RNA Binding Proteins Posttranscriptionally Regulate Genes Involved In Oncogenesis

DTIC Science & Technology

2010-06-01

whose steady state mRNA levels may not significantly change, but which are tr anslationally active inside cancer cells. Potentially the...techniques have the potential to better delineate gene s whose steady state mRNA levels may not significantly change, but which are translationally active ...significantly change, but which are tr anslationally active inside cancer cells. Potentially the identification of such genes m ay offer novel therapeutic

Voltage-gated sodium channel expression and action potential generation in differentiated NG108-15 cells.

PubMed

Liu, Jinxu; Tu, Huiyin; Zhang, Dongze; Zheng, Hong; Li, Yu-Long

2012-10-25

The generation of action potential is required for stimulus-evoked neurotransmitter release in most neurons. Although various voltage-gated ion channels are involved in action potential production, the initiation of the action potential is mainly mediated by voltage-gated Na+ channels. In the present study, differentiation-induced changes of mRNA and protein expression of Na+ channels, Na+ currents, and cell membrane excitability were investigated in NG108-15 cells. Whole-cell patch-clamp results showed that differentiation (9 days) didn't change cell membrane excitability, compared to undifferentiated state. But differentiation (21 days) induced the action potential generation in 45.5% of NG108-15 cells (25/55 cells). In 9-day-differentiated cells, Na+ currents were mildly increased, which was also found in 21-day differentiated cells without action potential. In 21-day differentiated cells with action potential, Na+ currents were significantly enhanced. Western blot data showed that the expression of Na+ channels was increased with differentiated-time dependent manner. Single-cell real-time PCR data demonstrated that the expression of Na+ channel mRNA was increased by 21 days of differentiation in NG108-15 cells. More importantly, the mRNA level of Na+ channels in cells with action potential was higher than that in cells without action potential. Differentiation induces expression of voltage-gated Na+ channels and action potential generation in NG108-15 cells. A high level of the Na+ channel density is required for differentiation-triggered action potential generation.

The Hippo pathway: regulators and regulations

PubMed Central

Yu, Fa-Xing; Guan, Kun-Liang

2013-01-01

Control of cell number is crucial in animal development and tissue homeostasis, and its dysregulation may result in tumor formation or organ degeneration. The Hippo pathway in both Drosophila and mammals regulates cell number by modulating cell proliferation, cell death, and cell differentiation. Recently, numerous upstream components involved in the Hippo pathway have been identified, such as cell polarity, mechanotransduction, and G-protein-coupled receptor (GPCR) signaling. Actin cytoskeleton or cellular tension appears to be the master mediator that integrates and transmits upstream signals to the core Hippo signaling cascade. Here, we review regulatory mechanisms of the Hippo pathway and discuss potential implications involved in different physiological and pathological conditions. PMID:23431053

Role of substrate biomechanics in controlling (stem) cell fate: Implications in regenerative medicine.

PubMed

Macri-Pellizzeri, Laura; De-Juan-Pardo, Elena M; Prosper, Felipe; Pelacho, Beatriz

2018-04-01

Tissue-specific stem cells reside in a specialized environment known as niche. The niche plays a central role in the regulation of cell behaviour and, through the concerted action of soluble molecules, supportive somatic cells, and extracellular matrix components, directs stem cells to proliferate, differentiate, or remain quiescent. Great efforts have been done to decompose and separately analyse the contribution of these cues in the in vivo environment. Specifically, the mechanical properties of the extracellular matrix influence many aspects of cell behaviour, including self-renewal and differentiation. Deciphering the role of biomechanics could thereby provide important insights to control the stem cells responses in a more effective way with the aim to promote their therapeutic potential. In this review, we provide a wide overview of the effect that the microenvironment stiffness exerts on the control of cell behaviour with a particular focus on the induction of stem cells differentiation. We also describe the process of mechanotransduction and the molecular effectors involved. Finally, we critically discuss the potential involvement of tissue biomechanics in the design of novel tissue engineering strategies. Copyright © 2017 John Wiley & Sons, Ltd.

Galectin-3 as a Potential Target to Prevent Cancer Metastasis

PubMed Central

Ahmed, Hafiz; AlSadek, Dina M. M.

2015-01-01

Interactions between two cells or between cell and extracellular matrix mediated by protein–carbohydrate interactions play pivotal roles in modulating various biological processes such as growth regulation, immune function, cancer metastasis, and apoptosis. Galectin-3, a member of the β-galactoside-binding lectin family, is involved in fibrosis as well as cancer progression and metastasis, but the detailed mechanisms of its functions remain elusive. This review discusses its structure, carbohydrate-binding properties, and involvement in various aspects of tumorigenesis and some potential carbohydrate ligands that are currently investigated to block galectin-3 activity. PMID:26640395

Cellular and Molecular Effect of MEHP Involving LXRα in Human Fetal Testis and Ovary

PubMed Central

Muczynski, Vincent; Lecureuil, Charlotte; Messiaen, Sébastien; Guerquin, Marie-Justine; N’Tumba-Byn, Thierry; Moison, Delphine; Hodroj, Wassim; Benjelloun, Hinde; Baijer, Jan; Livera, Gabriel; Frydman, René; Benachi, Alexandra; Habert, René; Rouiller-Fabre, Virginie

2012-01-01

Background Phthalates have been shown to have reprotoxic effects in rodents and human during fetal life. Previous studies indicate that some members of the nuclear receptor (NR) superfamilly potentially mediate phthalate effects. This study aimed to assess if expression of these nuclear receptors are modulated in the response to MEHP exposure on the human fetal gonads in vitro. Methodology/Principal Findings Testes and ovaries from 7 to 12 gestational weeks human fetuses were exposed to 10−4M MEHP for 72 h in vitro. Transcriptional level of NRs and of downstream genes was then investigated using TLDA (TaqMan Low Density Array) and qPCR approaches. To determine whether somatic or germ cells of the testis are involved in the response to MEHP exposure, we developed a highly efficient cytometric germ cell sorting approach. In vitro exposure of fetal testes and ovaries to MEHP up-regulated the expression of LXRα, SREBP members and of downstream genes involved in the lipid and cholesterol synthesis in the whole gonad. In sorted testicular cells, this effect is only observable in somatic cells but not in the gonocytes. Moreover, the germ cell loss induced by MEHP exposure, that we previously described, is restricted to the male gonad as oogonia density is not affected in vitro. Conclusions/Significance We evidenced for the first time that phthalate increases the levels of mRNA for LXRα, and SREBP members potentially deregulating lipids/cholesterol synthesis in human fetal gonads. Interestingly, this novel effect is observable in both male and female whereas the germ cell apoptosis is restricted to the male gonad. Furthermore, we presented here a novel and potentially very useful flow cytometric cell sorting method to analyse molecular changes in germ cells versus somatic cells. PMID:23118965

Cellular and molecular effect of MEHP Involving LXRα in human fetal testis and ovary.

PubMed

Muczynski, Vincent; Lecureuil, Charlotte; Messiaen, Sébastien; Guerquin, Marie-Justine; N'tumba-Byn, Thierry; Moison, Delphine; Hodroj, Wassim; Benjelloun, Hinde; Baijer, Jan; Livera, Gabriel; Frydman, René; Benachi, Alexandra; Habert, René; Rouiller-Fabre, Virginie

2012-01-01

Phthalates have been shown to have reprotoxic effects in rodents and human during fetal life. Previous studies indicate that some members of the nuclear receptor (NR) superfamilly potentially mediate phthalate effects. This study aimed to assess if expression of these nuclear receptors are modulated in the response to MEHP exposure on the human fetal gonads in vitro. Testes and ovaries from 7 to 12 gestational weeks human fetuses were exposed to 10(-4)M MEHP for 72 h in vitro. Transcriptional level of NRs and of downstream genes was then investigated using TLDA (TaqMan Low Density Array) and qPCR approaches. To determine whether somatic or germ cells of the testis are involved in the response to MEHP exposure, we developed a highly efficient cytometric germ cell sorting approach. In vitro exposure of fetal testes and ovaries to MEHP up-regulated the expression of LXRα, SREBP members and of downstream genes involved in the lipid and cholesterol synthesis in the whole gonad. In sorted testicular cells, this effect is only observable in somatic cells but not in the gonocytes. Moreover, the germ cell loss induced by MEHP exposure, that we previously described, is restricted to the male gonad as oogonia density is not affected in vitro. We evidenced for the first time that phthalate increases the levels of mRNA for LXRα, and SREBP members potentially deregulating lipids/cholesterol synthesis in human fetal gonads. Interestingly, this novel effect is observable in both male and female whereas the germ cell apoptosis is restricted to the male gonad. Furthermore, we presented here a novel and potentially very useful flow cytometric cell sorting method to analyse molecular changes in germ cells versus somatic cells.

Cell wall and enzyme changes during the graviresponse of the leaf-sheath pulvinus of oat (Avena sativa)

NASA Technical Reports Server (NTRS)

Gibeaut, David M.; Karuppiah, Nadarajah; Chang, S.-R.; Brock, Thomas G.; Vadlamudi, Babu; Kim, Donghern; Ghosheh, Najati S.; Rayle, David L.; Carpita, Nicholas C.; Kaufman, Peter B.

1990-01-01

The graviresponse of the leaf-sheath pulvinus of oat (Avena sativa) involves an asymmetric growth response and asymmetric processes involving degradation of starch and cell wall synthesis. Cellular and biochemical events were studied by investigation of the activities of related enzymes and changes in cell walls and their constituents. It is suggested that an osmotic potential gradient acts as the driving factor for growth, while wall extensibility is a limiting factor in pulvinus growth.

Th9 cells: differentiation and disease

PubMed Central

Kaplan, Mark H.

2014-01-01

Summary CD4+ T-helper cells regulate immunity and inflammation through the acquisition of potential to secrete specific cytokines. The acquisition of cytokine-secreting potential, in a process termed T-helper cell differentiation, is a response to multiple environmental signals including the cytokine milieu. The most recently defined subset of T-helper cells are termed Th9 and are identified by the potent production of interleukin-9 (IL-9). Given the pleiotropic functions of IL-9, Th9 cells might be involved in pathogen immunity and immune-mediated disease. In this review, I focus on recent developments in understanding the signals that promote Th9 differentiation, the transcription factors that regulate IL-9 expression, and finally the potential roles for Th9 cells in immunity in vivo. PMID:23405898

Cell Cycle Regulation of Stem Cells by MicroRNAs.

PubMed

Mens, Michelle M J; Ghanbari, Mohsen

2018-06-01

MicroRNAs (miRNAs) are a class of small non-coding RNA molecules involved in the regulation of gene expression. They are involved in the fine-tuning of fundamental biological processes such as proliferation, differentiation, survival and apoptosis in many cell types. Emerging evidence suggests that miRNAs regulate critical pathways involved in stem cell function. Several miRNAs have been suggested to target transcripts that directly or indirectly coordinate the cell cycle progression of stem cells. Moreover, previous studies have shown that altered expression levels of miRNAs can contribute to pathological conditions, such as cancer, due to the loss of cell cycle regulation. However, the precise mechanism underlying miRNA-mediated regulation of cell cycle in stem cells is still incompletely understood. In this review, we discuss current knowledge of miRNAs regulatory role in cell cycle progression of stem cells. We describe how specific miRNAs may control cell cycle associated molecules and checkpoints in embryonic, somatic and cancer stem cells. We further outline how these miRNAs could be regulated to influence cell cycle progression in stem cells as a potential clinical application.

Generators of the brainstem auditory evoked potential in cat. II. Correlating lesion sites with waveform changes.

PubMed

Melcher, J R; Guinan, J J; Knudson, I M; Kiang, N Y

1996-04-01

Brainstem regions involved in generating the brainstem auditory evoked potential (BAEP) were identified by examining the effects of lesions on the click-evoked BAEP in cats. An excitotoxin, kainic acid, was injected into various parts of the cochlear nucleus (CN) or into the superior olivary complex (SOC). The locations of the resulting lesions were correlated with the changes produced in the various extrema of the BAEP waveforms. The results indicate that: (1) the earliest BAEP extrema (P1, N1 (recorded between vertex and the earbar ipsilateral to the stimulus) and P1a, P1b, (vertex to contralateral earbar)) are generated by cells with somata peripheral to the CN; (2) P2 is primarily generated by posterior anteroventral CN (AVCNp) and anterior posteroventral CN (PVCNa) cells; (3) SOC, anterior anteroventral CN (AVCNa), AVCNp, and PVCNa cells are involved in generating P3; (4) AVCNa cells are the main CN cells involved in P4, N4, and P5 generation; (5) both ipsilateral and contralateral SOC cells have a role in generating monaurally evoked P4 and P5; and (6) P5 is generated by cells with characteristic frequencies below 10 kHz. From (2) and (4), it is clear that P2 and P4-P5 are generated by cells in distinct, parallel pathways.

The neuroprotection of cannabidiol against MPP⁺-induced toxicity in PC12 cells involves trkA receptors, upregulation of axonal and synaptic proteins, neuritogenesis, and might be relevant to Parkinson's disease.

PubMed

Santos, Neife Aparecida Guinaim; Martins, Nádia Maria; Sisti, Flávia Malvestio; Fernandes, Laís Silva; Ferreira, Rafaela Scalco; Queiroz, Regina Helena Costa; Santos, Antônio Cardozo

2015-12-25

Cannabidiol (CBD) is a non-psychoactive constituent of Cannabis sativa with potential to treat neurodegenerative diseases. Its neuroprotection has been mainly associated with anti-inflammatory and antioxidant events; however, other mechanisms might be involved. We investigated the involvement of neuritogenesis, NGF receptors (trkA), NGF, and neuronal proteins in the mechanism of neuroprotection of CBD against MPP(+) toxicity in PC12 cells. CBD increased cell viability, differentiation, and the expression of axonal (GAP-43) and synaptic (synaptophysin and synapsin I) proteins. Its neuritogenic effect was not dependent or additive to NGF, but it was inhibited by K252a (trkA inhibitor). CBD did not increase the expression of NGF, but protected against its decrease induced by MPP(+), probably by an indirect mechanism. We also evaluated the neuritogenesis in SH-SY5Y cells, which do not express trkA receptors. CBD did not induce neuritogenesis in this cellular model, which supports the involvement of trkA receptors. This is the first study to report the involvement of neuronal proteins and trkA in the neuroprotection of CBD. Our findings suggest that CBD has a neurorestorative potential independent of NGF that might contribute to its neuroprotection against MPP(+), a neurotoxin relevant to Parkinson's disease. Copyright © 2015 Elsevier B.V. All rights reserved.

Interaction of Mycobacterium leprae with Human Airway Epithelial Cells: Adherence, Entry, Survival, and Identification of Potential Adhesins by Surface Proteome Analysis

PubMed Central

Silva, Carlos A. M.; Danelishvili, Lia; McNamara, Michael; Berredo-Pinho, Márcia; Bildfell, Robert; Biet, Franck; Rodrigues, Luciana S.; Oliveira, Albanita V.

2013-01-01

This study examined the in vitro interaction between Mycobacterium leprae, the causative agent of leprosy, and human alveolar and nasal epithelial cells, demonstrating that M. leprae can enter both cell types and that both are capable of sustaining bacterial survival. Moreover, delivery of M. leprae to the nasal septum of mice resulted in macrophage and epithelial cell infection in the lung tissue, sustaining the idea that the airways constitute an important M. leprae entry route into the human body. Since critical aspects in understanding the mechanisms of infection are the identification and characterization of the adhesins involved in pathogen-host cell interaction, the nude mouse-derived M. leprae cell surface-exposed proteome was studied to uncover potentially relevant adhesin candidates. A total of 279 cell surface-exposed proteins were identified based on selective biotinylation, streptavidin-affinity purification, and shotgun mass spectrometry; 11 of those proteins have been previously described as potential adhesins. In vitro assays with the recombinant forms of the histone-like protein (Hlp) and the heparin-binding hemagglutinin (HBHA), considered to be major mycobacterial adhesins, confirmed their capacity to promote bacterial attachment to epithelial cells. Taking our data together, they suggest that the airway epithelium may act as a reservoir and/or portal of entry for M. leprae in humans. Moreover, our report sheds light on the potentially critical adhesins involved in M. leprae-epithelial cell interaction that may be useful in designing more effective tools for leprosy control. PMID:23670556

Expression and role of DJ-1 in leukemia

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

Liu Hang; Wang Min; Li Min

2008-10-24

DJ-1 is a multifunctional protein that has been implicated in pathogenesis of some solid tumors. In this study, we found that DJ-1 was overexpressed in acute leukemia (AL) patient samples and leukemia cell lines, which gave the first clue that DJ-1 overexpression might be involved in leukemogenesis and/or disease progression of AL. Inactivation of DJ-1 by RNA-mediated interference (RNAi) in leukemia cell lines K562 and HL60 resulted in inhibition of the proliferation potential and enhancement of the sensitivity of leukemia cells to chemotherapeutic drug etoposide. Further investigation of DJ-1 activity revealed that phosphatase and tensin homolog (PTEN), as well asmore » some proliferation and apoptosis-related genes, was regulated by DJ-1. Thus, DJ-1 might be involved in leukemogesis through regulating cell growth, proliferation, and apoptosis. It could be a potential therapeutic target for leukemia.« less

ER-mediated stress induces mitochondrial-dependent caspases activation in NT2 neuron-like cells.

PubMed

Arduino, Daniela M; Esteves, A Raquel; Domingues, A Filipa; Pereira, Claudia M F; Cardoso, Sandra M; Oliveira, Catarina R

2009-11-30

Recent studies have revealed that endoplasmic reticulum (ER) disturbance is involved in the pathophysiology of neurodegenerative disorders, contributing to the activation of the ER stress-mediated apoptotic pathway. Therefore, we investigated here the molecular mechanisms underlying the ER-mitochondria axis, focusing on calcium as a potential mediator of cell death signals. Using NT2 cells treated with brefeldin A or tunicamycin, we observed that ER stress induces changes in the mitochondrial function, impairing mitochondrial membrane potential and distressing mitochondrial respiratory chain complex Moreover, stress stimuli at ER level evoked calcium fluxes between ER and mitochondria. Under these conditions, ER stress activated the unfolded protein response by an overexpression of GRP78, and also caspase-4 and-2, both involved upstream of caspase-9. Our findings show that ER and mitochondria interconnection plays a prominent role in the induction of neuronal cell death under particular stress circumstances.

Transcriptional profiling of CD31(+) cells isolated from murine embryonic stem cells.

PubMed

Mariappan, Devi; Winkler, Johannes; Chen, Shuhua; Schulz, Herbert; Hescheler, Jürgen; Sachinidis, Agapios

2009-02-01

Identification of genes involved in endothelial differentiation is of great interest for the understanding of the cellular and molecular mechanisms involved in the development of new blood vessels. Mouse embryonic stem (mES) cells serve as a potential source of endothelial cells for transcriptomic analysis. We isolated endothelial cells from 8-days old embryoid bodies by immuno-magnetic separation using platelet endothelial cell adhesion molecule-1 (also known as CD31) expressed on both early and mature endothelial cells. CD31(+) cells exhibit endothelial-like behavior by being able to incorporate DiI-labeled acetylated low-density lipoprotein as well as form tubular structures on matrigel. Quantitative and semi-quantitative PCR analysis further demonstrated the increased expression of endothelial transcripts. To ascertain the specific transcriptomic identity of the CD31(+) cells, large-scale microarray analysis was carried out. Comparative bioinformatic analysis reveals an enrichment of the gene ontology categories angiogenesis, blood vessel morphogenesis, vasculogenesis and blood coagulation in the CD31(+) cell population. Based on the transcriptomic signatures of the CD31(+) cells, we conclude that this ES cell-derived population contains endothelial-like cells expressing a mesodermal marker BMP2 and possess an angiogenic potential. The transcriptomic characterization of CD31(+) cells enables an in vitro functional genomic model to identify genes required for angiogenesis.

Skin appendage-derived stem cells: cell biology and potential for wound repair.

PubMed

Xie, Jiangfan; Yao, Bin; Han, Yutong; Huang, Sha; Fu, Xiaobing

2016-01-01

Stem cells residing in the epidermis and skin appendages are imperative for skin homeostasis and regeneration. These stem cells also participate in the repair of the epidermis after injuries, inducing restoration of tissue integrity and function of damaged tissue. Unlike epidermis-derived stem cells, comprehensive knowledge about skin appendage-derived stem cells remains limited. In this review, we summarize the current knowledge of skin appendage-derived stem cells, including their fundamental characteristics, their preferentially expressed biomarkers, and their potential contribution involved in wound repair. Finally, we will also discuss current strategies, future applications, and limitations of these stem cells, attempting to provide some perspectives on optimizing the available therapy in cutaneous repair and regeneration.

New high-efficiency silicon solar cells

NASA Technical Reports Server (NTRS)

Daud, T.; Crotty, G. T.

1985-01-01

A design for silicon solar cells was investigated as an approach to increasing the cell open-circuit voltage and efficiency for flat-plate terrestrial photovoltaic applications. This deviates from past designs, where either the entire front surface of the cell is covered by a planar junction or the surface is textured before junction formation, which results in an even greater (up to 70%) junction area. The heavily doped front region and the junction space charge region are potential areas of high recombination for generated and injected minority carriers. The design presented reduces junction area by spreading equidiameter dot junctions across the surface of the cell, spaced about a diffusion length or less from each other. Various dot diameters and spacings allowed variations in total junction area. A simplified analysis was done to obtain a first-order design optimization. Efficiencies of up to 19% can be obtained. Cell fabrication involved extra masking steps for selective junction diffusion, and made surface passivation a key element in obtaining good collection. It also involved photolithography, with line widths down to microns. A method is demonstrated for achieving potentially high open-circuit voltages and solar-cell efficiencies.

Stem cell regenerative potential for plastic and reconstructive surgery.

PubMed

Boháč, Martin; Csöbönyeiová, Mária; Kupcová, Ida; Zamborský, Radoslav; Fedeleš, Jozef; Koller, Ján

2016-12-01

Stem cells represent heterogeneous population of undifferentiated cells with unique characteristics of long term self renewal and plasticity. Moreover, they are capable of active migration to diseased tissues, secretion of different bioactive molecules, and they have immunosuppressive potential as well. They occur in all tissues through life and are involved in process of embryogenesis and regeneration. During last decades stem cells attracted significant attention in each field of medicine, including plastic and reconstructive surgery. The main goal of the present review article is to present and discuss the potential of stem cells and to provide information about their safe utilization in chronic wounds and fistulae healing, scar management, breast reconstruction, as well as in bone, tendon and peripheral nerve regeneration.

O2⋅- and H2O2-Mediated Disruption of Fe Metabolism Causes the Differential Susceptibility of NSCLC and GBM Cancer Cells to Pharmacological Ascorbate.

PubMed

Schoenfeld, Joshua D; Sibenaller, Zita A; Mapuskar, Kranti A; Wagner, Brett A; Cramer-Morales, Kimberly L; Furqan, Muhammad; Sandhu, Sonia; Carlisle, Thomas L; Smith, Mark C; Abu Hejleh, Taher; Berg, Daniel J; Zhang, Jun; Keech, John; Parekh, Kalpaj R; Bhatia, Sudershan; Monga, Varun; Bodeker, Kellie L; Ahmann, Logan; Vollstedt, Sandy; Brown, Heather; Shanahan Kauffman, Erin P; Schall, Mary E; Hohl, Ray J; Clamon, Gerald H; Greenlee, Jeremy D; Howard, Matthew A; Schultz, Michael K; Smith, Brian J; Riley, Dennis P; Domann, Frederick E; Cullen, Joseph J; Buettner, Garry R; Buatti, John M; Spitz, Douglas R; Allen, Bryan G

2017-04-10

Pharmacological ascorbate has been proposed as a potential anti-cancer agent when combined with radiation and chemotherapy. The anti-cancer effects of ascorbate are hypothesized to involve the autoxidation of ascorbate leading to increased steady-state levels of H 2 O 2 ; however, the mechanism(s) for cancer cell-selective toxicity remain unknown. The current study shows that alterations in cancer cell mitochondrial oxidative metabolism resulting in increased levels of O 2 ⋅- and H 2 O 2 are capable of disrupting intracellular iron metabolism, thereby selectively sensitizing non-small-cell lung cancer (NSCLC) and glioblastoma (GBM) cells to ascorbate through pro-oxidant chemistry involving redox-active labile iron and H 2 O 2 . In addition, preclinical studies and clinical trials demonstrate the feasibility, selective toxicity, tolerability, and potential efficacy of pharmacological ascorbate in GBM and NSCLC therapy. Copyright © 2017 Elsevier Inc. All rights reserved.

Genetic analysis of indefinite division in human cells: Evidence for a cell senescence-related gene(s) on human chromosome 4

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

Yi Ning; Ledbetter, D.H.; Smith, J.R.

1991-07-01

Earlier studies had demonstrated that fusion of normal with immortal human cells yielded hybrids having limited division potential. This indicated that the phenotype of limited proliferation (cellular senescence) is dominant and that immortal cells result from recessive changes in normal growth-regulatory genes. In additional studies, the authors exploited the fact that the immortal phenotype is recessive and, by fusing various immortal human cell lines with each other, identified four complementation groups for indefinite division. Assignment of cell lines to specific groups allowed us to take a focused approach to identify the chromosomes and genes involved in growth regulation that havemore » been modified in immortal cells. They report here that introduction of a normal human chromosome 4 into three immortal cell lines (HeLa, J82, T98G) assigned to complementation group B resulted in loss of proliferation and reversal of the immortal phenotype. No effect on the proliferation potential of cell lines representative of the other complementation groups was observed. This result suggests that a gene(s) involved in cellular senescence and normal growth regulation resides on chromosome 4.« less

Lysophosphatidylcholine-induced cytotoxicity in osteoblast-like MG-63 cells: involvement of transient receptor potential vanilloid 2 (TRPV2) channels.

PubMed

Fallah, Abdallah; Pierre, Rachel; Abed, Elie; Moreau, Robert

2013-01-01

Epidemiological studies indicate that patients suffering from atherosclerosis are predisposed to develop osteoporosis. Accordingly, atherogenic determinants such as oxidized low density lipoprotein (OxLDL) particles have been shown to alter bone cell functions. In this work, we investigated the cytotoxicity of lysophosphatidylcholine (lysoPC), a major phospholipid component generated upon LDL oxidation, on bone-forming MG-63 osteoblast-like cells. Cell viability was reduced by lysoPC in a concentration-dependent manner with a LC50 of 18.7±0.7 μM. LysoPC-induced cell death was attributed to induction of both apoptosis and necrosis. Since impairment of intracellular calcium homeostasis is often involved in mechanism of cell death, we determined the involvement of calcium in lysoPC-induced cytotoxicity. LysoPC promoted a rapid and transient increase in intracellular calcium attributed to mobilization from calcium stores, followed by a sustained influx. Intracellular calcium mobilization was associated to phospholipase C (PLC)-dependent mobilization of calcium from the endoplasmic reticulum since inhibition of PLC or calcium depletion of reticulum endoplasmic with thapsigargin prevented the calcium mobilization. The calcium influx induced by lysoPC was abolished by inhibition of transient receptor potential vanilloid (TRPV) channels with ruthenium red whereas gadolinium, which inhibits canonical TRP (TRPC) channels, was without effect. Accordingly, expression of TRPV2 and TRPV4 were shown in MG-63 cells. The addition of TRPV2 inhibitor Tranilast in the incubation medium prevent the calcium influx triggered by lysoPC and reduced lysoPC-induced cytotoxicity whereas TRPV4 inhibitor RN 1734 was without effect, which confirms the involvement of TRPV2 activation in lysoPC-induced cell death.

Molecular Action of a Potential Tumor Suppression in Mammary Carcinogenesis

DTIC Science & Technology

2006-05-01

translocation in MDA-MB231 cells, as shown in Fig. 5D , indicating that Tid1 inhibits FVII -induced IL-8 production and cell migration by blocking NF-nB...tissue factor - FVIIa pathway modulates the migratory potential of cancer cells through IL-8 production (7). As Tid1 blocks the IL-8 production of...Introduction: ErbB family of growth factor receptors (ErbB1-4) are critically involved in the derivation of certain mammary cancers [1-3]. Among them

Inorganic Electrolytes

DTIC Science & Technology

1977-10-01

relatively low pressure increase (N15 lb/in. g), but could result in failure of the case in a prismatic cell configuration. Further heating to near the...Potential of a Li/S0C1 2 F Ceail vs. Temperature 77 17 Controlled Potential Electrolysis 80 18 SO2 in SOCl 2 , Infrared Spectrum 83 19 Beer’s Law Plot...variables involved. If the resistance of the contact area is near the design load range of the cell, the cell will discharge harmlessly (except at the end of

Intracellular recording of action potentials by nanopillar electroporation.

PubMed

Xie, Chong; Lin, Ziliang; Hanson, Lindsey; Cui, Yi; Cui, Bianxiao

2012-02-12

Action potentials have a central role in the nervous system and in many cellular processes, notably those involving ion channels. The accurate measurement of action potentials requires efficient coupling between the cell membrane and the measuring electrodes. Intracellular recording methods such as patch clamping involve measuring the voltage or current across the cell membrane by accessing the cell interior with an electrode, allowing both the amplitude and shape of the action potentials to be recorded faithfully with high signal-to-noise ratios. However, the invasive nature of intracellular methods usually limits the recording time to a few hours, and their complexity makes it difficult to simultaneously record more than a few cells. Extracellular recording methods, such as multielectrode arrays and multitransistor arrays, are non-invasive and allow long-term and multiplexed measurements. However, extracellular recording sacrifices the one-to-one correspondence between the cells and electrodes, and also suffers from significantly reduced signal strength and quality. Extracellular techniques are not, therefore, able to record action potentials with the accuracy needed to explore the properties of ion channels. As a result, the pharmacological screening of ion-channel drugs is usually performed by low-throughput intracellular recording methods. The use of nanowire transistors, nanotube-coupled transistors and micro gold-spine and related electrodes can significantly improve the signal strength of recorded action potentials. Here, we show that vertical nanopillar electrodes can record both the extracellular and intracellular action potentials of cultured cardiomyocytes over a long period of time with excellent signal strength and quality. Moreover, it is possible to repeatedly switch between extracellular and intracellular recording by nanoscale electroporation and resealing processes. Furthermore, vertical nanopillar electrodes can detect subtle changes in action potentials induced by drugs that target ion channels.

Intracellular recording of action potentials by nanopillar electroporation

NASA Astrophysics Data System (ADS)

Xie, Chong; Lin, Ziliang; Hanson, Lindsey; Cui, Yi; Cui, Bianxiao

2012-03-01

Action potentials have a central role in the nervous system and in many cellular processes, notably those involving ion channels. The accurate measurement of action potentials requires efficient coupling between the cell membrane and the measuring electrodes. Intracellular recording methods such as patch clamping involve measuring the voltage or current across the cell membrane by accessing the cell interior with an electrode, allowing both the amplitude and shape of the action potentials to be recorded faithfully with high signal-to-noise ratios. However, the invasive nature of intracellular methods usually limits the recording time to a few hours, and their complexity makes it difficult to simultaneously record more than a few cells. Extracellular recording methods, such as multielectrode arrays and multitransistor arrays, are non-invasive and allow long-term and multiplexed measurements. However, extracellular recording sacrifices the one-to-one correspondence between the cells and electrodes, and also suffers from significantly reduced signal strength and quality. Extracellular techniques are not, therefore, able to record action potentials with the accuracy needed to explore the properties of ion channels. As a result, the pharmacological screening of ion-channel drugs is usually performed by low-throughput intracellular recording methods. The use of nanowire transistors, nanotube-coupled transistors and micro gold-spine and related electrodes can significantly improve the signal strength of recorded action potentials. Here, we show that vertical nanopillar electrodes can record both the extracellular and intracellular action potentials of cultured cardiomyocytes over a long period of time with excellent signal strength and quality. Moreover, it is possible to repeatedly switch between extracellular and intracellular recording by nanoscale electroporation and resealing processes. Furthermore, vertical nanopillar electrodes can detect subtle changes in action potentials induced by drugs that target ion channels.

A Survey of Italian Physicians' Opinion about Stem Cells Research: What Doctors Prefer and What the Law Requires

PubMed Central

Frati, Paola; Pacchiarotti, Arianna; D'Errico, Stefano

2014-01-01

To evaluate the Italian physicians' knowledge/information level about the therapeutic potential of stem cells, the research choice between embryonic and cordonal stem cells, and the preference between autologous and heterologous storage of cordonal stem cells, we performed a national survey. The questionnaire—distributed to 3361 physicians—involved physicians of different religious orientations and of different medical specialities. Most of the physicians involved (67%) were Catholics, and the majority were gynaecologists and paediatricians (43%) who are mainly in charge to inform future mothers about the possibility of cordonal stem cells conservation. The majority of the physicians interviewed do not have specific knowledge about stem cells (59%), most of them having only generic information (92%). The largest part of physicians prefer to use umbilical cord blood cells rather than embryonic stem cells. Nevertheless, a large percentage of physicians were in favour of embryo research, especially when embryos are supernumerary (44% versus 34%). Eighty-seven % of the physicians interviewed proved to have a general knowledge about stem cells and believe in their therapeutic potential. They prefer research on cordonal stem cells rather than on embryo stem cells. Although they are in favour of heterologous stem cells donation, they still prefer cryopreservation for personal use. PMID:24877099

Posttransplantation lymphoproliferative disease involving the pituitary gland.

PubMed

Meriden, Zina; Bullock, Grant C; Bagg, Adam; Bonatti, Hugo; Cousar, John B; Lopes, M Beatriz; Robbins, Mark K; Cathro, Helen P

2010-11-01

Posttransplantation lymphoproliferative disorders (PTLD) are heterogeneous lesions with variable morphology, immunophenotype, and molecular characteristics. Multiple distinct primary lesions can occur in PTLD, rarely with both B-cell and T-cell characteristics. Lesions can involve both grafted organs and other sites; however, PTLD involving the pituitary gland has not been previously reported. We describe a patient who developed Epstein-Barr virus-negative PTLD 13 years posttransplantation involving the terminal ileum and pituitary, which was simultaneously involved by a pituitary adenoma. Immunohistochemistry of the pituitary lesion showed expression of CD79a, CD3, and CD7 with clonal rearrangements of both T-cell receptor gamma chain (TRG@) and immunoglobulin heavy chain (IGH@) genes. The terminal ileal lesion was immunophenotypically and molecularly distinct. This is the first report of pituitary PTLD and illustrates the potentially complex nature of PTLD. Copyright © 2010 Elsevier Inc. All rights reserved.

Giant cell phlebitis: a potentially lethal clinical entity.

PubMed

Kunieda, Takeshige; Murayama, Masanori; Ikeda, Tsuneko; Yamakita, Noriyoshi

2012-08-01

An 83-year-old woman presented to us with a 4-week history of general malaise, subjective fever and lower abdominal pain. Despite the intravenous infusion of antibiotics, her blood results and physical condition worsened, resulting in her sudden death. Autopsy study revealed that the medium-sized veins of the mesentery were infiltrated by eosinophil granulocytes, lymphocytes, macrophages and multinucleated giant cells; however, the arteries were not involved. Microscopically, venous giant cell infiltration was observed in the gastrointestinal tract, bladder, retroperitoneal tissues and myocardium. The final diagnosis was giant cell phlebitis, a rare disease of unknown aetiology. This case demonstrates for the first time that giant cell phlebitis involving extra-abdominal organs, including hearts, can cause serious morbidity.

Identified Cellular Correlates of Neocortical Ripple and High-Gamma Oscillations during Spindles of Natural Sleep.

PubMed

Averkin, Robert G; Szemenyei, Viktor; Bordé, Sándor; Tamás, Gábor

2016-11-23

Ultra-high-frequency network events in the hippocampus are instrumental in a dialogue with the neocortex during memory formation, but the existence of transient ∼200 Hz network events in the neocortex is not clear. Our recordings from neocortical layer II/III of freely behaving rats revealed field potential events at ripple and high-gamma frequencies repeatedly occurring at troughs of spindle oscillations during sleep. Juxtacellular recordings identified subpopulations of fast-spiking, parvalbumin-containing basket cells with epochs of firing at ripple (∼200 Hz) and high-gamma (∼120 Hz) frequencies detected during spindles and centered with millisecond precision at the trough of spindle waves in phase with field potential events but phase shifted relative to pyramidal cell firing. The results suggest that basket cell subpopulations are involved in spindle-nested, high-frequency network events that hypothetically provide repeatedly occurring neocortical temporal reference states potentially involved in mnemonic processes. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Process for control of cell division

NASA Technical Reports Server (NTRS)

Cone, C. D., Jr. (Inventor)

1977-01-01

A method of controlling mitosis of biological cells was developed, which involved inducing a change in the intracellular ionic hierarchy accompanying the cellular electrical transmembrane potential difference (Esubm) of the cells. The ionic hierarchy may be varied by imposing changes on the relative concentrations of Na(+), K(+) and Cl(-), or by directly imposing changes in the physical Esubm level across the cell surface.

Mediators involved in the immunomodulatory effects of apoptotic cells.

PubMed

Saas, Philippe; Bonnefoy, Francis; Kury-Paulin, Stephanie; Kleinclauss, François; Perruche, Sylvain

2007-07-15

Immunomodulatory properties are attributed to apoptotic cells. These properties have been used to modulate allogeneic immune responses in experimental transplantation settings. In independent studies, apoptotic cell infusion has been shown to favor hematopoietic cell engraftment, to increase heart graft survival, and to delay the lethal onset of graft-versus-host disease (GVHD). The goal of this review was to discuss how apoptotic cell infusion interferes with graft rejection or host rejection (i.e., GVHD) and to focus on the potential mediators or "perpetuators" involved in apoptotic cell-induced immunomodulation. Particular emphasis on apoptotic cell phagocytosis, transforming growth factor (TGF)-beta secretion, and regulatory T cell induction was performed. Stimulating "naturally" immunosuppressive molecules (i.e., TGF-beta) or immunomodulatory cells ("alternatively-activated" macrophages, certain dendritic cell subsets, or regulatory T cells) in a physiological manner by using apoptotic cell infusion can be a promising way to induce tolerance.

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

Peres, Elodie A.; Valable, Samuel; Guillamo, Jean-Sebastien

Hypoxia has been shown to be one of the major events involved in EPO expression. Accordingly, EPO might be expressed by cerebral neoplastic cells, especially in glioblastoma, known to be highly hypoxic tumours. The expression of EPOR has been described in glioma cells. However, data from the literature remain descriptive and controversial. On the basis of an endogenous source of EPO in the brain, we have focused on a potential role of EPOR in brain tumour growth. In the present study, with complementary approaches to target EPO/EPOR signalling, we demonstrate the presence of a functional EPO/EPOR system on glioma cellsmore » leading to the activation of the ERK pathway. This EPO/EPOR system is involved in glioma cell proliferation in vitro. In vivo, we show that the down-regulation of EPOR expression on glioma cells reduces tumour growth and enhances animal survival. Our results support the hypothesis that EPOR signalling in tumour cells is involved in the control of glioma growth.« less

Earmuff restricts progenitor cell potential by attenuating the competence to respond to self-renewal factors.

PubMed

Janssens, Derek H; Komori, Hideyuki; Grbac, Daniel; Chen, Keng; Koe, Chwee Tat; Wang, Hongyan; Lee, Cheng-Yu

2014-03-01

Despite expressing stem cell self-renewal factors, intermediate progenitor cells possess restricted developmental potential, which allows them to give rise exclusively to differentiated progeny rather than stem cell progeny. Failure to restrict the developmental potential can allow intermediate progenitor cells to revert into aberrant stem cells that might contribute to tumorigenesis. Insight into stable restriction of the developmental potential in intermediate progenitor cells could improve our understanding of the development and growth of tumors, but the mechanisms involved remain largely unknown. Intermediate neural progenitors (INPs), generated by type II neural stem cells (neuroblasts) in fly larval brains, provide an in vivo model for investigating the mechanisms that stably restrict the developmental potential of intermediate progenitor cells. Here, we report that the transcriptional repressor protein Earmuff (Erm) functions temporally after Brain tumor (Brat) and Numb to restrict the developmental potential of uncommitted (immature) INPs. Consistently, endogenous Erm is detected in immature INPs but undetectable in INPs. Erm-dependent restriction of the developmental potential in immature INPs leads to attenuated competence to respond to all known neuroblast self-renewal factors in INPs. We also identified that the BAP chromatin-remodeling complex probably functions cooperatively with Erm to restrict the developmental potential of immature INPs. Together, these data led us to conclude that the Erm-BAP-dependent mechanism stably restricts the developmental potential of immature INPs by attenuating their genomic responses to stem cell self-renewal factors. We propose that restriction of developmental potential by the Erm-BAP-dependent mechanism functionally distinguishes intermediate progenitor cells from stem cells, ensuring the generation of differentiated cells and preventing the formation of progenitor cell-derived tumor-initiating stem cells.

ER Stress-Mediated Signaling: Action Potential and Ca(2+) as Key Players.

PubMed

Bahar, Entaz; Kim, Hyongsuk; Yoon, Hyonok

2016-09-15

The proper functioning of the endoplasmic reticulum (ER) is crucial for multiple cellular activities and survival. Disturbances in the normal ER functions lead to the accumulation and aggregation of unfolded proteins, which initiates an adaptive response, the unfolded protein response (UPR), in order to regain normal ER functions. Failure to activate the adaptive response initiates the process of programmed cell death or apoptosis. Apoptosis plays an important role in cell elimination, which is essential for embryogenesis, development, and tissue homeostasis. Impaired apoptosis can lead to the development of various pathological conditions, such as neurodegenerative and autoimmune diseases, cancer, or acquired immune deficiency syndrome (AIDS). Calcium (Ca(2+)) is one of the key regulators of cell survival and it can induce ER stress-mediated apoptosis in response to various conditions. Ca(2+) regulates cell death both at the early and late stages of apoptosis. Severe Ca(2+) dysregulation can promote cell death through apoptosis. Action potential, an electrical signal transmitted along the neurons and muscle fibers, is important for conveying information to, from, and within the brain. Upon the initiation of the action potential, increased levels of cytosolic Ca(2+) (depolarization) lead to the activation of the ER stress response involved in the initiation of apoptosis. In this review, we discuss the involvement of Ca(2+) and action potential in ER stress-mediated apoptosis.

Role of Platelet-Derived Microvesicles As Crosstalk Mediators in Atherothrombosis and Future Pharmacology Targets: A Link between Inflammation, Atherosclerosis, and Thrombosis

PubMed Central

Badimon, Lina; Suades, Rosa; Fuentes, Eduardo; Palomo, Iván; Padró, Teresa

2016-01-01

Reports in the last decade have suggested that the role of platelets in atherosclerosis and its thrombotic complications may be mediated, in part, by local secretion of platelet-derived microvesicles (pMVs), small cell blebs released during the platelet activation process. MVs are the most abundant cell-derived microvesicle subtype in the circulation. High concentrations of circulating MVs have been reported in patients with atherosclerosis, acute vascular syndromes, and/or diabetes mellitus, suggesting a potential correlation between the quantity of microvesicles and the clinical severity of the atherosclerotic disease. pMVs are considered to be biomarkers of disease but new information indicates that pMVs are also involved in signaling functions. pMVs evoke or promote haemostatic and inflammatory responses, neovascularization, cell survival, and apoptosis, processes involved in the pathophysiology of cardiovascular disease. This review is focused on the complex cross-talk between platelet-derived microvesicles, inflammatory cells and vascular elements and their relevance in the development of the atherosclerotic disease and its clinical outcomes, providing an updated state-of-the art of pMV involvement in atherothrombosis and pMV potential use as therapeutic agent influencing cardiovascular biomedicine in the future. PMID:27630570

Generation of Cardiomyocytes from Pluripotent Stem Cells.

PubMed

Nakahama, Hiroko; Di Pasquale, Elisa

2016-01-01

The advent of pluripotent stem cells (PSCs) enabled a multitude of studies for modeling the development of diseases and testing pharmaceutical therapeutic potential in vitro. These PSCs have been differentiated to multiple cell types to demonstrate its pluripotent potential, including cardiomyocytes (CMs). However, the efficiency and efficacy of differentiation vary greatly between different cell lines and methods. Here, we describe two different methods for acquiring CMs from human pluripotent lines. One method involves the generation of embryoid bodies, which emulates the natural developmental process, while the other method chemically activates the canonical Wnt signaling pathway to induce a monolayer of cardiac differentiation.

Potential involvement of leptin in carcinogenesis of hepatocellular carcinoma.

PubMed

Wang, Xiu-Jie; Yuan, Shu-Lan; Lu, Qing; Lu, Yan-Rong; Zhang, Jie; Liu, Yan; Wang, Wen-Dong

2004-09-01

To investigate the potential involvement of leptin in carcinogenesis of hepatocellular carcinoma (HCC) and to elucidate the etiology, carcinogenesis and progress of HCC. Expressions of Ob gene product, leptin and its receptor, Ob-R were investigated in 36 cases of HCC specimens and corresponding adjacent non-tumorous liver tissues with immunohistochemical staining. The effect of leptin on proliferation of Chang liver cell line and liver cancer cell line SMMC-7721 was studied with cell proliferation assay (MTT). Leptin expression was detected in 36 cases of adjacent non-tumorous liver tissues (36/36, 100%) with moderate (++) to strong (+++) intensity; and in 72.22%(26/36) of HCC with weaker (+) intensity (P<0.05). Thirty of 36 (83.33%) cases of adjacent non-tumorous liver tissues were positive for Ob-R, with moderate (++) to strong (+++) intensity. In HCC, 11/36 (30.56%) cases were positive, with weak (+) intensity (P<0.05). In cell proliferation assay, leptin inhibited the proliferation of Chang liver cells. The cell survival rate was 10-13% lower than that of the untreated cells (P>0.05). Leptin had little effect on the proliferation of liver cancer cells (P>0.05). High level expression and decreased or absent expression of leptin and its receptor in adjacent non-tumorous liver cells and HCC cells, inhibitory effect of leptin on the proliferation of normal Chang liver cells and no effect of leptin on proliferation of liver cancer cells, may provide new insights into the carcinogenesis and progression of human HCC. It could be assumed that leptin acting as an inhibitor and/or promoter, is involved in the process of carcinogenesis and progress of human HCC. Copyright 2004 The WJG Press ISSN

Mediators involved in the immunomodulatory effects of apoptotic cells

PubMed Central

Saas, Philippe; Bonnefoy, Francis; Kury-Paulin, Stephanie; Kleinclauss, François M.; Perruche, Sylvain

2007-01-01

Immunomodulatory properties are attributed to apoptotic cells. These properties have been used to modulate allogeneic immune responses in experimental transplantation settings. In independent studies, apoptotic cell infusion has been shown to favor hematopoietic cell engraftment, to increase heart graft survival and to delay the lethal onset of graft-versus-host disease (GVHD). The goal of this review was to discuss how apoptotic cell infusion interferes with graft rejection or host rejection (i.e., GVHD) and to focus on the potential mediators or “perpetuators” involved in apoptotic cell-induced immunomodulation. Particular emphasis on apoptotic cell phagocytosis, TGF-β secretion and regulatory T cell induction was performed. Stimulating “naturally” immunosuppressive molecules (i.e., TGF-β) or immunomodulatory cells (“alternatively-activated” macrophages, certain DC subsets or regulatory T cells) in a physiological manner by using apoptotic cell infusion can be a promising way to induce tolerance. PMID:17632410

Evidence for Pipecolate Oxidase in Mediating Protection Against Hydrogen Peroxide Stress.

PubMed

Natarajan, Sathish Kumar; Muthukrishnan, Ezhumalai; Khalimonchuk, Oleh; Mott, Justin L; Becker, Donald F

2017-07-01

Pipecolate, an intermediate of the lysine catabolic pathway, is oxidized to Δ 1 -piperideine-6-carboxylate (P6C) by the flavoenzyme l-pipecolate oxidase (PIPOX). P6C spontaneously hydrolyzes to generate α-aminoadipate semialdehyde, which is then converted into α-aminoadipate acid by α-aminoadipatesemialdehyde dehydrogenase. l-pipecolate was previously reported to protect mammalian cells against oxidative stress. Here, we examined whether PIPOX is involved in the mechanism of pipecolate stress protection. Knockdown of PIPOX by small interference RNA abolished pipecolate protection against hydrogen peroxide-induced cell death in HEK293 cells suggesting a critical role for PIPOX. Subcellular fractionation analysis showed that PIPOX is localized in the mitochondria of HEK293 cells consistent with its role in lysine catabolism. Signaling pathways potentially involved in pipecolate protection were explored by treating cells with small molecule inhibitors. Inhibition of both mTORC1 and mTORC2 kinase complexes or inhibition of Akt kinase alone blocked pipecolate protection suggesting the involvement of these signaling pathways. Phosphorylation of the Akt downstream target, forkhead transcription factor O3 (FoxO3), was also significantly increased in cells treated with pipecolate, further implicating Akt in the protective mechanism and revealing FoxO3 inhibition as a potentially key step. The results presented here demonstrate that pipecolate metabolism can influence cell signaling during oxidative stress to promote cell survival and suggest that the mechanism of pipecolate protection parallels that of proline, which is also metabolized in the mitochondria. J. Cell. Biochem. 118: 1678-1688, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Calorimetric determination of thermal parameters for the Li/BrCl in SOCl2 (BCX) chemistry

NASA Technical Reports Server (NTRS)

Darcy, Eric C.; Kalu, Eric E.; White, Ralph E.

1990-01-01

The heat capacity of a Li-BCX DD-cell was found to be dependent on its state of charge by drop calorimetry measurements. The method of drop calorimetry involves measuring the energy (joules) gained or lost from a sample that is transferred from a bath at temperature A to one at temperature B. The thermoneutral potential is defined as the cell potential where the cell electrochemical reactions are neither exothermic nor endothermic. A Hart scientific calorimeter system, Model No. S77XX, designed for heat conduction calorimetry and drop calorimetry was used. Calorimetric analysis yielded a thermoneutral potential of 4.14 volts and a cell heat capacity dependent on the state of charge.

Modulation of vinblastine cytotoxicity by dilantin (phenytoin) or the protein phosphatase inhibitor okadaic acid involves the potentiation of anti-mitotic effects and induction of apoptosis in human tumour cells.

PubMed Central

Kawamura, K. I.; Grabowski, D.; Weizer, K.; Bukowski, R.; Ganapathi, R.

1996-01-01

Cellular insensitivity to vinca alkaloids is suggested to be primarily due to drug efflux by P-glycoprotein (P-gp). The anti-epileptic phenytoin (DPH), which does not bind to P-gp, can selectively enhance vincristine (VCR) cytotoxicity in wild-type (WT) or multidrug-resistant (MDR) cells. We now demonstrate that the protein phosphatase inhibitor okadaic acid (OKA) can mimic the effect of DPH by selectively enhancing cytotoxicity of vinblastine (VBL), but not taxol and doxorubicin, in human leukaemia HL-60 cells. Both DPH and OKA potentiate the anti-mitotic effects of VBL by enhanced damage to the mitotic spindle, resulting in prolonged growth arrest. Also, unlike VBL alone, in human leukaemia or non-small-cell lung carcinoma cells treated with VBL plus DPH, recovery from damage to the mitotic spindle is compromised in drug-free medium and cell death by apoptosis in interphase ensues. Since protein phosphatases are involved with the regulation of metaphase to anaphase transit of cells during the mitotic cycle, enhanced VBL cytotoxicity in the presence of DPH or OKA may involve effects during metaphase on the mitotic spindle tubulin leading to growth arrest and apoptosis in interphase. These novel results suggest that DPH or OKA could be powerful tools to study cellular effects of vinca alkaloids and possibly for the development of novel therapeutic strategies. Images Figure 6 PMID:8546904

Proteomic analysis of plasma membranes isolated from undifferentiated and differentiated HepaRG cells

PubMed Central

2012-01-01

Liver infection with hepatitis B virus (HBV), a DNA virus of the Hepadnaviridae family, leads to severe disease, such as fibrosis, cirrhosis and hepatocellular carcinoma. The early steps of the viral life cycle are largely obscure and the host cell plasma membrane receptors are not known. HepaRG is the only proliferating cell line supporting HBV infection in vitro, following specific differentiation, allowing for investigation of new host host-cell factors involved in viral entry, within a more robust and reproducible environment. Viral infection generally begins with receptor recognition at the host cell surface, following highly specific cell-virus interactions. Most of these interactions are expected to take place at the plasma membrane of the HepaRG cells. In the present study, we used this cell line to explore changes between the plasma membrane of undifferentiated (−) and differentiated (+) cells and to identify differentially-regulated proteins or signaling networks that might potentially be involved in HBV entry. Our initial study identified a series of proteins that are differentially expressed in the plasma membrane of (−) and (+) cells and are good candidates for potential cell-virus interactions. To our knowledge, this is the first study using functional proteomics to study plasma membrane proteins from HepaRG cells, providing a platform for future experiments that will allow us to understand the cell-virus interaction and mechanism of HBV viral infection. PMID:22857383

Are Th17 cells and their cytokines a therapeutic target in Guillain-Barré syndrome?

PubMed

Wu, Xiujuan; Wang, Juan; Liu, Kangding; Zhu, Jie; Zhang, Hong-Liang

2016-01-01

Guillain-Barré syndrome (GBS) is an immune-mediated inflammatory disorder of the peripheral nervous system (PNS). Experimental autoimmune neuritis (EAN) is a useful animal model for studying GBS. Currently, GBS remains a life-threatening disorder and more effective therapeutic strategies are in urgent need. Accumulating evidence has revealed that T helper (Th) 17 cells and their cytokines are pathogenic in GBS/EAN. Drugs attenuated clinical signs of GBS/EAN, in part, by decreasing Th17 cells or IL-17A. Th17 cells and their cytokines might be potential therapeutic targets. Approaches targeting Th17 cells or their cytokines are in development in treating Th17 cells-involved disorders. In this review, we summarize the up-to-date knowledge on roles of Th17 cells and their cytokines in GBS/EAN, as well potential approaches targeting Th17 cells and their cytokines as clinical applications. As Th17 cells produce different sets of pro-inflammatory cytokines and Th17-related cytokines are not exclusively produced by Th17 cells, targeting Th17 cell development may be superior to blocking a single Th17 cytokine to treat Th17 cells-involved disorders. Considering the essential role of retinoic acid-related orphan receptor γT (RORγT) and IL-23 in Th17 cell development, RORγT inhibitors or IL-23 antagonists may provide better clinical efficacy in treating GBS/EAN.

Cytotoxicity and Genotoxicity of Cypermethrin in Hepatocarcinoma Cells: A Dose- and Time-Dependent Study

PubMed Central

AlKahtane, Abdullah A.; Alarifi, Saud; Al-Qahtani, Ahmed A.; Ali, Daoud; Alomar, Suliman Y.; Aleissia, Mohammed S.; Alkahtani, Saad

2018-01-01

Most of the agricultural workers are potentially exposed to pesticides through different routes. Inhalation exposures may result in numerous diseases that can adversely affect an individual’s health and capacity to perform at work. The aim of this study was to determine the cytotoxic potential of cypermethrin pesticide on cultured human hepatocarcinoma (HepG2) cells. The HepG2 cells were exposed to cypermethrin (0, 5, 15, 40 ng/mL) for 24 and 48 hours. We observed that cypermethrin caused cell death of HepG2 cells using 3-(4, 5-dimethylthiozolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) and lactate dehydrogenase tests. Furthermore, cypermethrin reduced HepG2 cells viability in a time and dose dependent basis, that was probably mediated through the induction of reactive oxygen species (ROS) and apoptosis. An increase in ROS generation with a concomitant increase in expression of the proapoptotic protein Bcl-2 and cytochrome c and decrease in the antiapoptosis protein Bax suggested that a mitochondria-mediated pathway was involved in cypermethrin-induced apoptosis. These findings provide insights into the underlying mechanisms involved in cytotoxicity of cypermethrin in HepG2 cells. PMID:29686591

Cytotoxicity and Genotoxicity of Cypermethrin in Hepatocarcinoma Cells: A Dose- and Time-Dependent Study.

PubMed

AlKahtane, Abdullah A; Alarifi, Saud; Al-Qahtani, Ahmed A; Ali, Daoud; Alomar, Suliman Y; Aleissia, Mohammed S; Alkahtani, Saad

2018-01-01

Most of the agricultural workers are potentially exposed to pesticides through different routes. Inhalation exposures may result in numerous diseases that can adversely affect an individual's health and capacity to perform at work. The aim of this study was to determine the cytotoxic potential of cypermethrin pesticide on cultured human hepatocarcinoma (HepG2) cells. The HepG2 cells were exposed to cypermethrin (0, 5, 15, 40 ng/mL) for 24 and 48 hours. We observed that cypermethrin caused cell death of HepG2 cells using 3-(4, 5-dimethylthiozolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) and lactate dehydrogenase tests. Furthermore, cypermethrin reduced HepG2 cells viability in a time and dose dependent basis, that was probably mediated through the induction of reactive oxygen species (ROS) and apoptosis. An increase in ROS generation with a concomitant increase in expression of the proapoptotic protein Bcl-2 and cytochrome c and decrease in the antiapoptosis protein Bax suggested that a mitochondria-mediated pathway was involved in cypermethrin-induced apoptosis. These findings provide insights into the underlying mechanisms involved in cytotoxicity of cypermethrin in HepG2 cells.

Improving the outcome of adoptive cell transfer by targeting tumor escape

PubMed Central

Kaluza, Karen M.; Vile, Richard

2013-01-01

Adoptive T-cell transfer is among the most promising immunotherapies against cancer. To continue increasing the potential of this therapy, our studies focus on the inhibition of tumor recurrence. Recently, we have demonstrated several ways in which combination therapies involving multiple T-cell populations and immunostimulatory chemotherapy can enhance long-term survival. PMID:23483796

Surface-modified polymers for cardiac tissue engineering.

PubMed

Moorthi, Ambigapathi; Tyan, Yu-Chang; Chung, Tze-Wen

2017-09-26

Cardiovascular disease (CVD), leading to myocardial infarction and heart failure, is one of the major causes of death worldwide. The physiological system cannot significantly regenerate the capabilities of a damaged heart. The current treatment involves pharmacological and surgical interventions; however, less invasive and more cost-effective approaches are sought. Such new approaches are developed to induce tissue regeneration following injury. Hence, regenerative medicine plays a key role in treating CVD. Recently, the extrinsic stimulation of cardiac regeneration has involved the use of potential polymers to stimulate stem cells toward the differentiation of cardiomyocytes as a new therapeutic intervention in cardiac tissue engineering (CTE). The therapeutic potentiality of natural or synthetic polymers and cell surface interactive factors/polymer surface modifications for cardiac repair has been demonstrated in vitro and in vivo. This review will discuss the recent advances in CTE using polymers and cell surface interactive factors that interact strongly with stem cells to trigger the molecular aspects of the differentiation or formulation of cardiomyocytes for the functional repair of heart injuries or cardiac defects.

Activation of the Ubiquitin Proteasome Pathway by Silk Fibroin Modified Chitosan Nanoparticles in Hepatic Cancer Cells

PubMed Central

Yang, Ming-Hui; Chung, Tze-Wen; Lu, Yi-Shan; Chen, Yi-Ling; Tsai, Wan-Chi; Jong, Shiang-Bin; Yuan, Shyng-Shiou; Liao, Pao-Chi; Lin, Po-Chiao; Tyan, Yu-Chang

2015-01-01

Silk fibroin (SF) is a protein with bulky hydrophobic domains and can be easily purified as sericin-free silk-based biomaterial. Silk fibroin modified chitosan nanoparticle (SF-CSNP), a biocompatible material, has been widely used as a potential drug delivery system. Our current investigation studied the bio-effects of the SF-CSNP uptake by liver cells. In this experiment, the characterizations of SF-CSNPs were measured by particle size analysis and protein assay. The average size of the SF-CSNP was 311.9 ± 10.7 nm, and the average zeta potential was +13.33 ± 0.3 mV. The SF coating on the SF-CSNP was 6.27 ± 0.17 μg/mL. Moreover, using proteomic approaches, several proteins involved in the ubiquitin proteasome pathway were identified by analysis of differential protein expressions of HepG2 cell uptake the SF-CSNP. Our experimental results have demonstrated that the SF-CSNP may be involved in liver cancer cell survival and proliferation. PMID:25588218

Extracellular vesicles at the cross-line between basic science and clinical needs.

PubMed

Sasso, Luana; Hosamuddin, Huma; Emanueli, Costanza

2017-01-01

MiRNAs are small noncoding RNAs vital for protein regulation and gene expression. Since their discovery in the early nineties, many of their intracellular roles have been characterized. However, it is only recently that EVs loaded with miRNAs and other molecular types have started to be appreciated for their substantial involvement in cell-to-cell communication and signaling in physiological and pathological processes. EVs cell-to-cell signaling functions are complex and largely unknown, which still hampers the direct use of endogenous engineered EVs as therapeutics. However, ad hoc engineered synthetic EVs could represent new therapeutics. The potential of EV-inspired delivery carriers has now attracted the interest of the pharmaceutical industry and has challenged drug delivery researchers with new questions. This review will focus on EVs and EV-inspired drug delivery carriers, on their potential and on the challenges involved in the use of EV-inspired drug delivery systems. © 2016 John Wiley & Sons Ltd.

Giant cell phlebitis: a potentially lethal clinical entity

PubMed Central

Kunieda, Takeshige; Murayama, Masanori; Ikeda, Tsuneko; Yamakita, Noriyoshi

2012-01-01

An 83-year-old woman presented to us with a 4-week history of general malaise, subjective fever and lower abdominal pain. Despite the intravenous infusion of antibiotics, her blood results and physical condition worsened, resulting in her sudden death. Autopsy study revealed that the medium-sized veins of the mesentery were infiltrated by eosinophil granulocytes, lymphocytes, macrophages and multinucleated giant cells; however, the arteries were not involved. Microscopically, venous giant cell infiltration was observed in the gastrointestinal tract, bladder, retroperitoneal tissues and myocardium. The final diagnosis was giant cell phlebitis, a rare disease of unknown aetiology. This case demonstrates for the first time that giant cell phlebitis involving extra-abdominal organs, including hearts, can cause serious morbidity. PMID:22859384

Intercellular signaling through secreted proteins induces free-energy gradient-directed cell movement.

PubMed

Kravchenko-Balasha, Nataly; Shin, Young Shik; Sutherland, Alex; Levine, R D; Heath, James R

2016-05-17

Controlling cell migration is important in tissue engineering and medicine. Cell motility depends on factors such as nutrient concentration gradients and soluble factor signaling. In particular, cell-cell signaling can depend on cell-cell separation distance and can influence cellular arrangements in bulk cultures. Here, we seek a physical-based approach, which identifies a potential governed by cell-cell signaling that induces a directed cell-cell motion. A single-cell barcode chip (SCBC) was used to experimentally interrogate secreted proteins in hundreds of isolated glioblastoma brain cancer cell pairs and to monitor their relative motions over time. We used these trajectories to identify a range of cell-cell separation distances where the signaling was most stable. We then used a thermodynamics-motivated analysis of secreted protein levels to characterize free-energy changes for different cell-cell distances. We show that glioblastoma cell-cell movement can be described as Brownian motion biased by cell-cell potential. To demonstrate that the free-energy potential as determined by the signaling is the driver of motion, we inhibited two proteins most involved in maintaining the free-energy gradient. Following inhibition, cell pairs showed an essentially random Brownian motion, similar to the case for untreated, isolated single cells.

Cell Signaling Experiments Driven by Optical Manipulation

PubMed Central

Difato, Francesco; Pinato, Giulietta; Cojoc, Dan

2013-01-01

Cell signaling involves complex transduction mechanisms in which information released by nearby cells or extracellular cues are transmitted to the cell, regulating fundamental cellular activities. Understanding such mechanisms requires cell stimulation with precise control of low numbers of active molecules at high spatial and temporal resolution under physiological conditions. Optical manipulation techniques, such as optical tweezing, mechanical stress probing or nano-ablation, allow handling of probes and sub-cellular elements with nanometric and millisecond resolution. PicoNewton forces, such as those involved in cell motility or intracellular activity, can be measured with femtoNewton sensitivity while controlling the biochemical environment. Recent technical achievements in optical manipulation have new potentials, such as exploring the actions of individual molecules within living cells. Here, we review the progress in optical manipulation techniques for single-cell experiments, with a focus on force probing, cell mechanical stimulation and the local delivery of active molecules using optically manipulated micro-vectors and laser dissection. PMID:23698758

Role of cationic channel TRPV2 in promoting prostate cancer migration and progression to androgen resistance.

PubMed

Monet, Michaël; Lehen'kyi, V'yacheslav; Gackiere, Florian; Firlej, Virginie; Vandenberghe, Matthieu; Roudbaraki, Morad; Gkika, Dimitra; Pourtier, Albin; Bidaux, Gabriel; Slomianny, Christian; Delcourt, Philippe; Rassendren, François; Bergerat, Jean-Pierre; Ceraline, Jocelyn; Cabon, Florence; Humez, Sandrine; Prevarskaya, Natalia

2010-02-01

Castration resistance in prostate cancer (PCa) constitutes an advanced, aggressive disease with poor prognosis, associated with uncontrolled cell proliferation, resistance to apoptosis, and enhanced invasive potential. The molecular mechanisms involved in the transition of PCa to castration resistance are obscure. Here, we report that the nonselective cationic channel transient receptor potential vanilloid 2 (TRPV2) is a distinctive feature of castration-resistant PCa. TRPV2 transcript levels were higher in patients with metastatic cancer (stage M1) compared with primary solid tumors (stages T2a and T2b). Previous studies of the TRPV2 channel indicated that it is primarily involved in cancer cell migration and not in cell growth. Introducing TRPV2 into androgen-dependent LNCaP cells enhanced cell migration along with expression of invasion markers matrix metalloproteinase (MMP) 9 and cathepsin B. Consistent with the likelihood that TRPV2 may affect cancer cell aggressiveness by influencing basal intracellular calcium levels, small interfering RNA-mediated silencing of TRPV2 reduced the growth and invasive properties of PC3 prostate tumors established in nude mice xenografts, and diminished expression of invasive enzymes MMP2, MMP9, and cathepsin B. Our findings establish a role for TRPV2 in PCa progression to the aggressive castration-resistant stage, prompting evaluation of TRPV2 as a potential prognostic marker and therapeutic target in the setting of advanced PCa.

Mechanisms of crystalline silica-induced pulmonary toxicity revealed by global gene expression profiling

PubMed Central

Sellamuthu, Rajendran; Umbright, Christina; Li, Shengqiao; Kashon, Michael; Joseph, Pius

2015-01-01

A proper understanding of the mechanisms underlying crystalline silica-induced pulmonary toxicity has implications in the management and potential prevention of the adverse health effects associated with silica exposure including silicosis, cancer and several auto-immune diseases. Human lung type II epithelial cells and rat lungs exposed to crystalline silica were employed as experimental models to determine global gene expression changes in order to understand the molecular mechanisms underlying silica-induced pulmonary toxicity. The differential gene expression profile induced by silica correlated with its toxicity in the A549 cells. The biological processes perturbed by silica exposure in the A549 cells and rat lungs, as identified by the bioinformatics analysis of the differentially expressed genes, demonstrated significant similarity. Functional categorization of the differentially expressed genes identified cancer, cellular movement, cellular growth and proliferation, cell death, inflammatory response, cell cycle, cellular development, and genetic disorder as top ranking biological functions perturbed by silica exposure in A549 cells and rat lungs. Results of our study, in addition to confirming several previously identified molecular targets and mechanisms involved in silica toxicity, identified novel molecular targets and mechanisms potentially involved in silica-induced pulmonary toxicity. Further investigations, including those focused on the novel molecular targets and mechanisms identified in the current study may result in better management and, possibly, reduction and/or prevention of the potential adverse health effects associated with crystalline silica exposure. PMID:22087542

Mitochondrial electron transport chain is involved in microcystin-RR induced tobacco BY-2 cells apoptosis.

PubMed

Huang, Wenmin; Li, Dunhai; Liu, Yongding

2014-09-01

Microcystin-RR (MC-RR) has been suggested to induce apoptosis in tobacco BY-2 cells through mitochondrial dysfunction including the loss of mitochondrial membrane potential (ΔΨm). To further elucidate the mechanisms involved in MC-RR induced apoptosis in tobacco BY-2 cells, we have investigated the role of mitochondrial electron transport chain (ETC) as a potential source for reactive oxygen species (ROS). Tobacco BY-2 cells after exposure to MC-RR (60mg/L) displayed apoptotic changes in association with an increased production of ROS and loss of ΔΨm. All of these adverse effects were significantly attenuated by ETC inhibitors including Rotenone (2μmol/L, complex I inhibitor) and antimycin A (0.01μmol/L, complex III inhibitor), but not by thenoyltrifluoroacetone (5μmol/L, complex II inhibitor). These results suggest that mitochondrial ETC plays a key role in mediating MC-RR induced apoptosis in tobacco BY-2 cells through an increased mitochondrial production of ROS. Copyright © 2014. Published by Elsevier B.V.

Activation of formylmethanofuran synthesis in cell extracts of Methanobacterium thermoautotrophicum.

PubMed Central

Bobik, T A; Wolfe, R S

1989-01-01

In cell extracts of Methanobacterium thermoautotrophicum, formylmethanofuran (formyl-MFR) synthesis (an essential CO2 fixation reaction that is an early step in CO2 reduction to methane) is subject to a complex activation that involves a heterodisulfide of coenzyme M and N-(7-mercaptoheptanoyl)threonine O3-phosphate (CoM-S-S-HTP). In this paper we report that titanium(III) citrate, a low-potential reducing agent, stimulated CO2 reduction to methane and activated formyl-MFR synthesis in cell extracts. Titanium(III) citrate functioned as the sole source of electrons for formyl-MFR synthesis and enabled this reaction to occur independently of CoM-S-S-HTP. In addition, CoM-S-S-HTP was found to activate an unknown electron carrier that reduced metronidazole. The activation of formyl-MFR synthesis by CoM-S-S-HTP may involve the activation of a low-potential electron carrier. PMID:2921239

Autophagy induced by baicalin involves downregulation of CD147 in SMMC-7721 cells in vitro

PubMed Central

ZHANG, XIANJIAO; TANG, XU; LIU, HANQIANG; LI, LIANXIANG; HOU, QIAN; GAO, JIANMIN

2012-01-01

Baicalin has been demonstrated to exert anticancer effects mainly through induction of tumor cell apoptosis and cell cycle arrest. However, the precise mechanisms underlying its anticancer role remain to be elucidated. In the present study, we investigated whether autophagy was involved in the anticancer activity of baicalin in the human hepatocellular carcinoma (HCC) cell line SMMC-7721 and the possible molecular mechanisms. Our data showed that the viability of SMMC-7721 cells was significantly inhibited by baicalin in a dose- and time-dependent manner. Alongside apoptosis, autophagy was also induced by baicalin dose- and time-dependently with the involvement of the autophagy-associated protein Beclin 1. Moreover, we demonstrated that cell death induced by baicalin was significantly inhibited by the apoptosis inhibitor z-DEVD-fmk or the autophagy inhibitor 3-MA, respectively. In addition, we found that CD147, a key molecule related both to apoptosis and autophagy, was markedly downregulated at the protein level in SMMC-7721 cells treated with baicalin. Collectively, this is the first study to suggest that baicalin induces autophagic cell death in SMMC-7721 cells, which involves the downregulation of CD147. Our study reveals a new mechanism for the anticancer effects of baicalin and puts forward a potential crucial role of CD147 in baicalin-induced cancer cell death. PMID:22200845

Identification of potential target genes of ROR-alpha in THP1 and HUVEC cell lines.

PubMed

Gulec, Cagri; Coban, Neslihan; Ozsait-Selcuk, Bilge; Sirma-Ekmekci, Sema; Yildirim, Ozlem; Erginel-Unaltuna, Nihan

2017-04-01

ROR-alpha is a nuclear receptor, activity of which can be modulated by natural or synthetic ligands. Due to its possible involvement in, and potential therapeutic target for atherosclerosis, we aimed to identify ROR-alpha target genes in monocytic and endothelial cell lines. We performed chromatin immunoprecipitation (ChIP) followed by tiling array (ChIP-on-chip) for ROR-alpha in monocytic cell line THP1 and endothelial cell line HUVEC. Following bioinformatic analysis of the array data, we tested four candidate genes in terms of dependence of their expression level on ligand-mediated ROR-alpha activity, and two of them in terms of promoter occupancy by ROR-alpha. Bioinformatic analyses of ChIP-on-chip data suggested that ROR-alpha binds to genomic regions near the transcription start site (TSS) of more than 3000 genes in THP1 and HUVEC. Potential ROR-alpha target genes in both cell types seem to be involved mainly in membrane receptor activity, signal transduction and ion transport. While SPP1 and IKBKA were shown to be direct target genes of ROR-alpha in THP1 monocytes, inflammation related gene HMOX1 and heat shock protein gene HSPA8 were shown to be potential target genes of ROR-alpha. Our results suggest that ROR-alpha may regulate signaling receptor activity, and transmembrane transport activity through its potential target genes. ROR-alpha seems also to play role in cellular sensitivity to environmental substances like arsenite and chloroprene. Although, the expression analyses have shown that synthetic ROR-alpha ligands can modulate some of potential ROR-alpha target genes, functional significance of ligand-dependent modulation of gene expression needs to be confirmed with further analyses. Copyright © 2017 Elsevier Inc. All rights reserved.

Peroxide accumulation and cell death in filamentous fungi induced by contact with a contestant.

PubMed

Silar, Philippe

2005-02-01

Podospora anserina and Coprinopsis cinerea (syn. Coprinus cinereus) are endowed with a defence system able to differentiate self vs. non-self and involving the generation of peroxide. Indeed, they produce peroxide when confronted with a filamentous fungus, only in non-self confrontations. Both species are not able to recognize yeasts and show a differential response to bacteria. The accumulation of peroxides in the ascomycete Podospora anserina requires an NADPH oxidase and a MAP kinase cascade, previously shown to be involved in fruit body formation, cell differentiation and cell degeneration. Confrontation is accompanied by the death of the contestant hyphae only in specific combinations of species. As in animals and plants, data suggest that peroxide is likely involved in signalling rather than playing a direct toxic role. Fungi display more complex behaviours than generally acknowledged, i.e. they are able to recognize potential contestants and built up defence reactions involving evolutionary conserved enzymes.

Increased metastatic potential of tumor cells in von Willebrand factor-deficient mice.

PubMed

Terraube, V; Pendu, R; Baruch, D; Gebbink, M F B G; Meyer, D; Lenting, P J; Denis, C V

2006-03-01

The key role played by von Willebrand factor (VWF) in platelet adhesion suggests a potential implication in various pathologies, where this process is involved. In cancer metastasis development, tumor cells interact with platelets and the vessel wall to extravasate from the circulation. As a potential mediator of platelet-tumor cell interactions, VWF could influence this early step of tumor spread and therefore play a role in cancer metastasis. To investigate whether VWF is involved in metastasis development. In a first step, we characterized the interaction between murine melanoma cells B16-BL6 and VWF in vitro. In a second step, an experimental metastasis model was used to compare the formation of pulmonary metastatic foci in C57BL/6 wild-type and VWF-null mice following the injection of B16-BL6 cells or Lewis lung carcinoma cells. In vitro adhesion assays revealed that VWF is able to promote a dose-dependent adhesion of B16-BL6 cells via its Arg-Gly-Asp (RGD) sequence. In the experimental metastasis model, we found a significant increase in the number of pulmonary metastatic foci in VWF-null mice compared with the wild-type mice, a phenotype that could be corrected by restoring VWF plasma levels. We also showed that increased survival of the tumor cells in the lungs during the first 24 h in the absence of VWF was the cause of this increased metastasis. These findings suggest that VWF plays a protective role against tumor cell dissemination in vivo. Underlying mechanisms remain to be investigated.

A characterization of the ZFL cell line and primary hepatocytes as in vitro liver cell models for the zebrafish (Danio rerio).

PubMed

Eide, Marta; Rusten, Marte; Male, Rune; Jensen, Knut Helge Midtbø; Goksøyr, Anders

2014-02-01

The zebrafish (Danio rerio) is a widely used model species in biomedical research. The ZFL cell line, established from zebrafish liver, and freshly isolated primary hepatocytes from zebrafish have been used in several toxicological studies. However, no previous report has compared and characterized these two systems at the level of gene expression. The aim of this study was to evaluate the ZFL cell line in comparison to primary hepatocytes as in vitro models for studying effects of environmental contaminants in zebrafish liver. Using quantitative real-time PCR, the basal level and transcriptional induction potential of key genes involved in toxic responses in the ZFL cell line, primary hepatocytes and whole liver from zebrafish were compared. The study showed that the ZFL cells have lower levels of mRNA of most selected genes compared to zebrafish liver. The induced gene transcription following exposure to ligand was much lower in ZFL cells compared to zebrafish primary hepatocytes at the doses tested. Importantly, oestrogen receptor and vitellogenin genes showed low basal transcription and no induction response in the ZFL cell line. In conclusion, it appears that primary hepatocytes are well suited for studying environmental contaminants including xenoestrogens, but may show large sex-dependent differences in gene transcription. The ZFL cell line shows potential in toxicological studies involving the aryl hydrocarbon receptor pathway. However, low potential for transcriptional induction of genes in general should be expected, especially notable when studying estrogenic responses. Copyright © 2013 Elsevier B.V. All rights reserved.

Expansion and hepatic differentiation of rat multipotent adult progenitor cells in microcarrier suspension culture.

PubMed

Park, Y; Subramanian, K; Verfaillie, C M; Hu, W S

2010-10-01

Many potential applications of stem cells require large quantities of cells, especially those involving large organs such as the liver. For such applications, a scalable reactor system is desirable to ensure a reliable supply of sufficient quantities of differentiation competent or differentiated cells. We employed a microcarrier culture system for the expansion of undifferentiated rat multipotent adult progenitor cells (rMAPC) as well as for directed differentiation of these cells to hepatocyte-like cells. During the 4-day expansion culture, cell concentration increased by 85-fold while expression level of pluripotency markers were maintained, as well as the MAPC differentiation potential. Directed differentiation into hepatocyte-like cells on the microcarriers themselves gave comparable results as observed with cells cultured in static cultures. The cells expressed several mature hepatocyte-lineage genes and asialoglycoprotein receptor-1 (ASGPR-1) surface protein, and secreted albumin and urea. Microcarrier culture thus offers the potential of large-scale expansion and differentiation of stem cells in a more controlled bioreactor environment. Copyright © 2010 Elsevier B.V. All rights reserved.

[Stem cells--cloning, plasticity, bioethic].

PubMed

Pflegerl, Pamina; Keller, Thomas; Hantusch, Brigitte; Hoffmann, Thomas Sören; Kenner, Lukas

2008-01-01

Stem cells with certain characteristics have become promising tools for molecular medicine. They have the potential to self-regenerate and to differentiate into specific tissues. Besides their great potential, embryonic stem cells (ESC) run the risk of enhanced tumorigenesis. The use of human embryonic stem cells (hESC) is ethically problematic because their isolation involves the destruction of human embryos. Recently developed methods generate are able to pluripotent stem cells from fibroblasts. Alternatives for ESC are adult stem cells (ASC) derived from bone marrow, cord blood, amniotic fluid and other tissues. The following article is on the basis of testimony of Lukas Kenner for the German Bundestag about the use of ESC for research, therapy and drug development. Ethical aspects are taken into consideration.

LPA Induces Colon Cancer Cell Proliferation through a Cooperation between the ROCK and STAT-3 Pathways.

PubMed

Leve, Fernanda; Peres-Moreira, Rubem J; Binato, Renata; Abdelhay, Eliana; Morgado-Díaz, José A

2015-01-01

Lysophosphatidic acid (LPA) plays a critical role in the proliferation and migration of colon cancer cells; however, the downstream signaling events underlying these processes remain poorly characterized. The aim of this study was to investigate the signaling pathways triggered by LPA to regulate the mechanisms involved in the progression of colorectal cancer (CRC). We have used three cell line models of CRC, and initially analyzed the expression profile of LPA receptors (LPAR). Then, we treated the cells with LPA and events related to their tumorigenic potential, such as migration, invasion, anchorage-independent growth, proliferation as well as apoptosis and cell cycle were evaluated. We used the Chip array technique to analyze the global gene expression profiling that occurs after LPA treatment, and we identified cell signaling pathways related to the cell cycle. The inhibition of these pathways verified the conclusions of the transcriptomic analysis. We found that the cell lines expressed LPAR1, -2 and -3 in a differential manner and that 10 μM LPA did not affect cell migration, invasion and anchorage-independent growth, but it did induce proliferation and cell cycle progression in HCT-116 cells. Although LPA in this concentration did not induce transcriptional activity of β-catenin, it promoted the activation of Rho and STAT-3. Moreover, ROCK and STAT-3 inhibitors prevented LPA-induced proliferation, but ROCK inhibition did not prevent STAT-3 activation. Finally, we observed that LPA regulates the expression of genes related to the cell cycle and that the combined inhibition of ROCK and STAT-3 prevented cell cycle progression and increased the LPA-induced expression of cyclins E1, A2 and B1 to a greater degree than either inhibitor alone. Overall, these results demonstrate that LPA increases the proliferative potential of colon adenocarcinoma HCT-116 cells through a mechanism involving cooperation between the Rho-ROCK and STAT3 pathways involved in cell cycle control.

Apoptosis and autophagy induced by pyropheophorbide-α methyl ester-mediated photodynamic therapy in human osteosarcoma MG-63 cells.

PubMed

Huang, Qiu; Ou, Yun-Sheng; Tao, Yong; Yin, Hang; Tu, Ping-Hua

2016-06-01

Pyropheophorbide-α methyl ester (MPPa) was a second-generation photosensitizer with many potential applications. Here, we explored the impact of MPPa-mediated photodynamic therapy (MPPa-PDT) on the apoptosis and autophagy of human osteosarcoma (MG-63) cells as well as the relationships between apoptosis and autophagy of the cells, and investigated the related molecular mechanisms. We found that MPPa-PDT demonstrated the ability to inhibit MG-63 cell viability in an MPPa concentration- and light dose-dependent manner, and to induce apoptosis via the mitochondrial apoptosis pathway. Additionally, MPPa-PDT could also induce autophagy of MG-63 cell. Meanwhile, the ROS scavenger N-acetyl-L-cysteine (NAC) and the Jnk inhibitor SP600125 were found to inhibit the MPPa-PDT-induced autophagy, and NAC could also inhibit Jnk phosphorylation. Furthermore, pretreatment with the autophagy inhibitor 3-methyladenine or chloroquine showed the potential in reducing the apoptosis rate induced by MPPa-PDT in MG-63 cells. Our results indicated that the mitochondrial pathway was involved in MPPa-PDT-induced apoptosis of MG-63 cells. Meanwhile the ROS-Jnk signaling pathway was involved in MPPa-PDT-induced autophagy, which further promoted the apoptosis in MG-63 cells.

Alternative Sources of Adult Stem Cells: Human Amniotic Membrane

NASA Astrophysics Data System (ADS)

Wolbank, Susanne; van Griensven, Martijn; Grillari-Voglauer, Regina; Peterbauer-Scherb, Anja

Human amniotic membrane is a highly promising cell source for tissue engineering. The cells thereof, human amniotic epithelial cells (hAEC) and human amniotic mesenchymal stromal cells (hAMSC), may be immunoprivileged, they represent an early developmental status, and their application is ethically uncontroversial. Cell banking strategies may use freshly isolated cells or involve in vitro expansion to increase cell numbers. Therefore, we have thoroughly characterized the effect of in vitro cultivation on both phenotype and differentiation potential of hAEC. Moreover, we present different strategies to improve expansion including replacement of animal-derived supplements by human platelet products or the introduction of the catalytic subunit of human telomerase to extend the in vitro lifespan of amniotic cells. Characterization of the resulting cultures includes phenotype, growth characteristics, and differentiation potential, as well as immunogenic and immunomodulatory properties.

Enhanced susceptibility of cancer cells to oncolytic rhabdo-virotherapy by expression of Nodamura virus protein B2 as a suppressor of RNA interference.

PubMed

Bastin, Donald; Aitken, Amelia S; Pelin, Adrian; Pikor, Larissa A; Crupi, Mathieu J F; Huh, Michael S; Bourgeois-Daigneault, Marie-Claude; Bell, John C; Ilkow, Carolina S

2018-06-19

Antiviral responses are barriers that must be overcome for efficacy of oncolytic virotherapy. In mammalian cells, antiviral responses involve the interferon pathway, a protein-signaling cascade that alerts the immune system and limits virus propagation. Tumour-specific defects in interferon signaling enhance viral infection and responses to oncolytic virotherapy, but many human cancers are still refractory to oncolytic viruses. Given that invertebrates, fungi and plants rely on RNA interference pathways for antiviral protection, we investigated the potential involvement of this alternative antiviral mechanism in cancer cells. Here, we detected viral genome-derived small RNAs, indicative of RNAi-mediated antiviral responses, in human cancer cells. As viruses may encode suppressors of the RNA interference pathways, we engineered an oncolytic vesicular stomatitis virus variant to encode the Nodamura virus protein B2, a known inhibitor of RNAi-mediated immune responses. B2-expressing oncolytic virus showed enhanced viral replication and cytotoxicity, impaired viral genome cleavage and altered microRNA processing in cancer cells. Our data establish the improved therapeutic potential of our novel virus which targets the RNAi-mediated antiviral defense of cancer cells.

Antimetabolic Effects of Polyphenols in Breast Cancer Cells: Focus on Glucose Uptake and Metabolism.

PubMed

Keating, Elisa; Martel, Fátima

2018-01-01

In the last years, metabolic reprogramming became a new key hallmark of tumor cells. One of its components is a deviant energetic metabolism, known as Warburg effect-an aerobic lactatogenesis- characterized by elevated rates of glucose uptake and consumption with high-lactate production even in the presence of oxygen. Because many cancer cells display a greater sensitivity to glucose deprivation-induced cytotoxicity than normal cells, inhibitors of glucose cellular uptake (facilitative glucose transporter 1 inhibitors) and oxidative metabolism (glycolysis inhibitors) are potential therapeutic targets in cancer treatment. Polyphenols, abundantly contained in fruits and vegetables, are dietary components with an established protective role against cancer. Several molecular mechanisms are involved in the anticancer effect of polyphenols, including effects on apoptosis, cell cycle regulation, plasma membrane receptors, signaling pathways, and epigenetic mechanisms. Additionally, inhibition of glucose cellular uptake and metabolism in cancer cell lines has been described for several polyphenols, and this effect was shown to be associated with their anticarcinogenic effect. This work will review data showing an antimetabolic effect of polyphenols and its involvement in the chemopreventive/chemotherapeutic potential of these dietary compounds, in relation to breast cancer.

BMI-1, a promising therapeutic target for human cancer

PubMed Central

WANG, MIN-CONG; LI, CHUN-LI; CUI, JIE; JIAO, MIN; WU, TAO; JING, LI; NAN, KE-JUN

2015-01-01

BMI-1 oncogene is a member of the polycomb-group gene family and a transcriptional repressor. Overexpression of BMI-1 has been identified in various human cancer tissues and is known to be involved in cancer cell proliferation, cell invasion, distant metastasis, chemosensitivity and patient survival. Accumulating evidence has revealed that BMI-1 is also involved in the regulation of self-renewal, differentiation and tumor initiation of cancer stem cells (CSCs). However, the molecular mechanisms underlying these biological processes remain unclear. The present review summarized the function of BMI-1 in different human cancer types and CSCs, and discussed the signaling pathways in which BMI-1 is potentially involved. In conclusion, BMI-1 may represent a promising target for the prevention and therapy of various cancer types. PMID:26622537

Cutting Edge: TCR Revision Affects Predominantly Foxp3− Cells and Skews Them toward the Th17 Lineage1

PubMed Central

Zehn, Dietmar; Bevan, Michael J.; Fink, Pamela J.

2009-01-01

CD4+ T cells respond to peripheral endogenous superantigen stimulation by undergoing deletion or TCR revision. The latter involves RAG re-expression, TCR gene rearrangement, and expression of a novel TCR. TCR-revised T cells are functional and express a diverse TCR repertoire. Because TCR revision harbors the potential to create self-reactivity, it is important to explore whether T cells known to be self-reactive (regulatory T cells) or those involved in autoimmunity (Th17 cells) arise from TCR revision. Interestingly, we observed that Foxp3+ cells are excluded from revising their TCR and that only a small fraction of postrevision cells expresses Foxp3. In contrast, Th17 cells are 20 times more frequent among revised than among C57BL/6 CD4+ T cells, indicating that postrevision cells are biased toward the Th17 lineage. The link between Th17 differentiation and TCR revision might be highly relevant to the role of Th17 cells in promoting autoimmunity. PMID:17947636

Visualization of gene expression in the live subject using the Na/I symporter as a reporter gene: applications in biotherapy.

PubMed

Baril, Patrick; Martin-Duque, Pilar; Vassaux, Georges

2010-02-01

Biotherapies involve the utilization of antibodies, genetically modified viruses, bacteria or cells for therapeutic purposes. Molecular imaging has the potential to provide unique information that will guarantee their biosafety in humans and provide a rationale for the future development of new generations of reagents. In this context, non-invasive imaging of gene expression is an attractive prospect, allowing precise, spacio-temporal measurements of gene expression in longitudinal studies involving gene transfer vectors. With the emergence of cell therapies in regenerative medicine, it is also possible to track cells injected into subjects. In this context, the Na/I symporter (NIS) has been used in preclinical studies. Associated with a relevant radiotracer ((123)I(-), (124)I(-), (99m)TcO4(-)), NIS can be used to monitor gene transfer and the spread of selectively replicative viruses in tumours as well as in cells with a therapeutic potential. In addition to its imaging potential, NIS can be used as a therapeutic transgene through its ability to concentrate therapeutic doses of radionuclides in target cells. This dual property has applications in cancer treatment and could also be used to eradicate cells with therapeutic potential in the case of adverse events. Through experience acquired in preclinical studies, we can expect that non-invasive molecular imaging using NIS as a transgene will be pivotal for monitoring in vivo the exact distribution and pharmacodynamics of gene expression in a precise and quantitative way. This review highlights the applications of NIS in biotherapy, with a particular emphasis on image-guided radiotherapy, monitoring of gene and vector biodistribution and trafficking of stem cells.

The secret life of ion channels: Kv1.3 potassium channels and proliferation.

PubMed

Pérez-García, M Teresa; Cidad, Pilar; López-López, José R

2018-01-01

Kv1.3 channels are involved in the switch to proliferation of normally quiescent cells, being implicated in the control of cell cycle in many different cell types and in many different ways. They modulate membrane potential controlling K + fluxes, sense changes in potential, and interact with many signaling molecules through their intracellular domains. From a mechanistic point of view, we can describe the role of Kv1.3 channels in proliferation with at least three different models. In the "membrane potential model," membrane hyperpolarization resulting from Kv1.3 activation provides the driving force for Ca 2+ influx required to activate Ca 2+ -dependent transcription. This model explains most of the data obtained from several cells from the immune system. In the "voltage sensor model," Kv1.3 channels serve mainly as sensors that transduce electrical signals into biochemical cascades, independently of their effect on membrane potential. Kv1.3-dependent proliferation of vascular smooth muscle cells (VSMCs) could fit this model. Finally, in the "channelosome balance model," the master switch determining proliferation may be related to the control of the Kv1.3 to Kv1.5 ratio, as described in glial cells and also in VSMCs. Since the three mechanisms cannot function independently, these models are obviously not exclusive. Nevertheless, they could be exploited differentially in different cells and tissues. This large functional flexibility of Kv1.3 channels surely gives a new perspective on their functions beyond their elementary role as ion channels, although a conclusive picture of the mechanisms involved in Kv1.3 signaling to proliferation is yet to be reached.

The Antiproliferative Effect of Cyclodipeptides from Pseudomonas aeruginosa PAO1 on HeLa Cells Involves Inhibition of Phosphorylation of Akt and S6k Kinases.

PubMed

Hernández-Padilla, Laura; Vázquez-Rivera, Dolores; Sánchez-Briones, Luis A; Díaz-Pérez, Alma L; Moreno-Rodríguez, José; Moreno-Eutimio, Mario A; Meza-Carmen, Victor; Cruz, Homero Reyes-De la; Campos-García, Jesús

2017-06-20

Pseudomonas aeruginosa PAO1, a potential pathogen of plants and animals, produces the cyclodipeptides cyclo(l-Pro-l-Tyr), cyclo(l-Pro-l-Phe), and cyclo(l-Pro-l-Val) (PAO1-CDPs), whose effects have been implicated in inhibition of human tumor cell line proliferation. Our purpose was to investigate in depth in the mechanisms of HeLa cell proliferation inhibition by the PAO1-CDPs. The results indicate that PAO1-CDPs, both purified individually and in mixtures, inhibited HeLa cell proliferation by arresting the cell cycle at the G0-G1 transition. The crude PAO1-CDPs mixture promoted cell death in HeLa cells in a dose-dependent manner, showing efficacy similar to that of isolated PAO1-CDPs (LD 50 of 60-250 µM) and inducing apoptosis with EC 50 between 0.6 and 3.0 µM. Moreover, PAO1-CDPs showed a higher proapoptotic activity (~10³-10⁵ fold) than their synthetic analogs did. Subsequently, the PAO1-CDPs affected mitochondrial membrane potential and induced apoptosis by caspase-9-dependent pathway. The mechanism of inhibition of cells proliferation in HeLa cells involves inhibition of phosphorylation of both Akt-S473 and S6k-T389 protein kinases, showing a cyclic behavior of their expression and phosphorylation in a time and concentration-dependent fashion. Taken together our findings indicate that PI3K-Akt-mTOR-S6k signaling pathway blockage is involved in the antiproliferative effect of the PAO1-CDPs.

Immunomodulatory Properties of Induced Pluripotent Stem Cell-Derived Mesenchymal Cells.

PubMed

Ng, Jia; Hynes, Kim; White, Gregory; Sivanathan, Kisha Nandini; Vandyke, Kate; Bartold, Peter Mark; Gronthos, Stan

2016-12-01

MSC-like populations derived from induced pluripotent stem cells (iPSC-MSC) serve as an alternative stem cell source due to their high proliferative capacity. In this study, we assessed the immunomodulatory potential of iPSC-MSC generated from periodontal ligament (PDL) and gingival (GF) tissue. The iPSC-MSC lines exhibited a similar level of suppression of mitogen-stimulated peripheral blood mononuclear cells (PBMNC) proliferation compared to their respective parental fibroblast populations in vitro. Moreover, iPSC-MSC demonstrated the ability to suppress T-cells effector cells, Th1/Th2/Th17 populations, and increase levels of Treg cells. In order to investigate the mechanisms involved, expression of common MSC-derived soluble factors known to supress lymphocyte proliferation were assessed in iPSC-MSC cultured with PBMNC with direct cell-cell contact or separated in transwells. Real-time PCR analysis of factors known to be involved in MSC mediated immune regulation, found a general trend of elevated IDO1 and IL6 transcript levels in iPSC-MSC lines and their respective primary cells co-cultured with activated PBMNC, with a wide range of gene expression levels between the different mesenchymal cell types. The results suggest that different iPSC-MSC may be useful as a potential alternative source of cells for future clinical use in therapeutic applications because of their potent immunosuppressive properties. J. Cell. Biochem. 117: 2844-2853, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Annexin V-induced rat Leydig cell proliferation involves Ect2 via RhoA/ROCK signaling pathway.

PubMed

Jing, Jun; Chen, Li; Fu, Hai-Yan; Fan, Kai; Yao, Qi; Ge, Yi-Feng; Lu, Jin-Chun; Yao, Bing

2015-03-24

This study investigated the effect of annexin V on the proliferation of primary rat Leydig cells and the potential mechanism. Our results showed that annexin V promoted rat Leydig cell proliferation and cell cycle progression in a dose- and time-dependent manner. Increased level of annexin V also enhanced Ect2 protein expression. However, siRNA knockdown of Ect2 attenuated annexin V-induced proliferation of rat Leydig cells. Taken together, these data suggest that increased level of annexin V induced rat Leydig cell proliferation and cell cycle progression via Ect2. Since RhoA activity was increased following Ect2 activation, we further investigated whether Ect2 was involved in annexin V-induced proliferation via the RhoA/ROCK pathway, and the results showed that annexin V increased RhoA activity too, and this effect was abolished by the knockdown of Ect2. Moreover, inhibition of the RhoA/ROCK pathway by a ROCK inhibitor, Y27632, also attenuated annexin V-induced proliferation and cell cycle progression. We thus conclude that Ect2 is involved in annexin V-induced rat Leydig cell proliferation through the RhoA/ROCK pathway.

Adult-onset systemic Langerhans cell histiocytosis mimicking inflammatory bowel disease: the value of skin biopsy and review of cases of Langerhans cell histiocytosis with cutaneous involvement seen at the Mayo Clinic.

PubMed

Podjasek, Joshua O; Loftus, Conor G; Smyrk, Thomas C; Wieland, Carilyn N

2014-03-01

Langerhans cell histiocytosis (LCH) is frequently known to involve multiple organ systems. However, gastrointestinal involvement by LCH is rare. We describe a 68-year-old woman with a 3-year history of intermittent diarrhea initially diagnosed as inflammatory bowel disease. She was subsequently found to have systemic LCH with involvement of the gastrointestinal tract, lungs, liver, and skin after skin biopsy was performed. A retrospective review of patients with cutaneous involvement of LCH seen at the Mayo Clinic over the past 15 years was conducted. The presence of systemic disease as well as specific organ system involvement was reviewed. Twenty-four patients with cutaneous LCH were identified. Besides our case, one other patient with both gastrointestinal and cutaneous involvement was identified. This patient died at six months of age. No other adult-onset cases were identified. Gastrointestinal involvement with LCH is rare, can be easily misdiagnosed, and likely portends a poor prognosis. In patients with ill-defined systemic symptoms, cutaneous exam and biopsy have the potential to diagnose systemic disease. © 2013 The International Society of Dermatology.

Leptin-mediated ion channel regulation: PI3K pathways, physiological role, and therapeutic potential.

PubMed

Gavello, Daniela; Carbone, Emilio; Carabelli, Valentina

2016-07-03

Leptin is produced by adipose tissue and identified as a "satiety signal," informing the brain when the body has consumed enough food. Specific areas of the hypothalamus express leptin receptors (LEPRs) and are the primary site of leptin action for body weight regulation. In response to leptin, appetite is suppressed and energy expenditure allowed. Beside this hypothalamic action, leptin targets other brain areas in addition to neuroendocrine cells. LEPRs are expressed also in the hippocampus, neocortex, cerebellum, substantia nigra, pancreatic β-cells, and chromaffin cells of the adrenal gland. It is intriguing how leptin is able to activate different ionic conductances, thus affecting excitability, synaptic plasticity and neurotransmitter release, depending on the target cell. Most of the intracellular pathways activated by leptin and directed to ion channels involve PI3K, which in turn phosphorylates different downstream substrates, although parallel pathways involve AMPK and MAPK. In this review we will describe the effects of leptin on BK, KATP, KV, CaV, TRPC, NMDAR and AMPAR channels and clarify the landscape of pathways involved. Given the ability of leptin to influence neuronal excitability and synaptic plasticity by modulating ion channels activity, we also provide a short overview of the growing potentiality of leptin as therapeutic agent for treating neurological disorders.

Ophiobolin A, a sesterpenoid fungal phytotoxin, displays different mechanisms of cell death in mammalian cells depending upon the cancer cell origin.

PubMed

Morrison, Rachel; Lodge, Tiffany; Evidente, Antonio; Kiss, Robert; Townley, Helen

2017-03-01

Herein we have undertaken a systematic analysis of the effects of the fungal derivative ophiobolin A (OphA) on eight cancer cell lines from different tissue types. The LD50 for each cell line was determined and the change in cell size determined. Flow cytometric analysis and western blotting were used to assess the cell death markers for early apoptosis, late apoptosis and necrosis, and the involvement of the caspase signalling pathway. Alterations in calcium levels and reactive oxygen species were assessed due to their integral involvement in intracellular signalling. Subsequently, the endoplasmic reticulum (ER) and mitochondrial responses were investigated more closely. The extent of ER swelling, and the upregulation of proteins involved in the unfolded protein responses (UPR) were seen to vary according to cell line. The mitochondria were also shown to behave differently in response to the OphA in the different cell lines in terms of the change in membrane potential, the total area of mitochondria in the cell and the number of mitochondrial bifurcations. The data obtained in the present study indicate that the cancer cell lines tested are unable to successfully activate the ER stress/UPR responses, and that the mitochondria appear to be a central player in OphA-induced cancer cell death.

NITRIC OXIDE, MITOCHONDRIAL HYPERPOLARIZATION AND T-CELL ACTIVATION

PubMed Central

Nagy, Gyorgy; Koncz, Agnes; Fernandez, David; Perl, Andras

2007-01-01

T lymphocyte activation is associated with nitric oxide (NO) production that plays an essential role in multiple T cell functions. NO acts as a messenger, activating soluble guanyl cyclase and participating in the transduction signaling pathways involving cyclic GMP. NO modulates mitochondrial events that are involved in apoptosis and regulates mitochondrial membrane potential and mitochondrial biogenesis in many cell types, including lymphocytes. Mitochondrial hyperpolarization (MHP), an early and reversible event during both T lymphocyte activation and apoptosis, is regulated by NO. Here, we discuss recent evidence that NO-induced MHP represents a molecular switch in multiple T cell signaling pathways. Overproduction of NO in systemic lupus erythematosus (SLE) induces mitochondrial biogenesis and alters Ca2+ signaling. Thus, while NO plays a physiological role in lymphocyte cell signaling, its overproduction may disturb normal T cell function, contributing to the pathogenesis of autoimmunity. PMID:17462531

Effects of salinity on the transcriptome of growing maize leaf cells point at cell-age specificity in the involvement of the antioxidative response in cell growth restriction

PubMed Central

2013-01-01

Background Salinity inhibits growth and development of most plants. The response to salinity is complex and varies between plant organs and stages of development. It involves challenges of ion toxicities and deficiencies as well as osmotic and oxidative stresses. The range of functions affected by the stress is reflected in elaborate changes to the transcriptome. The mechanisms involved in the developmental-stage specificity of the inhibitory responses are not fully understood. The present study took advantage of the well characterized developmental progression that exists along the maize leaf, for identification of salinity induced, developmentally-associated changes to the transcriptome. Differential subtraction screening was conducted for cells of two developmental stages: from the center of the growth zone where the expansion rate is highest, and from older cells at a more distal location of the growing zone where the expansion rate is lower and the salinity restrictive effects are more pronounced. Real-Time PCR analysis was used for validation of the expression of selected genes. Results The salinity-induced changes demonstrated an age-related response of the growing tissue, with elevation of salinity-damages with increased age. Growth reduction, similar to the elevation of percentage dry matter (%DM), and Na and Cl concentrations were more pronounced in the older cells. The differential subtraction screening identified genes encoding to proteins involved in antioxidant defense, electron transfer and energy, structural proteins, transcription factors and photosynthesis proteins. Of special interest is the higher induced expression of genes involved in antioxidant protection in the young compared to older cells, which was accompanied by suppressed levels of reactive oxygen species (H2O2 and O2-). This was coupled with heightened expression in the older cells of genes that enhance cell-wall rigidity, which points at reduced potential for cell expansion. Conclusions The results demonstrate a cell-age specificity in the salinity response of growing cells, and point at involvement of the antioxidative response in cell growth restriction. Processes involved in reactive oxygen species (ROS) scavenging are more pronounced in the young cells, while the higher growth sensitivity of older cells is suggested to involve effects on cell-wall rigidity and lower protein protection. PMID:23324477

In vitro toxicological effects of estrogenic mycotoxins on human placental cells: Structure activity relationships

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

Prouillac, Caroline, E-mail: c.prouillac@vetagro-sup.fr; Koraichi, Farah; Videmann, Bernadette

2012-03-15

Zearalenone (ZEN) is a non-steroid estrogen mycotoxin produced by numerous strains of Fusarium which commonly contaminate cereals. After oral administration, ZEN is reduced via intestinal and hepatic metabolism to α- and β-zearalenol (αZEL and βZEL). These reduced metabolites possess estrogenic properties, αZEL showing the highest affinity for ERs. ZEN and reduced metabolites cause hormonal effects in animals, such as abnormalities in the development of the reproductive tract and mammary gland in female offspring, suggesting a fetal exposure to these contaminants. In our previous work, we have suggested the potential impact of ZEN on placental cells considering this organ as amore » potential target of xenobiotics. In this work, we first compared the in vitro effects of αZEL and βΖΕL on cell differentiation to their parental molecule on human trophoblast (BeWo cells). Secondly, we investigated their molecular mechanisms of action by investigating the expression of main differentiation biomarkers and the implication of nuclear receptor by docking prediction. Conversely to ZEN, reduced metabolites did not induce trophoblast differentiation. They also induced significant changes in ABC transporter expression by potential interaction with nuclear receptors (LXR, PXR, PR) that could modify the transport function of placental cells. Finally, the mechanism of ZEN differentiation induction seemed not to involve nuclear receptor commonly involved in the differentiation process (PPARγ). Our results demonstrated that in spite of structure similarities between ZEN, αZEL and βZEL, toxicological effects and toxicity mechanisms were significantly different for the three molecules. -- Highlights: ► ZEN and metabolites have differential effect on trophoblast differentiation. ► ZEN and metabolites have differential effect on ABC transporter expression. ► ZEN and metabolites effects involved nuclear receptors interaction.« less

Bioinformatics analysis for evaluation of the diagnostic potentialities of miR-19b, -125b and -205 as liquid biopsy markers of prostate cancer

NASA Astrophysics Data System (ADS)

Bryzgunova, O. E.; Lekchnov, E. A.; Zaripov, M. M.; Yurchenko, Yu. B.; Yarmoschuk, S. V.; Pashkovskaya, O. A.; Rykova, E. Yu.; Zheravin, A. A.; Laktionov, P. P.

2017-09-01

Presence of tumor-derived cell-free miRNA in biological fluids as well as simplicity and robustness of cell-free miRNA quantification makes them suitable markers for cancer diagnostics. Based on previously published data demonstrating diagnostic potentialities of miR-205 in blood and miR-19b as well as miR-125b in urine of prostate cancer patients, bioinformatics analysis was carried out to follow their involvement in prostate cancer development and select additional miRNA-markers for prostate cancer diagnostics. Studied miRNAs are involved in different signaling pathways and regulate a number of genes involved in cancer development. Five of their targets (CCND1, BRAF, CCNE1, CCNE2, RAF1), according to the STRING database, act as part of the same signaling pathway. RAF1 is regulated by miR-19b and miR-125b, and it was shown to be involved in prostate cancer development by DIANA and STRING databases. Thus, other microRNAs regulating RAF1 expression such as miR-16, -195, -497, and -7 (suggested by DIANA, TargetScan, MiRTarBase and miRDB databases) can potentially be regarded as prostate cancer markers.

Regulatory T cells in the actinic cheilitis.

PubMed

Gasparoto, Thaís Helena; de Souza Malaspina, Tatiana Salles; Damante, José Humberto; de Mello, Edgard Franco; Ikoma, Maura Rosane Valério; Garlet, Gustavo Pompermaier; Costa, Maria Renata Sales Nogueira; Cavassani, Karen Angélica; da Silva, João Santana; Campanelli, Ana Paula

2014-11-01

Actinic cheilitis (AC) is an oral potentially malignant lesion which is the counterpart of actinic keratosis of the skin and has potential to develop into squamous cell carcinoma. Regulatory T cells (Tregs) have a critical role in modulating the antitumor immune responses. The presence of regulatory T cells in potentially malignant lesions has not been described. We chose investigate the involvement of regulatory T cells in potentially malignant lesions. The frequency, phenotype, and activity of CD4+CD25+ T cells isolated from blood and lesion of AC patients were analyzed by flow cytometry. Cytokines were quantified by ELISA. Data were compared with samples from healthy subjects. The frequency and suppressor activity of circulating CD4+CD25+ T cells was similar in AC patients and control subjects. However, the frequencies of IL-10-positive Tregs were higher in AC patients, and these cells inhibited interferon-gamma (IFN-γ) and increased interleukin (IL)-10 productions in co-cultures. Furthermore, CD4+CD25+ T cells accumulate in AC lesions. Lesions-derived regulatory T cells suppressed lymphocyte proliferation and pro-inflammatory cytokine production. Moreover, high levels of IL-10 and transforming growth factor-β (TGF-β), and low IFN-γ were detected in the potentially malignant lesions. Therefore, our data show that Tregs accumulate in AC lesions, and these cells could be suppressing immune responses in a potentially malignant microenvironment. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

TRPM channels phosphorylation as a potential bridge between old signals and novel regulatory mechanisms of insulin secretion.

PubMed

Diaz-Garcia, Carlos Manlio; Sanchez-Soto, Carmen; Hiriart, Marcia

2013-03-01

Transient receptor potential channels, especially the members of the melastatin family (TRPM), participate in insulin secretion. Some of them are substrates for protein kinases, which are involved in several neurotransmitter, incretin and hormonal signaling cascades in β cells. The functional relationships between protein kinases and TRPM channels in systems of heterologous expression and native tissues rise issues about novel regulation pathways of pancreatic β-cell excitability. The aim of the present work is to review the evidences about phosphorylation of TRPM channels in β cells and to discuss the perspectives on insulin secretion.

Mesenchymal stem cells in cardiac regeneration: a detailed progress report of the last 6 years (2010-2015).

PubMed

Singh, Aastha; Singh, Abhishek; Sen, Dwaipayan

2016-06-04

Mesenchymal stem cells have been used for cardiovascular regenerative therapy for decades. These cells have been established as one of the potential therapeutic agents, following several tests in animal models and clinical trials. In the process, various sources of mesenchymal stem cells have been identified which help in cardiac regeneration by either revitalizing the cardiac stem cells or revascularizing the arteries and veins of the heart. Although mesenchymal cell therapy has achieved considerable admiration, some challenges still remain that need to be overcome in order to establish it as a successful technique. This in-depth review is an attempt to summarize the major sources of mesenchymal stem cells involved in myocardial regeneration, the significant mechanisms involved in the process with a focus on studies (human and animal) conducted in the last 6 years and the challenges that remain to be addressed.

Establishing a stem cell culture laboratory for clinical trials

PubMed Central

Sekiya, Elíseo Joji; Forte, Andresa; Kühn, Telma Ingrid Borges de Bellis; Janz, Felipe; Bydlowski, Sérgio Paulo; Alves, Adelson

2012-01-01

Adult stem/progenitor cells are found in different human tissues. An in vitro cell culture is needed for their isolation or for their expansion when they are not available in a sufficient quantity to regenerate damaged organs and tissues. The level of complexity of these new technologies requires adequate facilities, qualified personnel with experience in cell culture techniques, assessment of quality and clear protocols for cell production. The rules for the implementation of cell therapy centers involve national and international standards of good manufacturing practices. However, such standards are not uniform, reflecting the diversity of technical and scientific development. Here standards from the United States, the European Union and Brazil are analyzed. Moreover, practical solutions encountered for the implementation of a cell therapy center appropriate for the preparation and supply of cultured cells for clinical studies are described. Development stages involved the planning and preparation of the project, the construction of the facility, standardization of laboratory procedures and development of systems to prevent cross contamination. Combining the theoretical knowledge of research centers involved in the study of cells with the practical experience of blood therapy services that manage structures for cell transplantation is presented as the best potential for synergy to meet the demands to implement cell therapy centers. PMID:23049427

Acetate alters expression of genes involved in beige adipogenesis in 3T3-L1 cells and obese KK-Ay mice

PubMed Central

Hanatani, Satoko; Motoshima, Hiroyuki; Takaki, Yuki; Kawasaki, Shuji; Igata, Motoyuki; Matsumura, Takeshi; Kondo, Tatsuya; Senokuchi, Takafumi; Ishii, Norio; Kawashima, Junji; Kukidome, Daisuke; Shimoda, Seiya; Nishikawa, Takeshi; Araki, Eiichi

2016-01-01

The induction of beige adipogenesis within white adipose tissue, known as “browning”, has received attention as a novel potential anti-obesity strategy. The expression of some characteristic genes including PR domain containing 16 is induced during the browning process. Although acetate has been reported to suppress weight gain in both rodents and humans, its potential effects on beige adipogenesis in white adipose tissue have not been fully characterized. We examined the effects of acetate treatment on 3T3-L1 cells and in obese diabetic KK-Ay mice. The mRNA expression levels of genes involved in beige adipocyte differentiation and genes selectively expressed in beige adipocytes were significantly elevated in both 3T3-L1 cells incubated with 1.0 mM acetate and the visceral white adipose tissue from mice treated with 0.6% acetate for 16 weeks. In KK-Ay mice, acetate reduced the food efficiency ratio and increased the whole-body oxygen consumption rate. Additionally, reduction of adipocyte size and uncoupling protein 1-positive adipocytes and interstitial areas with multilocular adipocytes appeared in the visceral white adipose tissue of acetate-treated mice, suggesting that acetate induced initial changes of “browning”. In conclusion, acetate alters the expression of genes involved in beige adipogenesis and might represent a potential therapeutic agent to combat obesity. PMID:27895388

Potentiation of lead-induced cell death in PC12 cells by glutamate: Protection by N-acetylcysteine amide (NACA), a novel thiol antioxidant

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

Penugonda, Suman; Mare, Suneetha; Lutz, P.

2006-10-15

Oxidative stress has been implicated as an important factor in many neurological diseases. Oxidative toxicity in a number of these conditions is induced by excessive glutamate release and subsequent glutamatergic neuronal stimulation. This, in turn, causes increased generation of reactive oxygen species (ROS), oxidative stress, excitotoxicity, and neuronal damage. Recent studies indicate that the glutamatergic neurotransmitter system is involved in lead-induced neurotoxicity. Therefore, this study aimed to (1) investigate the potential effects of glutamate on lead-induced PC12 cell death and (2) elucidate whether the novel thiol antioxidant N-acetylcysteine amide (NACA) had any protective abilities against such cytotoxicity. Our results suggestmore » that glutamate (1 mM) potentiates lead-induced cytotoxicity by increased generation of ROS, decreased proliferation (MTS), decreased glutathione (GSH) levels, and depletion of cellular adenosine-triphosphate (ATP). Consistent with its ability to decrease ATP levels and induce cell death, lead also increased caspase-3 activity, an effect potentiated by glutamate. Exposure to glutamate and lead elevated the cellular malondialdehyde (MDA) levels and phospholipase-A{sub 2} (PLA{sub 2}) activity and diminished the glutamine synthetase (GS) activity. NACA protected PC12 cells from the cytotoxic effects of glutamate plus lead, as evaluated by MTS assay. NACA reduced the decrease in the cellular ATP levels and restored the intracellular GSH levels. The increased levels of ROS and MDA in glutamate-lead treated cells were significantly decreased by NACA. In conclusion, our data showed that glutamate potentiated the effects of lead-induced PC12 cell death by a mechanism involving mitochondrial dysfunction (ATP depletion) and oxidative stress. NACA had a protective role against the combined toxic effects of glutamate and lead by inhibiting lipid peroxidation and scavenging ROS, thus preserving intracellular GSH.« less

Engineering a 3D microfluidic culture platform for tumor-treating field application

NASA Astrophysics Data System (ADS)

Pavesi, Andrea; Adriani, Giulia; Tay, Andy; Warkiani, Majid Ebrahimi; Yeap, Wei Hseun; Wong, Siew Cheng; Kamm, Roger D.

2016-05-01

The limitations of current cancer therapies highlight the urgent need for a more effective therapeutic strategy. One promising approach uses an alternating electric field; however, the mechanisms involved in the disruption of the cancer cell cycle as well as the potential adverse effects on non-cancerous cells must be clarified. In this study, we present a novel microfluidic device with embedded electrodes that enables the application of an alternating electric field therapy to cancer cells in a 3D extracellular matrix. To demonstrate the potential of our system to aid in designing and testing new therapeutic approaches, cancer cells and cancer cell aggregates were cultured individually or co-cultured with endothelial cells. The metastatic potential of the cancer cells was reduced after electric field treatment. Moreover, the proliferation rate of the treated cancer cells was lower compared with that of the untreated cells, whereas the morphologies and proliferative capacities of the endothelial cells were not significantly affected. These results demonstrate that our novel system can be used to rapidly screen the effect of an alternating electric field on cancer and normal cells within an in vivo-like microenvironment with the potential to optimize treatment protocols and evaluate synergies between tumor-treating field treatment and chemotherapy.

Rectification of Acetylcholine-Elicited Currents in PC12 Pheochromocytoma Cells

NASA Astrophysics Data System (ADS)

Ifune, C. K.; Steinbach, J. H.

1990-06-01

The current-voltage (I-V) relationship for acetylcholine-elicited currents in the rat pheochromocytoma cell line PC12 is nonlinear. Two voltage-dependent processes that could account for the whole-cell current rectification were examined, receptor channel gating and single receptor channel permeation. We found that both factors are involved in the rectification of the whole-cell currents. The voltage dependence of channel gating determines the shape of the I-V curve at negative potentials. The single-channel I-V relationship is inwardly rectifying and largely responsible for the characteristic shape of the whole-cell I-V curve at positive potentials. The rectification of the single-channel currents is produced by the voltage-dependent block of outward currents by intracellular Mg2+ ions.

Proteome Analysis for Downstream Targets of Oncogenic KRAS - the Potential Participation of CLIC4 in Carcinogenesis in the Lung

PubMed Central

Okudela, Koji; Katayama, Akira; Woo, Tetsukan; Mitsui, Hideaki; Suzuki, Takehisa; Tateishi, Yoko; Umeda, Shigeaki; Tajiri, Michihiko; Masuda, Munetaka; Nagahara, Noriyuki; Kitamura, Hitoshi; Ohashi, Kenichi

2014-01-01

This study investigated the proteome modulated by oncogenic KRAS in immortalized airway epithelial cells. Chloride intracellular channel protein 4 (CLIC4), S100 proteins (S100A2 and S100A11), tropomyosin 2, cathepsin L1, integrinsα3, eukaryotic elongation factor 1, vimentin, and others were discriminated. We here focused on CLIC4 to investigate its potential involvement in carcinogenesis in the lung because previous studies suggested that some chloride channels and chloride channel regulators could function as tumor suppressors. CILC4 protein levels were reduced in some lung cancer cell lines. The restoration of CLIC4 in lung cancer cell lines in which CLIC4 expression was reduced attenuated their growth activity. The immunohistochemical expression of the CLIC4 protein was weaker in primary lung cancer cells than in non-tumorous airway epithelial cells and was occasionally undetectable in some tumors. CLIC4 protein levels were significantly lower in a subtype of mucinous ADC than in others, and were also significantly lower in KRAS-mutated ADC than in EGFR-mutated ADC. These results suggest that the alteration in CLIC4 could be involved in restrictedly the development of a specific fraction of lung adenocarcinomas. The potential benefit of the proteome modulated by oncogenic KRAS to lung cancer research has been demonstrated. PMID:24503901

Possible Muscle Repair in the Human Cardiovascular System.

PubMed

Sommese, Linda; Zullo, Alberto; Schiano, Concetta; Mancini, Francesco P; Napoli, Claudio

2017-04-01

The regenerative potential of tissues and organs could promote survival, extended lifespan and healthy life in multicellular organisms. Niches of adult stemness are widely distributed and lead to the anatomical and functional regeneration of the damaged organ. Conversely, muscular regeneration in mammals, and humans in particular, is very limited and not a single piece of muscle can fully regrow after a severe injury. Therefore, muscle repair after myocardial infarction is still a chimera. Recently, it has been recognized that epigenetics could play a role in tissue regrowth since it guarantees the maintenance of cellular identity in differentiated cells and, therefore, the stability of organs and tissues. The removal of these locks can shift a specific cell identity back to the stem-like one. Given the gradual loss of tissue renewal potential in the course of evolution, in the last few years many different attempts to retrieve such potential by means of cell therapy approaches have been performed in experimental models. Here we review pathways and mechanisms involved in the in vivo repair of cardiovascular muscle tissues in humans. Moreover, we address the ongoing research on mammalian cardiac muscle repair based on adult stem cell transplantation and pro-regenerative factor delivery. This latter issue, involving genetic manipulations of adult cells, paves the way for developing possible therapeutic strategies in the field of cardiovascular muscle repair.

Endoplasmic reticulum calcium release potentiates the ER stress and cell death caused by an oxidative stress in MCF-7 cells.

PubMed

Dejeans, Nicolas; Tajeddine, Nicolas; Beck, Raphaël; Verrax, Julien; Taper, Henryk; Gailly, Philippe; Calderon, Pedro Buc

2010-05-01

Increase in cytosolic calcium concentration ([Ca2+](c)), release of endoplasmic reticulum (ER) calcium ([Ca2+](er)) and ER stress have been proposed to be involved in oxidative toxicity. Nevertheless, their relative involvements in the processes leading to cell death are not well defined. In this study, we investigated whether oxidative stress generated during ascorbate-driven menadione redox cycling (Asc/Men) could trigger these three events, and, if so, whether they contributed to Asc/Men cytoxicity in MCF-7 cells. Using microspectrofluorimetry, we demonstrated that Asc/Men-generated oxidative stress was associated with a slow and moderate increase in [Ca2+](c), largely preceding permeation of propidium iodide, and thus cell death. Asc/Men treatment was shown to partially deplete ER calcium stores after 90 min (decrease by 45% compared to control). This event was associated with ER stress activation, as shown by analysis of eIF2 phosphorylation and expression of the molecular chaperone GRP94. Thapsigargin (TG) was then used to study the effect of complete [Ca2+](er) emptying during the oxidative stress generated by Asc/Men. Surprisingly, the combination of TG and Asc/Men increased ER stress to a level considerably higher than that observed for either treatment alone, suggesting that [Ca2+](er) release alone is not sufficient to explain ER stress activation during oxidative stress. Finally, TG-mediated [Ca2+](er) release largely potentiated ER stress, DNA fragmentation and cell death caused by Asc/Men, supporting a role of ER stress in the process of Asc/Men cytotoxicity. Taken together, our results highlight the involvement of ER stress and [Ca2+](er) decrease in the process of oxidative stress-induced cell death in MCF-7 cells. 2009 Elsevier Inc. All rights reserved.

Arsenic-induced malignant transformation of human keratinocytes: Involvement of Nrf2

PubMed Central

Pi, Jingbo; Diwan, Bhalchandra A.; Sun, Yang; Liu, Jie; Qu, Wei; He, Yuying; Styblo, Miroslav; Waalkes, Michael P.

2009-01-01

Arsenic is a well-known human skin carcinogen but the underlying mechanisms of carcinogenesis are unclear. Transcription factor Nrf2-mediated antioxidant response represents a critical cellular defense mechanism, and emerging data suggest that constitutive activation of Nrf2 contributes to malignant phenotype. In the present study when an immortalized, non-tumorigenic human keratinocyte cell line (HaCaT) was continuously exposed to environmentally relevant level of inorganic arsenite (100 nM) for 28 weeks, malignant transformation occurred as evidenced by the formation of highly aggressive squamous cell carcinoma after inoculation into nude mice. To investigate the mechanisms involved, a broad array of biomarkers for transformation were assessed in these arsenic-transformed cells (termed As-TM). In addition to increased secretion of matrix metalloproteinase-9 (MMP-9), a set of markers for squamous differentiation and skin keratinization, including keratin-1, keratin-10, involucrin, and loricrin, were significantly elevated in As-TM cells. Furthermore, As-TM cells showed increased intracellular glutathione, elevated expression of Nrf2 and its target genes, as well as generalized apoptotic resistance. In contrast to increased basal Nrf2 activity in As-TM cells, a diminished Nrf2-mediated antioxidant response induced by acute exposure to high dose of arsenite or tert-butyl hydroxyquinone occurred. The findings that multiple biomarkers for malignant transformation observed in As-TM cells, including MMP-9 and cytokeratins, are potentially regulated by Nrf2 suggest constitutive Nrf2 activation may be involved in arsenic carcinogenesis of skin. The weakened Nrf2 activation in response to oxidative stressors observed in As-TM cells, coupled with acquired apoptotic resistance, would potentially have increased the likelihood of transmittable oxidative DNA damage and fixation of mutational/DNA damage events. PMID:18572023

KIT D816V-mutated bone marrow mesenchymal stem cells in indolent systemic mastocytosis are associated with disease progression.

PubMed

Garcia-Montero, Andres C; Jara-Acevedo, Maria; Alvarez-Twose, Ivan; Teodosio, Cristina; Sanchez-Muñoz, Laura; Muñiz, Carmen; Muñoz-Gonzalez, Javier I; Mayado, Andrea; Matito, Almudena; Caldas, Carolina; Morgado, Jose M; Escribano, Luis; Orfao, Alberto

2016-02-11

Multilineage involvement of bone marrow (BM) hematopoiesis by the somatic KIT D816V mutation is present in a subset of adult indolent systemic mastocytosis (ISM) patients in association with a poorer prognosis. Here, we investigated the potential involvement of BM mesenchymal stem cells (MSCs) from ISM patients by the KIT D816V mutation and its potential impact on disease progression and outcome. This mutation was investigated in highly purified BM MSCs and other BM cell populations from 83 ISM patients followed for a median of 116 months. KIT D816V-mutated MSCs were detected in 22 of 83 cases. All MSC-mutated patients had multilineage KIT mutation (100% vs 30%, P = .0001) and they more frequently showed involvement of lymphoid plus myeloid BM cells (59% vs 22%; P = .03) and a polyclonal pattern of inactivation of the X-chromosome of KIT-mutated BM mast cells (64% vs 0%; P = .01) vs other multilineage ISM cases. Moreover, presence of KIT-mutated MSCs was associated with more advanced disease features, a greater rate of disease progression (50% vs 17%; P = .04), and a shorter progression-free survival (P ≤ .003). Overall, these results support the notion that ISM patients with mutated MSCs may have acquired the KIT mutation in a common pluripotent progenitor cell, prior to differentiation into MSCs and hematopoietic precursor cells, before the X-chromosome inactivation process occurs. From a clinical point of view, acquisition of the KIT mutation in an earlier BM precursor cell confers a significantly greater risk for disease progression and a poorer outcome. © 2016 by The American Society of Hematology.

The Potential Therapeutic Agent Mepacrine Protects Caco-2 Cells against Clostridium perfringens Enterotoxin Action.

PubMed

Freedman, John C; Hendricks, Matthew R; McClane, Bruce A

2017-01-01

Clostridium perfringens enterotoxin (CPE) causes the diarrhea associated with a common bacterial food poisoning and many antibiotic-associated diarrhea cases. The severity of some CPE-mediated disease cases warrants the development of potential therapeutics. A previous study showed that the presence of mepacrine inhibited CPE-induced electrophysiology effects in artificial lipid bilayers lacking CPE receptors. However, that study did not assess whether mepacrine inactivates CPE or, instead, inhibits a step in CPE action. Furthermore, CPE action in host cells is complex, involving the toxin binding to receptors, receptor-bound CPE oligomerizing into a prepore on the membrane surface, and β-hairpins in the CPE prepore inserting into the membrane to form a pore that induces cell death. Therefore, the current study evaluated the ability of mepacrine to protect cells from CPE. This drug was found to reduce CPE-induced cytotoxicity in Caco-2 cells. This protection did not involve mepacrine inactivation of CPE, indicating that mepacrine affects one or more steps in CPE action. Western blotting then demonstrated that mepacrine decreases CPE pore levels in Caco-2 cells. This mepacrine-induced reduction in CPE pore levels did not involve CPE binding inhibition but rather an increase in CPE monomer dissociation due to mepacrine interactions with Caco-2 membranes. In addition, mepacrine was also shown to inhibit CPE pores when already present in Caco-2 cells. These in vitro studies, which identified two mepacrine-sensitive steps in CPE-induced cytotoxicity, add support to further testing of the therapeutic potential of mepacrine against CPE-mediated disease. IMPORTANCE Clostridium perfringens enterotoxin (CPE) causes the gastrointestinal (GI) symptoms of a common bacterial food poisoning and several nonfoodborne human GI diseases. A previous study showed that, via an undetermined mechanism, the presence of mepacrine blocks CPE-induced electrophysiologic activity in artificial membranes. The current study now demonstrates that mepacrine also inhibits CPE-induced cytotoxicity in human enterocyte-like Caco-2 cells and that mepacrine does not directly inactivate CPE. Instead, this drug reduces both CPE pore formation and CPE pore activity in Caco-2 cells. These results suggest mepacrine as a therapeutic candidate for treating CPE-mediated GI diseases.

Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential.

PubMed

Song, Kai; Song, Yong; Zhao, Xiao-Ping; Shen, Hui; Wang, Meng; Yan, Ting-Lin; Liu, Ke; Shang, Zheng-Jun

2014-10-15

Most previous studies have linked cancer-macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed that SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression. Copyright © 2014 Elsevier Inc. All rights reserved.

The interplay between genetic and bioelectrical signaling permits a spatial regionalisation of membrane potentials in model multicellular ensembles

PubMed Central

Cervera, Javier; Meseguer, Salvador; Mafe, Salvador

2016-01-01

The single cell-centred approach emphasises ion channels as specific proteins that determine individual properties, disregarding their contribution to multicellular outcomes. We simulate the interplay between genetic and bioelectrical signals in non-excitable cells from the local single-cell level to the long range multicellular ensemble. The single-cell genetic regulation is based on mean-field kinetic equations involving the mRNA and protein concentrations. The transcription rate factor is assumed to depend on the absolute value of the cell potential, which is dictated by the voltage-gated cell ion channels and the intercellular gap junctions. The interplay between genetic and electrical signals may allow translating single-cell states into multicellular states which provide spatio-temporal information. The model results have clear implications for biological processes: (i) bioelectric signals can override slightly different genetic pre-patterns; (ii) ensembles of cells initially at the same potential can undergo an electrical regionalisation because of persistent genetic differences between adjacent spatial regions; and (iii) shifts in the normal cell electrical balance could trigger significant changes in the genetic regulation. PMID:27731412

The interplay between genetic and bioelectrical signaling permits a spatial regionalisation of membrane potentials in model multicellular ensembles.

PubMed

Cervera, Javier; Meseguer, Salvador; Mafe, Salvador

2016-10-12

The single cell-centred approach emphasises ion channels as specific proteins that determine individual properties, disregarding their contribution to multicellular outcomes. We simulate the interplay between genetic and bioelectrical signals in non-excitable cells from the local single-cell level to the long range multicellular ensemble. The single-cell genetic regulation is based on mean-field kinetic equations involving the mRNA and protein concentrations. The transcription rate factor is assumed to depend on the absolute value of the cell potential, which is dictated by the voltage-gated cell ion channels and the intercellular gap junctions. The interplay between genetic and electrical signals may allow translating single-cell states into multicellular states which provide spatio-temporal information. The model results have clear implications for biological processes: (i) bioelectric signals can override slightly different genetic pre-patterns; (ii) ensembles of cells initially at the same potential can undergo an electrical regionalisation because of persistent genetic differences between adjacent spatial regions; and (iii) shifts in the normal cell electrical balance could trigger significant changes in the genetic regulation.

Crataegus azarolus Leaves Induce Antiproliferative Activity, Cell Cycle Arrest, and Apoptosis in Human HT-29 and HCT-116 Colorectal Cancer Cells.

PubMed

Mustapha, Nadia; Pinon, Aline; Limami, Youness; Simon, Alain; Ghedira, Kamel; Hennebelle, Thierry; Chekir-Ghedira, Leila

2016-05-01

Limited success has been achieved in extending the survival of patients with metastatic colorectal cancer (CRC). There is a strong need for novel agents in the treatment and prevention of CRC. Therefore, in the present study we evaluated the antiproliferative and pro-apoptotic potential of Crataegus azarolus ethyl acetate extract in HCT-116 and HT-29 human colorectal cancer cell lines. Moreover, we attempted to investigate the signaling pathways that should be involved in its cytotoxic effect. The Crataegus azarolus ethyl acetate extract-induced growth inhibitory effect was associated with DNA fragmentation, sub-G1 peak, loss of mitochondrial potential, and poly (ADP-ribose) polymerase (PARP) cleavage. In addition, ethyl acetate extract of Crataegus azarolus induced the cleavage of caspase-8. It has no effect on steady-state levels of total Bcl-2 protein. Whereas Bax levels decreased significantly in a dose-dependent manner in both tested cell lines. Taken together, these findings confirm the involvement of the extrinsic pathway of apoptosis. The apoptotic cell death induced by ethyl acetate extract of Crataegus azarolus was accompanied by an enhancement of the p21 expression but not through p53 activation in human colorectal cancer cells. The above-mentioned data provide insight into the molecular mechanisms of Crataegus azarolus ethyl acetate extract-induced apoptosis in CRC. Therefore, this compound should be a potential anticancer agent for the treatment of CRC. © 2015 Wiley Periodicals, Inc.

Broad Ligament Perivascular Epithelioid Cell Tumor (PEComa) of Uncertain Malignant Potential.

PubMed

Mathew, Mary; Nayal, Bhavna; Rao, Lakshmi; Nagel, Bhawna

2016-01-01

PEComas are uncommon mesenchymal tumors often involving the pelvic organs. They have an unpredictable behavior. Accurate diagnosis and long-term follow-up is therefore essential in these patients. We report this case of PEComa of uncertain malignant potential in an unusual location with excellent prognosis.

Genome-wide effects of MELK-inhibitor in triple-negative breast cancer cells indicate context-dependent response with p53 as a key determinant

PubMed Central

Simon, Marisa; Mesmar, Fahmi; Helguero, Luisa

2017-01-01

Triple-negative breast cancer (TNBC) is an aggressive, highly recurrent breast cancer subtype, affecting approximately one-fifth of all breast cancer patients. Subpopulations of treatment-resistant cancer stem cells within the tumors are considered to contribute to disease recurrence. A potential druggable target for such cells is the maternal embryonic leucine-zipper kinase (MELK). MELK expression is upregulated in mammary stem cells and in undifferentiated cancers, where it correlates with poor prognosis and potentially mediates treatment resistance. Several MELK inhibitors have been developed, of which one, OTSSP167, is currently in clinical trials. In order to better understand how MELK and its inhibition influence TNBC, we verified its anti-proliferative and apoptotic effects in claudin-low TNBC cell lines MDA-MB-231 and SUM-159 using MTS assays and/or trypan blue viability assays together with analysis of PARP cleavage. Then, using microarrays, we explored which genes were affected by OTSSP167. We demonstrate that different sets of genes are regulated in MDA-MB-231 and SUM-159, but in both cell lines genes involved in cell cycle, mitosis and protein metabolism and folding were regulated. We identified p53 (TP53) as a potential upstream regulator of the regulated genes. Using western blot we found that OTSSP167 downregulates mutant p53 in all tested TNBC cell lines (MDA-MB-231, SUM-159, and BT-549), but upregulates wild-type p53 in the luminal A subtype MCF-7 cell line. We propose that OTSSP167 might have context-dependent or off-target effects, but that one consistent mechanism of action could involve the destabilization of mutant p53. PMID:28235006

Chemopreventive potential of in vitro fermented nuts in LT97 colon adenoma and primary epithelial colon cells.

PubMed

Schlörmann, Wiebke; Lamberty, Julia; Lorkowski, Stefan; Ludwig, Diana; Mothes, Henning; Saupe, Christian; Glei, Michael

2017-05-01

Due to their beneficial nutritional profile the consumption of nuts contributes to a healthy diet and might reduce colon cancer risk. To get closer insights into potential mechanisms, the chemopreventive potential of different in vitro fermented nut varieties regarding the modulation of genes involved in detoxification (CAT, SOD2, GSTP1, GPx1) and cell cycle (p21, cyclin D2) as well as proliferation and apoptosis was examined in LT97 colon adenoma and primary epithelial colon cells. Fermentation supernatants (FS) of nuts significantly induced mRNA expression of CAT (up to 4.0-fold), SOD2 (up to 2.5-fold), and GSTP1 (up to 2.3-fold), while GPx1 expression was significantly reduced by all nut FS (0.8 fold on average). Levels of p21 mRNA were significantly enhanced (up to 2.6-fold), whereas all nut FS significantly decreased cyclin D2 expression (0.4-fold on average). In primary epithelial cells, expression of CAT (up to 3.5-fold), GSTP1 (up to 3.0-fold), and GPx1 (up to 3.9-fold) was increased, whereas p21 and cyclin D2 levels were not influenced. Nut FS significantly inhibited growth of LT97 cells and increased levels of early apoptotic cells (8.4% on average) and caspase 3 activity (4.6-fold on average), whereas caspase 3 activity was not modulated in primary colon cells. The differential modulation of genes involved in detoxification and cell cycle together with an inhibition of proliferation and induction of apoptosis in adenoma cells might contribute to chemopreventive effects of nuts regarding colon cancer. © 2017 Wiley Periodicals, Inc.

Hematopoietic stem cell transplantation in Niemann-Pick disease type B monitored by chitotriosidase activity.

PubMed

Quarello, Paola; Spada, Marco; Porta, Francesco; Vassallo, Elena; Timeus, Fabio; Fagioli, Franca

2018-02-01

Here, we report a patient with Niemann-Pick disease type B, with early severe onset of disease and pulmonary involvement, treated with hematopoietic stem cell transplant (HSCT) from a bone marrow matched unrelated donor. We confirm that HSCT is feasible and potentially beneficial for patients with severe phenotype. Noteworthy, we discussed the potential usefulness of the activity of peripheral chitotriosidase for the longitudinal evaluation of HSCT success and effectiveness. © 2017 Wiley Periodicals, Inc.

Mechanistic Exploration of Cancer Stem Cell Marker Voltage-Dependent Calcium Channel α2δ1 Subunit-mediated Chemotherapy Resistance in Small-Cell Lung Cancer.

PubMed

Yu, Jiangyong; Wang, Shuhang; Zhao, Wei; Duan, Jianchun; Wang, Zhijie; Chen, Hanxiao; Tian, Yanhua; Wang, Di; Zhao, Jun; An, Tongtong; Bai, Hua; Wu, Meina; Wang, Jie

2018-05-01

Purpose: Chemoresistance in small-cell lung cancer (SCLC) is reportedly attributed to the existence of resistant cancer stem cells (CSC). Studies involving CSC-specific markers and related mechanisms in SCLC remain limited. This study explored the role of the voltage-dependent calcium channel α2δ1 subunit as a CSC marker in chemoresistance of SCLC, and explored the potential mechanisms of α2δ1-mediated chemoresistance and strategies of overcoming the resistance. Experimental Design: α2δ1-positive cells were identified and isolated from SCLC cell lines and patient-derived xenograft (PDX) models, and CSC-like properties were subsequently verified. Transcriptome sequencing and Western blotting were carried out to identify pathways involved in α2δ1-mediated chemoresistance in SCLC. In addition, possible interventions to overcome α2δ1-mediated chemoresistance were examined. Results: Different proportions of α2δ1 + cells were identified in SCLC cell lines and PDX models. α2δ1 + cells exhibited CSC-like properties (self-renewal, tumorigenic, differentiation potential, and high expression of genes related to CSCs and drug resistance). Chemotherapy induced the enrichment of α2δ1 + cells instead of CD133 + cells in PDXs, and an increased proportion of α2δ1 + cells corresponded to increased chemoresistance. Activation and overexpression of ERK in the α2δ1-positive H1048 cell line was identified at the protein level. mAb 1B50-1 was observed to improve the efficacy of chemotherapy and delay relapse as maintenance therapy in PDX models. Conclusions: SCLC cells expressing α2δ1 demonstrated CSC-like properties, and may contribute to chemoresistance. ERK may play a key role in α2δ1-mediated chemoresistance. mAb 1B50-1 may serve as a potential anti-SCLC drug. Clin Cancer Res; 24(9); 2148-58. ©2018 AACR . ©2018 American Association for Cancer Research.

Rice Bran Extract Inhibits TMEM16A-Involved Activity in the Neonatal Rat Cochlea.

PubMed

Sharm, Kushal; Sung, Jiwon; Kim, Hyun-Jung; Oak, Min-Ho; Yi, Eunyoung

2017-04-01

TMEM16A is a Ca²⁺-activated Cl⁻ channel found in secretory glands, GI and respiratory tracts, and sensory organs, playing a major physiological role in fluid secretion, autonomous GI motility, and sensory transduction. In addition, overexpression of TMEM16A has been associated with cancer cell proliferation and invasion. Suppression of upregulated TMEM16A has been proposed as an effective anti-cancer strategy. While searching for a potential TMEM16A inhibitor, components of rice bran attracted our attention due to their anti-cancer potential in colon cancer cells, a type of cells known to overexpressing TMEM16A. Here, it was tested whether rice bran extract exhibits anti-TMEM16A activity. Rice bran extract was tested in the neonatal rat cochlear tissues where TMEM16A-involved spontaneous activity is generated as a part of normal development of the auditory pathway. Rice bran extract readily inhibited the TMEM16A-involved activity in the cochlear tissues and the effect was reversible upon washout. Taken together, rice bran extract appears to contain a putative TMEM16A inhibitor and the rice byproduct might serve as a source of a new anti-cancer agent.

The potential of mesenchymal stromal cells as a novel cellular therapy for multiple sclerosis

PubMed Central

Auletta, Jeffery J; Bartholomew, Amelia M; Maziarz, Richard T; Deans, Robert J; Miller, Robert H; Lazarus, Hillard M; Cohen, Jeffrey A

2012-01-01

Multiple sclerosis (MS) is an inflammatory neurodegenerative disease of the CNS for which only partially effective therapies exist. Intense research defining the underlying immune pathophysiology is advancing both the understanding of MS as well as revealing potential targets for disease intervention. Mesenchymal stromal cell (MSC) therapy has the potential to modulate aberrant immune responses causing demyelination and axonal injury associated with MS, as well as to repair and restore damaged CNS tissue and cells. This article reviews the pathophysiology underlying MS, as well as providing a cutting-edge perspective into the field of MSC therapy based upon the experience of authors intrinsically involved in MS and MSC basic and translational science research. PMID:22642335

Voltage-Dependent Anion Channel 1 As an Emerging Drug Target for Novel Anti-Cancer Therapeutics

PubMed Central

Shoshan-Barmatz, Varda; Krelin, Yakov; Shteinfer-Kuzmine, Anna; Arif, Tasleem

2017-01-01

Cancer cells share several properties, high proliferation potential, reprogramed metabolism, and resistance to apoptotic cues. Acquiring these hallmarks involves changes in key oncogenes and non-oncogenes essential for cancer cell survival and prosperity, and is accompanied by the increased energy requirements of proliferating cells. Mitochondria occupy a central position in cell life and death with mitochondrial bioenergetics, biosynthesis, and signaling are critical for tumorigenesis. Voltage-dependent anion channel 1 (VDAC1) is situated in the outer mitochondrial membrane (OMM) and serving as a mitochondrial gatekeeper. VDAC1 allowing the transfer of metabolites, fatty acid ions, Ca2+, reactive oxygen species, and cholesterol across the OMM and is a key player in mitochondrial-mediate apoptosis. Moreover, VDAC1 serves as a hub protein, interacting with diverse sets of proteins from the cytosol, endoplasmic reticulum, and mitochondria that together regulate metabolic and signaling pathways. The observation that VDAC1 is over-expressed in many cancers suggests that the protein may play a pivotal role in cancer cell survival. However, VDAC1 is also important in mitochondria-mediated apoptosis, mediating release of apoptotic proteins and interacting with anti-apoptotic proteins, such as B-cell lymphoma 2 (Bcl-2), Bcl-xL, and hexokinase (HK), which are also highly expressed in many cancers. Strategically located in a “bottleneck” position, controlling metabolic homeostasis and apoptosis, VDAC1 thus represents an emerging target for anti-cancer drugs. This review presents an overview on the multi-functional mitochondrial protein VDAC1 performing several functions and interacting with distinct sets of partners to regulate both cell life and death, and highlights the importance of the protein for cancer cell survival. We address recent results related to the mechanisms of VDAC1-mediated apoptosis and the potential of associated proteins to modulate of VDAC1 activity, with the aim of developing VDAC1-based approaches. The first strategy involves modification of cell metabolism using VDAC1-specific small interfering RNA leading to inhibition of cancer cell and tumor growth and reversed oncogenic properties. The second strategy involves activation of cancer cell death using VDAC1-based peptides that prevent cell death induction by anti-apoptotic proteins. Finally, we discuss the potential therapeutic benefits of treatments and drugs leading to enhanced VDAC1 expression or targeting VDAC1 to induce apoptosis. PMID:28824871

DUBbing Cancer: Deubiquitylating Enzymes Involved in Epigenetics, DNA Damage and the Cell Cycle As Therapeutic Targets.

PubMed

Pinto-Fernandez, Adan; Kessler, Benedikt M

2016-01-01

Controlling cell proliferation is one of the hallmarks of cancer. A number of critical checkpoints ascertain progression through the different stages of the cell cycle, which can be aborted when perturbed, for instance by errors in DNA replication and repair. These molecular checkpoints are regulated by a number of proteins that need to be present at the right time and quantity. The ubiquitin system has emerged as a central player controlling the fate and function of such molecules such as cyclins, oncogenes and components of the DNA repair machinery. In particular, proteases that cleave ubiquitin chains, referred to as deubiquitylating enzymes (DUBs), have attracted recent attention due to their accessibility to modulation by small molecules. In this review, we describe recent evidence of the critical role of DUBs in aspects of cell cycle checkpoint control, associated DNA repair mechanisms and regulation of transcription, representing pathways altered in cancer. Therefore, DUBs involved in these processes emerge as potentially critical targets for the treatment of not only hematological, but potentially also solid tumors.

DUBbing Cancer: Deubiquitylating Enzymes Involved in Epigenetics, DNA Damage and the Cell Cycle As Therapeutic Targets

PubMed Central

Pinto-Fernandez, Adan; Kessler, Benedikt M.

2016-01-01

Controlling cell proliferation is one of the hallmarks of cancer. A number of critical checkpoints ascertain progression through the different stages of the cell cycle, which can be aborted when perturbed, for instance by errors in DNA replication and repair. These molecular checkpoints are regulated by a number of proteins that need to be present at the right time and quantity. The ubiquitin system has emerged as a central player controlling the fate and function of such molecules such as cyclins, oncogenes and components of the DNA repair machinery. In particular, proteases that cleave ubiquitin chains, referred to as deubiquitylating enzymes (DUBs), have attracted recent attention due to their accessibility to modulation by small molecules. In this review, we describe recent evidence of the critical role of DUBs in aspects of cell cycle checkpoint control, associated DNA repair mechanisms and regulation of transcription, representing pathways altered in cancer. Therefore, DUBs involved in these processes emerge as potentially critical targets for the treatment of not only hematological, but potentially also solid tumors. PMID:27516771

Chemical Constituents from Hericium erinaceus Promote Neuronal Survival and Potentiate Neurite Outgrowth via the TrkA/Erk1/2 Pathway.

PubMed

Zhang, Cheng-Chen; Cao, Chen-Yu; Kubo, Miwa; Harada, Kenichi; Yan, Xi-Tao; Fukuyama, Yoshiyasu; Gao, Jin-Ming

2017-07-30

Hericium erinaceus is a culinary-medicinal mushroom used traditionally in Eastern Asia to improve memory. In this work, we investigated the neuroprotective and neuritogenic effects of the secondary metabolites isolated from the MeOH extract of cultured mycelium of H. erinaceus and the primary mechanisms involved. One new dihydropyridine compound ( 6 ) and one new natural product ( 2 ) together with five known compounds ( 1 , 3 - 5 , 7 ) were obtained and their structures were elucidated by spectroscopic analysis, including 2D NMR and HRMS. The cell-based screening for bioactivity showed that 4-chloro-3,5-dimethoxybenzoic methyl ester ( 1 ) and a cyathane diterpenoid, erincine A ( 3 ), not only potentiated NGF-induced neurite outgrowth but also protected neuronally-differentiated cells against deprivation of NGF in PC12 pheochromocytoma cells. Additionally, compound 3 induced neuritogenesis in primary rat cortex neurons. Furthermore, our results revealed that TrkA-mediated and Erk1/2-dependant pathways could be involved in 1 and 3 -promoted NGF-induced neurite outgrowth in PC12 cells.

Chemical Constituents from Hericium erinaceus Promote Neuronal Survival and Potentiate Neurite Outgrowth via the TrkA/Erk1/2 Pathway

PubMed Central

Cao, Chen-Yu; Kubo, Miwa; Harada, Kenichi; Yan, Xi-Tao; Fukuyama, Yoshiyasu; Gao, Jin-Ming

2017-01-01

Hericium erinaceus is a culinary-medicinal mushroom used traditionally in Eastern Asia to improve memory. In this work, we investigated the neuroprotective and neuritogenic effects of the secondary metabolites isolated from the MeOH extract of cultured mycelium of H. erinaceus and the primary mechanisms involved. One new dihydropyridine compound (6) and one new natural product (2) together with five known compounds (1,3–5,7) were obtained and their structures were elucidated by spectroscopic analysis, including 2D NMR and HRMS. The cell-based screening for bioactivity showed that 4-chloro-3,5-dimethoxybenzoic methyl ester (1) and a cyathane diterpenoid, erincine A (3), not only potentiated NGF-induced neurite outgrowth but also protected neuronally-differentiated cells against deprivation of NGF in PC12 pheochromocytoma cells. Additionally, compound 3 induced neuritogenesis in primary rat cortex neurons. Furthermore, our results revealed that TrkA-mediated and Erk1/2-dependant pathways could be involved in 1 and 3-promoted NGF-induced neurite outgrowth in PC12 cells. PMID:28758954

Peroxiredoxin Expression of Human Osteosarcoma Cells Is Influenced by Cold Atmospheric Plasma Treatment.

PubMed

Gümbel, Denis; Gelbrich, Nadine; Napp, Matthias; Daeschlein, Georg; Kramer, Axel; Sckell, Axel; Burchardt, Martin; Ekkernkamp, Axel; Stope, Matthias B

2017-03-01

To evaluate the potential involvement of redox-specific signalling pathways in cold atmospheric plasma (CAP)-induced apoptosis on human osteosarcoma cells. Osteosarcoma cell lines were treated with CAP with or without antioxidative agents and seeded in cell culture plates. Cell proliferation was determined by counting viable cells. Carrier gas-treated cells served as control. Peroxiredoxin (PRX) 1-3 expression and secretion were assessed. CAP treatment exhibited strongly attenuated proliferation rates. This effect was significantly attenuated by the addition of N-acetylcysteine (NAC). CAP-treated cells exhibited an increase of PRX 1 and 2 10 sec after treatment. The ratio of oxidized to reduced PRX1 and PRX2 was significantly altered with increasing cellular concentration of the oxidized dimer. Antioxidant supplementation with NAC increases proliferation of CAP-treated osteosarcoma cells, implicating an involvement of redox signalling. Activation of PRX1 and -2 indicate CAP affects redox homeostasis. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

The intranuclear PEX domain of MMP involves proliferation, migration, and metastasis of aggressive adenocarcinoma cells.

PubMed

Okusha, Yuka; Eguchi, Takanori; Sogawa, Chiharu; Okui, Tatsuo; Nakano, Keisuke; Okamoto, Kuniaki; Kozaki, Ken-Ichi

2018-05-15

Members of matrix metalloproteinase (MMP) family promote cancer cell migration, invasion, and metastasis through alteration of the tumor milieu, intracellular signaling pathways, and transcription. We examined gene expression signatures of colon adenocarcinoma cell lines with different metastatic potentials and found that rapidly metastatic cells powerfully expressed genes encoding MMP3 and MMP9. The non-proteolytic PEX isoform and proteolytic isoforms of MMPs were significantly expressed in the metastatic cells in vitro. Knockdown of MMP3 attenuated cancer cell migration and invasion in vitro and lung metastasis in vivo. Profound nuclear localization of MMP3/PEX was found in tumor-stroma marginal area. In contrast, MMP9 was localized in central area of subcutaneous tumors. Overexpression of the PEX isoform of MMP3 promoted proliferation and migration of the rapidly metastatic cells in vitro. Taken together, the non-proteolytic PEX isoform of MMPs locating in cell nuclei involves proliferation, migration, and subsequent metastasis of aggressive adenocarcinoma cells. © 2018 Wiley Periodicals, Inc.

Depletion of histone demethylase KDM2A enhanced the adipogenic and chondrogenic differentiation potentials of stem cells from apical papilla

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

Dong, Rui; Yao, Rui; Du, Juan

Mesenchymal stem cells (MSCs) are a reliable resource for tissue regeneration, but the molecular mechanism underlying directed differentiation remains unclear; this has restricted potential MSC applications. The histone demethylase, lysine (K)-specific demethylase 2A (KDM2A), is evolutionarily conserved and ubiquitously expressed members of the JmjC-domain-containing histone demethylase family. A previous study determined that KDM2A can regulate the cell proliferation and osteo/dentinogenic differentiation of MSCs. It is not known whether KDM2A is involved in the other cell lineages differentiation of MSCs. Here, we show that depletion of KDM2A by short hairpin RNAs can enhance adipogenic and chondrogenic differentiation potentials in human stemmore » cells from apical papilla (SCAPs). We found that the stemness-related genes, SOX2, and the embryonic stem cell master transcription factor, NANOG were significantly increased after silence of KDM2A in SCAPs. Moreover, we found that knock-down of the KDM2A co-factor, BCOR also up-regulated the mRNA levels of SOX2 and NANOG. Furthermore, Chromatin immunoprecipitation assays demonstrate that silence of KDM2A increased the histone H3 Lysine 4 (H3K4) trimethylation in the SOX2 and NANOG locus and regulates its expression. In conclusion, our results suggested that depletion of KDM2A enhanced the adipogenic and chondrogenic differentiation potentials of SCAPs by up-regulated SOX2 and NANOG, BCOR also involved in this regulation as co-factor, and provided useful information to understand the molecular mechanism underlying directed differentiation in MSCs. - Highlights: • Depletion of KDM2A enhances adipogenic/chondrogenic differentiation in SCAPs. • Depletion of KDM2A enhances the differentiation of SCAPs by activate SOX2 and NANOG. • Silence of KDM2A increases histone H3 Lysine 4 trimethylation in SOX2 and NANOG. • BCOR is co-factor of KDM2A involved in the differentiation regulation.« less

Targeting Cancer Cells with Reactive Oxygen and Nitrogen Species Generated by Atmospheric-Pressure Air Plasma

PubMed Central

Hoan, Nguyen Ngoc; Kim, Churl Ho; Moon, Eunpyo; Choi, Kyeong Sook; Yang, Sang Sik; Lee, Jong-Soo

2014-01-01

The plasma jet has been proposed as a novel therapeutic method for cancer. Anticancer activity of plasma has been reported to involve mitochondrial dysfunction. However, what constituents generated by plasma is linked to this anticancer process and its mechanism of action remain unclear. Here, we report that the therapeutic effects of air plasma result from generation of reactive oxygen/nitrogen species (ROS/RNS) including H2O2, Ox, OH−, •O2, NOx, leading to depolarization of mitochondrial membrane potential and mitochondrial ROS accumulation. Simultaneously, ROS/RNS activate c-Jun NH2-terminal kinase (JNK) and p38 kinase. As a consequence, treatment with air plasma jets induces apoptotic death in human cervical cancer HeLa cells. Pretreatment of the cells with antioxidants, JNK and p38 inhibitors, or JNK and p38 siRNA abrogates the depolarization of mitochondrial membrane potential and impairs the air plasma-induced apoptotic cell death, suggesting that the ROS/RNS generated by plasma trigger signaling pathways involving JNK and p38 and promote mitochondrial perturbation, leading to apoptosis. Therefore, administration of air plasma may be a feasible strategy to eliminate cancer cells. PMID:24465942

Targeting the Regulatory Machinery of BIM for Cancer Therapy

PubMed Central

Harada, Hisashi; Grant, Steven

2013-01-01

BIM represents a BH3-only proapoptotic member of the BCL-2 family of apoptotic regulatory proteins. Recent evidence suggests that in addition to its involvement in normal homeostasis, BIM plays a critical role in tumor cell biology, including the regulation of tumorigenesis through activities as a tumor suppressor, tumor metastasis, and tumor cell survival. Consequently, BIM has become the focus of intense interest as a potential target for cancer chemotherapy. The control of BIM expression is complex, and involves multiple factors, including epigenetic events (i.e., promoter acetylation or methylation, miRNA), transcription factors, posttranscriptional regulation, and posttranslational modifications, most notably phosphorylation. Significantly, the expression of BIM by tumor cells has been shown to play an important role in determining the response of transformed cells to not only conventional cytotoxic agents, but also to a broad array of targeted agents that interrupt cell signaling and survival pathways. Furthermore, modifications in BIM expression may be exploited to improve the therapeutic activity and potentially the selectivity of such agents. It is likely that evolving insights into the factors that regulate BIM expression will ultimately lead to novel BIM-based therapeutic strategies in the future. PMID:22856430

New MKLP-2 inhibitors in the paprotrain series: Design, synthesis and biological evaluations.

PubMed

Labrière, Christophe; Talapatra, Sandeep K; Thoret, Sylviane; Bougeret, Cécile; Kozielski, Frank; Guillou, Catherine

2016-02-15

Members of the kinesin superfamily are involved in key functions during intracellular transport and cell division. Their involvement in cell division makes certain kinesins potential targets for drug development in cancer chemotherapy. The two most advanced kinesin targets are Eg5 and CENP-E with inhibitors in clinical trials. Other mitotic kinesins are also being investigated for their potential as prospective drug targets. One recently identified novel potential cancer therapeutic target is the Mitotic kinesin-like protein 2 (MKLP-2), a member of the kinesin-6 family, which plays an essential role during cytokinesis. Previous studies have shown that inhibition of MKLP-2 leads to binucleated cells due to failure of cytokinesis. We have previously identified compound 1 (paprotrain) as the first selective inhibitor of MKLP-2. Herein we describe the synthesis and biological evaluation of new analogs of 1. Our structure-activity relationship (SAR) study reveals the key chemical elements in the paprotrain family necessary for MKLP-2 inhibition. We have successfully identified one MKLP-2 inhibitor 9a that is more potent than paprotrain. In addition, in vitro analysis of a panel of kinesins revealed that this compound is selective for MKLP-2 compared to other kinesins tested and also does not have an effect on microtubule dynamics. Upon testing in different cancer cell lines, we find that the more potent paprotrain analog is also more active than paprotrain in 10 different cancer cell lines. Increased selectivity and higher potency is therefore a step forward toward establishing MKLP-2 as a potential cancer drug target. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cancer treatment in childhood and testicular function: the importance of the somatic environment.

PubMed

Stukenborg, Jan-Bernd; Jahnukainen, Kirsi; Hutka, Marsida; Mitchell, Rod T

2018-02-01

Testicular function and future fertility may be affected by cancer treatment during childhood. Whilst survival of the germ (stem) cells is critical for ensuring the potential for fertility in these patients, the somatic cell populations also play a crucial role in providing a suitable environment to support germ cell maintenance and subsequent development. Regulation of the spermatogonial germ-stem cell niche involves many signalling pathways with hormonal influence from the hypothalamo-pituitary-gonadal axis. In this review, we describe the somatic cell populations that comprise the testicular germ-stem cell niche in humans and how they may be affected by cancer treatment during childhood. We also discuss the experimental models that may be utilized to manipulate the somatic environment and report the results of studies that investigate the potential role of somatic cells in the protection of the germ cells in the testis from cancer treatment. © 2018 The authors.

Cancer treatment in childhood and testicular function: the importance of the somatic environment

PubMed Central

Stukenborg, Jan-Bernd; Jahnukainen, Kirsi; Hutka, Marsida

2018-01-01

Testicular function and future fertility may be affected by cancer treatment during childhood. Whilst survival of the germ (stem) cells is critical for ensuring the potential for fertility in these patients, the somatic cell populations also play a crucial role in providing a suitable environment to support germ cell maintenance and subsequent development. Regulation of the spermatogonial germ-stem cell niche involves many signalling pathways with hormonal influence from the hypothalamo-pituitary-gonadal axis. In this review, we describe the somatic cell populations that comprise the testicular germ-stem cell niche in humans and how they may be affected by cancer treatment during childhood. We also discuss the experimental models that may be utilized to manipulate the somatic environment and report the results of studies that investigate the potential role of somatic cells in the protection of the germ cells in the testis from cancer treatment. PMID:29351905

The osteoblastic niche in the context of multiple myeloma.

PubMed

Toscani, Denise; Bolzoni, Marina; Accardi, Fabrizio; Aversa, Franco; Giuliani, Nicola

2015-01-01

The osteoblastic niche has a critical role in the regulation of hemopoietic stem cell (HSC) quiescence and self-renewal and in the support of hematopoiesis. Several mechanisms are involved in the crosstalk between stem cells and osteoblasts, including soluble cytokines, adhesion molecules, and signal pathways such as the wingless-Int (Wnt), Notch, and parathyroid hormone pathways. According to the most recent evidence, there is an overlap between osteoblastic and perivascular niches that affects HSC function involving mesenchymal stromal and endothelial cells and a gradient of oxygen regulated by hypoxia inducible factor (HIF)-1α. Derived from plasma cells, multiple myeloma (MM) is a hematopoietic malignancy characterized by a peculiar dependency on the bone microenvironment. Quiescent MM cells may reside in the osteoblastic niche for protection from apoptotic stimuli; in turn, MM cells suppress osteoblast formation and function, leading to impairment of bone formation and the development of osteolytic lesions. Several recent studies have investigated the mechanisms involved in the relationship between osteoblasts and MM cells and identified potential therapeutic targets in the osteoblastic niche, including the HIF-1α, Runx2, and Wnt (both canonical and noncanonical) signaling pathways. © 2014 New York Academy of Sciences.

PRC2 inhibition counteracts the culture-associated loss of engraftment potential of human cord blood-derived hematopoietic stem and progenitor cells.

PubMed

Varagnolo, Linda; Lin, Qiong; Obier, Nadine; Plass, Christoph; Dietl, Johannes; Zenke, Martin; Claus, Rainer; Müller, Albrecht M

2015-07-22

Cord blood hematopoietic stem cells (CB-HSCs) are an outstanding source for transplantation approaches. However, the amount of cells per donor is limited and culture expansion of CB-HSCs is accompanied by a loss of engraftment potential. In order to analyze the molecular mechanisms leading to this impaired potential we profiled global and local epigenotypes during the expansion of human CB hematopoietic stem and progenitor cells (HPSCs). Human CB-derived CD34+ cells were cultured in serum-free medium together with SCF, TPO, FGF, with or without Igfbp2 and Angptl5 (STF/STFIA cocktails). As compared to the STF cocktail, the STFIA cocktail maintains in vivo repopulation capacity of cultured CD34+ cells. Upon expansion, CD34+ cells genome-wide remodel their epigenotype and depending on the cytokine cocktail, cells show different H3K4me3 and H3K27me3 levels. Expanding cells without Igfbp2 and Angptl5 leads to higher global H3K27me3 levels. ChIPseq analyses reveal a cytokine cocktail-dependent redistribution of H3K27me3 profiles. Inhibition of the PRC2 component EZH2 counteracts the culture-associated loss of NOD scid gamma (NSG) engraftment potential. Collectively, our data reveal chromatin dynamics that underlie the culture-associated loss of engraftment potential. We identify PRC2 component EZH2 as being involved in the loss of engraftment potential during the in vitro expansion of HPSCs.

Cadmium induces carcinogenesis in BEAS-2B cells through ROS-dependent activation of PI3K/AKT/GSK-3β/β-catenin signaling

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

Son, Young-Ok; Wang, Lei; Poyil, Pratheeshkumar

Cadmium has been widely used in industry and is known to be carcinogenic to humans. Although it is widely accepted that chronic exposure to cadmium increases the incidence of cancer, the mechanisms underlying cadmium-induced carcinogenesis are unclear. The main aim of this study was to investigate the role of reactive oxygen species (ROS) in cadmium-induced carcinogenesis and the signal transduction pathways involved. Chronic exposure of human bronchial epithelial BEAS-2B cells to cadmium induced cell transformation, as evidenced by anchorage-independent growth in soft agar and clonogenic assays. Chronic cadmium treatment also increased the potential of these cells to invade and migrate.more » Injection of cadmium-stimulated cells into nude mice resulted in the formation of tumors. In contrast, the cadmium-mediated increases in colony formation, cell invasion and migration were prevented by transfection with catalase, superoxide dismutase-1 (SOD1), or SOD2. In particular, chronic cadmium exposure led to activation of signaling cascades involving PI3K, AKT, GSK-3β, and β-catenin and transfection with each of the above antioxidant enzymes markedly inhibited cadmium-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the cadmium-mediated increase in total and active β-catenin proteins and colony formation. Moreover, there was a marked induction of AKT, GSK-3β, β-catenin, and carcinogenic markers in tumor tissues formed in mice after injection with cadmium-stimulated cells. Collectively, our findings suggest a direct involvement of ROS in cadmium-induced carcinogenesis and implicate a role of AKT/GSK-3β/β-catenin signaling in this process. -- Highlights: ► Chronic exposure to cadmium induces carcinogenic properties in BEAS-2B cells. ► ROS involved in cadmium-induced tumorigenicity of BEAS-2B cells. ► Cadmium activates ROS-dependent AKT/GSK-3β/β-catenin-mediated signaling. ► ROS-dependent signaling as potential therapeutic targets in cadmium carcinogenesis.« less

Involvement of intracellular Zn2+ signaling in LTP at perforant pathway-CA1 pyramidal cell synapse.

PubMed

Tamano, Haruna; Nishio, Ryusuke; Takeda, Atsushi

2017-07-01

Physiological significance of synaptic Zn 2+ signaling was examined at perforant pathway-CA1 pyramidal cell synapses. In vivo long-term potentiation (LTP) at perforant pathway-CA1 pyramidal cell synapses was induced using a recording electrode attached to a microdialysis probe and the recording region was locally perfused with artificial cerebrospinal fluid (ACSF) via the microdialysis probe. Perforant pathway LTP was not attenuated under perfusion with CaEDTA (10 mM), an extracellular Zn 2+ chelator, but attenuated under perfusion with ZnAF-2DA (50 μM), an intracellular Zn 2+ chelator, suggesting that intracellular Zn 2+ signaling is required for perforant pathway LTP. Even in rat brain slices bathed in CaEDTA in ACSF, intracellular Zn 2+ level, which was measured with intracellular ZnAF-2, was increased in the stratum lacunosum-moleculare where perforant pathway-CA1 pyramidal cell synapses were contained after tetanic stimulation. These results suggest that intracellular Zn 2+ signaling, which originates in internal stores/proteins, is involved in LTP at perforant pathway-CA1 pyramidal cell synapses. Because the influx of extracellular Zn 2+ , which originates in presynaptic Zn 2+ release, is involved in LTP at Schaffer collateral-CA1 pyramidal cell synapses, synapse-dependent Zn 2+ dynamics may be involved in plasticity of postsynaptic CA1 pyramidal cells. © 2017 Wiley Periodicals, Inc.

Novel clinical uses for cord blood derived mesenchymal stromal cells.

PubMed

Olson, Amanda L; McNiece, Ian K

2015-06-01

Regenerative medicine offers new hope for many debilitating diseases that result in damage to tissues and organs. The concept is straightforward with replacement of damaged cells with new functional cells. However, most tissues and organs are complex structures involving multiple cell types, supportive structures, a microenvironment producing cytokines and growth factors and a vascular system to supply oxygen and other nutrients. Therefore repair, particularly in the setting of ischemic damage, may require delivery of multiple cell types providing new vessel formation, a new microenvironment and functional cells. The field of stem cell biology has identified a number of stem cell sources including embryonic stem cells and adult stem cells that offer the potential to replace virtually all functional cells of the body. The focus of this article is a discussion of the potential of mesenchymal stromal cells (MSCs) from cord blood (CB) for regenerative medicine approaches. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Soluble Factors from Human Olfactory Neural Stem/Progenitor Cells Influence the Fate Decisions of Hippocampal Neural Precursor Cells.

PubMed

Gómez-Virgilio, Laura; Ramírez-Rodríguez, Gerardo Bernabé; Sánchez-Torres, Carmen; Ortiz-López, Leonardo; Meraz-Ríos, Marco Antonio

2018-03-01

Neurogenesis plays a significant role during adulthood, and the observation that neural stem cells reside in the central nervous system and the olfactory epithelium has attracted attention due to their importance in neuronal regeneration. In addition, soluble factors (SFs) release by neural stem cells may modulate the neurogenic process. Thus, in this study, we identified the SFs released by olfactory human neural stem/progenitor cells (hNS/PCs-OE). These cells express Ki67, nestin, and βIII-tubulin, indicating their neural lineage. The hNS/PCs-OE also express PSD95 and tau proteins during proliferation, but increased levels are observed after differentiation. Thus, we evaluated the effects of SFs from hNS/PCs-OE on the viability, proliferation, and differentiation potential of adult murine hippocampal neural precursor cells (AHPCs). SFs from hNS/PCs-OE maintain cells in the precursor and proliferative stages and mainly promote the astrocytic differentiation of AHPCs. These effects involved the activation, as measured by phosphorylation, of several proteins (Erk1/2; Akt/PRAS40/GSK3β and JAK/STAT) involved in key events of the neurogenic process. Moreover, according to the results from the antibody-based microarray approach, among the soluble factors, hNS/PCs-OE produce interleukin-6 (IL-6) and neurotrophin 4 (NT4). However, residual epidermal growth factor (EGF) was also detected. These proteins partially reproduced the effects of SFs from hNS/PCs-OE on AHPCs, and the mechanism underlying these effects is mediated by Src proteins, which have been implicated in EGF-induced transactivation of TrkB receptor. The results of the present study suggest the potential use of SFs from hNS/PCs-OE in controlling the differentiation potential of AHPCs. Thus, the potential clinical relevance of hNS/PCs-OE is worth pursuing.

PaDef defensin from avocado (Persea americana var. drymifolia) is cytotoxic to K562 chronic myeloid leukemia cells through extrinsic apoptosis.

PubMed

Flores-Alvarez, Luis José; Guzmán-Rodríguez, Jaquelina Julia; López-Gómez, Rodolfo; Salgado-Garciglia, Rafael; Ochoa-Zarzosa, Alejandra; López-Meza, Joel E

2018-06-01

Plant defensins, a group of antimicrobial peptides, show selective cytotoxicity toward cancer cells. However, their mechanisms of action remain poorly understood. Here, we evaluated the cytotoxicity of PaDef defensin from avocado (Persea americana var. drymifolia) on K562 chronic myeloid leukemia cells and analyzed the pathway involved in the induction of cell death. The defensin PaDef was not cytotoxic against human PBMCs; however, it was cytotoxic for K562 cell line (IC 50  = 97.3 μg/ml) activating apoptosis at 12 h. PaDef did not affect the mitochondrial membrane potential (ΔΨm), neither the transmembranal potential or the release of intracellular calcium. Also, PaDef induced gene expression of caspase 8 (∼2 fold), TNF-α (∼4 fold) and TNFR1 (∼10 fold). In addition, the activation of caspase 8 was detected at 24 h, whereas caspase 9 activity was not modified, suggesting that the extrinsic apoptosis pathway could be activated. In conclusion, PaDef induces apoptosis on K562 cells, which is related to the activation of caspase 8 and involves the participation of TNF-α, which is a novel property for a plant defensin. Copyright © 2018 Elsevier Ltd. All rights reserved.

GSTM3 and GSTP1: novel players driving tumor progression in cervical cancer.

PubMed

Checa-Rojas, Alberto; Delgadillo-Silva, Luis Fernando; Velasco-Herrera, Martín Del Castillo; Andrade-Domínguez, Andrés; Gil, Jeovanis; Santillán, Orlando; Lozano, Luis; Toledo-Leyva, Alfredo; Ramírez-Torres, Alberto; Talamas-Rohana, Patricia; Encarnación-Guevara, Sergio

2018-04-24

The molecular processes and proteomic markers leading to tumor progression (TP) in cervical cancer (CC) are either unknown or only partially understood. TP affects metabolic and regulatory mechanisms that can be identified as proteomic changes. To identify which proteins are differentially expressed and to understand the mechanisms of cancer progression, we analyzed the dynamics of the tumor proteome in CC cell lines. This analysis revealed two proteins that are up-regulated during TP, GSTM3 and GSTP1. These proteins are involved in cell maintenance, cell survival and the cellular stress response via the NF-κB and MAP kinase pathways during TP. Furthermore, GSTM3 and GSTP1 knockdown showed that evasion of apoptosis was affected, and tumor proliferation was significantly reduced. Our data indicate the critical role of GST proteins in the regulation and progression of cervical cancer cells. Hence, we suggest GSTM3 and GSTP1 as novel biomarkers and potential therapeutic targets for treating cervical cancer. CC is particularly hazardous in the advanced stages, and there are few therapeutic strategies specifically targeting these stages. We performed analyses on CC tumor proteome dynamics and identified GSTM3 and GSTP1 as novel potential therapeutic targets. Knockdown of these proteins showed that they are involved in cell survival, cell proliferation and cellular evasion of apoptosis.

Metabolomic study of corticosterone-induced cytotoxicity in PC12 cells by ultra performance liquid chromatography-quadrupole/time-of-flight mass spectrometry.

PubMed

Zhang, Hongye; Zheng, Hua; Zhao, Gan; Tang, Chaoling; Lu, Shiyin; Cheng, Bang; Wu, Fang; Wei, Jinbin; Liang, Yonghong; Ruan, Junxiang; Song, Hui; Su, Zhiheng

2016-03-01

Glucocorticoids (GCs) have been proved to be an important pathogenic factor of some neuropsychiatric disorders. Usually, a classical injury model based on corticosterone-induced cytotoxicity of differentiated rat pheochromocytoma (PC12) cells was used to stimulate the state of GC damage of hippocampal neurons and investigate its potential mechanisms involved. However, up to now, the mechanism of corticosterone-induced cytotoxicity in PC12 cells was still looking forward to further elucidation. In this work, the metabolomic study of the biochemical changes caused by corticosterone-induced cytotoxicity in differentiated PC12 cells with different corticosterone concentrations was performed for the first time, using the ultra performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF MS). Partial least squares-discriminate analysis (PLS-DA) indicated that metabolic profiles of different corticosterone treatment groups deviated from the control group. A total of fifteen metabolites were characterized as potential biomarkers involved in corticosterone-induced cytotoxicity, which were corresponding to the dysfunctions of five pathways including glycerophospholipid metabolism, sphingolipid metabolism, oxidation of fatty acids, glycerolipid metabolism and sterol lipid metabolism. This study indicated that the rapid and holistic cell metabolomics approach might be a powerful tool to further study the pathogenesis mechanism of corticosterone-induced cytotoxicity in PC12 cells.

Microarray-based detection of Salmonella enterica serovar Enteritidis genes involved in chicken reproductive tract colonization.

PubMed

Raspoet, R; Appia-Ayme, C; Shearer, N; Martel, A; Pasmans, F; Haesebrouck, F; Ducatelle, R; Thompson, A; Van Immerseel, F

2014-12-01

Salmonella enterica serovar Enteritidis has developed the potential to contaminate table eggs internally, by colonization of the chicken reproductive tract and internalization in the forming egg. The serotype Enteritidis has developed mechanisms to colonize the chicken oviduct more successfully than other serotypes. Until now, the strategies exploited by Salmonella Enteritidis to do so have remained largely unknown. For that reason, a microarray-based transposon library screen was used to identify genes that are essential for the persistence of Salmonella Enteritidis inside primary chicken oviduct gland cells in vitro and inside the reproductive tract in vivo. A total of 81 genes with a potential role in persistence in both the oviduct cells and the oviduct tissue were identified. Major groups of importance include the Salmonella pathogenicity islands 1 and 2, genes involved in stress responses, cell wall, and lipopolysaccharide structure, and the region-of-difference genomic islands 9, 21, and 40. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Ophiobolin A, a sesterpenoid fungal phytotoxin, displays different mechanisms of cell death in mammalian cells depending upon the cancer cell origin

PubMed Central

Morrison, Rachel; Lodge, Tiffany; Evidente, Antonio; Kiss, Robert; Townley, Helen

2017-01-01

Herein we have undertaken a systematic analysis of the effects of the fungal derivative ophiobolin A (OphA) on eight cancer cell lines from different tissue types. The LD50 for each cell line was determined and the change in cell size determined. Flow cytometric analysis and western blotting were used to assess the cell death markers for early apoptosis, late apoptosis and necrosis, and the involvement of the caspase signalling pathway. Alterations in calcium levels and reactive oxygen species were assessed due to their integral involvement in intracellular signalling. Subsequently, the endoplasmic reticulum (ER) and mitochondrial responses were investigated more closely. The extent of ER swelling, and the upregulation of proteins involved in the unfolded protein responses (UPR) were seen to vary according to cell line. The mitochondria were also shown to behave differently in response to the OphA in the different cell lines in terms of the change in membrane potential, the total area of mitochondria in the cell and the number of mitochondrial bifurcations. The data obtained in the present study indicate that the cancer cell lines tested are unable to successfully activate the ER stress/UPR responses, and that the mitochondria appear to be a central player in OphA-induced cancer cell death. PMID:28112374

Efficient generation of hPSC-derived midbrain dopaminergic neurons in a fully defined, scalable, 3D biomaterial platform

PubMed Central

Adil, Maroof M.; Rodrigues, Gonçalo M. C.; Kulkarni, Rishikesh U.; Rao, Antara T.; Chernavsky, Nicole E.; Miller, Evan W.; Schaffer, David V.

2017-01-01

Pluripotent stem cells (PSCs) have major potential as an unlimited source of functional cells for many biomedical applications; however, the development of cell manufacturing systems to enable this promise faces many challenges. For example, there have been major recent advances in the generation of midbrain dopaminergic (mDA) neurons from stem cells for Parkinson’s Disease (PD) therapy; however, production of these cells typically involves undefined components and difficult to scale 2D culture formats. Here, we used a fully defined, 3D, thermoresponsive biomaterial platform to rapidly generate large numbers of action-potential firing mDA neurons after 25 days of differentiation (~40% tyrosine hydroxylase (TH) positive, maturing into 25% cells exhibiting mDA neuron-like spiking behavior). Importantly, mDA neurons generated in 3D exhibited a 30-fold increase in viability upon implantation into rat striatum compared to neurons generated on 2D, consistent with the elevated expression of survival markers FOXA2 and EN1 in 3D. A defined, scalable, and resource-efficient cell culture platform can thus rapidly generate high quality differentiated cells, both neurons and potentially other cell types, with strong potential to accelerate both basic and translational research. PMID:28091566

Stochastic simulation tools and continuum models for describing two-dimensional collective cell spreading with universal growth functions

NASA Astrophysics Data System (ADS)

Jin, Wang; Penington, Catherine J.; McCue, Scott W.; Simpson, Matthew J.

2016-10-01

Two-dimensional collective cell migration assays are used to study cancer and tissue repair. These assays involve combined cell migration and cell proliferation processes, both of which are modulated by cell-to-cell crowding. Previous discrete models of collective cell migration assays involve a nearest-neighbour proliferation mechanism where crowding effects are incorporated by aborting potential proliferation events if the randomly chosen target site is occupied. There are two limitations of this traditional approach: (i) it seems unreasonable to abort a potential proliferation event based on the occupancy of a single, randomly chosen target site; and, (ii) the continuum limit description of this mechanism leads to the standard logistic growth function, but some experimental evidence suggests that cells do not always proliferate logistically. Motivated by these observations, we introduce a generalised proliferation mechanism which allows non-nearest neighbour proliferation events to take place over a template of r≥slant 1 concentric rings of lattice sites. Further, the decision to abort potential proliferation events is made using a crowding function, f(C), which accounts for the density of agents within a group of sites rather than dealing with the occupancy of a single randomly chosen site. Analysing the continuum limit description of the stochastic model shows that the standard logistic source term, λ C(1-C), where λ is the proliferation rate, is generalised to a universal growth function, λ C f(C). Comparing the solution of the continuum description with averaged simulation data indicates that the continuum model performs well for many choices of f(C) and r. For nonlinear f(C), the quality of the continuum-discrete match increases with r.

Potential role of p21 Activated Kinase 1 (PAK1) in the invasion and motility of oral cancer cells.

PubMed

Parvathy, Muraleedharan; Sreeja, Sreeharshan; Kumar, Rakesh; Pillai, Madhavan Radhakrishna

2016-05-16

Oral cancer malignancy consists of uncontrolled division of cells primarily in and around the floor of the oral cavity, gingiva, oropharynx, lower lip and base of the tongue. According to GLOBOCAN 2012 report, oral cancer is one of the most common cancers among males and females in India. Even though significant advancements have been made in the field of oral cancer treatment modalities, the overall prognosis for the patients has not improved in the past few decades and hence, this demands a new thrust for the identification of novel therapeutic targets in oral cancer. p21 Activated Kinases (PAKs) are potential therapeutic targets that are involved in numerous physiological functions. PAKs are serine-threonine kinases and they serve as important regulators of cytoskeletal dynamics and cell motility, transcription through MAP kinase cascades, death and survival signalling, and cell-cycle progression. Although PAKs are known to play crucial roles in cancer progression, the role and clinical significance of PAKs in oral cancer remains poorly understood. Our results suggest that PAK1 is over-expressed in oral cancer cell lines. Stimulation of Oral Squamous Cell Carcinoma (OSCC) cells with serum growth factors leads to PAK1 re-localization and might cause a profound cytoskeletal remodelling. PAK1 was also found to be involved in the invasion, migration and cytoskeletal remodelling of OSCC cells. Our study revealed that PAK1 may play a crucial role in the progression of OSCC. Studying the role of PAK1 and its substrates is likely to enhance our understanding of oral carcinogenesis and potential therapeutic value of PAKs in oral cancer.

Stochastic simulation tools and continuum models for describing two-dimensional collective cell spreading with universal growth functions.

PubMed

Jin, Wang; Penington, Catherine J; McCue, Scott W; Simpson, Matthew J

2016-10-07

Two-dimensional collective cell migration assays are used to study cancer and tissue repair. These assays involve combined cell migration and cell proliferation processes, both of which are modulated by cell-to-cell crowding. Previous discrete models of collective cell migration assays involve a nearest-neighbour proliferation mechanism where crowding effects are incorporated by aborting potential proliferation events if the randomly chosen target site is occupied. There are two limitations of this traditional approach: (i) it seems unreasonable to abort a potential proliferation event based on the occupancy of a single, randomly chosen target site; and, (ii) the continuum limit description of this mechanism leads to the standard logistic growth function, but some experimental evidence suggests that cells do not always proliferate logistically. Motivated by these observations, we introduce a generalised proliferation mechanism which allows non-nearest neighbour proliferation events to take place over a template of [Formula: see text] concentric rings of lattice sites. Further, the decision to abort potential proliferation events is made using a crowding function, f(C), which accounts for the density of agents within a group of sites rather than dealing with the occupancy of a single randomly chosen site. Analysing the continuum limit description of the stochastic model shows that the standard logistic source term, [Formula: see text], where λ is the proliferation rate, is generalised to a universal growth function, [Formula: see text]. Comparing the solution of the continuum description with averaged simulation data indicates that the continuum model performs well for many choices of f(C) and r. For nonlinear f(C), the quality of the continuum-discrete match increases with r.

Tolerogenic dendritic cells in autoimmune diseases: crucial players in induction and prevention of autoimmunity.

PubMed

Torres-Aguilar, Honorio; Blank, Miri; Jara, Luis J; Shoenfeld, Yehuda

2010-11-01

The immune system has evolved to coordinate responses against numerous invading pathogens and simultaneously remain silent facing self-antigens and those derived from commensal organisms. But, if both processes are not maintained in strict balance, a potential threat can emerge due to the risk of chronic inflammation and/or autoimmunity development. Therefore, there is a negative immune regulation where tolerogenic dendritic cells (tDCs) participate actively. Under steady-state conditions, tDC are notably involved in the elimination of autoreactive T cells at the thymus, and in the control of T cells specific to self and harmless antigens in the periphery. But in the presence of foreign antigens in an inflammatory milieu, dendritic cells (DCs) mature and induce T cells activation and their migration to B cell areas to assist in antibody production. Additionally, there are other factors such as infections, anti tumoral immune responses, trauma-mediated disruption, etc. that may induce alterations in the balance between tolerogenic and immunogenic functions of DCs and instigate the development of autoimmune diseases (ADs). Therefore, in recent years, DCs have emerged as therapeutic targets to control of ADs. Diverse strategies in vitro and/or in animal models of ADs have explored the tolerogenic functions of DCs and demonstrated their feasibility to prevent or control an autoimmune process, but still leaving a void in their application in clinical assays. The purpose of this paper is to give a general overview of the current literature on the significance of tDCs in tolerance maintenance to self and innocuous antigens, the most relevant alterations involved in the pathophysiology of ADs, the cellular and molecular mechanisms involved in their tolerogenic function and the current strategies used to exploit their tolerogenic potential. Published by Elsevier B.V.

PTK 7 Is a Transforming Gene and Prognostic Marker for Breast Cancer and Nodal Metastasis Involvement

PubMed Central

Knyazeva, Tatiana; Wolf, Petra; Högel, Bernhard; Eiermann, Wolfgang; Ullrich, Axel; Knyazev, Pjotr; Ataseven, Beyhan

2014-01-01

Protein Tyrosin Kinase 7 (PTK7) is upregulated in several human cancers; however, its clinical implication in breast cancer (BC) and lymph node (LN) is still unclear. In order to investigate the function of PTK7 in mediating BC cell motility and invasivity, PTK7 expression in BC cell lines was determined. PTK7 signaling in highly invasive breast cancer cells was inhibited by a dominant-negative PTK7 mutant, an antibody against the extracellular domain of PTK7, and siRNA knockdown of PTK7. This resulted in decreased motility and invasivity of BC cells. We further examined PTK7 expression in BC and LN tissue of 128 BC patients by RT-PCR and its correlation with BC related genes like HER2, HER3, PAI1, MMP1, K19, and CD44. Expression profiling in BC cell lines and primary tumors showed association of PTK7 with ER/PR/HER2-negative (TNBC-triple negative BC) cancer. Oncomine data analysis confirmed this observation and classified PTK7 in a cluster with genes associated with agressive behavior of primary BC. Furthermore PTK7 expression was significantly different with respect to tumor size (ANOVA, p = 0.033) in BC and nodal involvement (ANOVA, p = 0.007) in LN. PTK7 expression in metastatic LN was related to shorter DFS (Cox Regression, p = 0.041). Our observations confirmed the transforming potential of PTK7, as well as its involvement in motility and invasivity of BC cells. PTK7 is highly expressed in TNBC cell lines. It represents a novel prognostic marker for BC patients and has potential therapeutic significance. PMID:24409301

Status of and candidates for cell therapy in liver cirrhosis: overcoming the "point of no return" in advanced liver cirrhosis.

PubMed

Terai, Shuji; Tsuchiya, Atsunori

2017-02-01

The treatment of liver cirrhosis is currently being standardized and developed specifically to reduce activation of hepatic stellate cells (HSCs), inhibit fibrosis, increase degradation of matrix components, and reduce activated myofibroblasts. Cell therapy can be applied in the treatment of liver cirrhosis; however, the characteristic features of this therapy differ from those of other treatments because of the involvement of a living body origin and production of multiple cytokines, chemokines, matrix metalloproteinases (MMPs), and growth factors. Thus, cell therapies can potentially have multiple effects on the damaged liver, including alleviating liver cirrhosis and stimulating liver regeneration with affecting the host cells. Cell therapies initially involved autologous bone marrow cell infusion, and have recently developed to include the use of specific cells such as mesenchymal stem cells and macrophages. The associated molecular mechanisms, routes of administration, possibility of allogeneic cell therapy, and host conditions appropriate for cell therapies are now being extensively analyzed. In this review, we summarize the status and future prospects of cell therapy for liver cirrhosis.

Distal Regeneration Involves the Age Dependent Activity of Branchial Sac Stem Cells in the Ascidian Ciona intestinalis.

PubMed

Jeffery, William R

2015-02-01

Tunicates have high capacities for regeneration but the underlying mechanisms and their relationship to life cycle progression are not well understood. Here we investigate the regeneration of distal structures in the ascidian tunicate Ciona intestinalis . Analysis of regenerative potential along the proximal-distal body axis indicated that distal organs, such as the siphons, their pigmented sensory organs, and the neural complex, could only be replaced from body fragments containing the branchial sac. Distal regeneration involves the formation of a blastema composed of cells that undergo cell proliferation prior to differentiation and cells that differentiate without cell proliferation. Both cell types originate in the branchial sac and appear in the blastema at different times after distal injury. Whereas the branchial sac stem cells are present in young animals, they are depleted in old animals that have lost their regeneration capacity. Thus Ciona adults contain a population of age-related stem cells located in the branchial sac that are a source of precursors for distal body regeneration.

Modeling the Hydraulics of Root Growth in Three Dimensions with Phloem Water Sources1[C][OA

PubMed Central

Wiegers, Brandy S.; Cheer, Angela Y.; Silk, Wendy K.

2009-01-01

Primary growth is characterized by cell expansion facilitated by water uptake generating hydrostatic (turgor) pressure to inflate the cell, stretching the rigid cell walls. The multiple source theory of root growth hypothesizes that root growth involves transport of water both from the soil surrounding the growth zone and from the mature tissue higher in the root via phloem and protophloem. Here, protophloem water sources are used as boundary conditions in a classical, three-dimensional model of growth-sustaining water potentials in primary roots. The model predicts small radial gradients in water potential, with a significant longitudinal gradient. The results improve the agreement of theory with empirical studies for water potential in the primary growth zone of roots of maize (Zea mays). A sensitivity analysis quantifies the functional importance of apical phloem differentiation in permitting growth and reveals that the presence of phloem water sources makes the growth-sustaining water relations of the root relatively insensitive to changes in root radius and hydraulic conductivity. Adaptation to drought and other environmental stresses is predicted to involve more apical differentiation of phloem and/or higher phloem delivery rates to the growth zone. PMID:19542299

Modeling the hydraulics of root growth in three dimensions with phloem water sources.

PubMed

Wiegers, Brandy S; Cheer, Angela Y; Silk, Wendy K

2009-08-01

Primary growth is characterized by cell expansion facilitated by water uptake generating hydrostatic (turgor) pressure to inflate the cell, stretching the rigid cell walls. The multiple source theory of root growth hypothesizes that root growth involves transport of water both from the soil surrounding the growth zone and from the mature tissue higher in the root via phloem and protophloem. Here, protophloem water sources are used as boundary conditions in a classical, three-dimensional model of growth-sustaining water potentials in primary roots. The model predicts small radial gradients in water potential, with a significant longitudinal gradient. The results improve the agreement of theory with empirical studies for water potential in the primary growth zone of roots of maize (Zea mays). A sensitivity analysis quantifies the functional importance of apical phloem differentiation in permitting growth and reveals that the presence of phloem water sources makes the growth-sustaining water relations of the root relatively insensitive to changes in root radius and hydraulic conductivity. Adaptation to drought and other environmental stresses is predicted to involve more apical differentiation of phloem and/or higher phloem delivery rates to the growth zone.

Cytotoxicity of citral against melanoma cells: The involvement of oxidative stress generation and cell growth protein reduction.

PubMed

Sanches, Larissa Juliani; Marinello, Poliana Camila; Panis, Carolina; Fagundes, Tatiane Renata; Morgado-Díaz, José Andrés; de-Freitas-Junior, Julio Cesar Madureira; Cecchini, Rubens; Cecchini, Alessandra Lourenço; Luiz, Rodrigo Cabral

2017-03-01

Citral is a natural compound that has shown cytotoxic and antiproliferative effects on breast and hematopoietic cancer cells; however, there are few studies on melanoma cells. Oxidative stress is known to be involved in all stages of melanoma development and is able to modulate intracellular pathways related to cellular proliferation and death. In this study, we hypothesize that citral exerts its cytotoxic effect on melanoma cells by the modulation of cellular oxidative status and/or intracellular signaling. To test this hypothesis, we investigated the antiproliferative and cytotoxic effects of citral on B16F10 murine melanoma cells evaluating its effects on cellular oxidative stress, DNA damage, cell death, and important signaling pathways, as these pathways, namely, extracellular signal-regulated kinases 1/2 (ERK1/2), AKT, and phosphatidylinositol-3 kinase, are involved in cell proliferation and differentiation. The p53 and nuclear factor kappa B were also investigated due to their ability to respond to intracellular stress. We observed that citral exerted antiproliferative and cytotoxic effects in B16F10; induced oxidative stress, DNA lesions, and p53 nuclear translocation; and reduced nitric oxide levels and nuclear factor kappa B, ERK1/2, and AKT. To investigate citral specificity, we used non-neoplastic human and murine cells, HaCaT (human skin keratinocytes) and NIH-3T3 cells (murine fibroblasts), and observed that although citral effects were not specific for cancer cells, non-neoplastic cells were more resistant to citral than B16F10. These findings highlight the potential clinical utility of citral in melanoma, with a mechanism of action involving the oxidative stress generation, nitric oxide depletion, and interference in signaling pathways related to cell proliferation.

Sodium and calcium currents shape action potentials in immature mouse inner hair cells

PubMed Central

Marcotti, Walter; Johnson, Stuart L; Rüsch, Alfons; Kros, Corné J

2003-01-01

Before the onset of hearing at postnatal day 12, mouse inner hair cells (IHCs) produce spontaneous and evoked action potentials. These spikes are likely to induce neurotransmitter release onto auditory nerve fibres. Since immature IHCs express both α1D (Cav1.3) Ca2+ and Na+ currents that activate near the resting potential, we examined whether these two conductances are involved in shaping the action potentials. Both had extremely rapid activation kinetics, followed by fast and complete voltage-dependent inactivation for the Na+ current, and slower, partially Ca2+-dependent inactivation for the Ca2+ current. Only the Ca2+ current is necessary for spontaneous and induced action potentials, and 29 % of cells lacked a Na+ current. The Na+ current does, however, shorten the time to reach the action-potential threshold, whereas the Ca2+ current is mainly involved, together with the K+ currents, in determining the speed and size of the spikes. Both currents increased in size up to the end of the first postnatal week. After this, the Ca2+ current reduced to about 30 % of its maximum size and persisted in mature IHCs. The Na+ current was downregulated around the onset of hearing, when the spiking is also known to disappear. Although the Na+ current was observed as early as embryonic day 16.5, its role in action-potential generation was only evident from just after birth, when the resting membrane potential became sufficiently negative to remove a sizeable fraction of the inactivation (half inactivation was at −71 mV). The size of both currents was positively correlated with the developmental change in action-potential frequency. PMID:12937295

Canonical transient receptor potential channel 2 (TRPC2): old name-new games. Importance in regulating of rat thyroid cell physiology.

PubMed

Törnquist, Kid; Sukumaran, Pramod; Kemppainen, Kati; Löf, Christoffer; Viitanen, Tero

2014-11-01

In addition to the TSH-cyclic AMP signalling pathway, calcium signalling is of crucial importance in thyroid cells. Although the importance of calcium signalling has been thoroughly investigated for several decades, the nature of the calcium channels involved in signalling is unknown. In a recent series of investigations using the well-studied rat thyroid FRTL-5 cell line, we showed that these cells exclusively express the transient receptor potential canonical 2 (TRPC2) channel. Our results suggested that the TRPC2 channel is of significant importance in regulating thyroid cell function. These investigations were the first to show that thyroid cells express a member of the TRPC family of ion channels. In this review, we will describe the importance of the TRPC2 channel in regulating TSH receptor expression, thyroglobulin maturation, intracellular calcium and iodide homeostasis and that the channel also regulates thyroid cell proliferation.

Elongation Factor-1a is a novel protein associated with host cell invasion and a potential protective antigen of Cryptosporidium parvum*

USDA-ARS?s Scientific Manuscript database

The phylum Apicomplexa comprises obligate intracellular parasites that infect vertebrates. All invasive forms of Apicomplexa possess a unique complex of organelles at the anterior end, referred to as the apical complex, which is involved in host cell invasion. Previously, we generated the chicken m...

Production Of Human Antibodies

NASA Technical Reports Server (NTRS)

Sammons, David W.; Neil, Garry A.

1993-01-01

Process for making human monoclonal antibodies based on combination of techniques. Antibodies made active against specific antigen. Process involves in vivo immunization of human B lymphocyte cells in mice. B cells of interest enriched in vitro before fusion. Method potentially applicable to any antigen. Does not rely on use of Epstein-Barr virus at any step. Human lymphocytes taken from any source.

"Cancer Cell Biology:" A Student-Centered Instructional Module Exploring the Use of Multimedia to Enrich Interactive, Constructivist Learning of Science

ERIC Educational Resources Information Center

Bockholt, Susanne M.; West, J. Paige; Bollenbacher, Walter E.

2003-01-01

Multimedia has the potential of providing bioscience education novel learning environments and pedagogy applications to foster student interest, involve students in the research process, advance critical thinking/problem-solving skills, and develop conceptual understanding of biological topics. "Cancer Cell Biology," an interactive, multimedia,…

Accommodation of powdery mildew fungi in intact plant cells.

PubMed

Eichmann, Ruth; Hückelhoven, Ralph

2008-01-01

Parasitic powdery mildew fungi have to overcome basic resistance and manipulate host cells to establish a haustorium as a functional feeding organ in a host epidermal cell. Currently, it is of central interest how plant factors negatively regulate basal defense or whether they even support fungal development in compatible interactions. Additionally, creation of a metabolic sink in infected cells may involve host activity. Here, we review the current progress in understanding potential fungal targets for host reprogramming and nutrient acquisition.

Neurotoxicity Induced by Bupivacaine via T-Type Calcium Channels in SH-SY5Y Cells

PubMed Central

Wen, Xianjie; Xu, Shiyuan; Liu, Hongzhen; Zhang, Quinguo; Liang, Hua; Yang, Chenxiang; Wang, Hanbing

2013-01-01

There is concern regarding neurotoxicity induced by the use of local anesthetics. A previous study showed that an overload of intracellular calcium is involved in the neurotoxic effect of some anesthetics. T-type calcium channels, which lower the threshold of action potentials, can regulate the influx of calcium ions. We hypothesized that T-type calcium channels are involved in bupivacaine-induced neurotoxicity. In this study, we first investigated the effects of different concentrations of bupivacaine on SH-SY5Y cell viability, and established a cell injury model with 1 mM bupivacaine. The cell viability of SH-SY5Y cells was measured following treatment with 1 mM bupivacaine and/or different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride, an antagonist of T-type calcium channels for 24 h. In addition, we monitored the release of lactate dehydrogenase, cytosolic Ca2+ ([Ca2+]i), cell apoptosis and caspase-3 expression. SH-SY5Y cells pretreated with different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride improved cell viability, reduced lactate dehydrogenase release, inhibited apoptosis, and reduced caspase-3 expression following bupivacaine exposure. However, the protective effect of NNC 55-0396 dihydrochloride plateaued. Overall, our results suggest that T-type calcium channels may be involved in bupivacaine neurotoxicity. However, identification of the specific subtype of T calcium channels involved requires further investigation. PMID:23658789

Distinct pH regulation of slow and rapid anion channels at the plasma membrane of Arabidopsis thaliana hypocotyl cells.

PubMed

Colcombet, Jean; Lelièvre, Françoise; Thomine, Sébastien; Barbier-Brygoo, Hélène; Frachisse, Jean-Marie

2005-07-01

Variations in both intracellular and extracellular pH are known to be involved in a wealth of physiological responses. Using the patch-clamp technique on Arabidopsis hypocotyl cells, it is shown that rapid-type and slow-type anion channels at the plasma membrane are both regulated by pH via distinct mechanisms. Modifications of pH modulate the voltage-dependent gating of the rapid channel. While intracellular alkalinization facilitates channel activation by shifting the voltage gate towards negative potentials, extracellular alkalinization shifts the activation threshold to more positive potentials, away from physiological resting membrane potentials. By contrast, pH modulates slow anion channel activity in a voltage-independent manner. Intracellular acidification and extracellular alkalinization increase slow anion channel currents. The possible role of these distinct modulations in physiological processes involving anion efflux and modulation of extracellular and/or intracellular pH, such as elicitor and ABA signalling, are discussed.

Cellular Aspects of Prion Replication In Vitro

PubMed Central

Grassmann, Andrea; Wolf, Hanna; Hofmann, Julia; Graham, James; Vorberg, Ina

2013-01-01

Prion diseases or transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative disorders in mammals that are caused by unconventional agents predominantly composed of aggregated misfolded prion protein (PrP). Prions self-propagate by recruitment of host-encoded PrP into highly ordered β-sheet rich aggregates. Prion strains differ in their clinical, pathological and biochemical characteristics and are likely to be the consequence of distinct abnormal prion protein conformers that stably replicate their alternate states in the host cell. Understanding prion cell biology is fundamental for identifying potential drug targets for disease intervention. The development of permissive cell culture models has greatly enhanced our knowledge on entry, propagation and dissemination of TSE agents. However, despite extensive research, the precise mechanism of prion infection and potential strain effects remain enigmatic. This review summarizes our current knowledge of the cell biology and propagation of prions derived from cell culture experiments. We discuss recent findings on the trafficking of cellular and pathologic PrP, the potential sites of abnormal prion protein synthesis and potential co-factors involved in prion entry and propagation. PMID:23340381

Arginine dependence of tumor cells: targeting a chink in cancer’s armor

PubMed Central

Patil, MD; Bhaumik, J; Babykutty, S; Banerjee, UC; Fukumura, D

2017-01-01

Arginine, one among the 20 most common natural amino acids, has a pivotal role in cellular physiology as it is being involved in numerous cellular metabolic and signaling pathways. Dependence on arginine is diverse for both tumor and normal cells. Because of decreased expression of argininosuccinate synthetase and/or ornithine transcarbamoylase, several types of tumor are auxotrophic for arginine. Deprivation of arginine exploits a significant vulnerability of these tumor cells and leads to their rapid demise. Hence, enzyme-mediated arginine depletion is a potential strategy for the selective destruction of tumor cells. Arginase, arginine deiminase and arginine decarboxylase are potential enzymes that may be used for arginine deprivation therapy. These arginine catabolizing enzymes not only reduce tumor growth but also make them susceptible to concomitantly administered anti-cancer therapeutics. Most of these enzymes are currently under clinical investigations and if successful will potentially be advanced as anti-cancer modalities. PMID:27109103

Involvement of matrix metalloproteinase-13 in stromal-cell-derived factor 1 alpha-directed invasion of human basal cell carcinoma cells.

PubMed

Chu, C-Y; Cha, S-T; Chang, C-C; Hsiao, C-H; Tan, C-T; Lu, Y-C; Jee, S-H; Kuo, M-L

2007-04-12

Basal cell carcinoma (BCC) is one of the most common skin neoplasms in humans and is usually characterized by local aggressiveness with little metastatic potential, although deep invasion, recurrence, and regional and distant metastases may occur. Here, we studied the mechanism of BCC invasion. We found that human BCC tissues and a BCC cell line had significant expression of CXCR4, which was higher in invasive than non-invasive BCC types. Further, of 19 recurrent tumors among 390 BCCs diagnosed during the past 12 years, 17/19 (89.5%) had high CXCR4 expression. We found that the CXCR4 ligand, stromal-cell-derived factor 1alpha (SDF-1alpha), directed BCC invasion and that this was mediated by time-dependent upregulation of mRNA expression and gelatinase activity of matrix metalloproteinase-13 (MMP-13). The transcriptional regulation of MMP-13 by SDF-1alpha was mediated by phosphorylation of extracellular signal-related kinase 1/2 and activation of the AP-1 component c-Jun. Finally, CXCR4-transfected BCC cells injected into nude mice induced aggressive BCCs that co-expressed CXCR4 and MMP-13. The identification of SDF-1alpha/CXCR4 as an important factor in BCC invasiveness may contribute insight into mechanisms involved in the aggressive potential of human BCC and may improve therapy for invasive BCCs.

Electromotive force measurements on cells involving beta-alumina solid electrolyte

NASA Technical Reports Server (NTRS)

Choudhury, N. S.

1973-01-01

Open-circuit emf measurements have been made to demonstrate that a two-phase, polycrystalline mixture of beta-alumina and alpha-alumina could be used as a solid electrolyte in galvanic cells with reversible electrodes fixing oxygen or aluminum chemical potentials. These measurements indicate that such a two-phase solid electrolyte may be used to monitor oxygen chemical potentials as low as that corresponding to Al and Al2O3 coexistence (potentials of about 10 to the minus 47th power atm at 1000 K). The activity of Na2O in beta-alumina in coexistence with alpha-alumina was also determined by emf measurements.

The impact of oil spill to lung health – insights from an RNA-seq study of human airway epithelial cells

PubMed Central

Liu, Yao-Zhong; Roy-Engel, Astrid M; Baddoo, Melody C; Flemington, Erik K; Wang, Guangdi; Wang, He

2015-01-01

The Deepwater Horizon oil spill (BP oil spill) in the Gulf of Mexico was a unique disaster event, where a huge amount of oil spilled from the sea bed and a large volume of dispersants were applied to clean the spill. The operation lasted for almost three months and involved >50,000 workers. The potential health hazards to these workers may be significant as previous research suggested an association of persistent respiratory symptoms with exposure to oil and oil dispersants. To reveal the potential effects of oil and oil dispersants on the respiratory system at the molecular level, we evaluated the transcriptomic profile of human airway epithelial cells grown under treatment of crude oil, the dispersants Corexit 9500 and Corexit 9527 and oil-dispersant mixtures. We identified a very strong effect of Corexit 9500 treatment, with 84 genes (response genes) differentially expressed in treatment vs. control samples. We discovered an interactive effect of oil-dispersant mixtures; while no response gene was found for Corexit 9527 treatment alone, cells treated with Corexit 9527 + oil mixture showed an increased number of response genes (46 response genes), suggesting a synergic effect of 9527 with oil on airway epithelial cells. Through GO (gene ontology) functional term and pathway-based analysis, we identified upregulation of gene sets involved in angiogenesis and immune responses and downregulation of gene sets involved in cell junctions and steroid synthesis as the prevailing transcriptomic signatures in the cells treated with Corexit 9500, oil or Corexit 9500 + oil mixture. Interestingly, these key molecular signatures coincide with important pathological features observed in common lung diseases, such as asthma, cystic fibrosis and chronic obstructive pulmonary disease. Our study provides mechanistic insights into the detrimental effects of oil and oil dispersants to the respiratory system and suggests significant health impacts of the recent BP oil spill to those people involved in the cleaning operation. PMID:26692141

Circular RNAs: Regulators of Cancer-Related Signaling Pathways and Potential Diagnostic Biomarkers for Human Cancers

PubMed Central

Yang, Zuozhang; Xie, Lin; Han, Lei; Qu, Xin; Yang, Yihao; Zhang, Ya; He, Zewei; Wang, Yu; Li, Jing

2017-01-01

Circular RNAs (circRNAs) are newly discovered endogenous non-coding RNAs featuring structural stability, high abundance, and tissue-specific expression. CircRNAs are prevalent and conserved in mammalian cells. They are involved in cellular processes and regulate gene expression at the transcriptional or post-transcriptional level by interacting with microRNAs (miRNAs) and other molecules. Recent studies have shown that circRNAs play an important role in the progression of various human diseases including atherosclerosis, nervous system disorders, diabetes, and cancer. In this review, we summarize the advances on endogenous circRNAs in eukaryotic cells and elucidate their diagnostic and prognostic significance in human cancers. Especially, we highlight the involvement of circRNAs in signal transduction pathways as well as their clinical potential to serve as biomarkers. PMID:28839467

Down-regulation of 21A Alu RNA as a tool to boost proliferation maintaining the tissue regeneration potential of progenitor cells

PubMed Central

Gigoni, Arianna; Costa, Delfina; Gaetani, Massimiliano; Tasso, Roberta; Villa, Federico; Florio, Tullio; Pagano, Aldo

2016-01-01

ABSTRACT 21A is an Alu non-coding (nc) RNA transcribed by RNA polymerase (pol) III. While investigating the biological role of 21A ncRNA we documented an inverse correlation between its expression level and the rate of cell proliferation. The downregulation of this ncRNA not only caused a boost in cell proliferation, but was also associated to a transient cell dedifferentiation, suggesting a possible involvement of this RNA in cell dedifferentiation/reprogramming. In this study, we explored the possibility to enhance proliferation and dedifferentiation of cells of interest, by 21A down-regulation, using a mixture of chemically modified Anti-21A RNAs. Our results confirmed the validity of this approach that allows the amplification of specific cell populations, in a controlled manner and without inducing permanent effects. In addition to induce cell proliferation, the procedure did not decrease the tissue regeneration potential of progenitor cells in two different cell systems. PMID:27494068

Down-regulation of 21A Alu RNA as a tool to boost proliferation maintaining the tissue regeneration potential of progenitor cells.

PubMed

Gigoni, Arianna; Costa, Delfina; Gaetani, Massimiliano; Tasso, Roberta; Villa, Federico; Florio, Tullio; Pagano, Aldo

2016-09-16

21A is an Alu non-coding (nc) RNA transcribed by RNA polymerase (pol) III. While investigating the biological role of 21A ncRNA we documented an inverse correlation between its expression level and the rate of cell proliferation. The downregulation of this ncRNA not only caused a boost in cell proliferation, but was also associated to a transient cell dedifferentiation, suggesting a possible involvement of this RNA in cell dedifferentiation/reprogramming. In this study, we explored the possibility to enhance proliferation and dedifferentiation of cells of interest, by 21A down-regulation, using a mixture of chemically modified Anti-21A RNAs. Our results confirmed the validity of this approach that allows the amplification of specific cell populations, in a controlled manner and without inducing permanent effects. In addition to induce cell proliferation, the procedure did not decrease the tissue regeneration potential of progenitor cells in two different cell systems.

Calcium-Induced Calcium Release during Action Potential Firing in Developing Inner Hair Cells

PubMed Central

Iosub, Radu; Avitabile, Daniele; Grant, Lisa; Tsaneva-Atanasova, Krasimira; Kennedy, Helen J.

2015-01-01

In the mature auditory system, inner hair cells (IHCs) convert sound-induced vibrations into electrical signals that are relayed to the central nervous system via auditory afferents. Before the cochlea can respond to normal sound levels, developing IHCs fire calcium-based action potentials that disappear close to the onset of hearing. Action potential firing triggers transmitter release from the immature IHC that in turn generates experience-independent firing in auditory neurons. These early signaling events are thought to be essential for the organization and development of the auditory system and hair cells. A critical component of the action potential is the rise in intracellular calcium that activates both small conductance potassium channels essential during membrane repolarization, and triggers transmitter release from the cell. Whether this calcium signal is generated by calcium influx or requires calcium-induced calcium release (CICR) is not yet known. IHCs can generate CICR, but to date its physiological role has remained unclear. Here, we used high and low concentrations of ryanodine to block or enhance CICR to determine whether calcium release from intracellular stores affected action potential waveform, interspike interval, or changes in membrane capacitance during development of mouse IHCs. Blocking CICR resulted in mixed action potential waveforms with both brief and prolonged oscillations in membrane potential and intracellular calcium. This mixed behavior is captured well by our mathematical model of IHC electrical activity. We perform two-parameter bifurcation analysis of the model that predicts the dependence of IHCs firing patterns on the level of activation of two parameters, the SK2 channels activation and CICR rate. Our data show that CICR forms an important component of the calcium signal that shapes action potentials and regulates firing patterns, but is not involved directly in triggering exocytosis. These data provide important insights into the calcium signaling mechanisms involved in early developmental processes. PMID:25762313

Calcium-Induced calcium release during action potential firing in developing inner hair cells.

PubMed

Iosub, Radu; Avitabile, Daniele; Grant, Lisa; Tsaneva-Atanasova, Krasimira; Kennedy, Helen J

2015-03-10

In the mature auditory system, inner hair cells (IHCs) convert sound-induced vibrations into electrical signals that are relayed to the central nervous system via auditory afferents. Before the cochlea can respond to normal sound levels, developing IHCs fire calcium-based action potentials that disappear close to the onset of hearing. Action potential firing triggers transmitter release from the immature IHC that in turn generates experience-independent firing in auditory neurons. These early signaling events are thought to be essential for the organization and development of the auditory system and hair cells. A critical component of the action potential is the rise in intracellular calcium that activates both small conductance potassium channels essential during membrane repolarization, and triggers transmitter release from the cell. Whether this calcium signal is generated by calcium influx or requires calcium-induced calcium release (CICR) is not yet known. IHCs can generate CICR, but to date its physiological role has remained unclear. Here, we used high and low concentrations of ryanodine to block or enhance CICR to determine whether calcium release from intracellular stores affected action potential waveform, interspike interval, or changes in membrane capacitance during development of mouse IHCs. Blocking CICR resulted in mixed action potential waveforms with both brief and prolonged oscillations in membrane potential and intracellular calcium. This mixed behavior is captured well by our mathematical model of IHC electrical activity. We perform two-parameter bifurcation analysis of the model that predicts the dependence of IHCs firing patterns on the level of activation of two parameters, the SK2 channels activation and CICR rate. Our data show that CICR forms an important component of the calcium signal that shapes action potentials and regulates firing patterns, but is not involved directly in triggering exocytosis. These data provide important insights into the calcium signaling mechanisms involved in early developmental processes. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Cellular and molecular mechanisms of pomegranate juice-induced anti-metastatic effect on prostate cancer cells.

PubMed

Wang, Lei; Alcon, Andre; Yuan, Hongwei; Ho, Jeffrey; Li, Qi-Jing; Martins-Green, M

2011-07-01

Prostate cancer is the second leading cause of cancer-related deaths among US males. Pomegranate juice (PJ), a natural product, was shown in a clinical trial to inhibit progression of this disease. However, the underlying mechanisms involved in the anti-progression effects of PJ on prostate cancer remain unclear. Here we show that, in addition to causing cell death of hormone-refractory prostate cancer cells, PJ also increases cell adhesion and decreases cell migration of the cells that do not die. We hypothesized that PJ does so by stimulating the expression and/or activation of molecules that alter the cytoskeleton and the adhesion machinery of prostate cancer cells, resulting in enhanced cell adhesion and reduced cell migration. We took an integrative approach to these studies by using Affimetrix gene arrays to study gene expression, microRNA arrays to study the non-coding RNAs, molecules known to be disregulated in cancer cells, and Luminex Multiplex array assays to study the level of secreted pro-inflammatory cytokines/chemokines. PJ up-regulates genes involved in cell adhesion such as E-cadherin, intercellular adhesion molecule 1 (ICAM-1) and down-regulates genes involved in cell migration such as hyaluranan-mediated motility receptor (HMMR) and type I collagen. In addition, anti-invasive microRNAs such as miR-335, miR-205, miR-200, and miR-126, were up-regulated, whereas pro-invasive microRNA such as miR-21 and miR-373, were down-regulated. Moreover, PJ significantly reduced the level of secreted pro-inflammatory cytokines/chemokines such as IL-6, IL-12p40, IL-1β and RANTES, thereby having the potential to decrease inflammation and its impact on cancer progression. PJ also inhibits the ability of the chemokine SDF1α to chemoattract these cancer cells. SDF1α and its receptor CXCR4 are important in metastasis of cancer cells to the bone. Discovery of the mechanisms by which this enhanced adhesion and reduced migration are accomplished can lead to sophisticated and effective prevention of metastasis in prostate and potentially other cancers. This journal is © The Royal Society of Chemistry 2011

Sticholysin II-mediated cytotoxicity involves the activation of regulated intracellular responses that anticipates cell death.

PubMed

Soto, Carmen; Bergado, Gretchen; Blanco, Rancés; Griñán, Tania; Rodríguez, Hermis; Ros, Uris; Pazos, Fabiola; Lanio, María Eliana; Hernández, Ana María; Álvarez, Carlos

2018-05-01

Sticholysin II (StII) is a pore-forming toxin of biomedical interest that belongs to the actinoporin protein family. Sticholysins are currently under examination as an active immunomodulating component of a vaccinal platform against tumoral cells and as a key element of a nucleic acids delivery system to cell cytosol. These proteins form pores in the plasma membrane leading to ion imbalance and cell lysis. However, the intracellular mechanisms triggered by actinoporins upon binding to membranes and its consequences for cell death are barely understood. Here, we have examined the cytotoxicity and intracellular responses induced by StII upon binding to human B-cell lymphoma Raji in vitro. StII cytotoxicity involves a functional actin cytoskeleton, induces cellular swelling, lysis and the concomitant release of cytosol content. In addition, StII induces calcium release mainly from the Endoplasmic Reticulum, activates Mitogen-Activated Protein Kinase ERK and impairs mitochondrial membrane potential. Furthermore, StII stimulates the expression of receptor interacting protein kinase 1 (RIP1), normally related to different forms of regulated cell death such as apoptosis and necroptosis. In correspondence, necrostatin-1, an inhibitor of this kinase, reduces StII cytotoxicity. However, the mechanism of cell death activated by StII does not involve caspases activation, typical molecular features of apoptosis and pyroptosis. Our results suggest that, beyond pore-formation and cell lysis, StII-induced cytotoxicity could involve other regulated intracellular mechanisms connected to RIP1-MEK1/2 -ERK1/2- pathways. This opens new perspectives and challenges the general point of view that these toxins induce a completely unregulated mechanism of necrotic cell death. This study contributes to a better understanding of the molecular mechanisms involved in toxin-cell interaction and the implications for cell functioning, with connotation for the exploitations of these toxins in clinical settings. Copyright © 2018 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

Apoptosis of leukemia K562 and Molt-4 cells induced by emamectin benzoate involving mitochondrial membrane potential loss and intracellular Ca2+ modulation.

PubMed

Yun, Xinming; Rao, Wenbing; Xiao, Ciying; Huang, Qingchun

2017-06-01

Leukemia threatens millions of people's health and lives, and the pesticide-induced leukemia has been increasingly concerned because of the etiologic exposure. In this paper, cytotoxic effect of emamectin benzoate (EMB), an excellent natural-product insecticide, was evaluated through monitoring cell viability, cell apoptosis, mitochondrial membrane potential and intracellular Ca 2+ concentration ([Ca 2+ ] i ) in leukemia K562 and Molt-4 cells. Following the exposure to EMB, cell viability was decreased and positive apoptosis of K562 and Molt-4 cells was increased in a concentration- and time- dependent fashion. In the treatment of 10μM EMB, apoptotic cells accounted for 93.0% to K562 cells and 98.9% to Molt-4 cells based on the control, meanwhile, 63.47% of K562 cells and 81.15% of Molt-4 cells exhibited late apoptotic and necrotic features with damaged cytoplasmic membrane. 48h exposure to 10μM EMB increased significantly the great number of cells with mitochondrial membrane potential (MMP) loss, and the elevation of [Ca 2+ ] i level was peaked and persisted within 70s in K562 cells whilst 50s in Molt-4 cells. Moreover, a stronger cytotoxicity of EMB was further observed than that of imatinib. The results authenticate the efficacious effect of EMB as a potential anti-leukemia agent and an inconsistency with regard to insecticide-induced leukemia. Copyright © 2017 Elsevier B.V. All rights reserved.

The role of lysosomes in BDE 47-mediated activation of mitochondrial apoptotic pathway in HepG2 cells.

PubMed

Liu, Xiaohui; Wang, Jian; Lu, Chengquan; Zhu, Chunyan; Qian, Bo; Li, Zhenwei; Liu, Chang; Shao, Jing; Yan, Jinsong

2015-04-01

Polybrominated diphenyl ethers (PBDEs) are a group of widely used flame retardants. The rising presence of PBDEs in human tissues has received considerable concerns with regard to potential health risks. While the mitochondrial-apoptotic pathway has been suggested in PBDEs-induced apoptosis, the role of lysosomes is yet to be understood. In the present study, HepG2 cells were exposed to BDE 47 at various concentrations and durations to establish the causal and temporal relationships among various cellular events, such as cell viability, reactive oxygen species (ROS), mitochondrial membrane potential (MMP), apoptosis, and expression of cytochrome C and caspase 3. The involvement of lysosomes was simultaneously studied by evaluating lysosomal membrane permeability (LMP) and changes in the expression of cathepsin B, a lysosome hydrolase. In addition, a cathepsin B inhibitor (10 μM CA-074) was used to determine the involvement of lysosomes and potential interactions between lysosomes and mitochondria. Our results showed that ROS production was an initial response of HepG2 to BDE 47 exposure, followed by a decreased MMP; a loss of MMP caused additional ROS generation which acted to induce LMP; an increased LMP resulted in a release of cathepsin B which aggravated the loss of MMP leading to release of cytochrome C and caspase 3 and subsequent apoptosis. Pretreatment with CA-074 did not abolish the initial ROS generation, however, all downstream events were dramatically alleviated. Taken together, our data indicate that lysosomes might be involved in BDE 47-mediated mitochondrial-apoptotic pathway in HepG2 cells, possibly through feedback interactions between mitochondria and lysosomes. Copyright © 2014 Elsevier Ltd. All rights reserved.

Distinct Responses of Stem Cells to Telomere Uncapping-A Potential Strategy to Improve the Safety of Cell Therapy.

PubMed

Liu, Chang Ching; Ma, Dong Liang; Yan, Ting-Dong; Fan, XiuBo; Poon, Zhiyong; Poon, Lai-Fong; Goh, Su-Ann; Rozen, Steve G; Hwang, William Ying Khee; Tergaonkar, Vinay; Tan, Patrick; Ghosh, Sujoy; Virshup, David M; Goh, Eyleen L K; Li, Shang

2016-10-01

In most human somatic cells, the lack of telomerase activity results in progressive telomere shortening during each cell division. Eventually, DNA damage responses triggered by critically short telomeres induce an irreversible cell cycle arrest termed replicative senescence. However, the cellular responses of human pluripotent stem cells to telomere uncapping remain unknown. We generated telomerase knockout human embryonic stem (ES) cells through gene targeting. Telomerase inactivation in ES cells results in progressive telomere shortening. Telomere DNA damage in ES cells and neural progenitor cells induces rapid apoptosis when telomeres are uncapped, in contrast to fibroblast cells that enter a state of replicative senescence. Significantly, telomerase inactivation limits the proliferation capacity of human ES cells without affecting their pluripotency. By targeting telomerase activity, we can functionally separate the two unique properties of human pluripotent stem cells, namely unlimited self-renewal and pluripotency. We show that the potential of ES cells to form teratomas in vivo is dictated by their telomere length. By controlling telomere length of ES cells through telomerase inactivation, we can inhibit teratoma formation and potentially improve the safety of cell therapies involving terminally differentiated cells as well as specific progenitor cells that do not require sustained cellular proliferation in vivo, and thus sustained telomerase activity. Stem Cells 2016;34:2471-2484. © 2016 AlphaMed Press.

S-layers: principles and applications

PubMed Central

Sleytr, Uwe B; Schuster, Bernhard; Egelseer, Eva-Maria; Pum, Dietmar

2014-01-01

Monomolecular arrays of protein or glycoprotein subunits forming surface layers (S-layers) are one of the most commonly observed prokaryotic cell envelope components. S-layers are generally the most abundantly expressed proteins, have been observed in species of nearly every taxonomical group of walled bacteria, and represent an almost universal feature of archaeal envelopes. The isoporous lattices completely covering the cell surface provide organisms with various selection advantages including functioning as protective coats, molecular sieves and ion traps, as structures involved in surface recognition and cell adhesion, and as antifouling layers. S-layers are also identified to contribute to virulence when present as a structural component of pathogens. In Archaea, most of which possess S-layers as exclusive wall component, they are involved in determining cell shape and cell division. Studies on structure, chemistry, genetics, assembly, function, and evolutionary relationship of S-layers revealed considerable application potential in (nano)biotechnology, biomimetics, biomedicine, and synthetic biology. PMID:24483139

Systemic Regulation of the Age-Related Decline of Pancreatic β-Cell Replication

PubMed Central

Salpeter, Seth J.; Khalaileh, Abed; Weinberg-Corem, Noa; Ziv, Oren; Glaser, Benjamin; Dor, Yuval

2013-01-01

The frequency of pancreatic β-cell replication declines dramatically with age, potentially contributing to the increased risk of type 2 diabetes in old age. Previous studies have shown the involvement of cell-autonomous factors in this phenomenon, particularly the decline of polycomb genes and accumulation of p16/INK4A. Here, we demonstrate that a systemic factor found in the circulation of young mice is able to increase the proliferation rate of old pancreatic β-cells. Old mice parabiosed to young mice have increased β-cell replication compared with unjoined old mice or old mice parabiosed to old mice. In addition, we demonstrate that old β-cells transplanted into young recipients have increased replication rate compared with cells transplanted into old recipients; conversely, young β-cells transplanted into old mice decrease their replication rate compared with young cells transplanted into young recipients. The expression of p16/INK4A mRNA did not change in heterochronic parabiosis, suggesting the involvement of other pathways. We conclude that systemic factors contribute to the replicative decline of old pancreatic β-cells. PMID:23630298

[Cellular mechanism of the generation of spontaneous activity in gastric muscle].

PubMed

Nakamura, Eri; Kito, Yoshihiko; Fukuta, Hiroyasu; Yanai, Yoshimasa; Hashitani, Hikaru; Yamamoto, Yoshimichi; Suzuki, Hikaru

2004-03-01

In gastric smooth muscles, interstitial cells of Cajal (ICC) might be the pacemaker cells of spontaneous activities since ICC are rich in mitochondria and are connected with smooth muscle cells via gap junctions. Several types of ICC are distributed widely in the stomach wall. A group of ICC distributed in the myenteric layer (ICC-MY) were the pacemaker cells of gastrointestinal smooth muscles. Pacemaker potentials were generated in ICC-MY, and the potentials were conducted to circular smooth muscles to trigger slow waves and also conducted to longitudinal muscles to form follower potentials. In circular muscle preparations, interstitial cells distributed within muscle bundles (ICC-IM) produced unitary potentials, which were conducted to circular muscles to form slow potentials by summation. In mutant mice lacking inositol trisphosphate (IP(3)) receptor, slow waves were absent in gastric smooth muscles. The generation of spontaneous activity was impaired by the inhibition of Ca(2+)-release from internal stores through IP(3) receptors, inhibition of mitochondrial Ca(2+)-handling with proton pump inhibitors, and inhibition of ATP-sensitive K(+)-channels at the mitochondrial inner membrane. These results suggested that mitochondrial Ca(2+)-handling causes the generation of spontaneous activity in pacemaker cells. Possible involvement of protein kinase C (PKC) in the Ca(2+) signaling system was also suggested.

Two-dimensional computer simulation of EMVJ and grating solar cells under AMO illumination

NASA Technical Reports Server (NTRS)

Gray, J. L.; Schwartz, R. J.

1984-01-01

A computer program, SCAP2D (Solar Cell Analysis Program in 2-Dimensions), is used to evaluate the Etched Multiple Vertical Junction (EMVJ) and grating solar cells. The aim is to demonstrate how SCAP2D can be used to evaluate cell designs. The cell designs studied are by no means optimal designs. The SCAP2D program solves the three coupled, nonlinear partial differential equations, Poisson's Equation and the hole and electron continuity equations, simultaneously in two-dimensions using finite differences to discretize the equations and Newton's Method to linearize them. The variables solved for are the electrostatic potential and the hole and electron concentrations. Each linear system of equations is solved directly by Gaussian Elimination. Convergence of the Newton Iteration is assumed when the largest correction to the electrostatic potential or hole or electron quasi-potential is less than some predetermined error. A typical problem involves 2000 nodes with a Jacobi matrix of order 6000 and a bandwidth of 243.

Involvement of lateral septum in alcohol's dopamine-elevating effect in the rat.

PubMed

Jonsson, Susanne; Morud, Julia; Stomberg, Rosita; Ericson, Mia; Söderpalm, Bo

2017-01-01

Drugs of abuse share the ability to increase extracellular dopamine (DA) levels in the mesolimbic DA system. This effect has been linked to positive and reinforcing experiences of drug consumption and is presumed to be of importance for continued use, as well as for the development of dependence and addiction. Previous rat studies from our lab have implicated a neuronal circuitry involving glycine receptors in nucleus accumbens (nAc) and, secondarily, nicotinic acetylcholine receptors in the ventral tegmental area (VTA) in ethanol's (EtOH) DA-elevating effect. The work presented here, performed in male Wistar rats, suggests that the lateral septum (LS), which has previously been associated with different aspects of EtOH-related behaviour, is involved as well. In vivo microdialysis methodology demonstrated that blocking the generation of action potentials in LS using tetrodotoxin prevented a DA increase in nAc after accumbal EtOH perfusion. Retrograde tracing and polymerase chain reaction (PCR) were used to identify and characterize cells projecting to VTA from nAc/LS and from LS to nAc. Based on the PCR results, cells projecting from both LS/nAc to anterior VTA and from LS to nAc were mainly GABAergic neurons expressing glycine receptors, and these cells are presumed to be involved in mediating the DA-elevating effect of EtOH. These results provide further evidence implicating LS in the reinforcing effects of EtOH. Additional studies are needed to investigate LS involvement in EtOH consumption behaviour and its potential role in the development of dependence and addiction. © 2015 Society for the Study of Addiction.

Activation of the germ-cell potential of human bone marrow-derived cells by a chemical carcinogen

PubMed Central

Liu, Chunfang; Ma, Zhan; Xu, Songtao; Hou, Jun; Hu, Yao; Yu, Yinglu; Liu, Ruilai; Chen, Zhihong; Lu, Yuan

2014-01-01

Embryonic/germ cell traits are common in malignant tumors and are thought to be involved in malignant tumor behaviors. The reasons why tumors show strong embryonic/germline traits (displaced germ cells or gametogenic programming reactivation) are controversial. Here, we show that a chemical carcinogen, 3-methyl-cholanthrene (3-MCA), can trigger the germ-cell potential of human bone marrow-derived cells (hBMDCs). 3-MCA promoted the generation of germ cell-like cells from induced hBMDCs that had undergone malignant transformation, whereas similar results were not observed in the parallel hBMDC culture at the same time point. The malignant transformed hBMDCs spontaneously and more efficiently generated into germ cell-like cells even at the single-cell level. The germ cell-like cells from induced hBMDCs were similar to natural germ cells in many aspects, including morphology, gene expression, proliferation, migration, further development, and teratocarcinoma formation. Therefore, our results demonstrate that a chemical carcinogen can reactivate the germline phenotypes of human somatic tissue-derived cells, which might provide a novel idea to tumor biology and therapy. PMID:24998261

microRNA-449a functions as a tumor suppressor in neuroblastoma through inducing cell differentiation and cell cycle arrest

PubMed Central

Zhao, Zhenze; Ma, Xiuye; Sung, Derek; Li, Monica; Kosti, Adam; Lin, Gregory; Chen, Yidong; Pertsemlidis, Alexander; Hsiao, Tzu-Hung; Du, Liqin

2015-01-01

microRNA-449a (miR-449a) has been identified to function as a tumor suppressor in several types of cancers. However, the role of miR-449a in neuroblastoma has not been intensively investigated. We recently found that the overexpression of miR-449a significantly induces neuroblastoma cell differentiation, suggesting its potential tumor suppressor function in neuroblastoma. In this study, we further investigated the mechanisms underlying the tumor suppressive function of miR-449a in neuroblastoma. We observed that miR-449a inhibits neuroblastoma cell survival and growth through 2 mechanisms—inducing cell differentiation and cell cycle arrest. Our comprehensive investigations on the dissection of the target genes of miR-449a revealed that 3 novel targets- MFAP4, PKP4 and TSEN15 -play important roles in mediating its differentiation-inducing function. In addition, we further found that its function in inducing cell cycle arrest involves down-regulating its direct targets CDK6 and LEF1. To determine the clinical significance of the miR-449a-mediated tumor suppressive mechanism, we examined the correlation between the expression of these 5 target genes in neuroblastoma tumor specimens and the survival of neuroblastoma patients. Remarkably, we noted that high tumor expression levels of all the 3 miR-449a target genes involved in regulating cell differentiation, but not the target genes involved in regulating cell cycle, are significantly correlated with poor survival of neuroblastoma patients. These results suggest the critical role of the differentiation-inducing function of miR-449a in determining neuroblastoma progression. Overall, our study provides the first comprehensive characterization of the tumor-suppressive function of miR-449a in neuroblastoma, and reveals the potential clinical significance of the miR-449a-mediated tumor suppressive pathway in neuroblastoma prognosis. PMID:25760387

Visualization of gene expression in the live subject using the Na/I symporter as a reporter gene: applications in biotherapy

PubMed Central

Baril, Patrick; Martin-Duque, Pilar; Vassaux, Georges

2010-01-01

Biotherapies involve the utilization of antibodies, genetically modified viruses, bacteria or cells for therapeutic purposes. Molecular imaging has the potential to provide unique information that will guarantee their biosafety in humans and provide a rationale for the future development of new generations of reagents. In this context, non-invasive imaging of gene expression is an attractive prospect, allowing precise, spacio-temporal measurements of gene expression in longitudinal studies involving gene transfer vectors. With the emergence of cell therapies in regenerative medicine, it is also possible to track cells injected into subjects. In this context, the Na/I symporter (NIS) has been used in preclinical studies. Associated with a relevant radiotracer (123I-, 124I-, 99mTcO4-), NIS can be used to monitor gene transfer and the spread of selectively replicative viruses in tumours as well as in cells with a therapeutic potential. In addition to its imaging potential, NIS can be used as a therapeutic transgene through its ability to concentrate therapeutic doses of radionuclides in target cells. This dual property has applications in cancer treatment and could also be used to eradicate cells with therapeutic potential in the case of adverse events. Through experience acquired in preclinical studies, we can expect that non-invasive molecular imaging using NIS as a transgene will be pivotal for monitoring in vivo the exact distribution and pharmacodynamics of gene expression in a precise and quantitative way. This review highlights the applications of NIS in biotherapy, with a particular emphasis on image-guided radiotherapy, monitoring of gene and vector biodistribution and trafficking of stem cells. This article is part of a themed section on Imaging in Pharmacology. To view the editorial for this themed section visit http://dx.doi.org/10.1111/j.1476-5381.2010.00685.x PMID:19814733

Reduction of the tumorigenic potential of human retinoblastoma cell lines by TFF1 overexpression involves p53/caspase signaling and miR-18a regulation.

PubMed

Busch, Maike; Große-Kreul, Jan; Wirtz, Janina Jasmin; Beier, Manfred; Stephan, Harald; Royer-Pokora, Brigitte; Metz, Klaus; Dünker, Nicole

2017-08-01

Trefoil factor family (TFF) peptides have been shown to play a pivotal role in oncogenic transformation, tumorigenesis and metastasis by changing cell proliferation, apoptosis, migration and invasion behavior of various cancer cell lines. In the study presented, we investigated the effect of TFF1 overexpression on cell growth, viability, migration and tumorigenicity of different retinoblastoma (RB) cell lines. Transient TFF1 overexpression significantly increases RB cell apoptosis levels. Stable, lentiviral TFF1 overexpression likewise decreases RB cell viability, proliferation and growth and significantly increases apoptosis as revealed by WST-1 assays, BrdU and DAPI cell counts. TFF1-induced apoptosis is executed via cleaved caspase-3 activation as revealed by caspase blockage experiments and caspase-3 immunocytochemistry. Results from pG13-luciferase reporter assays and Western blot analyses indicate that TFF1-induced apoptosis is mediated through transcriptional activity of p53 with concurrently downregulated miR-18a expression. In ovo chicken chorioallantoic membrane (CAM) assays revealed that TFF1 overexpression significantly decreases the size of tumors forming from Y79 and RB355 cells and reduces the migration potential of RB355 cells. Differentially expressed genes and pathways involved in cancer progression were identified after TFF1 overexpression in Y79 cells by gene expression array analysis, underlining the effects on reduced tumorigenicity. TFF1 knockdown in RBL30 cells revealed caspase-3/7-independent apoptosis induction, but no changes on cell proliferation level. In summary, the in vitro and in vivo data demonstrate for the first time a tumor suppressor function of TFF1 in RB cells which is at least partly mediated by p53 activation and miR-18a downregulation. © 2017 UICC.

Complementation of non-tumorigenicity of HPV18-positive cervical carcinoma cells involves differential mRNA expression of cellular genes including potential tumor suppressor genes on chromosome 11q13.

PubMed

Kehrmann, Angela; Truong, Ha; Repenning, Antje; Boger, Regina; Klein-Hitpass, Ludger; Pascheberg, Ulrich; Beckmann, Alf; Opalka, Bertram; Kleine-Lowinski, Kerstin

2013-01-01

The fusion between human tumorigenic cells and normal human diploid fibroblasts results in non-tumorigenic hybrid cells, suggesting a dominant role for tumor suppressor genes in the generated hybrid cells. After long-term cultivation in vitro, tumorigenic segregants may arise. The loss of tumor suppressor genes on chromosome 11q13 has been postulated to be involved in the induction of the tumorigenic phenotype of human papillomavirus (HPV)18-positive cervical carcinoma cells and their derived tumorigenic hybrid cells after subcutaneous injection in immunocompromised mice. The aim of this study was the identification of novel cellular genes that may contribute to the suppression of the tumorigenic phenotype of non-tumorigenic hybrid cells in vivo. We used cDNA microarray technology to identify differentially expressed cellular genes in tumorigenic HPV18-positive hybrid and parental HeLa cells compared to non-tumorigenic HPV18-positive hybrid cells. We detected several as yet unknown cellular genes that play a role in cell differentiation, cell cycle progression, cell-cell communication, metastasis formation, angiogenesis, antigen presentation, and immune response. Apart from the known differentially expressed genes on 11q13 (e.g., phosphofurin acidic cluster sorting protein 1 (PACS1) and FOS ligand 1 (FOSL1 or Fra-1)), we detected novel differentially expressed cellular genes located within the tumor suppressor gene region (e.g., EGF-containing fibulin-like extracellular matrix protein 2 (EFEMP2) and leucine rich repeat containing 32 (LRRC32) (also known as glycoprotein-A repetitions predominant (GARP)) that may have potential tumor suppressor functions in this model system of non-tumorigenic and tumorigenic HeLa x fibroblast hybrid cells. Copyright © 2013 Elsevier Inc. All rights reserved.

The therapeutic potential of cell cycle targeting in multiple myeloma.

PubMed

Maes, Anke; Menu, Eline; Veirman, Kim De; Maes, Ken; Vand Erkerken, Karin; De Bruyne, Elke

2017-10-27

Proper cell cycle progression through the interphase and mitosis is regulated by coordinated activation of important cell cycle proteins (including cyclin-dependent kinases and mitotic kinases) and several checkpoint pathways. Aberrant activity of these cell cycle proteins and checkpoint pathways results in deregulation of cell cycle progression, which is one of the key hallmarks of cancer. Consequently, intensive research on targeting these cell cycle regulatory proteins identified several candidate small molecule inhibitors that are able to induce cell cycle arrest and even apoptosis in cancer cells. Importantly, several of these cell cycle regulatory proteins have also been proposed as therapeutic targets in the plasma cell malignancy multiple myeloma (MM). Despite the enormous progress in the treatment of MM the past 5 years, MM still remains most often incurable due to the development of drug resistance. Deregulated expression of the cyclins D is observed in virtually all myeloma patients, emphasizing the potential therapeutic interest of cyclin-dependent kinase inhibitors in MM. Furthermore, other targets have also been identified in MM, such as microtubules, kinesin motor proteins, aurora kinases, polo-like kinases and the anaphase promoting complex/cyclosome. This review will provide an overview of the cell cycle proteins and checkpoint pathways deregulated in MM and discuss the therapeutic potential of targeting proteins or protein complexes involved in cell cycle control in MM.

Thinking outside the liver: Induced pluripotent stem cells for hepatic applications

PubMed Central

Subba Rao, Mekala; Sasikala, Mitnala; Reddy, D Nageshwar

2013-01-01

The discovery of induced pluripotent stem cells (iPSCs) unraveled a mystery in stem cell research, after identification of four re-programming factors for generating pluripotent stem cells without the need of embryos. This breakthrough in generating iPSCs from somatic cells has overcome the ethical issues and immune rejection involved in the use of human embryonic stem cells. Hence, iPSCs form a great potential source for developing disease models, drug toxicity screening and cell-based therapies. These cells have the potential to differentiate into desired cell types, including hepatocytes, under in vitro as well as under in vivo conditions given the proper microenvironment. iPSC-derived hepatocytes could be useful as an unlimited source, which can be utilized in disease modeling, drug toxicity testing and producing autologous cell therapies that would avoid immune rejection and enable correction of gene defects prior to cell transplantation. In this review, we discuss the induction methods, role of reprogramming factors, and characterization of iPSCs, along with hepatocyte differentiation from iPSCs and potential applications. Further, we discuss the location and detection of liver stem cells and their role in liver regeneration. Although tumor formation and genetic mutations are a cause of concern, iPSCs still form a promising source for clinical applications. PMID:23801830

Thinking outside the liver: induced pluripotent stem cells for hepatic applications.

PubMed

Subba Rao, Mekala; Sasikala, Mitnala; Nageshwar Reddy, D

2013-06-14

The discovery of induced pluripotent stem cells (iPSCs) unraveled a mystery in stem cell research, after identification of four re-programming factors for generating pluripotent stem cells without the need of embryos. This breakthrough in generating iPSCs from somatic cells has overcome the ethical issues and immune rejection involved in the use of human embryonic stem cells. Hence, iPSCs form a great potential source for developing disease models, drug toxicity screening and cell-based therapies. These cells have the potential to differentiate into desired cell types, including hepatocytes, under in vitro as well as under in vivo conditions given the proper microenvironment. iPSC-derived hepatocytes could be useful as an unlimited source, which can be utilized in disease modeling, drug toxicity testing and producing autologous cell therapies that would avoid immune rejection and enable correction of gene defects prior to cell transplantation. In this review, we discuss the induction methods, role of reprogramming factors, and characterization of iPSCs, along with hepatocyte differentiation from iPSCs and potential applications. Further, we discuss the location and detection of liver stem cells and their role in liver regeneration. Although tumor formation and genetic mutations are a cause of concern, iPSCs still form a promising source for clinical applications.

Epstein-Barr virus-infected Akata cells are sensitive to histone deacetylase inhibitor TSA-provoked apoptosis.

PubMed

Kook, Sung-Ho; Son, Young-Ok; Han, Seong-Kyu; Lee, Hyung-Soon; Kim, Beom-Tae; Jang, Yong-Suk; Choi, Ki-Choon; Lee, Keun-Soo; Kim, So-Soon; Lim, Ji-Young; Jeon, Young-Mi; Kim, Jong-Ghee; Lee, Jeong-Chae

2005-11-30

Epstein-Barr virus (EBV) infects more than 90 % of the world's population and has a potential oncogenic nature. A histone deacetylase (HDAC) inhibitor, trichostatin A (TSA), has shown potential ability in cancer chemoprevention and treatment, but its effect on EBV-infected Akata cells has not been examined. This study investigated the effect of TSA on the proliferation and apoptosis of the cells. TSA inhibited cell growth and induced cytotoxicity in the EBV-infected Akata cells. TSA treatment sensitively induced apoptosis in the cell, which was demonstrated by the increased number of positively stained cells in the TUNEL assay, the migration of many cells to the sub-G0/G1 phase in flow cytometric analysis, and the ladder formation of genomic DNA. Western blot analysis showed that caspase-dependent pathways are involved in the TSA-induced apoptosis of EBV-infected Akata cells. Overall, this study shows that EBV-infected B lymphomas are quite sensitive to TSA-provoked apoptosis.

Changes in vacuolation in the root apex cells of soybean seedlings in microgravity

NASA Technical Reports Server (NTRS)

Klymchuk, D. O.; Kordyum, E. L.; Vorobyova, T. V.; Chapman, D. K.; Brown, C. S.

2003-01-01

Changes in the vacuolation in root apex cells of soybean (Glycine max L. [Merr.]) seedlings grown in microgravity were investigated. Spaceflight and ground control seedlings were grown in the absence or presence of KMnO4 (to remove ethylene) for 6 days. After landing, in order to study of cell ultrastructure and subcellular free calcium ion distribution, seedling root apices were fixed in 2.5% (w/v) glutaraldehyde in 0.1 M cacodylate buffer and 2% (w/v) glutaraldehyde, 2.5% (w/v) formaldehyde, 2% (w/v) potassium antimonate K[Sb(OH)6] in 0.1 M K2HPO4 buffer with an osmolarity (calculated theoretically) of 0.45 and 1.26 osmol. The concentrations of ethylene in all spaceflight canisters were significantly higher than in the ground control canisters. Seedling growth was reduced in the spaceflight-exposed plants. Additionally, the spaceflight-exposed plants exhibited progressive vacuolation in the root apex cells, particularly in the columella cells, to a greater degree than the ground controls. Plasmolysis was observed in columella cells of spaceflight roots fixed in solutions with relatively high osmolarity (1.26 osmol). The appearance of plasmolysis permitted the evaluation of the water status of cells. The water potential of the spaceflight cells was higher than the surrounding fixative solution. A decrease in osmotic potential and/or an increase in turgor potential may have induced increases in cell water potential. However, the plasmolysed (i.e. non-turgid) cells implied that increases in water potential were accompanied with a decrease in osmotic potential. In such cells changes in vacuolation may have been involved to maintain turgor pressure or may have been a result of intensification of other vacuolar functions like digestion and storage. c2003 COSPAR. Published by Elsevier Ltd. All rights reserved.

In Vitro Cardiomyogenic Potential of Human Amniotic Fluid Stem Cells

PubMed Central

Guan, Xuan; Delo, Dawn M.; Atala, Anthony; Soker, Shay

2010-01-01

Stem cell therapy for damaged cardiac tissue is currently limited by a number of factors, including the inability to obtain sufficient cell numbers, the potential tumorigenicity of certain types of stem cells, and the possible link between stem cell therapy and the development of malignant arrhythmias. In this study, we investigated whether human amniotic fluid-derived stem (hAFS) cells could be a potential source of cells for cardiac cell therapy by testing the in vitro differentiation capabilities. Undifferentiated hAFS cells express several cardiac genes, including the transcription factor mef2, the gap junction connexin43, and H- and N-cadherin. A 24-hour incubation with 5-aza-2′–deoxycytidine (5-AZA-dC) induced hAFS cell differentiation along the cardiac lineage. Evidence for this differentiation included morphological changes, up-regulation of cardiac-specific genes (cardiac troponin I and cardiac troponin T) and redistribution of connexin43, as well as down-regulation of the stem cell marker SRY-box 2 (sox2). When co-cultured with neonatal rat cardiomyocytes (NRCs), hAFS cells formed both mechanical and electrical connections with the NRCs. Dye transfer experiments showed that calcein dye could be transferred from NRCs to hAFS cells through cellular connections. The gap junction connexin 43 likely involved in the communication between the two cell types, because 12-O-Tetradecanoylphorbol 13-acetate (TPA) could partially block cellular crosstalk. We conclude that hAFS cells can be differentiated into a cardiomyocyte-like phenotype and can establish functional communication with NRCs. Thus, hAFS cells may potentially be used for cardiac cell therapy. PMID:20687122

In vitro cardiomyogenic potential of human amniotic fluid stem cells.

PubMed

Guan, Xuan; Delo, Dawn M; Atala, Anthony; Soker, Shay

2011-03-01

Stem cell therapy for damaged cardiac tissue is currently limited by a number of factors, including inability to obtain sufficient cell numbers, the potential tumorigenicity of certain types of stem cells and the possible link between stem cell therapy and the development of malignant arrhythmias. In this study, we investigated whether human amniotic fluid-derived stem (hAFS) cells could be a potential source of cells for cardiac cell therapy, by testing the in vitro differentiation capabilities. Undifferentiated hAFS cells express several cardiac genes, including the transcription factor mef2, the gap junction connexin43, and H- and N-cadherin. A 24 h incubation with 5-aza-2'-deoxycytidine (5-AZA-dC) induced hAFS cell differentiation along the cardiac lineage. Evidence for this differentiation included morphological changes, upregulation of cardiac-specific genes (cardiac troponin I and cardiac troponin T) and redistribution of connexin43, as well as downregulation of the stem cell marker SRY-box 2 (sox2). When co-cultured with neonatal rat cardiomyocytes (NRCs), hAFS cells formed both mechanical and electrical connections with the NRCs. Dye transfer experiments showed that calcein dye could be transferred from NRCs to hAFS cells through cellular connections. The gap junction connexin43 likely involved in the communication between the two cell types, because 12-O-tetradecanoylphorbol 13-acetate (TPA) could partially block cellular crosstalk. We conclude that hAFS cells can be differentiated into a cardiomyocyte-like phenotype and can establish functional communication with NRCs. Thus, hAFS cells may potentially be used for cardiac cell therapy. Copyright © 2010 John Wiley & Sons, Ltd.

Evasion of anti-growth signaling: a key step in tumorigenesis and potential target for treatment and prophylaxis by natural compounds

PubMed Central

Amin, A.R.M. Ruhul; Karpowicz, Phillip A.; Carey, Thomas E.; Arbiser, Jack; Nahta, Rita; Chen, Zhuo G.; Dong, Jin-Tang; Kucuk, Omer; Khan, Gazala N.; Huang, Gloria S.; Mi, Shijun; Lee, Ho-Young; Reichrath, Joerg; Honoki, Kanya; Georgakilas, Alexandros G.; Amedei, Amedeo; Amin, Amr; Helferich, Bill; Boosani, Chandra S.; Ciriolo, Maria Rosa; Chen, Sophie; Mohammed, Sulma I.; Azmi, Asfar S.; Keith, W Nicol; Bhakta, Dipita; Halicka, Dorota; Niccolai, Elena; Fujii, Hiromasa; Aquilano, Katia; Ashraf, S. Salman; Nowsheen, Somaira; Yang, Xujuan; Bilsland, Alan; Shin, Dong M.

2015-01-01

The evasion of anti-growth signaling is an important characteristic of cancer cells. In order to continue to proliferate, cancer cells must somehow uncouple themselves from the many signals that exist to slow down cell growth. Here, we define the anti-growth signaling process, and review several important pathways involved in growth signaling: p53, phosphatase and tensin homolog (PTEN), retinoblastoma protein (Rb), Hippo, growth differentiation factor 15 (GDF15), AT-rich interactive domain 1A (ARID1A), Notch, insulin-like growth factor (IGF), and Krüppel-like factor 5 (KLF5) pathways. Aberrations in these processes in cancer cells involve mutations and thus the suppression of genes that prevent growth, as well as mutation and activation of genes involved in driving cell growth. Using these pathways as examples, we prioritize molecular targets that might be leveraged to promote anti-growth signaling in cancer cells. Interestingly, naturally-occurring phytochemicals found in human diets (either singly or as mixtures) may promote anti-growth signaling, and do so without the potentially adverse effects associated with synthetic chemicals. We review examples of naturally-occurring phytochemicals that may be applied to prevent cancer by antagonizing growth signaling, and propose one phytochemical for each pathway. These are: epigallocatechin-3-gallate (EGCG) for the Rb pathway, luteolin for p53, curcumin for PTEN, porphyrins for Hippo, genistein for GDF15, resveratrol for ARID1A, withaferin A for Notch and diguelin for the IGF1-receptor pathway. The coordination of anti-growth signaling and natural compound studies will provide insight into the future application of these compounds in the clinical setting. PMID:25749195

Suppression of KV7/KCNQ potassium channel enhances neuronal differentiation of PC12 cells.

PubMed

Zhou, Najing; Huang, Sha; Li, Li; Huang, Dongyang; Yan, Yunli; Du, Xiaona; Zhang, Hailin

2016-10-01

Membrane potential shift driven by electrical activity is critical in determining the cell fate of proliferation or differentiation. As such, the ion channels that underlie the membrane electrical activity play an important role in cell proliferation/differentiation. KV7/KCNQ potassium channels are critical in determining the resting membrane potentials in many neuronal cells. However, the role of these channels in cell differentiation is not well studied. In the present study, we used PC12 cells as well as primary cultured rat cortical neurons to study the role and mechanism of KV7/KCNQ in neuronal differentiation. NGF induced PC12 cell differentiation into neuron-like cells with growth of neurites showing typical growth cone-like extensions. The Kv7/KCNQ blocker XE991 promoted NGF-induced neurite outgrowth, whereas Kv7/KCNQ opener retigabine (RTG) inhibited outgrowth. M-type Kv7 channels are likely involved in regulating neurite growth because overexpression of KCNQ2/Q3 inhibited neurite growth whereas suppression of KCNQ2/Q3 with shRNA promoted neurite growth. Membrane depolarization possibly underpins enhanced neurite growth induced by the suppression of Kv7/KCNQ. Additionally, high extracellular K(+) likely induced membrane depolarization and also promoted neurite growth. Finally, T-type Ca(2+) channels may be involved in membrane-depolarization-induced neurite growth. This study provides a new perspective for understanding neuronal differentiation as well as KV7/KCNQ channel function. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

The role of EMMPRIN in T cell biology and immunological diseases.

PubMed

Hahn, Jennifer Nancy; Kaushik, Deepak Kumar; Yong, V Wee

2015-07-01

EMMPRIN (CD147), originally described as an inducer of the expression of MMPs, has gained attention in its involvement in various immunologic diseases, such that anti-EMMPRIN antibodies are considered as potential therapeutic medications. Given that MMPs are involved in the pathogenesis of various disease states, it is relevant that targeting an upstream inducer would make for an effective therapeutic strategy. Additionally, EMMPRIN is now appreciated to have multiple roles apart from MMP induction, including in cellular functions, such as migration, adhesion, invasion, energy metabolism, as well as T cell activation and proliferation. Here, we review what is known about EMMPRIN in numerous immunologic/inflammatory disease conditions with a particular focus on its complex roles in T cell biology. © Society for Leukocyte Biology.

Human embryonic stem cell-derived oligodendrocyte progenitor cell transplants remyelinate and restore locomotion after spinal cord injury.

PubMed

Keirstead, Hans S; Nistor, Gabriel; Bernal, Giovanna; Totoiu, Minodora; Cloutier, Frank; Sharp, Kelly; Steward, Oswald

2005-05-11

Demyelination contributes to loss of function after spinal cord injury, and thus a potential therapeutic strategy involves replacing myelin-forming cells. Here, we show that transplantation of human embryonic stem cell (hESC)-derived oligodendrocyte progenitor cells (OPCs) into adult rat spinal cord injuries enhances remyelination and promotes improvement of motor function. OPCs were injected 7 d or 10 months after injury. In both cases, transplanted cells survived, redistributed over short distances, and differentiated into oligodendrocytes. Animals that received OPCs 7 d after injury exhibited enhanced remyelination and substantially improved locomotor ability. In contrast, when OPCs were transplanted 10 months after injury, there was no enhanced remyelination or locomotor recovery. These studies document the feasibility of predifferentiating hESCs into functional OPCs and demonstrate their therapeutic potential at early time points after spinal cord injury.

Protein kinase Cβ as a therapeutic target stabilizing blood–brain barrier disruption in experimental autoimmune encephalomyelitis

PubMed Central

Lanz, Tobias V.; Becker, Simon; Osswald, Matthias; Bittner, Stefan; Schuhmann, Michael K.; Opitz, Christiane A.; Gaikwad, Sadanand; Wiestler, Benedikt; Litzenburger, Ulrike M.; Sahm, Felix; Ott, Martina; Iwantscheff, Simeon; Grabitz, Carl; Mittelbronn, Michel; von Deimling, Andreas; Winkler, Frank; Meuth, Sven G.; Wick, Wolfgang; Platten, Michael

2013-01-01

Disruption of the blood–brain barrier (BBB) is a hallmark of acute inflammatory lesions in multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis. This disruption may precede and facilitate the infiltration of encephalitogenic T cells. The signaling events that lead to this BBB disruption are incompletely understood but appear to involve dysregulation of tight-junction proteins such as claudins. Pharmacological interventions aiming at stabilizing the BBB in MS might have therapeutic potential. Here, we show that the orally available small molecule LY-317615, a synthetic bisindolylmaleimide and inhibitor of protein kinase Cβ, which is clinically under investigation for the treatment of cancer, suppresses the transmigration of activated T cells through an inflamed endothelial cell barrier, where it leads to the induction of the tight-junction molecules zona occludens-1, claudin 3, and claudin 5 and other pathways critically involved in transendothelial leukocyte migration. Treatment of mice with ongoing experimental autoimmune encephalomyelitis with LY-317615 ameliorates inflammation, demyelination, axonal damage, and clinical symptoms. Although LY-317615 dose-dependently suppresses T-cell proliferation and cytokine production independent of antigen specificity, its therapeutic effect is abrogated in a mouse model requiring pertussis toxin. This abrogation indicates that the anti-inflammatory and clinical efficacy is mainly mediated by stabilization of the BBB, thus suppressing the transmigration of encephalitogenic T cells. Collectively, our data suggest the involvement of endothelial protein kinase Cβ in stabilizing the BBB in autoimmune neuroinflammation and imply a therapeutic potential of BBB-targeting agents such as LY-317615 as therapeutic approaches for MS. PMID:23959874

PAXIP1 potentiates the combination of WEE1 inhibitor AZD1775 and platinum agents in lung cancer

PubMed Central

Jhuraney, Ankita; Woods, Nicholas T.; Wright, Gabriela; Rix, Lily; Kinose, Fumi; Kroeger, Jodi L.; Remily-Wood, Elizabeth; Cress, W. Douglas; Koomen, John M.; Brantley, Stephen G.; Gray, Jhanelle E.; Haura, Eric B.; Rix, Uwe; Monteiro, Alvaro N.

2016-01-01

The DNA damage response (DDR) involves a complex network of signaling events mediated by modular protein domains such as the BRCT (BRCA1 C-terminal) domain. Thus, proteins that interact with BRCT domains and are a part of the DDR constitute potential targets for sensitization to DNA damaging chemotherapy agents. We performed a pharmacological screen to evaluate seventeen kinases, identified in a BRCT-mediated interaction network as targets to enhance platinum-based chemotherapy in lung cancer. Inhibition of mitotic kinase WEE1 was found to have the most effective response in combination with platinum compounds in lung cancer cell lines. In the BRCT-mediated interaction network, WEE1 was found in complex with PAXIP1, a protein containing six BRCT domains involved in transcription and in the cellular response to DNA damage. We show that PAXIP1 BRCT domains regulate WEE1-mediated phosphorylation of CDK1. Further, ectopic expression of PAXIP1 promotes enhanced caspase 3-mediated apoptosis in cells treated with WEE1 inhibitor AZD1775 (formerly, MK-1775) and cisplatin compared with cells treated with AZD1775 alone. Cell lines and patient-derived xenograft models expressing both PAXIP1 and WEE1 exhibited synergistic effects of AZD1775 and cisplatin. In summary, PAXIP1 is involved in sensitizing lung cancer cells to the WEE1 inhibitor AZD1775 in combination with platinum-based treatment. We propose that WEE1 and PAXIP1 levels may be used as mechanism-based biomarkers of response when WEE1 inhibitor AZD1775 is combined with DNA damaging agents. PMID:27196765

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

Gulec, Cagri, E-mail: cagri.gulec@gmail.com; Coban, Neslihan, E-mail: neslic@istanbul.edu.tr; Ozsait-Selcuk, Bilge, E-mail: ozsaitb@istanbul.edu.tr

ROR-alpha is a nuclear receptor, activity of which can be modulated by natural or synthetic ligands. Due to its possible involvement in, and potential therapeutic target for atherosclerosis, we aimed to identify ROR-alpha target genes in monocytic and endothelial cell lines. We performed chromatin immunoprecipitation (ChIP) followed by tiling array (ChIP-on-chip) for ROR-alpha in monocytic cell line THP1 and endothelial cell line HUVEC. Following bioinformatic analysis of the array data, we tested four candidate genes in terms of dependence of their expression level on ligand-mediated ROR-alpha activity, and two of them in terms of promoter occupancy by ROR-alpha. Bioinformatic analysesmore » of ChIP-on-chip data suggested that ROR-alpha binds to genomic regions near the transcription start site (TSS) of more than 3000 genes in THP1 and HUVEC. Potential ROR-alpha target genes in both cell types seem to be involved mainly in membrane receptor activity, signal transduction and ion transport. While SPP1 and IKBKA were shown to be direct target genes of ROR-alpha in THP1 monocytes, inflammation related gene HMOX1 and heat shock protein gene HSPA8 were shown to be potential target genes of ROR-alpha. Our results suggest that ROR-alpha may regulate signaling receptor activity, and transmembrane transport activity through its potential target genes. ROR-alpha seems also to play role in cellular sensitivity to environmental substances like arsenite and chloroprene. Although, the expression analyses have shown that synthetic ROR-alpha ligands can modulate some of potential ROR-alpha target genes, functional significance of ligand-dependent modulation of gene expression needs to be confirmed with further analyses.« less

The semisynthetic flavonoid monoHER sensitises human soft tissue sarcoma cells to doxorubicin-induced apoptosis via inhibition of nuclear factor-κB

PubMed Central

Jacobs, H; Bast, A; Peters, G J; van der Vijgh, W J F; Haenen, G R M M

2011-01-01

Background: Despite therapeutic advances, the prognosis of patients with metastatic soft tissue sarcoma (STS) remains extremely poor. The results of a recent clinical phase II study, evaluating the protective effects of the semisynthetic flavonoid 7-mono-O-(β-hydroxyethyl)-rutoside (monoHER) on doxorubicin-induced cardiotoxicity, suggest that monoHER enhances the antitumour activity of doxorubicin in STSs. Methods: To molecularly explain this unexpected finding, we investigated the effect of monoHER on the cytotoxicity of doxorubicin, and the potential involvement of glutathione (GSH) depletion and nuclear factor-κB (NF-κB) inactivation in the chemosensitising effect of monoHER. Results: MonoHER potentiated the antitumour activity of doxorubicin in the human liposarcoma cell line WLS-160. Moreover, the combination of monoHER with doxorubicin induced more apoptosis in WLS-160 cells compared with doxorubicin alone. MonoHER did not reduce intracellular GSH levels. On the other hand, monoHER pretreatment significantly reduced doxorubicin-induced NF-κB activation. Conclusion: These results suggest that reduction of doxorubicin-induced NF-κB activation by monoHER, which sensitises cancer cells to apoptosis, is involved in the chemosensitising effect of monoHER in human liposarcoma cells. PMID:21245867

BetaIg-h3 is involved in the HAb18G/CD147-mediated metastasis process in human hepatoma cells.

PubMed

Tang, Juan; Zhou, Hong-wei; Jiang, Jian-li; Yang, Xiang-min; Li, Yu; Zhang, Hong-xin; Chen, Zhi-nan; Guo, Wei-ping

2007-03-01

HAb18G/CD147, a new hepatoma-associated antigen cloned and screened from human hepatocellular carcinoma cDNA library, is closely correlated with metastasis process in human hepatoma cells. In the present study we aimed to identify the pivotal molecules of the HAb18G/CD147 signal transduction pathway. The investigation showed that betaig-h3, a secretory extracellular matrix (ECM) protein, was upregulated in HAb18G/CD147-expressing human hepatoma T7721 cells and was downregulated by depressing HAb18G/CD147 expression. The expression of betaig-h3, upregulated in human hepatoma cells, was positively relative to the expression of HAb18G/CD147 in different human hepatoma cell lines. By overexpressing betaig-h3 in human SMMC-7721 hepatoma cells, we discovered that betaig-h3 promoted cell adhesion, invasion, and matrix metalloproteinase (MMP) secretion potential. HAb18G/CD147-induced invasion and metastasis potential of human hepatoma cells can be attenuated by antibodies specific for betaig-h3, and no significant differences on inhibitory effects were observed among T7721 cells incubated with antibodies for betaig-h3 or HAb18G/CD147 or both types together. Taken together, our study suggests that betaig-h3, regulated by the expression of HAb18G/CD147, is involved in the HAb18G/CD147 signal transduction pathway and mediates the HAb18G/CD147-induced invasion and metastasis process of human hepatoma cells.

Anion exchanger 2 is critical for CD8(+) T cells to maintain pHi homeostasis and modulate immune responses.

PubMed

Concepcion, Axel R; Salas, January T; Sarvide, Sarai; Sáez, Elena; Ferrer, Alex; López, María; Portu, Ainhoa; Banales, Jesús M; Hervás-Stubbs, Sandra; Oude Elferink, Ronald P J; Prieto, Jesús; Medina, Juan F

2014-05-01

Mitogenic stimulation of lymphocytes involves alkalinization of intracellular pH (pHi ). Subsequent pHi regulation may involve HCO3 (-) extrusion through Cl(-) /HCO3 (-) exchangers and/or Na(+) -HCO3 (-) co-transporters with acid-loading capability. Abnormalities in these mechanisms could result in immune dysfunctions, as suggested by the CD8(+) T-cell expansion encountered in mice lacking Ae2 (a widely expressed acid loader with electroneutral and Na(+) -independent Cl(-) /HCO3 (-) anion-exchange activity). Here we report that CD8(+) T cells but not CD4(+) T cells or other lymphocyte populations, are crucially dependent on Ae2 for pHi regulation. While total lymphocytes (including isolated CD4(+) T cells) exhibit Ae1 expression and Na(+) -HCO3 (-) co-transport with acidifying potential, CD8(+) T cells lack these acid-loading mechanisms. In Ae2-KO mice, CD4(+) but not CD8(+) T cells upregulate these potential Ae2 surrogates. As a consequence, Ae2-KO CD8(+) T cells exhibit alkalinized pHi , and dramatically increase their pHi upon CD3 stimulation. Moreover, stimulated Ae2-deficient CD8(+) T cells show enhanced intracellular production of IL-2 and membrane expression of its receptor IL-2Rα, together with increased cell proliferation and activation. These findings demonstrate that CD8(+) T cells are critically dependent on Ae2 for pHi homeostasis and tuning of cell proliferation and activation. Ae2 thus constitutes a novel target to modulate CD8(+) T-cell responses. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Enhancement of P53-Mutant Human Colorectal Cancer Cells Radiosensitivity by Flavonoid Fisetin

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

Chen Wenshu; Lee Yijang; Yu Yichu

Purpose: The aim of this study was to investigate whether fisetin is a potential radiosensitizer for human colorectal cancer cells, which are relatively resistant to radiotherapy. Methods and Materials: Cell survival was examined by clonogenic survival assay, and DNA fragmentation was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. The effects of treatments on cell cycle distribution and apoptosis were examined by flow cytometry. Western blot analysis was performed to ascertain the protein levels of {gamma}-H2AX, phospho-Chk2, active caspase-3, PARP cleavage, phospho-p38, phospho-AKT, and phospho-ERK1/2. Results: Fisetin pretreatment enhanced the radiosensitivity of p53-mutant HT-29 human colorectal cancer cellsmore » but not human keratocyte HaCaT cells; it also prolonged radiation-induced G{sub 2}/M arrest, enhanced radiation-induced cell growth arrest in HT-29 cells, and suppressed radiation-induced phospho-H2AX (Ser-139) and phospho-Chk2 (Thr-68) in p53-mutant HT-29 cells. Pretreatment with fisetin enhanced radiation-induced caspase-dependent apoptosis in HT-29 cells. Fisetin pretreatment augmented radiation-induced phosphorylation of p38 mitogen-activated protein kinase, which is involved in caspase-mediated apoptosis, and SB202190 significantly reduced apoptosis and radiosensitivity in fisetin-pretreated HT-29 cells. By contrast, both phospho-AKT and phospho-ERK1/2, which are involved in cell proliferation and antiapoptotic pathways, were suppressed after irradiation combined with fisetin pretreatment. Conclusions: To our knowledge, this study is the first to provide evidence that fisetin exerts a radiosensitizing effect in p53-mutant HT-29 cells. Fisetin could potentially be developed as a novel radiosensitizer against radioresistant human cancer cells.« less

Differential induction of CD94 and NKG2 in CD4 helper T cells. A consequence of influenza virus infection and interferon-γ?

PubMed Central

Graham, Christine M; Christensen, Jillian R; Thomas, D Brian

2007-01-01

Influenza A virus causes worldwide epidemics and pandemics and the investigation of memory T helper (Th) cells that help maintain serological memory following infection is important for vaccine design. In this study we investigated CD94 and NKG2 gene expression in memory CD4 T-cell clones established from the spleens of C57BL/10 (H-2b) and BALB/c (H-2d) mice infected with influenza A virus (H3N2). CD94 and NKG2A/C/E proteins form heterodimeric membrane receptors that are involved in virus recognition. CD94 and NKG2 expression have been well characterized in natural killer (NK) and cytotoxic T cells. Despite CD94 being potentially an important marker for Th1 cells involved in virus infection, however, there has been little investigation of its expression or function in the CD4 T-cell lineage and no studies have looked at in-vivo-generated Th cells or memory cells. We show in this study that in-vivo-generated CD4 Th1 cells, but not Th2 cells, exhibited full-length CD94 and NKG2A gene expression following activation with viral peptide. For NKG2A, a novel ‘short’ (possibly redundant) truncated isoform was detectable in a Th2 cell clone. Another member of the NK receptor family, NKG2D, but not NKG2C or E, was also differentially expressed in Th1 cells. We show here that CD94 and NKG2A may exist as multiple isoforms with the potential to distinguish helper T-cell subsets. PMID:17462078

AURKA promotes cancer metastasis by regulating epithelial-mesenchymal transition and cancer stem cell properties in hepatocellular carcinoma

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

Chen, Chenlin; Song, Guangyuan; Xiang, Jue

AURKA (aurora kinase A) has been confirmed as an oncogene in cancer development; however, its role and underlying mechanisms in the metastasis of hepatocellular carcinoma (HCC) remain unknown. In this study, We found that AURKA was up-regulated in HCC tissues and correlated with pathological stage and distant metastasis. Further found that AURKA was involved in the cancer metastases after radiation in HCC. While overexpression of AURKA induced epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) behaviors though PI3K/AKT pathway, silencing AURKA suppressed radiation-enhanced cell invasiveness of HCC. Taken together, our results suggested that AURKA contributed in metastasis of irradiated residulmore » HCC though facilitating EMT and CSC properties, suggesting the potential clinical application of AURKA inhibitors in radiotherapy for patients with HCC. - Highlights: • First reported overexpression of AURKA in HCC and correlation with poor OS. • AURKA was involved in the cancer metastases after radiation in HCC. • Further found AURKA promoted EMT and CSC behaviors though PI3K/AKT pathway. • Silencing AURKA suppressed radiation-enhanced cell invasiveness of HCC. • AURKA may be potential therapeutic target of HCC.« less

Purinergic signaling in hypothalamic tanycytes: potential roles in chemosensing.

PubMed

Dale, Nicholas

2011-04-01

Hypothalamic tanycytes are cells that line the walls of the 3rd ventricle. Their cell bodies contact the cerebrospinal fluid and give rise to an inwardly directed process. The more dorsally located (α1 and α2) tanycytes project to areas of the brain involved in the control of feeding and energy balance (the arcuate nucleus and ventromedial hypothalamic nucleus). Although their functions are poorly understood, they have some similarities to glial cells. Recent evidence shows that they express key molecules involved in purinergic signaling and at least some tanycytes may act as adult multipotent stem cells. Emerging evidence suggests that tanycytes signal through changes in intracellular Ca(2+) and that they can respond with large Ca(2+) signals to ATP and transmitters associated with wakefulness and the drive to feed. They are also glucosensitive and this response is dependent on release of ATP from tanycytes and the activation of P2Y1 receptors. Their ability to release ATP gives potential for their integration into the hypothalamic circuitry controlling energy balance and feeding, but many fundamental questions about their possible functions and roles remain unanswered. Copyright © 2011 Elsevier Ltd. All rights reserved.

Neural cell adhesion molecule-deficient beta-cell tumorigenesis results in diminished extracellular matrix molecule expression and tumour cell-matrix adhesion.

PubMed

Håkansson, Joakim; Xian, Xiaojie; He, Liqun; Ståhlberg, Anders; Nelander, Sven; Samuelsson, Tore; Kubista, Mikael; Semb, Henrik

2005-01-01

To understand by which mechanism neural cell adhesion molecule (N-CAM) limits beta tumour cell disaggregation and dissemination, we searched for potential downstream genes of N-CAM during beta tumour cell progression by gene expression profiling. Here, we show that N-CAM-deficient beta-cell tumorigenesis is associated with changes in the expression of genes involved in cell-matrix adhesion and cytoskeletal dynamics, biological processes known to affect the invasive and metastatic behaviour of tumour cells. The extracellular matrix (ECM) molecules emerged as the primary target, i.e. N-CAM deficiency resulted in down-regulated mRNA expression of a broad range of ECM molecules. Consistent with this result, deficient deposition of major ECM stromal components, such as fibronectin, laminin 1 and collagen IV, was observed. Moreover, N-CAM-deficient tumour cells displayed defective matrix adhesion. These results offer a potential mechanism for tumour cell disaggregation during N-CAM-deficient beta tumour cell progression. Prospective consequences of these findings for the role of N-CAM in beta tumour cell dissemination are discussed.

Epigenomic analysis of primary human T cells reveals enhancers associated with TH2 memory cell differentiation and asthma susceptibility

PubMed Central

Seumois, Grégory; Chavez, Lukas; Gerasimova, Anna; Lienhard, Matthias; Omran, Nada; Kalinke, Lukas; Vedanayagam, Maria; Ganesan, Asha Purnima V; Chawla, Ashu; Djukanović, Ratko; Ansel, K Mark; Peters, Bjoern; Rao, Anjana; Vijayanand, Pandurangan

2014-01-01

A characteristic feature of asthma is the aberrant accumulation, differentiation or function of memory CD4+ T cells that produce type 2 cytokines (TH2 cells). By mapping genome-wide histone modification profiles for subsets of T cells isolated from peripheral blood of healthy and asthmatic individuals, we identified enhancers with known and potential roles in the normal differentiation of human TH1 cells and TH2 cells. We discovered disease-specific enhancers in T cells that differ between healthy and asthmatic individuals. Enhancers that gained the histone H3 Lys4 dimethyl (H3K4me2) mark during TH2 cell development showed the highest enrichment for asthma-associated single nucleotide polymorphisms (SNPs), which supported a pathogenic role for TH2 cells in asthma. In silico analysis of cell-specific enhancers revealed transcription factors, microRNAs and genes potentially linked to human TH2 cell differentiation. Our results establish the feasibility and utility of enhancer profiling in well-defined populations of specialized cell types involved in disease pathogenesis. PMID:24997565

Epigenomic analysis of primary human T cells reveals enhancers associated with TH2 memory cell differentiation and asthma susceptibility.

PubMed

Seumois, Grégory; Chavez, Lukas; Gerasimova, Anna; Lienhard, Matthias; Omran, Nada; Kalinke, Lukas; Vedanayagam, Maria; Ganesan, Asha Purnima V; Chawla, Ashu; Djukanović, Ratko; Ansel, K Mark; Peters, Bjoern; Rao, Anjana; Vijayanand, Pandurangan

2014-08-01

A characteristic feature of asthma is the aberrant accumulation, differentiation or function of memory CD4(+) T cells that produce type 2 cytokines (TH2 cells). By mapping genome-wide histone modification profiles for subsets of T cells isolated from peripheral blood of healthy and asthmatic individuals, we identified enhancers with known and potential roles in the normal differentiation of human TH1 cells and TH2 cells. We discovered disease-specific enhancers in T cells that differ between healthy and asthmatic individuals. Enhancers that gained the histone H3 Lys4 dimethyl (H3K4me2) mark during TH2 cell development showed the highest enrichment for asthma-associated single nucleotide polymorphisms (SNPs), which supported a pathogenic role for TH2 cells in asthma. In silico analysis of cell-specific enhancers revealed transcription factors, microRNAs and genes potentially linked to human TH2 cell differentiation. Our results establish the feasibility and utility of enhancer profiling in well-defined populations of specialized cell types involved in disease pathogenesis.

Prostaglandin E₂ regulates cellular migration via induction of vascular endothelial growth factor receptor-1 in HCA-7 human colon cancer cells.

PubMed

Fujino, Hiromichi; Toyomura, Kaori; Chen, Xiao-bo; Regan, John W; Murayama, Toshihiko

2011-02-01

An important event in the development of tumors is angiogenesis, or the formation of new blood vessels. Angiogenesis is also known to be involved in tumor cell metastasis and is dependent upon the activity of the vascular endothelial growth factor (VEGF) signaling pathway. Studies of mice in which the EP3 prostanoid receptors have been genetically deleted have shown a role for these receptors in cancer growth and angiogenesis. In the present study, human colon cancer HCA-7 cells were used as a model system to understand the potential role of EP3 receptors in tumor cell migration. We now show that stimulation of HCA-7 cells with PGE₂ enhanced the up-regulation of VEGF receptor-1 (VEGFR-1) expression by a mechanism involving EP3 receptor-mediated activation of phosphatidylinositol 3-kinase and the extracellular signal-regulated kinases. Moreover, the PGE₂ stimulated increase in VEGFR-1 expression was accompanied by an increase in the cellular migration of HCA-7 cells. Given the known involvement of VEGFR-1 in cellular migration, our results suggest that EP3 receptors may contribute to tumor cell metastasis by increasing cellular migration through the up-regulation of VEGFR-1 signaling. Copyright © 2010 Elsevier Inc. All rights reserved.

Myeloid-derived suppressor cells modulate B-cell responses.

PubMed

Lelis, Felipe J N; Jaufmann, Jennifer; Singh, Anurag; Fromm, Katja; Teschner, Annkathrin Chiara; Pöschel, Simone; Schäfer, Iris; Beer-Hammer, Sandra; Rieber, Nikolaus; Hartl, Dominik

2017-08-01

Myeloid-derived suppressor cells (MDSCs) are key regulators of adaptive immunity by suppressing T-cell functions. However, their potential action on or interaction with B cells remained poorly understood. Here we demonstrate that human polymorphonuclear MDSCs differentially modulate B-cell function by suppressing B-cell proliferation and antibody production. We further demonstrate that this MDSC-mediated effect is cell contact dependent and involves established mediators such as arginase-1, nitric oxide (NO), reactive oxygen species (ROS) as well as B-cell death. Collectively, our studies provide novel evidence that human MDSCs modulate B cells, which could have future implications for immunotherapy approaches. Copyright © 2017 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Muscle Stem Cells: A Model System for Adult Stem Cell Biology.

PubMed

Cornelison, Ddw; Perdiguero, Eusebio

2017-01-01

Skeletal muscle stem cells, originally termed satellite cells for their position adjacent to differentiated muscle fibers, are absolutely required for the process of skeletal muscle repair and regeneration. In the last decade, satellite cells have become one of the most studied adult stem cell systems and have emerged as a standard model not only in the field of stem cell-driven tissue regeneration but also in stem cell dysfunction and aging. Here, we provide background in the field and discuss recent advances in our understanding of muscle stem cell function and dysfunction, particularly in the case of aging, and the potential involvement of muscle stem cells in genetic diseases such as the muscular dystrophies.

Bitter triggers acetylcholine release from polymodal urethral chemosensory cells and bladder reflexes.

PubMed

Deckmann, Klaus; Filipski, Katharina; Krasteva-Christ, Gabriela; Fronius, Martin; Althaus, Mike; Rafiq, Amir; Papadakis, Tamara; Renno, Liane; Jurastow, Innokentij; Wessels, Lars; Wolff, Miriam; Schütz, Burkhard; Weihe, Eberhard; Chubanov, Vladimir; Gudermann, Thomas; Klein, Jochen; Bschleipfer, Thomas; Kummer, Wolfgang

2014-06-03

Chemosensory cells in the mucosal surface of the respiratory tract ("brush cells") use the canonical taste transduction cascade to detect potentially hazardous content and trigger local protective and aversive respiratory reflexes on stimulation. So far, the urogenital tract has been considered to lack this cell type. Here we report the presence of a previously unidentified cholinergic, polymodal chemosensory cell in the mammalian urethra, the potential portal of entry for bacteria and harmful substances into the urogenital system, but not in further centrally located parts of the urinary tract, such as the bladder, ureter, and renal pelvis. Urethral brush cells express bitter and umami taste receptors and downstream components of the taste transduction cascade; respond to stimulation with bitter (denatonium), umami (monosodium glutamate), and uropathogenic Escherichia coli; and release acetylcholine to communicate with other cells. They are approached by sensory nerve fibers expressing nicotinic acetylcholine receptors, and intraurethral application of denatonium reflexively increases activity of the bladder detrusor muscle in anesthetized rats. We propose a concept of urinary bladder control involving a previously unidentified cholinergic chemosensory cell monitoring the chemical composition of the urethral luminal microenvironment for potential hazardous content.

Potential radiological impact of tornadoes on the safety of Nuclear Fuel Services' West Valley Fuel Reprocessing Plant. Volume I. Tornado effects on head-end cell airflow

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

Holloway, L.J.; Andrae, R.W.

1981-09-01

This report describes results of a parametric study of the impacts of a tornado-generated depressurization on airflow in the contaminated process cells within the presently inoperative Nuclear Fuel Services fuel reprocessing facility near West Valley, NY. The study involved the following tasks: (1) mathematical modeling of installed ventilation and abnormal exhaust pathways from the cells and prediction of tornado-induced airflows in these pathways; (2) mathematical modeling of individual cell flow characteristics and prediction of in-cell velocities induced by flows from step 1; and (3) evaluation of the results of steps 1 and 2 to determine whether any of the pathwaysmore » investigated have the potential for releasing quantities of radioactively contaminated air from the main process cells. The study has concluded that in the event of a tornado strike, certain pathways from the cells have the potential to release radioactive materials of the atmosphere. Determination of the quantities of radioactive material released from the cells through pathways identified in step 3 is presented in Part II of this report.« less

Very small embryonic-like (VSEL) stem cells in adult organs and their potential role in rejuvenation of tissues and longevity

PubMed Central

Ratajczak, Mariusz Z.; Zuba-Surma, Ewa K.; Shin, Dong-Myung; Ratajczak, Janina; Kucia, Magda

2011-01-01

Recently, we purified rare CXC chemokine receptor 4 expressing (CXCR4+) small stem cells (SCs) from the murine bone marrow (BM) that express markers characteristic for embryonic (E)SCs, epiblast (EP)SCs, and primordial germ cells (PGCs). We named these primitive cells very small embryonic-like (VSEL) SCs (VSELs). Our data indicate that VSELs are also present in many other organs in mice and that they may differentiate into cells from all three germ layers. Similar SCs were also isolated from human cord blood (CB) and mobilized peripheral blood (mPB). We hypothesize that VSELs are deposited during gastrulation and organogenesis in developing organs/tissues of mammals as a population of pluripotent stem cells (PSCs) that give rise to tissue committed monopotent SCs and that their number decreases with age. Therefore VSELs could play a pivotal role in normal rejuvenation of adult tissues as well as involvement in regeneration of damaged organs. Thus, these cells are potential SCs candidates for regenerative medicine and we envision that the regenerative potential of these cells could be harnessed to decelerate the aging processes. PMID:18601995

CRKL overexpression suppresses in vitro proliferation, invasion and migration of murine hepatocarcinoma Hca-P cells.

PubMed

Lin, Qiuyue; Sun, Ming-Zhong; Guo, Chunmei; Shi, Ji; Chen, Xin; Liu, Shuqing

2015-02-01

The signal adaptor CRK family protein play important roles in cancer cell progression, proliferation, migration and invasion. Previously, we showed that CRK was involved in lymphatic metastatic potential of murine hepatocarcinoma cells. In current work, as a member of CRK family, chicken tumour virus number 10 regulator of kinase-like protein (CRKL) was revealed to be associated with malignant behaviors of Hca-P, a murine HCC cell with lymph node metastatic (LNM) rate of ∼25%. CRKL overexpression in Hca-P by a constructed eukaryotic expression vector of pcDNA3.1/V5-HisB-CRKL significantly ameliorated its malignant biological properties. CCK-8 and soft agar colony formation assays indicated CRKL overexpression significantly inhibits the cell proliferation and colony formation abilities of Hca-P. Additionally, transwell assays indicated that the Hca-P cell migration and invasion capacities were apparently reduced following CRKL overexpression. As Hca-P is an ideal hepatocarcinoma cell model with low (initial) LNM potential, CRKL is shown to act as a potential suppressor and to provide new insight for both the malignant behaviors of hepatocarcinoma cells and lymphatic metastasis mechanism of hepatocarcinoma. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

The soy-derived peptide Vglycin inhibits the growth of colon cancer cells in vitro and in vivo.

PubMed

Gao, Chang; Sun, Rui; Xie, Ya-Rong; Jiang, An-Li; Lin, Mei; Li, Min; Chen, Zheng-Wang; Zhang, Ping; Jin, Honglin; Feng, Jue-Ping

2017-05-01

Vglycin, a novel natural polypeptide isolated from pea seeds, possesses antidiabetic properties. Our previous studies have shown that Vglycin can induce the differentiation of human colon adenocarcinoma cells. We aimed to determine the anticancer activity of Vglycin against colon cancer cells and to elucidate related apoptosis-inducing mechanisms. Treatment with purified Vglycin significantly reduced growth, viability, and colony formation of CT-26, SW480, and NCL-H716 colon cancer cells in a dose-dependent manner while down-regulating the expression of proliferating cell nuclear antigen. Mouse xenograft studies showed a 38% inhibition of colon cancer growth in mice treated with Vglycin (20 mg/kg/day) at day 21. Furthermore, the potential mechanisms involved in Vglycin-induced cell apoptosis were examined using cell cycle studies, ultrastructural examination, as well as apoptosis-associated pathway analysis. The results showed that Vglycin significantly promoted apoptosis and G1/S phase cell cycle arrest. As revealed by Western blot, the expression of CDK2 and Cyclin D1 was down-regulated in all three Vglycin-treated colon cancer cells, indicating that the CDK2/Cyclin D1 cell cycle pathway involved in the initiation and progression of colon cancer. Moreover, the inhibition of Vglycin-induced cell proliferation in colon cancer cells was accompanied by alteration of the expression levels of the apoptosis-related proteins Bax, Bcl-2 and Mcl-1, and an increase of caspase-3 activity. Together, our results suggest that Vglycin may be another plant-derived peptide that suppresses colon cancer, supporting the continued investigation of Vglycin as therapeutic agent for colon cancer. Impact statement The antidiabetic properties and the capability of inducing differentiation of human colon adenocarcinoma cells of Vglycin have been reported in our previous studies. However, the anticancer potential of Vglycin on colon cancer cells and its possible related mechanisms were still unknown. In this study, we found that Vglycin could reduce growth, viability, and colony formation or colony size of CT-26, SW480, and NCL-H716 colon cancer cells. Moreover, Vglycin decreased tumor volume by 38% in xenograft mice transplanted with CT-26 cells. The mechanisms of these phenomena may be due to the down-regulated CDK2 and Cyclin D1, G1/S phase cell cycle arrest, and the dysregulated expression of Bax, Bcl-2, and Mcl-1. The findings highlight the anticancer potential of Vglycin against colon cancer cells, and suggest Vglycin may be another colon cancer potential suppressive component of plant-derived peptides.

The Effects of Storage on Irradiated Red Blood Cells: An In Vitro and In Vivo Study

DTIC Science & Technology

1991-08-01

nerve impulses, and is involved with cellular menbrane potential. It also influences and is influenced by the acid base balance. 19 Normal serum...maintained by active transport of scdium and potassium across the cell menbrane . Sodium is punped out and potassium pumped into the cell. The body’s...insufficiency or failure; renal dialysis is often required to remove 4 accumulated plasma potassium. Increased extr.cellular potassium causes changes in muscle

Characterization of increasing stages of invasiveness identifies stromal/cancer cell crosstalk in rat models of mesothelioma

PubMed Central

Nader, Joëlle S.; Abadie, Jérôme; Deshayes, Sophie; Boissard, Alice; Blandin, Stéphanie; Blanquart, Christophe; Boisgerault, Nicolas; Coqueret, Olivier; Guette, Catherine; Grégoire, Marc; Pouliquen, Daniel L.

2018-01-01

Sarcomatoid mesothelioma (SM) is a devastating cancer associated with one of the poorest outcome. Therefore, representative preclinical models reproducing different tumor microenvironments (TME) observed in patients would open up new prospects for the identification of markers and evaluation of innovative therapies. Histological analyses of four original models of rat SM revealed their increasing infiltrative and metastatic potential were associated with differences in Ki67 index, blood-vessel density, and T-lymphocyte and macrophage infiltration. In comparison with the noninvasive tumor M5-T2, proteomic analysis demonstrated the three invasive tumors F4-T2, F5-T1 and M5-T1 shared in common a very significant increase in the abundance of the multifunctional proteins galectin-3, prohibitin and annexin A5, and a decrease in proteins involved in cell adhesion, tumor suppression, or epithelial differentiation. The increased metastatic potential of the F5-T1 tumor, relative to F4-T2, was associated with an increased macrophage vs T-cell infiltrate, changes in the levels of expression of a panel of cytokine genes, an increased content of proteins involved in chromatin organization, ribosome structure, splicing, or presenting anti-adhesive properties, and a decreased content of proteins involved in protection against oxidative stress, normoxia and intracellular trafficking. The most invasive tumor, M5-T1, was characterized by a pattern of specific phenotypic and molecular features affecting the presentation of MHC class I-mediated antigens and immune cell infiltration, or involved in the reorganization of the cytoskeleton and composition of the extracellular matrix. These four preclinical models and data represent a new resource available to the cancer research community to catalyze further investigations on invasiveness. PMID:29662647

GSTM3 and GSTP1: novel players driving tumor progression in cervical cancer

PubMed Central

Checa-Rojas, Alberto; Delgadillo-Silva, Luis Fernando; Velasco-Herrera, Martín del Castillo; Andrade-Domínguez, Andrés; Gil, Jeovanis; Santillán, Orlando; Lozano, Luis; Toledo-Leyva, Alfredo; Ramírez-Torres, Alberto; Talamas-Rohana, Patricia; Encarnación-Guevara, Sergio

2018-01-01

The molecular processes and proteomic markers leading to tumor progression (TP) in cervical cancer (CC) are either unknown or only partially understood. TP affects metabolic and regulatory mechanisms that can be identified as proteomic changes. To identify which proteins are differentially expressed and to understand the mechanisms of cancer progression, we analyzed the dynamics of the tumor proteome in CC cell lines. This analysis revealed two proteins that are up-regulated during TP, GSTM3 and GSTP1. These proteins are involved in cell maintenance, cell survival and the cellular stress response via the NF-κB and MAP kinase pathways during TP. Furthermore, GSTM3 and GSTP1 knockdown showed that evasion of apoptosis was affected, and tumor proliferation was significantly reduced. Our data indicate the critical role of GST proteins in the regulation and progression of cervical cancer cells. Hence, we suggest GSTM3 and GSTP1 as novel biomarkers and potential therapeutic targets for treating cervical cancer. Significance CC is particularly hazardous in the advanced stages, and there are few therapeutic strategies specifically targeting these stages. We performed analyses on CC tumor proteome dynamics and identified GSTM3 and GSTP1 as novel potential therapeutic targets. Knockdown of these proteins showed that they are involved in cell survival, cell proliferation and cellular evasion of apoptosis. PMID:29774096

Porphyromonas gingivalis-mediated shedding of extracellular matrix metalloproteinase inducer (EMMPRIN) by oral epithelial cells: a potential role in inflammatory periodontal disease.

PubMed

Feldman, Mark; La, Vu Dang; Lombardo Bedran, Telma Blanca; Palomari Spolidorio, Denise Madalena; Grenier, Daniel

2011-12-01

Extracellular matrix metalloproteinase inducer (EMMPRIN) or CD147 is a transmembrane glycoprotein expressed by various cell types, including oral epithelial cells. Recent studies have brought evidence that EMMPRIN plays a role in periodontitis. In the present study, we investigated the effect of Porphyromonas gingivalis, a major pathogen in chronic periodontitis, on the shedding of membrane-anchored EMMPRIN and on the expression of the EMMPRIN gene by oral epithelial cells. A potential contribution of shed EMMPRIN to the inflammatory process of periodontitis was analyzed by evaluating the effect of recombinant EMMPRIN on cytokine and matrix metalloproteinase (MMP) secretion by human gingival fibroblasts. ELISA and immunofluorescence analyses revealed that P. gingivalis mediated the shedding of epithelial cell-surface EMMPRIN in a dose- and time-dependent manner. Cysteine proteinase (gingipain)-deficient P. gingivalis mutants were used to demonstrate that both Arg- and Lys-gingipain activities are involved in EMMPRIN shedding. Real-time PCR showed that P. gingivalis had no significant effect on the expression of the EMMPRIN gene in epithelial cells. Recombinant EMMPRIN induced the secretion of IL-6 and MMP-3 by gingival fibroblasts, a phenomenon that appears to involve mitogen activated protein kinases. The present study brought to light a new mechanism by which P. gingivalis can promote the inflammatory response during periodontitis. Copyright © 2011 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Pro-inflammatory Cytokines Are Involved in Fluoride-Induced Cytotoxic Potential in HeLa Cells.

PubMed

Wang, Hong-Wei; Zhou, Bian-Hua; Cao, Jian-Wen; Zhao, Jing; Zhao, Wen-Peng; Tan, Pan-Pan

2017-01-01

This study was designed to investigate the pro-inflammatory cytokines and their involvement in the cytotoxic potential of fluoride (F) in HeLa cells. HeLa cells were cultured with varying F concentrations (1-50 mg/L) for 48 h, and treatment effects were analyzed. The viability of HeLa cells was determined with a colorimetric method. The concentrations of IL-1β, IL-2, IL-6, and TNF-a in culture supernatant were measured through enzyme linked immunosorbent assay (ELISA). The mRNA expression levels of IL-1β, IL-2, IL-6 and TNF-a were subjected to transcript analysis and quantified through reverse transcription real-time PCR. Results showed that 10, 20 and 50 mg/L F significantly decreased the viability of HeLa cells incubated for 24 and 48 h. With their cytotoxic effect, the concentrations of IL-1β, IL-2, IL-6, and TNF-a decreased significantly in response to F, especially at 20 and 50 mg/L for 48 h. The mRNA expression levels of IL-1β, IL-2, IL-6, and TNF-a were downregulated at 50 mg/L F for 48 h. Therefore, F inhibited HeLa cell growth; as such, F could be used to alleviate the inhibition of pro-inflammatory cytokine expression.

Silencing of karyopherin α2 inhibits cell growth and survival in human hepatocellular carcinoma

PubMed Central

Yang, Yunfeng; Guo, Jian; Hao, Yuxia; Wang, Fuhua; Li, Fengxia; Shuang, Shaomin; Wang, Junping

2017-01-01

Karyopherin α2 (KPNA2), involved in nucleocytoplasmic transport, has been reported to be upregulated in hepatocellular carcinoma and considered as a biomarker for poor prognosis. However, comprehensive studies of KPNA2 functions in hepatocellular carcinogenesis are still lacking. Our study examine the roles and related molecular mechanisms of KPNA2 in hepatocellular carcinoma development. Results show that KPNA2 knockdown inhibited the proliferation and growth of hepatocellular carcinoma cells in vitro and in vivo. KPNA2 knockdown also inhibited colony formation ability, induced cell cycle arrest and cellular apoptosis in two hepatocellular carcinoma cell lines, HepG2 and SMMC-7721. Furthermore, gene expression microarray analysis in HepG2 cells with KPNA2 knockdown revealed that critical signaling pathways involved in cell proliferation and survival were deregulated. In conclusion, this study provided systematic evidence that KPNA2 was an essential factor promoting hepatocellular carcinoma and unraveled potential molecular pathways and networks underlying KPNA2-induced hepatocellular carcinogenesis. PMID:28422734

Silencing of karyopherin α2 inhibits cell growth and survival in human hepatocellular carcinoma.

PubMed

Yang, Yunfeng; Guo, Jian; Hao, Yuxia; Wang, Fuhua; Li, Fengxia; Shuang, Shaomin; Wang, Junping

2017-05-30

Karyopherin α2 (KPNA2), involved in nucleocytoplasmic transport, has been reported to be upregulated in hepatocellular carcinoma and considered as a biomarker for poor prognosis. However, comprehensive studies of KPNA2 functions in hepatocellular carcinogenesis are still lacking. Our study examine the roles and related molecular mechanisms of KPNA2 in hepatocellular carcinoma development. Results show that KPNA2 knockdown inhibited the proliferation and growth of hepatocellular carcinoma cells in vitro and in vivo. KPNA2 knockdown also inhibited colony formation ability, induced cell cycle arrest and cellular apoptosis in two hepatocellular carcinoma cell lines, HepG2 and SMMC-7721. Furthermore, gene expression microarray analysis in HepG2 cells with KPNA2 knockdown revealed that critical signaling pathways involved in cell proliferation and survival were deregulated. In conclusion, this study provided systematic evidence that KPNA2 was an essential factor promoting hepatocellular carcinoma and unraveled potential molecular pathways and networks underlying KPNA2-induced hepatocellular carcinogenesis.

P16/p53 expression and telomerase activity in immortalized human dental pulp cells

PubMed Central

Egbuniwe, Obi; Idowu, Bernadine D; Funes, Juan M; Grant, Andrew D; Renton, Tara

2011-01-01

Introduction Residing within human dental pulp are cells of an ectomesenchymal origin that have the potential to differentiate into odontoblast-like cells. These cells have a limited growth potential owing to the effects of cell senescence. This study examines the effects of immortalizing odontoblast-like cells on cell proliferation and mineralization by comparing transformed dental pulp stem cells (tDPSCs) and non-transformed dental pulp stem cells (nDPSCs). Results With the exogenous expression of hTERT, tDPSCs maintained a continued expression of odontogenic markers for cell proliferation and mineralization (ALP, COL-1, DMP-1, DSPP, OCN and OPN), as did nDPSCs. Oncoprotein expression was seen in both groups except for a noted absence of p16 in the tDPSCs. nDPSCs also showed lower levels of total ALP and DNA activity in comparison to tDPSCs when assayed, as well as low telomerase activity readings. Methods Using a retroviral vector, exogenous human telomerase reverse transcriptase (hTERT) was expressed in tDPSCs. Both cell groups were cultured, and their telomerase activities were determined using a telomerase quantification assay. Also examined, were the expression of genes involved in proliferation and mineralization, such as human alkaline phosphatase (ALP), β-actin, collagen I (col-1), core binding factor (cbfa)-1, dentin matrix protein (DMP-1), dentin sialophosphoprotein (DSPP), GAPDH, hTERT, osteocalcin (OCN), osteopontin (OPN) as well as oncoproteins involved in senescence (p16, p21 and p53) using RT-PCR. DNA and alkaline phosphate activity was also assayed in both cell groups. Conclusion These results indicate maintenance of odontoblast-like differentiation characteristics after retroviral transformation with hTERT and suggest a possible link with a reduced p16 expression. PMID:22067611

Reduced immunomodulation potential of bone marrow-derived mesenchymal stem cells induced CCR4+CCR6+ Th/Treg cell subset imbalance in ankylosing spondylitis

PubMed Central

2011-01-01

Introduction Ankylosing spondylitis (AS) is a chronic autoimmune disease, and the precise pathogenesis is largely unknown at present. Bone marrow-derived mesenchymal stem cells (BMSCs) with immunosuppressive and anti-inflammatory potential and Th17/Treg cells with a reciprocal relationship regulated by BMSCs have been reported to be involved in some autoimmune disorders. Here we studied the biological and immunological characteristics of BMSCs, the frequency and phenotype of CCR4+CCR6+ Th/Treg cells and their interaction in vitro in AS. Methods The biological and immunomodulation characteristics of BMSCs were examined by induced multiple-differentiation and two-way mixed peripheral blood mononuclear cell (PBMC) reactions or after stimulation with phytohemagglutinin, respectively. The interactions of BMSCs and PBMCs were detected with a direct-contact co-culturing system. CCR4+CCR6+ Th/Treg cells and surface markers of BMSCs were assayed using flow cytometry. Results The AS-BMSCs at active stage showed normal proliferation, cell viability, surface markers and multiple differentiation characteristics, but significantly reduced immunomodulation potential (decreased 68 ± 14%); the frequencies of Treg and Fox-P3+ cells in AS-PBMCs decreased, while CCR4+CCR6+ Th cells increased, compared with healthy donors. Moreover, the AS-BMSCs induced imbalance in the ratio of CCR4+CCR6+ Th/Treg cells by reducing Treg/PBMCs and increasing CCR4+CCR6+ Th/PBMCs, and also reduced Fox-P3+ cells when co-cultured with PBMCs. Correlation analysis showed that the immunomodulation potential of BMSCs has significant negative correlations with the ratio of CCR4+CCR6+ Th to Treg cells in peripheral blood. Conclusions The immunomodulation potential of BMSCs is reduced and the ratio of CCR4+CCR6+ Th/Treg cells is imbalanced in AS. The BMSCs with reduced immunomodulation potential may play a novel role in AS pathogenesis by inducing CCR4+CCR6+ Th/Treg cell imbalance. PMID:21338515

Fisetin and hesperetin induced apoptosis and cell cycle arrest in chronic myeloid leukemia cells accompanied by modulation of cellular signaling.

PubMed

Adan, Aysun; Baran, Yusuf

2016-05-01

Fisetin and hesperetin, naturally occurring flavonoids, have been reported as novel antioxidants with chemopreventive/chemotherapeutic potential against various types of cancer. However, their mechanism of action in CML is still unknown. This particular study aims to evaluate the therapeutic potentials of fisetin and hesperetin and their effects on cell proliferation, apoptosis, and cell cycle progression in human K562 CML cells. The results indicated that fisetin and hesperetin inhibited cell proliferation and triggered programmed cell death in these cells. The latter was confırmed by mitochondrial membrane depolarization and an increase in caspase-3 activation. In addition to that, we have detected S and G2/M cell cycle arrests and G0/G1 arrest upon fisetin and hesperetin treatment, respectively. To identify the altered genes and genetic networks in response to fisetin and hesperetin, whole-genome microarray analysis was performed. The microarray gene profiling analysis revealed some important signaling pathways including JAK/STAT pathway, KIT receptor signaling, and growth hormone receptor signaling that were altered upon fisetin and hesperetin treatment. Moreover, microarray data suggested potential candidate genes for targeted CML therapy. Fisetin and hesperetin significantly modulated the expression of genes involved in cell proliferation and division, apoptosis, cell cycle regulation, and other significant cellular processes such as replication, transcription, and translation. In conclusion, our results suggest that fisetin and hesperetin as potential natural agents for CML therapy.

HOXA13 is a potential GBM diagnostic marker and promotes glioma invasion by activating the Wnt and TGF-β pathways

PubMed Central

Duan, Ran; Han, Lei; Wang, Qixue; Wei, Jianwei; Chen, Luyue; Zhang, Jianning; Kang, Chunsheng; Wang, Lei

2015-01-01

Homeobox (HOX) genes, including HOXA13, are involved in human cancer. We found that HOXA13 expression was associated with glioma grade and prognosis. Bioinformatics analysis revealed that most of the HOXA13-associated genes were enriched in cancer-related signaling pathways and mainly involved in the regulation of transcription. We transfected four glioma cell lines with Lenti-si HOXA13. HOXA13 increased cell proliferation and invasion and inhibited apoptosis. HOXA13 decreased β-catenin, phospho-SMAD2, and phospho-SMAD3 in the nucleus and increased phospho-β-catenin in the cytoplasm. Furthermore, downregulation of HOXA13 in orthotopic tumors decreased tumor growth. We suggest that HOXA13 promotes glioma progression in part via Wnt- and TGF-β-induced EMT and is a potential diagnostic biomarker for glioblastoma and an independent prognostic factor in high-grade glioma. PMID:26356815

Mycoparasitism studies of Trichoderma harzianum against Sclerotinia sclerotiorum: evaluation of antagonism and expression of cell wall-degrading enzymes genes.

PubMed

Troian, Rogério Fraga; Steindorff, Andrei Stecca; Ramada, Marcelo Henrique Soller; Arruda, Walquiria; Ulhoa, Cirano José

2014-10-01

Trichoderma spp. are known for their biocontrol activity against several plant pathogens. A specific isolate of Trichoderma harzianum, 303/02, has the potential to inhibit the growth of Sclerotinia sclerotiorum, an important agent involved in several crop diseases. In this study, the interaction between T. harzianum 303/02 and mycelia, sclerotia and apothecia of S. sclerotiorum was studied by scanning electron microscopy. RT-qPCR was used to examine the expression of 11 genes potentially involved in biocontrol. T. harzianum 303/02 parasitizes S. sclerotiorum by forming branches that coil around the hyphae. The fungus multiplied abundantly at the sclerotia and apothecia surface, forming a dense mycelium that penetrated the inner surface of these structures. The levels of gene expression varied according to the type of structure with which T. harzianum was interacting. The data also showed the presence of synergistic action between the cell-wall degrading enzymes.

Molecular expression in transfected corneal endothelial cells

NASA Astrophysics Data System (ADS)

Wang, Fan; Miao, Zhuang; Lu, Chengwei; Hao, Jilong

2017-10-01

To investigate the capability of human corneal endothelial cells serving as immunological cells. Expression of HLA-DP, -DQ, -DR, CD40, CD80, and CD86 was determined by immunohistochemical methods. Meanwhile, purified peripheral blood mononuclear cells were cocultured with human corneal endothelial cells which were pre-treated with and without -IFN respectively, activation of lymphocytes was determined by FACS analysis. In coculture system, T lymphocyte was activated by corneal endothelial cells, HLA-DP, -DQ, -DR and CD40 expression were increased by - IFN induction. Costimulatory molecular CD80 was shown on the endothelial cells. Human corneal endothelial cells were assumed to be involved in the corneal transplantation rejection process as potential antigen presenting cells.

Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential

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

Song, Kai; The State Key Laboratory Breeding Base of Basic Science of Stomatology; Song, Yong

Most previous studies have linked cancer–macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed thatmore » SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression. - Highlights: • The fusion events between oral cancer and endothelial cells undergo nuclear fusion. • The resulting hybrid cells acquire a new property of drug resistance. • The resulting hybrid cells express the markers of both parental cells (i.e. vimentin and cytokeratin 18). • The hybrid cells contribute to tumor repopulation in vivo.« less

Modelling effects on grid cells of sensory input during self‐motion

PubMed Central

Raudies, Florian; Hinman, James R.

2016-01-01

Abstract The neural coding of spatial location for memory function may involve grid cells in the medial entorhinal cortex, but the mechanism of generating the spatial responses of grid cells remains unclear. This review describes some current theories and experimental data concerning the role of sensory input in generating the regular spatial firing patterns of grid cells, and changes in grid cell firing fields with movement of environmental barriers. As described here, the influence of visual features on spatial firing could involve either computations of self‐motion based on optic flow, or computations of absolute position based on the angle and distance of static visual cues. Due to anatomical selectivity of retinotopic processing, the sensory features on the walls of an environment may have a stronger effect on ventral grid cells that have wider spaced firing fields, whereas the sensory features on the ground plane may influence the firing of dorsal grid cells with narrower spacing between firing fields. These sensory influences could contribute to the potential functional role of grid cells in guiding goal‐directed navigation. PMID:27094096

DR-nm23 expression affects neuroblastoma cell differentiation, integrin expression, and adhesion characteristics.

PubMed

Amendola, R; Martinez, R; Negroni, A; Venturelli, D; Tanno, B; Calabretta, B; Raschellà, G

2001-01-01

Nm23 gene family has been associated with metastasis suppression and differentiation. We studied DR-nm23 during neuroblastoma cells differentiation. DR-nm23 expression increased after retinoic acid induction of differentiation in human cell lines SK-N-SH and LAN-5. In several cell lines, overexpression of DR-nm23 was associated with more differentiated phenotypes. SK-N-SH cells increased vimentin expression, increased deposition of collagen type IV, modulated integrin expression, and underwent growth arrest; the murine neuroblastoma cell line N1E-115 showed neurite outgrowth and a striking enhancement of beta1 integrin expression. Up-regulation of beta1 integrin was specifically responsible for the increase in the adhesion to collagen type I-coated plates. Finally, cells overexpressing DR-nm23 were unable to growth in soft agar. In conclusion, DR-nm23 expression is directly involved in differentiation of neuroblastoma cells, and its ability to affects the adhesion to extracellular substrates and to inhibit growth in soft agar suggests an involvement in the metastatic potential of neuroblastoma.

Distal regeneration involves the age dependent activity of branchial sac stem cells in the ascidian Ciona intestinalis

PubMed Central

2014-01-01

Abstract Tunicates have high capacities for regeneration but the underlying mechanisms and their relationship to life cycle progression are not well understood. Here we investigate the regeneration of distal structures in the ascidian tunicate Ciona intestinalis. Analysis of regenerative potential along the proximal−distal body axis indicated that distal organs, such as the siphons, their pigmented sensory organs, and the neural complex, could only be replaced from body fragments containing the branchial sac. Distal regeneration involves the formation of a blastema composed of cells that undergo cell proliferation prior to differentiation and cells that differentiate without cell proliferation. Both cell types originate in the branchial sac and appear in the blastema at different times after distal injury. Whereas the branchial sac stem cells are present in young animals, they are depleted in old animals that have lost their regeneration capacity. Thus Ciona adults contain a population of age‐related stem cells located in the branchial sac that are a source of precursors for distal body regeneration. PMID:25893097

EPHRIN-B1 Mosaicism Drives Cell Segregation in Craniofrontonasal Syndrome hiPSC-Derived Neuroepithelial Cells.

PubMed

Niethamer, Terren K; Larson, Andrew R; O'Neill, Audrey K; Bershteyn, Marina; Hsiao, Edward C; Klein, Ophir D; Pomerantz, Jason H; Bush, Jeffrey O

2017-03-14

Although human induced pluripotent stem cells (hiPSCs) hold great potential for the study of human diseases affecting disparate cell types, they have been underutilized in seeking mechanistic insights into the pathogenesis of congenital craniofacial disorders. Craniofrontonasal syndrome (CFNS) is a rare X-linked disorder caused by mutations in EFNB1 and characterized by craniofacial, skeletal, and neurological anomalies. Heterozygous females are more severely affected than hemizygous males, a phenomenon termed cellular interference that involves mosaicism for EPHRIN-B1 function. Although the mechanistic basis for cellular interference in CFNS has been hypothesized to involve Eph/ephrin-mediated cell segregation, no direct evidence for this has been demonstrated. Here, by generating hiPSCs from CFNS patients, we demonstrate that mosaicism for EPHRIN-B1 expression induced by random X inactivation in heterozygous females results in robust cell segregation in human neuroepithelial cells, thus supplying experimental evidence that Eph/ephrin-mediated cell segregation is relevant to pathogenesis in human CFNS patients. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

A novel PHD-finger protein 14/KIF4A complex overexpressed in lung cancer is involved in cell mitosis regulation and tumorigenesis.

PubMed

Zhang, Lin; Huang, Qin; Lou, Jiatao; Zou, Liangjian; Wang, Yiguo; Zhang, Peng; Yang, Guang; Zhang, Junyi; Yu, Lan; Yan, Dai; Zhang, Chenyi; Qiao, Jing; Wang, Shuting; Wang, Sai; Xu, Yongdong; Ji, Hongbin; Chen, Zhengjun; Zhang, Zhe

2017-03-21

The plant homeodomain (PHD) finger-containing proteins have been implicated in many human diseases including cancer. In this study, we found that PHF14, a newly identified PHD finger protein, is highly expressed in lung cancer. The high expression level of PHF14 was associated with adenocarcinoma and poor survival in lung cancer patients. Knocking down PHF14 suppressed cancer cell growth and carcinogenesis, while over-expressing PHF14 promoted cell proliferation. During cell division, PHF14 directly bound to and co-localized with KIF4A (a nuclear motor protein involved in lung carcinogenesis) to form a functional complex. Similarly to the effect of KIF4A depletion, silencing PHF14 in several cell lines caused cell mitotic defects, prolonged M phase, and inhibited cell proliferation. What's more, these two proteins had a synergistic effect on cell proliferation and were significantly co-overexpressed in lung cancer tissues. Our data provide new insights into the biological significance of PHD finger proteins and imply that PHF14 may be a potential biomarker for lung cancer.

Human trabecular meshwork cell volume decrease by NO-independent soluble guanylate cyclase activators YC-1 and BAY-58-2667 involves the BKCa ion channel.

PubMed

Dismuke, William M; Sharif, Najam A; Ellis, Dorette Z

2009-07-01

There is a correlation between cell volume changes and changes in the rate of aqueous humor outflow; agents that decrease trabecular meshwork (TM) cell volume increase the rate of aqueous humor outflow. This study investigated the effects of the nitric oxide (NO)-independent activators of soluble guanylate cyclase (sGC), YC-1, and BAY-58-2667 on TM cell volume and the signal transduction pathways and ion channel involved. Cell volume was measured with the use of calcein AM fluorescent dye, detected by confocal microscopy. Inhibitors and activators of sGC, 3',5'-cyclic guanosine monophosphate (cGMP), protein kinase G (PKG), and the BK(Ca) channel were used to characterize their involvement in the YC-1- and BAY-58-2667-induced regulation of TM cell volume. cGMP was assayed by an enzyme immunoassay. YC-1 (10 nM-200 microM) and BAY-58-2667 (10 nM-100 microM) each elicited a biphasic effect on TM cell volume. YC-1 (1 microM) increased TM cell volume, but higher concentrations decreased TM cell volume. Similarly, BAY-58-2667 (100 nM) increased TM cell volume, but higher concentrations decreased cell volume. The YC-1-induced cell volume decrease was mimicked by 8-Br-cGMP and abolished by the sGC inhibitor ODQ, the PKG inhibitor (RP)-8-Br-PET-cGMP-S, and the BK(Ca) channel inhibitor IBTX. The BAY-58-2667-induced cell volume decrease was mimicked by 8-Br-cGMP and was abolished by the PKG inhibitor and the BK(Ca) channel inhibitor. Unlike the YC-1 response, ODQ potentiated the BAY-58-2667-induced decreases in cell volume. These data suggest that the NO-independent decrease in TM cell volume is mediated by the sGC/cGMP/PKG pathway and involves K(+) efflux.

FOXO1 is a TXN- and p300-dependent sensor and effector of oxidative stress in diffuse large B-cell lymphomas characterized by increased oxidative metabolism.

PubMed

Sewastianik, T; Szydlowski, M; Jablonska, E; Bialopiotrowicz, E; Kiliszek, P; Gorniak, P; Polak, A; Prochorec-Sobieszek, M; Szumera-Cieckiewicz, A; Kaminski, T S; Markowicz, S; Nowak, E; Grygorowicz, M A; Warzocha, K; Juszczynski, P

2016-11-17

Molecular profiling has led to identification of subtypes of diffuse large B-cell lymphomas (DLBCLs) differing in terms of oncogenic signaling and metabolic programs. The OxPhos-DLBCL subtype is characterized by enhanced mitochondrial oxidative phosphorylation. As increased oxidative metabolism leads to overproduction of potentially toxic reactive oxygen species (ROS), we sought to identify mechanisms responsible for adaptation of OxPhos cells to these conditions. Herein, we describe a mechanism involving the FOXO1-TXN-p300 redox-dependent circuit protecting OxPhos-DLBCL cells from ROS toxicity. We identify a BCL6-dependent transcriptional mechanism leading to relative TXN overexpression in OxPhos cells. We found that OxPhos cells lacking TXN were uniformly more sensitive to ROS and doxorubicin than control cells. Consistent with this, the overall survival of patients with high TXN mRNA expression, treated with doxorubicin-containing regimens, is significantly shorter than of those with low TXN mRNA expression. TXN overexpression curtails p300-mediated FOXO1 acetylation and its nuclear translocation in response to oxidative stress, thus attenuating FOXO1 transcriptional activity toward genes involved in apoptosis and cell cycle inhibition. We also demonstrate that FOXO1 knockdown in cells with silenced TXN expression markedly reduces ROS-induced apoptosis, indicating that FOXO1 is the major sensor and effector of oxidative stress in OxPhos-DLBCLs. These data highlight dynamic, context-dependent modulation of FOXO1 tumor-suppressor functions via acetylation and reveal potentially targetable vulnerabilities in these DLBCLs.

RNA-Sequencing Analyses Demonstrate the Involvement of Canonical Transient Receptor Potential Channels in Rat Tooth Germ Development

PubMed Central

Yang, Jun; Cai, Wenping; Lu, Xi; Liu, Shangfeng; Zhao, Shouliang

2017-01-01

Tooth development depends on multiple molecular interactions between the dental epithelium and mesenchyme, which are derived from ectodermal and ectomesenchymal cells, respectively. We report on a systematic RNA sequencing analysis of transcriptional expression levels from the bud to hard tissue formation stages of rat tooth germ development. We found that GNAO1, ENO1, EFNB1, CALM1, SIAH2, ATP6V0A1, KDELR2, GTPBP1, POLR2C, SORT1, and members of the canonical transient receptor potential (TRPC) channel family are involved in tooth germ development. Furthermore, Cell Counting Kit 8 (CCK8) and Transwell migration assays were performed to explore the effects of these differentially expressed genes (DEGs) on the proliferation and migration of dental pulp stem cells. Immunostaining revealed that TRPC channels are expressed at varying levels during odontogenesis. The identified genes represent novel candidates that are likely to be vital for rat tooth germ development. Together, the results provide a valuable resource to elucidate the gene regulatory mechanisms underlying mammalian tooth germ development. PMID:28706494

Immunohistochemical profile of cytokines and growth factors expressed in vestibular schwannoma and in normal vestibular nerve tissue.

PubMed

Taurone, Samanta; Bianchi, Enrica; Attanasio, Giuseppe; Di Gioia, Cira; Ierinó, Rocco; Carubbi, Cecilia; Galli, Daniela; Pastore, Francesco Saverio; Giangaspero, Felice; Filipo, Roberto; Zanza, Christian; Artico, Marco

2015-07-01

Vestibular schwannomas, also known as acoustic neuromas, are benign tumors, which originate from myelin-forming Schwann cells. They develop in the vestibular branch of the eighth cranial nerve in the internal auditory canal or cerebellopontine angle. The clinical progression of the condition involves slow and progressive growth, eventually resulting in brainstem compression. The objective of the present study was to investigate the expression level and the localization of the pro-inflammatory cytokines, transforming growth factor-β1 (TGF-β1) interleukin (IL)-1β, IL-6 and tumor necrosis factor-α (TNF-α), as well as the adhesion molecules, intracellular adhesion molecule-1 and vascular endothelial growth factor (VEGF), in order to determine whether these factors are involved in the transformation and development of human vestibular schwannoma. The present study investigated whether changes in inflammation are involved in tumor growth and if so, the mechanisms underlying this process. The results of the current study demonstrated that pro-inflammatory cytokines, including TGF-β1, IL-1β and IL-6 exhibited increased expression in human vestibular schwannoma tissue compared with normal vestibular nerve samples. TNF-α was weakly expressed in Schwann cells, confirming that a lower level of this cytokine is involved in the proliferation of Schwann cells. Neoplastic Schwann cells produce pro-inflammatory cytokines that may act in an autocrine manner, stimulating cellular proliferation. In addition, the increased expression of VEGF in vestibular schwannoma compared with that in normal vestibular nerve tissue, suggests that this factor may induce neoplastic growth via the promotion of angiogenesis. The present findings suggest that inflammation may promote angiogenesis and consequently contribute to tumor progression. In conclusion, the results of the present study indicated that VEGF and pro-inflammatory cytokines may be potential therapeutic targets in vestibular schwannoma. Further studies are necessary to confirm the involvement of these factors in the growth of neoplasms and to develop inhibitors of pro-inflammatory cytokines as a potential treatment option in the future.

Spaceflight Enhances Cell Aggregation and Random Budding in Candida albicans

PubMed Central

Woolley, Christine M.; Barrila, Jennifer; Buchanan, Kent; McCracken, James; Inglis, Diane O.; Searles, Stephen C.; Nelman-Gonzalez, Mayra A.; Ott, C. Mark; Wilson, James W.; Pierson, Duane L.; Stefanyshyn-Piper, Heidemarie M.; Hyman, Linda E.; Nickerson, Cheryl A.

2013-01-01

This study presents the first global transcriptional profiling and phenotypic characterization of the major human opportunistic fungal pathogen, Candida albicans, grown in spaceflight conditions. Microarray analysis revealed that C. albicans subjected to short-term spaceflight culture differentially regulated 452 genes compared to synchronous ground controls, which represented 8.3% of the analyzed ORFs. Spaceflight-cultured C. albicans–induced genes involved in cell aggregation (similar to flocculation), which was validated by microscopic and flow cytometry analysis. We also observed enhanced random budding of spaceflight-cultured cells as opposed to bipolar budding patterns for ground samples, in accordance with the gene expression data. Furthermore, genes involved in antifungal agent and stress resistance were differentially regulated in spaceflight, including induction of ABC transporters and members of the major facilitator family, downregulation of ergosterol-encoding genes, and upregulation of genes involved in oxidative stress resistance. Finally, downregulation of genes involved in actin cytoskeleton was observed. Interestingly, the transcriptional regulator Cap1 and over 30% of the Cap1 regulon was differentially expressed in spaceflight-cultured C. albicans. A potential role for Cap1 in the spaceflight response of C. albicans is suggested, as this regulator is involved in random budding, cell aggregation, and oxidative stress resistance; all related to observed spaceflight-associated changes of C. albicans. While culture of C. albicans in microgravity potentiates a global change in gene expression that could induce a virulence-related phenotype, no increased virulence in a murine intraperitoneal (i.p.) infection model was observed under the conditions of this study. Collectively, our data represent an important basis for the assessment of the risk that commensal flora could play during human spaceflight missions. Furthermore, since the low fluid-shear environment of microgravity is relevant to physical forces encountered by pathogens during the infection process, insights gained from this study could identify novel infectious disease mechanisms, with downstream benefits for the general public. PMID:24324620

Involvement of PUMA in pericyte migration induced by methamphetamine.

PubMed

Zhang, Yanhong; Zhang, Yuan; Bai, Ying; Chao, Jie; Hu, Gang; Chen, Xufeng; Yao, Honghong

2017-07-01

Mounting evidence indicates that methamphetamine causes blood-brain barrier damage, with emphasis on endothelial cells. The role of pericytes in methamphetamine-induced BBB damage remains unknown. Our study demonstrated that methamphetamine increased the migration of pericytes from the endothelial basement membrane. However, the detailed mechanisms underlying this process remain poorly understood. Thus, we examined the molecular mechanisms involved in methamphetamine-induced pericyte migration. The results showed that exposure of C3H/10T1/2 cells and HBVPs to methamphetamine increased PUMA expression via activation of the sigma-1 receptor, MAPK and Akt/PI3K pathways. Moreover, methamphetamine treatment resulted in the increased migration of C3H/10T1/2 cells and HBVPs. Knockdown of PUMA in pericytes transduced with PUMA siRNA attenuated the methamphetamine-induced increase in cell migration through attenuation of integrin and tyrosine kinase mechanisms, implicating a role of PUMA in the migration of C3H/10T1/2 cells and HBVPs. This study has demonstrated that methamphetamine-mediated pericytes migration involves PUMA up-regulation. Thus, targeted studies of PUMA could provide insights to facilitate the development of a potential therapeutic approach for alleviation of methamphetamine-induced pericyte migration. Copyright © 2017. Published by Elsevier Inc.

VBNC Legionella pneumophila cells are still able to produce virulence proteins.

PubMed

Alleron, Laëtitia; Khemiri, Arbia; Koubar, Mohamad; Lacombe, Christian; Coquet, Laurent; Cosette, Pascal; Jouenne, Thierry; Frere, Jacques

2013-11-01

Legionella pneumophila is the agent responsible for legionellosis. Numerous bacteria, including L. pneumophila, can enter into a viable but not culturable (VBNC) state under unfavorable environmental conditions. In this state, cells are unable to form colonies on standard medium but are still alive. Here we show that VBNC L. pneumophila cells, obtained by monochloramine treatment, were still able to synthesize proteins, some of which are involved in virulence. Protein synthesis was measured using (35)S-labeling and the proteomes of VBNC and culturable cells then compared. This analysis allowed the identification of nine proteins that were accumulated in the VBNC state. Among them, four were involved in virulence, i.e., the macrophage infectivity potentiator protein, the hypothetical protein lpl2247, the ClpP protease proteolytic subunit and the 27 kDa outer membrane protein. Others, i.e., the enoyl reductase, the electron transfer flavoprotein (alpha and beta subunits), the 50S ribosomal proteins (L1 and L25) are involved in metabolic and energy production pathways. However, resuscitation experiments performed with Acanthamoeba castellanii failed, suggesting that the accumulation of virulence factors by VBNC cells is not sufficient to maintain their virulence. Copyright © 2013 Elsevier Ltd. All rights reserved.

Cell separation using electric fields

NASA Technical Reports Server (NTRS)

Eppich, Henry M. (Inventor); Mangano, Joseph A. (Inventor)

2003-01-01

The present invention involves methods and devices which enable discrete objects having a conducting inner core, surrounded by a dielectric membrane to be selectively inactivated by electric fields via irreversible breakdown of their dielectric membrane. One important application of the invention is in the selection, purification, and/or purging of desired or undesired biological cells from cell suspensions. According to the invention, electric fields can be utilized to selectively inactivate and render non-viable particular subpopulations of cells in a suspension, while not adversely affecting other desired subpopulations. According to the inventive methods, the cells can be selected on the basis of intrinsic or induced differences in a characteristic electroporation threshold, which can depend, for example, on a difference in cell size and/or critical dielectric membrane breakdown voltage. The invention enables effective cell separation without the need to employ undesirable exogenous agents, such as toxins or antibodies. The inventive method also enables relatively rapid cell separation involving a relatively low degree of trauma or modification to the selected, desired cells. The inventive method has a variety of potential applications in clinical medicine, research, etc., with two of the more important foreseeable applications being stem cell enrichment/isolation, and cancer cell purging.

Adult T-cell leukemia/lymphoma with EBV-positive Hodgkin-like cells

PubMed Central

Venkataraman, Girish; Berkowitz, Jonathan; Morris, John C.; Janik, John E.; Raffeld, Mark A.; Pittaluga, Stefania

2011-01-01

SUMMARY Hodgkin-like cells (HLC) have been described in a variety of non-Hodgkin lymphomas (NHL) including chronic lymphocytic leukemia (CLL) and peripheral T-cell lymphoma (PTCL). There have been rare reports in the Japanese population of human T-cell lymphotrophic virus-1 (HTLV-1)-associated adult T-cell leukemia/lymphoma (ATLL) harboring HLC; however, no similar cases have been described in western patients. We report a 53-year-old African-American man that presented with progressive weakness and lethargy, and was found to have generalized lymphadenopathy and hypercalcemia. A lymph node biopsy showed involvement by ATLL with scattered Epstein-Barr virus (EBV)-positive cells, some of which resembled Hodgkin cells that had a B-cell phenotype, consistent with an Epstein-Barr virus-lymphoproliferative disorder (LPD). The patient had stage 4 disease with bone marrow involvement. In light of the associated B-cell lymphoproliferative process, the patient was treated with six cycles of intensive chemotherapy that targeted both the ATLL and the EBV-LPD that resulted in a complete response. An awareness of the association of EBV-LPD with Hodgkin-like cells in the context of ATLL is necessary to avoid potential misdiagnosis and to aid in therapeutic decisions. PMID:21315416

Role of hepatocyte growth factor in the development of dendritic cells from CD34+ bone marrow cells.

PubMed

Ovali, E; Ratip, S; Kibaroglu, A; Tekelioglu, Y; Cetiner, M; Karti, S; Aydin, F; Bayik, M; Akoglu, T

2000-05-01

Hepatocyte growth factor (HGF) is known to augment the effects of stem cell factor, interleukin-3, granulocyte-macrophage colony-stimulating factor (GM-CSF), erythropoetin, and granulocyte colony-stimulating factor, all of which are involved in hematopoiesis. HGF is also known to have a role in immune responses. The aim of this study was to investigate whether HGF is involved in the development of dendritic cells (DC) from CD34+ bone marrow cells. CD34+ cells obtained from three healthy donors were incubated in various combinations of HGF, GM-CSF, and tumor necrosis factor (TNF) for 12 days. Developing cell populations were analyzed for surface markers, morphology and functional capacities by flow cytometry, light microscopy and mixed lymphocyte reaction, respectively. Incubation with HGF alone generated greater number of dendritic cells from CD34+ bone marrow cells than incubation with GM-CSF, or a combination of GM-CSF with TNF. HGF was also found to potentiate the effect of GM-CSF on DC and monocyte development. The effects of HGF were inhibited by the concurrent use of TNF. HGF appears to be a significant factor in the development of dendritic cells from CD34+ bone marrow cells.

Cell separation using electric fields

NASA Technical Reports Server (NTRS)

Mangano, Joseph (Inventor); Eppich, Henry (Inventor)

2009-01-01

The present invention involves methods and devices which enable discrete objects having a conducting inner core, surrounded by a dielectric membrane to be selectively inactivated by electric fields via irreversible breakdown of their dielectric membrane. One important application of the invention is in the selection, purification, and/or purging of desired or undesired biological cells from cell suspensions. According to the invention, electric fields can be utilized to selectively inactivate and render non-viable particular subpopulations of cells in a suspension, while not adversely affecting other desired subpopulations. According to the inventive methods, the cells can be selected on the basis of intrinsic or induced differences in a characteristic electroporation threshold, which can depend, for example, on a difference in cell size and/or critical dielectric membrane breakdown voltage. The invention enables effective cell separation without the need to employ undesirable exogenous agents, such as toxins or antibodies. The inventive method also enables relatively rapid cell separation involving a relatively low degree of trauma or modification to the selected, desired cells. The inventive method has a variety of potential applications in clinical medicine, research, etc., with two of the more important foreseeable applications being stem cell enrichment/isolation, and cancer cell purging.

The role of the obestatin/GPR39 system in human gastric adenocarcinomas.

PubMed

Alén, Begoña O; Leal-López, Saúl; Alén, María Otero; Viaño, Patricia; García-Castro, Victoria; Mosteiro, Carlos S; Beiras, Andrés; Casanueva, Felipe F; Gallego, Rosalía; García-Caballero, Tomás; Camiña, Jesús P; Pazos, Yolanda

2016-02-02

Obestatin, a 23-amino acid peptide encoded by the ghrelin gene, and the GPR39 receptor were reported to be involved in the control of mitogenesis of gastric cancer cell lines; however, the relationship between the obestatin/GPR39 system and gastric cancer progression remains unknown. In the present study, we determined the expression levels of the obestatin/GPR39 system in human gastric adenocarcinomas and explored their potential functional roles. Twenty-eight patients with gastric adenocarcinomas were retrospectively studied, and clinical data were obtained. The role of obestatin/GPR39 in gastric cancer progression was studied in vitro using the human gastric adenocarcinoma AGS cell line. Obestatin exogenous administration in these GPR39-bearing cells deregulated the expression of several hallmarks of the epithelial-mesenchymal transition (EMT) and angiogenesis. Moreover, obestatin signaling promoted phenotypic changes via GPR39, increasingly impacting on the cell morphology, proliferation, migration and invasion of these cells. In healthy human stomachs, obestatin expression was observed in the neuroendocrine cells and GPR39 expression was localized mainly in the chief cells of the oxyntic glands. In human gastric adenocarcinomas, no obestatin expression was found; however, an aberrant pattern of GPR39 expression was discovered, correlating to the dedifferentiation of the tumor. Altogether, our data strongly suggest the involvement of the obestatin/GPR39 system in the pathogenesis and/or clinical outcome of human gastric adenocarcinomas and highlight the potential usefulness of GPR39 as a prognostic marker in gastric cancer.

The role of the obestatin/GPR39 system in human gastric adenocarcinomas

PubMed Central

Alén, Begoña O.; Leal-López, Saúl; Alén, María Otero; Viaño, Patricia; García-Castro, Victoria; Mosteiro, Carlos S.; Beiras, Andrés; Casanueva, Felipe F.; Gallego, Rosalía; García-Caballero, Tomás; Camiña, Jesús P.; Pazos, Yolanda

2016-01-01

Obestatin, a 23-amino acid peptide encoded by the ghrelin gene, and the GPR39 receptor were reported to be involved in the control of mitogenesis of gastric cancer cell lines; however, the relationship between the obestatin/GPR39 system and gastric cancer progression remains unknown. In the present study, we determined the expression levels of the obestatin/GPR39 system in human gastric adenocarcinomas and explored their potential functional roles. Twenty-eight patients with gastric adenocarcinomas were retrospectively studied, and clinical data were obtained. The role of obestatin/GPR39 in gastric cancer progression was studied in vitro using the human gastric adenocarcinoma AGS cell line. Obestatin exogenous administration in these GPR39-bearing cells deregulated the expression of several hallmarks of the epithelial-mesenchymal transition (EMT) and angiogenesis. Moreover, obestatin signaling promoted phenotypic changes via GPR39, increasingly impacting on the cell morphology, proliferation, migration and invasion of these cells. In healthy human stomachs, obestatin expression was observed in the neuroendocrine cells and GPR39 expression was localized mainly in the chief cells of the oxyntic glands. In human gastric adenocarcinomas, no obestatin expression was found; however, an aberrant pattern of GPR39 expression was discovered, correlating to the dedifferentiation of the tumor. Altogether, our data strongly suggest the involvement of the obestatin/GPR39 system in the pathogenesis and/or clinical outcome of human gastric adenocarcinomas and highlight the potential usefulness of GPR39 as a prognostic marker in gastric cancer. PMID:26716511

Characterization of CD34+ hematopoietic cells in systemic mastocytosis: Potential role in disease dissemination.

PubMed

Mayado, A; Teodosio, C; Dasilva-Freire, N; Jara-Acevedo, M; Garcia-Montero, A C; Álvarez-Twose, I; Sánchez-Muñoz, L; Matito, A; Caldas, C; Muñoz-González, J I; Henriques, A; Sánchez-Gallego, J I; Escribano, L; Orfao, A

2018-01-13

Recent studies show that most systemic mastocytosis (SM) patients, including indolent SM (ISM) with (ISMs+) and without skin lesions (ISMs-), carry the KIT D816V mutation in PB leukocytes. We investigated the potential association between the degree of involvement of BM hematopoiesis by the KIT D816V mutation and the distribution of different maturation-associated compartments of bone marrow (BM) and peripheral blood (PB) CD34 + hematopoietic precursors (HPC) in ISM and identified the specific PB cell compartments that carry this mutation. The distribution of different maturation-associated subsets of BM and PB CD34 + HPC from 64 newly diagnosed (KIT-mutated) ISM patients and 14 healthy controls was analyzed by flow cytometry. In 18 patients, distinct FACS-purified PB cell compartments were also investigated for the KIT mutation. ISM patients showed higher percentages of both BM and PB MC-committed CD34 + HPC vs controls, particularly among ISM cases with MC-restricted KIT mutation (ISM MC ); this was associated with progressive blockade of maturation of CD34 + HPC to the neutrophil lineage from ISM MC to multilineage KIT-mutated cases (ISM ML ). Regarding the frequency of KIT-mutated cases and cell populations in PB, variable patterns were observed, the percentage of KIT-mutated PB CD34 + HPC, eosinophils, neutrophils, monocytes and T cells increasing from ISMs- MC and ISMs+ MC to ISM ML patients. The presence of the KIT D816V mutation in PB of ISM patients is associated with (early) involvement of circulating CD34 + HPC and multiple myeloid cell subpopulations, KIT-mutated PB CD34 + HPC potentially contributing to early dissemination of the disease. © 2018 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.

Temporal Profiling and Pulsed SILAC Labeling Identify Novel Secreted Proteins During Ex Vivo Osteoblast Differentiation of Human Stromal Stem Cells*

PubMed Central

Kristensen, Lars P.; Chen, Li; Nielsen, Maria Overbeck; Qanie, Diyako W.; Kratchmarova, Irina; Kassem, Moustapha; Andersen, Jens S.

2012-01-01

It is well established that bone forming cells (osteoblasts) secrete proteins with autocrine, paracrine, and endocrine function. However, the identity and functional role for the majority of these secreted and differentially expressed proteins during the osteoblast (OB) differentiation process, is not fully established. To address these questions, we quantified the temporal dynamics of the human stromal (mesenchymal, skeletal) stem cell (hMSC) secretome during ex vivo OB differentiation using stable isotope labeling by amino acids in cell culture (SILAC). In addition, we employed pulsed SILAC labeling to distinguish genuine secreted proteins from intracellular contaminants. We identified 466 potentially secreted proteins that were quantified at 5 time-points during 14-days ex vivo OB differentiation including 41 proteins known to be involved in OB functions. Among these, 315 proteins exhibited more than 2-fold up or down-regulation. The pulsed SILAC method revealed a strong correlation between the fraction of isotope labeling and the subset of proteins known to be secreted and involved in OB differentiation. We verified SILAC data using qRT-PCR analysis of 9 identified potential novel regulators of OB differentiation. Furthermore, we studied the biological effects of one of these proteins, the hormone stanniocalcin 2 (STC2) and demonstrated its autocrine effects in enhancing osteoblastic differentiation of hMSC. In conclusion, combining complete and pulsed SILAC labeling facilitated the identification of novel factors produced by hMSC with potential role in OB differentiation. Our study demonstrates that the secretome of osteoblastic cells is more complex than previously reported and supports the emerging evidence that osteoblastic cells secrete proteins with endocrine functions and regulate cellular processes beyond bone formation. PMID:22801418

Grewia tiliaefolia and its active compound vitexin regulate the expression of glutamate transporters and protect Neuro2a cells from glutamate toxicity.

PubMed

Malar, Dicson Sheeja; Prasanth, Mani Iyer; Shafreen, Rajamohamed Beema; Balamurugan, Krishnaswamy; Devi, Kasi Pandima

2018-04-25

Glutamate is a major neurotransmitter involved in several brain functions and glutamate excitotoxicity is involved in Alzheimer's disease (AD). In the current study, the neuroprotective effect of the Indian medicinal plant Grewia tiliaefolia (GT) and its active component vitexin was evaluated in Neuro-2a cells against glutamate toxicity. Neuro-2a cells were exposed to glutamate to cause excitotoxicity and the neuroprotective effect of GT and vitexin were evaluated using biochemical studies (estimation of reactive oxygen species, reactive nitrogen species, protein carbonyl content, lipid peroxidation level, mitochondrial membrane potential and caspase-3 activity), molecular docking studies, gene expression and western blot analysis. Glutamate exposure to Neuro-2a cells induced oxidative stress, loss of membrane potential, suppressed the expression of antioxidant response genes (Nrf-2, HO-1, NQO-1), glutamate transporters (GLAST-1, GLT-1) and induced the expression of NMDAR, Calpain. However, pre-treatment of cells with GT/vitexin inhibited oxidative stress mediated damage by augmenting the expression of Nrf-2/HO-1 pathway, inducing the expression of glutamate transporters and downregulating Calpain, NMDAR. Molecular docking showed that vitexin effectively binds to NMDAR and GSK-3β and thereby can inhibit their activation. GT/vitexin also inhibited glutamate induced Bax expression. Methanol extract of G. tiliaefolia and its active component vitexin can act in an antioxidant dependent mechanism as well as by regulating glutamate in mitigating the toxicity exerted by glutamate in Neuro-2a cells. Our results conclude that GT/vitexin can act as potential drug leads for the therapeutic intervention of AD. Copyright © 2017. Published by Elsevier Inc.

Involvement of nitric oxide synthase in matrix metalloproteinase-9- and/or urokinase plasminogen activator receptor-mediated glioma cell migration

PubMed Central

2013-01-01

Background Src tyrosine kinase activates inducible nitric oxide synthase (iNOS) and, in turn, nitric oxide production as a means to transduce cell migration. Src tyrosine kinase plays a key proximal role to control α9β1 signaling. Our recent studies have clearly demonstrated the role of α9β1 integrin in matrix metalloproteinase-9 (MMP-9) and/or urokinase plasminogen activator receptor (uPAR)-mediated glioma cell migration. In the present study, we evaluated the involvement of α9β1 integrin-iNOS pathway in MMP-9- and/or uPAR-mediated glioma cell migration. Methods MMP-9 and uPAR shRNAs and overexpressing plasmids were used to downregulate and upregulate these molecules, respectively in U251 glioma cells and 5310 glioma xenograft cells. The effect of treatments on migration and invasion potential of these glioma cells were assessed by spheroid migration, wound healing, and Matrigel invasion assays. In order to attain the other objectives we also performed immunocytochemical, immunohistochemical, RT-PCR, Western blot and fluorescence-activated cell sorting (FACS) analysis. Results Immunohistochemical analysis revealed the prominent association of iNOS with glioblastoma multiforme (GBM). Immunofluorescence analysis showed prominent expression of iNOS in glioma cells. MMP-9 and/or uPAR knockdown by respective shRNAs reduced iNOS expression in these glioma cells. RT-PCR analysis revealed elevated iNOS mRNA expression in either MMP-9 or uPAR overexpressed glioma cells. The migration potential of MMP-9- and/or uPAR-overexpressed U251 glioma cells was significantly inhibited after treatment with L-NAME, an inhibitor of iNOS. Similarly, a significant inhibition of the invasion potential of the control or MMP-9/uPAR-overexpressed glioma cells was noticed after L-NAME treatment. A prominent reduction of iNOS expression was observed in the tumor regions of nude mice brains, which were injected with 5310 glioma cells, after MMP-9 and/or uPAR knockdown. Protein expressions of cSrc, phosphoSrc and p130Cas were reduced with simultaneous knockdown of both MMP-9 and uPAR. Conclusions Taken together, our results from the present and earlier studies clearly demonstrate that α9β1 integrin-mediated cell migration utilizes the iNOS pathway, and inhibition of the migratory potential of glioma cells by simultaneous knockdown of MMP-9 and uPAR could be attributed to the reduced α9β1 integrin and iNOS levels. PMID:24325546

Stimulators of Mineralization Limit the Invasive Phenotype of Human Osteosarcoma Cells by a Mechanism Involving Impaired Invadopodia Formation

PubMed Central

Cmoch, Anna; Podszywalow-Bartnicka, Paulina; Palczewska, Malgorzata; Piwocka, Katarzyna; Groves, Patrick; Pikula, Slawomir

2014-01-01

Background Osteosarcoma (OS) is a highly aggressive bone cancer affecting children and young adults. Growing evidence connects the invasive potential of OS cells with their ability to form invadopodia (structures specialized in extracellular matrix proteolysis). Results In this study, we tested the hypothesis that commonly used in vitro stimulators of mineralization limit the invadopodia formation in OS cells. Here we examined the invasive potential of human osteoblast-like cells (Saos-2) and osteolytic-like (143B) OS cells treated with the stimulators of mineralization (ascorbic acid and B-glycerophosphate) and observed a significant difference in response of the tested cells to the treatment. In contrast to 143B cells, osteoblast-like cells developed a mineralization phenotype that was accompanied by a decreased proliferation rate, prolongation of the cell cycle progression and apoptosis. On the other hand, stimulators of mineralization limited osteolytic-like OS cell invasiveness into collagen matrix. We are the first to evidence the ability of 143B cells to degrade extracellular matrix to be driven by invadopodia. Herein, we show that this ability of osteolytic-like cells in vitro is limited by stimulators of mineralization. Conclusions Our study demonstrates that mineralization competency determines the invasive potential of cancer cells. A better understanding of the molecular mechanisms by which stimulators of mineralization regulate and execute invadopodia formation would reveal novel clinical targets for treating osteosarcoma. PMID:25314307

Embryonic Stem Cells Promoting Macrophage Survival and Function are Crucial for Teratoma Development

PubMed Central

Chen, Tianxiang; Wang, Xi; Guo, Lei; Wu, Mingmei; Duan, Zhaoxia; Lv, Jing; Tai, Wenjiao; Renganathan, Hemamalini; Didier, Ruth; Li, Jinhua; Sun, Dongming; Chen, Xiaoming; He, Xijing; Fan, Jianqing; Young, Wise; Ren, Yi

2014-01-01

Stem cell therapies have had tremendous potential application for many diseases in recent years. However, the tumorigenic properties of stem cells restrict their potential clinical application; therefore, strategies for reducing the tumorigenic potential of stem cells must be established prior to transplantation. We have demonstrated that syngeneic transplantation of embryonic stem cells (ESCs) provokes an inflammatory response that involves the rapid recruitment of bone marrow-derived macrophages (BMDMs). ESCs are able to prevent mature macrophages from macrophage colony-stimulating factor (M-CSF) withdrawal-induced apoptosis, and thus prolong macrophage lifespan significantly by blocking various apoptotic pathways in an M-CSF-independent manner. ESCs express and secrete IL-34, which may be responsible for ESC-promoted macrophage survival. This anti-apoptotic effect of ESCs involves activation of extracellular signal-regulated kinase (ERK)1/2 and PI3K/Akt pathways and thus, inhibition of ERK1/2 and PI3K/AKT activation decreases ESC-induced macrophage survival. Functionally, ESC-treated macrophages also showed a higher level of phagocytic activity. ESCs further serve to polarize BMDMs into M2-like macrophages that exhibit most tumor-associated macrophage phenotypic and functional features. ESC-educated macrophages produce high levels of arginase-1, Tie-2, and TNF-α, which participate in angiogenesis and contribute to teratoma progression. Our study suggests that induction of M2-like macrophage activation is an important mechanism for teratoma development. Strategies targeting macrophages to inhibit teratoma development would increase the safety of ESC-based therapies, inasmuch as the depletion of macrophages completely inhibits ESC-induced angiogenesis and teratoma development. PMID:25071759

Frontrunners of T cell activation: Initial, localized Ca2+ signals mediated by NAADP and the type 1 ryanodine receptor.

PubMed

Wolf, Insa M A; Diercks, Björn-Philipp; Gattkowski, Ellen; Czarniak, Frederik; Kempski, Jan; Werner, René; Schetelig, Daniel; Mittrücker, Hans-Willi; Schumacher, Valéa; von Osten, Manuel; Lodygin, Dimitri; Flügel, Alexander; Fliegert, Ralf; Guse, Andreas H

2015-10-13

The activation of T cells is the fundamental on switch for the adaptive immune system. Ca(2+) signaling is essential for T cell activation and starts as initial, short-lived, localized Ca(2+) signals. The second messenger nicotinic acid adenine dinucleotide phosphate (NAADP) forms rapidly upon T cell activation and stimulates early Ca(2+) signaling. We developed a high-resolution imaging technique using multiple fluorescent Ca(2+) indicator dyes to characterize these early signaling events and investigate the channels involved in NAADP-dependent Ca(2+) signals. In the first seconds of activation of either primary murine T cells or human Jurkat cells with beads coated with an antibody against CD3, we detected Ca(2+) signals with diameters close to the limit of detection and that were close to the activation site at the plasma membrane. In Jurkat cells in which the ryanodine receptor (RyR) was knocked down or in primary T cells from RyR1(-/-) mice, either these early Ca(2+) signals were not detected or the number of signals was markedly reduced. Local Ca(2+) signals observed within 20 ms upon microinjection of Jurkat cells with NAADP were also sensitive to RyR knockdown. In contrast, TRPM2 (transient receptor potential channel, subtype melastatin 2), a potential NAADP target channel, was not required for the formation of initial Ca(2+) signals in primary T cells. Thus, through our high-resolution imaging method, we characterized early Ca(2+) release events in T cells and obtained evidence for the involvement of RyR and NAADP in such signals. Copyright © 2015, American Association for the Advancement of Science.

Roles of specific Kv channel types in repolarization of the action potential in genetically identified subclasses of pyramidal neurons in mouse neocortex

PubMed Central

Pathak, Dhruba; Guan, Dongxu

2016-01-01

The action potential (AP) is a fundamental feature of excitable cells that serves as the basis for long-distance signaling in the nervous system. There is considerable diversity in the appearance of APs and the underlying repolarization mechanisms in different neuronal types (reviewed in Bean BP. Nat Rev Neurosci 8: 451–465, 2007), including among pyramidal cell subtypes. In the present work, we used specific pharmacological blockers to test for contributions of Kv1, Kv2, or Kv4 channels to repolarization of single APs in two genetically defined subpopulations of pyramidal cells in layer 5 of mouse somatosensory cortex (etv1 and glt) as well as pyramidal cells from layer 2/3. These three subtypes differ in AP properties (Groh A, Meyer HS, Schmidt EF, Heintz N, Sakmann B, Krieger P. Cereb Cortex 20: 826–836, 2010; Guan D, Armstrong WE, Foehring RC. J Neurophysiol 113: 2014–2032, 2015) as well as laminar position, morphology, and projection targets. We asked what the roles of Kv1, Kv2, and Kv4 channels are in AP repolarization and whether the underlying mechanisms are pyramidal cell subtype dependent. We found that Kv4 channels are critically involved in repolarizing neocortical pyramidal cells. There are also pyramidal cell subtype-specific differences in the role for Kv1 channels. Only Kv4 channels were involved in repolarizing the narrow APs of glt cells. In contrast, in etv1 cells and layer 2/3 cells, the broader APs are partially repolarized by Kv1 channels in addition to Kv4 channels. Consistent with their activation in the subthreshold range, Kv1 channels also regulate AP voltage threshold in all pyramidal cell subtypes. PMID:26864770

Single-Nucleotide Polymorphisms of the MSH2 and MLH1 Genes, Potential Molecular Markers for Susceptibility to the Development of Basal Cell Carcinoma in the Brazilian Population.

PubMed

da Silva Calixto, Poliane; Lopes, Otávio Sérgio; Dos Santos Maia, Mayara; Herrero, Sylvia Satomi Takeno; Longui, Carlos Alberto; Melo, Cynthia Germoglio Farias; de Carvalho Filho, Ivan Rodrigues; Soares, Leonardo Ferreira; de Medeiros, Arnaldo Correia; Delatorre, Plínio; Khayat, André Salim; Burbano, Rommel Rodriguez; Lima, Eleonidas Moura

2018-07-01

Basal cell carcinoma - BCC is considered a multifactorial neoplasm involving genetic, epigenetic and environmental factors. Where UVB radiation is considered the main physical agent involved in BCC carcinogenesis. The Brazil and state of Paraíba are exposed to high levels of UVB rays. The mismatch repair - MMR is important DNA repair mechanisms to maintain replication fidelity. Therefore, single nucleotide polymorphisms (SNPs) in genes encoding proteins involved in MMR may be potential molecular markers of susceptibility to BCC. The objective of this study was to evaluate and describe for the first time the SNPs rs560246973, rs2303425 and rs565410865 and risk of developing BCC. The present study analyzed 100 samples of paraffin-embedded tissue from patients with histopathological diagnosis of BCC and 100 control samples. The results were obtained by genotyping method, Dideoxy Unique Allele Specific - PCR (DSASP). The SNPs rs2303425 were not associated with Basal Cell Carcinoma. However, the SNPs rs560246973 and rs565410865 was shown to be associated with the development of BCC when compared to control samples (P < 0.0001). The SNPs rs565410865 was also statistical significance between the genotypes of and the age group (p = 0.0027) and tumor location (p = 0,0191). The result suggests that SNPs rs2303425 and rs565410865 are associated with susceptibility to the development of BCC in the Brazilian population and may be considered as potential molecular markers for BCC.

The natural compound forskolin synergizes with dexamethasone to induce cell death in myeloma cells via BIM.

PubMed

Follin-Arbelet, Virginie; Misund, Kristine; Naderi, Elin Hallan; Ugland, Hege; Sundan, Anders; Blomhoff, Heidi Kiil

2015-08-26

We have previously demonstrated that activation of the cyclic adenosine monophosphate (cAMP) pathway kills multiple myeloma (MM) cells both in vitro and in vivo. In the present study we have investigated the potential of enhancing the killing of MM cell lines and primary MM cells by combining the cAMP-elevating compound forskolin with the commonly used MM therapeutic drugs melphalan, cyclophosphamide, doxorubicin, bortezomib and dexamethasone. We observed that forskolin potentiated the killing induced by all the tested agents as compared to treatment with the single agents alone. In particular, forskolin had a synergistic effect on the dexamethasone-responsive cell lines H929 and OM-2. By knocking down the proapoptotic BCL-2 family member BIM, we proved this protein to be involved in the synergistic induction of apoptosis by dexamethasone and forskolin. The ability of forskolin to maintain the killing of MM cells even at lower concentrations of the conventional agents suggests that forskolin may be used to diminish treatment-associated side effects. Our findings support a potential role of forskolin in combination with current conventional agents in the treatment of MM.

The natural compound forskolin synergizes with dexamethasone to induce cell death in myeloma cells via BIM

PubMed Central

Follin-Arbelet, Virginie; Misund, Kristine; Hallan Naderi, Elin; Ugland, Hege; Sundan, Anders; Kiil Blomhoff, Heidi

2015-01-01

We have previously demonstrated that activation of the cyclic adenosine monophosphate (cAMP) pathway kills multiple myeloma (MM) cells both in vitro and in vivo. In the present study we have investigated the potential of enhancing the killing of MM cell lines and primary MM cells by combining the cAMP-elevating compound forskolin with the commonly used MM therapeutic drugs melphalan, cyclophosphamide, doxorubicin, bortezomib and dexamethasone. We observed that forskolin potentiated the killing induced by all the tested agents as compared to treatment with the single agents alone. In particular, forskolin had a synergistic effect on the dexamethasone-responsive cell lines H929 and OM-2. By knocking down the proapoptotic BCL-2 family member BIM, we proved this protein to be involved in the synergistic induction of apoptosis by dexamethasone and forskolin. The ability of forskolin to maintain the killing of MM cells even at lower concentrations of the conventional agents suggests that forskolin may be used to diminish treatment-associated side effects. Our findings support a potential role of forskolin in combination with current conventional agents in the treatment of MM. PMID:26306624

Compact, Automated, Frequency-Agile Microspectrofluorimeter

NASA Technical Reports Server (NTRS)

Fernandez, Salvador M.; Guignon, Ernest F.

1995-01-01

Compact, reliable, rugged, automated cell-culture and frequency-agile microspectrofluorimetric apparatus developed to perform experiments involving photometric imaging observations of single live cells. In original application, apparatus operates mostly unattended aboard spacecraft; potential terrestrial applications include automated or semiautomated diagnosis of pathological tissues in clinical laboratories, biomedical instrumentation, monitoring of biological process streams, and portable instrumentation for testing biological conditions in various environments. Offers obvious advantages over present laboratory instrumentation.

Human embryonic stem cell-derived mesodermal progenitors display substantially increased tissue formation compared to human mesenchymal stem cells under dynamic culture conditions in a packed bed/column bioreactor.

PubMed

de Peppo, Giuseppe Maria; Sladkova, Martina; Sjövall, Peter; Palmquist, Anders; Oudina, Karim; Hyllner, Johan; Thomsen, Peter; Petite, Hervé; Karlsson, Camilla

2013-01-01

Bone tissue engineering represents a promising strategy to obviate bone deficiencies, allowing the ex vivo construction of bone substitutes with unprecedented potential in the clinical practice. Considering that in the human body cells are constantly stimulated by chemical and mechanical stimuli, the use of bioreactor is emerging as an essential factor for providing the proper environment for the reproducible and large-scale production of the engineered substitutes. Human mesenchymal stem cells (hMSCs) are experimentally relevant cells but, regardless the encouraging results reported after culture under dynamic conditions in bioreactors, show important limitations for tissue engineering applications, especially considering their limited proliferative potential, loss of functionality following protracted expansion, and decline in cellular fitness associated with aging. On the other hand, we previously demonstrated that human embryonic stem cell-derived mesodermal progenitors (hES-MPs) hold great potential to provide a homogenous and unlimited source of cells for bone engineering applications. Based on prior scientific evidence using different types of stem cells, in the present study we hypothesized that dynamic culture of hES-MPs in a packed bed/column bioreactor had the potential to affect proliferation, expression of genes involved in osteogenic differentiation, and matrix mineralization, therefore resulting in increased bone-like tissue formation. The reported findings suggest that hES-MPs constitute a suitable alternative cell source to hMSCs and hold great potential for the construction of bone substitutes for tissue engineering applications in clinical settings.

Burn Wound gammadelta T-Cells Support a Th2 and Th17 Immune Response

DTIC Science & Technology

2014-02-01

disorder (rheumatoid arthritis), psoriasis , and graft vs host disease.24–27 Gamma-δ T-cells are functionally specialized and are involved in...Mathers AR, Ferris LK. Anti-cytokine therapy in the treatment of psoriasis . Cytokine 2013;61:704–12. 26. Greenblatt MB, Vrbanac V, Vbranac V, et al...Meglio P, Perera GK, et al. Identification of a novel proinflammatory human skin-homing V?9Vd2 T cell subset with a potential role in psoriasis . J

Differential Alterations in Excitatory and Inhibitory Networks Involving Dentate Granule Cells Following Chronic Treatment with Distinct Classes of NMDAR Antagonists in Hippocampal Slice Cultures

DTIC Science & Technology

2010-03-08

1992; Jung and McNaughton, 1993); (2) low incidence of recurrent excitatory synapses between granule cells (Molnar and Nadler, 1999; Okazaki et al...neurons, dentate granule cells have a relatively more negative resting membrane potential and exhibit low-frequency firing (Staley et al., 1992; Jung ...inhibition plays a dual role in brain function and possibly seizure occurrence through balancing excitation and synchronizing neuronal firing. An

EVEN VISITING SCIENTISTS COULD MAKE DISCOVERIES IN MONTREAL.

PubMed

Lázár, György

2014-03-30

This publication summarizes the scientific adventure with Professor Selye, and focuses on the specific effect of rare metal salts on reticuloendothelial functions. Rare earth metal ions markedly affect the functions of cells involved in inflammatory and immunological phenomena. The Kupffer cell blockade induced by GdCl3 is a generally accepted method for investigation of the physiological and pathophysiological roles of Kupffer cells. Potential beneficial effects of macrophage blockade have been demonstrated in different shock states, liver injury and obstructive jaundice.

Influences of innate immunity, autophagy, and fibroblast activation in the pathogenesis of lung fibrosis

PubMed Central

O'Dwyer, David N.; Ashley, Shanna L.

2016-01-01

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease characterized by accumulation of extracellular matrix (ECM) and impaired gas exchange. The pathobiological mechanisms that account for disease progression are poorly understood but likely involve alterations in innate inflammatory cells, epithelial cells, and fibroblasts. Thus we seek to review the most recent literature highlighting the complex roles of neutrophils and macrophages as both promoters of fibrosis and defenders against infection. With respect to epithelial cells and fibroblasts, we review the data suggesting that defective autophagy promotes the fibrogenic potential of both cell types and discuss new evidence related to matrix metalloproteinases, growth factors, and cellular metabolism in the form of lactic acid generation that may have consequences for promoting fibrogenesis. We discuss potential cross talk between innate and structural cell types and also highlight literature that may help explain the limitations of current IPF therapies. PMID:27474089

Influences of innate immunity, autophagy, and fibroblast activation in the pathogenesis of lung fibrosis.

PubMed

O'Dwyer, David N; Ashley, Shanna L; Moore, Bethany B

2016-09-01

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease characterized by accumulation of extracellular matrix (ECM) and impaired gas exchange. The pathobiological mechanisms that account for disease progression are poorly understood but likely involve alterations in innate inflammatory cells, epithelial cells, and fibroblasts. Thus we seek to review the most recent literature highlighting the complex roles of neutrophils and macrophages as both promoters of fibrosis and defenders against infection. With respect to epithelial cells and fibroblasts, we review the data suggesting that defective autophagy promotes the fibrogenic potential of both cell types and discuss new evidence related to matrix metalloproteinases, growth factors, and cellular metabolism in the form of lactic acid generation that may have consequences for promoting fibrogenesis. We discuss potential cross talk between innate and structural cell types and also highlight literature that may help explain the limitations of current IPF therapies. Copyright © 2016 the American Physiological Society.

Channels, pumps, and exchangers in the gill and kidney of freshwater fishes: their role in ionic and acid-base regulation.

PubMed

Perry, S F; Shahsavarani, A; Georgalis, T; Bayaa, M; Furimsky, M; Thomas, S L Y

2003-11-01

In freshwater fishes, the gill and kidney are intricately involved in ionic and acid-base regulation owing to the presence of numerous ion channels, pumps, or exchangers. This review summarizes recent developments in branchial and renal ion transport physiology and presents several models that integrate epithelial ion and acid-base movements in freshwater fishes. At the gill, three cell types are potentially involved in ionic uptake: pavement cells, mitochondria-rich (MR) PNA(+) cells, and MR PNA(-) cells. The transfer of acidic or basic equivalents between the fish and its environment is accomplished largely by the gill and is appropriately regulated to correct acid-base imbalances. The kidney, while less important than the gill in overall acid or base excretion, has an essential role in regulating systemic acid-base balance by controlling HCO(3) (-) reabsorption from the filtrate. Copyright 2003 Wiley-Liss, Inc.

Molecular pathogenesis of splenic and nodal marginal zone lymphoma.

PubMed

Spina, Valeria; Rossi, Davide

Genomic studies have improved our understanding of the biological basis of splenic (SMZL) and nodal (NMZL) marginal zone lymphoma by providing a comprehensive and unbiased view of the genes/pathways that are deregulated in these diseases. Consistent with the physiological involvement of NOTCH, NF-κB, B-cell receptor and toll-like receptor signaling in mature B-cells differentiation into the marginal zone B-cells, many oncogenic mutations of genes involved in these pathways have been identified in SMZL and NMZL. Beside genetic lesions, also epigenetic and post-transcriptional modifications contribute to the deregulation of marginal zone B-cell differentiation pathways in SMZL and NMZL. This review describes the progress in understanding the molecular mechanism underlying SMZL and NMZL, including molecular and post-transcriptional modifications, and discusses how information gained from these efforts has provided new insights on potential targets of diagnostic, prognostic and therapeutic relevance in SMZL and NMZL. Copyright © 2016 Elsevier Ltd. All rights reserved.

The meaning of PIWI proteins in cancer development.

PubMed

Litwin, Monika; Szczepańska-Buda, Anna; Piotrowska, Aleksandra; Dzięgiel, Piotr; Witkiewicz, Wojciech

2017-05-01

Cancer is a histologically and genetically heterogeneous population of tumor cells that exhibits distinct molecular profiles determined by epigenetic alterations. P-element-induced wimpy testis (PIWI) proteins in complex with PIWI-interacting RNA (piRNA) have been previously demonstrated to be involved in epigenetic regulation in germline cells. Recently, reactivation of PIWI expression, primarily PIWI-like protein 1 and 2, through aberrant DNA methylation resulting in genomic silencing has been identified in various types of tumors. It has been suggested that the PIWI-piRNA complex contributes to cancer development and progression by promoting a stem-like state of cancer cells, or cancer stem cells (CSCs). It has been identified that CSCs represent the cells that have undergone epithelial-mesenchymal transition (EMT) and acquired metastatic capacities. However, the molecular association between the EMT process and the stem-cell state remains unclear. Further extensive characterization of CSCs in individual types of tumors is required to identify specific markers for the heterogeneous population of CSCs and therefore selectively target CSCs. Previous studies indicate a reciprocal regulation between PIWI proteins and a complex signaling network linking markers characterized for CSCs and transcription factors involved in EMT. In the present review, studies of PIWI function are summarized, and the potential involvement of PIWI proteins in cancer development and progression is discussed.

The meaning of PIWI proteins in cancer development

PubMed Central

Litwin, Monika; Szczepańska-Buda, Anna; Piotrowska, Aleksandra; Dzięgiel, Piotr; Witkiewicz, Wojciech

2017-01-01

Cancer is a histologically and genetically heterogeneous population of tumor cells that exhibits distinct molecular profiles determined by epigenetic alterations. P-element-induced wimpy testis (PIWI) proteins in complex with PIWI-interacting RNA (piRNA) have been previously demonstrated to be involved in epigenetic regulation in germline cells. Recently, reactivation of PIWI expression, primarily PIWI-like protein 1 and 2, through aberrant DNA methylation resulting in genomic silencing has been identified in various types of tumors. It has been suggested that the PIWI-piRNA complex contributes to cancer development and progression by promoting a stem-like state of cancer cells, or cancer stem cells (CSCs). It has been identified that CSCs represent the cells that have undergone epithelial-mesenchymal transition (EMT) and acquired metastatic capacities. However, the molecular association between the EMT process and the stem-cell state remains unclear. Further extensive characterization of CSCs in individual types of tumors is required to identify specific markers for the heterogeneous population of CSCs and therefore selectively target CSCs. Previous studies indicate a reciprocal regulation between PIWI proteins and a complex signaling network linking markers characterized for CSCs and transcription factors involved in EMT. In the present review, studies of PIWI function are summarized, and the potential involvement of PIWI proteins in cancer development and progression is discussed. PMID:28529570

Diamond-Nanoneedle-Array-Facilitated Intracellular Delivery and the Potential Influence on Cell Physiology.

PubMed

Zhu, Xiaoyue; Yuen, Muk Fung; Yan, Li; Zhang, Zhenyu; Ai, Fujin; Yang, Yang; Yu, Peter K N; Zhu, Guangyu; Zhang, Wenjun; Chen, Xianfeng

2016-05-01

Vertical arrays of nanostructures can provide access to the cell cytoplasma and probe intracellular molecules. Here, the simple combination of diamond nanoneedle arrays with centrifugation-induced supergravity is shown to efficiently deliver drugs and biomaterials into the cytosol within several minutes, negotiating the endocytososomal system. The potential influence of the technique on cell metabolism is thoroughly studied. By detecting the phosphorylated histone variant H2AX (pH2AX) in the nucleus, it is proved that the operating process will not lead to DNA double-strand breaks. However, the mechanical disruption can temporarily improve the permeability of the cell membranes. Nanoneedle treatment affects cell metabolism at multiple points. The treatment can slightly elevate the apoptotic signal in A549 cells and can significantly increase the production of reactive oxygen species (ROS) in cells, particularly if combined with anticancer drugs. Meanwhile, the activity of cytosolic glucose 6-phosphate dehydrogenase (G6PD) is also raised to counterbalance the elevated ROS content. A detected depolarization of the mitochondrial membrane potential suggests mitochondrial involvement in the intracellular redox reactions and cell apoptosis which are induced by diamond nanoneedle treatment. Overall this study provides a novel understanding on the intracellular delivery mediated by nanoneedles, especially the impact on cell physiology. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Indole-3- carbinol enhances sorafenib cytotoxicity in hepatocellular carcinoma cells: A mechanistic study.

PubMed

Abdelmageed, Mai M; El-Naga, Reem N; El-Demerdash, Ebtehal; Elmazar, Mohamed M

2016-09-09

Sorafenib is the only chemotherapeutic agent currently approved for unresectable hepatocellular carcinoma (HCC). However, poor response rates have been widely reported. Indole-3-carbinol (I3C) is a potential chemopreventive phytochemical. The present study aimed to explore the potential chemomodulatory effects of I3C on sorafenib in HCC cells as well as the possible underlying mechanisms. I3C exhibited a greater cytotoxicity in HepG2 cells compared to Huh-7 cells (p < 0.0001). Moreover, the co-treatment of HepG2 cells with I3C and sorafenib was more effective (p = 0.002). Accordingly, subsequent mechanistic studies were carried on HepG2 cells. The results show that the ability of I3C to enhance sorafenib cytotoxicity in HCC cells could be partially attributed to increasing the apoptotic activity and decreasing the angiogenic potentials. The combination had a negative effect on epithelial-mesenchymal transition (EMT). Increased NOX-1 expression was also observed which may indicate the involvement of NOX-1 in I3C chemomodulatory effects. Additionally, the combination induced cell cycle arrest at the G0/G1 phase. In conclusion, these findings provide evidence that I3C enhances sorafenib anti-cancer activity in HCC cells.

Indole-3- carbinol enhances sorafenib cytotoxicity in hepatocellular carcinoma cells: A mechanistic study

PubMed Central

Abdelmageed, Mai M.; El-Naga, Reem N.; El-Demerdash, Ebtehal; Elmazar, Mohamed M.

2016-01-01

Sorafenib is the only chemotherapeutic agent currently approved for unresectable hepatocellular carcinoma (HCC). However, poor response rates have been widely reported. Indole-3-carbinol (I3C) is a potential chemopreventive phytochemical. The present study aimed to explore the potential chemomodulatory effects of I3C on sorafenib in HCC cells as well as the possible underlying mechanisms. I3C exhibited a greater cytotoxicity in HepG2 cells compared to Huh-7 cells (p < 0.0001). Moreover, the co-treatment of HepG2 cells with I3C and sorafenib was more effective (p = 0.002). Accordingly, subsequent mechanistic studies were carried on HepG2 cells. The results show that the ability of I3C to enhance sorafenib cytotoxicity in HCC cells could be partially attributed to increasing the apoptotic activity and decreasing the angiogenic potentials. The combination had a negative effect on epithelial-mesenchymal transition (EMT). Increased NOX-1 expression was also observed which may indicate the involvement of NOX-1 in I3C chemomodulatory effects. Additionally, the combination induced cell cycle arrest at the G0/G1 phase. In conclusion, these findings provide evidence that I3C enhances sorafenib anti-cancer activity in HCC cells. PMID:27612096

Exosomes: vehicles for the transfer of toxic proteins associated with neurodegenerative diseases?

PubMed

Bellingham, Shayne A; Guo, Belinda B; Coleman, Bradley M; Hill, Andrew F

2012-01-01

Exosomes are small membranous vesicles secreted by a number of cell types including neurons and can be isolated from conditioned cell media or bodily fluids such as urine and plasma. Exosome biogenesis involves the inward budding of endosomes to form multivesicular bodies (MVB). When fused with the plasma membrane, the MVB releases the vesicles into the extracellular environment as exosomes. Proposed functions of these vesicles include roles in cell-cell signaling, removal of unwanted proteins, and the transfer of pathogens between cells. One such pathogen which exploits this pathway is the prion, the infectious particle responsible for the transmissible neurodegenerative diseases such as Creutzfeldt-Jakob disease (CJD) of humans or bovine spongiform encephalopathy (BSE) of cattle. Similarly, exosomes are also involved in the processing of the amyloid precursor protein (APP) which is associated with Alzheimer's disease. Exosomes have been shown to contain full-length APP and several distinct proteolytically cleaved products of APP, including Aβ. In addition, these fragments can be modulated using inhibitors of the proteases involved in APP cleavage. These observations provide further evidence for a novel pathway in which PrP and APP fragments are released from cells. Other proteins such as superoxide dismutase I and alpha-synuclein (involved in amyotrophic lateral sclerosis and Parkinson's disease, respectively) are also found associated with exosomes. This review will focus on the role of exosomes in neurodegenerative disorders and discuss the potential of these vesicles for the spread of neurotoxicity, therapeutics, and diagnostics for these diseases.

Estrogen signalling in the pathogenesis of age-related macular degeneration.

PubMed

Kaarniranta, Kai; Machalińska, Anna; Veréb, Zoltán; Salminen, Antero; Petrovski, Goran; Kauppinen, Anu

2015-02-01

Age-related macular degeneration (AMD) is a multifactorial eye disease that is associated with aging, family history, smoking, obesity, cataract surgery, arteriosclerosis, hypertension, hypercholesterolemia and unhealthy diet. Gender has commonly been classified as a weak or inconsistent risk factor for AMD. This disease is characterized by degeneration of retinal pigment epithelial (RPE) cells, Bruch's membrane, and choriocapillaris, which secondarily lead to damage and death of photoreceptor cells and central visual loss. Pathogenesis of AMD involves constant oxidative stress, chronic inflammation, and increased accumulation of lipofuscin and drusen. Estrogen has both anti-oxidative and anti-inflammatory capacity and it regulates signaling pathways that are involved in the pathogenesis of AMD. In this review, we discuss potential cellular signaling targets of estrogen in retinal cells and AMD pathology.

Micrococcus luteus mediated dual mode synthesis of gold nanoparticles: involvement of extracellular α-amylase and cell wall teichuronic acid.

PubMed

Arunkumar, Pichaimani; Thanalakshmi, Muthukrishnan; Kumar, Priyadarsini; Premkumar, Kumpati

2013-03-01

In the present study we have utilized the bioreductive potential of Micrococcus luteus for the synthesis of gold nanoparticles. Biochemical and physiological analysis indicate that the biosynthesized GNPs were achieved by dual mode, involving extracellular α-amylase and cell wall teichuronic acid (TUA) of M. luteus. The biosynthetic potential of both α-amylase and TUA, after isolation from bacterium, was examined. Under optimum conditions, these biomolecules reduces Au(3+) into Au(0) and the resulting GNPs were found to be stable for 1 month. The synthesized GNPs were characterized by UV-VIS spectrometry, transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FTIR) and dynamic light scattering (DLS). Results demonstrated that the synthesized GNPs were found to be monodispersive and spherical in shape with an average size of ∼6 nm and ∼50 nm for α-amylase and teichuronic acid, respectively. These findings suggest that M. luteus can be exploited as a potential biosource for the eco-friendly synthesis of gold nanoparticles. Copyright © 2012 Elsevier B.V. All rights reserved.

Mastocytosis of the female breast.

PubMed

Setia, Namrata; Crisi, Giovanna M; Pantanowitz, Liron

2010-10-01

A subset of patients with systemic mastocytosis may manifest with extracutaneous involvement. To the best of our knowledge, mastocytosis of the human breast has not been described. This study reports a case with mastocytosis involving the breasts of a 33-year-old woman associated with mammary hypertrophy (breast mastocytosis). The potential for infiltrating mast cells to mimic lobular carcinoma is emphasized and the relationship to breast hypertrophy in this case is discussed.

[Effect of 3-bromopyruvate on mitochondrial membrane potential and apoptosis of human breast carcinoma SK-BR-3 cells].

PubMed

Zhang, Yuanyuan; Liu, Zhe; Zhang, Qianwen; Chao, Zhenhua; Zhang, Pei; Xia, Fei; Jiang, Chenchen; Liu, Hao; Jiang, Zhiwen

2013-09-01

To study the effect of glycolysis inhibitor 3-bromopyruvate (3-BrPA) in inducing apoptosis of human breast carcinoma cells SK-BR-3 and the possible mechanism. MTT assay was used to detect the growth inhibition induced by 3-BrPA in breast cancer cells SK-BR-3. The apoptotic cells were detected by flow cytometry with propidium iodide (PI). ATP levels in the cells were detected by ATP assay kit, and DHE fluorescent probe technique was used to determine superoxide anion levels; the mitochondrial membrane potential was assessed using JC-1 staining assay. MTT assay showed that the proliferation of SK-BR-3 cells was inhibited by 3-BrPA in a time- and concentration-dependent manner. Exposure to 80, 160, and 320 µmol·L(-1) 3-BrPA for 24 h resulted in cell apoptosis rates of 6.7%, 22.3%, and 79.6%, respectively, and the intracellular ATP levels of SK-BR-3 cells treated with 80, 160, 320 µmol·L(-1) 3-BrPA for 5 h were 87.7%, 60.6%, and 23.7% of the control levels. 3-BrPA at 160 µmol·L(-1) increased reactive oxygen levels and lowered mitochondrial membrane potential of SK-BR-3 cells. 3-BrPA can inhibit cell proliferation, reduce the mitochondrial membrane potential and induce apoptosis in SK-BR-3 cells, the mechanism of which may involve a reduced ATP level by inhibiting glycolysis and increasing the reactive oxygen level in the cells.

Angiogenic properties of endometrial mesenchymal stromal cells in endothelial co-culture: an in vitro model of endometriosis.

PubMed

Canosa, S; Moggio, A; Brossa, A; Pittatore, G; Marchino, G L; Leoncini, S; Benedetto, C; Revelli, A; Bussolati, B

2017-03-01

Can endometrial mesenchymal stromal cells (E-MSCs) differentiate into endothelial cells in an in vitro co-culture system with human umbilical vein endothelial cells (HUVECs)? E-MSCs can acquire endothelial markers and function in a direct co-culture system with HUVECs. E-MSCs have been identified in the human endometrium as well as in endometriotic lesions. E-MSCs appear to be involved in formation of the endometrial stromal vascular tissue and the support of tissue growth and vascularization. The use of anti-angiogenic drugs appears as a possible therapeutic strategy against endometriosis. This is an in vitro study comprising patients receiving surgical treatment of ovarian endometriosis (n = 9). E-MSCs were isolated from eutopic and ectopic endometrial tissue and were characterized for the expression of mesenchymal and endothelial markers by FACS analysis and Real-Time PCR. CD31 acquisition was evaluated by FACS analysis and immunofluorescence after a 48 h-direct co-culture with green fluorescent protein +-HUVECs. A tube-forming assay was set up in order to analyze the functional potential of their interaction. Finally, co-cultures were treated with the anti-angiogenic agent Cabergoline. A subpopulation of E-MSCs acquired CD31 expression and integrated into tube-like structures when directly in contact with HUVECs, as observed by both FACS analysis and immunofluorescence. The isolation of CD31+ E-MSCs revealed significant increases in CD31, vascular endothelial growth factor receptor 2, TEK receptor tyrosine kinase and vascular endothelial-Cadherin mRNA expression levels with respect to basal and to CD31neg cells (P < 0.05). On the other hand, the expression of mesenchymal genes such as c-Myc, Vimentin, neuronal-Cadherin and sushi domain containing 2 remained unchanged. Cabergoline treatment induced a significant reduction of the E-MSC angiogenic potential (P < 0.05 versus control). Not applicable. Further studies are necessary to investigate the cellular and molecular mechanisms underlying the endothelial cell differentiation. E-MSCs may undergo endothelial differentiation, and be potentially involved in the development of endometriotic implants. Cell culture systems that more closely mimic the cellular complexity typical of endometriotic tissues in vivo are required to develop novel strategies for treatment. This study was supported by the 'Research Fund ex-60%', University of Turin, Turin, Italy. All authors declare that their participation in the study did not involve actual or potential conflicts of interests. © The Author 2017. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com

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

Laulicht, Freda; Brocato, Jason; Cartularo, Laura

Metals such as arsenic, cadmium, beryllium, and nickel are known human carcinogens; however, other transition metals, such as tungsten (W), remain relatively uninvestigated with regard to their potential carcinogenic activity. Tungsten production for industrial and military applications has almost doubled over the past decade and continues to increase. Here, for the first time, we demonstrate tungsten's ability to induce carcinogenic related endpoints including cell transformation, increased migration, xenograft growth in nude mice, and the activation of multiple cancer-related pathways in transformed clones as determined by RNA sequencing. Human bronchial epithelial cell line (Beas-2B) exposed to tungsten developed carcinogenic properties. Inmore » a soft agar assay, tungsten-treated cells formed more colonies than controls and the tungsten-transformed clones formed tumors in nude mice. RNA-sequencing data revealed that the tungsten-transformed clones altered the expression of many cancer-associated genes when compared to control clones. Genes involved in lung cancer, leukemia, and general cancer genes were deregulated by tungsten. Taken together, our data show the carcinogenic potential of tungsten. Further tests are needed, including in vivo and human studies, in order to validate tungsten as a carcinogen to humans. - Highlights: • Tungsten (W) induces cell transformation and increases migration in vitro. • W increases xenograft growth in nude mice. • W altered the expression of cancer-related genes such as those involved in leukemia. • Some of the dysregulated leukemia genes include, CD74, CTGF, MST4, and HOXB5. • For the first time, data is presented that demonstrates tungsten's carcinogenic potential.« less

Apoptotic effect of novel Schiff based CdCl₂(C₁₄H₂₁N₃O₂) complex is mediated via activation of the mitochondrial pathway in colon cancer cells.

PubMed

Hajrezaie, Maryam; Paydar, Mohammadjavad; Looi, Chung Yeng; Moghadamtousi, Soheil Zorofchian; Hassandarvish, Pouya; Salga, Muhammad Saleh; Karimian, Hamed; Shams, Keivan; Zahedifard, Maryam; Majid, Nazia Abdul; Ali, Hapipah Mohd; Abdulla, Mahmood Ameen

2015-03-13

The development of metal-based agents has had a tremendous role in the present progress in cancer chemotherapy. One well-known example of metal-based agents is Schiff based metal complexes, which hold great promise for cancer therapy. Based on the potential of Schiff based complexes for the induction of apoptosis, this study aimed to examine the cytotoxic and apoptotic activity of a CdCl2(C14H21N3O2) complex on HT-29 cells. The complex exerted a potent suppressive effect on HT-29 cells with an IC50 value of 2.57 ± 0.39 after 72 h of treatment. The collapse of the mitochondrial membrane potential and the elevated release of cytochrome c from the mitochondria to the cytosol indicate the involvement of the intrinsic pathway in the induction of apoptosis. The role of the mitochondria-dependent apoptotic pathway was further proved by the significant activation of the initiator caspase-9 and the executioner caspases-3 and -7. In addition, the activation of caspase-8, which is associated with the suppression of NF-κB translocation to the nucleus, also revealed the involvement of the extrinsic pathway in the induced apoptosis. The results suggest that the CdCl2(C14H21N3O2) complex is able to induce the apoptosis of colon cancer cells and is a potential candidate for future cancer studies.

Predicting gene regulatory networks by combining spatial and temporal gene expression data in Arabidopsis root stem cells

PubMed Central

de Luis Balaguer, Maria Angels; Fisher, Adam P.; Clark, Natalie M.; Fernandez-Espinosa, Maria Guadalupe; Möller, Barbara K.; Weijers, Dolf; Williams, Cranos; Lorenzo, Oscar; Sozzani, Rosangela

2017-01-01

Identifying the transcription factors (TFs) and associated networks involved in stem cell regulation is essential for understanding the initiation and growth of plant tissues and organs. Although many TFs have been shown to have a role in the Arabidopsis root stem cells, a comprehensive view of the transcriptional signature of the stem cells is lacking. In this work, we used spatial and temporal transcriptomic data to predict interactions among the genes involved in stem cell regulation. To accomplish this, we transcriptionally profiled several stem cell populations and developed a gene regulatory network inference algorithm that combines clustering with dynamic Bayesian network inference. We leveraged the topology of our networks to infer potential major regulators. Specifically, through mathematical modeling and experimental validation, we identified PERIANTHIA (PAN) as an important molecular regulator of quiescent center function. The results presented in this work show that our combination of molecular biology, computational biology, and mathematical modeling is an efficient approach to identify candidate factors that function in the stem cells. PMID:28827319

Tissue engineering and regenerative medicine approaches to enhance the functional response to skeletal muscle injury.

PubMed

Sicari, Brian M; Dearth, Christopher L; Badylak, Stephen F

2014-01-01

The well-recognized ability of skeletal muscle for functional and structural regeneration following injury is severely compromised in degenerative diseases and in volumetric muscle loss. Tissue engineering and regenerative medicine strategies to support muscle reconstruction have typically been cell-centric with approaches that involve the exogenous delivery of cells with myogenic potential. These strategies have been limited by poor cell viability and engraftment into host tissue. Alternative approaches have involved the use of biomaterial scaffolds as substrates or delivery vehicles for exogenous myogenic progenitor cells. Acellular biomaterial scaffolds composed of mammalian extracellular matrix (ECM) have also been used as an inductive niche to promote the recruitment and differentiation of endogenous myogenic progenitor cells. An acellular approach, which activates or utilizes endogenous cell sources, obviates the need for exogenous cell administration and provides an advantage for clinical translation. The present review examines the state of tissue engineering and regenerative medicine therapies directed at augmenting the skeletal muscle response to injury and presents the pros and cons of each with respect to clinical translation. Copyright © 2013 Wiley Periodicals, Inc.

Involvement of Receptor Activator of Nuclear Factor-κB Ligand (RANKL)-induced Incomplete Cytokinesis in the Polyploidization of Osteoclasts.

PubMed

Takegahara, Noriko; Kim, Hyunsoo; Mizuno, Hiroki; Sakaue-Sawano, Asako; Miyawaki, Atsushi; Tomura, Michio; Kanagawa, Osami; Ishii, Masaru; Choi, Yongwon

2016-02-12

Osteoclasts are specialized polyploid cells that resorb bone. Upon stimulation with receptor activator of nuclear factor-κB ligand (RANKL), myeloid precursors commit to becoming polyploid, largely via cell fusion. Polyploidization of osteoclasts is necessary for their bone-resorbing activity, but the mechanisms by which polyploidization is controlled remain to be determined. Here, we demonstrated that in addition to cell fusion, incomplete cytokinesis also plays a role in osteoclast polyploidization. In in vitro cultured osteoclasts derived from mice expressing the fluorescent ubiquitin-based cell cycle indicator (Fucci), RANKL induced polyploidy by incomplete cytokinesis as well as cell fusion. Polyploid cells generated by incomplete cytokinesis had the potential to subsequently undergo cell fusion. Nuclear polyploidy was also observed in osteoclasts in vivo, suggesting the involvement of incomplete cytokinesis in physiological polyploidization. Furthermore, RANKL-induced incomplete cytokinesis was reduced by inhibition of Akt, resulting in impaired multinucleated osteoclast formation. Taken together, these results reveal that RANKL-induced incomplete cytokinesis contributes to polyploidization of osteoclasts via Akt activation. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Involvement of Receptor Activator of Nuclear Factor-κB Ligand (RANKL)-induced Incomplete Cytokinesis in the Polyploidization of Osteoclasts*

PubMed Central

Takegahara, Noriko; Kim, Hyunsoo; Mizuno, Hiroki; Sakaue-Sawano, Asako; Miyawaki, Atsushi; Tomura, Michio; Kanagawa, Osami; Ishii, Masaru; Choi, Yongwon

2016-01-01

Osteoclasts are specialized polyploid cells that resorb bone. Upon stimulation with receptor activator of nuclear factor-κB ligand (RANKL), myeloid precursors commit to becoming polyploid, largely via cell fusion. Polyploidization of osteoclasts is necessary for their bone-resorbing activity, but the mechanisms by which polyploidization is controlled remain to be determined. Here, we demonstrated that in addition to cell fusion, incomplete cytokinesis also plays a role in osteoclast polyploidization. In in vitro cultured osteoclasts derived from mice expressing the fluorescent ubiquitin-based cell cycle indicator (Fucci), RANKL induced polyploidy by incomplete cytokinesis as well as cell fusion. Polyploid cells generated by incomplete cytokinesis had the potential to subsequently undergo cell fusion. Nuclear polyploidy was also observed in osteoclasts in vivo, suggesting the involvement of incomplete cytokinesis in physiological polyploidization. Furthermore, RANKL-induced incomplete cytokinesis was reduced by inhibition of Akt, resulting in impaired multinucleated osteoclast formation. Taken together, these results reveal that RANKL-induced incomplete cytokinesis contributes to polyploidization of osteoclasts via Akt activation. PMID:26670608

MicroRNA profiling of antler stem cells in potentiated and dormant states and their potential roles in antler regeneration.

PubMed

Ba, Hengxing; Wang, Datao; Li, Chunyi

2016-04-01

MicroRNAs (miRNAs) can effectively regulate gene expression at the post-transcriptional level and play a critical role in tissue growth, development and regeneration. Our previous studies showed that antler regeneration is a stem cell-based process and antler stem cells reside in the periosteum of a pedicle, the permanent bony protuberance, from which antler regeneration takes place. Antlers are the only mammalian organ that can fully regenerate and hence provide a unique opportunity to identify miRNAs that are involved in organ regeneration. In the present study, we used next generation sequencing technology sequenced miRNAs of the stem cells derived from either the potentiated or the dormant pedicle periosteum. A population of both conserved and 20 deer-specific miRNAs was identified. These conserved miRNAs were derived from 453 homologous hairpin precursors across 88 animal species, and were further grouped into 167 miRNA families. Among them, the miR-296 is embryonic stem cell-specific. The potentiation process resulted in the significant regulation (>±2 Fold, q value <0.05) of conserved miRNAs; 8 miRNA transcripts were down- and 6 up-regulated. Several GO biology processes and the Wnt, MAPK and TGF-beta signaling pathways were found to be up-regulated as part of antlerogenic stem cell potentiation process. This research has identified miRNAs that are associated either with the dormant or the potentiated antler stem cells and identified some target miRNAs for further research into their role played in mammalian organ regeneration.

Lectin array and glycogene expression analyses of ovarian cancer cell line A2780 and its cisplatin-resistant derivate cell line A2780-cp.

PubMed

Zhao, Ran; Qin, Wenjun; Qin, Ruihuan; Han, Jing; Li, Can; Wang, Yisheng; Xu, Congjian

2017-01-01

Ovarian cancer is one of the most lethal gynecological malignancies, in which platinum resistance is a common cause of its relapse and death. Glycosylation has been reported to be involved in drug resistance, and glycomic analyses of ovarian cancer may improve our understanding of the mechanisms underlying cancer cell drug resistance and provide potential biomarkers and therapeutic targets. The serous ovarian cancer cell line A2780 and its platinum-resistant counterpart A2780-cp were used in this study. We performed a lectin array analysis to compare the glycosylation patterns of the two cell lines, a gene expression array was employed to probe the differences in glycogenes. Furthermore, the results were verified by lectin blots. A2780-cp cell exhibited stronger intensities of Lens culinaris (LCA) Canavalia ensiformis (ConA), and Lycopersicon esculentum (LEL) and weaker intensities of Sambucus nigra (SNA) lectins. The gene expression array analysis revealed increased expression of Fut8, B3gnt4, B3gnt5, B4galt2 and decreased expression of Fut1 and ST6GalNAc 6 expression were evident in the A2780-cp cells. The lectin blot confirmed the differences in LCA, ConA, SNA and LEL between the A2780 and A2780-cp cells. The combination of the lectin and gene expression analyses showed that the levels of core fucosylation and poly-LacNAc were increased in the A2780-cp cells and the levels of Fuc α1-2(gal β1-4) GlcNAc and α2-6-linked sialic structures were decreased in the A2780-cp cells. These glycans represent potential biomarkers and might be involved in the mechanism of drug resistance in ovarian cancer.

The Voltage-dependent Anion Channel 1 Mediates Amyloid β Toxicity and Represents a Potential Target for Alzheimer Disease Therapy.

PubMed

Smilansky, Angela; Dangoor, Liron; Nakdimon, Itay; Ben-Hail, Danya; Mizrachi, Dario; Shoshan-Barmatz, Varda

2015-12-25

The voltage-dependent anion channel 1 (VDAC1), found in the mitochondrial outer membrane, forms the main interface between mitochondrial and cellular metabolisms, mediates the passage of a variety of molecules across the mitochondrial outer membrane, and is central to mitochondria-mediated apoptosis. VDAC1 is overexpressed in post-mortem brains of Alzheimer disease (AD) patients. The development and progress of AD are associated with mitochondrial dysfunction resulting from the cytotoxic effects of accumulated amyloid β (Aβ). In this study we demonstrate the involvement of VDAC1 and a VDAC1 N-terminal peptide (VDAC1-N-Ter) in Aβ cell penetration and cell death induction. Aβ directly interacted with VDAC1 and VDAC1-N-Ter, as monitored by VDAC1 channel conductance, surface plasmon resonance, and microscale thermophoresis. Preincubated Aβ interacted with bilayer-reconstituted VDAC1 and increased its conductance ∼ 2-fold. Incubation of cells with Aβ resulted in mitochondria-mediated apoptotic cell death. However, the presence of non-cell-penetrating VDAC1-N-Ter peptide prevented Aβ cellular entry and Aβ-induced mitochondria-mediated apoptosis. Likewise, silencing VDAC1 expression by specific siRNA prevented Aβ entry into the cytosol as well as Aβ-induced toxicity. Finally, the mode of Aβ-mediated action involves detachment of mitochondria-bound hexokinase, induction of VDAC1 oligomerization, and cytochrome c release, a sequence of events leading to apoptosis. As such, we suggest that Aβ-mediated toxicity involves mitochondrial and plasma membrane VDAC1, leading to mitochondrial dysfunction and apoptosis induction. The VDAC1-N-Ter peptide targeting Aβ cytotoxicity is thus a potential new therapeutic strategy for AD treatment. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

PHTS, a novel putative tumor suppressor, is involved in the transformation reversion of HeLaHF cells independently of the p53 pathway

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

Yu Dehua; Fan, Wufang; Liu, Guohong

2006-04-01

HeLaHF is a non-transformed revertant of HeLa cells, likely resulting from the activation of a putative tumor suppressor(s). p53 protein was stabilized in this revertant and reactivated for certain transactivation functions. Although p53 stabilization has not conclusively been linked to the reversion, it is clear that the genes in p53 pathway are involved. The present study confirms the direct role of p53 in HeLaHF reversion by demonstrating that RNAi-mediated p53 silencing partially restores anchorage-independent growth potential of the revertant through the suppression of anoikis. In addition, we identified a novel gene, named PHTS, with putative tumor suppressor properties, and showedmore » that this gene is also involved in HeLaHF reversion independently of the p53 pathway. Expression profiling revealed that PHTS is one of the genes that is up-regulated in HeLaHF but not in HeLa. It encodes a putative protein with CD59-like domains. RNAi-mediated PHTS silencing resulted in the partial restoration of transformation (anchorage-independent growth) in HeLaHF cells, similar to that of p53 gene silencing, implying its tumor suppressor effect. However, the observed increased transformation potential by PHTS silencing appears to be due to an increased anchorage-independent proliferation rate rather than suppression of anoikis, unlike the effect of p53 silencing. p53 silencing did not affect PHTS gene expression, and vice versa, suggesting PHTS may function in a new and p53-independent tumor suppressor pathway. Furthermore, over-expression of PHTS in different cancer cell lines, in addition to HeLa, reduces cell growth likely via induced apoptosis, confirming the broad PHTS tumor suppressor properties.« less

Sevoflurane post-conditioning protects primary rat cortical neurons against oxygen-glucose deprivation/resuscitation via down-regulation in mitochondrial apoptosis axis of Bid, Bim, Puma-Bax and Bak mediated by Erk1/2.

PubMed

Zhang, Li-Min; Zhao, Xiao-Chun; Sun, Wen-Bo; Li, Rui; Jiang, Xiao-Jing

2015-10-15

Temporal post-conditioning helps provide neuroprotection against brain injury secondary to ischemia-reperfusion and is considered an effective intervention, but the exact mechanism of sevoflurane post-conditioning is unclear. The essential axis involves activator Bid, Bim, Puma (BH3s), Bax, and Bak; activates the mitochondrial death program; and might be involved in a cell death signal. Extracellular signal-related kinases 1/2 (Erk1/2) play a pivotal role in cell growth and proliferation. We hypothesized that sevoflurane post-conditioning might inhibit Bid, Bim, Puma, Bax, and Bak expression and is activated by phosphor-Erk1/2 to decrease neuronal death. To test this hypothesis, we exposed primary cortical neuron cultures to oxygen-glucose deprivation for 1h, along with resuscitation for 24h (OGD/R). MTT assays, propidium iodide uptake (PI), JC-1 fluorescence, and Western blot indicated the following: decreased cell viability (P<0.05); increased cell death (P<0.05); decreased mitochondrial membrane potential (P<0.05); and decreased Bid, Bim, Puma, Bax, and Bak expression with OGD/R exposure. Inhibition of Erk1/2 phosphorylation could attenuate sevoflurane post-conditioning that mediated an increase in neuronal viability and mitochondrial membrane potential, as well as a decrease in cell death and Bid, Bim, Puma, Bax, and Bak expression after OGD/R treatment. The results demonstrated that sevoflurane post-conditioning caused a marked decrease in cortical neuronal death secondary to OGD/R exposure through the downregulation of the mitochondrial apoptosis axis involving Bid, Bim, Puma, Bax, and Bak that was mediated by the phosphorylation/activation of Erk1/2. Copyright © 2015 Elsevier B.V. All rights reserved.

Involvement of pre- and postsynaptic NMDA receptors at local circuit interneuron connections in rat neocortex

PubMed Central

De-May, C.L.; Ali, A.B.

2013-01-01

To investigate the involvement of N-Methyl-D-aspartate (NMDA) receptors in local neocortical synaptic transmission, dual whole-cell recordings – combined with biocytin labelling – were obtained from bitufted adapting, multipolar adapting or multipolar non-adapting interneurons and pyramidal cells in layers II–V of rat (postnatal days 17–22) sensorimotor cortex. The voltage dependency of the amplitude of Excitatory postsynaptic potentials (EPSPs) received by the three types of interneuron appeared to coincide with the interneuron subclass; upon depolarisation, EPSPs received by multipolar non-adapting interneurons either decreased in amplitude or appeared insensitive, multipolar adapting interneuron EPSP amplitudes increased or appeared insensitive, whereas bitufted interneuron EPSP amplitudes increased or decreased. Connections were challenged with the NMDA receptor antagonist d-(−)-2-amino-5-phosphonopentanoic acid (d-AP5) (50 μM) revealing NMDA receptors to contribute to EPSPs received by all cell types, this also abolished the non-conventional voltage dependency. Reciprocal connections were frequent between pyramidal cells and multipolar interneurons, and inhibitory postsynaptic potentials (IPSPs) elicited in pyramidal cells by both multipolar adapting and multipolar non-adapting interneurons were sensitive to a significant reduction in amplitude by d-AP5. The involvement of presynaptic NMDA receptors was indicated by coefficient of variation analysis and an increase in the failures of transmission. Furthermore, by loading MK-801 into the pre- or postsynaptic neurons, we observed that a reduction in inhibition requires presynaptic and not postsynaptic NMDA receptors. These results suggest that NMDA receptors possess pre- and postsynaptic roles at selective neocortical synapses that are probably important in governing spike-timing and information flow. PMID:23079623

Polyploidization of murine mesenchymal cells is associated with suppression of the long noncoding RNA H19 and reduced tumorigenicity.

PubMed

Shoshani, Ofer; Massalha, Hassan; Shani, Nir; Kagan, Sivan; Ravid, Orly; Madar, Shalom; Trakhtenbrot, Luba; Leshkowitz, Dena; Rechavi, Gideon; Zipori, Dov

2012-12-15

Mesenchymal stromal cells (MSC) are used extensively in clinical trials; however, the possibility that MSCs have a potential for malignant transformation was raised. We examined the genomic stability versus the tumor-forming capacity of multiple mouse MSCs. Murine MSCs have been shown to be less stable and more prone to malignant transformation than their human counterparts. A large series of independently isolated MSC populations exhibited low tumorigenic potential under syngeneic conditions, which increased in immunocompromised animals. Unexpectedly, higher ploidy correlated with reduced tumor-forming capacity. Furthermore, in both cultured MSCs and primary hepatocytes, polyploidization was associated with a dramatic decrease in the expression of the long noncoding RNA H19. Direct knockdown of H19 expression in diploid cells resulted in acquisition of polyploid cell traits. Moreover, artificial tetraploidization of diploid cancer cells led to a reduction of H19 levels, as well as to an attenuation of the tumorigenic potential. Polyploidy might therefore serve as a protective mechanism aimed at reducing malignant transformation through the involvement of the H19 regulatory long noncoding RNA.

Characterization and identification of a chymotryptic hydrolysate of alpha-lactalbumin stimulating cholecystokinin release in STC-1 cells.

PubMed

Catiau, Lucie; Delval-Dubois, Véronique; Guillochon, Didier; Nedjar-Arroume, Naïma

2011-11-01

Alpha-lactalbumin hydrolysate is of significant interest, due to its potential application as a source of bioactive peptides in nutraceutical and pharmaceutical domains. This study was focused on the cholecystokinin (CCK) family compounds which are small peptides involved in the satiety control. The action of chymotryptic hydrolysate of alpha-lactalbumin on cholecystokinin release from intestinal endocrine STC-1 cells was investigated. We demonstrated for the first time that a chymotryptic hydrolysate of alpha-lactalbumin was able to highly stimulate CCK-releasing activity from STC-1 cells. The peptidic hydrolysate was characterized by LC/MS and MS/MS, thus highlighting the presence of 11 fractions containing 21 peptides, each potentially having the desired activity.

Translocation of cell-penetrating peptides into Candida fungal pathogens.

PubMed

Gong, Zifan; Karlsson, Amy J

2017-09-01

Cell-penetrating peptides (CPPs) are small peptides capable of crossing cellular membranes while carrying molecular cargo. Although they have been widely studied for their ability to translocate nucleic acids, small molecules, and proteins into mammalian cells, studies of their interaction with fungal cells are limited. In this work, we evaluated the translocation of eleven fluorescently labeled peptides into the important human fungal pathogens Candida albicans and C. glabrata and explored the mechanisms of translocation. Seven of these peptides (cecropin B, penetratin, pVEC, MAP, SynB, (KFF) 3 K, and MPG) exhibited substantial translocation (>80% of cells) into both species in a concentration-dependent manner, and an additional peptide (TP-10) exhibiting strong translocation into only C. glabrata. Vacuoles were involved in translocation and intracellular trafficking of the peptides in the fungal cells and, for some peptides, escape from the vacuoles and localization in the cytosol were correlated to toxicity toward the fungal cells. Endocytosis was involved in the translocation of cecropin B, MAP, SynB, MPG, (KFF) 3 K, and TP-10, and cecropin B, penetratin, pVEC, and MAP caused membrane permeabilization during translocation. These results indicate the involvement of multiple translocation mechanisms for some CPPs. Although high levels of translocation were typically associated with toxicity of the peptides toward the fungal cells, SynB was translocated efficiently into Candida cells at concentrations that led to minimal toxicity. Our work highlights the potential of CPPs in delivering antifungal molecules and other bioactive cargo to Candida pathogens. © 2017 The Protein Society.

mRNA and methylation profiling of radioresistant esophageal cancer cells: the involvement of Sall2 in acquired aggressive phenotypes

PubMed Central

Luo, Judong; Wang, Wenjie; Tang, Yiting; Zhou, Dandan; Gao, Yi; Zhang, Qi; Zhou, Xifa; Zhu, Hui; Xing, Ligang; Yu, Jinming

2017-01-01

Esophageal squamous cell carcinoma (ESCC) is one of the deadliest malignancies worldwide. Radiotherapy plays a critical role in the curative management of inoperable ESCC patients. However, radioresistance restricts the efficacy of radiotherapy for ESCC patients. The molecules involved in radioresistance remain largely unknown, and new approaches to sensitize cells to irradiation are in demand. Technical advances in analysis of mRNA and methylation have enabled the exploration of the etiology of diseases and have the potential to broaden our understanding of the molecular pathways of ESCC radioresistance. In this study, we constructed radioresistant TE-1 and Eca-109 cell lines (TE-1/R and Eca-109/R, respectively). The radioresistant cells showed an increased migration ability but reduced apoptosis and cisplatin sensitivity compared with their parent cells. mRNA and methylation profiling by microarray revealed 1192 preferentially expressed mRNAs and 8841 aberrantly methylated regions between TE-1/R and TE-1 cells. By integrating the mRNA and methylation profiles, we related the decreased expression of transcription factor Sall2 with a corresponding increase in its methylation in TE-1/R cells, indicating its involvement in radioresistance. Upregulation of Sall2 decreased the growth and migration advantage of radioresistant ESCC cells. Taken together, our present findings illustrate the mRNA and DNA methylation changes during the radioresistance of ESCC and the important role of Sall2 in esophageal cancer malignancy. PMID:28367244

Benzene and its metabolite decreases cell proliferation via LncRNA-OBFC2A-mediated anti-proliferation effect involving NOTCH1 and KLF15

PubMed Central

Sun, Pengling; Wang, Jing; Guo, Xiaoli; Chen, Yujiao; Xing, Caihong; Gao, Ai

2017-01-01

LncRNA has been considered to play a crucial role in the progression of several diseases by affecting cell proliferation. However, its role in benzene toxicity remains unclear. Our study showed that the expression of lncRNA-OBFC2A increased accompanied with the change of cell proliferation related-genes in benzene-exposed workers. In vitro experiments, 1,4-Benzoquinone dose-dependently inhibited cell proliferation and simultaneously caused the decrease of NOTCH1 expression and the increase of KLF15 in AHH-1 cell lines. Meanwhile, 1, 4-Benzoquinone obviously increased the expression of lncRNA-OBFC2A, which was consistent with our previous population results. Therefore, we propose that lncRNA-OBFC2A is involved in benzene toxicity by regulating cell proliferation. Further, we successfully constructed a lentivirus model of interfering the expression of lncRNA-OBFC2A. After interfering lncRNA-OBFC2A, the cell proliferation inhibition and the expression of NOTCH1 and KLF15 induced by 1, 4-Benzoquinone were reversed. Subsequently, RNA fluorescence in situ Hybridization assay showed that lncRNA-OBFC2A was located in cell nuclei. These results suggest that benzene and its metabolite decreases cell proliferation via LncRNA-OBFC2A-mediated anti-proliferation effect involving NOTCH1 and KLF15. LncRNA-OBFC2A can be a potential biomarker for benzene toxicity. PMID:28388563

Benzene and its metabolite decreases cell proliferation via LncRNA-OBFC2A-mediated anti-proliferation effect involving NOTCH1 and KLF15.

PubMed

Sun, Pengling; Wang, Jing; Guo, Xiaoli; Chen, Yujiao; Xing, Caihong; Gao, Ai

2017-06-20

LncRNA has been considered to play a crucial role in the progression of several diseases by affecting cell proliferation. However, its role in benzene toxicity remains unclear. Our study showed that the expression of lncRNA-OBFC2A increased accompanied with the change of cell proliferation related-genes in benzene-exposed workers. In vitro experiments, 1,4-Benzoquinone dose-dependently inhibited cell proliferation and simultaneously caused the decrease of NOTCH1 expression and the increase of KLF15 in AHH-1 cell lines. Meanwhile, 1, 4-Benzoquinone obviously increased the expression of lncRNA-OBFC2A, which was consistent with our previous population results. Therefore, we propose that lncRNA-OBFC2A is involved in benzene toxicity by regulating cell proliferation. Further, we successfully constructed a lentivirus model of interfering the expression of lncRNA-OBFC2A. After interfering lncRNA-OBFC2A, the cell proliferation inhibition and the expression of NOTCH1 and KLF15 induced by 1, 4-Benzoquinone were reversed. Subsequently, RNA fluorescence in situ Hybridization assay showed that lncRNA-OBFC2A was located in cell nuclei. These results suggest that benzene and its metabolite decreases cell proliferation via LncRNA-OBFC2A-mediated anti-proliferation effect involving NOTCH1 and KLF15. LncRNA-OBFC2A can be a potential biomarker for benzene toxicity.

Function-blocking antibodies to human vascular adhesion protein-1: a potential anti-inflammatory therapy.

PubMed

Kirton, Christopher M; Laukkanen, Marja-Leena; Nieminen, Antti; Merinen, Marika; Stolen, Craig M; Armour, Kathryn; Smith, David J; Salmi, Marko; Jalkanen, Sirpa; Clark, Michael R

2005-11-01

Human vascular adhesion protein-1 (VAP-1) is a homodimeric 170-kDa sialoglycoprotein that is expressed on the surface of endothelial cells and functions as a semicarbazide-sensitive amine oxidase and as an adhesion molecule. Blockade of VAP-1 has been shown to reduce leukocyte adhesion and transmigration in in vivo and in vitro models, suggesting that VAP-1 is a potential target for anti-inflammatory therapy. In this study we have constructed mouse-human chimeric antibodies by genetic engineering in order to circumvent the potential problems involved in using murine antibodies in man. Our chimeric anti-VAP-1 antibodies, which were designed to lack Fc-dependent effector functions, bound specifically to cell surface-expressed recombinant human VAP-1 and recognized VAP-1 in different cell types in tonsil. Furthermore, the chimeric antibodies prevented leukocyte adhesion and transmigration in vitro and in vivo. Hence, these chimeric antibodies have the potential to be used as a new anti-inflammatory therapy.

Dysregulated MicroRNA Involvement in Multiple Sclerosis by Induction of T Helper 17 Cell Differentiation

PubMed Central

Chen, Chen; Zhou, Yifan; Wang, Jingqi; Yan, Yaping; Peng, Lisheng; Qiu, Wei

2018-01-01

Multiple sclerosis (MS) is an immune-mediated demyelinating disease of the central nervous system. Growing evidence has proven that T helper 17 (Th17) cells are one of the regulators of neuroinflammation mechanisms in MS disease. Researchers have demonstrated that some microRNAs (miRNAs) are associated with disease activity and duration, even with different MS patterns. miRNAs regulate CD4+ T cells to differentiate toward various T cell subtypes including Th17 cells. In this review, we discuss the possible mechanisms of miRNAs in MS pathophysiology by regulating CD4+ T cell differentiation into Th17 cells, and potential miRNA targets for current disease-modifying treatments.

DOSE ASSESSMENTS FROM THE DISPOSAL OF LOW ...

EPA Pesticide Factsheets

Modeling the long-term performance of the RCRA-C disposal cell and potential doses to off-site receptors is used to derive maximum radionuclide specific concentrations in the wastes that would enable these wastes to be disposed of safely using the RCRA-C disposal cell technology. Modeling potential exposures to derive these waste acceptance concentrations involves modeling exposures to workers during storage, treatment and disposal of the wastes, as well as exposures to individuals after disposal operations have ceased. Post facility closure exposures can result from the slow expected degradation of the disposal cell over long time periods (one thousand years after disposal) and in advertent human intrusion. Provide a means of determining waste acceptance radionuclide concentrations for disposal of debris from radiological dispersal device incidents as well as low-activity wastes generated in commercial, medical and research activities, potentially serve as the technical basis for guidance on disposal of these materials.

Cell wall glycans and soluble factors determine the interactions between the hyphae of Candida albicans and Pseudomonas aeruginosa.

PubMed

Brand, Alexandra; Barnes, Julia D; Mackenzie, Kevin S; Odds, Frank C; Gow, Neil A R

2008-10-01

The fungus, Candida albicans, and the bacterium, Pseudomonas aeruginosa, are opportunistic human pathogens that have been coisolated from diverse body sites. Pseudomonas aeruginosa suppresses C. albicans proliferation in vitro and potentially in vivo but it is the C. albicans hyphae that are killed while yeast cells are not. We show that hyphal killing involves both contact-mediated and soluble factors. Bacterial culture filtrates contained heat-labile soluble factors that killed C. albicans hyphae. In cocultures, localized points of hyphal lysis were observed, suggesting that adhesion and subsequent bacteria-mediated cell wall lysis is involved in the killing of C. albicans hyphae. The glycosylation status of the C. albicans cell wall affected the rate of contact-dependent killing because mutants with severely truncated O-linked, but not N-linked, glycans were hypersensitive to Pseudomonas-mediated killing. Deletion of HWP1, ALS3 or HYR1, which encode major hypha-associated cell wall proteins, had no effect on fungal susceptibility.

Deciphering early events involved in hyperosmotic stress-induced programmed cell death in tobacco BY-2 cells.

PubMed

Monetti, Emanuela; Kadono, Takashi; Tran, Daniel; Azzarello, Elisa; Arbelet-Bonnin, Delphine; Biligui, Bernadette; Briand, Joël; Kawano, Tomonori; Mancuso, Stefano; Bouteau, François

2014-03-01

Hyperosmotic stresses represent one of the major constraints that adversely affect plants growth, development, and productivity. In this study, the focus was on early responses to hyperosmotic stress- (NaCl and sorbitol) induced reactive oxygen species (ROS) generation, cytosolic Ca(2+) concentration ([Ca(2+)]cyt) increase, ion fluxes, and mitochondrial potential variations, and on their links in pathways leading to programmed cell death (PCD). By using BY-2 tobacco cells, it was shown that both NaCl- and sorbitol-induced PCD seemed to be dependent on superoxide anion (O2·(-)) generation by NADPH-oxidase. In the case of NaCl, an early influx of sodium through non-selective cation channels participates in the development of PCD through mitochondrial dysfunction and NADPH-oxidase-dependent O2·(-) generation. This supports the hypothesis of different pathways in NaCl- and sorbitol-induced cell death. Surprisingly, other shared early responses, such as [Ca(2+)]cyt increase and singlet oxygen production, do not seem to be involved in PCD.

Involvement of regulatory T cells in the immunosuppression characteristic of patients with paracoccidioidomycosis.

PubMed

Ferreira, Maria Carolina; de Oliveira, Rômulo Tadeu Dias; da Silva, Rosiane Maria; Blotta, Maria Heloisa Souza Lima; Mamoni, Ronei Luciano

2010-10-01

Patients with paracoccidioidomycosis (PCM) exhibit a suppression of the cellular immune response characterized by negative delayed-type hypersensitivity (DTH) to Paracoccidioides brasiliensis antigens, the apoptosis of lymphocytes, and high levels of expression of cytotoxic-T-lymphocyte-associated antigen 4 (CTLA-4), interleukin-10 (IL-10), and transforming growth factor β (TGF-β). The aim of this study was to investigate whether and how regulatory T cells (Treg cells) are involved in this immunosuppression by analyzing the number, phenotype, and activity of these cells in patients with active disease (AD group) and patients who had received treatment (TD group). Our results showed that the AD patients had more Treg cells than the TD patients or controls (C group) and also had elevated levels of expression of regulatory markers (glucocorticoid-induced tumor necrosis factor [TNF] receptor-related protein [GITR], CTLA-4, CD95L, LAP-1, and CD38). An analysis of regulatory activity showed that Treg cells from the AD group had greater activity than did cells from the other groups and that cell-cell contact is mandatory for this activity in the C group but was only partially involved in the regulatory activity of cells from AD patients. The addition of anti-IL-10 and anti-TGF-β neutralizing antibodies to the cultures showed that the production of cytokines may be another mechanism used by Treg cells. In conclusion, the elevated numbers of these cells with an increased regulatory phenotype and strong suppressive activity suggest a potential role for them in the immunosuppression characteristic of paracoccidioidomycosis. In addition, our results indicate that while Treg cells act by cell-cell contact, cytokine production also plays an important role.

Regenerative Endodontics in light of the stem cell paradigm

PubMed Central

Rosa, Vinicius; Botero, Tatiana M.; Nör, Jacques E.

2013-01-01

Stem cells play a critical role in development and in tissue regeneration. The dental pulp contains a small sub-population of stem cells that are involved in the response of the pulp to caries progression. Specifically, stem cells replace odontoblasts that have undergone cell death as a consequence of the cariogenic challenge. Stem cells also secrete factors that have the potential to enhance pulp vascularization and provide the oxygen and nutrients required for the dentinogenic response that is typically observed in teeth with deep caries. However, the same angiogenic factors that are required for dentin regeneration may ultimately contribute to the demise of the pulp by enhancing vascular permeability and interstitial pressure. Recent studies focused on the biology of dental pulp stem cells revealed that the multipotency and angiogenic capacity of these cells could be exploited therapeutically in dental pulp tissue engineering. Collectively, these findings suggest new treatment paradigms in the field of Endodontics. The goal of this review is to discuss the potential impact of dental pulp stem cells to Regenerative Endodontics. PMID:21726222

Experimental Proof for the Role of Nonlinear Photoionization in Plasmonic Phototherapy.

PubMed

Minai, Limor; Zeidan, Adel; Yeheskely-Hayon, Daniella; Yudovich, Shimon; Kviatkovsky, Inna; Yelin, Dvir

2016-07-13

Targeting individual cells within a heterogeneous tissue is a key challenge in cancer therapy, encouraging new approaches for cancer treatment that complement the shortcomings of conventional therapies. The highly localized interactions triggered by focused laser beams promise great potential for targeting single cells or small cell clusters; however, most laser-tissue interactions often involve macroscopic processes that may harm healthy nearby tissue and reduce specificity. Specific targeting of living cells using femtosecond pulses and nanoparticles has been demonstrated promising for various potential therapeutic applications including drug delivery via optoporation, drug release, and selective cell death. Here, using an intense resonant femtosecond pulse and cell-specific gold nanorods, we show that at certain irradiation parameters cell death is triggered by nonlinear plasmonic photoionization and not by thermally driven processes. The experimental results are supported by a physical model for the pulse-particle-medium interactions. A good correlation is found between the calculated total number and energy of the generated free electrons and the observed cell death, suggesting that femtosecond photoionization plays the dominant role in cell death.

Alternative sources of pluripotency: science, ethics, and stem cells.

PubMed

Kastenberg, Zachary J; Odorico, Jon S

2008-07-01

Despite many advances in human embryonic stem cell (hESC) technology the ethical dilemma involving the destruction of a human embryo is one factor that has limited the development of hESC based clinical therapies. Two recent reports describing the production of pluripotent stem cells following the in vitro reprogramming of human somatic cells with certain defined factors illustrate one potential method of bypassing the ethical debate surrounding hESCs (Yu J, Vodyanik MA, Smuga-Otto K, et al. Induced pluripotent stem cell lines derived from human somatic cells. Science. 2007 Dec;318(5858):1917-1920; Takahashi K, Tanabe K, Ohnuki M, et al. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell. 2007 Nov;131(5): 861-872.). Other alternative methods include nuclear transfer, altered nuclear transfer, and parthenogenesis; each with its own set of advantages and disadvantages. This review discusses recent advances in these technologies with specific focus on the issues of embryo destruction, oocyte recovery, and the potential of each technology to produce large scale, patient specific cell transplantation therapies that would require little or no immunosuppression.

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

Orfali, Nina; Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA.; McKenna, Sharon L.

Retinoids are a family of signaling molecules derived from vitamin A with well established roles in cellular differentiation. Physiologically active retinoids mediate transcriptional effects on cells through interactions with retinoic acid (RARs) and retinoid-X (RXR) receptors. Chromosomal translocations involving the RARα gene, which lead to impaired retinoid signaling, are implicated in acute promyelocytic leukemia (APL). All-trans-retinoic acid (ATRA), alone and in combination with arsenic trioxide (ATO), restores differentiation in APL cells and promotes degradation of the abnormal oncogenic fusion protein through several proteolytic mechanisms. RARα fusion-protein elimination is emerging as critical to obtaining sustained remission and long-term cure in APL.more » Autophagy is a degradative cellular pathway involved in protein turnover. Both ATRA and ATO also induce autophagy in APL cells. Enhancing autophagy may therefore be of therapeutic benefit in resistant APL and could broaden the application of differentiation therapy to other cancers. Here we discuss retinoid signaling in hematopoiesis, leukemogenesis, and APL treatment. We highlight autophagy as a potential important regulator in anti-leukemic strategies. - Highlights: • Normal and aberrant retinoid signaling in hematopoiesis and leukemia is reviewed. • We suggest a novel role for RARα in the development of X-RARα gene fusions in APL. • ATRA therapy in APL activates transcription and promotes onco-protein degradation. • Autophagy may be involved in both onco-protein degradation and differentiation. • Pharmacologic autophagy induction may potentiate ATRA's therapeutic effects.« less

Endothelial effects of emission source particles: acute toxic response gene expression profiles.

PubMed

Nadadur, Srikanth S; Haykal-Coates, Najwa; Mudipalli, Anuradha; Costa, Daniel L

2009-02-01

Air pollution epidemiology has established a strong association between exposure to ambient particulate matter (PM) and cardiovascular outcomes. Experimental studies in both humans and laboratory animals support varied biological mechanisms including endothelial dysfunction as potentially a central step to the elicitation of cardiovascular events. We therefore hypothesized that relevant early molecular alterations on endothelial cells should be assessable in vitro upon acute exposure to PM components previously shown to be involved in health outcomes. Using a model emission PM, residual oil fly ash and one of its predominant constituents (vanadium-V), we focused on the development of gene expression profiles to fingerprint that particle and its constituents to explore potential biomarkers for PM-induced endothelial dysfunction. Here we present differential gene expression and transcription factor activation profiles in human vascular endothelial cells exposed to a non-cytotoxic dose of fly ash or V following semi-global gene expression profiling of approximately 8000 genes. Both fly ash and it's prime constituent, V, induced alterations in genes involved in passive and active transport of solutes across the membrane; voltage-dependent ion pumps; induction of extracellular matrix proteins and adhesion molecules; and activation of numerous kinases involved in signal transduction pathways. These preliminary data suggest that cardiovascular effects associated with exposure to PM may be mediated by perturbations in endothelial cell permeability, membrane integrity; and ultimately endothelial dysfunction.

Numbers, Neurons and Tides, Oh My!

ERIC Educational Resources Information Center

Ortiz, Mary Theresa

2006-01-01

Mathematical applications to biology are presented in Anatomy & Physiology, General and Marine Biology. Body measurements and anatomical terminology are integrated, and problems involving neuron conduction speed, red blood cells, hemoglobin and glomerular filtration presented. General Biology applications include trans-membrane potential and…

Of mice and men: how animal models advance our understanding of T-cell function in RA.

PubMed

Kobezda, Tamás; Ghassemi-Nejad, Sheida; Mikecz, Katalin; Glant, Tibor T; Szekanecz, Zoltán

2014-03-01

The involvement of autoreactive T cells in the pathogenesis of rheumatoid arthritis (RA) as well as in autoimmune animal models of arthritis has been well established; however, unanswered questions, such as the role of joint-homing T cells, remain. Animal models of arthritis are superb experimental tools in demonstrating how T cells trigger joint inflammation, and thus can help to further our knowledge of disease mechanisms and potential therapies. In this Review, we discuss the similarities and differences in T-cell subsets and functions between RA and mouse arthritis models. For example, various T-cell subsets are involved in both human and mouse arthritis, but differences might exist in the cytokine regulation and plasticity of these cells. With regard to joint-homing T cells, an abundance of synovial T cells is present in humans compared with mice. On the other hand, local expansion of type 17 T-helper (TH17) cells is observed in some animal models, but not in RA. Finally, whereas T-cell depletion therapy essentially failed in RA, antibody targeting of T cells can work, at least preventatively, in most arthritis models. Clearly, additional human and animal studies are needed to fill the gap in our understanding of the specific contribution of T-cell subsets to arthritis in mice and men.

Satellite cell therapy – from mice to men

PubMed Central

2013-01-01

Satellite cells are rare mononuclear skeletal muscle-resident cells that are the chief contributors to regenerative myogenesis following muscle injury. Although first identified more than 50 years ago, it is only recently that the murine satellite cell has become molecularly defined with the ability to prospectively isolate these cells from their niche. Human satellite cells are considerably less well understood with relatively few studies having been performed on them. In this review, a critical evaluation of this literature is provided along with a discussion of the practical and methodological issues involved with research on human satellite cells. The therapeutic potential of these and other cells types is also discussed, and the various challenges that face satellite cell therapy are addressed. PMID:23369649

Surface heat shock protein 90 serves as a potential receptor for calcium oxalate crystal on apical membrane of renal tubular epithelial cells.

PubMed

Fong-Ngern, Kedsarin; Sueksakit, Kanyarat; Thongboonkerd, Visith

2016-07-01

Adhesion of calcium oxalate monohydrate (COM) crystals on renal tubular epithelial cells is a crucial step in kidney stone formation. Finding potential crystal receptors on the apical membrane of the cells may lead to a novel approach to prevent kidney stone disease. Our previous study identified a large number of crystal-binding proteins on the apical membrane of MDCK cells. However, their functional role as potential crystal receptors had not been validated. The present study aimed to address the potential role of heat shock protein 90 (HSP90) as a COM crystal receptor. The apical membrane was isolated from polarized MDCK cells by the peeling method and recovered proteins were incubated with COM crystals. Western blot analysis confirmed the presence of HSP90 in the apical membrane and the crystal-bound fraction. Immunofluorescence staining without permeabilization and laser-scanning confocal microscopy confirmed the surface HSP90 expression on the apical membrane of the intact cells. Crystal adhesion assay showed that blocking surface HSP90 by specific anti-HSP90 antibody and knockdown of HSP90 by small interfering RNA (siRNA) dramatically reduced crystal binding on the apical surface of MDCK cells (by approximately 1/2 and 2/3, respectively). Additionally, crystal internalization assay revealed the presence of HSP90 on the membrane of endocytic vesicle containing the internalized COM crystal. Moreover, pretreatment of MDCK cells with anti-HSP90 antibody significantly reduced crystal internalization (by approximately 1/3). Taken together, our data indicate that HSP90 serves as a potential receptor for COM crystals on the apical membrane of renal tubular epithelial cells and is involved in endocytosis/internalization of the crystals into the cells.

GTSE1: a novel TEAD4-E2F1 target gene involved in cell protrusions formation in triple-negative breast cancer cell models

PubMed Central

Stelitano, Debora; Leticia, Yamila Peche; Dalla, Emiliano; Monte, Martin; Piazza, Silvano; Schneider, Claudio

2017-01-01

GTSE1 over-expression has been reported as a potential marker for metastasis in various types of malignancies, including breast cancer. Despite this, the transcriptional regulation of this protein and the causes of its misregulation in tumors remain largely unknown. The aims of this work were to elucidate how GTSE1 is regulated at the transcriptional level and to clarify the mechanism underlying GTSE1-dependent cell functions in triple-negative breast cancer (TNBC). Here, we identified GTSE1 as a novel target gene of the TEAD4 transcription factor, highlighting a role for the YAP and TAZ coactivators in the transcriptional regulation of GTSE1. Moreover, we found that TEAD4 controls the formation of cell protrusions required for cell migration through GTSE1, unveiling a relevant effector role for this protein in the TEAD-dependent cellular functions and confirming TEAD4 role in promoting invasion and metastasis in breast cancer. Finally, we highlighted a role for the pRb-E2F1 pathway in the control of GTSE1 transcription and observed that treatment with drugs targeting the pRb-E2F1 or YAP/TAZ-TEAD pathways dramatically downregulated the expression levels of GTSE1 and of other genes involved in the formation of metastasis, suggesting their potential use in the treatment of TNBC. PMID:28978043

Transient Receptor Potential Channels in the Vasculature

PubMed Central

Earley, Scott; Brayden, Joseph E.

2015-01-01

The mammalian genome encodes 28 distinct members of the transient receptor potential (TRP) superfamily of cation channels, which exhibit varying degrees of selectivity for different ionic species. Multiple TRP channels are present in all cells and are involved in diverse aspects of cellular function, including sensory perception and signal transduction. Notably, TRP channels are involved in regulating vascular function and pathophysiology, the focus of this review. TRP channels in vascular smooth muscle cells participate in regulating contractility and proliferation, whereas endothelial TRP channel activity is an important contributor to endothelium-dependent vasodilation, vascular wall permeability, and angiogenesis. TRP channels are also present in perivascular sensory neurons and astrocytic endfeet proximal to cerebral arterioles, where they participate in the regulation of vascular tone. Almost all of these functions are mediated by changes in global intracellular Ca2+ levels or subcellular Ca2+ signaling events. In addition to directly mediating Ca2+ entry, TRP channels influence intracellular Ca2+ dynamics through membrane depolarization associated with the influx of cations or through receptor- or store-operated mechanisms. Dysregulation of TRP channels is associated with vascular-related pathologies, including hypertension, neointimal injury, ischemia-reperfusion injury, pulmonary edema, and neurogenic inflammation. In this review, we briefly consider general aspects of TRP channel biology and provide an in-depth discussion of the functions of TRP channels in vascular smooth muscle cells, endothelial cells, and perivascular cells under normal and pathophysiological conditions. PMID:25834234

Determination of the transforming activities of adenovirus oncogenes.

PubMed

Speiseder, Thomas; Nevels, Michael; Dobner, Thomas

2014-01-01

The last 50 years of molecular biological investigations into human adenoviruses (Ads) have contributed enormously to our understanding of the basic principles of normal and malignant cell growth. Much of this knowledge stems from analyses of the Ad productive infection cycle in permissive host cells. Also, initial observations concerning the transforming potential of human Ads subsequently revealed decisive insights into the molecular mechanisms of the origins of cancer and established Ads as a model system for explaining virus-mediated transformation processes. Today it is well established that cell transformation by human Ads is a multistep process involving several gene products encoded in early transcription units 1A (E1A) and 1B (E1B). Moreover, a large body of evidence now indicates that alternative or additional mechanisms are engaged in Ad-mediated oncogenic transformation involving gene products encoded in early region 4 (E4) as well as epigenetic changes resulting from viral DNA integration. In particular, studies on the transforming potential of several E4 gene products have now revealed new pathways that point to novel general mechanisms of virus-mediated oncogenesis. In this chapter we describe in vitro and in vivo assays to determine the transforming and oncogenic activities of the E1A, E1B, and E4 oncoproteins in primary baby rat kidney cells, human amniotic fluid cells and athymic nude mice.

Serotonin Shapes the Migratory Potential of NK Cells - An in vitro Approach.

PubMed

Zimmer, Philipp; Bloch, Wilhelm; Kieven, Markus; Lövenich, Lukas; Lehmann, Jonas; Holthaus, Michelle; Theurich, Sebastian; Schenk, Alexander

2017-10-01

Increased serotonin (5-HT) levels have been shown to influence natural killer cell (NK cell) function. Acute exercise mobilizes and activates NK cells and further increases serum 5-HT concentrations in a dose-dependent manner. The aim of this study was to investigate the impact of different serum 5-HT concentrations on NK cell migratory potential and cytotoxicity. The human NK cell line KHYG-1 was assigned to 4 conditions, including 3 physiological concentrations of 5-HT (100, 130 or 170 µg/l 5-HT) and one control condition. NK cells were analyzed regarding cytotoxicity, migratory potential and expression of adhesion molecules. No treatment effect on NK cell cytotoxicity and expression of integrin subunits was detected. Migratory potential was increased in a dose dependent manner, indicating the highest protease activity in cells that were incubated with 170 µg/l 5-HT (170 µg/l vs. control, p<0.001, 170 µg/l vs. 100 µg/l, p<0.001; 170 µg/l vs. 130 µg/l, p=0.003; 130 µg/l vs. control, p<0.001, 130 µg/l vs. 100 µg/l, p<0.001). These results suggest that elevated 5-HT serum levels play a mediating role in NK cell function. As exercise has been shown to be involved in NK cell mobilization and redistribution, the influence of 5-HT should be investigated in ex vivo and in vivo experiments. © Georg Thieme Verlag KG Stuttgart · New York.

Generators of the brainstem auditory evoked potential in cat. I. An experimental approach to their identification.

PubMed

Melcher, J R; Knudson, I M; Fullerton, B C; Guinan, J J; Norris, B E; Kiang, N Y

1996-04-01

This paper is the first in a series aimed at identifying the cellular generators of the brainstem auditory evoked potential (BAEP) in cats. The approach involves (1) developing experimental procedures for making small selective lesions and determining the corresponding changes in BAEP waveforms, (2) identifying brainstem regions involved in BAEP generation by examining the effects of lesions on the BAEP and (3) identifying specific cell populations involved by combining the lesion results with electrophysiological and anatomical information from other kinds of studies. We created lesions in the lower brainstem by injecting kainic acid which is generally toxic for neuronal cell bodies but not for axons and terminals. This first paper describes the justifications for using kainic acid, explains the associated problems, and develops a methodology that addresses the main difficulties. The issues and aspects of the specific methods are generally applicable to physiological and anatomical studies using any neurotoxin, as well as to the present BAEP study. The methods chosen involved (1) measuring the BAEP at regular intervals until it reached a post-injection steady state and perfusing the animals with fixative shortly after the last BAEP recordings were made, (2) using objective criteria to distinguish injection-related BAEP changes from unrelated ones, (3) making control injections to identify effects not due to kainic acid toxicity, (4) verifying the anatomical and functional integrity of axons in lesioned regions, and (5) examining injected brainstems microscopically for cell loss and cellular abnormalities indicating dysfunction. This combination of methods enabled us to identify BAEP changes which are clearly correlated with lesion locations.

The Network of Epithelial-mesenchymal transition: potential new targets for tumor resistance

PubMed Central

Nantajit, Danupon; Lin, Dong; Li, Jian Jian

2014-01-01

Purpose In multiple cell metazoans, the ability of polarized epithelial cells to convert to motile mesenchymal cells in order to relocate to another location is governed by a unique process termed epithelial-mesenchymal transition (EMT). While being an essential process of cellular plasticity for normal tissue and organ developments, EMT is found to be involved in an array of malignant phenotypes of tumor cells including proliferation and invasion, angiogenesis, stemness of cancer cells and resistance to chemo-radiotherapy. Although EMT is being extensively studied and demonstrated to play a key role in tumor metastasis and in sustaining tumor hallmarks, there is a lack of clear picture of the overall EMT signaling network, wavering the potential clinical trials targeting EMT. Methods In this review, we highlight the potential key therapeutic targets of EMT linked with tumor aggressiveness, hypoxia, angiogenesis and cancer stem cells, emphasizing on an emerging EMT-associated NF-κB/HER2/STAT3 pathway in radioresistance of breast cancer stem cells. Results Further definition of cancer stem cell repopulation due to EMT-controlled tumor microenvironment will help to understand how tumors exploit the EMT mechanisms for their survival and expansion advantages. Conclusions The knowledge of EMT will offer more effective targets in clinical trials to treat therapy-resistant metastatic lesions. PMID:25270087

Possible involvement of persistent activity of the mammalian target of rapamycin pathway in the cisplatin resistance of AFP-producing gastric cancer cells.

PubMed

Kamata, Shigeyuki; Kishimoto, Takashi; Kobayashi, Soichi; Miyazaki, Masaru; Ishikura, Hiroshi

2007-07-01

AFP-producing gastric carcinoma (AFPGC) is a highly malignant variant of gastric cancer. An effective chemotherapy is needed to improve on the poor outcome of this disease. Survival signals activated by intracellular kinase networks could be involved in chemoresistance in malignant tumors. We investigated the role of a pivotal kinase pathway, the mammalian target of rapamycin complex 1 (mTORC1) pathway, in the effectiveness of chemotherapeutic agents in three AFPGC cell lines (GCIY, FU97 and Takigawa) as well as in four cell lines of conventional-type gastric carcinoma (CGC). AFPGC cells were generally resistant to multiple chemotherapeutic agents, including cisplatin, while CGC cells were generally sensitive. Downstream targets of mTORC1, including p70S6K and 4EBP1, were phosphorylated in all cell lines. Interestingly, cisplatin virtually abolished phosphorylation of p70S6K and 4EBP1 in CGC cells, while phosphorylation was maintained in cisplatin-treated AFPGC cells. The addition of rapamycin, an inhibitor of mTORC1, diminished the remaining activity of mTORC1 and significantly intensified the cytotoxic action of cisplatin in AFPGC cells. These results suggested that persistent activity of mTORC1 signals in cisplatin-treated AFPGC cells is involved in the mechanisms of cisplatin resistance in AFPGC. Finally, combined treatment of rapamycin and cisplatin significantly suppressed the subcutaneously implanted GCIY cells. In conclusion rapamycin may be a potential supplemental agent for the treatment of AFPGC when used in combination with cisplatin.

ATP release from freshly isolated guinea-pig bladder urothelial cells: a quantification and study of the mechanisms involved.

PubMed

McLatchie, Linda M; Fry, Christopher H

2015-06-01

To quantify the amount of ATP released from freshly isolated bladder urothelial cells, study its control by intracellular and extracellular calcium and identify the pathways responsible for its release. Urothelial cells were isolated from male guinea-pig urinary bladders and stimulated to release ATP by imposition of drag forces by repeated pipetting. ATP was measured using a luciferin-luciferase assay and the effects of modifying internal and external calcium concentration and blockers of potential release pathways studied. Freshly isolated guinea-pig urothelial cells released ATP at a mean (sem) rate of 1.9 (0.1) pmoles/mm(2) cell membrane, corresponding to about 700 pmoles/g of tissue, and about half [49 (6)%, n = 9) of the available cell ATP. This release was reduced to a mean (sem) of 0.46 (0.08) pmoles/mm(2) (160 pmoles/g) with 1.8 mm external calcium, and was increased about two-fold by increasing intracellular calcium. The release from umbrella cells was not significantly different from a mixed intermediate and basal cell population, suggesting that all three groups of cells release a similar amount of ATP per unit area. ATP release was reduced by ≈ 50% by agents that block pannexin and connexin hemichannels. It is suggested that the remainder may involve vesicular release. A significant fraction of cellular ATP is released from isolated urothelial cells by imposing drag forces that cause minimal loss of cell viability. This release involves multiple release pathways, including hemichannels and vesicular release. © 2014 The Authors BJU International © 2014 BJU International.

ABCB1 regulation through LRPPRC is influenced by the methylation status of the GC -100 box in its promoter

PubMed Central

Corrêa, Stephany; Binato, Renata; Du Rocher, Bárbara; Ferreira, Gerson; Cappelletti, Paola; Soares-Lima, Sheila; Pinto, Luis Felipe; Mencalha, André; Abdelhay, Eliana

2014-01-01

One of the potential mechanisms of imatinib mesylate (IM) resistance in chronic myeloid leukemia (CML) is increased level of P-glycoprotein (Pgp). Pgp is an efflux pump capable of activating the multidrug resistance (MDR) phenotype. The gene encoding Pgp (ABCB1) has several binding sites in its promoter region, along with CpG islands and GC boxes, involved in its epigenetic control. In previous work, we performed a proteomic study to identify proteins involved in IM cross-resistance in acute leukemia. Among these proteins, we identified LRPPRC as a potential regulator of ABCB1 transcription via an invMED1 binding site in ABCB1. Interestingly, this invMED1 binding site overlaps with the GC -100 box. In this work, we investigated the potential role of LRPPRC in the regulation of ABCB1 transcriptional activity in CML resistance. In addition, we evaluated the potential connection between this regulation and the methylation status of the ABCB1 promoter in its GC -100 box. Our results show that LRPPRC binds prominently to the ABCB1 promoter in Lucena cells, an IM-resistant cell line. Luciferase assays showed that ABCB1 transcription is positively regulated by LRPPRC upon its knockdown. Pyrosequencing analysis showed that the ABCB1 promoter is differentially methylated at its GC -100 box in K562 cells compared with Lucena cells, and in CML patients with different response to IM. Chromatin immunoprecipitation and Pgp expression after DNA demethylation treatment showed that LRPPRC binding is affected by the methylation status of ABCB1 GC -100 box. Taken together, our findings indicate that LRPPRC is a transcription factor related to ABCB1 expression and highlight the importance of epigenetic regulation in CML resistance. PMID:25089713

ABCB1 regulation through LRPPRC is influenced by the methylation status of the GC -100 box in its promoter.

PubMed

Corrêa, Stephany; Binato, Renata; Du Rocher, Bárbara; Ferreira, Gerson; Cappelletti, Paola; Soares-Lima, Sheila; Pinto, Luis Felipe; Mencalha, André; Abdelhay, Eliana

2014-08-01

One of the potential mechanisms of imatinib mesylate (IM) resistance in chronic myeloid leukemia (CML) is increased level of P-glycoprotein (Pgp). Pgp is an efflux pump capable of activating the multidrug resistance (MDR) phenotype. The gene encoding Pgp (ABCB1) has several binding sites in its promoter region, along with CpG islands and GC boxes, involved in its epigenetic control. In previous work, we performed a proteomic study to identify proteins involved in IM cross-resistance in acute leukemia. Among these proteins, we identified LRPPRC as a potential regulator of ABCB1 transcription via an invMED1 binding site in ABCB1. Interestingly, this invMED1 binding site overlaps with the GC -100 box. In this work, we investigated the potential role of LRPPRC in the regulation of ABCB1 transcriptional activity in CML resistance. In addition, we evaluated the potential connection between this regulation and the methylation status of the ABCB1 promoter in its GC -100 box. Our results show that LRPPRC binds prominently to the ABCB1 promoter in Lucena cells, an IM-resistant cell line. Luciferase assays showed that ABCB1 transcription is positively regulated by LRPPRC upon its knockdown. Pyrosequencing analysis showed that the ABCB1 promoter is differentially methylated at its GC -100 box in K562 cells compared with Lucena cells, and in CML patients with different response to IM. Chromatin immunoprecipitation and Pgp expression after DNA demethylation treatment showed that LRPPRC binding is affected by the methylation status of ABCB1 GC -100 box. Taken together, our findings indicate that LRPPRC is a transcription factor related to ABCB1 expression and highlight the importance of epigenetic regulation in CML resistance.

An intracellular analysis of the visual responses of neurones in cat visual cortex.

PubMed Central

Douglas, R J; Martin, K A; Whitteridge, D

1991-01-01

1. Extracellular and intracellular recordings were made from neurones in the visual cortex of the cat in order to compare the subthreshold membrane potentials, reflecting the input to the neurone, with the output from the neurone seen as action potentials. 2. Moving bars and edges, generated under computer control, were used to stimulate the neurones. The membrane potential was digitized and averaged for a number of trials after stripping the action potentials. Comparison of extracellular and intracellular discharge patterns indicated that the intracellular impalement did not alter the neurones' properties. Input resistance of the neurone altered little during stable intracellular recordings (30 min-2 h 50 min). 3. Intracellular recordings showed two distinct patterns of membrane potential changes during optimal visual stimulation. The patterns corresponded closely to the division of S-type (simple) and C-type (complex) receptive fields. Simple cells had a complex pattern of membrane potential fluctuations, involving depolarizations alternating with hyperpolarizations. Complex cells had a simple single sustained plateau of depolarization that was often followed but not preceded by a hyperpolarization. In both simple and complex cells the depolarizations led to action potential discharges. The hyperpolarizations were associated with inhibition of action potential discharge. 4. Stimulating simple cells with non-optimal directions of motion produced little or no hyperpolarization of the membrane in most cases, despite a lack of action potential output. Directional complex cells always produced a single plateau of depolarization leading to action potential discharge in both the optimal and non-optimal directions of motion. The directionality could not be predicted on the basis of the position of the hyperpolarizing inhibitory potentials found in the optimal direction. 5. Stimulation of simple cells with non-optimal orientations occasionally produced slight hyperpolarizations and inhibition of action potential discharge. Complex cells, which had broader orientation tuning than simple cells, could show marked hyperpolarization for non-optimal orientations, but this was not generally the case. 6. The data do not support models of directionality and orientation that rely solely on strong inhibitory mechanisms to produce stimulus selectivity. PMID:1804981

TRPM4 non-selective cation channels influence action potentials in rabbit Purkinje fibres.

PubMed

Hof, Thomas; Sallé, Laurent; Coulbault, Laurent; Richer, Romain; Alexandre, Joachim; Rouet, René; Manrique, Alain; Guinamard, Romain

2016-01-15

The transient receptor potential melastatin 4 (TRPM4) inhibitor 9-phenanthrol reduces action potential duration in rabbit Purkinje fibres but not in ventricle. TRPM4-like single channel activity is observed in isolated rabbit Purkinje cells but not in ventricular cells. The TRPM4-like current develops during the notch and early repolarization phases of the action potential in Purkinje cells. Transient receptor potential melastatin 4 (TRPM4) Ca(2+)-activated non-selective cation channel activity has been recorded in cardiomyocytes and sinus node cells from mammals. In addition, TRPM4 gene mutations are associated with human diseases of cardiac conduction, suggesting that TRPM4 plays a role in this aspect of cardiac function. Here we evaluate the TRPM4 contribution to cardiac electrophysiology of Purkinje fibres. Ventricular strips with Purkinje fibres were isolated from rabbit hearts. Intracellular microelectrodes recorded Purkinje fibre activity and the TRPM4 inhibitor 9-phenanthrol was applied to unmask potential TRPM4 contributions to the action potential. 9-Phenanthrol reduced action potential duration measured at the point of 50 and 90% repolarization with an EC50 of 32.8 and 36.1×10(-6) mol l(-1), respectively, but did not modulate ventricular action potentials. Inside-out patch-clamp recordings were used to monitor TRPM4 activity in isolated Purkinje cells. TRPM4-like single channel activity (conductance = 23.8 pS; equal permeability for Na(+) and K(+); sensitivity to voltage, Ca(2+) and 9-phenanthrol) was observed in 43% of patches from Purkinje cells but not from ventricular cells (0/16). Action potential clamp experiments performed in the whole-cell configuration revealed a transient inward 9-phenanthrol-sensitive current (peak density = -0.65 ± 0.15 pA pF(-1); n = 5) during the plateau phases of the Purkinje fibre action potential. These results show that TRPM4 influences action potential characteristics in rabbit Purkinje fibres and thus could modulate cardiac conduction and be involved in triggering arrhythmias. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Possible involvement of TLRs and hemichannels in stress-induced CNS dysfunction via mastocytes, and glia activation.

PubMed

Aguirre, Adam; Maturana, Carola J; Harcha, Paloma A; Sáez, Juan C

2013-01-01

In the central nervous system (CNS), mastocytes and glial cells (microglia, astrocytes and oligodendrocytes) function as sensors of neuroinflammatory conditions, responding to stress triggers or becoming sensitized to subsequent proinflammatory challenges. The corticotropin-releasing hormone and glucocorticoids are critical players in stress-induced mastocyte degranulation and potentiation of glial inflammatory responses, respectively. Mastocytes and glial cells express different toll-like receptor (TLR) family members, and their activation via proinflammatory molecules can increase the expression of connexin hemichannels and pannexin channels in glial cells. These membrane pores are oligohexamers of the corresponding protein subunits located in the cell surface. They allow ATP release and Ca(2+) influx, which are two important elements of inflammation. Consequently, activated microglia and astrocytes release ATP and glutamate, affecting myelinization, neuronal development, and survival. Binding of ligands to TLRs induces a cascade of intracellular events leading to activation of several transcription factors that regulate the expression of many genes involved in inflammation. During pregnancy, the previous responses promoted by viral infections and other proinflammatory conditions are common and might predispose the offspring to develop psychiatric disorders and neurological diseases. Such disorders could eventually be potentiated by stress and might be part of the etiopathogenesis of CNS dysfunctions including autism spectrum disorders and schizophrenia.

Extract of Ginkgo biloba exacerbates liver metastasis in a mouse colon cancer Xenograft model.

PubMed

Wang, Huan; Wu, Xia; Lezmi, Stephane; Li, Qian; Helferich, William G; Xu, Yueqing; Chen, Hong

2017-12-02

Metastasis refers to the spread of a primary tumor cell from the primary site to other locations in the body and it is generally associated with the severity of a tumor. Extract of Ginkgo biloba (EGb) contains various bioactive compounds and it exerts beneficial effects including improvements in brain function and reduced risk of cardiovascular diseases. On the other hand, increased risk of thyroid and liver cancers by EGb have been reported in animals. A colon cancer metastasis model was established using intrasplenic injection of a human colon cancer cell line, SW620-luc in athymic mice to investigate the potential impact of EGb on colon cancer progression. After tumor establishment, EGb was intraperitonically injected daily for 5 wks. EGb significantly increased the rate of metastasis in mouse liver and decreased the number of necrotic and apoptotic cells in the metastatic liver when compared to the control. Meanwhile, EGb significantly induced proliferation of tumor cells in the metastatic liver, indicated by increased staining of Ki67 and H3S10p. mRNA expression of genes involved in cell cycle, metastasis, apoptosis, and oxidative stress were altered by EGb treatment in livers with tumors. Moreover, EGb activated the stress-responsive MAPK pathways in the liver with metastatic tumors. EGb exacerbated liver metastasis in a mouse colon cancer metastasis model. This is potentially due to the increased tumor cell proliferation involving stimulated MAPK pathways.

Human bone marrow-derived mesenchymal cells differentiate and mature into endocrine pancreatic lineage in vivo.

PubMed

Phadnis, Smruti M; Joglekar, Mugdha V; Dalvi, Maithili P; Muthyala, Sudhakar; Nair, Prabha D; Ghaskadbi, Surendra M; Bhonde, Ramesh R; Hardikar, Anandwardhan A

2011-03-01

The scarcity of human islets for transplantation remains a major limitation of cell replacement therapy for diabetes. Bone marrow-derived progenitor cells are of interest because they can be isolated, expanded and offered for such therapy under autologous/allogeneic settings. We characterized and compared human bone marrow-derived mesenchymal cells (hBMC) obtained from (second trimester), young (1-24 years) and adult (34-81 years) donors. We propose a novel protocol that involves assessment of paracrine factors from regenerating pancreas in differentiation and maturation of hBMC into endocrine pancreatic lineage in vivo. We observed that donor age was inversely related to growth potential of hBMC. Following in vitro expansion and exposure to specific growth factors involved in pancreatic development, hBMC migrated and formed islet-like cell aggregates (ICA). ICA show increased abundance of pancreatic transcription factors (Ngn3, Brn4, Nkx6.1, Pax6 and Isl1). Although efficient differentiation was not achieved in vitro, we observed significant maturation and secretion of human c-peptide (insulin) upon transplantation into pancreactomized and Streptozotocin (STZ)-induced diabetic mice. Transplanted ICA responded to glucose and maintained normoglycemia in diabetic mice. Our data demonstrate that hBMC have tremendous in vitro expansion potential and can be differentiated into multiple lineages, including the endocrine pancreatic lineage. Paracrine factors secreted from regenerating pancreas help in efficient differentiation and maturation of hBMC, possibly via recruiting chromatin modulators, to generate glucose-responsive insulin-secreting cells.

Protective Effect of Neuropeptide Apelin-13 on 6-Hydroxydopamine-Induced Neurotoxicity in SH-SY5Y Dopaminergic Cells: Involvement of Its Antioxidant and Antiapoptotic Properties.

PubMed

Pouresmaeili-Babaki, Elham; Esmaeili-Mahani, Saeed; Abbasnejad, Mehdi; Ravan, Hadi

2018-04-01

Parkinson's disease (PD) is a severe neurodegenerative disorder characterized by the loss of brain dopaminergic neurons. Beside pharmacologic and symptomatic treatment of PD the neuroprotective therapy has recently attracted more attention. Apelin, a novel neuropeptide, and its receptors have numerous reported roles in regulating brain functions. In addition, this peptide has potent neuroprotective effects in some neurodegenerative situations. In this study, the effects of apelin-13 were investigated in a cell model of PD. Human neuroblastoma SH-SY5Y cell damage was induced by 150 μM 6-hydroxydopamine (6-OHDA) and the cells viability was examined by MTT assay. Intracellular reactive oxygen species (ROS) and mitochondrial membrane potential were determined by fluorescence spectrophotometry method. Immunoblotting analysis was also employed to evaluate cytochrome c release and caspase-3 activity. Data showed that 6-OHDA could decrease cell viability and mitochondrial membrane potential and increase intracellular ROS, cytochrome c, and cleaved caspase-3 levels. Pretreatment of SH-SY5Y cells with apelin-13 (5 and 10 nM) significantly prevented the mentioned biochemical and molecular markers of 6-OHDA-induced neurotoxicity. Furthermore, the results showed that apelin receptor and PI3K signaling contributed to the observed protective effects of apelin. The results suggest that apelin-13 has protective effects against dopaminergic neural toxicity and its antioxidant and antiapoptotic properties are involved, at least in part, in such protection.

Involvement of multiple stressors induced by non-thermal plasma-charged aerosols during inactivation of airborne bacteria

PubMed Central

Vaze, Nachiket D.; Park, Sin; Brooks, Ari D.; Fridman, Alexander; Joshi, Suresh G.

2017-01-01

A lab-scale, tunable, single-filament, point-to-point nonthermal dieletric-barrier discharge (DBD) plasma device was built to study the mechanisms of inactivation of aerosolized bacterial pathogens. The system inactivates airborne antibiotic-resistant pathogens efficiently. Nebulization mediated pre-optimized (4 log and 7 log) bacterial loads were challenged to plasma-charged aerosols, and lethal and sublethal doses determined using colony assay, and cell viability assay; and the loss of membrane potential and cellular respiration were determined using cell membrane potential assay and XTT assay. Using the strategies of Escherichia coli wildtype, over-expression mutant, deletion mutants, and peroxide and heat stress scavenging, we analyzed activation of intracellular reactive oxygen species (ROS) and heat shock protein (hsp) chaperons. Superoxide dismutase deletion mutants (ΔsodA, ΔsodB, ΔsodAΔsodB) and catalase mutants ΔkatG and ΔkatEΔkatG did not show significant difference from wildtype strain, and ΔkatE and ΔahpC was found significantly more susceptible to cell death than wildtype. The oxyR regulon was found to mediate plasma-charged aerosol-induced oxidative stress in bacteria. Hsp deficient E. coli (ΔhtpG, ΔgroEL, ΔclpX, ΔgrpE) showed complete inactivation of cells at ambient temperature, and the treatment at cold temperature (4°C) significantly protected hsp deletion mutants and wildtype cells, and indicate a direct involvement of hsp in plasma-charged aerosol mediated E. coli cell death. PMID:28166240

Silver Nanoparticles: Synthetic Routes, In Vitro Toxicity and Theranostic Applications for Cancer Disease.

PubMed

De Matteis, Valeria; Cascione, Mariafrancesca; Toma, Chiara Cristina; Leporatti, Stefano

2018-05-10

The large use of nanomaterials in many fields of application and commercial products highlights their potential toxicity on living organisms and the environment, despite their physico-chemical properties. Among these, silver nanoparticles (Ag NPs) are involved in biomedical applications such as antibacterial agents, drug delivery vectors and theranostics agents. In this review, we explain the common synthesis routes of Ag NPs using physical, chemical, and biological methods, following their toxicity mechanism in cells. In particular, we analyzed the physiological cellular pathway perturbations in terms of oxidative stress induction, mitochondrial membrane potential alteration, cell death, apoptosis, DNA damage and cytokines secretion after Ag NPs exposure. In addition, their potential anti-cancer activity and theranostic applications are discussed.

The kin17 Protein in Murine Melanoma Cells

PubMed Central

Ramos, Anelise C.; Gaspar, Vanessa P.; Kelmer, Sabrina M. G.; Sellani, Tarciso A.; Batista, Ana G. U.; De Lima Neto, Quirino A.; Rodrigues, Elaine G.; Fernandez, Maria A.

2015-01-01

kin17 has been described as a protein involved in the processes of DNA replication initiation, DNA recombination, and DNA repair. kin17 has been studied as a potential molecular marker of breast cancer. This work reports the detection and localization of this protein in the murine melanoma cell line B16F10-Nex2 and in two derived subclones with different metastatic potential, B16-8HR and B16-10CR. Nuclear and chromatin-associated protein fractions were analyzed, and kin17 was detected in all fractions, with an elevated concentration observed in the chromatin-associated fraction of the clone with low metastatic potential, suggesting that the kin17 expression level could be a marker of melanoma. PMID:26610484

Plasma rich in growth factors (PRGF-Endoret) stimulates tendon and synovial fibroblasts migration and improves the biological properties of hyaluronic acid.

PubMed

Anitua, E; Sanchez, M; De la Fuente, M; Zalduendo, M M; Orive, G

2012-09-01

Cell migration plays an essential role in development, wound healing, and tissue regeneration. Plasma rich in growth factors (PRGF-Endoret) technology offers a potential source of growth factors involved in tissue regeneration. Here, we evaluate the potential of PRGF-Endoret over tendon cells and synovial fibroblasts migration and study whether the combination of this autologous technology with hyaluronic acid (HA) improves the effect and potential of the biomaterials over the motility of both types of fibroblasts. Migration of primary tendon cells and synovial fibroblasts after culturing with either PRGF or PPGF (plasma poor in growth factors) at different doses was evaluated. Furthermore, the migratory capacity induced by the combination of PPGF and PRGF with HA was tested. PPGF stimulated migration of both types of cells but this effect was significantly higher when PRGF was used. Tendon cells showed an increase of 212% in migratory ability when HA was combined with PPGF and of 335% in the case of HA + PRGF treatment compared with HA alone. PRGF-Endoret stimulates migration of tendon cells and synovial fibroblasts and improves the biological properties of HA.

Magnetic field-controlled gene expression in encapsulated cells

PubMed Central

Ortner, Viktoria; Kaspar, Cornelius; Halter, Christian; Töllner, Lars; Mykhaylyk, Olga; Walzer, Johann; Günzburg, Walter H.; Dangerfield, John A.; Hohenadl, Christine; Czerny, Thomas

2012-01-01

Cell and gene therapies have an enormous range of potential applications, but as for most other therapies, dosing is a critical issue, which makes regulated gene expression a prerequisite for advanced strategies. Several inducible expression systems have been established, which mainly rely on small molecules as inducers, such as hormones or antibiotics. The application of these inducers is difficult to control and the effects on gene regulation are slow. Here we describe a novel system for induction of gene expression in encapsulated cells. This involves the modification of cells to express potential therapeutic genes under the control of a heat inducible promoter and the co-encapsulation of these cells with magnetic nanoparticles. These nanoparticles produce heat when subjected to an alternating magnetic field; the elevated temperatures in the capsules then induce gene expression. In the present study we define the parameters of such systems and provide proof-of-principle using reporter gene constructs. The fine-tuned heating of nanoparticles in the magnetic field allows regulation of gene expression from the outside over a broad range and within short time. Such a system has great potential for advancement of cell and gene therapy approaches. PMID:22197778

PITX2 Enhances the Regenerative Potential of Dystrophic Skeletal Muscle Stem Cells.

PubMed

Vallejo, Daniel; Hernández-Torres, Francisco; Lozano-Velasco, Estefanía; Rodriguez-Outeiriño, Lara; Carvajal, Alejandra; Creus, Carlota; Franco, Diego; Aránega, Amelia Eva

2018-04-10

Duchenne muscular dystrophy (DMD), one of the most lethal genetic disorders, involves progressive muscle degeneration resulting from the absence of DYSTROPHIN. Lack of DYSTROPHIN expression in DMD has critical consequences in muscle satellite stem cells including a reduced capacity to generate myogenic precursors. Here, we demonstrate that the c-isoform of PITX2 transcription factor modifies the myogenic potential of dystrophic-deficient satellite cells. We further show that PITX2c enhances the regenerative capability of mouse DYSTROPHIN-deficient satellite cells by increasing cell proliferation and the number of myogenic committed cells, but importantly also increasing dystrophin-positive (revertant) myofibers by regulating miR-31. These PITX2-mediated effects finally lead to improved muscle function in dystrophic (DMD/mdx) mice. Our studies reveal a critical role for PITX2 in skeletal muscle repair and may help to develop therapeutic strategies for muscular disorders. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Exosomes: From Garbage Bins to Promising Therapeutic Targets

PubMed Central

H. Rashed, Mohammed; Bayraktar, Emine; K. Helal, Gouda; Abd-Ellah, Mohamed F.; Amero, Paola; Chavez-Reyes, Arturo; Rodriguez-Aguayo, Cristian

2017-01-01

Intercellular communication via cell-released vesicles is a very important process for both normal and tumor cells. Cell communication may involve exosomes, small vesicles of endocytic origin that are released by all types of cells and are found in abundance in body fluids, including blood, saliva, urine, and breast milk. Exosomes have been shown to carry lipids, proteins, mRNAs, non-coding RNAs, and even DNA out of cells. They are more than simply molecular garbage bins, however, in that the molecules they carry can be taken up by other cells. Thus, exosomes transfer biological information to neighboring cells and through this cell-to-cell communication are involved not only in physiological functions such as cell-to-cell communication, but also in the pathogenesis of some diseases, including tumors and neurodegenerative conditions. Our increasing understanding of why cells release exosomes and their role in intercellular communication has revealed the very complex and sophisticated contribution of exosomes to health and disease. The aim of this review is to reveal the emerging roles of exosomes in normal and pathological conditions and describe the controversial biological role of exosomes, as it is now understood, in carcinogenesis. We also summarize what is known about exosome biogenesis, composition, functions, and pathways and discuss the potential clinical applications of exosomes, especially as biomarkers and novel therapeutic agents. PMID:28257101

Evidence for the involvement of lung-specific γδ T cell subsets in local responses to Streptococcus pneumoniae infection

PubMed Central

Kirby, Alun C; Newton, Darren J; Carding, Simon R; Kaye, Paul M

2007-01-01

Although γδ T cells are involved in the response to many pathogens, the dynamics and heterogeneity of the local γδ T cell response remains poorly defined. We recently identified γδ T cells as regulators of macrophages and dendritic cells during the resolution of Streptococcus pneumoniae-mediated lung inflammation. Here, using PCR, spectratype analysis and flow cytometry, we show that multiple γδ T cell subsets, including those bearing Vγ1, Vγ4 and Vγ6 TCR, increase in number in the lungs of infected mice, but not in associated lymphoid tissue. These γδ T cells displayed signs of activation, as defined by CD69 and CD25 expression. In vivo BrdU incorporation suggested that local expansion, rather than recruitment, was the principal mechanism underlying this increase in γδ T cells. This conclusion was supported by the finding that pulmonary γδ T cells, but not αβ T cells, isolated from mice that had resolved infection exhibited lung-homing capacity in both naive and infected recipients. Together, these data provide novel insights into the origins of the heterogeneous γδ T cell response that accompanies lung infection, and the first evidence that inflammation-associated γδ T cells may exhibit distinct tissue-homing potential. PMID:18022862

Involvement of Ubiquitin-Editing Protein A20 in Modulating Inflammation in Rat Cochlea Associated with Silver Nanoparticle-Induced CD68 Upregulation and TLR4 Activation

NASA Astrophysics Data System (ADS)

Feng, Hao; Pyykkö, Ilmari; Zou, Jing

2016-05-01

Silver nanoparticles (AgNPs) were shown to temporarily impair the biological barriers in the skin of the external ear canal, mucosa of the middle ear, and inner ear, causing partially reversible hearing loss after delivery into the middle ear. The current study aimed to elucidate the molecular mechanism, emphasizing the TLR signaling pathways in association with the potential recruitment of macrophages in the cochlea and the modulation of inflammation by ubiquitin-editing protein A20. Molecules potentially involved in these signaling pathways were thoroughly analysed using immunohistochemistry in the rat cochlea exposed to AgNPs at various concentrations through intratympanic injection. The results showed that 0.4 % AgNPs but not 0.02 % AgNPs upregulated the expressions of CD68, TLR4, MCP1, A20, and RNF11 in the strial basal cells, spiral ligament fibrocytes, and non-sensory supporting cells of Corti's organ. 0.4 % AgNPs had no effect on CD44, TLR2, MCP2, Rac1, myosin light chain, VCAM1, Erk1/2, JNK, p38, IL-1β, TNF-α, TNFR1, TNFR2, IL-10, or TGF-β. This study suggested that AgNPs might confer macrophage-like functions on the strial basal cells and spiral ligament fibrocytes and enhance the immune activities of non-sensory supporting cells of Corti's organ through the upregulation of CD68, which might be involved in TLR4 activation. A20 and RNF11 played roles in maintaining cochlear homeostasis via negative regulation of the expressions of inflammatory cytokines.

Testosterone decreases urinary bladder smooth muscle excitability via novel signaling mechanism involving direct activation of the BK channels

PubMed Central

Hristov, Kiril L.; Parajuli, Shankar P.; Provence, Aaron

2016-01-01

In addition to improving sexual function, testosterone has been reported to have beneficial effects in ameliorating lower urinary tract symptoms by increasing bladder capacity and compliance, while decreasing bladder pressure. However, the cellular mechanisms by which testosterone regulates detrusor smooth muscle (DSM) excitability have not been elucidated. Here, we used amphotericin-B perforated whole cell patch-clamp and single channel recordings on inside-out excised membrane patches to investigate the regulatory role of testosterone in guinea pig DSM excitability. Testosterone (100 nM) significantly increased the depolarization-induced whole cell outward currents in DSM cells. The selective pharmacological inhibition of the large-conductance voltage- and Ca2+-activated K+ (BK) channels with paxilline (1 μM) completely abolished this stimulatory effect of testosterone, suggesting a mechanism involving BK channels. At a holding potential of −20 mV, DSM cells exhibited transient BK currents (TBKCs). Testosterone (100 nM) significantly increased TBKC activity in DSM cells. In current-clamp mode, testosterone (100 nM) significantly hyperpolarized the DSM cell resting membrane potential and increased spontaneous transient hyperpolarizations. Testosterone (100 nM) rapidly increased the single BK channel open probability in inside-out excised membrane patches from DSM cells, clearly suggesting a direct BK channel activation via a nongenomic mechanism. Live-cell Ca2+ imaging showed that testosterone (100 nM) caused a decrease in global intracellular Ca2+ concentration, consistent with testosterone-induced membrane hyperpolarization. In conclusion, the data provide compelling mechanistic evidence that under physiological conditions, testosterone at nanomolar concentrations directly activates BK channels in DSM cells, independent from genomic testosterone receptors, and thus regulates DSM excitability. PMID:27605581

Receptor for advanced glycation end products (RAGE) functions as receptor for specific sulfated glycosaminoglycans, and anti-RAGE antibody or sulfated glycosaminoglycans delivered in vivo inhibit pulmonary metastasis of tumor cells.

PubMed

Mizumoto, Shuji; Takahashi, Jun; Sugahara, Kazuyuki

2012-06-01

Altered expression of chondroitin sulfate (CS) and heparan sulfate (HS) at the surfaces of tumor cells plays a key role in malignant transformation and tumor metastasis. Previously we demonstrated that a Lewis lung carcinoma (LLC)-derived tumor cell line with high metastatic potential had a higher proportion of E-disaccharide units, GlcUA-GalNAc(4,6-O-disulfate), in CS chains than low metastatic LLC cells and that such CS chains are involved in the metastatic process. The metastasis was markedly inhibited by the pre-administration of CS-E from squid cartilage rich in E units or by preincubation with a phage display antibody specific for CS-E. However, the molecular mechanism of the inhibition remains to be investigated. In this study the receptor molecule for CS chains containing E-disaccharides expressed on LLC cells was revealed to be receptor for advanced glycation end products (RAGE), which is a member of the immunoglobulin superfamily predominantly expressed in the lung. Interestingly, RAGE bound strongly to not only E-disaccharide, but also HS-expressing LLC cells. Furthermore, the colonization of the lungs by LLC cells was effectively inhibited by the blocking of CS or HS chains at the tumor cell surface with an anti-RAGE antibody through intravenous injections in a dose-dependent manner. These results provide the clear evidence that RAGE is at least one of the critical receptors for CS and HS chains expressed at the tumor cell surface and involved in experimental lung metastasis and that CS/HS and RAGE are potential molecular targets in the treatment of pulmonary metastasis.

Emerging treatments in alopecia.

PubMed

Falto-Aizpurua, Leyre; Choudhary, Sonal; Tosti, Antonella

2014-12-01

Alopecia is a common concern encountered in the medical practice. Treatment approach varies according to the type and severity of alopecia. However, available treatment options have limited efficacy and several adverse effects. Presently, there are different treatment options being studied to overcome these limitations. Additionally, cellular pathways involved in the pathophysiology of alopecia are further being clarified to potentially target pathogenic molecules. We searched the literature for recently published articles discussing new treatment options as well as mechanisms involved in alopecia. We discuss the use of stem cells, growth factors, cellular pathways and robotic hair transplant, among other emerging therapies used for alopecia. Future looks very promising and new effective treatments such as janus kinase inhibitors could possibly be available for alopecia areata. The stem-cell technology is advancing and companies involved in hair follicle neogenesis are starting clinical trials on patients with androgenetic alopecia.

Metastatic potential of melanoma cells is not affected by electrochemotherapy.

PubMed

Todorovic, Vesna; Sersa, Gregor; Mlakar, Vid; Glavac, Damjan; Flisar, Karel; Cemazar, Maja

2011-06-01

Electrochemotherapy is a local treatment combining chemotherapy and application of electric pulses to the tumour. Electrochemotherapy with bleomycin and cisplatin has shown its effectiveness in controlling local tumour growth in the treatment of malignant melanoma. However, the effect of electrochemotherapy on the metastatic potential of tumour cells is not known. Prevention of metastasis is an important aspect of successful treatment; however, it is known that metastasis can be induced by different treatment modalities. Therefore, the aim of this study was to evaluate the effect of electrochemotherapy with cisplatin on the metastatic potential of human malignant melanoma cells. Cells treated by electrochemotherapy with cisplatin were tested for their ability to migrate and invade through Matrigel-coated porous membrane. In addition, RNA was isolated from cells after treatment and differentially expressed genes were investigated by microarray analysis to evaluate the effect of electrochemotherapy with cisplatin on gene expression. There were no significant changes observed in cell migration and invasion of melanoma cells after electrochemotherapy. In addition, there were no changes observed in cell adhesion on Matrigel. Gene expression analysis showed that a very low number of genes were differentially expressed after electrochemotherapy with cisplatin. Two genes, LAMB3 and CD63 involved in cell migration, were both downregulated after electrochemotherapy with cisplatin and the expression of metastasis promoting genes was not increased after electrochemotherapy. Our data suggest that electrochemotherapy does not increase the metastatic behaviour of human melanoma cells.

Focal giant cell cardiomyopathy with Beckwith-Wiedemann syndrome.

PubMed

Kapur, S; Kuehl, K S; Midgely, F M; Chandra, R S

1985-01-01

Cardiac involvement in Beckwith-Wiedemann syndrome is mostly limited to mild cardiomegaly. Although these patients have visceromegaly, macroglossia, gigantism, and adrenal cytomegaly, no significant myocardial changes have been described. An infant with dysmorphic features of this syndrome had supraventricular tachycardia since birth. Nodular lesions were present in the right atrium. Morphologically these lesions were composed of hypertrophic myocardial fibers admixed with multinucleated giant cells of myogenic origin. The exact nature of these lesions remains undetermined. It is postulated that hypertrophic myocardial cells may represent cardiac cytomegaly as a manifestation of the accelerated growth potential of cells seen with this syndrome.

Low Doses of Curcuma longa Modulates Cell Migration and Cell-Cell Adhesion.

PubMed

de Campos, Paloma Santos; Matte, Bibiana Franzen; Diel, Leonardo Francisco; Jesus, Luciano Henrique; Bernardi, Lisiane; Alves, Alessandro Menna; Rados, Pantelis Varvaki; Lamers, Marcelo Lazzaron

2017-09-01

Cell invasion and metastasis are involved in clinical failures in cancer treatment, and both events require the acquisition of a migratory behavior by tumor cells. Curcumin is a promising natural product with anti-proliferative activity, but its effects on cell migration are still unclear. We evaluated the effects of curcumin on the proliferation, apoptosis, migration, and cell-cell adhesion of keratinocyte, oral squamous cell carcinoma (OSCC), and fibroblast cell lines, as well as in a xenograft model of OSCC. Curcumin (2 μM) decreased cell proliferation in cell lines with mesenchymal characteristics, while cell death was detected only at 50 μM. We observed that highly migratory cells showed a decrease on migration speed and directionality when treated with 2 or 5 μM of curcumin (50% and 40%, respectively, p < 0.05). Using spheroids, we observed that curcumin dose dependently decreased cell-cell adhesion, especially on tumor-derived spheroids. Also, in a xenograft model with patient-derived OSCC cells, the administration of curcumin decreased tumor growth and aggressiveness when compared with untreated tumors, indicating the potential antitumor effect in oral cancer. These results suggest that lower doses of curcumin can influence several steps involved in tumorigenesis, including migration properties, suggesting a possible use in cancer therapy. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

The emotional cerebellum.

PubMed

Strata, Piergiorgio

2015-10-01

Great attention has been given so far to cerebellar control of posture and of skilled movements despite the well-demonstrated interconnections between the cerebellum and the autonomic nervous system. Here is a review of the link between these two structures and a report on the recently acquired evidence for its involvement in the world of emotions. In rodents, the reversible inactivation of the vermis during the consolidation or the reconsolidation period hampers the retention of the fear memory trace. In this region, there is a long-term potentiation of both the excitatory synapses between the parallel fibres and the Purkinje cells and of the feed-forward inhibition mediated by molecular layer interneurons. This concomitant potentiation ensures the temporal fidelity of the system. Additional contacts between mossy fibre terminals and Golgi cells provide morphological evidence of the potentiation of another feed-forward inhibition in the granular layer. Imaging experiments show that also in humans the cerebellum is activated during mental recall of emotional personal episodes and during learning of a conditioned or unconditioned association involving emotions. The vermis participates in fear learning and memory mechanisms related to the expression of autonomic and motor responses of emotions. In humans, the cerebellar hemispheres are also involved at a higher emotional level. The importance of these findings is evident when considering the cerebellar malfunctioning in psychiatric diseases like autism and schizophrenia which are characterized behaviourally by emotion processing impairments.

Critical role of zinc finger protein 521 in the control of growth, clonogenicity and tumorigenic potential of medulloblastoma cells

PubMed Central

Iaccino, Enrico; Scicchitano, Stefania; Lupia, Michela; Chiarella, Emanuela; Mega, Tiziana; Bernaudo, Francesca; Pelaggi, Daniela; Mesuraca, Maria; Pazzaglia, Simonetta; Semenkow, Samantha; Bar, Eli E.; Kool, Marcel; Pfister, Stefan; Bond, Heather M.; Eberhart, Charles G.; Steinkühler, Christian; Morrone, Giovanni

2013-01-01

The stem cell-associated transcription co-factor ZNF521 has been implicated in the control of hematopoietic, osteo-adipogenic and neural progenitor cells. ZNF521 is highly expressed in cerebellum and in particular in the neonatal external granule layer that contains candidate medulloblastoma cells-of-origin, and in the majority of human medulloblastomas. Here we have explored its involvement in the control of human and murine medulloblastoma cells. The effect of ZNF521 on growth and tumorigenic potential of human medulloblastoma cell lines as well as primary Ptc1−/+ mouse medulloblastoma cells was investigated in a variety of in vitro and in vivo assays, by modulating its expression using lentiviral vectors carrying the ZNF521 cDNA, or shRNAs that silence its expression. Enforced overexpression of ZNF521 in DAOY medulloblastoma cells significantly increased their proliferation, growth as spheroids and ability to generate clones in single-cell cultures and semisolid media, and enhanced their migratory ability in wound-healing assays. Importantly, ZNF521-expressing cells displayed a greatly enhanced tumorigenic potential in nude mice. All these activities required the ZNF521 N-terminal motif that recruits the nucleosome remodeling and histone deacetylase complex, which might therefore represent an appealing therapeutic target. Conversely, silencing of ZNF521 in human UW228 medulloblastoma cells that display high baseline expression decreased their proliferation, clonogenicity, sphere formation and wound-healing ability. Similarly, Zfp521 silencing in mouse Ptc1−/+ medulloblastoma cells drastically reduced their growth and tumorigenic potential. Our data strongly support the notion that ZNF521, through the recruitment of the NuRD complex, contributes to the clonogenic growth, migration and tumorigenicity of medulloblastoma cells. PMID:23907569

Critical role of zinc finger protein 521 in the control of growth, clonogenicity and tumorigenic potential of medulloblastoma cells.

PubMed

Spina, Raffaella; Filocamo, Gessica; Iaccino, Enrico; Scicchitano, Stefania; Lupia, Michela; Chiarella, Emanuela; Mega, Tiziana; Bernaudo, Francesca; Pelaggi, Daniela; Mesuraca, Maria; Pazzaglia, Simonetta; Semenkow, Samantha; Bar, Eli E; Kool, Marcel; Pfister, Stefan; Bond, Heather M; Eberhart, Charles G; Steinkühler, Christian; Morrone, Giovanni

2013-08-01

The stem cell-associated transcription co-factor ZNF521 has been implicated in the control of hematopoietic, osteo-adipogenic and neural progenitor cells. ZNF521 is highly expressed in cerebellum and in particular in the neonatal external granule layer that contains candidate medulloblastoma cells-of-origin, and in the majority of human medulloblastomas. Here we have explored its involvement in the control of human and murine medulloblastoma cells. The effect of ZNF521 on growth and tumorigenic potential of human medulloblastoma cell lines as well as primary Ptc1-/+ mouse medulloblastoma cells was investigated in a variety of in vitro and in vivo assays, by modulating its expression using lentiviral vectors carrying the ZNF521 cDNA, or shRNAs that silence its expression. Enforced overexpression of ZNF521 in DAOY medulloblastoma cells significantly increased their proliferation, growth as spheroids and ability to generate clones in single-cell cultures and semisolid media, and enhanced their migratory ability in wound-healing assays. Importantly, ZNF521-expressing cells displayed a greatly enhanced tumorigenic potential in nude mice. All these activities required the ZNF521 N-terminal motif that recruits the nucleosome remodeling and histone deacetylase complex, which might therefore represent an appealing therapeutic target. Conversely, silencing of ZNF521 in human UW228 medulloblastoma cells that display high baseline expression decreased their proliferation, clonogenicity, sphere formation and wound-healing ability. Similarly, Zfp521 silencing in mouse Ptc1-/+ medulloblastoma cells drastically reduced their growth and tumorigenic potential. Our data strongly support the notion that ZNF521, through the recruitment of the NuRD complex, contributes to the clonogenic growth, migration and tumorigenicity of medulloblastoma cells.

DIOXIN 2003 BOSTON

EPA Science Inventory

DIOXON 2003 BOSTON
SESSION SUMMARY REPORT

Toxicology of BFRs

This oral session is comprised of seven presentations dealing with the potential health effects of BFRs. Talks involve both in vivo and in vitro studies in mice, rats, and cultured cells, and are from...

Cancer-derived extracellular vesicles: friend and foe of tumour immunosurveillance.

PubMed

Dörsam, Bastian; Reiners, Kathrin S; von Strandmann, Elke Pogge

2018-01-05

Extracellular vesicles (EVs) are important players of intercellular signalling mechanisms, including communication with and among immune cells. EVs can affect the surrounding tissue as well as peripheral cells. Recently, EVs have been identified to be involved in the aetiology of several diseases, including cancer. Tumour cell-released EVs or exosomes have been shown to promote a tumour-supporting environment in non-malignant tissue and, thus, benefit metastasis. The underlying mechanisms are numerous: loss of antigen expression, direct suppression of immune effector cells, exchange of nucleic acids, alteration of the recipient cells' transcription and direct suppression of immune cells. Consequently, tumour cells can subvert the host's immune detection as well as suppress the immune system. On the contrary, recent studies reported the existence of EVs able to activate immune cells, thus promoting the tumour-directed immune response. In this article, the immunosuppressive capabilities of EVs, on the one hand, and their potential use in immunoactivation and therapeutic potential, on the other hand, are discussed.This article is part of the discussion meeting issue 'Extracellular vesicles and the tumour microenvironment'. © 2017 The Authors.

A gene involved in control of human cellular senescence on human chromosome 1q

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

Hensler, P.J.; Pereira-Smith, O.M.; Annab, L.A.

1994-04-01

Normal cells in culture exhibit limited division potential and have been used as a model for cellular senescence. In contrast, tumor-derived or carcinogen- or virus-transformed cells are capable of indefinite division. Fusion of normal human diploid fibroblasts with immortal human cells yielded hybrids having limited life spans, indicating that cellular senescence was dominant. Fusions of various immortal human cell lines with each other led to the identification of four complementation groups for indefinite division. The purpose of this study was to determine whether human chromosome 1 could complement the recessive immortal defect of human cell lines assigned to one ofmore » the four complementation groups. Using microcell fusion, the authors introduced a single normal human chromosome 1 into immortal human cell lines representing the complementation groups and determined that it caused loss of proliferative potential of an osteosarcoma-derived cell line (TE85), a cytomegalovirus-transformed lung fibroblast cell line (CMV-Mj-HEL-1), and a Ki-ras[sup +]-transformed derivative of TE85 (143B TK[sup [minus

Activation of antioxidant pathways in ras-mediated oncogenic transformation of human surface ovarian epithelial cells revealed by functional proteomics and mass spectrometry.

PubMed

Young, Travis W; Mei, Fang C; Yang, Gong; Thompson-Lanza, Jennifer A; Liu, Jinsong; Cheng, Xiaodong

2004-07-01

Cellular transformation is a complex process involving genetic alterations associated with multiple signaling pathways. Development of a transformation model using defined genetic elements has provided an opportunity to elucidate the role of oncogenes and tumor suppressor genes in the initiation and development of ovarian cancer. To study the cellular and molecular mechanisms of Ras-mediated oncogenic transformation of ovarian epithelial cells, we used a proteomic approach involving two-dimensional electrophoresis and mass spectrometry to profile two ovarian epithelial cell lines, one immortalized with SV40 T/t antigens and the human catalytic subunit of telomerase and the other transformed with an additional oncogenic ras(V12) allele. Of approximately 2200 observed protein spots, we have identified >30 protein targets that showed significant changes between the immortalized and transformed cell lines using peptide mass fingerprinting. Among these identified targets, one most notable group of proteins altered significantly consists of enzymes involved in cellular redox balance. Detailed analysis of these protein targets suggests that activation of Ras-signaling pathways increases the threshold of reactive oxidative species (ROS) tolerance by up-regulating the overall antioxidant capacity of cells, especially in mitochondria. This enhanced antioxidant capacity protects the transformed cells from high levels of ROS associated with the uncontrolled growth potential of tumor cells. It is conceivable that an enhanced antioxidation capability may constitute a common mechanism for tumor cells to evade apoptosis induced by oxidative stresses at high ROS levels.

Potential involvement of F0F1-ATP(synth)ase and reactive oxygen species in apoptosis induction by the antineoplastic agent erucylphosphohomocholine in glioblastoma cell lines : a mechanism for induction of apoptosis via the 18 kDa mitochondrial translocator protein.

PubMed

Veenman, Leo; Alten, Julia; Linnemannstöns, Karen; Shandalov, Yulia; Zeno, Sivan; Lakomek, Max; Gavish, Moshe; Kugler, Wilfried

2010-07-01

Erucylphosphohomocholine (ErPC3, Erufosine) was reported previously to induce apoptosis in otherwise highly apoptosis-resistant malignant glioma cell lines while sparing their non-tumorigenic counterparts. We also previously found that the mitochondrial 18 kDa Translocator Protein (TSPO) is required for apoptosis induction by ErPC3. These previous studies also suggested involvement of reactive oxygen species (ROS). In the present study we further investigated the potential involvement of ROS generation, the participation of the mitochondrial respiration chain, and the role of the mitochondrial F(O)F(1)-ATP(synth)ase in the pro-apoptotic effects of ErPC3 on U87MG and U118MG human glioblastoma cell lines. For this purpose, cells were treated with the ROS chelator butylated hydroxyanisole (BHA), the mitochondrial respiration chain inhibitors rotenone, antimycin A, myxothiazol, and the uncoupler CCCP. Also oligomycin and piceatannol were studied as inhibitors of the F(O) and F(1) subunits of the mitochondrial F(O)F(1)-ATP(synth)ase, respectively. BHA was able to attenuate apoptosis induction by ErPC3, including mitochondrial ROS generation as determined with cardiolipin oxidation, as well as collapse of the mitochondrial membrane potential (Deltapsi(m)). Similarly, we found that oligomycin attenuated apoptosis and collapse of the Deltapsi(m), normally induced by ErPC3, including the accompanying reductions in cellular ATP levels. Other inhibitors of the mitochondrial respiration chain, as well as piceatannol, did not show such effects. Consequently, our findings strongly point to a role for the F(O) subunit of the mitochondrial F(O)F(1)-ATP(synth)ase in ErPC3-induced apoptosis and dissipation of Deltapsi(m) as well as ROS generation by ErPC3 and TSPO.

Toxicity of the mycotoxin fumonisin B1 on the insect Sf9 cell line.

PubMed

Zhang, He; Zhang, Liyang; Diao, Xue; Li, Na; Liu, Chenglan

2017-04-01

Fumonisins are a type of mycotoxin produced by Fusarium spp., mainly F. proliferatum and F. vertieilliodes, and represent a potential hazard to the health of animals and human beings. The toxicity and mechanism of action of fumonisins is ambiguous, and it is unclear whether fumonisins are toxic to insect cells. This study examines the toxicity of fumonisin B 1 (FB 1 ) and its mechanism of action in the Spodoptera frugiperda Sf9 cell line. We found that FB 1 inhibited Sf9 cellular proliferation and arrested cell growth at the G 2 /M phase. Morphological observation showed that FB 1 induced swelling, vacuole formation, and loss of adhesion in Sf9 cells. Flow cytometry analysis showed that FB 1 caused depolarization of the cell membrane potential and hyperpolarization of the mitochondrial membrane potential. To uncover potential genes associated with the molecular mechanisms of FB 1 , 41 differentially expressed genes were identified by transcriptome analyses after FB 1 treatment. These genes are putatively involved in detoxification metabolism, insect hormone regulation, cell apoptosis, and other related processes. Finally, six differentially expressed genes were chosen and validated by quantitative real-time PCR (QRT-PCR). Our test could provide a reference for other kinds of insect cells studies on FB 1 stress. At the same time, our studies try to provide a possible for FB 1 as a precursor compounds of biological insecticide. Copyright © 2017 Elsevier Ltd. All rights reserved.

ROCK Inhibition Extends Passage of Pluripotent Stem Cell-Derived Retinal Pigmented Epithelium

PubMed Central

Croze, Roxanne H.; Buchholz, David E.; Radeke, Monte J.; Thi, William J.; Hu, Qirui; Coffey, Peter J.

2014-01-01

Human embryonic stem cells (hESCs) offer a potentially unlimited supply of cells for emerging cell-based therapies. Unfortunately, the process of deriving distinct cell types can be time consuming and expensive. In the developed world, age-related macular degeneration (AMD) is the leading cause of blindness in the elderly, with more than 7.2 million people afflicted in the U.S. alone. Both hESC-derived retinal pigmented epithelium (hESC-RPE) and induced pluripotent stem cell-derived RPE (iPSC-RPE) are being developed for AMD therapies by multiple groups, but their potential for expansion in culture is limited. To attempt to overcome this passage limitation, we examined the involvement of Rho-associated, coiled-coil protein kinase (ROCK) in hESC-RPE and iPSC-RPE culture. We report that inhibiting ROCK1/2 with Y-27632 allows extended passage of hESC-RPE and iPSC-RPE. Microarray analysis suggests that ROCK inhibition could be suppressing an epithelial-to-mesenchymal transition through various pathways. These include inhibition of key ligands of the transforming growth factor-β pathway (TGFB1 and GDF6) and Wnt signaling. Two important processes are affected, allowing for an increase in hESC-RPE expansion. First, ROCK inhibition promotes proliferation by inducing multiple components that are involved in cell cycle progression. Second, ROCK inhibition affects many pathways that could be converging to suppress RPE-to-mesenchymal transition. This allows hESC-RPE to remain functional for an extended but finite period in culture. PMID:25069775

Differentiation of endosperm transfer cells of barley: a comprehensive analysis at the micro-scale.

PubMed

Thiel, Johannes; Riewe, David; Rutten, Twan; Melzer, Michael; Friedel, Swetlana; Bollenbeck, Felix; Weschke, Winfriede; Weber, Hans

2012-08-01

Barley endosperm cells differentiate into transfer cells (ETCs) opposite the nucellar projection. To comprehensively analyse ETC differentiation, laser microdissection-based transcript and metabolite profiles were obtained from laser microdissected tissues and cell morphology was analysed. Flange-like secondary-wall ingrowths appeared between 5 and 7 days after pollination within the three outermost cell layers. Gene expression analysis indicated that ethylene-signalling pathways initiate ETC morphology. This is accompanied by gene activity related to cell shape control and vesicle transport, with abundant mitochondria and endomembrane structures. Gene expression analyses indicate predominant formation of hemicelluloses, glucuronoxylans and arabinoxylans, and transient formation of callose, together with proline and 4-hydroxyproline biosynthesis. Activation of the methylation cycle is probably required for biosynthesis of phospholipids, pectins and ethylene. Membrane microdomains involving sterols/sphingolipids and remorins are potentially involved in ETC development. The transcriptional activity of assimilate and micronutrient transporters suggests ETCs as the main uptake organs of solutes into the endosperm. Accordingly, the endosperm grows maximally after ETCs are fully developed. Up-regulated gene expression related to amino acid catabolism, C:N balances, carbohydrate oxidation, mitochondrial activity and starch degradation meets high demands for respiratory energy and carbohydrates, required for cell proliferation and wall synthesis. At 10 days after pollination, ETCs undergo further differentiation, potentially initiated by abscisic acid, and metabolism is reprogrammed as shown by activated storage and stress-related processes. Overall, the data provide a comprehensive view of barley ETC differentiation and development, and identify candidate genes and associated pathways. © 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.

ROCK Inhibition Extends Passage of Pluripotent Stem Cell-Derived Retinal Pigmented Epithelium.

PubMed

Croze, Roxanne H; Buchholz, David E; Radeke, Monte J; Thi, William J; Hu, Qirui; Coffey, Peter J; Clegg, Dennis O

2014-09-01

Human embryonic stem cells (hESCs) offer a potentially unlimited supply of cells for emerging cell-based therapies. Unfortunately, the process of deriving distinct cell types can be time consuming and expensive. In the developed world, age-related macular degeneration (AMD) is the leading cause of blindness in the elderly, with more than 7.2 million people afflicted in the U.S. alone. Both hESC-derived retinal pigmented epithelium (hESC-RPE) and induced pluripotent stem cell-derived RPE (iPSC-RPE) are being developed for AMD therapies by multiple groups, but their potential for expansion in culture is limited. To attempt to overcome this passage limitation, we examined the involvement of Rho-associated, coiled-coil protein kinase (ROCK) in hESC-RPE and iPSC-RPE culture. We report that inhibiting ROCK1/2 with Y-27632 allows extended passage of hESC-RPE and iPSC-RPE. Microarray analysis suggests that ROCK inhibition could be suppressing an epithelial-to-mesenchymal transition through various pathways. These include inhibition of key ligands of the transforming growth factor-β pathway (TGFB1 and GDF6) and Wnt signaling. Two important processes are affected, allowing for an increase in hESC-RPE expansion. First, ROCK inhibition promotes proliferation by inducing multiple components that are involved in cell cycle progression. Second, ROCK inhibition affects many pathways that could be converging to suppress RPE-to-mesenchymal transition. This allows hESC-RPE to remain functional for an extended but finite period in culture. ©AlphaMed Press.

Galactosylated polyaspartamide copolymers for siRNA targeted delivery to hepatocellular carcinoma cells.

PubMed

Cavallaro, Gennara; Farra, Rossella; Craparo, Emanuela Fabiola; Sardo, Carla; Porsio, Barbara; Giammona, Gaetano; Perrone, Francesca; Grassi, Mario; Pozzato, Gabriele; Grassi, Gabriele; Dapas, Barbara

2017-06-20

The limited efficacy of available treatments for hepatocellular carcinoma (HCC) requires the development of novel therapeutic approaches. We synthesized a novel cationic polymer based on α,β-poly-(N-2-hydroxyethyl)-d,L-aspartamide (PHEA) for drug delivery to HCC cells. The copolymer was synthesized by subsequent derivatization of PHEA with diethylene triamine (DETA) and with a polyethylene glycol (PEG) derivative bearing galactose (GAL) molecules, obtaining the cationic derivative PHEA-DETA-PEG-GAL. PHEA-DETA-PEG-GAL has suitable chemical-physical characteristics for a potential systemic use and can effectively deliver a siRNA (siE2F1) targeted against the transcription factor E2F1, a gene product involved in HCC. The presence of GAL residues in the polyplexes allows the targeting of HCC cells that express the asialo-glycoprotein receptor (ASGP-R). In these cells, but not in ASGP-R non-expressing cells, PHEA-DETA-PEG-GAL/siE2F1 polyplexes induce the reduction of the mRNA and protein levels of E2F1 and of E2F1-regulated genes, all involved in the promotion of the G1/S phase transition. This results in a decrease of cell proliferation with a G1/G0 phase cells accumulation. Notably, removal of GAL residue almost completely abrogates the targeting capacity of the developed polyplexes. In conclusion, the generated polyplexes demonstrate the potential to effectively contributing to the development of novel anti-HCC therapeutic approaches via a siRNA-targeted delivery. Copyright © 2017 Elsevier B.V. All rights reserved.

Phenolic extract from oleaster (Olea europaea var. Sylvestris) leaves reduces colon cancer growth and induces caspase-dependent apoptosis in colon cancer cells via the mitochondrial apoptotic pathway.

PubMed

Zeriouh, Wafa; Nani, Abdelhafid; Belarbi, Meriem; Dumont, Adélie; de Rosny, Charlotte; Aboura, Ikram; Ghanemi, Fatima Zahra; Murtaza, Babar; Patoli, Danish; Thomas, Charles; Apetoh, Lionel; Rébé, Cédric; Delmas, Dominique; Khan, Naim Akhtar; Ghiringhelli, François; Rialland, Mickael; Hichami, Aziz

2017-01-01

Dietary polyphenols, derived from natural products, have received a great interest for their chemopreventive properties against cancer. In this study, we investigated the effects of phenolic extract of the oleaster leaves (PEOL) on tumor growth in mouse model and on cell death in colon cancer cell lines. We assessed the effect of oleaster leaf infusion on HCT116 (human colon cancer cell line) xenograft growth in athymic nude mice. We observed that oleaster leaf polyphenol-rich infusion limited HCT116 tumor growth in vivo. Investigations of PEOL on two human CRC cell lines showed that PEOL induced apoptosis in HCT116 and HCT8 cells. We demonstrated an activation of caspase-3, -7 and -9 by PEOL and that pre-treatment with the pan-caspase inhibitor, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (z-VAD-fmk), prevented PEOL-induced cell death. We observed an involvement of the mitochondrial pathway in PEOL-induced apoptosis evidenced by reactive oxygen species (ROS) production, a decrease of mitochondrial membrane potential, and cytochrome c release. Increase in intracellular Ca2+ concentration induced by PEOL represents the early event involved in mitochondrial dysfunction, ROS-induced endoplasmic reticulum (ER) stress and apoptosis induced by PEOL, as ruthenium red, an inhibitor of mitochondrial calcium uptake inhibited apoptotic effect of PEOL, BAPTA/AM inhibited PEOL-induced ROS generation and finally, N-acetyl-L-cysteine reversed ER stress and apoptotic effect of PEOL. These results demonstrate that polyphenols from oleaster leaves might have a strong potential as chemopreventive agent in colorectal cancer.

G protein-coupled receptor kinase 2 positively regulates epithelial cell migration

PubMed Central

Penela, Petronila; Ribas, Catalina; Aymerich, Ivette; Eijkelkamp, Niels; Barreiro, Olga; Heijnen, Cobi J; Kavelaars, Annemieke; Sánchez-Madrid, Francisco; Mayor, Federico

2008-01-01

Cell migration requires integration of signals arising from both the extracellular matrix and messengers acting through G protein-coupled receptors (GPCRs). We find that increased levels of G protein-coupled receptor kinase 2 (GRK2), a key player in GPCR regulation, potentiate migration of epithelial cells towards fibronectin, whereas such process is decreased in embryonic fibroblasts from hemizygous GRK2 mice or upon knockdown of GRK2 expression. Interestingly, the GRK2 effect on fibronectin-mediated cell migration involves the paracrine/autocrine activation of a sphingosine-1-phosphate (S1P) Gi-coupled GPCR. GRK2 positively modulates the activity of the Rac/PAK/MEK/ERK pathway in response to adhesion and S1P by a mechanism involving the phosphorylation-dependent, dynamic interaction of GRK2 with GIT1, a key scaffolding protein in cell migration processes. Furthermore, decreased GRK2 levels in hemizygous mice result in delayed wound healing rate in vivo, consistent with a physiological role of GRK2 as a regulator of coordinated integrin and GPCR-directed epithelial cell migration. PMID:18369319

Targeting Metastasis with Snake Toxins: Molecular Mechanisms

PubMed Central

Urra, Félix A.

2017-01-01

Metastasis involves the migration of cancer cells from a primary tumor to invade and establish secondary tumors in distant organs, and it is the main cause for cancer-related deaths. Currently, the conventional cytostatic drugs target the proliferation of malignant cells, being ineffective in metastatic disease. This highlights the need to find new anti-metastatic drugs. Toxins isolated from snake venoms are a natural source of potentially useful molecular scaffolds to obtain agents with anti-migratory and anti-invasive effects in cancer cells. While there is greater evidence concerning the mechanisms of cell death induction of several snake toxin classes on cancer cells; only a reduced number of toxin classes have been reported (i.e., disintegrins/disintegrin-like proteins, C-type lectin-like proteins, C-type lectins, serinproteases, cardiotoxins, snake venom cystatins) as inhibitors of adhesion, migration, and invasion of cancer cells. Here, we discuss the anti-metastatic mechanisms of snake toxins, distinguishing three targets, which involve (1) inhibition of extracellular matrix components-dependent adhesion and migration, (2) inhibition of epithelial-mesenchymal transition, and (3) inhibition of migration by alterations in the actin/cytoskeleton network. PMID:29189742

Relevance of Endoplasmic Reticulum Stress Cell Signaling in Liver Cold Ischemia Reperfusion Injury

PubMed Central

Folch-Puy, Emma; Panisello, Arnau; Oliva, Joan; Lopez, Alexandre; Castro Benítez, Carlos; Adam, René; Roselló-Catafau, Joan

2016-01-01

The endoplasmic reticulum (ER) is involved in calcium homeostasis, protein folding and lipid biosynthesis. Perturbations in its normal functions lead to a condition called endoplasmic reticulum stress (ERS). This can be triggered by many physiopathological conditions such as alcoholic steatohepatitis, insulin resistance or ischemia-reperfusion injury. The cell reacts to ERS by initiating a defensive process known as the unfolded protein response (UPR), which comprises cellular mechanisms for adaptation and the safeguarding of cell survival or, in cases of excessively severe stress, for the initiation of the cell death program. Recent experimental data suggest the involvement of ERS in ischemia/reperfusion injury (IRI) of the liver graft, which has been considered as one of major problems influencing outcome after liver transplantation. The purpose of this review is to summarize updated data on the molecular mechanisms of ERS/UPR and the consequences of this pathology, focusing specifically on solid organ preservation and liver transplantation models. We will also discuss the potential role of ERS, beyond the simple adaptive response and the regulation of cell death, in the modification of cell functional properties and phenotypic changes. PMID:27231901

Redox signaling regulated by an electrophilic cyclic nucleotide and reactive cysteine persulfides.

PubMed

Fujii, Shigemoto; Sawa, Tomohiro; Nishida, Motohiro; Ihara, Hideshi; Ida, Tomoaki; Motohashi, Hozumi; Akaike, Takaaki

2016-04-01

Reactive oxygen (oxidant) and free radical species are known to cause nonspecific damage of various biological molecules. The oxidant toxicology is developing an emerging concept of the physiological functions of reactive oxygen species in cell signaling regulation. Redox signaling is precisely modulated by endogenous electrophilic substances that are generated from reactive oxygen species during cellular oxidative stress responses. Among diverse electrophilic molecular species that are endogenously generated, 8-nitroguanosine 3',5'-cyclic monophosphate (8-nitro-cGMP) is a unique second messenger whose formation, signaling, and metabolism in cells was recently clarified. Most important, our current studies revealed that reactive cysteine persulfides that are formed abundantly in cells are critically involved in the metabolism of 8-nitro-cGMP. Modern redox biology involves frontiers of cell research and stem cell research; medical and clinical investigations of infections, cancer, metabolic syndrome, aging, and neurodegenerative diseases; and other fields. 8-Nitro-cGMP-mediated signaling and metabolism in cells may therefore be potential targets for drug development, which may lead to discovery of new therapeutic agents for many diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

Environmental, health, and safety issues of sodium-sulfur batteries for electric and hybrid vehicles. Volume 1, Cell and battery safety

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

Ohi, J M

1992-09-01

This report is the first of four volumes that identify and assess the environmental, health, and safety issues involved in using sodium-sulfur (Na/S) battery technology as the energy source in electric and hybrid vehicles that may affect the commercialization of Na/S batteries. This and the other reports on recycling, shipping, and vehicle safety are intended to help the Electric and Hybrid Propulsion Division of the Office of Transportation Technologies in the US Department of Energy (DOE/EHP) determine the direction of its research, development, and demonstration (RD&D) program for Na/S battery technology. The reports review the status of Na/S battery RD&Dmore » and identify potential hazards and risks that may require additional research or that may affect the design and use of Na/S batteries. This volume covers cell design and engineering as the basis of safety for Na/S batteries and describes and assesses the potential chemical, electrical, and thermal hazards and risks of Na/S cells and batteries as well as the RD&D performed, under way, or to address these hazards and risks. The report is based on a review of the literature and on discussions with experts at DOE, national laboratories and agencies, universities, and private industry. Subsequent volumes will address environmental, health, and safety issues involved in shipping cells and batteries, using batteries to propel electric vehicles, and recycling and disposing of spent batteries. The remainder of this volume is divided into two major sections on safety at the cell and battery levels. The section on Na/S cells describes major component and potential failure modes, design, life testing and failure testing, thermal cycling, and the safety status of Na/S cells. The section on batteries describes battery design, testing, and safety status. Additional EH&S information on Na/S batteries is provided in the appendices.« less

The graphic cell method: a new look at digitizing geologic maps

USGS Publications Warehouse

Hanley, J.T.

1982-01-01

The graphic cell method is an alternative method of digitizing areal geologic information. It involves a discrete-point sampling scheme in which the computer establishes a matrix of cells over the map. Each cell and the whole cell is assigned the identity or value of the geologic information that is recognized at its center. Cell size may be changed to suit the needs of the user. The computer program resolves the matrix and identifies potential errors such as multiple assignments. Input includes the digitized boundaries of each geologic formation. This method should eliminate a primary bottleneck in the creation and testing of geomathematical models in such disciplines as resource appraisal. ?? 1982.

Role of estrogen receptors alpha, beta and GPER1/GPR30 in pancreatic beta-cells.

PubMed

Nadal, Angel; Alonso-Magdalena, Paloma; Soriano, Sergi; Ripoll, Cristina; Fuentes, Esther; Quesada, Ivan; Ropero, Ana Belen

2011-01-01

Estrogen receptors (ER) are emerging as important molecules involved in the adaptation of beta-cells to insulin resistance. The onset of type 2 diabetes is marked by insulin secretory dysfunction and decreased beta-cell mass. During pregnancy, puberty and obesity there is increased metabolic demand and insulin resistance is developed. This metabolic state increases the demand on beta-cells to augment insulin biosynthesis and release. In this respect, ERalpha is directly implicated in the E2-regulation of insulin content and secretion, while ERbeta is in the E2-potentiation of glucose-induced insulin release. Both receptors develop their actions within the physiological range of E2. In addition, the G protein-coupled estrogen receptor (GPER1/GPR30) seems to be implicated in the E2-regulation of stimulus-secretion coupling in the three cell types of the islet. The increased demand of insulin production for long time may lead to beta-cell stress and apoptosis. ERalpha, ERbeta and GPER1/GPR30 are involved in preventing beta-cell apoptosis, impeding the loss of critical beta-cell mass. Therefore, estrogen receptors may play an essential role in the adaptation of the pancreas to insulin resistant periods.

KHYG-1 and NK-92 represent different subtypes of LFA-1-mediated NK cell adhesiveness.

PubMed

Suck, Garnet; Tan, Suet-Mien; Chu, Sixian; Niam, Madelaine; Vararattanavech, Ardcharaporn; Lim, Tsyr Jong; Koh, Mickey B C

2011-01-01

Novel cancer cellular therapy approaches involving long-term ex vivo IL-2 stimulated highly cytotoxic natural killer (NK) cells are emerging. However, adhesion properties of such NK cells are not very well understood. Herein, we describe the novel observation of permanently activated alphaLbeta2 integrin leukocyte function-associated antigen (LFA)-1 adhesion receptor in long-term IL-2 activated NK cells and the permanent NK cell lines KHYG-1 and NK-92. We show that such cytokine activated NK effectors constitutively adhered to the LFA-1-ligand ICAM-1, whereas binding to the lower affinity ligand ICAM-3 required additional exogenous activating conditions. The results demonstrate an extended conformation and an intermediate affinity state for the LFA-1 population expressed by the NK cells. Interestingly, adhesion to ICAM-1 or K562 induced pronounced cell spreading in KHYG-1, but not in NK-92, and partially in long-term IL-2 stimulated primary NK cells. It is conceivable that such differential adhesion characteristics may impact motility potential of such NK effectors with relevance to clinical tumor targeting. KHYG-1 could be a useful model in planning future targeted therapeutic approaches involving NK effectors with augmented functions.

CO2 laser increases the regenerative capacity of human adipose-derived stem cells by a mechanism involving the redox state and enhanced secretion of pro-angiogenic molecules.

PubMed

Constantin, Alina; Dumitrescu, Madalina; Mihai Corotchi, Maria Cristina; Jianu, Dana; Simionescu, Maya

2017-01-01

CO 2 laser has a beneficial effect on stem cells by mechanisms that are not clearly elucidated. We hypothesize that the effect of fractional CO 2 laser on human adipose-derived stem cells (ADSC) could be due to changes in redox homeostasis and secretion of factors contributing to cellular proliferation and angiogenic potential. ADSC incubated in medium containing 0.5 or 10 % FBS were exposed to a single irradiation of a 10,600-nm fractional CO 2 laser; non-irradiated ADSC were used as control. Viability/proliferation of ADSC was assessed by MTT assay; the intracellular reactive oxygen species (ROS) levels and the mitochondrial membrane potential (∆Ψ m ) were determined with DCFH-DA and JC-1 fluorescent probes, respectively. Molecules secreted by ADSC in the medium were determined by ELISA assay, and their capacity to support endothelial tube-like formation by the Matrigel assay. The results showed that compared to controls, ADSC kept in low FBS medium and irradiated with CO 2 laser at 9 W exhibited: (a) increased proliferation (∼20 %), (b) transient increase of mitochondrial ROS and the capacity to restore Δψ m after rotenone induced depolarization, and (c) augmented secretion in the conditioned medium of MMP-2 (twofold), MMP-9 (eightfold), VEGF (twofold), and adiponectin (∼50 %) that have the capacity to support angiogenesis of endothelial progenitor cells. In conclusion, the mechanisms underlying the benefic effect of CO 2 laser on ADSC are the activation of the redox pathways which increases cell proliferation and enhances secretion of angiogenic molecules. These results explain, in part, the mechanisms involved in the increased regenerative potential of CO 2 laser-exposed ADSC that could be exploited for clinical applications.

MicroRNA and receptor mediated signaling pathways as potential therapeutic targets in heart failure.

PubMed

Tuttolomondo, Antonino; Simonetta, Irene; Pinto, Antonio

2016-11-01

Cardiac remodelling is a complex pathogenetic pathway involving genome expression, molecular, cellular, and interstitial changes that cause changes in size, shape and function of the heart after cardiac injury. Areas covered: We will review recent advances in understanding the role of several receptor-mediated signaling pathways and micro-RNAs, in addition to their potential as candidate target pathways in the pathogenesis of heart failure. The myocyte is the main target cell involved in the remodelling process via ischemia, cell necrosis and apoptosis (by means of various receptor pathways), and other mechanisms mediated by micro-RNAs. We will analyze the role of some receptor mediated signaling pathways such as natriuretic peptides, mediators of glycogen synthase kinase 3 and ERK1/2 pathways, beta-adrenergic receptor subtypes and relaxin receptor signaling mechanisms, TNF/TNF receptor family and TWEAK/Fn14 axis, and some micro-RNAs as candidate target pathways in pathogenesis of heart failure. These mediators of receptor-mediated pathways and micro-RNA are the most addressed targets of emerging therapies in modern heart failure treatment strategies. Expert opinion: Future treatment strategies should address mediators involved in multiple steps within heart failure pathogenetic pathways.

A novel adoptive transfer model of chronic lymphocytic leukemia suggests a key role for T lymphocytes in the disease

PubMed Central

Bagnara, Davide; Kaufman, Matthew S.; Calissano, Carlo; Marsilio, Sonia; Patten, Piers E. M.; Simone, Rita; Chum, Philip; Yan, Xiao-Jie; Allen, Steven L.; Kolitz, Jonathan E.; Baskar, Sivasubramanian; Rader, Christoph; Mellstedt, Hakan; Rabbani, Hodjattallah; Lee, Annette; Gregersen, Peter K.; Rai, Kanti R.

2011-01-01

Chronic lymphocytic leukemia (CLL) is an incurable adult disease of unknown etiology. Understanding the biology of CLL cells, particularly cell maturation and growth in vivo, has been impeded by lack of a reproducible adoptive transfer model. We report a simple, reproducible system in which primary CLL cells proliferate in nonobese diabetes/severe combined immunodeficiency/γcnull mice under the influence of activated CLL-derived T lymphocytes. By cotransferring autologous T lymphocytes, activated in vivo by alloantigens, the survival and growth of primary CFSE-labeled CLL cells in vivo is achieved and quantified. Using this approach, we have identified key roles for CD4+ T cells in CLL expansion, a direct link between CD38 expression by leukemic B cells and their activation, and support for CLL cells preferentially proliferating in secondary lymphoid tissues. The model should simplify analyzing kinetics of CLL cells in vivo, deciphering involvement of nonleukemic elements and nongenetic factors promoting CLL cell growth, identifying and characterizing potential leukemic stem cells, and permitting preclinical studies of novel therapeutics. Because autologous activated T lymphocytes are 2-edged swords, generating unwanted graph-versus-host and possibly autologous antitumor reactions, the model may also facilitate analyses of T-cell populations involved in immune surveillance relevant to hematopoietic transplantation and tumor cytoxicity. PMID:21385850

Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

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

Zhang, Fenxi, E-mail: fxzhang0824@gmail.com; Hong, Yan; Liang, Wenmei

Highlights: Black-Right-Pointing-Pointer Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). Black-Right-Pointing-Pointer Presence of SCs dramatically increased proliferation and migration of UCMSCs. Black-Right-Pointing-Pointer Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of 'nurse' cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neuralmore » stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.« less

Differential expression of extracellular-signal-regulated kinase 5 (ERK5) in normal and degenerated human nucleus pulposus tissues and cells

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

Liang, Weiguo, E-mail: liangweiguo@tom.com; Fang, Dejian; Ye, Dongping

2014-07-11

Highlights: • ERK5 involved in NP cells. • ERK5 involved in NP tissue. • It was important modulator. - Abstract: Extracellular-signal-regulated kinase 5 (ERK5) is a member of the mitogen-activated protein kinase (MAPK) family and regulates a wide variety of cellular processes such as proliferation, differentiation, necrosis, apoptosis and degeneration. However, the expression of ERK5 and its role in degenerated human nucleus pulposus (NP) is hitherto unknown. In this study, we observed the differential expression of ERK5 in normal and degenerated human nucleus pulposus tissues by using immunohistochemical staining and Western blot. Treatment of NP cells with Pro-inflammatory cytokine, TNF-αmore » decreased ERK5 gene expression as well as NP marker gene expression; including the type II collagen and aggrecan. Suppression of ERK5 gene expression in NP cells by ERK5 siRNA resulted in decreased gene expression of type II collagen and aggrecan. Furthermore, inhibition of ERK5 activation by BIX02188 (5 μM) decreased the gene expression of type II collagen and aggrecan in NP cells. Our results document the expression of ERK5 in degenerated nucleus pulposus tissues, and suggest a potential involvement of ERK5 in human degenerated nucleus pulposus.« less

Review of the Potential of the Ni/Cu Plating Technique for Crystalline Silicon Solar Cells

PubMed Central

Rehman, Atteq ur; Lee, Soo Hong

2014-01-01

Developing a better method for the metallization of silicon solar cells is integral part of realizing superior efficiency. Currently, contact realization using screen printing is the leading technology in the silicon based photovoltaic industry, as it is simple and fast. However, the problem with metallization of this kind is that it has a lower aspect ratio and higher contact resistance, which limits solar cell efficiency. The mounting cost of silver pastes and decreasing silicon wafer thicknesses encourages silicon solar cell manufacturers to develop fresh metallization techniques involving a lower quantity of silver usage and not relying pressing process of screen printing. In recent times nickel/copper (Ni/Cu) based metal plating has emerged as a metallization method that may solve these issues. This paper offers a detailed review and understanding of a Ni/Cu based plating technique for silicon solar cells. The formation of a Ni seed layer by adopting various deposition techniques and a Cu conducting layer using a light induced plating (LIP) process are appraised. Unlike screen-printed metallization, a step involving patterning is crucial for opening the masking layer. Consequently, experimental procedures involving patterning methods are also explicated. Lastly, the issues of adhesion, back ground plating, process complexity and reliability for industrial applications are also addressed. PMID:28788516

Review of the Potential of the Ni/Cu Plating Technique for Crystalline Silicon Solar Cells.

PubMed

Rehman, Atteq Ur; Lee, Soo Hong

2014-02-18

Developing a better method for the metallization of silicon solar cells is integral part of realizing superior efficiency. Currently, contact realization using screen printing is the leading technology in the silicon based photovoltaic industry, as it is simple and fast. However, the problem with metallization of this kind is that it has a lower aspect ratio and higher contact resistance, which limits solar cell efficiency. The mounting cost of silver pastes and decreasing silicon wafer thicknesses encourages silicon solar cell manufacturers to develop fresh metallization techniques involving a lower quantity of silver usage and not relying pressing process of screen printing. In recent times nickel/copper (Ni/Cu) based metal plating has emerged as a metallization method that may solve these issues. This paper offers a detailed review and understanding of a Ni/Cu based plating technique for silicon solar cells. The formation of a Ni seed layer by adopting various deposition techniques and a Cu conducting layer using a light induced plating (LIP) process are appraised. Unlike screen-printed metallization, a step involving patterning is crucial for opening the masking layer. Consequently, experimental procedures involving patterning methods are also explicated. Lastly, the issues of adhesion, back ground plating, process complexity and reliability for industrial applications are also addressed.

Proteome Analysis and Serological Characterization of Surface-Exposed Proteins of Rickettsia heilongjiangensis

PubMed Central

Qi, Yong; Xiong, Xiaolu; Wang, Xile; Duan, Changsong; Jia, Yinjun; Jiao, Jun; Gong, Wenping; Wen, Bohai

2013-01-01

Background Rickettsia heilongjiangensis, the agent of Far-Eastern spotted fever (FESF), is an obligate intracellular bacterium. The surface-exposed proteins (SEPs) of rickettsiae are involved in rickettsial adherence to and invasion of host cells, intracellular bacterial growth, and/or interaction with immune cells. They are also potential molecular candidates for the development of diagnostic reagents and vaccines against rickettsiosis. Methods R. heilongjiangensis SEPs were identified by biotin-streptavidin affinity purification and 2D electrophoreses coupled with ESI-MS/MS. Recombinant SEPs were probed with various sera to analyze their serological characteristics using a protein microarray and an enzyme-linked immune sorbent assay (ELISA). Results Twenty-five SEPs were identified, most of which were predicted to reside on the surface of R. heilongjiangensis cells. Bioinformatics analysis suggests that these proteins could be involved in bacterial pathogenesis. Eleven of the 25 SEPs were recognized as major seroreactive antigens by sera from R. heilongjiangensis-infected mice and FESF patients. Among the major seroreactive SEPs, microarray assays and/or ELISAs revealed that GroEL, OmpA-2, OmpB-3, PrsA, RplY, RpsB, SurA and YbgF had modest sensitivity and specificity for recognizing R. heilongjiangensis infection and/or spotted fever. Conclusions Many of the SEPs identified herein have potentially important roles in R. heilongjiangensis pathogenicity. Some of them have potential as serodiagnostic antigens or as subunit vaccine antigens against the disease. PMID:23894656

Distinctive expression pattern of OCT4 variants in different types of breast cancer.

PubMed

Soheili, Saamaaneh; Asadi, Malek Hossein; Farsinejad, Alireza

2017-01-01

OCT4 is a key regulator of self-renewal and pluripotency in embryonic stem cells which can potentially encode three spliced variants designated OCT4A, OCT4B and OCT4B1. Based on cancer stem cell concept, it is suggested that the stemness factors misexpressed in cancer cells and potentially is involved in tumorigenesis. Accordingly, in this study, we investigated the potential expression of OCT4 variants in breast cancer tissues. A total of 94 tumoral and peritumoral breast specimens were evaluated with respect to the expression of OCT4 variants using quantitative RT-PCR and immunohistochemical (IHC) analysis. We detected the expression of OCT4 variants in breast tumor tissues with no or very low levels of expression in peritumoral samples of the same patients. While OCT4B was highly expressed in lobular type of breast cancer, OCT4A and OCTB1 variants are highly expressed in low grade (I and II) ductal tumors. Furthermore, the results of this study revealed a considerable association between the expression level of OCT4 variants and the expression of ER, PR, Her2 and P53 factors. All data demonstrated a distinctive expression pattern of OCT4 spliced variants in different types of breast cancer and provide further evidence for the involvement of embryonic genes in carcinogenesis.

Strategies to enhance the bioavailability of curcumin: a potential antitumor drug

NASA Astrophysics Data System (ADS)

Kumar, Abhishek; Chittigori, Joshna; Li, Lian; Samuelson, Lynne; Sandman, Daniel; Kumar, Jayant

2012-02-01

Curcumin is a polyphenol which has elicited considerable interest for its antioxidant and anti tumor properties. Although curcumin may be used as potential therapeutic drug, it is very sparingly soluble in water which makes it less bioavailable under physiological conditions. We report two approaches to make curcumin more bioavailable. The first approach involves fabricating colloidal dispersions of curcumin in the range of tens of nanometers. The second approach involves functionalization of curcumin with polyethylene glycol (PEG) to render it water dispersible or soluble. Since curcumin is a fluorescent molecule as well as a potential drug, its interactions with cells have been investigated using one and two photon confocal fluorescence imaging. We have also observed strong interaction between curcumin and metal ions, which may have physiological implications.

The potential of induced pluripotent stem cell derived hepatocytes.

PubMed

Hannoun, Zara; Steichen, Clara; Dianat, Noushin; Weber, Anne; Dubart-Kupperschmitt, Anne

2016-07-01

Orthotopic liver transplantation remains the only curative treatment for liver disease. However, the number of patients who die while on the waiting list (15%) has increased in recent years as a result of severe organ shortages; furthermore the incidence of liver disease is increasing worldwide. Clinical trials involving hepatocyte transplantation have provided encouraging results. However, transplanted cell function appears to often decline after several months, necessitating liver transplantation. The precise aetiology of the loss of cell function is not clear, but poor engraftment and immune-mediated loss appear to be important factors. Also, primary human hepatocytes (PHH) are not readily available, de-differentiate, and die rapidly in culture. Hepatocytes are available from other sources, such as tumour-derived human hepatocyte cell lines and immortalised human hepatocyte cell lines or porcine hepatocytes. However, all these cells suffer from various limitations such as reduced or differences in functions or risk of zoonotic infections. Due to their significant potential, one possible inexhaustible source of hepatocytes is through the directed differentiation of human induced pluripotent stem cells (hiPSCs). This review will discuss the potential applications and existing limitations of hiPSC-derived hepatocytes in regenerative medicine, drug screening, in vitro disease modelling and bioartificial livers. Copyright © 2016 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Mechanistic insight of bivalent compound 21MO as potential neuroprotectant for Alzheimer’s disease

PubMed Central

Saathoff, John M.; Liu, Kai; Chojnacki, Jeremy E.; He, Liu; Chen, Qun; Lesnefsky, Edward J.; Zhang, Shijun

2016-01-01

We have recently developed a bivalent strategy to provide novel compounds that potentially target multiple risk factors involved in the development of Alzheimer’s disease (AD). Our previous studies employing a bivalent compound with a shorter spacer (17MN) implicated that this compound can localize into mitochondria and endoplasmic reticulum (ER), thus interfering with the change of mitochondria membrane potential (ΔΨm) and Ca2+ levels in MC65 cells upon removal of tetracycline (TC). In this report, we examined the effects by a bivalent compound with a longer spacer (21MO) in MC65 cells. Our results demonstrated that 21MO suppressed the change of ΔΨm, possibly via interaction with the mitochondrial complex I in MC65 cells. Interestingly, 21MO did not show any effects on the Ca2+ level upon TC removal in MC65 cells. Our previous studies suggested that the mobilization of Ca2+ in MC65 cells, upon withdraw of TC, is originated from ER, so the results implicated that 21MO may preferentially interact with mitochondria in MC65 cells under the current experimental conditions. Collectively, the results suggest that bivalent compounds with varied spacer length and cell membrane anchor moiety may exhibit neuroprotective activities via different mechanisms of action. PMID:27023508

Blood characterization using UV/vis spectroscopy

NASA Astrophysics Data System (ADS)

Mattley, Yvette D.; Mitrani-Gold, F.; Orton, S.; Bacon, Christina P.; Leparc, German F.; Bayona, M.; Potter, Robert L.; Garcia-Rubio, Luis H.

1995-05-01

The current methods used for typing blood involve an agglutination reaction which results from the association of specific antibodies with antigens present on the erythrocyte cell surface. While this method is effective, it requires involved laboratory procedures to detect the cell surface antigens. As an alternative technique, uv/vis spectroscopy has been investigated as a novel way to characterize and differentiate the blood types. Typing with this technique is based on spectral differences which appear throughout portions of both the ultraviolet and visible range. The origin of these spectral differences is unknown and presently under investigation. They may be due to intrinsic absorption differences at the molecular level, and/or they may be due to scattering differences brought about by either subtle variation in cell surface characteristics, cell shape or state of aggregation. As the background optical density in these samples is identified and accounted for, the spectral differences become more defined. This work and the continuation of this project will be included in a general database encompassing a wide range of blood samples. In addition, long term goals involve the investigation of diseased blood with the potential of providing a more rapid diagnosis for blood borne pathogens.

Transcriptome-wide targets of alternative splicing by RBM4 and possible role in cancer.

PubMed

Markus, M Andrea; Yang, Yee Hwa J; Morris, Brian J

2016-04-01

This study determined transcriptome-wide targets of the splicing factor RBM4 using Affymetrix GeneChip(®) Human Exon 1.0 ST Arrays and HeLa cells treated with RBM4-specific siRNA. This revealed 238 transcripts that were targeted for alternative splicing. Cross-linking and immunoprecipitation experiments identified 945 RBM4 targets in mouse HEK293 cells, 39% of which were ascribed to "alternative splicing" by in silico pathway analysis. Mouse embryonic stem cells transfected with Rbm4 siRNA hairpins exhibited reduced colony numbers and size consistent with involvement of RBM4 in cell proliferation. RBM4 cDNA probing of a cancer cDNA array involving 18 different tumor types from 13 different tissues and matching normal tissue found overexpression of RBM4 mRNA (p<0.01) in cervical, breast, lung, colon, ovarian and rectal cancers. Many RBM4 targets we identified have been implicated in these cancers. In conclusion, our findings reveal transcriptome-wide targets of RBM4 and point to potential cancer-related targets and mechanisms that may involve RBM4. Copyright © 2016 Elsevier Inc. All rights reserved.

COMPREHENSIVE MOLECULAR CHARACTERIZATION OF CLEAR CELL RENAL CELL CARCINOMA

PubMed Central

2013-01-01

Genetic changes underlying clear cell renal cell carcinoma (ccRCC) include alterations in genes controlling cellular oxygen sensing (e.g. VHL) and the maintenance of chromatin states (e.g. PBRM1). We surveyed more than 400 tumors using different genomic platforms and identified 19 significantly mutated genes. The PI3K/Akt pathway was recurrently mutated, suggesting this pathway as a potential therapeutic target. Widespread DNA hypomethylation was associated with mutation of the H3K36 methyltransferase SETD2, and integrative analysis suggested that mutations involving the SWI/SNF chromatin remodeling complex (PBRM1, ARID1A, SMARCA4) could have far-reaching effects on other pathways. Aggressive cancers demonstrated evidence of a metabolic shift, involving down-regulation of genes involved in the TCA cycle, decreased AMPK and PTEN protein levels, up-regulation of the pentose phosphate pathway and the glutamine transporter genes, increased acetyl-CoA carboxylase protein, and altered promoter methylation of miR-21 and GRB10. Remodeling cellular metabolism thus constitutes a recurrent pattern in ccRCC that correlates with tumor stage and severity and offers new views on the opportunities for disease treatment. PMID:23792563

Targeting mast cells in gastric cancer with special reference to bone metastases

PubMed Central

Leporini, Christian; Ammendola, Michele; Marech, Ilaria; Sammarco, Giuseppe; Sacco, Rosario; Gadaleta, Cosmo Damiano; Oakley, Caroline; Russo, Emilio; De Sarro, Giovambattista; Ranieri, Girolamo

2015-01-01

Bone metastases from gastric cancer (GC) are considered a relatively uncommon finding; however, they are related to poorer prognosis. Both primary GC and its metastatic progression rely on angiogenesis. Several lines of evidence from GC patients strongly support the involvement of mast cells (MCs) positive to tryptase (MCPT) in primary gastric tumor angiogenesis. Recently, we analyzed infiltrating MCs and neovascularization in bone tissue metastases from primary GC patients, and observed a significant correlation between infiltrating MCPT and angiogenesis. Such a finding suggested the involvement of peritumoral MCPT by infiltrating surrounding tumor cells, and in bone metastasis angiogenesis from primary GC. Thus, an MCPT-stimulated angiogenic process could support the development of metastases in bone tissue. From this perspective, we aim to review the hypothetical involvement of tumor-infiltrating, peritumoral MCPT in angiogenesis-mediated GC cell growth in the bone microenvironment and in tumor-induced osteoclastic bone resorption. We also focus on the potential use of MCPT targeting agents, such as MCs tryptase inhibitors (gabexate mesylate, nafamostat mesylate) or c-KitR tyrosine kinase inhibitors (imatinib, masitinib), as possible new anti-angiogenic and anti-resorptive strategies for the treatment of GC patients affected by bone metastases. PMID:26457010

Lens fibre cell differentiation and organelle loss: many paths lead to clarity

PubMed Central

Wride, Michael A.

2011-01-01

The programmed removal of organelles from differentiating lens fibre cells contributes towards lens transparency through formation of an organelle-free zone (OFZ). Disruptions in OFZ formation are accompanied by the persistence of organelles in lens fibre cells and can contribute towards cataract. A great deal of work has gone into elucidating the nature of the mechanisms and signalling pathways involved. It is apparent that multiple, parallel and redundant pathways are involved in this process and that these pathways form interacting networks. Furthermore, it is possible that the pathways can functionally compensate for each other, for example in mouse knockout studies. This makes sense given the importance of lens clarity in an evolutionary context. Apoptosis signalling and proteolytic pathways have been implicated in both lens fibre cell differentiation and organelle loss, including the Bcl-2 and inhibitor of apoptosis families, tumour necrosis factors, p53 and its regulators (such as Mdm2) and proteolytic enzymes, including caspases, cathepsins, calpains and the ubiquitin–proteasome pathway. Ongoing approaches being used to dissect the molecular pathways involved, such as transgenics, lens-specific gene deletion and zebrafish mutants, are discussed here. Finally, some of the remaining unresolved issues and potential areas for future studies are highlighted. PMID:21402582

Role of HCA₂ (GPR109A) in nicotinic acid and fumaric acid ester-induced effects on the skin.

PubMed

Hanson, Julien; Gille, Andreas; Offermanns, Stefan

2012-10-01

Nicotinic acid (NA) and fumaric acid esters (FAE) such as monomethyl fumarate or dimethyl fumarate are drugs that elicit a cutaneous reaction called flushing as a side effect. NA is used to reduce progression of atherosclerosis through its anti-dyslipidemic activity and lipid-independent mechanisms involving immune cells, whereas FAE are used to treat psoriasis via largely unknown mechanisms. Both, NA and FAE, induce flushing by the activation of the G-protein-coupled receptor (GPCR) Hydroxy-carboxylic acid receptor 2 (HCA₂, GPR109A) in cells of the epidermis. While the wanted effects of NA are at least in part also mediated by HCA₂, it is currently not clear whether this receptor is also involved in the anti-psoriatic effects of FAE. The HCA₂-mediated flushing response to these drugs involves the formation of prostaglandins D₂ and E₂ by Langerhans cells and keratinocytes via COX-1 in Langerhans cells and COX-2 in keratinocytes. This review summarizes recent progress in the understanding of the mechanisms underlying HCA₂-mediated flushing, describes strategies to mitigate it and discusses the potential link between flushing, HCA₂ and the anti-psoriatic effects of FAE. Copyright © 2012 Elsevier Inc. All rights reserved.

The human endonuclease III enzyme is a relevant target to potentiate cisplatin cytotoxicity in Y-box-binding protein-1 overexpressing tumor cells.

PubMed

Guay, David; Garand, Chantal; Reddy, Shanti; Schmutte, Chris; Lebel, Michel

2008-04-01

Y-box-binding protein-1 (YB-1) is a multifunctional protein involved in the regulation of transcription, translation, and mRNA splicing. In recent years, several laboratories have demonstrated that YB-1 is directly involved in the cellular response to genotoxic stress. Importantly, YB-1 is increased in tumor cell lines resistant to cisplatin, and the level of nuclear expression of YB-1 is predictive of drug resistance and patient outcome in breast tumors, ovarian cancers, and synovial sarcomas. YB-1 binds to several DNA repair enzymes in vitro including human endonuclease III (hNTH1). Human NTH1 is a bifunctional DNA glycosylase/apurinic/apyrimidinic lyase involved in base excision repair. In this study, we show that YB-1 binds specifically to the auto-inhibitory domain of hNTH1, providing a mechanism by which YB-1 stimulates hNTH1 activity. Indeed, YB-1 strongly stimulates in vitro the activity of hNTH1 toward DNA duplex probes containing oxidized bases, lesions prone to be present in cisplatin treated cells. We also observed an increase in YB-1/hNTH1 complex formation in the mammary adenocarcinoma MCF7 cell line treated with UV light and cisplatin. Such an increase was not observed with mitomycin C or the topoisomerase I inhibitor camptothecin. Accordingly, antisense RNAs against either YB-1 or hNTH1 increased cellular sensitivity to UV and cisplatin but not to mitomycin C. An antisense RNA against YB-1 increased camptothecin sensitivity. In contrast, an antisense against hNTH1 did not. Finally, siRNA against hNTH1 re-established cytotoxicity in otherwise cisplatin-resistant YB-1 overexpressing MCF7 cells. These data indicate that hNTH1 is a relevant target to potentiate cisplatin cytotoxicity in YB-1 overexpressing tumor cells.

Biphasic voltage-dependent inactivation of human NaV 1.3, 1.6 and 1.7 Na+ channels expressed in rodent insulin-secreting cells.

PubMed

Godazgar, Mahdieh; Zhang, Quan; Chibalina, Margarita V; Rorsman, Patrik

2018-05-01

Na + current inactivation is biphasic in insulin-secreting cells, proceeding with two voltage dependences that are half-maximal at ∼-100 mV and -60 mV. Inactivation of voltage-gated Na + (Na V ) channels occurs at ∼30 mV more negative voltages in insulin-secreting Ins1 and primary β-cells than in HEK, CHO or glucagon-secreting αTC1-6 cells. The difference in inactivation between Ins1 and non-β-cells persists in the inside-out patch configuration, discounting an involvement of a diffusible factor. In Ins1 cells and primary β-cells, but not in HEK cells, inactivation of a single Na V subtype is biphasic and follows two voltage dependences separated by 30-40 mV. We propose that Na V channels adopt different inactivation behaviours depending on the local membrane environment. Pancreatic β-cells are equipped with voltage-gated Na + channels that undergo biphasic voltage-dependent steady-state inactivation. A small Na + current component (10-15%) inactivates over physiological membrane potentials and contributes to action potential firing. However, the major Na + channel component is completely inactivated at -90 to -80 mV and is therefore inactive in the β-cell. It has been proposed that the biphasic inactivation reflects the contribution of different Na V α-subunits. We tested this possibility by expression of TTX-resistant variants of the Na V subunits found in β-cells (Na V 1.3, Na V 1.6 and Na V 1.7) in insulin-secreting Ins1 cells and in non-β-cells (including HEK and CHO cells). We found that all Na V subunits inactivated at 20-30 mV more negative membrane potentials in Ins1 cells than in HEK or CHO cells. The more negative inactivation in Ins1 cells does not involve a diffusible intracellular factor because the difference between Ins1 and CHO persisted after excision of the membrane. Na V 1.7 inactivated at 15--20 mV more negative membrane potentials than Na V 1.3 and Na V 1.6 in Ins1 cells but this small difference is insufficient to solely explain the biphasic inactivation in Ins1 cells. In Ins1 cells, but never in the other cell types, widely different components of Na V inactivation (separated by 30 mV) were also observed following expression of a single type of Na V α-subunit. The more positive component exhibited a voltage dependence of inactivation similar to that found in HEK and CHO cells. We propose that biphasic Na V inactivation in insulin-secreting cells reflects insertion of channels in membrane domains that differ with regard to lipid and/or membrane protein composition. © 2018 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.

TAZ is required for metastatic activity and chemoresistance of breast cancer stem cells.

PubMed

Bartucci, M; Dattilo, R; Moriconi, C; Pagliuca, A; Mottolese, M; Federici, G; Benedetto, A Di; Todaro, M; Stassi, G; Sperati, F; Amabile, M I; Pilozzi, E; Patrizii, M; Biffoni, M; Maugeri-Saccà, M; Piccolo, S; De Maria, R

2015-02-05

Metastatic growth in breast cancer (BC) has been proposed as an exclusive property of cancer stem cells (CSCs). However, formal proof of their identity as cells of origin of recurrences at distant sites and the molecular events that may contribute to tumor cell dissemination and metastasis development are yet to be elucidated. In this study, we analyzed a set of patient-derived breast cancer stem cell (BCSC) lines. We found that in vitro BCSCs exhibit a higher chemoresistance and migratory potential when compared with differentiated, nontumorigenic, breast cancer cells (dBCCs). By developing an in vivo metastatic model simulating the disease of patients with early BC, we observed that BCSCs is the only cell population endowed with metastatic potential. Gene-expression profile studies comparing metastagenic and non-metastagenic cells identified TAZ, a transducer of the Hippo pathway and biomechanical cues, as a central mediator of BCSCs metastatic ability involved in their chemoresistance and tumorigenic potential. Overexpression of TAZ in low-expressing dBCCs induced cell transformation and conferred tumorigenicity and migratory activity. Conversely, loss of TAZ in BCSCs severely impaired metastatic colonization and chemoresistance. In clinical data from 99 BC patients, high expression levels of TAZ were associated with shorter disease-free survival in multivariate analysis, thus indicating that TAZ may represent a novel independent negative prognostic factor. Overall, this study designates TAZ as a novel biomarker and a possible therapeutic target for BC.

Bcl-2 Family Members and Functional Electron Transport Chain Regulate Oxygen Deprivation-Induced Cell Death

PubMed Central

McClintock, David S.; Santore, Matthew T.; Lee, Vivian Y.; Brunelle, Joslyn; Budinger, G. R. Scott; Zong, Wei-Xing; Thompson, Craig B.; Hay, Nissim; Chandel, Navdeep S.

2002-01-01

The mechanisms underlying cell death during oxygen deprivation are unknown. We report here a model for oxygen deprivation-induced apoptosis. The death observed during oxygen deprivation involves a decrease in the mitochondrial membrane potential, followed by the release of cytochrome c and the activation of caspase-9. Bcl-XL prevented oxygen deprivation-induced cell death by inhibiting the release of cytochrome c and caspase-9 activation. The ability of Bcl-XL to prevent cell death was dependent on allowing the import of glycolytic ATP into the mitochondria to generate an inner mitochondrial membrane potential through the F1F0-ATP synthase. In contrast, although activated Akt has been shown to inhibit apoptosis induced by a variety of apoptotic stimuli, it did not prevent cell death during oxygen deprivation. In addition to Bcl-XL, cells devoid of mitochondrial DNA (ρ° cells) that lack a functional electron transport chain were resistant to oxygen deprivation. Further, murine embryonic fibroblasts from bax−/− bak−/− mice did not die in response to oxygen deprivation. These data suggest that when subjected to oxygen deprivation, cells die as a result of an inability to maintain a mitochondrial membrane potential through the import of glycolytic ATP. Proapoptotic Bcl-2 family members and a functional electron transport chain are required to initiate cell death in response to oxygen deprivation. PMID:11739725

The impact of the IGF-1 system of cancer cells on radiation response - An in vitro study.

PubMed

Venkatachalam, Senthiladipan; Mettler, Esther; Fottner, Christian; Miederer, Matthias; Kaina, Bernd; Weber, Matthias M

2017-12-01

Overexpression of the insulin-like growth factor-1 receptor (IGF-1R) is associated with increased cell proliferation, differentiation, transformation, and tumorigenicity. Additionally, signaling involved in the resistance of cancer cells to radiotherapy originates from IGF-1R. The purpose of this study was to investigate the role of the IGF-1 system in the radiation response and further evaluate its effect on the expression of DNA repair pathway genes. To inhibit the IGF-1 system, we stably transfected the Caco-2 cell line to express a kinase-deficient IGF-1R mutant. We then studied the effects of this mutation on cell growth, the response to radiation, and clonogenic survival, as well as using a cell viability assay to examine DNA damage and repair. Finally, we performed immunofluorescence for γ-H2AX to examine double-strand DNA breaks and evaluated the expression of 84 key genes involved in DNA repair with a real-time PCR array. Mutant IGF-1R cells exhibited significantly blunted cell growth and viability, compared to wild-type cells, as well as reduced clonogenic survival after γ-irradiation. However, mutant IGF-1R cells did not show any significant delays in the repair of radiation-induced DNA double-strand breaks. Furthermore, expression of mutant IGF-1R significantly down-regulated the mRNA levels of BRCA2, a major protein involved in homologous recombination DNA repair. These results indicate that blocking the IGF-1R-mediated signaling cascade, through the expression of a kinase-deficient IGF-1R mutant, reduces cell growth and sensitizes cancer cells to ionizing radiation. Therefore, the IGF-1R system could be a potential target to enhance radio-sensitivity and the efficacy of cancer treatments.

Aging of marrow stromal (skeletal) stem cells and their contribution to age-related bone loss.

PubMed

Bellantuono, Ilaria; Aldahmash, Abdullah; Kassem, Moustapha

2009-04-01

Marrow stromal cells (MSC) are thought to be stem cells with osteogenic potential and therefore responsible for the repair and maintenance of the skeleton. Age related bone loss is one of the most prevalent diseases in the elder population. It is controversial whether MSC undergo a process of aging in vivo, leading to decreased ability to form and maintain bone homeostasis with age. In this review we summarize evidence of MSC involvement in age related bone loss and suggest new emerging targets for intervention.

Late Maturation Steps Preceding Selective Nuclear Export and Egress of Progeny Parvovirus

PubMed Central

Wolfisberg, Raphael; Kempf, Christoph

2016-01-01

ABSTRACT Although the mechanism is not well understood, growing evidence indicates that the nonenveloped parvovirus minute virus of mice (MVM) may actively egress before passive release through cell lysis. We have dissected the late maturation steps of the intranuclear progeny with the aims of confirming the existence of active prelytic egress and identifying critical capsid rearrangements required to initiate the process. By performing anion-exchange chromatography (AEX), we separated intranuclear progeny particles by their net surface charges. Apart from empty capsids (EC), two distinct populations of full capsids (FC) arose in the nuclei of infected cells. The earliest population of FC to appear was infectious but, like EC, could not be actively exported from the nucleus. Further maturation of this early population, involving the phosphorylation of surface residues, gave rise to a second, late population with nuclear export potential. While capsid surface phosphorylation was strictly associated with nuclear export capacity, mutational analysis revealed that the phosphoserine-rich N terminus of VP2 (N-VP2) was dispensable, although it contributed to passive release. The reverse situation was observed for the incoming particles, which were dephosphorylated in the endosomes. Our results confirm the existence of active prelytic egress and reveal a late phosphorylation event occurring in the nucleus as a selective factor for initiating the process. IMPORTANCE In general, the process of egress of enveloped viruses is active and involves host cell membranes. However, the release of nonenveloped viruses seems to rely more on cell lysis. At least for some nonenveloped viruses, an active process before passive release by cell lysis has been reported, although the underlying mechanism remains poorly understood. By using the nonenveloped model parvovirus minute virus of mice, we could confirm the existence of an active process of nuclear export and further characterize the associated capsid maturation steps. Following DNA packaging in the nucleus, capsids required further modifications, involving the phosphorylation of surface residues, to acquire nuclear export potential. Inversely, those surface residues were dephosphorylated on entering capsids. These spatially controlled phosphorylation-dephosphorylation events concurred with the nuclear export-import potential required to complete the infectious cycle. PMID:27009963

Late Maturation Steps Preceding Selective Nuclear Export and Egress of Progeny Parvovirus.

PubMed

Wolfisberg, Raphael; Kempf, Christoph; Ros, Carlos

2016-06-01

Although the mechanism is not well understood, growing evidence indicates that the nonenveloped parvovirus minute virus of mice (MVM) may actively egress before passive release through cell lysis. We have dissected the late maturation steps of the intranuclear progeny with the aims of confirming the existence of active prelytic egress and identifying critical capsid rearrangements required to initiate the process. By performing anion-exchange chromatography (AEX), we separated intranuclear progeny particles by their net surface charges. Apart from empty capsids (EC), two distinct populations of full capsids (FC) arose in the nuclei of infected cells. The earliest population of FC to appear was infectious but, like EC, could not be actively exported from the nucleus. Further maturation of this early population, involving the phosphorylation of surface residues, gave rise to a second, late population with nuclear export potential. While capsid surface phosphorylation was strictly associated with nuclear export capacity, mutational analysis revealed that the phosphoserine-rich N terminus of VP2 (N-VP2) was dispensable, although it contributed to passive release. The reverse situation was observed for the incoming particles, which were dephosphorylated in the endosomes. Our results confirm the existence of active prelytic egress and reveal a late phosphorylation event occurring in the nucleus as a selective factor for initiating the process. In general, the process of egress of enveloped viruses is active and involves host cell membranes. However, the release of nonenveloped viruses seems to rely more on cell lysis. At least for some nonenveloped viruses, an active process before passive release by cell lysis has been reported, although the underlying mechanism remains poorly understood. By using the nonenveloped model parvovirus minute virus of mice, we could confirm the existence of an active process of nuclear export and further characterize the associated capsid maturation steps. Following DNA packaging in the nucleus, capsids required further modifications, involving the phosphorylation of surface residues, to acquire nuclear export potential. Inversely, those surface residues were dephosphorylated on entering capsids. These spatially controlled phosphorylation-dephosphorylation events concurred with the nuclear export-import potential required to complete the infectious cycle. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Nanog reverses the effects of organismal aging on mesenchymal stem cell proliferation and myogenic differentiation potential

PubMed Central

Han, Juhee; Mistriotis, Panagiotis; Lei, Pedro; Wang, Dan; Liu, Song; Andreadis, Stelios T.

2012-01-01

Although the therapeutic potential of mesenchymal stem cells (MSC) is widely accepted, loss of cell function due to donor aging or culture senescence are major limiting factors hampering their clinical application. Our laboratory recently showed that MSC originating from older donors suffer from limited proliferative capacity and significantly reduced myogenic differentiation potential. This is a major concern, as the patients most likely to suffer from cardiovascular disease are elderly. Here we tested the hypothesis that a single pluripotency associated transcription factor, namely Nanog, may reverse the proliferation and differentiation potential of BM-MSC from adult donors. Microarray analysis showed that adult (a)BM-MSC expressing Nanog clustered close to Nanog-expressing neonatal cells. Nanog markedly upregulated genes involved in cell cycle, DNA replication and DNA damage repair and enhanced the proliferation rate and clonogenic capacity of aBM-MSC. Notably, Nanog reversed the myogenic differentiation potential and restored the contractile function of aBM-MSC to a similar level as that of neonatal (n)BM-MSC. The effect of Nanog on contractility was mediated – at least in part - through activation of the TGF-β pathway by diffusible factors secreted in the conditioned medium of Nanog-expressing BM-MSC. Overall, our results suggest that Nanog may be used to overcome the effects of organismal aging on aBM-MSC, thereby increasing the potential of MSC from aged donors for cellular therapy and tissue regeneration. PMID:22949105

Flavonoid-induced glutathione depletion: Potential implications for cancer treatment☆

PubMed Central

Kachadourian, Remy; Day, Brian J.

2014-01-01

The ability of a number of flavonoids to induce glutathione (GSH) depletion was measured in lung (A549), myeloid (HL-60), and prostate (PC-3) human tumor cells. The hydroxychalcone (2′-HC) and the dihydroxychalcones (2′,2-, 2′,3-, 2′,4-, and 2′,5′-DHC) were the most effective in A549 and HL-60 cells, depleting more than 50% of intracellular GSH within 4 h of exposure at 25 µM. In contrast, the flavones chrysin and apigenin were the most effective in PC-3 cells, depleting 50–70% of intracellular GSH within 24 h of exposure at 25 µM. In general, these flavonoids were more effective than three classical substrates of multidrug resistance protein 1 (MK-571, indomethacin, and verapamil). Prototypic flavonoids (2′,5′-DHC and chrysin) were subsequently tested for their abilities to potentiate the toxicities of prooxidants (etoposide, rotenone, 2-methoxyestradiol, and curcumin). In A549 cells, 2′,5′-DHC potentiated the cytotoxicities of rotenone, 2-methoxyestradiol, and curcumin, but not etoposide. In HL-60 and PC-3 cells, chrysin potentiated the cytotoxicity of curcumin, cytotoxicity that was attenuated by the catalytic antioxidant manganese(III) meso-tetrakis(N-ethylpyridinium-2-yl)porphyrin (MnTE-2-PyP). Assessments of mitochondrial GSH levels mitochondrial membrane potential and cytochrome c release showed that the potentiation effects induced by 2′,5′-DHC and chrysin involve mitochondrial dysfunction. PMID:16781454

The extrinsic and intrinsic apoptotic pathways are involved in manganese toxicity in rat astrocytoma C6 cells.

PubMed

Alaimo, Agustina; Gorojod, Roxana M; Kotler, Mónica L

2011-08-01

Manganese (Mn) is a trace element known to be essential for maintaining the proper function and regulation of many biochemical and cellular reactions. However, chronic exposure to high levels of Mn in occupational or environmental settings can lead to its accumulation in the brain resulting in a degenerative brain disorder referred to as Manganism. Astrocytes are the main Mn store in the central nervous system and several lines of evidence implicate these cells as major players in the role of Manganism development. In the present study, we employed rat astrocytoma C6 cells as a sensitive experimental model for investigating molecular mechanisms involved in Mn neurotoxicity. Our results show that C6 cells undergo reactive oxygen species-mediated apoptotic cell death involving caspase-8 and mitochondrial-mediated pathways in response to Mn. Exposed cells exhibit typical apoptotic features, such as chromatin condensation, cell shrinkage, membrane blebbing, caspase-3 activation and caspase-specific cleavage of the endogenous substrate poly (ADP-ribose) polymerase. Participation of the caspase-8 dependent pathway was assessed by increased levels of FasL, caspase-8 activation and Bid cleavage. The involvement of the mitochondrial pathway was demonstrated by the disruption of the mitochondrial membrane potential, the opening of the mitochondrial permeability transition pore, cytochrome c release, caspase-9 activation and the increased mitochondrial levels of the pro-apoptotic Bcl-2 family proteins. In addition, our data also shows for the first time that mitochondrial fragmentation plays a relevant role in Mn-induced apoptosis. Taking together, these findings contribute to a deeper elucidation of the molecular signaling mechanisms underlying Mn-induced apoptosis. Copyright © 2011 Elsevier B.V. All rights reserved.

Characterization of Apoptosis Induced by Emodin and Related Regulatory Mechanisms in Human Neuroblastoma Cells

PubMed Central

Huang, Fu-Jen; Hsuuw, Yan-Der; Chan, Wen-Hsiung

2013-01-01

Emodin (1,3,8-trihydroxy-6-methylanthraquinone), a major constituent of rhubarb, has a wide range of therapeutic applications. Recent studies have shown that emodin can induce or prevent cell apoptosis, although the precise molecular mechanisms underlying these effects are unknown. Experiments from the current study revealed that emodin (10–20 μM) induces apoptotic processes in the human neuroblastoma cell line, IMR-32, but exerts no injury effects at treatment doses below 10 μM. Treatment with emodin at concentrations of 10–20 μM led to a direct increase in the reactive oxygen species (ROS) content in IMR-32 cells, along with significant elevation of cytoplasmic free calcium and nitric oxide (NO) levels, loss of mitochondrial membrane potential (MMP), activation of caspases-9 and -3, and cell death. Pretreatment with nitric oxide (NO) scavengers suppressed the apoptotic biochemical changes induced by 20 μM emodin, and attenuated emodin-induced p53 and p21 expression involved in apoptotic signaling. Our results collectively indicate that emodin at concentrations of 10–20 μM triggers apoptosis of IMR-32 cells via a mechanism involving both ROS and NO. Based on the collective results, we propose a model for an emodin-triggered apoptotic signaling cascade that sequentially involves ROS, Ca2+, NO, p53, caspase-9 and caspase-3. PMID:24113589

The PTPN14 Tumor Suppressor Is a Degradation Target of Human Papillomavirus E7.

PubMed

Szalmás, Anita; Tomaić, Vjekoslav; Basukala, Om; Massimi, Paola; Mittal, Suruchi; Kónya, József; Banks, Lawrence

2017-04-01

Activation of signaling pathways ensuring cell growth is essential for the proliferative competence of human papillomavirus (HPV)-infected cells. Tyrosine kinases and phosphatases are key regulators of cellular growth control pathways. A recently identified potential cellular target of HPV E7 is the cytoplasmic protein tyrosine phosphatase PTPN14, which is a potential tumor suppressor and is linked to the control of the Hippo and Wnt/beta-catenin signaling pathways. In this study, we show that the E7 proteins of both high-risk and low-risk mucosal HPV types can interact with PTPN14. This interaction is independent of retinoblastoma protein (pRb) and involves residues in the carboxy-terminal region of E7. We also show that high-risk E7 induces proteasome-mediated degradation of PTPN14 in cells derived from cervical tumors. This degradation appears to be independent of cullin-1 or cullin-2 but most likely involves the UBR4/p600 ubiquitin ligase. The degree to which E7 downregulates PTPN14 would suggest that this interaction is important for the viral life cycle and potentially also for the development of malignancy. In support of this we find that overexpression of PTPN14 decreases the ability of HPV-16 E7 to cooperate with activated EJ-ras in primary cell transformation assays. IMPORTANCE This study links HPV E7 to the deregulation of protein tyrosine phosphatase signaling pathways. PTPN14 is classified as a potential tumor suppressor protein, and here we show that it is very susceptible to HPV E7-induced proteasome-mediated degradation. Intriguingly, this appears to use a mechanism that is different from that employed by E7 to target pRb. Therefore, this study has important implications for our understanding of the molecular basis for E7 function and also sheds important light on the potential role of PTPN14 as a tumor suppressor. Copyright © 2017 American Society for Microbiology.

Oocytes express an endogenous red fluorescent protein in a stony coral, Euphyllia ancora: a potential involvement in coral oogenesis

PubMed Central

Shikina, Shinya; Chiu, Yi-Ling; Chung, Yi-Jou; Chen, Chieh-Jhen; Lee, Yan-Horn; Chang, Ching-Fong

2016-01-01

To date,the molecular and cellular mechanisms underlying coral sexual reproduction remain largely unknown. We then performed a differential screen to identify genes related to oogenesis in the stony coral Euphyllia ancora. We identified a clone encoding a novel red fluorescent protein cDNA of E. ancora (named EaRFP). Microscopic observation and quantitative RT-PCR revealed that EaRFP is almost exclusively expressed in the ovary of the adult coral. The combination of the ovarian-cell separation method and the RT-PCR analysis revealed that the oocytes, but not the ovarian somatic cells, are the cells expressing EaRFP. Immunohistochemical analysis revealed that the expression of EaRFP starts in the early stage of the oocyte and continues until the maturation period. Furthermore, recombinant EaRFP was shown to possess an H2O2 degradation activity. These results raise the possibility that EaRFP plays a role in protecting the oocytes from oxidative stress from the early to late stages of oogenesis. The present study provides not only the first evidence for the potential involvement of FPs in coral oogenesis but also an insight into a cellular strategy underlying coral sexual reproduction. PMID:27167722

Oocytes express an endogenous red fluorescent protein in a stony coral, Euphyllia ancora: a potential involvement in coral oogenesis.

PubMed

Shikina, Shinya; Chiu, Yi-Ling; Chung, Yi-Jou; Chen, Chieh-Jhen; Lee, Yan-Horn; Chang, Ching-Fong

2016-05-11

To date,the molecular and cellular mechanisms underlying coral sexual reproduction remain largely unknown. We then performed a differential screen to identify genes related to oogenesis in the stony coral Euphyllia ancora. We identified a clone encoding a novel red fluorescent protein cDNA of E. ancora (named EaRFP). Microscopic observation and quantitative RT-PCR revealed that EaRFP is almost exclusively expressed in the ovary of the adult coral. The combination of the ovarian-cell separation method and the RT-PCR analysis revealed that the oocytes, but not the ovarian somatic cells, are the cells expressing EaRFP. Immunohistochemical analysis revealed that the expression of EaRFP starts in the early stage of the oocyte and continues until the maturation period. Furthermore, recombinant EaRFP was shown to possess an H2O2 degradation activity. These results raise the possibility that EaRFP plays a role in protecting the oocytes from oxidative stress from the early to late stages of oogenesis. The present study provides not only the first evidence for the potential involvement of FPs in coral oogenesis but also an insight into a cellular strategy underlying coral sexual reproduction.

Involvement of cell surface TG2 in the aggregation of K562 cells triggered by gluten.

PubMed

Feriotto, G; Calza, R; Bergamini, C M; Griffin, M; Wang, Z; Beninati, S; Ferretti, V; Marzola, E; Guerrini, R; Pagnoni, A; Cavazzini, A; Casciano, F; Mischiati, C

2017-03-01

Gluten-induced aggregation of K562 cells represents an in vitro model reproducing the early steps occurring in the small bowel of celiac patients exposed to gliadin. Despite the clear involvement of TG2 in the activation of the antigen-presenting cells, it is not yet clear in which compartment it occurs. Herein we study the calcium-dependent aggregation of these cells, using either cell-permeable or cell-impermeable TG2 inhibitors. Gluten induces efficient aggregation when calcium is absent in the extracellular environment, while TG2 inhibitors do not restore the full aggregating potential of gluten in the presence of calcium. These findings suggest that TG2 activity is not essential in the cellular aggregation mechanism. We demonstrate that gluten contacts the cells and provokes their aggregation through a mechanism involving the A-gliadin peptide 31-43. This peptide also activates the cell surface associated extracellular TG2 in the absence of calcium. Using a bioinformatics approach, we identify the possible docking sites of this peptide on the open and closed TG2 structures. Peptide docks with the closed TG2 structure near to the GTP/GDP site, by establishing molecular interactions with the same amino acids involved in stabilization of GTP binding. We suggest that it may occur through the displacement of GTP, switching the TG2 structure from the closed to the active open conformation. Furthermore, docking analysis shows peptide binding with the β-sandwich domain of the closed TG2 structure, suggesting that this region could be responsible for the different aggregating effects of gluten shown in the presence or absence of calcium. We deduce from these data a possible mechanism of action by which gluten makes contact with the cell surface, which could have possible implications in the celiac disease onset.

Rocky Mountain spotted fever.

PubMed

Lacz, N L; Schwartz, R A; Kapila, R

2006-04-01

Rocky Mountain spotted fever (RMSF) is an unusual but important dermatological condition to identify without hesitation. The classic triad of headache, fever, and a rash that begins on the extremities and travels proximally to involve the trunk is found in a majority of patients. The cutaneous centripetal pattern is a result of cell to cell migration by the causative organism Rickettsia rickettsii. Such individuals should receive prompt antimicrobial therapy and supportive care to avoid serious and potentially fatal complications.

Alleviating the Common Confusion Caused by Polarity in Electrochemistry.

ERIC Educational Resources Information Center

Moran, P. J.; Gileadi, E.

1989-01-01

Discussed is some of the confusion encountered in electrochemistry due to misunderstandings of sign conventions and simple mathematical errors. Clarified are issues involving emf series, IUPAC sign conventions, calculation of cell potentials, reference electrodes, the polarity of electrodes in electrochemical devices, and overpotential. (CW)

How do CARs work?

PubMed Central

Davila, Marco L.; Brentjens, Renier; Wang, Xiuyan; Rivière, Isabelle; Sadelain, Michel

2012-01-01

Second-generation chimeric antigen receptors (CARs) are powerful tools to redirect antigen-specific T cells independently of HLA-restriction. Recent clinical studies evaluating CD19-targeted T cells in patients with B-cell malignancies demonstrate the potency of CAR-engineered T cells. With results from 28 subjects enrolled by five centers conducting studies in patients with chronic lymphocytic leukemia (CLL) or lymphoma, some insights into the parameters that determine T-cell function and clinical outcome of CAR-based approaches are emerging. These parameters involve CAR design, T-cell production methods, conditioning chemotherapy as well as patient selection. Here, we discuss the potential relevance of these findings and in particular the interplay between the adoptive transfer of T cells and pre-transfer patient conditioning. PMID:23264903

TRPV2.

PubMed

Kojima, Itaru; Nagasawa, Masahiro

2014-01-01

Transient receptor potential vanilloid type 2, TRPV2, is a calcium-permeable cation channel belonging to the TRPV channel family. This channel is activated by heat (>52 °C), various ligands, and mechanical stresses. In most of the cells, a large portion of TRPV2 is located in the endoplasmic reticulum under unstimulated conditions. Upon stimulation of the cells with phosphatidylinositol 3-kinase-activating ligands, TRPV2 is translocated to the plasma membrane and functions as a cation channel. Mechanical stress may also induce translocation of TRPV2 to the plasma membrane. The expression of TRPV2 is high in some types of cells including neurons, neuroendocrine cells, immune cells involved in innate immunity, and certain types of cancer cells. TRPV2 may modulate various cellular functions in these cells.

Reactive astrocytes and therapeutic potential in focal ischemic stroke

PubMed Central

Choudhury, Gourav Roy; Ding, Shinghua

2015-01-01

Astrocytes are specialized and the most abundant cell type in the central nervous system (CNS). They play important roles in the physiology of the brain. Astrocytes are also critically involved in many CNS disorders including focal ischemic stroke, the leading cause of brain injury and death in patients. One of the prominent pathological features of a focal ischemic stroke is reactive astrogliosis and glial scar formation. Reactive astrogliosis is accompanied with changes in morphology, proliferation and gene expression in the reactive astrocytes. This study provides an overview of the most recent advances in astrocytic Ca2+ signaling, spatial and temporal dynamics of the morphology and proliferation of reactive astrocytes as well as signaling pathways involved in the reactive astrogliosis after ischemic stroke based on results from experimental studies performed in various animal models. This review also discusses the therapeutic potential of reactive astrocytes in a focal ischemic stroke. As reactive astrocytes exhibit high plasticity, we suggest that modulation of local reactive astrocytes is a promising strategy for cell-based stroke therapy. PMID:25982835

Calcium and ROS: A mutual interplay

PubMed Central

Görlach, Agnes; Bertram, Katharina; Hudecova, Sona; Krizanova, Olga

2015-01-01

Calcium is an important second messenger involved in intra- and extracellular signaling cascades and plays an essential role in cell life and death decisions. The Ca2+ signaling network works in many different ways to regulate cellular processes that function over a wide dynamic range due to the action of buffers, pumps and exchangers on the plasma membrane as well as in internal stores. Calcium signaling pathways interact with other cellular signaling systems such as reactive oxygen species (ROS). Although initially considered to be potentially detrimental byproducts of aerobic metabolism, it is now clear that ROS generated in sub-toxic levels by different intracellular systems act as signaling molecules involved in various cellular processes including growth and cell death. Increasing evidence suggests a mutual interplay between calcium and ROS signaling systems which seems to have important implications for fine tuning cellular signaling networks. However, dysfunction in either of the systems might affect the other system thus potentiating harmful effects which might contribute to the pathogenesis of various disorders. PMID:26296072

New insights of T cells in the pathogenesis of psoriasis

PubMed Central

Cai, Yihua; Fleming, Chris; Yan, Jun

2012-01-01

Psoriasis is one of the most common immune-mediated chronic, inflammatory skin diseases characterized by hyperproliferative keratinocytes and infiltration of T cells, dendritic cells, macrophages and neutrophils. Although the pathogenesis of psoriasis is not fully understood, there is ample evidence suggesting that the dysregulation of immune cells in the skin, particularly T cells, plays a critical role in psoriasis development. In this review, we mainly focus on the pathogenic T cells and discuss how these T cells are activated and involved in the disease pathogenesis. Newly identified ‘professional' IL-17-producing dermal γδ T cells and their potential role in psoriasis will also be included. Finally, we will briefly summarize the recent progress on the T cell and its related cytokine-targeted therapy for psoriasis treatment. PMID:22705915

Microfluidics separation reveals the stem-cell-like deformability of tumor-initiating cells.

PubMed

Zhang, Weijia; Kai, Kazuharu; Choi, Dong Soon; Iwamoto, Takayuki; Nguyen, Yen H; Wong, Helen; Landis, Melissa D; Ueno, Naoto T; Chang, Jenny; Qin, Lidong

2012-11-13

Here we report a microfluidics method to enrich physically deformable cells by mechanical manipulation through artificial microbarriers. Driven by hydrodynamic forces, flexible cells or cells with high metastatic propensity change shape to pass through the microbarriers and exit the separation device, whereas stiff cells remain trapped. We demonstrate the separation of (i) a mixture of two breast cancer cell types (MDA-MB-436 and MCF-7) with distinct deformabilities and metastatic potentials, and (ii) a heterogeneous breast cancer cell line (SUM149), into enriched flexible and stiff subpopulations. We show that the flexible phenotype is associated with overexpression of multiple genes involved in cancer cell motility and metastasis, and greater mammosphere formation efficiency. Our observations support the relationship between tumor-initiating capacity and cell deformability, and demonstrate that tumor-initiating cells are less differentiated in terms of cell biomechanics.

The role of Fas ligand and transforming growth factor beta in tumor progression: molecular mechanisms of immune privilege via Fas-mediated apoptosis and potential targets for cancer therapy.

PubMed

Kim, Ryungsa; Emi, Manabu; Tanabe, Kazuaki; Uchida, Yoko; Toge, Tetsuya

2004-06-01

Despite the fact that expression of Fas ligand (FasL) in cytotoxic T lymphocytes (CTLs) and in natural killer (NK) cells plays an important role in Fas-mediated tumor killing, During tumor progression FasL-expressing tumor cells are involved in counterattacking to kill tumor-infiltrating lymphocytes (TILs). Soluble FasL levels also increase with tumor progression in solid tumors, and this increase inhibits Fas-mediated tumor killing by CTLs and NK cells. The increased expression of FasL in tumor cells is associated with decreased expression of Fas; and the promoter region of the FASL gene is regulated by transcription factors, such as neuronal factor kappaB (NF-kappaB) and AP-1, in the tumor microenvironment. Although the ratio of FasL expression to Fas expression in tumor cells is not strongly related to the induction of apoptosis in TILs, increased expression of FasL is associated with decreased Fas levels in tumor cells that can escape immune surveillance and facilitate tumor progression and metastasis. Transforming growth factor beta (TGF-beta) is a potent growth inhibitor and has tumor-suppressing activity in the early phases of carcinogenesis. During subsequent tumor progression, the increased secretion of TGF-beta by both tumor cells and, in a paracrine fashion, stromal cells, is involved in the enhancement of tumor invasion and metastasis accompanied by immunosuppression. Herein, the authors review the clinical significance of FasL and TGF-beta expression patterns as features of immune privilege accompanying tumor progression in the tumor microenvironment. Potential strategies for identifying which molecules can serve as targets for effective antitumor therapy also are discussed. Copyright 2004 American Cancer Society.

Assessment of the potential utility of different regions of Streptococcus uberis adhesion molecule (SUAM) for mastitis subunit vaccine development.

PubMed

Perrig, Melina Soledad; Veaute, Carolina; Renna, María Sol; Pujato, Nazarena; Calvinho, Luis; Marcipar, Iván; Barbagelata, María Sol

2017-04-01

Streptococcus uberis is one of the most prevalent pathogens causing clinical and subclinical mastitis worldwide. Among bacterial factors involved in intramammary infections caused by this organism, S. uberis adhesion molecule (SUAM) is one of the main virulence factors identified. This molecule is involved in S. uberis internalization to mammary epithelial cells through lactoferrin (Lf) binding. The objective of this study was to evaluate SUAM properties as a potential subunit vaccine component for prevention of S. uberis mastitis. B epitope prediction analysis of SUAM sequence was used to identify potentially immunogenic regions. Since these regions were detected all along the gene, this criterion did not allow selecting a specific region as a potential immunogen. Hence, four fractions of SUAM (-1fr, 2fr, 3fr and 4fr), comprising most of the protein, were cloned and expressed. Every fraction elicited a humoral immune response in mice as predicted by bioinformatics analysis. SUAM-1fr generated antibodies with the highest recognition ability towards SUAM native protein. Moreover, antibodies against SUAM-1fr produced the highest proportion of internalization inhibition of S. uberis to mammary epithelial cells. In conclusion, SUAM immunogenic and functionally relevant regions were identified and allowed to propose SUAM-1fr as a potential candidate for a subunit vaccine for S. uberis mastitis prevention. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Functional Role of RB-Dependent Pathway in the Control of Quiescence in Adult Epidermal Stem Cells Revealed by Genomic Profiling

PubMed Central

Lorz, Corina; García-Escudero, Ramón; Segrelles, Carmen; Garín, Marina I.; Ariza, José M.; Santos, Mirentxu; Ruiz, Sergio; Lara, María F.; Martínez-Cruz, Ana B.; Costa, Clotilde; Buitrago-Pérez, Águeda; Saiz-Ladera, Cristina; Dueñas, Marta

2010-01-01

Continuous cell renewal in mouse epidermis is at the expense of a pool of pluripotent cells that lie in a well defined niche in the hair follicle known as the bulge. To identify mechanisms controlling hair follicle stem cell homeostasis, we developed a strategy to isolate adult bulge stem cells in mice and to define their transcriptional profile. We observed that a large number of transcripts are underexpressed in hair follicle stem cells when compared to non-stem cells. Importantly, the majority of these downregulated genes are involved in cell cycle. Using bioinformatics tools, we identified the E2F transcription factor family as a potential element involved in the regulation of these transcripts. To determine their functional role, we used engineered mice lacking Rb gene in epidermis, which showed increased expression of most E2F family members and increased E2F transcriptional activity. Experiments designed to analyze epidermal stem cell functionality (i.e.: hair regrowth and wound healing) imply a role of the Rb-E2F axis in the control of stem cell quiescence in epidermis. Electronic supplementary material The online version of this article (doi:10.1007/s12015-010-9139-0) contains supplementary material, which is available to authorized users. PMID:20376578

MicroRNAs: From Female Fertility, Germ Cells, and Stem Cells to Cancer in Humans

PubMed Central

Virant-Klun, Irma; Ståhlberg, Anders; Kubista, Mikael; Skutella, Thomas

2016-01-01

MicroRNAs are a family of naturally occurring small noncoding RNA molecules that play an important regulatory role in gene expression. They are suggested to regulate a large proportion of protein encoding genes by mediating the translational suppression and posttranscriptional control of gene expression. Recent findings show that microRNAs are emerging as important regulators of cellular differentiation and dedifferentiation, and are deeply involved in developmental processes including human preimplantation development. They keep a balance between pluripotency and differentiation in the embryo and embryonic stem cells. Moreover, it became evident that dysregulation of microRNA expression may play a fundamental role in progression and dissemination of different cancers including ovarian cancer. The interest is still increased by the discovery of exosomes, that is, cell-derived vesicles, which can carry different proteins but also microRNAs between different cells and are involved in cell-to-cell communication. MicroRNAs, together with exosomes, have a great potential to be used for prognosis, therapy, and biomarkers of different diseases including infertility. The aim of this review paper is to summarize the existent knowledge on microRNAs related to female fertility and cancer: from primordial germ cells and ovarian function, germinal stem cells, oocytes, and embryos to embryonic stem cells. PMID:26664407

Mast cells in rheumatoid arthritis: friends or foes?

PubMed

Rivellese, Felice; Nerviani, Alessandra; Rossi, Francesca Wanda; Marone, Gianni; Matucci-Cerinic, Marco; de Paulis, Amato; Pitzalis, Costantino

2017-06-01

Mast cells are tissue-resident cells of the innate immunity, implicated in the pathogenesis of many autoimmune diseases, including rheumatoid arthritis (RA). They are present in synovia and their activation has been linked to the potentiation of inflammation in the course of RA. However, recent investigations questioned the role of mast cells in arthritis. In particular, animal models generated conflicting results, so that many of their pro-inflammatory, i.e. pro-arthritogenic functions, even though supported by robust experimental evidence, have been labelled as redundant. At the same time, a growing body of evidence suggests that mast cells can act as tunable immunomodulatory cells. These characteristics, not yet fully understood in the context of RA, could partially explain the inconsistent results obtained with experimental models, which do not account for the pro- and anti-inflammatory functions exerted in more chronic heterogeneous conditions such as RA. Here we present an overview of the current knowledge on mast cell involvement in RA, including the intriguing hypothesis of mast cells acting as subtle immunomodulatory cells and the emerging concept of synovial mast cells as potential biomarkers for patient stratification. Copyright © 2017 Elsevier B.V. All rights reserved.

Mesenchymal Stem Cell Therapy for Nonhealing Cutaneous Wounds

PubMed Central

Hanson, Summer E.; Bentz, Michael L.; Hematti, Peiman

2014-01-01

Summary Chronic wounds remain a major challenge in modern medicine and represent a significant burden, affecting not only physical and mental health, but also productivity, health care expenditure, and long-term morbidity. Even under optimal conditions, the healing process leads to fibrosis or scar. One promising solution, cell therapy, involves the transplantation of progenitor/stem cells to patients through local or systemic delivery, and offers a novel approach to many chronic diseases, including nonhealing wounds. Mesenchymal stem cells are multipotent, adult progenitor cells of great interest because of their unique immunologic properties and regenerative potential. A variety of preclinical and clinical studies have shown that mesenchymal stem cells may have a useful role in wound-healing and tissue-engineering strategies and both aesthetic and reconstructive surgery. Recent advances in stem cell immunobiology can offer insight into the multiple mechanisms through which mesenchymal stem cells could affect underlying pathophysiologic processes associated with nonhealing mesenchymal stem cells. Critical evaluation of the current literature is necessary for understanding how mesenchymal stem cells could potentially revolutionize our approach to skin and soft-tissue defects and designing clinical trials to address their role in wound repair and regeneration. PMID:20124836

Herbal Formulation C168 Attenuates Proliferation and Induces Apoptosis in HCT 116 Human Colorectal Carcinoma Cells: Role of Oxidative Stress and DNA Damage

PubMed Central

Leong, Lek Mun; Chan, Kok Meng; Hamid, Asmah; Latip, Jalifah; Rajab, Nor Fadilah

2016-01-01

The use of herbal formulations has gained scientific interest, particularly in cancer treatment. In this study, the herbal formulation of interest, denoted as C168, is a mixture of eight genera of plants. This study aims to investigate the antiproliferative effect of C168 methanol extract (CME) on various cancer cells and its underlying mechanism of action on the most responsive cell line, namely, HCT 116 cells. CME exerted antiproliferative activities on HCT 116 colorectal carcinoma cells and HepG2 hepatocellular carcinoma cells but not on CCD-841-CoN normal colon epithelial cells, Jurkat E6.1 lymphoblastic leukemic cells, and V79-4 Chinese hamster lung fibroblasts. Further investigation on HCT 116 cells showed that CME induced G2/M cell-cycle arrest and apoptosis. Treatment of CME induced oxidative stress in HCT 116 cells by increasing the superoxide anion level and decreasing the intracellular glutathione. CME also increased tail moment value and H2AX phosphorylation in HCT 116 cells, suggesting DNA damage as an early signal of CME induced apoptosis. Loss of mitochondrial membrane potential in CME-treated cells also indicated the involvement of mitochondria in CME induced apoptosis. This study indicated the selectivity of CME toward colon cancer cells with the involvement of oxidative damage as its possible mechanism of action. PMID:26884792

The oncogenic transforming potential of the passage of single α particles through mammalian cell nuclei

PubMed Central

Miller, Richard C.; Randers-Pehrson, Gerhard; Geard, Charles R.; Hall, Eric J.; Brenner, David J.

1999-01-01

Domestic, low-level exposure to radon gas is considered a major environmental lung-cancer hazard involving DNA damage to bronchial cells by α particles from radon progeny. At domestic exposure levels, the relevant bronchial cells are very rarely traversed by more than one α particle, whereas at higher radon levels—at which epidemiological studies in uranium miners allow lung-cancer risks to be quantified with reasonable precision—these bronchial cells are frequently exposed to multiple α-particle traversals. Measuring the oncogenic transforming effects of exactly one α particle without the confounding effects of multiple traversals has hitherto been unfeasible, resulting in uncertainty in extrapolations of risk from high to domestic radon levels. A technique to assess the effects of single α particles uses a charged-particle microbeam, which irradiates individual cells or cell nuclei with predefined exact numbers of particles. Although previously too slow to assess the relevant small oncogenic risks, recent improvements in throughput now permit microbeam irradiation of large cell numbers, allowing the first oncogenic risk measurements for the traversal of exactly one α particle through a cell nucleus. Given positive controls to ensure that the dosimetry and biological controls were comparable, the measured oncogenicity from exactly one α particle was significantly lower than for a Poisson-distributed mean of one α particle, implying that cells traversed by multiple α particles contribute most of the risk. If this result applies generally, extrapolation from high-level radon risks (involving cellular traversal by multiple α particles) may overestimate low-level (involving only single α particles) radon risks. PMID:9874764

Feedback regulation of mitochondria by caspase-9 in the B cell receptor-mediated apoptosis.

PubMed

Eeva, J; Nuutinen, U; Ropponen, A; Mättö, M; Eray, M; Pellinen, R; Wahlfors, J; Pelkonen, J

2009-12-01

During the germinal centre reaction (GC), B cells with non-functional or self-reactive antigen receptors are negatively selected by apoptosis to generate B cell repertoire with appropriate antigen specificities. We studied the molecular mechanism of Fas/CD95- and B cell receptor (BCR)-induced apoptosis to shed light on the signalling events involved in the negative selection of GC B cells. As an experimental model, we used human follicular lymphoma (FL) cell line HF1A3, which originates from a GC B cell, and transfected HF1A3 cell lines overexpressing Bcl-x(L), c-FLIP(long) or dominant negative (DN) caspase-9. Fas-induced apoptosis was dependent on the caspase-8 activation, since the overexpression of c-FLIP(long), a natural inhibitor of caspase-8 activation, blocked apoptosis induced by Fas. In contrast, caspase-9 activation was not involved in Fas-induced apoptosis. BCR-induced apoptosis showed the typical characteristics of mitochondria-dependent (intrinsic) apoptosis. Firstly, the activation of caspase-9 was involved in BCR-induced DNA fragmentation, while caspase-8 showed only marginal role. Secondly, overexpression of Bcl-x(L) could block all apoptotic changes induced by BCR. As a novel finding, we demonstrate that caspase-9 can enhance the cytochrome-c release and collapse of mitochondrial membrane potential (DeltaPsi(m)) during BCR-induced apoptosis. The requirement of different signalling pathways in apoptosis induced by BCR and Fas may be relevant, since Fas- and BCR-induced apoptosis can thus be regulated independently, and targeted to different subsets of GC B cells.

Transport characteristics of L-citrulline in renal apical membrane of proximal tubular cells.

PubMed

Mitsuoka, Keisuke; Shirasaka, Yoshiyuki; Fukushi, Akimasa; Sato, Masanobu; Nakamura, Toshimichi; Nakanishi, Takeo; Tamai, Ikumi

2009-04-01

L-Citrulline has diagnostic potential for renal function, because its plasma concentration increases with the progression of renal failure. Although L-citrulline extracted by glomerular filtration in kidney is mostly reabsorbed, the mechanism involved is not clearly understood. The present study was designed to characterize L-citrulline transport across the apical membranes of renal epithelial tubular cells, using primary-cultured rat renal proximal tubular cells, as well as the human kidney proximal tubular cell line HK-2. L-Citrulline was transported in a Na(+)-dependent manner from the apical side of both cell types cultured on permeable supports with a microporous membrane. Kinetic analysis indicated that the transport involves two distinct Na(+)-dependent saturable systems and one Na(+)-independent saturable system in HK-2 cells. The uptake was competitively inhibited by neutral and cationic, but not anionic amino acids. Relatively large cationic and anionic compounds inhibited the uptake, but smaller ones did not. In HK-2 cells, mRNA expression of SLC6A19 and SLC7A9, which encode B(0)AT1 and b(0,+)AT, respectively, was detected by RT-PCR. In addition, L-citrulline transport was significantly decreased in HK-2 cells in which either SLC6A19 or SLC7A9 was silenced. Hence, these results suggest that amino acid transporters B(0)AT1 and b(0,+)AT are involved in the reabsorption of L-citrulline in the kidney, at least in part, by mediating the apical membrane transport of L-citrulline in renal tubule cells.

Apoptotic effect of novel Schiff Based CdCl2(C14H21N3O2) complex is mediated via activation of the mitochondrial pathway in colon cancer cells

PubMed Central

Hajrezaie, Maryam; Paydar, Mohammadjavad; Looi, Chung Yeng; Moghadamtousi, Soheil Zorofchian; Hassandarvish, Pouya; Salga, Muhammad Saleh; Karimian, Hamed; Shams, Keivan; Zahedifard, Maryam; Majid, Nazia Abdul; Ali, Hapipah Mohd; Abdulla, Mahmood Ameen

2015-01-01

The development of metal-based agents has had a tremendous role in the present progress in cancer chemotherapy. One well-known example of metal-based agents is Schiff based metal complexes, which hold great promise for cancer therapy. Based on the potential of Schiff based complexes for the induction of apoptosis, this study aimed to examine the cytotoxic and apoptotic activity of a CdCl2(C14H21N3O2) complex on HT-29 cells. The complex exerted a potent suppressive effect on HT-29 cells with an IC50 value of 2.57 ± 0.39 after 72 h of treatment. The collapse of the mitochondrial membrane potential and the elevated release of cytochrome c from the mitochondria to the cytosol indicate the involvement of the intrinsic pathway in the induction of apoptosis. The role of the mitochondria-dependent apoptotic pathway was further proved by the significant activation of the initiator caspase-9 and the executioner caspases-3 and -7. In addition, the activation of caspase-8, which is associated with the suppression of NF-κB translocation to the nucleus, also revealed the involvement of the extrinsic pathway in the induced apoptosis. The results suggest that the CdCl2(C14H21N3O2) complex is able to induce the apoptosis of colon cancer cells and is a potential candidate for future cancer studies. PMID:25764970

TRPV2 activation induces apoptotic cell death in human T24 bladder cancer cells: a potential therapeutic target for bladder cancer.

PubMed

Yamada, Takahiro; Ueda, Takashi; Shibata, Yasuhiro; Ikegami, Yosuke; Saito, Masaki; Ishida, Yusuke; Ugawa, Shinya; Kohri, Kenjiro; Shimada, Shoichi

2010-08-01

To investigate the functional expression of the transient receptor potential vanilloid 2 (TRPV2) channel protein in human urothelial carcinoma (UC) cells and to determine whether calcium influx into UC cells through TRPV2 is involved in apoptotic cell death. The expression of TRPV2 mRNA in bladder cancer cell lines (T24, a poorly differentiated UC cell line and RT4, a well-differentiated UC cell line) was analyzed using reverse transcriptase-polymerase chain reaction. The calcium permeability of TRPV2 channels in T24 cells was investigated using a calcium imaging assay that used cannabidiol (CBD), a relatively selective TRPV2 agonist, and ruthenium red (RuR), a nonselective TRPV channel antagonist. The death of T24 or RT4 cells in the presence of CBD was evaluated using a cellular viability assay. Apoptosis of T24 cells caused by CBD was confirmed using an annexin-V assay and small interfering RNA (siRNA) silencing of TRPV2. TRPV2 mRNA was abundantly expressed in T24 cells. The expression level in UC cells was correlated with high-grade disease. The administration of CBD increased intracellular calcium concentrations in T24 cells. In addition, the viability of T24 cells progressively decreased with increasing concentrations of CBD, whereas RT4 cells were mostly unaffected. Cell death occurred via apoptosis caused by continuous influx of calcium through TRPV2. TRPV2 channels in UC cells are calcium-permeable and the regulation of calcium influx through these channels leads directly to the death of UC cells. TRPV2 channels in UC cells may be a potential new therapeutic target, especially in higher-grade UC cells. Copyright 2010 Elsevier Inc. All rights reserved.

Radiation-induced genomic instability and bystander effects: related inflammatory-type responses to radiation-induced stress and injury? A review.

PubMed

Lorimore, S A; Wright, E G

2003-01-01

To review studies of radiation responses in the haemopoietic system in the context of radiation-induced genomic instability, bystander effects and inflammatory-type processes. There is considerable evidence that cells that themselves are not exposed to ionizing radiation but are the progeny of cells irradiated many cell divisions previously may express a high frequency of gene mutations, chromosomal aberrations and cell death. These effects are collectively known as radiation-induced genomic instability. A second untargeted effect results in non-irradiated cells exhibiting responses typically associated with direct radiation exposure but occurs as a consequence of contact with irradiated cells or by receiving soluble signals from irradiated cells. These effects are collectively known as radiation-induced bystander effects. Reported effects include increases or decreases in damage-inducible and stress-related proteins; increases or decreases in reactive oxygen species, cell death or cell proliferation, and induction of mutations and chromosome aberrations. This array of responses is reminiscent of effects mediated by cytokines and other similar regulatory factors that may involve, but do not necessarily require, gap junction-mediated transfer, have multiple inducers and a variety of context-dependent consequences in different cell systems. That chromosomal instability in haemopoietic cells can be induced by an indirect bystander-type mechanism both in vitro and in vivo provides a potential link between these two untargeted effects and there are radiation responses in vivo consistent with the microenvironment contributing secondary cell damage as a consequence of an inflammatory-type response to radiation-induced injury. Intercellular signalling, production of cytokines and free radicals are features of inflammatory responses that have the potential for both bystander-mediated and persisting damage as well as for conferring a predisposition to malignancy. The induction of bystander effects and instabilities may reflect interrelated aspects of a non-specific inflammatory-type response to radiation-induced stress and injury and be involved in a variety of the pathological consequences of radiation exposures.

Lactobacillus delbrueckii subsp lactis CIDCA 133 modulates response of human epithelial and dendritic cells infected with Bacillus cereus.

PubMed

Rolny, I S; Tiscornia, I; Racedo, S M; Pérez, P F; Bollati-Fogolín, M

2016-11-30

It is known that probiotic microorganisms are able to modulate pathogen virulence. This ability is strain dependent and involves multiple interactions between microorganisms and relevant host's cell populations. In the present work we focus on the effect of a potentially probiotic lactobacillus strain (Lactobacillus delbrueckii subsp. lactis CIDCA 133) in an in vitro model of Bacillus cereus infection. Our results showed that infection of intestinal epithelial HT-29 cells by B. cereus induces nuclear factor kappa B (NF-κB) pathway. Noteworthy, the presence of strain L. delbrueckii subsp.lactis CIDCA 133 increases stimulation. However, B. cereus-induced interleukin (IL)-8 production by epithelial cells is partially abrogated by L. delbrueckii subsp. lactis CIDCA 133. These findings suggest that signalling pathways other than that of NF-κB are involved. In a co-culture system (HT-29 and monocyte-derived dendritic cells), B. cereus was able to translocate from the epithelial (upper) to the dendritic cell compartment (lower). This translocation was partially abrogated by the presence of lactobacilli in the upper compartment. In addition, infection of epithelial cells in the co-culture model, led to an increase in the expression of CD86 by dendritic cells. This effect could not be modified in the presence of lactobacilli. Interestingly, infection of enterocytes with B. cereus triggers production of proinflammatory cytokines by dendritic cells (IL-8, IL-6 and tumour necrosis factor alpha (TNF-α)). The production of TNF-α (a protective cytokine in B. cereus infections) by dendritic cells was increased in the presence of lactobacilli. The present work demonstrates for the first time the effect of L. delbrueckii subsp. lactis CIDCA 133, a potentially probiotic strain, in an in vitro model of B. cereus infection. The presence of the probiotic strain modulates cell response both in infected epithelial and dendritic cells thus suggesting a possible beneficial effect of selected lactobacilli strains on the course of B. cereus infection.

The chalcone 2'-hydroxy-4',5'-dimethoxychalcone activates death receptor 5 pathway and leads to apoptosis in human nonsmall cell lung cancer cells.

PubMed

Yang, Lina; Su, Ling; Cao, Congmei; Xu, Linyan; Zhong, Diansheng; Xu, Lijia; Liu, Xiangguo

2013-06-01

Natural chalcones have been proved to inhibit cancer cells with therapeutic potential, but the underlying molecular mechanism is still largely unexplored. Here, we identified a novel chalcone, 2'-hydroxy-4',5'-dimethoxychalcone (HDMC) and demonstrated that HDMC induced apoptosis in various nonsmall cell lung cancer cells. Further study showed that HDMC elevated cellular reactive oxygen species (ROS) levels, thus inducing expressions of ATF4 and C/EBP homologous protein (CHOP). Then, death receptor 5 (DR5) was upregulated through ATF4-CHOP axis and eventually resulted in apoptosis. We also found that downregulation of c-FLIPL contributed to HDMC-induced apoptosis. In conclusion, HDMC induces apoptosis in human nonsmall cell lung cancer cells via activation of DR5 signaling pathway, and ROS-mediated ATF4-CHOP axis is involved in the process. Our results further supported the potential for HDMC to be developed as a new antitumor agent for cancer therapy or chemoprevention. Copyright © 2013 International Union of Biochemistry and Molecular Biology, Inc.

Monoacylglycerol lipase promotes metastases in nasopharyngeal carcinoma.

PubMed

Hu, Wen-Rong; Lian, Yi-Fan; Peng, Li-Xia; Lei, Jin-Ju; Deng, Cheng-Cheng; Xu, Miao; Feng, Qi-Sheng; Chen, Li-Zhen; Bei, Jin-Xin; Zeng, Yi-Xin

2014-01-01

Monoacylglycerol lipase (MAGL) is a serine hydrolase that hydrolyzes monoacylglycerides into free fatty acids and glycerol. It has recently been found to be involved in cancer progression through the free fatty acid or endocannabinoid network after studies on its function in the endocannabinoid system. Here, we determined a role for MAGL in nasopharyngeal carcinoma (NPC), which is known for its high metastatic potential. Among the different NPC cells we tested, MAGL was highly expressed in high metastatic NPC cells, whereas low metastatic potential NPC cells exhibited lower expression of MAGL. Overexpression of MAGL in low metastatic NPC cells enhanced their motile behavior and metastatic capacity in vivo. Conversely, knockdown of MAGL reduced the motility of highly metastatic cells, reducing their metastatic capacity in vivo. Growth rate was not influenced by MAGL in either high or low metastatic cells. MAGL expression was associated with the epithelial-mesenchymal transition (EMT) proteins, such as E-cadherin, vimentin and Snail. It was also related to the sidepopulation (SP) of NPC cells. Our findings establish that MAGL promotes metastases in NPC through EMT, and it may serve as a target for the prevention of NPC metastases.

Life and death of female gametes during oogenesis and folliculogenesis.

PubMed

Krysko, Dmitri V; Diez-Fraile, Araceli; Criel, Godelieve; Svistunov, Andrei A; Vandenabeele, Peter; D'Herde, Katharina

2008-09-01

The vertebrate ovary is an extremely dynamic organ in which excessive or defective follicles are rapidly and effectively eliminated early in ontogeny and thereafter continuously throughout reproductive life. More than 99% of follicles disappear, primarily due to apoptosis of granulosa cells, and only a minute fraction of the surviving follicles successfully complete the path to ovulation. The balance between signals for cell death and survival determines the destiny of the follicles. An abnormally high rate of cell death followed by atresia can negatively affect fertility and eventually lead irreversibly to premature ovarian failure. In this review we provide a short overview of the role of programmed cell death in prenatal differentiation of the primordial germ cells and in postnatal folliculogenesis. We also discuss the issue of neo-oogenesis. Next, we highlight molecules involved in regulation of granulosa cell apoptosis. We further discuss the potential use of scores for apoptosis in granulosa cells and characteristics of follicular fluid as prognostic markers for predicting the outcome of assisted reproduction. Potential therapeutic strategies for combating premature ovarian failure are also addressed.

CD6 as a potential target for treating multiple sclerosis

PubMed Central

Singer, Nora G.; Whitbred, Joy; Bowen, Michael A.; Lin, Feng

2017-01-01

CD6 was established as a marker of T cells more than three decades ago, and recent studies have identified CD6 as a risk gene for multiple sclerosis (MS), a disease in which autoreactive T cells are integrally involved. Nevertheless, the precise role of CD6 in regulating T-cell responses is controversial and its significance in the pathogenesis of various diseases remains elusive, partly due to the lack of animals engineered to alter expression of the CD6 gene. In this report, we found that CD6 KO mice showed decreased pathogenic T-cell responses, reduced spinal cord T-cell infiltration, and attenuated disease severity in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. CD6-deficient T cells exhibited augmented activation, but also significantly reduced survival and proliferation after activation, leading to overall decreased Th1 and Th17 polarization. Activated CD6-deficient T cells also showed impaired infiltration through brain microvascular endothelial cell monolayers. Furthermore, by developing CD6 humanized mice, we identified a mouse anti-human CD6 monoclonal antibody that is highly effective in treating established EAE without depleting T cells. These results suggest that (i) CD6 is a negative regulator of T-cell activation, (ii) at the same time, CD6 is a positive regulator of activated T-cell survival/proliferation and infiltration; and (iii) CD6 is a potential new target for treating MS and potentially other T-cell–driven autoimmune conditions. PMID:28209777

3,3'-diindolylmethane potentiates tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis of gastric cancer cells.

PubMed

Ye, Yang; Miao, Shuhan; Wang, Yan; Zhou, Jianwei; Lu, Rongzhu

2015-05-01

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) specifically kills cancer cells without destroying the majority of healthy cells. However, numerous types of cancer cell, including gastric cancer cells, tend to be resistant to TRAIL. The bioactive product 3,3'-diindolylmethane (DIM), which is derived from cruciferous vegetables, is also currently recognized as a candidate anticancer agent. In the present study, a Cell Counting Kit 8 cell growth assay and an Annexin V-fluorescein isothiocyanate apoptosis assay were performed to investigate the potentiating effect of DIM on TRAIL-induced apoptosis in gastric cancer cells, and the possible mechanisms of this potentiation. The results obtained demonstrated that, compared with TRAIL or DIM treatment alone, co-treatment with TRAIL (25 or 50 ng/ml) and DIM (10 µmol/l) induced cytotoxic and apoptotic effects in BGC-823 and SGC-7901 gastric cancer cells. Furthermore, western blot analysis revealed that the protein expression levels of death receptor 5 (DR5), CCAAT/enhancer binding protein homologous protein (CHOP) and glucose-regulated protein 78 (GRP78) were upregulated in the co-treated gastric cancer cells. To the best of our knowledge, the present study is the first to provide evidence that DIM sensitizes TRAIL-induced inhibition of proliferation and apoptosis in gastric cancer cells, accompanied by the upregulated expression of DR5, CHOP and GRP78 proteins, which may be involved in endoplasmic reticulum stress mechanisms.

Molecular mechanism of cardol, isolated from Trigona incisa stingless bee propolis, induced apoptosis in the SW620 human colorectal cancer cell line.

PubMed

Kustiawan, Paula Mariana; Lirdprapamongkol, Kriengsak; Palaga, Tanapat; Puthong, Songchan; Phuwapraisirisan, Preecha; Svasti, Jisnuson; Chanchao, Chanpen

2017-05-04

Cardol is a major bioactive constituent in the Trigona incisa propolis from Indonesia, with a strong in vitro antiproliferative activity against the SW620 colorectal adenocarcinoma cell line (IC 50 of 4.51 ± 0.76 μg/mL). Cardol induced G 0 /G 1 cell cycle arrest and apoptotic cell death. The present study was designed to reveal the mechanism of cardol's antiproliferative effect and induction of apoptosis. Changes in cell morphology were observed by light microscopy. To determine whether the mitochondrial apoptotic pathway was involved in cell death, caspase-3 and caspase-9 activities, western blot analysis, mitochondrial membrane potential, and intracellular reactive oxygen species (ROS) levels were assayed. Changes in the cell morphology and the significantly increased caspase-3 and caspase-9 activities, plus the cleavage of pro-caspase-3, pro-caspase-9 and PARP, supported that cardol caused apoptosis in SW620 cells within 2 h after treatment by cardol. In addition, cardol decreased the mitochondrial membrane potential while increasing the intracellular ROS levels in a time- and dose-dependent manner. Antioxidant treatment supported that the cardol-induced cell death was dependent on ROS production. Cardol induced cell death in SW620 cells was mediated by oxidative stress elevation and the mitochondrial apoptotic pathway, and these could be the potential molecular mechanism for the antiproliferative effect of cardol.

Red blood cell aggregation, aggregate strength and oxygen transport potential of blood are abnormal in both homozygous sickle cell anemia and sickle-hemoglobin C disease.

PubMed

Tripette, Julien; Alexy, Tamas; Hardy-Dessources, Marie-Dominique; Mougenel, Daniele; Beltan, Eric; Chalabi, Tawfik; Chout, Roger; Etienne-Julan, Maryse; Hue, Olivier; Meiselman, Herbert J; Connes, Philippe

2009-08-01

Recent evidence suggests that red blood cell aggregation and the ratio of hematocrit to blood viscosity (HVR), an index of the oxygen transport potential of blood, might considerably modulate blood flow dynamics in the microcirculation. It thus seems likely that these factors could play a role in sickle cell disease. We compared red blood cell aggregation characteristics, blood viscosity and HVR at different shear rates between sickle cell anemia and sickle cell hemoglobin C disease (SCC) patients, sickle cell trait carriers (AS) and control individuals (AA). Blood viscosity determined at high shear rate was lower in sickle cell anemia (n=21) than in AA (n=52), AS (n=33) or SCC (n=21), and was markedly increased in both SCC and AS. Despite differences in blood viscosity, both sickle cell anemia and SCC had similar low HVR values compared to both AA and AS. Sickle cell anemia (n=21) and SCC (n=19) subjects had a lower red blood cell aggregation index and longer time for red blood cell aggregates formation than AA (n=16) and AS (n=15), and a 2 to 3 fold greater shear rate required to disperse red blood cell aggregates. The low HVR levels found in sickle cell anemia and SCC indicates a comparable low oxygen transport potential of blood in both genotypes. Red blood cell aggregation properties are likely to be involved in the pathophysiology of sickle cell disease: the increased shear forces needed to disperse red blood cell aggregates may disturb blood flow, especially at the microcirculatory level, since red blood cell are only able to pass through narrow capillaries as single cells rather than as aggregates.

Inhibitory Effects of Leptin on Pancreatic α-Cell Function

PubMed Central

Tudurí, Eva; Marroquí, Laura; Soriano, Sergi; Ropero, Ana B.; Batista, Thiago M.; Piquer, Sandra; López-Boado, Miguel A.; Carneiro, Everardo M.; Gomis, Ramón; Nadal, Angel; Quesada, Ivan

2009-01-01

OBJECTIVE Leptin released from adipocytes plays a key role in the control of food intake, energy balance, and glucose homeostasis. In addition to its central action, leptin directly affects pancreatic β-cells, inhibiting insulin secretion, and, thus, modulating glucose homeostasis. However, despite the importance of glucagon secretion in glucose homeostasis, the role of leptin in α-cell function has not been studied in detail. In the present study, we have investigated this functional interaction. RESEARCH DESIGN AND METHODS The presence of leptin receptors (ObR) was demonstrated by RT-PCR analysis, Western blot, and immunocytochemistry. Electrical activity was analyzed by patch-clamp and Ca2+ signals by confocal microscopy. Exocytosis and glucagon secretion were assessed using fluorescence methods and radioimmunoassay, respectively. RESULTS The expression of several ObR isoforms (a–e) was detected in glucagon-secreting αTC1-9 cells. ObRb, the main isoform involved in leptin signaling, was identified at the protein level in αTC1-9 cells as well as in mouse and human α-cells. The application of leptin (6.25 nmol/l) hyperpolarized the α-cell membrane potential, suppressing the electrical activity induced by 0.5 mmol/l glucose. Additionally, leptin inhibited Ca2+ signaling in αTC1-9 cells and in mouse and human α-cells within intact islets. A similar result occurred with 0.625 nmol/l leptin. These effects were accompanied by a decrease in glucagon secretion from mouse islets and were counteracted by the phosphatidylinositol 3-kinase inhibitor, wortmannin, suggesting the involvement of this pathway in leptin action. CONCLUSIONS These results demonstrate that leptin inhibits α-cell function, and, thus, these cells are involved in the adipoinsular communication. PMID:19401420

Involvement of Bim in Photofrin-mediated photodynamically induced apoptosis.

PubMed

Wang, Xianwang; He, Xiaobing; Hu, Shujuan; Sun, Anbang; Lu, Chengbiao

2015-01-01

Photodynamic therapy (PDT) is a promising noninvasive technique, which has been successfully applied to the treatment of human cancers. Studies have shown that the Bcl-2 family proteins play important roles in PDT-induced apoptosis. However, whether Bcl-2-interacting mediator of cell death (Bim) is involved in photodynamic treatment remains unknown. In this study, we attempt to determine the effect of Bim on Photofrin photodynamic treatment (PPT)-induced apoptosis in human lung adenocarcinoma ASTC-a-1 cells. The translocation of Bim/Bax of the cells were monitored by laser confocal scanning microscope. The levels of Bim protein and activated caspase-3 in cells were detected by western blot assay. Caspase-3 activities were measured by Caspase-3 Fluorogenic Substrate (Ac-DEVD-AFC) analysis. The induction of apoptosis was detected by Hoechst 33258 and PI staining as well as flow cytometry analysis. The effect of Bim on PPT-induced apoptosis was determined by RNAi. BimL translocated to mitochondria in response to PPT, similar to the downstream pro-apoptotic protein Bax activation. PPT increased the level of Bim and activated caspase-3 in cells and that knockdown of Bim by RNAi significantly protected against caspase-3 activity. PPT-induced apoptosis were suppressed in cells transfected with shRNA-Bim. We demonstrated the involvement of Bim in PPT-induced apoptosis in human ASTC-a-1 lung adenocarcinoma cells and suggested that enhancing Bim activity might be a potential strategy for treating human cancers. © 2015 S. Karger AG, Basel.

Modulation of gene expression and cell cycle by botryosphaeran, a (1→3)(1→6)-β-d-glucan in human lymphocytes.

PubMed

Malini, Maressa; Souza, Marilesia Ferreira de; Oliveira, Marcelo Tempesta de; Antunes, Lusânia Maria Greggi; Figueiredo, Suely Gomes de; Barbosa, Aneli M; Dekker, Robert F H; Cólus, Ilce Mara de Syllos

2015-01-01

There is growing interest in the anticancer and immunomodulatory potential of fungal β-d-glucans. In the present study, the modulation of gene expression via RT-qPCR and cell cycle kinetics via flow cytometry were assessed in human normal and tumor (Jurkat) lymphocytes after treatment with botryosphaeran (a fungal (1→3)(1→6)-β-d-glucan) from Botryosphaeria rhodina MAMB-05. Cell cultures were treated with botryosphaeran either alone, or in combination with doxorubicin (DXR), in a post-treatment protocol. The expression of genes involved in immunomodulatory processes, apoptosis and cell cycle control, as well as β-d-glucans cell receptors were assessed. Flow cytometry analysis identified tetraploid Jurkat cells in G1 phase when treated with botryosphaeran combined with DXR. This antiproliferative effect in G1 may be associated with down-regulation of the expression of genes involved in the G1 checkpoint. The repression of the CCR5 gene following botryosphaeran treatment, either alone or in combination with DXR, in tumor lymphocytes indicates a possible affinity of this particular (1→3)(1→6)-β-d-glucan for the receptor CCR5. Therefore, botryosphaeran action appears to be involved in the repression of genes related to the G1 phase of the cell cycle and possibly in the interaction of the botryosphaeran, either alone, or in combination with DXR, with the CCR5 receptor. Copyright © 2015 Elsevier B.V. All rights reserved.

Intracellular recordings of action potentials by an extracellular nanoscale field-effect transistor.

PubMed

Duan, Xiaojie; Gao, Ruixuan; Xie, Ping; Cohen-Karni, Tzahi; Qing, Quan; Choe, Hwan Sung; Tian, Bozhi; Jiang, Xiaocheng; Lieber, Charles M

2011-12-18

The ability to make electrical measurements inside cells has led to many important advances in electrophysiology. The patch clamp technique, in which a glass micropipette filled with electrolyte is inserted into a cell, offers both high signal-to-noise ratio and temporal resolution. Ideally, the micropipette should be as small as possible to increase the spatial resolution and reduce the invasiveness of the measurement, but the overall performance of the technique depends on the impedance of the interface between the micropipette and the cell interior, which limits how small the micropipette can be. Techniques that involve inserting metal or carbon microelectrodes into cells are subject to similar constraints. Field-effect transistors (FETs) can also record electric potentials inside cells, and because their performance does not depend on impedance, they can be made much smaller than micropipettes and microelectrodes. Moreover, FET arrays are better suited for multiplexed measurements. Previously, we have demonstrated FET-based intracellular recording with kinked nanowire structures, but the kink configuration and device design places limits on the probe size and the potential for multiplexing. Here, we report a new approach in which a SiO2 nanotube is synthetically integrated on top of a nanoscale FET. This nanotube penetrates the cell membrane, bringing the cell cytosol into contact with the FET, which is then able to record the intracellular transmembrane potential. Simulations show that the bandwidth of this branched intracellular nanotube FET (BIT-FET) is high enough for it to record fast action potentials even when the nanotube diameter is decreased to 3 nm, a length scale well below that accessible with other methods. Studies of cardiomyocyte cells demonstrate that when phospholipid-modified BIT-FETs are brought close to cells, the nanotubes can spontaneously penetrate the cell membrane to allow the full-amplitude intracellular action potential to be recorded, thus showing that a stable and tight seal forms between the nanotube and cell membrane. We also show that multiple BIT-FETs can record multiplexed intracellular signals from both single cells and networks of cells.

Intracellular recordings of action potentials by an extracellular nanoscale field-effect transistor

NASA Astrophysics Data System (ADS)

Duan, Xiaojie; Gao, Ruixuan; Xie, Ping; Cohen-Karni, Tzahi; Qing, Quan; Choe, Hwan Sung; Tian, Bozhi; Jiang, Xiaocheng; Lieber, Charles M.

2012-03-01

The ability to make electrical measurements inside cells has led to many important advances in electrophysiology. The patch clamp technique, in which a glass micropipette filled with electrolyte is inserted into a cell, offers both high signal-to-noise ratio and temporal resolution. Ideally, the micropipette should be as small as possible to increase the spatial resolution and reduce the invasiveness of the measurement, but the overall performance of the technique depends on the impedance of the interface between the micropipette and the cell interior, which limits how small the micropipette can be. Techniques that involve inserting metal or carbon microelectrodes into cells are subject to similar constraints. Field-effect transistors (FETs) can also record electric potentials inside cells, and because their performance does not depend on impedance, they can be made much smaller than micropipettes and microelectrodes. Moreover, FET arrays are better suited for multiplexed measurements. Previously, we have demonstrated FET-based intracellular recording with kinked nanowire structures, but the kink configuration and device design places limits on the probe size and the potential for multiplexing. Here, we report a new approach in which a SiO2 nanotube is synthetically integrated on top of a nanoscale FET. This nanotube penetrates the cell membrane, bringing the cell cytosol into contact with the FET, which is then able to record the intracellular transmembrane potential. Simulations show that the bandwidth of this branched intracellular nanotube FET (BIT-FET) is high enough for it to record fast action potentials even when the nanotube diameter is decreased to 3 nm, a length scale well below that accessible with other methods. Studies of cardiomyocyte cells demonstrate that when phospholipid-modified BIT-FETs are brought close to cells, the nanotubes can spontaneously penetrate the cell membrane to allow the full-amplitude intracellular action potential to be recorded, thus showing that a stable and tight seal forms between the nanotube and cell membrane. We also show that multiple BIT-FETs can record multiplexed intracellular signals from both single cells and networks of cells.

Complete human serum maintains viability and chondrogenic potential of human synovial stem cells: suitable conditions for transplantation.

PubMed

Mizuno, Mitsuru; Katano, Hisako; Otabe, Koji; Komori, Keiichiro; Kohno, Yuji; Fujii, Shizuka; Ozeki, Nobutake; Horie, Masafumi; Tsuji, Kunikazu; Koga, Hideyuki; Muneta, Takeshi; Sekiya, Ichiro

2017-06-13

In our clinical practice, we perform transplantations of autologous synovial mesenchymal stem cells (MSCs) for cartilage and meniscus regenerative medicine. One of the most important issues to ensuring clinical efficacy involves the transport of synovial MSCs from the processing facility to the clinic. Complete human serum (100% human serum) is an attractive candidate material in which to suspend synovial MSCs for their preservation during transport. The purpose of this study was to investigate whether complete human serum maintained MSC viability and chondrogenic potential and to examine the optimal temperature conditions for the preservation of human synovial MSCs. Human synovium was harvested from the knees of 14 donors with osteoarthritis during total knee arthroplasty. Passage 2 synovial MSCs were suspended at 2 million cells/100 μL in Ringer's solution or complete human serum at 4, 13, and 37 °C for 48 h. These cells were analyzed for live cell rates, cell surface marker expression, metabolic activity, proliferation, and adipogenic, calcification, and chondrogenic differentiation potentials before and after preservation. After preservation, synovial MSCs maintained higher live cell rates in human serum than in Ringer's solution at 4 and 13 °C. Synovial MSCs preserved in human serum at 4 and 13 °C also maintained high ratios of propidium iodide - and annexin V - cells. MSC surface marker expression was not altered in cells preserved at 4 and 13 °C. The metabolic activities of cells preserved in human serum at 4 and 13 °C was maintained, while significantly reduced in other conditions. Replated MSCs retained their proliferation ability when preserved in human serum at 4 and 13 °C. Adipogenesis and calcification potential could be observed in cells preserved in each condition, whereas chondrogenic potential was retained only in cells preserved in human serum at 4 and 13 °C. The viability and chondrogenic potential of synovial MSCs were maintained when the cells were suspended in human serum at 4 and 13 °C.

Effects of inhibitors of vascular endothelial growth factor receptor 2 and downstream pathways of receptor tyrosine kinases involving phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin or mitogen-activated protein kinase in canine hemangiosarcoma cell lines.

PubMed

Adachi, Mami; Hoshino, Yuki; Izumi, Yusuke; Sakai, Hiroki; Takagi, Satoshi

2016-07-01

Canine hemangiosarcoma (HSA) is a progressive malignant neoplasm with no current effective treatment. Previous studies showed that receptor tyrosine kinases and molecules within their downstream pathways involving phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (m-TOR) or mitogen-activated protein kinase (MAPK) were overexpressed in canine, human, and murine tumors, including HSA. The present study investigated the effects of inhibitors of these pathways in canine splenic and hepatic HSA cell lines using assays of cell viability and apoptosis. Inhibitors of the MAPK pathway did not affect canine HSA cell viability. However, cell viability was significantly reduced by exposure to inhibitors of vascular endothelial growth factor receptor 2 and the PI3K/Akt/m-TOR pathway; these inhibitors also induced apoptosis in these cell lines. These results suggest that these inhibitors reduce the proliferation of canine HSA cells by inducing apoptosis. Further study of these inhibitors, using xenograft mouse models of canine HSA, are warranted to explore their potential for clinical application.

Effects of inhibitors of vascular endothelial growth factor receptor 2 and downstream pathways of receptor tyrosine kinases involving phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin or mitogen-activated protein kinase in canine hemangiosarcoma cell lines

PubMed Central

Adachi, Mami; Hoshino, Yuki; Izumi, Yusuke; Sakai, Hiroki; Takagi, Satoshi

2016-01-01

Canine hemangiosarcoma (HSA) is a progressive malignant neoplasm with no current effective treatment. Previous studies showed that receptor tyrosine kinases and molecules within their downstream pathways involving phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (m-TOR) or mitogen-activated protein kinase (MAPK) were overexpressed in canine, human, and murine tumors, including HSA. The present study investigated the effects of inhibitors of these pathways in canine splenic and hepatic HSA cell lines using assays of cell viability and apoptosis. Inhibitors of the MAPK pathway did not affect canine HSA cell viability. However, cell viability was significantly reduced by exposure to inhibitors of vascular endothelial growth factor receptor 2 and the PI3K/Akt/m-TOR pathway; these inhibitors also induced apoptosis in these cell lines. These results suggest that these inhibitors reduce the proliferation of canine HSA cells by inducing apoptosis. Further study of these inhibitors, using xenograft mouse models of canine HSA, are warranted to explore their potential for clinical application. PMID:27408334

Fuel Cells: Power System Option for Space Research

NASA Astrophysics Data System (ADS)

Shaneeth, M.; Mohanty, Surajeet

2012-07-01

Fuel Cells are direct energy conversion devices and, thereby, they deliver electrical energy at very high efficiency levels. Hydrogen and Oxygen gases are electrochemically processed, producing clean electric power with water as the only by product. A typical, Fuel Cell based power system involve a Electrochemical power converter, gas storage and management systems, thermal management systems and relevant control units. While there exists different types of Fuel cells, Proton Exchange Membrane (PEM) Fuel Cells are considered as the most suitable one for portable applications. Generally, Fuel Cells are considered as the primary power system option in space missions requiring high power ( > 5kW) and long durations and also where water is a consumable, such as manned missions. This is primarily due to the advantage that fuel cell based power systems offer, in terms of specific energy. Fuel cells have the potential to attain specific energy > 500Wh/kg, specific power >500W/kg, energy density > 400Whr/L and also power density > 200 W/L. This apart, a fuel cell system operate totally independent of sun light, whereas as battery based system is fully dependent on the same. This uniqueness provides added flexibility and capabilities to the missions and modularity for power system. High power requiring missions involving reusable launch vehicles, manned missions etc are expected to be richly benefited from this. Another potential application of Fuel Cell would be interplanetary exploration. Unpredictable and dusty atmospheres of heavenly bodies limits sun light significantly and there fuel cells of different types, eg, Bio-Fuel Cells, PEMFC, DMFCs would be able to work effectively. Manned or unmanned lunar out post would require continuous power even during extra long lunar nights and high power levels are expected. Regenerative Fuel Cells, a combination of Fuel Cells and Electrolysers, are identified as strong candidate. While application of Fuel Cells in high power requiring missions is well established, as exemplified in Apollo and Space Shuttles, use in low power missions for science probes/rovers form a relatively newer area. Low power small fuel cells of this class are expected to bring in lot of operational convenience and freedom on onboard / extra terrestrial environment. Technological improvisations in the area, especially with regard to miniaturisation, and extra capabilities that the system offers, make it a strong candidate. The paper outlines features of fuel cells power systems, different types and their potential application scenarios, in the present context. It elucidates the extra capabilities and advantages, due to fuel cells, for different missions. Specific case analyses are also included.

A novel molten-salt electrochemical cell for investigating the reduction of uranium dioxide to uranium metal by lithium using in situ synchrotron radiation

PubMed Central

Brown, Leon D.; Abdulaziz, Rema; Jervis, Rhodri; Bharath, Vidal; Mason, Thomas J.; Reinhard, Christina; Connor, Leigh D.; Inman, Douglas; Brett, Daniel J. L.; Shearing, Paul R.

2017-01-01

A novel electrochemical cell has been designed and built to allow for in situ energy-dispersive X-ray diffraction measurements to be made during reduction of UO2 to U metal in LiCl–KCl at 500°C. The electrochemical cell contains a recessed well at the bottom of the cell into which the working electrode sits, reducing the beam path for the X-rays through the molten-salt and maximizing the signal-to-noise ratio from the sample. Lithium metal was electrodeposited onto the UO2 working electrode by exposing the working electrode to more negative potentials than the Li deposition potential of the LiCl–KCl eutectic electrolyte. The Li metal acts as a reducing agent for the chemical reduction of UO2 to U, which appears to proceed to completion. All phases were fitted using Le Bail refinement. The cell is expected to be widely applicable to many studies involving molten-salt systems. PMID:28244437

A novel molten-salt electrochemical cell for investigating the reduction of uranium dioxide to uranium metal by lithium using in situ synchrotron radiation.

PubMed

Brown, Leon D; Abdulaziz, Rema; Jervis, Rhodri; Bharath, Vidal; Mason, Thomas J; Atwood, Robert C; Reinhard, Christina; Connor, Leigh D; Inman, Douglas; Brett, Daniel J L; Shearing, Paul R

2017-03-01

A novel electrochemical cell has been designed and built to allow for in situ energy-dispersive X-ray diffraction measurements to be made during reduction of UO 2 to U metal in LiCl-KCl at 500°C. The electrochemical cell contains a recessed well at the bottom of the cell into which the working electrode sits, reducing the beam path for the X-rays through the molten-salt and maximizing the signal-to-noise ratio from the sample. Lithium metal was electrodeposited onto the UO 2 working electrode by exposing the working electrode to more negative potentials than the Li deposition potential of the LiCl-KCl eutectic electrolyte. The Li metal acts as a reducing agent for the chemical reduction of UO 2 to U, which appears to proceed to completion. All phases were fitted using Le Bail refinement. The cell is expected to be widely applicable to many studies involving molten-salt systems.

Novel Passivating/Antireflective Coatings for Space Solar Cells

NASA Technical Reports Server (NTRS)

Faur, Mircea; Faur, Maria; Bailey, S. G.; Flood, D. J.; Faur, H. M.; Mateescu, C. G.; Alterovitz, S. A.; Scheiman, D.; Jenkins, P. P.; Brinker, D. J.

2005-01-01

We are developing a novel process to grow passivating/antireflective (AR) coatings for terrestrial and space solar cells. Our approach involves a Room Temperature Wet Chemical Growth (RTWCG) process, which was pioneered, and is under development at SPECMAT, Inc., under a Reimbursable Space Act Agreement with NASA Glenn Research Center. The RTWCG passivating/AR coatings with graded index of refraction are applied in one easy step on finished (bare) cells. The RTWCG coatings grown on planar, textured and porous Si, as well as on poly-Si, CuInSe2, and III-V substrates, show excellent uniformity irrespective of surface topography, crystal orientation, size and shape. In this paper we present some preliminary results of the RTWCG coatings on Si and III-V substrates that show very good potential for use as a passivation/AR coating for space solar cell applications. Compared to coatings grown using conventional techniques, the RTWCG coatings have the potential to reduce reflection losses and improve current collection near the illuminated surface of space solar cells, while reducing the fabrication costs.

Labeling Human Mesenchymal Stem Cells with Gold Nanocages for in vitro and in vivo Tracking by Two-Photon Microscopy and Photoacoustic Microscopy

PubMed Central

Zhang, Yu Shrike; Wang, Yu; Wang, Lidai; Wang, Yucai; Cai, Xin; Zhang, Chi; Wang, Lihong V.; Xia, Younan

2013-01-01

Stem cell tracking is a highly important subject. Current techniques based on nanoparticle-labeling, such as magnetic resonance imaging, fluorescence microscopy, and micro-computed tomography, are plagued by limitations including relatively low sensitivity or penetration depth, involvement of ionizing irradiation, and potential cytotoxicity of the nanoparticles. Here we introduce a new class of contrast agents based on gold nanocages (AuNCs) with hollow interiors and porous walls to label human mesenchymal stem cells (hMSCs) for both in vitro and in vivo tracking using two-photon microscopy and photoacoustic microscopy. As demonstrated by the viability assay, the AuNCs showed negligible cytotoxicity under a reasonable dose, and did not alter the differentiation potential of the hMSCs into desired lineages. We were able to image the cells labeled with AuNCs in vitro for at least 28 days in culture, as well as to track the cells that homed to the tumor region in nude mice in vivo. PMID:23946820

Co-culture systems and technologies: taking synthetic biology to the next level.

PubMed

Goers, Lisa; Freemont, Paul; Polizzi, Karen M

2014-07-06

Co-culture techniques find myriad applications in biology for studying natural or synthetic interactions between cell populations. Such techniques are of great importance in synthetic biology, as multi-species cell consortia and other natural or synthetic ecology systems are widely seen to hold enormous potential for foundational research as well as novel industrial, medical and environmental applications with many proof-of-principle studies in recent years. What is needed for co-cultures to fulfil their potential? Cell-cell interactions in co-cultures are strongly influenced by the extracellular environment, which is determined by the experimental set-up, which therefore needs to be given careful consideration. An overview of existing experimental and theoretical co-culture set-ups in synthetic biology and adjacent fields is given here, and challenges and opportunities involved in such experiments are discussed. Greater focus on foundational technology developments for co-cultures is needed for many synthetic biology systems to realize their potential in both applications and answering biological questions. © 2014 The Author(s) Published by the Royal Society. All rights reserved.

In Vitro Activity of Lactobacilli with Probiotic Potential Isolated from Cocoa Fermentation against Gardnerella vaginalis

PubMed Central

Pessoa, Wallace Felipe Blohem; Melgaço, Ana Clara Correia; Ramos, Louise Pereira; Rezende, Rachel Passos

2017-01-01

Study of the probiotic potential of microorganisms isolated from fermented foods has been increasing, especially studies related to lactobacilli. In intestinal models, lactobacilli have demonstrated beneficial properties, such as anti-inflammatory activity and increased antibody production, but the molecular mechanisms involving probiotic and antagonistic action as well as their effect on human vaginal cells have not yet been fully elucidated. The aim of this study was to evaluate the functional and antagonistic properties of three strains of lactobacilli isolated from cocoa fermentation (Lactobacillus fermentum 5.2, L. plantarum 6.2, and L. plantarum 7.1) against Gardnerella vaginalis. Our results show that the lactobacilli have potential use as probiotics, since they have high hydrophobicity and autoaggregation properties and effectively adhere to vaginal cells. Metabolites secreted into the culture medium and whole cells of the strains under study are capable of interfering with the growth of G. vaginalis to different degrees. The elucidation of the antagonistic mechanisms as well as their effect on human cells may be useful in the development of a product containing such microorganisms or products secreted by them. PMID:29226130

Neuron membrane trafficking and protein kinases involved in autism and ADHD.

PubMed

Kitagishi, Yasuko; Minami, Akari; Nakanishi, Atsuko; Ogura, Yasunori; Matsuda, Satoru

2015-01-30

A brain-enriched multi-domain scaffolding protein, neurobeachin has been identified as a candidate gene for autism patients. Mutations in the synaptic adhesion protein cell adhesion molecule 1 (CADM1) are also associated with autism spectrum disorder, a neurodevelopmental disorder of uncertain molecular origin. Potential roles of neurobeachin and CADM1 have been suggested to a function of vesicle transport in endosomal trafficking. It seems that protein kinase B (AKT) and cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) have key roles in the neuron membrane trafficking involved in the pathogenesis of autism. Attention deficit hyperactivity disorder (ADHD) is documented to dopaminergic insufficiencies, which is attributed to synaptic dysfunction of dopamine transporter (DAT). AKT is also essential for the DAT cell-surface redistribution. In the present paper, we summarize and discuss the importance of several protein kinases that regulate the membrane trafficking involved in autism and ADHD, suggesting new targets for therapeutic intervention.

Aqueous Ethanolic Extract of Tinospora cordifolia as a Potential Candidate for Differentiation Based Therapy of Glioblastomas

PubMed Central

Mishra, Rachana; Kaur, Gurcharan

2013-01-01

Glioblastomas are the most aggressive primary brain tumors and their heterogeneity and complexity often renders them non responsive to various conventional treatments. Search for herbal products having potential anti-cancer activity is an active area of research in the Indian traditional system of medicine i.e., Ayurveda. Tinospora cordifolia, also named as ‘heavenly elixir’ is used in various ayurvedic decoctions as panacea to treat several body ailments. The current study investigated the anti-brain cancer potential of 50% ethanolic extract of Tinospora cordifolia (TCE) using C6 glioma cells. TCE significantly reduced cell proliferation in dose-dependent manner and induced differentiation in C6 glioma cells, resulting in astrocyte-like morphology as indicated by phase contrast images, GFAP expression and process outgrowth data of TCE treated cells which exhibited higher number and longer processes than untreated cells. Reduced proliferation of cells was accompanied by enhanced expression of senescence marker, mortalin and its translocation from perinuclear to pancytoplasmic spaces. Further, TCE showed anti-migratory and anti-invasive potential as depicted by wound scratch assay and reduced expression of plasticity markers NCAM and PSA-NCAM along with MMP-2 and 9. On analysis of the cell cycle and apoptotic markers, TCE treatment was seen to arrest the C6 cells in G0/G1 and G2/M phase, suppressing expression of G1/S phase specific protein cyclin D1 and anti-apoptotic protein Bcl-xL, thus supporting its anti-proliferative and apoptosis inducing potential. Present study provides the first evidence for the presence of anti-proliferative, differentiation-inducing and anti-migratory/anti-metastatic potential of TCE in glioma cells and possible signaling pathways involved in its mode of action. Our primary data suggests that TCE and its active components may prove to be promising phytotherapeutic interventions in gliobalstoma multiformae.  PMID:24205314

Streptomyces artemisiae MCCB 248 isolated from Arctic fjord sediments has unique PKS and NRPS biosynthetic genes and produces potential new anticancer natural products.

PubMed

Dhaneesha, M; Benjamin Naman, C; Krishnan, K P; Sinha, Rupesh Kumar; Jayesh, P; Joseph, Valsamma; Bright Singh, I S; Gerwick, William H; Sajeevan, T P

2017-05-01

After screening marine actinomycetes isolated from sediment samples collected from the Arctic fjord Kongsfjorden for potential anticancer activity, an isolate identified as Streptomyces artemisiae MCCB 248 exhibited promising results against the NCI-H460 human lung cancer cell line. H460 cells treated with the ethyl acetate extract of strain MCCB 248 and stained with Hoechst 33342 showed clear signs of apoptosis, including shrinkage of the cell nucleus, DNA fragmentation and chromatin condensation. Further to this treated cells showed indications of early apoptotic cell death, including a significant proportion of Annexin V positive staining and evidence of DNA damage as observed in the TUNEL assay. Amplified PKS 1 and NRPS genes involved in secondary metabolite production showed only 82% similarity to known biosynthetic genes of Streptomyces, indicating the likely production of a novel secondary metabolite in this extract. Additionally, chemical dereplication efforts using LC-MS/MS molecular networking suggested the presence of a series of undescribed tetraene polyols. Taken together, these results revealed that this Arctic S. artemisiae strain MCCB 248 is a promising candidate for natural products drug discovery and genome mining for potential anticancer agents.

Applications of Mesenchymal Stem Cells in Sinus Lift Augmentation as a Dental Implant Technology.

PubMed

Parnia, Feridoun; Yazdani, Javad; Maleki Dizaj, Solmaz

2018-01-01

The potential application of stem cell biology in human dentistry is a new and emerging field of research. The objective of the current review was to study the efficiency of mesenchymal stem cells (MSCs) in sinus lift augmentation (SLA). A literature review was performed in PubMed Central using MeSH keywords such as sinus lift, MSCs, dental implants, and augmentation. The searches involved full-text papers written in English, published in the past 10 years (2007-2017). The review included in vitro and in vivo studies on the use of MSCs in SLA. Electronic searching provided 45 titles, and among them, 8 papers were chosen as suitable based on the inclusion requirements of this review. The reviewed studies have revealed the potential of MSCs in SLA. According to these papers, stem cell therapy combined with different biomaterials may considerably improve bone regeneration in previous steps of dental implantation and may veritably lead to efficient clinical usages in the recent future. However, the identification of an ideal source of stem cells as well as long-term studies is vital to assess the success rate of this technology. Further clinical trials are also needed to approve the potential of MSCs in SLA.

Purification and immunolocalization of an annexin-like protein in pea seedlings

NASA Technical Reports Server (NTRS)

Clark, G. B.; Dauwalder, M.; Roux, S. J.

1992-01-01

As part of a study to identify potential targets of calcium action in plant cells, a 35-kDa, annexin-like protein was purified from pea (Pisum sativum L.) plumules by a method used to purify animal annexins. This protein, called p35, binds to a phosphatidylserine affinity column in a calcium-dependent manner and binds 45Ca2+ in a dot-blot assay. Preliminary sequence data confirm a relationship for p35 with the annexin family of proteins. Polyclonal antibodies have been raised which recognize p35 in Western and dot blots. Immunofluorescence and immunogold techniques were used to study the distribution and subcellular localization of p35 in pea plumules and roots. The highest levels of immunostain were found in young developing vascular cells producing wall thickenings and in peripheral root-cap cells releasing slime. This localization in cells which are actively involved in secretion is of interest because one function suggested for the animal annexins is involvement in the mediation of exocytosis.

Epigenetic Research of Neurodegenerative Disorders Using Patient iPSC-Based Models

PubMed Central

2016-01-01

Epigenetic mechanisms play a role in human disease but their involvement in pathologies from the central nervous system has been hampered by the complexity of the brain together with its unique cellular architecture and diversity. Until recently, disease targeted neural types were only available as postmortem materials after many years of disease evolution. Current in vitro systems of induced pluripotent stem cells (iPSCs) generated by cell reprogramming of somatic cells from patients have provided valuable disease models recapitulating key pathological molecular events. Yet whether cell reprogramming on itself implies a truly epigenetic reprogramming, the epigenetic mechanisms governing this process are only partially understood. Moreover, elucidating epigenetic regulation using patient-specific iPSC-derived neural models is expected to have a great impact to unravel the pathophysiology of neurodegenerative diseases and to hopefully expand future therapeutic possibilities. Here we will critically review current knowledge of epigenetic involvement in neurodegenerative disorders focusing on the potential of iPSCs as a promising tool for epigenetic research of these diseases. PMID:26697081

Activation of raphe nuclei triggers rapid and distinct effects on parallel olfactory bulb output channels

PubMed Central

Kapoor, Vikrant; Provost, Allison; Agarwal, Prateek; Murthy, Venkatesh N.

2015-01-01

The serotonergic raphe nuclei are involved in regulating brain states over time-scales of minutes and hours. We examined more rapid effects of serotonergic activation on two classes of principal neurons in the mouse olfactory bulb, mitral and tufted cells, which send olfactory information to distinct targets. Brief stimulation of the raphe nuclei led to excitation of tufted cells at rest and potentiation of their odor responses. While mitral cells at rest were also excited by raphe activation, their odor responses were bidirectionally modulated, leading to improved pattern separation of odors. In vitro whole-cell recordings revealed that specific optogenetic activation of raphe axons affected bulbar neurons through dual release of serotonin and glutamate. Therefore, the raphe nuclei, in addition to their role in neuromodulation of brain states, are also involved in fast, sub-second top-down modulation, similar to cortical feedback. This modulation can selectively and differentially sensitize or decorrelate distinct output channels. PMID:26752161

Suppression of Cartilage Degradation by Zingerone Involving the p38 and JNK MAPK Signaling Pathway.

PubMed

Ruangsuriya, Jetsada; Budprom, Piyaporn; Viriyakhasem, Nawarat; Kongdang, Patiwat; Chokchaitaweesuk, Chatchadawalai; Sirikaew, Nutnicha; Chomdej, Siriwadee; Nganvongpanit, Korakot; Ongchai, Siriwan

2017-02-01

Zingerone, an active compound that is present in cooked ginger, has been claimed to be a bioactive ingredient that holds the potential of preventing and/or treating diseases involving inflammation. In this study, zingerone was used to discover its properties against joint inflammation using interleukin-1 β -induced osteoarthritis in cartilage explant and cell culture models. Zingerone was supplemented into the cartilage explant and cell culture media at different concentrations along with the presence of interleukin-1 β , an inducer of osteoarthritis. Markers indicating cartilage degradation, inflammation, and the signaling molecules involved in the inflammatory induction were investigated. Diacerien, an anti-osteoarthritic drug, was used as a positive control. Zingerone at a concentration of 40 µM reduced the level of matrix metalloproteinase-13 to about 31.95 ± 4.33 % compared with the interleukin-1 β -treated group and halted cartilage explant degradation as indicated by reducing the accumulative release of sulfated glycosaminoglycans by falling to the control concomitantly with an elevation of the remaining contents of uronic acid and collagen in the explant tissues when zingerone was added. In the SW1353 cell line model, zingerone efficiently suppressed the expression of TNF- α , interleukin-6, and interleukin-8 mRNA levels and tended to reduce the levels of both p38 and c-Jun N-terminal kinase phosphorylation. From the results of this study, it can be concluded that zingerone potentially reduced cartilage degradation, which is partially involved in p38 and c-Jun N-terminal kinases of the mitogen activator protein kinase signaling pathway leading to the reduction of proinflammatory cytokine amplification effects and cartilage-degrading enzyme syntheses. This finding supports the contention that ginger holds positive pharmaceutical effects against osteoarthritis. Georg Thieme Verlag KG Stuttgart · New York.

Implication of oxidative stress in size-dependent toxicity of silica nanoparticles in kidney cells.

PubMed

Passagne, Isabelle; Morille, Marie; Rousset, Marine; Pujalté, Igor; L'azou, Béatrice

2012-09-28

Silica nanoparticles (nano-SiO(2)) are one of the most popular nanomaterials used in industrial manufacturing, synthesis, engineering and medicine. While inhalation of nanoparticles causes pulmonary damage, nano-SiO(2) can be transported into the blood and deposit in target organs where they exert potential toxic effects. Kidney is considered as such a secondary target organ. However, toxicological information of their effect on renal cells and the mechanisms involved remain sparse. In the present study, the cytotoxicity of nano-SiO(2) of different sizes was investigated on two renal proximal tubular cell lines (human HK-2 and porcine LLC-PK(1)). The molecular pathways involved were studied with a focus on the involvement of oxidative stress. Nanoparticle characterization was performed (primary nanoparticle size, surface area, dispersion) in order to investigate a potential relationship between their physical properties and their toxic effects. Firstly, evidence of particle internalization was obtained by transmission electron microscopy and conventional flux cytometry techniques. The use of specific inhibitors of endocytosis pathways showed an internalization process by macropinocytosis and clathrin-mediated endocytosis for 100 nm nano-SiO(2) nanoparticles. These nanoparticles were localized in vesicles. Toxicity was size- and time-dependent (24h, 48 h, 72 h). Indeed, it increased as nanoparticles became smaller. Secondly, analysis of oxidative stress based on the assessment of ROS (reactive oxygen species) production (DHE, dihydroethidium) or lipid peroxidation (MDA, malondialdehyde) clearly demonstrated the involvement of oxidative stress in the toxicity of 20 nm nano-SiO(2). The induction of antioxidant enzymes (catalase, GSTpi, thioredoxin reductase) could explain their lesser toxicity with 100 nm nano-SiO(2). Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Current Status and Future Development of Cell Transplantation Therapy for Periodontal Tissue Regeneration

PubMed Central

Yoshida, Toshiyuki; Washio, Kaoru; Iwata, Takanori; Okano, Teruo; Ishikawa, Isao

2012-01-01

It has been shown that stem cell transplantation can regenerate periodontal tissue, and several clinical trials involving transplantation of stem cells into human patients have already begun or are in preparation. However, stem cell transplantation therapy is a new technology, and the events following transplantation are poorly understood. Several studies have reported side effects and potential risks associated with stem cell transplantation therapy. To protect patients from such risks, governments have placed regulations on stem cell transplantation therapies. It is important for the clinicians to understand the relevant risks and governmental regulations. This paper describes the ongoing clinical studies, basic research, risks, and governmental controls related to stem cell transplantation therapy. Then, one clinical study is introduced as an example of a government-approved periodontal cell transplantation therapy. PMID:22315604

Delivery of chemotherapeutic drugs in tumour cell-derived microparticles.

PubMed

Tang, Ke; Zhang, Yi; Zhang, Huafeng; Xu, Pingwei; Liu, Jing; Ma, Jingwei; Lv, Meng; Li, Dapeng; Katirai, Foad; Shen, Guan-Xin; Zhang, Guimei; Feng, Zuo-Hua; Ye, Duyun; Huang, Bo

2012-01-01

Cellular microparticles are vesicular plasma membrane fragments with a diameter of 100-1,000 nanometres that are shed by cells in response to various physiological and artificial stimuli. Here we demonstrate that tumour cell-derived microparticles can be used as vectors to deliver chemotherapeutic drugs. We show that tumour cells incubated with chemotherapeutic drugs package these drugs into microparticles, which can be collected and used to effectively kill tumour cells in murine tumour models without typical side effects. We describe several mechanisms involved in this process, including uptake of drug-containing microparticles by tumour cells, synthesis of additional drug-packaging microparticles by these cells that contribute to the cytotoxic effect and the inhibition of drug efflux from tumour cells. This study highlights a novel drug delivery strategy with potential clinical application.

Rapid fusion between mesenchymal stem cells and cardiomyocytes yields electrically active, non-contractile hybrid cells.

PubMed

Shadrin, Ilya Y; Yoon, Woohyun; Li, Liqing; Shepherd, Neal; Bursac, Nenad

2015-07-10

Cardiac cell therapies involving bone marrow-derived human mesenchymal stem cells (hMSCs) have shown promising results, although their mechanisms of action are still poorly understood. Here, we investigated direct interactions between hMSCs and cardiomyocytes in vitro. Using a genetic Ca(2+) indicator gCaMP3 to efficiently label hMSCs in co-cultures with neonatal rat ventricular myocytes (NRVMs), we determined that 25-40% of hMSCs (from 4 independent donors) acquired periodic Ca(2+) transients and cardiac markers through spontaneous fusion with NRVMs. Sharp electrode and voltage-clamp recordings in fused cells showed action potential properties and Ca(2+) current amplitudes in between those of non-fused hMSCs and NRVMs. Time-lapse video-microscopy revealed the first direct evidence of active fusion between hMSCs and NRVMs within several hours of co-culture. Application of blebbistatin, nifedipine or verapamil caused complete and reversible inhibition of fusion, suggesting potential roles for actomyosin bridging and Ca(2+) channels in the fusion process. Immunostaining for Cx43, Ki67, and sarcomeric α-actinin showed that fused cells remain strongly coupled to surrounding NRVMs, but downregulate sarcomeric structures over time, acquiring a non-proliferative and non-contractile phenotype. Overall, these results describe the phenotype and mechanisms of hybrid cell formation via fusion of hMSCs and cardiomyocytes with potential implications for cardiac cell therapy.

Neural cell adhesion molecule potentiates invasion and metastasis of melanoma cells through CAMP-dependent protein kinase and phosphatidylinositol 3-kinase pathways.

PubMed

Shi, Yu; Liu, Rui; Zhang, Si; Xia, Yin-Yan; Yang, Hai-Jie; Guo, Ke; Zeng, Qi; Feng, Zhi-Wei

2011-04-01

Neural cell adhesion molecule (NCAM) has been implicated in tumor metastasis yet its function in melanoma progression remains unclear. Here, we demonstrate that stably silencing NCAM expression in mouse melanoma B16F0 cells perturbs their cellular invasion and metastatic dissemination in vivo. The pro-invasive function of NCAM is exerted via dual mechanisms involving both cAMP-dependent protein kinase (PKA) and phosphatidylinositol 3-kinase (PI3K) pathways. Pharmacologic inhibition of PKA and PI3K leads to impaired cellular invasion. In contrast, forced expression of constitutively activated Akt, the major downstream target of PI3K, restores the defective cellular invasiveness of NCAM knock-down (KD) B16F0 cells. Furthermore, attenuation of either PKA or Akt activity in NCAM KD cells is shown to affect their common downstream target, transcription factor cAMP response element binding protein (CREB), which in turn down-regulates mRNA expression of matrix metalloproteinase-2 (MMP-2), thus contributes to impaired cellular invasion and metastasis of melanoma cells. Together, these findings indicate that NCAM potentiates cellular invasion and metastasis of melanoma cells through stimulation of PKA and PI3K signaling pathways thus suggesting the potential implication of anti-NCAM strategy in melanoma treatment. Copyright © 2011 Elsevier Ltd. All rights reserved.

Colon Cancer-Upregulated Long Non-Coding RNA lincDUSP Regulates Cell Cycle Genes and Potentiates Resistance to Apoptosis.

PubMed

Forrest, Megan E; Saiakhova, Alina; Beard, Lydia; Buchner, David A; Scacheri, Peter C; LaFramboise, Thomas; Markowitz, Sanford; Khalil, Ahmad M

2018-05-09

Long non-coding RNAs (lncRNAs) are frequently dysregulated in many human cancers. We sought to identify candidate oncogenic lncRNAs in human colon tumors by utilizing RNA sequencing data from 22 colon tumors and 22 adjacent normal colon samples from The Cancer Genome Atlas (TCGA). The analysis led to the identification of ~200 differentially expressed lncRNAs. Validation in an independent cohort of normal colon and patient-derived colon cancer cell lines identified a novel lncRNA, lincDUSP, as a potential candidate oncogene. Knockdown of lincDUSP in patient-derived colon tumor cell lines resulted in significantly decreased cell proliferation and clonogenic potential, and increased susceptibility to apoptosis. The knockdown of lincDUSP affects the expression of ~800 genes, and NCI pathway analysis showed enrichment of DNA damage response and cell cycle control pathways. Further, identification of lincDUSP chromatin occupancy sites by ChIRP-Seq demonstrated association with genes involved in the replication-associated DNA damage response and cell cycle control. Consistent with these findings, lincDUSP knockdown in colon tumor cell lines increased both the accumulation of cells in early S-phase and γH2AX foci formation, indicating increased DNA damage response induction. Taken together, these results demonstrate a key role of lincDUSP in the regulation of important pathways in colon cancer.

The Role of Mitochondria in Cancer Induction, Progression and Changes in Metabolism.

PubMed

Rogalinska, Malgorzata

2016-01-01

Mitochondria play important roles as energetic centers. Mutations in mitochondrial DNA (mtDNA) were found in several diseases, including cancers. Studies on cytoplasmic hybrids (cybrids) confirm that directed mutation introduced into mtDNA could be a reason for cancer induction. Mitochondria could also be a factor linking cancer transformation and progression. The importance of mitochondria in cancer also confirms their involvement in the resistance to treatment. Resistance to treatment of cancer cells can frequently be a reason for glycolysis acceleration. It could be explained by cancer cells' high proliferation index and high energy request. The involvement of mitochondria in metabolic disturbances of several metabolic diseases, including cancers, was reported. These data confirm that cancer induction, as well as cancer progression, could have metabolic roots. The aberrant products observed in prostate cells involved in the Krebs cycle could promote cancer progression. These multiple relationships between alterations on a genetic level translated into disturbances in cellular metabolism and their potential relation with epigenetic control of gene expression make cancerogenesis more complicated and prognoses' success in studies on cancer etiology more distant in time.

Induction of apoptosis in cancer cell lines by the Red Sea brine pool bacterial extracts.

PubMed

Sagar, Sunil; Esau, Luke; Holtermann, Karie; Hikmawan, Tyas; Zhang, Guishan; Stingl, Ulrich; Bajic, Vladimir B; Kaur, Mandeep

2013-12-05

Marine microorganisms are considered to be an important source of bioactive molecules against various diseases and have great potential to increase the number of lead molecules in clinical trials. Progress in novel microbial culturing techniques as well as greater accessibility to unique oceanic habitats has placed the marine environment as a new frontier in the field of natural product drug discovery. A total of 24 microbial extracts from deep-sea brine pools in the Red Sea have been evaluated for their anticancer potential against three human cancer cell lines. Downstream analysis of these six most potent extracts was done using various biological assays, such as Caspase-3/7 activity, mitochondrial membrane potential (MMP), PARP-1 cleavage and expression of γH2Ax, Caspase-8 and -9 using western blotting. In general, most of the microbial extracts were found to be cytotoxic against one or more cancer cell lines with cell line specific activities. Out of the 13 most active microbial extracts, six extracts were able to induce significantly higher apoptosis (>70%) in cancer cells. Mechanism level studies revealed that extracts from Chromohalobacter salexigens (P3-86A and P3-86B(2)) followed the sequence of events of apoptotic pathway involving MMP disruption, caspase-3/7 activity, caspase-8 cleavage, PARP-1 cleavage and Phosphatidylserine (PS) exposure, whereas another Chromohalobacter salexigens extract (K30) induced caspase-9 mediated apoptosis. The extracts from Halomonas meridiana (P3-37B), Chromohalobacter israelensis (K18) and Idiomarina loihiensis (P3-37C) were unable to induce any change in MMP in HeLa cancer cells, and thus suggested mitochondria-independent apoptosis induction. However, further detection of a PARP-1 cleavage product, and the observed changes in caspase-8 and -9 suggested the involvement of caspase-mediated apoptotic pathways. Altogether, the study offers novel findings regarding the anticancer potential of several halophilic bacterial species inhabiting the Red Sea (at the depth of 1500-2500 m), which constitute valuable candidates for further isolation and characterization of bioactive molecules.

Evidence for a potential tumor suppressor role for the Na,K-ATPase ß1-subunit

PubMed Central

Inge, Landon J.; Rajasekaran, Sigrid A.; Yoshimoto, Koji; Mischel, Paul S.; McBride, William; Landaw, Elliot; Rajasekaran, Ayyappan K.

2009-01-01

Summary The Na,K-ATPase, consisting of two essential subunits (α, ß), plays a critical role in the regulation of ion homeostasis in mammalian cells. Recent studies indicate that reduced expression of the ß1 isoform (NaK-ß1) is commonly observed in carcinoma and is associated with events involved in cancer progression. In this study, we present evidence that repletion of NaK-ß1 in Moloney sarcoma virus-transformed Madin-Darby canine kidney cells (MSV-MDCK), a highly tumorigenic cell line, inhibits anchorage independent growth and suppresses tumor formation in immunocompromised mice. Additionally, using an in vitro cell-cell aggregation assay, we showed that cell aggregates of NaK-ß1 subunit expressing MSV-MDCK cells have reduced extracellular regulated kinase (ERK) 1/2 activity compared with parental MSV-MDCK cells. Finally, using immunohistochemistry and fully quantitative image analysis approaches, we showed that the levels of phosphorylated ERK 1/2 are inversely correlated to the NaK-ß1 levels in the tumors. These findings reveal for the first time that NaK-ß1 has a potential tumor-suppressor function in epithelial cells. PMID:18228203

Graphene supports in vitro proliferation and osteogenic differentiation of goat adult mesenchymal stem cells: potential for bone tissue engineering.

PubMed

Elkhenany, Hoda; Amelse, Lisa; Lafont, Andersen; Bourdo, Shawn; Caldwell, Marc; Neilsen, Nancy; Dervishi, Enkeleda; Derek, Oshin; Biris, Alexandru S; Anderson, David; Dhar, Madhu

2015-04-01

Current treatments for bone loss injuries involve autologous and allogenic bone grafts, metal alloys and ceramics. Although these therapies have proved useful, they suffer from inherent challenges, and hence, an adequate bone replacement therapy has not yet been found. We hypothesize that graphene may be a useful nanoscaffold for mesenchymal stem cells and will promote proliferation and differentiation into bone progenitor cells. In this study, we evaluate graphene, a biocompatible inert nanomaterial, for its effect on in vitro growth and differentiation of goat adult mesenchymal stem cells. Cell proliferation and differentiation are compared between polystyrene-coated tissue culture plates and graphene-coated plates. Graphitic materials are cytocompatible and support cell adhesion and proliferation. Importantly, cells seeded on to oxidized graphene films undergo osteogenic differentiation in fetal bovine serum-containing medium without the addition of any glucocorticoid or specific growth factors. These findings support graphene's potential to act as an osteoinducer and a vehicle to deliver mesenchymal stem cells, and suggest that the combination of graphene and goat mesenchymal stem cells provides a promising construct for bone tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.

Cafestol, a diterpene molecule found in coffee, induces leukemia cell death.

PubMed

Lima, Cauê S; Spindola, Daniel G; Bechara, Alexandre; Garcia, Daniel M; Palmeira-Dos-Santos, Caroline; Peixoto-da-Silva, Janaina; Erustes, Adolfo G; Michelin, Luis F G; Pereira, Gustavo J S; Smaili, Soraya S; Paredes-Gamero, Edgar; Calgarotto, Andrana K; Oliveira, Carlos R; Bincoletto, Claudia

2017-08-01

To evaluate the antitumor properties of Cafestol four leukemia cell lines were used (NB4, K562, HL60 and KG1). Cafestol exhibited the highest cytotoxicity against HL60 and KG1 cells, as evidenced by the accumulation of cells in the sub-G1 fraction, mitochondrial membrane potential reduction, accumulation of cleaved caspase-3 and phosphatidylserine externalization. An increase in CD11b and CD15 differentiation markers with attenuated ROS generation was also observed in Cafestol-treated HL60 cells. These results were similar to those obtained following exposure of the same cell line to cytarabine (Ara-C), an antileukemic drug. Cafestol and Ara-C reduced the clonogenic potential of HL60 cells by 100%, but Cafestol spared murine colony forming unit- granulocyte/macrophage (CFU-GM), which retained their clonogenicity. The co-treatment of Cafestol and Ara-C reduced HL60 cell viability compared with both drugs administered alone. In conclusion, despite the distinct molecular mechanisms involved in the activity of Cafestol and Ara-C, a similar cytotoxicity towards leukemia cells was observed, which suggests a need for prophylactic-therapeutic pre-clinical studies regarding the anticancer properties of Cafestol. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Emerging Importance of Phytochemicals in Regulation of Stem Cells Fate via Signaling Pathways.

PubMed

Dadashpour, Mehdi; Pilehvar-Soltanahmadi, Younes; Zarghami, Nosratollah; Firouzi-Amandi, Akram; Pourhassan-Moghaddam, Mohammad; Nouri, Mohammad

2017-11-01

To reach ideal therapeutic potential of stem cells for regenerative medicine purposes, it is essential to retain their self-renewal and differentiation capacities. Currently, biological factors are extensively used for stemness maintaining and differentiation induction of stem cells. However, low stability, high cost, complicated production process, and risks associated with viral/endotoxin infection hamper the widespread use of biological factors in the stem cell biology. Moreover, regarding the modulation of several signaling cascades, which lead to a distinct fate, phytochemicals are preferable in the stem cells biology because of their efficiency. Considering the issues related to the application of biological factors and potential advantages of phytochemicals in stem cell engineering, there is a considerable increasing trend in studies associated with the application of novel alternative molecules in the stem cell biology. In support of this statement, we aimed to highlight the various effects of phytochemicals on signaling cascades involved in commitment of stem cells. Hence, in this review, the current trends in the phytochemicals-based modulation of stem cell fate have been addressed. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Manufacturing Cell Therapies Using Engineered Biomaterials.

PubMed

Abdeen, Amr A; Saha, Krishanu

2017-10-01

Emerging manufacturing processes to generate regenerative advanced therapies can involve extensive genomic and/or epigenomic manipulation of autologous or allogeneic cells. These cell engineering processes need to be carefully controlled and standardized to maximize safety and efficacy in clinical trials. Engineered biomaterials with smart and tunable properties offer an intriguing tool to provide or deliver cues to retain stemness, direct differentiation, promote reprogramming, manipulate the genome, or select functional phenotypes. This review discusses the use of engineered biomaterials to control human cell manufacturing. Future work exploiting engineered biomaterials has the potential to generate manufacturing processes that produce standardized cells with well-defined critical quality attributes appropriate for clinical testing. Copyright © 2017 Elsevier Ltd. All rights reserved.

Small Molecules Affect Human Dental Pulp Stem Cell Properties Via Multiple Signaling Pathways

PubMed Central

Al-Habib, Mey; Yu, Zongdong

2013-01-01

One fundamental issue regarding stem cells for regenerative medicine is the maintenance of stem cell stemness. The purpose of the study was to test whether small molecules can enhance stem cell properties of mesenchymal stem cells (MSCs) derived from human dental pulp (hDPSCs), which have potential for multiple clinical applications. We identified the effects of small molecules (Pluripotin (SC1), 6-bromoindirubin-3-oxime and rapamycin) on the maintenance of hDPSC properties in vitro and the mechanisms involved in exerting the effects. Primary cultures of hDPSCs were exposed to optimal concentrations of these small molecules. Treated hDPSCs were analyzed for their proliferation, the expression levels of pluripotent and MSC markers, differentiation capacities, and intracellular signaling activations. We found that small molecule treatments decreased cell proliferation and increased the expression of STRO-1, NANOG, OCT4, and SOX2, while diminishing cell differentiation into odonto/osteogenic, adipogenic, and neurogenic lineages in vitro. These effects involved Ras-GAP-, ERK1/2-, and mTOR-signaling pathways, which may preserve the cell self-renewal capacity, while suppressing differentiation. We conclude that small molecules appear to enhance the immature state of hDPSCs in culture, which may be used as a strategy for adult stem cell maintenance and extend their capacity for regenerative applications. PMID:23573877

Transport of BMAA into Neurons and Astrocytes by System xc.

PubMed

Albano, Rebecca; Lobner, Doug

2018-01-01

The study of the mechanism of β-N-methylamino-L-alanine (BMAA) neurotoxicity originally focused on its effects at the N-methyl-D-aspartate (NMDA) receptor. In recent years, it has become clear that its mechanism of action is more complicated. First, there are certain cell types, such as motor neurons and cholinergic neurons, where the dominate mechanism of toxicity is through action at AMPA receptors. Second, even in cortical neurons where the primary mechanism of toxicity appears to be activation of NMDA receptors, there are other mechanisms involved. We found that along with NMDA receptors, activation of mGLuR5 receptors and effects on the cystine/glutamate antiporter (system x c -) were involved in the toxicity. The effects on system x c - are of particular interest. System x c - mediates the transport of cystine into the cell in exchange for releasing glutamate into the extracellular fluid. By releasing glutamate, system x c - can potentially cause excitotoxicity. However, through providing cystine to the cell, it regulates the levels of cellular glutathione (GSH), the main endogenous intracellular antioxidant, and in this way may protect cells against oxidative stress. We have previously published that BMAA inhibits cystine uptake leading to GSH depletion and had indirect evidence that BMAA is transported into the cells by system x c -. We now present direct evidence that BMAA is transported into both astrocytes and neurons through system x c -. The fact that BMAA is transported by system x c - also provides a mechanism for BMAA to enter brain cells potentially leading to misincorporation into proteins and protein misfolding.

Streptococcus suis DivIVA Protein Is a Substrate of Ser/Thr Kinase STK and Involved in Cell Division Regulation

PubMed Central

Ni, Hua; Fan, Weiwei; Li, Chaolong; Wu, Qianqian; Hou, Hongfen; Hu, Dan; Zheng, Feng; Zhu, Xuhui; Wang, Changjun; Cao, Xiangrong; Shao, Zhu-Qing; Pan, Xiuzhen

2018-01-01

Streptococcus suis serotype 2 is an important swine pathogen and an emerging zoonotic agent that causes severe infections. Recent studies have reported a eukaryotic-like Ser/Thr protein kinase (STK) gene and characterized its role in the growth and virulence of different S. suis 2 strains. In the present study, phosphoproteomic analysis was adopted to identify substrates of the STK protein. Seven proteins that were annotated to participate in different cell processes were identified as potential substrates, which suggests the pleiotropic effects of stk on S. suis 2 by targeting multiple pathways. Among them, a protein characterized as cell division initiation protein (DivIVA) was further investigated. In vitro analysis demonstrated that the recombinant STK protein directly phosphorylates threonine at amino acid position 199 (Thr-199) of DivIVA. This effect could be completely abolished by the T199A mutation. To determine the specific role of DivIVA in growth and division, a divIVA mutant was constructed. The ΔdivIVA strain exhibited impaired growth and division, including lower viability, enlarged cell mass, asymmetrical division caused by aberrant septum, and extremely weak pathogenicity in a mouse infection model. Collectively, our results reveal that STK regulates the cell growth and virulence of S. suis 2 by targeting substrates that are involved in different biological pathways. The inactivation of DivIVA leads to severe defects in cell division and strongly attenuates pathogenicity, thereby indicating its potential as a molecular drug target against S. suis. PMID:29616196

Regulation of TRP channels by steroids: Implications in physiology and diseases.

PubMed

Kumar, Ashutosh; Kumari, Shikha; Majhi, Rakesh Kumar; Swain, Nirlipta; Yadav, Manoj; Goswami, Chandan

2015-09-01

While effects of different steroids on the gene expression and regulation are well established, it is proven that steroids can also exert rapid non-genomic actions in several tissues and cells. In most cases, these non-genomic rapid effects of steroids are actually due to intracellular mobilization of Ca(2+)- and other ions suggesting that Ca(2+) channels are involved in such effects. Transient Receptor Potential (TRP) ion channels or TRPs are the largest group of non-selective and polymodal ion channels which cause Ca(2+)-influx in response to different physical and chemical stimuli. While non-genomic actions of different steroids on different ion channels have been established to some extent, involvement of TRPs in such functions is largely unexplored. In this review, we critically analyze the literature and summarize how different steroids as well as their metabolic precursors and derivatives can exert non-genomic effects by acting on different TRPs qualitatively and/or quantitatively. Such effects have physiological repercussion on systems such as in sperm cells, immune cells, bone cells, neuronal cells and many others. Different TRPs are also endogenously expressed in diverse steroid-producing tissues and thus may have importance in steroid synthesis as well, a process which is tightly controlled by the intracellular Ca(2+) concentrations. Tissue and cell-specific expression of TRP channels are also regulated by different steroids. Understanding of the crosstalk between TRP channels and different steroids may have strong significance in physiological, endocrinological and pharmacological context and in future these compounds can also be used as potential biomedicine. Copyright © 2014 Elsevier Inc. All rights reserved.

Redox-Sensitive Cerium Oxide Nanoparticles Protect Human Keratinocytes from Oxidative Stress Induced by Glutathione Depletion.

PubMed

Singh, Ragini; Karakoti, Ajay S; Self, William; Seal, Sudipta; Singh, Sanjay

2016-11-22

Cerium oxide nanoparticles (CeNPs) have gathered much attention in the biomedical field due to its unique antioxidant property. It can protect cells and tissues from oxidative stress induced damage due to its autoregenerative redox cycle. Our study explores the antioxidant and antigenotoxic behavior of PEGylated CeNPs toward oxidative insult produced by buthionine sulfoximine (BSO) in human keratinocytes (HaCaT cells). BSO inhibits the γ-glutamylcysteinesynthetase (γ-GCS) enzyme and thus acts as a glutathione (GSH) depleting agent to modulate the cellular redox potential. GSH is a natural ROS scavenger present in the mammalian cells, and its depletion causes generation of reactive oxygen species (ROS). In this study, we challenged HaCaT cells (keratinocytes) with BSO to alter the redox potential within the cell and monitored toxicity, ROS generation, and nuclear fragmentation. We also followed changes in expressions of related proteins and genes. We found that PEGylated CeNPs can protect HaCaT cells from BSO-induced oxidative damage. BSO-exposed cells, preincubated with PEGylated CeNPs, showed better cell survival and significant decrease in the intracellular levels of ROS. We also observed decrease in lactate dehydrogenase (LDH) release and nuclear fragmentation in CeNP-treated cells that were challenged with BSO as compared to treatment with BSO alone. Exposure of HaCaT cells with BSO leads to altered expression of antioxidant genes and proteins, i.e., thioredoxin reductase (TrxR) and peroxiredoxin 6 (Prx6) whereas, in our study, pretreatment of PEGylated CeNPs reduces the need for induction of genes that produce enzymes involved in the defense against oxidative stress. Since, growing evidence argued the involvement of ROS in mediating death of mammalian cells in several ailments, our finding reinforces the use of PEGylated CeNPs as a potent pharmacological agent under the lower cellular GSH/GSSG ratios for the treatment of diseases mediated by free radicals.

Bifunctional Alkylating Agent-Induced p53 and Nonclassical Nuclear Factor kB Responses and Cell Death are Altered by Caffeic Acid Phenethyl Ester: A Potential Role for Antioxidant/Electrophilic Response-Element Signaling

DTIC Science & Technology

2007-01-01

Methods. To establish the maximal LDH 2,3,7,8-tetrachloro- activity achievable from these cells, untreated control cells were grown dibenzo-p- dioxin to the...to modulated by AhR (Miao et al., 2005). Curcumin has been induce apoptosis. CAPE may be involved in anoikis, that is, shown to compete with dioxin ...the AhR directly binds (Miao et al., 2005). adhesion kinase (Weyant et al., 2000). Also, CAPE-induced Exposure of Hepalclc7 cells to dioxin results in

Regenerating new heart with stem cells

PubMed Central

Anversa, Piero; Kajstura, Jan; Rota, Marcello; Leri, Annarosa

2013-01-01

This article discusses current understanding of myocardial biology, emphasizing the regeneration potential of the adult human heart and the mechanisms involved. In the last decade, a novel conceptual view has emerged. The heart is no longer considered a postmitotic organ, but is viewed as a self-renewing organ characterized by a resident stem cell compartment responsible for tissue homeostasis and cardiac repair following injury. Additionally, HSCs possess the ability to transdifferentiate and acquire the cardiomyocyte, vascular endothelial, and smooth muscle cell lineages. Both cardiac and hematopoietic stem cells may be used therapeutically in an attempt to reverse the devastating consequences of chronic heart failure of ischemic and nonischemic origin. PMID:23281411

Light energy conservation processes in Halobacterium halobium cells

NASA Technical Reports Server (NTRS)

Bogomolni, R. A.

1977-01-01

Proton pumping driven by light or by respiration generates an electrochemical potential difference across the membrane in Halobacterium halobium. The pH changes induced by light or by respiration in cell suspensions are complicated by proton flows associated with the functioning of the cellular energy transducers. A proton-per-ATP ratio of about 3 is calculated from simultaneous measurements of phosphorylation and the proton inflow. This value is compatible with the chemiosmotic coupling hypothesis. The time course of the light-induced changes in membrane potential indicates that light-driven pumping increases a dark pre-existing potential of about 130 mV only by a small amount (20 to 30 mV). The complex kinetic features of the membrane potential changes do not closely follow those of the pH changes, which suggests that flows of ions other than protons are involved. A qualitative model consistent with the available data is presented.

Bax Translocation Mediated Mitochondrial Apoptosis and Caspase Dependent Photosensitizing Effect of Ficus religiosa on Cancer Cells

PubMed Central

Thankayyan R, Santhosh Kumar; Sithul, Hima; Sreeharshan, Sreeja

2012-01-01

The main aim of the present work was to investigate the potential effect of acetone extract of Ficus religosa leaf (FAE) in multiple apoptosis signalling in human breast cancer cells. FAE treatment significantly induced dose and time dependent, irreversible inhibition of breast cancer cell growth with moderate toxicity to normal breast epithelial cells. This observation was validated using Sulforhodamine B assay. Cell cycle analysis by Flow cytometry showed cell cycle arrest in G1 phase and induction of sub-G0 peak. FAE induced chromatin condensation and displayed an increase in apoptotic population in Annexin V-FITC/PI (Fluorescein isothiocyanate/Propidium iodide) double staining. FAE stimulated the loss of mitochondrial membrane potential in multiple breast cancer cell lines when compared to normal diploid cells. To understand the role of Bax in FAE induced apoptosis, we employed a sensitive cell based platform of MCF-7 cells expressing Bax-EGFP. Bax translocation to mitochondria was accompanied by the disruption of mitochondrial membrane potential and marked elevation in LEHDase activity (Caspase 9). Consistent with this data, FAE induced Caspase activation as evidenced by ratio change in FRET Caspase sensor expressing MCF-7 cell line and cleavage of prominent Caspases and PARP. Interestingly, FAE accelerated cell death in a mitochondrial dependent manner in continuous live cell imaging mode indicating its possible photosensitizing effect. Intracellular generation of reactive oxygen species (ROS) by FAE played a critical role in mediating apoptotic cell death and photosensitizing activity. FAE induced dose and time dependent inhibition of cancer cell growth which was associated with Bax translocation and mitochondria mediated apoptosis with the activation of Caspase 9 dependent Caspase cascade. FAE also possessed strong photosensitizing effect on cancer cell line that was mediated through rapid mitochondrial transmembrane potential loss and partial Caspase activation involving generation of intracellular ROS. PMID:22792212

The O-methylated isoflavone, formononetin, inhibits human ovarian cancer cell proliferation by sub G0/G1 cell phase arrest through PI3K/AKT and ERK1/2 inactivation.

PubMed

Park, Sunwoo; Bazer, Fuller W; Lim, Whasun; Song, Gwonhwa

2018-05-15

Formononetin is an isoflavone that is extracted from red clovers or soy. It has anti-oxidant, anti-proliferative, and anti-tumor effects against cells in various diseases. Several cohort studies have indicated that phytoestrogen intake, including formononetin, could reduce the risk of various carcinogenesis. In fact, many case-control studies have indicated the potential value of flavonoids as drug supplements in the treatment of many cancer patients. However, the toxic effects and the anti-cancer mechanism of formononetin in ovarian cancer are unknown. We investigated the toxicological mechanism of formononetin in ES2 and OV90 ovarian cancer cells. Formononetin suppressed cell proliferation through sub G0/G1 phase arrest and increased apoptosis in both cell lines. Furthermore, it induced loss of mitochondrial membrane potential and generation of reactive oxygen species in ES2 and OV90 cells. The formononetin-mediated regulation of cell proliferation and apoptosis involved decreased phosphorylation of ERK1/2, P90RSK, AKT, P70S6K, and S6 proteins, and increased phosphorylation of P38 protein in ES2 and OV90 cells. Co-treatment of formononetin with pharmacological inhibitors (LY294002 or U0126) revealed additional anti-proliferative effects on the two human ovarian cancer cell types. Conclusively, the results indicate the potential value of formononetin as an anti-cancer agent in human ovarian cancer. © 2018 Wiley Periodicals, Inc.

Sensitive molecular detection of small nodal metastasis in uterine cervical cancer using HPV16-E6/CK19/MUC1 cancer biomarkers.

PubMed

Samouëlian, Vanessa; Mechtouf, Nawel; Leblanc, Eric; Cardin, Guillaume B; Lhotellier, Valérie; Querleu, Denis; Révillion, Françoise; Rodier, Francis

2018-04-24

Metastatic nodal involvement is a critical prognostic factor in uterine cervical cancer (UCC). To improve current methods of detecting UCC metastases in lymph nodes (LNs), we used quantitative PCR (qPCR) to assess mRNA expression of potential metastatic biomarkers. We found that expression of HPV16-E6, cytokeratin19 (CK19), and mucin1 (MUC1) is consistently upregulated in tumors and metastatic tissues, supporting a role for these genes in UCC progression. These putative biomarkers were able to predict the presence of histologically positive metastatic LNs with respective sensitivities and specificities of 82% and 99% (CK19), 76% and 95% (HPV16-E6), and 76% and 78% (MUC1). While the biomarkers failed to detect 1.7% to 2.2% of the histologically positive LNs when used individually, combining CK19 and HPV16-E6 enhanced sensitivity and specificity to 100% and 94%, respectively. To explore the sensitivity of qPCR-based detection of varying proportions of invading HPV16-positive UCC cells, we designed a LN metastasis model that achieved a fresh cell detection limit of 0.008% (1:12500 HPV16-positive to HPV16-negative cells), and a paraffin-embedded, formalin-fixed (PEFF) detection limit of 0.02% (1:5000 HPV16-positive to HPV16-negative cells), both of which are within the theoretical detection limit for micrometastasis. Thus, HPV E6/E7 oncogenes may be useful targets for the ultrasensitive detection of nodal involvements like micrometastases in fresh or archived tissue samples. Moreover, our results suggest that the biomarker combination of CK19/HPV-E6 could support a real-time intraoperative strategy for the detection of small, but potentially lethal, metastatic nodal involvements in fresh UCC tissues.

Induction of apoptosis in melanoma A375 cells by a chloroform fraction of Centratherum anthelminticum (L.) seeds involves NF-kappaB, p53 and Bcl-2-controlled mitochondrial signaling pathways.

PubMed

Looi, Chung Yeng; Moharram, Bushra; Paydar, Mohammadjavad; Wong, Yi Li; Leong, Kok Hoong; Mohamad, Khalit; Arya, Aditya; Wong, Won Fen; Mustafa, Mohd Rais

2013-07-10

Centratherum anthelminticum (L.) Kuntze (scientific synonyms: Vernonia anthelmintica; black cumin) is one of the ingredients of an Ayurvedic preparation, called "Kayakalp", commonly applied to treat skin disorders in India and Southeast Asia. Despite its well known anti-inflammatory property on skin diseases, the anti-cancer effect of C. anthelminticum seeds on skin cancer is less documented. The present study aims to investigate the anti-cancer effect of Centratherum anthelminticum (L.) seeds chloroform fraction (CACF) on human melanoma cells and to elucidate the molecular mechanism involved. A chloroform fraction was extracted from C. anthelminticum (CACF). Bioactive compounds of the CACF were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Human melanoma cell line A375 was treated with CACF in vitro. Effects of CACF on growth inhibition, morphology, stress and survival of the cell were examined with MTT, high content screening (HSC) array scan and flow cytometry analyses. Involvement of intrinsic or extrinsic pathways in the CACF-induced A375 cell death mechanism was examined using a caspase luminescence assay. The results were further verified with different caspase inhibitors. In addition, Western blot analysis was performed to elucidate the changes in apoptosis-associated molecules. Finally, the effect of CACF on the NF-κB nuclear translocation ability was assayed. The MTT assay showed that CACF dose-dependently inhibited cell growth of A375, while exerted less cytotoxic effect on normal primary epithelial melanocytes. We demonstrated that CACF induced cell growth inhibition through apoptosis, as evidenced by cell shrinkage, increased annexin V staining and formation of membrane blebs. CACF treatment also resulted in higher reactive oxygen species (ROS) production and lower Bcl-2 expression, leading to decrease mitochondrial membrane potential (MMP). Disruption of the MMP facilitated the release of mitochondrial cytochrome c, which activates caspase-9 and downstream caspase-3/7, resulting in DNA fragmentation and up-regulation of p53 in melanoma cells. Moreover, CACF prevented TNF-α-induced NF-κB nuclear translocation, which further committed A375 cells toward apoptosis. Together, our findings suggest CACF as a potential therapeutic agent against human melanoma malignancy.

Targeted therapy against EGFR and VEGFR using ZD6474 enhances the therapeutic potential of UV-B phototherapy in breast cancer cells.

PubMed

Sarkar, Siddik; Rajput, Shashi; Tripathi, Amit Kumar; Mandal, Mahitosh

2013-10-20

The hypoxic environment of tumor region stimulated the up regulation of growth factors responsible for angiogenesis and tumor proliferation. Thus, targeting the tumor vasculature along with the proliferation by dual tyrosine kinase inhibitor may be the efficient way of treating advanced breast cancers, which can be further enhanced by combining with radiotherapy. However, the effectiveness of radiotherapy may be severely compromised by toxicities and tumor resistance due to radiation-induced adaptive response contributing to recurrence and metastases of breast cancer. The rational of using ZD6474 is to evaluate the feasibility and efficacy of combined VEGFR2 and EGFR targeting with concurrent targeted and localized UV-B phototherapy in vitro breast cancer cells with the anticipation to cure skin lesions infiltrated with breast cancer cells. Breast cancer cells were exposed to UV-B and ZD6474 and the cell viability, apoptosis, invasion and motility studies were conducted for the combinatorial effect. Graphs and statistical analyses were performed using Graph Pad Prism 5.0. ZD6474 and UV-B decreased cell viability in breast cancers in combinatorial manner without affecting the normal human mammary epithelial cells. ZD6474 inhibited cyclin E expression and induced p53 expression when combined with UV-B. It activated stress induced mitochondrial pathway by inducing translocation of bax and cytochrome-c. The combination of ZD6474 with UV-B vs. either agent alone also more potently down-regulated the anti-apoptotic bcl-2 protein, up-regulated pro-apoptotic signaling events involving expression of bax, activation of caspase-3 and caspase-7 proteins, and induced poly (ADP-ribose) polymerase resulting in apoptosis. ZD6474 combined with UV-B inhibited invasion of breast cancer cells in vitro as compared to either single agent, indicating a potential involvement of pro-angiogenic growth factors in regulating the altered expression and reorganization of cytoskeletal proteins in combinatorial treated breast cancer cells. Involvement of combination therapy in reducing the expression of matrix metalloprotease was also observed. Collectively, our studies indicate that incorporating an anti-EGFR plus VEGFR strategy (ZD6474) with phototherapy (UV-B), an alternative approach to the ongoing conventional radiotherapy for the treatment of infiltrating metastatic breast cancer cells in the skin and for locally recurrence breast cancer than either approach alone.

Essential Role of DAP12 Signaling in Macrophage Programming into a Fusion-Competent State

PubMed Central

Helming, Laura; Tomasello, Elena; Kyriakides, Themis R.; Martinez, Fernando O.; Takai, Toshiyuki; Gordon, Siamon; Vivier, Eric

2009-01-01

Multinucleated giant cells, formed by fusion of macrophages, are a hallmark of granulomatous inflammation. With a genetic approach, we show that signaling through the adaptor protein DAP12 (DNAX activating protein of 12 kD), its associated receptor triggering receptor expressed by myeloid cells 2 (TREM-2), and the downstream protein tyrosine kinase Syk is required for the cytokine-induced formation of giant cells and that overexpression of DAP12 potentiates macrophage fusion. We also present evidence that DAP12 is a general macrophage fusion regulator and is involved in modulating the expression of several macrophage-associated genes, including those encoding known mediators of macrophage fusion, such as DC-STAMP and Cadherin 1. Thus, DAP12 is involved in programming of macrophages through the regulation of gene and protein expression to induce a fusion-competent state. PMID:18957693

Mycobacteria exploit nitric oxide-induced transformation of macrophages into permissive giant cells.

PubMed

Gharun, Kourosh; Senges, Julia; Seidl, Maximilian; Lösslein, Anne; Kolter, Julia; Lohrmann, Florens; Fliegauf, Manfred; Elgizouli, Magdeldin; Vavra, Martina; Schachtrup, Kristina; Illert, Anna L; Gilleron, Martine; Kirschning, Carsten J; Triantafyllopoulou, Antigoni; Henneke, Philipp

2017-12-01

Immunity to mycobacteria involves the formation of granulomas, characterized by a unique macrophage (MΦ) species, so-called multinucleated giant cells (MGC). It remains unresolved whether MGC are beneficial to the host, that is, by prevention of bacterial spread, or whether they promote mycobacterial persistence. Here, we show that the prototypical antimycobacterial molecule nitric oxide (NO), which is produced by MGC in excessive amounts, is a double-edged sword. Next to its antibacterial capacity, NO propagates the transformation of MΦ into MGC, which are relatively permissive for mycobacterial persistence. The mechanism underlying MGC formation involves NO-induced DNA damage and impairment of p53 function. Moreover, MGC have an unsurpassed potential to engulf mycobacteria-infected apoptotic cells, which adds a further burden to their antimycobacterial capacity. Accordingly, mycobacteria take paradoxical advantage of antimicrobial cellular efforts by driving effector MΦ into a permissive MGC state. © 2017 The Authors.

CD151-mediated adhesion is crucial to osteosarcoma pulmonary metastasis

PubMed Central

Sun, Mengxiong; Zhou, Chenghao; Chen, Jian; Yin, Fei; Wang, Hongsheng; Lin, Binhui; Zuo, Dongqing; Li, Suoyuan; Feng, Lijin; Duan, Zhenfeng; Cai, Zhengdong; Hua, Yingqi

2016-01-01

CD151, a tetraspanin family protein involved in cell-cell and cell-extracellular matrix interaction, is differentially expressed in osteosarcoma cell membranes. Thus, this study aimed to investigate the role of CD151 in osteosarcoma metastasis. We analyzed CD151 expression in patient tissue samples using immunohistochemistry. CD151 expression was also silenced with shRNA in osteosarcoma cells of high metastatic potential, and cell adhesion, migration and invasion were evaluated in vitro and pulmonary metastasis was investigated in vivo. Mediators of cell signaling pathways were also examined following suppression of CD151 expression. Overall survival for patients with low versus high CD151 expression level was 94 vs. 41 months (p=0.0451). CD151 expression in osteosarcoma cells with high metastatic potential was significantly higher than in those with low metastatic potential (p<0.001). shRNA-mediated silencing of CD151 did not influence cell viability or proliferation; however, cell adhesion, migration and invasion were all inhibited (all p<0.001). In mice inoculated with shRNA-transduced osteosarcoma cells, the number and size of lung metastatic lesions were reduced compared to the mice inoculated with control-shRNA transduced cells (p<0.001). In addition, CD151 knockdown significantly reduced Akt, p38, and p65 phosphorylation as well as focal adhesion kinase, integrin β1, p70s6, and p-mTOR levels. Taken together, CD151 induced osteosarcoma metastasis likely by regulating cell function through adhesion signaling. Further studies are necessary to fully explore the diagnostic and prognostic value of determining CD151 expression in osteosarcoma patients. PMID:27556355

Cell-geometry-dependent changes in plasma membrane order direct stem cell signalling and fate

NASA Astrophysics Data System (ADS)

von Erlach, Thomas C.; Bertazzo, Sergio; Wozniak, Michele A.; Horejs, Christine-Maria; Maynard, Stephanie A.; Attwood, Simon; Robinson, Benjamin K.; Autefage, Hélène; Kallepitis, Charalambos; del Río Hernández, Armando; Chen, Christopher S.; Goldoni, Silvia; Stevens, Molly M.

2018-03-01

Cell size and shape affect cellular processes such as cell survival, growth and differentiation1-4, thus establishing cell geometry as a fundamental regulator of cell physiology. The contributions of the cytoskeleton, specifically actomyosin tension, to these effects have been described, but the exact biophysical mechanisms that translate changes in cell geometry to changes in cell behaviour remain mostly unresolved. Using a variety of innovative materials techniques, we demonstrate that the nanostructure and lipid assembly within the cell plasma membrane are regulated by cell geometry in a ligand-independent manner. These biophysical changes trigger signalling events involving the serine/threonine kinase Akt/protein kinase B (PKB) that direct cell-geometry-dependent mesenchymal stem cell differentiation. Our study defines a central regulatory role by plasma membrane ordered lipid raft microdomains in modulating stem cell differentiation with potential translational applications.

The islet beta-cell: fuel responsive and vulnerable.

PubMed

Nolan, Christopher J; Prentki, Marc

2008-10-01

The pancreatic beta-cell senses blood nutrient levels and is modulated by neurohormonal signals so that it secretes insulin according to the need of the organism. Nutrient sensing involves marked metabolic activation, resulting in the production of coupling signals that promote insulin biosynthesis and secretion. The beta-cell's high capacity for nutrient sensing, however, necessitates reduced protection to nutrient toxicity. This potentially explains why in susceptible individuals, chronic fuel surfeit results in beta-cell failure and type 2 diabetes. Here we discuss recent insights into first, the biochemical basis of beta-cell signaling in response to glucose, amino acids and fatty acids, and second, beta-cell nutrient detoxification. We emphasize the emerging role of glycerolipid/fatty acid cycling in these processes.

Specialized proteasome subunits play an essential role in thymic selection of CD8+ T cells

PubMed Central

Kincaid, Eleanor Z.; Murata, Shigeo; Tanaka, Keiji; Rock, Kenneth L.

2016-01-01

The cells that stimulate positive selection express different specialized proteasome β-subunits than all other cells, including those involved in negative selection. Mice that lack all four specialized proteasome β-subunits, and therefore express only constitutive proteasomes in all cells, had a profound defect in the generation of CD8+ T cells. While a defect in positive selection would reflect an inability to generate the appropriate positively selecting peptides, a block at negative selection would point to the potential need to switch peptides between positive and negative selection to avoid the two processes often cancelling each other out. We found that the block in T cell development occurred around the checkpoints of positive and, surprisingly, also negative selection. PMID:27294792

Cell Surface Translocation of Annexin A2 Facilitates Glutamate-induced Extracellular Proteolysis*

PubMed Central

Valapala, Mallika; Maji, Sayantan; Borejdo, Julian; Vishwanatha, Jamboor K.

2014-01-01

Glutamate-induced elevation in intracellular Ca2+ has been implicated in excitotoxic cell death. Neurons respond to increased glutamate levels by activating an extracellular proteolytic cascade involving the components of the plasmin-plasminogen system. AnxA2 is a Ca2+-dependent phospholipid binding protein and serves as an extracellular proteolytic center by recruiting the tissue plasminogen activator and plasminogen and mediating the localized generation of plasmin. Ratiometric Ca2+ imaging and time-lapse confocal microscopy demonstrated glutamate-induced Ca2+ influx. We showed that glutamate translocated both endogenous and AnxA2-GFP to the cell surface in a process dependent on the activity of the NMDA receptor. Glutamate-induced translocation of AnxA2 is dependent on the phosphorylation of tyrosine 23 at the N terminus, and mutation of tyrosine 23 to a non-phosphomimetic variant inhibits the translocation process. The cell surface-translocated AnxA2 forms an active plasmin-generating complex, and this activity can be neutralized by a hexapeptide directed against the N terminus. These results suggest an involvement of AnxA2 in potentiating glutamate-induced cell death processes. PMID:24742684

Copper activates HIF-1α/GPER/VEGF signalling in cancer cells

PubMed Central

Rigiracciolo, Damiano Cosimo; Scarpelli, Andrea; Lappano, Rosamaria; Pisano, Assunta; Santolla, Maria Francesca; De Marco, Paola; Cirillo, Francesca; Cappello, Anna Rita; Dolce, Vincenza; Belfiore, Antonino; Maggiolini, Marcello; De Francesco, Ernestina Marianna

2015-01-01

Copper promotes tumor angiogenesis, nevertheless the mechanisms involved remain to be fully understood. We have recently demonstrated that the G-protein estrogen receptor (GPER) cooperates with hypoxia inducible factor-1α (HIF-1α) toward the regulation of the pro-angiogenic factor VEGF. Here, we show that copper sulfate (CuSO4) induces the expression of HIF-1α as well as GPER and VEGF in breast and hepatic cancer cells through the activation of the EGFR/ERK/c-fos transduction pathway. Worthy, the copper chelating agent TEPA and the ROS scavenger NAC prevented the aforementioned stimulatory effects. We also ascertained that HIF-1α and GPER are required for the transcriptional activation of VEGF induced by CuSO4. In addition, in human endothelial cells, the conditioned medium from breast cancer cells treated with CuSO4 promoted cell migration and tube formation through HIF-1α and GPER. The present results provide novel insights into the molecular mechanisms involved by copper in triggering angiogenesis and tumor progression. Our data broaden the therapeutic potential of copper chelating agents against tumor angiogenesis and progression. PMID:26415222

Chronic inhibition of glycogen synthase kinase-3 protects against rotenone-induced cell death in human neuron-like cells by increasing BDNF secretion.

PubMed

Giménez-Cassina, Alfredo; Lim, Filip; Díaz-Nido, Javier

2012-12-07

Mitochondrial dysfunction is a common feature of many neurodegenerative disorders. Likewise, activation of glycogen synthase kinase-3 (GSK-3) has been proposed to play an important role in neurodegeneration. This multifunctional protein kinase is involved in a number of cellular functions and we previously showed that chronic inhibition of GSK-3 protects neuronal cells against mitochondrial dysfunction-elicited cell death, through a mechanism involving increased glucose metabolism and the translocation of hexokinase II (HKII) to mitochondria. Here, we sought to gain deeper insight into the molecular basis of this neuroprotection. We found that chronic inhibition of GSK-3, either genetically or pharmacologically, elicited a marked increase in brain-derived neurotrophic factor (BDNF) secretion, which in turn conferred resistance to mitochondrial dysfunction through subcellular re-distribution of HKII. These results define a molecular pathway through which chronic inhibition of GSK-3 may protect neuronal cells from death. Moreover, they highlight the potential benefits of enhanced neurotrophic factor secretion as a therapeutic approach to treat neurodegenerative diseases. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Kif18A is involved in human breast carcinogenesis.

PubMed

Zhang, Chunpeng; Zhu, Changjun; Chen, Hongyan; Li, Linwei; Guo, Liping; Jiang, Wei; Lu, Shih Hsin

2010-09-01

Microtubule (MT) kinesin motor proteins orchestrate various cellular processes (e.g. mitosis, motility and organelle transportation) and have been implicated in human carcinogenesis. Kif18A, a plus-end directed MT depolymerase kinesin, regulates MT dynamics, chromosome congression and cell division. In this study, we report that Kif18A is overexpressed in human breast cancers and Kif18A overexpression is associated with tumor grade, metastasis and poor survival. Functional analyses reveal that ectopic overexpression of Kif18A results in cell multinucleation, whereas ablation of Kif18A expression significantly inhibits the proliferative capability of breast cancer cells in vitro and in vivo. Inhibition of Kif18A not only affects the critical mitotic function of Kif18A but also decreases cancer cell migration by stabilizing MTs at leading edges and ultimately induces anoikis of cells with inactivation of the phosphatidylinositol 3-kinase-Akt signaling pathway. Together, our results indicate that Kif18A is involved in human breast carcinogenesis and may serve as a potential therapeutic target for human breast cancer.

Deciphering early events involved in hyperosmotic stress-induced programmed cell death in tobacco BY-2 cells

PubMed Central

Monetti, Emanuela; Kadono, Takashi; Bouteau, François

2014-01-01

Hyperosmotic stresses represent one of the major constraints that adversely affect plants growth, development, and productivity. In this study, the focus was on early responses to hyperosmotic stress- (NaCl and sorbitol) induced reactive oxygen species (ROS) generation, cytosolic Ca2+ concentration ([Ca2+]cyt) increase, ion fluxes, and mitochondrial potential variations, and on their links in pathways leading to programmed cell death (PCD). By using BY-2 tobacco cells, it was shown that both NaCl- and sorbitol-induced PCD seemed to be dependent on superoxide anion (O2·–) generation by NADPH-oxidase. In the case of NaCl, an early influx of sodium through non-selective cation channels participates in the development of PCD through mitochondrial dysfunction and NADPH-oxidase-dependent O2·– generation. This supports the hypothesis of different pathways in NaCl- and sorbitol-induced cell death. Surprisingly, other shared early responses, such as [Ca2+]cyt increase and singlet oxygen production, do not seem to be involved in PCD. PMID:24420571