-1 cells (a human ESCC cell line) were investigated with Boyden chambers. The results showed that transfection of Stat3 not only promoted invasion of EC-1 cells but also significantly induced MMP2 expression in a dose-dependent manner. In contrast, suppressing expression of endogenous Stat3 mRNA and protein by Stat3 siRNA significantly reduced EC-1 cell invasion and MMP2 expression. A high-affinity Stat3-binding element was localized to the positions of 648-641 bp (TTCTCGAA) in the MMP2 promoter with electrophoretic mobility shift assay. Our results suggest that Stat3, p-Stat3, and MMP2 were overexpressed in ESCC and associated with invasion of ESCC; and Stat3 up-regulated expression of MMP2 in ESCC through directly binding to the MMP2 promoter.

  12. Gallium arsenide selectively up-regulates inflammatory cytokine expression at exposure site.

    PubMed

    Becker, Stephen M; McCoy, Kathleen L

    2003-12-01

    Gallium arsenide (GaAs), a technologically and economically important semiconductor, is widely utilized in both military and commercial applications. This chemical is a potential health hazard as a carcinogen and immunotoxicant. We previously reported that macrophages at the exposure site exhibit characteristics of activation. In vitro culture of macrophages with GaAs fails to recapitulate the in vivo phenotype, suggesting that complete GaAs-mediated activation in vivo may require other cells or components found in the body's microenvironment. Our present study examined the role of cytokines upon GaAs-mediated macrophage activation. Intraperitoneal administration of GaAs elicited rapid specific recruitment of blood monocytes to the exposure site. This recruitment occurred concomitant with up-regulation of 17 chemokine and inflammatory cytokine mRNAs, while transcripts of three inhibitory cytokines diminished. Administration of latex beads caused less cytokine induction than GaAs, indicating that changes in mRNA levels could not be attributed to phagocytosis. Four representative chemokines and cytokines were selected for further analysis. Increased cytokine mRNA expression was paralleled by similar increases in cytokine protein levels, and secreted protein products were detected in peritoneal fluid. Cytokine protein expression was constrained to myeloid cells, and to a lesser extent to B cells. Alterations in patterns of cytokine gene expression elucidate mechanisms for increased cellular activation and antigen processing, and modulation of the inflammatory response. Our findings indicate that in vivo GaAs exposure alters cytokine gene expression, which may lead to an inflammatory reaction and contribute to pathological tissue damage.

  13. Modified AS1411 Aptamer Suppresses Hepatocellular Carcinoma by Up-Regulating Galectin-14

    PubMed Central

    Lee, Jeong-Hoon; Lee, Dong Hyeon; Cho, Eun Ju; Yu, Su Jong; Kim, Yoon Jun; Kim, Jong In; Im, Jong Hun; Lee, Jung Hwan; Oh, Eun Ju; Yoon, Jung-Hwan

    2016-01-01

    Aptamers are small synthetic oligonucleotides that bind to target proteins with high specificity and affinity. AS1411 is an aptamer that binds to nucleolin, which is overexpressed in the cytoplasm and occurs on the surface of cancer cells. We investigated the therapeutic potential of aptamers in hepatocellular carcinoma (HCC) by evaluating anti-tumor effects and confirming the affinity and specificity of AS1411- and modified AS1411-aptamers in HCC cells. Cell growth was assessed using the MTS assay, and cell death signaling was explored by immunoblot analysis. Fluorescence-activated cell sorting was performed to evaluate the affinity and specificity of AS1411-aptamers in SNU-761 HCC cells. We investigated the in vivo effects of the AS1411-aptamer using BALB/c nude mice in a subcutaneous xenograft model with SNU-761 cells. Treatment with a modified AS1411-aptamer significantly decreased in vitro (under normoxic [P = 0.035] and hypoxic [P = 0.018] conditions) and in vivo (under normoxic conditions, P = 0.041) HCC cell proliferation compared to control aptamers. AS1411- and control aptamers failed to control HCC cell proliferation. However, AS1411- and the modified AS1411-aptamer did not induce caspase activation. Decrease in cell growth by AS1411 or modified AS1411 was not prevented by caspase or necrosis inhibitors. In a microarray, AS1411 significantly enhanced galectin-14 expression. Suppression of HCC cell proliferation by the modified AS1411-aptamer was attenuated by galectin-14 siRNA transfection. Modified AS1411-aptamer suppressed HCC cell growth in vitro and in vivo by up-regulating galectin-14 expressions. Modified AS1411-aptamers may have therapeutic potential as a novel targeted therapy for HCC. PMID:27494117

  14. The Wnt Inhibitor Sclerostin Is Up-regulated by Mechanical Unloading in Osteocytes in Vitro.

    PubMed

    Spatz, Jordan M; Wein, Marc N; Gooi, Jonathan H; Qu, Yili; Garr, Jenna L; Liu, Shawn; Barry, Kevin J; Uda, Yuhei; Lai, Forest; Dedic, Christopher; Balcells-Camps, Mercedes; Kronenberg, Henry M; Babij, Philip; Pajevic, Paola Divieti

    2015-07-01

    Although bone responds to its mechanical environment, the cellular and molecular mechanisms underlying the response of the skeleton to mechanical unloading are not completely understood. Osteocytes are the most abundant but least understood cells in bones and are thought to be responsible for sensing stresses and strains in bone. Sclerostin, a product of the SOST gene, is produced postnatally primarily by osteocytes and is a negative regulator of bone formation. Recent studies show that SOST is mechanically regulated at both the mRNA and protein levels. During prolonged bed rest and immobilization, circulating sclerostin increases both in humans and in animal models, and its increase is associated with a decrease in parathyroid hormone. To investigate whether SOST/sclerostin up-regulation in mechanical unloading is a cell-autonomous response or a hormonal response to decreased parathyroid hormone levels, we subjected osteocytes to an in vitro unloading environment achieved by the NASA rotating wall vessel system. To perform these studies, we generated a novel osteocytic cell line (Ocy454) that produces high levels of SOST/sclerostin at early time points and in the absence of differentiation factors. Importantly, these osteocytes recapitulated the in vivo response to mechanical unloading with increased expression of SOST (3.4 ± 1.9-fold, p < 0.001), sclerostin (4.7 ± 0.1-fold, p < 0.001), and the receptor activator of nuclear factor κΒ ligand (RANKL)/osteoprotegerin (OPG) (2.5 ± 0.7-fold, p < 0.001) ratio. These data demonstrate for the first time a cell-autonomous increase in SOST/sclerostin and RANKL/OPG ratio in the setting of unloading. Thus, targeted osteocyte therapies could hold promise as novel osteoporosis and disuse-induced bone loss treatments by directly modulating the mechanosensing cells in bone. PMID:25953900

  15. Cathepsin D is up-regulated in inflammatory bowel disease macrophages

    PubMed Central

    HAUSMANN, M; OBERMEIER, F; SCHREITER, K; SPOTTL, T; FALK, W; SCHÖLMERICH, J; HERFARTH, H; SAFTIG, P; ROGLER, G

    2004-01-01

    Down-regulation of receptors involved in the recognition or transmission of inflammatory signals and a reduced responsiveness support the concept that macrophages are ‘desensitized’ during their differentiation in the intestinal mucosa. During inflammatory bowel disease (IBD) intestinal macrophages (IMACs) change to a reactive or ‘aggressive’ type. After having established a method of isolation and purification of IMACs, message for cathepsin D was one of the mRNAs we found to be up-regulated in a subtractive hybridization of Crohn's disease (CD) macrophages versus IMACs from control mucosa. The expression of cathepsin D in intestinal mucosa was analysed by immunohistochemistry in biopsies from IBD and control patients and in a mouse model of dextran sulphate sodium (DSS)-induced acute and chronic colitis. IMACs were isolated and purified from normal and inflamed mucosa by immunomagnetic beads armed with a CD33 antibody. RT-PCR was performed for cathepsin D mRNA. Results were confirmed by Northern blot and flow cytometrical analysis. Immunohistochemistry revealed a significant increase in the cathepsin D protein expression in inflamed intestinal mucosa from IBD patients compared to non-inflamed mucosa. No cathepsin D polymerase chain reaction (PCR) product could be obtained with mRNA from CD33-positive IMACs from normal mucosa. Reverse transcription (RT)-PCR showed an induction of mRNA for cathepsin D in purified IMACs from IBD patients. Northern blot and flow cytometry analysis confirmed these results. Cathepsin D protein was also found in intestinal mucosa in acute and chronic DSS-colitis but was absent in normal mucosa. This study shows that expression of cathepsin D is induced in inflammation-associated IMACs. The presence of cathepsin D might contribute to the mucosal damage in IBD. PMID:15030527

  16. Up-regulation of apolipoprotein E by leptin in the hypothalamus of mice and rats

    PubMed Central

    Shen, Ling; Tso, Patrick; Wang, David Q.-H.; Woods, Stephen C.; Davidson, W. Sean; Sakai, Randall; Liu, Min

    2009-01-01

    Apolipoprotein E (apoE) is a satiation factor, playing an important role in the regulation of food intake and body weight. We previously reported that apoE was present in the hypothalamus, but it is unclear which type of the cells in this brain area expressing apoE. In addition, hypothalamic apoE mRNA levels were significantly reduced in both genetically obese ob/ob (leptin deficient) mice and high-fat diet-induced obese (leptin resistant) rats, raising the possibility that deficient leptin signaling might be related to the change in apoE gene expression. In the present studies, using double-staining immunohistochemistry, we demonstrated that apoE is mainly present in astrocytes. To characterize the effect of leptin on apoE gene expression, ob/ob and db/db mice were treated with recombinant mouse leptin (3 μg/g daily, i.p.) or vehicle for 5 days. We found that the increased hypothalamic apoE mRNA levels occurred only in leptin-treated ob/ob, but not in pair-fed ob/ob, or db/db, mice, indicating that leptin up-regulated hypothalamic apoE gene expression depends upon an intact leptin receptor, and this effect is not related to the changes in food intake and body weight. The reduced apoE gene expression caused by fasting, which also results in relatively lower leptin level, is restored by intracerebroventricular administration of leptin. In addition, leptin was significantly less efficacious in apoE KO mice because these animals consumed more food and lost less weight following leptin treatment, compared with wild-type controls. These observations imply that apoE signaling, at least partially, mediates the inhibitory effects of leptin on feeding. PMID:19481557

  17. Homeobox A7 stimulates breast cancer cell proliferation by up-regulating estrogen receptor-alpha

    SciTech Connect

    Zhang, Yu; Cheng, Jung-Chien; Huang, He-Feng; Leung, Peter C.K.

    2013-11-01

    Highlights: •HOXA7 regulates MCF7 cell proliferation. •HOXA7 up-regulates ERα expression. •HOXA7 mediates estrogen-induced MCF7 cell proliferation. -- Abstract: Breast cancer is the most common hormone-dependent malignancy in women. Homeobox (HOX) transcription factors regulate many cellular functions, including cell migration, proliferation and differentiation. The aberrant expression of HOX genes has been reported to be associated with human reproductive cancers. Estradiol (E2) and its nuclear receptors, estrogen receptor (ER)-alpha and ER-beta, are known to play critical roles in the regulation of breast cancer cell growth. However, an understanding of the potential relationship between HOXA7 and ER in breast cancer cells is limited. In this study, our results demonstrate that knockdown of HOXA7 in MCF7 cells significantly decreased cell proliferation and ERα expression. In addition, HOXA7 knockdown attenuated E2-induced cell proliferation as well as progesterone receptor (PR) expression. The stimulatory effects of E2 on cell proliferation and PR expression were abolished by co-treatment with ICI 182780, a selective ERα antagonist. In contrast, overexpression of HOXA7 significantly stimulated cell proliferation and ERα expression. Moreover, E2-induced cell proliferation, as well as PR expression, was enhanced by the overexpression of HOXA7. Neither knockdown nor overexpression of HOXA7 affected the ER-beta levels. Our results demonstrate a novel mechanistic role for HOXA7 in modulating breast cancer cell proliferation via regulation of ERα expression. This finding contributes to our understanding of the role HOXA7 plays in regulating the proliferation of ER-positive cancer cells.

  18. The Wnt Inhibitor Sclerostin Is Up-regulated by Mechanical Unloading in Osteocytes in Vitro*

    PubMed Central

    Spatz, Jordan M.; Wein, Marc N.; Gooi, Jonathan H.; Qu, Yili; Garr, Jenna L.; Liu, Shawn; Barry, Kevin J.; Uda, Yuhei; Lai, Forest; Dedic, Christopher; Balcells-Camps, Mercedes; Kronenberg, Henry M.; Babij, Philip; Pajevic, Paola Divieti

    2015-01-01

    Although bone responds to its mechanical environment, the cellular and molecular mechanisms underlying the response of the skeleton to mechanical unloading are not completely understood. Osteocytes are the most abundant but least understood cells in bones and are thought to be responsible for sensing stresses and strains in bone. Sclerostin, a product of the SOST gene, is produced postnatally primarily by osteocytes and is a negative regulator of bone formation. Recent studies show that SOST is mechanically regulated at both the mRNA and protein levels. During prolonged bed rest and immobilization, circulating sclerostin increases both in humans and in animal models, and its increase is associated with a decrease in parathyroid hormone. To investigate whether SOST/sclerostin up-regulation in mechanical unloading is a cell-autonomous response or a hormonal response to decreased parathyroid hormone levels, we subjected osteocytes to an in vitro unloading environment achieved by the NASA rotating wall vessel system. To perform these studies, we generated a novel osteocytic cell line (Ocy454) that produces high levels of SOST/sclerostin at early time points and in the absence of differentiation factors. Importantly, these osteocytes recapitulated the in vivo response to mechanical unloading with increased expression of SOST (3.4 ± 1.9-fold, p < 0.001), sclerostin (4.7 ± 0.1-fold, p < 0.001), and the receptor activator of nuclear factor κΒ ligand (RANKL)/osteoprotegerin (OPG) (2.5 ± 0.7-fold, p < 0.001) ratio. These data demonstrate for the first time a cell-autonomous increase in SOST/sclerostin and RANKL/OPG ratio in the setting of unloading. Thus, targeted osteocyte therapies could hold promise as novel osteoporosis and disuse-induced bone loss treatments by directly modulating the mechanosensing cells in bone. PMID:25953900

  19. Rosiglitazone ameliorates diffuse axonal injury by reducing loss of tau and up-regulating caveolin-1 expression

    PubMed Central

    Zhao, Yong-lin; Song, Jin-ning; Ma, Xu-dong; Zhang, Bin-fei; Li, Dan-dong; Pang, Hong-gang

    2016-01-01

    Rosiglitazone up-regulates caveolin-1 levels and has neuroprotective effects in both chronic and acute brain injury. Therefore, we postulated that rosiglitazone may ameliorate diffuse axonal injury via its ability to up-regulate caveolin-1, inhibit expression of amyloid-beta precursor protein, and reduce the loss and abnormal phosphorylation of tau. In the present study, intraperitoneal injection of rosiglitazone significantly reduced the levels of amyloid-beta precursor protein and hyperphosphorylated tau (phosphorylated at Ser404(p-tau (S404)), and it increased the expression of total tau and caveolin-1 in the rat cortex. Our results show that rosiglitazone inhibits the expression of amyloid-beta precursor protein and lowers p-tau (S404) levels, and it reduces the loss of total tau, possibly by up-regulating caveolin-1. These actions of rosiglitazone may underlie its neuroprotective effects in the treatment of diffuse axonal injury. PMID:27482223

  20. HGF/MET signaling promotes glioma growth via up-regulation of Cox-2 expression and PGE2 production

    PubMed Central

    Zhao, Yan; Sun, Ying; Zhang, Haiyan; Liu, Xing; Du, Wenzong; Li, Yongli; Zhang, Junhe; Chen, Lingchao; Jiang, Chuanlu

    2015-01-01

    Cyclooxygenase2 (Cox-2) is well known for glioma growth through up-regulation of prostaglandin E2 (PGE2) levels. MET, a hepatocyte growth factor (HGF) receptor, is also frequently high expressed in glioma, which promotes glioma growth and invasion. Here, we demonstrate that HGF/MET signaling can promote PGE2 production in glioma cells via Cox-2 up-regulation. RNA inhibition of MET suggested that MET signaling is essential for Cox-2 up-regulation. Moreover, HGF could enhance Cox-2 expression and PGE2 release. Knockdown of Cox-2 inhibited growth-promoting effects of HGF, suggesting that HGF/MET functioned via Cox-2/PGE2 pathway. Therefore, our work reveals a critical relationship of Cox-2/PGE2 and HGF/MET signaling in promoting glioma cells proliferation. Further, targeting MET and Cox-2 may represent an attractive target therapy for glioma. PMID:26097553

  1. Meta-analysis of gene expression profiles indicates genes in spliceosome pathway are up-regulated in hepatocellular carcinoma (HCC).

    PubMed

    Xu, Weijin; Huang, Huixing; Yu, Long; Cao, Lihuan

    2015-04-01

    Hepatocellular carcinoma (HCC) is among the commonest kind of malignant tumors, which accounts for more than 500,000 cases of newly diagnosed cancer annually. Many microarray studies for identifying differentially expressed genes (DEGs) in HCC have been conducted, but results have varied across different studies. Here, we performed a meta-analysis of publicly available microarray Gene Expression Omnibus datasets, which covers five independent studies, containing 753 HCC samples and 638 non-tumor liver samples. We identified 192 DEGs that were consistently up-regulated in HCC vs. normal liver tissue. For the 192 up-regulated genes, we performed Kyoto Encyclopedia of Genes and Genomes pathway analysis. To our surprise, besides several cell growth-related pathways, spliceosome pathway was also up-regulated in HCC. For further exploring the relationship between spliceosome pathway and HCC, we investigated the expression data of spliceosome pathway genes in 15 independent studies in Nextbio database ( https://www.nextbio.com/b/nextbioCorp.nb ). It was found that many genes of spliceosome pathway such as HSPA1A, SNRPE, SF3B2, SF3B4 and TRA2A genes which we identified to be up-regulated in our meta-analysis were generally overexpressed in HCC. At last, using real-time PCR, we also found that BUD31, SF3B2, SF3B4, SNRPE, SPINK1, TPA2A and HSPA1A genes are significantly up-regulated in clinical HCC samples when compared to the corresponding non-tumorous liver tissues. Our study for the first time indicates that many genes of spliceosome pathway are up-regulated in HCC. This finding might put new insights for people's understanding about the relationship of spliceosome pathway and HCC.

  2. Up-regulation of divalent metal transporter 1 in 6-hydroxydopamine intoxication is IRE/IRP dependent.

    PubMed

    Jiang, Hong; Song, Ning; Xu, Huamin; Zhang, Shuzhen; Wang, Jun; Xie, Junxia

    2010-03-01

    Iron plays a key role in Parkinson's disease (PD). Increased iron content of the substantia nigra (SN) has been found in PD patients, and divalent metal transporter 1 (DMT1) has been shown to be up-regulated in the SN of both MPTP-induced PD models and PD patients. However, the mechanisms underlying DMT1 up-regulation are largely unknown. In the present study, we observed that in the SN of 6-hydroxydopamine (6-OHDA)-induced PD rats, DMT1 with the iron responsive element (IRE, DMT1+IRE), but not DMT1 without IRE (DMT1-IRE), was up-regulated, suggesting that increased DMT1+IRE expression might account for nigral iron accumulation in PD rats. This possibility was further assessed in an in vitro study using 6-OHDA-treated and DMT1+IRE-over-expressing MES23.5 cells. In 6-OHDA-treated MES23.5 cells, increased iron regulatory protein (IRP) 1 and IRP2 expression was observed, while silencing of IRPs dramatically diminished 6-OHDA-induced DMT1+IRE up-regulation. Pretreatment with N-acetyl-L-cysteine fully suppressed IRPs up-regulation by inhibition of 6-OHDA-induced oxidative stress. Increased DMT1+IRE expression resulted in increased iron influx by MES23.5 cells. Our data provide direct evidence that DMT1+IRE up-regulation can account for IRE/IRP-dependent 6-OHDA-induced iron accumulation initiated by 6-OHDA-induced intracellular oxidative stress and that increased levels of intracellular iron result in aggravated oxidative stress. The results of this study provide novel evidence supporting the use of anti-oxidants in the treatment of PD, with the goal of inhibiting iron accumulation by regulation of DMT1 expression. PMID:20125122

  3. Dietary iron-deficiency up-regulates hephaestin mRNA level in small intestine of rats.

    PubMed

    Sakakibara, Shoji; Aoyama, Yoritaka

    2002-05-17

    Hephaestin is a protein, recently found from the study of sla (sex-linked anemia) mouse. Hephaestin is suggested to transport iron from intestinal enterocytes into the circulation. Iron is essential for living and for humans to maintain a constant total iron concentration in whole body. In this study, it was found that dietary iron-deficiency up-regulated hephaestin mRNA level in the proximal small intestine of rats. Therefore, it is suggested that in dietary iron-deficiency, hephaestin gene expression in proximal small intestine is up-regulated to absorb more iron from diet.

  4. Up-Regulation of Claudin-6 in the Distal Lung Impacts Secondhand Smoke-Induced Inflammation

    PubMed Central

    Lewis, Joshua B.; Milner, Dallin C.; Lewis, Adam L.; Dunaway, Todd M.; Egbert, Kaleb M.; Albright, Scott C.; Merrell, Brigham J.; Monson, Troy D.; Broberg, Dallin S.; Gassman, Jason R.; Thomas, Daniel B.; Arroyo, Juan A.; Reynolds, Paul R.

    2016-01-01

    It has long been understood that increased epithelial permeability contributes to inflammation observed in many respiratory diseases. Recently, evidence has revealed that environmental exposure to noxious material such as cigarette smoke reduces tight junction barrier integrity, thus enhancing inflammatory conditions. Claudin-6 (Cldn6) is a tetraspanin transmembrane protein found within the tight junctional complex and is implicated in maintaining lung epithelial barriers. To test the hypothesis that increased Cldn6 ameliorates inflammation at the respiratory barrier, we utilized the Tet-On inducible transgenic system to conditionally over-express Clnd6 in the distal lung. Cldn6 transgenic (TG) and control mice were continuously provided doxycycline from postnatal day (PN) 30 until euthanasia date at PN90. A subset of Cldn6 TG and control mice were also subjected to daily secondhand tobacco smoke (SHS) via a nose only inhalation system from PN30-90 and compared to room air (RA) controls. Animals were euthanized on PN90 and lungs were harvested for histological and molecular characterization. Bronchoalveolar lavage fluid (BALF) was procured for the assessment of inflammatory cells and molecules. Quantitative RT-PCR and immunoblotting revealed increased Cldn6 expression in TG vs. control animals and SHS decreased Cldn6 expression regardless of genetic up-regulation. Histological evaluations revealed no adverse pulmonary remodeling via Hematoxylin and Eosin (H&E) staining or any qualitative alterations in the abundance of type II pneumocytes or proximal non-ciliated epithelial cells via staining for cell specific propeptide of Surfactant Protein-C (proSP-C) or Club Cell Secretory Protein (CCSP), respectively. Immunoblotting and qRT-PCR confirmed the differential expression of Cldn6 and the pro-inflammatory cytokines TNF-α and IL-1β. As a general theme, inflammation induced by SHS exposure was influenced by the availability of Cldn6. These data reveal captivating

  5. Aberrant Alternative Polyadenylation is Responsible for Survivin Up-regulation in Ovarian Cancer

    PubMed Central

    He, Xiang-Jun; Zhang, Qi; Ma, Li-Ping; Li, Na; Chang, Xiao-Hong; Zhang, Yu-Jun

    2016-01-01

    RNAs and is responsible for survivin up-regulation. PMID:27174320

  6. Antagonism between MyD88- and TRIF-dependent signals in B7RP-1 up-regulation.

    PubMed

    Zhou, Zuping; Hoebe, Kasper; Du, Xin; Jiang, Zhengfan; Shamel, Louis; Beutler, Bruce

    2005-06-01

    Type I interferons (IFN) play a critical role in the Toll-like receptor (TLR)-mediated expression of B7 costimulatory family members. For example, LPS-induced up-regulation of CD80 (B7.1) and CD86 (B7.2) is abrogated in antigen-presenting cells (APC) deficient in TRIF or TRAM, two adaptors that are responsible for TLR4-mediated production of Type I IFN. In this report, we demonstrate that LPS-induced up-regulation of B7-related protein 1 (B7RP-1), a ligand for ICOS, is dependent primarily upon the MyD88-dependent signaling pathway. Signaling via the TRIF pathway sharply limits MyD88-dependent B7RP-1 up-regulation. Hence, LPS induces significantly higher B7RP-1 expression on TRIF- or TRAM-deficient mouse peritoneal macrophages and on TRIF-deficient mouse splenic B cells as compared to wild-type cells. Further studies reveal that Type I IFN are general suppressors of TLR-mediated up-regulation of B7RP-1. These data indicate that Type I IFN play a dual role in the TLR-mediated expression of B7 costimulatory family members and suggest that they may act to limit B7RP-1 expression and thus limit signals derived from B7RP-1-ICOS interaction.

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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) ...

  8. Sildenafil prevents the up-regulation of transient receptor potential canonical channels in the development of cardiomyocyte hypertrophy

    SciTech Connect

    Kiso, Hironori; Ohba, Takayoshi; Iino, Kenji; Sato, Kazuhiro; Terata, Yutaka; Murakami, Manabu; Ono, Kyoichi; Watanabe, Hiroyuki; Ito, Hiroshi

    2013-07-05

    Highlights: •Transient receptor potential canonical (TRPC1, 3 and 6) are up-regulated by ET-1. •Sildenafil inhibited hypertrophic responses (BNP, Ca entry, NFAT activation). •Sildenafil suppressed TRPC1, 3 and 6 expression. -- Abstract: Background: Transient receptor potential canonical (TRPCs) channels are up-regulated in the development of cardiac hypertrophy. Sildenafil inhibits TRPC6 activation and expression, leading to the prevention of cardiac hypertrophy. However, the effects of sildenafil on the expression of other TRPCs remain unknown. We hypothesized that in addition to its effects of TRPC6, sildenafil blocks the up-regulation of other TRPC channels to suppress cardiomyocyte hypertrophy. Methods and results: In cultured neonatal rat cardiomyocytes, a 48 h treatment with 10 nM endothelin (ET)-1 induced hypertrophic responses characterized by nuclear factor of activated T cells activation and enhancement of brain natriuretic peptide expression and cell surface area. Co-treatment with sildenafil (1 μM, 48 h) inhibited these ET-1-induced hypertrophic responses. Although ET-1 enhanced the gene expression of TRPCs, sildenafil inhibited the enhanced gene expression of TRPC1, C3 and C6. Moreover, co-treatment with sildenafil abolished the augmentation of SOCE in the hypertrophied cardiomyocytes. Conclusions: These results suggest that sildenafil inhibits cardiomyocyte hypertrophy by suppressing the up-regulation of TRPC expression.

  9. Cotton Benzoquinon Reductase: Up-Regulation During Early Fiber Development and Heterologous Expresson and Characterization in Pichia Pastoris

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Benzoquinone reductase (BR) is an enzyme which catalyzes the bivalent redox reactions of quinones without the production of free radical intermediates. Using 2-D PAGE, two proteins were found to be up-regulated in wild-type cotton ovules during the fiber initiation stage. These proteins were excis...

  10. Up-Regulation of Nerve Growth Factor in Cholestatic Livers and Its Hepatoprotective Role against Oxidative Stress

    PubMed Central

    Tsai, Ming-Shian; Lin, Yu-Chun; Sun, Cheuk-Kwan; Huang, Shih-Che; Lee, Po-Huang; Kao, Ying-Hsien

    2014-01-01

    The role of nerve growth factor (NGF) in liver injury induced by bile duct ligation (BDL) remains elusive. This study aimed to investigate the relationship between inflammation and hepatic NGF expression, to explore the possible upstream molecules up-regulating NGF, and to determine whether NGF could protect hepatocytes from oxidative liver injury. Biochemical and molecular detection showed that NGF was up-regulated in cholestatic livers and plasma, and well correlated with systemic and hepatic inflammation. Conversely, systemic immunosuppression reduced serum NGF levels and resulted in higher mortality in BDL-treated mice. Immunohistochemistry showed that the up-regulated NGF was mainly localized in parenchymal hepatocytes. In vitro mechanistic study further demonstrated that TGF-β1 up-regulated NGF expression in clone-9 and primary rat hepatocytes. Exogenous NGF supplementation and endogenous NGF overexpression effectively protected hepatocytes against TGF-β1- and oxidative stress-induced cell death in vitro, along with reduced formation of oxidative adducted proteins modified by 4-HNE and 8-OHdG. TUNEL staining confirmed the involvement of anti-apoptosis in the NGF-exhibited hepatoprotection. Moreover, NGF potently induced Akt phosphorylation and increased Bcl-2 to Bax ratios, whereas these molecular alterations by NGF were only seen in the H2O2-, but not TGF-β1-treated hepatocytes. In conclusion, NGF exhibits anti-oxidative and hepatoprotective effects and is suggested to be therapeutically applicable in treating cholestatic liver diseases. PMID:25397406

  11. Localization of a filarial phosphate permease that is up-regulated in response to depletion of essential Wolbachia endobacteria.

    PubMed

    Arumugam, Sridhar; Hoerauf, Achim; Pfarr, Kenneth M

    2014-03-01

    Wolbachia of filarial nematodes are essential, obligate endobacteria. When depleted by doxycycline worm embryogenesis, larval development and worm survival are inhibited. The molecular basis governing the endosymbiosis between Wolbachia and their filarial host is still being deciphered. In rodent filarial nematode Litomosoides sigmodontis, a nematode encoded phosphate permease gene (Ls-ppe-1) was up-regulated at the mRNA level in response to Wolbachia depletion and this gene promises to have an important role in Wolbachia-nematode endosymbiosis. To further characterize this gene, the regulation of phosphate permease during Wolbachia depletion was studied at the protein level in L. sigmodontis and in the human filaria Onchocerca volvulus. And the localization of phosphate permease (PPE) and Wolbachia in L. sigmodontis and O. volvulus was investigated in untreated and antibiotic treated worms. Depletion of Wolbachia by tetracycline (Tet) resulted in up-regulation of Ls-ppe-1 in L. sigmodontis. On day 36 of Tet treatment, compared to controls (Con), >98% of Wolbachia were depleted with a 3-fold increase in mRNA levels of Ls-ppe-1. Anti-Ls-PPE serum used in Western blots showed up-regulation of Ls-PPE at the protein level in Tet worms on day 15 and 36 of treatment. Immunohistology revealed the localization of Wolbachia and Ls-PPE in the embryos, microfilariae and hypodermis of L. sigmodontis female worms and up-regulation of Ls-PPE in response to Wolbachia depletion. Expression of O. volvulus phosphate permease (Ov-PPE) studied using anti-Ov-PPE serum, showed up-regulation of Ov-PPE at the protein level in doxycycline treated Wolbachia depleted O. volvulus worms and immunohistology revealed localization of Ov-PPE and Wolbachia and up-regulation of Ov-PPE in the hypodermis and embryos of doxycycline treated worms. Ls-PPE and Ov-PPE are upregulated upon Wolbachia depletion in same tissues and regions where Wolbachia are located in untreated worms, reinforcing a link

  12. [Low dose benzo(a)pyrene up-regulated the transcription of HSP70 and HSP90 in Eisenia fetida].

    PubMed

    Zheng, Sen-Lin; Sun, Tie-Heng; Xiao, Hong; Qiu, Xiao-Yan; Song, Yu-Fang

    2008-02-01

    To search for the molecular biomarkers of sub-lethal polycyclic aromatic hydrocarbons (PAHs)-contamination of soil, the subtractive cDNA libraries of earthworm Eisenia fetida exposed to benzo(a)pyrene (BaP) in artificial soil were constructed by suppression subtractive hybridization. After sequencing and analyzing with basic local alignment search tool (BLAST), two clones matching heat shock protein 70 k Da (HSP70) and one clone matching heat shock protein 90 k Da (HSP90) were isolated from the up-regulated library, and subsequently, the up-regulation of HSP70 and HSP90 was verified by real-time PCR in E. fetida exposed to 0.1 mg x kg(-1) and 1.0 mg x kg(-1) BaP. It was indicated that these two newly identified HSPs in E. fetida were the potential molecular biomarkers for soil contamination monitoring.

  13. {beta}-Catenin up-regulates Nanog expression through interaction with Oct-3/4 in embryonic stem cells

    SciTech Connect

    Takao, Yukinari; Yokota, Takashi; Koide, Hiroshi . E-mail: hkoide@med.kanazawa-u.ac.jp

    2007-02-16

    It is well known that mouse embryonic stem (ES) cells can be maintained by the presence of leukemia inhibitory factor (LIF). Recent studies have revealed that Wnt also exhibits activity similar to LIF. The molecular mechanism behind the maintenance of ES cells by these factors, however, is not fully understood. In this study, we found that LIF enhances level of nuclear {beta}-catenin, a component of the Wnt signaling pathway. Expression of an activated mutant of {beta}-catenin led to the long-term proliferation of ES cells, even in the absence of LIF. Furthermore, it was found that {beta}-catenin up-regulates Nanog in an Oct-3/4-dependent manner and that {beta}-catenin physically associates with Oct-3/4. These results suggest that up-regulating Nanog through interaction with Oct-3/4 involves {beta}-catenin in the LIF- and Wnt-mediated maintenance of ES cell self-renewal.

  14. Ischemic postconditioning protects against ischemic brain injury by up-regulation of acid-sensing ion channel 2a

    PubMed Central

    Duanmu, Wang-sheng; Cao, Liu; Chen, Jing-yu; Ge, Hong-fei; Hu, Rong; Feng, Hua

    2016-01-01

    Ischemic postconditioning renders brain tissue tolerant to brain ischemia, thereby alleviating ischemic brain injury. However, the exact mechanism of action is still unclear. In this study, a rat model of global brain ischemia was subjected to ischemic postconditioning treatment using the vessel occlusion method. After 2 hours of ischemia, the bilateral common carotid arteries were blocked immediately for 10 seconds and then perfused for 10 seconds. This procedure was repeated six times. Ischemic postconditioning was found to mitigate hippocampal CA1 neuronal damage in rats with brain ischemia, and up-regulate acid-sensing ion channel 2a expression at the mRNA and protein level. These findings suggest that ischemic postconditioning up-regulates acid-sensing ion channel 2a expression in the rat hippocampus after global brain ischemia, which promotes neuronal tolerance to ischemic brain injury. PMID:27212927

  15. HDAC up-regulation in early colon field carcinogenesis is involved in cell tumorigenicity through regulation of chromatin structure.

    PubMed

    Stypula-Cyrus, Yolanda; Damania, Dhwanil; Kunte, Dhananjay P; Cruz, Mart Dela; Subramanian, Hariharan; Roy, Hemant K; Backman, Vadim

    2013-01-01

    Normal cell function is dependent on the proper maintenance of chromatin structure. Regulation of chromatin structure is controlled by histone modifications that directly influence chromatin architecture and genome function. Specifically, the histone deacetylase (HDAC) family of proteins modulate chromatin compaction and are commonly dysregulated in many tumors, including colorectal cancer (CRC). However, the role of HDAC proteins in early colorectal carcinogenesis has not been previously reported. We found HDAC1, HDAC2, HDAC3, HDAC5, and HDAC7 all to be up-regulated in the field of human CRC. Furthermore, we observed that HDAC2 up-regulation is one of the earliest events in CRC carcinogenesis and observed this in human field carcinogenesis, the azoxymethane-treated rat model, and in more aggressive colon cancer cell lines. The universality of HDAC2 up-regulation suggests that HDAC2 up-regulation is a novel and important early event in CRC, which may serve as a biomarker. HDAC inhibitors (HDACIs) interfere with tumorigenic HDAC activity; however, the precise mechanisms involved in this process remain to be elucidated. We confirmed that HDAC inhibition by valproic acid (VPA) targeted the more aggressive cell line. Using nuclease digestion assays and transmission electron microscopy imaging, we observed that VPA treatment induced greater changes in chromatin structure in the more aggressive cell line. Furthermore, we used the novel imaging technique partial wave spectroscopy (PWS) to quantify nanoscale alterations in chromatin. We noted that the PWS results are consistent with the biological assays, indicating a greater effect of VPA treatment in the more aggressive cell type. Together, these results demonstrate the importance of HDAC activity in early carcinogenic events and the unique role of higher-order chromatin structure in determining cell tumorigenicity.

  16. Up-regulation of stomatin expression by hypoxia and glucocorticoid stabilizes membrane-associated actin in alveolar epithelial cells

    PubMed Central

    Chen, Ji-Cheng; Cai, Hao-Yu; Wang, Yan; Ma, Yuan-Yuan; Song, Liang-Nian; Yin, Li-Juan; Cao, Dong-Mei; Diao, Fei; Li, Yi-Dong; Lu, Jian

    2013-01-01

    Stomatin is an important lipid raft-associated protein which interacts with membrane proteins and plays a role in the membrane organization. However, it is unknown whether it is involved in the response to hypoxia and glucocorticoid (GC) in alveolar epithelial cells (AEC). In this study we found that hypoxia and dexamethasone (dex), a synthetic GC not only up-regulated the expression of stomatin alone, but also imposed additive effect on the expression of stomatin in A549 cells, primary AEC and lung of rats. Then we investigated whether hypoxia and dex transcriptionally up-regulated the expression of stomatin by reporter gene assay, and found that dex, but not hypoxia could increase the activity of a stomatin promoter-driven reporter gene. Further deletion and mutational studies demonstrated that a GC response element (GRE) within the promoter region mainly contributed to the induction of stomatin by dex. Moreover, we found that hypoxia exposure did not affect membrane-associated actin, but decreased actin in cytoplasm in A549 cells. Inhibiting stomatin expression by stomatin siRNA significantly decreased dense of peripheral actin ring in hypoxia or dex treated A549 cells. Taken all together, these data indicated that dex and/or hypoxia significantly up-regulated the expression of stomatin in vivo and in vitro, which could stabilize membrane-associated actin in AEC. We suppose that the up-regulation of stomatin by hypoxia and dex may enhance the barrier function of alveolar epithelia and mediate the adaptive role of GC to hypoxia. PMID:23672602

  17. Isolation and characterization of a novel gene sfig in rat skeletal muscle up-regulated by spaceflight (STS-90)

    NASA Technical Reports Server (NTRS)

    Kano, Mihoko; Kitano, Takako; Ikemoto, Madoka; Hirasaka, Katsuya; Asanoma, Yuki; Ogawa, Takayuki; Takeda, Shinichi; Nonaka, Ikuya; Adams, Gregory R.; Baldwin, Kenneth M.; Oarada, Motoko; Kishi, Kyoichi; Nikawa, Takeshi

    2003-01-01

    We obtained the skeletal muscle of rats exposed to weightless conditions during a 16-day-spaceflight (STS-90). By using a differential display technique, we identified 6 up-regulated and 3 down-regulated genes in the gastrocnemius muscle of the spaceflight rats, as compared to the ground control. The up-regulated genes included those coding Casitas B-lineage lymphoma-b, insulin growth factor binding protein-1, titin and mitochondrial gene 16 S rRNA and two novel genes (function unknown). The down-regulated genes included those encoding RNA polymerase II elongation factor-like protein, NADH dehydrogenase and one novel gene (function unknown). In the present study, we isolated and characterized one of two novel muscle genes that were remarkably up-regulated by spaceflight. The deduced amino acid sequence of the spaceflight-induced gene (sfig) comprises 86 amino acid residues and is well conserved from Drosophila to Homo sapiens. A putative leucine-zipper structure located at the N-terminal region of sfig suggests that this gene may encode a transcription factor. The up-regulated expression of this gene, confirmed by Northern blot analysis, was observed not only in the muscles of spaceflight rats but also in the muscles of tail-suspended rats, especially in the early stage of tail-suspension when gastrocnemius muscle atrophy initiated. The gene was predominantly expressed in the kidney, liver, small intestine and heart. When rat myoblastic L6 cells were grown to 100% confluence in the cell culture system, the expression of sfig was detected regardless of the cell differentiation state. These results suggest that spaceflight has many genetic effects on rat skeletal muscle.

  18. Thrombin-mediated IL-10 up-regulation involves protease-activated receptor (PAR)-1 expression in human mononuclear leukocytes.

    PubMed

    Naldini, Antonella; Bernini, Claudia; Pucci, Annalisa; Carraro, Fabio

    2005-09-01

    Thrombin, the key enzyme of the coagulation cascade, exerts cellular effects through activation of the protease-activated receptors (PARs). Interleukin (IL)-10, besides its anti-inflammatory properties, is considered a major denominator of the immunosuppressive effect during human endotoxemia. We have recently shown that thrombin inhibits IL-12 production in human mononuclear cells and that such inhibition is accompanied by IL-10 up-regulation. To our knowledge, there are no data available to show that thrombin mediates IL-10 production by its interactions with PAR-1. We here report that human alpha-thrombin enhances IL-10 expression in human peripheral blood mononuclear cells and in established monocytic cell lines and that this up-regulation requires PAR-1 expression. The use of proteolytically inactive thrombin reveals that such enhancement requires thrombin proteolytic activity. Addition of PAR-1 agonist peptides, such as SFLLRN, results in a significant increase of IL-10 production. PAR-1 expression is required for thrombin-induced IL-10 production, as shown by experiments performed with antisense or sense PAR-1 oligonucleotides. Treatment with thrombin or SFLLRN of monocytic cell lines, such as U937 and Mono Mac-6, results in an increased IL-10 production. This suggests that the observed IL-10 up-regulation may be the result of a direct interaction with monocytes. The observation that thrombin-mediated up-regulation of IL-10 may require the expression of the PAR-1 receptor identifies a new, functional link between inflammation and coagulation. Our results may also contribute to better design therapeutic strategies to treat several disorders, characterized by the presence of inflammatory as well as coagulant responses. PMID:15961578

  19. Antitumor Effects of a Sirtuin Inhibitor, Tenovin-6, against Gastric Cancer Cells via Death Receptor 5 Up-Regulation

    PubMed Central

    Hirai, Sachiko; Endo, Shinji; Saito, Rie; Hirose, Mitsuaki; Ueno, Takunori; Suzuki, Hideo; Yamato, Kenji; Abei, Masato; Hyodo, Ichinosuke

    2014-01-01

    Up-regulated sirtuin 1 (SIRT1), an NAD+-dependent class III histone deacetylase, deacetylates p53 and inhibits its transcriptional activity, leading to cell survival. SIRT1 overexpression has been reported to predict poor survival in some malignancies, including gastric cancer. However, the antitumor effect of SIRT1 inhibition remains elusive in gastric cancer. Here, we investigated the antitumor mechanisms of a sirtuin inhibitor, tenovin-6, in seven human gastric cancer cell lines (four cell lines with wild-type TP53, two with mutant-type TP53, and one with null TP53). Interestingly, tenovin-6 induced apoptosis in all cell lines, not only those with wild-type TP53, but also mutant-type and null versions, accompanied by up-regulation of death receptor 5 (DR5). In the KatoIII cell line (TP53-null), DR5 silencing markedly attenuated tenovin-6-induced apoptosis, suggesting that the pivotal mechanism behind its antitumor effects is based on activation of the death receptor signal pathway. Although endoplasmic reticulum stress caused by sirtuin inhibitors was reported to induce DR5 up-regulation in other cancer cell lines, we could not find marked activation of its related molecules, such as ATF6, PERK, and CHOP, in gastric cancer cells treated with tenovin-6. Tenovin-6 in combination with docetaxel or SN-38 exerted a slight to moderate synergistic cytotoxicity against gastric cancer cells. In conclusion, tenovin-6 has potent antitumor activity against human gastric cancer cells via DR5 up-regulation. Our results should be helpful for the future clinical development of sirtuin inhibitors. PMID:25033286

  20. Antitumor effects of a sirtuin inhibitor, tenovin-6, against gastric cancer cells via death receptor 5 up-regulation.

    PubMed

    Hirai, Sachiko; Endo, Shinji; Saito, Rie; Hirose, Mitsuaki; Ueno, Takunori; Suzuki, Hideo; Yamato, Kenji; Abei, Masato; Hyodo, Ichinosuke

    2014-01-01

    Up-regulated sirtuin 1 (SIRT1), an NAD+-dependent class III histone deacetylase, deacetylates p53 and inhibits its transcriptional activity, leading to cell survival. SIRT1 overexpression has been reported to predict poor survival in some malignancies, including gastric cancer. However, the antitumor effect of SIRT1 inhibition remains elusive in gastric cancer. Here, we investigated the antitumor mechanisms of a sirtuin inhibitor, tenovin-6, in seven human gastric cancer cell lines (four cell lines with wild-type TP53, two with mutant-type TP53, and one with null TP53). Interestingly, tenovin-6 induced apoptosis in all cell lines, not only those with wild-type TP53, but also mutant-type and null versions, accompanied by up-regulation of death receptor 5 (DR5). In the KatoIII cell line (TP53-null), DR5 silencing markedly attenuated tenovin-6-induced apoptosis, suggesting that the pivotal mechanism behind its antitumor effects is based on activation of the death receptor signal pathway. Although endoplasmic reticulum stress caused by sirtuin inhibitors was reported to induce DR5 up-regulation in other cancer cell lines, we could not find marked activation of its related molecules, such as ATF6, PERK, and CHOP, in gastric cancer cells treated with tenovin-6. Tenovin-6 in combination with docetaxel or SN-38 exerted a slight to moderate synergistic cytotoxicity against gastric cancer cells. In conclusion, tenovin-6 has potent antitumor activity against human gastric cancer cells via DR5 up-regulation. Our results should be helpful for the future clinical development of sirtuin inhibitors. PMID:25033286

  1. Up-regulated uridine kinase gene identified by RLCS in the ventral horn after crush injury to rat sciatic nerves.

    PubMed

    Yuh, I; Yaoi, T; Watanabe, S; Okajima, S; Hirasawa, Y; Fushiki, S

    1999-12-01

    Rat sciatic nerve crush injury is one of the models commonly employed for studying the mechanisms of nerve regeneration. In this study, we analyzed the temporal change of gene expression after injury in this model, to elucidate the molecular mechanisms involved in nerve regeneration. First, a cDNA analysis method, Restriction Landmark cDNA Scanning (RLCS), was applied to cells in the ventral horn of the spinal cord during a 7-day period after the crush injury. A total of 1991 cDNA species were detected as spots on gels, and 37 of these were shown to change after the injury. Temporally changed patterns were classified into three categories: the continuously up-regulated type (10 species), the transiently up-regulated type (22 species), and the down-regulated type (5 species). These complex patterns of gene expression demonstrated after the injury suggest that precise regulation in molecular pathways is required for accomplishing nerve regeneration. Secondly, the rat homologue of uridine kinase gene was identified as one of the up-regulated genes. Northern blot analysis on rat ventral horn tissue and brain revealed that the UK gene had three transcripts with different sizes (4.3, 1. 4, and 1.35 kb, respectively). All of the transcripts, especially the 4.3 kb one, were up-regulated mainly in a bimodal fashion during the 28-day period after the injury. The RLCS method that we employed in the present study shows promise as a means to fully analyze molecular changes in nerve regeneration in detail. PMID:10581173

  2. Up-regulation of the embryonic self-renewal network through reversible polyploidy in irradiated p53-mutant tumour cells

    SciTech Connect

    Salmina, Kristine; Jankevics, Eriks; Huna, Anda; Perminov, Dmitry; Radovica, Ilze; Klymenko, Tetyana; Ivanov, Andrey; Jascenko, Elina; Scherthan, Harry; Cragg, Mark; Erenpreisa, Jekaterina

    2010-08-01

    We have previously documented that transient polyploidy is a potential cell survival strategy underlying the clonogenic re-growth of tumour cells after genotoxic treatment. In an attempt to better define this mechanism, we recently documented the key role of meiotic genes in regulating the DNA repair and return of the endopolyploid tumour cells (ETC) to diploidy through reduction divisions after irradiation. Here, we studied the role of the pluripotency and self-renewal stem cell genes NANOG, OCT4 and SOX2 in this polyploidy-dependent survival mechanism. In irradiation-resistant p53-mutated lymphoma cell-lines (Namalwa and WI-L2-NS) but not sensitive p53 wild-type counterparts (TK6), low background expression of OCT4 and NANOG was up-regulated by ionising radiation with protein accumulation evident in ETC as detected by OCT4/DNA flow cytometry and immunofluorescence (IF). IF analysis also showed that the ETC generate PML bodies that appear to concentrate OCT4, NANOG and SOX2 proteins, which extend into complex nuclear networks. These polyploid tumour cells resist apoptosis, overcome cellular senescence and undergo bi- and multi-polar divisions transmitting the up-regulated OCT4, NANOG and SOX2 self-renewal cassette to their descendents. Altogether, our observations indicate that irradiation-induced ETC up-regulate key components of germ-line cells, which potentially facilitate survival and propagation of the tumour cell population.

  3. Non-Thermal Plasma Treatment Diminishes Fungal Viability and Up-Regulates Resistance Genes in a Plant Host

    PubMed Central

    Panngom, Kamonporn; Lee, Sang Hark; Park, Dae Hoon; Sim, Geon Bo; Kim, Yong Hee; Uhm, Han Sup; Park, Gyungsoon; Choi, Eun Ha

    2014-01-01

    Reactive oxygen and nitrogen species can have either harmful or beneficial effects on biological systems depending on the dose administered and the species of organism exposed, suggesting that application of reactive species can possibly produce contradictory effects in disease control, pathogen inactivation and activation of host resistance. A novel technology known as atmospheric-pressure non-thermal plasma represents a means of generating various reactive species that adversely affect pathogens (inactivation) while simultaneously up-regulating host defense genes. The anti-microbial efficacy of this technology was tested on the plant fungal pathogen Fusarium oxysporum f.sp. lycopersici and its susceptible host plant species Solanum lycopercicum. Germination of fungal spores suspended in saline was decreased over time after exposed to argon (Ar) plasma for 10 min. Although the majority of treated spores exhibited necrotic death, apoptosis was also observed along with the up-regulation of apoptosis related genes. Increases in the levels of peroxynitrite and nitrite in saline following plasma treatment may have been responsible for the observed spore death. In addition, increased transcription of pathogenesis related (PR) genes was observed in the roots of the susceptible tomato cultivar (S. lycopercicum) after exposure to the same Ar plasma dose used in fungal inactivation. These data suggest that atmospheric-pressure non-thermal plasma can be efficiently used to control plant fungal diseases by inactivating fungal pathogens and up-regulating mechanisms of host resistance. PMID:24911947

  4. Acetylation of p53 Protein at Lysine 120 Up-regulates Apaf-1 Protein and Sensitizes the Mitochondrial Apoptotic Pathway.

    PubMed

    Yun, Tao; Yu, Kaiwen; Yang, ShuangShuang; Cui, Yifan; Wang, Zixi; Ren, Huiyu; Chen, She; Li, Lin; Liu, Xiaoyun; Fang, Min; Jiang, Xuejun

    2016-04-01

    The p53 tumor suppressor controls cell growth, metabolism, and death by regulating the transcription of various target genes. The target-specific transcriptional activity of p53 is highly regulated. Here we demonstrate that acetylation of p53 at Lys-120 up-regulates its transcriptional activity toward Apaf-1, a core component in the mitochondrial apoptotic pathway, and thus sensitizes caspase activation and apoptosis. We found that histone deacetylase (HDAC) inhibitors, including butyrate, augment Lys-120 acetylation of p53 and thus Apaf-1 expression by inhibiting HDAC1. In p53-null cells, transfection of wild-type but not K120R mutant p53 can restore the p53-dependent sensitivity to butyrate. Strikingly, transfection of acetylation-mimicking K120Q mutant p53 is sufficient to up-regulates Apaf-1 in a manner independent of butyrate treatment. Therefore, HDAC inhibitors can induce p53 acetylation at lysine 120, which in turn enhances mitochondrion-mediated apoptosis through transcriptional up-regulation of Apaf-1. PMID:26851285

  5. Protracted treatment with MDMA induces heteromeric nicotinic receptor up-regulation in the rat brain: an autoradiography study.

    PubMed

    Ciudad-Roberts, Andrés; Camarasa, Jorge; Pubill, David; Escubedo, Elena

    2014-08-01

    Previous studies indicate that 3,4-methylenedioxy-methamphetamine (MDMA, ecstasy) can induce a heteromeric nicotinic acetylcholine receptor (nAChR, mainly of α4β2 subtype) up-regulation. In this study we treated male Sprague-Dawley rats twice-daily for 10 days with either saline or MDMA (7 mg/kg) and sacrificed them the day after to perform [(125)I]Epibatidine binding autoradiograms on serial coronal slices. MDMA induced significant increases in nAChR density in the substantia nigra, ventral tegmental area, nucleus accumbens, olfactory tubercle, anterior caudate-putamen, somatosensory, motor, auditory and retrosplenial cortex, laterodorsal thalamus nuclei, amygdala, postsubiculum and pontine nuclei. These increases ranged from 3% (retrosplenial cortex) to 30 and 34% (amygdala and substantia nigra). No increased α4 subunit immunoreactivity was found in up-regulated areas compared with saline-treated rats, suggesting a post-translational mechanism as occurs with nicotine. The heteromeric nAChR up-regulation in certain areas could account, at least in part, for the reinforcing, sensitizing and psychiatric disorders observed after long-term consumption of MDMA.

  6. Isoform-Specific Up-Regulation of Plasma Membrane Ca2+ATPase Expression During Colon and Gastric Cancer Cell Differentiation

    PubMed Central

    Ribiczey, Polett; Tordai, Attila; Andrikovics, Hajnalka; Filoteo, Adelaida G.; Penniston, John T.; Enouf, Jocelyne; Enyedi, Ágnes; Papp, Béla; Kovács, Tünde

    2007-01-01

    Summary In this work we demonstrate a differentiation-induced up-regulation of the expression of plasma membrane Ca2+ATPase (PMCA) isoforms being present in various gastric/colon cancer cell types. We found PMCA1b as the major isoform in non-differentiated cancer cell lines, whereas the expression level of PMCA4b was significantly lower. Cell differentiation initiated with short chain fatty acids (SCFAs) and trichostatin A, or spontaneous differentiation of post-confluent cell cultures resulted in a marked induction of PMCA4b expression, while only moderately increased PMCA1b levels. Up-regulation of PMCA4b expression was demonstrated both at the protein and mRNA levels, and closely correlated with the induction of established differentiation markers. In contrast, the expression level of the Na+/K+-ATPase or that of the sarco/endoplasmic reticulum Ca2+ATPase 2 protein did not change significantly under these conditions. In membrane vesicles obtained from SCFA-treated gastric/colon cancer cells a marked increase in the PMCA-dependent Ca2+ transport activity was observed, indicating a general increase of PMCA function during the differentiation of these cancer cells. Because various PMCA isoforms display distinct functional characteristics, we suggest that up-regulated PMCA expression, together with a major switch in PMCA isoform pattern may significantly contribute to the differentiation of gastric/colon cancer cells. The analysis of PMCA expression may provide a new diagnostic tool for monitoring the tumor phenotype. PMID:17433436

  7. Pregnancy-associated plasma protein A up-regulated by progesterone promotes adhesion and proliferation of trophoblastic cells.

    PubMed

    Wang, Jiao; Liu, Shuai; Qin, Hua-Min; Zhao, Yue; Wang, Xiao-Qi; Yan, Qiu

    2014-01-01

    Embryo implantation and development is a complex biological process for the establishment of the successful pregnancy. Progesterone is a critical factor in the regulation of embryo adhesion to uterine endometrium and proliferation. Although it has been reported that pregnancy-associated plasma protein A (PAPPA) is increased in pregnant women, the relationship between progesterone and PAPPA, and the effects of PAPPA on embryo adhesion and proliferation are still not clear. The present results showed that the serum level of progesterone and PAPPA was closely correlated by ELISA assay (p<0.01). PAPPA was detected in the villi of early embryo by RT-PCR, Western blot, immunohistochemistry and immunofluorescent staining. Moreover, PAPPA was significantly up-regulated by progesterone in trophoblastic (JAR) cells by Real-time PCR and ELISA assay (p<0.01); while the expression was decreased by the progesterone receptor inhibitor RU486. The down-regulation of PAPPA by siRNA transfection or up-regulation of PAPPA by progesterone treatment significantly decreased or increased the adhesion rate of trophoblastic cells to human uterine epithelial cell lines (RL95-2 and HEC-1A), respectively (p<0.01), as well as the proliferation of trophoblastic cells. In conclusion, PAPPA is up-regulated by progesterone, which promotes the adhesion and proliferation potential of trophoblastic cells. PMID:24817938

  8. The juxtamembrane domain in ETV6/FLT3 is critical for PIM-1 up-regulation and cell proliferation

    SciTech Connect

    Vu, Hoang Anh; Xinh, Phan Thi; Kano, Yasuhiko; Tokunaga, Katsushi; Sato, Yuko

    2009-06-05

    We recently reported that the ETV6/FLT3 fusion protein conferred interleukin-3-independent growth on Ba/F3 cells. The present study has been conducted to assess role of the juxtamembrane domain of FLT3 for signal transduction and cell transformation. The wild-type ETV6/FLT3 fusion protein in transfected cells was a constitutively activated tyrosine kinase that led to up-regulation of PIM-1 and activations of STAT5, AKT, and MAPK. Deletion of the juxtamembrane domain abrogated interleukin-3-independent growth of the transfected cells and PIM-1 up-regulation, whereas it retained compatible levels of phosphorylations of STAT5, AKT, and MAPK. Further deletion of N-terminal region of the tyrosine kinase I domain of FLT3 completely abolished these phosphorylations. Our data indicate that the juxtamembrane domain of FLT3 in ETV6/FLT3 fusion protein is critical for cell proliferation and PIM-1 up-regulation that might be independent of a requirement for signaling through STAT5, MAPK, and AKT pathways.

  9. Proteolytic fragments of laminin promote excitotoxic neurodegeneration by up-regulation of the KA1 subunit of the kainate receptor.

    PubMed

    Chen, Zu-Lin; Yu, Huaxu; Yu, Wei-Ming; Pawlak, Robert; Strickland, Sidney

    2008-12-29

    Degradation of the extracellular matrix (ECM) protein laminin contributes to excitotoxic cell death in the hippocampus, but the mechanism of this effect is unknown. To study this process, we disrupted laminin gamma1 (lamgamma1) expression in the hippocampus. Lamgamma1 knockout (KO) and control mice had similar basal expression of kainate (KA) receptors, but the lamgamma1 KO mice were resistant to KA-induced neuronal death. After KA injection, KA1 subunit levels increased in control mice but were unchanged in lamgamma1 KO mice. KA1 levels in tissue plasminogen activator (tPA)-KO mice were also unchanged after KA, indicating that both tPA and laminin were necessary for KA1 up-regulation after KA injection. Infusion of plasmin-digested laminin-1 into the hippocampus of lamgamma1 or tPA KO mice restored KA1 up-regulation and KA-induced neuronal degeneration. Interfering with KA1 function with a specific anti-KA1 antibody protected against KA-induced neuronal death both in vitro and in vivo. These results demonstrate a novel pathway for neurodegeneration involving proteolysis of the ECM and KA1 KA receptor subunit up-regulation.

  10. Up-regulation of Fas (CD95) and induction of apoptosis in intestinal epithelial cells by nematode-derived molecules.

    PubMed

    Kuroda, Akio; Uchikawa, Ryuichi; Matsuda, Shinji; Yamada, Minoru; Tegoshi, Tatsuya; Arizono, Naoki

    2002-08-01

    Infection by the intestinal nematode Nippostrongylus brasiliensis induces acceleration of apoptosis in the small intestinal villus epithelial cells in vivo. In the present study, we examined whether worm extract or excretory-secretory product induces apoptosis in the rat intestinal epithelial cell line IEC-6 in vitro. In the presence of worm extract or excretory-secretory product (> or =6 microg/ml), IEC-6 cell growth was significantly suppressed, and there was a concomitant increase in the number of detached cells in culture dishes. Detached cells showed nuclear fragmentation, activation of caspase-3, and specific cleavage of poly(ADP-ribose) polymerase, suggesting that apoptosis was induced in these cells. Semiquantitative reverse transcription-PCR showed that expression of Fas (CD95) mRNA was up-regulated as early as 6 h after addition of excretory-secretory product, while Fas ligand expression and p53 expression were not up-regulated. Fluorescence-activated cell sorter analyses revealed a significant increase in Fas expression and a slight increase in FasL expression in IEC-6 cells cultured in the presence of excretory-secretory product, while control IEC-6 cells expressed neither Fas or FasL. These results indicated that N. brasiliensis worms produce and secrete biologically active molecules that trigger apoptosis in intestinal epithelial cells together with up-regulation of Fas expression, although the mechanism of induction of apoptosis remains to be elucidated. PMID:12117905

  11. Up-regulation of the iC3b receptor (CR3) is neither necessary nor sufficient to promote neutrophil aggregation.

    PubMed Central

    Philips, M R; Buyon, J P; Winchester, R; Weissmann, G; Abramson, S B

    1988-01-01

    The iC3b receptor (CR3) is required for neutrophil adhesive functions, including homotypic aggregation. Because stimuli that enhance neutrophil adhesion also induce up-regulation of surface CR3, it is widely held that these two responses are causally related. We have dissociated CR3 display (immunofluorescence) from CR3 function (aggregation). Neutrophils isolated at 4 degrees C and rewarmed to 37 degrees C up-regulated surface CR3 twofold, but did not aggregate. The kinetics of FMLP-induced CR3 up-regulation were discordant with those of aggregation. In the absence of extracellular divalent cations, CR3 expression increased twofold after exposure to FMLP, but neutrophils did not aggregate. FMLP elicited 3.5-fold more aggregation than the ionophore A23187, yet less than one-half as much CR3 up-regulation. 3 mM sodium salicylate inhibited aggregation 55 +/- 4%, but had no effect on CR3 up-regulation. Conversely, 1 mM tetracaine completely inhibited CR3 up-regulation, while significantly enhancing aggregation. Neutroplasts expressed CR3, but did not up-regulate the receptor; in contrast, FMLP induced CR3-dependent aggregation of neutroplasts. We conclude that, although constitutive surface CR3 is required for neutrophil aggregation, the up-regulation of CR3 is neither necessary nor sufficient to promote cell-cell adhesion. Images PMID:2841354

  12. Up-regulation of hypoxia-inducible factor-1α enhanced the cardioprotective effects of ischemic postconditioning in hyperlipidemic rats.

    PubMed

    Li, Xiaoyu; Zhao, Huanxin; Wu, Ye; Zhang, Suli; Zhao, Xiaoqin; Zhang, Yan; Wang, Jin; Wang, Jie; Liu, Huirong

    2014-02-01

    Hyperlipidemia is an independent risk factor in the development of ischemic heart disease, which can increase myocardial susceptibility to ischemia/reperfusion (I/R) injury. Ischemic postconditioning (PostC) has now been demonstrated as a novel strategy to harness nature's protection against myocardial I/R injury in normal conditions. However, the effect of PostC on hyperlipidemic animals remains elusive. It has been shown in our previous study that PostC reduces the myocardial I/R injury, and hypoxia-inducible factor-1α (HIF-1α) may play an important role in the cardioprotective mechanisms of PostC on normal rats. Here, we tested the hypothesis that the cardioprotection of PostC on hyperlipidemic rats is associated with the up-regulated HIF-1α expression. Male Wistar rats were fed with a high-fat diet for 8 weeks, and then randomly divided into five groups: sham, I/R, dimethyloxalylglycine (DMOG) + I/R, PostC, and DMOG + PostC group. The detrimental indices induced by I/R injury included infarct size, plasma creatine kinase (CK) activity and caspase-3 activity. The results showed that PostC could reduce the infarct size, when compared with the I/R group, which was consistent with the significant lower levels of plasma CK activity and caspase-3 activity, and that it increased the expression of HIF-1α in hyperlipidemic rats. When DMOG was given before PostC to up-regulate HIF-1α protein level, the degree of I/R injury was attenuated. In conclusion, these data suggested that the up-regulation of HIF-1α may be one of the cardioprotective mechanisms of PostC against I/R injury in hyperlipidemic rats.

  13. Involvement of the Up-regulated FoxO1 Expression in Follicular Granulosa Cell Apoptosis Induced by Oxidative Stress*

    PubMed Central

    Shen, Ming; Lin, Fei; Zhang, Jiaqing; Tang, Yiting; Chen, Wei-Kang; Liu, Honglin

    2012-01-01

    Follicular atresia is common in female mammalian ovaries, where most follicles undergo degeneration at any stage of growth and development. Oxidative stress gives rise to triggering granulosa cell apoptosis, which has been suggested as a major cause of follicular atresia. However, the underlying mechanism by which the oxidative stress induces follicular atresia remains unclear. FoxO transcription factors are known as critical mediators in the regulation of oxidative stress and apoptosis. In this study, the involvement of FoxO1 in oxidative stress-induced apoptosis of mouse follicular granulosa cells (MGCs) was investigated in vivo and in vitro. It was observed that increased apoptotic signals correlated with elevated expression of FoxO1 in MGCs when mice were treated with the oxidant. Correspondingly, the expressions of FoxO1 target genes, such as proapoptotic genes and antioxidative genes, were also up-regulated. In primary cultured MGCs, treatment with H2O2 led to FoxO1 nuclear translocation. Further studies with overexpression and knockdown of FoxO1 demonstrated the critical role of FoxO1 in the induction of MGC apoptosis by oxidative stress. Finally, inactivation of FoxO1 by insulin treatment confirmed that FoxO1 induced by oxidative stress played a pivotal role in up-regulating the expression of downstream apoptosis-related genes in MGCs. Our results suggest that up-regulation of FoxO1 by oxidative stress leads to apoptosis of granulosa cells, which eventually results in follicular atresia in mice. PMID:22669940

  14. Galectin-9 Protein Is Up-regulated in Astrocytes by Tumor Necrosis Factor and Promotes Encephalitogenic T-cell Apoptosis*

    PubMed Central

    Steelman, Andrew J.; Smith, Roger; Welsh, C. Jane; Li, Jianrong

    2013-01-01

    Demyelination and axonal damage in multiple sclerosis (MS) are thought to be a consequence of inflammatory processes that are perpetuated by activated glia and infiltrating leukocytes. Galectin-9 is a β-galactoside binding lectin capable of modulating immune responses and appears to be up-regulated in MS. However, its role in the pathogenesis of MS has yet to be determined. Here, we report that proinflammatory cytokines induce galectin-9 (Gal-9) expression in primary astrocytes and the mechanism by which TNF up-regulates Gal-9. Astrocytes did not express Gal-9 under basal conditions nor did IL-6, IL-10, or IL-13 trigger Gal-9 expression. In contrast, IL-1β, IFN-γ, and particularly TNF up-regulated Gal-9 in astrocytes. TNF-induced Gal-9 expression was dependent on TNF receptor 1 (TNFR1) as TNF failed to induce Gal-9 in TNFR1−/− astrocytes. Blockade of the JNK MAP kinase pathway with the JNK inhibitor SP600125 abrogated TNF-induced Gal-9, whereas p38 and MEK inhibitors had minimal effects. Furthermore, specific knockdown of c-Jun via siRNA in astrocytes before TNF treatment greatly suppressed Gal-9 transcription, suggesting that TNF induces astroglial Gal-9 through the TNF/TNFR1/JNK/cJun signaling pathway. Finally, utilizing astrocytes from Lgals9 mutant (Gal-9−/−) mice as well as a myelin basic protein-specific Tim-3+ encephalitogenic T-cell clone (LCN-8), we found that conditioned medium from TNF-stimulated Gal-9+/+ but not Gal-9−/− astrocytes increased the percentage of apoptotic encephalitogenic T-cells. Together, our results suggest that Gal-9 is induced in astrocytes by TNF via the JNK/c-Jun pathway and that astrocyte-derived Gal-9 may function as an immunoregulatory protein in response to ongoing neuroinflammation. PMID:23836896

  15. Activation of neurokinin-1 receptors up-regulates substance P and neurokinin-1 receptor expression in murine pancreatic acinar cells.

    PubMed

    Koh, Yung-Hua; Moochhala, Shabbir; Bhatia, Madhav

    2012-07-01

    Acute pancreatitis (AP) has been associated with an up-regulation of substance P (SP) and neurokinin-1 receptor (NK1R) in the pancreas. Increased SP-NK1R interaction was suggested to be pro-inflammatory during AP. Previously, we showed that caerulein treatment increased SP/NK1R expression in mouse pancreatic acinar cells, but the effect of SP treatment was not evaluated. Pancreatic acinar cells were obtained from pancreas of male swiss mice (25-30 g). We measured mRNA expression of preprotachykinin-A (PPTA) and NK1R following treatment of SP (10(-6) M). SP treatment increased PPTA and NK1R expression in isolated pancreatic acinar cells, which was abolished by pretreatment of a selective NK1R antagonist, CP96,345. SP also time dependently increased protein expression of NK1R. Treatment of cells with a specific NK1R agonist, GR73,632, up-regulated SP protein levels in the cells. Using previously established concentrations, pre-treatment of pancreatic acinar cells with Gö6976 (10 nM), rottlerin (5 μM), PD98059 (30 μM), SP600125 (30 μM) or Bay11-7082 (30 μM) significantly inhibited up-regulation of SP and NK1R. These observations suggested that the PKC-ERK/JNK-NF-κB pathway is necessary for the modulation of expression levels. In comparison, pre-treatment of CP96,345 reversed gene expression in SP-induced cells, but not in caerulein-treated cells. Overall, the findings in this study suggested a possible auto-regulatory mechanism of SP/NK1R expression in mouse pancreatic acinar cells, via activation of NK1R. Elevated SP levels during AP might increase the occurrence of a positive feedback loop that contributes to abnormally high expression of SP and NK1R.

  16. Oncogenic viral protein HPV E7 up-regulates the SIRT1 longevity protein in human cervical cancer cells.

    PubMed

    Allison, Simon J; Jiang, Ming; Milner, Jo

    2009-03-02

    Senescence is blocked in human cervical keratinocytes infected with high risk human papillomavirus (e.g. HPV type16). Viral oncoproteins HPV E6 and HPV E7 access the cell cycle via cellular p53 and retinoblastoma proteins respectively. Previously we have shown that HPV E7, not HPV E6, is also responsible for cervical cancer cell survival (SiHa cells; HPV type16). We now present evidence that SIRT1, an aging-related NAD-dependent deacetylase, mediates HPV E7 survival function in SiHa cervical cancer cells. Moreover, HPV E7 up-regulates SIRT1 protein when expressed in primary human keratinocytes. Conversely, SIRT1 levels decrease following RNAi-mediated silencing of HPV E7 in SiHa cells. Silencing HPV E6 has no effect on SIRT1 but, as expected, causes marked accumulation of p53 protein accompanied by p53-mediated up-regulation of p21. However, p53 acetylation (K382Ac) was barely detectable. Since p53 is a known SIRT1 substrate we propose that elevated SIRT1 levels (induced by HPV E7) attenuate p53 pro-apoptotic capacity via its de-acetylation. Our discovery that HPV E7 up-regulates SIRT1 links a clinically important oncogenic virus with the multi-functional SIRT1 protein. This link may open the way for a more in-depth understanding of the process of HPV-induced malignant transformation and also of the inter-relationships between aging and cancer.

  17. Up-regulation of endothelial monocyte chemoattractant protein-1 by coplanar PCB77 is caveolin-1-dependent

    SciTech Connect

    Majkova, Zuzana; Smart, Eric; Toborek, Michal; Hennig, Bernhard

    2009-05-15

    Atherosclerosis, the primary cause of heart disease and stroke is initiated in the vascular endothelium, and risk factors for its development include environmental exposure to persistent organic pollutants. Caveolae are membrane microdomains involved in regulation of many signaling pathways, and in particular in endothelial cells. We tested the hypothesis that intact caveolae are required for coplanar PCB77-induced up-regulation of monocyte chemoattractant protein-1 (MCP-1), an endothelium-derived chemokine that attracts monocytes into sub-endothelial space in early stages of the atherosclerosis development. Atherosclerosis-prone LDL-R{sup -/-} mice (control) or caveolin-1{sup -/-}/LDL-R{sup -/-} mice were treated with PCB77. PCB77 induced aortic mRNA expression and plasma protein levels of MCP-1 in control, but not caveolin-1{sup -/-}/LDL-R{sup -/-} mice. To study the mechanism of this effect, primary endothelial cells were used. PCB77 increased MCP-1 levels in endothelial cells in a time- and concentration-dependent manner. This effect was abolished by caveolin-1 silencing using siRNA. Also, MCP-1 up-regulation by PCB77 was prevented by inhibiting p38 and c-Jun N-terminal kinase (JNK), but not ERK1/2, suggesting regulatory functions via p38 and JNK MAPK pathways. Finally, pre-treatment of endothelial cells with the aryl hydrocarbon receptor (AhR) inhibitor {alpha}-naphthoflavone ({alpha}-NF) partially blocked MCP-1 up-regulation. Thus, our data demonstrate that coplanar PCB77 can induce MCP-1 expression by endothelial cells and that this effect is mediated by AhR, as well as p 38 and JNK MAPK pathways. Intact caveolae are required for these processes both in vivo and in vitro. This further supports a key role for caveolae in vascular inflammation induced by persistent organic pollutants.

  18. Mitochondrial genes for heme-dependent respiratory chain complexes are up-regulated after depletion of Wolbachia from filarial nematodes.

    PubMed

    Strübing, Uta; Lucius, Richard; Hoerauf, Achim; Pfarr, Kenneth M

    2010-08-15

    The filarial nematodes Brugia malayi, Wuchereria bancrofti and Onchocerca volvulus cause elephantiasis or dermatitis and blindness resulting in severe morbidity. Annually, 1.3 billion people are at risk of infection. Targeting the essential Wolbachia endobacteria of filarial nematodes with doxycycline has proven to be an effective therapy resulting in a block in embryogenesis, worm development and macrofilaricidal effects. However, doxycycline is contraindicated for a large portion of the at risk population. To identify new targets for anti-wolbachial therapy, understanding the molecular basis of the Wolbachia-filaria symbiosis is required. Using the B. malayi microarray we identified differentially expressed genes in the rodent filaria Litomosoides sigmodontis after depletion of Wolbachia which might have a role in symbiosis. The microarray data were filtered for regulated genes with a false discovery rate <5% and a > or = 2-fold-change. Most of the genes were differentially expressed at day 36 of tetracycline treatment, when 99.8% of Wolbachia were depleted. Several classes of genes were affected, including genes for translation, transcription, folding/sorting of proteins, motility, structure and metabolic and signalling pathways. Quantitative PCR validated 60% of the genes found to be regulated in the microarray. A nuclear encoded heme-binding protein of the globin family was up-regulated upon loss of Wolbachia. Interestingly, mitochondrial encoded subunits of respiratory chain complexes containing heme and riboflavin were also up-regulated. No change in the expression of these genes was seen in tetracycline treated Wolbachia-free Acanthocheilonema viteae. As Wolbachia synthesise heme and filaria do not, we hypothesise that without the endosymbionts no functional heme-containing enzymes can be formed, leading to loss of energy metabolism which then results in up-regulation of the mitochondrial encoded subunits in an attempt to correct the deviation from

  19. A peptide that ameliorates lupus up-regulates the diminished expression of early growth response factors 2 and 3.

    PubMed

    Sela, Uri; Dayan, Molly; Hershkoviz, Rami; Lider, Ofer; Mozes, Edna

    2008-02-01

    Expansion of autoreactive T cells and their resistance to anergy was demonstrated in systemic lupus erythematosus (SLE). A pair of transcription factors, early growth response 2 (Egr-2) and 3 (Egr-3), are negative regulators of T cell activation that were shown to be important in anergy. A peptide (designated hCDR1 for human CDR1) based on the CDR-1 of an anti-DNA Ab ameliorated SLE in both induced and spontaneous lupus models. Our objectives were to determine the expression levels of Egr-2 and Egr-3 in autoreactive T cells following immunization with the lupus-inducing anti-DNA Ab that bears a common Id designated 16/6Id and also in a full-blown SLE and to determine the effect of hCDR1 on these transcription factors. We demonstrated diminished expression levels of Egr-2 and Egr-3 mRNA both early after immunization with the 16/6Id and in SLE-afflicted (NZB x NZW)F1 (New Zealand Black and New Zealand White) mice. Furthermore, by down-regulating Akt phosphorylation and up-regulating TGFbeta secretion, treatment with hCDR1 significantly up-regulated Egr-2 and Egr-3 expression. This was associated with an increased expression of the E3 ligase Cbl-b. Inhibition of Akt in T cells of immunized mice decreased, whereas silencing of the Egr-2 and Egr-3 in T cells of hCDR1-treated mice increased IFN-gamma secretion. Thus, hCDR1 down-regulates Akt phosphorylation, which leads to up-regulated expression of T cell Egr-2 and Egr-3, resulting in the inhibition of IFN-gamma secretion that is required for the maintenance of SLE. PMID:18209054

  20. Signal pathways in up-regulation of chemokines by tyrosine kinase MER/NYK in prostate cancer cells.

    PubMed

    Wu, Yi-Mi; Robinson, Dan R; Kung, Hsing-Jien

    2004-10-15

    The AXL/UFO family of tyrosine kinases is characterized by a common N-CAM (neural adhesion molecule)-related extracellular domain and a common ligand, GAS6 (growth arrest-specific protein 6). Family members are prone to transcriptional regulation and carry out diverse functions including the regulation of cell adhesion, migration, phagocytosis, and survival. In this report, we describe a new role of MER/N-CAM-related kinase (NYK), a member of the AXL family of kinases, in the up-regulation of chemokines in prostate cancer cells. We show that NYK has elevated expression in a subset of tumor specimens and prostate cancer cell lines. Activation of NYK in the prostate cancer cell line DU145 does not cause a mitogenic effect; instead, it causes a differentiation phenotype. Microarray analysis revealed that NYK is a strong inducer of endocrine factors including interleukin (IL)-8 and several other angiogenic CXC chemokines as well as bone morphogenic factors. The dramatic increase of IL-8 expression is seen at both transcriptional and posttranscriptional levels. The downstream signals engaged by NYK were characterized, and those responsible for the up-regulation of IL-8 transcription were defined. In contrast to IL-1alpha, NYK-induced up-regulation of IL-8 in DU145 depends on the mitogen-activated protein kinase kinase/extracellular signal-regulated kinase/Jun/Fos pathway, but not phosphoinositide 3'-kinase/nuclear factor-kappaB. These data define a new function of the AXL family of kinases and suggest a potential role of NYK in prostate cancer progression. PMID:15492251

  1. Hypoxia Suppresses Spontaneous Mineralization and Osteogenic Differentiation of Mesenchymal Stem Cells via IGFBP3 Up-Regulation

    PubMed Central

    Kim, Ji Hye; Yoon, Sei Mee; Song, Sun U.; Park, Sang Gyu; Kim, Won-Serk; Park, In Guk; Lee, Jinu; Sung, Jong-Hyuk

    2016-01-01

    Hypoxia has diverse stimulatory effects on human adipose-derived stem cells (ASCs). In the present study, we investigated whether hypoxic culture conditions (2% O2) suppress spontaneous mineralization and osteogenic differentiation of ASCs. We also investigated signaling pathways and molecular mechanisms involved in this process. We found that hypoxia suppressed spontaneous mineralization and osteogenic differentiation of ASCs, and up-regulated mRNA and protein expression of Insulin-like growth factor binding proteins (IGFBPs) in ASCs. Although treatment with recombinant IGFBPs did not affect osteogenic differentiation of ASCs, siRNA-mediated inhibition of IGFBP3 attenuated hypoxia-suppressed osteogenic differentiation of ASCs. In contrast, overexpression of IGFBP3 via lentiviral vectors inhibited ASC osteogenic differentiation. These results indicate that hypoxia suppresses spontaneous mineralization and osteogenic differentiation of ASCs via intracellular IGFBP3 up-regulation. We determined that reactive oxygen species (ROS) generation followed by activation of the MAPK and PI3K/Akt pathways play pivotal roles in IGFBP3 expression under hypoxia. For example, ROS scavengers and inhibitors for MAPK and PI3K/Akt pathways attenuated the hypoxia-induced IGFBP3 expression. Inhibition of Elk1 and NF-κB through siRNA transfection also led to down-regulation of IGFBP3 mRNA expression. We next addressed the proliferative potential of ASCs with overexpressed IGFBP3, but IGFBP3 overexpression reduced the proliferation of ASCs. In addition, hypoxia reduced the osteogenic differentiation of bone marrow-derived clonal mesenchymal stem cells. Collectively, our results indicate that hypoxia suppresses the osteogenic differentiation of mesenchymal stem cells via IGFBP3 up-regulation. PMID:27563882

  2. The prevention and treatment of hypoadiponectinemia-associated human diseases by up-regulation of plasma adiponectin.

    PubMed

    Hossain, Md Murad; Mukheem, Abdul; Kamarul, Tunku

    2015-08-15

    Hypoadiponectinemia is characterized by low plasma adiponectin levels that can be caused by genetic factors, such as single nucleotide polymorphisms (SNPs) and mutations in the adiponectin gene or by visceral fat deposition/obesity. Reports have suggested that hypoadiponectinemia is associated with dyslipidemia, hypertension, hyperuricemia, metabolic syndrome, atherosclerosis, type 2 diabetes mellitus and various cardiovascular diseases. Previous studies have highlighted several potential strategies to up-regulate adiponectin secretion and function, including visceral fat reduction through diet therapy and exercise, administration of exogenous adiponectin, treatment with peroxisome proliferator-activating receptor gamma (PPARγ) agonists (e.g., thiazolidinediones (TZDs)) and ligands (e.g., bezafibrate and fenofibrate) or the blocking of the renin-angiotensin system. Likewise, the up-regulation of the expression and stimulation of adiponectin receptors by using adiponectin receptor agonists would be an effective method to treat obesity-related conditions. Notably, adiponectin is an abundantly expressed bioactive protein that also exhibits a wide spectrum of biological properties, such as insulin-sensitizing, anti-diabetic, anti-inflammatory and anti-atherosclerotic activities. Although targeting adiponectin and its receptors has been useful for treating diabetes and other metabolic-related diseases in experimental studies, current drug development based on adiponectin/adiponectin receptors for clinical applications is scarce, and there is a lack of available clinical trial data. This comprehensive review discusses the strategies that are presently being pursued to harness the potential of adiponectin up-regulation. In addition, we examined the current status of drug development and its potential for clinical applications. PMID:25818192

  3. Estrogen-dependent up-regulation of TRPA1 and TRPV1 receptor proteins in the rat endometrium.

    PubMed

    Pohóczky, Krisztina; Kun, József; Szalontai, Bálint; Szőke, Éva; Sághy, Éva; Payrits, Maja; Kajtár, Béla; Kovács, Krisztina; Környei, József László; Garai, János; Garami, András; Perkecz, Anikó; Czeglédi, Levente; Helyes, Zsuzsanna

    2016-02-01

    Transient receptor potential ankyrin 1 (TRPA1) and vanilloid 1 (TRPV1) receptors expressed predominantly in sensory nerves are activated by inflammatory stimuli and mediate inflammation and pain. Although they have been shown in the human endometrium, their regulation and function are unknown. Therefore, we investigated their estrogen- and progesterone-dependent alterations in the rat endometrium in comparison with the estrogen-regulated inflammatory cytokine macrophage migration inhibitory factor (MIF). Four-week-old (sexually immature) and four-month-old (sexually mature) female rats were treated with the non-selective estrogen receptor (ER) agonist diethylstilboestrol (DES), progesterone and their combination, or ovariectomized. RT-PCR and immunohistochemistry were performed to determine mRNA and protein expression levels respectively. Channel function was investigated with ratiometric [Ca(2+)]i measurement in cultured primary rat endometrial cells. Both TRP receptors and MIF were detected in the endometrium at mRNA and protein levels, and their localizations were similar. Immunostaining was observed in the immature epithelium, while stromal, glandular and epithelial positivity were observed in adults. Functionally active TRP receptor proteins were shown in endometrial cells by activation-induced calcium influx. In adults, Trpa1 and Trpv1 mRNA levels were significantly up-regulated after DES treatment. TRPA1 increased after every treatment, but TRPV1 remained unchanged following the combined treatment and ovariectomy. In immature rats, DES treatment resulted in increased mRNA expression of both channels and elevated TRPV1 immunopositivity. MIF expression changed in parallel with TRPA1/TRPV1 in most cases. DES up-regulated Trpa1, Trpv1 and Mif mRNA levels in endometrial cell cultures, but 17β-oestradiol having ERα-selective potency increased only the expression of Trpv1. We provide the first evidence for TRPA1/TRPV1 expression and their estrogen-induced up-regulation

  4. Up regulated expression of tumour necrosis factor α converting enzyme in peripheral monocytes of patients with early systemic sclerosis

    PubMed Central

    Bohgaki, T; Amasaki, Y; Nishimura, N; Bohgaki, M; Yamashita, Y; Nishio, M; Sawada, K; Jodo, S; Atsumi, T; Koike, T

    2005-01-01

    Background: Systemic sclerosis (SSc) is accompanied by abnormalities in humoral and cellular immune systems. Objective: To determine the genes specifically expressed in the immune system in SSc by analysis of the gene expression profile of peripheral blood mononuclear cells (PBMC) from patients with SSc, including those treated with haematopoietic stem cell transplantation (HSCT). Additionally, to investigate the clinical significance of the up regulation of tumour necrosis factor α (TNFα) converting enzyme (TACE). Methods: PBMC from patients with SSc (n = 23) and other autoimmune diseases (systemic lupus erythematosus (SLE, n = 16), rheumatoid arthritis (RA, n = 29)), and from disease-free controls (n = 36) were examined. Complementary DNA arrays were used to evaluate gene expression of PBMC, in combination with real time quantitative polymerase chain reactions. TACE protein expression in PBMC was examined by fluorescence activated cell sorter (FACS). Results: In patients with SSc 118 genes were down regulated after HSCT. Subsequent comparative analysis of SSc without HSCT and healthy controls indicated SSc-specific up regulation for three genes: monocyte chemoattractant protein-3 (p = 0.0015), macrophage inflammatory protein 3α (p = 0.0339), and TACE (p = 0.0251). In the FACS analysis, TACE protein was mainly expressed on CD14+ monocytes both in patients with SSc and controls. TACE expression on CD14+ cells was significantly increased in patients with early SSc (p = 0.0096), but not in those with chronic SSc, SLE, or RA. TACE protein levels in SSc monocytes correlated with the intracellular CD68 levels (p = 0.0016). Conclusions: Up regulation of TACE expression was a unique profile in early SSc, and may affect the function of TNFα and other immunoregulatory molecules. PMID:16014681

  5. Activation of neurokinin-1 receptors up-regulates substance P and neurokinin-1 receptor expression in murine pancreatic acinar cells

    PubMed Central

    Koh, Yung-Hua; Moochhala, Shabbir; Bhatia, Madhav

    2012-01-01

    Abstract Acute pancreatitis (AP) has been associated with an up-regulation of substance P (SP) and neurokinin-1 receptor (NK1R) in the pancreas. Increased SP-NK1R interaction was suggested to be pro-inflammatory during AP. Previously, we showed that caerulein treatment increased SP/NK1R expression in mouse pancreatic acinar cells, but the effect of SP treatment was not evaluated. Pancreatic acinar cells were obtained from pancreas of male swiss mice (25–30 g). We measured mRNA expression of preprotachykinin-A (PPTA) and NK1R following treatment of SP (10−6M). SP treatment increased PPTA and NK1R expression in isolated pancreatic acinar cells, which was abolished by pretreatment of a selective NK1R antagonist, CP96,345. SP also time dependently increased protein expression of NK1R. Treatment of cells with a specific NK1R agonist, GR73,632, up-regulated SP protein levels in the cells. Using previously established concentrations, pre-treatment of pancreatic acinar cells with Gö6976 (10 nM), rottlerin (5 μM), PD98059 (30 μM), SP600125 (30 μM) or Bay11-7082 (30 μM) significantly inhibited up-regulation of SP and NK1R. These observations suggested that the PKC-ERK/JNK-NF-κB pathway is necessary for the modulation of expression levels. In comparison, pre-treatment of CP96,345 reversed gene expression in SP-induced cells, but not in caerulein-treated cells. Overall, the findings in this study suggested a possible auto-regulatory mechanism of SP/NK1R expression in mouse pancreatic acinar cells, via activation of NK1R. Elevated SP levels during AP might increase the occurrence of a positive feedback loop that contributes to abnormally high expression of SP and NK1R. PMID:22040127

  6. 3'5'-cyclic adenosine monophosphate-dependent up-regulation of phosphodiesterase type 3A in porcine cumulus cells.

    PubMed

    Sasseville, Maxime; Côté, Nancy; Vigneault, Christian; Guillemette, Christine; Richard, François J

    2007-04-01

    The means by which cumulus cells react to gonadotropin stimulation and regulate the subsequent production and degradation of cAMP are largely unknown. In this article, we report that cyclic nucleotide phosphodiesterase (PDE) type 3A (Pde3a) is transcriptionally regulated in porcine cumulus cells by a cAMP-dependent pathway during in vitro maturation (IVM). cAMP-PDE activity was increased in the cumulus-oocyte complex (COC) after 10 h of IVM, and 78% of this increase was sensitive to a Pde3-specific inhibitor, cilostamide. Although no variation was observed in the oocyte, cilostamide-sensitive cAMP-PDE activity increased in the cumulus cells after IVM. This was supported by Western blotting, which showed that the intensity of a 135-kDa anti-Pde3a immunoreactive band was increased in COC after IVM. The Pde3a mRNA level was up-regulated 28-fold in the COC after 4 h of IVM and remained high up to 12 h. The mRNA up-regulation and increased activity were inhibited by an RNA synthesis inhibitor, alpha-amanitin. The cilostamide-sensitive increase in PDE activity was inhibited by a protein synthesis inhibitor, cycloheximide. Pregnant mare serum gonadotropin (PMSG) caused dose-dependent activation of Pde3. The PMSG-dependent increase in Pde3 activity and Pde3a mRNA were mimicked by the adenylyl cyclase activator forskolin or prostaglandin E2. PMSG-dependent Pde3 activation was inhibited by the protein kinase A-specific inhibitor H89. Collectively, our results show for the first time that degradation of the intracellular cyclic nucleotide by Pde3a is transcriptionally up-regulated via a cAMP-dependent pathway in cumulus cells, suggesting that it has a functional role during the ovulatory gonadotropin surge.

  7. Hypoxia Suppresses Spontaneous Mineralization and Osteogenic Differentiation of Mesenchymal Stem Cells via IGFBP3 Up-Regulation.

    PubMed

    Kim, Ji Hye; Yoon, Sei Mee; Song, Sun U; Park, Sang Gyu; Kim, Won-Serk; Park, In Guk; Lee, Jinu; Sung, Jong-Hyuk

    2016-01-01

    Hypoxia has diverse stimulatory effects on human adipose-derived stem cells (ASCs). In the present study, we investigated whether hypoxic culture conditions (2% O₂) suppress spontaneous mineralization and osteogenic differentiation of ASCs. We also investigated signaling pathways and molecular mechanisms involved in this process. We found that hypoxia suppressed spontaneous mineralization and osteogenic differentiation of ASCs, and up-regulated mRNA and protein expression of Insulin-like growth factor binding proteins (IGFBPs) in ASCs. Although treatment with recombinant IGFBPs did not affect osteogenic differentiation of ASCs, siRNA-mediated inhibition of IGFBP3 attenuated hypoxia-suppressed osteogenic differentiation of ASCs. In contrast, overexpression of IGFBP3 via lentiviral vectors inhibited ASC osteogenic differentiation. These results indicate that hypoxia suppresses spontaneous mineralization and osteogenic differentiation of ASCs via intracellular IGFBP3 up-regulation. We determined that reactive oxygen species (ROS) generation followed by activation of the MAPK and PI3K/Akt pathways play pivotal roles in IGFBP3 expression under hypoxia. For example, ROS scavengers and inhibitors for MAPK and PI3K/Akt pathways attenuated the hypoxia-induced IGFBP3 expression. Inhibition of Elk1 and NF-κB through siRNA transfection also led to down-regulation of IGFBP3 mRNA expression. We next addressed the proliferative potential of ASCs with overexpressed IGFBP3, but IGFBP3 overexpression reduced the proliferation of ASCs. In addition, hypoxia reduced the osteogenic differentiation of bone marrow-derived clonal mesenchymal stem cells. Collectively, our results indicate that hypoxia suppresses the osteogenic differentiation of mesenchymal stem cells via IGFBP3 up-regulation. PMID:27563882

  8. Adenosine A2A Receptor Up-Regulates Retinal Wave Frequency via Starburst Amacrine Cells in the Developing Rat Retina

    PubMed Central

    Huang, Pin-Chien; Hsiao, Yu-Tien; Kao, Shao-Yen; Chen, Ching-Feng; Chen, Yu-Chieh; Chiang, Chung-Wei; Lee, Chien-fei; Lu, Juu-Chin; Chern, Yijuang; Wang, Chih-Tien

    2014-01-01

    Background Developing retinas display retinal waves, the patterned spontaneous activity essential for circuit refinement. During the first postnatal week in rodents, retinal waves are mediated by synaptic transmission between starburst amacrine cells (SACs) and retinal ganglion cells (RGCs). The neuromodulator adenosine is essential for the generation of retinal waves. However, the cellular basis underlying adenosine's regulation of retinal waves remains elusive. Here, we investigated whether and how the adenosine A2A receptor (A2AR) regulates retinal waves and whether A2AR regulation of retinal waves acts via presynaptic SACs. Methodology/Principal Findings We showed that A2AR was expressed in the inner plexiform layer and ganglion cell layer of the developing rat retina. Knockdown of A2AR decreased the frequency of spontaneous Ca2+ transients, suggesting that endogenous A2AR may up-regulate wave frequency. To investigate whether A2AR acts via presynaptic SACs, we targeted gene expression to SACs by the metabotropic glutamate receptor type II promoter. Ca2+ transient frequency was increased by expressing wild-type A2AR (A2AR-WT) in SACs, suggesting that A2AR may up-regulate retinal waves via presynaptic SACs. Subsequent patch-clamp recordings on RGCs revealed that presynaptic A2AR-WT increased the frequency of wave-associated postsynaptic currents (PSCs) or depolarizations compared to the control, without changing the RGC's excitability, membrane potentials, or PSC charge. These findings suggest that presynaptic A2AR may not affect the membrane properties of postsynaptic RGCs. In contrast, by expressing the C-terminal truncated A2AR mutant (A2AR-ΔC) in SACs, the wave frequency was reduced compared to the A2AR-WT, but was similar to the control, suggesting that the full-length A2AR in SACs is required for A2AR up-regulation of retinal waves. Conclusions/Significance A2AR up-regulates the frequency of retinal waves via presynaptic SACs, requiring its full

  9. Methamphetamine acutely inhibits voltage-gated calcium channels but chronically up-regulates L-type channels.

    PubMed

    Andres, Marilou A; Cooke, Ian M; Bellinger, Frederick P; Berry, Marla J; Zaporteza, Maribel M; Rueli, Rachel H; Barayuga, Stephanie M; Chang, Linda

    2015-07-01

    In neurons, calcium (Ca(2+) ) channels regulate a wide variety of functions ranging from synaptic transmission to gene expression. They also induce neuroplastic changes that alter gene expression following psychostimulant administration. Ca(2+) channel blockers have been considered as potential therapeutic agents for the treatment of methamphetamine (METH) dependence because of their ability to reduce drug craving among METH users. Here, we studied the effects of METH exposure on voltage-gated Ca(2+) channels using SH-SY5Y cells as a model of dopaminergic neurons. We found that METH has different short- and long-term effects. A short-term effect involves immediate (< 5 min) direct inhibition of Ca(2+) ion movements through Ca(2+) channels. Longer exposure to METH (20 min or 48 h) selectively up-regulates the expression of only the CACNA1C gene, thus increasing the number of L-type Ca(2+) channels. This up-regulation of CACNA1C is associated with the expression of the cAMP-responsive element-binding protein (CREB), a known regulator of CACNA1C gene expression, and the MYC gene, which encodes a transcription factor that putatively binds to a site proximal to the CACNA1C gene transcription initiation site. The short-term inhibition of Ca(2+) ion movement and later, the up-regulation of Ca(2+) channel gene expression together suggest the operation of cAMP-responsive element-binding protein- and C-MYC-mediated mechanisms to compensate for Ca(2+) channel inhibition by METH. Increased Ca(2+) current density and subsequent increased intracellular Ca(2+) may contribute to the neurodegeneration accompanying chronic METH abuse. Methamphetamine (METH) exposure has both short- and long-term effects. Acutely, methamphetamine directly inhibits voltage-gated calcium channels. Chronically, neurons compensate by up-regulating the L-type Ca(2+) channel gene, CACNA1C. This compensatory mechanism is mediated by transcription factors C-MYC and CREB, in which CREB is linked to the

  10. Cadmium at high dose perturbs growth, photosynthesis and nitrogen metabolism while at low dose it up regulates sulfur assimilation and antioxidant machinery in garden cress (Lepidium sativum L.).

    PubMed

    Gill, Sarvajeet Singh; Khan, Nafees A; Tuteja, Narendra

    2012-01-01

    Metal contamination of soils has become a worldwide problem and great environmental threat, as these metals accumulate in soils and plants in excess, and enter the food chain. Increased cadmium (Cd) uptake from contaminated soils leads to altered plant metabolism and limits the crop productivity. The experimental crop, Lepidium sativum L. (Garden Cress, Family: Brassicaceae) is a medicinally and economically important plant. An experiment was conducted to examine the effect of different concentrations of Cd (0, 25, 50 or 100 mg kg(-1) soil) on the performance of L. sativum. Cd accumulation in roots and leaves (roots>leaves) increased with the increaseing Cd concentration in soil. High Cd concentration (100mg Cd kg(-1) soil) inhibited the leaf area and plant dry mass and significant decline in net photosynthetic rate (P(N)), stomatal conductance (gs), intercellular CO(2) (Ci), chlorophyll (Chl a, Chl b, total Chl) content, carbonic anhydrase (CA; E.C. 4.2.1.1) activity, nitrate reductase (NR; E.C. 1.6.6.1) activity and nitrogen (N) content was also observed. However, ATP-sulfurylase (ATP-S; EC. 2.7.7.4) activity, sulfur (S) content and activities of antioxidant enzymes such as superoxide dismutase (SOD; E.C. 1.15.1.1); catalase (CAT; E.C. 1.11.1.6); ascorbate peroxidase (APX; E.C. 1.11.1.11) and glutathione reductase (GR; E.C. 1.6.4.2) and glutathione (GSH) content were increased. Specifically, the decrease in NR activity and N content showed that Cd affects N metabolism negatively; whereas, the increase in ATP-S activity and S content suggests the up-regulation of S assimilation pathway for possible Cd tolerance in coordination with enhanced activities of antioxidant enzymes and GSH. High Cd concentration (100mg Cd kg(-1) soil) perturbs the L. sativum growth by interfering with the photosynthetic machinery and disrupting the coordination between carbon, N and S metabolism. On the other hand, at low Cd concentration (25mg Cd kg(-1) soil) co-ordination of S and N

  11. Up-Regulation of microRNA-210 is Associated with Spermatogenesis by Targeting IGF2 in Male Infertility

    PubMed Central

    Tang, Dongdong; Huang, Yuanyuan; Liu, Weiqun; Zhang, Xiansheng

    2016-01-01

    Background MicroRNAs (miRNAs) play pivotal roles in spermatogenesis. MicroRNA-210 (miR-210) expression was up-regulated in the testes of sterile men with non-obstructive azoospermia (NOA). However, the underlying mechanisms of miR-210 involved in the spermatogenesis in patients with NOA are unknown. Material/Methods Expression of miR-210 and insulin-like growth factor II (IGF2) in the testes of NOA cases (only including maturation arrest and hypospermatogenesis) were detected in this study. We carried out in vitro experiments to determine if IGF2 was directly targeted by miR-210 in NT2 cells. Results Compared with obstructive azoospermia (OA) as normal control, our results suggest that miR-210 was significantly up-regulated in testis of patients with NOA (P<0.05), and IGF2 was down-regulated, but without a significant difference. The results also indicated that IGF2 was directly targeted by miR-210 in NT2 cells. Conclusions The results showed that miR-210 was involved in spermatogenesis by targeting IGF2 in male infertility. PMID:27535712

  12. Up-regulation of a cysteine protease accompanies the ethylene-insensitive senescence of daylily (Hemerocallis) flowers.

    PubMed

    Valpuesta, V; Lange, N E; Guerrero, C; Reid, M S

    1995-06-01

    The flowers of daylily (Hemerocallis x hybrida cv. Cradle Song) open at midnight, start to senesce 12 h later, and are completely senescent by the following midnight. Differential screening of a cDNA library constructed from tepals of flowers showing incipient senescence revealed 25 clones that were strongly up-regulated in senescent tepals. Re-screening and interactive Southern analysis of these clones revealed 3 families of up-regulated clones. Transcripts of one clone, SEN10, were not detectable at midnight, but increased dramatically as senescence proceeded. The derived amino acid sequence of the full-length cDNA (SEN102) has strong homology with cysteine proteases that have been reported from other plant tissues. The sequence contains a secretory signal peptide and a probable prosequence upstream of the mature protein. Amino acids critical to the active site and structure of cysteine proteases are conserved, and the C-terminus of the polypeptide has a unique putative endoplasmic reticulum retention signal -RDEL. PMID:7632925

  13. Axotomy does not up-regulate expression of sodium channel Na(v)1.8 in Purkinje cells.

    PubMed

    Black, J A; Dusart, I; Sotelo, C; Waxman, S G

    2002-05-30

    Aberrant expression of the sensory neuron specific (SNS) sodium channel Na(v)1.8 has been demonstrated in cerebellar Purkinje cells in experimental models of multiple sclerosis (MS) and in human MS. The aberrant expression of Na(v)1.8, which is normally present in primary sensory neurons but not in the CNS, may perturb cerebellar function, but the mechanisms that trigger it are not understood. Because axotomy can provoke changes in Na(v)1.8 expression in dorsal root ganglion (DRG) neurons, we tested the hypothesis that axotomy can provoke an up-regulation of Na(v)1.8 expression in Purkinje cells, using a surgical model that transects axons of Purkinje cells in lobules IIIb-VII in the rat. In situ hybridization and immunocytochemistry did not reveal an up-regulation of Na(v)1.8 mRNA or protein in axotomized Purkinje cells. Hybridization and immunostaining signals for the sodium channel Na(v)1.6 were clearly present, demonstrating that sodium channel transcripts and protein were present in experimental cerebella. These results demonstrate that axotomy does not trigger the expression of Na(v)1.8 in Purkinje cells. PMID:12007840

  14. Nitrogen mustard up-regulates Bcl-2 and GSH and increases NTP and PCr in HT-29 colon cancer cells.

    PubMed Central

    Boddie, A. W.; Constantinou, A.; Williams, C.; Reed, A.

    1998-01-01

    We hypothesized that unexplained increases in nucleoside triphosphates (NTP) observed by 31P magnetic resonance spectroscopy (MRS) after treatment of tumours by DNA-damaging agents were related to chemotherapy-induced up-regulation of the bcl-2 gene and DNA damage prevention and repair processes. To test this hypothesis, we treated HT-29 cells with 10(-4) M nitrogen mustard (HN2) and performed sequential perchloric acid extractions in replicate over 0-18 h. By reference to an internal standard (methylene diphosphonic acid), absolute changes in 31P-detectable high-energy phosphates in these extracts were determined and correlated with changes in bcl-2 protein levels, cell viability, cell cycle, apoptosis and total cellular glutathione (GSH) (an important defence against DNA damage from alkylating agents). After HN2 administration, bcl-2 protein levels in the HT-29 cell line rose at 2 h. Cell viability declined to 25% within 18 h, but apoptosis measured using fluorescence techniques remained in the 1-4% range. Increased cell division was noted at 4 h. Two high-energy interconvertible phosphates, NTP (P < or = 0.006) and phosphocreatine (PCr) (P < or = 0.0002), increased at 2 h concurrently with increased levels of bcl-2 protein and glutathione. This study demonstrates that bcl-2 and glutathione are up-regulated by HN2 and links this to a previously unexplained 31P MRS phenomenon: increased NTP after chemotherapy. Images Figure 6 PMID:9652754

  15. Chronic up-regulation of the SHH pathway normalizes some developmental effects of trisomy in Ts65Dn mice.

    PubMed

    Dutka, Tara; Hallberg, Dorothy; Reeves, Roger H

    2015-02-01

    Down Syndrome (DS) is a highly complex developmental genetic disorder caused by trisomy for human chromosome 21 (Hsa21). All individuals with DS exhibit some degree of brain structural changes and cognitive impairment; mouse models such as Ts65Dn have been instrumental in understanding the underlying mechanisms. Several phenotypes of DS might arise from a reduced response of trisomic cells to the Sonic Hedgehog (SHH) growth factor. If all trisomic cells show a similar reduced response to SHH, then up-regulation of the pathway in trisomic cells might ameliorate multiple DS phenotypes. We crossed Ptch1tm1Mps/+ mice, in which the canonical SHH pathway is expected to be up-regulated in every SHH-responsive cell due to the loss of function of one allele of the pathway suppressor, Ptch1, to the Ts65Dn DS model and assessed the progeny for possible rescue of multiple DS-related phenotypes. Down-regulation of Ptch produced several previously unreported effects on development by itself, complicating interpretation of some phenotypes, and a number of structural or behavioral effects of trisomy were not compensated by SHH signaling. However, a deficit in a nest-building task was partially restored in Ts;Ptch+/- mice, as were the structural anomalies of the cerebellum seen in Ts65Dn mice. These results extend the body of evidence indicating that reduced response to SHH in trisomic cells and tissues contributes to various aspects of the trisomic phenotype. PMID:25511459

  16. A specialist herbivore pest adaptation to xenobiotics through up-regulation of multiple Cytochrome P450s.

    PubMed

    Zhu, Fang; Moural, Timothy W; Nelson, David R; Palli, Subba R

    2016-01-01

    The adaptation of herbivorous insects to their host plants is hypothesized to be intimately associated with their ubiquitous development of resistance to synthetic pesticides. However, not much is known about the mechanisms underlying the relationship between detoxification of plant toxins and synthetic pesticides. To address this knowledge gap, we used specialist pest Colorado potato beetle (CPB) and its host plant, potato, as a model system. Next-generation sequencing (454 pyrosequencing) was performed to reveal the CPB transcriptome. Differential expression patterns of cytochrome P450 complement (CYPome) were analyzed between the susceptible (S) and imidacloprid resistant (R) beetles. We also evaluated the global transcriptome repertoire of CPB CYPome in response to the challenge by potato leaf allelochemicals and imidacloprid. The results showed that more than half (51.2%) of the CBP cytochrome P450 monooxygenases (P450s) that are up-regulated in the R strain are also induced by both host plant toxins and pesticide in a tissue-specific manner. These data suggest that xenobiotic adaptation in this specialist herbivore is through up-regulation of multiple P450s that are potentially involved in detoxifying both pesticide and plant allelochemicals. PMID:26861263

  17. Fibrinogen up-regulates the expression of monocyte chemoattractant protein 1 in human saphenous vein endothelial cells.

    PubMed Central

    Harley, S L; Powell, J T

    1999-01-01

    High concentrations of fibrinogen in plasma have been associated with an increased risk of saphenous vein graft pathology. We have investigated the ability of fibrinogen to up-regulate the expression of monocyte chemoattractant protein 1 (MCP-1) in cultured human saphenous vein endothelial cells (HSVEC) isolated from saphenous vein. Increasing concentrations of fibrinogen (0-4 microM) stimulated a 20-fold increase in MCP-1 secretion within 4 h. Incubation of HSVEC with 2 microM fibrinogen for 4 h also caused a 2-fold increase in the MCP-1-to-glyceraldehyde-3-phosphate dehydrogenase mRNA ratio. The fibrinogen-mediated MCP-1 secretion fell to basal levels after preincubation of HSVEC with the complex of fibrinogen fragments D and E but remained unchanged after preincubation of HSVEC with either fibrinogen fragment E, s-ICAM-1 or the pentapeptide GRGDV. In contrast, fibrinogen fragment D acted as a potent inhibitor of fibrinogen-mediated MCP-1 secretion. Labelled fibrinogen fragment D bound to HSVEC with a K(d) of 6.5 microM. These findings indicate that fibrinogen, at physiological concentrations, uses an epitope on the fibrinogen D domain to bind to a receptor on HSVEC to up-regulate MCP-1 expression and secretion. This receptor seems to be distinct from intercellular adhesion molecule 1 and the integrins previously recognized as fibrinogen receptors. PMID:10417339

  18. Novel role of neuronal Ca2+ sensor-1 as a survival factor up-regulated in injured neurons.

    PubMed

    Nakamura, Tomoe Y; Jeromin, Andreas; Smith, George; Kurushima, Hideaki; Koga, Hitoshi; Nakabeppu, Yusaku; Wakabayashi, Shigeo; Nabekura, Junichi

    2006-03-27

    A molecular basis of survival from neuronal injury is essential for the development of therapeutic strategy to remedy neurodegenerative disorders. In this study, we demonstrate that an EF-hand Ca2+-binding protein neuronal Ca2+ sensor-1 (NCS-1), one of the key proteins for various neuronal functions, also acts as an important survival factor. Overexpression of NCS-1 rendered cultured neurons more tolerant to cell death caused by several kinds of stressors, whereas the dominant-negative mutant (E120Q) accelerated it. In addition, NCS-1 proteins increased upon treatment with glial cell line-derived neurotrophic factor (GDNF) and mediated GDNF survival signal in an Akt (but not MAPK)-dependent manner. Furthermore, NCS-1 is significantly up-regulated in response to axotomy-induced injury in the dorsal motor nucleus of the vagus neurons of adult rats in vivo, and adenoviral overexpression of E120Q resulted in a significant loss of surviving neurons, suggesting that NCS-1 is involved in an antiapoptotic mechanism in adult motor neurons. We propose that NCS-1 is a novel survival-promoting factor up-regulated in injured neurons that mediates the GDNF survival signal via the phosphatidylinositol 3-kinase-Akt pathway.

  19. An essential microRNA maturing microprocessor complex component DGCR8 is up-regulated in colorectal carcinomas.

    PubMed

    Kim, Bora; Lee, Jae-Ho; Park, Jong Wook; Kwon, Taeg Kyu; Baek, Seong Kyu; Hwang, Ilseon; Kim, Shin

    2014-08-01

    MicroRNAs (miRNAs) regulate gene expression through degradation and/or translational repression of target mRNAs. Dysregulations in the miRNA machinery may be involved in carcinogenesis of colorectal cancer (CRC). The purpose of the current study was to evaluate the DiGeorge syndrome critical region gene 8 (DGCR8) and argonaute 2 (AGO2) mRNA expression in CRC and to evaluate the value of clinical parameters on their expression. We investigated the mRNA expressions of DGCR8 and AGO2 in 60 CRC tissues and adjacent histologically non-neoplastic tissues by using quantitative real-time PCR. Our study revealed that the mRNA expression level of DGCR8 is up-regulated in CRC. However, AGO2 mRNA expression was not significantly altered in CRC tissues. Neither DGCR8 nor AGO2 mRNA expression level was not associated with any clinical parameters, including age, tumor stage, CEA titer, and BMI in CRC cases. However, the mRNA expression levels of DGCR8 and AGO2 were positively correlated to each other. This study demonstrated for the first time that the DGCR8 mRNA expression level was up-regulated in CRC, suggesting its important role in pathobiology of colorectal carcinogenesis.

  20. Up-regulation of alpha1-microglobulin by hemoglobin and reactive oxygen species in hepatoma and blood cell lines.

    PubMed

    Olsson, Magnus G; Allhorn, Maria; Olofsson, Tor; Akerström, Bo

    2007-03-15

    alpha(1)-Microglobulin is a 26-kDa glycoprotein synthesized in the liver, secreted to the blood, and rapidly distributed to the extravascular compartment of all tissues. Recent results show that alpha(1)-microglobulin has heme-binding and heme-degrading properties and it has been suggested that the protein is involved in the defense against oxidation by heme and reactive oxygen species. In the present study the influence of hemoglobin and reactive oxygen species (ROS) on the cellular expression of alpha(1)-microglobulin was investigated. Oxy- and methemoglobin, free heme, and Fenton reaction-induced hydroxyl radicals induced a dose-dependent up-regulation of alpha(1)-microglobulin on both mRNA and protein levels in hepatoma cells and an increased secretion of alpha(1)-microglobulin. The up-regulation was reversed by the addition of catalase and ascorbate, and by reacting hemoglobin with cyanide which prevents redox reactions. Furthermore, the blood cell lines U937 and K562 expressed alpha(1)-microglobulin at low levels, and this expression increased up to 11-fold by the addition of hemoglobin. These results suggest that alpha(1)-microglobulin expression is induced by ROS, arising from redox reactions of hemoglobin or from other sources and are consistent with the hypothesis that alpha(1)-microglobulin participates in the defense against oxidation by hemoglobin, heme, and reactive oxygen species.

  1. TGEV infection up-regulates FcRn expression via activation of NF-κB signaling

    PubMed Central

    Guo, Jinyue; Li, Fei; Qian, Shaoju; Bi, Dingren; He, Qigai; Jin, Hui; Luo, Rui; Li, Shaowen; Meng, Xianrong; Li, Zili

    2016-01-01

    It has been well characterized that the neonatal Fc receptor (FcRn) transports maternal IgG to a fetus or newborn and protects IgG from degradation. We previously reported that FcRn is expressed in a model of normal porcine intestinal epithelial cells (IPEC-J2). Transmissible gastroenteritis is an acute enteric disease of swine that is caused by transmissible gastroenteritis virus (TGEV). How porcine FcRn (pFcRn) expression is regulated by pathogenic infection remains unknown. Our research shows that IPEC-J2 cells infected with TGEV had up-regulated pFcRn expression. In addition, the NF-κB signaling pathway was activated in IPEC-J2 cells by TGEV infection. Furthermore, treatment of TGEV-infected IPEC-J2 cells with the NF-κB-specific inhibitor BAY 11-7082 resulted in down-regulation of pFcRn expression. Transient transfection of pFcRn promoter luciferase report plasmids with overexpression of NF-κB p65 transcription factor enhanced the activation of the luciferase report plasmids. We identified four NF-κB transcription factor binding sites in the promoter region of this gene using luciferase reporter system, chromatin immunoprecipitation, electromobility shift assay, and supershift analysis. Together, the data provide the first evidence that TGEV infection up-regulates pFcRn expression via activation of NF-κB signaling. PMID:27555521

  2. Transcutaneous electrical nerve stimulation (TENS) improves the diabetic cytopathy (DCP) via up-regulation of CGRP and cAMP.

    PubMed

    Ding, Liucheng; Song, Tao; Yi, Chaoran; Huang, Yi; Yu, Wen; Ling, Lin; Dai, Yutian; Wei, Zhongqing

    2013-01-01

    The objective of this study was to investigate the effects and mechanism of Transcutaneous Electrical Nerve Stimulation (TENS) on the diabetic cytopathy (DCP) in the diabetic bladder. A total of 45 rats were randomly divided into diabetes mellitus (DM)/TENS group (n=15), DM group (n=15) and control group (n=15). The rats in the DM/TENS and TENS groups were electronically stimulated (stimulating parameters: intensity-31 V, frequency-31 Hz, and duration of stimulation of 15 min) for three weeks. Bladder histology, urodynamics and contractile responses to field stimulation and carbachol were determined. The expression of calcitonin gene-related peptide (CGRP) was analyzed by RT-PCR and Western blotting. The results showed that contractile responses of the DM rats were ameliorated after 3 weeks of TENS. Furthermore, TENS significantly increased bladder wet weight, volume threshold for micturition and reduced PVR, V% and cAMP content of the bladder. The mRNA and protein levels of CGRP in dorsal root ganglion (DRG) in the DM/TENS group were higher than those in the DM group. TENS also significantly up-regulated the cAMP content in the bladder body and base compared with diabetic rats. We conclude that TENS can significantly improve the urine contractility and ameliorate the feeling of bladder fullness in DM rats possibly via up-regulation of cAMP and CGRP in DRG.

  3. Uncoupling Protein-2 is an Antioxidant that is Up-Regulated in the Enamel Organ of Fluoride-Treated Rats*

    PubMed Central

    Suzuki, Maiko; Sierant, Megan L.; Antone, Jerry V.; Everett, Eric T.; Whitford, Gary M.; Bartlett, John D.

    2014-01-01

    Dental fluorosis is characterized by subsurface hypomineralization and retention of enamel matrix proteins. Fluoride (F−) exposure generates reactive oxygen species (ROS) that can cause ER-stress. We therefore screened oxidative stress arrays to identify genes regulated by F− exposure. Vitamin E is an antioxidant so we asked if a diet high in vitamin E would attenuate dental fluorosis. Maturation stage incisor enamel organs (EO) were harvested from F− treated rats and mice were assessed to determine if vitamin E ameliorates dental fluorosis. Uncoupling protein-2 (Ucp2) was significantly up-regulated by F− (~1.5 & 2.0 fold for the 50 or 100 ppm F− treatment groups respectively). Immunohistochemical results on maturation stage rat incisors demonstrated that UCP2 protein levels increased with F− treatment. UCP2 down-regulates mitochondrial production of ROS, which decreases ATP production. Thus, in addition to reduced protein translation caused by ER-stress, a reduction in ATP production by UCP2 may contribute to the inability of ameloblasts to remove protein from the hardening enamel. Fluoride treated mouse enamel had significantly higher quantitative fluorescence (QF) than the untreated controls. No significant QF difference was observed between control and vitamin E enriched diets within a given F− treatment group. Therefore, a diet rich in vitamin E did not attenuate dental fluorosis. We have identified a novel oxidative stress response gene that is up-regulated in vivo by F− and activation of this gene may adversely affect ameloblast function. PMID:25158175

  4. TGF-β1 promotes scar fibroblasts proliferation and transdifferentiation via up-regulating MicroRNA-21.

    PubMed

    Liu, Ying; Li, Yue; Li, Ning; Teng, Wen; Wang, Min; Zhang, Yingbo; Xiao, Zhibo

    2016-01-01

    TGF-β1, upregulated in keloid tissue, promotes the proliferation, collagen formation and differentiation of dermal fibroblasts. miR-21 is one of microRNAs first found in human genome. The aim of our study is to explore the mechanisms of miR-21 in TGF-β1-induced scar fibroblasts proliferation and transdifferentiation. In the present study, first we found that TGF-β1 promoted scar fibroblasts proliferation and transdifferentiation via up-regulating miR-21 expression, which could be attenuated when miR-21 was inhibited. Overexpression of miR-21 had similar effect as TGF-β1 on proliferation and transdifferentiation. Additionally, TGF-β1 increased the expressions and activities of MMP2 and MMP9 in keloid fibroblasts, which was suppressed by miR-21 inhibition. Finally, the results demonstrated that PTEN/AKT signaling pathway played important role in TGF-β1-induced transdifferentiation. In conclusion, our study suggests that TGF-β1 promotes keloid fibroblasts proliferation and transdifferentiation via up-regulation of miR-21 and PTEN/AKT signalling pathway plays important role in this process, which provides a potential theoretical basis for clinical treatment of skin scars. PMID:27554193

  5. Vitamin D analogues up-regulate p21 and p27 during growth inhibition of pancreatic cancer cell lines.

    PubMed Central

    Kawa, S.; Nikaido, T.; Aoki, Y.; Zhai, Y.; Kumagai, T.; Furihata, K.; Fujii, S.; Kiyosawa, K.

    1997-01-01

    To obtain information regarding the growth-inhibitory effect of 1,25-dihydroxyvitamin D3 and its non-calcaemic analogue 22-oxa-1,25-dihydroxyvitamin D3 on pancreatic cancer cell lines, differences in the effects of G1-phase cell cycle-regulating factors were studied in vitamin D-responsive and non-responsive cell lines. Levels of expression of cyclins (D1, E and A), cyclin-dependent kinases (2 and 4) and cyclin-dependent kinase inhibitors (p21 and p27) were analysed by Western blotting after treatment with these compounds. In the responsive cells (BxPC-3, Hs 700T and SUP-1), our observations were: (1) marked up-regulation of p21 and p27 after 24 h treatment with 10(-7) mol l(-1) 1,25-dihydroxyvitamin D3 and 22-oxa-1,25-dihydroxyvitamin D3; and (2) marked down-regulation of cyclins, cyclin-dependent kinases and cyclin-dependent kinase inhibitors after 7 days' treatment. In non-responsive cells (Hs 766T and Capan-1), no such changes were observed. In conclusion, vitamin D analogues up-regulate p21 and p27 as an early event, which in turn could block the G1/S transition and induce growth inhibition in responsive cells. Images Figure 3 Figure 5 Figure 6 PMID:9328147

  6. A specialist herbivore pest adaptation to xenobiotics through up-regulation of multiple Cytochrome P450s

    PubMed Central

    Zhu, Fang; Moural, Timothy W.; Nelson, David R.; Palli, Subba R.

    2016-01-01

    The adaptation of herbivorous insects to their host plants is hypothesized to be intimately associated with their ubiquitous development of resistance to synthetic pesticides. However, not much is known about the mechanisms underlying the relationship between detoxification of plant toxins and synthetic pesticides. To address this knowledge gap, we used specialist pest Colorado potato beetle (CPB) and its host plant, potato, as a model system. Next-generation sequencing (454 pyrosequencing) was performed to reveal the CPB transcriptome. Differential expression patterns of cytochrome P450 complement (CYPome) were analyzed between the susceptible (S) and imidacloprid resistant (R) beetles. We also evaluated the global transcriptome repertoire of CPB CYPome in response to the challenge by potato leaf allelochemicals and imidacloprid. The results showed that more than half (51.2%) of the CBP cytochrome P450 monooxygenases (P450s) that are up-regulated in the R strain are also induced by both host plant toxins and pesticide in a tissue-specific manner. These data suggest that xenobiotic adaptation in this specialist herbivore is through up-regulation of multiple P450s that are potentially involved in detoxifying both pesticide and plant allelochemicals. PMID:26861263

  7. Carboxypeptidase E protects hippocampal neurons during stress in male mice by up-regulating prosurvival BCL2 protein expression.

    PubMed

    Murthy, S R K; Thouennon, E; Li, W-S; Cheng, Y; Bhupatkar, J; Cawley, N X; Lane, M; Merchenthaler, I; Loh, Y P

    2013-09-01

    Prolonged chronic stress causing elevated plasma glucocorticoids leads to neurodegeneration. Adaptation to stress (allostasis) through neuroprotective mechanisms can delay this process. Studies on hippocampal neurons have identified carboxypeptidase E (CPE) as a novel neuroprotective protein that acts extracellularly, independent of its enzymatic activity, although the mechanism of action is unclear. Here, we aim to determine if CPE plays a neuroprotective role in allostasis in mouse hippocampus during chronic restraint stress (CRS), and the molecular mechanisms involved. Quantitative RT-PCR/in situ hybridization and Western blots were used to assay for mRNA and protein. After mild CRS (1 h/d for 7 d), CPE protein and mRNA were significantly elevated in the hippocampal CA3 region, compared to naïve littermates. In addition, luciferase reporter assays identified a functional glucocorticoid regulatory element within the cpe promoter that mediated the up-regulation of CPE expression in primary hippocampal neurons following dexamethasone treatment, suggesting that circulating plasma glucocorticoids could evoke a similar effect on CPE in the hippocampus in vivo. Overexpression of CPE in hippocampal neurons, or CRS in mice, resulted in elevated prosurvival BCL2 protein/mRNA and p-AKT levels in the hippocampus; however, CPE(-/-) mice showed a decrease. Thus, during mild CRS, CPE expression is up-regulated, possibly contributed by glucocorticoids, to mediate neuroprotection of the hippocampus by enhancing BCL2 expression through AKT signaling, and thereby maintaining allostasis.

  8. Candida albicans up-regulates the Fas-L expression in liver Natural Killer and Natural Killer T cells.

    PubMed

    Renna, María Sol; Figueredo, Carlos Mauricio; Rodríguez-Galán, María Cecilia; Icely, Paula Alejandra; Cejas, Hugo; Cano, Roxana; Correa, Silvia Graciela; Sotomayor, Claudia Elena

    2015-11-01

    After Candida albicans arrival to the liver, the local production of proinflammatory cytokines and the expanded intrahepatic lymphocytes (IHL) can be either beneficial or detrimental to the host. Herein we explored the balance between protective inflammatory reaction and liver damage, focusing our study on the contribution of TNF-α and Fas-Fas-L pathways in the hepatocellular apoptosis associated to C. albicans infection. A robust tissue reaction and a progressive increase of IL-1β, IL-6 and TNF-α were observed in infected animals. Blocking the biological activity of TNF-α did not modify the number of apoptotic cells observed in C. albicans infected animals. Fas-L molecule was up regulated on purified hepatic mononuclear cells and its expression progressed with the infection. In the IHL compartment, the absolute number of Fas-L+ NK and NKT cells increased on days 1 and 3 of the infection. C. albicans was also able to up regulate Fas-L expression in normal liver NK and NKT cells after in vitro contact. The innate receptor TLR2 was involved in this phenomenon. In the interplay between host factors and evasion strategies exploited by pathogens, the mechanism supported here could represent an additional way that allows this fungus to circumvent protective immune responses in the liver.

  9. TGF-β1 promotes scar fibroblasts proliferation and transdifferentiation via up-regulating MicroRNA-21

    PubMed Central

    Liu, Ying; Li, Yue; Li, Ning; Teng, Wen; Wang, Min; Zhang, Yingbo; Xiao, Zhibo

    2016-01-01

    TGF-β1, upregulated in keloid tissue, promotes the proliferation, collagen formation and differentiation of dermal fibroblasts. miR-21 is one of microRNAs first found in human genome. The aim of our study is to explore the mechanisms of miR-21 in TGF-β1-induced scar fibroblasts proliferation and transdifferentiation. In the present study, first we found that TGF-β1 promoted scar fibroblasts proliferation and transdifferentiation via up-regulating miR-21 expression, which could be attenuated when miR-21 was inhibited. Overexpression of miR-21 had similar effect as TGF-β1 on proliferation and transdifferentiation. Additionally, TGF-β1 increased the expressions and activities of MMP2 and MMP9 in keloid fibroblasts, which was suppressed by miR-21 inhibition. Finally, the results demonstrated that PTEN/AKT signaling pathway played important role in TGF-β1-induced transdifferentiation. In conclusion, our study suggests that TGF-β1 promotes keloid fibroblasts proliferation and transdifferentiation via up-regulation of miR-21 and PTEN/AKT signalling pathway plays important role in this process, which provides a potential theoretical basis for clinical treatment of skin scars. PMID:27554193

  10. Chronic up-regulation of the SHH pathway normalizes some developmental effects of trisomy in Ts65Dn mice

    PubMed Central

    Dutka, Tara; Hallberg, Dorothy; Reeves, Roger H.

    2014-01-01

    Down Syndrome (DS) is a highly complex developmental genetic disorder caused by trisomy for human chromosome 21 (Hsa21). All individuals with DS exhibit some degree of brain structural changes and cognitive impairment; mouse models such as Ts65Dn have been instrumental in understanding the underlying mechanisms. Several phenotypes of DS might arise from a reduced response of trisomic cells to the Sonic Hedgehog (SHH) growth factor. If all trisomic cells show a similar reduced response to SHH, then up-regulation of the pathway in trisomic cells might ameliorate multiple DS phenotypes. We crossed Ptch1tm1Mps/+ mice, in which the canonical SHH pathway is expected to be up-regulated in every SHH-responsive cell due to the loss of function of one allele of the pathway suppressor, Ptch1, to the Ts65Dn DS model and assessed the progeny for possible rescue of multiple DS-related phenotypes. Down-regulation of Ptch produced several previously unreported effects on development by itself, complicating interpretation of some phenotypes, and a number structural or behavioral effects of trisomy were not compensated by SHH signaling. However, a deficit in a nest-building task was partially restored in Ts;Ptch+/− mice, as were structural anomalies of the cerebellum in Ts65Dn mice. These results extend the body of evidence indicating that reduced response to SHH in trisomic cells and tissues contributes to various aspects of the trisomic phenotype. PMID:25511459

  11. Convergence of multiple signaling pathways is required to coordinately up-regulate mtDNA and mitochondrial biogenesis during T cell activation.

    PubMed

    D'Souza, Anthony D; Parikh, Neal; Kaech, Susan M; Shadel, Gerald S

    2007-12-01

    The quantity and activity of mitochondria vary dramatically in tissues and are modulated in response to changing cellular energy demands and environmental factors. The amount of mitochondrial DNA (mtDNA), which encodes essential subunits of the oxidative phosphorylation complexes required for cellular ATP production, is also tightly regulated, but by largely unknown mechanisms. Using murine T cells as a model system, we have addressed how specific signaling pathways influence mitochondrial biogenesis and mtDNA copy number. T cell receptor (TCR) activation results in a large increase in mitochondrial mass and membrane potential and a corresponding amplification of mtDNA, consistent with a vital role for mitochondrial function for growth and proliferation of these cells. Independent activation of protein kinase C (via PMA) or calcium-related pathways (via ionomycin) had differential and sub-maximal effects on these mitochondrial parameters, as did activation of naïve T cells with proliferative cytokines. Thus, the robust mitochondrial biogenesis response observed upon TCR activation requires synergy of multiple downstream signaling pathways. One such pathway involves AMP-activated protein kinase (AMPK), which we show has an unprecedented role in negatively regulating mitochondrial biogenesis that is mammalian target of rapamycin (mTOR)-dependent. That is, inhibition of AMPK after TCR signaling commences results in excessive, but uncoordinated mitochondrial proliferation. Thus mitochondrial biogenesis is not under control of a single master regulatory circuit, but rather requires the convergence of multiple signaling pathways with distinct downstream consequences on the organelle's structure, composition, and function.

  12. Convergence of multiple signaling pathways is required to coordinately up-regulate mtDNA and mitochondrial biogenesis during T cell activation

    PubMed Central

    D’Souza, Anthony D.; Parikh, Neal; Kaech, Susan M.; Shadel, Gerald S.

    2009-01-01

    The quantity and activity of mitochondria vary dramatically in tissues and are modulated in response to changing cellular energy demands and environmental factors. The amount of mitochondrial DNA (mtDNA), which encodes essential subunits of the oxidative phosphorylation complexes required for cellular ATP production, is also tightly regulated, but by largely unknown mechanisms. Using murine T cells as a model system, we have addressed how specific signaling pathways influence mitochondrial biogenesis and mtDNA levels. T cell receptor (TCR) activation results in a large increase in mitochondrial mass and membrane potential and a corresponding increase of mtDNA copy number, indicating the vital role for mitochondrial function for the growth and proliferation of these cells. Independent activation of protein kinase C (via PMA) or calcium-related pathways (via ionomycin) had differential and sub-maximal effects on these mitochondrial parameters, as did activation of naïve T cells with proliferative cytokines. Thus, the robust mitochondrial biogenesis response observed upon TCR activation requires synergy of multiple downstream signaling pathways. One such pathway involves AMP-activated protein kinase (AMPK), which we show has an unprecedented role in negatively regulating mitochondrial biogenesis that is mammalian target of rapamycin (mTOR)-dependent. That is, inhibition of AMPK after TCR signaling commences results in excessive, but uncoordinated mitochondrial proliferation. We propose that mitochondrial biogenesis is not under control of a master regulatory circuit, but rather requires the convergence of multiple signaling pathways with distinct downstream consequences on the organelle’s structure, composition, and function. PMID:17890163

  13. Up-Regulation of miR-21 Is Associated with Cervicitis and Human Papillomavirus Infection in Cervical Tissues

    PubMed Central

    Bumrungthai, Sureewan; Ekalaksananan, Tipaya; Evans, Mark Francis; Chopjitt, Peechanika; Tangsiriwatthana, Thumwadee; Patarapadungkit, Natcha; Kleebkaow, Pilaiwan; Luanratanakorn, Sanguanchoke; Kongyingyoes, Bunkerd; Worawichawong, Suchin; Pientong, Chamsai

    2015-01-01

    MicroRNA-21 (miR-21) is recognized as an oncomir and shows up-regulation in many types of human malignancy. The aim of this study was to investigate the association of miR-21 expression associated with HPV infection in normal and abnormal cervical tissues. Cervical tissue samples with different cytological or histopathological grades were investigated for HPV by PCR and for miR-21 and programmed cell death, protein 4 (PDCD4) expression using quantitative real-time PCR (qRT-PCR). Laser capture microdissection (LCM) of stromal and epithelial tissues and in situ hybridization (ISH) using locked nucleic acid (LNA) probes were performed on a subset of fixed specimens. Cell line experiments were conducted on fibroblasts stimulated in culture media from HeLa cells, which were then assessed for miR-21, PDCD4, IL-6 and α-SMA expression by qRT-PCR. Twenty normal cervical cell, 12 cervicitis, 14 cervical intraepithelial neoplastic I (CIN I), 22 CIN II-III and 43 cervical squamous cell carcinoma (SCC) specimens were investigated. miR-21 levels were significantly lower in normal than in abnormal tissues. The expression of miR-21 in HPV negative normal cytology was significantly lower than in HPV positive samples in abnormal tissue and SCC. The miR-21 expression was significantly higher in HPV negative cervicitis than HPV negative normal cells. LCM and ISH data showed that miR-21 is primarily expressed in the tumor-associated stromal cell microenvironment. Fibroblasts treated with HeLa cell culture media showed up-regulated expression of miR-21, which correlated with increased expression of α-SMA and IL-6 and with down-regulation of PDCD4. These results demonstrate that miR-21 is associated with HPV infection and involved in cervical lesions as well as cervicitis and its up-regulation in tumor-stroma might be involved in the inflammation process and cervical cancer progression. PMID:26010154

  14. Caveolin-1 mediates tissue plasminogen activator-induced MMP-9 up-regulation in cultured brain microvascular endothelial cells.

    PubMed

    Jin, Xinchun; Sun, Yanyun; Xu, Ji; Liu, Wenlan

    2015-03-01

    Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates blood-brain barrier injury and increases the risk of symptomatic cerebral hemorrhage. The mechanism through which tPA enhances MMP-9 activity is not well understood. Here we report an important role of caveolin-1 in mediating tPA-induced MMP-9 synthesis. Brain microvascular endothelial cell line bEnd3 cells were incubated with 5 or 20 μg/ml tPA for 24 hrs before analyzing MMP-9 levels in the conditioned media and cellular extracts by gelatin zymography. tPA at a dose of 20 μg/mL tPA, but not 5 μg/mL, significantly increased MMP-9 level in cultured media while decreasing it in cellular extracts. Concurrently, tPA treatment induced a 2.3-fold increase of caveolin-1 protein levels in endothelial cells. Interestingly, knockdown of Cav-1 with siRNA inhibited tPA-induced MMP-9 mRNA up-regulation and MMP-9 increase in the conditioned media, but did not affect MMP-9 decrease in cellular extracts. These results suggest that caveolin-1 critically contributes to tPA-mediated MMP-9 up-regulation, but may not facilitate MMP-9 secretion in endothelial cells. Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates ischemic blood brain barrier (BBB) injury and increases the risk of symptomatic cerebral hemorrhage. Our results suggest a novel mechanism underlying this tPA-MMP 9 axis. In response to tPA treatment, caveolin-1 protein levels increased in endothelial cells, which mediate MMP-9 mRNA up-regulation and its secretion into extracellular space. Caveolin-1 may, however, not facilitate MMP-9 secretion in endothelial cells. Our data suggest caveolin-1 as a novel therapeutic target for protecting the BBB against ischemic damage. The schematic outlines tPA-induced MMP-9 upreguation.

  15. Exposure to diesel exhaust up-regulates iNOS expression in ApoE knockout mice

    SciTech Connect

    Bai Ni; Kido, Takashi; Kavanagh, Terrance J.; Kaufman, Joel D.; Rosenfeld, Michael E.; Breemen, Cornelis van; Eeden, Stephan F. van

    2011-09-01

    Traffic related particulate matter air pollution is a risk factor for cardiovascular events; however, the biological mechanisms are unclear. We hypothesize that diesel exhaust (DE) inhalation induces up-regulation of inducible nitric oxide synthase (iNOS), which is known to contribute to vascular dysfunction, progression of atherosclerosis and ultimately cardiovascular morbidity and mortality. Methods: ApoE knockout mice (30-week) were exposed to DE (at 200 {mu}g/m{sup 3} of particulate matter) or filtered-air (control) for 7 weeks (6 h/day, 5 days/week). iNOS expression in the blood vessels and heart was evaluated by immunohistochemistry and western blotting analysis. To examine iNOS activity, thoracic aortae were mounted in a wire myograph, and vasoconstriction stimulated by phenylephrine (PE) was measured with and without the presence of the specific inhibitor for iNOS (1400 W). NF-{kappa}B (p65) activity was examined by ELISA. The mRNA expression of iNOS and NF-{kappa}B (p65) was determined by real-time PCR. Results: DE exposure significantly enhanced iNOS expression in the thoracic aorta (4-fold) and heart (1.5 fold). DE exposure significantly attenuated PE-stimulated vasoconstriction by {approx} 20%, which was partly reversed by 1400 W. The mRNA expression of iNOS and NF-{kappa}B was significantly augmented after DE exposure. NF-{kappa}B activity was enhanced 2-fold after DE inhalation, and the augmented NF-{kappa}B activity was positively correlated with iNOS expression (R{sup 2} = 0.5998). Conclusions: We show that exposure to DE increases iNOS expression and activity possibly via NF-{kappa}B-mediated pathway. We suspect that DE exposure-caused up-regulation of iNOS contributes to vascular dysfunction and atherogenesis, which could ultimately lead to urban air pollution-associated cardiovascular morbidity and mortality. - Highlights: > Exposed ApoE knockout mice (30-week) to diesel exhaust (DE) for 7 weeks. > Examine iNOS expression and activity in the

  16. Caveolin-1 mediates tissue plasminogen activator-induced MMP-9 up-regulation in cultured brain microvascular endothelial cells.

    PubMed

    Jin, Xinchun; Sun, Yanyun; Xu, Ji; Liu, Wenlan

    2015-03-01

    Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates blood-brain barrier injury and increases the risk of symptomatic cerebral hemorrhage. The mechanism through which tPA enhances MMP-9 activity is not well understood. Here we report an important role of caveolin-1 in mediating tPA-induced MMP-9 synthesis. Brain microvascular endothelial cell line bEnd3 cells were incubated with 5 or 20 μg/ml tPA for 24 hrs before analyzing MMP-9 levels in the conditioned media and cellular extracts by gelatin zymography. tPA at a dose of 20 μg/mL tPA, but not 5 μg/mL, significantly increased MMP-9 level in cultured media while decreasing it in cellular extracts. Concurrently, tPA treatment induced a 2.3-fold increase of caveolin-1 protein levels in endothelial cells. Interestingly, knockdown of Cav-1 with siRNA inhibited tPA-induced MMP-9 mRNA up-regulation and MMP-9 increase in the conditioned media, but did not affect MMP-9 decrease in cellular extracts. These results suggest that caveolin-1 critically contributes to tPA-mediated MMP-9 up-regulation, but may not facilitate MMP-9 secretion in endothelial cells. Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates ischemic blood brain barrier (BBB) injury and increases the risk of symptomatic cerebral hemorrhage. Our results suggest a novel mechanism underlying this tPA-MMP 9 axis. In response to tPA treatment, caveolin-1 protein levels increased in endothelial cells, which mediate MMP-9 mRNA up-regulation and its secretion into extracellular space. Caveolin-1 may, however, not facilitate MMP-9 secretion in endothelial cells. Our data suggest caveolin-1 as a novel therapeutic target for protecting the BBB against ischemic damage. The schematic outlines tPA-induced MMP-9 upreguation. PMID:25683686

  17. Up-Regulated Expression of LAMP2 and Autophagy Activity during Neuroendocrine Differentiation of Prostate Cancer LNCaP Cells

    PubMed Central

    Vara-Ciruelos, Diana; Ramos-Torres, Ágata; Altamirano-Dimas, Manuel; Díaz-Laviada, Inés; Rodríguez-Henche, Nieves

    2016-01-01

    Neuroendocrine (NE) prostate cancer (PCa) is a highly aggressive subtype of prostate cancer associated with resistance to androgen ablation therapy. In this study, we used LNCaP prostate cancer cells cultured in a serum-free medium for 6 days as a NE model of prostate cancer. Serum deprivation increased the expression of NE markers such as neuron-specific enolase (NSE) and βIII tubulin (βIII tub) and decreased the expression of the androgen receptor protein in LNCaP cells. Using cDNA microarrays, we compared gene expression profiles of NE cells and non-differentiated LNCaP cells. We identified up-regulation of 155 genes, among them LAMP2, a lysosomal membrane protein involved in lysosomal stability and autophagy. We then confirmed up-regulation of LAMP2 in NE cells by qRT-PCR, Western blot and confocal microscopy assays, showing that mRNA up-regulation correlated with increased levels of LAMP2 protein. Subsequently, we determined autophagy activity in NE cells by assessing the protein levels of SQSTM/p62 and LC3 by Western blot and LC3 and Atg5 mRNAs content by qRT-PCR. The decreased levels of SQSTM/p62 was accompanied by an enhanced expression of LC3 and ATG5, suggesting activation of autophagy in NE cells. Blockage of autophagy with 1μM AKT inhibitor IV, or by silencing Beclin 1 and Atg5, prevented NE cell differentiation, as revealed by decreased levels of the NE markers. In addition, AKT inhibitor IV as well as Beclin1 and Atg5 kwockdown attenuated LAMP2 expression in NE cells. On the other hand, LAMP2 knockdown by siRNA led to a marked blockage of autophagy, prevention of NE differentiation and decrease of cell survival. Taken together, these results suggest that LAMP2 overexpression assists NE differentiation of LNCaP cells induced by serum deprivation and facilitates autophagy activity in order to attain the NE phenotype and cell survival. LAMP2 could thus be a potential biomarker and potential target for NE prostate cancer. PMID:27627761

  18. Rapamycin regulates the proliferation of Huh7, a hepatocellular carcinoma cell line, by up-regulating p53 expression.

    PubMed

    Kwon, Sora; Jeon, Ji-Sook; Ahn, Curie; Sung, Jung-Suk; Choi, Inho

    2016-10-01

    Rapamycin, a specific inhibitor of mTOR used extensively as an immunosuppressant, has been expanded recently to cancer therapy, because the mTOR signal is known to be up-regulated in various cancer cells including hepatocellular carcinoma (HCC) cells. In spite of extensive efforts to employ mTOR inhibitors as anti-HCC therapy, they have not yet been approved by the FDA. Because of the heterogeneity and complexity of molecular signaling in HCC, suitable biomarkers should be identified or discovered to improve clinical efficacy of mTOR-specific inhibitors to HCC cells. In this study, the effect of rapamycin was investigated on two different HCC cell lines, Huh7 cells and HepG2 cells. Rapamycin was found to inhibit the proliferation of Huh7 cells but not of HepG2 cells. Moreover, it was found that rapamycin can up-regulate p53 at the protein level, but not affect its transcript. To understand the critical role of p53 in the rapamycin effect, knock-down experiments were performed using small-interfering RNAs (siRNAs). The anti-proliferative effect of rapamycin on Huh7 cells clearly disappeared after blocking p53 production with siRNA, which indicates that p53 is a critical factor in the anti-proliferative effect of rapamycin in HCC cells. The over-expression system of p53 was also employed to mimic the effect of rapamycin and found that cell proliferation was clearly down-regulated by p53 over-expression. Finally, we found that the extracellular signal-regulated kinase 1/2 (ERK1/2) signal was regulated by p53 whose expression was induced by rapamycin. Overall, this study demonstrates that rapamycin inhibited the proliferation of Huh7 cells by up-regulating the expression of p53 and down-regulating the ERK1/2 signal, indicating that p53 is a useful biomarker for anti-cancer therapy using the specific inhibitor of mTOR signal, rapamycin, against hepatocellular carcinoma cells.

  19. Up-Regulated Expression of LAMP2 and Autophagy Activity during Neuroendocrine Differentiation of Prostate Cancer LNCaP Cells.

    PubMed

    Morell, Cecilia; Bort, Alicia; Vara-Ciruelos, Diana; Ramos-Torres, Ágata; Altamirano-Dimas, Manuel; Díaz-Laviada, Inés; Rodríguez-Henche, Nieves

    2016-01-01

    Neuroendocrine (NE) prostate cancer (PCa) is a highly aggressive subtype of prostate cancer associated with resistance to androgen ablation therapy. In this study, we used LNCaP prostate cancer cells cultured in a serum-free medium for 6 days as a NE model of prostate cancer. Serum deprivation increased the expression of NE markers such as neuron-specific enolase (NSE) and βIII tubulin (βIII tub) and decreased the expression of the androgen receptor protein in LNCaP cells. Using cDNA microarrays, we compared gene expression profiles of NE cells and non-differentiated LNCaP cells. We identified up-regulation of 155 genes, among them LAMP2, a lysosomal membrane protein involved in lysosomal stability and autophagy. We then confirmed up-regulation of LAMP2 in NE cells by qRT-PCR, Western blot and confocal microscopy assays, showing that mRNA up-regulation correlated with increased levels of LAMP2 protein. Subsequently, we determined autophagy activity in NE cells by assessing the protein levels of SQSTM/p62 and LC3 by Western blot and LC3 and Atg5 mRNAs content by qRT-PCR. The decreased levels of SQSTM/p62 was accompanied by an enhanced expression of LC3 and ATG5, suggesting activation of autophagy in NE cells. Blockage of autophagy with 1μM AKT inhibitor IV, or by silencing Beclin 1 and Atg5, prevented NE cell differentiation, as revealed by decreased levels of the NE markers. In addition, AKT inhibitor IV as well as Beclin1 and Atg5 kwockdown attenuated LAMP2 expression in NE cells. On the other hand, LAMP2 knockdown by siRNA led to a marked blockage of autophagy, prevention of NE differentiation and decrease of cell survival. Taken together, these results suggest that LAMP2 overexpression assists NE differentiation of LNCaP cells induced by serum deprivation and facilitates autophagy activity in order to attain the NE phenotype and cell survival. LAMP2 could thus be a potential biomarker and potential target for NE prostate cancer. PMID:27627761

  20. Interleukin (IL)-12 and IL-12 gene transfer up-regulate Fas expression in human osteosarcoma and breast cancer cells.

    PubMed

    Lafleur, E A; Jia, S F; Worth, L L; Zhou, Z; Owen-Schaub, L B; Kleinerman, E S

    2001-05-15

    Expression of Fas (CD95, APO-1), a cell surface receptor capable of inducing ligand-mediated apoptosis, is involved in tissue homeostasis and elimination of targeted cells by natural killer and T cells. Corruption of this pathway, such as reduced Fas expression, can allow tumor cells to escape elimination and promote metastatic potential. In this study, the status of Fas expression has been examined in the parental SAOS human osteosarcoma cells that do not metastasize and in selected variants that cause lung metastases in 16 weeks (LM2) or 8 weeks (LM6) after i.v. injection into nude mice. Fas expression correlated with the metastatic potentials of the three cell lines. Northern and fluorescence-activated cell-sorting analyses indicated that LM6 cells expressed Fas at a lower level than seen in the parental cells. Infection of the LM6 cells with an adenoviral vector containing the murine interleukin (IL)-12 gene (AD:mIL-12) or treatment with recombinant murine IL-12 resulted in a dose-dependent up-regulation of FAS: The up-regulation of Fas by IL-12 was also demonstrated in human etoposide-resistant MDA-MB-231 breast cancer cells. [(3)H]Thymidine growth inhibition studies indicated that the cell surface Fas induced after IL-12 exposure was functional and able to mediate cell death on cross-linking with anti-FAS: We also demonstrate that this effect is independent of IFN-gamma. Whereas these cell lines are sensitive to IFN-gamma, incubation with IFN-gamma does not increase susceptibility to Fas-mediated cell death, nor do these cells produce IFN-gamma with or without IL-12 treatment. We hypothesize that expression of Fas may play a role in the elimination of metastatic tumor cells in the lung, an organ in which Fas ligand is expressed. The antitumor activity of IL-12 may be secondary in part to its ability to up-regulate Fas expression on tumor cells, which subsequently increases immune-mediated destruction of osteosarcoma cells.

  1. L-Acetylcarnitine induces analgesia by selectively up-regulating mGlu2 metabotropic glutamate receptors.

    PubMed

    Chiechio, S; Caricasole, A; Barletta, E; Storto, M; Catania, M V; Copani, A; Vertechy, M; Nicolai, R; Calvani, M; Melchiorri, D; Nicoletti, F

    2002-05-01

    L-Acetylcarnitine (LAC, 100 mg/kg, s.c.), a drug commonly used for the treatment of painful neuropathies, substantially reduced mechanical allodynia in rats subjected to monolateral chronic constriction injury (CCI) of the sciatic nerve and also attenuated acute thermal pain in intact rats. In both cases, induction of analgesia required repeated injections of LAC, suggesting that the drug induces plastic changes within the nociceptive pathway. In both CCI- and sham-operated rats, a 24-day treatment with LAC increased the expression of metabotropic glutamate (mGlu) receptors 2 and 3 in the lumbar segment of the spinal cord, without changing the expression of mGlu1a or -5 receptors. A similar up-regulation of mGlu2/3 receptors was detected in the dorsal horns and dorsal root ganglia of intact rats treated with LAC for 5-7 days, a time sufficient for the induction of thermal analgesia. Immunohistochemical analysis showed that LAC treatment enhanced mGlu2/3 immunoreactivity in the inner part of lamina II and in laminae III and IV of the spinal cord. An increased mGlu2/3 receptor expression was also observed in the cerebral cortex but not in the hippocampus or cerebellum of LAC-treated animals. Reverse transcription-polymerase chain reaction combined with Northern blot analysis showed that repeated LAC injections selectively induced mGlu2 mRNA in the dorsal horns and cerebral cortex (but not in the hippocampus). mGlu3 mRNA levels did not change in any brain region of LAC-treated animals. To examine whether the selective up-regulation of mGlu2 receptors had any role in LAC-induced analgesia, we have used the novel compound LY 341495, which is a potent and systemically active mGlu2/3 receptor antagonist. LAC-induced analgesia was largely reduced 45 to 75 min after a single injection of LY 341495 (1 mg/kg, i.p.) in both CCI rats tested for mechanical allodynia and intact rats tested for thermal pain. We conclude that LAC produces analgesia against chronic pain produced not

  2. Methamphetamine and 3,4-methylenedioxymethamphetamine interact with central nicotinic receptors and induce their up-regulation

    SciTech Connect

    Garcia-Rates, Sara; Camarasa, Jordi; Escubedo, Elena; Pubill, David

    2007-09-15

    Previous work from our group indicated that {alpha}7 nicotinic acetylcholine receptors ({alpha}7 nAChR) potentially play a role in methamphetamine (METH) and 3,4-methylenedioxymethamphetamine (MDMA) neurotoxicity. The aims of the present study were two-fold: (1) to demonstrate the interaction of METH and MDMA with homomeric {alpha}7 nAChR ([{sup 3}H]methyllycaconitine binding) and other heteromeric subtypes ([{sup 3}H]epibatidine binding); and (2) to show the effects of amphetamine derivative pretreatment on the density of binding sites. METH and MDMA displaced [{sup 3}H]methyllycaconitine and [{sup 3}H]epibatidine binding in membranes from NGF-differentiated PC 12 cells and mouse brain, with K{sub i} values in the micromolar range, MDMA revealing a greater affinity than METH. In addition, METH and MDMA induced a time- and concentration-dependent increase in [{sup 3}H]methyllycaconitine and [{sup 3}H]epibatidine binding; which had already been apparent after 6 h of pretreatment, and which peaked in differentiated PC 12 cells after 48 h. The highest increases were found in [{sup 3}H]epibatidine binding, with MDMA inducing higher increases than METH. Treatment with METH and MDMA increased B{sub max} of high-affinity sites for both radioligands without affecting K{sub d}. The heightened binding was inhibited by pretreatment with cycloheximide, suggesting the participation of newly synthesised proteins while inhibition of protein trafficking to plasma membrane did not block up-regulation. The effects of protein kinase and cyclophilin inhibitors on such up-regulation were explored, revealing a rapid, differential and complex regulation, similar to that described for nicotinic ligands. All of these results demonstrate that METH and MDMA have affinity for, and can interact with, nAChR, inducing their up-regulation, specially when higher doses are used. Such effects may have a role in METH- and MDMA-induced neurotoxicity, cholinergic neurotransmission, and in processes

  3. Mannitol facilitates neurotrophic factor up-regulation and behavioural recovery in neonatal hypoxic-ischaemic rats with human umbilical cord blood grafts

    PubMed Central

    Yasuhara, T; Hara, K; Maki, M; Xu, L; Yu, G; Ali, M M; Masuda, T; Yu, S J; Bae, E K; Hayashi, T; Matsukawa, N; Kaneko, Y; Kuzmin-Nichols, N; Ellovitch, S; Cruz, E L; Klasko, S K; Sanberg, C D; Sanberg, P R; Borlongan, C V

    2010-01-01

    Abstract We recently demonstrated that blood–brain barrier permeabilization using mannitol enhances the therapeutic efficacy of systemically administered human umbilical cord blood (HUCB) by facilitating the entry of neurotrophic factors from the periphery into the adult stroke brain. Here, we examined whether the same blood–brain barrier manipulation approach increases the therapeutic effects of intravenously delivered HUCB in a neonatal hypoxic-ischaemic (HI) injury model. Seven-day-old Sprague–Dawley rats were subjected to unilateral HI injury and then at day 7 after the insult, animals intravenously received vehicle alone, mannitol alone, HUCB cells (15k mononuclear fraction) alone or a combination of mannitol and HUCB cells. Behavioural tests at post-transplantation days 7 and 14 showed that HI animals that received HUCB cells alone or when combined with mannitol were significantly less impaired in motor asymmetry and motor coordination compared with those that received vehicle alone or mannitol alone. Brain tissues from a separate animal cohort from the four treatment conditions were processed for enzyme-linked immunosorbent assay at day 3 post-transplantation, and revealed elevated levels of GDNF, NGF and BDNF in those that received HUCB cells alone or when combined with mannitol compared with those that received vehicle or mannitol alone, with the combined HUCB cells and mannitol exhibiting the most robust neurotropic factor up-regulation. Histological assays revealed only sporadic detection of HUCB cells, suggesting that the trophic factor–mediated mechanism, rather than cell replacement per se, principally contributed to the behavioural improvement. These findings extend the utility of blood–brain barrier permeabilization in facilitating cell therapy for treating neonatal HI injury. PMID:20569276

  4. Epithelial cell proliferation in nasal polyps could be up-regulated by platelet-derived growth factor.

    PubMed

    Coste, A; Wang, Q P; Roudot-Thoraval, F; Chapelin, C; Bedbeder, P; Poron, F; Peynègre, R; Escudier, E

    1996-05-01

    The modifications of epithelial differentiation and proliferation observed in nasal polyps (NP) could be related to local secretion of growth factors, among which platelet-derived growth factor (PDGF) could play a key role. We therefore prospectively studied, by immunohistochemistry, proliferating cell nuclear antigen (PCNA, an S-phase cell marker), PDGF, and CD-68 (activated macrophages marker) expression in NP and inferior turbinate mucosa (NM) in 11 patients. Our data show that PCNA and PDGF expression are increased in NP epithelium, while CD-68 expression is increased in NP epithelium and lamina propria when compared to NM. Increased local PDGF secretion by numerous activated macrophages could therefore be involved in epithelial cell proliferation up-regulation in NP. PDGF could also be involved in the pathogenesis of NP via its connective tissue remodeling actions.

  5. Glucose metabolism activation by SHIP2 inhibitors via up-regulation of GLUT1 gene in L6 myotubes.

    PubMed

    Suwa, Akira; Kurama, Takeshi; Yamamoto, Tadashi; Sawada, Akihiko; Shimokawa, Teruhiko; Aramori, Ichiro

    2010-09-10

    Lipid phosphatase SH2 domain-containing inositol 5'-phosphatase 2 (SHIP2) plays an important role in the regulation of insulin signaling. In this report, we identified AS1938909, a novel small-molecule SHIP2 inhibitor. AS1938909 showed potent inhibition of SHIP2 (Ki=0.44 microuM) and significant selectivity over other related phosphatases. Further, AS1938909 increased Akt phosphorylation, glucose consumption, and glucose uptake in L6 myotubes. Treatment of L6 myotubes with SHIP2 inhibitors for 48 h significantly induced expression of GLUT1 mRNA, but not that of GLUT4. These results suggest that pharmacological inhibition of SHIP2 activates glucose metabolism due, at least in part, to up-regulation of GLUT1 gene expression.

  6. Differences in the time course of haloperidol-induced up-regulation of rat striatal and mesolimbic dopamine receptors

    SciTech Connect

    Prosser, E.S.; Csernansky, J.G.; Hollister, L.E.

    1988-01-01

    Regional differences in the onset and persistence of increased dopamine D2 receptor density in rat brain were studied following daily injections of haloperidol for 3, 7, 14, or 28 days. Striatal (/sup 3/H)-spiroperidol Bmax values were significantly increased following 3 - 28 days of haloperidol treatment, as compared to saline controls. Olfactory tubercle Bmax values were significantly increased only after 14 or 28 days of haloperidol treatment. Nucleus accumbens Bmax values were significantly increased only in the 14-day drug treatment group, suggesting that dopamine D2 receptor up-regulation in nucleus accumbens may reverse during ongoing neuroleptic treatment. These findings suggest that important differences in adaptive responses to chronic dopamine blockade may exist between dopaminergic synapses located in various rat brain regions.

  7. Berberine up-regulates the BDNF expression in hippocampus and attenuates corticosterone-induced depressive-like behavior in mice.

    PubMed

    Shen, Ji-Duo; Ma, Li-Gang; Hu, Chun-Yue; Pei, Yang-Yi; Jin, Shuang-Li; Fang, Xiao-Yan; Li, Yu-Cheng

    2016-02-12

    Depression is increasingly become a global public healthy problem. This study was to investigate whether berberine could attenuate the depressive-like behavior induced by repeated corticosterone injection and explore the possible mechanisms. The present results showed that exogenous corticosterone injection caused depressive-like behaviors in mice, such as decreased sucrose intake in sucrose preference test (SPT) and increased immobility time in forced swimming test (FST). These behavioral alterations were accompanying with the decreased BDNF mRNA and protein levels in hippocampus and the elevated serum corticosterone levels. Treatment with berberine prevented these changes above. Our findings confirmed the antidepressant-like effect of berberine and suggested its mechanisms might be partially mediated by up-regulation of BDNF in hippocampus.

  8. Isoreserpine promotes {beta}-catenin degradation via Siah-1 up-regulation in HCT116 colon cancer cells

    SciTech Connect

    Gwak, Jungsug; Song, Taeyun; Song, Jie-Young; Yun, Yeon-Sook; Choi, Il-Whan; Jeong, Yongsu; Shin, Jae-Gook; Oh, Sangtaek

    2009-09-25

    Aberrant accumulation of intracellular {beta}-catenin in intestinal epithelial cells is a frequent early event during the development of colon cancer. To identify small molecules that decrease the level of intracellular {beta}-catenin, we performed cell-based chemical screening using genetically engineered HEK293 reporter cells to detect compounds that inhibit TOPFlash reporter activity, which was stimulated by Wnt3a-conditioned medium. We found that isoreserpine promoted the degradation of intracellular {beta}-catenin by up-regulation of Siah-1 in HEK293 and HCT116 colon cancer cells. Moreover, isoreserpine repressed the expression of {beta}-catenin/T-cell factor (TCF)-dependent genes, such as cyclin D1 and c-myc, resulting in the suppression of HCT116 cell proliferation. Our findings suggest that isoreserpine can potentially be used as a chemotherapeutic agent against colon cancer.

  9. Mannose receptor induces T-cell tolerance via inhibition of CD45 and up-regulation of CTLA-4.

    PubMed

    Schuette, Verena; Embgenbroich, Maria; Ulas, Thomas; Welz, Meike; Schulte-Schrepping, Jonas; Draffehn, Astrid M; Quast, Thomas; Koch, Katharina; Nehring, Melanie; König, Jessica; Zweynert, Annegret; Harms, Frederike L; Steiner, Nancy; Limmer, Andreas; Förster, Irmgard; Berberich-Siebelt, Friederike; Knolle, Percy A; Wohlleber, Dirk; Kolanus, Waldemar; Beyer, Marc; Schultze, Joachim L; Burgdorf, Sven

    2016-09-20

    The mannose receptor (MR) is an endocytic receptor involved in serum homeostasis and antigen presentation. Here, we identify the MR as a direct regulator of CD8(+) T-cell activity. We demonstrate that MR expression on dendritic cells (DCs) impaired T-cell cytotoxicity in vitro and in vivo. This regulatory effect of the MR was mediated by a direct interaction with CD45 on the T cell, inhibiting its phosphatase activity, which resulted in up-regulation of cytotoxic T-lymphocyte-associated Protein 4 (CTLA-4) and the induction of T-cell tolerance. Inhibition of CD45 prevented expression of B-cell lymphoma 6 (Bcl-6), a transcriptional inhibitor that directly bound the CTLA-4 promoter and regulated its activity. These data demonstrate that endocytic receptors expressed on DCs contribute to the regulation of T-cell functionality. PMID:27601670

  10. Growth differentiation factor 8 suppresses cell proliferation by up-regulating CTGF expression in human granulosa cells.

    PubMed

    Chang, Hsun-Ming; Pan, Hui-Hui; Cheng, Jung-Chien; Zhu, Yi-Min; Leung, Peter C K

    2016-02-15

    Connective tissue growth factor (CTGF) is a matricellular protein that plays a critical role in the development of ovarian follicles. Growth differentiation factor 8 (GDF8) is mainly, but not exclusively, expressed in the mammalian musculoskeletal system and is a potent negative regulator of skeletal muscle growth. The aim of this study was to investigate the effects of GDF8 and CTGF on the regulation of cell proliferation in human granulosa cells and to examine its underlying molecular determinants. Using dual inhibition approaches (inhibitors and small interfering RNAs), we have demonstrated that GDF8 induces the up-regulation of CTGF expression through the activin receptor-like kinase (ALK)4/5-mediated SMAD2/3-dependent signaling pathways. In addition, the increase in CTGF expression contributes to the GDF8-induced suppressive effect on granulosa cell proliferation. Our findings suggest that GDF8 and CTGF may play critical roles in the regulation of proliferative events in human granulosa cells.

  11. Body fat mass reduction and up-regulation of uncoupling protein by novel lipolysis-promoting plant extract.

    PubMed

    Mori, Shinobu; Satou, Mayumi; Kanazawa, Satoshi; Yoshizuka, Naonobu; Hase, Tadashi; Tokimitsu, Ichiro; Takema, Yoshinori; Nishizawa, Yoshinori; Yada, Toshihiko

    2009-01-01

    We have found natural products exhibiting lipolysis-promoting activity in subcutaneous adipocytes, which are less sensitive to hormones than visceral adipocytes. The activities and a action mechanisms of a novel plant extract of Cirsium oligophyllum (CE) were investigated in isolated adipocytes from rat subcutaneous fat, and its fat-reducing effects by peroral administration and topical application were evaluated in vivo. CE-induced lipolysis was synergistically enhanced by caffeine, a phosphodiesterase inhibitor, and was reduced by propranolol, a beta adrenergic antagonist. The peroral administration of 10% CE solution to Wistar rats for 32 days reduced body weight gain, subcutaneous, and visceral fat weights by 6.6, 26.2, and 3.0%, respectively, as compared to the control group. By the topical application of 2% of this extract to rats for 7 days, weight of subcutaneous fat in the treated skin was reduced by 23.2%. This fat mass reduction was accompanied by the up-regulation of uncoupling protein 1 (UCP), a principal thermogenic mitochondrial molecule related to energy dissipating, in subcutaneous fat and UCP3 in skin except for the fat layer. These results indicate that CE promotes lipolysis via a mechanism involving the beta adrenergic receptor, and affects the body fat mass. This fat reduction may be partially due to UCP up-regulation in the skin including subcutaneous fat. This is the first report showing that repeated lipolysis promotion through CE administration may be beneficial for the systematic suppression of body fat accumulation or the control of fat distribution in obesity.

  12. Exposure to Diesel Exhaust Up-regulates iNOS Expression in ApoE Knockout Mice

    PubMed Central

    Bai, Ni; Kido, Takashi; Kavanagh, Terrance J.; Kaufman, Joel D.; Rosenfeld, Michael E.; van Breemen, Cornelis; van Eeden, Stephan F.

    2012-01-01

    Traffic related particulate matter air pollution is a risk factor for cardiovascular events; however, the biological mechanisms are unclear. We hypothesize that diesel exhaust (DE) inhalation induces up-regulation of inducible nitric oxide synthase (iNOS), which is known to contribute to vascular dysfunction, progression of atherosclerosis and ultimately cardiovascular morbidity and mortality. Methods ApoE knockout mice (30-week) were exposed to DE (at 200µg/m3 of particulate matter) or filtered-air (control) for 7 weeks (6h/day, 5days/week). iNOS expression in the blood vessels and heart was evaluated by immunohistochemistry and western blotting analysis. To examine iNOS activity, thoracic aortae were mounted in a wire myograph, and vasoconstriction stimulated by phenylephrine (PE) was measured with and without the presence of the specific inhibitor for iNOS (1400W). NF-κB (p65) activity was examined by ELISA. The mRNA expression of iNOS and NF-κB (p65) was determined by real-time PCR. Results DE exposure significantly enhanced iNOS expression in the thoracic aorta (4-fold) and heart (1.5 fold). DE exposure significantly attenuated PE-stimulated vasoconstriction by ~20%, which was partly reversed by 1400W. The mRNA expression of iNOS and NF-κB was significantly augmented after DE exposure. NF-κB activity was enhanced 2-fold after DE inhalation, and the augmented NF-κB activity was positively correlated with iNOS expression (R2= 0.5998). Conclusions We show that exposure to DE increases iNOS expression and activity possibly via NF-κB-mediated pathway. We suspect that DE exposure-caused up-regulation of iNOS contributes to vascular dysfunction and atherogenesis, which could ultimately lead to urban air pollution-associated cardiovascular morbidity and mortality. PMID:21722660

  13. Direct interaction of CaVβ with actin up-regulates L-type calcium currents in HL-1 cardiomyocytes.

    PubMed

    Stölting, Gabriel; de Oliveira, Regina Campos; Guzman, Raul E; Miranda-Laferte, Erick; Conrad, Rachel; Jordan, Nadine; Schmidt, Silke; Hendriks, Johnny; Gensch, Thomas; Hidalgo, Patricia

    2015-02-20

    Expression of the β-subunit (CaVβ) is required for normal function of cardiac L-type calcium channels, and its up-regulation is associated with heart failure. CaVβ binds to the α1 pore-forming subunit of L-type channels and augments calcium current density by facilitating channel opening and increasing the number of channels in the plasma membrane, by a poorly understood mechanism. Actin, a key component of the intracellular trafficking machinery, interacts with Src homology 3 domains in different proteins. Although CaVβ encompasses a highly conserved Src homology 3 domain, association with actin has not yet been explored. Here, using co-sedimentation assays and FRET experiments, we uncover a direct interaction between CaVβ and actin filaments. Consistently, single-molecule localization analysis reveals streaklike structures composed by CaVβ2 that distribute over several micrometers along actin filaments in HL-1 cardiomyocytes. Overexpression of CaVβ2-N3 in HL-1 cells induces an increase in L-type current without altering voltage-dependent activation, thus reflecting an increased number of channels in the plasma membrane. CaVβ mediated L-type up-regulation, and CaVβ-actin association is prevented by disruption of the actin cytoskeleton with cytochalasin D. Our study reveals for the first time an interacting partner of CaVβ that is directly involved in vesicular trafficking. We propose a model in which CaVβ promotes anterograde trafficking of the L-type channels by anchoring them to actin filaments in their itinerary to the plasma membrane.

  14. Direct Interaction of CaVβ with Actin Up-regulates L-type Calcium Currents in HL-1 Cardiomyocytes*

    PubMed Central

    Stölting, Gabriel; de Oliveira, Regina Campos; Guzman, Raul E.; Miranda-Laferte, Erick; Conrad, Rachel; Jordan, Nadine; Schmidt, Silke; Hendriks, Johnny; Gensch, Thomas; Hidalgo, Patricia

    2015-01-01

    Expression of the β-subunit (CaVβ) is required for normal function of cardiac L-type calcium channels, and its up-regulation is associated with heart failure. CaVβ binds to the α1 pore-forming subunit of L-type channels and augments calcium current density by facilitating channel opening and increasing the number of channels in the plasma membrane, by a poorly understood mechanism. Actin, a key component of the intracellular trafficking machinery, interacts with Src homology 3 domains in different proteins. Although CaVβ encompasses a highly conserved Src homology 3 domain, association with actin has not yet been explored. Here, using co-sedimentation assays and FRET experiments, we uncover a direct interaction between CaVβ and actin filaments. Consistently, single-molecule localization analysis reveals streaklike structures composed by CaVβ2 that distribute over several micrometers along actin filaments in HL-1 cardiomyocytes. Overexpression of CaVβ2-N3 in HL-1 cells induces an increase in L-type current without altering voltage-dependent activation, thus reflecting an increased number of channels in the plasma membrane. CaVβ mediated L-type up-regulation, and CaVβ-actin association is prevented by disruption of the actin cytoskeleton with cytochalasin D. Our study reveals for the first time an interacting partner of CaVβ that is directly involved in vesicular trafficking. We propose a model in which CaVβ promotes anterograde trafficking of the L-type channels by anchoring them to actin filaments in their itinerary to the plasma membrane. PMID:25533460

  15. Plant stanols induce intestinal tumor formation by up-regulating Wnt and EGFR signaling in Apc Min mice.

    PubMed

    Marttinen, Maija; Päivärinta, Essi; Storvik, Markus; Huikko, Laura; Luoma-Halkola, Heli; Piironen, Vieno; Pajari, Anne-Maria; Mutanen, Marja

    2013-01-01

    The rate of APC mutations in the intestine increases in middle-age. At the same period of life, plant sterol and stanol enriched functional foods are introduced to diet to lower blood cholesterol. This study examined the effect of plant stanol enriched diet on intestinal adenoma formation in the Apc(Min) mouse. Apc(Min) mice were fed 0.8% plant stanol diet or control diet for nine weeks. Cholesterol, plant sterols and plant stanols were analyzed from the caecum content and the intestinal mucosa. Levels of β-catenin, cyclin D1, epidermal growth factor receptor (EGFR) and extracellular signal-regulated kinase 1/2 (ERK1/2) were measured from the intestinal mucosa by Western blotting. Gene expression was determined from the intestinal mucosa using Affymetrix and the data were analyzed for enriched categories and pathways. Plant stanols induced adenoma formation in the small intestine, however, the adenoma size was not affected. We saw increased levels of nuclear β-catenin, phosphorylated β-catenin (Ser675 and Ser552), nuclear cyclin D1, total and phosphorylated EGFR and phosphorylated ERK1/2 in the intestinal mucosa after plant stanol feeding. The Affymetrix data demonstrate that several enzymes of cholesterol synthesis pathway were up-regulated, although the cholesterol level in the intestinal mucosa was not altered. We show that plant stanols induce adenoma formation by activating Wnt and EGFR signaling. EGFR signaling seems to have promoted β-catenin phosphorylation and its translocation into the nucleus, where the expression of cyclin D1 was increased. Up-regulated cholesterol synthesis may partly explain the increased EGFR signaling in the plant stanol-fed mice.

  16. Nitrosative stress leads to protein glutathiolation, increased s-nitrosation, and up-regulation of peroxiredoxins in the heart.

    PubMed

    Reinartz, Michael; Ding, Zhaoping; Flögel, Ulrich; Gödecke, Axel; Schrader, Jürgen

    2008-06-20

    Nitric oxide (NO) is produced by different isoforms of nitric oxide synthases (NOSs) and operates as a mediator of important cell signaling pathways, such as the cGMP signaling cascade. Another mechanism by which NO exerts biological effects is mediated through S-nitrosation of target proteins. To explore thiol-based protein modifications in a situation of defined nitrosative stress, we used a transgenic mouse model with cardiac specific overexpression of inducible nitric oxide synthase (iNOS) and concomitant myoglobin deficiency (iNOS(+)/myo(-/-)). In comparison with the wild type hearts, protein glutathiolation detected by immunoblotting was significantly enhanced in iNOS(+)/myo(-/-) hearts, whereas protein S-nitrosation as measured by the biotin switch assay and two-dimensional PAGE revealed that nearly all of the detected proteins ( approximately 60) remained unchanged with the exception of three proteins. Tandem mass spectrometry revealed these proteins to be peroxiredoxins (Prxs), which are known to possess peroxidase activity, whereby hydrogen peroxide, peroxynitrite, and a wide range of organic hydroperoxides are reduced and detoxified. Immunoblotting with specific antibodies revealed up-regulation of Prx VI in the iNOS(+)/myo(-/-) hearts, whereas expression of Prx II and Prx III remained unchanged. Furthermore, the analysis of the cardiac S-nitrososubproteome identified several new proteins possibly being involved in NO-signaling pathways. Our data indicate that S-nitrosation and glutathiolation of cardiac proteins may contribute to the phenotype of NO-induced heart failure. The up-regulation of antioxidant proteins like Prx VI appears to be an additional mechanism to antagonize an excess of reactive oxygen/nitrogen species. Furthermore, S-nitrosation of Prxs may serve a new function in the signaling cascade of nitrosative stress. PMID:18426799

  17. Improved myocardial perfusion in chronic diabetic mice by the up-regulation of pLKB1 and AMPK signaling.

    PubMed

    Kusmic, Claudia; L'abbate, Antonio; Sambuceti, Gianmario; Drummond, George; Barsanti, Cristina; Matteucci, Marco; Cao, Jian; Piccolomini, Francesco; Cheng, Jennifer; Abraham, Nader G

    2010-04-01

    Previous studies related impaired myocardial microcirculation in diabetes to oxidative stress and endothelial dysfunction. Thus, this study was aimed to determine the effect of up-regulating pAMPK-pAKT signaling on coronary microvascular reactivity in the isolated heart of diabetic mice. We measured coronary resistance in wild-type and streptozotocin (STZ)-treated mice, during perfusion pressure changes. Glucose, insulin, and adiponectin levels in plasma and superoxide formation, NOx levels and heme oxygenase (HO) activity in myocardial tissue were determined. In addition, the expression of HO-1, 3-nitrotyrosine, pLKB1, pAMPK, pAKT, and peNOS proteins in control and diabetic hearts were measured. Coronary response to changes in perfusion pressure diverged from control in a time-dependent manner following STZ administration. The responses observed at 28 weeks of diabetes (the maximum time examined) were mimicked by L-NAME administration to control animals and were associated with a decrease in serum adiponectin and myocardial pLKB1, pAMPK, pAKT, and pGSK-3 expression. Cobalt protoporphyrin treatment to induce HO-1 expression reversed the microvascular reactivity seen in diabetes towards that of controls. Up-regulation of HO-1 was associated with an increase in adiponectin, pLKB1, pAKT, pAMPK, pGSK-3, and peNOS levels and a decrease in myocardial superoxide and 3-nitrotyrosine levels. In the present study we describe the time course of microvascular functional changes during the development of diabetes and the existence of a unique relationship between the levels of serum adiponectin, pLKB1, pAKT, and pAMPK activation in diabetic hearts. The restoration of microvascular function suggests a new therapeutic approach to even advanced cardiac microvascular derangement in diabetes.

  18. Green tea diet decreases PCB 126-induced oxidative stress in mice by up-regulating antioxidant enzymes.

    PubMed

    Newsome, Bradley J; Petriello, Michael C; Han, Sung Gu; Murphy, Margaret O; Eske, Katryn E; Sunkara, Manjula; Morris, Andrew J; Hennig, Bernhard

    2014-02-01

    Superfund chemicals such as polychlorinated biphenyls pose a serious human health risk due to their environmental persistence and link to multiple diseases. Selective bioactive food components such as flavonoids have been shown to ameliorate PCB toxicity, but primarily in an in vitro setting. Here, we show that mice fed a green tea-enriched diet and subsequently exposed to environmentally relevant doses of coplanar PCB exhibit decreased overall oxidative stress primarily due to the up-regulation of a battery of antioxidant enzymes. C57BL/6 mice were fed a low-fat diet supplemented with green tea extract (GTE) for 12 weeks and exposed to 5 μmol PCB 126/kg mouse weight (1.63 mg/kg-day) on weeks 10, 11 and 12 (total body burden: 4.9 mg/kg). F2-isoprostane and its metabolites, established markers of in vivo oxidative stress, measured in plasma via HPLC-MS/MS exhibited fivefold decreased levels in mice supplemented with GTE and subsequently exposed to PCB compared to animals on a control diet exposed to PCB. Livers were collected and harvested for both messenger RNA and protein analyses, and it was determined that many genes transcriptionally controlled by aryl hydrocarbon receptor and nuclear factor (erythroid-derived 2)-like 2 proteins were up-regulated in PCB-exposed mice fed the green tea-supplemented diet. An increased induction of genes such as SOD1, GSR, NQO1 and GST, key antioxidant enzymes, in these mice (green tea plus PCB) may explain the observed decrease in overall oxidative stress. A diet supplemented with green tea allows for an efficient antioxidant response in the presence of PCB 126, which supports the emerging paradigm that healthful nutrition may be able to bolster and buffer a physiological system against the toxicities of environmental pollutants.

  19. Characteristics of genes up-regulated and down-regulated after 24 h starvation in the head of Drosophila.

    PubMed

    Fujikawa, Kazuyo; Takahashi, Aya; Nishimura, Azusa; Itoh, Masanobu; Takano-Shimizu, Toshiyuki; Ozaki, Mamiko

    2009-10-01

    Starvation is a common experience under fluctuating food conditions in nature, and response to it is vital for many organisms. Many studies have investigated the response at physiological and behavioral level, whereas the studies on starvation-induced transcriptional changes in the brain and the surrounding tissues are still limited. We here investigated global changes in transcript abundance in the head after 24 h starvation by microarray expression profiling of 2 wild-derived inbred strains of Drosophila melanogaster, and identified a core set of 65 up-regulated and 48 down-regulated genes upon starvation. Among these up-regulated genes, 22 genes were circadian oscillating genes previously identified in the head of Drosophila. Interestingly, most (86%) of these circadian genes show their expression peak in a narrow time range of ZT7.0-12.0, when flies are relatively restless and less feeding in the normal condition. Among the down-regulated genes, 2 genes with highest fold-differences, fit and CG8147, are known to have female-biased expression in the head, and 1 gene, Obp99b, is known to be male-biased. Together with the realtime qPCR experiments on female and male transcripts, our data suggest that these sex-specific genes are candidate genes mediating a possible trade-off between starvation resistance and reproduction. Eleven down-regulated genes are known to be involved in the immune response. These changes in head transcriptome upon starvation reflect modulation of expression in some normally oscillating rhythmic genes and reduction in the resource allocation toward sexual activity and immunity.

  20. Simvastatin alleviates cardiac fibrosis induced by infarction via up-regulation of TGF-β receptor III expression

    PubMed Central

    Sun, Fei; Duan, Wenqi; Zhang, Yu; Zhang, Lingling; Qile, Muge; Liu, Zengyan; Qiu, Fang; Zhao, Dan; Lu, Yanjie; Chu, Wenfeng

    2015-01-01

    Background and Purpose Statins decrease heart disease risk, but their mechanisms are not completely understood. We examined the role of the TGF-β receptor III (TGFBR3) in the inhibition of cardiac fibrosis by simvastatin. Experimental Approach Myocardial infarction (MI) was induced by ligation of the left anterior descending coronary artery in mice given simvastatin orally for 7 days. Cardiac fibrosis was measured by Masson staining and electron microscopy. Heart function was evaluated by echocardiography. Signalling through TGFBR3, ERK1/2, JNK and p38 pathways was measured using Western blotting. Collagen content and cell viability were measured in cultures of neonatal mouse cardiac fibroblasts (NMCFs). Interactions between TGFBR3 and the scaffolding protein, GAIP-interacting protein C-terminus (GIPC) were detected using co-immunoprecipitation (co-IP). In vivo, hearts were injected with lentivirus carrying shRNA for TGFBR3. Key Results Simvastatin prevented fibrosis following MI, improved heart ultrastructure and function, up-regulated TGFBR3 and decreased ERK1/2 and JNK phosphorylation. Simvastatin up-regulated TGFBR3 in NMCFs, whereas silencing TGFBR3 reversed inhibitory effects of simvastatin on cell proliferation and collagen production. Simvastatin inhibited ERK1/2 and JNK signalling while silencing TGFBR3 opposed this effect. Co-IP demonstrated TGFBR3 binding to GIPC. Overexpressing TGFBR3 inhibited ERK1/2 and JNK signalling which was abolished by knock-down of GIPC. In vivo, suppression of cardiac TGFBR3 abolished anti-fibrotic effects, improvement of cardiac function and changes in related proteins after simvastatin. Conclusions and Implications TGFBR3 mediated the decreased cardiac fibrosis, collagen deposition and fibroblast activity, induced by simvastatin, following MI. These effects involved GIPC inhibition of the ERK1/2/JNK pathway. PMID:25884615

  1. Ampelopsin Improves Insulin Resistance by Activating PPARγ and Subsequently Up-Regulating FGF21-AMPK Signaling Pathway

    PubMed Central

    Qin, Yu; Liu, Lei; Wan, Jing; Zou, Lingyun; Zhang, Qianyong; Zhu, Jundong; Mi, Mantian

    2016-01-01

    Ampelopsin (APL), a major bioactive constituent of Ampelopsis grossedentata, exerts a number of biological effects. Here, we explored the anti-diabetic activity of APL and elucidate the underlying mechanism of this action. In palmitate-induced insulin resistance of L6 myotubes, APL treatment markedly up- regulated phosphorylated insulin receptor substrate-1 and protein kinase B, along with a corresponding increase of glucose uptake capacity. APL treatment also increased expressions of fibroblast growth factor (FGF21) and phosphorylated adenosine 5’-monophosphate -activated protein kinase (p-AMPK), however inhibiting AMPK by Compound C or AMPK siRNA, or blockage of FGF21 by FGF21 siRNA, obviously weakened APL -induced increases of FGF21 and p-AMPK as well as glucose uptake capacity in palmitate -pretreated L6 myotubes. Furthermore, APL could activate PPAR γ resulting in increases of glucose uptake capacity and expressions of FGF21 and p-AMPK in palmitate -pretreated L6 myotubes, whereas all those effects were obviously abolished by addition of GW9662, a specific inhibitor of peroxisome proliferator- activated receptor –γ (PPARγ), and PPARγsiRNA. Using molecular modeling and the luciferase reporter assays, we observed that APL could dock with the catalytic domain of PPARγ and dose-dependently up-regulate PPARγ activity. In summary, APL maybe a potential agonist of PPARγ and promotes insulin sensitization by activating PPARγ and subsequently regulating FGF21- AMPK signaling pathway. These results provide new insights into the protective health effects of APL, especially for the treatment of Type 2 diabetes mellitus. PMID:27391974

  2. Uterine Expression of NDRG4 Is Induced by Estrogen and Up-Regulated during Embryo Implantation Process in Mice

    PubMed Central

    Zhang, Xuan; Wang, Jian-Mei; He, Ya-Ping; Shi, Yan; Sun, Zhao-Gui; Shi, Hui-Juan; Wang, Jian

    2016-01-01

    Embryo implantation is an essential step for the establishment of pregnancy and dynamically regulated by estrogen and progesterone. NDRG4 (N-myc down-regulated gene 4) is a tumor suppressor that participates in cell survival, tumor invasion and angiogenesis. The objective of this study was to preliminarily explore the role of NDRG4 in embryo implantation. By immunohistochemistry (IHC) and quantitive RT-PCR (qRT-PCR), we found that uterine expression of NDRG4 was increased along with puberal development, and its expression in adult females reached the peak at the estrus stage during the estrus cycle. Furthermore, uterine NDRG4 expression was significantly induced by the treatment of estradiol (E2) both in pre-puberty females and ovariectomized adult females. Uterine expression pattern of NDRG4 during the peri-implantation period in mice was determined by IHC, qRT-PCR and Western blot. It was observed that NDRG4 expression was up-regulated during the implantation process, and its expression level at the implantation sites was significantly higher than that at the inter-implantation sites. Meanwhile, an increased expression in NDRG4 was associated with artificial decidualization as well as the activation of delayed implantation. By qRT-PCR and Western blot, we found that the in vitro decidualization of endometrial stromal cells (ESCs) was accompanied by up-regulation of NDRG4 expression, whereas knockdown of its expression in these cells by siRNA inhibited the decidualization process. In addition, Western blot analysis showed that NDRG4 protein expression was decreased in human villus tissues of recurrent miscarriage (RM) patients compared to normal pregnant women. Collectively, these data suggested that uterine NDRG4 expression could be induced by estrogen, and NDRG4 might play an important role during early pregnancy. PMID:27175791

  3. Up-regulation of C1GALT1 promotes breast cancer cell growth through MUC1-C signaling pathway

    PubMed Central

    Chou, Chih-Hsing; Huang, Miao-Juei; Chen, Chi-Hau; Shyu, Ming-Kwang; Huang, John; Hung, Ji-Shiang; Huang, Chiun-Sheng; Huang, Min-Chuan

    2015-01-01

    Aberrant glycosylation is frequently observed in cancers. Core 1 β1,3-galactosyltransferase (C1GALT1) is an exclusive enzyme in humans that catalyzes the biosynthesis of core 1 O-glycan structure, Gal-GalNAc-O-Ser/Thr, whose expression is commonly up-regulated during tumorigenesis. Little is known about the function of C1GALT1 in breast cancer. This study aims to determine the correlation between C1GALT1 expression and breast cancer clinicopathological features and roles of C1GALT1 in breast cancer malignant phenotypes. Public databases and our data showed that C1GALT1 mRNA and C1GALT1 protein are frequently up-regulated in breast cancer; and increased C1GALT1 expression correlates with higher histological grade and advanced tumor stage. Overexpression of C1GALT1 enhanced breast cancer cell growth, migration, and invasion in vitro as well as tumor growth in vivo. Conversely, C1GALT1 knockdown suppressed these malignant phenotypes. Furthermore, C1GALT1 modulates O-glycan structures on Mucin (MUC) 1 and promotes MUC1-C/β-catenin signaling in breast cancer cells. These findings suggest that C1GALT1 enhances breast cancer malignant progression through promoting MUC1-C/β-catenin signaling pathway. Unveiling the function of C1GALT1 in breast cancer opens new insights to the roles of C1GALT1 and O-glycosylation in tumorigenesis and renders the potential of C1GALT1 as a target of novel therapeutic agent development. PMID:25762620

  4. Tissue specific up regulation of ACE2 in rabbit model of atherosclerosis by atorvastatin: role of epigenetic histone modifications.

    PubMed

    Tikoo, Kulbhushan; Patel, Gaurang; Kumar, Sandeep; Karpe, Pinakin Arun; Sanghavi, Maitri; Malek, Vajir; Srinivasan, K

    2015-02-01

    Growing body of evidence points out the crucial role of ACE2 in preventing atherosclerosis. However, data on how atherosclerosis affects ACE2 expression in heart and kidney remains unknown. Atherosclerosis was induced by feeding New Zealand White rabbits with high cholesterol diet (HCD - 2%) for 12 weeks and atorvastatin was administered (5mg/kg/day p.o) in last 3 weeks. ACE2 mRNA and protein expression was assessed by Western blotting and real time PCR. HCD fed rabbits developed atherosclerosis as confirmed by increase in plasma total cholesterol, LDL and triglycerides as well as formation atherosclerotic plaques in arch of aorta. The ACE2 protein but not mRNA expression was reduced in heart and kidney of HCD rabbits. Interestingly, atorvastatin increased the ACE2 protein expression in heart and kidney of HCD rabbits. However, atorvastatin increased ACE2 mRNA in heart but not in kidney of HCD rabbits. Atorvastatin increased the occupancy of histone H3 acetylation (H3-Ac) mark on ACE2 promoter region in heart of HCD rabbits indicating direct or indirect epigenetic up-regulation of ACE2 by atorvastatin. Further, atorvastatin suppressed Ang II-induced contractile responses and enhanced AT2 receptor mediated relaxant responses in atherosclerotic aorta. We propose that atherosclerosis is associated with reduced ACE2 expression in heart and kidney. We also show an unexplored potential of atorvastatin to up-regulate ACE2 via epigenetic histone modifications. Our data suggest a novel way of replenishing ACE2 expression for preventing not only atherosclerosis but also other cardiovascular disorders. PMID:25482567

  5. MicroRNA miR-1 is up-regulated in remote myocardium in patients with myocardial infarction.

    PubMed

    Bostjancic, E; Zidar, N; Stajner, D; Glavac, D

    2010-01-01

    MicroRNAs are small regulatory RNA molecules that mediate regulation of gene expression, thus affecting a variety of physiological, developmental and pathological conditions. They are believed to be new promising therapeutic targets. In recent studies two muscle-specific microRNAs were discovered to contribute to heart diseases and development: miR-1 and miR-133, but there is little data on their expression patterns in human myocardial infarction. We performed simultaneous expression analysis of miR-1, miR-133a, miR-133b in samples of infarcted tissue and remote myocardium from twenty- four patients with acute myocardial infarction. MicroRNA expression was analysed using quantitative real-time PCR and compared to the expression patterns in myocardium of eight healthy adults who died in accidents. We found ~3.8-fold miR-1 up-regulation in remote myocardium when compared to infarcted tissue or healthy adult hearts. As miR-1 has been shown in animal models and clinical studies to contribute to arrhythmogenesis by regulating pacemaker channel genes, our finding of miR-1 up-regulation in patients with myocardial infarction indicates that it might be responsible for the higher risk for arrhythmias in these patients. In addition, miR-133a/b down-regulation in infarcted tissue and remote myocardium was observed, indicating miR-133a/b involvement in the heart response to myocardial infarction. We conclude that miR-1 and miR-133 seem to be important regulators of heart adaptation after ischaemic stress.

  6. An early ethylene up-regulated gene encoding a calmodulin-binding protein involved in plant senescence and death

    NASA Technical Reports Server (NTRS)

    Yang, T.; Poovaiah, B. W.

    2000-01-01

    35S-Labeled calmodulin (CaM) was used to screen a tobacco anther cDNA library. A positive clone (NtER1) with high homology to an early ethylene-up-regulated gene (ER66) in tomato, and an Arabidopsis homolog was isolated and characterized. Based on the helical wheel projection, a 25-mer peptide corresponding to the predicted CaM-binding region of NtER1 (amino acids 796-820) was synthesized. The gel-mobility shift assay showed that the peptide formed a stable complex with CaM only in the presence of Ca(2+). CaM binds to NtER1 with high affinity (K(d) approximately 12 nm) in a calcium-dependent manner. Tobacco flowers at different stages of development were treated with ethylene or with 1-methylcyclopropene for 2 h before treating with ethylene. Northern analysis showed that the NtER1 was rapidly induced after 15 min of exposure to ethylene. However, the 2-h 1-methylcyclopropene treatment totally blocked NtER1 expression in flowers at all stages of development, suggesting that NtER1 is an early ethylene-up-regulated gene. The senescing leaves and petals had significantly increased NtER1 induction as compared with young leaves and petals, implying that NtER1 is developmentally regulated and acts as a trigger for senescence and death. This is the first documented evidence for the involvement of Ca(2+)/CaM-mediated signaling in ethylene action.

  7. Increased curvature of hollow fiber membranes could up-regulate differential functions of renal tubular cell layers.

    PubMed

    Shen, Chong; Meng, Qin; Zhang, Guoliang

    2013-08-01

    Tissue engineering devices as in vitro cell culture systems in scaffolds has encountered the bottleneck due to their much lower cell functions than real tissues/organs in vivo. Such situation has been improved in some extent by mimicking the cell microenvironments in vivo from either chemical or physical ways. However, microenvironmental curvature, commonly seen in real tissues/organs, has never been manipulated to regulate the cell performance in vitro. In this regard, this paper fabricated polysulfone membranes with or without polyethylene glycol modification to investigate the impact of curvature on two renal tubular cells. Regardless the varying membrane curvatures among hollow fiber membranes of different diameters and flat membrane of zero curvature, both renal cells could well attach at 4 h of seeding and form similar confluent layers at 6 days on each membrane. Nevertheless, the renal cells on hollow fibers, though showing confluent morphology as those on flat membranes, expressed higher renal functions and, moreover, the renal functions significantly increased with the membrane curvature among hollow fibers. Such upregulation on functions was unassociated with mass transport barrier of hollow fibers, because the cultures on lengthwise cut hollow fibers without mass transfer barrier showed same curvature effect on renal functions as whole hollow fibers. It could be proposed that the curvature of hollow fiber membrane approaching to the large curvature in kidney tubules increased the mechanical stress in the renal cells and thus might up-regulate the renal cell functions. In conclusion, the increase of substrate curvature could up-regulate the cell functions without altering the confluent cell morphology and this finding will facilitate the design of functional tissue engineering devices.

  8. Hypoxic stress up-regulates Kir2.1 expression and facilitates cell proliferation in brain capillary endothelial cells.

    PubMed

    Yamamura, Hideto; Suzuki, Yoshiaki; Yamamura, Hisao; Asai, Kiyofumi; Imaizumi, Yuji

    2016-08-01

    The blood-brain barrier (BBB) is mainly composed of brain capillary endothelial cells (BCECs), astrocytes and pericytes. Brain ischemia causes hypoxic encephalopathy and damages BBB. However, it remains still unclear how hypoxia affects BCECs. In the present study, t-BBEC117 cells, an immortalized bovine brain endothelial cell line, were cultured under hypoxic conditions at 4-5% oxygen for 72 h. This hypoxic stress caused hyperpolarization of resting membrane potential. Patch-clamp recordings revealed a marked increase in Ba(2+)-sensitive inward rectifier K(+) current in t-BBEC117 cells after hypoxic culture. Western blot and real-time PCR analyses showed that Kir2.1 expression was significantly up-regulated at protein level but not at mRNA level after the hypoxic culture. Ca(2+) imaging study revealed that the hypoxic stress enhanced store-operated Ca(2+) (SOC) entry, which was significantly reduced in the presence of 100 μM Ba(2+). On the other hand, the expression of SOC channels such as Orai1, Orai2, and transient receptor potential channels was not affected by hypoxic stress. MTT assay showed that the hypoxic stress significantly enhanced t-BBEC117 cell proliferation, which was inhibited by approximately 60% in the presence of 100 μM Ba(2+). We first show here that moderate cellular stress by cultivation under hypoxic conditions hyperpolarizes membrane potential via the up-regulation of functional Kir2.1 expression and presumably enhances Ca(2+) entry, resulting in the facilitation of BCEC proliferation. These findings suggest potential roles of Kir2.1 expression in functional changes of BCECs in BBB following ischemia. PMID:27235552

  9. Uterine Expression of NDRG4 Is Induced by Estrogen and Up-Regulated during Embryo Implantation Process in Mice.

    PubMed

    Yang, Qian; Gu, Yan; Zhang, Xuan; Wang, Jian-Mei; He, Ya-Ping; Shi, Yan; Sun, Zhao-Gui; Shi, Hui-Juan; Wang, Jian

    2016-01-01

    Embryo implantation is an essential step for the establishment of pregnancy and dynamically regulated by estrogen and progesterone. NDRG4 (N-myc down-regulated gene 4) is a tumor suppressor that participates in cell survival, tumor invasion and angiogenesis. The objective of this study was to preliminarily explore the role of NDRG4 in embryo implantation. By immunohistochemistry (IHC) and quantitive RT-PCR (qRT-PCR), we found that uterine expression of NDRG4 was increased along with puberal development, and its expression in adult females reached the peak at the estrus stage during the estrus cycle. Furthermore, uterine NDRG4 expression was significantly induced by the treatment of estradiol (E2) both in pre-puberty females and ovariectomized adult females. Uterine expression pattern of NDRG4 during the peri-implantation period in mice was determined by IHC, qRT-PCR and Western blot. It was observed that NDRG4 expression was up-regulated during the implantation process, and its expression level at the implantation sites was significantly higher than that at the inter-implantation sites. Meanwhile, an increased expression in NDRG4 was associated with artificial decidualization as well as the activation of delayed implantation. By qRT-PCR and Western blot, we found that the in vitro decidualization of endometrial stromal cells (ESCs) was accompanied by up-regulation of NDRG4 expression, whereas knockdown of its expression in these cells by siRNA inhibited the decidualization process. In addition, Western blot analysis showed that NDRG4 protein expression was decreased in human villus tissues of recurrent miscarriage (RM) patients compared to normal pregnant women. Collectively, these data suggested that uterine NDRG4 expression could be induced by estrogen, and NDRG4 might play an important role during early pregnancy. PMID:27175791

  10. CX3CL1 is up-regulated in the rat hippocampus during memory-associated synaptic plasticity.

    PubMed

    Sheridan, Graham K; Wdowicz, Anita; Pickering, Mark; Watters, Orla; Halley, Paul; O'Sullivan, Niamh C; Mooney, Claire; O'Connell, David J; O'Connor, John J; Murphy, Keith J

    2014-01-01

    Several cytokines and chemokines are now known to play normal physiological roles in the brain where they act as key regulators of communication between neurons, glia, and microglia. In particular, cytokines and chemokines can affect cardinal cellular and molecular processes of hippocampal-dependent long-term memory consolidation including synaptic plasticity, synaptic scaling and neurogenesis. The chemokine, CX3CL1 (fractalkine), has been shown to modulate synaptic transmission and long-term potentiation (LTP) in the CA1 pyramidal cell layer of the hippocampus. Here, we confirm widespread expression of CX3CL1 on mature neurons in the adult rat hippocampus. We report an up-regulation in CX3CL1 protein expression in the CA1, CA3 and dentate gyrus (DG) of the rat hippocampus 2 h after spatial learning in the water maze task. Moreover, the same temporal increase in CX3CL1 was evident following LTP-inducing theta-burst stimulation in the DG. At physiologically relevant concentrations, CX3CL1 inhibited LTP maintenance in the DG. This attenuation in dentate LTP was lost in the presence of GABAA receptor/chloride channel antagonism. CX3CL1 also had opposing actions on glutamate-mediated rise in intracellular calcium in hippocampal organotypic slice cultures in the presence and absence of GABAA receptor/chloride channel blockade. Using primary dissociated hippocampal cultures, we established that CX3CL1 reduces glutamate-mediated intracellular calcium rises in both neurons and glia in a dose dependent manner. In conclusion, CX3CL1 is up-regulated in the hippocampus during a brief temporal window following spatial learning the purpose of which may be to regulate glutamate-mediated neurotransmission tone. Our data supports a possible role for this chemokine in the protective plasticity process of synaptic scaling. PMID:25161610

  11. Nitrosative stress leads to protein glutathiolation, increased s-nitrosation, and up-regulation of peroxiredoxins in the heart.

    PubMed

    Reinartz, Michael; Ding, Zhaoping; Flögel, Ulrich; Gödecke, Axel; Schrader, Jürgen

    2008-06-20

    Nitric oxide (NO) is produced by different isoforms of nitric oxide synthases (NOSs) and operates as a mediator of important cell signaling pathways, such as the cGMP signaling cascade. Another mechanism by which NO exerts biological effects is mediated through S-nitrosation of target proteins. To explore thiol-based protein modifications in a situation of defined nitrosative stress, we used a transgenic mouse model with cardiac specific overexpression of inducible nitric oxide synthase (iNOS) and concomitant myoglobin deficiency (iNOS(+)/myo(-/-)). In comparison with the wild type hearts, protein glutathiolation detected by immunoblotting was significantly enhanced in iNOS(+)/myo(-/-) hearts, whereas protein S-nitrosation as measured by the biotin switch assay and two-dimensional PAGE revealed that nearly all of the detected proteins ( approximately 60) remained unchanged with the exception of three proteins. Tandem mass spectrometry revealed these proteins to be peroxiredoxins (Prxs), which are known to possess peroxidase activity, whereby hydrogen peroxide, peroxynitrite, and a wide range of organic hydroperoxides are reduced and detoxified. Immunoblotting with specific antibodies revealed up-regulation of Prx VI in the iNOS(+)/myo(-/-) hearts, whereas expression of Prx II and Prx III remained unchanged. Furthermore, the analysis of the cardiac S-nitrososubproteome identified several new proteins possibly being involved in NO-signaling pathways. Our data indicate that S-nitrosation and glutathiolation of cardiac proteins may contribute to the phenotype of NO-induced heart failure. The up-regulation of antioxidant proteins like Prx VI appears to be an additional mechanism to antagonize an excess of reactive oxygen/nitrogen species. Furthermore, S-nitrosation of Prxs may serve a new function in the signaling cascade of nitrosative stress.

  12. Up-regulation of gelatinases and tissue type plasminogen activator by root canal sealers in human osteoblastic cells.

    PubMed

    Huang, Fu-Mei; Yang, Shun-Fa; Chang, Yu-Chao

    2008-03-01

    Histologic investigations have demonstrated that root canal sealers can induce mild to severe inflammatory alternations. However, there is little information on the precise mechanisms about root canal sealer-induced inflammatory reaction. The proteolysis of extracellular matrix by matrix metalloproteinases (MMPs) and plasminogen activators (PAs) seems to be a key initiating event for the progression of the inflammatory process. The aim of this study was to investigate the effects of epoxy resin-based root canal sealer AH26 and zinc oxide-eugenol-based root canal sealer Canals and one paste sealer N2 on the expression of MMPs and PAs in human osteoblastic cell line U2OS cells. The levels of gelatinolytic and caseinolytic activities were measured by gelatin and casein zymography. The results showed that AH26, Canals, and N2 were cytotoxic to U2OS cells in a concentration-dependent manner (P < .05). The gelatin zymograms revealed that MMP-2 (72 kd) and MMP-9 (92 kd) were secreted by U2OS cells. The exposure of U2OS cells to root canal sealers resulted in the up-regulation of MMP-2 and MMP-9 expression (P < .05). Casein zymography exhibited a caseinolytic band with a molecular weight of 70 kd, indicative of the presence of tissue type plasminogen activators (t-PA). t-PA was also found to be up-regulated by root canal sealers (P < .05). Taken together, the activation of gelatinases and t-PA might play an important role in the pathogenesis of root canal sealer-induced periapical inflammation.

  13. Immunomodulatory drugs act as inhibitors of DNA methyltransferases and induce PU.1 up-regulation in myeloma cells.

    PubMed

    Endo, Shinya; Amano, Masayuki; Nishimura, Nao; Ueno, Niina; Ueno, Shikiko; Yuki, Hiromichi; Fujiwara, Shiho; Wada, Naoko; Hirata, Shinya; Hata, Hiroyuki; Mitsuya, Hiroaki; Okuno, Yutaka

    2016-01-01

    Immunomodulatory drugs (IMiDs) such as thalidomide, lenalidomide, and pomalidomide are efficacious in the treatment of multiple myeloma and significantly prolong their survival. However, the mechanisms of such effects of IMiDs have not been fully elucidated. Recently, cereblon has been identified as a target binding protein of thalidomide. Lenalidomide-resistant myeloma cell lines often lose the expression of cereblon, suggesting that IMiDs act as an anti-myeloma agent through interacting with cereblon. Cereblon binds to damaged DNA-binding protein and functions as a ubiquitin ligase, inducing degradation of IKZF1 and IKZF3 that are essential transcription factors for B and T cell development. Degradation of both IKZF1 and IKZF3 reportedly suppresses myeloma cell growth. Here, we found that IMiDs act as inhibitors of DNA methyltransferases (DMNTs). We previously reported that PU.1, which is an ETS family transcription factor and essential for myeloid and lymphoid development, functions as a tumor suppressor in myeloma cells. PU.1 induces growth arrest and apoptosis of myeloma cell lines. In this study, we found that low-dose lenalidomide and pomalidomide up-regulate PU.1 expression through inducing demethylation of the PU.1 promoter. In addition, IMiDs inhibited DNMT1, DNMT3a, and DNMT3b activities in vitro. Furthermore, lenalidomide and pomalidomide decreased the methylation status of the whole genome in myeloma cells. Collectively, IMiDs exert demethylation activity through inhibiting DNMT1, 3a, and 3b, and up-regulating PU.1 expression, which may be one of the mechanisms of the anti-myeloma activity of IMiDs.

  14. Technology coordination

    NASA Technical Reports Server (NTRS)

    Hartman, Steven

    1992-01-01

    Viewgraphs on technology coordination are provided. Topics covered include: technology coordination process to date; goals; how the Office of Aeronautics and Space Technology (OAST) can support the Office of Space Science and Applications (OSSA); how OSSA can support OAST; steps to technology transfer; and recommendations.

  15. Nicotinic acetylcholine receptors containing alpha 7 subunits on rat cortical neurons do not undergo long-lasting inactivation even when up-regulated by chronic nicotine exposure.

    PubMed

    Kawai, H; Berg, D K

    2001-09-01

    Chronic exposure to (-)nicotine has been widely reported to up-regulate nicotinic acetylcholine receptors on neurons and induce long-term inactivation as a possible cause. Nicotinic receptors containing alpha 7 subunits are among the most abundant in brain and influence diverse cellular events. Whole-cell patch clamp recording from embryonic rat cortical neurons in culture was used to identify responses from alpha 7-containing receptors. Immunochemical staining for glutamic acid decarboxylase (GAD) indicated that both GABAergic and non-GABAergic neurons expressed the receptors. Exposure to micromolar concentrations of nicotine for 1-4 days caused up-regulation of the receptors as measured by [alpha-(125)I]-bungarotoxin binding. Carbachol produced the same up-regulation, and cell counts demonstrated that neuronal survival was unchanged. The up-regulation was accompanied by an increased whole-cell response; no evidence was found for long-lasting inactivation. Autonomic alpha 7-containing receptors also avoided long-lasting inactivation, even though the receptors were down-regulated by nicotine. Blocking protein synthesis or protein glycosylation prevented receptor up-regulation on cortical neurons, suggesting that new synthesis was required. No evidence was found for a pre-existing intracellular pool that supplied receptors to the surface. The results indicate that alpha 7-containing receptors differ from other receptor subtypes in their regulation by nicotine and demonstrate further that long-lasting inactivation is not an obligatory requirement for up-regulation in this case.

  16. Poisson Coordinates.

    PubMed

    Li, Xian-Ying; Hu, Shi-Min

    2013-02-01

    Harmonic functions are the critical points of a Dirichlet energy functional, the linear projections of conformal maps. They play an important role in computer graphics, particularly for gradient-domain image processing and shape-preserving geometric computation. We propose Poisson coordinates, a novel transfinite interpolation scheme based on the Poisson integral formula, as a rapid way to estimate a harmonic function on a certain domain with desired boundary values. Poisson coordinates are an extension of the Mean Value coordinates (MVCs) which inherit their linear precision, smoothness, and kernel positivity. We give explicit formulas for Poisson coordinates in both continuous and 2D discrete forms. Superior to MVCs, Poisson coordinates are proved to be pseudoharmonic (i.e., they reproduce harmonic functions on n-dimensional balls). Our experimental results show that Poisson coordinates have lower Dirichlet energies than MVCs on a number of typical 2D domains (particularly convex domains). As well as presenting a formula, our approach provides useful insights for further studies on coordinates-based interpolation and fast estimation of harmonic functions.

  17. Up-regulation of eEF1A2 promotes proliferation and inhibits apoptosis in prostate cancer

    SciTech Connect

    Sun, Yue; Du, Chengli; Wang, Bo; Zhang, Yanling; Liu, Xiaoyan; Ren, Guoping

    2014-07-18

    Highlights: • The expression of eEF1A2 is up-regulated in prostate cancer tissues. • Suppression of eEF1A2 inhibits the proliferation and promotes apoptosis. • Inhibition of eEF1A2 enhances the expression of apoptotic relevant proteins. • The expressions of eEF1A2 and cleavage-caspase3 are inversely correlated. - Abstract: Background: eEF1A2 is a protein translation factor involved in protein synthesis, which possesses important function roles in cancer development. This study aims at investigating the expression pattern of eEF1A2 in prostate cancer and its potential role in prostate cancer development. Methods: We examined the expression level of eEF1A2 in 30 pairs of prostate cancer tissues by using RT-PCR and immunohistochemical staining (IHC). Then we applied siRNA specifically targeting eEF1A2 to down-regulate its expression in DU-145 and PC-3 cells. Flow cytometer was used to explore apoptosis and Western-blot was used to detect the pathway proteins of apoptosis. Results: Our results showed that the expression level of eEF1A2 in prostate cancer tissues was significantly higher compared to their corresponding normal tissues. Reduction of eEF1A2 expression in DU-145 and PC-3 cells led to a dramatic inhibition of proliferation accompanied with enhanced apoptosis rate. Western blot revealed that apoptosis pathway proteins (caspase3, BAD, BAX, PUMA) were significantly up-regulated after suppression of eEF1A2. More importantly, the levels of eEF1A2 and caspase3 were inversely correlated in prostate cancer tissues. Conclusion: Our data suggests that eEF1A2 plays an important role in prostate cancer development, especially in inhibiting apoptosis. So eEF1A2 might serve as a potential therapeutic target in prostate cancer.

  18. TNF-α contributes to up-regulation of Nav1.3 and Nav1.8 in DRG neurons following motor fiber injury.

    PubMed

    He, Xin-Hua; Zang, Ying; Chen, Xi; Pang, Rui-Ping; Xu, Ji-Tian; Zhou, Xiang; Wei, Xu-Hong; Li, Yong-Yong; Xin, Wen-Jun; Qin, Zhi-Hai; Liu, Xian-Guo

    2010-11-01

    A large body of evidence has demonstrated that the ectopic discharges of action potentials in primary afferents, resulted from the abnormal expression of voltage gated sodium channels (VGSCs) in dorsal root ganglion (DRG) neurons following peripheral nerve injury are important for the development of neuropathic pain. However, how nerve injury affects the expression of VGSCs is largely unknown. Here, we reported that selective injury of motor fibers by L5 ventral root transection (L5-VRT) up-regulated Nav1.3 and Nav1.8 at both mRNA and protein level and increased current densities of TTX-S and TTX-R channels in DRG neurons, suggesting that nerve injury may up-regulate functional VGSCs in sensory neurons indirectly. As the up-regulated Nav1.3 and Nav1.8 were highly co-localized with TNF-α, we tested the hypothesis that the increased TNF-α may lead to over-expression of the sodium channels. Indeed, we found that peri-sciatic administration of recombinant rat TNF-α (rrTNF) without any nerve injury, which produced lasting mechanical allodynia, also up-regulated Nav1.3 and Nav1.8 in DRG neurons in vivo and that rrTNF enhanced the expression of Nav1.3 and Nav1.8 in cultured adult rat DRG neurons in a dose-dependent manner. Furthermore, inhibition of TNF-α synthesis, which prevented neuropathic pain, strongly inhibited the up-regulation of Nav1.3 and Nav1.8. The up-regulation of the both channels following L5-VRT was significantly lower in TNF receptor 1 knockout mice than that in wild type mice. These data suggest that increased TNF-α may be responsible for up-regulation of Nav1.3 and Nav1.8 in uninjured DRG neurons following nerve injury. PMID:20638792

  19. β₁-adrenergic receptor up-regulation induced by nadolol is mediated via signal transduction pathway coupled to α₁-adrenergic receptors.

    PubMed

    Mizuno, Koji; Kurokawa, Kazuhiro; Shibasaki, Masahiro; Ohkuma, Seitaro

    2011-09-26

    Although up-regulation of β-adrenergic receptors (β-ARs) occurs after long-term use of their antagonists in various tissues, the available data are little on mechanisms of β-AR up-regulation induced by their continuous blockade. The present study attempted to clarify mechanisms of β-AR up-regulation using mouse cerebral cortical neurons continuously exposed to nadolol (10 nM), a non-selective β-AR antagonist, for 24 h. Nadolol dose-dependently induced both subtypes of β-ARs, β₁- and β₂-ARs, which were not suppressed by protein A kinase inhibition with KT5720. On the other hand, blockade of α₁-ARs, which are immunohistochemically confirmed to be co-localized with β-ARs in the same neurons, significantly inhibited only β₁-AR up-regulation and the expression of β₂-ARs did not alter. In addition, phenylephrine, an agonist specific to α₁-ARs up-regulated β₁-ARs, but not β₂-ARs. Under the conditions with β-AR up-regulation, the level of phosphorylated protein kinase Cα (pPKCα) increased, which is significantly suppressed by prazosin, an α1-AR antagonist. Furthermore, nadolol decreased the degradation of mRNA of β₁-ARs, but not β₂-ARs. These results indicate that the nadolol-induced β₁-AR up-regulation is mediated via PKC-relating pathway via α₁-AR activation with stabilizing β₁-AR mRNA and that the increased expression of β₂-ARs is regulated by pathways different from those for β₁-AR expression.

  20. Pancreatic adenocarcinoma up-regulated factor expression is associated with disease-specific survival in cervical cancer patients.

    PubMed

    Choi, Chel Hun; Chung, Joon-Yong; Park, Ho-Seop; Jun, Minsik; Lee, Yoo-Young; Kim, Byung-Gie; Hewitt, Stephen M

    2015-06-01

    Pancreatic adenocarcinoma up-regulated factor (PAUF) is a novel soluble protein involved in tumor development and metastases. This study was to investigate the PAUF expression and its prognostic value in cervical cancer patients. The expression of PAUF was immunohistochemically determined in 345 formalin-fixed, paraffin-embedded cervical cancer tissues and 107 normal cervical epitheliums. Subsequently, its associations with clinicopathological characteristics and patient survival were assessed. PAUF protein was expressed both in cytoplasm and nucleus, and cytoplasmic expression was more frequent in cancers than normal tissues (32% versus 17%, P = .002), and the difference was prominent in glandular cells. Notably, the expression was more frequent in adenocarcinoma than in squamous cell carcinoma (57% versus 25%, respectively; P < .001), and the differential expression was also seen at the messenger RNA level (P = .014). Cox regression analysis showed that the cytoplasmic expression of PAUF protein was independently associated with poor disease-free (hazard ratio = 2.3; 95% confidence interval, 1.2-4.3; P = .008) and overall survival (hazard ratio = 2.9; 95% confidence interval, 1.2-7.5; P = .020). Detection of PAUF expression may aid current evaluation of prognosis in cervical adenocarcinoma.

  1. Chromatin remodelling and antisense-mediated up-regulation of the developmental switch gene eud-1 control predatory feeding plasticity

    PubMed Central

    Serobyan, Vahan; Xiao, Hua; Namdeo, Suryesh; Rödelsperger, Christian; Sieriebriennikov, Bogdan; Witte, Hanh; Röseler, Waltraud; Sommer, Ralf J.

    2016-01-01

    Phenotypic plasticity has been suggested to act through developmental switches, but little is known about associated molecular mechanisms. In the nematode Pristionchus pacificus, the sulfatase eud-1 was identified as part of a developmental switch controlling mouth-form plasticity governing a predatory versus bacteriovorous mouth-form decision. Here we show that mutations in the conserved histone-acetyltransferase Ppa-lsy-12 and the methyl-binding-protein Ppa-mbd-2 mimic the eud-1 phenotype, resulting in the absence of one mouth-form. Mutations in both genes cause histone modification defects and reduced eud-1 expression. Surprisingly, Ppa-lsy-12 mutants also result in the down-regulation of an antisense-eud-1 RNA. eud-1 and antisense-eud-1 are co-expressed and further experiments suggest that antisense-eud-1 acts through eud-1 itself. Indeed, overexpression of the antisense-eud-1 RNA increases the eud-1-sensitive mouth-form and extends eud-1 expression. In contrast, this effect is absent in eud-1 mutants indicating that antisense-eud-1 positively regulates eud-1. Thus, chromatin remodelling and antisense-mediated up-regulation of eud-1 control feeding plasticity in Pristionchus. PMID:27487725

  2. Repeated aripiprazole treatment causes dopamine D2 receptor up-regulation and dopamine supersensitivity in young rats.

    PubMed

    Varela, Fausto A; Der-Ghazarian, Taleen; Lee, Ryan J; Charntikov, Sergios; Crawford, Cynthia A; McDougall, Sanders A

    2014-04-01

    Aripiprazole is a second-generation antipsychotic that is increasingly being prescribed to children and adolescents. Despite this trend, little preclinical research has been done on the neural and behavioral actions of aripiprazole during early development. In the present study, young male and female Sprague-Dawley rats were pretreated with vehicle, haloperidol (1 mg/kg), or aripiprazole (10 mg/kg) once daily on postnatal days (PD) 10-20. After 1, 4, or 8 days (i.e. on PD 21, PD 24, or PD 28), amphetamine-induced locomotor activity and stereotypy, as well as dorsal striatal D2 receptor levels, were measured in separate groups of rats. Pretreating young rats with aripiprazole or haloperidol increased D2 binding sites in the dorsal striatum. Consistent with these results, dopamine supersensitivity was apparent when aripiprazole- and haloperidol-pretreated rats were given a test day injection of amphetamine (2 or 4 mg/kg). Increased D2 receptor levels and altered behavioral responding persisted for at least 8 days after conclusion of the pretreatment regimen. Contrary to what has been reported in adults, repeated aripiprazole treatment caused D2 receptor up-regulation and persistent alterations of amphetamine-induced behavior in young rats. These findings are consistent with human clinical studies showing that children and adolescents are more prone than adults to aripiprazole-induced side effects, including extrapyramidal symptoms.

  3. Effect of caffeine on erectile function via up-regulating cavernous cyclic guanosine monophosphate in diabetic rats.

    PubMed

    Yang, Rong; Wang, Jiuling; Chen, Yun; Sun, Zeyu; Wang, Run; Dai, Yutian

    2008-01-01

    Erectile dysfunction (ED) is a common complication of diabetes mellitus. Phosphodiesterase-5 (PDE5) inhibitors, which inhibit the breakdown of intracellular cyclic guanosine monophosphate (cGMP), are used to treat diabetic ED. Caffeine, a nonselective PDE inhibitor used in our daily diet, is controversial regarding its effect on erectile function. To investigate the effect of caffeine on erectile function in diabetic rat models and explore the mechanism, male Sprague-Dawley rats were injected with streptozotocin to induce diabetes mellitus. The rats with blood glucose levels above 300 mg/dL were selected for the study. The rats were divided into 4 groups: group A (normal control rats), group B (diabetic rats treated with normal saline), group C (diabetic rats treated with caffeine, 10 mg/kg per day), and group D (diabetic rats treated with caffeine, 20 mg/kg per day). After 8 weeks of treatment, intracavernous pressure (ICP) was measured to assess erectile function. The radioimmunoassay was used to evaluate the level of cGMP in the cavernosum. The ICP and the cavernous cGMP decreased significantly in the diabetic rats compared with normal controls. An 8-week administration of caffeine at the given dosages increased the ICP and cavernous cGMP in diabetic rats. Caffeine consumption improved the erectile function of diabetic rats by up-regulating cavernous cGMP.

  4. Transcript profiling reveals that cysteine protease inhibitors are up-regulated in tuber sprouts after extended darkness.

    PubMed

    Grandellis, Carolina; Giammaria, Veronica; Fantino, Elisa; Cerrudo, Ignacio; Bachmann, Sandra; Santin, Franco; Ulloa, Rita M

    2016-07-01

    Potato (Solanum tuberosum L.) tubers are an excellent staple food due to its high nutritional value. When the tuber reaches physiological competence, sprouting proceeds accompanied by changes at mRNA and protein levels. Potato tubers become a source of carbon and energy until sprouts are capable of independent growth. Transcript profiling of sprouts grown under continuous light or dark conditions was performed using the TIGR 10K EST Solanaceae microarray. The profiles analyzed show a core of highly expressed transcripts that are associated to the reactivation of growth. Under light conditions, the photosynthetic machinery was fully activated; the highest up-regulation was observed for the Rubisco activase (RCA), the glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and the Photosystem II 22 kDa protein (CP22) genes, among others. On the other hand, sprouts exposed to continuous darkness elongate longer, and after extended darkness, synthesis of chloroplast components was repressed, the expression of proteases was reduced while genes encoding cysteine protease inhibitors (CPIs) and metallocarboxypeptidase inhibitors (MPIs) were strongly induced. Northern blot and RT-PCR analysis confirmed that MPI levels correlated with the length of the dark period; however, CPI expression was strong only after longer periods of darkness, suggesting a feedback loop (regulation mechanism) in response to dark-induced senescence. Prevention of cysteine protease activity in etiolated sprouts exposed to extended darkness could delay senescence until they emerge to light.

  5. Cinnamon and its Components Suppress Vascular Smooth Muscle Cell Proliferation by Up-Regulating Cyclin-Dependent Kinase Inhibitors.

    PubMed

    Kwon, Hyeeun; Lee, Jung-Jin; Lee, Ji-Hye; Cho, Won-Kyung; Gu, Min Jung; Lee, Kwang Jin; Ma, Jin Yeul

    2015-01-01

    Cinnamomum cassia bark has been used in traditional herbal medicine to treat a variety of cardiovascular diseases. However, the antiproliferative effect of cinnamon extract on vascular smooth muscle cells (VSMCs) and the corresponding restenosis has not been explored. Hence, after examining the effect of cinnamon extract on VSMC proliferation, we investigated the possible involvement of signal transduction pathways associated with early signal and cell cycle analysis, including regulatory proteins. Besides, to identify the active components, we investigated the components of cinnamon extract on VSMC proliferation. Cinnamon extract inhibited platelet-derived growth factor (PDGF)-BB-induced VSMC proliferation and suppressed the PDGF-stimulated early signal transduction. In addition, cinnamon extract arrested the cell cycle and inhibited positive regulatory proteins. Correspondingly, the protein levels of p21 and p27 not only were increased in the presence of cinnamon extract, also the expression of proliferating cell nuclear antigen (PCNA) was inhibited by cinnamon extract. Besides, among the components of cinnamon extract, cinnamic acid (CA), eugenol (EG) and cinnamyl alcohol significantly inhibited the VSMC proliferation. Overall, the present study demonstrates that cinnamon extract inhibited the PDGF-BB-induced proliferation of VSMCs through a G0/G1 arrest, which down-regulated the expression of cell cycle positive regulatory proteins by up-regulating p21 and p27 expression.

  6. Apocynin improving cardiac remodeling in chronic renal failure disease is associated with up-regulation of epoxyeicosatrienoic acids.

    PubMed

    Zhang, Kun; Liu, Yu; Liu, Xiaoqiang; Chen, Jie; Cai, Qingqing; Wang, Jingfeng; Huang, Hui

    2015-09-22

    Cardiac remodeling is one of the most common cardiac abnormalities and associated with a high mortality in chronic renal failure (CRF) patients. Apocynin, a nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase inhibitor, has been showed cardio-protective effects. However, whether apocynin can improve cardiac remodeling in CRF and what is the underlying mechanism are unclear. In the present study, we enrolled 94 participants. In addition, we used 5/6 nephrectomized rats to mimic cardiac remodeling in CRF. Serum levels of epoxyeicosatrienoic acids (EETs) and its mainly metabolic enzyme-soluble epoxide hydrolase (sEH) were measured. The results showed that the serum levels of EETs were significantly decreased in renocardiac syndrome participants (P < 0.05). In 5/6 nephrectomized CRF model, the ratio of left ventricular weight / body weight, left ventricular posterior wall thickness, and cardiac interstitial fibrosis were significantly increased while ejection fraction significantly decreased (P < 0.05). All these effects could partly be reversed by apocynin. Meanwhile, we found during the process of cardiac remodeling in CRF, apocynin significantly increased the reduced serum levels of EETs and decreased the mRNA and protein expressions of sEH in the heart (P < 0.05). Our findings indicated that the protective effect of apocynin on cardiac remodeling in CRF was associated with the up-regulation of EETs. EETs may be a new mediator for the injury of kidney-heart interactions.

  7. Up-regulation of Na + expression in the area postrema of total sleep deprived rats by TOF-SIMS analysis

    NASA Astrophysics Data System (ADS)

    Mai, Fu-Der; Chen, Bo-Jung; Ling, Yong-Chien; Wu, Un-In; Huang, Yi-Lun; Chang, Hung-Ming

    2008-12-01

    Area postrema (AP) is a circumventricular organ plays an important role in sodium homeostasis and cardiovascular regulation. Since sleep deficiency will cause cardiovascular dysfunction, the present study aims to determine whether sodium level would significantly alter in AP following total sleep deprivation (TSD). Sodium level was investigated in vivo by time-of-flight secondary ion mass spectrometry (TOF-SIMS). Clinical manifestation of cardiovascular function was demonstrated by mean arterial pressure (MAP) values. Results indicated that in normal rats, TOF-SIMS spectrum revealed a major peak of sodium ion counting as 5.61 × 10 5 at m/ z 23. The sodium ions were homogeneous distributed in AP without specific localization. However, following TSD, the sodium intensity was relatively increased (6.73 × 10 5) and the signal for sodium image was strongly expressed throughout AP with definite spatial distribution. MAP of TSD rats is 138 ± 5 mmHg, which is significantly higher than that of normal ones (121 ± 3 mmHg). Regarding AP is an important area for sodium sensation and development of hypernatremic related sympatho-excitation; up-regulation of sodium expression following TSD suggests that high sodium level might over-activate AP, through complex neuronal networks involving in sympathetic regulation, which could lead to the formation of TSD relevant cardiovascular diseases.

  8. Coagulation factor Xa drives tumor cells into apoptosis through BH3-only protein Bim up-regulation

    SciTech Connect

    Borensztajn, Keren S. . E-mail: K.S.Borensztajn@amc.uva.nl; Bijlsma, Maarten F.; Groot, Angelique P.; Brueggemann, Lois W.; Versteeg, Henri H.; Reitsma, Pieter H.; Peppelenbosch, Maikel P.; Spek, C. Arnold

    2007-07-15

    Coagulation Factor (F)Xa is a serine protease that plays a crucial role during blood coagulation by converting prothrombin into active thrombin. Recently, however, it emerged that besides this role in coagulation, FXa induces intracellular signaling leading to different cellular effects. Here, we show that coagulation factor (F)Xa drives tumor cells of epithelial origin, but not endothelial cells or monocytes, into apoptosis, whereas it even enhances fibroblast survival. FXa signals through the protease activated receptor (PAR)-1 to activate extracellular-signal regulated kinase (ERK) 1/2 and p38. This activation is associated with phosphorylation of the transcription factor CREB, and in tumor cells with up-regulation of the BH3-only pro-apoptotic protein Bim, leading to caspase-3 cleavage, the main hallmark of apoptosis. Transfection of tumor cells with dominant negative forms of CREB or siRNA for either PAR-1, Bim, ERK1 and/or p38 inhibited the pro-apoptotic effect of FXa. In fibroblasts, FXa-induced PAR-1 activation leads to down-regulation of Bim and pre-treatment with PAR-1 or Bim siRNA abolishes proliferation. We thus provide evidence that beyond its role in blood coagulation, FXa plays a key role in cellular processes in which Bim is the central player in determining cell survival.

  9. The long noncoding RNA MALAT1 promotes tumor-driven angiogenesis by up-regulating pro-angiogenic gene expression

    PubMed Central

    Tee, Andrew E.; Liu, Bing; Song, Renhua; Li, Jinyan; Pasquier, Eddy; Cheung, Belamy B.; Jiang, Cizhong; Marshall, Glenn M.; Haber, Michelle; Norris, Murray D.; Fletcher, Jamie I.; Dinger, Marcel E.; Liu, Tao

    2016-01-01

    Neuroblastoma is the most common solid tumor during early childhood. One of the key features of neuroblastoma is extensive tumor-driven angiogenesis due to hypoxia. However, the mechanism through which neuroblastoma cells drive angiogenesis is poorly understood. Here we show that the long noncoding RNA MALAT1 was upregulated in human neuroblastoma cell lines under hypoxic conditions. Conditioned media from neuroblastoma cells transfected with small interfering RNAs (siRNA) targeting MALAT1, compared with conditioned media from neuroblastoma cells transfected with control siRNAs, induced significantly less endothelial cell migration, invasion and vasculature formation. Microarray-based differential gene expression analysis showed that one of the genes most significantly down-regulated following MALAT1 suppression in human neuroblastoma cells under hypoxic conditions was fibroblast growth factor 2 (FGF2). RT-PCR and immunoblot analyses confirmed that MALAT1 suppression reduced FGF2 expression, and Enzyme-Linked Immunosorbent Assays revealed that transfection with MALAT1 siRNAs reduced FGF2 protein secretion from neuroblastoma cells. Importantly, addition of recombinant FGF2 protein to the cell culture media reversed the effects of MALAT1 siRNA on vasculature formation. Taken together, our data suggest that up-regulation of MALAT1 expression in human neuroblastoma cells under hypoxic conditions increases FGF2 expression and promotes vasculature formation, and therefore plays an important role in tumor-driven angiogenesis. PMID:26848616

  10. MIP-1α enhances Jurkat cell transendothelial migration by up-regulating endothelial adhesion molecules VCAM-1 and ICAM-1.

    PubMed

    Ma, Yi-Ran; Ma, Ying-Huan

    2014-11-01

    The aim of this study is to evaluate the expression of macrophage inflammatory protein-1α (MIP-1α) in Jurkat cells and its effect on transendothelial migration. In the present study, human acute lymphoblastic leukemia Jurkat cells (Jurkat cells) were used as a model of T cells in human T-cell acute lymphoblastic leukemia (T-ALL), which demonstrated significantly higher MIP-1α expression compared with that in normal T-cell controls. The ability of Jurkat cells to cross a human brain microvascular endothelial cell (HBMEC) monolayer was almost completely abrogated by MIP-1α siRNA. In addition, the overexpression of MIP-1α resulted in the up-regulated expression of endothelial adhesion molecules, which enhanced the migration of Jurkat cells through a monolayer of HBMEC. MIP-1α levels in Jurkat cells appeared to be an important factor for its transendothelial migration, which may provide the theoretical basis to understand the mechanisms of brain metastases of T-ALL at cellular and molecular levels.

  11. Up-regulation of cholesterol associated genes as novel resistance mechanism in glioblastoma cells in response to archazolid B

    SciTech Connect

    Hamm, Rebecca; Zeino, Maen; Frewert, Simon; Efferth, Thomas

    2014-11-15

    Treatment of glioblastoma multiforme (GBM), the most common and aggressive lethal brain tumor, represents a great challenge. Despite decades of research, the survival prognosis of GBM patients is unfavorable and more effective therapeutics are sorely required. Archazolid B, a potent vacuolar H{sup +}-ATPase inhibitor influencing cellular pH values, is a promising new compound exerting cytotoxicity in the nanomolar range on wild-type U87MG glioblastoma cells and U87MG.∆EGFR cells transfected with a mutant epidermal growth factor receptor (EGFR) gene. Gene expression profiling using microarray technology showed that archazolid B caused drastic disturbances in cholesterol homeostasis. Cholesterol, a main component of cellular membranes, is known to be essential for GBM growth and cells bearing EGFRvIII mutation are highly dependent on exogenous cholesterol. Archazolid B caused excessive accumulation of free cholesterol within intracellular compartments thus depleting cellular cholesterol and leading to up-regulation of SREBP targeted genes, including LDLR and HMGCR, the key enzyme of cholesterol biosynthesis. This cholesterol response is considered to be a novel resistance mechanism induced by archazolid B. We surmise that re-elevation of cholesterol levels in archazolid B treated cells may be mediated by newly synthesized cholesterol, since the drug leads to endosomal/lysosomal malfunction and cholesterol accumulation.

  12. Prolonged Starvation Causes Up-Regulation of AQP1 in Adipose Tissue Capillaries of AQP7 Knock-Out Mice.

    PubMed

    Skowronski, Mariusz T; Skowronska, Agnieszka; Rojek, Aleksandra; Oklinski, Michal K; Nielsen, Søren

    2016-01-01

    Aquaporins (AQPs) are membrane proteins involved in the regulation of cellular transport and the balance of water and glycerol and cell volume in the white adipose tissue (WAT). In our previous study, we found the co-expression of the AQP1 water channel and AQP7 in the mouse WAT. In our present study, we aimed to find out whether prolonged starvation influences the AQP1 expression of AQP7 knock-out mice (AQP7 KO) in the WAT. To resolve this hypothesis, immunoperoxidase, immunoblot and immunogold microscopy were used. AQP1 expression was found with the use of immunohistochemistry and was confirmed by immunogold microscopy in the vessels of mouse WAT of all studied groups. Semi-quantitative immunoblot and quantitative immunogold microscopy showed a significant increase (by 2.5- to 3-fold) in the abundance of AQP1 protein expression in WAT in the 72 h starved AQP7 KO mice as compared to AQP7+/+ (p < 0.05) and AQP7-/- (p < 0.01) controls, respectively. In conclusion, the AQP1 water channel located in the vessels of WAT is up-regulated in response to prolonged starvation in the WAT of AQP7 KO mice. The present data suggest that an interaction of different AQP isoforms is required for maintaining proper water homeostasis within the mice WAT. PMID:27455244

  13. Interplay between up-regulation of cytochrome-c-oxidase and hemoglobin oxygenation induced by near-infrared laser.

    PubMed

    Wang, Xinlong; Tian, Fenghua; Soni, Sagar S; Gonzalez-Lima, F; Liu, Hanli

    2016-01-01

    Photobiomodulation, also known as low-level laser/light therapy (LLLT), refers to the use of red-to-near-infrared light to stimulate cellular functions for physiological or clinical benefits. The mechanism of LLLT is assumed to rely on photon absorption by cytochrome c oxidase (CCO), the terminal enzyme in the mitochondrial respiratory chain that catalyzes the reduction of oxygen for energy metabolism. In this study, we used broadband near-infrared spectroscopy (NIRS) to measure the LLLT-induced changes in CCO and hemoglobin concentrations in human forearms in vivo. Eleven healthy participants were administered with 1064-nm laser and placebo treatments on their right forearms. The spectroscopic data were analyzed and fitted with wavelength-dependent, modified Beer-Lambert Law. We found that LLLT induced significant increases of CCO concentration (Δ[CCO]) and oxygenated hemoglobin concentration (Δ[HbO]) on the treated site as the laser energy dose accumulated over time. A strong linear interplay between Δ[CCO] and Δ[HbO] was observed for the first time during LLLT, indicating a hemodynamic response of oxygen supply and blood volume closely coupled to the up-regulation of CCO induced by photobiomodulation. These results demonstrate the tremendous potential of broadband NIRS as a non-invasive, in vivo means to study mechanisms of photobiomodulation and perform treatment evaluations of LLLT. PMID:27484673

  14. Up-regulation of CHAF1A, a poor prognostic factor, facilitates cell proliferation of colon cancer

    SciTech Connect

    Wu, Zehua; Cui, Feifei; Yu, Fudong; Peng, Xiao; Jiang, Tao; Chen, Dawei; Lu, Su; Tang, Huamei; Peng, Zhihai

    2014-06-27

    Highlights: • We identified that CHAF1A was up-regulated in colon tumor mucosa in TMA. • The expression pattern of CHAF1A was validated with qPCR and western-blot. • CHAF1A overexpression is an independent indicator for poor colon cancer survival. • CHAF1A facilitates cell proliferation of colon cancer both in vitro and in vivo. - Abstract: Deregulation of chromatin assembly factor 1, p150 subunit A (CHAF1A) has recently been reported to be involved in the development of some cancer types. In this study, we identified that the frequency of positive CHAF1A staining in primary tumor mucosa (45.8%, 93 of 203 samples) was significantly elevated compared to that in paired normal mucosa (18.7%, 38 of 203 samples). The increased expression was strongly associated with cancer stage, tumor invasion, and histological grade. The five-year survival rate of patients with CHAF1A-positive tumors was remarkably lower than that of patients with CHAF1A-negative tumors. Colon cancer cells with CHAF1A knockdown exhibited decreased cell growth index, reduction in colony formation ability, elevated cell apoptosis rate as well as impaired colon tumorigenicity in nude mice. Hence, CHAF1A upregulation functions as a poor prognostic indicator of colon cancer, potentially contributing to its progression by mediating cancer cell proliferation.

  15. Chronic adventitial inflammation, vasa vasorum expansion, and 5-lipoxygenase up-regulation in irradiated arteries from cancer survivors

    PubMed Central

    Halle, Martin; Christersdottir, Tinna; Bäck, Magnus

    2016-01-01

    Radiation-induced cardiovascular disease is an emerging problem in a steadily increasing population of survivors of cancer. However, the underlying biology is poorly described, and the late onset, which occurs several years after exposure, precludes adequate investigations in animal and cell culture models. We investigated the role of the 5-lipoxygenase (5-LO)/leukotriene pathway in radiation-induced vascular changes. Use of paired samples of irradiated arteries and nonirradiated internal control arteries from the same patient that were harvested during surgery for cancer reconstruction ≤10 yr after radiotherapy provides a unique human model of chronic radiation–induced vascular changes. Immunohistochemical stainings and perioperative inspection revealed an adventitial inflammatory response, with vasa vasorum expansion and chronic infiltration of CD68+ macrophages. These macrophages stained positive for the leukotriene-forming enzyme 5-LO. Messenger RNA levels of 5-LO and leukotriene B4 receptor 1 were increased in irradiated arterial segments compared with control vessels. These results point to targeting the 5-LO/leukotriene pathway as a therapeutic adjunct to prevent late adverse vascular effects of radiotherapy.—Halle, M., Christersdottir, T., Bäck, M. Chronic adventitial inflammation, vasa vasorum expansion, and 5-lipoxygenase up-regulation in irradiated arteries from cancer survivors. PMID:27530979

  16. Interleukin 6 trigged ataxia-telangiectasia mutated activation facilitates lung cancer metastasis via MMP-3/MMP-13 up-regulation

    PubMed Central

    Huang, Xiao Bo; Wang, Yi Nan; Li, Qing; Gao, Feng Guang

    2015-01-01

    Our previous studies show that the phosphorylation of ataxia-telangiectasia mutated (ATM) induced by interleukin 6 (IL-6) treatment contributes to multidrug resistance formation in lung cancer cells, but the exact role of ATM activation in IL-6 increased metastasis is still elusive. In the present study, matrix metalloproteinase-3 (MMP-3) and MMP-13 were firstly demonstrated to be involved in IL-6 correlated cell migration. Secondly, IL-6 treatment not only increased MMP-3/MMP-13 expression but also augmented its activities. Thirdly, the inhibition of ATM phosphorylation efficiently abolished IL-6 up-regulating MMP-3/MMP-13 expression and increasing abilities of cell migration. Most importantly, the in vivo test showed that the inhibition of ATM abrogate the effect of IL-6 on lung cancer metastasis via MMP-3/MMP-13 down-regulation. Taken together, these findings demonstrate that IL-6 inducing ATM phosphorylation increases the expression of MMP-3/MMP-13, augments the abilities of cell migration, and promotes lung cancer metastasis, indicating that ATM is a potential target molecule to overcome IL-6 correlated lung cancer metastasis. PMID:26528698

  17. ras-induced Up-regulation of CTP:Phosphocholine Cytidylyltransferase α Contributes to Malignant Transformation of Intestinal Epithelial Cells*

    PubMed Central

    Arsenault, Daniel J.; Yoo, Byong H.; Rosen, Kirill V.; Ridgway, Neale D.

    2013-01-01

    Cancer cells have enhanced lipogenic capacity characterized by increased synthesis of fatty acids and complex lipids, including phosphatidylcholine (PC). As the rate-limiting enzyme in the CDP-choline pathway for PC synthesis, CTP:phosphocholine cytidylyltransferase α (CCTα) is implicated in the provision of membranes and bioactive lipids necessary of cell proliferation. In this study, we assessed the role of CCTα in malignant intestinal epithelial cells transformed with activated H-ras (IEC-ras). Three IEC-ras clones had significant up-regulation CCTα expression, but PC synthesis and in vitro activity of CCTα were similar to control IEC. RNA interference of CCTα in adherent IEC-ras did not affect PC synthesis, confirming that the enzyme was relatively inactive. However, CCTα silencing in ras-transformed IEC reduced anchorage-independent growth, a criterion for malignant transformation, as well as tumorigenicity in mice. Relative to their adherent counterparts, detached IEC-ras had increased PC synthesis that was attenuated by inducible CCTα silencing. Detachment of IEC-ras was accompanied by increased CCTα phosphorylation and cytosolic enzyme activity. We conclude that the expanded pool of CCTα in IEC-ras is activated by detachment. This provides the increased PC biosynthetic capacity that contributes to malignant transformation of intestinal epithelial cells when detached from the extracellular matrix. PMID:23155050

  18. Aniline exposure associated with up-regulated transcriptional responses of three glutathione S-transferase Delta genes in Drosophila melanogaster.

    PubMed

    Chan, Wen-Chiao; Chien, Yi-Chih; Chien, Cheng-I

    2015-03-01

    Complex transcriptional profile of glutathione S-transferase Delta cluster genes occurred in the developmental process of the fruit fly Drosophila melanogaster. The purpose of this project was to quantify the expression levels of Gst Delta class genes altered by aniline exposure and to understand the relationship between aniline dosages and the variation of Gst Delta genes expressed in D. melanogaster. Using RT-PCR expression assays, the expression patterns of the transcript mRNAs of the glutathione S-transferase Delta genes were revealed and their expression levels were measured at eggs, larvae, pupae and adults. The adult stage was selected for further dose-response assays. After analysis, the results indicated that three Gst Delta genes (Gst D2, Gst D5 and Gst D6) were found to show a peak of up-regulated transcriptional response at 6-8h of exposure of aniline. Furthermore, the dose-response relationship of their induction levels within the dose regiments (from 1.2 to 2.0 μl/tube) had been measured. The expression patterns and annotations of these genes were discussed in the context. PMID:25682008

  19. Aniline exposure associated with up-regulated transcriptional responses of three glutathione S-transferase Delta genes in Drosophila melanogaster.

    PubMed

    Chan, Wen-Chiao; Chien, Yi-Chih; Chien, Cheng-I

    2015-03-01

    Complex transcriptional profile of glutathione S-transferase Delta cluster genes occurred in the developmental process of the fruit fly Drosophila melanogaster. The purpose of this project was to quantify the expression levels of Gst Delta class genes altered by aniline exposure and to understand the relationship between aniline dosages and the variation of Gst Delta genes expressed in D. melanogaster. Using RT-PCR expression assays, the expression patterns of the transcript mRNAs of the glutathione S-transferase Delta genes were revealed and their expression levels were measured at eggs, larvae, pupae and adults. The adult stage was selected for further dose-response assays. After analysis, the results indicated that three Gst Delta genes (Gst D2, Gst D5 and Gst D6) were found to show a peak of up-regulated transcriptional response at 6-8h of exposure of aniline. Furthermore, the dose-response relationship of their induction levels within the dose regiments (from 1.2 to 2.0 μl/tube) had been measured. The expression patterns and annotations of these genes were discussed in the context.

  20. HSP70 increases extracellular matrix production by human vascular smooth muscle through TGF-β1 up-regulation.

    PubMed

    González-Ramos, Marta; Calleros, Laura; López-Ongil, Susana; Raoch, Viviana; Griera, Mercedes; Rodríguez-Puyol, Manuel; de Frutos, Sergio; Rodríguez-Puyol, Diego

    2013-02-01

    The circulating levels of heat shock proteins (HSP) are increased in cardiovascular diseases; however, the implication of this for the fibrotic process typical of such diseases remains unclear. HSP70 can interact with the vascular smooth muscle cells (SMC), the major producer of extracellular matrix (ECM) proteins, through the Toll-like receptors 4 (TLR4). The transforming growth factor type-β1 (TGF-β1) is a well known vascular pro-fibrotic cytokine that is regulated in part by AP-1-dependent transcriptional mechanisms. We hypothesized that extracellular HSP70 could interact with SMCs, inducing TGF-β1 synthesis and subsequent changes in the vascular ECM. We demonstrate that extracellular HSP70 binds to human aorta SMC TLR4, which up-regulates the AP-1-dependent transcriptional activity of the TGF-β1 promoter. This is achieved through the mitogen activated protein kinases JNK and ERK, as demonstrated by the use of specific blockers and the knockdown of TLR4 with specific small interfering RNAs. The TGF-β1 upregulation increase the expression of the ECM proteins type I collagen and fibronectin. This novel observation may elucidate the mechanisms by which HSP70 contributes in the inflammation and fibrosis present in atherosclerosis and other fibrosis-related diseases.

  1. Exosomes from hypoxic endothelial cells have increased collagen crosslinking activity through up-regulation of lysyl oxidase-like 2.

    PubMed

    de Jong, Olivier G; van Balkom, Bas W M; Gremmels, Hendrik; Verhaar, Marianne C

    2016-02-01

    Exosomes are important mediators of intercellular communication. Additionally, they contain a variety of components capable of interacting with the extracellular matrix (ECM), including integrins, matrix metalloproteinases and members of the immunoglobin superfamily. Despite these observations, research on exosome-ECM interactions is limited. Here, we investigate whether the exosome-associated lysyl oxidase family member lysyl oxidase-like 2 (LOXL2) is involved in ECM remodelling. We found that LOXL2 is present on the exterior of endothelial cell (EC)-derived exosomes, placing it in direct vicinity of the ECM. It is up-regulated twofold in EC-derived exosomes cultured under hypoxic conditions. Intact exosomes from hypoxic EC and LOXL2 overexpressing EC show increased activity in a fluorometric lysyl oxidase enzymatic activity assay as well as in a collagen gel contraction assay. Concordantly, knockdown of LOXL2 in exosome-producing EC in both normal and hypoxic conditions reduces activity of exosomes in both assays. Our findings show for the first time that ECM crosslinking by EC-derived exosomes is mediated by LOXL2 under the regulation of hypoxia, and implicate a role for exosomes in hypoxia-regulated focal ECM remodelling, a key process in both fibrosis and wound healing.

  2. Up-regulation of Interferon-inducible protein 16 contributes to psoriasis by modulating chemokine production in keratinocytes

    PubMed Central

    Cao, Tianyu; Shao, Shuai; Li, Bing; Jin, Liang; Lei, Jie; Qiao, Hongjiang; Wang, Gang

    2016-01-01

    Psoriasis is a common chronic inflammatory skin disease characterized by epidermal hyperplasia and dermal inflammation. Keratinocyte activation is known to play a critical role in psoriasis, but the underlying mechanism remains unclear. Interferon-inducible protein 16 (IFI16), an innate immune system sensor, is reported to affect keratinocyte function. We therefore hypothesized that IFI16 promotes psoriasis by modulating keratinocyte activation. In the present study, we cinfirmed that IFI16 was overexpressed in epidermal keratinocytes of psoriasis patients. In addition, psoriasis-related cytokines, including IFN-γ, TNF-α, IL-17 and IL-22, induced IFI16 up-regulation in keratinocytes via activation of STAT3 signaling. We also observed that IFI16 activated the TBK1-NF-κB signaling, leading to the production of CXCL10 and CCL20. Importantly, knocking down p204, which is reported as the mouse orthologous of human IFI16, inhibited epidermal hyperplasia in mice with imiquimod-induced psoriasiform dermatitis. These findings indicate that IFI16 plays a critical role in the pathogenesis of psoriasis and may be a potential therapeutic target. PMID:27137868

  3. Hepatic and Nephric NRF2 Pathway Up-Regulation, an Early Antioxidant Response, in Acute Arsenic-Exposed Mice

    PubMed Central

    Li, Jinlong; Duan, Xiaoxu; Dong, Dandan; Zhang, Yang; Li, Wei; Zhao, Lu; Nie, Huifang; Sun, Guifan; Li, Bing

    2015-01-01

    Inorganic arsenic (iAs), a proven human carcinogen, damages biological systems through multiple mechanisms, one of them being reactive oxygen species (ROS) production. NRF2 is a redox-sensitive transcription factor that positively regulates the genes of encoding antioxidant and detoxification enzymes to neutralize ROS. Although NRF2 pathway activation by iAs has been reported in various cell types, however, the experimental data in vivo are very limited and not fully elucidated in humans. The present investigation aimed to explore the hepatic and nephric NRF2 pathway upregulation in acute arsenic-exposed mice in vivo. Our results showed 10 mg/kg NaAsO2 elevated the NRF2 protein and increased the transcription of Nrf2 mRNA, as well as up-regulated NRF2 downstream targets HO-1, GST and GCLC time- and dose-dependently both in the liver and kidney. Acute NaAsO2 exposure also resulted in obvious imbalance of oxidative redox status represented by the increase of GSH and MDA, and the decrease of T-AOC. The present investigation reveals that hepatic and nephric NRF2 pathway expression is an early antioxidant defensive response upon iAs exposure. A better knowledge about the NRF2 pathway involvment in the cellular response against arsenic could help improve the strategies for reducing the cellular toxicity related to this metalloid. PMID:26473898

  4. Folic acid protects against arsenic-mediated embryo toxicity by up-regulating the expression of Dvr1.

    PubMed

    Ma, Yan; Zhang, Chen; Gao, Xiao-Bo; Luo, Hai-Yan; Chen, Yang; Li, Hui-hua; Ma, Xu; Lu, Cai-Ling

    2015-11-05

    As a nutritional factor, folic acid can prevent cardiac and neural defects during embryo development. Our previous study showed that arsenic impairs embryo development by down-regulating Dvr1/GDF1 expression in zebrafish. Here, we investigated whether folic acid could protect against arsenic-mediated embryo toxicity. We found that folic acid supplementation increases hatching and survival rates, decreases malformation rate and ameliorates abnormal cardiac and neural development of zebrafish embryos exposed to arsenite. Both real-time PCR analysis and whole in-mount hybridization showed that folic acid significantly rescued the decrease in Dvr1 expression caused by arsenite. Subsequently, our data demonstrated that arsenite significantly decreased cell viability and GDF1 mRNA and protein levels in HEK293ET cells, while folic acid reversed these effects. Folic acid attenuated the increase in subcellular reactive oxygen species (ROS) levels and oxidative adaptor p66Shc protein expression in parallel with the changes in GDF1 expression and cell viability. P66Shc knockdown significantly inhibited the production of ROS and the down-regulation of GDF1 induced by arsenite. Our data demonstrated that folic acid supplementation protected against arsenic-mediated embryo toxicity by up-regulating the expression of Dvr1/GDF1, and folic acid enhanced the expression of GDF1 by decreasing p66Shc expression and subcellular ROS levels.

  5. Low concentrations of salicylic acid delay methyl jasmonate-induced leaf senescence by up-regulating nitric oxide synthase activity.

    PubMed

    Ji, Yingbin; Liu, Jian; Xing, Da

    2016-09-01

    In plants, extensive efforts have been devoted to understanding the crosstalk between salicylic acid (SA) and jasmonic acid (JA) signaling in pathogen defenses, but this crosstalk has scarcely been addressed during senescence. In this study, the effect of SA application on methyl jasmonate (MeJA)-induced leaf senescence was assessed. We found that low concentrations of SA (1-50 μM) played a delayed role against the senescence promoted by MeJA. Furthermore, low concentrations of SA enhanced plant antioxidant defenses and restricted reactive oxygen species (ROS) accumulation in MeJA-treated leaves. When applied simultaneously with MeJA, low concentrations of SA triggered a nitric oxide (NO) burst, and the elevated NO levels were linked to the nitric oxide associated 1 (NOA1)-dependent pathway via nitric oxide synthase (NOS) activity. The ability of SA to up-regulate plant antioxidant defenses, reduce ROS accumulation, and suppress leaf senescence was lost in NO-deficient Atnoa1 plants. In a converse manner, exogenous addition of NO donors increased the plant antioxidant capacity and lowered the ROS levels in MeJA-treated leaves. Taken together, the results indicate that SA at low concentrations counteracts MeJA-induced leaf senescence through NOA1-dependent NO signaling and strengthening of the antioxidant defense. PMID:27440938

  6. Human ApoD, an apolipoprotein up-regulated in neurodegenerative diseases, extends lifespan and increases stress resistance in Drosophila

    PubMed Central

    Muffat, Julien; Walker, David W.; Benzer, Seymour

    2008-01-01

    Apolipoprotein D (ApoD) expression increases in several neurological disorders and in spinal cord injury. We provide a report of a physiological role for human ApoD (hApoD): Flies overexpressing hApoD are long-lived and protected against stress conditions associated with aging and neurodegeneration, including hyperoxia, dietary paraquat, and heat stress. We show that the fly ortholog, Glial Lazarillo, is strongly up-regulated in response to these extrinsic stresses and also can protect in vitro-cultured cells in situations modeling Alzheimer's disease (AD) and Parkinson's disease (PD). In adult flies, hApoD overexpression reduces age-associated lipid peroxide accumulation, suggesting a proximal mechanism of action. Similar data obtained in the mouse [Ganfornina, M.D., et al., (2008) Apolipoprotein D is involved in the mechanisms regulating protection from oxidative stress. Aging Cell 10.1111/j.1474-9726.2008.00395.] as well as in plants (Charron et al., personal communication) suggest that ApoD and its orthologs play an evolutionarily conserved role in response to stress, possibly managing or preventing lipid peroxidation. PMID:18458334

  7. Human ApoD, an apolipoprotein up-regulated in neurodegenerative diseases, extends lifespan and increases stress resistance in Drosophila.

    PubMed

    Muffat, Julien; Walker, David W; Benzer, Seymour

    2008-05-13

    Apolipoprotein D (ApoD) expression increases in several neurological disorders and in spinal cord injury. We provide a report of a physiological role for human ApoD (hApoD): Flies overexpressing hApoD are long-lived and protected against stress conditions associated with aging and neurodegeneration, including hyperoxia, dietary paraquat, and heat stress. We show that the fly ortholog, Glial Lazarillo, is strongly up-regulated in response to these extrinsic stresses and also can protect in vitro-cultured cells in situations modeling Alzheimer's disease (AD) and Parkinson's disease (PD). In adult flies, hApoD overexpression reduces age-associated lipid peroxide accumulation, suggesting a proximal mechanism of action. Similar data obtained in the mouse [Ganfornina, M.D., et al., (2008) Apolipoprotein D is involved in the mechanisms regulating protection from oxidative stress. Aging Cell 10.1111/j.1474-9726.2008.00395.] as well as in plants (Charron et al., personal communication) suggest that ApoD and its orthologs play an evolutionarily conserved role in response to stress, possibly managing or preventing lipid peroxidation.

  8. The long noncoding RNA MALAT1 promotes tumor-driven angiogenesis by up-regulating pro-angiogenic gene expression.

    PubMed

    Tee, Andrew E; Liu, Bing; Song, Renhua; Li, Jinyan; Pasquier, Eddy; Cheung, Belamy B; Jiang, Cizhong; Marshall, Glenn M; Haber, Michelle; Norris, Murray D; Fletcher, Jamie I; Dinger, Marcel E; Liu, Tao

    2016-02-23

    Neuroblastoma is the most common solid tumor during early childhood. One of the key features of neuroblastoma is extensive tumor-driven angiogenesis due to hypoxia. However, the mechanism through which neuroblastoma cells drive angiogenesis is poorly understood. Here we show that the long noncoding RNA MALAT1 was upregulated in human neuroblastoma cell lines under hypoxic conditions. Conditioned media from neuroblastoma cells transfected with small interfering RNAs (siRNA) targeting MALAT1, compared with conditioned media from neuroblastoma cells transfected with control siRNAs, induced significantly less endothelial cell migration, invasion and vasculature formation. Microarray-based differential gene expression analysis showed that one of the genes most significantly down-regulated following MALAT1 suppression in human neuroblastoma cells under hypoxic conditions was fibroblast growth factor 2 (FGF2). RT-PCR and immunoblot analyses confirmed that MALAT1 suppression reduced FGF2 expression, and Enzyme-Linked Immunosorbent Assays revealed that transfection with MALAT1 siRNAs reduced FGF2 protein secretion from neuroblastoma cells. Importantly, addition of recombinant FGF2 protein to the cell culture media reversed the effects of MALAT1 siRNA on vasculature formation. Taken together, our data suggest that up-regulation of MALAT1 expression in human neuroblastoma cells under hypoxic conditions increases FGF2 expression and promotes vasculature formation, and therefore plays an important role in tumor-driven angiogenesis.

  9. Farnesoid X receptor up-regulates expression of Lipid transfer inhibitor protein in liver cells and mice

    SciTech Connect

    Li, Liangpeng; Liu, Hong; Peng, Jiahe; Wang, Yongchao; Zhang, Yan; Dong, Jinyu; Liu, Xiaohua; Guo, Dongmei; Jiang, Yu

    2013-11-29

    Highlights: •FXR up-regulates apoF. •It binds to ER1 element. •It activates apoF gene promoter. -- Abstract: Apolipoprotein F is a component protein mainly secreted by liver and resides on several lipoprotein classes. It can inhibit lipids transfer between different lipoproteins. FXR is a member of the nuclear receptor superfamily which is also highly expressed in the liver. It modulates bile acids synthesis and lipids metabolism by transcriptional regulation. We aimed to determine whether apoF can be regulated by FXR. The FXR agonist Chenodeoxycholic acid (CDCA) and GW4064 both can activate the expression of apoF in liver cell lines and in C57/BL6 mouse liver. This is dependent on the binding of FXR to the FXR element ER1 (−2904 to −2892 bp) in the apoF gene promoter. Taken together, we have identified apoF as likely another target gene of FXR.

  10. Prolonged Starvation Causes Up-Regulation of AQP1 in Adipose Tissue Capillaries of AQP7 Knock-Out Mice

    PubMed Central

    Skowronski, Mariusz T.; Skowronska, Agnieszka; Rojek, Aleksandra; Oklinski, Michal K.; Nielsen, Søren

    2016-01-01

    Aquaporins (AQPs) are membrane proteins involved in the regulation of cellular transport and the balance of water and glycerol and cell volume in the white adipose tissue (WAT). In our previous study, we found the co-expression of the AQP1 water channel and AQP7 in the mouse WAT. In our present study, we aimed to find out whether prolonged starvation influences the AQP1 expression of AQP7 knock-out mice (AQP7 KO) in the WAT. To resolve this hypothesis, immunoperoxidase, immunoblot and immunogold microscopy were used. AQP1 expression was found with the use of immunohistochemistry and was confirmed by immunogold microscopy in the vessels of mouse WAT of all studied groups. Semi-quantitative immunoblot and quantitative immunogold microscopy showed a significant increase (by 2.5- to 3-fold) in the abundance of AQP1 protein expression in WAT in the 72 h starved AQP7 KO mice as compared to AQP7+/+ (p < 0.05) and AQP7−/− (p < 0.01) controls, respectively. In conclusion, the AQP1 water channel located in the vessels of WAT is up-regulated in response to prolonged starvation in the WAT of AQP7 KO mice. The present data suggest that an interaction of different AQP isoforms is required for maintaining proper water homeostasis within the mice WAT. PMID:27455244

  11. Up-Regulated Expression of AOS-LOXa and Increased Eicosanoid Synthesis in Response to Coral Wounding

    PubMed Central

    Lõhelaid, Helike; Teder, Tarvi; Tõldsepp, Kadri; Ekins, Merrick; Samel, Nigulas

    2014-01-01

    In octocorals, a catalase–like allene oxide synthase (AOS) and an 8R-lipoxygenase (LOX) gene are fused together encoding for a single AOS-LOX fusion protein. Although the AOS-LOX pathway is central to the arachidonate metabolism in corals, its biological function in coral homeostasis is unclear. Using an acute incision wound model in the soft coral Capnella imbricata, we here test whether LOX pathway, similar to its role in plants, can contribute to the coral damage response and regeneration. Analysis of metabolites formed from exogenous arachidonate before and after fixed time intervals following wounding indicated a significant increase in AOS-LOX activity in response to mechanical injury. Two AOS-LOX isoforms, AOS-LOXa and AOS-LOXb, were cloned and expressed in bacterial expression system as active fusion proteins. Transcription levels of corresponding genes were measured in normal and stressed coral by qPCR. After wounding, AOS-LOXa was markedly up-regulated in both, the tissue adjacent to the incision and distal parts of a coral colony (with the maximum reached at 1 h and 6 h post wounding, respectively), while AOS-LOXb was stable. According to mRNA expression analysis, combined with detection of eicosanoid product formation for the first time, the AOS-LOX was identified as an early stress response gene which is induced by mechanical injury in coral. PMID:24551239

  12. Up-regulation of BRAF activated non-coding RNA is associated with radiation therapy for lung cancer.

    PubMed

    Chen, Jian-xiang; Chen, Ming; Zheng, Yuan-da; Wang, Sheng-ye; Shen, Zhu-ping

    2015-04-01

    Radiation therapy has become more effective in treating primary tumors, such as lung cancer. Recent evidence suggested that BRAF activated non-coding RNAs (BANCR) play a critical role in cellular processes and are found to be dysregulated in a variety of cancers. The clinical significance of BANCR in radiation therapy, and its molecular mechanisms controlling tumor growth are unclear. In the present study, C57BL/6 mice were inoculated Lewis lung cancer cells and exposed to radiation therapy, then BANCR expression was analyzed using qPCR. Chromatin immunoprecipitation and western blot were performed to calculate the enrichment of histone acetylation and HDAC3 protein levels in Lewis lung cancer cells, respectively. MTT assay was used to evaluate the effects of BANCR on Lewis lung cancer cell viability. Finally, we found that BANCR expression was significantly increased in C57BL/6 mice receiving radiation therapy (P<0.05) compared with control group. Additionally, knockdown of BANCR expression was associated with larger tumor size in C57BL/6 mice inoculated Lewis lung cancer cells. Histone deacetylation was observed to involve in the regulation of BANCR in Lewis lung cancer cells. Moreover, over expression HDAC3 reversed the effect of rays on BANCR expression. MTT assay showed that knockdown of BANCR expression promoted cell viability surviving from radiation. In conclusion, these findings indicated that radiation therapy was an effective treatment for lung cancer, and it may exert function through up-regulation BANCR expression.

  13. Up-regulation of the clusterin gene after proteotoxic stress: implication of HSF1–HSF2 heterocomplexes

    PubMed Central

    Loison, Fabien; Debure, Laure; Nizard, Philippe; le Goff, Pascale; Michel, Denis; le Dréan, Yves

    2005-01-01

    Clusterin is a secreted protein chaperone up-regulated in several pathologies, including cancer and neurodegenerative diseases. The present study shows that accumulation of aberrant proteins, caused by the proteasome inhibitor MG132 or the incorporation of the amino acid analogue AZC (L-azetidine-2-carboxylic acid), increased both clusterin protein and mRNA levels in the human glial cell line U-251 MG. Consistently, MG132 treatment was capable of stimulating a 1.3 kb clusterin gene promoter. Promoter deletion and mutation studies revealed a critical MG132-responsive region between −218 and −106 bp, which contains a particular heat-shock element, named CLE for ‘clusterin element’. Gel mobility-shift assays demonstrated that MG132 and AZC treatments induced the formation of a protein complex that bound to CLE. As shown by supershift and chromatin-immunoprecipitation experiments, CLE is bound by HSF1 (heat-shock factor 1) and HSF2 upon proteasome inhibition. Furthermore, co-immunoprecipitation assays indicated that these two transcription factors interact. Gel-filtration analyses revealed that the HSF1–HSF2 heterocomplexes bound to CLE after proteasome inhibition have the same apparent mass as HSF1 homotrimers after heat shock, suggesting that HSF1 and HSF2 could heterotrimerize. Therefore these studies indicate that the clusterin is a good candidate to be part of a cellular defence mechanism against neurodegenerative diseases associated with misfolded protein accumulation or decrease in proteasome activity. PMID:16336210

  14. HSP70 increases extracellular matrix production by human vascular smooth muscle through TGF-β1 up-regulation.

    PubMed

    González-Ramos, Marta; Calleros, Laura; López-Ongil, Susana; Raoch, Viviana; Griera, Mercedes; Rodríguez-Puyol, Manuel; de Frutos, Sergio; Rodríguez-Puyol, Diego

    2013-02-01

    The circulating levels of heat shock proteins (HSP) are increased in cardiovascular diseases; however, the implication of this for the fibrotic process typical of such diseases remains unclear. HSP70 can interact with the vascular smooth muscle cells (SMC), the major producer of extracellular matrix (ECM) proteins, through the Toll-like receptors 4 (TLR4). The transforming growth factor type-β1 (TGF-β1) is a well known vascular pro-fibrotic cytokine that is regulated in part by AP-1-dependent transcriptional mechanisms. We hypothesized that extracellular HSP70 could interact with SMCs, inducing TGF-β1 synthesis and subsequent changes in the vascular ECM. We demonstrate that extracellular HSP70 binds to human aorta SMC TLR4, which up-regulates the AP-1-dependent transcriptional activity of the TGF-β1 promoter. This is achieved through the mitogen activated protein kinases JNK and ERK, as demonstrated by the use of specific blockers and the knockdown of TLR4 with specific small interfering RNAs. The TGF-β1 upregulation increase the expression of the ECM proteins type I collagen and fibronectin. This novel observation may elucidate the mechanisms by which HSP70 contributes in the inflammation and fibrosis present in atherosclerosis and other fibrosis-related diseases. PMID:23084979

  15. Characterization of a RacGTPase up-regulated in the large yellow croaker Pseudosciaena crocea immunity.

    PubMed

    Han, Fang; Wang, Xiaoqing; Yang, Qilian; Cai, Mingyi; Wang, Zhi Yong

    2011-02-01

    The Rac proteins are members of the Rho family of small G proteins and are implicated in the regulation of several pathways, including those leading to cytoskeleton reorganization, gene expression, cell proliferation, cell adhesion and cell migration and survival. In this investigation, a Rac gene (named as LycRac gene) was obtained from the large yellow croaker and it was expressed in Escherichia coli and purified. Subsequently the specific antibody was raised using the purified fusion protein (GST-LycRac). Moreover, the GTP-binding assay showed that the LycRac protein had GTP-binding activity. The LycRac gene was ubiquitously transcribed and expressed in 9 tissues. Quantitative real-time RT-PCR and Western blot analysis revealed the highest expression in gill and the weakest expression in spleen. Time-course analysis revealed that LycRac expression was obviously up-regulated in blood, spleen and liver after immunization with polyinosinic polycytidynic acid (poly I:C), formalin-inactive Gram-negative bacterium Vibrio parahemolyticus and bacterial lipopolysaccharides (LPS). These results suggested that LycRac protein might play an important role in the immune response against microorganisms in large yellow croaker. PMID:21130170

  16. Zirconium Ions Up-Regulate the BMP/SMAD Signaling Pathway and Promote the Proliferation and Differentiation of Human Osteoblasts

    PubMed Central

    Chen, Yongjuan; Roohani-Esfahani, Seyed-Iman; Lu, ZuFu; Zreiqat, Hala; Dunstan, Colin R.

    2015-01-01

    Zirconium (Zr) is an element commonly used in dental and orthopedic implants either as zirconia (ZrO2) or in metal alloys. It can also be incorporated into calcium silicate-based ceramics. However, the effects of in vitro culture of human osteoblasts (HOBs) with soluble ionic forms of Zr have not been determined. In this study, primary culture of human osteoblasts was conducted in the presence of medium containing either ZrCl4 or Zirconium (IV) oxynitrate (ZrO(NO3)2) at concentrations of 0, 5, 50 and 500 µM, and osteoblast proliferation, differentiation and calcium deposition were assessed. Incubation of human osteoblast cultures with Zr ions increased the proliferation of human osteoblasts and also gene expression of genetic markers of osteoblast differentiation. In 21 and 28 day cultures, Zr ions at concentrations of 50 and 500 µM increased the deposition of calcium phosphate. In addition, the gene expression of BMP2 and BMP receptors was increased in response to culture with Zr ions and this was associated with increased phosphorylation of SMAD1/5. Moreover, Noggin suppressed osteogenic gene expression in HOBs co-treated with Zr ions. In conclusion, Zr ions appear able to induce both the proliferation and the differentiation of primary human osteoblasts. This is associated with up-regulation of BMP2 expression and activation of BMP signaling suggesting this action is, at least in part, mediated by BMP signaling. PMID:25602473

  17. Up-regulation of CD44 in the development of metastasis, recurrence and drug resistance of ovarian cancer.

    PubMed

    Gao, Yan; Foster, Rosemary; Yang, Xiaoqian; Feng, Yong; Shen, Jacson K; Mankin, Henry J; Hornicek, Francis J; Amiji, Mansoor M; Duan, Zhenfeng

    2015-04-20

    The clinical significance of Cluster of Differentiation 44 (CD44) remains controversial in human ovarian cancer. The aim of this study is to evaluate the clinical significance of CD44 expression by using a unique tissue microarray, and then to determine the biological functions of CD44 in ovarian cancer. In this study, a unique ovarian cancer tissue microarray (TMA) was constructed with paired primary, metastatic, and recurrent tumor tissues from 26 individual patients. CD44 expression in TMA was assessed by immunohistochemistry. Both the metastatic and recurrent ovarian cancer tissues expressed higher level of CD44 than the patient-matched primary tumor. A significant association has been shown between CD44 expression and both the disease free survival and overall survival. A strong increase of CD44 was found in the tumor recurrence of mouse model. Finally, when CD44 was knocked down, proliferation, migration/invasion activity, and spheroid formation were significantly suppressed, while drug sensitivity was enhanced. Thus, up-regulation of CD44 represents a crucial event in the development of metastasis, recurrence, and drug resistance to current treatments in ovarian cancer. Developing strategies to target CD44 may prevent metastasis, recurrence, and drug resistance in ovarian cancer.

  18. Carboplatin-docetaxel-induced activity against ovarian cancer is dependent on up-regulated lncRNA PVT1.

    PubMed

    Liu, Enling; Liu, Zheng; Zhou, Yuxiu

    2015-01-01

    Ovarian cancer is the fourth most ordinary cause of cancer-related deaths in women. In recent, combination chemotherapy with carboplatin and docetaxel was developed as first-line drug to treat ovarian carcinoma. However, the detailed molecular mechanism, which accounts for the cells to apoptosis induced by administration of carboplatin and docetaxel, was unrecognized. In present study, we provide the mechanistic link between mixture of carboplatin plus docetaxel and its anticancer activity. Primarily, a majority of 30 cancer-related long non-coding RNA (lncRNA) showed differential alteration in carboplatin-docetaxel-treated 3AO cells. Among six up-regulating lncRNAs, we screened out carboplatin-docetaxel-induced lncRNA PVT1 which may be a central downstream target of carboplatin plus docetaxel because expression of PVT1 positively correlates with anticancer action of carboplatin plus docetaxel. Besides, p53 and tissue inhibitor of matrix metalloproteinases-1 (TIMP1) were mediated by lncRNA PVT1, which may explain partially the anticancer activity of lncRNA PVT1. Collectively, we have identified a potential mechanism by which PVT1 regulated by carboplatin plus docetaxel contributes to the carboplatin-docetaxel-induced anticancer action in ovarian cancer. These discoveries also give proof of the potential of PVT1 as significant downstream targets for therapeutic intervention in ovarian cancer.

  19. Transient up-regulation of a novel member of Spot 14 family in androgen-stimulated rat prostate.

    PubMed

    Nishi, Nozomu; Shoji, Hiroki; Miyanaka, Hiroshi; Nakamura, Takanori

    2008-01-01

    The rat prostate is dependent on androgen for growth and differentiation. In an effort to find novel genes involved in androgen-induced growth of the rat prostate, we carried out PCR-based subtractive hybridization and identified several factors that were transiently up-regulated after androgen stimulation in castrated rat prostate. Among them, a novel member of the Spot 14 family has been identified. This protein (Spot 14-like androgen-inducible protein, SLAP) exhibited the highest homology (about 50%) with zebrafish gastrulation-specific G12 protein and about 30% homology with rat Spot 14. The SLAP mRNA level decreased following castration and transiently increased after testosterone replacement, attaining a peak 3 days after the treatment. The change in the protein level was similar to that in mRNA except that it was low in both untreated and castrated rat prostate tissue. In normal adult rats, SLAP was expressed at relatively high levels in the lung, stomach and liver. Unlike the prostate, SLAP expression in the lung was not affected by the androgen status. Like other members of the Spot 14 family, SLAP has a leucine-zipper motif in its C-terminal region, making it possible to form a stable homodimer. Though the physiological role of SLAP remains to be clarified, the current results suggest that SLAP expression is associated with some growth-related processes in regrowing rat prostate.

  20. Azelastine hydrochloride (Azeptin) inhibits peplomycin (PLM)-induced pulmonary fibrosis by contradicting the up-regulation of signal transduction.

    PubMed

    Yoneda, K; Yamamoto, T; Ueta, E; Osaki, T

    1997-10-01

    Inhibition of peplomycin (PLM)-induced pulmonary fibrosis by azelastine hydrochloride (Azeptin) was examined using ICR mice, and the effects of both drugs on signal transduction were investigated. Microscopically, Azeptin (a total of 56 mg/kg for 28 days) suppressed pulmonary fibrosis in mice which received an i.p. injection of a total of 60 or 75 mg/kg PLM. In parallel with the microscopic findings, smaller amounts of collagen were synthesized in the lungs of Azeptin-injected mice. PLM enhanced the expression of interleukin-1 beta- and transforming growth factor-beta-mRNA in lungs. In contrast, Azeptin suppressed the expression. Compatible with these in vivo results, Azeptin and PLM contradictively regulated protein tyrosine phosphorylation and c-myc mRNA expression in human gingival and mouse pulmonary fibroblasts. In addition, NF-kappa B was activated by fibroblast treatment with 5 micrograms/ml PLM for 1 h, but intranuclear NF-kappa B was decreased by cell treatment with 10(-5) M Azeptin. From these results, it is concluded that Azeptin inhibits PLM-induced pulmonary fibrosis by antagonizing the up-regulation of signal transduction.

  1. Folic acid protects against arsenic-mediated embryo toxicity by up-regulating the expression of Dvr1

    PubMed Central

    Ma, Yan; Zhang, Chen; Gao, Xiao-Bo; Luo, Hai-Yan; Chen, Yang; Li, Hui-hua; Ma, Xu; Lu, Cai-Ling

    2015-01-01

    As a nutritional factor, folic acid can prevent cardiac and neural defects during embryo development. Our previous study showed that arsenic impairs embryo development by down-regulating Dvr1/GDF1 expression in zebrafish. Here, we investigated whether folic acid could protect against arsenic-mediated embryo toxicity. We found that folic acid supplementation increases hatching and survival rates, decreases malformation rate and ameliorates abnormal cardiac and neural development of zebrafish embryos exposed to arsenite. Both real-time PCR analysis and whole in-mount hybridization showed that folic acid significantly rescued the decrease in Dvr1 expression caused by arsenite. Subsequently, our data demonstrated that arsenite significantly decreased cell viability and GDF1 mRNA and protein levels in HEK293ET cells, while folic acid reversed these effects. Folic acid attenuated the increase in subcellular reactive oxygen species (ROS) levels and oxidative adaptor p66Shc protein expression in parallel with the changes in GDF1 expression and cell viability. P66Shc knockdown significantly inhibited the production of ROS and the down-regulation of GDF1 induced by arsenite. Our data demonstrated that folic acid supplementation protected against arsenic-mediated embryo toxicity by up-regulating the expression of Dvr1/GDF1, and folic acid enhanced the expression of GDF1 by decreasing p66Shc expression and subcellular ROS levels. PMID:26537450

  2. Resveratrol suppresses NTHi-induced inflammation via up-regulation of the negative regulator MyD88 short

    PubMed Central

    Andrews, Carla S.; Matsuyama, Shingo; Lee, Byung-Cheol; Li, Jian-Dong

    2016-01-01

    Upper respiratory tract inflammatory diseases such as asthma and chronic obstructive pulmonary diseases (COPD) affect more than one-half billion people globally and are characterized by chronic inflammation that is often exacerbated by respiratory pathogens such as nontypeable Haemophilus influenzae (NTHi). The increasing numbers of antibiotic-resistant bacterial strains and the limited success of currently available pharmaceuticals used to manage the symptoms of these diseases present an urgent need for the development of novel anti-inflammatory therapeutic agents. Resveratrol has long been thought as an interesting therapeutic agent for various diseases including inflammatory diseases. However, the molecular mechanisms underlying its anti-inflammatory properties remain largely unknown. Here we show for the first time that resveratrol decreases expression of pro-inflammatory mediators in airway epithelial cells and in the lung of mice by enhancing NTHi-induced MyD88 short, a negative regulator of inflammation, via inhibition of ERK1/2 activation. Furthermore, resveratrol inhibits NTHi-induced ERK1/2 phosphorylation by increasing MKP-1 expression via a cAMP-PKA-dependent signaling pathway. Finally, we show that resveratrol has anti-inflammatory effects post NTHi infection, thereby demonstrating its therapeutic potential. Together these data reveal a novel mechanism by which resveratrol alleviates NTHi-induced inflammation in airway disease by up-regulating the negative regulator of inflammation MyD88s. PMID:27677845

  3. Resveratrol up-regulates AMPA receptor expression via AMP-activated protein kinase-mediated protein translation.

    PubMed

    Wang, Guan; Amato, Stephen; Gilbert, James; Man, Heng-Ye

    2015-08-01

    Resveratrol is a phytoalexin that confers overall health benefits including positive regulation in brain function such as learning and cognition. However, whether and how resveratrol affects synaptic activity remains largely unknown. α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) are glutamatergic receptors that mediate the majority of fast excitatory transmission and synaptic plasticity, and thus play a critical role in higher brain functions, including learning and memory. We find that in rat primary neurons, resveratrol can rapidly increase AMPAR protein level, AMPAR synaptic accumulation and the strength of excitatory synaptic transmission. The resveratrol effect on AMPAR protein expression is independent of sirtuin 1 (SIRT1), the conventional downstream target of resveratrol, but rather is mediated by AMP-activated protein kinase (AMPK) and subsequent downstream phosphoinositide 3-kinase (PI3K)/Akt signaling. Application of the AMPK specific activator 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) mimics the effects of resveratrol on both signaling and AMPAR expression. The resveratrol-induced increase in AMPAR expression results from elevated protein synthesis via regulation of the eukaryotic initiation factor (eIF) 4E/4G complex. Disruption of the translation initiation complex completely blocks resveratrol-dependent AMPAR up-regulation. These findings indicate that resveratrol may regulate brain function through facilitation of AMPAR biogenesis and synaptic transmission.

  4. Folic acid-conjugated fluorescent polymer for up-regulation folate receptor expression study via targeted imaging of tumor cells.

    PubMed

    Qiao, Juan; Dong, Ping; Mu, Xiaoyu; Qi, Li; Xiao, Ran

    2016-04-15

    Thoroughly investigation of folate receptor (FR) expression related to targeting drug delivery in tumor cells has been intensively pursued in recent years. Herein, a simple and versatile strategy for determination of FR expression based on targeted imaging of tumor cells with fluorescent nano-conjugates was developed. The fluorescent nano-conjugates were composed of poly 2-vinyl-4,4-dimethyl azlactone (PVDMA) as the linker, folic acid as the targeting unit and amino-Rhodamine B as the fluorescent ligand. Owing to possessing dimethyl azlactone groups in polymer framework, PVDMA could easily reacted with amines or alcohols, and form water soluble materials. Fluorescent imaging studies indicated that the prepared nano-conjugates could specifically target tumor cells and monitor the over expressing of FR. Moreover, the FR expression up-regulation in HeLa cells through medicines regulation has been further explored. This new protocol opens an effective way through synthesis and design of novel fluorescent nano-conjugates for FR expression investigation in tumor cells via targeted imaging, showing great potential in drug delivery mechanism study and cancer therapy.

  5. The long noncoding RNA MALAT1 promotes tumor-driven angiogenesis by up-regulating pro-angiogenic gene expression.

    PubMed

    Tee, Andrew E; Liu, Bing; Song, Renhua; Li, Jinyan; Pasquier, Eddy; Cheung, Belamy B; Jiang, Cizhong; Marshall, Glenn M; Haber, Michelle; Norris, Murray D; Fletcher, Jamie I; Dinger, Marcel E; Liu, Tao

    2016-02-23

    Neuroblastoma is the most common solid tumor during early childhood. One of the key features of neuroblastoma is extensive tumor-driven angiogenesis due to hypoxia. However, the mechanism through which neuroblastoma cells drive angiogenesis is poorly understood. Here we show that the long noncoding RNA MALAT1 was upregulated in human neuroblastoma cell lines under hypoxic conditions. Conditioned media from neuroblastoma cells transfected with small interfering RNAs (siRNA) targeting MALAT1, compared with conditioned media from neuroblastoma cells transfected with control siRNAs, induced significantly less endothelial cell migration, invasion and vasculature formation. Microarray-based differential gene expression analysis showed that one of the genes most significantly down-regulated following MALAT1 suppression in human neuroblastoma cells under hypoxic conditions was fibroblast growth factor 2 (FGF2). RT-PCR and immunoblot analyses confirmed that MALAT1 suppression reduced FGF2 expression, and Enzyme-Linked Immunosorbent Assays revealed that transfection with MALAT1 siRNAs reduced FGF2 protein secretion from neuroblastoma cells. Importantly, addition of recombinant FGF2 protein to the cell culture media reversed the effects of MALAT1 siRNA on vasculature formation. Taken together, our data suggest that up-regulation of MALAT1 expression in human neuroblastoma cells under hypoxic conditions increases FGF2 expression and promotes vasculature formation, and therefore plays an important role in tumor-driven angiogenesis. PMID:26848616

  6. Zirconium ions up-regulate the BMP/SMAD signaling pathway and promote the proliferation and differentiation of human osteoblasts.

    PubMed

    Chen, Yongjuan; Roohani-Esfahani, Seyed-Iman; Lu, ZuFu; Zreiqat, Hala; Dunstan, Colin R

    2015-01-01

    Zirconium (Zr) is an element commonly used in dental and orthopedic implants either as zirconia (ZrO2) or in metal alloys. It can also be incorporated into calcium silicate-based ceramics. However, the effects of in vitro culture of human osteoblasts (HOBs) with soluble ionic forms of Zr have not been determined. In this study, primary culture of human osteoblasts was conducted in the presence of medium containing either ZrCl4 or Zirconium (IV) oxynitrate (ZrO(NO3)2) at concentrations of 0, 5, 50 and 500 µM, and osteoblast proliferation, differentiation and calcium deposition were assessed. Incubation of human osteoblast cultures with Zr ions increased the proliferation of human osteoblasts and also gene expression of genetic markers of osteoblast differentiation. In 21 and 28 day cultures, Zr ions at concentrations of 50 and 500 µM increased the deposition of calcium phosphate. In addition, the gene expression of BMP2 and BMP receptors was increased in response to culture with Zr ions and this was associated with increased phosphorylation of SMAD1/5. Moreover, Noggin suppressed osteogenic gene expression in HOBs co-treated with Zr ions. In conclusion, Zr ions appear able to induce both the proliferation and the differentiation of primary human osteoblasts. This is associated with up-regulation of BMP2 expression and activation of BMP signaling suggesting this action is, at least in part, mediated by BMP signaling.

  7. Interleukin-6 enhances cancer stemness and promotes metastasis of hepatocellular carcinoma via up-regulating osteopontin expression

    PubMed Central

    Wang, Chao-Qun; Sun, Hao-Ting; Gao, Xiao-Mei; Ren, Ning; Sheng, Yuan-Yuan; Wang, Zheng; Zheng, Yan; Wei, Jin-Wang; Zhang, Kai-Li; Yu, Xin-Xin; Zhu, Yin; Luo, Qin; Yang, Lu-Yu; Dong, Qiong-Zhu; Qin, Lun-Xiu

    2016-01-01

    Interleukin-6 (IL-6), one of the most important inflammatory cytokines, plays a pivotal role in metastasis and stemness of solid tumors. However, the underlying mechanisms of IL-6 in HCC metastasis remain unclear. In the present study, we demonstrated that stemness and metastatic potential of HCC cells were significantly enhanced after IL-6 stimulation. IL-6 could induce expression of osteopontin (OPN), along with other stemness-related genes, including HIF1α, BMI1, and HEY1. Block of OPN induction could significantly abrogate the effect of IL-6 on stemness and metastasis of HCC cells. Furthermore, IL-6 level was positively correlated with OPN in HCC. Patients with high plasma IL-6 or OPN level had poorer prognosis. In multivariate analysis, IL-6 and OPN were demonstrated to be independent prognostic indicators for HCC patients, and their combination had a better prognostic performance than IL-6 or OPN alone. Collectively, our findings indicate that IL-6 could enhance stemness and promote metastasis of HCC via up-regulating OPN expression, which can be a potential therapeutic target for combating HCC metastasis, and the combination of IL-6 and OPN serves as a promising prognostic predictor for HCC. PMID:27725896

  8. Interplay between up-regulation of cytochrome-c-oxidase and hemoglobin oxygenation induced by near-infrared laser

    PubMed Central

    Wang, Xinlong; Tian, Fenghua; Soni, Sagar S.; Gonzalez-Lima, F.; Liu, Hanli

    2016-01-01

    Photobiomodulation, also known as low-level laser/light therapy (LLLT), refers to the use of red-to-near-infrared light to stimulate cellular functions for physiological or clinical benefits. The mechanism of LLLT is assumed to rely on photon absorption by cytochrome c oxidase (CCO), the terminal enzyme in the mitochondrial respiratory chain that catalyzes the reduction of oxygen for energy metabolism. In this study, we used broadband near-infrared spectroscopy (NIRS) to measure the LLLT-induced changes in CCO and hemoglobin concentrations in human forearms in vivo. Eleven healthy participants were administered with 1064-nm laser and placebo treatments on their right forearms. The spectroscopic data were analyzed and fitted with wavelength-dependent, modified Beer-Lambert Law. We found that LLLT induced significant increases of CCO concentration (Δ[CCO]) and oxygenated hemoglobin concentration (Δ[HbO]) on the treated site as the laser energy dose accumulated over time. A strong linear interplay between Δ[CCO] and Δ[HbO] was observed for the first time during LLLT, indicating a hemodynamic response of oxygen supply and blood volume closely coupled to the up-regulation of CCO induced by photobiomodulation. These results demonstrate the tremendous potential of broadband NIRS as a non-invasive, in vivo means to study mechanisms of photobiomodulation and perform treatment evaluations of LLLT. PMID:27484673

  9. Up-regulation of interleukin-22 mediates liver fibrosis via activating hepatic stellate cells in patients with hepatitis C.

    PubMed

    Wu, Li-Yuan; Liu, Shuhong; Liu, Yuan; Guo, Chaonan; Li, Hanwei; Li, Wenshu; Jin, Xueyuan; Zhang, Keming; Zhao, Ping; Wei, Lai; Zhao, Jingmin

    2015-05-01

    Interleukin-22 (IL-22) is known to play a critical role in liver immunity. However, the role of IL-22 in HCV-associated liver fibrosis is poorly understood. In this study, patients with HCV infection disclosed significant increases in peripheral numbers of IL-22-producing cells as well as in IL-22 plasma levels. In the liver, the increased intrahepatic IL-22(+) cells were positively correlated with fibrotic staging scores and clinical progression from CHC to cirrhosis. Moreover, the majority of IL-22(+) cells were located in fibrotic areas in the liver of patients with cirrhosis and co-localized with α-smooth muscle actin (α-SMA) positive hepatic stellate cells (HSCs). In vitro, administration of IL-22 was accompanied with inhibited LX-2 cell apoptosis, promoted LX-2 cell proliferation, increased expression of α-SMA, and up-regulated collagen production by LX-2 cells. Collectively, our data provide evidence that IL-22 may contribute to the fibrogenesis of HCV-associated liver fibrosis by activating HSCs.

  10. Induction of release and up-regulated gene expression of interleukin (IL)-8 in A549 cells by serine proteinases

    PubMed Central

    Wang, Haiyan; Zheng, Yanshan; He, Shaoheng

    2006-01-01

    Background Hypersecretion of cytokines and serine proteinases has been observed in asthma. Since protease-activated receptors (PARs) are receptors of several serine proteinases and airway epithelial cells are a major source of cytokines, the influence of serine proteinases and PARs on interleukin (IL)-8 secretion and gene expression in cultured A549 cells was examined. Results A549 cells express all four PARs at both protein and mRNA levels as assessed by flow cytometry, immunofluorescence microscopy and reverse transcription polymerase chain reaction (PCR). Thrombin, tryptase, elastase and trypsin induce a up to 8, 4.3, 4.4 and 5.1 fold increase in IL-8 release from A549 cells, respectively following 16 h incubation period. The thrombin, elastase and trypsin induced secretion of IL-8 can be abolished by their specific inhibitors. Agonist peptides of PAR-1, PAR-2 and PAR-4 stimulate up to 15.6, 6.6 and 3.5 fold increase in IL-8 secretion, respectively. Real time PCR shows that IL-8 mRNA is up-regulated by the serine proteinases tested and by agonist peptides of PAR-1 and PAR-2. Conclusion The proteinases, possibly through activation of PARs can stimulate IL-8 release from A549 cells, suggesting that they are likely to contribute to IL-8 related airway inflammatory disorders in man. PMID:16696869

  11. Ischemia-Reperfusion Injury and Pregnancy Initiate Time-Dependent and Robust Signs of Up-Regulation of Cardiac Progenitor Cells

    PubMed Central

    Genead, Rami; Fischer, Helene; Hussain, Alamdar; Jaksch, Marie; Andersson, Agneta B.; Ljung, Karin; Bulatovic, Ivana; Franco-Cereceda, Anders; Elsheikh, Elzafir; Corbascio, Matthias; Smith, C. I. Edvard; Sylvén, Christer; Grinnemo, Karl-Henrik

    2012-01-01

    To explore how cardiac regeneration and cell turnover adapts to disease, different forms of stress were studied for their effects on the cardiac progenitor cell markers c-Kit and Isl1, the early cardiomyocyte marker Nkx2.5, and mast cells. Adult female rats were examined during pregnancy, after myocardial infarction and ischemia-reperfusion injury with/out insulin like growth factor-1(IGF-1) and hepatocyte growth factor (HGF). Different cardiac sub-domains were analyzed at one and two weeks post-intervention, both at the mRNA and protein levels. While pregnancy and myocardial infarction up-regulated Nkx2.5 and c-Kit (adjusted for mast cell activation), ischemia-reperfusion injury induced the strongest up-regulation which occurred globally throughout the entire heart and not just around the site of injury. This response seems to be partly mediated by increased endogenous production of IGF-1 and HGF. Contrary to c-Kit, Isl1 was not up-regulated by pregnancy or myocardial infarction while ischemia-reperfusion injury induced not a global but a focal up-regulation in the outflow tract and also in the peri-ischemic region, correlating with the up-regulation of endogenous IGF-1. The addition of IGF-1 and HGF did boost the endogenous expression of IGF and HGF correlating to focal up-regulation of Isl1. c-Kit expression was not further influenced by the exogenous growth factors. This indicates that there is a spatial mismatch between on one hand c-Kit and Nkx2.5 expression and on the other hand Isl1 expression. In conclusion, ischemia-reperfusion injury was the strongest stimulus with both global and focal cardiomyocyte progenitor cell marker up-regulations, correlating to the endogenous up-regulation of the growth factors IGF-1 and HGF. Also pregnancy induced a general up-regulation of c-Kit and early Nkx2.5+ cardiomyocytes throughout the heart. Utilization of these pathways could provide new strategies for the treatment of cardiac disease. PMID:22590612

  12. TGF-β1 Up-Regulates Connective Tissue Growth Factor Expression in Human Granulosa Cells through Smad and ERK1/2 Signaling Pathways

    PubMed Central

    Cheng, Jung-Chien; Chang, Hsun-Ming; Fang, Lanlan; Sun, Ying-Pu; Leung, Peter C. K.

    2015-01-01

    Connective tissue growth factor (CTGF), which is also called CCN2, is a secreted matricellular protein. CTGF regulates various important cellular functions by interacting with multiple molecules in the microenvironment. In the ovary, CTGF is mainly expressed in granulosa cells and involved in the regulation of follicular development, ovulation and luteinization. TGF-β1 has been shown to up-regulate CTGF expression in rat and hen granulosa cells. However, the underlying molecular mechanisms of this up-regulation remain undefined. More importantly, whether the stimulatory effect of TGF-β1 on CTGF expression can be observed in human granulosa cells remains unknown. In the present study, our results demonstrated that TGF-β1 treatment up-regulates CTGF expression in both immortalized human granulosa cells and primary human granulosa cells. Using a siRNA-mediated knockdown approach and a pharmacological inhibitor, we demonstrated that the inhibition of Smad2, Smad3 or ERK1/2 attenuates the TGF-β1-induced up-regulation of CTGF. This study provides important insights into the molecular mechanisms that mediate TGF-β1-up-regulated CTGF expression in human granulosa cells. PMID:25955392

  13. A novel class of antihyperlipidemic agents with low density lipoprotein receptor up-regulation via the adaptor protein autosomal recessive hypercholesterolemia.

    PubMed

    Asano, Shigehiro; Ban, Hitoshi; Tsuboya, Norie; Uno, Shinsaku; Kino, Kouichi; Ioriya, Katsuhisa; Kitano, Masafumi; Ueno, Yoshihide

    2010-04-22

    We have previously reported compound 2 as a inhibitor of acyl-coenzyme A:cholesterol O-acyltransferase (ACAT) and up-regulator of the low density lipoprotein receptor (LDL-R) expression. In this study we focused on compound 2, a unique LDL-R up-regulator, and describe the discovery of a novel class of up-regulators of LDL-R. Replacement the methylene urea linker in compound 2 with an acylsulfonamide linker kept a potent LDL-R up-regulatory activity, and subsequent optimization work gave compound 39 as a highly potent LDL-R up-regulator (39; EC(25) = 0.047 microM). Compound 39 showed no ACAT inhibitory activity even at 1 microM. The sodium salts of compound 39 reduced plasma total and LDL cholesterol levels in a dose-dependent manner in an experimental animal model of hyperlipidemia. Moreover, we revealed in this study using RNA interference that autosomal recessive hypercholesterolemia (ARH), an adaptor protein of LDL-R, is essential for compound 39 up-regulation of LDL-R expression. PMID:20356098

  14. TGF-β1 Up-Regulates Connective Tissue Growth Factor Expression in Human Granulosa Cells through Smad and ERK1/2 Signaling Pathways.

    PubMed

    Cheng, Jung-Chien; Chang, Hsun-Ming; Fang, Lanlan; Sun, Ying-Pu; Leung, Peter C K

    2015-01-01

    Connective tissue growth factor (CTGF), which is also called CCN2, is a secreted matricellular protein. CTGF regulates various important cellular functions by interacting with multiple molecules in the microenvironment. In the ovary, CTGF is mainly expressed in granulosa cells and involved in the regulation of follicular development, ovulation and luteinization. TGF-β1 has been shown to up-regulate CTGF expression in rat and hen granulosa cells. However, the underlying molecular mechanisms of this up-regulation remain undefined. More importantly, whether the stimulatory effect of TGF-β1 on CTGF expression can be observed in human granulosa cells remains unknown. In the present study, our results demonstrated that TGF-β1 treatment up-regulates CTGF expression in both immortalized human granulosa cells and primary human granulosa cells. Using a siRNA-mediated knockdown approach and a pharmacological inhibitor, we demonstrated that the inhibition of Smad2, Smad3 or ERK1/2 attenuates the TGF-β1-induced up-regulation of CTGF. This study provides important insights into the molecular mechanisms that mediate TGF-β1-up-regulated CTGF expression in human granulosa cells.

  15. Rac1-dependent transcriptional up-regulation of p27Kip1 by homophilic cell-cell contact in vascular endothelial cells.

    PubMed

    Hirano, Mayumi; Kanaide, Hideo; Hirano, Katsuya

    2007-10-01

    The mechanism for the transcriptional up-regulation of p27Kip1 due to the formation of the cell-cell contact was investigated in vascular endothelial cells. The induction of the cell-cell contact by adding an extra number of endothelial cells activated Rac1, up-regulated p27Kip1 mRNA and protein, and also facilitated the cell cycle arrest. Transduction of the Rac1 inhibitor protein using the cell-penetrating peptide or treatment with a Rac1 inhibitor NSC23766 inhibited the p27Kip1 up-regulation and delayed the cell cycle arrest. Rac1 was therefore suggested to mediate the contact-induced transcriptional up-regulation of p27Kip1. The role of Rac1 in the regulation of the p27Kip1 promoter activity was next examined with a luciferase reporter assay. The promoter activity was increased by inducing the cell-cell contact, which was significantly inhibited by the Rac1 inhibitory protein and NSC23766. The evaluation of various truncated promoter regions determined region -620 to -573 nucleotides from the initiation codon to be responsible for the contact-induced, Rac1-dependent activation of the p27Kip1 promoter. The present study thus demonstrated for the first time that the activation of Rac1 due to the cell-cell contact plays a critical role in the transcriptional up-regulation of p27Kip1 in vascular endothelial cells.

  16. Arabidopsis sodium dependent and independent phenotypes triggered by H+-PPase up-regulation are SOS1 dependent

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The goal of the study and research was to coordinate regulation of transporters at both the plasma membrane and vacuole contribute to plant cell’s ability to adapt to a changing environment and play a key role in the maintenance of the chemiosmotic circuits required for cellular growth. In this stud...

  17. Cyclic up-regulation fluorescence of pyrene excimer for studying polynucleotide kinase activity based on dual amplification.

    PubMed

    Xu, Jing; Gao, Yanfang; Li, Baoxin; Jin, Yan

    2016-06-15

    Due to its important biological and clinical roles of polynucleotide kinase (PNK), accurate monitoring of PNK activity and inhibition is highly desirable. Herein, a homogeneous and sensitive fluorescence assay has been proposed for the detection of PNK activity by integrating target recycling signal amplification of DNA toehold strand displacement reaction (TSDR) with gamma-cyclodextrin (γ-CD) enhancement of pyrene excimer. A label-free hairpin DNA1 (H1) and two singly pyrene-labelled DNA, H2 and H3, are designed. Accompanying the occurrence of the efficient enzyme reactions, namely phosphorylation-actuated λ exonuclease reaction, a single-stranded DNA as a trigger DNA (tDNA) of TSDR can be released from H1. Then, tDNA drives circulatory interactions between H2 and H3 to continuously form H2/H3 duplex, resulting in formation of pyrene excimer and a "turn on" fluorescence signal of pyrene excimer. Furthermore, the fluorescence of pyrene excimer is further amplified by introducing gamma-cyclodextrin (γ-CD), which can regulate the space proximity of two pyrene molecules. Thus, TSDR-induced cyclic formation of pyrene excimer and γ-CD enhancement can specifically up-regulate the fluorescence of pyrene excimer for detection of PNK activity, the detection limit is 9.3 × 10(-5)UmL(-1), which is superior to those of most existing approaches. Moreover, the proposed strategy can also be successfully utilized to study inhibition efficiency of different PNK inhibitors as well. Therefore, a dual amplification approach is provided for nucleic acid phosphorylation related researches. PMID:26807522

  18. Integrin αDβ2, an adhesion receptor up-regulated on macrophage foam cells, exhibits multiligand-binding properties

    PubMed Central

    Yakubenko, Valentin P.; Yadav, Satya P.; Ugarova, Tatiana P.

    2006-01-01

    Integrin αDβ2, the most recently discovered member of the β2 subfamily of integrin adhesion receptors, is up-regulated on macrophage foam cells. Although other members of the subfamily have been subjects of extensive research, the recognition specificity and the molecular basis for αDβ2 ligand binding remain unknown. Based on the high extent of structural homology between αDβ2 and the major myeloid-cell-specific integrin αMβ2 (Mac-1), noted for its capacity to bind multiple ligands, we considered that the 2 integrins have similar recognition specificity. In this study, using recombinant and natural αDβ2-expressing cells, we demonstrate that αDβ2 supports adhesion and migration to many extracellular matrix proteins in a fashion similar to αMβ2. Consistent with these data, the recombinant αDI-domain of the receptor bound selected ligands. The binding was activation-dependent because the αDI-domain with its C-terminal α7 helix truncated, but not the form with the C-terminal part extended, bound ligands. When the αDI-domain segment Lys244-Lys260 (highly homologous to its αMI-domain counterpart Lys245-Arg261 responsible for αMβ2 multiligand-binding properties) was inserted into the mono-specific αLI-domain, the chimeric protein bound many ligands with affinities similar to those of wild-type αDI-domain. These results establish integrin αDβ2 as a multiligand receptor and indicate that the mechanism whereby αDβ2 exhibits broad ligand specificity resembles that used by αMβ2, the most promiscuous member of the integrin family. PMID:16239428

  19. Increased NF-κB signalling up-regulates BACE1 expression and its therapeutic potential in Alzheimer's disease.

    PubMed

    Chen, Chia-Hsiung; Zhou, Weihui; Liu, Shengchun; Deng, Yu; Cai, Fang; Tone, Masahide; Tone, Yukiko; Tong, Yigang; Song, Weihong

    2012-02-01

    Elevated levels of β-site APP cleaving enzyme 1 (BACE1) were found in the brain of some sporadic Alzheimer's disease (AD) patients; however, the underlying mechanism is unknown. BACE1 cleaves β-amyloid precursor protein (APP) to generate amyloid β protein (Aβ), a central component of neuritic plaques in AD brains. Nuclear factor-kappa B (NF-κB) signalling plays an important role in gene regulation and is implicated in inflammation, oxidative stress and apoptosis. In this report we found that both BACE1 and NF-κB p65 levels were significantly increased in the brains of AD patients. Two functional NF-κB-binding elements were identified in the human BACE1 promoter region. We found that NF-κB p65 expression resulted in increased BACE1 promoter activity and BACE1 transcription, while disruption of NF-κB p65 decreased BACE1 gene expression in p65 knockout (RelA-knockout) cells. In addition, NF-κB p65 expression leads to up-regulated β-secretase cleavage and Aβ production, while non-steroidal anti-inflammatory drugs (NSAIDs) inhibited BACE1 transcriptional activation induced by strong NF-κB activator tumour necrosis factor-alpha (TNF-α). Taken together, our results clearly demonstrate that NF-κB signalling facilitates BACE1 gene expression and APP processing, and increased BACE1 expression mediated by NF-κB signalling in the brain could be one of the novel molecular mechanisms underlying the development of AD in some sporadic cases. Furthermore, NSAIDs could block the inflammation-induced BACE1 transcription and Aβ production. Our study suggests that inhibition of NF-κB-mediated BACE1 expression may be a valuable drug target for AD therapy.

  20. Poncirin Induces Apoptosis in AGS Human Gastric Cancer Cells through Extrinsic Apoptotic Pathway by up-Regulation of Fas Ligand

    PubMed Central

    Venkatarame Gowda Saralamma, Venu; Nagappan, Arulkumar; Hong, Gyeong Eun; Lee, Ho Jeong; Yumnam, Silvia; Raha, Suchismita; Heo, Jeong Doo; Lee, Sang Joon; Lee, Won Sup; Kim, Eun Hee; Kim, Gon Sup

    2015-01-01

    Poncirin, a natural bitter flavanone glycoside abundantly present in many species of citrus fruits, has various biological benefits such as anti-oxidant, anti-microbial, anti-inflammatory and anti-cancer activities. The anti-cancer mechanism of Poncirin remains elusive to date. In this study, we investigated the anti-cancer effects of Poncirin in AGS human gastric cancer cells (gastric adenocarcinoma). The results revealed that Poncirin could inhibit the proliferation of AGS cells in a dose-dependent manner. It was observed Poncirin induced accumulation of sub-G1 DNA content, apoptotic cell population, apoptotic bodies, chromatin condensation, and DNA fragmentation in a dose-dependent manner in AGS cells. The expression of Fas Ligand (FasL) protein was up-regulated dose dependently in Poncirin-treated AGS cells Moreover, Poncirin in AGS cells induced activation of Caspase-8 and -3, and subsequent cleavage of poly(ADP-ribose) polymerase (PARP). Inhibitor studies’ results confirm that the induction of caspase-dependent apoptotic cell death in Poncirin-treated AGS cells was led by the Fas death receptor. Interestingly, Poncirin did not show any effect on mitochondrial membrane potential (ΔΨm), pro-apoptotic proteins (Bax and Bak) and anti-apoptotic protein (Bcl-xL) in AGS-treated cells followed by no activation in the mitochondrial apoptotic protein caspase-9. This result suggests that the mitochondrial-mediated pathway is not involved in Poncirin-induced cell death in gastric cancer. These findings suggest that Poncirin has a potential anti-cancer effect via extrinsic pathway-mediated apoptosis, possibly making it a strong therapeutic agent for human gastric cancer. PMID:26393583

  1. Up- regulation of miR-328-3p sensitizes non-small cell lung cancer to radiotherapy

    PubMed Central

    Ma, Wei; Ma, Chao-nan; Zhou, Nan-nan; Li, Xian-dong; Zhang, Yi-jie

    2016-01-01

    MicroRNAs (miRNAs) are believed to be resistant against radiotherapy in certain types of cancers. The aim of our study was to determine the clinical application of miRNAs in non-small cell lung cancer (NSCLC). Sixty NSCLC tissue samples and adjacent histologically normal tissues were obtained for miRNAs microarray analysis and validated by RT-qPCR. Correlation between miRNA expression level and clinicopathological features was evaluated. Our study examined the influence of changed miRNA expression on the damaged DNA and its associated radio sensitivity. Luciferase assay was performed to determine potential effects on the targeted gene. Our study identified fifteen altered miRNAs in which miR-328-3p was down regulated in NSCLC tumour tissue as compared to normal tissues. Down-expression of miR-328-3p was positively associated with an enhanced lymph node metastasis, advanced clinical stage and a shortened survival rate. miR-328-3p expression was decreased in A549 cells compared to other NSCLC cell lines. Up-regulation of miR-328-3p demonstrated a survival inhibition effect in A549 and restored NSCLC cells’ sensitivity to radio therapy. An increased miR-328-3p expression promoted irradiation-induced DNA damage in cells. γ-H2AX was identified as the direct target of miR-328-3p. Over-expressed miR-328-3p can improve the radiosensitvity of cells by altering the DNA damage/repair signalling pathways in NSCLC. PMID:27530148

  2. Lysophosphatidic Acid Up-Regulates Hexokinase II and Glycolysis to Promote Proliferation of Ovarian Cancer Cells1

    PubMed Central

    Mukherjee, Abir; Ma, Yibao; Yuan, Fang; Gong, Yongling; Fang, Zhenyu; Mohamed, Esraa M.; Berrios, Erika; Shao, Huanjie; Fang, Xianjun

    2015-01-01

    Lysophosphatidic acid (LPA), a blood-borne lipid mediator, is present in elevated concentrations in ascites of ovarian cancer patients and other malignant effusions. LPA is a potent mitogen in cancer cells. The mechanism linking LPA signal to cancer cell proliferation is not well understood. Little is known about whether LPA affects glucose metabolism to accommodate rapid proliferation of cancer cells. Here we describe that in ovarian cancer cells, LPA enhances glycolytic rate and lactate efflux. A real time PCR-based miniarray showed that hexokinase II (HK2) was the most dramatically induced glycolytic gene to promote glycolysis in LPA-treated cells. Analysis of the human HK2 gene promoter identified the sterol regulatory element-binding protein as the primary mediator of LPA-induced HK2 transcription. The effects of LPA on HK2 and glycolysis rely on LPA2, an LPA receptor subtype overexpressed in ovarian cancer and many other malignancies. We further examined the general role of growth factor-induced glycolysis in cell proliferation. Like LPA, epidermal growth factor (EGF) elicited robust glycolytic and proliferative responses in ovarian cancer cells. Insulin-like growth factor 1 (IGF-1) and insulin, however, potently stimulated cell proliferation but only modestly induced glycolysis. Consistent with their differential effects on glycolysis, LPA and EGF-dependent cell proliferation was highly sensitive to glycolytic inhibition while the growth-promoting effect of IGF-1 or insulin was more resistant. These results indicate that LPA- and EGF-induced cell proliferation selectively involves up-regulation of HK2 and glycolytic metabolism. The work is the first to implicate LPA signaling in promotion of glucose metabolism in cancer cells. PMID:26476080

  3. Isoflurane Preconditioning Induces Neuroprotection by Up-Regulation of TREK1 in a Rat Model of Spinal Cord Ischemic Injury

    PubMed Central

    Wang, Kun; Kong, Xiangang

    2016-01-01

    This study aimed to explore the neuroprotection and mechanism of isoflurane on rats with spinal cord ischemic injury. Total 40 adult male Sprague-Dawley rats were divided into the four groups (n=10). Group A was sham-operation group; group B was ischemia group; group C was isoflurane preconditioning group; group D was isoflurane preconditioning followed by ischemia treatment group. Then the expressions of TWIK-related K+ channel 1 (TREK1) in the four groups were detected by immunofluorescent assay, real time-polymerase chain reactions (RT-PCR) and western blot. The primary neurons of rats were isolated and cultured under normal and hypoxic conditions. Besides, the neurons under two conditions were transfected with green fluorescent protein (GFP)-TREK1 and lentivirual to overexpress and silence TREK1. Additionally, the neurons were treated with isoflurane or not. Then caspase-3 activity and cell cycle of neurons under normal and hypoxic conditions were detected. Furthermore, nicotinamide adenine dinucleotide hydrate (NADH) was detected using NAD+/NADH quantification colorimetric kit. Results showed that the mRNA and protein expressions of TREK1 increased significantly in group C and D. In neurons, when TREK1 silenced, isoflurane treatment improved the caspase-3 activity. In hypoxic condition, the caspase-3 activity and sub-G1 cell percentage significantly increased, however, when TREK1 overexpressed the caspase-3 activity and sub-G1 cell percentage decreased significantly. Furthermore, both isoflurane treatment and overexpression of TREK1 significantly decreased NADH. In conclusion, isoflurane-induced neuroprotection in spinal cord ischemic injury may be associated with the up-regulation of TREK1. PMID:27469140

  4. Up-regulation of abscisic acid signaling pathway facilitates aphid xylem absorption and osmoregulation under drought stress.

    PubMed

    Guo, Huijuan; Sun, Yucheng; Peng, Xinhong; Wang, Qinyang; Harris, Marvin; Ge, Feng

    2016-02-01

    The activation of the abscisic acid (ABA) signaling pathway reduces water loss from plants challenged by drought stress. The effect of drought-induced ABA signaling on the defense and nutrition allocation of plants is largely unknown. We postulated that these changes can affect herbivorous insects. We studied the effects of drought on different feeding stages of pea aphids in the wild-type A17 of Medicago truncatula and ABA signaling pathway mutant sta-1. We examined the impact of drought on plant water status, induced plant defense signaling via the abscisic acid (ABA), jasmonic acid (JA), and salicylic acid (SA) pathways, and on the host nutritional quality in terms of leaf free amino acid content. During the penetration phase of aphid feeding, drought decreased epidermis/mesophyll resistance but increased mesophyll/phloem resistance of A17 but not sta-1 plants. Quantification of transcripts associated with ABA, JA and SA signaling indicated that the drought-induced up-regulation of ABA signaling decreased the SA-dependent defense but increased the JA-dependent defense in A17 plants. During the phloem-feeding phase, drought had little effect on the amino acid concentrations and the associated aphid phloem-feeding parameters in both plant genotypes. In the xylem absorption stage, drought decreased xylem absorption time of aphids in both genotypes because of decreased water potential. Nevertheless, the activation of the ABA signaling pathway increased water-use efficiency of A17 plants by decreasing the stomatal aperture and transpiration rate. In contrast, the water potential of sta-1 plants (unable to close stomata) was too low to support xylem absorption activity of aphids; the aphids on sta-1 plants had the highest hemolymph osmolarity and lowest abundance under drought conditions. Taken together this study illustrates the significance of cross-talk between biotic-abiotic signaling pathways in plant-aphid interaction, and reveals the mechanisms leading to alter

  5. Recurrent MALAT1-GLI1 oncogenic fusion and GLI1 up-regulation define a subset of plexiform fibromyxoma.

    PubMed

    Spans, Lien; Fletcher, Christopher Dm; Antonescu, Cristina R; Rouquette, Alexandre; Coindre, Jean-Michel; Sciot, Raf; Debiec-Rychter, Maria

    2016-07-01

    Plexiform fibromyxomas are rare neoplasms, being officially recognized as a distinct entity among benign mesenchymal gastric tumours in the 2010 WHO Classification of Tumours of the Digestive System. Characteristically, these tumours have a multinodular/plexiform growth pattern, and histologically contain variably cellular areas of bland myofibroblastic-type spindle cells embedded in an abundant myxoid matrix, rich in capillary-type vessels. As yet, the molecular and/or genetic features of these tumours are unknown. Here we describe a recurrent translocation, t(11;12)(q11;q13), involving the long non-coding gene metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and the gene glioma-associated oncogene homologue 1 (GLI1) in a subgroup of these tumours. The presence of the fusion transcript in our index case was confirmed using polymerase chain reaction (PCR) on genomic DNA, followed by Sanger sequencing. We showed that the truncated GLI1 protein is overexpressed and retains its capacity to transcriptionally activate its target genes. A specific FISH assay was developed to detect the novel MALAT1-GLI1 translocation in formalin-fixed, paraffin-embedded (FFPE) material. This resulted in the identification of two additional cases with this fusion and two cases with polysomy of the GLI1 gene. Finally, immunohistochemistry revealed that the GLI1 protein is exclusively overexpressed in those cases that harbour GLI1/12q13 genomic alterations. In conclusion, overexpression of GLI1 through a recurrent MALAT1-GLI1 translocation or GLI1 up-regulation delineates a pathogenically distinct subgroup of plexiform fibromyxomas with activation of the Sonic Hedgehog signalling pathway. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. PMID:27101025

  6. Norcantharidin Facilitates LPS-Mediated Immune Responses by Up-Regulation of AKT/NF-κB Signaling in Macrophages

    PubMed Central

    Li, Ruimei; Tan, Binghe; Han, Honghui; Liu, Mingyao; Qian, Min; Du, Bing

    2012-01-01

    Norcantharidin (NCTD), a demethylated analog of cantharidin, is a common used clinical drug to inhibit proliferation and metastasis of cancer cells. But the role of NCTD in modulating immune responses remains unknown. Here, we investigated the function and mechanism of NCTD in regulation of TLR4 associated immune response in macrophages. We evaluated the influence of NCTD on host defense against invaded pathogens by acute peritonitis mouse model, ELISA, Q-PCR, nitrite quantification, phagocytosis assay and gelatin zymography assay. Our data showed that the survival and the serum concentrations of IL-6 and TNF-α were all enhanced by NCTD significantly in peritonitis mouse model. Accordingly, LPS-induced cytokine, nitric oxide and MMP-9 production as well as the phagocytosis of bacteria were all up-regulated by NCTD in a dose dependent manner in both RAW264.7 cells and bone marrow-derived macrophages (BMMs). Then we further analyzed TLR4 associated signaling pathway by Western blot, Immunofluorescence and EMSA in the presence or absence of LPS. The phosphorylation of AKT and p65 at serine 536 but not serine 468 was enhanced obviously by NCTD in a dose dependent manner, whereas the degradation of IκBα was little effected. Consequently, the nuclear translocation and DNA binding ability of NF-κB was also increased by NCTD obviously in RAW264.7 cells. Our results demonstrated that NCTD could facilitate LPS-mediated immune response through promoting the phosphorylation of AKT/p65 and transcriptional activity of NF-κB, thus reprofiling the traditional anti-tumor drug NCTD as a novel immune regulator in promoting host defense against bacterial infection. PMID:22984593

  7. Up-regulation of transferrin receptor gene expression by granulocyte colony-stimulating factor in human myeloid leukemia cells.

    PubMed

    Morishita, Y; Kataoka, T; Towatari, M; Ito, T; Inoue, H; Ogura, M; Morishima, Y; Saito, H

    1990-12-15

    Granulocyte colony-stimulating factor (G-CSF) enhanced surface transferrin receptor (TfR) expression in two human myeloid leukemia cell lines, NKM-1 and NOMO-1, which possess G-CSF receptors. Radioligand-binding assay revealed that 10 ng/ml G-CSF significantly increased TfR to 186 +/- 20 and 276 +/- 38% of control for NKM-1 cells and NOMO-1 cells, respectively, in a 24-h culture. Scatchard analysis showed the increase of transferrin (Tf)-binding sites but no change in the receptor affinity. The enhanced TfR expression was not mediated either by the kinetic change of receptor cycling or by cellular iron content. Immunoprecipitation with anti-TfR antibody was used, and the increased biosynthesis of the receptor was demonstrated in G-CSF-stimulated cells. Northern blot analysis showed a 2- to 3-fold increase of TfR mRNA of NKM-1 cells cultured in medium containing Tf and G-CSF, whereas the mRNA declined without G-CSF. The effect of G-CSF on the TfR mRNA was observed within 2 h, which preceded the increase of surface TfR and the transition to the S phase of the cell cycle. G-CSF also potentiated TfR expression in freshly obtained myeloid leukemia cells. The present study shows up-regulation of TfR expression by G-CSF in myeloid leukemia cells and provides evidence that the regulation is mediated by controlling the steady-state level of the mRNA. PMID:1701357

  8. Natriuretic Peptide Receptor-C is Up-Regulated in the Intima of Advanced Carotid Artery Atherosclerosis

    PubMed Central

    Zayed, Mohamed A; Harring, Scott D; Abendschein, Dana R; Vemuri, Chandu; Lu, Dongsi; Detering, Lisa; Liu, Yongjian; Woodard, Pamela K

    2016-01-01

    Objective Natriuretic peptide receptor-C (NPR-C/NPR-3) is a cell surface protein involved in vascular remodelling that is up-regulated in atherosclerosis. NPR-C expression has not been well characterized in human carotid artery occlusive lesions. We hypothesized that NPR-C expression correlates with intimal features of vulnerable atherosclerotic carotid artery plaque. Methods To test this hypothesis, we evaluated NPR-C expression by immunohistochemistry (IHC) in carotid endarterectomy (CEA) specimens isolated from 18 patients. The grade, location, and co-localization of NPR-C in CEA specimens were evaluated using two tissue analysis techniques. Results Relative to minimally diseased CEA specimens, we observed avid NPR-C tissue staining in the intima of maximally diseased CEA specimens (65%; p=0.06). Specifically, maximally diseased CEA specimens demonstrated increased NPR-C expression in the superficial intima (61%, p=0.17), and deep intima (138% increase; p=0.05). In the superficial intima, NPR-C expression significantly co-localized with vascular smooth muscle cells (VSMCs) and macrophages. The intensity of NPR-C expression was also higher in the superficial intima plaque shoulder and cap regions, and significantly correlated with atheroma and fibroatheroma vulnerable plaque regions (β=1.04, 95% CI=0.46, 1.64). Conclusion These findings demonstrate significant NPR-C expression in the intima of advanced carotid artery plaques. Furthermore, NPR-C expression was higher in vulnerable carotid plaque intimal regions, and correlate with features of advanced disease. Our findings suggest that NPR-C may serve as a potential biomarker for carotid plaque vulnerability and progression, in patients with advanced carotid artery occlusive disease. PMID:27547837

  9. Six1 induces protein synthesis signaling expression in duck myoblasts mainly via up-regulation of mTOR

    PubMed Central

    Wang, Haohan; Li, Xinxin; Liu, Hehe; Sun, Lingli; Zhang, Rongping; Li, Liang; Wangding, Mincheng; Wang, Jiwen

    2016-01-01

    Abstract As a critical transcription factor, Six1 plays an important role in the regulation of myogenesis and muscle development. However, little is known about its regulatory mechanism associated with muscular protein synthesis. The objective of this study was to investigate the effects of overexpression ofSix1 on the expression of key protein metabolism-related genes in duck myoblasts. Through an experimental model where duck myoblasts were transfected with a pEGFP-duSix1 construct, we found that overexpression of duckSix1 could enhance cell proliferation activity and increase mRNA expression levels of key genes involved in the PI3K/Akt/mTOR signaling pathway, while the expression of FOXO1, MuRF1and MAFbx was not significantly altered, indicating thatSix1 could promote protein synthesis in myoblasts through up-regulating the expression of several related genes. Additionally, in duck myoblasts treated with LY294002 and rapamycin, the specific inhibitors ofPI3K and mTOR, respectively, the overexpression of Six1 could significantly ameliorate inhibitive effects of these inhibitors on protein synthesis. Especially, the mRNA expression levels of mTOR and S6K1 were observed to undergo a visible change, and a significant increase in protein expression of S6K1 was seen. These data suggested that Six1plays an important role in protein synthesis, which may be mainly due to activation of the mTOR signaling pathway. PMID:27007909

  10. Campylobacter jejuni Biofilms Up-Regulated in the Absence of the Stringent Response Utilize a Calcofluor White-Reactive Polysaccharide▿

    PubMed Central

    McLennan, Meghan K.; Ringoir, Danielle D.; Frirdich, Emilisa; Svensson, Sarah L.; Wells, Derek H.; Jarrell, Harold; Szymanski, Christine M.; Gaynor, Erin C.

    2008-01-01

    The enteric pathogen Campylobacter jejuni is a highly prevalent yet fastidious bacterium. Biofilms and surface polysaccharides participate in stress survival, transmission, and virulence in C. jejuni; thus, the identification and characterization of novel genes involved in each process have important implications for pathogenesis. We found that C. jejuni reacts with calcofluor white (CFW), indicating the presence of surface polysaccharides harboring β1-3 and/or β1-4 linkages. CFW reactivity increased with extended growth, under 42°C anaerobic conditions, and in a ΔspoT mutant defective for the stringent response (SR). Conversely, two newly isolated dim mutants exhibited diminished CFW reactivity as well as growth and serum sensitivity differences from the wild type. Genetic, biochemical, and nuclear magnetic resonance analyses suggested that differences in CFW reactivity between wild-type and ΔspoT and dim mutant strains were independent of well-characterized lipooligosaccharides, capsular polysaccharides, and N-linked polysaccharides. Targeted deletion of carB downstream of the dim13 mutation also resulted in CFW hyporeactivity, implicating a possible role for carbamoylphosphate synthase in the biosynthesis of this polysaccharide. Correlations between biofilm formation and production of the CFW-reactive polymer were demonstrated by crystal violet staining, scanning electron microscopy, and confocal microscopy, with the C. jejuni ΔspoT mutant being the first SR mutant in any bacterial species identified as up-regulating biofilms. Together, these results provide new insight into genes and processes important for biofilm formation and polysaccharide production in C. jejuni. PMID:17993532

  11. Dietary fish oil replacement with canola oil up-regulates glutathione peroxidase 1 gene expression in yellowtail kingfish (Seriola lalandi).

    PubMed

    Bowyer, Jenna N; Rout-Pitt, Nathan; Bain, Peter A; Stone, David A J; Schuller, Kathryn A

    2012-08-01

    The marine carnivore yellowtail kingfish (YTK, Seriola lalandi) was fed diets containing 5% residual fish oil (from the dietary fish meal) plus either 20% fish oil (FO), 20% canola oil (CO), 20% poultry oil (PO), 10% fish oil plus 10% canola oil (FO/CO) or 10% fish oil plus 10% poultry oil (FO/PO) and the effects on fish growth and hepatic expression of two glutathione peroxidase (GPx 1 and GPx 4) and two peroxiredoxin (Prx 1 and Prx 4) antioxidant genes were investigated. Partial (50%) replacement of the added dietary fish oil with poultry oil significantly improved fish growth whereas 100% replacement with canola oil significantly depressed fish growth. The fatty acid profiles of the fish fillets generally reflected those of the dietary oils except that there was apparent selective utilization of palmitic acid (16:0) and oleic acid (18:1n-9) and apparent selective retention of eicospentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3). The Prx 1 and 4 genes were expressed at 10- and 100-fold the level of the GPx 4 and 1 genes, respectively, and at one-tenth the level of the highly expressed β-actin reference gene. Dietary fish oil replacement with canola oil significantly up-regulated GPx 1 gene expression and there was a non-significant tendency towards down-regulation of Prx 1 and Prx 4. The results are discussed in terms of the effects of fish oil replacement on the peroxidation index of the diets and the resulting effects on the target antioxidant enzymes. PMID:22521527

  12. Up-regulation of galectin-9 induces cell migration in human dendritic cells infected with dengue virus.

    PubMed

    Hsu, Yu-Lin; Wang, Mei-Yi; Ho, Ling-Jun; Huang, Chuan-Yueh; Lai, Jenn-Haung

    2015-05-01

    Galectin-9 (Gal-9) exerts immunosuppressive effects by inducing apoptosis in T cells that produce interferon-γ and interleukin (IL)-17. However, Gal-9 can be pro-inflammatory in lipopolysaccharide-stimulated monocytes. Using microarray analysis, we observed that Gal-9 was up-regulated in human dendritic cells (DCs) after dengue virus (DV) infection. The investigation into the immunomodulatory effects and mechanisms of Gal-9 in DCs exposed to DV revealed that DV infection specifically increased mRNA and protein levels of Gal-9 but not those of Gal-1 or Gal-3. Blocking p38, but not c-Jun N-terminal kinase or extracellular signal-regulated kinase (ERK), inhibited DV-induced expression of Gal-9. Reduction in Gal-9 by small interference RNA treatment suppressed DV-stimulated migration of DCs towards the chemoattractants CCL19 and CCL21. In addition, DV-induced IL-12p40 production was reduced after knockdown of Gal-9 in DCs. Furthermore, Gal-9 deficiency suppressed DV-induced activation of nuclear factor-κB. Inhibition of DV-induced DC migration under conditions of Gal-9 deficiency was mediated through suppressing ERK activation but not by regulating the expression of CCR7, the receptor for CCL19 and CCL21. Both the reduction in IL-12 production and the suppression of ERK activity might account for the inhibition of DV-induced DC migration after knockdown of Gal-9. In summary, this study reveals the roles of Gal-9 in DV-induced migration of DCs. The findings indicate that Gal-9 might be a therapeutic target for preventing immunopathogenesis induced by DV infection. PMID:25754930

  13. NOS1AP modulates intracellular Ca2+ in cardiac myocytes and is up-regulated in dystrophic cardiomyopathy

    PubMed Central

    Treuer, Adriana V; Gonzalez, Daniel R

    2014-01-01

    NOS1AP gene (nitric oxide synthase 1-adaptor protein) is strongly associated with abnormalities in the QT interval of the electrocardiogram and with sudden cardiac death. To determine the role of NOS1AP in the physiology of the cardiac myocyte, we assessed the impact of silencing NOS1AP, using siRNA, on [Ca2+]i transients in neonatal cardiomyocytes. In addition, we examined the co-localization of NOS1AP with cardiac ion channels, and finally, evaluated the expression of NOS1AP in a mouse model of dystrophic cardiomyopathy. Using siRNA, NOS1AP levels were reduced to ~30% of the control levels (p<0.05). NOS1AP silencing in cardiac myocytes reduced significantly the amplitude of electrically evoked calcium transients (p<0.05) and the degree of S-nitrosylation of the cells (p<0.05). Using confocal microscopy, we evaluated NOS1AP subcellular location and interactions with other proteins by co-localization analysis. NOS1AP showed a high degree of co-localization with the L-type calcium channel and the inwardly rectifying potassium channel Kir3.1, a low degree of co-localization with the ryanodine receptor (RyR2) and alfa-sarcomeric actin and no co-localization with connexin 43, suggesting functionally relevant interactions with the ion channels that regulate the action potential duration. Finally, using immunofluorescence and Western blotting, we observed that in mice with dystrophic cardiomyopathy, NOS1AP was significantly up-regulated (p<0.05). These results suggest for a role of NOS1AP on cardiac arrhythmias, acting on the L-type calcium channel, and potassium channels, probably through S-nitrosylation. PMID:24665357

  14. Recurrent MALAT1-GLI1 oncogenic fusion and GLI1 up-regulation define a subset of plexiform fibromyxoma.

    PubMed

    Spans, Lien; Fletcher, Christopher Dm; Antonescu, Cristina R; Rouquette, Alexandre; Coindre, Jean-Michel; Sciot, Raf; Debiec-Rychter, Maria

    2016-07-01

    Plexiform fibromyxomas are rare neoplasms, being officially recognized as a distinct entity among benign mesenchymal gastric tumours in the 2010 WHO Classification of Tumours of the Digestive System. Characteristically, these tumours have a multinodular/plexiform growth pattern, and histologically contain variably cellular areas of bland myofibroblastic-type spindle cells embedded in an abundant myxoid matrix, rich in capillary-type vessels. As yet, the molecular and/or genetic features of these tumours are unknown. Here we describe a recurrent translocation, t(11;12)(q11;q13), involving the long non-coding gene metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and the gene glioma-associated oncogene homologue 1 (GLI1) in a subgroup of these tumours. The presence of the fusion transcript in our index case was confirmed using polymerase chain reaction (PCR) on genomic DNA, followed by Sanger sequencing. We showed that the truncated GLI1 protein is overexpressed and retains its capacity to transcriptionally activate its target genes. A specific FISH assay was developed to detect the novel MALAT1-GLI1 translocation in formalin-fixed, paraffin-embedded (FFPE) material. This resulted in the identification of two additional cases with this fusion and two cases with polysomy of the GLI1 gene. Finally, immunohistochemistry revealed that the GLI1 protein is exclusively overexpressed in those cases that harbour GLI1/12q13 genomic alterations. In conclusion, overexpression of GLI1 through a recurrent MALAT1-GLI1 translocation or GLI1 up-regulation delineates a pathogenically distinct subgroup of plexiform fibromyxomas with activation of the Sonic Hedgehog signalling pathway. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  15. Ethanol up-regulates nucleus accumbens neuronal activity dependent pentraxin (Narp): implications for alcohol-induced behavioral plasticity.

    PubMed

    Ary, Alexis W; Cozzoli, Debra K; Finn, Deborah A; Crabbe, John C; Dehoff, Marlin H; Worley, Paul F; Szumlinski, Karen K

    2012-06-01

    Neuronal activity dependent pentraxin (Narp) interacts with α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA) glutamate receptors to facilitate excitatory synapse formation by aggregating them at established synapses. Alcohol is well-characterized to influence central glutamatergic transmission, including AMPA receptor function. Herein, we examined the influence of injected and ingested alcohol upon Narp protein expression, as well as basal Narp expression in mouse lines selectively bred for high blood alcohol concentrations under limited access conditions. Alcohol up-regulated accumbens Narp levels, concomitant with increases in levels of the GluR1 AMPA receptor subunit. However, accumbens Narp or GluR1 levels did not vary as a function of selectively bred genotype. We next employed a Narp knock-out (KO) strategy to begin to understand the behavioral relevance of alcohol-induced changes in protein expression in several assays of alcohol reward. Compared to wild-type mice, Narp KO animals: fail to escalate daily intake of high alcohol concentrations under free-access conditions; shift their preference away from high alcohol concentrations with repeated alcohol experience; exhibit a conditioned place-aversion in response to the repeated pairing of 3 g/kg alcohol with a distinct environment and fail to exhibit alcohol-induced locomotor hyperactivity following repeated alcohol treatment. Narp deletion did not influence the daily intake of either food or water, nor did it alter any aspect of spontaneous or alcohol-induced motor activity, including the development of tolerance to its motor-impairing effects with repeated treatment. Taken together, these data indicate that Narp induction, and presumably subsequent aggregation of AMPA receptors, may be important for neuroplasticity within limbic subcircuits mediating or maintaining the rewarding properties of alcohol.

  16. Up-regulation of phosphoinositide metabolism in tobacco cells constitutively expressing the human type I inositol polyphosphate 5-phosphatase

    NASA Technical Reports Server (NTRS)

    Perera, Imara Y.; Love, John; Heilmann, Ingo; Thompson, William F.; Boss, Wendy F.; Brown, C. S. (Principal Investigator)

    2002-01-01

    To evaluate the impact of suppressing inositol 1,4,5-trisphosphate (InsP(3)) in plants, tobacco (Nicotiana tabacum) cells were transformed with the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), an enzyme which specifically hydrolyzes InsP(3). The transgenic cell lines showed a 12- to 25-fold increase in InsP 5-ptase activity in vitro and a 60% to 80% reduction in basal InsP(3) compared with wild-type cells. Stimulation with Mas-7, a synthetic analog of the wasp venom peptide mastoparan, resulted in an approximately 2-fold increase in InsP(3) in both wild-type and transgenic cells. However, even with stimulation, InsP(3) levels in the transgenic cells did not reach wild-type basal values, suggesting that InsP(3) signaling is compromised. Analysis of whole-cell lipids indicated that phosphatidylinositol 4,5-bisphosphate (PtdInsP(2)), the lipid precursor of InsP(3), was greatly reduced in the transgenic cells. In vitro assays of enzymes involved in PtdInsP(2) metabolism showed that the activity of the PtdInsP(2)-hydrolyzing enzyme phospholipase C was not significantly altered in the transgenic cells. In contrast, the activity of the plasma membrane PtdInsP 5 kinase was increased by approximately 3-fold in the transgenic cells. In vivo labeling studies revealed a greater incorporation of (32)P into PtdInsP(2) in the transgenic cells compared with the wild type, indicating that the rate of PtdInsP(2) synthesis was increased. These studies show that the constitutive expression of the human type I InsP 5-ptase in tobacco cells leads to an up-regulation of the phosphoinositide pathway and highlight the importance of PtdInsP(2) synthesis as a regulatory step in this system.

  17. Neurodegeneration in Autoimmune Optic Neuritis Is Associated with Altered APP Cleavage in Neurons and Up-Regulation of p53.

    PubMed

    Herold, Sabine; Kumar, Prateek; Wichert, Sven P; Kretzschmar, Benedikt; Bähr, Mathias; Rossner, Moritz J; Hein, Katharina

    2015-01-01

    Multiple Sclerosis (MS) is a chronic autoimmune inflammatory disease of the central nervous system (CNS). Histopathological and radiological analysis revealed that neurodegeneration occurs early in the disease course. However, the pathological mechanisms involved in neurodegeneration are poorly understood. Myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE) in Brown Norway rats (BN-rats) is a well-established animal model, especially of the neurodegenerative aspects of MS. Previous studies in this animal model indicated that loss of retinal ganglion cells (RGCs), the neurons that form the axons of the optic nerve, occurs in the preclinical phase of the disease and is in part independent of overt histopathological changes of the optic nerve. Therefore, the aim of this study was to identify genes which are involved in neuronal cell loss at different disease stages of EAE. Furthermore, genes that are highly specific for autoimmune-driven neurodegeneration were compared to those regulated in RGCs after optic nerve axotomy at corresponding time points. Using laser capture micro dissection we isolated RNA from unfixed RGCs and performed global transcriptome analysis of retinal neurons. In total, we detected 582 genes sequentially expressed in the preclinical phase and 1150 genes in the clinical manifest EAE (P < 0.05, fold-induction >1.5). Furthermore, using ingenuity pathway analysis (IPA), we identified amyloid precursor protein (APP) as a potential upstream regulator of changes in gene expression in the preclinical EAE but neither in clinical EAE, nor at any time point after optic nerve transection. Therefore, the gene pathway analysis lead to the hypothesis that altered cleavage of APP in neurons in the preclinical phase of EAE leads to the enhanced production of APP intracellular domain (AICD), which in turn acts as a transcriptional regulator and thereby initiates an apoptotic signaling cascade via up-regulation of the

  18. MiR-21 up-regulation mediates glioblastoma cancer stem cells apoptosis and proliferation by targeting FASLG.

    PubMed

    Shang, Chao; Guo, Yan; Hong, Yang; Liu, Yun-hui; Xue, Yi-xue

    2015-03-01

    To investigate whether miR-21 can affect the apoptosis and proliferation of glioblastoma cancer stem cells (GSCs) from down-regulating FASLG. The expression of miRNA-21 was detected by quantitative real-time PCR in normal brain tissue and glioblastoma samples, and the changes of miRNA-21 expression between GSCs and non-GSCs were also detected. The apoptosis and proliferation ability of miR-21 in GSCs were analyzed by MTT and flow cytometry assay after anti-miR-21 transfection. For the regulation mechanism analysis of miR-21, TargetScan, PicTar and microRNA were selected to predict some potential target genes of miR-21. The predicted gene was identified to be the direct and specific target gene of miR-21 by luciferase activities assay and western blot. RNA interference technology was used to confirm the apoptosis and proliferation effects of miR-21 were directly induced by FASLG. The expression of miR-21 increased significantly in glioblastoma contrast to normal brain tissue, and miR-21 up-regulated in GSCs remarkably. The proliferation of GSCs cell could be inhibited with high-expression of miR-21 and this effect could be restored by miR-21 knocked down. Mechanism analysis revealed that FASLG was a specific and direct target gene of miR-21. The advanced effects of anti-miR-21 on GSCs apoptosis and proliferation were mediated by expression of silenced FASLG. In summary, aberrantly expressed miR-21 regulates GSCs apoptosis and proliferation partly through directly down-regulating FASLG protein expression in GSCs and this might offer a new potential therapeutic stratagem for glioblastoma. PMID:25394756

  19. Isoflurane Preconditioning Induces Neuroprotection by Up-Regulation of TREK1 in a Rat Model of Spinal Cord Ischemic Injury.

    PubMed

    Wang, Kun; Kong, Xiangang

    2016-09-01

    This study aimed to explore the neuroprotection and mechanism of isoflurane on rats with spinal cord ischemic injury. Total 40 adult male Sprague-Dawley rats were divided into the four groups (n=10). Group A was sham-operation group; group B was ischemia group; group C was isoflurane preconditioning group; group D was isoflurane preconditioning followed by ischemia treatment group. Then the expressions of TWIK-related K⁺ channel 1 (TREK1) in the four groups were detected by immunofluorescent assay, real time-polymerase chain reactions (RT-PCR) and western blot. The primary neurons of rats were isolated and cultured under normal and hypoxic conditions. Besides, the neurons under two conditions were transfected with green fluorescent protein (GFP)-TREK1 and lentivirual to overexpress and silence TREK1. Additionally, the neurons were treated with isoflurane or not. Then caspase-3 activity and cell cycle of neurons under normal and hypoxic conditions were detected. Furthermore, nicotinamide adenine dinucleotide hydrate (NADH) was detected using NAD+/NADH quantification colorimetric kit. Results showed that the mRNA and protein expressions of TREK1 increased significantly in group C and D. In neurons, when TREK1 silenced, isoflurane treatment improved the caspase-3 activity. In hypoxic condition, the caspase-3 activity and sub-G1 cell percentage significantly increased, however, when TREK1 overexpressed the caspase-3 activity and sub-G1 cell percentage decreased significantly. Furthermore, both isoflurane treatment and overexpression of TREK1 significantly decreased NADH. In conclusion, isoflurane-induced neuroprotection in spinal cord ischemic injury may be associated with the up-regulation of TREK1. PMID:27469140

  20. Direct laser machining-induced topographic pattern promotes up-regulation of myogenic markers in human mesenchymal stem cells.

    PubMed

    Li, Huaqiong; Wen, Feng; Wong, Yee Shan; Boey, Freddy Yin Chiang; Subbu, Venkatraman S; Leong, David Tai; Ng, Kee Woei; Ng, Gary Ka Lai; Tan, Lay Poh

    2012-02-01

    The engineering of tissue is preferably done with stem cells, which can be differentiated into the tissue of interest using biochemical or physical cues. While much effort has been focused on using biological factors to regulate stem cell differentiation, recently interest in the contribution of physical factors has increased. In this work, three-dimensional (3-D) microchannels with topographic micropatterns were fabricated by femtosecond laser machining on a biodegradable polymer (poly(L-lactide-co-ε-caprolactone)) substrate. Two substrates with narrow and wide channels respectively were created. Human mesenchymal stem cells (hMSCs) were cultured on the scaffolds for cell proliferation and cellular organization. Gene expression and the immunostaining of myogenic and neurogenic markers were studied. Both scaffolds improved the cell alignment along the channels as compared to the control group. Microfilaments within hMSCs were more significantly aligned and elongated on the narrower microchannels. The gene expression study revealed significant up-regulation of several hallmark markers associated with myogenesis for hMSCs cultured on the scaffold with narrow microchannels, while osteogenic and neurogenic markers were down-regulated or remained similar to the control at day 14. Immunostaining of myogen- and neurogen-specific differentiation markers were used to further confirm the specific differentiation towards a myogenic lineage. This study demonstrates that femtosecond laser machining is a versatile tool for generating controllable 3-D microchannels with topographic features that can be used to induce specific myogenic differentiation of hMSCs in vitro, even in the absence of biological factors.

  1. Transcriptome Profiling Revealed Stress-Induced and Disease Resistance Genes Up-Regulated in PRSV Resistant Transgenic Papaya.

    PubMed

    Fang, Jingping; Lin, Aiting; Qiu, Weijing; Cai, Hanyang; Umar, Muhammad; Chen, Rukai; Ming, Ray

    2016-01-01

    Papaya is a productive and nutritious tropical fruit. Papaya Ringspot Virus (PRSV) is the most devastating pathogen threatening papaya production worldwide. Development of transgenic resistant varieties is the most effective strategy to control this disease. However, little is known about the genome-wide functional changes induced by particle bombardment transformation. We conducted transcriptome sequencing of PRSV resistant transgenic papaya SunUp and its PRSV susceptible progenitor Sunset to compare the transcriptional changes in young healthy leaves prior to infection with PRSV. In total, 20,700 transcripts were identified, and 842 differentially expressed genes (DEGs) randomly distributed among papaya chromosomes. Gene ontology (GO) category analysis revealed that microtubule-related categories were highly enriched among these DEGs. Numerous DEGs related to various transcription factors, transporters and hormone biosynthesis showed clear differences between the two cultivars, and most were up-regulated in transgenic papaya. Many known and novel stress-induced and disease-resistance genes were most highly expressed in SunUp, including MYB, WRKY, ERF, NAC, nitrate and zinc transporters, and genes involved in the abscisic acid, salicylic acid, and ethylene signaling pathways. We also identified 67,686 alternative splicing (AS) events in Sunset and 68,455 AS events in SunUp, mapping to 10,994 and 10,995 papaya annotated genes, respectively. GO enrichment for the genes displaying AS events exclusively in Sunset was significantly different from those in SunUp. Transcriptomes in Sunset and transgenic SunUp are very similar with noteworthy differences, which increased PRSV-resistance in transgenic papaya. No detrimental pathways and allergenic or toxic proteins were induced on a genome-wide scale in transgenic SunUp. Our results provide a foundation for unraveling the mechanism of PRSV resistance in transgenic papaya. PMID:27379138

  2. Neurodegeneration in Autoimmune Optic Neuritis Is Associated with Altered APP Cleavage in Neurons and Up-Regulation of p53

    PubMed Central

    Wichert, Sven P.; Kretzschmar, Benedikt; Bähr, Mathias; Rossner, Moritz J.; Hein, Katharina

    2015-01-01

    Multiple Sclerosis (MS) is a chronic autoimmune inflammatory disease of the central nervous system (CNS). Histopathological and radiological analysis revealed that neurodegeneration occurs early in the disease course. However, the pathological mechanisms involved in neurodegeneration are poorly understood. Myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE) in Brown Norway rats (BN-rats) is a well-established animal model, especially of the neurodegenerative aspects of MS. Previous studies in this animal model indicated that loss of retinal ganglion cells (RGCs), the neurons that form the axons of the optic nerve, occurs in the preclinical phase of the disease and is in part independent of overt histopathological changes of the optic nerve. Therefore, the aim of this study was to identify genes which are involved in neuronal cell loss at different disease stages of EAE. Furthermore, genes that are highly specific for autoimmune-driven neurodegeneration were compared to those regulated in RGCs after optic nerve axotomy at corresponding time points. Using laser capture micro dissection we isolated RNA from unfixed RGCs and performed global transcriptome analysis of retinal neurons. In total, we detected 582 genes sequentially expressed in the preclinical phase and 1150 genes in the clinical manifest EAE (P < 0.05, fold-induction >1.5). Furthermore, using ingenuity pathway analysis (IPA), we identified amyloid precursor protein (APP) as a potential upstream regulator of changes in gene expression in the preclinical EAE but neither in clinical EAE, nor at any time point after optic nerve transection. Therefore, the gene pathway analysis lead to the hypothesis that altered cleavage of APP in neurons in the preclinical phase of EAE leads to the enhanced production of APP intracellular domain (AICD), which in turn acts as a transcriptional regulator and thereby initiates an apoptotic signaling cascade via up-regulation of the

  3. Up-regulation of abscisic acid signaling pathway facilitates aphid xylem absorption and osmoregulation under drought stress

    PubMed Central

    Guo, Huijuan; Sun, Yucheng; Peng, Xinhong; Wang, Qinyang; Harris, Marvin; Ge, Feng

    2016-01-01

    The activation of the abscisic acid (ABA) signaling pathway reduces water loss from plants challenged by drought stress. The effect of drought-induced ABA signaling on the defense and nutrition allocation of plants is largely unknown. We postulated that these changes can affect herbivorous insects. We studied the effects of drought on different feeding stages of pea aphids in the wild-type A17 of Medicago truncatula and ABA signaling pathway mutant sta-1. We examined the impact of drought on plant water status, induced plant defense signaling via the abscisic acid (ABA), jasmonic acid (JA), and salicylic acid (SA) pathways, and on the host nutritional quality in terms of leaf free amino acid content. During the penetration phase of aphid feeding, drought decreased epidermis/mesophyll resistance but increased mesophyll/phloem resistance of A17 but not sta-1 plants. Quantification of transcripts associated with ABA, JA and SA signaling indicated that the drought-induced up-regulation of ABA signaling decreased the SA-dependent defense but increased the JA-dependent defense in A17 plants. During the phloem-feeding phase, drought had little effect on the amino acid concentrations and the associated aphid phloem-feeding parameters in both plant genotypes. In the xylem absorption stage, drought decreased xylem absorption time of aphids in both genotypes because of decreased water potential. Nevertheless, the activation of the ABA signaling pathway increased water-use efficiency of A17 plants by decreasing the stomatal aperture and transpiration rate. In contrast, the water potential of sta-1 plants (unable to close stomata) was too low to support xylem absorption activity of aphids; the aphids on sta-1 plants had the highest hemolymph osmolarity and lowest abundance under drought conditions. Taken together this study illustrates the significance of cross-talk between biotic-abiotic signaling pathways in plant-aphid interaction, and reveals the mechanisms leading to alter

  4. Up- regulation of miR-328-3p sensitizes non-small cell lung cancer to radiotherapy.

    PubMed

    Ma, Wei; Ma, Chao-Nan; Zhou, Nan-Nan; Li, Xian-Dong; Zhang, Yi-Jie

    2016-01-01

    MicroRNAs (miRNAs) are believed to be resistant against radiotherapy in certain types of cancers. The aim of our study was to determine the clinical application of miRNAs in non-small cell lung cancer (NSCLC). Sixty NSCLC tissue samples and adjacent histologically normal tissues were obtained for miRNAs microarray analysis and validated by RT-qPCR. Correlation between miRNA expression level and clinicopathological features was evaluated. Our study examined the influence of changed miRNA expression on the damaged DNA and its associated radio sensitivity. Luciferase assay was performed to determine potential effects on the targeted gene. Our study identified fifteen altered miRNAs in which miR-328-3p was down regulated in NSCLC tumour tissue as compared to normal tissues. Down-expression of miR-328-3p was positively associated with an enhanced lymph node metastasis, advanced clinical stage and a shortened survival rate. miR-328-3p expression was decreased in A549 cells compared to other NSCLC cell lines. Up-regulation of miR-328-3p demonstrated a survival inhibition effect in A549 and restored NSCLC cells' sensitivity to radio therapy. An increased miR-328-3p expression promoted irradiation-induced DNA damage in cells. γ-H2AX was identified as the direct target of miR-328-3p. Over-expressed miR-328-3p can improve the radiosensitvity of cells by altering the DNA damage/repair signalling pathways in NSCLC. PMID:27530148

  5. Transcriptional up-regulation of a novel ferritin homolog in abalone Haliotis discus hannai Ino by dietary iron.

    PubMed

    Wu, Chenglong; Zhang, Wenbing; Mai, Kangsen; Xu, Wei; Wang, Xiaojie; Ma, Hongming; Liufu, Zhiguo

    2010-11-01

    A novel cDNA encoding ferritin (HdhNFT) was cloned from the hepatopancreas of abalone, Haliotis discus hannai Ino. The deduced protein contains 171 amino acid residues with a predicted molecular mass (MW) about 19.8 kDa and theoretical isoelectric point (pI) of 4.792. Amino acid alignment revealed that HdhNFT shared high similarity with other known ferritins. The HdhNFT contained a highly conserved motif for the ferroxidase center, which consists of seven residues of a typical vertebrate heavy-chain ferritin with a typical stem-loop structure. HdhNFT mRNA contains a 27 bp iron-responsive element (IRE) in the 5'-untranslated region. This IRE exhibited 82.14% similarity with abalone H. discus discus and 78.57% similarity with Pacific oyster Crassostrea gigas IREs. By real-time PCR assays, the mRNA transcripts of HdhNFT were found to be higher expressed in kidney, hepatopancreas, gill, mantle and muscle than in haemocytes and gonad. Moreover, mRNA expression levels of HdhNFT in the hepatopancreas and haemocytes were measured by real-time PCR in abalone fed with graded levels of dietary iron (29.2, 65.7, 1267.2 and 6264.7 mg/kg). Results showed that the expression of the HdhNFT mRNA increased with dietary iron contents. Furthermore, the maximum value of the HdhNFT mRNA was found in the treatment with 6264.7 mg/kg of dietary iron. These data indicated that dietary iron can up-regulate HdhNFT at transcriptional level in abalone.

  6. Up-regulation of abscisic acid signaling pathway facilitates aphid xylem absorption and osmoregulation under drought stress.

    PubMed

    Guo, Huijuan; Sun, Yucheng; Peng, Xinhong; Wang, Qinyang; Harris, Marvin; Ge, Feng

    2016-02-01

    The activation of the abscisic acid (ABA) signaling pathway reduces water loss from plants challenged by drought stress. The effect of drought-induced ABA signaling on the defense and nutrition allocation of plants is largely unknown. We postulated that these changes can affect herbivorous insects. We studied the effects of drought on different feeding stages of pea aphids in the wild-type A17 of Medicago truncatula and ABA signaling pathway mutant sta-1. We examined the impact of drought on plant water status, induced plant defense signaling via the abscisic acid (ABA), jasmonic acid (JA), and salicylic acid (SA) pathways, and on the host nutritional quality in terms of leaf free amino acid content. During the penetration phase of aphid feeding, drought decreased epidermis/mesophyll resistance but increased mesophyll/phloem resistance of A17 but not sta-1 plants. Quantification of transcripts associated with ABA, JA and SA signaling indicated that the drought-induced up-regulation of ABA signaling decreased the SA-dependent defense but increased the JA-dependent defense in A17 plants. During the phloem-feeding phase, drought had little effect on the amino acid concentrations and the associated aphid phloem-feeding parameters in both plant genotypes. In the xylem absorption stage, drought decreased xylem absorption time of aphids in both genotypes because of decreased water potential. Nevertheless, the activation of the ABA signaling pathway increased water-use efficiency of A17 plants by decreasing the stomatal aperture and transpiration rate. In contrast, the water potential of sta-1 plants (unable to close stomata) was too low to support xylem absorption activity of aphids; the aphids on sta-1 plants had the highest hemolymph osmolarity and lowest abundance under drought conditions. Taken together this study illustrates the significance of cross-talk between biotic-abiotic signaling pathways in plant-aphid interaction, and reveals the mechanisms leading to alter

  7. Transcriptome Profiling Revealed Stress-Induced and Disease Resistance Genes Up-Regulated in PRSV Resistant Transgenic Papaya

    PubMed Central

    Fang, Jingping; Lin, Aiting; Qiu, Weijing; Cai, Hanyang; Umar, Muhammad; Chen, Rukai; Ming, Ray

    2016-01-01

    Papaya is a productive and nutritious tropical fruit. Papaya Ringspot Virus (PRSV) is the most devastating pathogen threatening papaya production worldwide. Development of transgenic resistant varieties is the most effective strategy to control this disease. However, little is known about the genome-wide functional changes induced by particle bombardment transformation. We conducted transcriptome sequencing of PRSV resistant transgenic papaya SunUp and its PRSV susceptible progenitor Sunset to compare the transcriptional changes in young healthy leaves prior to infection with PRSV. In total, 20,700 transcripts were identified, and 842 differentially expressed genes (DEGs) randomly distributed among papaya chromosomes. Gene ontology (GO) category analysis revealed that microtubule-related categories were highly enriched among these DEGs. Numerous DEGs related to various transcription factors, transporters and hormone biosynthesis showed clear differences between the two cultivars, and most were up-regulated in transgenic papaya. Many known and novel stress-induced and disease-resistance genes were most highly expressed in SunUp, including MYB, WRKY, ERF, NAC, nitrate and zinc transporters, and genes involved in the abscisic acid, salicylic acid, and ethylene signaling pathways. We also identified 67,686 alternative splicing (AS) events in Sunset and 68,455 AS events in SunUp, mapping to 10,994 and 10,995 papaya annotated genes, respectively. GO enrichment for the genes displaying AS events exclusively in Sunset was significantly different from those in SunUp. Transcriptomes in Sunset and transgenic SunUp are very similar with noteworthy differences, which increased PRSV-resistance in transgenic papaya. No detrimental pathways and allergenic or toxic proteins were induced on a genome-wide scale in transgenic SunUp. Our results provide a foundation for unraveling the mechanism of PRSV resistance in transgenic papaya. PMID:27379138

  8. Carnitine palmitoyltransferase-1 up-regulation by PPAR-β/δ prevents lipid-induced endothelial dysfunction.

    PubMed

    Toral, Marta; Romero, Miguel; Jiménez, Rosario; Mahmoud, Ayman Moawad; Barroso, Emma; Gómez-Guzmán, Manuel; Sánchez, Manuel; Cogolludo, Ángel; García-Redondo, Ana B; Briones, Ana M; Vázquez-Carrera, Manuel; Pérez-Vizcaíno, Francisco; Duarte, Juan

    2015-11-01

    Fatty acids cause endothelial dysfunction involving increased ROS (reactive oxygen species) and reduced NO (nitric oxide) bioavailability. We show that in MAECs (mouse aortic endothelial cells), the PPARβ/δ (peroxisome- proliferator-activated receptor β/δ) agonist GW0742 prevented the decreased A23187-stimulated NO production, phosphorylation of eNOS (endothelial nitric oxide synthase) at Ser1177 and increased intracellular ROS levels caused by exposure to palmitate in vitro. The impaired endothelium-dependent relaxation to acetylcholine in mouse aorta induced by palmitate was restored by GW0742. In vivo, GW0742 treatment prevented the reduced aortic relaxation, phosphorylation of eNOS at Ser1177, and increased ROS production and NADPH oxidase in mice fed on a high-fat diet. The PPARβ/δ antagonist GSK0660 abolished all of these protective effects induced by GW0742. This agonist enhanced the expression of CPT (carnitine palmitoyltransferase)-1. The effects of GW0742 on acetylcholine- induced relaxation in aorta and on NO and ROS production in MAECs exposed to palmitate were abolished by the CPT-1 inhibitor etomoxir or by siRNA targeting CPT-1. GW0742 also inhibited the increase in DAG (diacylglycerol), PKCα/βII (protein kinase Cα/βII) activation, and phosphorylation of eNOS at Thr495 induced by palmitate in MAECs, which were abolished by etomoxir. In conclusion, PPARβ/δ activation restored the lipid-induced endothelial dysfunction by up-regulation of CPT-1, thus reducing DAG accumulation and the subsequent PKC-mediated ROS production and eNOS inhibition.

  9. Electrical stimulation induces calcium-dependent up-regulation of neuregulin-1β in dystrophic skeletal muscle cell lines.

    PubMed

    Juretić, Nevenka; Jorquera, Gonzalo; Caviedes, Pablo; Jaimovich, Enrique; Riveros, Nora

    2012-01-01

    Duchenne muscular dystrophy (DMD) is a neuromuscular disease originated by reduced or no expression of dystrophin, a cytoskeletal protein that provides structural integrity to muscle fibres. A promising pharmacological treatment for DMD aims to increase the level of a structural dystrophin homolog called utrophin. Neuregulin-1 (NRG-1), a growth factor that potentiates myogenesis, induces utrophin expression in skeletal muscle cells. Microarray analysis of total gene expression allowed us to determine that neuregulin-1β (NRG-1β) is one of 150 differentially expressed genes in electrically stimulated (400 pulses, 1 ms, 45 Hz) dystrophic human skeletal muscle cells (RCDMD). We investigated the effect of depolarization, and the involvement of intracellular Ca(2+) and PKC isoforms on NRG-1β expression in dystrophic myotubes. Electrical stimulation of RCDMD increased NRG-1β mRNA and protein levels, and mRNA enhancement was abolished by actinomycin D. NRG-1β transcription was inhibited by BAPTA-AM, an intracellular Ca(2+) chelator, and by inhibitors of IP(3)-dependent slow Ca(2+) transients, like 2-APB, Ly 294002 and Xestospongin B. Ryanodine, a fast Ca(2+) signal inhibitor, had no effect on electrical stimulation-induced expression. BIM VI (general inhibitor of PKC isoforms) and Gö 6976 (specific inhibitor of Ca(2+)-dependent PKC isoforms) abolished NRG-1β mRNA induction. Our results suggest that depolarization induced slow Ca(2+) signals stimulate NRG-1β transcription in RCDMD cells, and that Ca(2+)-dependent PKC isoforms are involved in this process. Based on utrophin's ability to partially compensate dystrophin disfunction, knowledge on the mechanism involved on NRG-1 up-regulation could be important for new therapeutic strategies design. PMID:22613991

  10. Up-regulation of inducible nitric oxide synthase expression in cancer-prone p53 knockout mice.

    PubMed

    Ambs, S; Ogunfusika, M O; Merriam, W G; Bennett, W P; Billiar, T R; Harris, C C

    1998-07-21

    High concentrations of nitric oxide (NO) cause DNA damage and apoptosis in many cell types. Thus, regulation of NO synthase (NOS) activity is essential for minimizing effects of cytotoxic and genotoxic nitrogen oxide species. We have shown previously that NO-induced p53 protein accumulation down-regulates basal and cytokine-modulated inducible NOS (NOS2) expression in human cells in vitro. To further characterize the feedback loop between NOS2 and p53, we have investigated NO production, i.e., urinary nitrate plus nitrite excretion, and NOS2 expression in homozygous p53 knockout (KO) mice. We report here that untreated p53 KO mice excreted 70% more nitrite plus nitrate than mice with wild-type (wt) p53. NOS2 protein expression was constitutively detected in the spleen of untreated p53 KO mice, whereas it was undetectable in the spleen of wt p53 controls. Upon treatment with heat-inactivated Corynebacterium parvum, urinary nitrite plus nitrate excretion of p53 KO mice exceeded that of wt controls by approximately 200%. C. parvum treatment also induced p53 accumulation in the liver. Splenectomy reduced the NO output of C. parvum-treated p53 KO mice but not of wt p53 controls. Although NO production and NOS2 protein expression were increased similarly in KO and wt p53 mice 10 days after injection of C. parvum, NOS2 expression returned to baseline levels only in wt p53 controls while remaining up-regulated in p53 KO mice. These genetic and functional data indicate that p53 is an important transrepressor of NOS2 expression in vivo and attenuates excessive NO production in a regulatory negative feedback loop. PMID:9671763

  11. Rg1 protects the MPP+-treated MES23.5 cells via attenuating DMT1 up-regulation and cellular iron uptake.

    PubMed

    Xu, Huamin; Jiang, Hong; Wang, Jun; Xie, Junxia

    2010-02-01

    Ginsenoside-Rg1 is one of the pharmacologically active component isolated from ginseng. Our previous study observed the protective effect of Rg1 on iron accumulation in the substantia nigra (SN) in 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP)-treated Parkinson's disease (PD) mice. However, the mechanisms of this neuroprotective effect of Rg1 are unknown. In this study, we elucidated possible mechanisms for this effect using 1-methyl-4-phenylpyridinium (MPP(+))-treated MES23.5 cells. Previous study showed MPP+ treatment induced up-regulation of divalent metal transporter 1 without iron responsive element (DMT1-IRE) in MES23.5 cells. In the present study, we observed that pretreatment with Rg1 could inhibit MPP+-induced up-regulation of DMT1-IRE in MES23.5 cells. Up-regulation of DMT1-IRE by MPP+ treatment was associated with ROS production and translocation of nuclear factor-kappaB (NF-kappaB) to nuclei, both of which were significantly inhibited by Rg1 pretreatment. The role of ROS and NF-kappaB in the up-regulation of DMT1-IRE was supported by application of an antioxidant NAC and BAY 11-7082, an inhibitor of IkappaBalpha phosphorylation. Furthermore, we also showed Rg1 could decrease DMT1-mediated ferrous iron uptake and iron-induced cell damage by inhibiting the up-regulation of DMT1-IRE. These results indicate that Rg1 protected the MPP+-treated MES23.5 cells via attenuating DMT1-IRE up-regulation likely through inhibition of ROS-NF-kappaB pathway; Attenuation of DMT1-IRE expression decreased the iron influx and iron-induced oxidative stress.

  12. Association of CD69 Up-Regulation on CD4+ CLA+ T cells versus patch test, strip patch test and clinical history in nickel sensitization

    PubMed Central

    2010-01-01

    Objective The patch test (PT) with its modification - the strip patch test (SPT) - is the standard in vivo procedure to diagnose an allergic contact dermatitis (ACD). To date, none of the in vitro tests for the diagnosis of ACD fulfils the requirements of an easy, valid and reliable test. To investigate the prediction ability of a flow cytometric assay of CD69 up-regulation on CD4+ CLA+ T cells in nickel-sensitive and non-nickel-sensitive patients. Methods In a prospective, investigator-blinded, clinical study a total of 85 nickel-sensitive (n = 44; 51.8%) and non-nickel-sensitive patients (n = 41; 48.2%) were enrolled. The association between CD69 up-regulation on CD4+ CLA+ T cells on the one hand and PT, SPT, and clinical history on the other hand was measured. Association is expressed with c statistic values (receiver operating characteristic analysis) and corresponding 95% CIs. Results The associations were c = 0.57 (95% CI: 0.42-0.72) between CD69 up-regulation and PT, c = 0.49 (95% CI: 0.36-0.62) between CD69 up-regulation and SPT, and c = 0.51 (95% CI: 0.37-0.64) between CD69 up-regulation and clinical history. Conclusions CD69 up-regulation on CD4+ CLA+ T cells in vitro could not predict neither a positive PT or SPT result nor a positive clinical history to nickel sulfate. The combination of clinical history and patch testing still remains the basis for diagnosing ACD. PMID:20696642

  13. COORDINATED AV.

    ERIC Educational Resources Information Center

    CLEAVES, PAUL C.; AND OTHERS

    THE INSTRUCTIONAL MATERIALS CENTER IS LOCATED IN THE LOCAL HIGH SCHOOL AND SUPPLIES ALL SCHOOLS IN THE AREA. AUDIOVISUAL EQUIPMENT ORDERS, AFTER SELECTIONS ARE MADE BY THE CLASSROOM TEACHER, ARE PROCESSED BY THE CENTER, CONFIRMED AND DELIVERED BY TRUCK THREE TIMES EACH WEEK. EACH SCHOOL HAS A BUILDING COORDINATOR WHO CHECKS THE ORDERS INTO THE…

  14. Functional up-regulation of Nav1.8 sodium channel in Aβ afferent fibers subjected to chronic peripheral inflammation

    PubMed Central

    2014-01-01

    Background Functional alterations in the properties of Aβ afferent fibers may account for the increased pain sensitivity observed under peripheral chronic inflammation. Among the voltage-gated sodium channels involved in the pathophysiology of pain, Nav1.8 has been shown to participate in the peripheral sensitization of nociceptors. However, to date, there is no evidence for a role of Nav1.8 in controlling Aβ-fiber excitability following persistent inflammation. Methods Distribution and expression of Nav1.8 in dorsal root ganglia and sciatic nerves were qualitatively or quantitatively assessed by immunohistochemical staining and by real time-polymerase chain reaction at different time points following complete Freund’s adjuvant (CFA) administration. Using a whole-cell patch-clamp configuration, we further determined both total INa and TTX-R Nav1.8 currents in large-soma dorsal root ganglia (DRG) neurons isolated from sham or CFA-treated rats. Finally, we analyzed the effects of ambroxol, a Nav1.8-preferring blocker on the electrophysiological properties of Nav1.8 currents and on the mechanical sensitivity and inflammation of the hind paw in CFA-treated rats. Results Our findings revealed that Nav1.8 is up-regulated in NF200-positive large sensory neurons and is subsequently anterogradely transported from the DRG cell bodies along the axons toward the periphery after CFA-induced inflammation. We also demonstrated that both total INa and Nav1.8 peak current densities are enhanced in inflamed large myelinated Aβ-fiber neurons. Persistent inflammation leading to nociception also induced time-dependent changes in Aβ-fiber neuron excitability by shifting the voltage-dependent activation of Nav1.8 in the hyperpolarizing direction, thus decreasing the current threshold for triggering action potentials. Finally, we found that ambroxol significantly reduces the potentiation of Nav1.8 currents in Aβ-fiber neurons observed following intraplantar CFA injection and

  15. α-Hispanolol sensitizes hepatocellular carcinoma cells to TRAIL-induced apoptosis via death receptor up-regulation

    SciTech Connect

    Mota, Alba; Jiménez-Garcia, Lidia; Herránz, Sandra; Heras, Beatriz de las; Hortelano, Sonsoles

    2015-08-01

    Hispanolone derivatives have been previously described as anti-inflammatory and antitumoral agents. However, their effects on overcoming Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) resistance remain to be elucidated. In this study, we analyzed the cytotoxic effects of the synthetic hispanolone derivative α-hispanolol (α-H) in several tumor cell lines, and we evaluated the induction of apoptosis, as well as the TRAIL-sensitizing potential of α-H in the hepatocellular carcinoma cell line HepG2. Our data show that α-H decreased cell viability in a dose-dependent manner in HeLa, MDA-MB231, U87 and HepG2 cell lines, with a more prominent effect in HepG2 cells. Interestingly, α-H had no effect on non-tumoral cells. α-H induced activation of caspase-8 and caspase-9 and also increased levels of the proapoptotic protein Bax, decreasing antiapoptotic proteins (Bcl-2, X-IAP and IAP-1) in HepG2 cells. Specific inhibition of caspase-8 abrogated the cascade of caspase activation, suggesting that the extrinsic pathway has a critical role in the apoptotic events induced by α-H. Furthermore, combined treatment of α-H with TRAIL enhanced apoptosis in HepG2 cells, activating caspase-8 and caspase-9. This correlated with up-regulation of both the TRAIL death receptor DR4 and DR5. DR4 or DR5 neutralizing antibodies abolished the effect of α-H on TRAIL-induced apoptosis, suggesting that sensitization was mediated through the death receptor pathway. Our results demonstrate that α-H induced apoptosis in the human hepatocellular carcinoma cell line HepG2 through activation of caspases and induction of the death receptor pathway. In addition, we describe a novel function of α-H as a sensitizer on TRAIL-induced apoptotic cell death in HepG2 cells. - Highlights: • α-Hispanolol induced apoptosis in the human hepatocellular carcinoma cell line HepG2. • α-Hispanolol induced activation of caspases and the death receptor pathway. • α-Hispanolol enhanced

  16. Cysteamine-induced depletion of brain somatostatin is associated with up-regulation of cerebrocortical somatostatin receptors

    SciTech Connect

    Srikant, C.B.; Patel, Y.C.

    1984-09-01

    Cysteamine (CSH) administered as a single sc injection to rats produced rapid depletion of cerebrocortical Somatostatin-14 like immunoreactivity (S-14 LI) with a significant 48% reduction occurring within 5 min and maximum (72%) decrease at 4 h. The depletion of S-14 LI was associated with a 1.7 fold increase in Bmax of the cerebrocortical S-14 receptors 5 min after CSH administration and a concomitant but slower increase in the affinity of these receptors. Incubation of intact synaptosomes with 1 mM CSH at 37 C in vitro for 60 min also caused a rapid depletion of S-14 LI, but there was no change in the Bmax or Kd of the S-14 receptors for up to 30 min beyond which time a 2.8-fold decrease in the affinity of S-14 receptors was observed. Higher concentrations of CSH (greater than or equal to 10 mM) added during the incubation of synaptosomes in vitro completely abolished the specific binding of these receptors. The pituitary S-14 receptors were studied 30 min after CSH administration and unlike the cerebrocortical S-14 receptors at this time did not exhibit any change in Bmax or affinity. When added at the time of the binding assay CSH (1 mM) was without a direct effect on cerebrocortical as well as pituitary membrane S-14 receptors. Furthermore, addition of CSH at the time of binding assay did not destroy the integrity of (/sup 125/I-Tyr11)S-14. It is concluded that administration of CSH to rats in vivo depletes brain S-14 LI and up-regulates synaptosomal S-14 receptors. Exposure of synaptosomes to CSH in vitro for 30 min also depletes S-14 LI but has no effect on S-14 receptors. CSH has a direct inhibitory effect on S-14 receptor binding after prolonged in vitro incubation. Pituitary S-14 receptors unlike those in the brain are unaffected by S-14 LI depletion at least acutely.

  17. Cysteamine-induced depletion of brain somatostatin is associated with up-regulation of cerebrocortical somatostatin receptors.

    PubMed

    Srikant, C B; Patel, Y C

    1984-09-01

    Cysteamine (CSH) administered as a single sc injection to rats produced rapid depletion of cerebrocortical Somatostatin-14 like immunoreactivity (S-14 LI) with a significant 48% reduction occurring within 5 min and maximum (72%) decrease at 4 h. The depletion of S-14 LI was associated with a 1.7 fold increase in Bmax of the cerebrocortical S-14 receptors 5 min after CSH administration and a concomitant but slower increase in the affinity of these receptors [dissociation constant (Kd) being 1.8- and 1.6-fold lower than the control at 30 and 60 min, respectively, post CSH]. Incubation of intact synaptosomes with 1 mM CSH at 37 C in vitro for 60 min also caused a rapid depletion of S-14 LI, but in contrast to the in vivo data, there was no change in the Bmax or Kd of the S-14 receptors for up to 30 min beyond which time a 2.8-fold decrease in the affinity of S-14 receptors was observed. Higher concentrations of CSH (greater than or equal to 10 mM) added during the incubation of synaptosomes in vitro completely abolished the specific binding of these receptors. The pituitary S-14 receptors were studied 30 min after CSH administration and unlike the cerebrocortical S-14 receptors at this time did not exhibit any change in Bmax or affinity. When added at the time of the binding assay CSH (1 mM) was without a direct effect on cerebrocortical as well as pituitary membrane S-14 receptors. Furthermore, addition of CSH at the time of binding assay did not destroy the integrity of [125I-Tyr11]S-14. It is concluded that administration of CSH to rats in vivo depletes brain S-14 LI and up-regulates synaptosomal S-14 receptors. Exposure of synaptosomes to CSH in vitro for 30 min also depletes S-14 LI but has no effect on S-14 receptors suggesting that S-14 receptor regulation by S-14 is an in vivo phenomenon or requires the intact cell. CSH has a direct inhibitory effect on S-14 receptor binding after prolonged in vitro incubation. Pituitary S-14 receptors unlike those in the

  18. MiR-210 Up-Regulation Inhibits Proliferation and Induces Apoptosis in Glioma Cells by Targeting SIN3A

    PubMed Central

    Shang, Chao; Hong, Yang; Guo, Yan; Liu, Yun-hui; Xue, Yi-xue

    2014-01-01

    Background The aim of this study was to determine whether miR-210 can affect the apoptosis and proliferation of human U251 glioma cells from down-regulating SIN3A. Material/Methods The expression of miRNA-210 was detected by quantitative real-time PCR in normal brain tissue and glioma samples. The apoptosis and proliferation ability of U251 cells were analyzed by MTT and flow cytometry assay after anti-miR-210 transfection. For the regulation mechanism analysis of miR-210, TargetScan, PicTar, and microRNA were selected to predict some potential target genes of miR-210. The predicted gene was identified to be the direct and specific target gene of miR-210 by luciferase activities assay and Western blot. RNA interference technology was used to confirm that the apoptosis and proliferation effects of miR-210 were directly induced by SIN3A. Results The expression of miR-210 increased significantly in glioma in comparison with normal brain tissue. The silence of miR-210 expression could inhibit the proliferation of U251 cells and induce the apoptosis. Mechanism analysis revealed that SIN3A was a specific and direct target gene of miR-210. The siRNA-SIN3A could down-regulate the expression of SIN3A protein, which was up-regulated in U251 cells by anti-miR-210 transfection, and our experiments found that silence of SIN3A could inhibit the apoptosis and sharply increase the proliferation of U251 cells. The regulation effects of anti-miR-210 on apoptosis and proliferation can be reversed respectively by the expression silence of SIN3A. Conclusions Aberrantly expressed miR-210 regulates human U251 glioma cells apoptosis and proliferation partly through directly down-regulating SIN3A protein expression. This might offer a new potential therapeutic stratagem for glioma. PMID:25481483

  19. Beta-Adrenergic Receptor Population is Up-Regulated in Chicken Skeletal Muscle Cells Treated with Forskolin

    NASA Technical Reports Server (NTRS)

    Bridge, K. Y.; Young, R. B.; Vaughn, J. R.

    1998-01-01

    Skeletal muscle hypertrophy is promoted by in vivo administration of beta-adrenergic receptor (betaAR) agonists. These compounds presumably exert their physiological action through the betaAR, and alterations in the population of betaAR could potentially change the ability of the cell to respond to the betaAR agonists. Since the intracellular chemical signal generated by the betaAR is cyclic AMP (cAMP), experiments were initiated in primary chicken muscle cell cultures to determine if artificial elevation of intracellular cAMP by treatment with forskolin would alter the population of functional betaAR expressed on the surface of muscle cells. Chicken skeletal muscle cells after 7 days in culture were employed for the experiments because muscle cells have attained a steady state with respect to muscle protein metabolism at this stage. Cells were treated with 0-10 microM forskolin for a total of three days. At the end of the 1, 2, and 3 day treatment intervals, the concentration of cAMP and the betaAR population were measured. Receptor population was measured in intact muscle cell cultures as the difference between total binding of [H-3]CGP-12177 and non-specific binding of [H-3]CGP-12177 in the presence of 1 microM propranolol. Intracellular cAMP concentration was measured by radioimmunoassay. The concentration of cAMP in forskolin-treated cells increased up to 10-fold in a dose dependent manner. Increasing concentrations of forskolin also led to an increase in betaAR population, with a maximum increase of approximately 50% at 10 microM. This increase in PAR population was apparent after only 1 day of treatment, and the pattern of increase was maintained for all 3 days of the treatment period. Thus, increasing the intracellular concentration of cAMP leads to up-regulation of betaAR population. The effect of forskolin on the quantity and apparent synthesis rate of the heavy chain of myosin (mhc) were also investigated. A maximum increase of 50% in the quantity of mhc

  20. Up-regulation of miR-26a promotes neurite outgrowth and ameliorates apoptosis by inhibiting PTEN in bupivacaine injured mouse dorsal root ganglia.

    PubMed

    Cui, Changlei; Xu, Gong; Qiu, Jinpeng; Fan, Xiushuang

    2015-08-01

    Local anesthetic of bupivacaine may inhibit neurite outgrowth and induce apoptosis in mouse dorsal root ganglia (DRG) neurons. In this work, we intended to investigate the functional role of microRNA 26a (miR-26a) in regulating bupivacaine-induced nerve injury in DRG neurons. DRG neurons were extracted from C57BL/6 mice and cultured in vitro. Bupivacaine was applied in vitro and it induced apoptosis, inhibited neurite growth, and significantly down-regulated miR-26a gene in DRG neurons. MiR-26a mimic was then used to up-regulate miR-26a expression in DRG neurons. We found that miR-26a up-regulation promoted neurite outgrowth and reduced apoptosis in bupivacaine-injured DRG neurons. Luciferase assay and Western blot confirmed that Phosphatase and tensin homolog (PTEN) was down-stream target of miR-26a in DRG neurons. Ectopic PTEN up-regulation was then able to reverse the protective effect of miR-26a overexpression on bupivacaine-induced nerve injury in DRG neurons. Overall, this work demonstrated that miR-26a had a functional role in regulating bupivacaine-induced nerve injury in DRG neurons. Up-regulating miR-26a to suppress PTEN signaling pathway may be an effective method to protect local anesthetic-induced nerve injury in spinal cord.

  1. Activation of stress-activated MAP protein kinases up-regulates expression of transgenes driven by the cytomegalovirus immediate/early promoter.

    PubMed Central

    Bruening, W; Giasson, B; Mushynski, W; Durham, H D

    1998-01-01

    The immediate/early promoter/enhancer of cytomegalovirus (CMV promoter) is one of the most commonly used promoters for expression of transgenes in eukaryotic cells. In practice, the CMV promoter is often thought of as a constitutively active unregulated promoter. However, we have observed that transcription from the CMV promoter can be up-regulated by a variety of environmental stresses. Many forms of cellular stress stimulate MAP kinase signalling pathways, resulting in activation of stress-activated protein kinases [SAPKs, also called Jun N-terminal kinases (JNKs)] and p38 kinases. We have found that the same conditions that lead to activation of SAPK/JNKs and p38 kinases can also dramatically increase expression from the CMV promoter. Inhibitors of p38 kinases abolished basal transcription from the CMV promoter and completely blocked stress-induced up-regulation of the CMV promoter. Overexpression of a dominant negative JNK kinase had no effect on basal transcription, but significantly reduced up-regulation caused by stress. These results have grave implications for use of the CMV promoter. If the CMV promoter can be up-regulated by cellular stresses, inadvertent activation of the stress kinase pathways may complicate, if not invalidate, the interpretation of a wide range of experiments. PMID:9421504

  2. Chloroquine enhances TRAIL-mediated apoptosis through up-regulation of DR5 by stabilization of mRNA and protein in cancer cells

    PubMed Central

    Park, Eun Jung; Min, Kyoung-jin; Choi, Kyeong Sook; Kubatka, Peter; Kruzliak, Peter; Kim, Dong Eun; Kwon, Taeg Kyu

    2016-01-01

    Chloroquine (CQ), an anti-malarial drug, has immune-modulating activity and lysosomotropic activity. In this study, we investigated CQ sensitizes TRAIL-mediated apoptosis in human renal cancer Caki cells. Combination of CQ and TRAIL significantly induces apoptosis in human renal cancer Caki cells and various human cancer cells, but not in normal mouse kidney cells (TMCK-1) and human mesangial cells (MC). CQ up-regulates DR5 mRNA and protein expression in a dose- and time- dependent manner. Interestingly, CQ regulates DR5 expression through the increased stability in the mRNA and protein of DR5, rather than through the increased transcriptional activity of DR5. Moreover, we found that CQ decreased the expression of Cbl, an E3 ligase of DR5, and knock-down of Cbl markedly enhanced DR5 up-regulation. Other lysosomal inhibitors, including monensin and nigericin, also up-regulated DR5 and sensitized TRAIL-mediated apoptosis. Therefore, this study demonstrates that lysosomal inhibition by CQ may sensitize TRAIL-mediated apoptosis in human renal cancer Caki cells via DR5 up-regulation. PMID:26964637

  3. Chloroquine enhances TRAIL-mediated apoptosis through up-regulation of DR5 by stabilization of mRNA and protein in cancer cells.

    PubMed

    Park, Eun Jung; Min, Kyoung-jin; Choi, Kyeong Sook; Kubatka, Peter; Kruzliak, Peter; Kim, Dong Eun; Kwon, Taeg Kyu

    2016-01-01

    Chloroquine (CQ), an anti-malarial drug, has immune-modulating activity and lysosomotropic activity. In this study, we investigated CQ sensitizes TRAIL-mediated apoptosis in human renal cancer Caki cells. Combination of CQ and TRAIL significantly induces apoptosis in human renal cancer Caki cells and various human cancer cells, but not in normal mouse kidney cells (TMCK-1) and human mesangial cells (MC). CQ up-regulates DR5 mRNA and protein expression in a dose- and time- dependent manner. Interestingly, CQ regulates DR5 expression through the increased stability in the mRNA and protein of DR5, rather than through the increased transcriptional activity of DR5. Moreover, we found that CQ decreased the expression of Cbl, an E3 ligase of DR5, and knock-down of Cbl markedly enhanced DR5 up-regulation. Other lysosomal inhibitors, including monensin and nigericin, also up-regulated DR5 and sensitized TRAIL-mediated apoptosis. Therefore, this study demonstrates that lysosomal inhibition by CQ may sensitize TRAIL-mediated apoptosis in human renal cancer Caki cells via DR5 up-regulation. PMID:26964637

  4. INFLUENZA-INDUCED UP-REGULATION OF TLR3 IN RESPIRATORY EPITHELIAL CELLS MAY OCCUR THROUGH A POSITIVE FEEDBACK LOOP INVOLVING TYPE I INTERFERON

    EPA Science Inventory

    Toll-like receptor 3 (TLR3) plays an important role in the host defense responses against viral infections, including Influenza virus infections. Based on our previous observations showing that Influenza infection of respiratory epithelial cells results in an up-regulation of Tol...

  5. Elevation of Highly Up-regulated in Liver Cancer (HULC) by Hepatitis B Virus X Protein Promotes Hepatoma Cell Proliferation via Down-regulating p18*

    PubMed Central

    Du, Yumei; Kong, Guangyao; You, Xiaona; Zhang, Shuai; Zhang, Tao; Gao, Yuen; Ye, Lihong; Zhang, Xiaodong

    2012-01-01

    Long noncoding RNAs (lncRNAs) play crucial roles in human cancers. It has been reported that lncRNA highly up-regulated in liver cancer (HULC) is dramatically up-regulated in hepatocellular carcinoma (HCC). Hepatitis B virus X protein (HBx) contributes importantly to the development of HCC. However, the function of HULC in HCC mediated by HBx remains unclear. Here, we report that HULC is involved in HBx-mediated hepatocarcinogenesis. We found that the expression levels of HULC were positively correlated with those of HBx in clinical HCC tissues. Moreover, we revealed that HBx up-regulated HULC in human immortalized normal liver L-O2 cells and hepatoma HepG2 cells. Luciferase reporter gene assay and chromatin immunoprecipitation (ChIP) assay showed that HBx activated the HULC promoter via cAMP-responsive element-binding protein. We further demonstrated that HULC promoted cell proliferation by methyl thiazolyl tetrazolium, 5-ethynyl-2′-deoxyuridine, colony formation assay, and tumorigenicity assay. Next, we hypothesized that HULC might function through regulating a tumor suppressor gene p18 located near HULC in the same chromosome. We found that the mRNA levels of p18 were inversely correlated with those of HULC in the above clinical HCC specimens. Then, we validated that HULC down-regulated p18, which was involved in the HULC-enhanced cell proliferation in vitro and in vivo. Furthermore, we observed that knockdown of HULC could abolish the HBx-enhanced cell proliferation through up-regulating p18. Thus, we conclude that the up-regulated HULC by HBx promotes proliferation of hepatoma cells through suppressing p18. This finding provides new insight into the roles of lncRNAs in HBx-related hepatocarcinogenesis. PMID:22685290

  6. Synergistic effect of interleukin 1 alpha on nontypeable Haemophilus influenzae-induced up-regulation of human beta-defensin 2 in middle ear epithelial cells

    PubMed Central

    Moon, Sung-Kyun; Lee, Haa-Yung; Pan, Huiqi; Takeshita, Tamotsu; Park, Raekil; Cha, Kiweon; Andalibi, Ali; Lim, David J

    2006-01-01

    Background We recently showed that beta-defensins have antimicrobial activity against nontypeable Haemophilus influenzae (NTHi) and that interleukin 1 alpha (IL-1 alpha) up-regulates the transcription of beta-defensin 2 (DEFB4 according to new nomenclature of the Human Genome Organization) in human middle ear epithelial cells via a Src-dependent Raf-MEK1/2-ERK signaling pathway. Based on these observations, we investigated if human middle ear epithelial cells could release IL-1 alpha upon exposure to a lysate of NTHi and if this cytokine could have a synergistic effect on beta-defensin 2 up-regulation by the bacterial components. Methods The studies described herein were carried out using epithelial cell lines as well as a murine model of acute otitis media (OM). Human cytokine macroarray analysis was performed to detect the released cytokines in response to NTHi exposure. Real time quantitative PCR was done to compare the induction of IL-1 alpha or beta-defensin 2 mRNAs and to identify the signaling pathways involved. Direct activation of the beta-defensin 2 promoter was monitored using a beta-defensin 2 promoter-Luciferase construct. An IL-1 alpha blocking antibody was used to demonstrate the direct involvement of this cytokine on DEFB4 induction. Results Middle ear epithelial cells released IL-1 alpha when stimulated by NTHi components and this cytokine acted in an autocrine/paracrine synergistic manner with NTHi to up-regulate beta-defensin 2. This synergistic effect of IL-1 alpha on NTHi-induced beta-defensin 2 up-regulation appeared to be mediated by the p38 MAP kinase pathway. Conclusion We demonstrate that IL-1 alpha is secreted by middle ear epithelial cells upon exposure to NTHi components and that it can synergistically act with certain of these molecules to up-regulate beta-defensin 2 via the p38 MAP kinase pathway. PMID:16433908

  7. SARS coronavirus papain-like protease induces Egr-1-dependent up-regulation of TGF-β1 via ROS/p38 MAPK/STAT3 pathway.

    PubMed

    Li, Shih-Wein; Wang, Ching-Ying; Jou, Yu-Jen; Yang, Tsuey-Ching; Huang, Su-Hua; Wan, Lei; Lin, Ying-Ju; Lin, Cheng-Wen

    2016-01-01

    SARS coronavirus (SARS-CoV) papain-like protease (PLpro) has been identified in TGF-β1 up-regulation in human promonocytes (Proteomics 2012, 12: 3193-205). This study investigates the mechanisms of SARS-CoV PLpro-induced TGF-β1 promoter activation in human lung epithelial cells and mouse models. SARS-CoV PLpro dose- and time-dependently up-regulates TGF-β1 and vimentin in A549 cells. Dual luciferase reporter assays with TGF-β1 promoter plasmids indicated that TGF-β1 promoter region between -175 to -60, the Egr-1 binding site, was responsible for TGF-β1 promoter activation induced by SARS-CoV PLpro. Subcellular localization analysis of transcription factors showed PLpro triggering nuclear translocation of Egr-1, but not NF-κB and Sp-1. Meanwhile, Egr-1 silencing by siRNA significantly reduced PLpro-induced up-regulation of TGF-β1, TSP-1 and pro-fibrotic genes. Furthermore, the inhibitors for ROS (YCG063), p38 MAPK (SB203580), and STAT3 (Stattic) revealed ROS/p38 MAPK/STAT3 pathway involving in Egr-1 dependent activation of TGF-β1 promoter induced by PLpro. In a mouse model with a direct pulmonary injection, PLpro stimulated macrophage infiltration into lung, up-regulating Egr-1, TSP-1, TGF-β1 and vimentin expression in lung tissues. The results revealed that SARS-CoV PLpro significantly triggered Egr-1 dependent activation of TGF-β1 promoter via ROS/p38 MAPK/STAT3 pathway, correlating with up-regulation of pro-fibrotic responses in vitro and in vivo. PMID:27173006

  8. Transcriptional Up-Regulation of APE1/Ref-1 in Hepatic Tumor: Role in Hepatocytes Resistance to Oxidative Stress and Apoptosis

    PubMed Central

    Di Maso, Vittorio; Mediavilla, María Gabriela; Vascotto, Carlo; Lupo, Francesco; Baccarani, Umberto; Avellini, Claudio; Tell, Gianluca; Tiribelli, Claudio; Crocè, Lory Saveria

    2015-01-01

    Objective Human Hepatocellular Carcinoma (HCC) is the fifth most frequent neoplasm worldwide and the most serious complication of long-standing chronic liver diseases (CLD). Its development is associated with chronic inflammation and sustained oxidative stress. Deregulation of apurinic apyrimidinic endonuclease 1/redox effector factor 1 (APE1/Ref-1), a master regulator of cellular response to oxidative stress, has been associated with poor prognosis in several cancers including HCC. Design In the present study we investigated the APE1/Ref-1 mRNA levels in cirrhotic and HCC tissues obtained during HCC resection. The possible protective role of APE1/Ref-1 against oxidative stress and apoptosis was evaluated in vitro in immortalized human hepatocytes (IHH) over-expressing APE1/Ref-1. Results APE1/Ref-1 was up-regulated in HCC, regulation occurring at the transcriptional level. APE1/Ref-1 mRNA content increased with the progression of liver disease with the transcriptional up-regulation present in cirrhosis significantly increased in HCC. The up-regulation was higher in the less differentiated cancers. In vitro, over-expression of APE1/Ref-1 in normal hepatocytes conferred cell protection against oxidative stress and it was associated with BAX inhibition and escape from apoptosis. Conclusion APE1/Ref-1 is up-regulated in HCC and this over-expression correlates with cancer aggressiveness. The up-regulation occurs at the transcriptional level and it is present in the earliest phases of hepatocarcinogenesis. The APE-1/Ref-1 over-expression is associated with hepatocyte survival and inhibits BAX activation and apoptosis. These data suggest a possible role of APE1/Ref-1 over-expression both in hepatocyte survival and HCC development calling attention to this molecule as a promising marker for HCC diagnosis and treatment. PMID:26624999

  9. Cyanide-induced death of dopaminergic cells is mediated by uncoupling protein-2 up-regulation and reduced Bcl-2 expression

    SciTech Connect

    Zhang, X.; Li, L.; Zhang, L.; Borowitz, J.L.; Isom, G.E.

    2009-07-01

    Cyanide is a potent inhibitor of mitochondrial oxidative metabolism and produces mitochondria-mediated death of dopaminergic neurons and sublethal intoxications that are associated with a Parkinson-like syndrome. Cyanide toxicity is enhanced when mitochondrial uncoupling is stimulated following up-regulation of uncoupling protein-2 (UCP-2). In this study, the role of a pro-survival protein, Bcl-2, in cyanide-mediated cell death was determined in a rat dopaminergic immortalized mesencephalic cell line (N27 cells). Following pharmacological up-regulation of UCP-2 by treatment with Wy14,643, cyanide reduced cellular Bcl-2 expression by increasing proteasomal degradation of the protein. The increased turnover of Bcl-2 was mediated by an increase of oxidative stress following UCP-2 up-regulation. The oxidative stress involved depletion of mitochondrial glutathione (mtGSH) and increased H{sub 2}O{sub 2} generation. Repletion of mtGSH by loading cells with glutathione ethyl ester reduced H{sub 2}O{sub 2} generation and in turn blocked the cyanide-induced decrease of Bcl-2. To determine if UCP-2 mediated the response, RNAi knock down was conducted. The RNAi decreased cyanide-induced depletion of mtGSH, reduced H{sub 2}O{sub 2} accumulation, and inhibited down-regulation of Bcl-2, thus blocking cell death. To confirm the role of Bcl-2 down-regulation in the cell death, it was shown that over-expression of Bcl-2 by cDNA transfection attenuated the enhancement of cyanide toxicity after UCP-2 up-regulation. It was concluded that UCP-2 up-regulation sensitizes cells to cyanide by increasing cellular oxidative stress, leading to an increase of Bcl-2 degradation. Then the reduced Bcl-2 levels sensitize the cells to cyanide-mediated cell death.

  10. P2Y2 nucleotide receptor activation up-regulates vascular cell adhesion molecule-1 [corrected] expression and enhances lymphocyte adherence to a human submandibular gland cell line.

    PubMed

    Baker, Olga J; Camden, Jean M; Rome, Danny E; Seye, Cheikh I; Weisman, Gary A

    2008-01-01

    Sjögren's syndrome (SS) is a chronic inflammatory autoimmune disease that causes salivary and lacrimal gland tissue destruction resulting in impaired secretory function. Although lymphocytic infiltration of salivary epithelium is associated with SS, the mechanisms involved have not been adequately elucidated. Our previous studies have shown that the G protein-coupled P2Y2 nucleotide receptor (P2Y2R) is up-regulated in response to damage or stress of salivary gland epithelium, and in salivary glands of the NOD.B10 mouse model of SS-like autoimmune exocrinopathy. Additionally, we have shown that P2Y2R activation up-regulates vascular cell adhesion molecule-1 (VCAM-1) expression in endothelial cells leading to the binding of monocytes. The present study demonstrates that activation of the P2Y2R in dispersed cell aggregates from rat submandibular gland (SMG) and in human submandibular gland ductal cells (HSG) up-regulates the expression of VCAM-1. Furthermore, P2Y2R activation mediated the up-regulation of VCAM-1 expression in HSG cells leading to increased adherence of lymphocytic cells. Inhibitors of EGFR phosphorylation and metalloprotease activity abolished P2Y2R-mediated VCAM-1 expression and decreased lymphocyte binding to HSG cells. Moreover, silencing of EGFR expression abolished UTP-induced VCAM-1 up-regulation in HSG cells. These results suggest that P2Y2R activation in salivary gland cells increases the EGFR-dependent expression of VCAM-1 and the binding of lymphocytes, a pathway relevant to inflammation associated with SS.

  11. P2Y2 Nucleotide Receptor Activation Up-regulates Vascular Cell Adhesion Molecular-1 Expression and Enhances Lymphocyte Adherence to a Human Submandibular Gland Cell Line

    PubMed Central

    Baker, Olga J.; Camden, Jean M.; Rome, Danny E.; Seye, Cheikh I.; Weisman, Gary A.

    2007-01-01

    Sjögren’s syndrome (SS) is a chronic inflammatory autoimmune disease that causes salivary and lacrimal gland tissue destruction resulting in impaired secretory function. Although lymphocytic infiltration of salivary epithelium is associated with SS, the mechanisms involved have not been adequately elucidated. Our previous studies have shown that the G protein-coupled P2Y2 nucleotide receptor (P2Y2R) is up-regulated in response to damage or stress of salivary gland epithelium, and in salivary glands of the NOD.B10 mouse model of SS-like autoimmune exocrinopathy. Additionally, we have shown that P2Y2R activation up-regulates vascular cell adhesion molecule-1 (VCAM-1) expression in endothelial cells leading to the binding of monocytes. The present study demonstrates that activation of the P2Y2R in dispersed cell aggregates from rat submandibular gland (SMG) and in human submandibular gland ductal cells (HSG) up-regulates the expression of VCAM-1. Furthermore, P2Y2R activation mediated the up-regulation of VCAM-1 expression in HSG cells leading to increased adherence of lymphocytic cells. Inhibitors of EGFR phosphorylation and metalloprotease activity abolished P2Y2R-mediated VCAM-1 expression and decreased lymphocyte binding to HSG cells. Moreover, silencing of EGFR expression abolished UTP-induced VCAM-1 up-regulation in HSG cells. These results suggest that P2Y2R activation in salivary gland cells increases the EGFR-dependent expression of VCAM-1 and the binding of lymphocytes, a pathway relevant to inflammation associated with SS. PMID:17599409

  12. Interleukin-1 beta-induced up-regulation of opioid receptors in the untreated and morphine-desensitized U87 MG human astrocytoma cells

    PubMed Central

    2012-01-01

    Background Interleukin-1beta (IL-1β) is a pro-inflammatory cytokine that can be produced in the central nervous system during inflammatory conditions. We have previously shown that IL-1β expression is altered in the rat brain during a morphine tolerant state, indicating that this cytokine may serve as a convergent point between the immune challenge and opiate mediated biological pathways. We hypothesized that IL-1β up-regulates opioid receptors in human astrocytes in both untreated and morphine-desensitized states. Methods To test this hypothesis, we compared the basal expression of the mu (MOR), delta (DOR), and kappa (KOR) opioid receptors in the human U87 MG astrocytic cell line to SH-SY5Y neuronal and HL-60 immune cells using absolute quantitative real time RT-PCR (AQ-rt-RT-PCR). To demonstrate that IL-1β induced up-regulation of the MOR, DOR and KOR, U87 MG cells (2 x 105 cells/well) were treated with IL-1β (20 ng/mL or 40 ng/mL), followed by co-treatment with interleukin-1 receptor antagonist protein (IL-1RAP) (400 ng/mL or 400 ng/mL). The above experiment was repeated in the cells desensitized with morphine, where U87 MG cells were pre-treated with 100 nM morphine. The functionality of the MOR in U87 MG cells was then demonstrated using morphine inhibition of forksolin-induced intracellular cAMP, as determined by radioimmunoassay. Results U87 MG cells treated with IL-1β for 12 h showed a significant up-regulation of MOR and KOR. DOR expression was also elevated, although not significantly. Treatment with IL-1β also showed a significant up-regulation of the MOR in U87 MG cells desensitized with morphine. Co-treatment with IL-1β and interleukin-1 receptor antagonist protein (IL-1RAP) resulted in a significant decrease in IL-1β-mediated MOR up-regulation. Conclusion Our results indicate that the pro-inflammatory cytokine, IL-1β, affects opiate-dependent pathways by up-regulating the expression of the MOR in both untreated and morphine-desensitized U87

  13. Ursolic Acid Attenuates Diabetic Mesangial Cell Injury through the Up-Regulation of Autophagy via miRNA-21/PTEN/Akt/mTOR Suppression

    PubMed Central

    Lu, Xinxing; Fan, Qiuling; Xu, Li; Li, Lin; Yue, Yuan; Xu, Yanyan; Su, Yan; Zhang, Dongcheng; Wang, Lining

    2015-01-01

    Objective To investigate the effect of ursolic acid on autophagy mediated through the miRNA-21-targeted phosphoinositide 3 kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway in rat mesangial cells cultured under high glucose (HG) conditions. Methods Rat glomerular mesangial cells were cultured under normal glucose, HG, HG with the PI3K inhibitor LY294002 or HG with ursolic acid conditions. Cell proliferation and hypertrophy were assayed using an MTT assay and the ratio of total protein to cell number, respectively. The miRNA-21 expression was detected using RT-qPCR. The expression of phosphatase and tensin homolog (PTEN)/AKT/mTOR signaling signatures, autophagy-associated protein and collagen I was detected by western blotting and RT-qPCR. Autophagosomes were observed using electron microscopy. Results Compared with mesangial cells cultured under normal glucose conditions, the cells exposed to HG showed up-regulated miRNA-21 expression, down-regulated PTEN protein and mRNA expression, up-regulated p85PI3K, pAkt, pmTOR, p62/SQSTMI, and collagen I expression and down-regulated LC3II expression. Ursolic acid and LY294002 inhibited HG-induced mesangial cell hypertrophy and proliferation, down-regulated p85PI3K, pAkt, pmTOR, p62/SQSTMI, and collagen I expression and up-regulated LC3II expression. However, LY294002 did not affect the expression of miRNA-21 and PTEN. Ursolic acid down-regulated miRNA-21 expression and up-regulated PTEN protein and mRNA expression. Conclusions Ursolic acid inhibits the glucose-induced up-regulation of mesangial cell miRNA-21 expression, up-regulates PTEN expression, inhibits the activation of PI3K/Akt/mTOR signaling pathway, and enhances autophagy to reduce the accumulation of the extracellular matrix and ameliorate cell hypertrophy and proliferation. PMID:25689721

  14. Controlled bile acid exposure to oesophageal mucosa causes up-regulation of nuclear γ-H2AX possibly via iNOS induction

    PubMed Central

    Jiang, Bo; Zhao, Shengqian; Tao, Zhen; Wen, Jin; Yang, Yancheng; Zheng, Yin; Yan, Hongling; Sheng, Ying; Gao, Aimin

    2016-01-01

    Using an in vitro model in which flatmounts of oesophagus was periodically exposed to bile acids, we demonstrate, using multiple methods, that the bile acid receptor TGR5, inducible nitric oxide synthase (iNOS) and γ-histone family 2A variant (γ-H2AX) are up-regulated. This indicates that bile acids cause up-regulation of iNOS, which further causes genotoxic stress as evidenced by increase of the highly sensitive marker, phosphorylated histone. In vitro nitric oxide (NO) assays showed increased production of nitric acid in the oesophageal epithelium exposed to the bile acids. This increase was inhibited in the presence of the nonspecific iNOS inhibitor aminoguanidine (AG). Cumulatively, the results of the present study provide suggestion that not only acid reflux, but also non-acid reflux of bile may cause genotoxic stress. These aspects merit to be tested in wide spectrum of Barrett epithelial tissues. PMID:27247425

  15. Lectin purified from Musca domestica pupa up-regulates NO and iNOS production via TLR4/NF-κB signaling pathway in macrophages.

    PubMed

    Cao, Xiaohong; Zhou, Minghui; Wang, Chunling; Hou, Lihua; Zeng, Bin

    2011-04-01

    The present study reported that nitric oxide (NO) was up-regulated by the induction of lectin purified from Musca domestica pupa (MPL) in macrophages without cytotoxicity. The mRNA expression and protein secretion of inducible nitric oxide synthase (iNOS) were strongly induced by MPL treatments. Subsequent investigation revealed that the nuclear factor-κB (NF-κB) inhibitory κB (IκB) in endochylema was inhibited and NF-κB translocated into the nucleus after MPL treatment. Meanwhile, the IKKβ was strongly induced and the production of the toll-like receptor 4 (TLR4) was significantly up-regulated. Moreover, MPL increased NO production via inducing the expression of iNOS through the activation of NF-κB, which suggested that MPL probably acted as an activating agent of the NF-κB activation.

  16. The neuroprotection of hypoxic preconditioning on rat brain against traumatic brain injury by up-regulated transcription factor Nrf2 and HO-1 expression.

    PubMed

    Shu, Longfei; Wang, Chunlin; Wang, Jinbiao; Zhang, Yongming; Zhang, Xing; Yang, Yanyan; Zhuo, Jianwei; Liu, Jiachuan

    2016-01-12

    Hypoxic preconditioning (HPC) increases the inherent tolerance of brain tissue suffering from severe hypoxia or ischemia insult by stimulating the protective ability of the brain. However, little is known concerning the effect of HPC on traumatic brain injury (TBI). We designed this study to investigate the effect of HPC on TBI and explore its underlying mechanisms. We found that HPC significantly alleviates neurological dysfunction, lessens brain edema, reduces cell apoptosis, increases neuronal survival, up-regulates the expressions of Nrf2 and HO-1, and decreases the inducer of protein carbonyls, 4-hydroxy-2-nonenal, and 8-hydroxy-2-deoxyguanosine in the brain tissue of rats 24h after brain injury. However, no influence was observed in normal rats after only 3d of hypoxic training. Results further indicated that HPC protects the brain against traumatic damage. This protective effect may be achieved by up-regulating Nrf2 and HO-1 expression and alleviating oxidative stress damage. PMID:26590328

  17. Controlled bile acid exposure to oesophageal mucosa causes up-regulation of nuclear γ-H2AX possibly via iNOS induction.

    PubMed

    Jiang, Bo; Zhao, Shengqian; Tao, Zhen; Wen, Jin; Yang, Yancheng; Zheng, Yin; Yan, Hongling; Sheng, Ying; Gao, Aimin

    2016-08-01

    Using an in vitro model in which flatmounts of oesophagus was periodically exposed to bile acids, we demonstrate, using multiple methods, that the bile acid receptor TGR5, inducible nitric oxide synthase (iNOS) and γ-histone family 2A variant (γ-H2AX) are up-regulated. This indicates that bile acids cause up-regulation of iNOS, which further causes genotoxic stress as evidenced by increase of the highly sensitive marker, phosphorylated histone. In vitro nitric oxide (NO) assays showed increased production of nitric acid in the oesophageal epithelium exposed to the bile acids. This increase was inhibited in the presence of the nonspecific iNOS inhibitor aminoguanidine (AG). Cumulatively, the results of the present study provide suggestion that not only acid reflux, but also non-acid reflux of bile may cause genotoxic stress. These aspects merit to be tested in wide spectrum of Barrett epithelial tissues. PMID:27247425

  18. Iron-induced Local Complement Component 3 (C3) Up-regulation via Non-canonical Transforming Growth Factor (TGF)-β Signaling in the Retinal Pigment Epithelium*

    PubMed Central

    Li, Yafeng; Song, Delu; Song, Ying; Zhao, Liangliang; Wolkow, Natalie; Tobias, John W.; Song, Wenchao; Dunaief, Joshua L.

    2015-01-01

    Dysregulation of iron homeostasis may be a pathogenic factor in age-related macular degeneration (AMD). Meanwhile, the formation of complement-containing deposits under the retinal pigment epithelial (RPE) cell layer is a pathognomonic feature of AMD. In this study, we investigated the molecular mechanisms by which complement component 3 (C3), a central protein in the complement cascade, is up-regulated by iron in RPE cells. Modulation of TGF-β signaling, involving ERK1/2, SMAD3, and CCAAT/enhancer-binding protein-δ, is responsible for iron-induced C3 expression. The differential effects of spatially distinct SMAD3 phosphorylation sites at the linker region and at the C terminus determined the up-regulation of C3. Pharmacologic inhibition of either ERK1/2 or SMAD3 phosphorylation decreased iron-induced C3 expression levels. Knockdown of SMAD3 blocked the iron-induced up-regulation and nuclear accumulation of CCAAT/enhancer-binding protein-δ, a transcription factor that has been shown previously to bind the basic leucine zipper 1 domain in the C3 promoter. We show herein that mutation of this domain reduced iron-induced C3 promoter activity. In vivo studies support our in vitro finding of iron-induced C3 up-regulation. Mice with a mosaic pattern of RPE-specific iron overload demonstrated co-localization of iron-induced ferritin and C3d deposits. Humans with aceruloplasminemia causing RPE iron overload had increased RPE C3d deposition. The molecular events in the iron-C3 pathway represent therapeutic targets for AMD or other diseases exacerbated by iron-induced local complement dysregulation. PMID:25802332

  19. The soy isoflavone equol may increase cancer malignancy via up-regulation of eukaryotic protein synthesis initiation factor eIF4G.

    PubMed

    de la Parra, Columba; Otero-Franqui, Elisa; Martinez-Montemayor, Michelle; Dharmawardhane, Suranganie

    2012-12-01

    Dietary soy is thought to be cancer-preventive; however, the beneficial effects of soy on established breast cancer is controversial. We recently demonstrated that dietary daidzein or combined soy isoflavones (genistein, daidzein, and glycitein) increased primary mammary tumor growth and metastasis. Cancer-promoting molecules, including eukaryotic protein synthesis initiation factors (eIF) eIF4G and eIF4E, were up-regulated in mammary tumors from mice that received dietary daidzein. Herein, we show that increased eIF expression in tumor extracts of mice after daidzein diets is associated with protein expression of mRNAs with internal ribosome entry sites (IRES) that are sensitive to eIF4E and eIF4G levels. Results with metastatic cancer cell lines show that some of the effects of daidzein in vivo can be recapitulated by the daidzein metabolite equol. In vitro, equol, but not daidzein, up-regulated eIF4G without affecting eIF4E or its regulator, 4E-binding protein (4E-BP), levels. Equol also increased metastatic cancer cell viability. Equol specifically increased the protein expression of IRES containing cell survival and proliferation-promoting molecules and up-regulated gene and protein expression of the transcription factor c-Myc. Moreover, equol increased the polysomal association of mRNAs for p 120 catenin and eIF4G. The elevated eIF4G in response to equol was not associated with eIF4E or 4E-binding protein in 5' cap co-capture assays or co-immunoprecipitations. In dual luciferase assays, IRES-dependent protein synthesis was increased by equol. Therefore, up-regulation of eIF4G by equol may result in increased translation of pro-cancer mRNAs with IRESs and, thus, promote cancer malignancy.

  20. Human p38{delta} MAP kinase mediates UV irradiation induced up-regulation of the gene expression of chemokine BRAK/CXCL14

    SciTech Connect

    Ozawa, Shigeyuki; Ito, Shin; Kato, Yasumasa; Kubota, Eiro; Hata, Ryu-Ichiro

    2010-06-11

    The mitogen-activated protein kinase (MAPK) family comprises ERK, JNK, p38 and ERK5 (big-MAPK, BMK1). UV irradiation of squamous cell carcinoma cells induced up-regulation of gene expression of chemokine BRAK/CXCL14, stimulated p38 phosphorylation, and down-regulated the phosphorylation of ERK. Human p38 MAPKs exist in 4 isoforms: p38{alpha}, {beta}, {gamma} and {delta}. The UV stimulation of p38 phosphorylation was not inhibited by the presence of SB203580 or PD169316, inhibitors of p38{alpha} and {beta}, suggesting p38 phosphorylation was not dependent on these 2 isoforms and that p38{gamma} and/or {delta} was responsible for the phosphorylation. In fact, inhibition of each of these 4 p38 isoforms by the introduction of short hairpin (sh) RNAs for respective isoforms revealed that only shRNA for p38{delta} attenuated the UV-induced up-regulation of BRAK/CXCL14 gene expression. In addition, over-expression of p38 isoforms in the cells showed the association of p38{delta} with ERK1 and 2, concomitant with down-regulation of ERK phosphorylation. The usage of p38{delta} isoform by UV irradiation is not merely due to the abundance of this p38 isoform in the cells. Because serum deprivation of the cells also induced an increase in BRAK/CXCL14 gene expression, and in this case p38{alpha} and/or {beta} isoform is responsible for up-regulation of BRAK/CXCL14 gene expression. Taken together, the data indicate that the respective stress-dependent action of p38 isoforms is responsible for the up-regulation of the gene expression of the chemokine BRAK/CXCL14.

  1. Up-regulation of fibroblast growth factor (FGF) 9 expression and FGF-WNT/β-catenin signaling in laser-induced wound healing.

    PubMed

    Zheng, Zhenlong; Kang, Hye-Young; Lee, Sunha; Kang, Shin-Wook; Goo, Boncheol; Cho, Sung Bin

    2014-01-01

    Fibroblast growth factor (FGF) 9 is secreted by both mesothelial and epithelial cells, and plays important roles in organ development and wound healing via WNT/β-catenin signaling. The aim of this study was to evaluate FGF9 expression and FGF-WNT/β-catenin signaling during wound healing of the skin. We investigated FGF9 expression and FGF-WNT/β-catenin signaling after laser ablation of mouse skin and adult human skin, as well as in cultured normal human epidermal keratinocytes (NHEKs) upon stimulation with recombinant human (rh) FGF9 and rh-transforming growth factor (TGF)-β1. Our results showed that laser ablation of both mouse skin and human skin leads to marked overexpression of FGF9 and FGF9 mRNA. Control NHEKs constitutively expressed FGF9, WNT7b, WNT2, and β-catenin, but did not show Snail or FGF receptor (FGFR) 2 expression. We also found that FGFR2 was significantly induced in NHEKs by rhFGF9 stimulation, and observed that FGFR2 expression was slightly up-regulated on particular days during the wound healing process after ablative laser therapy. Both WNT7b and WNT2 showed up-regulated protein expression during the laser-induced wound healing process in mouse skin; moreover, we discerned that the stimulatory effect of rhFGF9 and rhTGF-β1 activates WNT/β-catenin signaling via WNT7b in cultured NHEKs. Our data indicated that rhFGF9 and/or rhTGF-β1 up-regulate FGFR2, WNT7b, and β-catenin, but not FGF9 and Snail; pretreatment with rh dickkopf-1 significantly inhibited the up-regulation of FGFR2, WNT7b, and β-catenin. Our results suggested that FGF9 and FGF-WNT/β-catenin signaling may play important roles in ablative laser-induced wound healing processes.

  2. Transcriptomic Analysis Reveals Wound Healing of Morus alba Root Extract by Up-Regulating Keratin Filament and CXCL12/CXCR4 Signaling.

    PubMed

    Kim, Kang-Hoon; Chung, Won-Seok; Kim, Yoomi; Kim, Ki-Suk; Lee, In-Seung; Park, Ji Young; Jeong, Hyeon-Soo; Na, Yun-Cheol; Lee, Chang-Hun; Jang, Hyeung-Jin

    2015-08-01

    Facilitation of the wound healing process is important because a prolonged wound site increases pain and the risk of infection. In oriental medicine, an extract of Morus alba root (MA) has usually been prescribed as traditional treatment for accelerating wound healing, and it has been proven to be safe for centuries. To study the molecular mechanism of MA-mediated skin wound healing, we performed a primary cell culture and a skin explant culture and observed significant difference between the groups with and without MA extract. In the cellular system, a real-time cell analysis and real-time quantitative PCR were performed. It was found that MA extract enhanced proliferation in a dose-dependent manner on Kera-308 cell line, and up-regulated keratin expression including wound-induced Krt6a. In skin explant culture, the mRNA level derived from cell outgrowth displayed a tendency toward more up-regulated mRNA associated keratin filaments and toward a more up-regulated mRNA level of C-X-C motif chemokine 12 (CXCL12) and a chemokine receptor 4 (CXCR4) axis signaling pathway downstream. In this process, we concluded that MA extract had a scientific possibility of wound repair by increasing intracellular and extracellular supports and by inducing a CXCL12/CXCR4 signaling pathway.

  3. Fighting food temptations: the modulating effects of short-term cognitive reappraisal, suppression and up-regulation on mesocorticolimbic activity related to appetitive motivation.

    PubMed

    Siep, Nicolette; Roefs, Anne; Roebroeck, Alard; Havermans, Remco; Bonte, Milene; Jansen, Anita

    2012-03-01

    The premise of cognitive therapy is that one can overcome the irresistible temptation of highly palatable foods by actively restructuring the way one thinks about food. Testing this idea, participants in the present study were instructed to passively view foods, up-regulate food palatability thoughts, apply cognitive reappraisal (e.g., thinking about health consequences), or suppress food palatability thoughts and cravings. We examined whether these strategies affect self-reported food craving and mesocorticolimbic activity as assessed by functional magnetic resonance imaging. It was hypothesized that cognitive reappraisal would most effectively inhibit the mesocorticolimbic activity and associated food craving as compared to suppression. In addition, it was hypothesized that suppression would lead to more prefrontal cortex activity, reflecting the use of more control resources, as compared to cognitive reappraisal. Self-report results indicated that up-regulation increased food craving compared to the other two conditions, but that there was no difference in craving between the suppression and cognitive reappraisal strategy. Corroborating self-report results, the neuroimaging results showed that up-regulation increased activity in important regions of the mesocorticolimbic circuitry, including the ventral tegmental area, ventral striatum, operculum, posterior insular gyrus, medial orbitofrontal cortex and ventromedial prefrontal cortex. Contrary to our hypothesis, suppression more effectively decreased activity in the core of the mesocorticolimbic circuitry (i.e., ventral tegmental area and ventral striatum) compared to cognitive reappraisal. Overall, the results support the contention that appetitive motivation can be modulated by the application of short-term cognitive control strategies.

  4. Cancer–Osteoblast Interaction Reduces Sost Expression in Osteoblasts and Up-Regulates lncRNA MALAT1 in Prostate Cancer

    PubMed Central

    Sebastian, Aimy; Hum, Nicholas R.; Hudson, Bryan D.; Loots, Gabriela G.

    2015-01-01

    Dynamic interaction between prostate cancer and the bone microenvironment is a major contributor to metastasis of prostate cancer to bone. In this study, we utilized an in vitro co-culture model of PC3 prostate cancer cells and osteoblasts followed by microarray based gene expression profiling to identify previously unrecognized prostate cancer–bone microenvironment interactions. Factors secreted by PC3 cells resulted in the up-regulation of many genes in osteoblasts associated with bone metabolism and cancer metastasis, including Mmp13, Il-6 and Tgfb2, and down-regulation of Wnt inhibitor Sost. To determine whether altered Sost expression in the bone microenvironment has an effect on prostate cancer metastasis, we co-cultured PC3 cells with Sost knockout (SostKO) osteoblasts and wildtype (WT) osteoblasts and identified several genes differentially regulated between PC3-SostKO osteoblast co-cultures and PC3-WT osteoblast co-cultures. Co-culturing PC3 cells with WT osteoblasts up-regulated cancer-associated long noncoding RNA (lncRNA) MALAT1 in PC3 cells. MALAT1 expression was further enhanced when PC3 cells were co-cultured with SostKO osteoblasts and treatment with recombinant Sost down-regulated MALAT1 expression in these cells. Our results suggest that reduced Sost expression in the tumor microenvironment may promote bone metastasis by up-regulating MALAT1 in prostate cancer. PMID:27600237

  5. Up-regulation of DRP-3 long isoform during the induction of neural progenitor cells by glutamate treatment in the ex vivo rat retina

    SciTech Connect

    Tokuda, Kazuhiro; Kuramitsu, Yasuhiro; Byron, Baron; Kitagawa, Takao; Tokuda, Nobuko; Kobayashi, Daiki; Nagayama, Megumi; Araki, Norie; Sonoda, Koh-Hei; Nakamura, Kazuyuki

    2015-08-07

    Glutamate has been shown to induce neural progenitor cells in the adult vertebrate retina. However, protein dynamics during progenitor cell induction by glutamate are not fully understood. To identify specific proteins involved in the process, we employed two-dimensional electrophoresis-based proteomics on glutamate untreated and treated retinal ex vivo sections. Rat retinal tissues were incubated with 1 mM glutamate for 1 h, followed by incubation in glutamate-free media for a total of 24 h. Consistent with prior reports, it was found that mitotic cells appeared in the outer nuclear layer without any histological damage. Immunohistological evaluations and immunoblotting confirmed the emergence of neuronal progenitor cells in the mature retina treated with glutamate. Proteomic analysis revealed the up-regulation of dihydropyrimidinase-related protein 3 (DRP-3), DRP-2 and stress-induced-phosphoprotein 1 (STIP1) during neural progenitor cell induction by glutamate. Moreover, mRNA expression of DRP-3, especially, its long isoform, robustly increased in the treated retina compared to that in the untreated retina. These results may indicate that glutamate induces neural progenitor cells in the mature rat retina by up-regulating the proteins which mediate cell mitosis and neurite growth. - Highlights: • Glutamate induced neuronal progenitor cells in the mature rat retina. • Proteomic analysis revealed the up-regulation of DRP-3, DRP-2 and STIP1. • mRNA expression of DRP-3, especially, its long isoform, robustly increased.

  6. CIAPIN1 confers multidrug resistance through up-regulation of MDR-1 and Bcl-L in LoVo/Adr cells and is independent of p53.

    PubMed

    Zhang, Ya-Fei; Li, Xiao-Hua; Shi, Yong-Quan; Wu, Yu-Yun; Li, Ning; He, Qiang; Ji, Qing; Wang, Rong-Quan; Yang, Shi-Ming; Fang, Dian-Chun

    2011-04-01

    Recent investigations discovered that CIAPIN1 might be another drug resistance-associated molecule in cancer cells. However, the underlying mechanisms of CIAPIN1-related multidrug resistance (MDR) remain elusive. In the present study, we investigated the role and possible mechanisms of CIAPIN1 in MDR of human colon carcinoma LoVo/Adr cells which express the wild-type p53 gene. By using small interference RNA and gene transfection techniques, we found that knockdown of CIAPIN1 expression re-sensitized LoVo/Adr cells to anti-cancer drugs and up-regulation of CIAPIN1 in sensitive LoVo cells resulted in a distinct MDR phenotype. We further revealed that CIAPIN1 conferred the MDR phenotype in LoVo/Adr cells through up-regulating expression of MDR-1 (P-gp) and Bcl-xL. Finally, by analyzing the effect of inactivation of wild-type p53 on CIAPIN1-induced up-regulation of P-gp and Bcl-xL, we determined that CIAPIN1 could exhibit its MDR-related function independently of the p53 signaling pathway. Overall, the results presented here further suggest that over-expression of CIAPIN1 is an important mechanism of drug resistance in human cancers, even if not the sole one.

  7. Ciliary Neurotrophic Factor Promotes the Migration of Corneal Epithelial Stem/progenitor Cells by Up-regulation of MMPs through the Phosphorylation of Akt

    PubMed Central

    Chen, Jialin; Chen, Peng; Backman, Ludvig J.; Zhou, Qingjun; Danielson, Patrik

    2016-01-01

    The migration of limbal epithelial stem cells is important for the homeostasis and regeneration of corneal epithelium. Ciliary neurotrophic factor (CNTF) has been found to promote corneal epithelial wound healing by activating corneal epithelial stem/progenitor cells. However, the possible effect of CNTF on the migration of corneal epithelial stem/progenitor cells is not clear. This study found the expression of CNTF in mouse corneal epithelial stem/progenitor cells (TKE2) to be up-regulated after injury, on both gene and protein level. CNTF promoted migration of TKE2 in a dose-dependent manner and the peak was seen at 10 ng/ml. The phosphorylation level of Akt (p-Akt), and the expression of MMP3 and MMP14, were up-regulated after CNTF treatment both in vitro and in vivo. Akt and MMP3 inhibitor treatment delayed the migration effect by CNTF. Finally, a decreased expression of MMP3 and MMP14 was observed when Akt inhibitor was applied both in vitro and in vivo. This study provides new insights into the role of CNTF on the migration of corneal epithelial stem/progenitor cells and its inherent mechanism of Up-regulation of matrix metalloproteinases through the Akt signalling pathway. PMID:27174608

  8. Differential screening of mutated SOD1 transgenic mice reveals early up-regulation of a fast axonal transport component in spinal cord motor neurons.

    PubMed

    Dupuis, L; de Tapia, M; René, F; Lutz-Bucher, B; Gordon, J W; Mercken, L; Pradier, L; Loeffler, J P

    2000-08-01

    In the present study we analyze the molecular mechanisms underlying motor neuron degeneration in familial amyotrophic lateral sclerosis (FALS). For this, we used a transgenic mouse model expressing the Cu/Zn superoxide dismutase (SOD1) gene with a Gly(86) to Arg (G86R) mutation equivalent to that found in a subset of human FALS. Using an optimized suppression subtractive hybridization method, a cDNA specifically up-regulated during the asymptomatic phase in the lumbar spinal cord of G86R mice was identified by sequence analysis as the KIF3-associated protein (KAP3), a regulator of fast axonal transport. RT-PCR analysis revealed that KAP3 induction was an early event arising long before axonal degeneration. Immunohistochemical studies further revealed that KAP3 protein predominantly accumulates in large motor neurons of the ventral spinal cord. We further demonstrated that KAP3 up-regulation occurs independent of any change in the other components of the kinesin II complex. However, since the ubiquitous KIF1A motor is up-regulated, our results show an early and complex rearrangement of the fast axonal transport machinery in the course of FALS pathology.

  9. Involvement of up-regulated Necl-5/Tage4/PVR/CD155 in the loss of contact inhibition in transformed NIH3T3 cells

    SciTech Connect

    Minami, Yukiko . E-mail: ytakai@molbio.med.osaka-u.ac.jp

    2007-01-26

    Normal cells show contact inhibition of cell movement and proliferation, but this is lost following transformation. We found that Necl-5, originally identified as a poliovirus receptor and up-regulated in many cancer cells, enhances growth factor-induced cell movement and proliferation. We showed that when cells contact other cells, Necl-5 interacts in trans with nectin-3 and is removed by endocytosis from the cell surface, resulting in a reduction of cell movement and proliferation. We show here that up-regulation of the gene encoding Necl-5 by the oncogene V12-Ki-Ras causes enhanced cell movement and proliferation. Upon cell-cell contact, de novo synthesis of Necl-5 exceeds the rate of Necl-5 endocytosis, eventually resulting in a net increase in the amount of Necl-5 at the cell surface. In addition, expression of the gene encoding nectin-3 is markedly reduced in transformed cells. Thus, up-regulation of Necl-5 following transformation contributes to the loss of contact inhibition in transformed cells.

  10. Cancer–Osteoblast Interaction Reduces Sost Expression in Osteoblasts and Up-Regulates lncRNA MALAT1 in Prostate Cancer

    PubMed Central

    Sebastian, Aimy; Hum, Nicholas R.; Hudson, Bryan D.; Loots, Gabriela G.

    2015-01-01

    Dynamic interaction between prostate cancer and the bone microenvironment is a major contributor to metastasis of prostate cancer to bone. In this study, we utilized an in vitro co-culture model of PC3 prostate cancer cells and osteoblasts followed by microarray based gene expression profiling to identify previously unrecognized prostate cancer–bone microenvironment interactions. Factors secreted by PC3 cells resulted in the up-regulation of many genes in osteoblasts associated with bone metabolism and cancer metastasis, including Mmp13, Il-6 and Tgfb2, and down-regulation of Wnt inhibitor Sost. To determine whether altered Sost expression in the bone microenvironment has an effect on prostate cancer metastasis, we co-cultured PC3 cells with Sost knockout (SostKO) osteoblasts and wildtype (WT) osteoblasts and identified several genes differentially regulated between PC3-SostKO osteoblast co-cultures and PC3-WT osteoblast co-cultures. Co-culturing PC3 cells with WT osteoblasts up-regulated cancer-associated long noncoding RNA (lncRNA) MALAT1 in PC3 cells. MALAT1 expression was further enhanced when PC3 cells were co-cultured with SostKO osteoblasts and treatment with recombinant Sost down-regulated MALAT1 expression in these cells. Our results suggest that reduced Sost expression in the tumor microenvironment may promote bone metastasis by up-regulating MALAT1 in prostate cancer.

  11. Quantitative Proteomic Analysis Revealed 4-(methylnitrosamino)-1-(3-pyridinyl)-1-butanone-induced Up-regulation of 20S Proteasome in Cultured Human Fibroblast Cells

    PubMed Central

    Prins, John M.; Wang, Yinsheng

    2012-01-01

    The tobacco-specific N-nitrosamine, 4-(methylnitrosamino)-1-(3-pyridinyl)-1-butanone (NNK), is a well-known carcinogen. Although the ability of the metabolically activated form of NNK to generate DNA adducts is well established, little is known about the cellular pathways perturbed by NNK in its native state. In this study, we utilized stable isotope labeling by amino acid in cell culture (SILAC), together with mass spectrometry, to assess the perturbation of protein expression in GM00637 human skin fibroblast cells upon NNK exposure. With this approach, we were able to quantify 1412 proteins and 137 of them were with significantly altered expression following NNK exposure, including the up-regulation of all subunits of the 20S proteasome core complex. The up-regulation of the 20S core complex was also reflected by a significant increase in 20S proteasome activities in GM00637, IMR90 and MCF-7 cells upon NNK treatment. Furthermore, the β-adrenergic receptor (β-AR) antagonist propranolol could attenuate significantly the NNK-induced increase in proteasome activity in all the three cell lines, suggesting that up-regulation of the 20S proteasome may be mediated through the β-AR. Additionally, we found that NNK treatment altered the expression levels of other important proteins including mitochondrial proteins, cytoskeleton-associated proteins, and proteins involved in glycolysis and gluconeogenesis. Results from the present study provided novel insights into the cellular mechanisms targeted by NNK. PMID:22369695

  12. Anaplasma phagocytophilum up-regulates some anti-apoptotic genes in neutrophils and pro-inflammatory genes in mononuclear cells of sheep.

    PubMed

    Woldehiwet, Z; Yavari, C

    2014-05-01

    Anaplasma phagocytophilum, the causative agent of tick-borne fever (TBF) in sheep and cattle and human granulocytic anaplasmosis, has the unique ability to selectively infect and multiply within the hostile environment of the neutrophil. Previous studies have shown that sheep with TBF are more susceptible to other infections and that infected neutrophils have reduced phagocytic ability and delayed apoptosis. This suggests that survival of A. phagocytophilum in these short-lived cells involves the ability to subvert or resist their bacterial killing, but also to modify the host cells such that the host cells survive long after infection. The present study shows that infection of sheep by A. phagocytophilum is characterized by up-regulation of some anti-apoptotic genes (BCL2, BIRC3 and CFLAR) in neutrophils and up-regulation of genes encoding the pro-inflammatory cytokines interferon-γ, interleukin (IL)-1β and IL-6 in mononuclear cells during the period of bacteraemia. Infection with A. phagocytophilum was also characterized by significant up-regulation of CYBB, which is associated with the respiratory burst of neutrophils. PMID:24602324

  13. SEIZURE ACTIVITY INVOLVED IN THE UP-REGULATION OF BDNF mRNA EXPRESSION BY ACTIVATION OF CENTRAL MU OPIOID RECEPTORS

    PubMed Central

    ZHANG, H. N.; KO, M. C.

    2009-01-01

    Chemical-induced seizures up-regulated brain-derived neurotrophic factor (BDNF) mRNA expression. Intracerebroventricular (i.c.v.) administration of endogenous opioids preferentially activating μ opioid receptor (MOR) could also increase BDNF mRNA expression. The aim of this study was to determine to what extent i.c.v. administration of synthetic MOR-selective agonists in rats can modulate both seizure activity and up-regulation of BDNF mRNA expression. Effects and potencies of i.c.v. administration of morphine and [D-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin (DAMGO), were directly investigated by scoring behavioral seizures and measuring BDNF mRNA expression. In addition, effects of the opioid receptor antagonist naloxone and antiepileptic drugs, diazepam, phenobarbital, and valproate, on i.c.v. MOR agonist-induced behavioral seizures and up-regulation of BDNF mRNA expression were determined. A single i.c.v. administration of morphine (10–100 μg) or DAMGO (0.15–1.5 μg) dose-dependently elicited behavioral seizures and increased BDNF mRNA expression in the widespread brain regions. However, subcutaneous administration of MOR agonists neither produced behavioral seizures nor increased BDNF mRNA expression. Pretreatment with naloxone 1 mg/kg significantly reduced behavioral seizure scores and the up-regulation of BDNF mRNA expression elicited by i.c.v. morphine or DAMGO. Similarly, diazepam 10 mg/kg and phenobarbital 40 mg/kg significantly blocked i.c.v. MOR agonist-induced actions. Pretreatment with valproate 300 mg/kg only attenuated behavioral seizures, but it did not affect morphine-induced increase of BDNF mRNA expression. This study provides supporting evidence that seizure activity plays an important role in the up-regulation of BDNF mRNA expression elicited by central MOR activation and that decreased inhibitory action of GABAergic system through the modulation on GABA receptor synaptic function by central MOR activation is involved in its regulation of BDNF m

  14. Serpinin: A Novel Chromogranin A-Derived, Secreted Peptide Up-Regulates Protease Nexin-1 Expression and Granule Biogenesis in Endocrine Cells

    PubMed Central

    Koshimizu, Hisatsugu; Cawley, Niamh X.; Kim, Taeyoon; Yergey, Alfred L.

    2011-01-01

    Previously we demonstrated that chromogranin A (CgA) promoted secretory granule biogenesis in endocrine cells by stabilizing and preventing granule protein degradation in the Golgi, through up-regulation of expression of the protease inhibitor, protease nexin-1 (PN-1). However, the mechanism by which CgA signals the increase of PN-1 expression is unknown. Here we identified a 2.9-kDa CgA-C-terminus peptide, which we named serpinin, in conditioned media from AtT-20 cells, a corticotroph cell line, which up-regulated PN-1 mRNA expression. Serpinin was secreted from AtT-20 cells upon high potassium stimulation and increased PN-1 mRNA transcription in these cells, in an actinomycin D-inhibitable manner. CgA itself and other CgA-derived peptides, when added to AtT-20 cell media, had no effect on PN-1 expression. Treatment of AtT-20 cells with 10 nm serpinin elevated cAMP levels and PN-1 mRNA expression, and this effect was inhibited by a protein kinase A inhibitor, 6–22 amide. Serpinin and a cAMP analog, 8-bromo-cAMP, promoted the translocation of the transcription factor Sp1 into the nucleus, which is known to drive PN-1 expression. Additionally, an Sp1 inhibitor, mithramycin A inhibited the serpinin-induced PN-1 mRNA up-regulation. Furthermore, a luciferase reporter assay demonstrated serpinin-induced up-regulation of PN-1 promoter activity in an Sp1-dependent manner. When added to CgB-transfected 6T3 cells, a mutant AtT20 cell line, serpinin induced granule biogenesis as evidenced by the presence of CgB puncta accumulation in the processes and tips. Our findings taken together show that serpinin, a novel CgA-derived peptide, is secreted upon stimulation of corticotrophs and plays an important autocrine role in up-regulating PN-1-dependent granule biogenesis via a cAMP-protein kinase A-Sp1 pathway to replenish released granules. PMID:21436258

  15. Up-regulation of K{sub ir}2.1 by ER stress facilitates cell death of brain capillary endothelial cells

    SciTech Connect

    Kito, Hiroaki; Yamazaki, Daiju; Ohya, Susumu; Yamamura, Hisao; Asai, Kiyofumi; Imaizumi, Yuji

    2011-07-29

    Highlights: {yields} We found that application of endoplasmic reticulum (ER) stress with tunicamycin to brain capillary endothelial cells (BCECs) induced cell death. {yields} The ER stress facilitated the expression of inward rectifier K{sup +} channel (K{sub ir}2.1) and induced sustained membrane hyperpolarization. {yields} The membrane hyperpolarization induced sustained Ca{sup 2+} entry through voltage-independent nonspecific cation channels and consequently facilitated cell death. {yields} The K{sub ir}2.1 up-regulation by ER stress is, at least in part, responsible for cell death of BCECs under pathological conditions. -- Abstract: Brain capillary endothelial cells (BCECs) form blood brain barrier (BBB) to maintain brain homeostasis. Cell turnover of BCECs by the balance of cell proliferation and cell death is critical for maintaining the integrity of BBB. Here we found that stimuli with tunicamycin, endoplasmic reticulum (ER) stress inducer, up-regulated inward rectifier K{sup +} channel (K{sub ir}2.1) and facilitated cell death in t-BBEC117, a cell line derived from bovine BCECs. The activation of K{sub ir} channels contributed to the establishment of deeply negative resting membrane potential in t-BBEC117. The deep resting membrane potential increased the resting intracellular Ca{sup 2+} concentration due to Ca{sup 2+} influx through non-selective cation channels and thereby partly but significantly regulated cell death in t-BBEC117. The present results suggest that the up-regulation of K{sub ir}2.1 is, at least in part, responsible for cell death/cell turnover of BCECs induced by a variety of cellular stresses, particularly ER stress, under pathological conditions.

  16. Non-adherent culture induces paclitaxel resistance in H460 lung cancer cells via ERK-mediated up-regulation of βIVa-tubulin.

    PubMed

    Atjanasuppat, Korakot; Lirdprapamongkol, Kriengsak; Jantaree, Phatcharida; Svasti, Jisnuson

    2015-10-23

    Circulating tumor cells (CTCs) are metastasizing epithelial cancer cells that adapt to survive when floating in bloodstream during metastasis. This condition can be mimicked in vitro by using non-adherent cell culture. The chemosensitivity of CTCs appears to correlate with the response of metastatic cancer patients to therapy, but chemoresistance is also frequently observed in advanced stage cancer patients, who have never previously received chemotherapy. We hypothesize that adaptation of epithelial cancer cells to become floating CTCs could lead to development of chemoresistance. Here, we explore whether chemoresistance is induced in epithelial cancer cells when cultured under non-adherent conditions. Increased paclitaxel-specific resistance was observed in floating cells compared to attached cells in H460, MCF-7, and HepG2 human cancer cell lines, by 15.6-, 3.9-, and 2.6-fold increases in IC50 values, respectively. qRT-PCR analysis showed that a paclitaxel-resistant β-tubulin isotype, βIVa-tubulin, was the most up-regulated gene compared with other β-tubulin isotypes in H460 floating cells, concomitant with elevated ERK activation. ERK inhibitor treatment could attenuate the up-regulation of βIVa-tubulin, and decreased the paclitaxel resistance of H460 floating cells, even though other β-tubulin isotypes were up-regulated when the ERK activation was blocked. In conclusion, we show induction of paclitaxel resistance in epithelial cancer cells, when floating in non-adherent culture, and this might occur with CTCs of cancer patients.

  17. Histone hyperacetylation up-regulates protein kinase Cδ in dopaminergic neurons to induce cell death: relevance to epigenetic mechanisms of neurodegeneration in Parkinson disease.

    PubMed

    Jin, Huajun; Kanthasamy, Arthi; Harischandra, Dilshan S; Kondru, Naveen; Ghosh, Anamitra; Panicker, Nikhil; Anantharam, Vellareddy; Rana, Ajay; Kanthasamy, Anumantha G

    2014-12-12

    The oxidative stress-sensitive protein kinase Cδ (PKCδ) has been implicated in dopaminergic neuronal cell death. However, little is known about the epigenetic mechanisms regulating PKCδ expression in neurons. Here, we report a novel mechanism by which the PKCδ gene can be regulated by histone acetylation. Treatment with histone deacetylase (HDAC) inhibitor sodium butyrate (NaBu) induced PKCδ expression in cultured neurons, brain slices, and animal models. Several other HDAC inhibitors also mimicked NaBu. The chromatin immunoprecipitation analysis revealed that hyperacetylation of histone H4 by NaBu is associated with the PKCδ promoter. Deletion analysis of the PKCδ promoter mapped the NaBu-responsive element to an 81-bp minimal promoter region. Detailed mutagenesis studies within this region revealed that four GC boxes conferred hyperacetylation-induced PKCδ promoter activation. Cotransfection experiments and Sp inhibitor studies demonstrated that Sp1, Sp3, and Sp4 regulated NaBu-induced PKCδ up-regulation. However, NaBu did not alter the DNA binding activities of Sp proteins or their expression. Interestingly, a one-hybrid analysis revealed that NaBu enhanced transcriptional activity of Sp1/Sp3. Overexpression of the p300/cAMP-response element-binding protein-binding protein (CBP) potentiated the NaBu-mediated transactivation potential of Sp1/Sp3, but expressing several HDACs attenuated this effect, suggesting that p300/CBP and HDACs act as coactivators or corepressors in histone acetylation-induced PKCδ up-regulation. Finally, using genetic and pharmacological approaches, we showed that NaBu up-regulation of PKCδ sensitizes neurons to cell death in a human dopaminergic cell model and brain slice cultures. Together, these results indicate that histone acetylation regulates PKCδ expression to augment nigrostriatal dopaminergic cell death, which could contribute to the progressive neuropathogenesis of Parkinson disease.

  18. The membrane-associated progesterone-binding protein 25-Dx: expression, cellular localization and up-regulation after brain and spinal cord injuries.

    PubMed

    Guennoun, R; Meffre, D; Labombarda, F; Gonzalez, S L; Gonzalez Deniselle, M C; Stein, D G; De Nicola, A F; Schumacher, M

    2008-03-01

    Progesterone has neuroprotective effects in the injured and diseased spinal cord and after traumatic brain injury (TBI). In addition to intracellular progesterone receptors (PR), membrane-binding sites of progesterone may be involved in neuroprotection. A first putative membrane receptor of progesterone, distinct from the classical intracellular PR isoforms, with a single membrane-spanning domain, has been cloned from porcine liver. Homologous proteins were cloned in rats (25-Dx), mice (PGRMC1) and humans (Hpr.6). We will refer to this progesterone-binding protein as 25-Dx. The distribution and regulation of 25-Dx in the nervous system may provide some clues to its functions. In spinal cord, 25-Dx is localized in cell membranes of dorsal horn neurons and ependymal cells lining the central canal. A role of 25-Dx in mediating the protective effects of progesterone in the spinal cord is supported by the observation that its mRNA and protein are up-regulated by progesterone in dorsal horn of the injured spinal cord. In contrast, the classical intracellular PRs were down-regulated under these conditions. In brain, 25-Dx is particularly abundant in the hypothalamic area, circumventricular organs, ependymal cells of the ventricular walls, and the meninges. Interestingly, it is co-expressed with vasopressin in neurons of the paraventricular, supraoptic and retrochiasmatic nuclei. In response to TBI, 25-Dx expression is up-regulated in neurons and induced in astrocytes. The expression of 25-Dx in structures involved in cerebrospinal fluid production and osmoregulation, and its up-regulation after brain damage, point to a potentially important role of this progesterone-binding protein in the maintenance of water homeostasis after TBI. Our observations suggest that progesterone's actions may involve different signaling mechanisms depending on the pathophysiological context, and that 25-Dx may be involved in the neuroprotective effect of progesterone in the injured brain and

  19. ERβ-dependent neuroglobin up-regulation impairs 17β-estradiol-induced apoptosis in DLD-1 colon cancer cells upon oxidative stress injury.

    PubMed

    Fiocchetti, Marco; Camilli, Giulia; Acconcia, Filippo; Leone, Stefano; Ascenzi, Paolo; Marino, Maria

    2015-05-01

    Besides other mechanism(s) 17β-estradiol (E2) facilitates neuronal survival by increasing, via estrogen receptor β (ERβ), the levels of neuroglobin (NGB) an anti-apoptotic protein. In contrast, E2 could exert protective effects in cancer cells by activating apoptosis when the ERβ level prevails on that of ERα as in colon cancer cell lines. These apparently contrasting results raise the possibility that E2-induced NGB up-regulation could regulate the ERβ activities shunning this receptor subtype to trigger an apoptotic cascade in neurons but not in non-neuronal cells. Here, human colorectal adenocarcinoma cell line (DLD-1) that only expresses ERβ and HeLa cells transiently transfected with ERβ encoding vector has been used to verify this hypothesis. In addition, neuroblastoma SK-N-BE cells were used as positive control. Surprisingly, E2 also induced NGB up-regulation, in a dose- and time-dependent manner, in DLD-1 cells. The ERβ-mediated activation of p38/MAPK was necessary for this E2 effect. E2 induced NGB re-allocation in mitochondria where, subsequently to an oxidative stress injury (i.e., 100μM H2O2), NGB interacted with cytochrome c preventing its release into the cytosol and the activation of an apoptotic cascade. As a whole, these results demonstrate that E2-induced NGB up-regulation could act as an oxidative stress sensor, which does not oppose to the pro-apoptotic E2 effect in ERβ-containing colon cancer cells unless a rise of oxidative stress occurs. These results support the concept that oxidative stress plays a critical role in E2-induced carcinogenesis and further open an important scenario to develop novel therapeutic strategies that target NGB against E2-related cancers.

  20. Monocarboxylate transporter 1 is up-regulated in Caco-2 cells by the methionine precursor DL-2-hydroxy-(4-methylthio)butanoic acid.

    PubMed

    Martín-Venegas, Raquel; Brufau, M Teresa; Mañas-Cano, Oriol; Mercier, Yves; Nonis, Magalie K; Ferrer, Ruth

    2014-12-01

    The methionine precursor, DL-2-hydroxy-(4-methylthio)butanoic acid (HMTBA), is a synthetic source of dietary methionine, which is widely used as a poultry nutritional supplement. In the intestinal epithelium, HMTBA transport across the apical membrane is mediated by monocarboxylate transporter 1 (MCT1). The first step in biological utilisation of this methionine precursor is the stereospecific conversion of HMTBA to the corresponding keto acid. In the present study, the regulation of trans-epithelial HMTBA transport was investigated in Caco-2 cell monolayers. Differentiated Caco-2 cells were maintained under control conditions (apical compartment: 0.2 mmol/L L-methionine) or in a HMTBA-enriched medium (2 mmol/L HMTBA). The effect of culture on HMTBA transport was evaluated from apical and basolateral kinetic parameters. MCT1 and MCT4 immuno-localisation and gene expression were investigated by confocal microscopy and real-time quantitative RT-PCR, respectively. The results indicated that apical MCT1 was up-regulated by exposure to HMTBA (1.4-fold increase in Vmax without changes in Km). Moreover, total monolayer MCT1 immunoreactivity increased 1.8-fold in HMTBA-supplemented cultures, this effect mainly being localised at the apical membrane. Functional and immuno-localisation data suggest involvement of MCT1 and MCT4 in basolateral HMTBA transport, although, in this case, no effect was observed for HMTBA-enrichment. Molecular analysis confirmed MCT1 mRNA up-regulation (1.8-fold), with no effect on MCT4 mRNA expression. Thus, exposure to HMTBA up-regulates the trans-epithelial transport of this methionine precursor by increasing the expression and the transport capacity of apical MCT1. PMID:25447800

  1. G protein-coupled receptor 30 expression is up-regulated by EGF and TGF alpha in estrogen receptor alpha-positive cancer cells.

    PubMed

    Vivacqua, Adele; Lappano, Rosamaria; De Marco, Paola; Sisci, Diego; Aquila, Saveria; De Amicis, Francesca; Fuqua, Suzanne A W; Andò, Sebastiano; Maggiolini, Marcello

    2009-11-01

    In the present study, we evaluated the regulation of G protein-coupled receptor (GPR)30 expression in estrogen receptor (ER)-positive endometrial, ovarian, and estrogen-sensitive, as well as tamoxifen-resistant breast cancer cells. We demonstrate that epidermal growth factor (EGF) and TGF alpha transactivate the GPR30 promoter and accordingly up-regulate GPR30 mRNA and protein levels only in endometrial and tamoxifen-resistant breast cancer cells. These effects exerted by EGF and TGF alpha were dependent on EGF receptor (EGFR) expression and activation and involved phosphorylation of the Tyr(1045) and Tyr(1173) EGFR sites. Using gene-silencing experiments and specific pharmacological inhibitors, we have ascertained that EGF and TGF alpha induce GPR30 expression through the EGFR/ERK transduction pathway, and the recruitment of c-fos to the activator protein-1 site located within GPR30 promoter sequence. Interestingly, we show that functional cross talk of GPR30 with both activated EGFR and ER alpha relies on a physical interaction among these receptors, further extending the potential of estrogen to trigger a complex stimulatory signaling network in hormone-sensitive tumors. Given that EGFR/HER2 overexpression is associated with tamoxifen resistance, our data may suggest that ligand-activated EGFR could contribute to the failure of tamoxifen therapy also by up-regulating GPR30, which in turn could facilitates the action of estrogen. In addition, important for resistance is the ability of tamoxifen to bind to and activate GPR30, the expression of which is up-regulated by EGFR activation. Our results emphasize the need for new endocrine agents able to block widespread actions of estrogen without exerting any stimulatory activity on transduction pathways shared by the steroid and growth factor-signaling networks.

  2. Ammonium increases Ca(2+) signalling and up-regulates expression of TRPC1 gene in astrocytes in primary cultures and in the in vivo brain.

    PubMed

    Liang, Chunguang; Du, Ting; Zhou, Jing; Verkhratsky, Alexei; Peng, Liang

    2014-11-01

    Rapid rise in ammonium concentration in the brain is the major pathogenic factor in hepatic encephalopathy that is manifested by state of confusion, forgetfulness and irritability, psychotic symptoms, delusions, lethargy, somnolence and, in the terminal stages, coma. Primary cultures of mouse astrocytes were used to investigate effects of chronic treatment (3 days) with ammonium chloride (ammonium) at 3 mM, this being a relevant concentration for hepatic encephalopathy condition, on metabotropic receptor agonist-induced increases in free cytosolic Ca(2+) concentration [(Ca(2+))i], measured with fura-2 based microfluorimetry and on store-operated Ca(2+) entry (SOCE) activated following treatment with the SERCA inhibitor thapsigargin. The agonists used were the β-adrenergic agonist isoproterenol, the α2-adrenergic agonist dexmedetomidine, the InsP3 receptor (InsP3R) agonist adenophostin A and ryanodine receptor agonist 4-Chloro-m-cresol (4-CMC). Agonist-induced [Ca(2+)]i responses were significantly increased in astrocytes chronically exposed to ammonium. Similarly, the SOCE, meditated by the transient receptor potential channel 1 (TRPC1), was significantly augmented. The ammonium-induced increase in SOCE was a result of an up-regulation of mRNA and protein expression of TRPC1 in astrocytes. Increase in TRPC1 expression and in SOCE were both prevented by ouabain antagonist canrenone. Similar up-regulation of TRPC1 gene expression was found in the brain of adult mice subjected to intraperitoneal injection of urease for 3 days. In transgenic mice tagged with an astrocyte-specific or a neurone-specific markers and treated with intraperitoneal injections of urease for 3 days, the fluorescence-activated cell sorting of neurones and astrocytes demonstrated that TRPC1 mRNA expression was up-regulated in astrocytes, but not in neurones. PMID:25113123

  3. Cultured lymphocytes from autistic children and non-autistic siblings up-regulate heat shock protein RNA in response to thimerosal challenge.

    PubMed

    Walker, Stephen J; Segal, Jeffrey; Aschner, Michael

    2006-09-01

    There are reports suggesting that some autistic children are unable to mount an adequate response following exposure to environmental toxins. This potential deficit, coupled with the similarity in clinical presentations of autism and some heavy metal toxicities, has led to the suggestion that heavy metal poisoning might play a role in the etiology of autism in uniquely susceptible individuals. Thimerosal, an anti-microbial preservative previously added routinely to childhood multi-dose vaccines, is composed of 49.6% ethyl mercury. Based on the levels of this toxin that children receive through routine immunization schedules in the first years of life, it has been postulated that thimerosal may be a potential triggering mechanism contributing to autism in susceptible individuals. One potential risk factor in these individuals may be an inability to adequately up-regulate metallothionein (MT) biosynthesis in response to presentation of a heavy metal challenge. To investigate this hypothesis, cultured lymphocytes (obtained from the Autism Genetic Resource Exchange, AGRE) from autistic children and non-autistic siblings were challenged with either 10 microM ethyl mercury, 150 microM zinc, or fresh media (control). Following the challenge, total RNA was extracted and used to query "whole genome" DNA microarrays. Cultured lymphocytes challenged with zinc responded with an impressive up-regulation of MT transcripts (at least nine different MTs were over-expressed) while cells challenged with thimerosal responded by up-regulating numerous heat shock protein transcripts, but not MTs. Although there were no apparent differences between autistic and non-autistic sibling responses in this very small sampling group, the differences in expression profiles between those cells treated with zinc versus thimerosal were dramatic. Determining cellular response, at the level of gene expression, has important implications for the understanding and treatment of conditions that result

  4. The Inhibitory Effects of Anti-Oxidants on Ultraviolet-Induced Up-Regulation of the Wrinkling-Inducing Enzyme Neutral Endopeptidase in Human Fibroblasts

    PubMed Central

    Nakajima, Hiroaki; Terazawa, Shuko; Niwano, Takao; Yamamoto, Yorihiro; Imokawa, Genji

    2016-01-01

    We recently reported that the over-expression of skin fibroblast-derived neutral endopeptidase (NEP) plays a pivotal role in impairing the three-dimensional architecture of dermal elastic fibers during the biological mechanism of ultraviolet (UV)-induced skin wrinkling. In that process, a UVB-associated epithelial-mesenchymal cytokine interaction as well as a direct UVA-induced cellular stimulation are associated with the up-regulation of NEP in human fibroblasts. In this study, we characterized the mode of action of ubiquinol10 which may abrogate the up-regulation of NEP by dermal fibroblasts, resulting in a reported in vivo anti-wrinkling action, and compared that with 3 other anti-oxidants, astaxanthin (AX), riboflavin (RF) and flavin mononucleotide (FMN). Post-irradiation treatment with all 4 of those anti-oxidants elicited an interrupting effect on the UVB-associated epithelial-mesenchymal cytokine interaction leading to the up-regulation of NEP in human fibroblasts but with different modes of action. While AX mainly served as an inhibitor of the secretion of wrinkle-inducing cytokines, such as interleukin-1α (IL-1α) and granulocyte macrophage colony stimulatory factor (GM-CSF) in UVB-exposed epidermal keratinocytes, ubiquinol10, RF and FMN predominantly interrupted the IL-1α and GM-CSF-stimulated expression of NEP in dermal fibroblasts. On the other hand, as for the UVA-associated mechanism, similar to the abrogating effects reported for AX and FMN, ubiquinol10 but not RF had the potential to abrogate the increased expression of NEP and matrix-metalloproteinase-1 in UVA-exposed human fibroblasts. Our findings strongly support the in vivo anti-wrinkling effects of ubiquinol10 and AX on human and animal skin and provide convincing proof of the UV-induced wrinkling mechanism that essentially focuses on the over-expression of NEP by dermal fibroblasts as an intrinsic causative factor. PMID:27648570

  5. Extremely Low-Frequency Electromagnetic Fields Promote In Vitro Neuronal Differentiation and Neurite Outgrowth of Embryonic Neural Stem Cells via Up-Regulating TRPC1

    PubMed Central

    Ma, Qinlong; Chen, Chunhai; Deng, Ping; Zhu, Gang; Lin, Min; Zhang, Lei; Xu, Shangcheng; He, Mindi; Lu, Yonghui; Duan, Weixia; Pi, Huifeng; Cao, Zhengwang; Pei, Liping; Li, Min; Liu, Chuan; Zhang, Yanwen; Zhong, Min; Zhou, Zhou; Yu, Zhengping

    2016-01-01

    Exposure to extremely low-frequency electromagnetic fields (ELF-EMFs) can enhance hippocampal neurogenesis in adult mice. However, little is focused on the effects of ELF-EMFs on embryonic neurogenesis. Here, we studied the potential effects of ELF-EMFs on embryonic neural stem cells (eNSCs). We exposed eNSCs to ELF-EMF (50 Hz, 1 mT) for 1, 2, and 3 days with 4 hours per day. We found that eNSC proliferation and maintenance were significantly enhanced after ELF-EMF exposure in proliferation medium. ELF-EMF exposure increased the ratio of differentiated neurons and promoted the neurite outgrowth of eNSC-derived neurons without influencing astrocyes differentiation and the cell apoptosis. In addition, the expression of the proneural genes, NeuroD and Ngn1, which are crucial for neuronal differentiation and neurite outgrowth, was increased after ELF-EMF exposure. Moreover, the expression of transient receptor potential canonical 1 (TRPC1) was significantly up-regulated accompanied by increased the peak amplitude of intracellular calcium level induced by ELF-EMF. Furthermore, silencing TRPC1 expression eliminated the up-regulation of the proneural genes and the promotion of neuronal differentiation and neurite outgrowth induced by ELF-EMF. These results suggest that ELF-EMF exposure promotes the neuronal differentiation and neurite outgrowth of eNSCs via up-regulation the expression of TRPC1 and proneural genes (NeuroD and Ngn1). These findings also provide new insights in understanding the effects of ELF-EMF exposure on embryonic brain development. PMID:26950212

  6. The freshwater Amazonian stingray, Potamotrygon motoro, up-regulates glutamine synthetase activity and protein abundance, and accumulates glutamine when exposed to brackish (15 per thousand) water.

    PubMed

    Ip, Y K; Loong, A M; Ching, B; Tham, G H Y; Wong, W P; Chew, S F

    2009-12-01

    This study aimed to examine whether the stenohaline freshwater stingray, Potamotrygon motoro, which lacks a functional ornithine-urea cycle, would up-regulate glutamine synthetase (GS) activity and protein abundance, and accumulate glutamine during a progressive transfer from freshwater to brackish (15 per thousand) water with daily feeding. Our results revealed that, similar to other freshwater teleosts, P. motoro performed hyperosmotic regulation, with very low urea concentrations in plasma and tissues, in freshwater. In 15 per thousand water, it was non-ureotelic and non-ureoosmotic, acting mainly as an osmoconformer with its plasma osmolality, [Na+] and [Cl-] comparable to those of the external medium. There were significant increases in the content of several free amino acids (FAAs), including glutamate, glutamine and glycine, in muscle and liver, but not in plasma, indicating that FAAs could contribute in part to cell volume regulation. Furthermore, exposure of P. motoro to 15 per thousand water led to up-regulation of GS activity and protein abundance in both liver and muscle. Thus, our results indicate for the first time that, despite the inability to synthesize urea and the lack of functional carbamoyl phosphate synthetase III (CPS III) which uses glutamine as a substrate, P. motoro retained the capacity to up-regulate the activity and protein expression of GS in response to salinity stress. Potamotrygon motoro was not nitrogen (N) limited when exposed to 15 per thousand water with feeding, and there were no significant changes in the amination and deamination activities of hepatic glutamate dehydrogenase. In contrast, P. motoro became N limited when exposed to 10 per thousand water with fasting and could not survive well in 15 per thousand water without food.

  7. Inhibition of the ERK phosphorylation plays a role in terbinafine-induced p21 up-regulation and DNA synthesis inhibition in human vascular endothelial cells

    SciTech Connect

    Ho, P.-Y.; Hsu, S.-P.; Liang, Y.-C.; Kuo, M.-L.; Ho, Y.-S.; Lee, W.-S.

    2008-05-15

    Previously, we showed that terbinafine (TB) induces cell-cycle arrest in cultured human umbilical vein endothelial cells (HUVEC) through an up-regulation of the p21 protein. The aim of this study is to delineate the molecular mechanisms underlying TB-induced increase of p21 protein. RT-PCR analysis demonstrated that the mRNA levels of p21 and p53 were increased in the TB-treated HUVEC. The p21 promoter activity was also increased by TB treatment. Transfection of HUVEC with p53 dominant negative (DN) abolished the TB-induced increases of p21 promoter activity and protein level, suggesting that the TB-induced increase of p21 is p53-dependent. Western blot analysis demonstrated that TB decreased the levels of phosphorylated extracellular signal-regulated kinase (ERK). Over-expression of mitogen-activated protein kinase (MEK)-1, the immediate upstream activator kinase of ERK, abolished the TB-induced increases of p21 and p53 protein and decrease of thymidine incorporation. The ERK inhibitor (PD98059) enhanced the TB-induced inhibition of thymidine incorporation into HUVEC. Taken together, these data suggest that the decrease of ERK activity plays a role in the TB-induced up-regulation of p21 in HUVEC. On the other hand, pretreatment of the cells with geranylgeraniol (GGOH), farnesol (FOH), or Ras inhibitor peptide did not affect the TB-induced decrease of thymidine incorporation. Taken together, our results suggest that TB might cause a decrease of MEK, which in turn up-regulates p53 through the inhibition of ERK phosphorylation, and finally causes an increase of p21 expression and cell-cycle arrest.

  8. The peptidyl-prolyl isomerase Pin1 up-regulation and proapoptotic function in dopaminergic neurons: relevance to the pathogenesis of Parkinson disease.

    PubMed

    Ghosh, Anamitra; Saminathan, Hariharan; Kanthasamy, Arthi; Anantharam, Vellareddy; Jin, Huajun; Sondarva, Gautam; Harischandra, Dilshan S; Qian, Ziqing; Rana, Ajay; Kanthasamy, Anumantha G

    2013-07-26

    Parkinson disease (PD) is a chronic neurodegenerative disease characterized by a slow and progressive degeneration of dopaminergic neurons in substantia nigra. The pathophysiological mechanisms underlying PD remain unclear. Pin1, a major peptidyl-prolyl isomerase, has recently been associated with certain diseases. Notably, Ryo et al. (Ryo, A., Togo, T., Nakai, T., Hirai, A., Nishi, M., Yamaguchi, A., Suzuki, K., Hirayasu, Y., Kobayashi, H., Perrem, K., Liou, Y. C., and Aoki, I. (2006) J. Biol. Chem. 281, 4117-4125) implicated Pin1 in PD pathology. Therefore, we sought to systematically characterize the role of Pin1 in PD using cell culture and animal models. To our surprise we observed a dramatic up-regulation of Pin1 mRNA and protein levels in dopaminergic MN9D neuronal cells treated with the parkinsonian toxicant 1-methyl-4-phenylpyridinium (MPP(+)) as well as in the substantia nigra of the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mouse model. Notably, a marked expression of Pin1 was also observed in the substantia nigra of human PD brains along with a high co-localization of Pin1 within dopaminergic neurons. In functional studies, siRNA-mediated knockdown of Pin1 almost completely prevented MPP(+)-induced caspase-3 activation and DNA fragmentation, indicating that Pin1 plays a proapoptotic role. Interestingly, multiple pharmacological Pin1 inhibitors, including juglone, attenuated MPP(+)-induced Pin1 up-regulation, α-synuclein aggregation, caspase-3 activation, and cell death. Furthermore, juglone treatment in the MPTP mouse model of PD suppressed Pin1 levels and improved locomotor deficits, dopamine depletion, and nigral dopaminergic neuronal loss. Collectively, our findings demonstrate for the first time that Pin1 is up-regulated in PD and has a pathophysiological role in the nigrostriatal dopaminergic system and suggest that modulation of Pin1 levels may be a useful translational therapeutic strategy in PD.

  9. Cancer-associated fibroblasts up-regulate CCL2, CCL26, IL6 and LOXL2 genes related to promotion of cancer progression in hepatocellular carcinoma cells.

    PubMed

    Lin, Zu-Yau; Chuang, Yen-Hwang; Chuang, Wan-Long

    2012-10-01

    Impact of different cancer-associated fibroblast (CAF) cell lines on proliferation, migration, invasion and differential expressions of genes in different hepatocellular carcinoma (HCC) cell lines was investigated. Two human CAF cell lines (F26/KMUH, F28/KMUH) and two human HCC cell lines (HCC24/KMUH, HCC38/KMUH) were studied. Influence of F28/KMUH cells on expressions of genes in HCC38/KMUH cells was detected by microarray to select genes for further analysis. Both CAF cell lines promoted proliferation (all P<0.05), migration (all P<0.05) and Matrigel invasion (all P<0.0001) of both HCC cell lines. F26/KMUH cells showed stronger promoted effects on, firstly, proliferation of HCC24/KMUH cells (P=0.0064) and, secondly, migration of both HCC cell lines than F28/KMUH cells did (all P<0.002). Ten up-regulated genes (APLN, CCL2, CCL26, CXCR4, IL6, MUC1, LOXL2, PDGFA, PGK1, VEGFA) related to proliferation, migration, invasion and angiogenesis of HCC detected by microarray were selected for quantitative reverse transcriptase-polymerase chain reaction analysis. Both CAF cell lines had same tendency of effects on differential expressions of genes in same HCC cell line, but expressions of genes between different HCC cell lines were not consistent. Only CCL2, CCL26, IL6 and LOXL2 genes were consistently up-regulated in both HCC cell lines. In conclusion, the effects of CAFs to promote proliferation, migration and invasion of HCC cells are influenced by the characteristics of both CAFs and HCC cells. Up-regulations of CCL2, CCL26, IL6 and LOXL2 genes in cancer cells are part of the common effects of CAFs on HCC cells.

  10. ERβ-dependent neuroglobin up-regulation impairs 17β-estradiol-induced apoptosis in DLD-1 colon cancer cells upon oxidative stress injury.

    PubMed

    Fiocchetti, Marco; Camilli, Giulia; Acconcia, Filippo; Leone, Stefano; Ascenzi, Paolo; Marino, Maria

    2015-05-01

    Besides other mechanism(s) 17β-estradiol (E2) facilitates neuronal survival by increasing, via estrogen receptor β (ERβ), the levels of neuroglobin (NGB) an anti-apoptotic protein. In contrast, E2 could exert protective effects in cancer cells by activating apoptosis when the ERβ level prevails on that of ERα as in colon cancer cell lines. These apparently contrasting results raise the possibility that E2-induced NGB up-regulation could regulate the ERβ activities shunning this receptor subtype to trigger an apoptotic cascade in neurons but not in non-neuronal cells. Here, human colorectal adenocarcinoma cell line (DLD-1) that only expresses ERβ and HeLa cells transiently transfected with ERβ encoding vector has been used to verify this hypothesis. In addition, neuroblastoma SK-N-BE cells were used as positive control. Surprisingly, E2 also induced NGB up-regulation, in a dose- and time-dependent manner, in DLD-1 cells. The ERβ-mediated activation of p38/MAPK was necessary for this E2 effect. E2 induced NGB re-allocation in mitochondria where, subsequently to an oxidative stress injury (i.e., 100μM H2O2), NGB interacted with cytochrome c preventing its release into the cytosol and the activation of an apoptotic cascade. As a whole, these results demonstrate that E2-induced NGB up-regulation could act as an oxidative stress sensor, which does not oppose to the pro-apoptotic E2 effect in ERβ-containing colon cancer cells unless a rise of oxidative stress occurs. These results support the concept that oxidative stress plays a critical role in E2-induced carcinogenesis and further open an important scenario to develop novel therapeutic strategies that target NGB against E2-related cancers. PMID:25683270

  11. The negative regulators Foxj1 and Foxo3a are up-regulated by a peptide that inhibits systemic lupus erythematosus-associated T cell responses.

    PubMed

    Sela, Uri; Dayan, Molly; Hershkoviz, Rami; Cahalon, Liora; Lider, Ofer; Mozes, Edna

    2006-11-01

    A peptide (hCDR1) based on the complementarity determining region-1 of an anti-DNA antibody ameliorates systemic lupus erythematosus (SLE) in induced and spontaneous lupus models. Our objectives were to determine the effects of hCDR1 on TCR signaling and on its negative regulators, Foxj1 and Foxo3a. BALB/c mice were immunized with the SLE-inducing anti-DNA antibody, designated 16/6Id, and treated with hCDR1. hCDR1 treatment specifically inhibited IFN-gamma secretion by T cells in association with down-regulated T-bet expression and NF-kappaB activation; however, GATA-3 expression was not affected. Furthermore, TCR signaling (ZAP-70 phosphorylation) was inhibited, and the mRNA expression of the two modulators of Th1 activation, Foxj1 and Foxo3a, was significantly up-regulated. The latter were also elevated in SLE-afflicted (NZBxNZW)F1 mice that were treated with hCDR1. Addition of TGF-beta, which was elevated following treatment with hCDR1, to T cells from 16/6Id immunized mice, up-regulated Foxj1 and Foxo3a mRNA expression, similarly to hCDR1. In contrast, anti-TGF-beta antibodies added to hCDR1-treated T cells abrogated its effect. Thus, hCDR1 elevates TGF-beta, which contributes to the up-regulation of T cell Foxj1 and Foxo3a expression, leading to inhibition of NF-kappaB activation and IFN-gamma secretion, which is required for the maintenance of SLE. PMID:17051618

  12. Prenatal Ethanol Exposure Up-Regulates the Cholesterol Transporters ATP-Binding Cassette A1 and G1 and Reduces Cholesterol Levels in the Developing Rat Brain

    PubMed Central

    Zhou, Chunyan; Chen, Jing; Zhang, Xiaolu; Costa, Lucio G.; Guizzetti, Marina

    2014-01-01

    Aims: Cholesterol plays a pivotal role in many aspects of brain development; reduced cholesterol levels during brain development, as a consequence of genetic defects in cholesterol biosynthesis, leads to severe brain damage, including microcephaly and mental retardation, both of which are also hallmarks of the fetal alcohol syndrome. We had previously shown that ethanol up-regulates the levels of two cholesterol transporters, ABCA1 (ATP binding cassette-A1) and ABCG1, leading to increased cholesterol efflux and decreased cholesterol content in astrocytes in vitro. In the present study we investigated whether similar effects could be seen in vivo. Methods: Pregnant Sprague-Dawley rats were fed liquid diets containing 36% of the calories from ethanol from gestational day (GD) 6 to GD 21. A pair-fed control groups and an ad libitum control group were included in the study. ABCA1 and ABCG1 protein expression and cholesterol and phospholipid levels were measured in the neocortex of female and male fetuses at GD 21. Results: Body weights were decreased in female fetuses as a consequence of ethanol treatments. ABCA1 and ABCG1 protein levels were increased, and cholesterol levels were decreased, in the neocortex of ethanol-exposed female, but not male, fetuses. Levels of phospholipids were unchanged. Control female fetuses fed ad libitum displayed an up-regulation of ABCA1 and a decrease in cholesterol content compared with pair-fed controls, suggesting that a compensatory up-regulation of cholesterol levels may occur during food restriction. Conclusion: Maternal ethanol consumption may affect fetal brain development by increasing cholesterol transporters’ expression and reducing brain cholesterol levels. PMID:25081040

  13. Tumour-derived interleukin 1alpha (IL-1alpha) up-regulates the release of soluble intercellular adhesion molecule-1 (sICAM-1) by endothelial cells.

    PubMed Central

    Fonsatti, E.; Altomonte, M.; Coral, S.; Cattarossi, I.; Nicotra, M. R.; Gasparollo, A.; Natali, P. G.; Maio, M.

    1997-01-01

    Levels of circulating soluble intercellular adhesion molecule-1 (sICAM-1) are elevated in patients affected by solid malignancies; however, the cellular sources generating high levels of sICAM-1 remain to be characterized. Using conditioned media (CM) from seven ICAM-1-positive or -negative neoplastic cells, we demonstrate that tumour-derived interleukin 1alpha (IL-1alpha) significantly (P < 0.05) up-regulates the release of sICAM-1 by human umbilical vein endothelial cells. The intensity of the effect correlated with the amounts of IL-1alpha detectable in CM. Levels of ICAM-1 mRNA were also up-regulated by tumour-secreted IL-1alpha. The up-regulation of the shedding of sICAM-1 and of its expression at protein and mRNA level were completely reversed by the addition of anti-IL-1alpha neutralizing antibodies. Consistent with the in vitro data, tumour endothelia were strongly stained for ICAM-1 compared with autologous normal tissue endothelia. Taken altogether, our observations reveal an IL-1alpha-mediated tumour-endothelium relationship sustaining the shedding of sICAM-1 by endothelial cells. This is a general phenomenon in solid malignancies that correlates with the ability of neoplastic cells to secrete IL-1alpha rather than with their expression of ICAM-1 and/or histological origin. sICAM-1 has been previously shown to inhibit LFA-1/ICAM-1-mediated cell-cell interactions; therefore, the ability of neoplastic cells to secrete IL-1alpha is likely to represent a mechanism for their escape from immune interaction. Images Figure 5 Figure 6 PMID:9374368

  14. hCG-induced Sprouty2 mediates amphiregulin-stimulated COX-2/PGE2 up-regulation in human granulosa cells: a potential mechanism for the OHSS

    PubMed Central

    Cheng, Jung-Chien; Fang, Lanlan; Chang, Hsun-Ming; Sun, Ying-Pu; Leung, Peter C. K.

    2016-01-01

    Sprouty2 (SPRY2) is an important intracellular regulator for epidermal growth factor receptor (EGFR)-mediated ERK1/2 signaling. In human granulosa cells, although SPRY2 is expressed, its regulation and function remains complete unknown and must be defined. Our previous study has shown that human chorionic gonadotropin (hCG)/luteinizing hormone (LH) up-regulates the expression levels of EGF-like growth factor, amphiregulin (AREG), which subsequently contributes to the hCG/LH-induced COX-2 expression and PGE2 production. The aim of the present study was to investigate the effect of hCG on SPRY2 expression and the role of hCG-induced SPRY2 in AREG-stimulated COX-2 expression and PGE2 production in human granulosa cells. Our results demonstrated that the expression of SPRY2 was up-regulated by hCG treatment. Using pharmacological inhibitors and siRNA knockdown, we showed that activation of ERK1/2 signaling was required for hCG-induced up-regulation of SPRY2 expression. Further, SPRY2 knockdown attenuated the AREG-induced COX-2 expression and PGE2 production by inhibiting AREG-activated ERK1/2 signaling. Interestingly, we showed that SPRY2 expression levels were significantly increased in granulosa cells of ovarian hyperstimulation syndrome (OHSS) patients. These results for the first time elucidate the physiological roles of SPRY2 in human granulosa cells and suggest that aberrant expression of SPRY2 may contribute to the pathogenesis of OHSS. PMID:27539669

  15. Gene transcription of TLR2, TLR4, LPS ligands and prostaglandin synthesis enzymes are up-regulated in canine uteri with cystic endometrial hyperplasia-pyometra complex.

    PubMed

    Silva, E; Leitão, S; Henriques, S; Kowalewski, M P; Hoffmann, B; Ferreira-Dias, G; da Costa, L Lopes; Mateus, L

    2010-01-01

    Escherichia coli (E. coli) is the most frequent bacterium isolated in cases of cystic endometrial hyperplasia-pyometra complex, the most frequent endometrial disorder in the bitch. Toll-like receptors (TLRs) play an essential role in the innate immune system. The aim of this study was to compare transcription of genes encoding TLR2, TLR4 and LPS ligands (CD14, MD-2, LBP), prostaglandin synthesis enzymes (COX1, COX2, PGES1 and PGFS), and to compare COX1 and COX2 protein expression and PGE(2) and PGF(2alpha) endometrial content in the endometrium of canine diestrous uteri with (n=7) or without (n=7) pyometra. All cases of pyometra were hyperplastic and E. coli was the only isolated bacteria, while diestrous normal uteri did not present signs of cystic endometrial hyperplasia and were negative for bacteriology. Except for COX1, transcription of all genes was significantly higher in pyometra than in normal endometria. COX1 protein was observed in both normal and pyometra uteri, but COX2 protein was only present in pyometra cases. Endometrial PGE(2) and PGF(2alpha) content were significantly higher in pyometra than in normal diestrous endometria. In conclusion, data obtained in this study provides evidence that pyometra-isolated E. coli induces the up-regulation of TLR2 and TLR4 genes in the canine diestrous endometrium. This up-regulation, which is probably the result of the stimulation by LPS and lipoprotein E. coli constituents, leads to the endometrial up-regulation of PG synthesis genes. This, in turn, results in a higher endometrial concentration of PGE(2) and PGF(2alpha), which may further regulate the local inflammatory response. PMID:19945173

  16. Copper deficiency alters cell bioenergetics and induces mitochondrial fusion through up-regulation of MFN2 and OPA1 in erythropoietic cells

    SciTech Connect

    Bustos, Rodrigo I.; Jensen, Erik L.; Ruiz, Lina M.; Rivera, Salvador; Ruiz, Sebastián; Simon, Felipe; Riedel, Claudia; Ferrick, David; Elorza, Alvaro A.

    2013-08-02

    Highlights: •In copper deficiency, cell proliferation is not affected. In turn, cell differentiation is impaired. •Enlarged mitochondria are due to up-regulation of MNF2 and OPA1. •Mitochondria turn off respiratory chain and ROS production. •Energy metabolism switch from mitochondria to glycolysis. -- Abstract: Copper is essential in cell physiology, participating in numerous enzyme reactions. In mitochondria, copper is a cofactor for respiratory complex IV, the cytochrome c oxidase. Low copper content is associated with anemia and the appearance of enlarged mitochondria in erythropoietic cells. These findings suggest a connection between copper metabolism and bioenergetics, mitochondrial dynamics and erythropoiesis, which has not been explored so far. Here, we describe that bathocuproine disulfonate-induced copper deficiency does not alter erythropoietic cell proliferation nor induce apoptosis. However it does impair erythroid differentiation, which is associated with a metabolic switch between the two main energy-generating pathways. That is, from mitochondrial function to glycolysis. Switching off mitochondria implies a reduction in oxygen consumption and ROS generation along with an increase in mitochondrial membrane potential. Mitochondrial fusion proteins MFN2 and OPA1 were up-regulated along with the ability of mitochondria to fuse. Morphometric analysis of mitochondria did not show changes in total mitochondrial biomass but rather bigger mitochondria because of increased fusion. Similar results were also obtained with human CD34+, which were induced to differentiate into red blood cells. In all, we have shown that adequate copper levels are important for maintaining proper mitochondrial function and for erythroid differentiation where the energy metabolic switch plus the up-regulation of fusion proteins define an adaptive response to copper deprivation to keep cells alive.

  17. Up-regulation of expression of tubulin genes and roles of microtubules in hypergravity-induced growth modification in Arabidopsis hypocotyls

    NASA Astrophysics Data System (ADS)

    Matsumoto, Shouhei; Saito, Yuka; Kumasaki, Saori; Soga, Kouichi; Wakabayashi, Kazuyuki; Hoson, Takayuki

    We examined the roles of microtubules in gravity-induced modification of growth and development in plants by analyzing the expression levels of the α- and β-tubulin gene family and growth behavior of Arabidopsis hypocotyls treated with the microtubule-disrupting reagents colchicine, oryzalin, and propyzamide. Expression of the majority of the examined α- and β-tubulin genes was up-regulated by hypergravity at 300 g, although the extent was variable among genes, indicating that up-regulation of the expression of tubulin genes is the universal response of Arabidopsis hypocotyls to hypergravity. Hypergravity suppressed elongation growth by decreasing the cell-wall extensibility, whereas it stimulated lateral thickening of hypocotyls. By treatment with colchicine, oryzalin, and propyzamide, the elongation growth was suppressed, lateral thickening was stimulated, and the cell-wall extensibility of hypocotyls decreased dose-dependently even under 1 g conditions. The degree of hypergravity-induced changes decreased with increasing concentration of microtubule-disrupting reagents. As a result, hypergravity affected neither the length, the thickness, nor the cell-wall extensibility of hypocotyls in the presence of high concentrations of microtubule-disrupting reagents. Moreover, colchicine-treated seedlings showed helical growth even under 1 g conditions, and this phenotype was intensified under hypergravity conditions. These results suggest that the up-regulation of the expression of tubulin genes is involved in gravity-induced modification of microtubule dynamics, which may play an important role in the resistance of plant organs to the gravitational force and maintenance of normal growth phenotype.

  18. The absence of core fucose up-regulates GnT-III and Wnt target genes: a possible mechanism for an adaptive response in terms of glycan function.

    PubMed

    Kurimoto, Ayako; Kitazume, Shinobu; Kizuka, Yasuhiko; Nakajima, Kazuki; Oka, Ritsuko; Fujinawa, Reiko; Korekane, Hiroaki; Yamaguchi, Yoshiki; Wada, Yoshinao; Taniguchi, Naoyuki

    2014-04-25

    Glycans play key roles in a variety of protein functions under normal and pathological conditions, but several glycosyltransferase-deficient mice exhibit no or only mild phenotypes due to redundancy or compensation of glycan functions. However, we have only a limited understanding of the underlying mechanism for these observations. Our previous studies indicated that 70% of Fut8-deficient (Fut8(-/-)) mice that lack core fucose structure die within 3 days after birth, but the remainder survive for up to several weeks although they show growth retardation as well as emphysema. In this study, we show that, in mouse embryonic fibroblasts (MEFs) from Fut8(-/-) mice, another N-glycan branching structure, bisecting GlcNAc, is specifically up-regulated by enhanced gene expression of the responsible enzyme N-acetylglucosaminyltransferase III (GnT-III). As candidate target glycoproteins for bisecting GlcNAc modification, we confirmed that level of bisecting GlcNAc on β1-integrin and N-cadherin was increased in Fut8(-/-) MEFs. Moreover using mass spectrometry, glycan analysis of IgG1 in Fut8(-/-) mouse serum demonstrated that bisecting GlcNAc contents were also increased by Fut8 deficiency in vivo. As an underlying mechanism, we found that in Fut8(-/-) MEFs Wnt/β-catenin signaling is up-regulated, and an inhibitor against Wnt signaling was found to abrogate GnT-III expression, indicating that Wnt/β-catenin is involved in GnT-III up-regulation. Furthermore, various oxidative stress-related genes were also increased in Fut8(-/-) MEFs. These data suggest that Fut8(-/-) mice adapted to oxidative stress, both ex vivo and in vivo, by inducing various genes including GnT-III, which may compensate for the loss of core fucose functions.

  19. Monocarboxylate transporter 1 is up-regulated in Caco-2 cells by the methionine precursor DL-2-hydroxy-(4-methylthio)butanoic acid.

    PubMed

    Martín-Venegas, Raquel; Brufau, M Teresa; Mañas-Cano, Oriol; Mercier, Yves; Nonis, Magalie K; Ferrer, Ruth

    2014-12-01

    The methionine precursor, DL-2-hydroxy-(4-methylthio)butanoic acid (HMTBA), is a synthetic source of dietary methionine, which is widely used as a poultry nutritional supplement. In the intestinal epithelium, HMTBA transport across the apical membrane is mediated by monocarboxylate transporter 1 (MCT1). The first step in biological utilisation of this methionine precursor is the stereospecific conversion of HMTBA to the corresponding keto acid. In the present study, the regulation of trans-epithelial HMTBA transport was investigated in Caco-2 cell monolayers. Differentiated Caco-2 cells were maintained under control conditions (apical compartment: 0.2 mmol/L L-methionine) or in a HMTBA-enriched medium (2 mmol/L HMTBA). The effect of culture on HMTBA transport was evaluated from apical and basolateral kinetic parameters. MCT1 and MCT4 immuno-localisation and gene expression were investigated by confocal microscopy and real-time quantitative RT-PCR, respectively. The results indicated that apical MCT1 was up-regulated by exposure to HMTBA (1.4-fold increase in Vmax without changes in Km). Moreover, total monolayer MCT1 immunoreactivity increased 1.8-fold in HMTBA-supplemented cultures, this effect mainly being localised at the apical membrane. Functional and immuno-localisation data suggest involvement of MCT1 and MCT4 in basolateral HMTBA transport, although, in this case, no effect was observed for HMTBA-enrichment. Molecular analysis confirmed MCT1 mRNA up-regulation (1.8-fold), with no effect on MCT4 mRNA expression. Thus, exposure to HMTBA up-regulates the trans-epithelial transport of this methionine precursor by increasing the expression and the transport capacity of apical MCT1.

  20. Transcription factor Ets-1 inhibits glucose-stimulated insulin secretion of pancreatic β-cells partly through up-regulation of COX-2 gene expression.

    PubMed

    Zhang, Xiong-Fei; Zhu, Yi; Liang, Wen-Biao; Zhang, Jing-Jing

    2014-08-01

    Increased cyclooxygenase-2 (COX-2) expression is associated with pancreatic β-cell dysfunction. We previously demonstrated that the transcription factor Ets-1 significantly up-regulated COX-2 gene promoter activity. In this report, we used the pancreatic β-cell line INS-1 and isolated rat islets to investigate whether Ets-1 could induce β-cell dysfunction through up-regulating COX-2 gene expression. We investigated the effects of ETS-1 overexpression and the effects of ETS-1 RNA interference on endogenous COX-2 expression in INS-1 cells. We used site-directed mutagenesis and a dual luciferase reporter assay to study putative Ets-1 binding sites in the COX-2 promoter. The effect of ETS-1 1 overexpression on the insulin secretion function of INS-1 cells and rat islets and the potential reversal of these effects by a COX-2 inhibitor were determined in a glucose-stimulated insulin secretion (GSIS) assay. ETS-1 overexpression significantly induces endogenous COX-2 expression, but ETS-1 RNA interference has no effect on basal COX-2 expression in INS-1 cells. Ets-1 protein significantly increases COX-2 promoter activity through the binding site located in the -195/-186 region of the COX-2 promoter. ETS-1 overexpression significantly inhibited the GSIS function of INS-1 cells and islet cells and COX-2 inhibitor treatment partly reversed this effect. These findings indicated that ETS-1 overexpression induces β-cell dysfunction partly through up-regulation of COX-2 gene expression. Moreover, Ets-1, the transcriptional regulator of COX-2 expression, may be a potential target for the prevention of β-cell dysfunction mediated by COX-2.

  1. Extremely Low-Frequency Electromagnetic Fields Promote In Vitro Neuronal Differentiation and Neurite Outgrowth of Embryonic Neural Stem Cells via Up-Regulating TRPC1.

    PubMed

    Ma, Qinlong; Chen, Chunhai; Deng, Ping; Zhu, Gang; Lin, Min; Zhang, Lei; Xu, Shangcheng; He, Mindi; Lu, Yonghui; Duan, Weixia; Pi, Huifeng; Cao, Zhengwang; Pei, Liping; Li, Min; Liu, Chuan; Zhang, Yanwen; Zhong, Min; Zhou, Zhou; Yu, Zhengping

    2016-01-01

    Exposure to extremely low-frequency electromagnetic fields (ELF-EMFs) can enhance hippocampal neurogenesis in adult mice. However, little is focused on the effects of ELF-EMFs on embryonic neurogenesis. Here, we studied the potential effects of ELF-EMFs on embryonic neural stem cells (eNSCs). We exposed eNSCs to ELF-EMF (50 Hz, 1 mT) for 1, 2, and 3 days with 4 hours per day. We found that eNSC proliferation and maintenance were significantly enhanced after ELF-EMF exposure in proliferation medium. ELF-EMF exposure increased the ratio of differentiated neurons and promoted the neurite outgrowth of eNSC-derived neurons without influencing astrocyes differentiation and the cell apoptosis. In addition, the expression of the proneural genes, NeuroD and Ngn1, which are crucial for neuronal differentiation and neurite outgrowth, was increased after ELF-EMF exposure. Moreover, the expression of transient receptor potential canonical 1 (TRPC1) was significantly up-regulated accompanied by increased the peak amplitude of intracellular calcium level induced by ELF-EMF. Furthermore, silencing TRPC1 expression eliminated the up-regulation of the proneural genes and the promotion of neuronal differentiation and neurite outgrowth induced by ELF-EMF. These results suggest that ELF-EMF exposure promotes the neuronal differentiation and neurite outgrowth of eNSCs via up-regulation the expression of TRPC1 and proneural genes (NeuroD and Ngn1). These findings also provide new insights in understanding the effects of ELF-EMF exposure on embryonic brain development.

  2. Testosterone protects female embryonic heart H9c2 cells against severe metabolic stress by activating estrogen receptors and up-regulating IES SUR2B.

    PubMed

    Ballantyne, Thomas; Du, Qingyou; Jovanović, Sofija; Neemo, Andrew; Holmes, Robert; Sinha, Sharabh; Jovanović, Aleksandar

    2013-02-01

    A recent clinical study demonstrated that a testosterone supplementation improves functional capacity in elderly female patients suffering from heart failure. These findings prompted us to consider possible mechanisms of testosterone-induced cardioprotection in females. To address this question we have used a pure female population of rat heart embryonic H9c2 cells. Pre-treatment of cells with testosterone for 24h significantly increased survival of H9c2 cells exposed to 2,4-dinitrophenol (DNP), an inhibitor of oxidative phosphorylation. These cells expressed low level of androgen receptors and the effect of testosterone was not modified by hydroxyflutamide, an antagonist of androgen receptor. In contrast, cyclohexamide, an inhibitor of protein biosynthesis, and tamoxifene, a partial agonist of estrogen receptors, abolished cardioprotection afforded by testosterone. In addition, finasteride, an inhibitor of 5α-reductase, and anastrazole, an inhibitor of α-aromatase, also blocked testosterone-induced cytoprotection. Real time RT-PCR revealed that testosterone did not regulate the expression of nine subunits and accessory proteins of sarcolemmal ATP-sensitive K(+) (K(ATP)) channels. On the other hand, testosterone, as well as 17β-estradiol, up-regulated a putative mitochondrial K(ATP) channel subunit, mitochondrial sulfonylurea receptor 2B intraexonics splice variant (IES SUR2B), without affecting expression of IES SUR2A. Tamoxifene inhibited testosterone-induced up-regulation of IES SUR2B without affecting IES SUR2A. In conclusion, this study has shown that testosterone protect female embryonic heart H9c2 cells against severe metabolic stress by its conversion into metabolites that activate estrogen receptors and up-regulate IES SUR2B.

  3. The Inhibitory Effects of Anti-Oxidants on Ultraviolet-Induced Up-Regulation of the Wrinkling-Inducing Enzyme Neutral Endopeptidase in Human Fibroblasts.

    PubMed

    Nakajima, Hiroaki; Terazawa, Shuko; Niwano, Takao; Yamamoto, Yorihiro; Imokawa, Genji

    2016-01-01

    We recently reported that the over-expression of skin fibroblast-derived neutral endopeptidase (NEP) plays a pivotal role in impairing the three-dimensional architecture of dermal elastic fibers during the biological mechanism of ultraviolet (UV)-induced skin wrinkling. In that process, a UVB-associated epithelial-mesenchymal cytokine interaction as well as a direct UVA-induced cellular stimulation are associated with the up-regulation of NEP in human fibroblasts. In this study, we characterized the mode of action of ubiquinol10 which may abrogate the up-regulation of NEP by dermal fibroblasts, resulting in a reported in vivo anti-wrinkling action, and compared that with 3 other anti-oxidants, astaxanthin (AX), riboflavin (RF) and flavin mononucleotide (FMN). Post-irradiation treatment with all 4 of those anti-oxidants elicited an interrupting effect on the UVB-associated epithelial-mesenchymal cytokine interaction leading to the up-regulation of NEP in human fibroblasts but with different modes of action. While AX mainly served as an inhibitor of the secretion of wrinkle-inducing cytokines, such as interleukin-1α (IL-1α) and granulocyte macrophage colony stimulatory factor (GM-CSF) in UVB-exposed epidermal keratinocytes, ubiquinol10, RF and FMN predominantly interrupted the IL-1α and GM-CSF-stimulated expression of NEP in dermal fibroblasts. On the other hand, as for the UVA-associated mechanism, similar to the abrogating effects reported for AX and FMN, ubiquinol10 but not RF had the potential to abrogate the increased expression of NEP and matrix-metalloproteinase-1 in UVA-exposed human fibroblasts. Our findings strongly support the in vivo anti-wrinkling effects of ubiquinol10 and AX on human and animal skin and provide convincing proof of the UV-induced wrinkling mechanism that essentially focuses on the over-expression of NEP by dermal fibroblasts as an intrinsic causative factor. PMID:27648570

  4. Tissue-specific up-regulation of arginase I and II induced by p38 MAPK mediates endothelial dysfunction in type 1 diabetes mellitus

    PubMed Central

    Pernow, J; Kiss, A; Tratsiakovich, Y; Climent, B

    2015-01-01

    Background and Purpose Emerging evidence suggests a selective up-regulation of arginase I in diabetes causing coronary artery disease; however, the mechanisms behind this up-regulation are still unknown. Activated p38 MAPK has been reported to increase arginase II in various cardiovascular diseases. We therefore tested the role of p38 MAPK in the regulation of arginase I and II expression and its effect on endothelial dysfunction in diabetes mellitus. Experimental Approach Endothelial function was determined in septal coronary (SCA), left anterior descending coronary (LAD) and mesenteric (MA) arteries from healthy and streptozotocin-induced diabetic Wistar rats by wire myographs. Arginase activity and protein levels of arginase I, II, phospho-p38 MAPK and phospho-endothelial NOS (eNOS) (Ser1177) were determined in these arteries from diabetic and healthy rats treated with a p38 MAPK inhibitor in vivo. Key Results Diabetic SCA and MA displayed impaired endothelium-dependent relaxation, which was prevented by arginase and p38 MAPK inhibition while LAD relaxation was not affected. Arginase I, phospho-p38 MAPK and eNOS protein expression was increased in diabetic coronary arteries. In diabetic MA, however, increased expression of arginase II and phospho-p38 MAPK, increased arginase activity and decreased expression of eNOS were observed. All these effects were reversed by p38 MAPK inhibition. Conclusions and Implications Diabetes-induced activation of p38 MAPK causes endothelial dysfunction via selective up-regulation of arginase I expression in coronary arteries and arginase II expression in MA. Therefore, regional differences appear to exist in the arginase isoforms contributing to endothelial dysfunction in type 1 diabetes mellitus. PMID:26140333

  5. A peptide based on the complementarity-determining region 1 of an autoantibody ameliorates lupus by up-regulating CD4+CD25+ cells and TGF-beta.

    PubMed

    Sharabi, Amir; Zinger, Heidy; Zborowsky, Maya; Sthoeger, Zev M; Mozes, Edna

    2006-06-01

    Systemic lupus erythematosus is an autoimmune disease characterized by autoantibodies and systemic clinical manifestations. A peptide, designated hCDR1, based on the complementarity-determining region (CDR) 1 of an autoantibody, ameliorated the serological and clinical manifestations of lupus in both spontaneous and induced murine models of lupus. The objectives of the present study were to determine the mechanism(s) underlying the beneficial effects induced by hCDR1. Adoptive transfer of hCDR1-treated cells to systemic lupus erythematosus-afflicted (NZBxNZW)F1 female mice down-regulated all disease manifestations. hCDR1 treatment up-regulated (by 30-40%) CD4+CD25+ cells in association with CD45RBlow, cytotoxic T lymphocyte antigen 4, and Foxp3 expression. Depletion of the CD25+ cells diminished significantly the therapeutic effects of hCDR1, whereas administration of the enriched CD4+CD25+ cell population was beneficial to the diseased mice. Amelioration of disease manifestations was associated with down-regulation of the pathogenic cytokines (e.g., IFN-gamma and IL-10) and up-regulation of the immunosuppressive cytokine TGF-beta, which substantially contributed to the suppressed autoreactivity. TGF-beta was secreted by CD4+ cells that were affected by hCDR1-induced immunoregulatory cells. The hCDR1-induced CD4+CD25+ cells suppressed autoreactive CD4+ cells, resulting in reduced rates of activation-induced apoptosis. Thus, hCDR1 ameliorates lupus through the induction of CD4+CD25+ cells that suppress activation of the autoreactive cells and trigger the up-regulation of TGF-beta.

  6. Antisense-mediated depletion of p300 in human cells leads to premature G1 exit and up-regulation of c-MYC.

    PubMed

    Kolli, S; Buchmann, A M; Williams, J; Weitzman, S; Thimmapaya, B

    2001-04-10

    The cAMP-response element-binding protein (CREB)-binding protein and p300 are two highly conserved transcriptional coactivators and histone acetyltransferases that integrate signals from diverse signal transduction pathways in the nucleus and also link chromatin remodeling with transcription. In this report, we have examined the role of p300 in the control of the G(1) phase of the cell cycle in nontransformed immortalized human breast epithelial cells (MCF10A) and fibroblasts (MSU) by using adenovirus vectors expressing p300-specific antisense sequences. Quiescent MCF10A and MSU cells expressing p300-specific antisense sequences synthesized p300 at much reduced levels and exited G(1) phase without serum stimulation. These cells also showed an increase in cyclin A and cyclin A- and E-associated kinase activities characteristic of S phase induction. Further analysis of the p300-depleted quiescent MCF10A cells revealed a 5-fold induction of c-MYC and a 2-fold induction of c-JUN. A direct target of c-MYC, CAD, which is required for DNA synthesis, was also found to be up-regulated, indicating that up-regulation of c-MYC functionally contributed to DNA synthesis. Furthermore, S phase induction in p300-depleted cells was reversed when antisense c-MYC was expressed in these cells, indicating that up-regulation of c-MYC may directly contribute to S phase induction. Adenovirus E1A also induced DNA synthesis and increased the levels of c-MYC and c-JUN in serum-starved MCF10A cells in a p300-dependent manner. Our results suggest an important role of p300 in cell cycle regulation at G(1) and raise the possibility that p300 may negatively regulate early response genes, including c-MYC and c-JUN, thereby preventing DNA synthesis in quiescent cells.

  7. Exposure of LS-180 Cells to Drugs of Diverse Physicochemical and Therapeutic Properties Up-regulates P-glycoprotein Expression and Activity

    PubMed Central

    Abuznait, Alaa H.; Patrick, Shawn G.; Kaddoumi, Amal

    2011-01-01

    Purpose Drug transporters are increasingly recognized as important determinants of variability in drug disposition and therapeutic response, both in pre-clinical and clinical stages of drug development process. The role P-glycoprotein (P-gp) plays in drug interactions via its inhibition is well established. However, much less knowledge is available about drugs effect on P-gp up-regulation. The objective of this work was to in vitro investigate and rank commonly used drugs according to their potencies to up-regulate P-gp activity utilizing the same experimental conditions. Methods The in vitro potencies of several drugs of diverse physicochemical and therapeutic properties including rifampicin, dexamethasone, caffeine, verapamil, pentylenetetrazole, hyperforin, and β-estradiol over broad concentration range to up-regulate P-gp expression and activity were examined. For dose-response studies, LS-180 cells were treated with different concentrations of the selected drugs followed by P-gp protein and gene expressions analyses. P-gp functionality was determined by uptake studies with rhodamine 123 as a P-gp substrate, followed by Emax/EC50 evaluation. Results The results demonstrated a dose-dependent increase in P-gp expression and activity following treatments. At 50 μM concentration (hyperforin, 0.1 μM), examined drugs increased P-gp protein and gene expressions by up to 5.5 and 6.2-fold, respectively, while enhanced P-gp activity by 1.8–4-fold. The rank order of these drugs potencies to up-regulate P-gp activity was as following: hyperforin ⋙ dexamethasone ≈ β-estradiol > caffeine > rifampicin ≈ pentylenetetrazole > verapamil. Conclusions These drugs have the potential to be involved in drug interactions when administered with other drugs that are P-gp substrates. Further studies are needed to in vivo evaluate these drugs and verify the consequences of such induction on P-gp activity for in vitro-in vivo correlation purposes. PMID:21733412

  8. The mitochondrial uncoupler DNP triggers brain cell mTOR signaling network reprogramming and CREB pathway up-regulation.

    PubMed

    Liu, Dong; Zhang, Yongqing; Gharavi, Robert; Park, Hee Ra; Lee, Jaewon; Siddiqui, Sana; Telljohann, Richard; Nassar, Matthew R; Cutler, Roy G; Becker, Kevin G; Mattson, Mark P

    2015-08-01

    Mitochondrial metabolism is highly responsive to nutrient availability and ongoing activity in neuronal circuits. The molecular mechanisms by which brain cells respond to an increase in cellular energy expenditure are largely unknown. Mild mitochondrial uncoupling enhances cellular energy expenditure in mitochondria and can be induced with 2,4-dinitrophenol (DNP), a proton ionophore previously used for weight loss. We found that DNP treatment reduces mitochondrial membrane potential, increases intracellular Ca(2+) levels and reduces oxidative stress in cerebral cortical neurons. Gene expression profiling of the cerebral cortex of DNP-treated mice revealed reprogramming of signaling cascades that included suppression of the mammalian target of rapamycin (mTOR) and insulin--PI3K - MAPK pathways, and up-regulation of tuberous sclerosis complex 2, a negative regulator of mTOR. Genes encoding proteins involved in autophagy processes were up-regulated in response to DNP. CREB (cAMP-response element-binding protein) signaling, Arc and brain-derived neurotrophic factor, which play important roles in synaptic plasticity and adaptive cellular stress responses, were up-regulated in response to DNP, and DNP-treated mice exhibited improved performance in a test of learning and memory. Immunoblot analysis verified that key DNP-induced changes in gene expression resulted in corresponding changes at the protein level. Our findings suggest that mild mitochondrial uncoupling triggers an integrated signaling response in brain cells characterized by reprogramming of mTOR and insulin signaling, and up-regulation of pathways involved in adaptive stress responses, molecular waste disposal, and synaptic plasticity. Physiological bioenergetic challenges such as exercise and fasting can enhance neuroplasticity and protect neurons against injury and neurodegeneration. Here, we show that the mitochondrial uncoupling agent 2,4-dinitrophenol (DNP) elicits adaptive signaling responses in the

  9. Substance P primes lipoteichoic acid- and Pam3CysSerLys4-mediated activation of human mast cells by up-regulating Toll-like receptor 2.

    PubMed

    Tancowny, Brian P; Karpov, Victor; Schleimer, Robert P; Kulka, Marianna

    2010-10-01

    Substance P (SP) is a neuropeptide with neuroimmunoregulatory activity that may play a role in susceptibility to infection. Human mast cells, which are important in innate immune responses, were analysed for their responses to pathogen-associated molecules via Toll-like receptors (TLRs) in the presence of SP. Human cultured mast cells (LAD2) were activated by SP and TLR ligands including lipopolysaccharide (LPS), Pam3CysSerLys4 (Pam3CSK4) and lipoteichoic acid (LTA), and mast cell leukotriene and chemokine production was assessed by enzyme-linked immunosorbent assay (ELISA) and gene expression by quantitative PCR (qPCR). Mast cell degranulation was determined using a β-hexosaminidase (β-hex) assay. SP treatment of LAD2 up-regulated mRNA for TLR2, TLR4, TLR8 and TLR9 while anti-immunoglobulin E (IgE) stimulation up-regulated expression of TLR4 only. Flow cytometry and western blot confirmed up-regulation of TLR2 and TLR8. Pretreatment of LAD2 with SP followed by stimulation with Pam3CSK4 or LTA increased production of leukotriene C4 (LTC(4) ) and interleukin (IL)-8 compared with treatment with Pam3CSK4 or LTA alone (>2-fold; P<0·01). SP alone activated 5-lipoxygenase (5-LO) nuclear translocation but also augmented Pam3CSK4 and LTA-mediated 5-LO translocation. Pam3CSK4, LPS and LTA did not induce LAD2 degranulation. SP primed LTA and Pam3CSK4-mediated activation of JNK, p38 and extracellular-signal-regulated kinase (ERK) and activated the nuclear translocation of c-Jun, nuclear factor (NF)-κB, activating transcription factor 2 (ATF-2) and cyclic-AMP-responsive element binding protein (CREB) transcription factors. Pretreatment with SP followed by LTA stimulation synergistically induced production of chemokine (C-X-C motif) ligand 8 (CXCL8)/IL-8, chemokine (C-C motif) ligand 2 (CCL2)/monocyte chemotactic protein 1 (MCP-1), tumour necrosis factor (TNF) and IL-6 protein. SP primes TLR2-mediated activation of human mast cells by up-regulating TLR expression and

  10. EsMlp, a muscle-LIM protein gene, is up-regulated during cold exposure in the freeze-avoiding larvae of Epiblema scudderiana.

    PubMed

    Bilgen, T; English, T E; McMullen, D C; Storey, K B

    2001-08-01

    Screening of a cDNA library identified transcripts that were up-regulated by cold (4 or -20 degrees C) exposure in larvae of the freeze-avoiding goldenrod gall moth, Epiblema scudderiana. One clone contained a full-length open reading frame encoding a protein of 94 amino acids. The gene product, with 79.1% of residues identical with the Drosophila LIM protein Mlp60A, was named EsMlp and contained a single LIM domain and consensus sequences characteristic of a LIM protein. Transcript levels rose approx twofold when larvae were shifted from 4 to -20 degrees C and approx threefold over the midwinter months compared with larvae sampled in October or April. EsMlp expression was high in larval head (possibly due to expression in pharyngeal muscles) and body wall but was not detected in fat body. Immunoblotting revealed a three- to fourfold increase in EsMlp protein in midwinter larvae (January-February) compared with November-collected animals and a further rise to eightfold higher than November values in larvae collected in April. Cold up-regulation of EsMlp and the pattern of EsMlp levels in the larvae suggest possible roles for the protein, such as in muscle maintenance over the winter or as a preparative function that could facilitate the rapid resumption of development and metamorphosis when environmental temperatures rise in the spring.

  11. The BPV-4 co-carcinogen quercetin induces cell cycle arrest and up-regulates transcription from the LCR of BPV-4.

    PubMed

    Connolly, J A; Morgan, I M; Jackson, M E; Campo, M S

    1998-05-28

    Bracken fern is the environmental co-carcinogen of BPV-4 in the induction of neoplasias of the upper alimentary canal of cattle. The flavonoid quercetin is one of the most potent and best characterised mutagens present in the fern. We have shown that transfection with BPV-4 DNA and exposure to a single dose of quercetin leads to tumorigenic transformation of primary bovine cells. We now show that quercetin induces cell cycle arrest and up-regulates transcription from the BPV-4 long control region (LCR). This up-regulation is mediated by a 21 nucleotide-long cis-element in the LCR, designated QRE-1, which is located immediately downstream of the TATA box. Cellular proteins bind to QRE-1 and removal or substitution of QRE-1 lead to the abrogation of the response to quercetin. As expression of the viral oncogenes is controlled by the LCR, perturbation in this control and increased oncoprotein expression are likely to contribute to fully malignant cell transformation by overcoming the cell cycle arrest induced by quercetin, thus forcing damaged cells to proliferate.

  12. TNF-alpha increases ubiquitin-conjugating activity in skeletal muscle by up-regulating UbcH2/E220k

    NASA Technical Reports Server (NTRS)

    Li, Yi-Ping; Lecker, Stewart H.; Chen, Yuling; Waddell, Ian D.; Goldberg, Alfred L.; Reid, Michael B.

    2003-01-01

    In some inflammatory diseases, TNF-alpha is thought to stimulate muscle catabolism via an NF-kappaB-dependent process that increases ubiquitin conjugation to muscle proteins. The transcriptional mechanism of this response has not been determined. Here we studied the potential role of UbcH2, a ubiquitin carrier protein and homologue of murine E220k. We find that UbcH2 is constitutively expressed by human skeletal and cardiac muscles, murine limb muscle, and cultured myotubes. TNF-alpha stimulates UbcH2 expression in mouse limb muscles in vivo and in cultured myotubes. The UbcH2 promoter region contains a functional NF-kappaB binding site; NF-kappaB binding to this sequence is increased by TNF-alpha stimulation. A dominant negative inhibitor of NF-kappaB activation blocks both UbcH2 up-regulation and the increase in ubiquitin-conjugating activity stimulated by TNF-alpha. In extracts from TNF-alpha-treated myotubes, ubiquitin-conjugating activity is limited by UbcH2 availability; activity is inhibited by an antiserum to UbcH2 or a dominant negative mutant of UbcH2 and is enhanced by wild-type UbcH2. Thus, UbcH2 up-regulation is a novel response to TNF-alpha/NF-kappaB signaling in skeletal muscle that appears to be essential for the increased ubiquitin conjugation induced by this cytokine.

  13. Ginsenoside Rg3 up-regulates the expression of vascular endothelial growth factor in human dermal papilla cells and mouse hair follicles.

    PubMed

    Shin, Dae Hyun; Cha, Youn Jeong; Yang, Kyeong Eun; Jang, Ik-Soon; Son, Chang-Gue; Kim, Bo Hyeon; Kim, Jung Min

    2014-07-01

    Crude Panax ginseng has been documented to possess hair growth activity and is widely used to treat alopecia, but the effects of ginsenoside Rg3 on hair growth have not to our knowledge been determined. The aim of the current study was to identify the molecules through which Rg3 stimulates hair growth. The thymidine incorporation for measuring cell proliferation was determined. We used DNA microarray analysis to measure gene expression levels in dermal papilla (DP) cells upon treatment with Rg3. The mRNA and protein expression levels of vascular endothelial growth factor (VEGF) in human DP cells were measured by real-time polymerase chain reaction and immunohistochemistry, respectively. We also used immunohistochemistry assays to detect in vivo changes in VEGF and 3-stemness marker expressions in mouse hair follicles. Reverse transcription polymerase chain reaction showed dose-dependent increases in VEGF mRNA levels on treatment with Rg3. Immunohistochemical analysis showed that expression of VEGF was significantly up-regulated by Rg3 in a dose-dependent manner in human DP cells and in mouse hair follicles. In addition, the CD8 and CD34 were also up-regulated by Rg3 in the mouse hair follicles. It may be concluded that Rg3 might increase hair growth through stimulation of hair follicle stem cells and it has the potential to be used in hair growth products.

  14. Dehydrin2 is a stress-inducible, whereas Dehydrin1 is constitutively expressed but up-regulated gene under varied cues in tea [Camellia sinensis (L.) O. Kuntze].

    PubMed

    Paul, Asosii; Kumar, Sanjay

    2013-05-01

    Two expressed sequence tags with similarity to dehydrin were cloned to full-length (CsDHN1 and CsDHN2) through rapid amplification of cDNA ends. CsDHN1 and CsDHN2 were 1,027 and 992 base pair long and encoded for predicted polypeptides of 260 and 201 amino acids, respectively. Deduced CsDHN1 protein had a S-segment and three lysine-rich consensus motifs (K-segments). The Y-segment was, however, absent in the deduced CsDHN1. CsDHN2 had three Y motifs, one S-segment and two K-segments. Expression of CsDHN1 was visible at all the time points of study, though up-regulation was observed in response to winter dormancy (WD) as well as abiotic stresses [low temperature, sodium chloride, polyethylene glycol, and hydrogen peroxide]. Expression of CsDHN2 was strongly up-regulated within 1 h of exposure to abiotic stresses as well in the tissue harvested during WD in contrast to the respective "control" for abiotic stresses and in tissue during the period of active growth, when the expression was not detectable.

  15. Up-regulation of the extracellular matrix glycoprotein tenascin-R during axonal reorganization and astrogliosis in the adult rat hippocampus.

    PubMed

    Brenneke, Franziska; Schachner, Melitta; Elger, Christian E; Lie, Ailing A

    2004-02-01

    Interactions between cells and extracellular matrix (ECM) molecules play a crucial role during brain development. The ECM glycoprotein tenascin-R (TN-R) has been implicated in the control of axon targeting, neural cell adhesion, migration and differentiation. Here, we have focused on the putative role of TN-R in chronic brain diseases involving increased neuronal excitability, as found in epilepsy. An episode of pilocarpine-induced status epilepticus (SE) led over a period of 3-30 days to neuron loss in the hippocampal hilus, CA3 and CA1 with reactive mossy fiber sprouting and astrogliosis in these regions. We found a focal up-regulation of granular TN-R immunoreactivity within the neuropil of segments of the CA3 pyramidal cell layer, the extent of this up-regulation paralleled the degree of pyramidal cell loss, mossy fiber sprouting and astrogliosis in these CA3 segments. In contrast, parvalbumin immunoreactivity and Wisteria floribundi agglutinin (WFA)-labeled perineuronal nets were reduced in CA3 segments with neuronal cell loss. The parallel development of increase in focal granular TN-R immunoreactivity, reactive mossy fiber sprouting and astrogliosis in CA3 implies a role for TN-R in axon targeting and synapse formation and/or in astrocytic targeting and interactions with the ECM during lesion-induced sprouting in the adult brain.

  16. Membrane lipid modification by polyunsaturated fatty acids sensitizes oligodendroglial OLN-93 cells against oxidative stress and promotes up-regulation of heme oxygenase-1 (HSP32).

    PubMed

    Brand, Annette; Bauer, Nina G; Hallott, Amanda; Goldbaum, Olaf; Ghebremeskel, Kebreab; Reifen, Ram; Richter-Landsberg, Christiane

    2010-04-01

    Polyunsaturated fatty acids (PUFA) are highly abundant in brain tissue, and docosahexaenoic acid (DHA) might protect cells from oxidative stress (OS) during inflammation and demyelinating disorders, but also might exert pro-oxidant effects. Here we investigated if PUFA supplements lead to heat shock protein induction, altered cell survival properties and stress responses to OS exerted by hydrogen peroxide in oligodendroglial OLN-93 cells. The data show that supplements of various fatty acids (FA) with 18-22 carbons chain length and 2-6 double bonds led to PUFA enrichment in cellular membranes. Depending on the degree of desaturation, FA-supplements caused the up-regulation of heme oxygenase-1 (HSP32), a stress protein inducible by OS, and an increase in sensitivity to hydrogen peroxide-treatment. DHA, with the highest number of double bonds, was most effective. Co-treatment with DHA and the lipophilic vitamin E analogue alpha-tocopherol, suppressed heme oxygenase-1 up-regulation and cell survival was restored. Analysis of the lipid profile demonstrates that alpha-tocopherol not only has antioxidant capacities, but also directly modified the PUFA profile in cell membranes. Enrichment with higher omega-3, -6 and -9 PUFA and an increase in the biosynthesis rate of very long chain fatty acids, mainly changed the FA profile of ethanolamine and serine phosphoglycerides.

  17. P-selectin glycoprotein ligand-1 (PSGL-1) is up-regulated on leucocytes from patients with chronic obstructive pulmonary disease

    PubMed Central

    Schumacher, A; Liebers, U; John, M; Gerl, V; Meyer, M; Witt, C; Wolff, G

    2005-01-01

    Chronic obstructive pulmonary disease (COPD) is characterized by a dysregulated recruitment of circulating leucocytes into the lung which is associated with the onset and progress of the disease. P-selectin glycoprotein ligand-1 (PSGL-1) is expressed on leucocytes and plays an essential role in primary leucocyte-endothelial cell adhesive contacts. The present study investigated if PSGL-1 is up-regulated on leucocytes of COPD patients. Peripheral blood samples were collected from COPD patients as well as controls (smoking, nonsmoking volunteers) and subjected to analysis of PSGL-1 expression on leucocytes, i.e. neutrophils, eosinophils, monocytes and lymphocytes by flow cytometry. No significant difference was observed between healthy nonsmoking and healthy smoking control subjects. In contrast, PSGL-1 expression was found to be significantly increased on the surface of all four leucocyte populations in COPD patients compared to both control groups. The finding that PSGL-1 surface expression is up-regulated on leucocytes of COPD patients as compared to leucocytes of controls suggests PSGL-1 as a potential target for anti-inflammatory treatment. PMID:16232226

  18. The Aldo-Keto Reductase AKR1B10 Is Up-Regulated in Keloid Epidermis, Implicating Retinoic Acid Pathway Dysregulation in the Pathogenesis of Keloid Disease.

    PubMed

    Jumper, Natalie; Hodgkinson, Tom; Arscott, Guyan; Har-Shai, Yaron; Paus, Ralf; Bayat, Ardeshir

    2016-07-01

    Keloid disease is a recurrent fibroproliferative cutaneous tumor of unknown pathogenesis for which clinical management remains unsatisfactory. To obtain new insights into hitherto underappreciated aspects of keloid pathobiology, we took a laser capture microdissection-based, whole-genome microarray analysis approach to identify distinct keloid disease-associated gene expression patterns within defined keloid regions. Identification of the aldo-keto reductase enzyme AKR1B10 as highly up-regulated in keloid epidermis suggested that an imbalance of retinoic acid metabolism is likely associated with keloid disease. Here, we show that AKR1B10 transfection into normal human keratinocytes reproduced the abnormal retinoic acid pathway expression pattern we had identified in keloid epidermis. Cotransfection of AKR1B10 with a luciferase reporter plasmid showed reduced retinoic acid response element activity, supporting the hypothesis of retinoic acid synthesis deficiency in keloid epidermis. Paracrine signals released by AKR1B10-overexpressing keratinocytes into conditioned medium resulted in up-regulation of transforming growth factor-β1, transforming growth factor-β2, and collagens I and III in both keloid and normal skin fibroblasts, mimicking the typical profibrotic keloid profile. Our study results suggest that insufficient retinoic acid synthesis by keloid epidermal keratinocytes may contribute to the pathogenesis of keloid disease. We refocus attention on the role of injured epithelium in keloid disease and identify AKR1B10 as a potential new target in future management of keloid disease. PMID:27025872

  19. Down-Regulation of Canonical and Up-Regulation of Non-Canonical Wnt Signalling in the Carcinogenic Process of Squamous Cell Lung Carcinoma

    PubMed Central

    Weich, Alexander; Kiss, Edit; Barko, Szilvia; Kovacs, Tamas; Avdicevic, Monika; D’Souza, Vijay K.; Rapp, Judit; Kvell, Krisztian; Jakab, Laszlo; Nyitrai, Miklos; Molnar, Tamas F.; Thickett, David R.; Laszlo, Terezia; Pongracz, Judit E.

    2013-01-01

    The majority of lung cancers (LC) belong to the non-small cell lung carcinoma (NSCLC) type. The two main NSCLC sub-types, namely adenocarcinoma (AC) and squamous cell carcinoma (SCC), respond differently to therapy. Whereas the link between cigarette smoke and lung cancer risk is well established, the relevance of non-canonical Wnt pathway up-regulation detected in SCC remains poorly understood. The present study was undertaken to investigate further the molecular events in canonical and non-canonical Wnt signalling during SCC development. A total of 20 SCC and AC samples with matched non-cancerous controls were obtained after surgery. TaqMan array analysis confirmed up-regulation of non-canonical Wnt5a and Wnt11 and identified down-regulation of canonical Wnt signalling in SCC samples. The molecular changes were tested in primary small airway epithelial cells (SAEC) and various lung cancer cell lines (e.g. A549, H157, etc). Our studies identified Wnt11 and Wnt5a as regulators of cadherin expression and potentiated relocation of β-catenin to the nucleus as an important step in decreased cellular adhesion. The presented data identifies additional details in the regulation of SCC that can aid identification of therapeutic drug targets in the future. PMID:23505429

  20. Exogenous NAD(+) decreases oxidative stress and protects H2O2-treated RPE cells against necrotic death through the up-regulation of autophagy.

    PubMed

    Zhu, Ying; Zhao, Ke-Ke; Tong, Yao; Zhou, Ya-Li; Wang, Yi-Xiao; Zhao, Pei-Quan; Wang, Zhao-Yang

    2016-05-31

    Increased oxidative stress, which can lead to the retinal pigment epithelium (RPE) cell death by inducing ATP depletion and DNA repair, is believed to be a prominent pathology in age-related macular degeneration (AMD). In the present study, we showed that and 0.1 mM nicotinamide adenine dinucleotide (NAD(+)) administration significantly blocked RPE cell death induced by 300 μM H2O2. Further investigation showed that H2O2 resulted in increased intracellular ROS level, activation of PARP-1 and subsequently necrotic death of RPE cells. Exogenous NAD(+) administration significantly decreased intracellular and intranuclear ROS levels in H2O2-treated RPE cells. In addition, NAD(+) administration to H2O2-treated RPE cells inhibited the activation of PARP-1 and protected the RPE cells against necrotic death. Moreover, exogenous NAD(+) administration up-regulated autophagy in the H2O2-treated RPE cells. Inhibition of autophagy by LY294002 blocked the decrease of intracellular and intranuclear ROS level. Besides, inhibition of autophagy by LY294002 abolished the protection of exogenous NAD(+) against H2O2-induced cell necrotic death. Taken together, our findings indicate that that exogenous NAD(+) administration suppresses H2O2-induced oxidative stress and protects RPE cells against PARP-1 mediated necrotic death through the up-regulation of autophagy. The results suggest that exogenous NAD(+) administration might be potential value for the treatment of AMD.

  1. Novel mechanism of harmaline on inducing G2/M cell cycle arrest and apoptosis by up-regulating Fas/FasL in SGC-7901 cells.

    PubMed

    Wang, Yihai; Wang, Chunhua; Jiang, Chenguang; Zeng, Hong; He, Xiangjiu

    2015-01-01

    Harmaline (HAR), a natural occurrence β-carboline alkaloid, was isolated from the seeds of Peganum harmala and exhibited potent antitumor effect. In this study, the anti-gastric tumor effects of HAR were firstly investigated in vitro and in vivo. The results strongly showed that HAR could inhibit tumor cell proliferation and induce G2/M cell cycle arrest accompanied by an increase in apoptotic cell death in SGC-7901 cancer cells. HAR could up-regulate the expressions of cell cycle-related proteins of p-Cdc2, p21, p-p53, Cyclin B and down-regulate the expression of p-Cdc25C. In addition, HAR could up-regulate the expressions of Fas/FasL, activated Caspase-8 and Caspase-3. Moreover, blocking Fas/FasL signaling could markedly inhibit the apoptosis caused by HAR, suggesting that Fas/FasL mediated pathways were involved in HAR-induced apoptosis. Interestingly, HAR could also exert on antitumor activity with a dose of 15 mg/kg/day in vivo, which was also related with cell cycle arrest. These new findings provided a framework for further exploration of HAR which possess the potential antitumor activity by inducing cell cycle arrest and apoptosis. PMID:26678950

  2. Up-regulation of carbon metabolism-related glyoxylate cycle and toxin production in Beauveria bassiana JEF-007 during infection of bean bug, Riptortus pedestris (Hemiptera: Alydidae).

    PubMed

    Yang, Yi-Ting; Lee, Se Jin; Nai, Yu-Shin; Kim, Sihyeon; Kim, Jae Su

    2016-10-01

    Beauveria bassiana (Bb) is used as an environment-friendly biopesticide. However, the molecular mechanisms of Bb-host interactions are not well understood. Herein, RNA isolated from B. bassiana (Bb JEF-007) and Riptortus pedestris (Hemiptera: Alydidae) infected with this strain were firstly subjected to high-throughput next generation sequencing (NGS) to analyze and compare transcriptomes. Due to lack of fungal and host genome information, fungal transcriptome was processed to partially exclude non-infection specific genes and host-flora. Differentially Expressed Gene (DEG) analysis showed that 2381 genes were up-regulated and 2303 genes were down-regulated upon infection. Most DEGs were classified into the categories of single-organism, cellular and metabolism processes by Gene Ontology analysis. Most DEGs were involved in metabolic pathways based on Kyoto Encyclopedia of Genes and Genomes pathway mapping. Carbon metabolism-related enzymes in the glyoxylate cycle were significantly up-regulated, suggesting a possible role for them in Bb growth in the host. Additionally, transcript levels of several fungal genes were dramatically increased after infection, such as cytotoxic lectin-like protein, bacterial-like toxin, proteins related to cell wall formation, hyphal growth, nutrient uptake, and halogenated compound synthesis. This work provides insight into how entomopathogenic B. bassiana grows in agriculturally harmful bean bug at 6 d post infection. PMID:27647240

  3. miR-6734 Up-Regulates p21 Gene Expression and Induces Cell Cycle Arrest and Apoptosis in Colon Cancer Cells

    PubMed Central

    Kang, Moo Rim; Park, Ki Hwan; Yang, Jeong-Ook; Lee, Chang Woo; Oh, Soo Jin; Yun, Jieun; Lee, Myeong Youl; Han, Sang-Bae; Kang, Jong Soon

    2016-01-01

    Recently, microRNAs have been implicated in the regulation of gene expression in terms of both gene silencing and gene activation. Here, we investigated the effects of miR-6734, which has a sequence homology with a specific region of p21WAF1/CIP1 (p21) promoter, on cancer cell growth and the mechanisms involved in this effect. miR-6734 up-regulated p21 expression at both mRNA and protein levels and chromatin immunoprecipitation analysis using biotin-labeled miR-6734 confirmed the association of miR-6734 with p21 promoter. Moreover, miR-6734 inhibited cancer cell growth and induced cell cycle arrest and apoptosis in HCT-116 cells, which was abolished by knockdown of p21. The phosphorylation of Rb and the cleavage of caspase 3 and PARP were suppressed by miR-6734 transfection in HCT-116 cells and these effects were also reversed by p21 knockdown. In addition, miR-6734 transfection caused prolonged induction of p21 gene and modification of histones in p21 promoter, which are typical aspects of a phenomenon referred to as RNA activation (RNAa). Collectively, our results demonstrated that miR-6734 inhibits the growth of colon cancer cells by up-regulating p21 gene expression and subsequent induction of cell cycle arrest and apoptosis, suggesting its role as an important endogenous regulator of cancer cell proliferation and survival. PMID:27509128

  4. Ergosterol purified from medicinal mushroom Amauroderma rude inhibits cancer growth in vitro and in vivo by up-regulating multiple tumor suppressors.

    PubMed

    Li, Xiangmin; Wu, Qingping; Xie, Yizhen; Ding, Yinrun; Du, William W; Sdiri, Mouna; Yang, Burton B

    2015-07-10

    We have previously screened thirteen medicinal mushrooms for their potential anti-cancer activities in eleven different cell lines and found that the extract of Amauroderma rude exerted the highest capacity in inducing cancer cell death. The current study aimed to purify molecules mediating the anti-cancer cell activity. The extract of Amauroderma rude was subject to fractionation, silica gel chromatography,and HPLC. We purified a compound and identified it as ergosterol by EI-MS and NMR,which was expressed at the highest level in Amauroderma rude compared with other medicinal mushrooms tested. We found that ergosterol induced cancer cell death,which was time and concentration dependent. In the in vivo experiment, normal mice were injected with murine cancer cell line B16 that is very aggressive and caused mouse death severely. We found that treatment with ergosterol prolonged mouse survival. We found that ergosterol-mediated suppression of breast cancer cell viability occurred through apoptosis and that ergosterol up-regulated expression of the tumor suppressor Foxo3. In addition, the Foxo3 down-stream signaling molecules Fas, FasL,BimL, and BimS were up-regulated leading to apoptosis in human breast cancer cells MDA-MB-231. Our results suggest that ergosterol is the main anti-cancer ingredient in Amauroderma rude, which activated the apoptotic signal pathway. Ergosterol may serve as a potential lead for cancer therapy. PMID:26098777

  5. A novel action mechanism for MPT0G013, a derivative of arylsulfonamide, inhibits tumor angiogenesis through up-regulation of TIMP3 expression.

    PubMed

    Wang, Chih-Ya; Liou, Jing-Ping; Tsai, An-Chi; Lai, Mei-Jung; Liu, Yi-Min; Lee, Hsueh-Yun; Wang, Jing-Chi; Pan, Shiow-Lin; Teng, Che-Ming

    2014-10-30

    Tissue inhibitors of metalloproteinases 3 (TIMP3) were originally characterized as inhibitors of matrix metalloproteinases (MMPs), acting as potent antiangiogenic proteins. In this study, we demonstrated that the arylsulfonamide derivative MPT0G013 has potent antiangiogenic activities in vitro and in vivo viainducing TIMP3 expression. Treatments with MPT0G013 significantly inhibited endothelial cell functions, such as cell proliferation, migration, and tube formation, as well as induced p21 and cell cycle arrest at the G0/G1 phase. Subsequent microarray analysis showed significant induction of TIMP3 gene expression by MPT0G013, and siRNA-mediated blockage of TIMP3 up-regulation abrogated the antiangiogenic activities of MPT0G013 and prevented inhibition of p-AKT and p-ERK proteins. Importantly, MPT0G013 exhibited antiangiogenic activities in in vivo Matrigel plug assays, inhibited tumor growth and up-regulated TIMP3 and p21 proteins in HCT116 mouse xenograft models. These data suggest potential therapeutic application of MPT0G013 for angiogenesis-related diseases such as cancer.

  6. Fucoidan protects hepatocytes from apoptosis and inhibits invasion of hepatocellular carcinoma by up-regulating p42/44 MAPK-dependent NDRG-1/CAP43.

    PubMed

    Cho, Yuri; Yoon, Jung-Hwan; Yoo, Jeong-Ju; Lee, Minjong; Lee, Dong Hyeon; Cho, Eun Ju; Lee, Jeong-Hoon; Yu, Su Jong; Kim, Yoon Jun; Kim, Chung Yong

    2015-11-01

    Fucoidan is a traditional Chinese medicine suggested to possess anti-tumor effects. In this study the anti-metastatic effects of fucoidan were investigated in vitro in human hepatocellular carcinoma (HCC) cells (Huh-7 and SNU-761) under normoxic and hypoxic conditions and in vivo using a distant liver metastasis model involving injection of MH134 cells into spleen via the portal vein. Its ability to protect hepatocytes against bile acid (BA)-induced apoptosis was investigated in primary hepatocytes. Fucoidan was found to suppress the invasion of HCC cells through up-regulation of p42/44 MAPK-dependent NDRG-1/CAP43 and partly, under normoxic conditions, through up-regulation of p42/44 MAPK-dependent VMP-1 expression. It also significantly decreased liver metastasis in vivo. As regards its hepatoprotective effect, fucoidan decreased BA-induced hepatocyte apoptosis as shown by the attenuation of caspase-8, and -7 cleavages and suppression of the mobilization of caspase-8 and Fas associated death domain (FADD) into the death-inducing signaling complex. In summary, fucoidan displays inhibitory effects on proliferation of HCC cells and protective effects on hepatocytes. The results suggest fucoidan is a potent suppressor of tumor invasion with hepatoprotective effects. PMID:26713269

  7. Up-regulation of anti-apoptotic genes confers resistance to the novel anti-leukaemic compound PEP005 in primary AML cells

    PubMed Central

    Hampson, Peter; Wang, Keqing; Ersvær, Elisabeth; McCormack, Emmet; Schüler, Julia; Fiebig, Heinz-Herbert; Gjertsen, Bjørn Tore; Bruserud, Øystein; Lord, Janet M.

    2014-01-01

    We showed previously that PEP005 induced apoptosis in leukaemic cell lines and blasts from patients with acute myeloid leukaemia (AML). Here we assess the anti-leukeamic effects of PEP005 in vivo and determine the mechanism of resistance of PEP005 non-responsive cells. We used 2 human xenograft mouse models of AML to assess the anti-leukaemic effects of PEP005 in vivo. Expression microarray analysis of primary AML blasts following treatment with PEP005 was used to determine patterns of gene expression that conferred resistance. PEP005 significantly reduced tumour burden in two human leukaemia mouse xenograft models. We also assessed responsiveness of 33 AML samples to PEP005, with 78% of the samples entering apoptosis at 100nM. Resistance to PEP005 was not restricted to a particular AML subtype. Expression microarray analysis of resistant samples following treatment with PEP005 revealed a significant up regulation of the anti-apoptotic genes Bcl-2A1, Mcl-1, and PHLDA1 which was verified using RT-PCR. We conclude that PEP005 shows broad efficacy against AML subtypes and that up regulation of anti-apoptotic genes underlies resistance to this agent and could be used to screen for patients unlikely to benefit from a therapeutic regime involving PEP005. PMID:25594060

  8. Exogenous NAD+ decreases oxidative stress and protects H2O2-treated RPE cells against necrotic death through the up-regulation of autophagy

    PubMed Central

    Zhu, Ying; Zhao, Ke-ke; Tong, Yao; Zhou, Ya-li; Wang, Yi-xiao; Zhao, Pei-quan; Wang, Zhao-yang

    2016-01-01

    Increased oxidative stress, which can lead to the retinal pigment epithelium (RPE) cell death by inducing ATP depletion and DNA repair, is believed to be a prominent pathology in age-related macular degeneration (AMD). In the present study, we showed that and 0.1 mM nicotinamide adenine dinucleotide (NAD+) administration significantly blocked RPE cell death induced by 300 μM H2O2. Further investigation showed that H2O2 resulted in increased intracellular ROS level, activation of PARP-1 and subsequently necrotic death of RPE cells. Exogenous NAD+ administration significantly decreased intracellular and intranuclear ROS levels in H2O2-treated RPE cells. In addition, NAD+ administration to H2O2-treated RPE cells inhibited the activation of PARP-1 and protected the RPE cells against necrotic death. Moreover, exogenous NAD+ administration up-regulated autophagy in the H2O2-treated RPE cells. Inhibition of autophagy by LY294002 blocked the decrease of intracellular and intranuclear ROS level. Besides, inhibition of autophagy by LY294002 abolished the protection of exogenous NAD+ against H2O2-induced cell necrotic death. Taken together, our findings indicate that that exogenous NAD+ administration suppresses H2O2-induced oxidative stress and protects RPE cells against PARP-1 mediated necrotic death through the up-regulation of autophagy. The results suggest that exogenous NAD+ administration might be potential value for the treatment of AMD. PMID:27240523

  9. Effects of Arg-Gly-Asp-modified elastin-like polypeptide on pseudoislet formation via up-regulation of cell adhesion molecules and extracellular matrix proteins.

    PubMed

    Lee, Kyeong-Min; Jung, Gwon-Soo; Park, Jin-Kyu; Choi, Seong-Kyoon; Jeon, Won Bae

    2013-03-01

    Extracellular matrix (ECM) plays an important role in controlling the β-cell morphology, survival and insulin secretary functions. An RGD-modified elastin-like polypeptide (RGD-ELP), TGPG[VGRGD(VGVPG)(6)](20)WPC, has been reported previously as a bioactive matrix. In this study, to investigate whether RGD-ELP affects β-cell growth characteristics and insulin secretion, β-TC6 cells were cultured on the RGD-ELP coatings prepared via thermally induced phase transition. On RGD-ELP, β-TC6 cells clustered into an islet-like architecture with high cell viability. Throughout 7days' culture, the proliferation rate of the cells within a pseudoislet was similar to that of monolayer culture. Under high glucose (25mM), β-TC6 pseudoislets showed up-regulated insulin gene expression and exhibited glucose-stimulated insulin secretion. Importantly, the mRNA and protein abundances of cell adhesion molecules (CAM) E-cadherin and connexin-36 were much higher in pseudoislets than in monolayer cells. The siRNA-mediated inhibition of E-cadherin or connexin-36 expression severely limited pseudoislet formation. In addition, the mRNA levels of collagen types I and IV, fibronectin and laminin were significantly elevated in pseudoislets. The results suggest that RGD-ELP promotes pseudoislet formation via up-regulation of the CAM and ECM components. The functional roles of RGD-ELP are discussed in respect of its molecular composition.

  10. Behavioral stress causes mitochondrial dysfunction via ABAD up-regulation and aggravates plaque pathology in the brain of a mouse model of Alzheimer disease.

    PubMed

    Seo, Ji-Seon; Lee, Kang-Woo; Kim, Tae-Kyung; Baek, In-Sun; Im, Joo-Young; Han, Pyung-Lim

    2011-06-01

    Basic and clinical studies have reported that behavioral stress worsens the pathology of Alzheimer disease (AD), but the underlying mechanism has not been clearly understood. In this study, we determined the mechanism by which behavioral stress affects the pathogenesis of AD using Tg-APPswe/PS1dE9 mice, a murine model of AD. Tg-APPswe/PS1dE9 mice that were restrained for 2h daily for 16 consecutive days (2-h/16-day stress) from 6.5months of age had significantly increased Aβ(1-42) levels and plaque deposition in the brain. The 2-h/16-day stress increased oxidative stress and induced mitochondrial dysfunction in the brain. Treatment with glucocorticoid (corticosterone) and Aβ in SH-SY5Y cells increased the expression of 17β-hydroxysteroid dehydrogenase (ABAD), mitochondrial dysfunction, and levels of ROS, whereas blockade of ABAD expression by siRNA-ABAD in SH-SY5Y cells suppressed glucocorticoid-enhanced mitochondrial dysfunction and ROS accumulation. The 2-h/16-day stress up-regulated ABAD expression in mitochondria in the brain of Tg-APPswe/PS1dE9 mice. Moreover, all visible Aβ plaques were costained with anti-ABAD in the brains of Tg-APPswe/PS1dE9 mice. Together, these results suggest that behavioral stress aggravates plaque pathology and mitochondrial dysfunction via up-regulation of ABAD in the brain of a mouse model of AD.

  11. Dietary fish oil up-regulates cholesterol 7alpha-hydroxylase mRNA in mouse liver leading to an increase in bile acid and cholesterol excretion.

    PubMed

    Bérard, Annie M; Dumon, Marie-France; Darmon, Michel

    2004-02-13

    To investigate the molecular events controlling reverse cholesterol transport, we compared gene expression of normal mouse liver to that of mice fed a long chain (LC) omega-3 fatty acid-enriched diet. Using cDNA microarrays, we assessed expression levels of 1176 genes, and we found that D-site binding protein (DBP) was three-fold increased in mice on a LC omega-3 fatty acid-rich diet compared to controls. DBP is known to increase transcriptional level of cholesterol 7alpha-hydroxylase (C7alpha), the rate-limiting enzyme for bile acid production and cholesterol excretion, and we found that C7alpha mRNA was also up-regulated by LC omega-3 fatty acids. Moreover, liver X receptor-alpha, another transcription factor up-regulating C7alpha, was three- to four-fold increased in liver of treated mice. On the other hand, we demonstrated that bile acid and cholesterol excretion were two-fold increased. These results show that LC omega-3 fatty acids control cholesterol metabolism in mice at a new endpoint.

  12. Fucoidan protects hepatocytes from apoptosis and inhibits invasion of hepatocellular carcinoma by up-regulating p42/44 MAPK-dependent NDRG-1/CAP43.

    PubMed

    Cho, Yuri; Yoon, Jung-Hwan; Yoo, Jeong-Ju; Lee, Minjong; Lee, Dong Hyeon; Cho, Eun Ju; Lee, Jeong-Hoon; Yu, Su Jong; Kim, Yoon Jun; Kim, Chung Yong

    2015-11-01

    Fucoidan is a traditional Chinese medicine suggested to possess anti-tumor effects. In this study the anti-metastatic effects of fucoidan were investigated in vitro in human hepatocellular carcinoma (HCC) cells (Huh-7 and SNU-761) under normoxic and hypoxic conditions and in vivo using a distant liver metastasis model involving injection of MH134 cells into spleen via the portal vein. Its ability to protect hepatocytes against bile acid (BA)-induced apoptosis was investigated in primary hepatocytes. Fucoidan was found to suppress the invasion of HCC cells through up-regulation of p42/44 MAPK-dependent NDRG-1/CAP43 and partly, under normoxic conditions, through up-regulation of p42/44 MAPK-dependent VMP-1 expression. It also significantly decreased liver metastasis in vivo. As regards its hepatoprotective effect, fucoidan decreased BA-induced hepatocyte apoptosis as shown by the attenuation of caspase-8, and -7 cleavages and suppression of the mobilization of caspase-8 and Fas associated death domain (FADD) into the death-inducing signaling complex. In summary, fucoidan displays inhibitory effects on proliferation of HCC cells and protective effects on hepatocytes. The results suggest fucoidan is a potent suppressor of tumor invasion with hepatoprotective effects.

  13. Novel mechanism of harmaline on inducing G2/M cell cycle arrest and apoptosis by up-regulating Fas/FasL in SGC-7901 cells.

    PubMed

    Wang, Yihai; Wang, Chunhua; Jiang, Chenguang; Zeng, Hong; He, Xiangjiu

    2015-12-18

    Harmaline (HAR), a natural occurrence β-carboline alkaloid, was isolated from the seeds of Peganum harmala and exhibited potent antitumor effect. In this study, the anti-gastric tumor effects of HAR were firstly investigated in vitro and in vivo. The results strongly showed that HAR could inhibit tumor cell proliferation and induce G2/M cell cycle arrest accompanied by an increase in apoptotic cell death in SGC-7901 cancer cells. HAR could up-regulate