Sample records for affect cell proliferation
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Li, Junqin; Li, Xinhua; Hou, Ruixia; Liu, Ruifeng; Zhao, Xincheng; Dong, Feng; Wang, Chunfang; Yin, Guohua; Zhang, Kaiming
2015-09-01
Psoriasis is mediated primarily by T cells, which reduce epidermal turnover time and affect keratinocyte proliferation. We aimed to identify differentially expressed genes (DEG) in T cells from normal, five pairs of monozygotic twins concordant or discordant for psoriasis, to determine whether these DEG may account for the influence to epidermal turnover time and keratinocyte proliferation. The impact of T cells on keratinocyte proliferation and epidermal turnover time were investigated separately by immunohistochemistry and cultured with (3) H-TdR. mRNA expression patterns were investigated by RNA sequencing and verified by real-time reverse transcription polymerase chain reaction. After co-culture with psoriatic T cells, the expression of Ki-67, c-Myc and p53 increased, while expression of Bcl-2 and epidermal turnover time decreased. There were 14 DEG which were found to participate in the regulation of cell proliferation or differentiation. Psoriatic T cells exhibited the ability to decrease epidermal turnover time and affect keratinocyte proliferation because of the differential expression of PPIL1, HSPH1, SENP3, NUP54, FABP5, PLEKHG3, SLC9A9 and CHCHD4. © 2015 Japanese Dermatological Association.
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Activation of cutaneous immune responses in complex regional pain syndrome
Birklein, Frank; Drummond, Peter D.; Li, Wenwu; Schlereth, Tanja; Albrecht, Nahid; Finch, Philip M.; Dawson, Linda F.; Clark, J. David; Kingery, Wade S.
2014-01-01
The pathogenesis of complex regional pain syndrome (CRPS) is unresolved, but TNF-α and IL-6 are elevated in experimental skin blister fluid from CRPS affected limbs, as is tryptase, a marker for mast cells. In the rat fracture model of CRPS exaggerated sensory and sympathetic neural signaling stimulate keratinocyte and mast cell proliferation, causing the local production of high levels of inflammatory cytokines leading to pain behavior. The current investigation used CRPS patient skin biopsies to determine whether keratinocyte and mast cell proliferation occur in CRPS skin and to identify the cellular source of the up-regulated TNF-α, IL-6, and tryptase observed in CRPS experimental skin blister fluid. Skin biopsies were collected from the affected skin and the contralateral mirror site in 55 CRPS patients and the biopsy sections were immunostained for keratinocyte, cell proliferation, mast cell markers, TNF-α, and IL-6. In early CRPS keratinocytes were activated in the affected skin, resulting in proliferation, epidermal thickening, and up-regulated TNF-α and IL-6 expression. In chronic CRPS there was reduced keratinocyte proliferation with epidermal thinning in the affected skin. Acute CRPS patients also had increased mast cell accumulation in the affected skin, but there was no increase in mast cell numbers in chronic CRPS. PMID:24462502
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Neuronal models for evaluation of proliferation in vitro using high content screening.
Mundy, William R; Radio, Nicholas M; Freudenrich, Theresa M
2010-04-11
In vitro test methods can provide a rapid approach for the screening of large numbers of chemicals for their potential to produce toxicity (hazard identification). In order to identify potential developmental neurotoxicants, a battery of in vitro tests for neurodevelopmental processes such as cell proliferation, differentiation, growth, and synaptogenesis has been proposed. The development of in vitro approaches for toxicity testing will require choosing a model system that is appropriate to the endpoint of concern. This study compared several cell lines as models for neuronal proliferation. The sensitivities of neuronal cell lines derived from three species (PC12, rat; N1E-115, mouse; SH-SY5Y, human) to chemicals known to affect cell proliferation were assessed using a high content screening system. After optimizing conditions for cell growth in 96-well plates, proliferation was measured as the incorporation of 5-bromo-2'-deoxyuridine (BrdU) into replicating DNA during S phase. BrdU-labeled cells were detected by immunocytochemistry and cell counts were obtained using automated image acquisition and analysis. The three cell lines showed approximately 30-40% of the population in S phase after a 4h pulse of BrdU. Exposure to the DNA polymerase inhibitor aphidicolin for 20 h prior to the 4h pulse of BrdU significantly decreased proliferation in all three cell lines. The sensitivities of the cell lines were compared by exposure to eight chemicals known to affect proliferation (positive controls) and determination of the concentration inhibiting proliferation by 50% of control (I(50)). PC12 cells were the most sensitive to chemicals; 6 out of 8 chemicals (aphidicolin, cadmium, cytosine arabinoside, dexamethasone, 5-fluorouracil, and methylmercury) inhibited proliferation at the concentrations tested. SH-SY5Y cells were somewhat less sensitive to chemical effects, with five out of eight chemicals inhibiting proliferation; dexamethasone had no effect, and cadmium inhibited proliferation only at concentrations that decreased cell viability. Data from the N1E-115 cell line was extremely variable between experiments, and only 4 out of 8 chemicals resulted in inhibition of proliferation. Chemicals that had not been previously shown to alter proliferation (negative controls) did not affect proliferation or cell viability in any cell line. The results show that high content screening can be used to rapidly assess chemical effects on proliferation. Three neuronal cell lines exhibited differential sensitivity to the effect of chemicals on this endpoint, with PC12 cells being the most sensitive to inhibition of proliferation. Published by Elsevier Ireland Ltd.
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Velleman, Sandra G; Clark, Daniel L; Tonniges, Jeffrey R
2018-09-01
Posthatch skeletal muscle growth requires myogenic satellite cells and the dynamic expression of cell membrane-associated proteins. The membrane associated heparan sulfate proteoglycans, syndecan-4 and glypican-1, link the satellite cell niche to the intracellular environment. Sydnecan-4 and glypican-1 are differentially expressed with age in turkey satellite cells and their over-expression impacts both satellite cell proliferation and differentiation, but their effect on satellite cells from lines with different growth potentials is not known. The objective of the current study was to determine if syndecan-4 and glypican-1 regulation of satellite cell proliferation and differentiation is affected by age and growth selection. Pectoralis major satellite cells isolated at 1 d, 7 and 16-wk of age from a Randombred Control 2 (RBC2) line and a 16-wk body weight (F) line selected from the RBC2 line turkeys were studied. Syndecan-4 and glypican-1 expression was knocked down in both lines. The F-line cells proliferated faster than RBC2 line cells regardless of age, while differentiation tended to be greater in RBC2 line cells than F-line cells at each age. Syndecan-4 knockdown decreased proliferation at 7- and 16-wk but not 1 d cells, and increased differentiation at 1 d and 7 wk but not 16 wk cells. Glypican-1 knockdown differentially affected proliferation depending on cell age, whereas differentiation was decreased for 7- and 16-wk but not 1 d cells. These data suggest syndecan-4 and glypican-1 differentially affected satellite cell function in an age-dependent manner, but had little impact on differences in proliferation and differentiation due to growth selection. Copyright © 2018. Published by Elsevier Inc.
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Quiescence does not affect p53 and stress response by irradiation in human lung fibroblasts
DOE Office of Scientific and Technical Information (OSTI.GOV)
Dai, Jiawen; Itahana, Koji, E-mail: koji.itahana@duke-nus.edu.sg; Baskar, Rajamanickam, E-mail: r.baskar@nccs.com.sg
Cells in many organs exist in both proliferating and quiescent states. Proliferating cells are more radio-sensitive, DNA damage pathways including p53 pathway are activated to undergo either G{sub 1}/S or G{sub 2}/M arrest to avoid entering S and M phase with DNA damage. On the other hand, quiescent cells are already arrested in G{sub 0}, therefore there may be fundamental difference of irradiation response between proliferating and quiescent cells, and this difference may affect their radiosensitivity. To understand these differences, proliferating and quiescent human normal lung fibroblasts were exposed to 0.10–1 Gy of γ-radiation. The response of key proteins involvedmore » in the cell cycle, cell death, and metabolism as well as histone H2AX phosphorylation were examined. Interestingly, p53 and p53 phosphorylation (Ser-15), as well as the cyclin-dependent kinase inhibitors p21 and p27, were induced similarly in both proliferating and quiescent cells after irradiation. Furthermore, the p53 protein half-life, and expression of cyclin A, cyclin E, proliferating cell nuclear antigen (PCNA), Bax, or cytochrome c expression as well as histone H2AX phosphorylation were comparable after irradiation in both phases of cells. The effect of radioprotection by a glycogen synthase kinase 3β inhibitor on p53 pathway was also similar between proliferating and quiescent cells. Our results showed that quiescence does not affect irradiation response of key proteins involved in stress and DNA damage at least in normal fibroblasts, providing a better understanding of the radiation response in quiescent cells, which is crucial for tissue repair and regeneration. - Highlights: • p53 response by irradiation was similar between proliferating and quiescent cells. • Quiescent cells showed similar profiles of cell cycle proteins after irradiation. • Radioprotection of GSK-3β inhibitor caused similar effects between these cells. • Quiescence did not affect p53 response despite its known role in radio-resistance.« less
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Rindsjö, E; Joerink, M; Johansson, C; Bremme, K; Malmström, V; Scheynius, A
2010-07-01
It is hypothesized that the in utero environment in allergic mothers can affect the neonatal immune responses. The aim of this study was to analyse the effect of maternal allergic disease on cord blood mononuclear cell (CBMC) phenotype and proliferative responses upon allergen stimulation. Peripheral blood mononuclear cells (PBMC) from 12 allergic and 14 nonallergic mothers and CBMC from their children were analysed. In the mothers, we determined cell proliferation, production of IL-4 and expression of FOXP3 in response to allergen stimulation. In the children, we evaluated cell proliferation and FOXP3 expression following allergen stimulation. Furthermore, expression of different homing markers on T cells and regulatory T cells and maturity of the T cells and B cell subsets were evaluated directly ex vivo. The timothy- and birch-allergic mothers responded with increased proliferation and/or IL-4 production towards timothy and birch extract, respectively, when compared to nonallergic mothers. This could not be explained by impairment of FOXP3(+) regulatory T cells in the allergic mothers. CBMC proliferation and FOXP3 expression in response to allergens were not affected by the allergic status of the mother. Also, phenotype of T cells, FOXP3(+) regulatory T cells and B cells was not affected by the allergic status of the mother. Our results suggest that maternal allergic disease has no effect on the neonatal response to allergens or the phenotype of neonatal lymphocytes. The factors studied here could, however, still affect later development of allergy.
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Pyruvate kinase isoform expression alters nucleotide synthesis to impact cell proliferation
Lunt, Sophia Y.; Muralidhar, Vinayak; Hosios, Aaron M.; Israelsen, William J.; Gui, Dan Y.; Newhouse, Lauren; Ogrodzinski, Martin; Hecht, Vivian; Xu, Kali; Acevedo, Paula N. Marín; Hollern, Daniel P.; Bellinger, Gary; Dayton, Talya L.; Christen, Stefan; Elia, Ilaria; Dinh, Anh T.; Stephanopoulos, Gregory; Manalis, Scott R.; Yaffe, Michael B.; Andrechek, Eran R.; Fendt, Sarah-Maria; Heiden, Matthew G. Vander
2014-01-01
SUMMARY Metabolic regulation influences cell proliferation. The influence of pyruvate kinase isoforms on tumor cells has been extensively studied, but whether PKM2 is required for normal cell proliferation is unknown. We examine how PKM2-deletion affects proliferation and metabolism in non-transformed, non-immortalized PKM2-expressing primary cells. We find that deletion of PKM2 in primary cells results in PKM1 expression and proliferation arrest. PKM1 expression, rather than PKM2 loss, is responsible for this effect, and proliferation arrest cannot be explained by cell differentiation, senescence, death, changes in gene expression, or prevention of cell growth. Instead, PKM1 expression impairs nucleotide production and the ability to synthesize DNA and progress through the cell cycle. Nucleotide biosynthesis is limiting, as proliferation arrest is characterized by severe thymidine depletion, and supplying exogenous thymine rescues both nucleotide levels and cell proliferation. Thus, PKM1 expression promotes a metabolic state that is unable to support DNA synthesis. PMID:25482511
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Anti-proliferative and gene expression actions of resveratrol in breast cancer cells in vitro
Yang, Sheng-Huei; Tsai, Po-Wei; Wang, Shwu-Huey; Wang, Ching-Chiung; Lee, Yee-Shin; Cheng, Guei-Yun; HuangFu, Wei-Chun; London, David; Tang, Heng-Yuan; Fu, Earl; Yen, Yun; Liu, Leroy F.; Lin, Hung-Yun; Davis, Paul J.
2014-01-01
We have used a perfusion bellows cell culture system to investigate resveratrolinduced anti-proliferation/apoptosis in a human estrogen receptor (ER)-negative breast cancer cell line (MDA-MB-231). Using an injection system to perfuse media with stilbene, we showed resveratrol (0.5 – 100 μM) to decrease cell proliferation in a concentration-dependent manner. Comparison of influx and medium efflux resveratrol concentrations revealed rapid disappearance of the stilbene, consistent with cell uptake and metabolism of the agent reported by others. Exposure of cells to 10 μM resveratrol for 4 h daily × 6 d inhibited cell proliferation by more than 60%. Variable extracellular acid-alkaline conditions (pH 6.8 – 8.6) affected basal cell proliferation rate, but did not alter anti-proliferation induced by resveratrol. Resveratrol-induced gene expression, including transcription of the most up-regulated genes and pro-apoptotic p53-dependent genes, was not affected by culture pH changes. The microarray findings in the context of induction of anti-proliferation with brief daily exposure of cells to resveratrol—and rapid disappearance of the compound in the perfusion system—are consistent with existence of an accessible initiation site for resveratrol actions on tumor cells, e.g., the cell surface receptor for resveratrol described on integrin αvβ3. PMID:25436977
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Saito, Yuki; Iwatsuki, Ken; Hanyu, Hikaru
We investigated the effects of essential amino acids on intestinal stem cell proliferation and differentiation using murine small intestinal organoids (enteroids) from the jejunum. By selectively removing individual essential amino acids from culture medium, we found that 24 h of methionine (Met) deprivation markedly suppressed cell proliferation in enteroids. This effect was rescued when enteroids cultured in Met deprivation media for 12 h were transferred to complete medium, suggesting that Met plays an important role in enteroid cell proliferation. In addition, mRNA levels of the stem cell marker leucine-rich repeat-containing G protein-coupled receptor 5 (Lgr5) decreased in enteroids grown in Met deprivationmore » conditions. Consistent with this observation, Met deprivation also attenuated Lgr5-EGFP fluorescence intensity in enteroids. In contrast, Met deprivation enhanced mRNA levels of the enteroendocrine cell marker chromogranin A (ChgA) and markers of K cells, enterochromaffin cells, goblet cells, and Paneth cells. Immunofluorescence experiments demonstrated that Met deprivation led to an increase in the number of ChgA-positive cells. These results suggest that Met deprivation suppresses stem cell proliferation, thereby promoting differentiation. In conclusion, Met is an important nutrient in the maintenance of intestinal stem cells and Met deprivation potentially affects cell differentiation. - Highlights: • Met influences the proliferation of enteroids. • Met plays a crucial role in the maintenance of stem cells. • Met deprivation potentially promotes differentiation into secretory cells.« less
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Low doses of TiO2-polyethylene glycol nanoparticles stimulate proliferation of hepatocyte cells
NASA Astrophysics Data System (ADS)
Sun, Qingqing; Kanehira, Koki; Taniguchi, Akiyoshi
2016-01-01
This paper describes the effect of low concentrations of 100 nm polyethylene glycol-modified TiO2 nanoparticles (TiO2-PEG NPs) on HepG2 hepatocellular carcinoma cells. Proliferation of HepG2 cells increased significantly when the cells were exposed to low doses (<100 μg ml-1) of TiO2-PEG NPs. These results were further confirmed by cell counting experiments and cell cycle assays. Cellular uptake assays were performed to determine why HepG2 cells proliferate with low-dose exposure to TiO2-PEG NPs. The results showed that exposure to lower doses of NPs led to less cellular uptake, which in turn decreased cytotoxicity. We therefore hypothesized that TiO2-PEG NPs could affect the activity of hepatocyte growth factor receptors (HGFRs), which bind to hepatocyte growth factor and stimulate cell proliferation. The localization of HGFRs on the surface of the cell membrane was detected via immunofluorescence staining and confocal microscopy. The results showed that HGFRs aggregate after exposure to TiO2-PEG NPs. In conclusion, our results indicate that TiO2-PEG NPs have the potential to promote proliferation of HepG2 cells through HGFR aggregation and suggest that NPs not only exhibit cytotoxicity but also affect cellular responses.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Reinbothe, Susann; Larsson, Anna-Maria; Vaapil, Marica
Highlights: • New anti-human EPOR antibody confirms full-length EPOR expression in breast cancer cells. • Proliferation of breast cancer cells is not affected by rhEPO treatment in vitro. • EPOR knockdown impairs proliferation of ERa positive breast cancer cells. • EPOR knockdown reduces AKT phosphorylation and ERa activity. - Abstract: The main function of Erythropoietin (EPO) and its receptor (EPOR) is the stimulation of erythropoiesis. Recombinant human EPO (rhEPO) is therefore used to treat anemia in cancer patients. However, clinical trials have indicated that rhEPO treatment might promote tumor progression and has a negative effect on patient survival. In addition,more » EPOR expression has been detected in several cancer forms. Using a newly produced anti-EPOR antibody that reliably detects the full-length isoform of the EPOR we show that breast cancer tissue and cells express the EPOR protein. rhEPO stimulation of cultured EPOR expressing breast cancer cells did not result in increased proliferation, overt activation of EPOR (receptor phosphorylation) or a consistent activation of canonical EPOR signaling pathway mediators such as JAK2, STAT3, STAT5, or AKT. However, EPOR knockdown experiments suggested functional EPO receptors in estrogen receptor positive (ERα{sup +}) breast cancer cells, as reduced EPOR expression resulted in decreased proliferation. This effect on proliferation was not seen in ERα negative cells. EPOR knockdown decreased ERα activity further supports a mechanism by which EPOR affects proliferation via ERα-mediated mechanisms. We show that EPOR protein is expressed in breast cancer cells, where it appears to promote proliferation by an EPO-independent mechanism in ERα expressing breast cancer cells.« less
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Parker, Aimee; Maclaren, Oliver J.; Fletcher, Alexander G.; Muraro, Daniele; Kreuzaler, Peter A.; Byrne, Helen M.; Maini, Philip K.; Watson, Alastair J. M.; Pin, Carmen
2017-01-01
The functional integrity of the intestinal epithelial barrier relies on tight coordination of cell proliferation and migration, with failure to regulate these processes resulting in disease. It is not known whether cell proliferation is sufficient to drive epithelial cell migration during homoeostatic turnover of the epithelium. Nor is it known precisely how villus cell migration is affected when proliferation is perturbed. Some reports suggest that proliferation and migration may not be related while other studies support a direct relationship. We used established cell-tracking methods based on thymine analog cell labeling and developed tailored mathematical models to quantify cell proliferation and migration under normal conditions and when proliferation is reduced and when it is temporarily halted. We found that epithelial cell migration velocities along the villi are coupled to cell proliferation rates within the crypts in all conditions. Furthermore, halting and resuming proliferation results in the synchronized response of cell migration on the villi. We conclude that cell proliferation within the crypt is the primary force that drives cell migration along the villus. This methodology can be applied to interrogate intestinal epithelial dynamics and characterize situations in which processes involved in cell turnover become uncoupled, including pharmacological treatments and disease models.—Parker, A., Maclaren, O. J., Fletcher, A. G., Muraro, D., Kreuzaler, P. A., Byrne, H. M., Maini, P. K., Watson, A. J. M., Pin, C. Cell proliferation within small intestinal crypts is the principal driving force for cell migration on villi. PMID:27811059
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Lew, Carolyn Ritterson; Tolan, Dean R.
2012-01-01
In cancer, glucose uptake and glycolysis are increased regardless of the oxygen concentration in the cell, a phenomenon known as the Warburg effect. Several (but not all) glycolytic enzymes have been investigated as potential therapeutic targets for cancer treatment using RNAi. Here, four previously untargeted glycolytic enzymes, aldolase A, glyceraldehyde 3-phosphate dehydrogenase, triose phosphate isomerase, and enolase 1, are targeted using RNAi in Ras-transformed NIH-3T3 cells. Of these enzymes, knockdown of aldolase causes the greatest effect, inhibiting cell proliferation by 90%. This defect is rescued by expression of exogenous aldolase. However, aldolase knockdown does not affect glycolytic flux or intracellular ATP concentration, indicating a non-metabolic cause for the cell proliferation defect. Furthermore, this defect could be rescued with an enzymatically dead aldolase variant that retains the known F-actin binding ability of aldolase. One possible model for how aldolase knockdown may inhibit transformed cell proliferation is through its disruption of actin-cytoskeleton dynamics in cell division. Consistent with this hypothesis, aldolase knockdown cells show increased multinucleation. These results are compared with other studies targeting glycolytic enzymes with RNAi in the context of cancer cell proliferation and suggest that aldolase may be a useful target in the treatment of cancer. PMID:23093405
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Warchol, Mark E
2002-04-01
Sensory hair cells in the inner ears of nonmammalian vertebrates can regenerate after injury. In many species, replacement hair cells are produced by the proliferation of epithelial supporting cells. Thus, the ability of supporting cells to undergo renewed proliferation is a key determinant of regenerative ability. The present study used cultures of isolated inner ear sensory epithelia to identify cellular signals that regulate supporting cell proliferation. Small pieces of sensory epithelia from the chicken utricle were cultured in glass microwells. Under those conditions, cell proliferation was inversely related to local cell density. The signaling molecules N-cadherin, beta-catenin, and focal adhesion kinase were immunolocalized in the cultured epithelial cells, and high levels of phosphotyrosine immunoreactivity were present at cell-cell junctions and focal contacts of proliferating cells. Binding of microbeads coated with a function-blocking antibody to N-cadherin inhibited ongoing proliferation. The growth of epithelial cells was also affected by the density of extracellular matrix molecules. The results suggest that cell density, cell-cell contact, and the composition of the extracellular matrix may be critical influences on the regulation of sensory regeneration in the inner ear.
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Lea, Michael A; Qureshi, Mehreen S; Buxhoeveden, Michael; Gengel, Nicolette; Kleinschmit, Jessica; Desbordes, Charles
2013-02-01
In previous studies performed by our group, we observed that 2-deoxyglucose blocked the acidification of the medium used for culture of colon cancer cells caused by incubation with biguanides and it had an additive inhibitory effect on growth. In the present work, we found that 3-bromopyruvate can also prevent the lowering of pH caused by biguanide treatment. 3-Bromopyruvate inhibited colonic cancer cell proliferation, but the effect was not always additive to that of biguanides and an additive effect was more notable in combined treatment with 3-bromopyruvate and 2-deoxyglucose. The induction of alkaline phosphatase activity by butyrate was not consistently affected by combination with other agents that modified glucose metabolism. The drug combinations that were examined inhibited proliferation of wild-type and p53-null cells and affected colonic cancer lines with different growth rates.
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Lea, Michael A.; Qureshi, Mehreen S.; Buxhoeveden, Michael; Gengel, Nicolette; Kleinschmit, Jessica; desBordes, Charles
2013-01-01
In previous studies we observed that 2-deoxyglucose blocked the acidification of the medium used for culture of colon cancer cells caused by incubation with biguanides and had an additive inhibitory effect on growth. In the present work, we found that 3-bromopyruvate can also prevent the lowering of pH caused by biguanide treatment. 3-Bromopyruvate inhibited colonic cancer cell proliferation but the effect was not always additive to that of biguanides and an additive effect was more notable in combined treatment with 3-bromopyruvate and 2-deoxyglucose. The induction of alkaline phosphatase activity by butyrate was not consistently affected by combination with other agents that modified glucose metabolism. The drug combinations that were examined inhibited proliferation of wild-type and P53 null cells and affected colonic cancer lines with different growth rates. PMID:23393330
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Zhou, Nan; Hao, Shuang; Huang, Zongqiang; Wang, Weiwei; Yan, Penghui; Zhou, Wei; Zhu, Qihang; Liu, Xiaokang
2018-01-01
Objective Neural stem cells play an important role in the recovery and regeneration of peripheral nerve injury, and the microRNA-7 (miR-7) regulates differentiation of neural stem cells. This study aimed to explore the role of miR-7 in neural stem cells homing and proliferation and its influence on peripheral nerve injury repair. Methods The mice model of peripheral nerve injury was created by segmental sciatic nerve defect (sciatic nerve injury), and neural stem cells treatment was performed with a gelatin hydrogel conduit containing neural stem cells inserted into the sciatic nerve injury mice. The Sciatic Function Index was used to quantify sciatic nerve functional recovery in the mice. The messenger RNA and protein expression were detected by reverse transcription polymerase chain reaction and Western blot, respectively. Luciferase reporter assay was used to confirm the binding between miR-7 and the 3'UTR of cell division cycle protein 42 (cdc42). The neural stem cells migration and proliferation were analyzed by transwell assay and a Cell-LightTM EdU DNA Cell Proliferation kit, respectively. Results Neural stem cells treatment significantly promoted nerve repair in sciatic nerve injury mice. MiR-7 expression was decreased in sciatic nerve injury mice with neural stem cells treatment, and miR-7 mimic transfected into neural stem cells suppressed migration and proliferation, while miR-7 inhibitor promoted migration and proliferation. The expression level and effect of cdc42 on neural stem cells migration and proliferation were opposite to miR-7, and the luciferase reporter assay proved that cdc42 was a target of miR-7. Using co-transfection into neural stem cells, we found pcDNA3.1-cdc42 and si-cdc42 could reverse respectively the role of miR-7 mimic and miR-7 inhibitor on neural stem cells migration and proliferation. In addition, miR-7 mimic-transfected neural stem cells could abolish the protective role of neural stem cells on peripheral nerve injury. Conclusion MiR-7 inhibited peripheral nerve injury repair by affecting neural stem cells migration and proliferation through cdc42.
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Inhibition of brain tumor cell proliferation by alternating electric fields
DOE Office of Scientific and Technical Information (OSTI.GOV)
Jeong, Hyesun; Oh, Seung-ick; Hong, Sunghoi, E-mail: shong21@korea.ac.kr, E-mail: radioyoon@korea.ac.kr
2014-11-17
This study was designed to investigate the mechanism by which electric fields affect cell function, and to determine the optimal conditions for electric field inhibition of cancer cell proliferation. Low-intensity (<2 V/cm) and intermediate-frequency (100–300 kHz) alternating electric fields were applied to glioblastoma cell lines. These electric fields inhibited cell proliferation by inducing cell cycle arrest and abnormal mitosis due to the malformation of microtubules. These effects were significantly dependent on the intensity and frequency of applied electric fields.
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Nitric oxide regulates stretch-induced proliferation in C2C12 myoblasts.
Soltow, Quinlyn A; Lira, Vitor A; Betters, Jenna L; Long, Jodi H D; Sellman, Jeff E; Zeanah, Elizabeth H; Criswell, David S
2010-09-01
Mechanical stretch of skeletal muscle activates nitric oxide (NO) production and is an important stimulator of satellite cell proliferation. Further, cyclooxygenase (COX) activity has been shown to promote satellite cell proliferation in response to stretch. Since COX-2 expression in skeletal muscle can be regulated by NO we sought to determine if NO is required for stretch-induced myoblast proliferation and whether supplemental NO can counter the effects of COX-2 and NF-kappaB inhibitors. C2C12 myoblasts were cultured for 24 h, then switched to medium containing either the NOS inhibitor, L-NAME (200 microM), the COX-2 specific inhibitor NS-398 (100 microM), the NF-kappaB inhibiting antioxidant, PDTC (5 mM), the nitric oxide donor, DETA-NONOate (10-100 microM) or no supplement (control) for 24 h. Subgroups of each treatment were exposed to 1 h of 15% cyclic stretch (1 Hz), and were then allowed to proliferate for 24 h before fixing. Proliferation was measured by BrdU incorporation during the last hour before fixing, and DAPI stain. Stretch induced a twofold increase in nuclear number compared to control, and this effect was completely inhibited by L-NAME, NS-398 or PDTC (P < 0.05). Although DETA-NONOate (10 microM) did not affect basal proliferation, the NO-donor augmented the stretch-induced increase in proliferation and rescued stretch-induced proliferation in NS-398-treated cells, but not in PDTC-treated cells. In conclusion, NO, COX-2, and NF-kappaB are necessary for stretch-induced proliferation of myoblasts. Although COX-2 and NF-kappaB are both involved in basal proliferation, NO does not affect basal growth. Thus, NO requires the synergistic effect of stretch in order to induce muscle cell proliferation.
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Jin, Xiu-Li; Sun, Qin-Sheng; Liu, Feng; Yang, Hong-Wei; Liu, Min; Liu, Hong-Xia; Xu, Wei; Jiang, Yu-Yang
2013-07-01
Transcriptional repressor Pokemon is a critical factor in embryogenesis, development, cell proliferation, differentiation, and oncogenesis, thus behaving as an oncogene. Oncomine database suggests a potential correlation between the expressions of Pokemon and Sprouty1. This study investigated the regulatory role of Pokemon in Sprouty1 expression and the effect on liver cancer cell growth and proliferation, revealing a novel miR-21-mediated regulatory circuit. In normal (HL-7702) and cancer (QGY-7703) liver cell lines, Sprouty1 expression is inversely correlated with Pokemon levels. Targeted expression or siRNA-mediated silencing showed that Pokemon is a repressor of Sprouty1 expression at both mRNA and protein levels, but Pokemon cannot affect the promoter activity of Sprouty1. Sprouty1 is a target of miR-21 and interestingly, we found that miR-21 is up-regulated by Pokemon in liver cancer cells. Luciferase reporter assays showed that Pokemon up-regulated miR-21 transcription in a dose-dependent manner, and ChIP assay exhibited a direct binding of Pokemon to the miR-21 promoter at -747 to -399 bp. Site-directed mutagenesis of the GC boxes at -684 to -679 bp and -652 to -647 bp of miR-21 promoter abolished the regulatory activity by Pokemon. Furthermore, we found that the modulation of Pokemon and miR-21 expression affected the growth and proliferation of liver cancer cells QGY-7703. In summary, our findings demonstrate that Pokemon suppresses Sprouty1 expression through a miR-21-mediated mechanism, affecting the growth and proliferation of liver cancer cells. This study recognized miR-21 and Sprouty1 as novel targets of the Pokemon regulatory network. Copyright © 2013 Wiley Periodicals, Inc.
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Dose-related cell proliferation in medaka (Oryzias latipes) after N-nitrosodiethylamine exposure
DOE Office of Scientific and Technical Information (OSTI.GOV)
Ortego, L.S.; Hawkins, W.E.; Walker, W.W.
1994-12-31
Cell proliferation is important in toxic and carcinogenic mechanisms. Carcinogens such as N-nitrosodiethylamine (DEN) that cause necrotizing injury stimulate cell proliferation as part of an injury-repair mechanism. A stimulus to cell division in an organ with a low rate of cell division, such as the liver, may initiate or enhance the carcinogenicity of a chemical. The authors examined the effect of DEN exposure on cell proliferation in the liver of medaka (Oryzias latipes). Two age groups (6 and 56 days post-hatch) were exposed to DEN continuously at 5 doses (0.0, 2.5, 5.0, 10.0, and 20.0 ppm) for 28 days. Cellmore » proliferation was measured using the proliferating cell nuclear antigen (PCNA) assay two months post-initiation of DEN exposure. The assay involves monoclonal antibody detection of PCNA, an auxiliary protein of DNA polymerase delta which is, expressed during cell division. Results suggested that cell proliferation paralleled the DEN dose and that age at initiation of exposure did not affect this relationship. The increase in cell proliferation appeared to be a sustained response from that initiated during DEN exposure. The study suggests that cell proliferation in medaka is an important component in carcinogenesis and is related to carcinogen exposure dose.« less
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Lee, Jong-Sun; Kim, Jeong-Yub; Kim, Hee-Jin; Kim, Jeong Cheol; Lee, Jae-Seon; Kim, Nam; Park, Myung-Jin
2016-01-01
Alzheimer's disease (AD) is the most common progressive and irreversible neurodegenerative disease and it is caused by neuronal death in the brain. Recent studies have shown that non-ionizing radiofrequency (RF) radiation has some beneficial cognitive effects in animal models of AD. In this study, we examined the effect of combined RF radiation on amyloid-beta (Aβ)–induced cytotoxicity in HT22 rat hippocampal neurons. Treatment with Aβ suppressed HT22 cell proliferation in a concentration-dependent manner. RF exposure did not affect cell proliferation, and also had a marginal effect on Aβ-induced suppression of growth in HT22 cells. Cell cycle analysis showed that Aβ decreased the G1 fraction and increased the subG1 fraction, indicating increased apoptosis. Accordingly, Aβ increased the annexin V/propidium iodide (PI)–positive cell fraction and the degradation of poly (ADP ribose) polymerase and caspase-3 in HT22 cells. However, RF alone and the combination of Aβ and RF did not affect these events significantly. Aβ increased reactive oxygen species (ROS) generation, thereby suppressing cell proliferation. This was abrogated by N-acetylcysteine (NAC) treatment, indicating that Aβ-induced ROS generation is the main cause of suppression of proliferation. NAC also restored Aβ-induced annexin V/PI–positive cell populations. However, RF did not have a significant impact on these events. Finally, Aβ stimulated the ataxia telangiectasia and Rad3-related protein/checkpoint kinase 1 DNA single-strand breakage pathway, and enhanced beta-site amyloid precursor protein expression; RF had no effect on them. Taken together, our results demonstrate that RF exposure did not significantly affect the Aβ-induced decrease of cell proliferation, increase of ROS production, or induction of cell death in these cells. PMID:27325640
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Sousa-Ferreira, Lígia; Aveleira, Célia; Botelho, Mariana; Álvaro, Ana Rita; Pereira de Almeida, Luís; Cavadas, Cláudia
2014-01-01
A significant number of children undergo maternal exposure to antidepressants and they often present low birth weight. Therefore, it is important to understand how selective serotonin reuptake inhibitors (SSRIs) affect the development of the hypothalamus, the key center for metabolism regulation. In this study we investigated the proliferative actions of fluoxetine in fetal hypothalamic neuroprogenitor cells and demonstrate that fluoxetine induces the proliferation of these cells, as shown by increased neurospheres size and number of proliferative cells (Ki-67+ cells). Moreover, fluoxetine inhibits the differentiation of hypothalamic neuroprogenitor cells, as demonstrated by decreased number of mature neurons (Neu-N+ cells) and increased number of undifferentiated cells (SOX-2+ cells). Additionally, fluoxetine-induced proliferation and maintenance of hypothalamic neuroprogenitor cells leads to changes in the mRNA levels of appetite regulator neuropeptides, including Neuropeptide Y (NPY) and Cocaine-and-Amphetamine-Regulated-Transcript (CART). This study provides the first evidence that SSRIs affect the development of hypothalamic neuroprogenitor cells in vitro with consequent alterations on appetite neuropeptides. PMID:24598761
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Cell proliferation of Paramecium tetraurelia on a slow rotating clinostat
NASA Astrophysics Data System (ADS)
Sawai, Satoe; Mogami, Yoshihiro; Baba, Shoji A.
Paramecium is known to proliferate faster under microgravity conditions, and slower under hypergravity. Experiments using axenic culture medium have demonstrated that hypergravity affected directly on the proliferation of Paramecium itself. In order to assess the mechanisms underlying the physiological effects of gravity on cell proliferation, Paramecium tetraurelia was grown under clinorotation (2.5 rpm) and the time course of the proliferation was investigated in detail on the basis of the logistic analysis. On the basis of the mechanical properties of Paramecium, this slow rate of the rotation appears to be enough to simulate microgravity in terms of the randomization of the cell orientation with respect to gravity. P. tetraurelia was cultivated in a closed chamber in which cells were confined without air bubbles, reducing the shear forces and turbulences under clinorotation. The chamber is made of quartz and silicone rubber film; the former is for the optically-flat walls for the measurement of cell density by means of a non-invasive laser optical-slice method, and the latter for gas exchange. Because of the small dimension for culture space, Paramecium does not accumulate at the top of the chamber in spite of its known negative gravitactic behavior. We measured the cell density at regular time intervals without breaking the configuration of the chamber, and analyzed the proliferation parameters by fitting the data to a logistic equation. As a result, P. tetraurelia showed reduced proliferation under slow clinorotation. The saturation of the cell density as well as the maximum proliferation rate decreased, although we found no significant changes on the half maximal time for proliferation. We also found that the mean swimming velocity decreased under slow clinorotation. These results were not consistent with those under microgravity and fast rotating clinostat. This may suggest that randomization of the cell orientation performed by slow rotating clinostat has not the same effect on Paramecium as that under microgravity that may affect the proliferation as the result of the reduced cost of propulsion.
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Low doses of TiO2-polyethylene glycol nanoparticles stimulate proliferation of hepatocyte cells
Sun, Qingqing; Kanehira, Koki; Taniguchi, Akiyoshi
2016-01-01
Abstract This paper describes the effect of low concentrations of 100 nm polyethylene glycol-modified TiO2 nanoparticles (TiO2-PEG NPs) on HepG2 hepatocellular carcinoma cells. Proliferation of HepG2 cells increased significantly when the cells were exposed to low doses (<100 μg ml–1) of TiO2-PEG NPs. These results were further confirmed by cell counting experiments and cell cycle assays. Cellular uptake assays were performed to determine why HepG2 cells proliferate with low-dose exposure to TiO2-PEG NPs. The results showed that exposure to lower doses of NPs led to less cellular uptake, which in turn decreased cytotoxicity. We therefore hypothesized that TiO2-PEG NPs could affect the activity of hepatocyte growth factor receptors (HGFRs), which bind to hepatocyte growth factor and stimulate cell proliferation. The localization of HGFRs on the surface of the cell membrane was detected via immunofluorescence staining and confocal microscopy. The results showed that HGFRs aggregate after exposure to TiO2-PEG NPs. In conclusion, our results indicate that TiO2-PEG NPs have the potential to promote proliferation of HepG2 cells through HGFR aggregation and suggest that NPs not only exhibit cytotoxicity but also affect cellular responses. PMID:27877913
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Lee, Yunkyoung; Lee, Bongkyu; Jeong, Sumin; Park, Ji-Won; Han, Inn-Oc; Lee, Chang-Joong
2016-08-26
Physostigmine, an acetylcholinesterase inhibitor, is known to affect the brain function in various aspects. This study was conducted to test whether physostigmine affects cell proliferation in the telencephalon of zebrafish. BrdU-labeled cells was prominently observed in the ventral zone of the ventral telencephalon of zebrafish. The increased number of BrdU- and proliferating cell nuclear antigen-labeled cells were shown in zebrafish treated with 200μM physostigmine, which was inhibited by pretreatment with 200μM scopolamine. iNOS mRNA expression was increased in the brain of zebrafish treated with 200μM physostigmine. Consistently, aminoguanidine, an iNOS inhibitor, attenuated the increase in the number of BrdU-labeled cells by physostigmine treatment. Zebrafish also showed seizure-like locomotor activity characterized by a rapid and abrupt movement during a 30min treatment with 200μM physostigmine. Neural activity in response to an electrical stimulus was increased in the isolated telencephalon of zebrafish continuously perfused with 200μM physostigmine. None of the number of BrdU-labeled cells, neural activity, or locomotor activity was affected by treatment with 20μM physostigmine. These results suggest that 200μM physostigmine increased neural activity and induced cell proliferation via nitric oxide production in zebrafish. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.
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Kitajima, Ken; Miura, Shin-Ichiro; Mastuo, Yoshino; Uehara, Yoshinari; Saku, Keijiro
2009-03-01
Peroxisome proliferator-activated receptor-alpha (PPAR-alpha) is a key regulator of lipid and glucose metabolism and has been implicated in inflammation. The vascular effects of activator for PPARs, particularly PPAR-alpha, on vascular cells remain to be fully elucidated. Therefore, we analyzed the hypothesis that newly developed (R)-K-13675 decreases the secretion of inflammatory markers without affecting cell proliferation or tube formation. Human coronary endothelial cells (HCECs) were maintained in different doses of (R)-K-13675 under serum starvation. After 20h, the levels of monocyte chemoattractant protein-1 (MCP-1), regulated on activation, normal T expressed and secreted (RANTES), interleukin-6 (IL-6) and interferon-gamma (INF-gamma) secreted in the medium and nuclear factor kappa B (NFkappaB) in cell lysate were analyzed using enzyme-linked immunosorbent assays (ELISA). Upon treatment with (R)-K-13675 at 0, 10, 20, 50 and 100nM, with the inflammatory markers at 0nM as 100 (arbitrary units), MCP-1 levels were significantly suppressed (94+/-9, 88+/-2, 80+/-5 and 74+/-11, respectively). RANTES, IL-6 and INF-gamma levels were also significantly suppressed (RANTES: 92+/-2, 74+/-9, 64+/-7 and 60+/-2, respectively, IL-6: 97+/-2, 89+/-10, 82+/-1 and 66+/-7, respectively, INF-gamma: 98+/-7, 94+/-3, 76+/-8 and 64+/-8, respectively). NFkappaB levels were also decreased to 91+/-5, 90+/-5, 84+/-7 and 82+/-8, respectively. In addition, (R)-K-13675 did not affect HCEC proliferation or tube formation at up to 100nM. Thus, (R)-K-13675 was associated with the inhibition of inflammatory responses without affecting cell proliferation or angiogenesis, and subsequently may induce an anti-atherosclerotic effect.
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Phenolic Compounds in Extra Virgin Olive Oil Stimulate Human Osteoblastic Cell Proliferation.
García-Martínez, Olga; De Luna-Bertos, Elvira; Ramos-Torrecillas, Javier; Ruiz, Concepción; Milia, Egle; Lorenzo, María Luisa; Jimenez, Brigida; Sánchez-Ortiz, Araceli; Rivas, Ana
2016-01-01
In this study, we aimed to clarify the effects of phenolic compounds and extracts from different extra virgin olive oil (EVOO) varieties obtained from fruits of different ripening stages on osteoblast cells (MG-63) proliferation. Cell proliferation was increased by hydroxytyrosol, luteolin, apigenin, p-coumaric, caffeic, and ferulic acids by approximately 11-16%, as compared with controls that were treated with one vehicle alone, while (+)-pinoresinol, oleuropein, sinapic, vanillic acid and derivative (vanillin) did not affect cell proliferation. All phenolic extracts stimulated MG-63 cell growth, and they induced higher cell proliferation rates than individual compounds. The most effective EVOO phenolic extracts were those obtained from the Picual variety, as they significantly increased cell proliferation by 18-22%. Conversely, Arbequina phenolic extracts increased cell proliferation by 9-13%. A decline in osteoblast proliferation was observed in oils obtained from olive fruits collected at the end of the harvest period, as their total phenolic content decreases at this late stage. Further research on the signaling pathways of olive oil phenolic compounds involved in the processes and their metabolism should be carried out to develop new interventions and adjuvant therapies using EVOO for bone health (i.e.osteoporosis) in adulthood and the elderly.
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Nitric acid passivation does not affect in vitro biocompatibility of titanium.
Faria, Adriana C L; Beloti, Márcio M; Rosa, Adalberto L
2003-01-01
In general, both chemical composition and surface features of implants affect cell response. The aim of this study was to evaluate the effect of titanium (Ti) passivation on the response of rat bone marrow cells, considering cell attachment, cell morphology, cell proliferation, total protein content, alkaline phosphatase (ALP) activity, and bonelike nodule formation. Cells were cultured on both commercially pure titanium (cpTi) and titanium-aluminium-vanadium alloy (Ti-6Al-4V) discs, either passivated or not. For attachment evaluation, cells were cultured for 4 and 24 hours. Cell morphology was evaluated after 4 days. After 7, 14, and 21 days, cell proliferation, total protein content, and ALP activity were evaluated. Bonelike nodule formation was evaluated after 21 days. Data were compared by analysis of variance and the Duncan multiple range test. Cell attachment, cell morphology, cell proliferation, total protein content, ALP activity, and bonelike nodule formation all were unaffected by Ti composition or passivation. Although the protocol for passivation used here could interfere with the pattern of ions released from Ti-6Al-4V and cpTi surfaces, the present study did not show any effect of this surface treatment on in vitro biocompatibility of Ti as evaluated by osteoblast attachment, proliferation, and differentiation.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Sato, Tsuyoshi; Abe, Takahiro; Nakamoto, Norimichi
Recent studies have suggested that nicotine critically affects bone metabolism. Many studies have examined the effects of nicotine on proliferation and differentiation, but the underlying molecular mechanisms remain unclear. We examined cell cycle regulators involved in the proliferation and differentiation of MC3T3-E1 cells. Nicotine induced cell proliferation in association with p53 down-regulation and cyclin D1 up-regulation. In differentiated cells, nicotine reduced alkaline phosphatase activity and mineralized nodule formation in dose-dependent manners. Furthermore, p53 expression was sustained in nicotine-treated cells during differentiation. These findings indicate that nicotine promotes the cell cycle and inhibits differentiation in association with p53 regulation in pre-osteoblasticmore » cells.« less
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Gutsch, Romina; Kandemir, Judith D; Pietsch, Daniel; Cappello, Christian; Meyer, Johann; Simanowski, Kathrin; Huber, René; Brand, Korbinian
2011-07-01
Monocytic differentiation is orchestrated by complex networks that are not fully understood. This study further elucidates the involvement of transcription factor CCAAT/enhancer-binding protein β (C/EBPβ). Initially, we demonstrated a marked increase in nuclear C/EBPβ-liver-enriched activating protein* (LAP*)/liver-enriched activating protein (LAP) levels and LAP/liver-enriched inhibiting protein (LIP) ratios in phorbol 12-myristate 13-acetate (PMA)-treated differentiating THP-1 premonocytic cells accompanied by reduced proliferation. To directly study C/EBPβ effects on monocytic cells, we generated novel THP-1-derived (low endogenous C/EBPβ) cell lines stably overexpressing C/EBPβ isoforms. Most importantly, cells predominantly overexpressing LAP* (C/EBPβ-long), but not those overexpressing LIP (C/EBPβ-short), exhibited a reduced proliferation, with no effect on morphology. PMA-induced inhibition of proliferation was attenuated in C/EBPβ-short cells. In C/EBPβ(WT) macrophage-like cells (high endogenous C/EBPβ), we measured a reduced proliferation/cycling index compared with C/EBPβ(KO). The typical macrophage morphology was only observed in C/EBPβ(WT), whereas C/EBPβ(KO) stayed round. C/EBPα did not compensate for C/EBPβ effects on proliferation/morphology. Serum reduction, an independent approach known to inhibit proliferation, induced macrophage morphology in C/EBPβ(KO) macrophage-like cells but not THP-1. In PMA-treated THP-1 and C/EBPβ-long cells, a reduced phosphorylation of cell cycle repressor retinoblastoma was found. In addition, C/EBPβ-long cells showed reduced c-Myc expression accompanied by increased CDK inhibitor p27 and reduced cyclin D1 levels. Finally, C/EBPβ-long and C/EBPβ(WT) cells exhibited low E2F1 and cyclin E levels, and C/EBPβ overexpression was found to inhibit cyclin E1 promoter-dependent transcription. Our results suggest that C/EBPβ reduces monocytic proliferation by affecting the retinoblastoma/E2F/cyclin E pathway and that it may contribute to, but is not directly required for, macrophage morphology. Inhibition of proliferation by C/EBPβ may be important for coordinated monocytic differentiation.
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Moon, Mi-Young; Kim, Hyun Jung; Choi, Bo Young; Sohn, Min; Chung, Tae Nyoung; Suh, Sang Won
2018-01-01
Zinc is an essential element required for cell division, migration, and proliferation. Under zinc-deficient conditions, proliferation and differentiation of neural progenitors are significantly impaired. Adipose-derived mesenchymal stem cells (AD-MSCs) are multipotent stem cells that can differentiate into neurons. The aim of this study was to evaluate the effect of zinc on AD-MSC proliferation and differentiation. We initially examined the effect of zinc on stem cell proliferation at the undifferentiated stage. AD-MSCs showed high proliferation rates on day 6 in 30 μ M and 100 μ M of ZnCl 2 . Zinc chelation inhibited AD-MSC proliferation via downregulation of ERK1/2 activity. We then assessed whether zinc was involved in cell migration and neurite outgrowth during differentiation. After three days of neuronal differentiation, TUJ-1-positive cells were observed, implying that AD-MSCs had differentiated into early neuron or neuron-like cells. Neurite outgrowth was increased in the zinc-treated group, while the CaEDTA-treated group showed diminished, shrunken neurites. Furthermore, we showed that zinc promoted neurite outgrowth via the inactivation of RhoA and led to the induction of neuronal gene expression (MAP2 and nestin) in differentiated stem cells. Taken together, zinc promoted AD-MSC proliferation and affected neuronal differentiation, mainly by increasing neurite outgrowth.
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JEDINAK, ANDREJ; SLIVA, DANIEL
2009-01-01
In spite of the global consumption of mushrooms, only two epidemiological studies demonstrated an inverse correlation between mushroom intake and the risk of cancer. Therefore, in the present study we evaluated whether extracts from edible mushrooms Agaricus bisporus (portabella), Flammulina velutipes (enoki), Lentinula edodes (shiitake) and Pleurotus ostreatus (oyster) affect the growth of breast and colon cancer cells. Here, we identified as the most potent, P. ostreatus (oyster mushroom) which suppressed proliferation of breast cancer (MCF-7, MDA-MB-231) and colon cancer (HT-29, HCT-116) cells, without affecting proliferation of epithelial mammary MCF-10A and normal colon FHC cells. Flow cytometry revealed that the inhibition of cell proliferation by P. ostreatus was associated with the cell cycle arrest at G0/G1 phase in MCF-7 and HT-29 cells. Moreover, P. ostreatus induced the expression of the tumor suppressor p53 and cyclin-dependent kinase inhibitor p21(CIP1/WAF1), whereas inhibited the phosphorylation of retinoblastoma Rb protein in MCF-7 cells. In addition, P. ostreatus also up-regulated expression of p21 and inhibited Rb phosphorylation in HT-29 cells, suggesting that that P. ostreatus suppresses the proliferation of breast and colon cancer cells via p53-dependent as well as p53-independent pathway. In conclusion, our results indicated that the edible oyster mushroom has potential therapeutic/preventive effects on breast and colon cancer. PMID:19020765
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Gaber, Timo; Tran, Cam Loan; Schellmann, Saskia; Hahne, Martin; Strehl, Cindy; Hoff, Paula; Radbruch, Andreas; Burmester, Gerd-Rüdiger; Buttgereit, Frank
2013-06-01
Inflamed areas are characterized by infiltration of immune cells, local hypoxia and alterations of cellular redox states. We investigated the impact of hypoxia on survival, proliferation, cytokine secretion, intracellular energy and redox state of human CD4(+) T cells. We found that pathophysiological hypoxia (<2% O2 ) significantly decreased CD4(+) T-cell survival after mitogenic stimulation. This effect was not due to an increased caspase-3/7-mediated apoptosis or adenosine-5'-triphosphate (ATP) consumption/depletion. However, the ability of stimulated T cells to proliferate was reduced under hypoxic conditions, despite increased expression of CD25. Pathophysiological hypoxia was also found to modify intracellular ROS (iROS) levels in stimulated T cells over time as compared with levels found in normoxia. Physiological hypoxia (5% O2 ) did not decrease CD4(+) T-cell survival and proliferation or modify iROS levels as compared with normoxia. We conclude that pathophysiological hypoxia affects T-cell proliferation and viability via disturbed IL-2R signalling downstream of STAT5a phosphorylation, but not as a result of impaired cellular energy homeostasis. We suggest iROS links early events in T-cell stimulation to the inhibition of the lymphoproliferative response under pathophysiological hypoxic conditions. The level of iROS may therefore act as a mediator of immune functions leading to down-regulation of long-term T-cell activity in inflamed tissues. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Effect of Physical Forces on the Metastatic Bone Microenvironment
2013-10-01
G.R., et al., Cell proliferation of cultured human cancer cells are affected by the elevated tumor pressures that exist in vivo. Ann Biomed Eng, 2005... cell lines. In vitro experiments have shown that increased pressure leads to decreased PCa proliferation. Osteoblasts also have inhibited...applied to tumor cells . Novel candidate genes with altered expression due to pressure have been identified and are currently undergoing further
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Homeobox A7 stimulates breast cancer cell proliferation by up-regulating estrogen receptor-alpha
DOE Office of Scientific and Technical Information (OSTI.GOV)
Zhang, Yu; Department of Obstetrics and Gynaecology, Child and Family Research Institute, University of British Columbia, Vancouver, British Columbia V5Z 4H4; Cheng, Jung-Chien
2013-11-01
Highlights: •HOXA7 regulates MCF7 cell proliferation. •HOXA7 up-regulates ERα expression. •HOXA7 mediates estrogen-induced MCF7 cell proliferation. -- Abstract: Breast cancer is the most common hormone-dependent malignancy in women. Homeobox (HOX) transcription factors regulate many cellular functions, including cell migration, proliferation and differentiation. The aberrant expression of HOX genes has been reported to be associated with human reproductive cancers. Estradiol (E2) and its nuclear receptors, estrogen receptor (ER)-alpha and ER-beta, are known to play critical roles in the regulation of breast cancer cell growth. However, an understanding of the potential relationship between HOXA7 and ER in breast cancer cells is limited.more » In this study, our results demonstrate that knockdown of HOXA7 in MCF7 cells significantly decreased cell proliferation and ERα expression. In addition, HOXA7 knockdown attenuated E2-induced cell proliferation as well as progesterone receptor (PR) expression. The stimulatory effects of E2 on cell proliferation and PR expression were abolished by co-treatment with ICI 182780, a selective ERα antagonist. In contrast, overexpression of HOXA7 significantly stimulated cell proliferation and ERα expression. Moreover, E2-induced cell proliferation, as well as PR expression, was enhanced by the overexpression of HOXA7. Neither knockdown nor overexpression of HOXA7 affected the ER-beta levels. Our results demonstrate a novel mechanistic role for HOXA7 in modulating breast cancer cell proliferation via regulation of ERα expression. This finding contributes to our understanding of the role HOXA7 plays in regulating the proliferation of ER-positive cancer cells.« less
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Tetraspanin CD9 modulates human lymphoma cellular proliferation via histone deacetylase activity
DOE Office of Scientific and Technical Information (OSTI.GOV)
Herr, Michael J.; Department of Medicine, The University of Tennessee Health Science Center, Memphis, TN 38163; Department of Molecular Sciences, The University of Tennessee Health Science Center, Memphis, TN 38163
2014-05-16
Highlights: • CD9 is differentially expressed in human Burkitt’s lymphoma cells. • We found that CD9 expression promotes these cells proliferation. • CD9 expression also increases HDAC activity. • HDAC inhibition decreased both cell proliferation and importantly CD9 expression. • CD9 may dictate HDAC efficacy and play a role in HDAC regulation. - Abstract: Non-Hodgkin Lymphoma (NHL) is a type of hematological malignancy that affects two percent of the overall population in the United States. Tetraspanin CD9 is a cell surface protein that has been thoroughly demonstrated to be a molecular facilitator of cellular phenotype. CD9 expression varies in twomore » human lymphoma cell lines, Raji and BJAB. In this report, we investigated the functional relationship between CD9 and cell proliferation regulated by histone deacetylase (HDAC) activity in these two cell lines. Introduction of CD9 expression in Raji cells resulted in significantly increased cell proliferation and HDAC activity compared to Mock transfected Raji cells. The increase in CD9–Raji cell proliferation was significantly inhibited by HDAC inhibitor (HDACi) treatment. Pretreatment of BJAB cells with HDAC inhibitors resulted in a significant decrease in endogenous CD9 mRNA and cell surface expression. BJAB cells also displayed decreased cell proliferation after HDACi treatment. These results suggest a significant relationship between CD9 expression and cell proliferation in human lymphoma cells that may be modulated by HDAC activity.« less
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Stress Increases the Negative Effects of Chronic Pain on Hippocampal Neurogenesis.
Romero-Grimaldi, Carmen; Berrocoso, Esther; Alba-Delgado, Cristina; Madrigal, Jose Luis M; Perez-Nievas, Beatriz G; Leza, Juan Carlos; Mico, Juan Antonio
2015-10-01
Patients with chronic pain often suffer from affective disorders and cognitive decline, which significantly impairs their quality of life. In addition, many of these patients also experience stress unrelated to their illness, which can aggravate their symptoms. These nociceptive inputs are received by the hippocampus, in which maladaptive neuroplastic changes may occur in the conditions of chronic pain. The hippocampus is a structure involved in emotionality, learning, and memory, and the proliferating cells in the granular layer of the hippocampal dentate gyrus respond to chronic pain by slowing their turnover. However, whether the maturation, survival, and integration of newborn cells in the hippocampus are affected by chronic pain remains unclear. In addition, it is unknown whether an added stress may increase this effect. We have evaluated the proliferation, differentiation, and survival of newborn hippocampal cells in a rat model of neuropathic pain (chronic constriction injury), with or without stress (chronic immobilization), by assessing the incorporation of bromodeoxyuridine into proliferating cells and immunostaining. The data obtained indicated that there was a decrease in the number of proliferating cells 8 days after nerve injury in animals subjected to neuropathic pain, an effect that was exacerbated by stress. Moreover, 4 weeks after nerve injury, neuropathic pain was associated with a loss of neuroblasts and the reduced survival of new mature neurons in the hippocampal granular layer, phenomena that also were increased by stress. By contrast, the rate of differentiation was not affected in this paradigm. Neuropathic pain negatively influences hippocampal neurogenesis (proliferation and survival), and this effect is exacerbated by stress. These neuroplastic changes may account for the affective and cognitive impairment seen in patients with chronic pain.
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Liao, Shan; Xiao, Songshu; Chen, Hongxiang; Zhang, Manying; Chen, Zhifang; Long, Yuehua; Gao, Lu; Zhu, Guangchao; He, Junyu; Peng, Shuping; Xiong, Wei; Zeng, Zhaoyang; Li, Zheng; Zhou, Ming; Li, Xiaoling; Ma, Jian; Wu, Minghua; Xiang, Juanjuan; Li, Guiyuan; Zhou, Yanhong
2017-10-01
Cervical cancer is one of the most common malignant tumors in women all over the world. The exact mechanism of occurrence and development of cervical cancer has not been fully elucidated. CD38 is a type II transmembrane glycoprotein, which was found to mediate diverse activities, including signal transduction, cell adhesion, and cyclic ADP-ribose synthesis. Here, we reported that CD38 promoted cell proliferation and inhibited cell apoptosis in cervical cancer cells by affecting the mitochondria functions. We established stable cervical cancer cell lines with CD38 over-expressed. CCK8 assay and colony formation assay indicated that CD38 promoted cervical cancer cell proliferation. Nude mouse tumorigenicity assay showed that CD38 significantly promotes tumor growth in vivo. CD38 also induced S phase accumulation in cell cycle analysis and suppressed cell apoptosis in cervical cancer cells. Meanwhile, flow cytometry analysis of mitochondria functions suggested that CD38 decreased intracellular Ca 2+ levels in cervical cancer cells and CD38 was involved in down-regulation of ROS levels and prevented mitochondrial apoptosis in cervical cancer cells. The percentage of cells with loss of mitochondrial membrane potential (Δψm) in CD38-overexpressed cervical cancer cells was less than control groups. Furthermore, we found an up-regulation of MDM2, cyclinA1, CDK4, cyclinD1, NF-kB P65, c-rel, and a downregulation of P53, P21, and P38 by Western blot analysis. These results indicated that CD38 enhanced the proliferation and inhibited the apoptosis of cervical cancer cells by affecting the mitochondria functions. © 2017 Wiley Periodicals, Inc.
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Recino, Asha; Barkan, Kerry; Wong, F Susan; Ladds, Graham; Cooke, Anne; Wallberg, Maja
2017-08-31
Metabolism is of central importance for T cell survival and differentiation. It is well known that T cells cannot function in the absence of glucose, but it is less clear how they respond to excessive levels of glucose. In the present study, we investigated how increasing levels of glucose affect T-cell-mediated immune responses. We examined the effects of increased levels of glucose on CD8 + T-cell behaviour in vitro by assessing activation and cytokine production, as well as oxygen consumption rate (OCR), extracellular acidification rate (ECAR) and intracellular signalling. In addition, we assessed in vivo proliferation, cytokine production and cytolytic activity of cells in chemically induced diabetic C57BL/6 mice. Elevated levels of glucose in in vitro cultures had modest effects on proliferation and cytokine production, while in vivo hyperglycaemia had no effect on CD8 + T-cell proliferation, interferon γ (IFNγ) production or cytolytic killing. © 2017 The Author(s).
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T-kininogen induces endothelial cell proliferation.
Pérez, Viviana; Leiva-Salcedo, Elías; Acuña-Castillo, Claudio; Aravena, Mauricio; Gómez, Christian; Sabaj, Valeria; Colombo, Alicia; Nishimura, Sumiyo; Pérez, Claudio; Walter, Robin; Sierra, Felipe
2006-03-01
Basal proliferation of endothelial cells increases with age, and this might play a role in the etiology of age-related vascular diseases, as well as angiogenesis. Serum kininogen levels increase during aging in rats and humans, and T-kininogen (T-KG) can affect proliferative homeostasis in several cell models. Both kinins and kininogens have been shown previously to be angiogenic through activation of endothelial cell proliferation, and here we show that exposure of endothelial cells to T-KG results in vigorous cell proliferation, accompanied by ERK/AKT activation. In our experiments, the proliferative response requires B1 and B2 kinin receptors, even though kinins are not released from the precursor. We hypothesize that the age-related increase in T-KG could play a significant role in the age-related dysregulation of vascular physiology and function.
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Role of Dicer1 in thyroid cell proliferation and differentiation.
Penha, Ricardo Cortez Cardoso; Sepe, Romina; De Martino, Marco; Esposito, Francesco; Pellecchia, Simona; Raia, Maddalena; Del Vecchio, Luigi; Decaussin-Petrucci, Myriam; De Vita, Gabriella; Pinto, Luis Felipe Ribeiro; Fusco, Alfredo
2017-01-01
DICER1 plays a central role in the biogenesis of microRNAs and it is important for normal development. Altered microRNA expression and DICER1 dysregulation have been described in several types of tumors, including thyroid carcinomas. Recently, our group identified a new somatic mutation (c.5438A>G; E1813G) within DICER1 gene of an unknown function. Herein, we show that DICER1 is overexpressed, at mRNA level, in a significant-relative number of papillary (70%) and anaplastic (42%) thyroid carcinoma samples, whereas is drastically downregulated in all the analyzed human thyroid carcinoma cell lines (TPC-1, BCPAP, FRO and 8505c) in comparison with normal thyroid tissue samples. Conversely, DICER1 is downregulated, at protein level, in PTC in comparison with normal thyroid tissues. Our data also reveals that DICER1 overexpression positively regulates thyroid cell proliferation, whereas its silencing impairs thyroid cell differentiation. The expression of DICER1 gene mutation (c.5438A>G; E1813G) negatively affects the microRNA machinery and cell proliferation as well as upregulates DICER1 protein levels of thyroid cells but has no impact on thyroid differentiation. In conclusion, DICER1 protein is downregulated in papillary thyroid carcinomas and affects thyroid proliferation and differentiation, while DICER1 gene mutation (c.5438A>G; E1813G) compromises the DICER1 wild-type-mediated microRNA processing and cell proliferation.
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Buckner, Carly A; Buckner, Alison L; Koren, Stan A; Persinger, Michael A; Lafrenie, Robert M
2018-04-01
Exposure to specific electromagnetic field (EMF) patterns can affect a variety of biological systems. We have shown that exposure to Thomas-EMF, a low-intensity, frequency-modulated (25-6 Hz) EMF pattern, inhibited growth and altered cell signaling in malignant cells. Exposure to Thomas-EMF for 1 h/day inhibited the growth of malignant cells including B16-BL6 mouse melanoma cells, MDA-MB-231, MDA-MB-468, BT-20, and MCF-7 human breast cancer and HeLa cervical cancer cells but did not affect non-malignant cells. The Thomas-EMF-dependent changes in cell proliferation were mediated by adenosine 3',5'-cyclic monophosphate (cAMP) and extracellular-signal-regulated kinase (ERK) signaling pathways. Exposure of malignant cells to Thomas-EMF transiently changed the level of cellular cAMP and promoted ERK phosphorylation. Pharmacologic inhibitors (SQ22536) and activators (forskolin) of cAMP production both blocked the ability of Thomas-EMF to inhibit cell proliferation, and an inhibitor of the MAP kinase pathway (PD98059) was able to partially block Thomas-EMF-dependent inhibition of cell proliferation. Genetic modulation of protein kinase A (PKA) in B16-BL6 cells also altered the effect of Thomas-EMF on cell proliferation. Cells transfected with the constitutively active form of PKA (PKA-CA), which interfered with ERK phosphorylation, also interfered with the Thomas-EMF effect on cell proliferation. The non-malignant cells did not show any EMF-dependent changes in cAMP levels, ERK phosphorylation, or cell growth. These data indicate that exposure to the specific Thomas-EMF pattern can inhibit the growth of malignant cells in a manner dependent on contributions from the cAMP and MAP kinase pathways. Bioelectromagnetics. 39;217-230, 2018. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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Phenolic Compounds in Extra Virgin Olive Oil Stimulate Human Osteoblastic Cell Proliferation
García-Martínez, Olga; De Luna-Bertos, Elvira; Ramos-Torrecillas, Javier; Ruiz, Concepción; Milia, Egle; Lorenzo, María Luisa; Jimenez, Brigida; Sánchez-Ortiz, Araceli; Rivas, Ana
2016-01-01
In this study, we aimed to clarify the effects of phenolic compounds and extracts from different extra virgin olive oil (EVOO) varieties obtained from fruits of different ripening stages on osteoblast cells (MG-63) proliferation. Cell proliferation was increased by hydroxytyrosol, luteolin, apigenin, p-coumaric, caffeic, and ferulic acids by approximately 11–16%, as compared with controls that were treated with one vehicle alone, while (+)-pinoresinol, oleuropein, sinapic, vanillic acid and derivative (vanillin) did not affect cell proliferation. All phenolic extracts stimulated MG-63 cell growth, and they induced higher cell proliferation rates than individual compounds. The most effective EVOO phenolic extracts were those obtained from the Picual variety, as they significantly increased cell proliferation by 18–22%. Conversely, Arbequina phenolic extracts increased cell proliferation by 9–13%. A decline in osteoblast proliferation was observed in oils obtained from olive fruits collected at the end of the harvest period, as their total phenolic content decreases at this late stage. Further research on the signaling pathways of olive oil phenolic compounds involved in the processes and their metabolism should be carried out to develop new interventions and adjuvant therapies using EVOO for bone health (i.e.osteoporosis) in adulthood and the elderly. PMID:26930190
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Karam, Manale; Legay, Christine; Auclair, Christian
2012-03-10
Protein kinase D1, PKD1, is a novel serine/threonine kinase whose altered expression and dysregulation in many tumors as well as its activation by several mitogens suggest that this protein could regulate proliferation and tumorigenesis. Nevertheless, the precise signaling pathways used are still unclear and the potential direct role of PKD1 in tumor development and progression has not been yet investigated. In order to clarify the role of PKD1 in cell proliferation and tumorigenesis, we studied the effects of PKD1 overexpression in a human adenocarcinoma breast cancer cell line, MCF-7 cells. We demonstrated that overexpression of PKD1 specifically promotes MCF-7 cellmore » proliferation through accelerating G0/G1 to S phase transition of the cell cycle. Moreover, inhibition of endogenous PKD1 significantly reduced cell proliferation. Taken together, these results clearly strengthen the regulatory role of PKD1 in cell growth. We also demonstrated that overexpression of PKD1 specifically diminished serum- and anchorage-dependence for proliferation and survival in vitro and allowed MCF-7 cells to form tumors in vivo. Thus, all these data highlight the central role of PKD1 in biological processes which are hallmarks of malignant transformation. Analysis of two major signaling pathways implicated in MCF-7 cell proliferation showed that PKD1 overexpression significantly increased ERK1/2 phosphorylation state without affecting Akt phosphorylation. Moreover, PKD1 overexpression-stimulated cell proliferation and anchorage-independent growth were totally impaired by inhibition of the MEK/ERK kinase cascade. However, neither of these effects was affected by blocking the PI 3-kinase/Akt signaling pathway. Thus, the MEK/ERK signaling appears to be a determining pathway mediating the biological effects of PKD1 in MCF-7 cells. Taken together, all these data demonstrate that PKD1 overexpression increases the aggressiveness of MCF-7 breast cancer cells through enhancing their oncogenic properties and would, therefore, define PKD1 as a potentially new promising anti-tumor therapeutic target.« less
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Endogenous Hydrogen Sulfide Enhances Cell Proliferation of Human Gastric Cancer AGS Cells.
Sekiguchi, Fumiko; Sekimoto, Teruki; Ogura, Ayaka; Kawabata, Atsufumi
2016-01-01
Hydrogen sulfide (H2S), the third gasotransmitter, is endogenously generated by certain H2S synthesizing enzymes, including cystathionine-γ-lyase (CSE) and cystathionine-β-synthase (CBS) from L-cysteine in the mammalian body. Several studies have shown that endogenous and exogenous H2S affects the proliferation of cancer cells, although the effects of H2S appear to vary with cell type, being either promotive or suppressive. In the present study, we determined whether endogenously formed H2S regulates proliferation in human gastric cancer AGS cells. CSE, but not CBS, was expressed in AGS cells. CSE inhibitors, DL-propargylglycine (PPG) and β-cyano-L-alanine (BCA), significantly suppressed the proliferation of AGS cells in a concentration-dependent manner. CSE inhibitors did not increase lactate dehydrogenase (LDH) release in the same concentration range. The inhibitory effects of PPG and BCA on cell proliferation were reversed by repetitive application of NaHS, a donor of H2S. Interestingly, nuclear condensation and fragmentation were detected in AGS cells treated with PPG or BCA. These results suggest that endogenous H2S produced by CSE may contribute to the proliferation of gastric cancer AGS cells, most probably through anti-apoptotic actions.
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Miyake, Asako; Takeda, Takashi; Isobe, Aki; Wakabayashi, Atsuko; Nishimoto, Fumihito; Morishige, Ken-Ichirou; Sakata, Masahiro; Kimura, Tadashi
2009-06-01
Uterine leiomyomas are the most common gynecological benign tumor and greatly affect reproductive health and well-being. They are the predominant indication for hysterectomy in premenopausal women. Current epidemiological study reported that soy products intake is inversely associated with diseases leading to hysterectomy. Genistein is a soy-derived phytoestrogen and its inhibitory effect on leiomyoma cell proliferation is reported. In this study, we investigated the siginificant inhibitory effect of genistein on estradiol (E(2))-induced leiomyoma cells proliferation. The Eker rat-derived uterine leiomyoma cell line ELT-3 cells were used. Cell proliferation was assessed by counting the number of cells. The expression of estrogen receptors and peroxisome proliferator-activated receptor-gamma (PPARgamma) was evaluated by Western blot analysis. PPARgamma was expressed in ELT-3 cells and genistein acted as PPARgamma ligand. This inhibitory effect of genistein was attenuated by the treatment of cells with PPARgamma antagonist bisphenol A diglycidyl ether (BADGE) or GW9662. These experimental findings in vitro show that the repressive effect of genistein on E(2)-induced ELT-3 cell proliferation is through the activation of PPARgamma. Genistein may be useful as an alternative therapy for leiomyoma.
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Naito, Tomoaki; Mulet, Céline; De Castro, Cristina; Molinaro, Antonio; Saffarian, Azadeh; Nigro, Giulia; Bérard, Marion; Clerc, Mélanie; Pedersen, Amy B; Sansonetti, Philippe J; Pédron, Thierry
2017-10-17
We identified a crypt-specific core microbiota (CSCM) dominated by strictly aerobic, nonfermentative bacteria in murine cecal and proximal colonic (PC) crypts and hypothesized that, among its possible functions, it may affect epithelial regeneration. In the present work, we isolated representative CSCM strains using selective media based upon our initial 16S rRNA-based molecular identification (i.e., Acinetobacter , Delftia , and Stenotrophomonas ). Their tropism for the crypt was confirmed, and their influence on epithelial regeneration was demonstrated in vivo by monocolonization of germfree mice. We also showed that lipopolysaccharide (LPS), through its endotoxin activity, was the dominant bacterial agonist controlling proliferation. The relevant molecular mechanisms were analyzed using colonic crypt-derived organoids exposed to bacterial sonicates or highly purified LPS as agonists. We identified a Toll-like receptor 4 (TLR4)-dependent program affecting crypts at different stages of epithelial differentiation. LPS played a dual role: it repressed cell proliferation through RIPK3-mediated necroptosis of stem cells and cells of the transit-amplifying compartment and concurrently enhanced cell differentiation, particularly the goblet cell lineage. IMPORTANCE The LPS from crypt-specific core microbiota controls intestinal epithelium proliferation through necroptosis of stem cells and enhances cell differentiation, mainly the goblet cell lineage. Copyright © 2017 Naito et al.
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Wang, Yueqi; Cao, Wenfeng; Yu, Zaicheng; Liu, Zhihua
2009-09-01
Results from tissue microarray in this study and our previous reports revealed that stomatin-like protein 2 (SLP-2) is notably associated with tumorigenesis and metastasis. Many members of stomatin family are involved in tumor as mitochondrial component, and recent study has revealed that SLP-2 may also function in mitochondria. To further investigate the function of SLP-2, we used siRNA target SLP-2. Data showed that knock-down of SLP-2 potently inhibited cell motility, proliferation and slightly altered cell cycle without any significant change of apoptosis. Moreover, by combined application with different chemotherapeutic reagents, we observed the enhancement of cell chemosensitivity by SLP-2 depletion. We also confirmed that, SLP-2 localizes in mitochondria, affects mitochondrial membrane potential (MMP) and ATP production. We conclude that, SLP-2 is a mitochondrial protein and therefore, functions in energy process by MMP maintenance, and subsequently affecting cell motility, proliferation and chemosensitivity.
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Cell proliferation is a key determinant of the outcome of FOXO3a activation
DOE Office of Scientific and Technical Information (OSTI.GOV)
Poulsen, Raewyn C., E-mail: raewyn.poulsen@gmail.com; Carr, Andrew J.; Hulley, Philippa A.
2015-06-19
The FOXO family of forkhead transcription factors have a pivotal role in determining cell fate in response to oxidative stress. FOXO activity can either promote cell survival or induce cell death. Increased FOXO-mediated cell death has been implicated in the pathogenesis of degenerative diseases affecting musculoskeletal tissues. The aim of this study was to determine the conditions under which one member of the FOXO family, FOXO3a, promotes cell survival as opposed to cell death. Treatment of primary human tenocytes with 1 pM hydrogen peroxide for 18 h resulted in increased protein levels of FOXO3a. In peroxide-treated cells cultured in low serum media,more » FOXO3a inhibited cell proliferation and protected against apoptosis. However in peroxide treated cells cultured in high serum media, cell proliferation was unchanged but level of apoptosis significantly increased. Similarly, in tenocytes transduced to over-express FOXO3a, cell proliferation was inhibited and level of apoptosis unchanged in cells cultured in low serum. However there was a robust increase in cell death in FOXO3a-expressing cells cultured in high serum. Inhibition of cell proliferation in either peroxide-treated or FOXO3a-expressing cells cultured in high serum protected against apoptosis induction. Conversely, addition of a Chk2 inhibitor to peroxide-treated or FOXO3a-expressing cells overrode the inhibitory effect of FOXO3a on cell proliferation and led to increased apoptosis in cells cultured in low serum. This study demonstrates that proliferating cells may be particularly susceptible to the apoptosis-inducing actions of FOXO3a. Inhibition of cell proliferation by FOXO3a may be a critical event in allowing the pro-survival rather than the pro-apoptotic activity of FOXO3a to prevail. - Highlights: • FOXO3a activity can result in either promotion of cell survival or apoptosis. • The outcome of FOXO3a activation differs in proliferating compared to non-proliferating cells. • Proliferating cells are susceptible to FOXO3a-mediated apoptosis. • Inhibition of cell proliferation by FOXO3a promotes cell survival.« less
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Zinc oxide nanoparticles as selective killers of proliferating cells
Taccola, Liuba; Raffa, Vittoria; Riggio, Cristina; Vittorio, Orazio; Iorio, Maria Carla; Vanacore, Renato; Pietrabissa, Andrea; Cuschieri, Alfred
2011-01-01
Background: It has recently been demonstrated that zinc oxide nanoparticles (ZnO NPs) induce death of cancerous cells whilst having no cytotoxic effect on normal cells. However, there are several issues which need to be resolved before translation of zinc oxide nanoparticles into medical use, including lack of suitable biocompatible dispersion protocols and a better understanding being needed of the mechanism of their selective cytotoxic action. Methods: Nanoparticle dose affecting cell viability was evaluated in a model of proliferating cells both experimentally and mathematically. The key issue of selective toxicity of ZnO NPs toward proliferating cells was addressed by experiments using a biological model of noncancerous cells, ie, mesenchymal stem cells before and after cell differentiation to the osteogenic lineage. Results: In this paper, we report a biocompatible protocol for preparation of stable aqueous solutions of monodispersed zinc oxide nanoparticles. We found that the threshold of intracellular ZnO NP concentration required to induce cell death in proliferating cells is 0.4 ± 0.02 mM. Finally, flow cytometry analysis revealed that the threshold dose of zinc oxide nanoparticles was lethal to proliferating pluripotent mesenchymal stem cells but exhibited negligible cytotoxic effects to osteogenically differentiated mesenchymal stem cells. Conclusion: Results confirm the ZnO NP selective cytotoxic action on rapidly proliferating cells, whether benign or malignant. PMID:21698081
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Zinc oxide nanoparticles as selective killers of proliferating cells.
Taccola, Liuba; Raffa, Vittoria; Riggio, Cristina; Vittorio, Orazio; Iorio, Maria Carla; Vanacore, Renato; Pietrabissa, Andrea; Cuschieri, Alfred
2011-01-01
It has recently been demonstrated that zinc oxide nanoparticles (ZnO NPs) induce death of cancerous cells whilst having no cytotoxic effect on normal cells. However, there are several issues which need to be resolved before translation of zinc oxide nanoparticles into medical use, including lack of suitable biocompatible dispersion protocols and a better understanding being needed of the mechanism of their selective cytotoxic action. Nanoparticle dose affecting cell viability was evaluated in a model of proliferating cells both experimentally and mathematically. The key issue of selective toxicity of ZnO NPs toward proliferating cells was addressed by experiments using a biological model of noncancerous cells, ie, mesenchymal stem cells before and after cell differentiation to the osteogenic lineage. In this paper, we report a biocompatible protocol for preparation of stable aqueous solutions of monodispersed zinc oxide nanoparticles. We found that the threshold of intracellular ZnO NP concentration required to induce cell death in proliferating cells is 0.4 ± 0.02 mM. Finally, flow cytometry analysis revealed that the threshold dose of zinc oxide nanoparticles was lethal to proliferating pluripotent mesenchymal stem cells but exhibited negligible cytotoxic effects to osteogenically differentiated mesenchymal stem cells. Results confirm the ZnO NP selective cytotoxic action on rapidly proliferating cells, whether benign or malignant.
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Bidad, Katayoon; Salehi, Eisa; Oraei, Mona; Saboor-Yaraghi, Ali-Akbar; Nicknam, Mohammad Hossein
2011-12-01
All-trans retinoic acid (ATRA), as an active metabolite of vitamin A, has been shown to affect immune cells. This study was performed to evaluate the effect of ATRA on viability, proliferation, activation and lineage-specific transcription factors of CD4+ T cells. CD4+ T cells were separated from heparinized blood of healthy donors and were cultured in conditions, some with, some without ATRA. Viability was assessed by PI flowcytometry and proliferation was measured by MTT assay. CD69 expression was determined by flowcytometry as a measure of cell activation. Lineage-specific transcription factors (FOXP3, RORγt and T-bet) were examined by intracellular staining and flowcytometry. High doses of ATRA (0.1-1 mM) caused extensive cell death in both PBMCs and CD4+ T cells. Doses of ATRA equal to or lower than 10 µM did not adversely affect cell viability and proliferation in comparison to culture medium without ATRA. Doses of ATRA between 10 µM and 1nM significantly increased cell activation when compared to culture medium without ATRA. ATRA could increase FOXP3+ and also FOXP3+RORγt+ T cells while it decreased RORγt+ and T-bet+ T cells. This study showed that doses of ATRA up to 10 µM are safe when using with CD4+ T cells in terms of cell viability, proliferation and activation. We could also show that ATRA diverts the human immune response in neutral conditions (without adding polarizing cytokines) by increasing FOXP3+ cells and decreasing RORγt+ cells. ATRA could be regarded as a potential therapy in inflammatory conditions and autoimmunities.
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Ultra-fast laser microprocessing of medical polymers for cell engineering applications.
Ortiz, R; Moreno-Flores, S; Quintana, I; Vivanco, MdM; Sarasua, J R; Toca-Herrera, J L
2014-04-01
Picosecond laser micromachining technology (PLM) has been employed as a tool for the fabrication of 3D structured substrates. These substrates have been used as supports in the in vitro study of the effect of substrate topography on cell behavior. Different micropatterns were PLM-generated on polystyrene (PS) and poly-L-lactide (PLLA) and employed to study cellular proliferation and morphology of breast cancer cells. The laser-induced microstructures included parallel lines of comparable width to that of a single cell (which in this case is roughly 20μm), and the fabrication of square-like compartments of a much larger area than a single cell (250,000μm(2)). The results obtained from this in vitro study showed that though the laser treatment altered substrate roughness, it did not noticeably affect the adhesion and proliferation of the breast cancer cells. However, pattern direction directly affected cell proliferation, leading to a guided growth of cell clusters along the pattern direction. When cultured in square-like compartments, cells remained confined inside these for eleven incubation days. According to these results, laser micromachining with ultra-short laser pulses is a suitable method to directly modify the cell microenvironment in order to induce a predefined cellular behavior and to study the effect of the physical microenvironment on cell proliferation. Copyright © 2013 Elsevier B.V. All rights reserved.
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E-cadherin interactions regulate beta-cell proliferation in islet-like structures.
Carvell, Melanie J; Marsh, Phil J; Persaud, Shanta J; Jones, Peter M
2007-01-01
Islet function is dependent on cells within the islet interacting with each other. E-cadherin (ECAD) mediates Ca(2+)-dependent homophilic cell adhesion between b-cells within islets and has been identified as a tumour suppressor. We generated clones of the MIN6 beta-cell line that stably over- (S) and under-express (alphaS) ECAD. Modified expression of ECAD was confirmed by quantitative RT-PCR, immunoblotting and immunocytochemistry. Preproinsulin mRNA, insulin content and basal rates of insulin secretion were higher in S cells compared to aS and control (V) cells. However, stimulated insulin secretory responses were unaffected by ECAD expression levels. ECAD expression did affect proliferation, with enhanced ECAD expression being associated with reduced proliferation and vice versa. Formation of islet-like structures was associated with a significant reduction in proliferation of V and S cells but not alphaS cells. These data suggest that ECAD expression levels do not modulate insulin secretory function but are consistent with a role for ECAD in the regulation of beta-cell proliferation. Copyright (c) 2007 S. Karger AG, Basel.
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Moon, Mi-Young; Kim, Hyun Jung; Choi, Bo Young; Sohn, Min
2018-01-01
Zinc is an essential element required for cell division, migration, and proliferation. Under zinc-deficient conditions, proliferation and differentiation of neural progenitors are significantly impaired. Adipose-derived mesenchymal stem cells (AD-MSCs) are multipotent stem cells that can differentiate into neurons. The aim of this study was to evaluate the effect of zinc on AD-MSC proliferation and differentiation. We initially examined the effect of zinc on stem cell proliferation at the undifferentiated stage. AD-MSCs showed high proliferation rates on day 6 in 30 μM and 100 μM of ZnCl2. Zinc chelation inhibited AD-MSC proliferation via downregulation of ERK1/2 activity. We then assessed whether zinc was involved in cell migration and neurite outgrowth during differentiation. After three days of neuronal differentiation, TUJ-1-positive cells were observed, implying that AD-MSCs had differentiated into early neuron or neuron-like cells. Neurite outgrowth was increased in the zinc-treated group, while the CaEDTA-treated group showed diminished, shrunken neurites. Furthermore, we showed that zinc promoted neurite outgrowth via the inactivation of RhoA and led to the induction of neuronal gene expression (MAP2 and nestin) in differentiated stem cells. Taken together, zinc promoted AD-MSC proliferation and affected neuronal differentiation, mainly by increasing neurite outgrowth. PMID:29765417
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Liu, Y; Wang, S N; Cui, C Y; Dong, Y M
2017-04-18
Positive effects of bioactive glass (BG) on proliferation, mineralization, and differentiation of human dental pulp cells (hDPCs) was already verified in various former studies. The Arg-Gly-Asp-Ser sequence (RGDS) was confirmed of affecting cell adhesion. Before further investigation, the objective of this study is to investigate whether RGDS can affect the effects of BG on the adhesion, proliferation and mineralization of hDPCs. hDPCs were harvested from third molars of 18-25-year-old individuals after informed consent. Enzyme digestion technique was used. The 4th to 6th generation of hDPCs were used for all experiments. The cells of the experimental groups were cultured in Dulbecco minimum essential medium (DMEM) containing ionic dissolution products of BG and RGDS of several concentrations (12.5 mg/L, 25.0 mg/L, 50.0 mg/L, 100.0 mg/L, 200.0 mg/L). DMEM containing ionic dissolution products of BG without RGDS was used for cell culture as control group. Cell adhesion was tested 4 h after cell seeding by MTT assay. Cell proliferation was examined at 1, 3, 5, 7, and 9 d after cell seeding by MTT assay. Cell mineralization was investigated on days 14 and 28 by alizarin red staining. After being stained and dried, mineralized nodules were dissolved by cetylpyridinium chloride (CPC) for semi-quantitative test. Results were statistically analyzed by one way ANOVA, SPSS (version 19.0) and P<0.05 was considered to be significant. Cell adhesion in BG group showed no difference from that in DMEM group. Compared with BG group, hDPCs in BG+RGDS groups suggested weaker cell adhesion.When the concentration of RGDS increased, the adhered cell number decreased. hDPCs cultured with BG and RGDS showed lower proliferation activity in the early stage, while no significant difference was observed after 3 d. BG group promoted the mineralization of hDPCs compared with positive control group, negative control group and RGDS group. No significant difference was observed between BG+RGDS group and BG group or between RGDS group and positive control group. BG promotes proliferation and mineralization without affecting cell adhesion of hDPCs. Unbounded RGDS inhibits cell adhesion, but has no influence on the positive effects of BG on the proliferation and mineralization of hDPCs.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Sun, Lei; Department of Physiology, Nankai University School of Medicine, Tianjin 300071; Carr, Aprell L.
2014-07-11
Highlights: • Stil is a human oncogene that is conserved in vertebrate species. • Stil functions in the Shh pathway in mammalian cells. • The expression of Stil is required for mammalian dopaminergic cell proliferation. - Abstract: The human oncogene SCL/TAL1 interrupting locus (Stil) is highly conserved in all vertebrate species. In humans, the expression of Stil is involved in cancer cell survival, apoptosis and proliferation. In this research, we investigated the roles of Stil expression in cell proliferation of mammalian dopaminergic (DA) PC12 cells. Stil functions through the Sonic hedgehog (Shh) signal transduction pathway. Co-immunoprecipitation tests revealed that STILmore » interacts with Shh downstream components, which include SUFU and GLI1. By examining the expression of Stil, Gli1, CyclinD2 (cell-cycle marker) and PCNA (proliferating cell nuclear antigen), we found that up-regulation of Stil expression (transfection with overexpression plasmids) increased Shh signaling transduction and PC12 cell proliferation, whereas down-regulation of Stil expression (by shRNA) inhibited Shh signaling transduction, and thereby decreased PC12 cell proliferation. Transient transfection of PC12 cells with Stil knockdown or overexpression plasmids did not affect PC12 cell neural differentiation, further indicating the specific roles of Stil in cell proliferation. The results from this research suggest that Stil may serve as a bio-marker for neurological diseases involved in DA neurons, such as Parkinson’s disease.« less
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Kwon, Joseph; Oh, Kyung Seo; Cho, Se-Young; Bang, Mi Ae; Kim, Hwan Seon; Vaidya, Bipin; Kim, Duwoon
2016-11-01
Hyperforin, a major active compound of St. John's wort extract, affects estrogenic activity. In this study, the compound evoked estrogen response element-dependent luciferase activity and cell proliferation in MCF-7 cells. Hyperforin-induced cell proliferation was significantly inhibited by the estrogen receptor antagonist ICI 182,780. These results suggested that hyperforin had estrogenic and cell proliferation activities, which were stimulated via the estrogen receptor. Compared to 17 β -estradiol, hyperforin showed significantly lower estrogenic activity and cell proliferation. The mechanism underlying the estrogenic activity of hyperforin was unknown, therefore, in this study, for the first time, the expression and post-translational modification of proteins were determined and compared among control, 17 β -estradiol-treated, and hyperforin-treated cells using proteomic techniques. A total of 453 proteins were identified, of which 282 proteins were significantly modulated in hyperforin-treated cells compared to 17 β -estradiol-treated cells. Ingenuity pathway analysis also demonstrated that hyperforin treatment induced less cell proliferation than 17 β -estradiol by downregulating estrogen receptor 1. Protein network analysis showed that cell proliferation was regulated mainly by cyclin D1 and extracellular signal-regulated kinases. In conclusion, although, hyperforin exhibited lower estrogenic activity than 17 β -estradiol, the compound induced lower levels of cancer cell proliferation in vitro . Georg Thieme Verlag KG Stuttgart · New York.
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Piura, Benjamin; Medina, Liat; Rabinovich, Alex; Dyomin, Victor; Huleihel, Mahmoud
2013-01-01
Thalidomide inhibits TNF-α production in lipopolysaccharide-stimulated monocytes. The aim of this study was to evaluate the effect of thalidomide on TNF-α, IL-6 and MMP secretion in epithelial ovarian carcinoma cells. SKOV-3 cells and primary epithelial ovarian carcinoma cells were cultured in the presence of various concentrations of thalidomide. Cell proliferation was examined by MTT proliferation assay. TNF-α and IL-6 levels were determined in the supernatants of the cell cultures by ELISA, and MMP activity was examined by gelatin zymography. Thalidomide did not significantly affect the proliferation and growth of SKOV-3 cells. However, it decreased significantly the capacity of SKOV-3 cells and primary epithelial ovarian carcinoma cells to secrete TNF-α. Thalidomide also significantly decreased the capacity of SKOV-3 cells, but not primary epithelial ovarian carcinoma cells, to secrete MMP-9 and MMP-2. However, thalidomide did not affect IL-6 secretion in SKOV-3 cells or primary epithelial ovarian carcinoma cells. Our study suggests that thalidomide distinctly affected TNF-α, IL-6 and MMPs secretion by an ovarian carcinoma cell line (SKOV-3) and primary ovarian cancer cells. This might suggest a different susceptibility of these two types of cells to thalidomide, and/or that the mechanisms of secretion of the factors examined are differently regulated in these cells. Our results may deepen our understanding the mechanism/s of action of thalidomide in ovarian carcinoma cells. The results might have important implications in future therapeutic strategies that will incorporate thalidomide and other cytokine inhibitors in the treatment of epithelial ovarian carcinoma.
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Harthan, Laura B; McFarland, Douglas C; Velleman, Sandra G
2014-01-01
Posthatch satellite cell mitotic activity is a critical component of muscle development and growth. Satellite cells are myogenic stem cells that can be induced by nutrition to follow other cellular developmental pathways, and whose mitotic activity declines with age. The objective of the current study was to determine the effect of restricting protein synthesis on the proliferation and differentiation, expression of myogenic transcriptional regulatory factors myogenic determination factor 1, myogenin, and myogenic regulatory factor 4, and expression of the heparan sulfate proteoglycans syndecan-4 and glypican-1 in satellite cells isolated from 1-d-, 7-wk-, and 16-wk-old turkey pectoralis major muscle (1 d, 7 wk, and 16 wk cells, respectively) by using variable concentrations of Met and Cys. Four Met concentrations-30 (control), 7.5, 3, or 0 mg/L with 3.2 mg/L of Cys per 1 mg/L of Met-were used for culture of satellite cells to determine the effect of nutrition and age on satellite cell behavior during proliferation and differentiation. Proliferation was reduced by lower Met and Cys concentrations in all ages at 96 h of proliferation. Differentiation was increased in the 1 d Met-restricted cells, whereas the 7 wk cells treated with 3 mg/L of Met had decreased differentiation. Reduced Met and Cys levels from the control did not significantly affect the 16 wk cells at 72 h of differentiation. However, medium with no Met or Cys suppressed differentiation at all ages. The expression of myogenic determination factor 1, myogenin, myogenic regulatory factor 4, syndecan-4, and glypican-1 was differentially affected by age and Met or Cys treatment. These data demonstrate the age-specific manner in which turkey pectoralis major muscle satellite cells respond to nutritional availability and the importance of defining optimal nutrition to maximize satellite cell proliferation and differentiation for subsequent muscle mass accretion.
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Effect of different wound dressings on cell viability and proliferation.
Paddle-Ledinek, Joanne E; Nasa, Zeyad; Cleland, Heather J
2006-06-01
Many new dressings have been developed since the early 1980s. Wound healing comprises cleansing, granulation/vascularization, and epithelialization phases. An optimum microenvironment and the absence of cytotoxic factors are essential for epithelialization. This study examines the effect of extracts of different wound dressings on keratinocyte survival and proliferation. Keratinocyte cultures were exposed for 40 hours to at least three extracts of each of the following wound dressings, which were tested in octuplicate: Acticoat, Aquacel-Ag, Aquacel, Algisite M, Avance, Comfeel Plus transparent, Contreet-H, Hydrasorb, and SeaSorb. Silicone extract provided the reference material. Controls were included of cells cultured in medium that had been incubated under conditions identical to those used with the extracts. Cell survival (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide reduction) and proliferation (5-bromo-2':-deoxyuridine incorporation) were measured. Extracts of silver-containing dressings (Acticoat, Aquacel-Ag, Contreet-H, and Avance) were most cytotoxic. Extracts of Hydrasorb were less cytotoxic but markedly affected keratinocyte proliferation and morphology. Extracts of alginate-containing dressings (Algisite M, SeaSorb, and Contreet-H) demonstrated high calcium concentrations, markedly reduced keratinocyte proliferation, and affected keratinocyte morphology. Extracts of Aquacel and Comfeel Plus transparent induced small but significant inhibition of keratinocyte proliferation. The principle of minimizing harm should be applied to the choice of wound dressing. Silver-based dressings are cytotoxic and should not be used in the absence of infection. Alginate dressings with high calcium content affect keratinocyte proliferation probably by triggering terminal differentiation of keratinocytes. Such dressings should be used with caution in cases in which keratinocyte proliferation is essential. All dressings should be tested in vitro before clinical application.
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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
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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.
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Primary cilia maintain corneal epithelial homeostasis by regulation of the Notch signaling pathway
Grisanti, Laura; Revenkova, Ekaterina; Gordon, Ronald E.
2016-01-01
Primary cilia have been linked to signaling pathways involved in cell proliferation, cell motility and cell polarity. Defects in ciliary function result in developmental abnormalities and multiple ciliopathies. Patients affected by severe ciliopathies, such as Meckel syndrome, present several ocular surface disease conditions of unclear pathogenesis. Here, we show that primary cilia are predominantly present on basal cells of the mouse corneal epithelium (CE) throughout development and in the adult. Conditional ablation of cilia in the CE leads to an increase in proliferation and vertical migration of basal corneal epithelial cells (CECs). A consequent increase in cell density of suprabasal layers results in a thicker than normal CE. Surprisingly, in cilia-deficient CE, cilia-mediated signaling pathways, including Hh and Wnt pathways, were not affected but the intensity of Notch signaling was severely diminished. Although Notch1 and Notch2 receptors were expressed normally, nuclear Notch1 intracellular domain (N1ICD) expression was severely reduced. Postnatal development analysis revealed that in cilia-deficient CECs downregulation of the Notch pathway precedes cell proliferation defects. Thus, we have uncovered a function of the primary cilium in maintaining homeostasis of the CE by balancing proliferation and vertical migration of basal CECs through modulation of Notch signaling. PMID:27122169
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Mainou-Fowler, T; Copplestone, J A; Prentice, A G
1995-01-01
AIMS--To investigate the effects of interleukin (IL) 1, 2, 4, and 5 on the proliferation and survival of peripheral blood B cells from patients with B chronic lymphocytic leukaemia (B-CLL) and compare them with the effects on normal peripheral blood B cells. METHODS--The proliferation and survival of pokeweed mitogen (PWM) activated B cells from B-CLL (n = 12) and normal peripheral blood (n = 5) were studied in vitro in response to IL-1, IL-2 IL-4, and IL-5. Survival of cells in cultures with or without added interleukins was studied by microscopic examination of cells and DNA agarose gel electrophoresis. RESULTS--Proliferation was observed in both B-CLL and normal peripheral blood cells on culture with IL-2 alone and also in some, but not all, B-CLL and normal peripheral blood cells with IL-1 and IL-4. However, there was greater variability in B-CLL cell responses than in normal peripheral blood cells. Il-5 did not affect normal peripheral blood cell proliferation but it increased proliferation in two B-CLL cases. Synergistic effects of these cytokines were not detected. IL-4 inhibited normal peripheral blood and B-CLL cell proliferation after the addition of IL-2. Inhibition of B-CLL cell responses to IL-2 was also observed with IL-5 and Il-1. Survival of B-CLL cells in cultures was enhanced with IL-4 not by an increase in proliferation but by reduced apoptosis. No such effect was seen in normal peripheral blood cells. IL-2 had a less noticeable antiapoptotic effect; IL-5 enhanced apoptosis in B-CLL cells. CONCLUSIONS--B-CLL and normal peripheral blood cells proliferated equally well in response to IL-2. IL-4 had a much lower effect on B-CLL cell proliferation, but had noticeable antiapoptotic activity. IL-5 enhanced cell death by apoptosis. Images PMID:7629299
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Kimbrough-Allah, Mawiyah N; Millena, Ana C; Khan, Shafiq A
2018-04-01
Transforming growth factor-β (TGF-β) acts as a tumor suppressor in normal epithelial cells but as a tumor promoter in advanced prostate cancer cells. PI3-kinase pathway mediates TGF-β effects on prostate cancer cell migration and invasion. PTEN inhibits PI3-kinase pathway and is frequently mutated in prostate cancers. We investigated possible role(s) of PTEN in TGF-β effects on proliferation and migration in prostate cancer cells. Expression of PTEN mRNA and proteins were determined using RT-PCR and Western blotting in RWPE1 and DU145 cells. We also studied the role of PTEN in TGF-β effects on cell proliferation and migration in DU145 cells after transient silencing of endogenous PTEN. Conversely, we determined the role of PTEN in cell proliferation and migration after over-expression of PTEN in PC3 cells which lack endogenous PTEN. TGF-β1 and TGF-β3 had no effect on PTEN mRNA levels but both isoforms increased PTEN protein levels in DU145 and RWPE1 cells indicating that PTEN may mediate TGF-β effects on cell proliferation. Knockdown of PTEN in DU145 cells resulted in significant increase in cell proliferation which was not affected by TGF-β isoforms. PTEN overexpression in PC3 cells inhibited cell proliferation. Knockdown of endogenous PTEN enhanced cell migration in DU145 cells, whereas PTEN overexpression reduced migration in PC3 cells and reduced phosphorylation of AKT in response to TGF-β. We conclude that PTEN plays a role in inhibitory effects of TGF-β on cell proliferation whereas its absence may enhance TGF-β effects on activation of PI3-kinase pathway and cell migration. © 2018 Wiley Periodicals, Inc.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Hou, Jianghong, E-mail: jianghonghou@163.com; Xue, Xiaolin; Li, Junnong
2016-01-22
Recently, the serum expression level of vasostatin-2 was found to be reduced and is being studied as an important indicator to assess the presence and severity of coronary artery disease; the functional properties of vasostatin-2 and its relationship with the development of atherosclerosis remains unclear. In this study, we attempted to detect the expression of vasostatin-2 and its impact on human vascular smooth muscle cells (VSMCs). Quantitative real-time PCR (qRT-PCR) and western blot were used to assess the expression level of vasostatin-2 in VSMCs between those from atherosclerosis and disease-free donors; we found that vasostatin-2 was significantly down-regulated in atherosclerosismore » patient tissues and cell lines. In addition, the over-expression of vasostatin-2 apparently inhibits cell proliferation and migration in VSMCs. Gain-of-function in vitro experiments further show that vasostatin-2 over-expression significantly inhibits inflammatory cytokines release in VSMCs. In addition, cell adhesion experimental analysis showed that soluble adhesion molecules (sICAM-1, sVCAM-1) had decreased expression when vasostatin-2 was over-expressed in VSMCs. Therefore, our results indicate that vasostatin-2 is an atherosclerosis-related factor that can inhibit cell proliferation, inflammatory response and cell adhesion in VSMCs. Taken together, our results indicate that vasostatin-2 could serve as a potential diagnostic biomarker and therapeutic option for human atherosclerosis in the near future. - Highlights: • Vasostatin-2 levels were down-regulated in atherosclerosis patient tissues and VSMCs. • Ectopic expression of vasostatin-2 directly affects cell proliferation and migration in vitro. • Ectopic expression of vasostatin-2 protein affects pro-inflammatory cytokines release in VSMCs. • Ectopic expression of vasostatin-2 protein affects cell adhesion in VSMCs.« less
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Crisp, Sarah E R H; Griffin, Jacob B; White, Brett R; Toombs, Candice F; Camporeale, Gabriela; Said, Hamid M; Zempleni, Janos
2004-02-01
Placental transfer of nutrients and secretion of hormones is essential for normal fetal development. To determine whether biotin supply affects biotin homeostasis, proliferation rates, and progesterone secretion in placenta cells. JAr choriocarcinoma cells were cultured in media containing deficient (25 pmol/L), physiological (250 pmol/L), or pharmacological concentrations (10,000 pmol/L) of biotin for three weeks; markers for biotin homeostasis, proliferation, and hormone secretion were quantified. Biotin concentrations in culture media correlated negatively with expression of the biotin transporter SMVT, as judged by cellular transport rates of biotin, abundance of SMVT protein, and transcriptional activity of SMVT reporter-gene constructs. Notwithstanding this homeostatic mechanism, biotin concentrations in media correlated positively with activities of biotin-dependent propionyl-CoA carboxylase, abundance of biotinylated carboxylases, and with biotinylation of histones. Biotin deficiency was associated with decreased rates of thymidine uptake into JAr cells [pmol thymidine/( 10(6) cells x 24 h)]: 1.6 +/- 0.1 (25 pmol/L biotin) versus 2.3 +/- 0.2 (250 pmol/L biotin) versus 3.7 +/- 0.4 (10,000 pmol/L biotin), suggesting that cell proliferation depends on biotin. Secretion of progesterone was reduced in biotin-deficient cells; this effect was caused by reduced generation of new cells in deficient media rather than by an immediate effect of biotin on progesterone secretion at the singlecell-level. This study provides evidence that choriocarcinoma cells cannot maintain normal activities of biotin-dependent metabolic pathways if biotin concentrations in culture media are low. It is uncertain whether activities of biotin-dependent pathways in placenta affect fetal development in vivo.
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The G1/S Specific Cyclin D2 Is a Regulator of HIV-1 Restriction in Non-proliferating Cells
Badia, Roger; Pujantell, Maria; Riveira-Muñoz, Eva; Puig, Teresa; Torres-Torronteras, Javier; Martí, Ramón; Clotet, Bonaventura; Ampudia, Rosa M.; Ballana, Ester
2016-01-01
Macrophages are a heterogeneous cell population strongly influenced by differentiation stimuli that become susceptible to HIV-1 infection after inactivation of the restriction factor SAMHD1 by cyclin-dependent kinases (CDK). Here, we have used primary human monocyte-derived macrophages differentiated through different stimuli to evaluate macrophage heterogeneity on cell activation and proliferation and susceptibility to HIV-1 infection. Stimulation of monocytes with GM-CSF induces a non-proliferating macrophage population highly restrictive to HIV-1 infection, characterized by the upregulation of the G1/S-specific cyclin D2, known to control early steps of cell cycle progression. Knockdown of cyclin D2, enhances HIV-1 replication in GM-CSF macrophages through inactivation of SAMHD1 restriction factor by phosphorylation. Co-immunoprecipitation experiments show that cyclin D2 forms a complex with CDK4 and p21, a factor known to restrict HIV-1 replication by affecting the function of the downstream cascade that leads to SAMHD1 deactivation. Thus, we demonstrate that cyclin D2 acts as regulator of cell cycle proteins affecting SAMHD1-mediated HIV-1 restriction in non-proliferating macrophages. PMID:27541004
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Mangiferin positively regulates osteoblast differentiation and suppresses osteoclast differentiation
Sekiguchi, Yuusuke; Mano, Hiroshi; Nakatani, Sachie; Shimizu, Jun; Kataoka, Aya; Ogura, Kana; Kimira, Yoshifumi; Ebata, Midori; Wada, Masahiro
2017-01-01
Mangiferin is a polyphenolic compound present in Salacia reticulata. It has been reported to reduce bone destruction and inhibit osteoclastic differentiation. This study aimed to determine whether mangiferin directly affects osteoblast and osteoclast proliferation and differentiation, and gene expression in MC3T3-E1 osteoblastic cells and osteoclast-like cells derived from primary mouse bone marrow macrophage cells. Mangiferin induced significantly greater WST-1 activity, indicating increased cell proliferation. Mangiferin induced significantly increased alkaline phosphatase staining, indicating greater cell differentiation. Reverse transcription-polymerase chain reaction (RT-PCR) demonstrated that mangiferin significantly increased the mRNA level of runt-related transcription factor 2 (RunX2), but did not affect RunX1 mRNA expression. Mangiferin significantly reduced the formation of tartrate-resistant acid phosphatase-positive multinuclear cells. RT-PCR demonstrated that mangiferin significantly increased the mRNA level of estrogen receptor β (ERβ), but did not affect the expression of other osteoclast-associated genes. Mangiferin may inhibit osteoclastic bone resorption by suppressing differentiation of osteoclasts and promoting expression of ERβ mRNA in mouse bone marrow macrophage cells. It also has potential to promote osteoblastic bone formation by promoting cell proliferation and inducing cell differentiation in preosteoblast MC3T3-E1 cells via RunX2. Mangiferin may therefore be useful in improving bone disease outcomes. PMID:28627701
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Sekiguchi, Yuusuke; Mano, Hiroshi; Nakatani, Sachie; Shimizu, Jun; Kataoka, Aya; Ogura, Kana; Kimira, Yoshifumi; Ebata, Midori; Wada, Masahiro
2017-08-01
Mangiferin is a polyphenolic compound present in Salacia reticulata. It has been reported to reduce bone destruction and inhibit osteoclastic differentiation. This study aimed to determine whether mangiferin directly affects osteoblast and osteoclast proliferation and differentiation, and gene expression in MC3T3‑E1 osteoblastic cells and osteoclast‑like cells derived from primary mouse bone marrow macrophage cells. Mangiferin induced significantly greater WST‑1 activity, indicating increased cell proliferation. Mangiferin induced significantly increased alkaline phosphatase staining, indicating greater cell differentiation. Reverse transcription‑polymerase chain reaction (RT‑PCR) demonstrated that mangiferin significantly increased the mRNA level of runt‑related transcription factor 2 (RunX2), but did not affect RunX1 mRNA expression. Mangiferin significantly reduced the formation of tartrate‑resistant acid phosphatase‑positive multinuclear cells. RT‑PCR demonstrated that mangiferin significantly increased the mRNA level of estrogen receptor β (ERβ), but did not affect the expression of other osteoclast‑associated genes. Mangiferin may inhibit osteoclastic bone resorption by suppressing differentiation of osteoclasts and promoting expression of ERβ mRNA in mouse bone marrow macrophage cells. It also has potential to promote osteoblastic bone formation by promoting cell proliferation and inducing cell differentiation in preosteoblast MC3T3‑E1 cells via RunX2. Mangiferin may therefore be useful in improving bone disease outcomes.
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Chen, Miaojiao; Zhang, Jingjing; Hu, Fang; Liu, Shiping; Zhou, Zhiguang
2015-11-01
Accumulating evidence suggests an association between diabetes and cancer. Inflammation is a key event that underlies the pathological processes of the two diseases. Metformin displays anti-cancer effects, but the mechanism is not completely clear. This study investigated whether metformin regulated the microenvironment of macrophage polarization to affect the characteristics of HepG2 cells and the possible role of the Notch-signalling pathway. RAW264.7 macrophages were cultured alone or co-cultured with HepG2 cells and treated with metformin. We analysed classical (M1) and alternative (M2) gene expression in RAW264.7 cells using quantitative real-time polymerase chain reaction. Changes in mRNA and protein expressions of Notch signalling in both cell types were also detected using quantitative real-time polymerase chain reaction and Western-blotting analyses. The proliferation, apoptosis and migration of HepG2 cells were detected using Cell Titer 96 AQueous One Solution Cell Proliferation Assay (MTS) (Promega Corporation, Fitchburg, WI, USA), Annexin V-FITC/PI (7SeaPharmTech, Shanghai, China) and the cell scratch assay, respectively. Metformin induced single-cultured RAW264.7 macrophages with an M2 phenotype but attenuated the M2 macrophage differentiation and inhibited monocyte chemoattractant protein-1 (MCP-1) secretion in a co-culture system. The co-cultured group of metformin pretreatment activated Notch signalling in macrophages but repressed it inHepG2 cells. Co-culture also promoted the proliferation and migration of HepG2 cells. However, along with the enhanced apoptosis, the proliferation and the migration of HepG2 cells were remarkably inhibited in another co-culture system with metformin pretreatment. Metformin can skew RAW264.7 macrophages toward different phenotypes according to changes in the microenvironment, which may affect the inflammatory conditions mediated by macrophages, induce apoptosis and inhibit the proliferation and migration of HepG2 cells. Notch signalling pathway is a potentially important mechanism in the regulation of metformin on macrophage polarization and the subsequent change of hepatoma cells. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.
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Hultsch, T; Martin, R; Hohman, R J
1992-01-01
The immunosuppressive drugs FK506 and cyclosporin A have an identical spectrum of activities with respect to IgE receptor (Fc epsilon RI)-mediated exocytosis from mast cells and T cell receptor-mediated transcription of IL-2. These findings suggest a common step in receptor-mediated signal transduction leading to exocytosis and transcription and imply that immunosuppressive drugs target specific signal transduction pathways, rather than specific cell types. This hypothesis is supported by studies on the effect of rapamycin on IL-3 dependent proliferation of the rodent mast cell line PT18. Rapamycin inhibits proliferation of PT18 cells, achieving a plateau of 80% inhibition at 1 nM. This inhibition is prevented in a competitive manner by FK506, a structural analogue of rapamycin. Proliferation of rat basophilic leukemia cells and WEHI-3 cells was also inhibited, at doses comparable to those shown previously to inhibit IL-2-dependent proliferation of cytotoxic T lymphocyte line (CTLL) cells. In contrast, proliferation of A-431 cells, a epidermoid cell line, was not affected by rapamycin. DNA histograms indicate that complexes formed between the rapamycin-FK506-binding protein (FKBP) and rapamycin arrest-proliferating PT18 cells in the G0/G1-phase. It is concluded that FKBP-rapamycin complexes may inhibit proliferative signals emanating from IL-3 receptors, resulting in growth arrest of cytokine-dependent, hematopoietic cells. PMID:1384815
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Li, Pei; Hu, Jing; Zhang, Ying; Li, Jianping; Dang, Yunzhi; Zhang, Rui; Wei, Lichun; Shi, Mei
2018-02-01
Objective To investigate the role and mechanism of microRNA-182 (miR-182) in the proliferation of cervical cancer cells. Methods With liposome-mediated transient transfection method, the level of miR-182 in HeLa and SiHa cells was increased or decreased. CCK-8 assay and colony formation assay were used to observe the effect of miR-182 on the proliferation of cervical cancer cells. Using bioinformatics predictions, real-time quantitative PCR, and dual luciferase reporter assay, we clarified the role of miR-182 in posttranscriptional regulation of adenomatous polyposis coli (APC) gene and its effect on the downstream molecules (c-Myc and cyclin D1) of Wnt singling pathway. Results Up-regulation of miR-182 significantly promoted the proliferation of cervical cancer cells, while down-regulation of miR-182 significantly inhibited the proliferation of cervical cancer cells. Over-expression of miR-182 inhibited the expression of APC gene in cervical cancer cells and the regulation of miR-182 affected the expression of canonical Wnt signaling pathway downstream molecules in cervical cancer cells. Conclusion The miR-182 stimulates canonical Wnt signaling pathway by targeting APC gene and enhances the proliferation of cervical cancer cells.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Fang, Dengfeng; Yang, Hui; Lin, Jing
2015-02-20
In bone, different concentration of estrogen leads to various of physiological processes in osteoblast, such as the proliferation, migration, and apoptosis in an estrogen receptor-dependent manner. But little was known about the estrogen effects on osteosarcoma (OS). In this study, OS cell MG-63 was treated with low (1 nM) or high (100 nM) dose of 17β-Estradiol (E2) with the presence or absence of estrogen receptor α (ERα), for evaluating the E2 effects on proliferation, migration, invasion, colony formation and apoptosis. Consistent with a previous study, high dose of E2 treatment dramatically downregulated expressing level of long non-coding RNA metastasis associated lung adenocarcinomamore » transcript 1 (MALAT-1). The observation of upregulation of miR-9 after a high dose of E2 treatment indicated the cause of MALAT-1 reduction. Downregulation of MALAT-1 promoted the combination of SFPQ/PTBP2 complex. It was also observed that the proliferation, migration, invasion, colony formation and apoptosis of OS cells were remarkably affected by high dose of E2 treatment, but not by low dose, in an ERα independent manner. Furthermore, the abolishment of the effects on these physiological processes caused by ectopic expression of miR-9 ASOs suggested the necessity of miR-9 in MALAT-1 regulation. Here we found that the high dose of E2 treatment upregulated miR-9 thus posttranscriptionally regulated MALAT-1 RNA level in OS cells, and then the downregulation of MALAT-1 inhibited cell proliferation, migration, invasion and epithelial–mesenchymal transition (EMT) processes in the E2-dose dependent and ER-independent ways. - Highlights: • E2 affects osteosarcoma cell MG-63 in an Estrogen receptor-independent way. • High dose of E2 treatment upregulates miR-9 which target to MALAT-1 RNA. • Upregulated miR-9 degrades MALAT-1 and thus affects combination of SFPQ/PTBP2. • E2 treatment block cell proliferation, colony formation, mobility, and enhance apoptosis.« less
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Rapamycin ameliorates IgA nephropathy via cell cycle-dependent mechanisms
Tian, Jihua; Wang, Yanhong; Liu, Xinyan; Zhou, Xiaoshuang
2014-01-01
IgA nephropathy is the most frequent type of glomerulonephritis worldwide. The role of cell cycle regulation in the pathogenesis of IgA nephropathy has been studied. The present study was designed to explore whether rapamycin ameliorates IgA nephropathy via cell cycle-dependent mechanisms. After establishing an IgA nephropathy model, rats were randomly divided into four groups. Coomassie Brilliant Blue was used to measure the 24-h urinary protein levels. Renal function was determined using an autoanalyzer. Proliferation was assayed via Proliferating Cell Nuclear Antigen (PCNA) immunohistochemistry. Rat mesangial cells were cultured and divided into the six groups. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) and flow cytometry were used to detect cell proliferation and the cell cycle phase. Western blotting was performed to determine cyclin E, cyclin-dependent kinase 2, p27Kip1, p70S6K/p-p70S6K, and extracellular signal-regulated kinase 1/2/p- extracellular signal-regulated kinase 1/2 protein expression. A low dose of the mammalian target of rapamycin (mTOR) inhibitor rapamycin prevented an additional increase in proteinuria, protected kidney function, and reduced IgA deposition in a model of IgA nephropathy. Rapamycin inhibited mesangial cell proliferation and arrested the cell cycle in the G1 phase. Rapamycin did not affect the expression of cyclin E and cyclin-dependent kinase 2. However, rapamycin upregulated p27Kip1 at least in part via AKT (also known as protein kinase B)/mTOR. In conclusion, rapamycin can affect cell cycle regulation to inhibit mesangial cell proliferation, thereby reduce IgA deposition, and slow the progression of IgAN. PMID:25349217
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Rapamycin ameliorates IgA nephropathy via cell cycle-dependent mechanisms.
Tian, Jihua; Wang, Yanhong; Liu, Xinyan; Zhou, Xiaoshuang; Li, Rongshan
2015-07-01
IgA nephropathy is the most frequent type of glomerulonephritis worldwide. The role of cell cycle regulation in the pathogenesis of IgA nephropathy has been studied. The present study was designed to explore whether rapamycin ameliorates IgA nephropathy via cell cycle-dependent mechanisms. After establishing an IgA nephropathy model, rats were randomly divided into four groups. Coomassie Brilliant Blue was used to measure the 24-h urinary protein levels. Renal function was determined using an autoanalyzer. Proliferation was assayed via Proliferating Cell Nuclear Antigen (PCNA) immunohistochemistry. Rat mesangial cells were cultured and divided into the six groups. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) and flow cytometry were used to detect cell proliferation and the cell cycle phase. Western blotting was performed to determine cyclin E, cyclin-dependent kinase 2, p27(Kip1), p70S6K/p-p70S6K, and extracellular signal-regulated kinase 1/2/p- extracellular signal-regulated kinase 1/2 protein expression. A low dose of the mammalian target of rapamycin (mTOR) inhibitor rapamycin prevented an additional increase in proteinuria, protected kidney function, and reduced IgA deposition in a model of IgA nephropathy. Rapamycin inhibited mesangial cell proliferation and arrested the cell cycle in the G1 phase. Rapamycin did not affect the expression of cyclin E and cyclin-dependent kinase 2. However, rapamycin upregulated p27(Kip1) at least in part via AKT (also known as protein kinase B)/mTOR. In conclusion, rapamycin can affect cell cycle regulation to inhibit mesangial cell proliferation, thereby reduce IgA deposition, and slow the progression of IgAN. © 2014 by the Society for Experimental Biology and Medicine.
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Luo, Xia; Yin, Ping; Coon V, John S; Cheng, You-Hong; Wiehle, Ronald D; Bulun, Serdar E
2010-05-15
To evaluate the effects of selective P receptor (PR) modulator CDB4124 on cell proliferation and apoptosis in cultured human uterine leiomyoma smooth muscle (LSM) cells and control myometrial smooth muscle (MSM) cells in matched uteri. Laboratory research. Academic medical center. Premenopausal women (n = 12) undergoing hysterectomy for leiomyoma-related symptoms. Treatment of primary LSM and MSM cells with CDB4124 (10(-8)-10(-6) M) or vehicle for 24, 48, or 72 hours. Western blot for protein expression of proliferating cell nuclear antigen, cleaved polyadenosine 5'-diphosphate-ribose polymerase, Bcl-2, and Krüppel-like transcription factor 11; 93-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide (MTT) assay to evaluate viable cell numbers; and real-time polymerase chain reaction (PCR) to quantify messenger RNA (mRNA) levels. Treatment with CDB4124 significantly decreased levels of the proliferation marker proliferating cell nuclear antigen, the number of viable LSM cells, and the antiapoptotic protein Bcl-2. On the other hand, treatment with CDB4124 increased levels of the apoptosis marker cleaved polyadenosine 5'-diphosphate-ribose polymerase and the tumor suppressor Krüppel-like transcription factor 11 in a dose- and time-dependent manner in LSM cells. In matched MSM cells, however, CDB4124 did not affect cell proliferation or apoptosis. CDB4124 selectively inhibits proliferation and induces apoptosis in LSM but not in MSM cells. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.
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Activities for leptin in bovine trophoblast cells.
Hughes, C K; Xie, M M; McCoski, S R; Ealy, A D
2017-01-01
Leptin is involved in various reproductive processes in humans and rodents, including placental development and function. The specific ways that leptin influences placental development and function in cattle are poorly understood. This work was completed to explore how leptin regulates hormone, cytokine and metalloprotease transcript abundance, and cell proliferation in cultured bovine trophoblast cells. In the first set of studies, cells were cultured in the presence of graded recombinant bovine leptin concentrations (0, 10, 50, 250 ng/mL) for 6 or 24 h. Transcript profiles were examined from extracted RNA. Leptin supplementation did not affect abundance of the maternal recognition of pregnancy factor, interferon-tau (IFNT), but leptin increased (P < 0.05) abundance of chorionic somatomammotropin hormone 2 (CSH2; ie, placental lactogen) at both 6 and 24 h at each concentration tested. At 24 h, the greatest CSH2 abundance (P < 0.05) was detected in cells supplemented with 50 ng/mL leptin. Transcript abundance of the remodeling factor, metalloprotease 2 (MMP2), was greater (P < 0.05) in leptin-treated cells at 24 h but not at 6 h. The 24 h MMP2 response was greatest (P < 0.05) at 250 ng/mL. Transcript abundance for MMP9 was not altered by leptin treatment. In a separate set of studies, cell proliferation assays were completed. Leptin supplementation did not affect bovine trophoblast cell line proliferation at any dose tested. In conclusion, leptin supplementation did not affect bovine trophoblast cell proliferation or IFNT expression, but leptin increases CSH2 and MMP2 transcript abundance. Both of these factors are involved with peri-implantation and postimplantation placental development and function, and this implicates leptin as a potential mediator of early placental development and function in cattle. Copyright © 2016 Elsevier Inc. All rights reserved.
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Buckner, Carly A; Buckner, Alison L; Koren, Stan A; Persinger, Michael A; Lafrenie, Robert M
2015-01-01
Electromagnetic field (EMF) exposures affect many biological systems. The reproducibility of these effects is related to the intensity, duration, frequency, and pattern of the EMF. We have shown that exposure to a specific time-varying EMF can inhibit the growth of malignant cells. Thomas-EMF is a low-intensity, frequency-modulated (25-6 Hz) EMF pattern. Daily, 1 h, exposures to Thomas-EMF inhibited the growth of malignant cell lines including B16-BL6, MDA-MB-231, MCF-7, and HeLa cells but did not affect the growth of non-malignant cells. Thomas-EMF also inhibited B16-BL6 cell proliferation in vivo. B16-BL6 cells implanted in syngeneic C57b mice and exposed daily to Thomas-EMF produced smaller tumours than in sham-treated controls. In vitro studies showed that exposure of malignant cells to Thomas-EMF for > 15 min promoted Ca(2+) influx which could be blocked by inhibitors of voltage-gated T-type Ca(2+) channels. Blocking Ca(2+) uptake also blocked Thomas-EMF-dependent inhibition of cell proliferation. Exposure to Thomas-EMF delayed cell cycle progression and altered cyclin expression consistent with the decrease in cell proliferation. Non-malignant cells did not show any EMF-dependent changes in Ca(2+) influx or cell growth. These data confirm that exposure to a specific EMF pattern can affect cellular processes and that exposure to Thomas-EMF may provide a potential anti-cancer therapy.
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Buckner, Carly A.; Buckner, Alison L.; Koren, Stan A.; Persinger, Michael A.; Lafrenie, Robert M.
2015-01-01
Electromagnetic field (EMF) exposures affect many biological systems. The reproducibility of these effects is related to the intensity, duration, frequency, and pattern of the EMF. We have shown that exposure to a specific time-varying EMF can inhibit the growth of malignant cells. Thomas-EMF is a low-intensity, frequency-modulated (25-6 Hz) EMF pattern. Daily, 1 h, exposures to Thomas-EMF inhibited the growth of malignant cell lines including B16-BL6, MDA-MB-231, MCF-7, and HeLa cells but did not affect the growth of non-malignant cells. Thomas-EMF also inhibited B16-BL6 cell proliferation in vivo. B16-BL6 cells implanted in syngeneic C57b mice and exposed daily to Thomas-EMF produced smaller tumours than in sham-treated controls. In vitro studies showed that exposure of malignant cells to Thomas-EMF for > 15 min promoted Ca2+ influx which could be blocked by inhibitors of voltage-gated T-type Ca2+ channels. Blocking Ca2+ uptake also blocked Thomas-EMF-dependent inhibition of cell proliferation. Exposure to Thomas-EMF delayed cell cycle progression and altered cyclin expression consistent with the decrease in cell proliferation. Non-malignant cells did not show any EMF-dependent changes in Ca2+ influx or cell growth. These data confirm that exposure to a specific EMF pattern can affect cellular processes and that exposure to Thomas-EMF may provide a potential anti-cancer therapy. PMID:25875081
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Effects of Extracellular pH on Dental Pulp Cells In Vitro.
Hirose, Yujiro; Yamaguchi, Masaya; Kawabata, Shigetada; Murakami, Masashi; Nakashima, Misako; Gotoh, Momokazu; Yamamoto, Tokunori
2016-05-01
The proliferation and migration of dental pulp stem cells (DPSCs), a population comprised of dental pulp cells (DPCs), are important processes for pulp tissue repair. Dental pulp is exposed to changes in extracellular pH under various conditions, such as acidosis and exposure to caries-associated bacteria or a pulp capping agent. The objective of this study was to investigate the effects of extracellular pH on DPC proliferation and migration in vitro. To evaluate the proliferation potency of DPCs in various extracellular pH conditions, 2 × 10(4) cells were seeded into 35-mm dishes. The following day, we changed to NaHCO3-free medium, which was adjusted to different extracellular pH levels. After 120 hours, DPCs cultured in media from a pH of 3.5 to 5.5 showed cell death, those cultured in conditions from a pH of 6.5 to 7.5 showed growth arrest or cell death, and those grown at a pH of 9.5 showed mild proliferation. The migratory activity of living DPCs was not affected by extracellular pH. For histologic analysis, human teeth possessing a small abscess in the coronal pulp chamber were sliced for histologic analysis. Proliferating cell nuclear antigen (PCNA) immunolocalization was used as an index of cell proliferation for the sections and cultured cells. Acidic extracellular pH conditions resulted in reduced numbers of PCNA-positive DPCs in the dishes. As for pulp tissue affected by a small abscess, a PCNA-negative pulp cell layer was observed in close proximity to the infectious lesion. Together, these results suggest that an acidic extracellular pH condition is associated with DPC growth arrest or cell death. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.
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Metformin selectively affects human glioblastoma tumor-initiating cell viability
Würth, Roberto; Pattarozzi, Alessandra; Gatti, Monica; Bajetto, Adirana; Corsaro, Alessandro; Parodi, Alessia; Sirito, Rodolfo; Massollo, Michela; Marini, Cecilia; Zona, Gianluigi; Fenoglio, Daniela; Sambuceti, Gianmario; Filaci, Gilberto; Daga, Antonio; Barbieri, Federica; Florio, Tullio
2013-01-01
Cancer stem cell theory postulates that a small population of tumor-initiating cells is responsible for the development, progression and recurrence of several malignancies, including glioblastoma. In this perspective, tumor-initiating cells represent the most relevant target to obtain effective cancer treatment. Metformin, a first-line drug for type II diabetes, was reported to possess anticancer properties affecting the survival of cancer stem cells in breast cancer models. We report that metformin treatment reduced the proliferation rate of tumor-initiating cell-enriched cultures isolated from four human glioblastomas. Metformin also impairs tumor-initiating cell spherogenesis, indicating a direct effect on self-renewal mechanisms. Interestingly, analyzing by FACS the antiproliferative effects of metformin on CD133-expressing subpopulation, a component of glioblastoma cancer stem cells, a higher reduction of proliferation was observed as compared with CD133-negative cells, suggesting a certain degree of cancer stem cell selectivity in its effects. In fact, glioblastoma cell differentiation strongly reduced sensitivity to metformin treatment. Metformin effects in tumor-initiating cell-enriched cultures were associated with a powerful inhibition of Akt-dependent cell survival pathway, while this pathway was not affected in differentiated cells. The specificity of metformin antiproliferative effects toward glioblastoma tumor-initiating cells was confirmed by the lack of significant inhibition of normal human stem cells (umbilical cord-derived mesenchymal stem cells) in vitro proliferation after metformin exposure. Altogether, these data clearly suggest that metformin exerts antiproliferative activity on glioblastoma cells, showing a higher specificity toward tumor-initiating cells, and that the inhibition of Akt pathway may represent a possible intracellular target of this effect. PMID:23255107
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Novak, Barbara; Vatzia, Eleni; Springler, Alexandra; Pierron, Alix; Reisinger, Nicole; Hessenberger, Sabine; Mayer, Elisabeth
2018-01-01
Deoxynivalenol (DON) is one of the most prevalent mycotoxins, contaminating cereals and cereal-derived products. Its derivative deepoxy-deoxynivalenol (DOM-1) is produced by certain bacteria, which either occur naturally or are supplemented in feed additive. DON-induced impairments in protein synthesis are particularly problematic for highly proliferating immune cells. This study provides the first comparison of the effects of DON and DOM-1 on the concanavalin A-induced proliferation of porcine, chicken, and bovine peripheral blood mononuclear cells (PBMCs). Therefore, isolated PBMCs were treated with DON (0.01–3.37 µM) and DOM-1 (1.39–357 µM) separately, and proliferation was measured using a bromodeoxyuridine (BrdU) assay. Although pigs are considered highly sensitive to DON, the present study revealed a substantially higher sensitivity of bovine (IC50 = 0.314 µM) PBMCs compared to chicken (IC50 = 0.691 µM) and porcine (IC50 = 0.693 µM) PBMCs. Analyses on the proliferation of bovine T-cell subsets showed that all major subsets, namely, CD4+, CD8β+, and γδ T cells, were affected to a similar extent. In contrast, DOM-1 did not affect bovine PBMCs, but reduced the proliferation of chicken and porcine PBMCs at the highest tested concentration (357 µM). Results confirm the necessity of feed additives containing DON-to-DOM-1-transforming bacteria and highlights species-specific differences in the DON sensitivity of immune cells. PMID:29641442
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Effect of arachidonic acid metabolites on thymocyte proliferation.
Delebassée, S; Gualde, N
1988-01-01
The influences of prostaglandin E2 (PGE2), 15-hydroxyeicosatetraenoic acid (15-HETE) and leukotrienes (LT) on the proliferative response of mature (PNA-) and immature (PNA+) mouse thymocytes was investigated. Both PNA+ and PNA- thymocytes proliferated when cultured with concanavalin A plus interleukin-2. PGE2 in concentrations of 10(-6) to 10(-9) M caused significant inhibition of proliferation of both PNA+ and PNA- thymocytes in these cultures. In contrast, the lipoxygenase products 15-HETE, LTB4, LTC4 and LTD4 caused marked increases in proliferation of PNA+ thymocytes while having no effect on PNA- cells. Therefore, the effect of leukotrienes on thymocyte proliferation depends upon the level of cell maturation and mainly affects immature PNA+ thymocytes.
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Controllable bioeffects of laser-generated intracellular microbubbles
NASA Astrophysics Data System (ADS)
Zohdy, Marwa Joy
Laser-induced optical breakdown (LIOB) is a nonlinear energy absorption process that can generate precise damage in biological tissues. With femtosecond laser pulses, disruption is highly localized with minimal thermal and mechanical effects to the surrounding region. Cavitation bubbles are produced as a result of LIOB, and these bubbles can be detected and monitored with high-frequency ultrasound. In this work, the controllable viability effects of LIOB bubbles in single cells were characterized. Using a high-frequency acoustic transducer synchronized with a 793 nm, 100 fs laser pulsed at 250 kHz, thermal effects in the vicinity of an LIOB event were directly assessed. Temperaturedependent pulse-echo displacements were calculated using phase-sensitive correlation tracking and fit to a finite-element heat transfer model to estimate thermal distribution. Results indicate a minimal temperature increase (<1 degree C) within 100 microns of a bubble created with multiple laser pulses, confirming that LIOB can be controlled to be thermally noninvasive in the bubble vicinity. Acoustically detectable microbubbles were generated in individual cells with femtosecond LIOB. By adjusting laser fluence, exposure time, and focal location, LIOB could be controlled to produce distinctly different cellular effects. Small (1-2 micron) bubbles with short lifetimes (10100 ms) could be generated in cells without affecting their viability; and, alternatively, large (510 micron) bubbles with long lifetimes (1-5 s) could be generated for selective cell killing without affecting immediately neighboring cells. Experiments were performed in Chinese hamster ovary (CHO) cells in vitro, and LIOB was detected with both optical and acoustic microscopy. A long-term proliferation assay was also performed using green-fluorescent MCA207 mouse sarcoma cells targeted for LIOB. This assay confirmed that nondestructive bubbles did not affect target cell proliferation over several generations, and that destructive bubbles could indeed eliminate target cells and prevent further proliferation with no effect on immediately neighboring cells. These studies help to outline future applications for site-activated, acoustically monitored intracellular microbubbles. Nondestructive bubbles can potentially be used for functional cell measurements without introducing exogenous agents or affecting subsequent cell proliferation, and destructive bubbles can be used for highly precise biologically-targeted cancer cell therapy with real-time acoustic validation.
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Nuclear orphan receptor TLX affects gene expression, proliferation and cell apoptosis in beta cells.
Shi, Xiaoli; Xiong, Xiaokan; Dai, Zhe; Deng, Haohua; Sun, Li; Hu, Xuemei; Zhou, Feng; Xu, Yancheng
Nuclear orphan receptor TLX is an essential regulator of the growth of neural stem cells. However, its exact function in pancreatic islet cells is still unknown. In the present study, gene expression profiling analysis revealed that overexpression of TLX in beta cell line MIN6 causes suppression of 176 genes and upregulation of 49 genes, including a cadre of cell cycle, cell proliferation and cell death control genes, such as Btg2, Ddit3 and Gadd45a. We next examined the effects of TLX overexpression on proliferation, apoptosis and insulin secretion in MIN6 cells. Proliferation analysis using EdU assay showed that overexpression of TLX increased percentage of EdU-positive cells. Cell cycle and apoptosis analysis revealed that overexpression of TLX in MIN6 cells resulted in higher percentage of cells exiting G1 into S-phase, and a 58.8% decrease of cell apoptosis induced by 0.5 mM palmitate. Moreover, TLX overexpression did not cause impairment of insulin secretion. Together, we conclude that TLX is among factors capable of controlling beta cell proliferation and survival, which may serve as a target for the development of novel therapies for diabetes. Copyright © 2015 Elsevier Inc. All rights reserved.
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Luo, Xia; Yin, Ping; Coon V., John S.; Cheng, You-Hong; Wiehle, Ronald D.; Bulun, Serdar E.
2009-01-01
Objective To evaluate the effects of selective progesterone receptor modulator CDB4124 on cell proliferation and apoptosis in cultured human uterine leiomyoma smooth muscle (LSM) cells and control myometrial smooth muscle (MSM) cells in matched uteri. Design Laboratory research. Setting Academic medical center. Patient(s) Premenopausal women (n=12) undergoing hysterectomy for leiomyoma-related symptoms. Intervention(s) Treatment of primary LSM and MSM cells with CDB4124 (10-8-10-6M) or vehicle for 24, 48 or 72 hours. Main Outcome Measure(s) Western blot for protein expression of proliferating cell nuclear antigen (PCNA), cleaved poly-adenosine 5’-diphosphate-ribose polymerase (PARP), Bcl-2 and Krüppel-like transcription factor 11 (KLF11); MTT assay to evaluate viable cell numbers; and real-time polymerase chain reaction to quantify mRNA levels. Result(s) Treatment with CDB4124 significantly decreased levels of the proliferation marker PCNA, the number of viable LSM cells, and the anti-apoptotic protein Bcl-2. On the other hand, treatment with CDB4124 increased levels of the apoptosis marker cleaved PARP and the tumor suppressor KLF11 in a dose- and time-dependent manner in LSM cells. In matched MSM cells, however, CDB4124 did not affect cell proliferation or apoptosis. Conclusion(s) CDB4124 selectively inhibits proliferation and induces apoptosis in LSM but not in MSM cells. PMID:20056218
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Nuclear orphan receptor TLX affects gene expression, proliferation and cell apoptosis in beta cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Shi, Xiaoli; Xiong, Xiaokan; Dai, Zhe
2015-12-04
Nuclear orphan receptor TLX is an essential regulator of the growth of neural stem cells. However, its exact function in pancreatic islet cells is still unknown. In the present study, gene expression profiling analysis revealed that overexpression of TLX in beta cell line MIN6 causes suppression of 176 genes and upregulation of 49 genes, including a cadre of cell cycle, cell proliferation and cell death control genes, such as Btg2, Ddit3 and Gadd45a. We next examined the effects of TLX overexpression on proliferation, apoptosis and insulin secretion in MIN6 cells. Proliferation analysis using EdU assay showed that overexpression of TLXmore » increased percentage of EdU-positive cells. Cell cycle and apoptosis analysis revealed that overexpression of TLX in MIN6 cells resulted in higher percentage of cells exiting G1 into S-phase, and a 58.8% decrease of cell apoptosis induced by 0.5 mM palmitate. Moreover, TLX overexpression did not cause impairment of insulin secretion. Together, we conclude that TLX is among factors capable of controlling beta cell proliferation and survival, which may serve as a target for the development of novel therapies for diabetes. - Highlights: • TLX overexpression in MIN6 cell causes significant expression changes of 225 genes. • TLX overexpression promotes MIN6 cell proliferation and decreases cell apoptosis. • TLX overexpression does not cause impairment of insulin secretion.« less
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Ribeiro, Mariana P.C.; Laboratory of Biochemistry, Faculty of Pharmacy, University of Coimbra, 3000-548 Coimbra; Nunes-Correia, Isabel
Recent reports suggest that N-methyl-D-aspartate receptor (NMDAR) blockade by MK-801 decreases tumor growth. Thus, we investigated whether other ionotropic glutamate receptor (iGluR) antagonists were also able to modulate the proliferation of melanoma cells. On the other hand, the antiestrogen tamoxifen (TAM) decreases the proliferation of melanoma cells, and is included in combined therapies for melanoma. As the efficacy of TAM is limited by its metabolism, we investigated the effects of the NMDAR antagonist MK-801 in combination with TAM and its active metabolites, 4-hydroxytamoxifen (OHTAM) and endoxifen (EDX). The NMDAR blockers MK-801 and memantine decreased mouse melanoma K1735-M2 cell proliferation. Inmore » contrast, the NMDAR competitive antagonist APV and the AMPA and kainate receptor antagonist NBQX did not affect cell proliferation, suggesting that among the iGluR antagonists only the NMDAR channel blockers inhibit melanoma cell proliferation. The combination of antiestrogens with MK-801 potentiated their individual effects on cell biomass due to diminished cell proliferation, since it decreased the cell number and DNA synthesis without increasing cell death. Importantly, TAM metabolites combined with MK-801 promoted cell cycle arrest in G1. Therefore, the data obtained suggest that the activity of MK-801 and antiestrogens in K1735-M2 cells is greatly enhanced when used in combination. - Highlights: • MK-801 and memantine decrease melanoma cell proliferation. • The combination of MK-801 with antiestrogens inhibits melanoma cell proliferation. • These combinations greatly enhance the effects of the compounds individually. • MK-801 combined with tamoxifen active metabolites induces cell cycle arrest in G1. • The combination of MK-801 and antiestrogens is an innovative strategy for melanoma.« less
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Liang, Xinyue; Gu, Junlian; Yu, Dehai; Wang, Guanjun; Zhou, Lei; Zhang, Xiaoying; Zhao, Yuguang; Chen, Xiao; Zheng, Shirong; Liu, Qiang; Cai, Lu; Cui, Jiuwei; Li, Wei
2016-01-01
Hormesis and adaptive responses are 2 important biological effects of low-dose ionizing radiation (LDR). In normal tissue, LDR induces hormesis as evinced by increased cell proliferation; however, whether LDR also increases tumor cell proliferation needs to be investigated. In this study, cell proliferation was assayed by total cell numbers and the Cell Counting Kit 8 assay. Mitogen-activated protein kinases (MAPK)/extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3' -kinase(PI3K)-Akt (PI3K/AKT) phosphorylation were determined by Western blot analysis. Human embryonic lung fibroblast 2BS and lung cancer NCI-H446 cell lines were irradiated with LDR at different doses (20-100 mGy). In response to 20 to 75 mGy X-rays, cell proliferation was significantly increased in 2BS but not in NCI-H446 cells. In 2BS cells, LDR at 20 to 75 mGy also stimulated phosphorylation of MAPK/ERK pathway proteins including ERK, MEK, and Raf and of the PI3K/AKT pathway protein AKT. To test whether ERK1/2 and AKT pathway activation was involved in the stimulation of cell proliferation in 2BS cells, the MAPK/ERK and PI3K/AKT pathways were inhibited using their specific inhibitors, U0126 and LY294002. U0126 decreased the phosphorylation of ERK1/2, and LY294002 decreased the phosphorylation of AKT; each could significantly inhibit LDR-induced 2BS cell proliferation. However, LDR did not stimulate these kinases, and kinase inhibitors also did not affect cell proliferation in the NCI-H446 cells. These results suggest that LDR stimulates cell proliferation via the activation of both MAPK/ERK and PI3K/AKT signaling pathways in 2BS but not in NCI-H446 cells. This finding implies the potential for applying LDR to protect normal tissues from radiotherapy without diminishing the efficacy of tumor therapy.
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Liang, Xinyue; Gu, Junlian; Yu, Dehai; Wang, Guanjun; Zhou, Lei; Zhang, Xiaoying; Zhao, Yuguang; Chen, Xiao; Zheng, Shirong; Liu, Qiang; Cai, Lu
2016-01-01
Hormesis and adaptive responses are 2 important biological effects of low-dose ionizing radiation (LDR). In normal tissue, LDR induces hormesis as evinced by increased cell proliferation; however, whether LDR also increases tumor cell proliferation needs to be investigated. In this study, cell proliferation was assayed by total cell numbers and the Cell Counting Kit 8 assay. Mitogen-activated protein kinases (MAPK)/extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3′ -kinase(PI3K)-Akt (PI3K/AKT) phosphorylation were determined by Western blot analysis. Human embryonic lung fibroblast 2BS and lung cancer NCI-H446 cell lines were irradiated with LDR at different doses (20-100 mGy). In response to 20 to 75 mGy X-rays, cell proliferation was significantly increased in 2BS but not in NCI-H446 cells. In 2BS cells, LDR at 20 to 75 mGy also stimulated phosphorylation of MAPK/ERK pathway proteins including ERK, MEK, and Raf and of the PI3K/AKT pathway protein AKT. To test whether ERK1/2 and AKT pathway activation was involved in the stimulation of cell proliferation in 2BS cells, the MAPK/ERK and PI3K/AKT pathways were inhibited using their specific inhibitors, U0126 and LY294002. U0126 decreased the phosphorylation of ERK1/2, and LY294002 decreased the phosphorylation of AKT; each could significantly inhibit LDR-induced 2BS cell proliferation. However, LDR did not stimulate these kinases, and kinase inhibitors also did not affect cell proliferation in the NCI-H446 cells. These results suggest that LDR stimulates cell proliferation via the activation of both MAPK/ERK and PI3K/AKT signaling pathways in 2BS but not in NCI-H446 cells. This finding implies the potential for applying LDR to protect normal tissues from radiotherapy without diminishing the efficacy of tumor therapy. PMID:26788032
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Song, Rui; Zhang, Jia; Huang, Junhua; Hai, Tao
2018-05-11
Breast cancer is a common malignancy in women and long non-coding RNAs (lncRNAs) have been shown to play key roles in the development and progression of breast cancer. In the present study, we examined the biological role of lncRNA gastric carcinoma highly expressed transcript 1 (GHET1) in breast cancer. The expression of GHET1 was determined by qRT-PCR assay; CCK-8, colony formation, Transwell invasion and migration assays detected breast cancer cell proliferation, invasion and migration; cell apoptosis and cell cycle were determined by flow cytometry; protein levels were determined by western blot assay. GHET1 was up-regulated in breast cancer tissues and cell lines, and the up-regulation of GHET1 was positively correlated with larger tumor size, advanced clinical stage, lymph node metastasis and shorter overall survival. Knockdown of GHET1 suppressed cell proliferation, invasion and migration, and induced apoptosis and G0/G1 cell cycle arrest in MCF-cells. Knockdown of GHET1 also suppressed the protein levels of N-cadherin, vimentin, and decreased the protein level of E-cadherin in MCF-7 cells. On the other hand, overexpression of GHET1 promoted cell proliferation, invasion and migration, and inhibited cell apoptosis and increased cell population at S phase in BT-20 cells. Overexpression of GHET1 also promoted epithelial mesenchymal transition (EMT) in BT-20 cells. Furthermore, knockdown of GHET1 also suppressed in vivo tumor growth of MCF-7 cells, and also decreased the protein levels of N-cadherin and vimentin, and increased the protein levels of E-cadherin in the tumor tissues from the nude mice. Our results demonstrated that GHET1 was up-regulated in breast cancer tissues and cell lines, and promoted breast cancer cell proliferation, invasion and migration by affecting EMT. Our study for the first time revealed the biological functions of GHET1 in breast cancer.
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Zuma, Aline Araujo; Mendes, Isabela Cecília; Reignault, Lissa Catherine; Elias, Maria Carolina; de Souza, Wanderley; Machado, Carlos Renato; Motta, Maria Cristina M
2014-02-01
The protozoan Trypanosoma cruzi is the etiological agent of Chagas disease, which affects approximately 8 million people in Latin America. This parasite contains a single nucleus and a kinetoplast, which harbors the mitochondrial DNA (kDNA). DNA topoisomerases act during replication, transcription and repair and modulate DNA topology by reverting supercoiling in the DNA double-strand. In this work, we evaluated the effects promoted by camptothecin, a topoisomerase I inhibitor that promotes protozoan proliferation impairment, cell cycle arrest, ultrastructure alterations and DNA lesions in epimastigotes of T. cruzi. The results showed that inhibition of cell proliferation was reversible only at the lowest drug concentration (1μM) used. The unpacking of nuclear heterochromatin and mitochondrion swelling were the main ultrastructural modifications observed. Inhibition of parasite proliferation also led to cell cycle arrest, which was most likely caused by nuclear DNA lesions. Following camptothecin treatment, some of the cells restored their DNA, whereas others entered early apoptosis but did not progress to late apoptosis, indicating that the protozoa stay alive in a "senescence-like" state. This programmed cell death may be associated with a decrease in mitochondrial membrane potential and an increase in the production of reactive oxygen species. Taken together, these results indicate that the inhibition of T. cruzi proliferation is related to events capable of affecting cell cycle, DNA organization and mitochondrial activity. Copyright © 2014. Published by Elsevier B.V.
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Hoppe, T; Kraus, D; Novak, N; Probstmeier, R; Frentzen, M; Wenghoefer, M; Jepsen, S; Winter, J
2016-10-01
The impact of oral pathogens onto the generation and variability of oral tumors has only recently been investigated. To get further insights, oral cancer cells were treated with pathogens and additionally, as a result of this bacterial cellular infection, with human defensins, which are as anti-microbial peptide members of the innate immune system. After cell stimulation, proliferation behavior, expression analysis of oncogenic relevant defensin genes, and effects on EGFR signaling were investigated. The expression of oncogenic relevant anti-microbial peptides was analyzed with real-time PCR and immunohistochemistry. Cell culture experiments were performed to examine cellular impacts caused by stimulation, i.e., altered gene expression, proliferation rate, and EGF receptor-dependent signaling. Incubation of oral tumor cells with an oral pathogen (Porphyromonas gingivalis) and human α-defensins led to an increase in cell proliferation. In contrast, another oral bacterium used, Aggregatibacter actinomycetemcomitans, enhanced cell death. The bacteria and anti-microbial peptides exhibited diverse effects on the transcript levels of oncogenic relevant defensin genes and epidermal growth factor receptor signaling. These two oral pathogens exhibited opposite primary effects on the proliferation behavior of oral tumor cells. Nevertheless, both microbe species led to similar secondary impacts on the proliferation rate by modifying expression levels of oncogenic relevant α-defensin genes. In this respect, oral pathogens exerted multiplying effects on tumor cell proliferation. Additionally, human defensins were shown to differently influence epidermal growth factor receptor signaling, supporting the hypothesis that these anti-microbial peptides serve as ligands of EGFR, thus modifying the proliferation behavior of oral tumor cells.
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Experimental study on the estrogen-like effect of boric Acid.
Wang, Yadong; Zhao, Yingzheng; Chen, Xiaoyu
2008-02-01
There are now considerable evidences that boric acid has reproductive and developmental toxicity, but it is uncertain whether such toxicity is caused by estrogen-like effect. Our objective is to determine the estrogen-like effect of boric acid. Proliferation assay of MCF-7 human breast cancer cells, uterotrophic assay, measure assay of the estradiol (E2), proliferation assay of mucous membrane cells, and assay of estrogen receptor were conducted in this study. Boric acid could increase the weight of uterus of ovariectomized SD rats and the height of epithelium cells of mucous membrane, enhance the expression of the proliferating cell nucleus antigen, and reduce the density of estrogen receptors. However, boric acid could not affect the level of estradiol in serum and stimulate the proliferation of MCF-7 human breast cancer cells. In this study, boric acid exhibited the estrogen-like effect in vivo.
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Caroprese, M; Ciliberti, M G; De Palo, P; Santillo, A; Sevi, A; Albenzio, M
2018-06-27
The present experiment aimed at understanding the effects of cortisol levels on sheep peripheral blood mononuclear cell (PBMC) proliferation and cytokine production during hyperthermia. To mimic stress related to the exposition of high ambient temperatures, PBMC were cultured at 43°C for 12 h, and subsequently at 39°C for additional 12 h. Cells in normothermia were cultured at 39°C for 24 h. Phytohemagglutinin-stimulated PBMC were cultured with different cortisol levels: 0 ng/mL; 100 ng/mL, representing the physiological cortisol concentration simulating stress condition (Cort100); and 1,000 ng/mL, representing the hyperactivated hypothalamic-pituitary-adrenal axis (Cort1000). Phytohemagglutinin-stimulated PBMC with 0 ng/mL of cortisol concentration represented the positive control, whereas nonstimulated PBMC without cortisol represented the negative control (NC). The free cell supernatants were collected for the determination of IL-6, IL-1β, and IL-10 by ELISA. Bromodeoxyuridine assay was performed on cells to determine cell proliferation. Exposition to hyperthermia negatively affected cell proliferation, IL-6, IL-1β, and IL-10 concentrations in cell supernatants. The interaction of hyperthermia and cortisol level affected both cell proliferation and IL-10 production. Both PBMC proliferation and IL-10 production in positive control, Cort100, and Cort100 decreased at 43°C as compared with 39°C NC. On average, the Cort100 treatment displayed higher concentrations of IL-6 than NC. The present experiment demonstrated that the action of cortisol concentration simulating stress condition on cell proliferation and cytokine production was a permissive/stimulatory action during normothermia, whereas it was a suppressive action during hyperthermia. These data confirmed that cortisol concentration simulating stress condition could have a role in the immune system of sheep via mediating cellular homeostasis in the condition of hyperthermia. The negative effects of hyperthermia on sheep immune responses were apparent when performing an immunological challenge. The Authors. Published by FASS Inc. and Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).
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Cell proliferation of Paramecium tetraurelia under simulated microgravity
NASA Astrophysics Data System (ADS)
Sawai, S.; Mogami, Y.; Baba, S. A.
Paramecium is known to proliferate faster under microgravity in space and slower under hypergravity Experiments using axenic culture medium have demonstrated that the hypergravity affected directly on the proliferation of Paramecium itself Kato et al 2003 In order to assess the mechanisms underlying the physiological effects of gravity on cell proliferation Paramecium tetraurelia was grown under simulated microgravity performed by clinorotation and the time course of the proliferation was investigated in detail on the basis of the logistic analysis P tetraurelia was cultivated in a closed chamber in which cells were confined without air babbles reducing the shear stresses and turbulence under the rotation The chamber is made of quartz and silicone rubber film the former is for the optically-flat walls for the measurement of cell density by means of a non-invasive laser optical-slice method and the latter for gas exchange Because the closed chamber has an inner dimension of 3 times 3 times 60 mm Paramecium does not accumulate at the top of the chamber despite its negative gravitactic behavior We measured the cell density at regular time intervals without breaking the configuration of the chamber and analyzed the proliferation parameters by fitting the data to a logistic equation Clinorotation had the effects of reducing the proliferation of P tetraurelia It reduced both the saturation cell density and the maximum proliferation rate although it had little effect on the
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Ma, Meng; Wang, Qian; Li, Zhanjie; Cheng, Huihui; Li, Zhaojie; Liu, Xiangli; Song, Weining; Appels, Rudi; Zhao, Huixian
2015-07-01
Several studies have described quantitative trait loci (QTL) for seed size in wheat, but the relevant genes and molecular mechanisms remain largely unknown. Here we report the functional characterization of the wheat TaCYP78A3 gene and its effect on seed size. TaCYP78A3 encoded wheat cytochrome P450 CYP78A3, and was specifically expressed in wheat reproductive organs. TaCYP78A3 activity was positively correlated with the final seed size. Its silencing caused a reduction of cell number in the seed coat, resulting in an 11% decrease in wheat seed size, whereas TaCYP78A3 over-expression induced production of more cells in the seed coat, leading to an 11-48% increase in Arabidopsis seed size. In addition, the cell number in the final seed coat was determined by the TaCYP78A3 expression level, which affected the extent of integument cell proliferation in the developing ovule and seed. Unfortunately, TaCYP78A3 over-expression in Arabidopsis caused a reduced seed set due to an ovule developmental defect. Moreover, TaCYP78A3 over-expression affected embryo development by promoting embryo integument cell proliferation during seed development, which also ultimately affected the final seed size in Arabidopsis. In summary, our results indicated that TaCYP78A3 plays critical roles in influencing seed size by affecting the extent of integument cell proliferation. The present study provides direct evidence that TaCYP78A3 affects seed size in wheat, and contributes to an understanding of the cellular basis of the gene influencing seed development. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Park, Hyejin; Yoon, Min-Sik; Ryu, Kwon-Yul, E-mail: kyryu@uos.ac.kr
Highlights: •Proliferation capacity of Ubc{sup −/−} FLCs was reduced during culture in vitro. •Ubc is required for proliferation of both hepatocytes and bipotent FLEPCs. •Bipotent FLEPCs exhibit highest Ubc transcription and proliferation capacity. •Cell types responsible for Ubc{sup −/−} fetal liver developmental defect were identified. -- Abstract: We have previously demonstrated that disruption of polyubiquitin gene Ubc leads to mid-gestation embryonic lethality most likely due to a defect in fetal liver development, which can be partially rescued by ectopic expression of Ub. In a previous study, we assessed the cause of embryonic lethality with respect to the fetal liver hematopoieticmore » system. We confirmed that Ubc{sup −/−} embryonic lethality could not be attributed to impaired function of hematopoietic stem cells, which raises the question of whether or not FLECs such as hepatocytes and bile duct cells, the most abundant cell types in the liver, are affected by disruption of Ubc and contribute to embryonic lethality. To answer this, we isolated FLCs from E13.5 embryos and cultured them in vitro. We found that proliferation capacity of Ubc{sup −/−} cells was significantly reduced compared to that of control cells, especially during the early culture period, however we did not observe the increased number of apoptotic cells. Furthermore, levels of Ub conjugate, but not free Ub, decreased upon disruption of Ubc expression in FLCs, and this could not be compensated for by upregulation of other poly- or mono-ubiquitin genes. Intriguingly, the highest Ubc expression levels throughout the entire culture period were observed in bipotent FLEPCs. Hepatocytes and bipotent FLEPCs were most affected by disruption of Ubc, resulting in defective proliferation as well as reduced cell numbers in vitro. These results suggest that defective proliferation of these cell types may contribute to severe reduction of fetal liver size and potentially mid-gestation lethality of Ubc{sup −/−} embryos.« less
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Keratinocyte Motility Is Affected by UVA Radiation-A Comparison between Normal and Dysplastic Cells.
Niculiţe, Cristina M; Nechifor, Marina T; Urs, Andreea O; Olariu, Laura; Ceafalan, Laura C; Leabu, Mircea
2018-06-07
UVA radiation induces multiple and complex changes in the skin, affecting epidermal cell behavior. This study reports the effects of UVA exposure on normal (HaCaT) and dysplastic (DOK) keratinocytes. The adherence, spreading and proliferation were investigated by time-lapse measurement of cell layer impedance on different matrix proteins. Prior to UVA exposure, the time required for adherence and spreading did not differ significantly for HaCaT and DOK cells, while spreading areas were larger for HaCaT cells. Under UVA exposure, HaCaT and DOK cells behavior differed in terms of movement and proliferation. The cells' ability to cover the denuded surface and individual cell trajectories were recorded by time-lapse videomicroscopy, during wound healing experiments. Dysplastic keratinocytes showed more sensitivity to UVA, exhibiting transient deficiencies in directionality of movement and a delay in re-coating the denuded area. The actin cytoskeleton displayed a cortical organization immediately after irradiation, in both cell lines, similar to mock-irradiated cells. Post-irradiation, DOK cells displayed a better organization of stress fibers, persistent filopodia, and new, stronger focal contacts. In conclusion, after UVA exposure HaCaT and DOK cells showed a different behavior in terms of adherence, spreading, motility, proliferation, and actin cytoskeleton dynamics, with the dyplastic keratinocytes being more sensitive.
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The cell proliferation antigen Ki-67 organises heterochromatin
Sobecki, Michal; Mrouj, Karim; Camasses, Alain; Parisis, Nikolaos; Nicolas, Emilien; Llères, David; Gerbe, François; Prieto, Susana; Krasinska, Liliana; David, Alexandre; Eguren, Manuel; Birling, Marie-Christine; Urbach, Serge; Hem, Sonia; Déjardin, Jérôme; Malumbres, Marcos; Jay, Philippe; Dulic, Vjekoslav; Lafontaine, Denis LJ; Feil, Robert; Fisher, Daniel
2016-01-01
Antigen Ki-67 is a nuclear protein expressed in proliferating mammalian cells. It is widely used in cancer histopathology but its functions remain unclear. Here, we show that Ki-67 controls heterochromatin organisation. Altering Ki-67 expression levels did not significantly affect cell proliferation in vivo. Ki-67 mutant mice developed normally and cells lacking Ki-67 proliferated efficiently. Conversely, upregulation of Ki-67 expression in differentiated tissues did not prevent cell cycle arrest. Ki-67 interactors included proteins involved in nucleolar processes and chromatin regulators. Ki-67 depletion disrupted nucleologenesis but did not inhibit pre-rRNA processing. In contrast, it altered gene expression. Ki-67 silencing also had wide-ranging effects on chromatin organisation, disrupting heterochromatin compaction and long-range genomic interactions. Trimethylation of histone H3K9 and H4K20 was relocalised within the nucleus. Finally, overexpression of human or Xenopus Ki-67 induced ectopic heterochromatin formation. Altogether, our results suggest that Ki-67 expression in proliferating cells spatially organises heterochromatin, thereby controlling gene expression. DOI: http://dx.doi.org/10.7554/eLife.13722.001 PMID:26949251
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Lin, Alexander Y T; Pearson, Bret J
2014-03-01
During adult homeostasis and regeneration, the freshwater planarian must accomplish a constant balance between cell proliferation and cell death, while also maintaining proper tissue and organ size and patterning. How these ordered processes are precisely modulated remains relatively unknown. Here we show that planarians use the downstream effector of the Hippo signaling cascade, yorkie (yki; YAP in vertebrates) to control a diverse set of pleiotropic processes in organ homeostasis, stem cell regulation, regeneration and axial patterning. We show that yki functions to maintain the homeostasis of the planarian excretory (protonephridial) system and to limit stem cell proliferation, but does not affect the differentiation process or cell death. Finally, we show that Yki acts synergistically with WNT/β-catenin signaling to repress head determination by limiting the expression domains of posterior WNT genes and that of the WNT-inhibitor notum. Together, our data show that yki is a key gene in planarians that integrates stem cell proliferation control, organ homeostasis, and the spatial patterning of tissues.
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p62 Regulates the Proliferation of Molecular Apocrine Breast Cancer Cells
Nozaki, Fumi; Hirotani, Yukari; Nakanishi, Yoko; Yamaguchi, Hiromi; Nishimaki, Haruna; Noda, Hiroko; Tang, Xiaoyan; Yamamoto, Hisae; Suzuki, Atsuko; Seki, Toshimi; Masuda, Shinobu
2016-01-01
p62, also called sequestosome 1 (SQSTM1), is a multifunctional signaling molecule that affects cell proliferation. Recently, we found accumulation of p62 in apocrine carcinoma of the breast, however, the biological role of p62 expression in apocrine carcinoma still remains unclear. To investigate whether p62 might contribute to tumor cell proliferation in apocrine carcinomas, we used the MDA-MB-453 (androgen receptor-positive, HER2-type) and MFM223 (androgen receptor-positive, triple-negative type) breast cancer cell lines as models of molecular apocrine carcinoma. Both MDA-MB-453 and MFM223 showed strong and d high p62 protein expression than MCF7 cells (androgen receptor-negative, luminal A type). Knockdown of p62 resulted in significant reduction of the cell proliferative activity in both MDA-MB-453 (P<0.01) and MFM223 (P<0.05). In conclusion, p62 could contribute to cell proliferation and represent a therapeutic target in apocrine carcinoma. PMID:27682016
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p62 Regulates the Proliferation of Molecular Apocrine Breast Cancer Cells.
Nozaki, Fumi; Hirotani, Yukari; Nakanishi, Yoko; Yamaguchi, Hiromi; Nishimaki, Haruna; Noda, Hiroko; Tang, Xiaoyan; Yamamoto, Hisae; Suzuki, Atsuko; Seki, Toshimi; Masuda, Shinobu
2016-08-30
p62, also called sequestosome 1 (SQSTM1), is a multifunctional signaling molecule that affects cell proliferation. Recently, we found accumulation of p62 in apocrine carcinoma of the breast, however, the biological role of p62 expression in apocrine carcinoma still remains unclear. To investigate whether p62 might contribute to tumor cell proliferation in apocrine carcinomas, we used the MDA-MB-453 (androgen receptor-positive, HER2-type) and MFM223 (androgen receptor-positive, triple-negative type) breast cancer cell lines as models of molecular apocrine carcinoma. Both MDA-MB-453 and MFM223 showed strong and d high p62 protein expression than MCF7 cells (androgen receptor-negative, luminal A type). Knockdown of p62 resulted in significant reduction of the cell proliferative activity in both MDA-MB-453 (P<0.01) and MFM223 (P<0.05). In conclusion, p62 could contribute to cell proliferation and represent a therapeutic target in apocrine carcinoma.
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Kalyanaraman, Balaraman; Cheng, Gang; Hardy, Micael; Ouari, Olivier; Lopez, Marcos; Joseph, Joy; Zielonka, Jacek; Dwinell, Michael B
2018-04-01
The present review is a sequel to the previous review on cancer metabolism published in this journal. This review focuses on the selective antiproliferative and cytotoxic effects of mitochondria-targeted therapeutics (MTTs) in cancer cells. Emerging research reveals a key role of mitochondrial respiration on tumor proliferation. Previously, a mitochondria-targeted nitroxide was shown to selectively inhibit colon cancer cell proliferation at submicromolar levels. This review is centered on the therapeutic use of MTTs and their bioenergetic profiling in cancer cells. Triphenylphosphonium cation conjugated to a parent molecule (e.g., vitamin-E or chromanol, ubiquinone, and metformin) via a linker alkyl chain is considered an MTT. MTTs selectively and potently inhibit proliferation of cancer cells and, in some cases, induce cytotoxicity. MTTs inhibit mitochondrial complex I activity and induce mitochondrial stress in cancer cells through generation of reactive oxygen species. MTTs in combination with glycolytic inhibitors synergistically inhibit tumor cell proliferation. This review discusses how signaling molecules traditionally linked to tumor cell proliferation affect tumor metabolism and bioenergetics (glycolysis, TCA cycle, and glutaminolysis). Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.
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Lack of effect of a granulocyte proliferation inhibitor or their committed precursor cells.
Lord, B I; Testa, N G; Wright, E G; Banerjee, R K
1977-05-01
Using the agar culture technique, we have measured the effect of granulocyte extracts GCE (and of erythrocyte-RCE and lymph node extracts-LNE) on the growth and proliferation of the committed granulocytic precursor cells, CFU-C. In addition we have determined their effects on the proliferation of the developing colony cells and on the ultimate cell production in the colonies. The results show that GCE has no effect on the growth or proliferative activity on the CFU-C. It does, however, reduce both the autoradiographic labelling indices of the developing colony cells and the net colony cellularities, acting as a cell cycle modulator. These are effects specific to the GCE since at the dose levels used, neither RCE nor LNE affected these measurements.
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Tan, Thomas C J; Knight, John; Sbarrato, Thomas; Dudek, Kate; Willis, Anne E; Zamoyska, Rose
2017-07-25
Global transcriptomic and proteomic analyses of T cells have been rich sources of unbiased data for understanding T-cell activation. Lack of full concordance of these datasets has illustrated that important facets of T-cell activation are controlled at the level of translation. We undertook translatome analysis of CD8 T-cell activation, combining polysome profiling and microarray analysis. We revealed that altering T-cell receptor stimulation influenced recruitment of mRNAs to heavy polysomes and translation of subsets of genes. A major pathway that was compromised, when TCR signaling was suboptimal, was linked to ribosome biogenesis, a rate-limiting factor in both cell growth and proliferation. Defective TCR signaling affected transcription and processing of ribosomal RNA precursors, as well as the translation of specific ribosomal proteins and translation factors. Mechanistically, IL-2 production was compromised in weakly stimulated T cells, affecting the abundance of Myc protein, a known regulator of ribosome biogenesis. Consequently, weakly activated T cells showed impaired production of ribosomes and a failure to maintain proliferative capacity after stimulation. We demonstrate that primary T cells respond to various environmental cues by regulating ribosome biogenesis and mRNA translation at multiple levels to sustain proliferation and differentiation.
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Chen, Hongxin; Goodus, Matthew T; de Toledo, Sonia M; Azzam, Edouard I; Levison, Steven W
2015-01-01
Damage to normal human brain cells from exposure to ionizing radiation may occur during the course of radiotherapy or from accidental exposure. Delayed effects may complicate the immediate effects resulting in neurodegeneration and cognitive decline. We examined cellular and molecular changes associated with exposure of neural stem/progenitor cells (NSPs) to 137Cs γ-ray doses in the range of 0 to 8 Gy. Subventricular zone NSPs isolated from newborn mouse pups were analyzed for proliferation, self-renewal, and differentiation, shortly after irradiation. Strikingly, there was no apparent increase in the fraction of dying cells after irradiation, and the number of single cells that formed neurospheres showed no significant change from control. Upon differentiation, irradiated neural precursors did not differ in their ability to generate neurons, astrocytes, and oligodendrocytes. By contrast, progression of NSPs through the cell cycle decreased dramatically after exposure to 8 Gy (p < .001). Mice at postnatal day 10 were exposed to 8 Gy of γ rays delivered to the whole body and NSPs of the subventricular zone were analyzed using a four-color flow cytometry panel combined with ethynyl deoxyuridine incorporation. Similar flow cytometric analyses were performed on NSPs cultured as neurospheres. These studies revealed that neither the percentage of neural stem cells nor their proliferation was affected. By contrast, γ-irradiation decreased the proliferation of two classes of multipotent cells and increased the proliferation of a specific glial-restricted precursor. Altogether, these results support the conclusion that primitive neural precursors are radioresistant, but their proliferation is slowed down as a consequence of γ-ray exposure. PMID:26056396
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Chen, Hui; Du, Shanshan; Zhen, Ping; Li, Xusheng; Liang, Xiaoyan; Fan, Lijuan; Jiang, Jie; Yang, Haiou; Liu, Jun
2016-12-28
To evaluate the effect of laser solid forming (LSF) of porous titanium on receptor activator of NF-κB ligand (RANKL)/osteoprorotegerin (OPG) expression and osteoblast cells growth. Methods: The DMEM and sterile saline were used for porous titanium extract. The osteoblast cells were cultured in the extract while equal amount of DMEM and sterile saline were added to the control group. The growth of the cells were observed under an inverted phase contrast microscope. MTT was used to detect the growth inhibitory rates. The adhesion capacity of osteoblasts were measured. The growth in the material surface was examined by the electron microscope, and the expressions of RANKL and OPG were determined by Westen blot. Results: At the first day, the osteoblast proliferation rate was significantly different (P<0.05), at the fourth and seventh day, the osteoblast proliferation rate was not significantly affected in the LSF group (P>0.05); at each time point, the osteoblast proliferation rate were significantly different between the two groups (P<0.05). Compared with the control group, RANKL and OPG protein expression were not significantly different (P>0.05). The laser solid forming of porous titanium showed well bone compatibility. Conclusion: The porous titanium did not affect osteoblast proliferation due to its well bone compatibility. It did not affect the OPG/RANKL/RANK-axis system of bone metabolism, exibiting a wide applicable prospect for tissue engineering.
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Li, Dan; Li, Yufeng; Hernandez, Jessica A.; Patenia, Rebecca; Kim, Tae Kon; Khalili, Jahan; Dougherty, Mark C.; Hanley, Patrick J.; Bollard, Catherine M.; Komanduri, Krishna V.; Hwu, Patrick; Champlin, Richard E.; Radvanyi, Laszlo G.; Molldrem, Jeffrey J.; Ma, Qing
2016-01-01
Statin treatment has been shown to reduce graft-versus-host disease (GVHD) while preserving graft-versus-tumor (GVT) effect in allogeneic stem cell transplantation (allo-HCT). Herein, we investigated whether lovastatin treatment affects the function of human cytolytic T lymphocytes (CTLs). Upon TCR stimulation, lovastatin significantly inhibited the proliferation of both CD4+ and CD8+ T cells from healthy donors while their intracellular cytokine production including IFN-γ and TNF-α remained the same with a slight decrease of IL-2. Moreover, the specific lysis of target cells by CTL lines derived from patients and normal donors specific for EBV-encoded antigen LMP2 or CMV-encoded antigen pp65 was uncompromised in the presence of lovastatin. In addition, we evaluated the effect of lovastatin on the proliferation and effector function of the CD8+ tumor–infiltrating lymphocytes (TILs) derived from melanoma patients specific for MART-1 antigen. Lovastatin significantly reduced the expansion of antigen-specific TILs upon MART-1 stimulation. However, the effector function of TILs, including the specific lysis of target cells and secretion of cytokine IFN-γ, remained intact with lovastatin treatment. Taken together, these data demonstrated that lovastatin inhibits the proliferation of EBV-, CMV- and MART-1-specific CTLs without affecting cytolytic capacity. The differential effect of lovastatin on the proliferation versus cytoxicity of CTLs might shed some light on elucidating the possible mechanisms of GVHD and GVT effect elicited by alloimmune responses. PMID:20948439
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Naito, Tomoaki; Mulet, Céline; De Castro, Cristina; Molinaro, Antonio; Saffarian, Azadeh; Nigro, Giulia; Bérard, Marion; Clerc, Mélanie; Pedersen, Amy B.; Pédron, Thierry
2017-01-01
ABSTRACT We identified a crypt-specific core microbiota (CSCM) dominated by strictly aerobic, nonfermentative bacteria in murine cecal and proximal colonic (PC) crypts and hypothesized that, among its possible functions, it may affect epithelial regeneration. In the present work, we isolated representative CSCM strains using selective media based upon our initial 16S rRNA-based molecular identification (i.e., Acinetobacter, Delftia, and Stenotrophomonas). Their tropism for the crypt was confirmed, and their influence on epithelial regeneration was demonstrated in vivo by monocolonization of germfree mice. We also showed that lipopolysaccharide (LPS), through its endotoxin activity, was the dominant bacterial agonist controlling proliferation. The relevant molecular mechanisms were analyzed using colonic crypt-derived organoids exposed to bacterial sonicates or highly purified LPS as agonists. We identified a Toll-like receptor 4 (TLR4)-dependent program affecting crypts at different stages of epithelial differentiation. LPS played a dual role: it repressed cell proliferation through RIPK3-mediated necroptosis of stem cells and cells of the transit-amplifying compartment and concurrently enhanced cell differentiation, particularly the goblet cell lineage. PMID:29042502
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Kimira, Yoshifumi, E-mail: kimira@josai.ac.jp; Ogura, Kana; Taniuchi, Yuri
Highlights: • Pro-Hyp did not affect MC3T3-E1 cell proliferation and matrix mineralization. • Pro-Hyp significantly increased alkaline phosphatase activity. • Pro-Hyp significantly upregulated gene expression of Runx2, Osterix, and Col1α1. - Abstract: Prolyl-hydroxyproline (Pro-Hyp) is one of the major constituents of collagen-derived dipeptides. The objective of this study was to investigate the effects of Pro-Hyp on the proliferation and differentiation of MC3T3-E1 osteoblastic cells. Addition of Pro-Hyp did not affect MC3T3-E1 cell proliferation and matrix mineralization but alkaline phosphatase activity was significantly increased. Furthermore, cells treated with Pro-Hyp significantly upregulated gene expression of Runx2, Osterix, and Col1α1. These results indicatemore » that Pro-Hyp promotes osteoblast differentiation. This study demonstrates for the first time that Pro-Hyp has a positive effect on osteoblast differentiation with upregulation of Runx2, Osterix, and Collα1 gene expression.« less
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Würth, Roberto; Pattarozzi, Alessandra; Gatti, Monica; Bajetto, Adirano; Corsaro, Alessandro; Parodi, Alessia; Sirito, Rodolfo; Massollo, Michela; Marini, Cecilia; Zona, Gianluigi; Fenoglio, Daniela; Sambuceti, Gianmario; Filaci, Gilberto; Daga, Antonio; Barbieri, Federica; Florio, Tullio
2013-01-01
Cancer stem cell theory postulates that a small population of tumor-initiating cells is responsible for the development, progression and recurrence of several malignancies, including glioblastoma. In this perspective, tumor-initiating cells represent the most relevant target to obtain effective cancer treatment. Metformin, a first-line drug for type II diabetes, was reported to possess anticancer properties affecting the survival of cancer stem cells in breast cancer models. We report that metformin treatment reduced the proliferation rate of tumor-initiating cell-enriched cultures isolated from four human glioblastomas. Metformin also impairs tumor-initiating cell spherogenesis, indicating a direct effect on self-renewal mechanisms. Interestingly, analyzing by FACS the antiproliferative effects of metformin on CD133-expressing subpopulation, a component of glioblastoma cancer stem cells, a higher reduction of proliferation was observed as compared with CD133-negative cells, suggesting a certain degree of cancer stem cell selectivity in its effects. In fact, glioblastoma cell differentiation strongly reduced sensitivity to metformin treatment. Metformin effects in tumor-initiating cell-enriched cultures were associated with a powerful inhibition of Akt-dependent cell survival pathway, while this pathway was not affected in differentiated cells. The specificity of metformin antiproliferative effects toward glioblastoma tumor-initiating cells was confirmed by the lack of significant inhibition of normal human stem cells (umbilical cord-derived mesenchymal stem cells) in vitro proliferation after metformin exposure. Altogether, these data clearly suggest that metformin exerts antiproliferative activity on glioblastoma cells, showing a higher specificity toward tumor-initiating cells, and that the inhibition of Akt pathway may represent a possible intracellular target of this effect.
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Cell culture density affects the proliferation activity of human adipose tissue stem cells.
Kim, Dae Seong; Lee, Myoung Woo; Ko, Young Jong; Chun, Yong Hoon; Kim, Hyung Joon; Sung, Ki Woong; Koo, Hong Hoe; Yoo, Keon Hee
2016-01-01
In this study, we investigated the effect of cell density on the proliferation activity of human mesenchymal stem cells (MSCs) derived from adipose tissue (AT-MSCs) over time in culture. Passage #4 (P4) and #12 (P12) AT-MSCs from two donors were plated at a density of 200 (culture condition 1, CC1) or 5000 (culture condition 2, CC2) cells cm(-2) . After 7 days of incubation, P4 and P12 AT-MSCs cultured in CC1 were thin and spindle-shaped, whereas those cultured in CC2 had extensive cell-to-cell contacts and an expanded cell volume. In addition, P4 and P12 AT-MSCs in CC1 divided more than three times, while those in CC2 divided less than once on average. Flow cytometric analysis using 5(6)-carboxyfluorescein diacetate N-succinimidyl ester dye showed that the fluorescence intensity of AT-MSCs was lower in CC1 than in CC2. Furthermore, expression of proliferation-associated genes, such as CDC45L, CDC20A and KIF20A, in P4 AT-MSCs was higher in CC1 than in CC2, and this difference was also observed in P12 AT-MSCs. These data demonstrated that cell culture density affects the proliferation activity of MSCs, suggesting that it is feasible to design a strategy to prepare suitable MSCs using specific culture conditions. Copyright © 2016 John Wiley & Sons, Ltd.
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Abdullah, Kay M; Abdullah, Ahmed; Johnson, Mary Lynn; Bilski, Jerzy J; Petry, Kimberly; Redmer, Dale A; Reynolds, Lawrence P; Grazul-Bilska, Anna T
2003-10-01
To evaluate the effects of Aloe vera on gap junctional intercellular communication (GJIC) and proliferation of human skin fibroblasts in the presence or absence of basic fibroblast growth factor (FGF-2). In vitro study using human type II diabetic and nondiabetic skin fibroblast cell lines. Diabetic (n = 4) and nondiabetic (n = 4) human skin fibroblast cell lines were purchased from Coriell Institute for Medical Research (Camden, NJ). The cells were cultured with or without Aloe vera extract in increasing concentrations (0%, 0.625%, 1.25%, 2.5%, 5%, 10%, and 20%; v/v) in culture medium and with or without FGF-2 (30 ng/mL). GJIC was evaluated after 48-hour incubation with treatments by laser cytometry. Cells were counted after 72-hour incubation with treatments by using a Coulter counter. The rate of GJIC was greater (p < 0.01) for diabetic than for nondiabetic fibroblasts (3.5 +/- 0.1 versus 3.0 +/- 0.1% per minute during the first 4 minutes after photobleaching). GJIC was increased ( p < 0.05) for diabetic fibroblasts in the presence of 2.5% and 5% of Aloe vera extract (4.2 +/- 0.1 and 4.0 +/- 0.2 versus 3.5 +/- 0.1% per minute for control, respectively). FGF-2 stimulated (p < 0.01) GJIC for diabetic (4.0 +/- 0.1 versus 3.5 +/- 0.1% per minute for control) and nondiabetic (3.5 +/- 0.1 versus 3.0 +/- 0.1% per minute for control) fibroblasts. Aloe vera extract did not affect GJIC of nondiabetic fibroblast cultured without FGF-2. However, Aloe vera extract decreased (p < 0.05) FGF-2 stimulatory effects on GJIC of diabetic and nondiabetic fibroblasts. Proliferation of diabetic fibroblasts was increased (p < 0.05) by 1.25% and 2.5% Aloe vera extract in medium. Proliferation of nondiabetic fibroblasts was not affected by Aloe vera extract. FGF-2 increased (p < 0.05) proliferation of nondiabetic fibroblasts and FGF-2 did not affect proliferation of diabetic fibroblasts. Aloe vera extract decreased (p < 0.05) FGF-2 stimulatory effects on proliferation of nondiabetic fibroblasts. These data demonstrate that Aloe vera has the ability to stimulate GJIC and proliferation of human skin fibroblasts in diabetes mellitus. Furthermore, these results indicate that Aloe vera contains a compound(s) that neutralizes, binds with FGF-2 receptor, or otherwise alters signaling pathways for FGF-2. By affecting both GJIC and proliferation of diabetic fibroblasts, Aloe vera may improve wound healing in diabetes mellitus.
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Kang, Kyung Shin; Lee, Seung-Jae; Lee, Haksue; Moon, Wonkyu
2011-01-01
We observed how combined mechanical stimuli affect the proliferation and differentiation of pre-osteoblasts. For this research, a bioreactor system was developed that can simultaneously stimulate cells with cyclic strain and ultrasound, each of which is known to effectively stimulate bone tissue regeneration. MC3T3-E1 pre-osteoblasts were chosen for bone tissue engineering due to their osteoblast-like characteristics. 3-D scaffolds were fabricated with polycaprolactone and poly-L-lactic acid using the salt leaching method. The cells were stimulated by the bioreactor with cyclic strain and ultrasound. The bioreactor was set at a frequency of 1.0 Hz and 10% strain for cyclic strain and 1.0 MHz and 30 mW/cm2 for ultrasound. Three experimental groups (ultrasound, cyclic strain, and combined stimulation) and a control group were examined. Each group was stimulated for 20 min/day. Mechanical stimuli did not affect MC3T3-E1 cell proliferation significantly up to 10 days when measured with the cell counting kit-8. However, gene expression analysis of collagen type-I, osteocalcin, RUNX2, and osterix revealed that the combined mechanical stimulation accelerated the matrix maturation of MC3T3-E1 cells. These results indicate that the combined mechanical stimulation can enhance the differentiation of pre-osteoblasts more efficiently than simple stimuli, in spite of no effect on cell proliferation. PMID:21532314
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Ahola, Tytti M; Manninen, Tommi; Alkio, Niina; Ylikomi, Timo
2002-09-01
The issue of how progesterone affects mammary gland growth is controversial, and the mechanism governing the effects of the hormone remains mostly unknown. We have previously shown that G protein-coupled receptor 30 (GPR30) is a progestin target gene whose expression correlates with progestin-induced growth inhibition in breast cancer cells. In this study, we investigate the role of GPR30 in regulating cell proliferation and mediating progestin-induced growth inhibition. When progestin failed to inhibit the growth of MCF-7 cells and instead stimulated growth, GPR30 was down-regulated. In this way, the inhibitory or stimulatory affects that progestin has on proliferation correlated with the level of expression of GPR30. Transient expression of GPR30 resulted in a marked inhibition of cell proliferation independent of estrogen treatment. GPR30 antisense was used to evaluate the role of GPR30 expression in progestin-induced growth inhibition. A diminished GPR30 mRNA expression by the antisense stimulated growth. Interestingly, GPR30 antisense abrogated the growth inhibitory effect of progestin and progesterone. Indeed, progestin induced 1) a reduction in cell proliferation, 2) G1-phase arrest, and 3) down-regulation of cyclin D1 was diminished. These data suggest that the orphan receptor, GPR30, is important for the inhibitory effect of progestin on growth.
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Kamal, Khaled Y; Herranz, Raúl; van Loon, Jack J W A; Medina, F Javier
2018-04-23
Gravity is the only component of Earth environment that remained constant throughout the entire process of biological evolution. However, it is still unclear how gravity affects plant growth and development. In this study, an in vitro cell culture of Arabidopsis thaliana was exposed to different altered gravity conditions, namely simulated reduced gravity (simulated microgravity, simulated Mars gravity) and hypergravity (2g), to study changes in cell proliferation, cell growth, and epigenetics. The effects after 3, 14, and 24-hours of exposure were evaluated. The most relevant alterations were found in the 24-hour treatment, being more significant for simulated reduced gravity than hypergravity. Cell proliferation and growth were uncoupled under simulated reduced gravity, similarly, as found in meristematic cells from seedlings grown in real or simulated microgravity. The distribution of cell cycle phases was changed, as well as the levels and gene transcription of the tested cell cycle regulators. Ribosome biogenesis was decreased, according to levels and gene transcription of nucleolar proteins and the number of inactive nucleoli. Furthermore, we found alterations in the epigenetic modifications of chromatin. These results show that altered gravity effects include a serious disturbance of cell proliferation and growth, which are cellular functions essential for normal plant development.
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Wang, Wenchao; Li, Zhiyuan; Liu, Juanjuan; Yang, Xingxing; Ji, Xinmiao; Luo, Yan; Hu, Chen; Hou, Yubin; He, Qianqian; Fang, Jun; Wang, Junfeng; Liu, Qingsong; Li, Guohui; Lu, Qingyou; Zhang, Xin
2016-01-01
Static magnetic fields (SMFs) can affect cell proliferation in a cell-type and intensity-dependent way but the mechanism remains unclear. At the same time, although the diamagnetic anisotropy of proteins has been proposed decades ago, the behavior of isolated proteins in magnetic fields has not been directly observed. Here we show that SMFs can affect isolated proteins at the single molecular level in an intensity-dependent manner. We found that Epidermal Growth Factor Receptor (EGFR), a protein that is overexpressed and highly activated in multiple cancers, can be directly inhibited by SMFs. Using Liquid-phase Scanning Tunneling Microscopy (STM) to examine pure EGFR kinase domain proteins at the single molecule level in solution, we observed orientation changes of these proteins in response to SMFs. This may interrupt inter-molecular interactions between EGFR monomers, which are critical for their activation. In molecular dynamics (MD) simulations, 1-9T SMFs caused increased probability of EGFR in parallel with the magnetic field direction in an intensity-dependent manner. A superconducting ultrastrong 9T magnet reduced proliferation of CHO-EGFR cells (Chinese Hamster Ovary cells with EGFR overexpression) and EGFR-expressing cancer cell lines by ~35%, but minimally affected CHO cells. We predict that similar effects of magnetic fields can also be applied to some other proteins such as ion channels. Our paper will help clarify some dilemmas in this field and encourage further investigations in order to achieve a better understanding of the biological effects of SMFs. PMID:27223425
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Density-Dependent Regulation of Glioma Cell Proliferation and Invasion Mediated by miR-9.
Katakowski, Mark; Charteris, Nicholas; Chopp, Michael; Khain, Evgeniy
2016-12-01
The phenotypic axis of invasion and proliferation in malignant glioma cells is a well-documented phenomenon. Invasive glioma cells exhibit a decreased proliferation rate and a resistance to apoptosis, and invasive tumor cells dispersed in brain subsequently revert to proliferation and contribute to secondary tumor formation. One miRNA can affect dozens of mRNAs, and some miRNAs are potent oncogenes. Multiple miRNAs are implicated in glioma malignancy, and several of which have been identified to regulate tumor cell motility and division. Using rat 9 L gliosarcoma and human U87 glioblastoma cell lines, we investigated miRNAs associated with the switch between glioma cell invasion and proliferation. Using micro-dissection of 9 L glioma tumor xenografts in rat brain, we identified disparate expression of miR-9 between cells within the periphery of the primary tumor, and those comprising tumor islets within the invasive zone. Modifying miR-9 expression in in vitro assays, we report that miR-9 controls the axis of glioma cell invasion/proliferation, and that its contribution to invasion or proliferation is biphasic and dependent upon local tumor cell density. In addition, immunohistochemistry revealed elevated hypoxia inducible factor 1 alpha (HIF-1α) in the invasive zone as compared to the primary tumor periphery. We also found that hypoxia promotes miR-9 expression in glioma cells. Based upon these findings, we propose a hypothesis for the contribution of miR-9 to the dynamics glioma invasion and satellite tumor formation in brain adjacent to tumor.
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Bao, Yong; Fan, Jian-Zhong; Li, Ke; Li, Chuan; Yang, Jun-Feng
2008-06-01
To investigate the effect of infrasound therapy on the proliferation, apoptosis and ultrastructure of human B lymphoma Raji cells. Human B lymphoma Raji cells were exposed to infrasound treatment for 15, 30, 60, 90 and 120 min and cultured subsequently for 24 or 48 h. MTT assay, flow cytometry analysis, and electron microscopy were performed to examine the proliferative status, cell apoptosis and ultrastructural changes of the exposed cells, respectively. MTT assay revealed no significant changes in the proliferation of the cells exposed to infrasound treatment (P>0.05), nor did flow cytometry analysis identified significant variation in the cell apoptosis (P>0.05). Scanning electron microscopy, however, identified shortened or reduced cell processes and microvilli on the surface of the cells with infrasound exposure and a subsequent 24-hour culture, and the cell membrane surface became smooth. Under transmission electron microscope, the cells with infrasound treatment presented with significantly reduced microvilli, and the cell nuclei appeared homogeneous, with cytoplasmic budding and losses after a 48-hour culture. Infrasound less than 90 dB does not obviously affect the proliferation and apoptosis of Raji cells, but may directly cause cell ultrastructural changes such as reduction of the cell processes.
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Borland, Michael G.; Foreman, Jennifer E.; Girroir, Elizabeth E.; Zolfaghari, Reza; Sharma, Arun K.; Amin, Shantu; Gonzalez, Frank J.; Ross, A. Catharine; Peters, Jeffrey M.
2009-01-01
Although there is strong evidence that ligand activation of peroxisome proliferator-activated receptor (PPAR)-β/δ induces terminal differentiation and attenuates cell growth, some studies suggest that PPARβ/δ actually enhances cell proliferation. For example, it was suggested recently that retinoic acid (RA) is a ligand for PPARβ/δ and potentiates cell proliferation by activating PPARβ/δ. The present study examined the effect of ligand activation of PPARβ/δ on cell proliferation, cell cycle kinetics, and target gene expression in human HaCaT keratinocytes using two highly specific PPARβ/δ ligands [4-[[[2-[3-fluoro-4-(trifluoromethyl)phenyl]-4-methyl-5-thiazolyl]methyl]thio]-2-methylphenoxy acetic acid (GW0742) and 2-methyl-4-((4-methyl-2-(4-trifluoromethylphenyl)-1,3-thiazol-5-yl)-methylsulfanyl)phenoxy-acetic acid (GW501516)] and RA. Both PPARβ/δ ligands and RA inhibited cell proliferation of HaCaT keratinocytes. GW0742 and GW501516 increased expression of known PPARβ/δ target genes, whereas RA did not; RA increased the expression of known retinoic acid receptor/retinoid X receptor target genes, whereas GW0742 did not affect these genes. GW0742, GW501516, and RA did not modulate the expression of 3-phosphoinositide-dependent protein kinase or alter protein kinase B phosphorylation. GW0742 and RA increased annexin V staining as quantitatively determined by flow cytometry. The effects of GW0742 and RA were also examined in wild-type and PPARβ/δ-null primary mouse keratinocytes to determine the specific role of PPARβ/δ in modulating cell growth. Although inhibition of keratinocyte proliferation by GW0742 was PPARβ/δ-dependent, inhibition of cell proliferation by RA occurred in both genotypes. Results from these studies demonstrate that ligand activation of PPARβ/δ inhibits keratinocyte proliferation through PPARβ/δ-dependent mechanisms. In contrast, the observed inhibition of cell proliferation in mouse and human keratinocytes by RA is mediated by PPARβ/δ-independent mechanisms and is inconsistent with the notion that RA potentiates cell proliferation by activating PPARβ/δ. PMID:18687807
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Borland, Michael G; Foreman, Jennifer E; Girroir, Elizabeth E; Zolfaghari, Reza; Sharma, Arun K; Amin, Shantu; Gonzalez, Frank J; Ross, A Catharine; Peters, Jeffrey M
2008-11-01
Although there is strong evidence that ligand activation of peroxisome proliferator-activated receptor (PPAR)-beta/delta induces terminal differentiation and attenuates cell growth, some studies suggest that PPARbeta/delta actually enhances cell proliferation. For example, it was suggested recently that retinoic acid (RA) is a ligand for PPARbeta/delta and potentiates cell proliferation by activating PPARbeta/delta. The present study examined the effect of ligand activation of PPARbeta/delta on cell proliferation, cell cycle kinetics, and target gene expression in human HaCaT keratinocytes using two highly specific PPARbeta/delta ligands [4-[[[2-[3-fluoro-4-(trifluoromethyl)phenyl]-4-methyl-5-thiazolyl]methyl]thio]-2-methylphenoxy acetic acid (GW0742) and 2-methyl-4-((4-methyl-2-(4-trifluoromethylphenyl)-1,3-thiazol-5-yl)-methylsulfanyl)phenoxy-acetic acid (GW501516)] and RA. Both PPARbeta/delta ligands and RA inhibited cell proliferation of HaCaT keratinocytes. GW0742 and GW501516 increased expression of known PPARbeta/delta target genes, whereas RA did not; RA increased the expression of known retinoic acid receptor/retinoid X receptor target genes, whereas GW0742 did not affect these genes. GW0742, GW501516, and RA did not modulate the expression of 3-phosphoinositide-dependent protein kinase or alter protein kinase B phosphorylation. GW0742 and RA increased annexin V staining as quantitatively determined by flow cytometry. The effects of GW0742 and RA were also examined in wild-type and PPARbeta/delta-null primary mouse keratinocytes to determine the specific role of PPARbeta/delta in modulating cell growth. Although inhibition of keratinocyte proliferation by GW0742 was PPARbeta/delta-dependent, inhibition of cell proliferation by RA occurred in both genotypes. Results from these studies demonstrate that ligand activation of PPARbeta/delta inhibits keratinocyte proliferation through PPARbeta/delta-dependent mechanisms. In contrast, the observed inhibition of cell proliferation in mouse and human keratinocytes by RA is mediated by PPARbeta/delta-independent mechanisms and is inconsistent with the notion that RA potentiates cell proliferation by activating PPARbeta/delta.
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Nutritional Factors Affecting Adult Neurogenesis and Cognitive Function
USDA-ARS?s Scientific Manuscript database
Adult neurogenesis, a complex process by which stem cells in the hippocampal brain region differentiate and proliferate into new neurons and other resident brain cells, is known to be affected by many intrinsic and extrinsic factors, including diet. Neurogenesis plays a critical role in neural plas...
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Menin determines K-RAS proliferative outputs in endocrine cells
Chamberlain, Chester E.; Scheel, David W.; McGlynn, Kathleen; Kim, Hail; Miyatsuka, Takeshi; Wang, Juehu; Nguyen, Vinh; Zhao, Shuhong; Mavropoulos, Anastasia; Abraham, Aswin G.; O’Neill, Eric; Ku, Gregory M.; Cobb, Melanie H.; Martin, Gail R.; German, Michael S.
2014-01-01
Endocrine cell proliferation fluctuates dramatically in response to signals that communicate hormone demand. The genetic alterations that override these controls in endocrine tumors often are not associated with oncogenes common to other tumor types, suggesting that unique pathways govern endocrine proliferation. Within the pancreas, for example, activating mutations of the prototypical oncogene KRAS drive proliferation in all pancreatic ductal adenocarcimomas but are never found in pancreatic endocrine tumors. Therefore, we asked how cellular context impacts K-RAS signaling. We found that K-RAS paradoxically suppressed, rather than promoted, growth in pancreatic endocrine cells. Inhibition of proliferation by K-RAS depended on antiproliferative RAS effector RASSF1A and blockade of the RAS-activated proproliferative RAF/MAPK pathway by tumor suppressor menin. Consistent with this model, a glucagon-like peptide 1 (GLP1) agonist, which stimulates ERK1/2 phosphorylation, did not affect endocrine cell proliferation by itself, but synergistically enhanced proliferation when combined with a menin inhibitor. In contrast, inhibition of MAPK signaling created a synthetic lethal interaction in the setting of menin loss. These insights suggest potential strategies both for regenerating pancreatic β cells for people with diabetes and for targeting menin-sensitive endocrine tumors. PMID:25133424
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Proliferation of the human urothelium is induced by atypical β1 -adrenoceptors.
Winder, M; Wasén, C; Aronsson, P; Giglio, D
2015-09-01
We wanted to assess whether β-adrenoceptors mediate proliferation in the normal and malignant urothelial cell lines UROtsa and T24, respectively. Urothelial cells were cultured for 24 h in the presence of the β-adrenoceptor agonists isoprenaline (β1/2/3 ), dobutamine (β1 ), salbutamol (β2 ), BRL 37344 (β3 ), CGP 12177 (a partial β-agonist) or β-adrenoceptor antagonists (metoprolol; β1 , propranolol; β1/2 ). Phosphorylation of kinases was screened with a Human Phospho-Kinase Array Kit (R&D systems). Intracellular pathways activated by proliferation of urothelial cells were characterized by incubating cells with the MEK1/2 inhibitor PD 98,059, the p38 kinase inhibitor losmapimod or with the Akt 1/2 kinase inhibitor. Proliferation was assessed with the MTT proliferation assay (ATCC). Western blot and immunocytochemistry were used for detection of the β1 -adrenoceptor. Isoprenaline and dobutamine induced proliferation, while salbutamol and BRL 37344 did not. Dobutamine-induced proliferation was not affected by metoprolol or propranolol but was instead antagonized by CGP 12177 in T24 but not in UROtsa. In response to stimulation with dobutamine, Akt1/2/3 was phosphorylated in UROtsa, while ERK1/2 and p38 were phosphorylated in T24. MEK1/2 inhibition blocked basal and dobutamine-induced proliferation in T24 but only basal proliferation in UROtsa. Losmapimod slightly inhibited basal proliferation in T24 but not dobutamine-induced proliferation. Akt 1/2 inhibitor blocked basal and dobutamine-induced proliferation in UROtsa. Immunocytochemistry and Western blot revealed expression of β1 -adrenoceptors in both urothelial cell lines. The present data show that the urothelium expresses atypical β1-adrenoceptors that activate intracellular kinases inducing urothelial proliferation. © 2016 John Wiley & Sons Ltd.
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Triclosan and bisphenol a affect decidualization of human endometrial stromal cells.
Forte, Maurizio; Mita, Luigi; Cobellis, Luigi; Merafina, Verdiana; Specchio, Raffaella; Rossi, Sergio; Mita, Damiano Gustavo; Mosca, Lavinia; Castaldi, Maria Antonietta; De Falco, Maria; Laforgia, Vincenza; Crispi, Stefania
2016-02-15
In recent years, impaired fertility and endometrium related diseases are increased. Many evidences suggest that environmental pollution might be considered a risk factor for endometrial physiopathology. Among environmental pollutants, endocrine disrupting chemicals (EDCs) act on endocrine system, causing hormonal imbalance which, in turn, leads to female and male reproductive dysfunctions. In this work, we studied the effects of triclosan (TCL) and bisphenol A (BPA), two widespread EDCs, on human endometrial stromal cells (ESCs), derived from endometrial biopsies from woman not affected by endometriosis. Cell proliferation, cell cycle, migration and decidualization mechanisms were investigated. Treatments have been performed with both the EDCs separately or in presence and in absence of progesterone used as decidualization stimulus. Both TCL and BPA did not affect cell proliferation, but they arrested ESCs at G2/M phase of cell cycle enhancing cell migration. TCL and BPA also increased gene expression and protein levels of some decidualization markers, such as insulin growth factor binding protein 1 (IGFBP1) and prolactin (PRL), amplifying the effect of progesterone alone. All together, our data strongly suggest that TCL and BPA might alter human endometrium physiology so affecting fertility and pregnancy outcome. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.
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Degryse, B; Maisonobe, F; Hovsépian, S; Fayet, G
1991-11-01
Insulin and Insulin-like Growth Factor I (IGF-I) are evaluated for their capacity to affect cell proliferation and plasminogen activator (PA) activity production in an ovine thyroid cell line OVNIS. Insulin at physiological and supraphysiological doses induces cell proliferation and increases PA activity. IGF-I, which is also clearly mitogenic for these cells, surprisingly does not modulate PA activity. The results indicate that the growth promoting effect is mediated through the insulin and IGF-I receptors whereas PA activity is solely regulated via the insulin receptors.
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Cold thyroid nodules show a marked increase in proliferation markers.
Krohn, Knut; Stricker, Ingo; Emmrich, Peter; Paschke, Ralf
2003-06-01
Thyroid follicular adenomas and adenomatous thyroid nodules are a frequent finding in geographical areas with iodine deficiency. They occur as hypofunctioning (scintigraphically cold) or hyperfunctioning (scintigraphically hot) nodules. Their predominant clonal origin suggests that they result from clonal expansion of a single cell, which is very likely the result of a prolonged increase in proliferation compared with non-affected surrounding cells. To test whether increased cell proliferation is detectable in cold thyroid nodules, we studied paraffin-embedded tissue from 40 cold thyroid nodules and their surrounding normal thyroid tissue for the occurrence of the proliferating cell nuclear antigen (PCNA) and Ki-67 (MIB-1 antibody) epitopes as markers for cell proliferation. All 40 thyroid nodules were histologically well characterized and have been studied for molecular characteristics before. The labeling index (number of labeled cells versus total cell number) for nodular and surrounding tissue was calculated. In 33 cold thyroid nodules a significant (p < or = 0.05) increase in the labeling index for PCNA was detectable. In 19 cold thyroid nodules a significant (p < or = 0.05) increase in the labeling index for Ki-67 was detectable. Moreover, surrounding tissues with lymphocyte infiltration showed a significantly higher labeling index for both PCNA and Ki-67 compared with normal surrounding tissue. These findings are first evidence that an increased thyroid epithelial cell proliferation is a uniform feature common to most cold nodules. However, the increase of proliferation markers shows a heterogeneity that is not correlated with histopathologic, molecular, or clinical characteristics.
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Changes in Reactivity In Vitro of CD4+CD25+ and CD4+CD25− T Cell Subsets in Transplant Tolerance
Hall, Bruce M.; Robinson, Catherine M.; Plain, Karren M.; Verma, Nirupama D.; Tran, Giang T.; Nomura, Masaru; Carter, Nicole; Boyd, Rochelle; Hodgkinson, Suzanne J.
2017-01-01
Transplant tolerance induced in adult animals is mediated by alloantigen-specific CD4+CD25+ T cells, yet in many models, proliferation of CD4+ T cells from hosts tolerant to specific-alloantigen in vitro is not impaired. To identify changes that may diagnose tolerance, changes in the patterns of proliferation of CD4+, CD4+CD25+, and CD4+CD25− T cells from DA rats tolerant to Piebald Virol Glaxo rat strain (PVG) cardiac allografts and from naïve DA rats were examined. Proliferation of CD4+ T cells from both naïve and tolerant hosts was similar to both PVG and Lewis stimulator cells. In mixed lymphocyte culture to PVG, proliferation of naïve CD4+CD25− T cells was greater than naïve CD4+ T cells. In contrast, proliferation of CD4+CD25− T cells from tolerant hosts to specific-donor PVG was not greater than CD4+ T cells, whereas their response to Lewis and self-DA was greater than CD4+ T cells. Paradoxically, CD4+CD25+ T cells from tolerant hosts did not proliferate to PVG, but did to Lewis, whereas naïve CD4+CD25+ T cells proliferate to both PVG and Lewis but not to self-DA. CD4+CD25+ T cells from tolerant, but not naïve hosts, expressed receptors for interferon (IFN)-γ and IL-5 and these cytokines promoted their proliferation to specific-alloantigen PVG but not to Lewis or self-DA. We identified several differences in the patterns of proliferation to specific-donor alloantigen between cells from tolerant and naïve hosts. Most relevant is that CD4+CD25+ T cells from tolerant hosts failed to proliferate or suppress to specific donor in the absence of either IFN-γ or IL-5. The proliferation to third-party and self of each cell population from tolerant and naïve hosts was similar and not affected by IFN-γ or IL-5. Our findings suggest CD4+CD25+ T cells that mediate transplant tolerance depend on IFN−γ or IL-5 from alloactivated Th1 and Th2 cells. PMID:28878770
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Grison, Alice; Gaiser, Carine; Bieder, Andrea; Baranek, Constanze; Atanasoski, Suzana
2018-03-23
Little is known about the molecular players driving proliferation of neural progenitor cells (NPCs) during embryonic mouse development. Here, we demonstrate that proliferation of NPCs in the developing forebrain depends on a particular combination of cell cycle regulators. We have analyzed the requirements for members of the cyclin-dependent kinase (cdk) family using cdk-deficient mice. In the absence of either cdk4 or cdk6, which are both regulators of the G1 phase of the cell cycle, we found no significant effects on the proliferation rate of cortical progenitor cells. However, concomitant loss of cdk4 and cdk6 led to a drastic decrease in the proliferation rate of NPCs, specifically the basal progenitor cells of both the dorsal and ventral forebrain at embryonic day 13.5 (E13.5). Moreover, basal progenitors in the forebrain of Cdk4;Cdk6 double mutant mice exhibited altered cell cycle characteristics. Cdk4;cdk6 deficiency led to an increase in cell cycle length and cell cycle exit of mutant basal progenitor cells in comparison to controls. In contrast, concomitant ablation of cdk2 and cdk6 had no effect on the proliferation of NCPs. Together, our data demonstrate that the expansion of the basal progenitor pool in the developing telencephalon is dependent on the presence of distinct combinations of cdk molecules. Our results provide further evidence for differences in the regulation of proliferation between apical and basal progenitors during cortical development. © 2018 Wiley Periodicals, Inc. Develop Neurobiol, 2018. © 2018 Wiley Periodicals, Inc.
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Burke, Mark W; Inyatkin, Alexey; Ptito, Maurice; Ervin, Frank R; Palmour, Roberta M
2016-10-27
Fetal alcohol exposure (FAE) alters hippocampal cell numbers in rodents and primates, and this may be due, in part, to a reduction in the number or migration of neuronal progenitor cells. The olfactory bulb exhibits substantial postnatal cellular proliferation and a rapid turnover of newly formed cells in the rostral migratory pathway, while production and migration of postnatal neurons into the dentate gyrus may be more complex. The relatively small size of the olfactory bulb, compared to the hippocampus, potentially makes this structure ideal for a rapid analysis. This study used the St. Kitts vervet monkey ( Chlorocebus sabeus ) to (1) investigate the normal developmental sequence of post-natal proliferation in the olfactory bulb and dentate gyrus and (2) determine the effects of naturalistic prenatal ethanol exposure on proliferation at three different ages (neonate, five months and two years). Using design-based stereology, we found an age-related decrease of actively proliferating cells in the olfactory bulb and dentate gyrus for both control and FAE groups. Furthermore, at the neonatal time point, the FAE group had fewer actively proliferating cells as compared to the control group. These data are unique with respect to fetal ethanol effects on progenitor proliferation in the primate brain and suggest that the olfactory bulb may be a useful structure for studies of cellular proliferation.
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The isolation and in vitro expansion of hepatic Sca-1 progenitor cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Clayton, Elizabeth, E-mail: Elizabeth.Clayton@ed.ac.uk; Forbes, Stuart J.
2009-04-17
The intra-hepatic population of liver progenitor cells expands during liver injury when hepatocyte proliferation is inhibited. These cells can be purified by density gradient centrifugation and cultured. Separated by size only this population contains small cells of hematopoietic, epithelial and endothelial lineages and is thought to contain liver stem cells. The identity of liver stem cells remains unknown although there is some evidence that tissue Sca1{sup +} CD45{sup -} cells display progenitor cell characteristics. We identified both intra-hepatic and gall bladder Sca1{sup +} cells following liver injury and expanded ex vivo Sca1 cells as part of heterogenous cell culture ormore » as a purified population. We found significant difference between the proliferation of Sca-1 cells when plated on laminin or collagen I while proliferation of heterogenous population was not affected by the extracellular matrix indicating the necessity for culture of Sca1{sup +} cells with laminin matrix or laminin producing cells in long term liver progenitor cell cultures.« less
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Catherino, William H.; Malik, Minnie; Driggers, Paul; Chappel, Scott; Segars, James; Davis, Joseph
2012-01-01
Context Uterine leiomyomas are highly prevalent and often symptomatic. Current medical therapies are limited. A novel, potent, selective, orally active therapy is needed. Objective and Methods To determine the progesterone receptor (PR) specificity and activation, endometrial response, and impact on proliferation and extracellular matrix (ECM) production of the novel non-steroidal selective progesterone receptor modulators (SPRMs) CP8863 and CP8947 in human immortalized leiomyoma and patient-matched myometrial cells. Receptor binding in vitro was assessed using LNCaP, Ishikawa, T-47D, and HeLa cell extracts for AR, ER-α, PR, and GR, respectively. Progestational activity assessed by alkaline phosphatase assay in T47D cells and ER-α expression in human leiomyoma and myometrial cells. In vivo progestational activity assayed by the McPhail assay. Proliferation and gene expression studies (q RT-PCR and western blot) were performed in immortalized leiomyoma and myometrial cells. Results Both CP8863 and CP8947 is highly selective for PR but not for ER-α, AR, and GR. Both induced alkaline phosphatase comparably to progesterone, while CP8947 induced ER-α in leiomyoma cells but not myometrial cells. CP8947 was progestational in rabbit endometrium. Nanomolar CP8947 treatment inhibited human leiomyoma but not myometrial cell proliferation. The decreased proliferation correlated with increased TRAIL and caspase -7, suggesting induction of apoptosis in leiomyoma cells. ECM components were decreased in leiomyoma cells, including COL1A1 and COL7A1 at nanomolar concentrations. Conclusions CP8947 was a potent novel non-steroidal SPRM that was selective for PR, showed progestational activity in endometrium, inhibited leiomyoma cell proliferation (potentially via induction of apoptosis), and decreased ECM component production, without disrupting myometrial cell proliferation. PMID:20493256
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Hedgehog signaling plays roles in epithelial cell proliferation in neonatal mouse uterus and vagina.
Nakajima, Tadaaki; Iguchi, Taisen; Sato, Tomomi
2012-04-01
Both the uterus and vagina develop from the Müllerian duct but are quite distinct in morphology and function. To investigate factors controlling epithelial differentiation and cell proliferation in neonatal uterus and vagina, we focused on Hedgehog (HH) signaling. In neonatal mice, Sonic hh (Shh) was localized in the vaginal epithelium and Indian hh (Ihh) was slightly expressed in the uterus and vagina, whereas all Glioma-associated oncogene homolog (Gli) genes were mainly expressed in the stroma. The expression of target genes of HH signaling was high in the neonatal vagina and in the uterus, it increased with growth. Thus, in neonatal mice, Shh in the vaginal epithelium and Ihh in the uterus and vagina activated HH signaling in the stroma. Tissue recombinants showed that vaginal Shh expression was inhibited by the vaginal stroma and uterine Ihh expression was stimulated by the uterine stroma. Addition of a HH signaling inhibitor decreased epithelial cell proliferation in organ-cultured uterus and vagina and increased stromal cell proliferation in organ-cultured uterus. However, it did not affect epithelial differentiation or the expression of growth factors in organ-cultured uterus and vagina. Thus, activated HH signaling stimulates epithelial cell proliferation in neonatal uterus and vagina but inhibits stromal cell proliferation in neonatal uterus.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Xu, Jun; Department of Experimental Hematology, Beijing Institute of Radiation Medicine, Beijing 100850; Sun, Hui-Yan
2015-05-01
SUMO/sentrin specific protease 1 (Senp1) is an important regulation protease in the protein sumoylation, which affects the cell cycle, proliferation and differentiation. The role of Senp1 mediated protein desumoylation in pathophysiological progression of multiple myeloma is unknown. In this study, we demonstrated that Senp1 is overexpressed and induced by IL-6 in multiple myeloma cells. Lentivirus-mediated Senp1 knockdown triggers apoptosis and reduces viability, proliferation and colony forming ability of MM cells. The NF-κB family members including P65 and inhibitor protein IkBα play important roles in regulation of MM cell survival and proliferation. We further demonstrated that Senp1 inhibition decreased IL-6-induced P65more » and IkBα phosphorylation, leading to inactivation of NF-kB signaling in MM cells. These results delineate a key role for Senp1in IL-6 induced proliferation and survival of MM cells, suggesting it may be a potential new therapeutic target in MM. - Highlights: • Senp1 is overexpressed and induced by IL-6 in multiple myeloma cells. • Senp1 knockdown triggers apoptosis and reduces proliferation of MM cells. • Senp1 inhibition decreased IL-6-induced P65 and IkBα phosphorylation.« less
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Andersson, Olov, E-mail: olov.andersson@ki.se
Glucose homeostasis, which is controlled by the endocrine cells of the pancreas, is disrupted in both type I and type II diabetes. Deficiency in the number of insulin-producing β cells – a primary cause of type I diabetes and a secondary contributor of type II diabetes – leads to hyperglycemia and hence an increase in the need for insulin. Although diabetes can be controlled with insulin injections, a curative approach is needed. A potential approach to curing diabetes involves regenerating the β-cell mass, e.g. by increasing β-cell proliferation, survival, neogenesis or transdifferentiation. The nucleoside adenosine and its cognate nucleotide ATPmore » have long been known to affect insulin secretion, but have more recently been shown to increase β-cell proliferation during homeostatic control and regeneration of the β-cell mass. Adenosine is also known to have anti-inflammatory properties, and agonism of adenosine receptors can promote the survival of β-cells in an inflammatory microenvironment. In this review, both intracellular and extracellular mechanisms of adenosine and ATP are discussed in terms of their established and putative effects on β-cell regeneration. - Highlights: • A potential way to cure diabetes is to regenerate the β-cell mass by promoting cell survival, proliferation or neogenesis. • Adenosine may promote β-cell regeneration through several cellular mechanisms. • Adenosine and its cognate nucleotide ATP can each promote β-cell proliferation. • Do adenosine and ATP interact in promoting β-cell proliferation?.« less
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Mukherjee, Abir; Ma, Yibao; Yuan, Fang; Gong, Yongling; Fang, Zhenyu; Mohamed, Esraa M.; Berrios, Erika; Shao, Huanjie; Fang, Xianjun
2015-01-01
Lysophosphatidic acid (LPA), a blood-borne lipid mediator, is present in elevated concentrations in ascites of ovarian cancer patients and other malignant effusions. LPA is a potent mitogen in cancer cells. The mechanism linking LPA signal to cancer cell proliferation is not well understood. Little is known about whether LPA affects glucose metabolism to accommodate rapid proliferation of cancer cells. Here we describe that in ovarian cancer cells, LPA enhances glycolytic rate and lactate efflux. A real time PCR-based miniarray showed that hexokinase II (HK2) was the most dramatically induced glycolytic gene to promote glycolysis in LPA-treated cells. Analysis of the human HK2 gene promoter identified the sterol regulatory element-binding protein as the primary mediator of LPA-induced HK2 transcription. The effects of LPA on HK2 and glycolysis rely on LPA2, an LPA receptor subtype overexpressed in ovarian cancer and many other malignancies. We further examined the general role of growth factor-induced glycolysis in cell proliferation. Like LPA, epidermal growth factor (EGF) elicited robust glycolytic and proliferative responses in ovarian cancer cells. Insulin-like growth factor 1 (IGF-1) and insulin, however, potently stimulated cell proliferation but only modestly induced glycolysis. Consistent with their differential effects on glycolysis, LPA and EGF-dependent cell proliferation was highly sensitive to glycolytic inhibition while the growth-promoting effect of IGF-1 or insulin was more resistant. These results indicate that LPA- and EGF-induced cell proliferation selectively involves up-regulation of HK2 and glycolytic metabolism. The work is the first to implicate LPA signaling in promotion of glucose metabolism in cancer cells. PMID:26476080
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Research progress on the proliferation and differentiation of
NASA Astrophysics Data System (ADS)
An, A.; Tan, B.
Space environments such as microgravity magnetic field radiation and heavy metal ions affects the development and functions of human and mammalian cells To study these influences and the corresponding metabolisms is in favour of knowing about the development and differentiation process of organism cells In recent years researches on the differentiation of stem cells induced in vitro provide a new pathway for the repair of tissue lesion and therapy of human diseases Stem cells are potential in capable of differentiating into different functional cells But there has no reliable methods to induce the stem cells differentiating forward specific cells and to gain enough cells for transplantation which limited their application on clinical therapy It has been indicated that microgravity influenced embryonic development hematopoietic and mesenchymal stem cells and so on Hematopoietic stem cell migration and its differentiation were affected by microgravity The specific differentiation of hematopoietic stem cells was inhibited under microgravity The expression of proteins regulating cell cycle period also changed Mesenchymal stem cells provide a source of cells for the repair of musculoskeletal tissue in ground experiment While under microgravity the proliferation and differentiation of mesenchymal stem cells were influenced along with the differentiated cells function changed Furthermore in the differentiation process of stem cells under microgravity the mechanism of signal transport was also affected and the specific differentiation
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Formulation Changes Affect Material Properties and Cell Behavior in HA-Based Hydrogels.
Lawyer, Thomas; McIntosh, Kristen; Clavijo, Cristian; Potekhina, Lydia; Mann, Brenda K
2012-01-01
To develop and optimize new scaffold materials for tissue engineering applications, it is important to understand how changes to the scaffold affect the cells that will interact with that scaffold. In this study, we used a hyaluronic acid- (HA-) based hydrogel as a synthetic extracellular matrix, containing modified HA (CMHA-S), modified gelatin (Gtn-S), and a crosslinker (PEGda). By varying the concentrations of these components, we were able to change the gelation time, enzymatic degradation, and compressive modulus of the hydrogel. These changes also affected fibroblast spreading within the hydrogels and differentially affected the proliferation and metabolic activity of fibroblasts and mesenchymal stem cells (MSCs). In particular, PEGda concentration had the greatest influence on gelation time, compressive modulus, and cell spreading. MSCs appeared to require a longer period of adjustment to the new microenvironment of the hydrogels than fibroblasts. Fibroblasts were able to proliferate in all formulations over the course of two weeks, but MSCs did not. Metabolic activity changed for each cell type during the two weeks depending on the formulation. These results highlight the importance of determining the effect of matrix composition changes on a particular cell type of interest in order to optimize the formulation for a given application.
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Development of bioengineering system for stem cell proliferation
NASA Astrophysics Data System (ADS)
Park, H. S.; Shah, R.; Shah, C.
2016-08-01
From last decades, intensive research in the field of stem cells proliferation had been promoted due to the unique property of stem cells to self-renew themselves into multiples and has potential to replicate into an organ or tissues and so it's highly demanding though challenging. Bioreactor, a mechanical device, works as a womb for stem cell proliferation by providing nutritious environment for the proper growth of stem cells. Various factors affecting stem cells growth are the bioreactor mechanism, feeding of continuous nutrients, healthy environment, etc., but it always remains a challenge for controlling biological parameters. The present paper unveils the design of mechanical device commonly known as bioreactor in tissues engineering and biotech field, use for proliferation of stem cells and imparts the proper growing condition for stem cells. This high functional bioreactor provides automation mixing of cell culture and stem cells. This design operates in conjunction with mechanism of reciprocating motion. Compare to commercial bioreactors, this proposed design is more convenient, easy to operate and less maintenance is required as bioreactor culture bag is made of polyethylene which is single use purpose. Development of this bioengineering system will be beneficial for better growth and expansion of stem cell
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Rauner, Gat, E-mail: gat.rauner@mail.huji.ac.il; The Robert H. Smith Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem; Barash, Itamar, E-mail: itamar.barash@mail.huji.ac.il
The challenge in manipulating the proportion of somatic stem cells lies in having to override tissue homeostasis. Xanthosine infusion via the teat canal has been reported to augment the number of label-retaining cells in the mammary gland of 3-month-old bovine calves. To further delineate xanthosine's effect on defined stem cells in the mammary gland of heifers—which are candidates for increased prospective milk production following such manipulation—bovine mammary parenchymal tissue was transplanted and integrated into the cleared mammary fat pad of immunodeficient mice. Xanthosine administration for 14 days did not affect the number of label-retaining cells after 10- and 11-week chases.more » No change in stem cell proportion, analyzed according to CD49f and CD24 expression, was noted. Clone formation and propagation rate of cultured cells, as well as expression of stem cell markers, were also unaffected. In contrast, a latent 50% decrease in bovine mammary cell proliferation rate was observed 11 weeks after xanthosine administration. Tumor development in mice was also limited by xanthosine administration. These effects may have resulted from an initial decrease in expression of the rate-limiting enzyme in guanine synthesis, IMPDH. The data indicate that caution should be exerted when considering xanthosine for stem cell manipulation. - Highlights: • Novel “bovinized“ mouse model for exogenous effects on bovine mammary gland. • Xanthosine did not affect stem cell number/function in bovine mammary gland. • Xanthosine caused an immediate decrease in IMPDH expression in bovine mammary gland. • Xanthosine had latent negative effect on cell proliferation in bovine mammary gland. • Xanthosine administration limited mammary tumor growth.« less
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Effects of electrical stimulation on cell proliferation and apoptosis.
Love, Maria R; Palee, Siripong; Chattipakorn, Siriporn C; Chattipakorn, Nipon
2018-03-01
The application of exogenous electrical stimulation (ES) to cells in order to manipulate cell apoptosis and proliferation has been widely investigated as a possible method of treatment in a number of diseases. Alteration of the transmembrane potential of cells via ES can affect various intracellular signaling pathways which are involved in the regulation of cellular function. Controversially, several types of ES have proved to be effective in both inhibiting or inducing apoptosis, as well as increasing proliferation. However, the mechanisms through which ES achieves this remain fairly unclear. The aim of this review was to comprehensively summarize current findings from in vitro and in vivo studies on the effects of different types of ES on cell apoptosis and proliferation, highlighting the possible mechanisms through which ES induced these effects and define the optimum parameters at which ES can be used. Through this we hope to provide a greater insight into how future studies can most effectively use ES at the clinical trial stage. © 2017 Wiley Periodicals, Inc.
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Sylman, Joanna L.; Ngo, Anh T. P.; Pang, Jiaqing; Sears, Rosalie C.; Williams, Craig D.; McCarty, Owen J. T.
2017-01-01
Aspirin, an anti-inflammatory and antithrombotic drug, has become the focus of intense research as a potential anticancer agent owing to its ability to reduce tumor proliferation in vitro and to prevent tumorigenesis in patients. Studies have found an anticancer effect of aspirin when used in low, antiplatelet doses. However, the mechanisms through which low-dose aspirin works are poorly understood. In this study, we aimed to determine the effect of aspirin on the cross talk between platelets and cancer cells. For our study, we used two colon cancer cell lines isolated from the same donor but characterized by different metastatic potential, SW480 (nonmetastatic) and SW620 (metastatic) cancer cells, and a pancreatic cancer cell line, PANC-1 (nonmetastatic). We found that SW480 and PANC-1 cancer cell proliferation was potentiated by human platelets in a manner dependent on the upregulation and activation of the oncoprotein c-MYC. The ability of platelets to upregulate c-MYC and cancer cell proliferation was reversed by an antiplatelet concentration of aspirin. In conclusion, we show for the first time that inhibition of platelets by aspirin can affect their ability to induce cancer cell proliferation through the modulation of the c-MYC oncoprotein. PMID:27903583
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Sierralta, Walter D; Epuñan, María J; Reyes, José M; Valladares, Luis E; Pino, Ana M
2008-01-01
A stable cyclized 9-mer peptide (cP) containing the active site of alpha-alpha fetoprotein (alphaFP) has been shown to be effective for prevention of estrogen-stimulated tumor cell proliferation in culture or of xenographt growth in immunodeficient mice. cP does not block 17beta-estradiol (E2) binding to its receptors, but rather appears to interfere with intracellular processing of the signal that supports growth. To obtain insight on that mechanism we studied the effect of cP on the proliferation of MCF-7 cells in culture. Proliferation in the presence of 2 microM E2 is decreased up to 40% upon addition of 2 microg ml(-1) cP to the medium; the presence of cP did not increase cell death, cP reduced also the proliferation of estrogen-dependent ZR75-1 cells but had no effect on autonomous MDA-MB-231 cells, cP did not modify the number of binding sites for labeled E2 or affected cell death. We detected increased nuclear p21Cip1 immunoreactivity after cP treatment. Our results suggest that cP acts via p21Cip1 to slow the process of MCF-7 cells through the cycle.
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NASA Astrophysics Data System (ADS)
Capri, Miriam; Mesirca, Pietro; Remondini, Daniel; Carosella, Simona; Pasi, Sara; Castellani, Gastone; Franceschi, Claudio; Bersani, Ferdinando
2004-12-01
In the last 30 years, an increasing public concern about the possible harmful effects of electromagnetic fields generated by power lines and domestic appliances has pushed the scientific community to search for a correct and comprehensive answer to this problem. In this work the effects of exposure to 50 Hz sinusoidal magnetic fields, with a magnetic flux density of 0.05 mT and 2.5 mT (peak values), were studied on human peripheral blood mononuclear cells (PBMCs) collected from healthy young and elderly donors. Cell activation and proliferation were investigated by using flow cytometry techniques and 3H-TdR incorporation assays, respectively. The results obtained indicated that exposure to the fields altered neither DNA synthesis nor the capacity of lymphocytes to enter the activation phase and progress into the cell cycle. Thus, the conclusions are that two important functional phases of human lymphocytes, such as activation and proliferation, are not affected by exposures to 50 Hz magnetic fields similar to those found under power lines.
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Dai, Jie-Min; Yu, Mu-Xue; Shen, Zhen-Yu; Guo, Chu-Yi; Zhuang, Si-Qi; Qiu, Xiao-Shan
2015-01-01
Signaling through the mammalian target of rapamycin (mTOR) in response to leucine modulates many cellular and developmental processes. However, in the context of satellite cell proliferation and differentiation, the role of leucine and mTORC1 is less known. This study investigates the role of leucine in the process of proliferation and differentiation of primary preterm rat satellite cells, and the relationship with mammalian target of rapamycin complex 1 (mTORC1) activation. Dissociation of primary satellite cells occurred with type I collagenase and trypsin, and purification, via different speed adherence methods. Satellite cells with positive expression of Desmin were treated with leucine and rapamycin. We observed that leucine promoted proliferation and differentiation of primary satellite cells and increased the phosphorylation of mTOR. Rapamycin inhibited proliferation and differentiation, as well as decreased the phosphorylation level of mTOR. Furthermore, leucine increased the expression of MyoD and myogenin while the protein level of MyoD decreased due to rapamycin. However, myogenin expressed no affect by rapamycin. In conclusion, leucine may up-regulate the activation of mTORC1 to promote proliferation and differentiation of primary preterm rat satellite cells. We have shown that leucine promoted the differentiation of myotubes in part through the mTORC1-MyoD signal pathway. PMID:26007333
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Zhang, Wenyao; Li, Xuezhong; Xu, Tong; Ma, Mengru; Zhang, Yong; Gao, Ming-Qing
2016-11-15
Hypernomic secretion of epithelial cytokines has several effects on stromal cells. The contributions of inflammatory epithelial cells to stromal fibroblasts in bovine mammary glands with mastitis remain poorly understood. Here, we established an inflammatory epithelial cell model of bovine mastitis with gram-negative lipopolysaccharide (LPS) and gram-positive lipoteichoic acid (LTA) bacterial cell wall components. We characterized immune responses of mammary stromal fibroblasts induced by inflammatory epithelial cells. Our results showed that inflammatory epithelial cells affected stromal fibroblast characteristics by increasing inflammatory mediator expression, elevating extracellular matrix protein deposition, decreasing proliferation capacity, and enhancing migration ability. The changes in stromal fibroblast proliferation and migration abilities were mediated by signal molecules, such as WNT signal pathway components. LPS- and LTA-induced inflammatory epithelial cells triggered different immune responses in stromal fibroblasts. Thus, in mastitis, bovine mammary gland stromal fibroblasts were affected by inflammatory epithelial cells and displayed inflammation-specific changes, suggesting that fibroblasts play crucial roles in bovine mastitis. Copyright © 2016 Elsevier Inc. All rights reserved.
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Xu, Lin; Xu, Qian; Li, Xiwen; Zhang, Xiaoling
2017-10-01
The proliferation and apoptosis of tumor cells are regulated by a variety of microRNAs (miRs). miR‑21 can inhibit the apoptosis of cancer cells in vitro. Tumor necrosis factor α (TNF‑α) serves an important role in the induction of proliferation of cervical cancer cells. Previous studies have demonstrated that the expression level of miR‑21 is associated with TNF‑α expression in alveolar macrophages. However, to the best of our knowledge, whether miR‑21 regulates TNF‑α in cervical cells has not been reported. The present study was designed to investigate whether miR‑21 regulates TNF‑α expression, proliferation and apoptosis of cervical cancer cells. miR‑21, miR‑21 inhibitor and control miRNA were synthesized and transfected into HeLa cervical cancer cells. Reverse transcription‑quantitative polymerase chain reaction was used to measure the expression levels of miR‑21 and TNF‑α at the mRNA level. Western blotting was used to measure the expression levels of TNF‑α at the protein level. MTT assay and Hoechest‑33342 staining were used to measure the proliferation and apoptosis of HeLa cells. miR‑21 was identified to upregulate the mRNA and protein expression levels of TNF‑α. Furthermore, upregulation of TNF‑α enhanced the proliferation capability of HeLa cells. Changes in the expression levels of miR‑21 and TNF‑α did not significantly affect the apoptosis of Hela cells. In conclusion, the present study demonstrated that miR‑21 regulates the expression of TNF‑α in HeLa cells. Additionally, the expression level of TNF‑α was positively associated with the proliferation capability of Hela cells, but not apoptosis. Therefore, miR‑21 regulates the proliferation of HeLa cells through regulation of TNF‑α. These results provide novel potential therapeutic targets for the treatment of cervical cancer.
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Popławski, Piotr; Wiśniewski, Jacek R; Rijntjes, Eddy; Richards, Keith; Rybicka, Beata; Köhrle, Josef; Piekiełko-Witkowska, Agnieszka
2017-01-01
Type 1 iodothyronine deiodinase (DIO1) contributes to deiodination of 3,5,3',5'-tetraiodo-L-thyronine (thyroxine, T4) yielding of 3,5,3'-triiodothyronine (T3), a powerful regulator of cell differentiation, proliferation, and metabolism. Our previous work showed that loss of DIO1 enhances proliferation and migration of renal cancer cells. However, the global effects of DIO1 expression in various tissues affected by cancer remain unknown. Here, the effects of stable DIO1 re-expression were analyzed on the proteome of renal cancer cells, followed by quantitative real-time PCR validation in two renal cancer-derived cell lines. DIO1-induced changes in intracellular concentrations of thyroid hormones were quantified by L-MS/MS and correlations between expression of DIO1 and potential target genes were determined in tissue samples from renal cancer patients. Stable re-expression of DIO1, resulted in 26 downregulated proteins while 59 proteins were overexpressed in renal cancer cells. The 'downregulated' group consisted mainly of oncoproteins (e.g. STAT3, ANPEP, TGFBI, TGM2) that promote proliferation, migration and invasion. Furthermore, DIO1 re-expression enhanced concentrations of two subunits of thyroid hormone transporter (SLC7A5, SLC3A2), enzymes of key pathways of cellular energy metabolism (e.g. TKT, NAMPT, IDH2), sex steroid metabolism and anti-oxidative response (AKR1C2, AKR1B10). DIO1 expression resulted in elevated intracellular concentration of T4. Expression of DIO1-affected genes strongly correlated with DIO1 transcript levels in tissue samples from renal cancer patients as well as with their poor survival. This first study addressing effects of deiodinase re-expression on proteome of cancer cells demonstrates that induced DIO1 re-expression in renal cancer robustly downregulates oncoproteins, affects key metabolic pathways, and triggers proteins involved in anti-oxidative protection. This data supports the notion that suppressed DIO1 expression and changes in local availability of thyroid hormones might favor a shift from a differentiated to a more proliferation-prone state of cancer tissues and cell lines.
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Rijntjes, Eddy; Richards, Keith; Rybicka, Beata; Köhrle, Josef
2017-01-01
Type 1 iodothyronine deiodinase (DIO1) contributes to deiodination of 3,5,3’,5’-tetraiodo-L-thyronine (thyroxine, T4) yielding of 3,5,3’-triiodothyronine (T3), a powerful regulator of cell differentiation, proliferation, and metabolism. Our previous work showed that loss of DIO1 enhances proliferation and migration of renal cancer cells. However, the global effects of DIO1 expression in various tissues affected by cancer remain unknown. Here, the effects of stable DIO1 re-expression were analyzed on the proteome of renal cancer cells, followed by quantitative real-time PCR validation in two renal cancer-derived cell lines. DIO1-induced changes in intracellular concentrations of thyroid hormones were quantified by L-MS/MS and correlations between expression of DIO1 and potential target genes were determined in tissue samples from renal cancer patients. Stable re-expression of DIO1, resulted in 26 downregulated proteins while 59 proteins were overexpressed in renal cancer cells. The ‘downregulated’ group consisted mainly of oncoproteins (e.g. STAT3, ANPEP, TGFBI, TGM2) that promote proliferation, migration and invasion. Furthermore, DIO1 re-expression enhanced concentrations of two subunits of thyroid hormone transporter (SLC7A5, SLC3A2), enzymes of key pathways of cellular energy metabolism (e.g. TKT, NAMPT, IDH2), sex steroid metabolism and anti-oxidative response (AKR1C2, AKR1B10). DIO1 expression resulted in elevated intracellular concentration of T4. Expression of DIO1-affected genes strongly correlated with DIO1 transcript levels in tissue samples from renal cancer patients as well as with their poor survival. This first study addressing effects of deiodinase re-expression on proteome of cancer cells demonstrates that induced DIO1 re-expression in renal cancer robustly downregulates oncoproteins, affects key metabolic pathways, and triggers proteins involved in anti-oxidative protection. This data supports the notion that suppressed DIO1 expression and changes in local availability of thyroid hormones might favor a shift from a differentiated to a more proliferation-prone state of cancer tissues and cell lines. PMID:29272308
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Kato, Hirohito; Taguchi, Yoichiro; Tominaga, Kazuya; Kimura, Daisuke; Yamawaki, Isao; Noguchi, Masahiro; Yamauchi, Nobuhiro; Tamura, Isao; Tanaka, Akio; Umeda, Makoto
2016-04-01
Diabetes mellitus (DM) is a major risk factor for periodontal disease and affects various cellular functions. Periodontal ligament stem cells (PDLSCs) play an important role in periodontal tissue regeneration; however, the effect of hyperglycemia on PDLSCs is unclear. The aim of this study is to investigate whether hyperglycemia affects periodontal tissue regeneration, using human PDLSCs and high-glucose medium as a model of DM. PDLSCs were obtained from healthy adult human mandibular third molars. Cell proliferation, osteoblastic differentiation, and proinflammatory cytokine expression were investigated by culturing PDLSCs in media supplemented with four different glucose concentrations representative of control patients (5.5 mM), patients with postprandial or controlled DM (8.0 mM), and patients with uncontrolled DM (12.0 and 24.0 mM). The molecular effects of hyperglycemia on PDLSC physiology were examined with a focus on the nuclear factor (NF)-(κB signaling pathway. The involvement of NF-κB was investigated with a specific NF-κB inhibitor in PDLSCs under hyperglycemic conditions. High glucose levels inhibited PDLSC proliferation and differentiation into osteoblasts but induced NF-κB activation and subsequent interleukin (IL)-6 and IL-8 expression. Treatment with an NF-κB inhibitor rescued the defects in cell proliferation and osteoblastic differentiation and inhibited the IL-6 expression caused by the high-glucose environment. The results of this study demonstrate that hyperglycemia inhibits human PDLSC proliferation and osteoblastic differentiation.
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Ghrelin promotes oral tumor cell proliferation by modifying GLUT1 expression.
Kraus, Dominik; Reckenbeil, Jan; Wenghoefer, Matthias; Stark, Helmut; Frentzen, Matthias; Allam, Jean-Pierre; Novak, Natalija; Frede, Stilla; Götz, Werner; Probstmeier, Rainer; Meyer, Rainer; Winter, Jochen
2016-03-01
In our study, ghrelin was investigated with respect to its capacity on proliferative effects and molecular correlations on oral tumor cells. The presence of all molecular components of the ghrelin system, i.e., ghrelin and its receptors, was analyzed and could be detected using real-time PCR and immunohistochemistry. To examine cellular effects caused by ghrelin and to clarify downstream-regulatory mechanisms, two different oral tumor cell lines (BHY and HN) were used in cell culture experiments. Stimulation of either cell line with ghrelin led to a significantly increased proliferation. Signal transduction occurred through phosphorylation of GSK-3β and nuclear translocation of β-catenin. This effect could be inhibited by blocking protein kinase A. Glucose transporter1 (GLUT1), as an important factor for delivering sufficient amounts of glucose to tumor cells having high requirements for this carbohydrate (Warburg effect) was up-regulated by exogenous and endogenous ghrelin. Silencing intracellular ghrelin concentrations using siRNA led to a significant decreased expression of GLUT1 and proliferation. In conclusion, our study describes the role for the appetite-stimulating peptide hormone ghrelin in oral cancer proliferation under the particular aspect of glucose uptake: (1) tumor cells are a source of ghrelin. (2) Ghrelin affects tumor cell proliferation through autocrine and/or paracrine activity. (3) Ghrelin modulates GLUT1 expression and thus indirectly enhances tumor cell proliferation. These findings are of major relevance, because glucose uptake is assumed to be a promising target for cancer treatment.
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Merluzzi, Sonia; Frossi, Barbara; Gri, Giorgia; Parusso, Serena; Tripodo, Claudio; Pucillo, Carlo
2010-04-08
The evidence of a tight spatial interaction between mast cells (MCs) and B lymphocytes in secondary lymphoid organs, along with the data regarding the abundance of MCs in several B-cell lymphoproliferative disorders prompted us to investigate whether MCs could affect the proliferation and differentiation of B cells. To this aim, we performed coculture assays using mouse splenic B cells and bone marrow-derived MCs. Both nonsensitized and activated MCs proved able to induce a significant inhibition of cell death and an increase in proliferation of naive B cells. Such proliferation was further enhanced in activated B cells. This effect relied on cell-cell contact and MC-derived interleukin-6 (IL-6). Activated MCs could regulate CD40 surface expression on unstimulated B cells and the interaction between CD40 with CD40 ligand (CD40L) on MCs, together with MC-derived cytokines, was involved in the differentiation of B cells into CD138(+) plasma cells and in selective immunoglobulin A (IgA) secretion. These data were corroborated by in vivo evidence of infiltrating MCs in close contact with IgA-expressing plasma cells within inflamed tissues. In conclusion, we reported here a novel role for MCs in sustaining B-cell expansion and driving the development of IgA-oriented humoral immune responses.
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Expression and function of activin receptors in human endometrial adenocarcinoma cells.
Tanaka, Tetsuji; Toujima, Saori; Otani, Tsutomu; Minami, Sawako; Yamoto, Mareo; Umesaki, Naohiko
2003-09-01
Menstrual cycle-dependent expressions of activin A in normal human endometrial tissues have been reported. Expression of activin receptor mRNAs and increased activin A production were also observed in human endometrial adenocarcinoma tissues, suggesting that activin A might enhance cell proliferation and inhibit apoptotic signaling in endometrial cancer cells. In this study, we have examined the effects of activin A on cell proliferation, anticancer drug-induced apoptosis and Fas-mediated apoptosis in 3 differentiated human endometrial adenocarcinoma cell lines, namely HEC-1, HHUA and Ishikawa. Flow cytometric analyses revealed moderate expressions of all 4 types of activin receptor subunits on the cell surfaces of the 3 cell lines. The proliferations of the 3 endometrial cancer cells were completely unaffected by activin A, whereas it suppressed the cell proliferation of a human ovarian endometrioid adenocarcinoma cell line, OVK-18, in a dose-dependent manner. Moreover, activin A did not affect the apoptotic changes in the 3 endometrial adenocarcinoma cells treated with 4 different anticancer drugs, namely CDDP, paclitaxel, etoposide and SN38. The apoptotic changes in HHUA cells treated with anti-Fas IgM were also unaffected by activin A. These results indicate that the increased activin A production in human endometrial adenocarcinoma tissues in vivo may not stimulate carcinoma cell proliferation or inhibit apoptotic signaling in carcinoma cells. Insensitivity to the usual growth suppression signals induced by activin A might be one of the mechanisms of immortality of human endometrial adenocarcinoma cells.
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Effect of Porosity of Alumina and Zirconia Ceramics toward Pre-Osteoblast Response
Hadjicharalambous, Chrystalleni; Prymak, Oleg; Loza, Kateryna; Buyakov, Ales; Kulkov, Sergei; Chatzinikolaidou, Maria
2015-01-01
It is acknowledged that cellular responses are highly affected by biomaterial porosity. The investigation of this effect is important for the development of implanted biomaterials that integrate with bone tissue. Zirconia and alumina ceramics exhibit outstanding mechanical properties and are among the most popular implant materials used in orthopedics, but few data exist regarding the effect of porosity on cellular responses to these materials. The present study investigates the effect of porosity on the attachment and proliferation of pre-osteoblastic cells on zirconia and alumina. For each composition, ceramics of three different porosities are fabricated by sintering, and characterized using scanning electron microscopy, energy dispersive X-ray spectroscopy and X-ray powder diffraction. Cell proliferation is quantified, and microscopy is employed to qualitatively support the proliferation results and evaluate cell morphology. Cell adhesion and metabolic activity are found comparable among low porosity zirconia and alumina. In contrast, higher porosity favors better cell spreading on zirconia and improves growth, but does not significantly affect cell response on alumina. Between the highest porosity materials, cell response on zirconia is found superior to alumina. Results show that an average pore size of ~150 μm and ~50% porosity can be considered beneficial to cellular growth on zirconia ceramics. PMID:26579516
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Calcitonin gene-related peptide stimulates proliferation of human endothelial cells.
Haegerstrand, A; Dalsgaard, C J; Jonzon, B; Larsson, O; Nilsson, J
1990-01-01
The effects of the vasoactive perivascular neuropeptides calcitonin gene-related peptide (CGRP), neurokinin A (NKA), neuropeptide Y (NPY), and vasoactive intestinal polypeptide (VIP) on proliferation of cultured human umbilical vein endothelial cells (HUVECs) were investigated. CGRP was shown to increase both cell number and DNA synthesis, whereas NKA, NPY, and VIP were ineffective. 125I-labeled CGRP was shown to bind to HUVECs and this binding was displaced by addition of unlabeled CGRP, suggesting the existence of specific CGRP receptors. The effect of CGRP on formation of adenosine 3',5'-cyclic monophosphate (cAMP) and inositol phosphates (InsP), two intracellular messengers known to be involved in regulation of cell proliferation, was investigated. CGRP stimulated cAMP formation but was without effect on the formation of InsP. Proliferation, as well as cAMP formation, was also stimulated by cholera toxin. Basic fibroblast growth factor stimulated growth without affecting cAMP or InsP formation, whereas thrombin, which increased InsP formation, did not stimulate proliferation. We thus suggest that CGRP may act as a local factor stimulating proliferation of endothelial cells; that the mechanism of action is associated with cAMP formation; and that this effect of CGRP may be important for formation of new vessels during physiological and pathophysiological events such as ischemia, inflammation, and wound healing. PMID:2159144
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Calcitonin gene-related peptide stimulates proliferation of human endothelial cells.
Haegerstrand, A; Dalsgaard, C J; Jonzon, B; Larsson, O; Nilsson, J
1990-05-01
The effects of the vasoactive perivascular neuropeptides calcitonin gene-related peptide (CGRP), neurokinin A (NKA), neuropeptide Y (NPY), and vasoactive intestinal polypeptide (VIP) on proliferation of cultured human umbilical vein endothelial cells (HUVECs) were investigated. CGRP was shown to increase both cell number and DNA synthesis, whereas NKA, NPY, and VIP were ineffective. 125I-labeled CGRP was shown to bind to HUVECs and this binding was displaced by addition of unlabeled CGRP, suggesting the existence of specific CGRP receptors. The effect of CGRP on formation of adenosine 3',5'-cyclic monophosphate (cAMP) and inositol phosphates (InsP), two intracellular messengers known to be involved in regulation of cell proliferation, was investigated. CGRP stimulated cAMP formation but was without effect on the formation of InsP. Proliferation, as well as cAMP formation, was also stimulated by cholera toxin. Basic fibroblast growth factor stimulated growth without affecting cAMP or InsP formation, whereas thrombin, which increased InsP formation, did not stimulate proliferation. We thus suggest that CGRP may act as a local factor stimulating proliferation of endothelial cells; that the mechanism of action is associated with cAMP formation; and that this effect of CGRP may be important for formation of new vessels during physiological and pathophysiological events such as ischemia, inflammation, and wound healing.
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Samaddar, Sreyashi; Vazquez, Kizzy; Ponkia, Dipen; Toruno, Pedro; Sahbani, Karim; Begum, Sultana; Abouelela, Ahmed; Mekhael, Wagdy; Ahmed, Zaghloul
2017-02-01
Direct current electrical fields have been shown to be a major factor in the regulation of cell proliferation, differentiation, migration, and survival, as well as in the maturation of dividing cells during development. During adulthood, spinal cord cells are continuously produced in both animals and humans, and they hold great potential for neural restoration following spinal cord injury. While the effects of direct current electrical fields on adult-born spinal cells cultured ex vivo have recently been reported, the effects of direct current electrical fields on adult-born spinal cells in vivo have not been characterized. Here, we provide convincing findings that a therapeutic form of transspinal direct current stimulation (tsDCS) affects the migration and proliferation of adult-born spinal cells in mice. Specifically, cathodal tsDCS attracted the adult-born spinal cells, while anodal tsDCS repulsed them. In addition, both tsDCS polarities caused a significant increase in cell number. Regarding the potential mechanisms involved, both cathodal and anodal tsDCS caused significant increases in expression of brain-derived neurotrophic factor, while expression of nerve growth factor increased and decreased, respectively. In the spinal cord, both anodal and cathodal tsDCS increased blood flow. Since blood flow and angiogenesis are associated with the proliferation of neural stem cells, increased blood flow may represent a major factor in the modulation of newly born spinal cells by tsDCS. Consequently, we propose that the method and novel findings presented in the current study have the potential to facilitate cellular, molecular, and/or bioengineering strategies to repair injured spinal cords. NEW & NOTEWORTHY Our results indicate that transspinal direct current stimulation (tsDCS) affects the migratory pattern and proliferation of adult newly born spinal cells, a cell population which has been implicated in learning and memory. In addition, our results suggest a potential mechanism of action regarding the functional effects of applying direct current. Thus tsDCS may represent a novel method by which to manipulate the migration and cell number of adult newly born cells and restore functions following brain or spinal cord injury. Copyright © 2017 the American Physiological Society.
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Upadhya, Dinesh; Ogata, Masato; Reneker, Lixing W.
2013-01-01
The mitogen-activated protein kinases (MAPKs; also known as ERKs) are key intracellular signaling molecules that are ubiquitously expressed in tissues and were assumed to be functionally equivalent. Here, we use the mouse lens as a model system to investigate whether MAPK1 plays a specific role during development. MAPK3 is known to be dispensable for lens development. We demonstrate that, although MAPK1 is uniformly expressed in the lens epithelium, its deletion significantly reduces cell proliferation in the peripheral region, an area referred to as the lens germinative zone in which most active cell division occurs during normal lens development. By contrast, cell proliferation in the central region is minimally affected by MAPK1 deletion. Cell cycle regulators, including cyclin D1 and survivin, are downregulated in the germinative zone of the MAPK1-deficient lens. Interestingly, loss of MAPK1 subsequently induces upregulation of phosphorylated MAPK3 (pMAPK3) levels in the lens epithelium; however, this increase in pMAPK3 is not sufficient to restore cell proliferation in the germinative zone. Additionally, MAPK1 plays an essential role in epithelial cell survival but is dispensable for fiber cell differentiation during lens development. Our data indicate that MAPK1/3 control cell proliferation in the lens epithelium in a spatially defined manner; MAPK1 plays a unique role in establishing the highly mitotic zone in the peripheral region, whereas the two MAPKs share a redundant role in controlling cell proliferation in the central region of the lens epithelium. PMID:23482492
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Wasinski, Frederick; Estrela, Gabriel R.; Arakaki, Aline M.; Bader, Michael; Alenina, Natalia; Klempin, Friederike; Araújo, Ronaldo C.
2016-01-01
Physical exercise positively affects the metabolism and induces proliferation of precursor cells in the adult brain. Maternal exercise likewise provokes adaptations early in the offspring. Using a high-intensity swimming protocol that comprises forced swim training before and during pregnancy, we determined the effect of maternal swimming on the mouse offspring's neurogenesis. Our data demonstrate decreased proliferation in sublayers of the postnatal dentate gyrus in offspring of swimming mother at postnatal day (P) 8 accompanied with decreased survival of newly generated cells 4 weeks later. The reduction in cell numbers was predominantly seen in the hilus and molecular layer. At P35, the reduced amount of cells was also reflected by a decrease in the population of newly generated immature and mature neurons of the granule cell layer. Our data suggest that forced maternal swimming at high-intensity has a negative effect on the neurogenic niche development in postnatal offspring. PMID:27621701
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In vitro biocorrosion of Co-Cr-Mo implant alloy by macrophage cells.
Lin, Hsin-Yi; Bumgardner, Joel D
2004-11-01
We hypothesized that macrophage cells and their released reactive chemical species (RCS) affect Co-Cr-Mo alloy's corrosion properties and that alloy corrosion products change macrophage cell behavior. A custom cell culture corrosion cell was used to evaluate how culture medium, cells, and RCS altered alloy corrosion in 3-day tests. Corrosion was evaluated by measuring total charge transfer at a constant potential using a potentiostat and metal ion release by atomic emission spectroscopy. Viability, proliferation, and NO (nitric oxide) and IL-1beta (interlukin-1beta) release were used to assess cellular response to alloy corrosion products. In the presence of activated cells, total charge transfers and Co ion release were the lowest (p < 0.05). This was attributed to an enhancement of the surface oxide by RCS. Cr and Mo release were not different between cells and activated cells. Low levels of metal ions did not affect cell viability, proliferation, or NO release, though IL-1beta released from the activated cells was higher on the alloy compared to the controls. These data support the hypothesis that macrophage cells and their RCS affect alloy corrosion. Changes in alloy corrosion by cells may be important to the development of host responses to the alloy and its corrosion products.
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miR-342-3p affects hepatocellular carcinoma cell proliferation via regulating NF-κB pathway
DOE Office of Scientific and Technical Information (OSTI.GOV)
Zhao, Liang; Zhang, Yubao, E-mail: zhyb880077@sina.com
2015-02-13
Recent research indicates that non-coding microRNAs (miRNAs) help regulate basic cellular processes in many types of cancer cells. We hypothesized that overexpression of miR-342-3p might affect proliferation of hepatocellular carcinoma (HCC) cells. After confirming overexpression of miR-342-3p with qRT-PCR, MTT assay showed that HCC cell proliferation was significantly inhibited by miR-342-3p, and that it significantly decreased BrdU-positive cell proliferation by nearly sixfold. Searching for targets using three algorithms we found that miR-342-3p is related to the NF-κB pathway and luciferase assay found that IKK-γ, TAB2 and TAB3 are miR-342-3p target genes. Results of western blot on extracted nuclear proteins ofmore » HepG2 and HCT-116 cells showed that miR-342-3p reduced and miR-342-3p-in increased p65 nuclear levels and qRT-PCR found that NF-κB pathway downstream genes were downregulated by miR-342-3p and upregulated by miR-342-3p-in, confirming that miR-342 targets NF-κB pathway. Overexpression of Ikk-γ, TAB2 and TAB3 partially rescued HCC cells proliferation inhibited by miR-342-3p. Using the GSE54751 database we evaluated expression from 10 HCC samples, which strongly suggested downregulation of miR-342-3p and we also found inverse expression between miR-342-3p and its targets IKK-γ, TAB2 and TAB3 from 71 HCC samples. Our results show that miR-342-3p has a significant role in HCC cell proliferation and is suitable for investigation of therapeutic targets. - Highlights: • MiR-342-3p suppresses hepatocellular carcinoma cell proliferation. • MiR-342-3p targets IKK-γ, TAB2 and TAB3 genes. • MiR-342-3p downregulates NF-kB signaling pathway. • MiR-342-3p is downregulated in clinical hepatocellular carcinoma samples. • The expression of miR-342-3p and its target gene is inversely related.« less
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Docosahexaenoic acid (DHA) effects on proliferation and steroidogenesis of bovine granulosa cells.
Maillard, Virginie; Desmarchais, Alice; Durcin, Maeva; Uzbekova, Svetlana; Elis, Sebastien
2018-04-26
Docosahexaenoic acid (DHA) is a n-3 polyunsaturated fatty acid (PUFA) belonging to a family of biologically active fatty acids (FA), which are known to have numerous health benefits. N-3 PUFAs affect reproduction in cattle, and notably directly affect follicular cells. In terms of reproduction in cattle, n-3 PUFA-enriched diets lead to increased follicle size or numbers. The objective of the present study was to analyze the effects of DHA (1, 10, 20 and 50 μM) on proliferation and steroidogenesis (parametric and/or non parametric (permutational) ANOVA) of bovine granulosa cells in vitro and mechanisms of action through protein expression (Kruskal-Wallis) and signaling pathways (non parametric ANOVA) and to investigate whether DHA could exert part of its action through the free fatty acid receptor 4 (FFAR4). DHA (10 and 50 μM) increased granulosa cell proliferation and DHA 10 μM led to a corresponding increase in proliferating cell nuclear antigen (PCNA) expression level. DHA also increased progesterone secretion at 1, 20 and 50 μM, and estradiol secretion at 1, 10 and 20 μM. Consistent increases in protein levels were also reported for the steroidogenic enzymes, cytochrome P450 family 11 subfamily A member 1 (CYP11A1) and hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid delta-isomerase 1 (HSD3B1), and of the cholesterol transporter steroidogenic acute regulatory protein (StAR), which are necessary for production of progesterone or androstenedione. FFAR4 was expressed in all cellular types of bovine ovarian follicles, and in granulosa cells it was localized close to the cellular membrane. TUG-891 treatment (1 and 50 μM), a FFAR4 agonist, increased granulosa cell proliferation and MAPK14 phosphorylation in a similar way to that observed with DHA treatment. However, TUG-891 treatment (1, 10 and 50 μM) showed no effect on progesterone or estradiol secretion. These data show that DHA stimulated proliferation and steroidogenesis of bovine granulosa cells and led to MAPK14 phosphorylation. FFAR4 involvement in DHA effects requires further investigation, even if our data might suggest FFAR4 role in DHA effects on granulosa cell proliferation. Other mechanisms of DHA action should be investigated as the steroidogenic effects seemed to be independent of FFAR4 activation.
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Glucocorticoid effects on immune cell activation by staphylococcal exotoxins and lipopolysaccharide
NASA Technical Reports Server (NTRS)
Chapes, S. K.; Kopydlowski, K. M.; Fleming, S. D.; Iandolo, J. J.; Spooner, B. S. (Principal Investigator)
1992-01-01
Experiments were conducted to determine the effects of physiologically elevated corticosterone on the activation of macrophages and T cells. These studies find that the elevation of corticosterone does not affect the expression of membrane receptors on macrophages and does not affect the activation of macrophages to produce cytokines. In contrast, elevated corticosterone levels correlate with enhanced T cell proliferation to both mitogens and superantigens.
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Amirikia, Mehdi; Shariatzadeh, Seyed Mohammad Ali; Jorsaraei, Seyed Gholam Ali; Soleimani Mehranjani, Malek
2017-12-01
Cell behaviours such as proliferation and attachment can be affected by the length of pre-incubation period of the scaffolds in the culture medium for long term. The aim of this study was to investigate the long term pre-incubation of 3D silk fibroin scaffolds in complete culture medium on cell attachment and proliferation. After the preparation of silk fibroin scaffolds by the technique of freeze drying, the scaffolds were pre-incubated in complete culture medium for 2 d, 6 d or 10 d before apical papilla stem cells (SCAP) seeding. Modifications of the scaffold surface and wettability were examined by FE-SEM and water contact angle, respectively. Results showed a decrease both in roughness and water contact angle as pre-incubation time increases. DNA measurement after 18h and 10 d cell seeding showed a significant increase of DNA concentration which represents better attachment and proliferation with pre-incubation time increase. Qualitative examination, live&dead assay or H&E staining method after 30h and 10 d cell seeding respectively, indicated that pre-incubation of scaffolds has time dependent effect on cell proliferation and attachment. This suggests that improvement of cell attachment and proliferation may be mediated by differences in the amount of wettability (decreased water contact angle) after exposure of scaffold to culture medium for long term which, in turn, causes more protein adsorption in the surface of silk fibroin scaffold (decreased roughness). Copyright © 2017. Published by Elsevier Ltd.
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Keohane, Aoife; Ryan, Sinead; Maloney, Eimer; Sullivan, Aideen M; Nolan, Yvonne M
2010-01-01
Tumour necrosis factor-alpha (TNFalpha) is a pro-inflammatory cytokine, which influences neuronal survival and function yet there is limited information available on its effects on hippocampal neural precursor cells (NPCs). We show that TNFalpha treatment during proliferation had no effect on the percentage of proliferating cells prepared from embryonic rat hippocampal neurosphere cultures, nor did it affect cell fate towards either an astrocytic or neuronal lineage when cells were then allowed to differentiate. However, when cells were differentiated in the presence of TNFalpha, significantly reduced percentages of newly born and post-mitotic neurons, significantly increased percentages of astrocytes and increased expression of TNFalpha receptors, TNF-R1 and TNF-R2, as well as expression of the anti-neurogenic Hes1 gene, were observed. These data indicate that exposure of hippocampal NPCs to TNFalpha when they are undergoing differentiation but not proliferation has a detrimental effect on their neuronal lineage fate, which may be mediated through increased expression of Hes1. Copyright 2009 Elsevier Inc. All rights reserved.
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Zhao, Jianzhi; Li, Hanjun; Zhou, Rujiang; Ma, Gang; Dekker, Joseph D.; Tucker, Haley O.; Yao, Zhengju; Guo, Xizhi
2015-01-01
Hair follicle stem cells (HFSCs) in the bugle circularly generate outer root sheath (ORS) through linear proliferation within limited cycles during anagen phases. However, the mechanisms controlling the pace of HFSC proliferation remain unclear. Here we revealed that Foxp1, a transcriptional factor, was dynamically relocated from the nucleus to the cytoplasm of HFSCs in phase transitions from anagen to catagen, coupled with the rise of oxidative stress. Mass spectrum analyses revealed that the S468 phosphorylation of Foxp1 protein was responsive to oxidative stress and affected its nucleocytoplasmic translocation. Foxp1 deficiency in hair follicles led to compromised ROS accrual and increased HFSC proliferation. And more, NAC treatment profoundly elongated the anagen duration and HFSC proliferation in Foxp1-deficient background. Molecularly, Foxp1 augmented ROS levels through suppression of Trx1-mediated reductive function, thereafter imposing the cell cycle arrest by modulating the activity of p19/p53 pathway. Our findings identify a novel role for Foxp1 in controlling HFSC proliferation with cellular dynamic location in response to oxidative stress during hair cycling. PMID:26171970
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Miyata, Sairi; Yada, Tomotaka; Ishikawa, Natsuko; Taheruzzaman, Kazi; Hara, Ryohei; Matsuzaki, Takashi; Nishikawa, Akio
2017-03-01
To understand the mechanism of muscle remodeling during Xenopus laevis metamorphosis, we examined the in vitro effect of insulin-like growth factor 1 (IGF-1) on growth and differentiation of three different-fate myogenic cell populations: tadpole tail, tadpole dorsal, and young adult leg muscle. IGF-1 promoted growth and differentiation of both tail and leg myogenic cells only under conditions where these cells could proliferate. Inhibition of cell proliferation by DNA synthesis inhibitor cytosine arabinoside completely canceled the IGF-1's cell differentiation promotion, suggesting the possibility that IGF-1's differentiation-promotion effect is an indirect effect via IGF-1's cell proliferation promotion. IGF-1 promoted differentiation dose dependently with maximum effect at 100-500 ng/ml. RT-PCR analysis revealed the upregulation (11-fold) of ifg1 mRNA expression in developing limbs, suggesting that IGF-1 plays a role in promoting muscle differentiation during limb development. The combined effect of triiodo-L-thyronine (T 3 ) and IGF-1 was also examined. In adult leg cells, IGF-1 promoted growth and differentiation irrespective of the presence of T 3 . In larval tail cells, cell count was 76% lower in the presence of T 3 , and IGF-1 did not promote proliferation and differentiation in T 3 -containing medium. In larval dorsal cells, cell count was also lower in the presence of T 3 , but IGF-1 enhanced proliferation and differentiation in T 3 -containing medium. This result is likely due to the presence among dorsal cells of both adult and larval types (1:1). Thus, IGF-1 affects only adult-type myogenic cells in the presence of T 3 and helps accelerate dorsal muscle remodeling during metamorphosis.
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Effects of nerve cells and adhesion molecules on nerve conduit for peripheral nerve regeneration
Fiorellini, Joseph P.
2017-01-01
Background For peripheral nerve regeneration, recent attentions have been paid to the nerve conduits made by tissue-engineering technique. Three major elements of tissue-engineering are cells, molecules, and scaffolds. Methods In this study, the attachments of nerve cells, including Schwann cells, on the nerve conduit and the effects of both growth factor and adhesion molecule on these attachments were investigated. Results The attachment of rapidly-proliferating cells, C6 cells and HS683 cells, on nerve conduit was better than that of slowly-proliferating cells, PC12 cells and Schwann cells, however, the treatment of nerve growth factor improved the attachment of slowly-proliferating cells. In addition, the attachment of Schwann cells on nerve conduit coated with fibronectin was as good as that of Schwann cells treated with glial cell line-derived neurotrophic factor (GDNF). Conclusions Growth factor changes nerve cell morphology and affects cell cycle time. And nerve growth factor or fibronectin treatment is indispensable for Schwann cell to be used for implantation in artificial nerve conduits. PMID:29090249
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Deng, Wenbin; Poretz, Ronald D
2002-03-01
Lead (Pb) is a common neurotoxicant of major public health concern. Previous studies revealed that cultured oligodendrocyte progenitor cells (OPCs) are highly vulnerable to Pb toxicity. The present study examines the effect of Pb on the survival, proliferation and differentiation of OPCs in vitro. Dose-response studies showed that> or = l5-10 microM Pb is cytotoxic to OPCs within 24 h. However, 1 microM of Pb was found to inhibit the proliferation and differentiation of OPCs without affecting cell viability. Pb markedly decreased the proliferative capability of OPCs and inhibited cell-intrinsic lineage progression of OPCs at a late progenitor stage. The Pb-induced decrease of proliferation and differentiation was abolished by inhibition of protein kinase C (PKC) with bisindolylmaleimide I, while the effect of the PKC-activating agent phorbol-12,13-didecanoate was potentiated by Pb. Furthermore, Pb exposure of OPCs caused the translocation of PKC from the cytoplasm to membrane without an increase in total cellular PKC enzymic activity. These results indicate that Pb inhibits the proliferation and differentiation of oligodendrocyte lineage cells in vitro through a mechanism requiring PKC activation.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Sekine, Yoshitaka; Furuya, Yosuke; Nishii, Masahiro
2008-07-25
Recently, statins have been being studied for their proapoptic and antimetastatic effects. However, the exact mechanisms of their anticancer action are still unclear. Dolichyl phosphate is a nonsterol isoprenoid derivative in the mevalonate pathway that affects the expression of the Insulin-like growth factor 1 receptor (IGF-1R). IGF-1R activation is required for prostate cell proliferation; therefore, IGF-1R inhibitory agents may be of preventive and/or therapeutic value. In this study, the effects of simvastatin on IGF-1R signaling in prostate cancer PC-3 cells were examined. Simvastatin suppressed proliferation and induced apoptosis of PC-3, and the expression of IGF-1R was suppressed by simvastatin. Knockdownmore » of IGF-1R by siRNA led to inhibition of proliferation of PC-3. Simvastatin also inhibited IGF-1-induced activation of both ERK and Akt signaling and IGF-1-induced PC-3 cell proliferation. Our results suggest statins are potent inhibitors of the IGF-1/IGF-1R system in prostate cancer cells and may be beneficial in prostate cancer treatment.« less
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Hu, Jiang-Tian; Li, Yan; Yu, Bing
2015-08-21
To explore how Girdin/GIV is regulated by cyclic tension and propagates downstream signals to affect cell proliferation and migration. Human osteoblast-like MG-63 cells were exposed to cyclic tension force at 4000 μstrain and 0.5 Hz for 6 h, produced by a four-point bending system. Cyclic tension force upregulated Girdin and Akt expression and phosphorylation in cultured MG-63 cells. Girdin and Akt each promoted the phosphorylation of the other under stimulated tension. In vitro MTT and transwell assays showed that Girdin and Akt are required for cell proliferation and migration during cellular quiescence. Moreover, STAT3 was determined to be essential for Girdin expression undermore » stimulated tension force in the physiological condition, as well as for osteoblast proliferation and migration during quiescence. These findings suggest that the STAT3/Girdin/Akt pathway activates in osteoblasts in response to mechanical stimulation and may play a significant role in triggering osteoblast proliferation and migration during orthodontic treatment. - Highlights: • Tension force upregulates Girdin and Akt expression and phosphorylation. • Girdin and Akt promotes the phosphorylation of each other under tension stimulation. • Girdin and Akt are required for MG-63 cell proliferation and migration. • STAT3 is essential for Girdin expression after application of the tension forces.« less
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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.
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The contribution of B-cell proliferation to spleen enlargement in Babesia microti-infected mice.
Inchley, C J
1987-01-01
Flow cytofluorimetric analysis showed that B-cell proliferation makes a major contribution to the enlargement and increased cellularity of the spleen, which are characteristic of Babesia microti infections in mice. Expansion of the B-cell population was accompanied by modulation of the cell surface, which affected most B lymphocytes, and which was detected as a reduction in the density of surface immunoglobulin. This effect was noted as early as Day 7, shortly after the appearance of parasites in the circulation and the onset of gross spleen changes. In contrast to the results for B cells, the frequency of splenic T cells declined, and when the data were transformed into absolute numbers it became clear that only limited T-cell proliferation had occurred. There was no evidence to suggest that the balance of T-cell subsets was shifted in favour of suppressor T cells. The relationships of these results to reports of immunosuppression by this parasite are discussed. Images Figure 2 Figure 5 PMID:3493207
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Ansell, J D; McDougall, C M; Micklem, H S; Inchley, C J
1980-01-01
Antigen-dependent localization of 51Cr-labelled lymphocytes, and the subsequent uptake of IUdR into lymphoid organs has been studied as a function of age. Measures of cell localization indicated that while old age can alter the patterns of entry of lymphocytes into lymph nodes and spleen, these changes are variable and probably not sufficient alone to explain decreased primary antibody responses in old animals. Proliferation of cells, however, was consistently affected in both organs and this phenomenon is discussed in terms of abnormal T-cell function. PMID:7429546
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RELM-β promotes human pulmonary artery smooth muscle cell proliferation via FAK-stimulated surviving
DOE Office of Scientific and Technical Information (OSTI.GOV)
Lin, Chunlong, E-mail: lclmd@sina.com; Li, Xiaohui; Luo, Qiong
Resistin-like molecule-β (RELM-β), focal adhesion kinase (FAK), and survivin may be involved in the proliferation of cultured human pulmonary artery smooth muscle cells (HPAMSCs), which is involved in pulmonary hypertension. HPAMSCs were treated with human recombinant RELM-β (rhRELM-β). siRNAs against FAK and survivin were transfected into cultured HPASMCs. Expression of FAK and survivin were examined by RT-PCR and western blot. Immunofluorescence was used to localize FAK. Flow cytometry was used to examine cell cycle distribution and cell death. Compared to the control group, all rhRELM-β-treated groups demonstrated significant increases in the expression of FAK and survivin (P<0.05). rhRELM-β significantly increasedmore » the proportion of HPASMCs in the S phase and decreased the proportion in G0/G1. FAK siRNA down-regulated survivin expression while survivin siRNA did not affect FAK expression. FAK siRNA effectively inhibited FAK and survivin expression in RELM-β-treated HPASMCs and partially suppressed cell proliferation. RELM-β promoted HPASMC proliferation and upregulated FAK and survivin expression. In conclusion, results suggested that FAK is upstream of survivin in the signaling pathway mediating cell proliferation. FAK seems to be important in RELM-β-induced HPASMC proliferation, partially by upregulating survivin expression. - Highlights: • rhRELM-β increased the expression of FAK and survivin. • rhRELM-β increased the proportion of HPASMCs in the S phase. • FAK is upstream of survivin in the signaling pathway mediating cell proliferation. • FAK is important in RELM-β-induced HPASMC proliferation, partly via survivin.« less
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O'Leary, Olivia F; O'Connor, Richard M; Cryan, John F
2012-01-01
Adult hippocampal neurogenesis is an important process in the regulation of cognition, stress responsivity, and sensitivity to antidepressant and mood stabiliser drugs. Increasing evidence suggests that the hippocampus is functionally divided along its axis with the ventral hippocampus (vHi) playing a preferential role in stress- and anxiety-related processes, while the dorsal hippocampus (dHi) is crucial for spatial learning and memory. However, it is currently unclear whether stress or the medications used to treat stress-related disorders, preferentially affect neurogenesis in the vHi rather than dHi. The aim of this study was to determine whether the mood stabiliser, lithium, preferentially affects cell proliferation and survival in the vHi rather than dHi under stress conditions. To this end, mice of the stress-sensitive strain, BALB/c, underwent chronic exposure to immobilisation stress plus lithium treatment (0.2% lithium-supplemented diet), and the rates of cell proliferation and survival were compared in the dHi and vHi. Lithium preferentially increased cell proliferation in the vHi under stress conditions only. This increase in cell proliferation was secondary to reductions in the survival of newly-born cells. Moreover, lithium-induced decreases in cell survival in the vHi were only observed under stress conditions. Taken together, the data suggest that the turnover of newly-born cells in response to chronic stress and lithium treatment occurs predominantly in the vHi rather than the dHi. This article is part of a Special Issue entitled 'Anxiety and Depression'. Copyright © 2011 Elsevier Ltd. All rights reserved.
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Inhibition of caspases prevents ototoxic and ongoing hair cell death
NASA Technical Reports Server (NTRS)
Matsui, Jonathan I.; Ogilvie, Judith M.; Warchol, Mark E.
2002-01-01
Sensory hair cells die after acoustic trauma or ototoxic insults, but the signal transduction pathways that mediate hair cell death are not known. Here we identify several important signaling events that regulate the death of vestibular hair cells. Chick utricles were cultured in media supplemented with the ototoxic antibiotic neomycin and selected pharmacological agents that influence signaling molecules in cell death pathways. Hair cells that were treated with neomycin exhibited classically defined apoptotic morphologies such as condensed nuclei and fragmented DNA. Inhibition of protein synthesis (via treatment with cycloheximide) increased hair cell survival after treatment with neomycin, suggesting that hair cell death requires de novo protein synthesis. Finally, the inhibition of caspases promoted hair cell survival after neomycin treatment. Sensory hair cells in avian vestibular organs also undergo continual cell death and replacement throughout mature life. It is unclear whether the loss of hair cells stimulates the proliferation of supporting cells or whether the production of new cells triggers the death of hair cells. We examined the effects of caspase inhibition on spontaneous hair cell death in the chick utricle. Caspase inhibitors reduced the amount of ongoing hair cell death and ongoing supporting cell proliferation in a dose-dependent manner. In isolated sensory epithelia, however, caspase inhibitors did not affect supporting cell proliferation directly. Our data indicate that ongoing hair cell death stimulates supporting cell proliferation in the mature utricle.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Shao, Lei; Jinan Central Hospital Affiliated to Shandong University, Jinan, 250012; Li, Huanjie
Irisin is involved in promoting metabolism, immune regulation, and affects chronic inflammation in many systemic diseases, including gastric cancer. However, the role of irisin in lung cancer is not well characterized. To determine whether irisin has a protective effect against lung cancer, we cultured A549 and NCI-H446 lung cancer cells and treated them with irisin. We detected the proliferation by MTT assay, and assessed the migration and invasion of the cells by scratch wound healing assay and Tran-swell assay. The expression levels of epithelial-to-mesenchymal transition (EMT) markers and the related signaling pathways were detected by western blot analysis. Meanwhile, anmore » inhibitor of PI3K was used to investigate the effect of irsin. Finally, the expression of Snail was detected. We demonstrated that irisin inhibits the proliferation, migration, and invasion of lung cancer cells, and has a novel role in mediating the PI3K/AKT pathway in the cells. Irisin can reverse the activity of EMT and inhibit the expression of Snail via mediating the PI3K/AKT pathway, which is a key regulator of Snail. These results revealed that irisin inhibited EMT and reduced the invasion of lung cancer cells via the PI3K/AKT/Snail pathway. - Highlights: • Irisin inhibits the proliferation of lung cancer cells. • Irisin inhibits the migration and invasion of lung cancer cells. • Irisin affects the expression of EMT markers via inhibiting the PI3K/AKT pathway in lung cancer cells. • Irisin induces Snail downregulation via PI3K/AKT pathway activation.« less
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Ramos-Solano, Moisés; Meza-Canales, Ivan D; Torres-Reyes, Luis A; Alvarez-Zavala, Monserrat; Alvarado-Ruíz, Liliana; Rincon-Orozco, Bladimiro; Garcia-Chagollan, Mariel; Ochoa-Hernández, Alejandra B; Ortiz-Lazareno, Pablo C; Rösl, Frank; Gariglio, Patricio; Jave-Suárez, Luis F; Aguilar-Lemarroy, Adriana
2015-07-01
According to the multifactorial model of cervical cancer (CC) causation, it is now recognized that other modifications, in addition to Human papillomavirus (HPV) infection, are necessary for the development of this neoplasia. Among these, it has been proposed that a dysregulation of the WNT pathway might favor malignant progression of HPV-immortalized keratinocytes. The aim of this study was to identify components of the WNT pathway differentially expressed in CC vs. non-tumorigenic, but immortalized human keratinocytes. Interestingly, WNT7A expression was found strongly downregulated in cell lines and biopsies derived from CC. Restoration of WNT7A in CC-derived cell lines using a lentiviral gene delivery system or after adding a recombinant human protein decreases cell proliferation. Likewise, WNT7A silencing in non-tumorigenic cells markedly accelerates proliferation. Decreased WNT7A expression was due to hypermethylation at particular CpG sites. To our knowledge, this is the first study reporting reduced WNT7A levels in CC-derived cells and that ectopic WNT7A restoration negatively affects cell proliferation and migration. Copyright © 2015 Elsevier Inc. All rights reserved.
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Distinct T helper cell dependence of memory B-cell proliferation versus plasma cell differentiation.
Zabel, Franziska; Fettelschoss, Antonia; Vogel, Monique; Johansen, Pål; Kündig, Thomas M; Bachmann, Martin F
2017-03-01
Several memory B-cell subclasses with distinct functions have been described, of which the most effective is the class-switched (CS) memory B-cell population. We have previously shown, using virus-like particles (VLPs), that the proliferative potential of these CS memory B cells is limited and they fail to re-enter germinal centres (GCs). However, VLP-specific memory B cells quickly differentiated into secondary plasma cells (PCs) with the virtue of elevated antibody production compared with primary PCs. Whereas the induction of VLP + memory B cells was strongly dependent on T helper cells, we were wondering whether re-stimulation of VLP + memory B cells and their differentiation into secondary PCs would also require T helper cells. Global absence of T helper cells led to strongly impaired memory B cell proliferation and PC differentiation. In contrast, lack of interleukin-21 receptor-dependent follicular T helper cells or CD40 ligand signalling strongly affected proliferation of memory B cells, but differentiation into mature secondary PCs exhibiting increased antibody production was essentially normal. This contrasts with primary B-cell responses, where a strong dependence on CD40 ligand but limited importance of interleukin-21 receptor was seen. Hence, T helper cell dependence differs between primary and secondary B-cell responses as well as between memory B-cell proliferation and PC differentiation. © 2016 John Wiley & Sons Ltd.
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SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Bonifati, Serena; Daly, Michele B.; St Gelais, Corine
SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G{sub 1}/G{sub 0} phase and reduces apoptosis. These alterationsmore » correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.« less
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Lapidot, Tair; Walker, Michael D; Kanner, Joseph
2002-05-22
It has recently been suggested that the ability of apple extracts to inhibit proliferation of tumor cells in vitro may be due to phenolic/flavonoid antioxidants. Our study demonstrates that this inhibition is caused indirectly by H(2)O(2) generated through interaction of the phenolics with the cell culture media. The results indicate that many previously reported effects of flavonoids and phenolic compounds on cultured cells may result from similar artifactual generation of oxidative stress. We suggest that in order to prevent such artifacts, the use of catalase and/or metmyoglobin in the presence of reducing agents should be considered as a method to decompose H(2)O(2) and prevent generation of other reactive oxygen species, which could affect cell proliferation. The use of tumor cells and "nontumor cells" in a bioassay to measure antioxidant activity, in this context, is potentially misleading and should be applied with caution.
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Linero, Itali M; Doncel, Adriana; Chaparro, Orlando
2014-01-01
The use of mesenchymal stem cells in clinical practice has increased considerably in the last decade because they play a supporting role in the processes of tissue repair and regeneration, becoming the main tool of cell therapy for the treatment of diseases functionally affecting bone and cartilage tissue . To evaluate in vitro the proliferative and osteogenic differentiation ability of mesenchymal stem cells derived from human adipose tissue in a blood plasma hydrogel. Mesenchymal stem cells were obtained from human adipose tissue explants and characterized by flow cytometry. Their multipotentiality was demonstrated by their ability to differentiate to adipogenic and osteogenic lineages. Cell proliferation and osteogenic differentiation ability of the cells cultured in blood plasma hydrogels were also evaluated. Mesenchymal stem cells derived from human adipose tissue growing in human blood plasma hydrogels showed a pattern of proliferation similar to that of the cells cultured in monolayer and also maintained their ability to differentiate to osteogenic lineage. Human blood plasma hydrogels are a suitable support for proliferation and osteogenic differentiation of mesenchymal stem cells derived from human adipose tissue and provides a substrate that is autologous, biocompatible, reabsorbable, easy to use, potentially injectable and economic, which could be used as a successful strategy for the management and clinical application of cell therapy in regenerative medicine.
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Harati, K; Behr, B; Daigeler, A; Hirsch, T; Jacobsen, F; Renner, M; Harati, A; Wallner, C; Lehnhardt, M; Becerikli, M
2017-01-01
The cytostatic effects of the polyphenol curcumin and Viscum album extract (VAE) were assessed in soft-tissue sarcoma (STS) cells. Eight human STS cell lines were used: fibrosarcoma (HT1080), liposarcoma (SW872, T778, MLS-402), synovial sarcoma (SW982, SYO1, 1273), and malignant fibrous histiocytoma (U2197). Primary human fibroblasts served as control cells. Cell proliferation, viability, and cell index (CI) were analyzed by BrdU assay, MTT assay, and real-time cell analysis (RTCA). As indicated by BrdU and MTT, curcumin significantly decreased the cell proliferation of five cell lines (HT1080, SW872, SYO1, 1273, and U2197) and the viability of two cell lines (SW872 and SW982). VAE led to significant decreases of proliferation in eight cell lines (HT1080, SW872, T778, MLS-402, SW982, SYO1, 1293, and U2197) and reduced viability in seven STS lines (HT1080, SW872, T778, MLS-402, SW982, SYO1, and 1273). As indicated by RTCA for 160 h, curcumin decreased the CI of all synovial sarcoma cell lines as well as T778 and HT1080. VAE diminished the CI in most of the synovial sarcoma (SW982, SYO1) and liposarcoma (SW872, T778) cell lines as well as HT1080. Primary fibroblasts were not affected adversely by the two compounds in RTCA. Curcumin and VAE can inhibit the proliferation and viability of STS cells.
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Lee, Suk Jun; Bae, Joonbeom; Kim, Sunhee; Jeong, Seonah; Choi, Chang-Yong; Choi, Sang-Pil; Kim, Hyun-Sook; Jung, Woon-Won; Imm, Jee-Young; Kim, Sae Hun; Chun, Taehoon
2013-02-01
Treatment of helper T (Th) cells with saponins from soy bean and mung bean prevented their activation by inhibiting cell proliferation and cytokine secretion. However, the saponins did not affect the expression of major histocompatibility complex class II (A(b)) and co-stimulatory molecule (CD86) on professional antigen-presenting cells. Instead, the saponins directly inhibited Th cell proliferation by blocking the G(1) to S phase cell cycle transition. Moreover, blocking of the cell cycle by the saponins was achieved by decreased expression of cyclin D1 and cyclin E, and constitutive expression of p27(KIP1). Saponins also increased stability of p27(KIP1) in Th cells after antigenic stimulation.
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Favoni, Roberto E; Pattarozzi, Alessandra; Lo Casto, Michele; Barbieri, Federica; Gatti, Monica; Paleari, Laura; Bajetto, Adriana; Porcile, Carola; Gaudino, Giovanni; Mutti, Luciano; Corte, Giorgio; Florio, Tullio
2010-03-01
Altered EGFR activity is a causal factor for human tumor development, including malignant pleural mesotheliomas. The aim of the present study was the evaluation of the effects of Gefitinib on EGF-induced mesothelioma cell proliferation and the intracellular mechanisms involved. Cell proliferation, DNA synthesis and apoptosis were measured by MTT, thymidine incorporation and FACS analysis; EGFR, ERK1/2 and Akt expression and phosphorylation by Western blot, whereas receptor sites were analyzed by binding studies. Gefitinib inhibited EGF-induced proliferation in two mesothelioma cell lines, derived from pleural effusion (IST-Mes2) or tumor biopsy (ZL55). The treatment with Gefitinib induced cell cycle arrest in both cell lines, while apoptosis was observed only for high concentrations and prolonged drug exposure. EGF-dependent mesothelioma cell proliferation was mediated by EGFR and ERK1/2 phosphorylation, while Akt was not affected. Gefitinib inhibited both EGFR and ERK1/2 activation, being maximal at drug concentrations that induce cytostatic effects, suggesting that the proapoptotic activity of Gefitinib is independent from EGFR inhibition. Gefitinib treatment increased EGFR Bmax, possibly through membrane stabilization of inactive receptor dimers that we show to be induced by the drug also in the absence of EGF. EGFR activation of ERK1/2 represents a key pathway for pleural mesothelioma cell proliferation. Low concentrations of Gefitinib cause mesothelioma cell cycle arrest through the blockade of EGFR activity while high concentrations induce apoptosis. Finally, we propose that the formation of inactive EGFR dimers may contribute to the antitumoral activity of Gefitinib.
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Abrogation of Cbl-PI3K interaction increases bone formation and osteoblast proliferation.
Brennan, Tracy; Adapala, Naga Suresh; Barbe, Mary F; Yingling, Vanessa; Sanjay, Archana
2011-11-01
Cbl is an adaptor protein and E3 ligase that plays both positive and negative roles in several signaling pathways that affect various cellular functions. Tyrosine 737 is unique to Cbl and phosphorylated by Src family kinases. Phosphorylated CblY737 creates a binding site for the p85 regulatory subunit of phosphatidylinositol 3 kinase (PI3K) that also plays an important role in the regulation of bone homeostasis. To investigate the role of Cbl-PI3K interaction in bone homeostasis, we examined knock-in mice in which the PI3K binding site on Cbl was ablated due to the substitution of tyrosine 737 to phenylalanine (Cbl(YF/YF), YF mice). We previously reported that bone volume in these mice is increased due to decreased osteoclast function (Adapala et al., J Biol Chem 285:36745-36758, 19). Here, we report that YF mice also have increased bone formation and osteoblast numbers. In ex vivo cultures bone marrow-derived YF osteoblasts showed increased Col1A expression and their proliferation was also significantly augmented. Moreover, proliferation of MC3T3-E1 cells was increased after treatment with conditioned medium generated by culturing YF bone marrow stromal cells. Expression of stromal derived factor-1 (SDF-1) was increased in YF bone marrow stromal cells compared to wild type. Increased immunostaining of SDF-1 and CXCR4 was observed in YF bone marrow stromal cells compared to wild type. Treatment of YF condition medium with neutralizing anti-SDF-1 and anti-CXCR4 antibodies attenuated MC3T3-E1 cell proliferation. Cumulatively, these results show that abrogation of Cbl-PI3K interaction perturbs bone homeostasis, affecting both osteoclast function and osteoblast proliferation.
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Jomaa, Barae; Aarts, Jac M M J G; de Haan, Laura H J; Peijnenburg, Ad A C M; Bovee, Toine F H; Murk, Albertinka J; Rietjens, Ivonne M C M
2013-01-01
This study investigates the in vitro effect of eleven thyroid-active compounds known to affect pituitary and/or thyroid weights in vivo, using the proliferation of GH3 rat pituitary cells in the so-called "T-screen," and of FRTL-5 rat thyroid cells in a newly developed test denoted "TSH-screen" to gain insight into the relative value of these in vitro proliferation tests for an integrated testing strategy (ITS) for thyroid activity. Pituitary cell proliferation in the T-screen was stimulated by three out of eleven tested compounds, namely thyrotropin releasing hormone (TRH), triiodothyronine (T3) and thyroxine (T4). Of these three compounds, only T4 causes an increase in relative pituitary weight, and thus T4 was the only compound for which the effect in the in vitro assay correlated with a reported in vivo effect. As to the newly developed TSH-screen, two compounds had an effect, namely, thyroid-stimulating hormone (TSH) induced and T4 antagonized FRTL-5 cell proliferation. These effects correlated with in vivo changes induced by these compounds on thyroid weight. Altogether, the results indicate that most of the selected compounds affect pituitary and thyroid weights by modes of action different from a direct thyroid hormone receptor (THR) or TSH receptor (TSHR)-mediated effect, and point to the need for additional in vitro tests for an ITS. Additional analysis of the T-screen revealed a positive correlation between the THR-mediated effects of the tested compounds in vitro and their effects on relative heart weight in vivo, suggesting that the T-screen may directly predict this THR-mediated in vivo adverse effect.
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Paredes, João A; Zhou, Xiaoshan; Höglund, Stefan; Karlsson, Anna
2013-01-01
Loss of thymidine kinase 2 (TK2) causes a heterogeneous myopathic form of mitochondrial DNA (mtDNA) depletion syndrome (MDS) in humans that predominantly affects skeletal muscle tissue. In mice, TK2 deficiency also affects several tissues in addition to skeletal muscle, including brain, heart, adipose tissue, kidneys and causes death about 3 weeks after birth. We analysed skeletal muscle and heart muscle tissues of Tk2 knockout mice at postnatal development phase and observed that TK2 deficient pups grew slower and their skeletal muscles appeared significantly underdeveloped, whereas heart was close to normal in size. Both tissues showed mtDNA depletion and mitochondria with altered ultrastructure, as revealed by transmission electron microscopy. Gene expression microarray analysis showed a strong down-regulation of genes involved in cell cycle and cell proliferation in both tissues, suggesting a lower pool of undifferentiated proliferating cells. Analysis of isolated primary myoblasts from Tk2 knockout mice showed slow proliferation, less ability to differentiate and signs of premature senescence, even in absence of mtDNA depletion. Our data demonstrate that TK2 deficiency disturbs myogenic progenitor cells function in postnatal skeletal muscle and we propose this as one of the causes of underdeveloped phenotype and myopathic characteristic of the TK2 deficient mice, in addition to the progressive mtDNA depletion, mitochondrial damage and respiratory chain deficiency in post-mitotic differentiated tissue.
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Paredes, João A.; Zhou, Xiaoshan; Höglund, Stefan; Karlsson, Anna
2013-01-01
Loss of thymidine kinase 2 (TK2) causes a heterogeneous myopathic form of mitochondrial DNA (mtDNA) depletion syndrome (MDS) in humans that predominantly affects skeletal muscle tissue. In mice, TK2 deficiency also affects several tissues in addition to skeletal muscle, including brain, heart, adipose tissue, kidneys and causes death about 3 weeks after birth. We analysed skeletal muscle and heart muscle tissues of Tk2 knockout mice at postnatal development phase and observed that TK2 deficient pups grew slower and their skeletal muscles appeared significantly underdeveloped, whereas heart was close to normal in size. Both tissues showed mtDNA depletion and mitochondria with altered ultrastructure, as revealed by transmission electron microscopy. Gene expression microarray analysis showed a strong down-regulation of genes involved in cell cycle and cell proliferation in both tissues, suggesting a lower pool of undifferentiated proliferating cells. Analysis of isolated primary myoblasts from Tk2 knockout mice showed slow proliferation, less ability to differentiate and signs of premature senescence, even in absence of mtDNA depletion. Our data demonstrate that TK2 deficiency disturbs myogenic progenitor cells function in postnatal skeletal muscle and we propose this as one of the causes of underdeveloped phenotype and myopathic characteristic of the TK2 deficient mice, in addition to the progressive mtDNA depletion, mitochondrial damage and respiratory chain deficiency in post-mitotic differentiated tissue. PMID:23341978
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FUNCTIONAL DEREGULATION OF KIT: LINK TO MAST CELL PROLIFERATIVE DISEASES AND OTHER NEOPLASMS
Cruse, Glenn; Metcalfe, Dean D.; Olivera, Ana
2014-01-01
SYNOPSIS Signaling through the receptor tyrosine kinase KIT mediates differentiation, proliferation and survival of hematopoietic precursor cells and mast cells. Constitutive KIT signaling due to somatic point mutations in c-Kit is an important occurrence in the development of mast cell proliferation disorders and other hematological malignancies. In this review, we discuss the common gain-of-function mutations found in these malignancies, particularly in mast cell proliferation disorders, and summarize the current understanding of the molecular mechanisms by which transforming point mutations in KIT may affect KIT structure and function and lead to altered downstream signaling and cellular transformation. Drugs targeting KIT have shown mixed success in the treatment of these diseases. A brief overview of the most common KIT inhibitors currently used, the reasons for the varied clinical results of such inhibitors and a discussion of potential new strategies are provided. PMID:24745671
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Chen, Shuang; Dong, Siyuan; Li, Zhao; Guo, Xiaofan; Zhang, Naijin; Yu, Bo; Sun, Yingxian
2017-01-01
Abnormal proliferation of vascular smooth muscle cells (VSMCs) is a hallmark of vascular lesions, such as atherosclerosis and restenosis. PDGF-ββ, an isoform of PDGF (platelet-derived growth factor), has been demonstrated to induce proliferation and migration of VSMCs. Atorvastatin calcium, a selective inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, has favorable protective effects on VSMCs. This study examined the effects of atorvastatin calcium on the proliferation and migration of PDGF-ββ-treated VSMCs, as well as its underlying mechanisms. MTT assays, Edu imaging, cell cycle analysis, wound healing assays, transwell migration assays, and western blot analysis were performed. Atorvastatin calcium significantly inhibited cell proliferation, DNA synthesis and cell migration of PDGF-ββ-treated VSMCs. We demonstrated that atorvastatin calcium induced cell cycle arrest in the G0/G1 phase in response to PDGF-ββ stimulation and decreased the expression of G0/G1-specific regulatory proteins, including proliferating cell nuclear antigen (PCNA), CDK2, cyclin D1, cyclin E and CDK4 in PDGF-ββ-treated VSMCs. Moreover, pretreatment with atorvastatin calcium inhibited the PDGF-ββ-treated phosphorylation of PDGFRβ and Akt, whereas atorvastatin calcium did not affect the phosphorylation of PLC-γ1 or (ERK) 1/2. Our data suggested that atorvastatin calcium inhibited abnormal proliferation and migration of VSMCs through G0/G1 cell cycle arrest and suppression of the PDGFRβ-Akt signaling cascade. © 2017 The Author(s). Published by S. Karger AG, Basel.
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Jiráková, Klára; Šeneklová, Monika; Jirák, Daniel; Turnovcová, Karolína; Vosmanská, Magda; Babič, Michal; Horák, Daniel; Veverka, Pavel; Jendelová, Pavla
2016-01-01
Introduction Magnetic resonance (MR) imaging is suitable for noninvasive long-term tracking. We labeled human induced pluripotent stem cell-derived neural precursors (iPSC-NPs) with two types of iron-based nanoparticles, silica-coated cobalt zinc ferrite nanoparticles (CZF) and poly-l-lysine-coated iron oxide superparamagnetic nanoparticles (PLL-coated γ-Fe2O3) and studied their effect on proliferation and neuronal differentiation. Materials and methods We investigated the effect of these two contrast agents on neural precursor cell proliferation and differentiation capability. We further defined the intracellular localization and labeling efficiency and analyzed labeled cells by MR. Results Cell proliferation was not affected by PLL-coated γ-Fe2O3 but was slowed down in cells labeled with CZF. Labeling efficiency, iron content and relaxation rates measured by MR were lower in cells labeled with CZF when compared to PLL-coated γ-Fe2O3. Cytoplasmic localization of both types of nanoparticles was confirmed by transmission electron microscopy. Flow cytometry and immunocytochemical analysis of specific markers expressed during neuronal differentiation did not show any significant differences between unlabeled cells or cells labeled with both magnetic nanoparticles. Conclusion Our results show that cells labeled with PLL-coated γ-Fe2O3 are suitable for MR detection, did not affect the differentiation potential of iPSC-NPs and are suitable for in vivo cell therapies in experimental models of central nervous system disorders. PMID:27920532
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Salton, Gabrielle Dias; Laurino, Claudia Cilene Fernandes Correia; Mega, Nicolás Oliveira; Delgado-Cañedo, Andrés; Setterblad, Niclas; Carmagnat, Maryvonnick; Xavier, Ricardo Machado; Cirne-Lima, Elizabeth; Lenz, Guido; Henriques, João Antonio Pêgas; Laurino, Jomar Pereira
2017-08-03
Eukaryote initiation factor 2 subunit β (eIF2β) plays a crucial role in regulation protein synthesis, which mediates the interaction of eIF2 with mRNA. eIF2β contains evolutionarily conserved polylysine stretches in amino-terminal region and a zinc finger motif in the carboxy-terminus. The gene eIF2β was cloned under tetracycline transcription control and the polylysine stretches were deleted by site-directed mutagenesis (eIF2βΔ3K). The plasmid was transfected into HEK 293 TetR cells. These cells were analyzed for their proliferative and translation capacities as well as cell death rate. Experiments were performed using gene reporter assays, western blotting, flow cytometry, cell sorting, cell proliferation assays and confocal immunofluorescence. eIF2βΔ3K affected negatively the protein synthesis, cell proliferation and cell survival causing G2 cell cycle arrest and increased cell death, acting in a negative dominant manner against the native protein. Polylysine stretches are also essential for eIF2β translocated from the cytoplasm to the nucleus, accumulating in the nucleolus and eIF2βΔ3K did not make this translocation. eIF2β is involved in the protein synthesis process and should act in nuclear processes as well. eIF2βΔ3K reduces cell proliferation and causes cell death. Since translation control is essential for normal cell function and survival, the development of drugs or molecules that inhibit translation has become of great interest in the scenario of proliferative disorders. In conclusion, our results suggest the dominant negative eIF2βΔ3K as a therapeutic strategy for the treatment of proliferative disorders and that eIF2β polylysine stretch domains are promising targets for this.
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Geiger, Pamina; Mayer, Barbara; Wiest, Irmi; Schulze, Sandra; Jeschke, Udo; Weissenbacher, Tobias
2016-11-08
Galectin-1 (gal-1) belongs to the family of β-galactoside-binding proteins which primarily recognizes the Galβ1-4GlcNAc sequences of oligosaccharides associated with several cell surface glycoconjugates. The lectin recognizes correspondent glycoepitopes on human breast cancer cells. Galectin-1 is expressed both in normal and malignant tissues. Lymphatic organs naturally possessing high rates of apoptotic cells, express high levels of Galectin-1. Furthermore galectin-1 can initiate T cell apoptosis. Binding of galectin-1 to trophoblast tumor cells presenting the oncofetal Thomsen-Friedenreich (TF) carbohydrate antigen inhibits tumor cell proliferation. In this study we examined the impact galectin-1 has in vitro on cell proliferation, apoptotic potential and metabolic activity of MCF-7 and T-47D breast cancer cells in dependence to their expression of the Thomsen-Friedenreich (TF) tumor antigen. For proliferation and apoptosis assays cells were grown in presence of 10, 30 and 60 μg gal-1/ml medium. Cell proliferation was determined by a BrdU uptake ELISA. Detection of apoptotic cells was done by M30 cyto death staining, in situ nick translation and by a nucleosome ELISA method. Furthermore we studied the impact galectin-1 has on the metabolic activity of MCF-7 and T-47D cells in a homotypic three-dimensional spheroid cell culture model mimicking a micro tumour environment. Gal-1 inhibited proliferation of MCF-7 cells (strong expression of the TF epitope) but did not significantly change proliferation of T-47D cells (weak expression of the TF epitope). The incubation of MCF-7 cells with gal-1 raised number of apoptotic cells significantly. Treating the spheroids with 30 μg/ml galectin-1 in addition to standard chemotherapeutic regimes (FEC, TAC) resulted in further suppression of the metabolic activity in MCF-7 cells whereas T-47D cells were not affected. Our results demonstrate that galectin-1 can inhibit proliferation und metabolic cell activity and induce apoptosis in breast tumor cell lines with high expression levels of the Thomsen-Friedenreich (TF) antigen in monolayer and spheroid cell culture models.
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Usage of Human Mesenchymal Stem Cells in Cell-based Therapy: Advantages and Disadvantages.
Kim, Hee Jung; Park, Jeong-Soo
2017-03-01
The use of human mesenchymal stem cells (hMSCs) in cell-based therapy has attracted extensive interest in the field of regenerative medicine, and it shows applications to numerous incurable diseases. hMSCs show several superior properties for therapeutic use compared to other types of stem cells. Different cell types are discussed in terms of their advantages and disadvantages, with focus on the characteristics of hMSCs. hMSCs can proliferate readily and produce differentiated cells that can substitute for the targeted affected tissue. To maximize the therapeutic effects of hMSCs, a substantial number of these cells are essential, requiring extensive ex vivo cell expansion. However, hMSCs have a limited lifespan in an in vitro culture condition. The senescence of hMSCs is a double-edged sword from the viewpoint of clinical applications. Although their limited cell proliferation potency protects them from malignant transformation after transplantation, senescence can alter various cell functions including proliferation, differentiation, and migration, that are essential for their therapeutic efficacy. Numerous trials to overcome the limited lifespan of mesenchymal stem cells are discussed.
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Usage of Human Mesenchymal Stem Cells in Cell-based Therapy: Advantages and Disadvantages
Kim, Hee Jung; Park, Jeong-Soo
2017-01-01
ABSTRACT The use of human mesenchymal stem cells (hMSCs) in cell-based therapy has attracted extensive interest in the field of regenerative medicine, and it shows applications to numerous incurable diseases. hMSCs show several superior properties for therapeutic use compared to other types of stem cells. Different cell types are discussed in terms of their advantages and disadvantages, with focus on the characteristics of hMSCs. hMSCs can proliferate readily and produce differentiated cells that can substitute for the targeted affected tissue. To maximize the therapeutic effects of hMSCs, a substantial number of these cells are essential, requiring extensive ex vivo cell expansion. However, hMSCs have a limited lifespan in an in vitro culture condition. The senescence of hMSCs is a double-edged sword from the viewpoint of clinical applications. Although their limited cell proliferation potency protects them from malignant transformation after transplantation, senescence can alter various cell functions including proliferation, differentiation, and migration, that are essential for their therapeutic efficacy. Numerous trials to overcome the limited lifespan of mesenchymal stem cells are discussed. PMID:28484739
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Omega-3 free fatty acids attenuate insulin-promoted breast cancer cell proliferation.
Guo, Yang; Zhu, Sheng-Long; Wu, Yi-Kuan; He, Zhao; Chen, Yong-Quan
2017-06-01
High insulin levels in obese people are considered as a risk factor to induce breast carcinogenesis. And consumption of fish oils which mainly contain omega-3 fatty acids is associated with a reduced risk of breast cancer. However, whether omega-3 free fatty acids (FFAs) modulate insulin signaling pathway to prevent breast cancer is poorly understood. The current study tested the hypothesis that omega-3 FFAs attenuate insulin-induced breast cancer cell proliferation and regulate insulin signaling pathway. We show here that omega-3 FFAs attenuate MCF-7 cell proliferation and Akt and Erk1/2 phosphorylation levels stimulated by insulin. Knockdown Shp2 by siRNA resulted in significantly elevated omega-3 FFAs-activated Akt phosphorylation but failed to change insulin-stimulated Akt and Erk1/2 phosphorylation. And viable cell number was not affected by either downregulation of Shp2 expression or Erk1/2 inhibitor U0126 treatment. These observations indicated that omega-3 FFAs attenuate insulin-promoted breast cancer cell proliferation and insulin-activated Akt phosphorylation. Copyright © 2017 Elsevier Inc. All rights reserved.
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[In vitro anti-angiogenic action of lambda-carrageenan oligosaccharides].
Chen, Hai-Min; Yan, Xiao-Jun; Wang, Feng; Lin, Jing; Xu, Wei-Feng
2007-06-01
This study was designed to evaluate the inhibition effect of lambda-carrageenan oligosaccharides on neovascularization in vitro by chick chorioallantoic membrane (CAM) model and human umbilical vein endothelial cell ( HUVEC). lambda-Carrageenan oligosaccharides caused a dose-dependent decrease of the vascular density of CAM, and adversely affected capillary plexus formation. At a high concentration of 1 mg x mL(-1), this compound inhibited the endothelial cell proliferation, while low concentration of lambda-carrageenan oligosaccharides (< 250 microg x mL(-1)) affected the cell survival slightly (> 95%). Different cytotoxic sensitivity of lambda-carrageenan oligosaccharides in three kinds of cells was observed, of which HUVEC is the most sensitive to this oligosaccharides. The inhibitory action of lambda-carrageenan oligosaccharides on the endothelial cell invasion and migration was also observed at relatively low concentration (150 - 300 microg x mL(-1)) through down-regulation of intracellular matrix metalloproteinases-2 (MMP-2) expression on endothelial cells. Current observations demonstrated that lambda-carrageenan oligosaccharides are potential angiogenesis inhibitor with combined effects of inhibiting invasion, migration and proliferation.
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Hu, Ke Yao; Wang, De Gui; Liu, Peng Fei; Cao, Yan Wei; Wang, Yong Hua; Yang, Xue Cheng; Hu, Cheng Xia; Sun, Li Jiang; Niu, Hai Tao
2016-08-01
Phosphofructokinase-2/fructose-2,6-bisphosphatase 3 (PFKFB3) and monocarboxylate transporter 1 (MCT1) play important roles in tumor endothelial cells (ECs) and several biological processes. The present study was conducted to study the effects of PFKFB3 and MCT1 on cell proliferation and apoptosis in the tumor microenvironment by co-culture of HUVECs and T24, a bladder cancer (BC) cell line, using a microfluidic device. Immunofluorescence assay showed that HUVEC activity was significantly enhanced under co-culture with T24 cells, according to the stronger fluorescence intensity of CD31 and CD105 than that in the signal‑cultured cells. Quercetin treatment inhibited MCT1 expression but did not affect PFKFB3 expression. Knockdown of MCT1 or/and PFKFB3 increased the apoptosis rate of the HUVECs under single-culture and co-culture situations by staining with calcein and propidium iodide. Meanwhile, cell proliferation and lactic concentration were significantly decreased after the blocking of MCT1 or/and PFKFB3, as compared with that in the control group. No obvious differences in the effects on apoptosis, proliferation and lactic concentration were found between cells treated with quercetin and siMCT1. Thus, we concluded that the targeting of MCT1 and PFKFB3 regulated cell proliferation and apoptosis in BC cells by altering the tumor microenvironment, and quercetin exhibited a potential antitumor effect by targeting MCT1.
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Lithium Causes G2 Arrest of Renal Principal Cells
de Groot, Theun; Alsady, Mohammad; Jaklofsky, Marcel; Otte-Höller, Irene; Baumgarten, Ruben; Giles, Rachel H.
2014-01-01
Vasopressin-regulated expression and insertion of aquaporin-2 channels in the luminal membrane of renal principal cells is essential for urine concentration. Lithium affects urine concentrating ability, and approximately 20% of patients treated with lithium develop nephrogenic diabetes insipidus (NDI), a disorder characterized by polyuria and polydipsia. Lithium-induced NDI is caused by aquaporin-2 downregulation and a reduced ratio of principal/intercalated cells, yet lithium induces principal cell proliferation. Here, we studied how lithium-induced principal cell proliferation can lead to a reduced ratio of principal/intercalated cells using two-dimensional and three-dimensional polarized cultures of mouse renal collecting duct cells and mice treated with clinically relevant lithium concentrations. DNA image cytometry and immunoblotting revealed that lithium initiated proliferation of mouse renal collecting duct cells but also increased the G2/S ratio, indicating G2/M phase arrest. In mice, treatment with lithium for 4, 7, 10, or 13 days led to features of NDI and an increase in the number of principal cells expressing PCNA in the papilla. Remarkably, 30%–40% of the PCNA-positive principal cells also expressed pHistone-H3, a late G2/M phase marker detected in approximately 20% of cells during undisturbed proliferation. Our data reveal that lithium treatment initiates proliferation of renal principal cells but that a significant percentage of these cells are arrested in the late G2 phase, which explains the reduced principal/intercalated cell ratio and may identify the molecular pathway underlying the development of lithium-induced renal fibrosis. PMID:24408872
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Hong, Wei; Peng, Gongyong; Hao, Binwei; Liao, Baoling; Zhao, Zhuxiang; Zhou, Yumin; Peng, Fang; Ye, Xiuqin; Huang, Lingmei; Zheng, Mengning; Pu, Jinding; Liang, Chunxiao; Yi, Erkang; Peng, Huanhuan; Li, Bing; Ran, Pixin
2017-01-01
The proliferation of human bronchial smooth muscle cells (HBSMCs) is a key pathophysiological component of airway remodeling in chronic obstructive pulmonary disease (COPD) for which pharmacotherapy is limited, and only slight improvements in survival have been achieved in recent decades. Cigarette smoke is a well-recognized risk factor for COPD; however, the pathogenesis of cigarette smoke-induced COPD remains incompletely understood. This study aimed to investigate the mechanisms by which nicotine affects HBSMC proliferation. Cell viability was assessed with a CCK-8 assay. Proliferation was measured by cell counting and EdU immunostaining. Fluorescence calcium imaging was performed to measure intracellular Ca2+ concentration ([Ca2+]i). The results showed that nicotine promotes HBSMC proliferation, which is accompanied by elevated store-operated calcium entry (SOCE), receptor-operated calcium entry (ROCE) and basal [Ca2+]i in HBSMCs. Moreover, we also confirmed that canonical transient receptor potential protein 6 (TRPC6) and α7 nicotinic acetylcholine receptor (α7 nAChR) are involved in nicotine-induced upregulation of cell proliferation. Furthermore, we verified that activation of the PI3K/Akt signaling pathway plays a pivotal role in nicotine-enhanced proliferation and calcium influx in HBSMCs. Inhibition of α7 nAChR significantly decreased Akt phosphorylation levels, and LY294002 inhibited the protein expression levels of TRPC6. Herein, these data provide compelling evidence that calcium entry via the α7 nAChR-PI3K/Akt-TRPC6 signaling pathway plays an important role in the physiological regulation of airway smooth muscle cell proliferation, representing an important target for augmenting airway remodeling. © 2017 The Author(s). Published by S. Karger AG, Basel.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Li Aihua; Cheng Guangli; Zhu Genghui
Ghrelin, a peptide hormone isolated from the stomach, releases growth hormone and stimulates appetite. Ghrelin is also expressed in pancreas, kidneys, cardiovascular system and in endothelial cells. The precise role of ghrelin in endothelial cell functions remains unknown. We examined the expression of ghrelin and its receptor (GHSR1) mRNAs and proteins in human microvascular endothelial cells (HMVEC) and determined whether ghrelin affects in these cells proliferation, migration and in vitro angiogenesis; and whether MAPK/ERK2 signaling is important for the latter action. We found that ghrelin and GHSR1 are constitutively expressed in HMVEC. Treatment of HMVEC with exogenous ghrelin significantly increasedmore » in these cells proliferation, migration, in vitro angiogenesis and ERK2 phosphorylation. MEK/ERK2 inhibitor, PD 98059 abolished ghrelin-induced in vitro angiogenesis. This is First demonstration that ghrelin and its receptor are expressed in human microvascular endothelial cells and that ghrelin stimulates HMVEC proliferation, migration, and angiogenesis through activation of ERK2 signaling.« less
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Kang, Chang-Won; Kim, Nan-Hee; Jung, Huyn Ah; Choi, Hyung-Wook; Kang, Min-Jae; Choi, Jae-Sue; Kim, Gun-Do
2016-04-01
This study is the first report of the antitumor activities of desmethylanhydroicaritin (DMAI) isolated from Sophora flavescens on U87MG cells. Human glioblastoma is one of the most aggressive malignant type of brain tumors and highly diffuses to around normal brain tissues. DMAI showed anti-proliferation effects on U87MG cells at the concentration of 30μM, however did not affect to HEK-293 cells. DMAI induced anti-proliferation effects via ERK/MAPK, PI3K/Akt/mTOR signal pathway and G2/M phase cell cycle arrest. DMAI led to morphological change and inhibition of filapodia formation through regulation of Rac 1 and Cdc 42. In addition, migration and invasion of U87MG cells were inhibited by DMAI via down-regulation of matrix metalloproteinase (MMP) -2 and MMP -9 expressions and activities. Our results suggest that DMAI has a potential as a therapeutic agent against glioblastoma cells. Copyright © 2016 Elsevier B.V. All rights reserved.
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Effect of nickel chloride on cell proliferation.
D'Antò, Vincenzo; Valletta, Rosa; Amato, Massimo; Schweikl, Helmut; Simeone, Michele; Paduano, Sergio; Rengo, Sandro; Spagnuolo, Gianrico
2012-01-01
Metal alloys used in dentistry and in other biomedical fields may release nickel ions in the oral environment. The release of nickel might influence the normal biological and physiological processes, including tissue wound healing, cell growth and proliferation. The aim of this study was to evaluate in vitro the effects of nickel ions on cell cycle, viability and proliferation. Human osteosarcoma cells (U2OS) and human keratinocytes (HaCat) were exposed to different nickel chloride (NiCl(2)) concentrations (0 - 5mM) for various periods exposure. The viability of cultured cells was estimated by flow cytometry using Annexin V-FITC and Propidium Iodide (PI). Cell proliferation was evaluated by using carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) and flow cytometry. Finally, the effects of NiCl(2) on cell cycle were assessed and quantified by flow cytometry. Statistical analysis was performed by means of ANOVA followed by Tukey's test. NiCl(2) induced a dose and time dependent decrease in cell viability. After 24h, 1mM NiCl(2) caused a similar and significant reduction of viability in U2OS and HaCat cells, while higher NiCl(2) concentrations and longer exposure times showed a reduced cytotoxic effect in HaCat as compared to U2OS cells. Exposure to NiCl(2) caused a dose- and time-dependent inhibition of cell proliferation in both cell lines tested, with a prominent effect on U2OS cells. Furthermore, both cell lines exposed to NiCl(2) exhibited significant changes in cell cycle distribution after 24h exposure 2mM NiCl2, as compared to untreated cells (p<0.05). Our results indicate that release of nickel ions may affect cell proliferation. The inhibition of cell growth by NiCl2 is mediated by both cell cycle arrest and by induction of cell death.
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Effect of Nickel Chloride on Cell Proliferation
D’Antò, Vincenzo; Valletta, Rosa; Amato, Massimo; Schweikl, Helmut; Simeone, Michele; Paduano, Sergio; Rengo, Sandro; Spagnuolo, Gianrico
2012-01-01
Objective: Metal alloys used in dentistry and in other biomedical fields may release nickel ions in the oral environment. The release of nickel might influence the normal biological and physiological processes, including tissue wound healing, cell growth and proliferation. The aim of this study was to evaluate in vitro the effects of nickel ions on cell cycle, viability and proliferation. Materials and Methods: Human osteosarcoma cells (U2OS) and human keratinocytes (HaCat) were exposed to different nickel chloride (NiCl2) concentrations (0 - 5mM) for various periods exposure. The viability of cultured cells was estimated by flow cytometry using Annexin V-FITC and Propidium Iodide (PI). Cell proliferation was evaluated by using carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) and flow cytometry. Finally, the effects of NiCl2 on cell cycle were assessed and quantified by flow cytometry. Statistical analysis was performed by means of ANOVA followed by Tukey’s test. Results: NiCl2 induced a dose and time dependent decrease in cell viability. After 24h, 1mM NiCl2 caused a similar and significant reduction of viability in U2OS and HaCat cells, while higher NiCl2 concentrations and longer exposure times showed a reduced cytotoxic effect in HaCat as compared to U2OS cells. Exposure to NiCl2 caused a dose- and time-dependent inhibition of cell proliferation in both cell lines tested, with a prominent effect on U2OS cells. Furthermore, both cell lines exposed to NiCl2 exhibited significant changes in cell cycle distribution after 24h exposure 2mM NiCl2, as compared to untreated cells (p<0.05). Conclusion: Our results indicate that release of nickel ions may affect cell proliferation. The inhibition of cell growth by NiCl2 is mediated by both cell cycle arrest and by induction of cell death. PMID:23198004
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Yoon, Jeongyeon; Ryoo, Sungwoo, E-mail: ryoosw08@kangwon.ac.kr
2013-06-07
Highlights: •Arginase inhibition suppressed proliferation of IL-1β-stimulated VSMCs in dose-dependent manner. •NO production from IL-1β-induced iNOS expression was augmented by arginase inhibition, reducing VSMC proliferation. •Incubation with cGMP analogues abolished IL-1β-dependent proliferation of VSMCs. -- Abstract: We investigated whether arginase inhibition suppressed interleukin (IL)-1β-stimulated proliferation in vascular smooth muscle cells (VSMCs) and the possible mechanisms involved. IL-1β stimulation increased VSMC proliferation, while the arginase inhibitor BEC and transfection of the antisense (AS) oligonucleotide against arginase I decreased VSMC proliferation and was associated with increased protein content of the cell cycle regulator p21Waf1/Cip1. IL-1β incubation induced inducible nitric oxide synthase (iNOS)more » mRNA expression and protein levels in a dose-dependent manner, but did not affect arginase I and II expression. Consistent with this data, IL-1β stimulation resulted in increase in NO production that was significantly augmented by arginase inhibition. The specific iNOS inhibitor 1400W abolished IL-1β-mediated NO production and further accentuated IL-1β-stimulated cell proliferation. Incubation with NO donors GSNO and DETA/NO in the presence of IL-1β abolished VSMCs proliferation and increased p21Waf1/Cip1 protein content. Furthermore, incubation with the cGMP analogue 8-Br-cGMP prevented IL-1β-induced VSMCs proliferation. In conclusion, arginase inhibition augmented iNOS-dependent NO production that resulted in suppression of IL-1β-induced VSMCs proliferation in a cGMP-dependent manner.« less
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Zdanowski, Robert; Skopińska-Różewska, Ewa; Wilczak, Jacek; Borecka, Anna; Lewicka, Aneta
2017-01-01
Angiogenesis plays an important role in many physiological processes, among them the formation of tissues and organs during embryogenesis. A lot of medicinal plants exhibit angiomodulatory properties. This creates the need for a thorough check of whether the plant extracts that we would like to give to pregnant women in order to increase their resistance to bacterial or viral infection will have negative effects on angiogenesis, and consequently on fetal development. This paper seeks to investigate the effect of serum of pregnant and nursing Balb/c mice that received aqueous (RKW) or hydro-alcoholic (RKW-A) R. kirilowii extracts (20 mg/kg), or epigallocatechin (0.2 mg/kg), on the in vitro proliferation and migration of mouse endothelial cell line Heca10. Of the 15 identified polyphenols in the extracts by HPLC, 8 were present in the sera. Chemical analysis revealed higher salidroside, kaempferol, chlorogenic acid, bFGF and VEGF concentration in RKW-A sera than in the sera of RKW group of mice. RKW-A and EGC sera did not affect migration of endothelial cells, however we noted some increase of migrating cells after RKW-sera treatment. RKW and EGC sera did not affect proliferation of endothelial cells. Sera of mothers from RKW-A group impaired the proliferation of endothelial cells in comparison to other groups. These data allow us to assume that Rhodiola kirilowii hydro-alcoholic extract (RKW-A) is potentially able to modulate pre- and post- natal angiogenesis what might influence the development of organs in progeny. Sera of RKW mothers have not harm the proliferation of endothelial cells, despite they also contain antiangiogenic catechins and salidroside. This suggests the existence in RKW-A extract and in RKW-A sera of some other, as yet unidentified substances influencing endothelial cells proliferation. PMID:28680327
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Zhang, Wenyao; Li, Xuezhong; Xu, Tong
Hypernomic secretion of epithelial cytokines has several effects on stromal cells. The contributions of inflammatory epithelial cells to stromal fibroblasts in bovine mammary glands with mastitis remain poorly understood. Here, we established an inflammatory epithelial cell model of bovine mastitis with gram-negative lipopolysaccharide (LPS) and gram-positive lipoteichoic acid (LTA) bacterial cell wall components. We characterized immune responses of mammary stromal fibroblasts induced by inflammatory epithelial cells. Our results showed that inflammatory epithelial cells affected stromal fibroblast characteristics by increasing inflammatory mediator expression, elevating extracellular matrix protein deposition, decreasing proliferation capacity, and enhancing migration ability. The changes in stromal fibroblast proliferationmore » and migration abilities were mediated by signal molecules, such as WNT signal pathway components. LPS- and LTA-induced inflammatory epithelial cells triggered different immune responses in stromal fibroblasts. Thus, in mastitis, bovine mammary gland stromal fibroblasts were affected by inflammatory epithelial cells and displayed inflammation-specific changes, suggesting that fibroblasts play crucial roles in bovine mastitis. - Highlights: • Inflammatory BMEs affect the properties of BMFs during mastitis. • BMEs inhibited the proliferation and promoted the migration of BMFs. • BMEs enhanced secretion of inflammatory mediators and deposition of ECM in BMFs. • Changes of the properties of BMFs were mediated by specific signal molecules.« less
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Park, Jeongyeon; Lee, Hyunyoung; Lee, Hae June; Kim, Gyoo Cheon; Kim, Do Young; Han, Sungbum; Song, Kiwon
2016-01-01
Non-thermal atmospheric pressure plasma (NTAPP) is defined as a partially ionized gas with electrically charged particles at atmospheric pressure. Our study showed that exposure to NTAPP generated in a helium-based dielectric barrier discharge (DBD) device increased the proliferation of adipose tissue-derived stem cells (ASCs) by 1.57-fold on an average, compared with untreated cells at 72 h after initial NTAPP exposure. NTAPP-exposed ASCs maintained their stemness, capability to differentiate into adipocytes but did not show cellular senescence. Therefore, we suggested that NTAPP can be used to increase the proliferation of ASCs without affecting their stem cell properties. When ASCs were exposed to NTAPP in the presence of a nitric oxide (NO) scavenger, the proliferation-enhancing effect of NTAPP was not obvious. Meanwhile, the proliferation of NTAPP-exposed ASCs was not much changed in the presence of scavengers for reactive oxygen species (ROS). Also, Akt, ERK1/2, and NF-κB were activated in ASCs after NTAPP exposure. These results demonstrated that NO rather than ROS is responsible for the enhanced proliferation of ASCs following NTAPP exposure. Taken together, this study suggests that NTAPP would be an efficient tool for use in the medical application of ASCs both in vitro and in vivo. PMID:27991548
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Kakiuchi, Kazue; Taniguchi, Kazumi; Kubota, Hiroshi
2018-05-16
Glial cell line-derived neurotrophic factor (GDNF) is essential for the self-renewal and proliferation of spermatogonial stem cells (SSCs) in mice, rats, and rabbits. Although the key extrinsic factors essential for spermatogonial proliferation in other mammals have not been determined, GDNF is one of the potential candidates. In this study, we isolated porcine GDNF (pGDNF) cDNAs from neonatal testis and generated recombinant pGDNF to investigate its biological activity on gonocytes/undifferentiated spermatogonia, including SSCs. In porcine testis, long and short forms of GDNF transcripts, the counterparts of pre-(α)pro and pre-(β)pro GDNF identified in humans and rodents, were expressed. The two transcripts encode identical mature proteins. Recombinant pGDNF supported proliferation of murine SSCs in culture, and their stem cell activity was confirmed by a transplantation assay. Subsequently, porcine gonocytes/undifferentiated spermatogonia were cultured with pGDNF; however, pGDNF did not affect their proliferation. Furthermore, GDNF expression was localised to the vascular smooth muscle cells, and its cognate receptor GFRA1 expression was negligible during spermatogonial proliferation in the testes. These results indicate that although pGDNF retains structural similarity with those of other mammals and conserves the biological activity on the self-renewal of murine SSCs, porcine SSCs likely require extrinsic factors other than GDNF for their proliferation.
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Scalloped and Yorkie are required for cell cycle re-entry of quiescent cells after tissue damage.
Meserve, Joy H; Duronio, Robert J
2015-08-15
Regeneration of damaged tissues typically requires a population of active stem cells. How damaged tissue is regenerated in quiescent tissues lacking a stem cell population is less well understood. We used a genetic screen in the developing Drosophila melanogaster eye to investigate the mechanisms that trigger quiescent cells to re-enter the cell cycle and proliferate in response to tissue damage. We discovered that Hippo signaling regulates compensatory proliferation after extensive cell death in the developing eye. Scalloped and Yorkie, transcriptional effectors of the Hippo pathway, drive Cyclin E expression to induce cell cycle re-entry in cells that normally remain quiescent in the absence of damage. Ajuba, an upstream regulator of Hippo signaling that functions as a sensor of epithelial integrity, is also required for cell cycle re-entry. Thus, in addition to its well-established role in modulating proliferation during periods of tissue growth, Hippo signaling maintains homeostasis by regulating quiescent cell populations affected by tissue damage. © 2015. Published by The Company of Biologists Ltd.
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Kanda, Shigeru; Miyata, Yasuyoshi; Kanetake, Hiroshi; Smithgall, Thomas E.
2009-01-01
The c-Fes protein-tyrosine kinase is associated with growth and differentiation of hematopoietic, neuronal, vascular endothelial and epithelial cell types. In this study, we investigated whether small interfering RNA (siRNA)-mediated knockdown of c-Fes expression affected proliferation of the human renal carcinoma cell lines, ACHN and VMRC-RCW. Immunofluorescence microscopy showed that c-Fes was expressed in both the cytosol and nuclei of these cells, and siRNA treatment preferentially downregulated c-Fes expression in the cytosol. Knock-down of c-Fes inhibited cellular proliferation in a dose-dependent manner with minimal increase in cell death. c-Fes siRNA treatment also downregulated the phosphorylation of Akt1 on S473 and IKKα on T23, and cyclin D1 expression, enhanced the expression of IκBα, and prevented the nuclear localization of NFκB. Treatment with an NFκB inhibitory peptide (SN50) also blocked the proliferation and nuclear localization of NFκB in these cells. The effect of SN50 treatment was not enhanced by c-Fes siRNA, suggesting that downregulation of c-Fes expression inhibited cell cycle progression through the Akt1/NFκB pathway. In contrast to siRNA-mediated knockdown, ectopic expression of either wild-type or kinase-inactive c-Fes in renal carcinoma cells failed to alter their proliferation in vitro and in vivo. Thus, suppression of proliferation resulting from siRNA-mediated knockdown may depend upon an expression of c-Fes protein rather than its kinase activity. Taken together, our results indicate that downregulation of c-Fes expression may be a potential therapeutic strategy for advanced human renal cell carcinoma and inhibition of its kinase activity as an antiangiogenic therapy does not seem to induce the growth of human renal carcinoma cells. PMID:19082481
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A novel PKC-ι inhibitor abrogates cell proliferation and induces apoptosis in neuroblastoma.
Pillai, Prajit; Desai, Shraddha; Patel, Rekha; Sajan, Mini; Farese, Robert; Ostrov, David; Acevedo-Duncan, Mildred
2011-05-01
Protein Kinase C-iota (PKC-ι), an atypical protein kinase C isoform manifests its potential as an oncogene by targeting various aspects of cancer cells such as growth, invasion and survival. PKC-ι confers resistance to drug-induced apoptosis in cancer cells. The acquisition of drug resistance is a major obstacle to good prognosis in neuroblastoma. The focus of this research was to identify the efficacy of [4-(5-amino-4-carbamoylimidazol-1-yl)-2,3-dihydroxycyclopentyl] methyl dihydrogen phosphate (ICA-1) as a novel PKC-ι inhibitor in neuroblastoma cell proliferation and apoptosis. ICA-1 specifically inhibits the activity of PKC-ι but not that of PKC-zeta (PKC-ζ), the closely related atypical PKC family member. The IC(50) for the kinase activity assay was approximately 0.1μM which is 1000 times less than that of aurothiomalate, a known PKC-ι inhibitor. Cyclin dependent kinase 7 (Cdk7) phosphorylates cyclin dependent kinases (cdks) and promotes cell proliferation. Our data shows that PKC-ι is an in vitro Cdk7 kinase and the phosphorylation of Cdk7 by PKC-ι was potently inhibited by ICA-1. Furthermore, our data shows that neuroblastoma cells proliferate via a PKC-ι/Cdk7/cdk2 cell signaling pathway and ICA-1 mediates its antiproliferative effects by inhibiting this pathway. ICA-1 (0.1μM) inhibited the in vitro proliferation of BE(2)-C neuroblastoma cells by 58% (P=0.01). Additionally, ICA-1 also induced apoptosis in neuroblastoma cells. Interestingly, ICA-1 did not affect the proliferation of normal neuronal cells suggesting its potential as chemotherapeutic with low toxicity. Hence, our results emphasize the potential of ICA-1 as a novel PKC-ι inhibitor and chemotherapeutic agent for neuroblastoma. Published by Elsevier Ltd.
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An, Mingrui; Zhu, Jianhui; Wu, Jing; Cuneo, Kyle C; Lubman, David M
2018-04-06
Circulating microvesicles are able to mediate long-distance cell-cell communications. It is essential to understand how microvesicles from pancreatic cancer act on other cells in the body. In this work, serum-derived microvesicles were isolated from 10 patients with locally advanced pancreatic cancer and healthy controls. Using Cell Transwell and WST-1 reagents, we found that microvesicles from pancreatic cancer accelerated migration and proliferation of PANC-1 cells. Meanwhile, the proliferation of these cancer-microvesicle-treated cells (CMTCs) was affected less by 10 μM of gemcitabine relative to healthy microvesicle-treated cells (HMTCs). Next, we optimized the filter-aided sample preparation method to increase the recovery of protein samples and then applied it to the quantification of the proteome of CMTCs and HMTCs. The peptides were labeled and analyzed by liquid chromatography-tandem mass spectrometry. In total, 4102 proteins were identified, where 35 proteins were up-regulated with 27 down-regulated in CMTCs. We verified the quantitative results of three key proteins CD44, PPP2R1A, and TP53 by Western blot. The Ingenuity Pathway Analysis revealed pathways that cancer microvesicles might participate in to promote cell migration and proliferation. These findings may provide novel clues of treatment for tumorigenesis and metastasis.
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Skeletal muscle satellite cells cultured in simulated microgravity
NASA Technical Reports Server (NTRS)
Molnar, Greg; Hartzell, Charles R.; Schroedl, Nancy A.; Gonda, Steve R.
1993-01-01
Satellite cells are postnatal myoblasts responsible for providing additional nuclei to growing or regenerating muscle cells. Satellite cells retain the capacity to proliferate and differentiate in vitro and therefore provide a useful model to study postnatal muscle development. Most culture systems used to study postnatal muscle development are limited by the two-dimensional (2-D) confines of the culture dish. Limiting proliferation and differentiation of satellite cells in 2-D could potentially limit cell-cell contacts important for developing the level of organization in skeletal muscle obtained in vivo. Culturing satellite cells on microcarrier beads suspended in the High-Aspect-Ratio-Vessel (HARV) designed by NASA provides a low shear, three-dimensional (3-D) environment to study muscle development. Primary cultures established from anterior tibialis muscles of growing rats (approximately 200 gm) were used for all studies and were composed of greater than 75 % satellite cells. Different inoculation densities did not affect the proliferative potential of satellite cells in the HARV. Plating efficiency, proliferation, and glucose utilization were compared between 2-D flat culture and 3-D HARV culture. Plating efficiency (cells attached - cells plated x 100) was similar between the two culture systems. Proliferation was reduced in HARV cultures and this reduction was apparent for both satellite cells and non-satellite cells. Furthermore, reduction in proliferation within the HARV could not be attributed to reduced substrate availability since glucose levels in media from HARV and 2-D cell culture were similar. Morphologically, microcarrier beads within the HARVS were joined together by cells into three-dimensional aggregates composed of greater than 10 beads/aggregate. Aggregation of beads did not occur in the absence of cells. Myotubes were often seen on individual beads or spanning the surface of two beads. In summary, proliferation and differentiation of satellite cells on microcarrier beads within the HARV bioreactor results in a three dimensional level of organization that could provide a more suitable model to study postnatal muscle development.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Alahuhta, Ilkka; Medical Research Center, Oulu University Hospital, Oulu; Aikio, Mari
The turnover of extracellular matrix liberates various cryptic molecules with novel biological activities. Endostatin is an endogenous angiogenesis inhibitor that is derived from the non-collagenous domain of collagen XVIII. Although there are a large number of studies on its anti-tumor effects, the molecular mechanisms are not yet completely understood, and the reasons why endostatin has not been successful in clinical trials are unclear. Research has mostly focused on its anti-angiogenic effect in tumors. Here, we aimed to elucidate how endostatin affects the behavior of aggressive tongue HSC-3 carcinoma cells that were transfected to overproduce endostatin. Endostatin inhibited the invasion ofmore » HSC-3 cells in a 3D collagen–fibroblast model. However, it had no effect on invasion in a human myoma organotypic model, which lacks vital fibroblasts. Recombinant endostatin was able to reduce the Transwell migration of normal fibroblasts, but had no effect on carcinoma associated fibroblasts. Surprisingly, endostatin increased the proliferation and decreased the apoptosis of cancer cells in organotypic models. Also subcutaneous tumors overproducing endostatin grew bigger, but showed less local invasion in nude mice xenografts. We conclude that endostatin affects directly to HSC-3 cells increasing their proliferation, but its net effect on cancer invasion seem to depend on the cellular composition and interactions of tumor microenvironment. - Highlights: • Endostatin affects not only angiogenesis, but also carcinoma cells and fibroblasts. • Endostatin increased carcinoma cell proliferation, but decreased 3D invasion. • The invasion inhibitory effect was sensitive to the microenvironment composition. • Fibroblasts may be a factor regulating the fluctuating roles of endostatin.« less
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Ramos-Solano, Moisés, E-mail: mrsolano84@gmail.com; Programa de Doctorado en Ciencias Biomédica, Centro Universitario de Ciencias de la Salud; Meza-Canales, Ivan D., E-mail: imezacanales@ice.mpg.de
According to the multifactorial model of cervical cancer (CC) causation, it is now recognized that other modifications, in addition to Human papillomavirus (HPV) infection, are necessary for the development of this neoplasia. Among these, it has been proposed that a dysregulation of the WNT pathway might favor malignant progression of HPV-immortalized keratinocytes. The aim of this study was to identify components of the WNT pathway differentially expressed in CC vs. non-tumorigenic, but immortalized human keratinocytes. Interestingly, WNT7A expression was found strongly downregulated in cell lines and biopsies derived from CC. Restoration of WNT7A in CC-derived cell lines using a lentiviralmore » gene delivery system or after adding a recombinant human protein decreases cell proliferation. Likewise, WNT7A silencing in non-tumorigenic cells markedly accelerates proliferation. Decreased WNT7A expression was due to hypermethylation at particular CpG sites. To our knowledge, this is the first study reporting reduced WNT7A levels in CC-derived cells and that ectopic WNT7A restoration negatively affects cell proliferation and migration. - Highlights: • WNT7A is expressed in normal keratinocytes or cervical cells without lesion. • WNT7A is significantly reduced in cervical cancer-derived cells. • Restoration of WNT7A expression in HeLa decreases proliferation and cell migration. • Silencing of WNT7A in HaCaT induces an increased proliferation and migration rate. • Decreased WNT7A expression in this model is due to hypermethylation.« less
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Repression of cell proliferation by miR319-regulated TCP4.
Schommer, Carla; Debernardi, Juan M; Bresso, Edgardo G; Rodriguez, Ramiro E; Palatnik, Javier F
2014-10-01
Leaf development has been extensively studied on a genetic level. However, little is known about the interplay between the developmental regulators and the cell cycle machinery--a link that ultimately affects leaf form and size. miR319 is a conserved microRNA that regulates TCP transcription factors involved in multiple developmental pathways, including leaf development and senescence, organ curvature, and hormone biosynthesis and signaling. Here, we analyze the participation of TCP4 in the control of cell proliferation. A small increase in TCP4 activity has an immediate impact on leaf cell number, by significantly reducing cell proliferation. Plants with high TCP4 levels have a strong reduction in the expression of genes known to be active in G2-M phase of the cell cycle. Part of these effects is mediated by induction of miR396, which represses Growth-Regulating Factor (GRF) transcription factors. Detailed analysis revealed TCP4 to be a direct regulator of MIR396b. However, we found that TCP4 can control cell proliferation through additional pathways, and we identified a direct connection between TCP4 and ICK1/KRP1, a gene involved in the progression of the cell cycle. Our results show that TCP4 can activate different pathways that repress cell proliferation. © The Author 2014. Published by the Molecular Plant Shanghai Editorial Office in association with Oxford University Press on behalf of CSPB and IPPE, SIBS, CAS.
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Regulators of Intestinal Epithelial Migration in Sepsis.
Meng, Mei; Klingensmith, Nathan J; Liang, Zhe; Lyons, John D; Fay, Katherine T; Chen, Ching-Wen; Ford, Mandy L; Coopersmith, Craig M
2018-02-08
The gut is a continuously renewing organ, with cell proliferation, migration and death occurring rapidly under basal conditions. Since the impact of critical illness on cell movement from crypt base to villus tip is poorly understood, the purpose of this study was to determine how sepsis alters enterocyte migration. Wild type, transgenic and knockout mice were injected with 5-bromo-2'deoxyuridine (BrdU) to label cells in S phase before and after the onset of cecal ligation and puncture and were sacrificed at pre-determined endpoints to determine distance proliferating cells migrated up the crypt-villus unit. Enterocyte migration rate was decreased from 24-96 hours following sepsis. BrdU was not detectable on villi 6 days after sham laparotomy, meaning all cells had migrated the length of the gut and been exfoliated into its lumen. However, BrdU positive cells were detectable on villi 10 days after sepsis. Multiple components of gut integrity altered enterocyte migration. Sepsis decreased crypt proliferation, which further slowed enterocyte transit as mice injected with BrdU after the onset of sepsis (decreased proliferation) had slower migration than mice injected with BrdU prior to the onset of sepsis (normal proliferation). Decreasing intestinal apoptosis via gut-specific overexpression of Bcl-2 prevented sepsis-induced slowing of enterocyte migration. In contrast, worsened intestinal hyperpermeability by genetic deletion of JAM-A increased enterocyte migration. Sepsis therefore significantly slows enterocyte migration, and intestinal proliferation, apoptosis and permeability all affect migration time, which can potentially be targeted both genetically and pharmacologically.
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DNA methyltransferase inhibitor CDA-II inhibits myogenic differentiation
DOE Office of Scientific and Technical Information (OSTI.GOV)
Chen, Zirong; Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610; Jin, Guorong
2012-06-08
Highlights: Black-Right-Pointing-Pointer CDA-II inhibits myogenic differentiation in a dose-dependent manner. Black-Right-Pointing-Pointer CDA-II repressed expression of muscle transcription factors and structural proteins. Black-Right-Pointing-Pointer CDA-II inhibited proliferation and migration of C2C12 myoblasts. -- Abstract: CDA-II (cell differentiation agent II), isolated from healthy human urine, is a DNA methyltransferase inhibitor. Previous studies indicated that CDA-II played important roles in the regulation of cell growth and certain differentiation processes. However, it has not been determined whether CDA-II affects skeletal myogenesis. In this study, we investigated effects of CDA-II treatment on skeletal muscle progenitor cell differentiation, migration and proliferation. We found that CDA-II blocked differentiationmore » of murine myoblasts C2C12 in a dose-dependent manner. CDA-II repressed expression of muscle transcription factors, such as Myogenin and Mef2c, and structural proteins, such as myosin heavy chain (Myh3), light chain (Mylpf) and MCK. Moreover, CDA-II inhibited C1C12 cell migration and proliferation. Thus, our data provide the first evidence that CDA-II inhibits growth and differentiation of muscle progenitor cells, suggesting that the use of CDA-II might affect skeletal muscle functions.« less
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Sakai, Chizuko; Arai, Mariko; Tanaka, Sachiko; Onda, Kenji; Sugiyama, Kentaro; Hirano, Toshihiko
2014-11-01
The effects of inorganic and organic arsenicals on proliferation, cell-cycle distribution, and apoptosis of all-transretinoic acid (ATRA)-resistant human promyelocytic leukemia HL-60 (HL-60-R2) cells were herein investigated. Cell proliferation was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell-cycle distribution and apoptotic cells were analyzed by flow cytometry. The 50% inhibitory concentrations (IC50 values) for As2O3 against proliferation of HL-60 and HL-60-R2 cells were 12.2 and 7.2 μM, while those for arsenate were >200 and 62.1 μM, respectively. In contrast, organic methylarsinic acid, dimethylarsonic acid, trimethylarsine oxide, and tetramethylarsonium did not exert any inhibitory effects even at 200 μM. As2O3 and arsenate increased the proportion of apoptotic cells dose-dependently at a concentration range of 5-200 μM. As2O3 did not activate caspase 3/7 in HL-60 and HL-60-R2 cells. As2O3 and arsenate inhibit cell proliferation, affect cell-cycle distribution, and induce apoptosis of ATRA-resistant HL-60-R2 cells. The apoptosis-inducing mechanism appears not to be mediated through caspase3/7. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.
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Effect of miRNA-203 on cervical cancer cells and its underlying mechanism.
Yin, X Z; Zhao, D M; Zhang, G X; Liu, L
2016-09-23
miRNA-203 is involved in the development and progression of various types of cancer. However, its role in cervical cancer remains unclear. The aim of this study was to investigate the effect of miRNA-203 on the proliferation and migration of HeLa cervical cancer cells, as well as survivin expression in these cells. A miRNA-203 primer probe was designed according to a sequence obtained from NCBI. The expression of miRNA-203 in cervical epithelial cells and cervical cancer cells was detected by quantitative reverse transcriptase-polymerase chain reaction. The miRNA-203 expression pattern was compared between these two cell lines. The cervical cancer cells were transfected with miRNA-203 mimic or inhibitor to determine their effects on proliferation and migration. The expression of the miRNA-203 target protein (survivin) was analyzed by western blot. Cervical cancer cells showed reduced miRNA-203 expression compared to cervical epithelial cells. Transfection of miRNA-203 mimic upregulated the expression of miRNA-203, suppressed cell proliferation and migration, and downregulated survivin expression (P < 0.05). However, downregulation of miRNA-203 expression did not affect proliferation, migration, and survivin expression in cervical cancer cells (P > 0.05). In conclusion, upregulation of miRNA-203 in cervical cancer cells inhibits the proliferative and migratory capacities of these cells by downregulating the expression of survivin.
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PRL-3 promotes breast cancer progression by downregulating p14ARF-mediated p53 expression.
Xie, Hua; Wang, Hao
2018-03-01
Prior studies have demonstrated that phosphatase of regenerating liver-3 (PRL-3) serves avital function in cell proliferation and metastasis in breast cancer. However, the molecular mechanisms underlying the function of PRL-3 in breast cancer remain unknown. PRL-3 expression was analyzed in 24 pairs of breast cancer and normal tissues using the reverse transcription-quantitative polymerase chain reaction assay. The results of the present study identified that the expression of PLR-3 in breast cancer tissues was increased 4.2-fold, compared with normal tissues. Notably, overexpression of PRL-3 significantly promoted the proliferation of cancer cells and inhibited endogenous p53 expression by downregulating the expression level of p14 alternate reading frame (p14 ARF ). In addition, decreased expression levels of PRL-3 resulted in decreased breast cancer cell proliferation and increased expression level of p14 ARF . These results suggested that PRL-3 enhances cell proliferation by downregulating p14 ARF expression, which results in decreased levels ofp53. The results of the present study demonstrated that PRL-3 promotes tumor proliferation by affecting the p14 ARF -p53 axis, and that it may serve as a prognostic marker for patients with breast cancer.
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Moraga, Ana; Pradillo, Jesús M; García-Culebras, Alicia; Palma-Tortosa, Sara; Ballesteros, Ivan; Hernández-Jiménez, Macarena; Moro, María A; Lizasoain, Ignacio
2015-05-10
Aging is not just a risk factor of stroke, but it has also been associated with poor recovery. It is known that stroke-induced neurogenesis is reduced but maintained in the aged brain. However, there is no consensus on how neurogenesis is affected after stroke in aged animals. Our objective is to determine the role of aging on the process of neurogenesis after stroke. We have studied neurogenesis by analyzing proliferation, migration, and formation of new neurons, as well as inflammatory parameters, in a model of cerebral ischemia induced by permanent occlusion of the middle cerebral artery in young- (2 to 3 months) and middle-aged mice (13 to 14 months). Aging increased both microglial proliferation, as shown by a higher number of BrdU(+) cells and BrdU/Iba1(+) cells in the ischemic boundary and neutrophil infiltration. Interestingly, aging increased the number of M1 monocytes and N1 neutrophils, consistent with pro-inflammatory phenotypes when compared with the alternative M2 and N2 phenotypes. Aging also inhibited (subventricular zone) SVZ cell proliferation by decreasing both the number of astrocyte-like type-B (prominin-1(+)/epidermal growth factor receptor (EGFR)(+)/nestin(+)/glial fibrillary acidic protein (GFAP)(+) cells) and type-C cells (prominin-1(+)/EGFR(+)/nestin(-)/Mash1(+) cells), and not affecting apoptosis, 1 day after stroke. Aging also inhibited migration of neuroblasts (DCX(+) cells), as indicated by an accumulation of neuroblasts at migratory zones 14 days after injury; consistently, aged mice presented a smaller number of differentiated interneurons (NeuN(+)/BrdU(+) and GAD67(+) cells) in the peri-infarct cortical area 14 days after stroke. Our data confirm that stroke-induced neurogenesis is maintained but reduced in aged animals. Importantly, we now demonstrate that aging not only inhibits proliferation of specific SVZ cell subtypes but also blocks migration of neuroblasts to the damaged area and decreases the number of new interneurons in the cortical peri-infarct area. Thus, our results highlight the importance of using aged animals for translation to clinical studies.
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Xu, Yu-Dong; Wang, Yu; Yin, Lei-Miao; Peng, Ling-Ling; Park, Gyoung-Hee; Yang, Yong-Qing
2017-06-21
Airway remodeling is a key feature of asthma, characterized by increased proliferation of airway smooth muscle cells (ASMCs). S100A8 is a calcium-binding protein with a potential to regulate cell proliferation. Here, the effect of exogenous S100A8 protein on the proliferation of ASMCs induced by platelet-derived growth factor (PDGF) and the underlying molecular mechanism was investigated. Rat ASMCs were cultured with or without a neutralizing antibody to the receptor for advanced glycation end-products (RAGE), a potential receptor for S100A8 protein. Purified recombinant rat S100A8 protein was then added into the cultured cells, and the proliferation of ASMCs induced by PDGF was detected by colorimetric-based WST-8 assay and ampedance-based xCELLigence proliferation assay. The expression levels of RAGE in ASMCs were analyzed using western blotting assay. Results showed that exogenous S100A8 inhibited the PDGF-induced proliferation of rat ASMCs in a dose-dependent manner with the maximal effect at 1 μg/ml in vitro. Furthermore, when ASMCs was pre-treated with anti-RAGE neutralizing antibody, the inhibitory effect of S100A8 on PDGF-induced proliferation was significantly suppressed. In addition, neither the treatment with S100A8 or PDGF alone nor the pre-treatment with rS100A8 followed by PDGF stimulation affected the expression levels of RAGE. Our study demonstrated that S100A8 inhibits PDGF-induced ASMCs proliferation in a manner dependent on membrane receptor RAGE.
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Hoose, Scott A.; Duran, Camille; Malik, Indranil; Eslamfam, Shabnam; Shasserre, Samantha C.; Downing, S. Sabina; Hoover, Evelyn M.; Dowd, Katherine E.; Smith, Roger; Polymenis, Michael
2012-01-01
Screening chemical libraries to identify compounds that affect overall cell proliferation is common. However, in most cases, it is not known whether the compounds tested alter the timing of particular cell cycle transitions. Here, we evaluated an FDA-approved drug library to identify pharmaceuticals that alter cell cycle progression in yeast, using DNA content measurements by flow cytometry. This approach revealed strong cell cycle effects of several commonly used pharmaceuticals. We show that the antilipemic gemfibrozil delays initiation of DNA replication, while cells treated with the antidepressant fluoxetine severely delay progression through mitosis. Based on their effects on cell cycle progression, we also examined cell proliferation in the presence of both compounds. We discovered a strong suppressive interaction between gemfibrozil and fluoxetine. Combinations of interest among diverse pharmaceuticals are difficult to identify, due to the daunting number of possible combinations that must be evaluated. The novel interaction between gemfibrozil and fluoxetine suggests that identifying and combining drugs that show cell cycle effects might streamline identification of drug combinations with a pronounced impact on cell proliferation. PMID:22567160
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Hoose, Scott A; Duran, Camille; Malik, Indranil; Eslamfam, Shabnam; Shasserre, Samantha C; Downing, S Sabina; Hoover, Evelyn M; Dowd, Katherine E; Smith, Roger; Polymenis, Michael
2012-01-01
Screening chemical libraries to identify compounds that affect overall cell proliferation is common. However, in most cases, it is not known whether the compounds tested alter the timing of particular cell cycle transitions. Here, we evaluated an FDA-approved drug library to identify pharmaceuticals that alter cell cycle progression in yeast, using DNA content measurements by flow cytometry. This approach revealed strong cell cycle effects of several commonly used pharmaceuticals. We show that the antilipemic gemfibrozil delays initiation of DNA replication, while cells treated with the antidepressant fluoxetine severely delay progression through mitosis. Based on their effects on cell cycle progression, we also examined cell proliferation in the presence of both compounds. We discovered a strong suppressive interaction between gemfibrozil and fluoxetine. Combinations of interest among diverse pharmaceuticals are difficult to identify, due to the daunting number of possible combinations that must be evaluated. The novel interaction between gemfibrozil and fluoxetine suggests that identifying and combining drugs that show cell cycle effects might streamline identification of drug combinations with a pronounced impact on cell proliferation.
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Yuasa, Masato; Yamada, Tsuyoshi; Taniyama, Takashi; Masaoka, Tomokazu; Xuetao, Wei; Yoshii, Toshitaka; Horie, Masaki; Yasuda, Hiroaki; Uemura, Toshimasa; Okawa, Atsushi; Sotome, Shinichi
2015-01-01
We evaluated whether dexamethasone augments the osteogenic capability of bone marrow-derived stromal cells (BMSCs) and muscle tissue-derived stromal cells (MuSCs), both of which are thought to contribute to ectopic bone formation induced by bone morphogenetic protein-2 (BMP-2), and determined the underlying mechanisms. Rat BMSCs and MuSCs were cultured in growth media with or without 10-7 M dexamethasone and then differentiated under osteogenic conditions with dexamethasone and BMP-2. The effects of dexamethasone on cell proliferation and osteogenic differentiation, and also on ectopic bone formation induced by BMP-2, were analyzed. Dexamethasone affected not only the proliferation rate but also the subpopulation composition of BMSCs and MuSCs, and subsequently augmented their osteogenic capacity during osteogenic differentiation. During osteogenic induction by BMP-2, dexamethasone also markedly affected cell proliferation in both BMSCs and MuSCs. In an in vivo ectopic bone formation model, bone formation in muscle-implanted scaffolds containing dexamethasone and BMP-2 was more than two fold higher than that in scaffolds containing BMP-2 alone. Our results suggest that dexamethasone potently enhances the osteogenic capability of BMP-2 and may thus decrease the quantity of BMP-2 required for clinical application, thereby reducing the complications caused by excessive doses of BMP-2. Highlights: 1. Dexamethasone induced selective proliferation of bone marrow- and muscle-derived cells with higher differentiation potential. 2. Dexamethasone enhanced the osteogenic capability of bone marrow- and muscle-derived cells by altering the subpopulation composition. 3. Dexamethasone augmented ectopic bone formation induced by bone morphogenetic protein-2. PMID:25659106
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Blum, Walter; Pecze, László; Felley-Bosco, Emanuela; Schwaller, Beat
2015-12-22
The Ca(2+)-binding protein calretinin is currently used as a positive marker for identifying epithelioid malignant mesothelioma (MM) and reactive mesothelium, but calretinin's likely role in mesotheliomagenesis remains unclear. Calretinin protects immortalized mesothelial cells in vitro from asbestos-induced cytotoxicity and thus might be implicated in mesothelioma formation. To further investigate calretinin's putative role in the early steps of MM generation, primary mesothelial cells from calretinin knockout (CR-/-) and wildtype (WT) mice were compared. Primary mouse mesothelial cells from WT and CR-/- mice were investigated with respect to morphology, marker proteins, proliferation, cell cycle parameters and mobility in vitro. Overexpression of calretinin or a nuclear-targeted variant was achieved by a lentiviral expression system. CR-/- mice have a normal mesothelium and no striking morphological abnormalities compared to WT animals were noted. Primary mouse mesothelial cells from both genotypes show a typical "cobblestone-like" morphology and express mesothelial markers including mesothelin. In cells from CR-/- mice in vitro, we observed more giant cells and a significantly decreased proliferation rate. Up-regulation of calretinin in mesothelial cells of both genotypes increases the proliferation rate and induces a cobblestone-like epithelial morphology. The length of the S/G2/M phase is unchanged, however the G1 phase is clearly prolonged in CR-/- cells. They are also much slower to close a scratch in a confluent cell layer (2D-wound assay). In addition to a change in cell morphology, an increase in proliferation and mobility is observed, if calretinin overexpression is targeted to the nucleus. Thus, both calretinin and nuclear-targeted calretinin increase mesothelial cell proliferation and consequently, speed up the scratch-closure time. The increased rate of scratch closure in WT cells is the result of two processes: an increased proliferation rate and augmented cell mobility of the border cells migrating towards the empty space. We hypothesize that the differences in proliferation and mobility between WT and CR-/- mesothelial cells are the likely result from differences in their developmental trajectories. The mechanistic understanding of the function of calretinin and its putative implication in signaling pathways in normal mesothelial cells may help understanding its role during the processes that lead to mesothelioma formation and could possibly open new avenues for mesothelioma therapy, either by directly targeting calretinin expression or indirectly by targeting calretinin-mediated downstream signaling.
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Wang, Jue; Huang, Weicong; Wu, Yihe; Hou, Jianfeng; Nie, Yu; Gu, Haiyong; Li, Jun; Hu, Shengshou
2012-01-01
The enhanced proliferation of mesenchymal stem cells (MSCs) can be helpful for the clinical translation of cell therapy. Low-level laser irradiation (LLLI) has been demonstrated as regulating MSC proliferation. MicroRNAs (miRNAs) are involved in various pathophysiologic processes in stem cells, but the role of miRNAs in the LLLI-based promotion of MSC proliferation remains unclear. We found that the proliferation level and cell cycle-associated genes in MSCs were increased after LLLI treatment in a time-dependent manner. Microarray assays revealed subsets of miRNAs to be differentially regulated, and these dynamic changes were confirmed by quantitative real-time polymerase chain reaction (qRT-PCR) after LLLI. miR-193 was the most highly up-regulated miRNA, and the change in it was related with the proliferation level. Gain-loss function experiments demonstrated that miR-193 could regulate the proliferation of MSCs, including human's and rat's, but could not affect the apoptosis and differentiation level. Blockade of miR-193 repressed the MSC proliferation induced by LLLI. By qRT-PCR, we found that miR-193, in particular, regulated cyclin-dependent kinase 2 (CDK2) expression. Bioinformatic analyses and luciferase reporter assays revealed that inhibitor of growth family, member 5 (ING5) could be the best target of miR-193 to functionally regulate proliferation and CDK2 activity, and the mRNA and protein level of ING5 was regulated by miR-193. Furthermore, the ING5 inhibited by small interfering RNA (siRNA) could up-regulate the proliferation of MSCs and the expression of CDK2. Taken together, these results strongly suggest that miR-193 plays a critical part in MSC proliferation in response to LLLI stimulation, which is potentially amenable to therapeutic manipulation for clinical application. PMID:22384930
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Umeda, Hirotsugu; Mano, Takamitsu; Harada, Koji; Tarannum, Ferdous; Ueyama, Yoshiya
2017-08-01
We have already reported that the apatite coating of titanium by the blast coating (BC) method could show a higher rate of bone contact from the early stages in vivo, when compared to the pure titanium (Ti) and the apatite coating of titanium by the flame spraying (FS) method. However, the detailed mechanism by which BC resulted in satisfactory bone contact is still unknown. In the present study, we investigated the importance of various factors including cell adhesion factor in osteoblast proliferation and differentiation that could affect the osteoconductivity of the BC disks. Cell proliferation assay revealed that Saos-2 could grow fastest on BC disks, and that a spectrophotometric method using a LabAssay TM ALP kit showed that ALP activity was increased in cells on BC disks compared to Ti disks and FS disks. In addition, higher expression of E-cadherin and Fibronectin was observed in cells on BC disks than Ti disks and FS disks by relative qPCR as well as Western blotting. These results suggested that the expression of cell-adhesion factors, proliferation and differentiation of osteoblast might be enhanced on BC disks, which might result higher osteoconductivity.
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Ethanol immunosuppression in vitro
DOE Office of Scientific and Technical Information (OSTI.GOV)
Kaplan, D.R.
Ethanol in concentrations equivalent to levels achieved by the ingestion of moderate to large amounts of alcoholic beverages has been shown to inhibit mitogen and anti-CD3 stimulated human T lymphocyte proliferation. This inhibition was monophasic suggesting that ethanol affected a single limiting component of T cell proliferation. In experiments designed to test the effect of ethanol on various aspects of proliferation, it was demonstrated that ethanol inhibited the capacity of exogenously supplied interleukin 2 to stimulate proliferation of T cells that had previously acquired interleukin 2 receptors in a monophasic, dose-dependent manner. Moreover, there was no suppression of interleukin 2more » production or interleukin 2 receptor acquisition. Thus, ethanol was shown to mediate immunosuppression by a mechanism specific to one component of proliferation. Additive inhibition of T cell proliferation was seen with ethanol plus cyclosporin A which inhibits interleukin 2 production. The level of inhibition with 250 ng/ml cyclosporin A alone was equivalent to the level seen with 62 ng/ml cyclosporin A plus 20 mM (94 mg%) ethanol. Ethanol also suppressed an immune effector mechanism. NK cytotoxicity was depressed in a monophasic, dose-dependent manner. Thus, ethanol might be considered as a possible adjunct in immunosuppressive therapy.« less
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CCCTC-binding Factor Mediates Effects of Glucose On Beta Cell Survival
Tsui, Shanli; Dai, Wei; Lu, Luo
2013-01-01
Objectives Pancreatic islet β-cell survival is important in regulating insulin activities and maintaining glucose homeostasis. Recently, Pax6 has been shown to be essential for many vital functions in β-cells, though the molecular mechanisms of its regulation in β-cells remain unclear. The present study investigates the novel effects of glucose- and insulin-induced CTCF activity on Pax6 gene expression as well as the subsequent effects of insulin-activated signaling pathways on β-cell proliferation. Material and methods Pancreatic β-TC-1-6 cells were cultured in DMEM medium and stimulated with high concentrations of glucose (5 to 125 mM) and cell viability was assessed by MTT assays. The effect of CTCF on Pax6 was evaluated in high glucose-induced and CCCTC-binding Factor (CTCF)/Erk suppressed cells by promoter reporter and Western analyses. Results Increases in glucose and insulin concentrations up-regulated CTCF and consequently down-regulated Pax6 in β-cell survival and proliferation. Knocking-down CTCF directly affected Pax6 transcription through CTCF binding and blocked the response to glucose. Altered Erk activity mediated the effects of CTCF on controlling Pax6 expression, which partially regulates β-cell proliferation. Conclusions CTCF functions as a molecular mediator between insulin-induced upstream Erk signaling and Pax6 expression in pancreatic β-cells. This pathway may contribute to regulation of β-cell survival and proliferation. PMID:24354619
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Liu, Quanliang; Wang, Xinghuan
2013-11-01
Transient receptor potential vanilloid 2 (TRPV2), a nonselective cation channel, has become an attractive target gene for tumor studies due to its wide range of physiological and pathological functions. However, its specific role in bladder cancer development and progression remains unclear. The aim of the present study was to investigate the effects of TRPV2 on the proliferation, migration and invasion of 5637 bladder cancer cells in vitro . Rat TRPV2 cDNA was transfected into 5637 bladder cancer cells and changes in the behavior of the cells were detected. It was observed that TRPV2 enhanced bladder cancer cell migration and invasion; however, it did not affect cell proliferation in vitro . TRPV2 activity, which may be mediated by direct matrix metalloproteinase 2 (MMP2) regulation, is important in bladder tumor development and progression. The results of this study suggest that TRPV2 channels are a potential therapeutic target for bladder carcinoma.
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LIU, QUANLIANG; WANG, XINGHUAN
2013-01-01
Transient receptor potential vanilloid 2 (TRPV2), a nonselective cation channel, has become an attractive target gene for tumor studies due to its wide range of physiological and pathological functions. However, its specific role in bladder cancer development and progression remains unclear. The aim of the present study was to investigate the effects of TRPV2 on the proliferation, migration and invasion of 5637 bladder cancer cells in vitro. Rat TRPV2 cDNA was transfected into 5637 bladder cancer cells and changes in the behavior of the cells were detected. It was observed that TRPV2 enhanced bladder cancer cell migration and invasion; however, it did not affect cell proliferation in vitro. TRPV2 activity, which may be mediated by direct matrix metalloproteinase 2 (MMP2) regulation, is important in bladder tumor development and progression. The results of this study suggest that TRPV2 channels are a potential therapeutic target for bladder carcinoma. PMID:24223658
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Thompson, Pamela D.; Sakwe, Amos; Koumangoye, Rainelli
2014-02-15
This study was performed to identify the potential role of Alpha-2 Heremans Schmid Glycoprotein (AHSG) in Head and Neck Squamous Cell Carcinoma (HNSCC) tumorigenesis using an HNSCC cell line model. HNSCC cell lines are unique among cancer cell lines, in that they produce endogenous AHSG and do not rely, solely, on AHSG derived from serum. To produce our model, we performed a stable transfection to down-regulate AHSG in the HNSCC cell line SQ20B, resulting in three SQ20B sublines, AH50 with 50% AHSG production, AH20 with 20% AHSG production and EV which is the empty vector control expressing wild-type levels ofmore » AHSG. Utilizing these sublines, we examined the effect of AHSG depletion on cellular adhesion, proliferation, migration and invasion in a serum-free environment. We demonstrated that sublines EV and AH50 adhered to plastic and laminin significantly faster than the AH20 cell line, supporting the previously reported role of exogenous AHSG in cell adhesion. As for proliferative potential, EV had the greatest amount of proliferation with AH50 proliferation significantly diminished. AH20 cells did not proliferate at all. Depletion of AHSG also diminished cellular migration and invasion. TGF-β was examined to determine whether levels of the TGF-β binding AHSG influenced the effect of TGF-β on cell signaling and proliferation. Whereas higher levels of AHSG blunted TGF-β influenced SMAD and ERK signaling, it did not clearly affect proliferation, suggesting that AHSG influences on adhesion, proliferation, invasion and migration are primarily due to its role in adhesion and cell spreading. The previously reported role of AHSG in potentiating metastasis via protecting MMP-9 from autolysis was also supported in this cell line based model system of endogenous AHSG production in HNSCC. Together, these data show that endogenously produced AHSG in an HNSCC cell line, promotes in vitro cellular properties identified as having a role in tumorigenesis. Highlights: • Head and neck squamous cell carcinoma cell lines synthesize and secret AHSG. • AHSG depleted cell lines are significantly inhibited in their ability to proliferate, adhere, migrate, invade and protect MMP-9. • Human AHSG and bovine fetuin-A are functionally equivalent in regards to growth promotion of cancer cell lines.« less
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Kwon, Sora; Jeon, Ji-Sook; Ahn, Curie
Rapamycin, a specific inhibitor of mTOR used extensively as an immunosuppressant, has been expanded recently to cancer therapy, because the mTOR signal is known to be up-regulated in various cancer cells including hepatocellular carcinoma (HCC) cells. In spite of extensive efforts to employ mTOR inhibitors as anti-HCC therapy, they have not yet been approved by the FDA. Because of the heterogeneity and complexity of molecular signaling in HCC, suitable biomarkers should be identified or discovered to improve clinical efficacy of mTOR-specific inhibitors to HCC cells. In this study, the effect of rapamycin was investigated on two different HCC cell lines,more » Huh7 cells and HepG2 cells. Rapamycin was found to inhibit the proliferation of Huh7 cells but not of HepG2 cells. Moreover, it was found that rapamycin can up-regulate p53 at the protein level, but not affect its transcript. To understand the critical role of p53 in the rapamycin effect, knock-down experiments were performed using small-interfering RNAs (siRNAs). The anti-proliferative effect of rapamycin on Huh7 cells clearly disappeared after blocking p53 production with siRNA, which indicates that p53 is a critical factor in the anti-proliferative effect of rapamycin in HCC cells. The over-expression system of p53 was also employed to mimic the effect of rapamycin and found that cell proliferation was clearly down-regulated by p53 over-expression. Finally, we found that the extracellular signal-regulated kinase 1/2 (ERK1/2) signal was regulated by p53 whose expression was induced by rapamycin. Overall, this study demonstrates that rapamycin inhibited the proliferation of Huh7 cells by up-regulating the expression of p53 and down-regulating the ERK1/2 signal, indicating that p53 is a useful biomarker for anti-cancer therapy using the specific inhibitor of mTOR signal, rapamycin, against hepatocellular carcinoma cells. - Highlights: • Rapamycin inhibits the proliferation of hepatocellular carcinoma cells depending on the expression of p53. • Rapamycin up-regulates p53 at the protein level, but not affect its transcript. • The up-regulation of p53 expression by rapamycin inhibits ERK signal.« less
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Duarte-Guterman, Paula; Lieblich, Stephanie E.; Chow, Carmen; Galea, Liisa A. M.
2015-01-01
Estradiol increases cell proliferation in the dentate gyrus of the female rodent but it is not known whether the G protein-coupled estrogen receptor (GPER), a membrane receptor, is involved in this process, nor whether there are regional differences in estradiol’s effects on cell proliferation. Thus, we investigated whether estradiol exerts its effects on cell proliferation in the dorsal and ventral dentate gyrus through GPER, using the GPER agonist, G1, and antagonist, G15. Ovariectomized adult female rats received a single injection of either: 17β-estradiol (10 μg), G1 (0.1, 5, 10 μg), G15 (40 μg), G15 and estradiol, or vehicle (oil, DMSO, or oil+DMSO). After 30 min, animals received an injection of bromodeoxyuridine (BrdU) and were perfused 24 h later. Acute treatment with estradiol increased, while the GPER agonist G1 (5 μg) decreased, the number of BrdU+ cells in the dentate gyrus relative to controls. The GPER antagonist, G15 increased the number of BrdU+ cells relative to control in the dorsal region and decreased the number of BrdU+ cells in the ventral region. However, G15 treatment in conjunction with estradiol partially eliminated the estradiol-induced increase in cell proliferation in the dorsal dentate gyrus. Furthermore, G1 decreased the expression of GPER in the dentate gyrus but not the CA1 and CA3 regions of the hippocampus. In summary, we found that activation of GPER decreased cell proliferation and GPER expression in the dentate gyrus of young female rats, presenting a potential and novel estrogen-independent role for this receptor in the adult hippocampus. PMID:26075609
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Duarte-Guterman, Paula; Lieblich, Stephanie E; Chow, Carmen; Galea, Liisa A M
2015-01-01
Estradiol increases cell proliferation in the dentate gyrus of the female rodent but it is not known whether the G protein-coupled estrogen receptor (GPER), a membrane receptor, is involved in this process, nor whether there are regional differences in estradiol's effects on cell proliferation. Thus, we investigated whether estradiol exerts its effects on cell proliferation in the dorsal and ventral dentate gyrus through GPER, using the GPER agonist, G1, and antagonist, G15. Ovariectomized adult female rats received a single injection of either: 17β-estradiol (10 μg), G1 (0.1, 5, 10 μg), G15 (40 μg), G15 and estradiol, or vehicle (oil, DMSO, or oil+DMSO). After 30 min, animals received an injection of bromodeoxyuridine (BrdU) and were perfused 24 h later. Acute treatment with estradiol increased, while the GPER agonist G1 (5 μg) decreased, the number of BrdU+ cells in the dentate gyrus relative to controls. The GPER antagonist, G15 increased the number of BrdU+ cells relative to control in the dorsal region and decreased the number of BrdU+ cells in the ventral region. However, G15 treatment in conjunction with estradiol partially eliminated the estradiol-induced increase in cell proliferation in the dorsal dentate gyrus. Furthermore, G1 decreased the expression of GPER in the dentate gyrus but not the CA1 and CA3 regions of the hippocampus. In summary, we found that activation of GPER decreased cell proliferation and GPER expression in the dentate gyrus of young female rats, presenting a potential and novel estrogen-independent role for this receptor in the adult hippocampus.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Ryu, Min Sook; Woo, Min-Yeong; Department of Biomedical Sciences, The Graduate School, Ajou University
2014-10-01
In vivo and in vitro effects of TIS21 gene on the mature T cell activation and antitumor activities were explored by employing MO5 melanoma orthograft and splenocytes isolated from the TIS21-knockout (KO) mice. Proliferation and survival of mature T cells were significantly increased in the KO than the wild type (WT) cells, indicating that TIS21 inhibits the rate of mature T cell proliferation and its survival. In MO5 melanoma orthograft model, the KO mice recruited much more CD8{sup +} T cells into the tumors at around day 14 after tumor cell injection along with reduced tumor volumes compared with themore » WT. The increased frequency of granzyme B{sup +} CD8{sup +} T cells in splenocytes of the KO mice compared with the WT may account for antitumor-immunity of TIS21 gene in the melanoma orthograft. In contrast, reduced frequencies of CD107a{sup +} CD8{sup +} T cells in the splenocytes of KO mice may affect the loss of CD8{sup +} T cell infiltration in the orthograft at around day 19. These results indicate that TIS21 exhibits antiproliferative and proapoptotic effects in mature T cells, and differentially affects the frequencies of granzyme B{sup +} CD8{sup +} T-cells and CD107a{sup +} CD8{sup +} T-cells, thus transiently regulating in vivo anti-tumor immunity. - Highlights: • Constitutive expression of TIS21 in splenocytes and upregulation by TCR stimulation. • Proliferation of mature T-cells in spleen of TIS21KO mice after TCR stimulation. • Inhibition of cell death in mature T-cells of TIS21KO mice compared with the wild type. • Inhibition of melanoma growth in TIS21KO mice and CD8{sup +} T cell infiltration in tumor. • Reduction of CD 107{sup +}CD8{sup +} T cells, but increased granzyme B{sup +} CD8{sup +} T cells in TIS21KO mice.« less
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Bouamama, Samia; Merzouk, Hafida; Medjdoub, Amel; Merzouk-Saidi, Amel; Merzouk, Sid Ahmed
2017-06-01
Aging is an inevitable biological event that is associated with immune alterations. These alterations are related to increased cellular oxidative stress and micronutrient deficiency. Antioxidant supplementation could improve these age-related abnormalities. The aim of this study was to determine in vitro effects of vitamin A, vitamin C, vitamin E, and nicotinamide adenine dinucleotide (NADH) on T cell proliferation, cytokine release, and cell redox status in the elderly compared with young adults. Peripheral blood lymphocytes were isolated using a density gradient of Histopaque. They were cultured in vitro and stimulated with concanavalin A in the presence or absence of vitamins. Cell proliferation was determined by conducting MTT assays, and based on interleukin-2 and interleukin-4 secretions. Cell oxidant/antioxidant balance was assessed by assaying reduced glutathione (GSH), malondialdehyde, carbonyl protein levels, and catalase activity. The present study demonstrated that T-lymphocyte proliferation was decreased with aging and was associated with cytokine secretion alterations, GSH depletion, and intracellular oxidative stress. In the elderly, vitamin C, vitamin E, and NADH significantly improved lymphocyte proliferation and mitigated cellular oxidative stress, whereas vitamin A did not affect cell proliferation or cell redox status. In conclusion, vitamin C, vitamin E, and NADH supplementation improved T-lymphocytes response in the elderly, and could contribute to the prevention of age-related immune alterations. Consumption of food items containing these vitamins is recommended, and further investigation is necessary to evaluate the effect of vitamin supplementation in vivo.
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Torres, Cristian Gabriel; Pino, Ana María; Sierralta, Walter Daniel
2009-06-01
The effects of estradiol (E2) and of an AFP-derived cyclized peptide (cP) on the proliferation of primary cultures of cancer cells isolated from spontaneous canine mammary tumors were studied. The cellular response to E2 and cP was related to the expression of estradiol receptor (isoforms alpha and beta). In ER-positive cells, 2 nM estradiol increased cell proliferation and the phosphorylation of ERK1/2; 2 microg/ml cP inhibited all these effects. Estradiol also increased HER2 immunoreactivity in ER-positive cells, an effect that was reverted to its basal values by cP. Estradiol stimulated in these cells the release of MMP2 and MMP9 and the shedding of HB-EGF, effects that the cP did not affect. ER-negative cells were refractory to estradiol or cP. All canine mammary tumor cells in culture responded to treatments analogously to human mammary cancer cells. Our results support the proposal of cP as a new, potentially effective therapeutic agent for the management of mammary cancer.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Yamamura, Hideto; Suzuki, Yoshiaki; Yamamura, Hisao
The blood-brain barrier (BBB) is mainly composed of brain capillary endothelial cells (BCECs), astrocytes and pericytes. Brain ischemia causes hypoxic encephalopathy and damages BBB. However, it remains still unclear how hypoxia affects BCECs. In the present study, t-BBEC117 cells, an immortalized bovine brain endothelial cell line, were cultured under hypoxic conditions at 4–5% oxygen for 72 h. This hypoxic stress caused hyperpolarization of resting membrane potential. Patch-clamp recordings revealed a marked increase in Ba{sup 2+}-sensitive inward rectifier K{sup +} current in t-BBEC117 cells after hypoxic culture. Western blot and real-time PCR analyses showed that Kir2.1 expression was significantly up-regulated at protein level butmore » not at mRNA level after the hypoxic culture. Ca{sup 2+} imaging study revealed that the hypoxic stress enhanced store-operated Ca{sup 2+} (SOC) entry, which was significantly reduced in the presence of 100 μM Ba{sup 2+}. On the other hand, the expression of SOC channels such as Orai1, Orai2, and transient receptor potential channels was not affected by hypoxic stress. MTT assay showed that the hypoxic stress significantly enhanced t-BBEC117 cell proliferation, which was inhibited by approximately 60% in the presence of 100 μM Ba{sup 2+}. We first show here that moderate cellular stress by cultivation under hypoxic conditions hyperpolarizes membrane potential via the up-regulation of functional Kir2.1 expression and presumably enhances Ca{sup 2+} entry, resulting in the facilitation of BCEC proliferation. These findings suggest potential roles of Kir2.1 expression in functional changes of BCECs in BBB following ischemia. -- Highlights: •Hypoxic culture of brain endothelial cells (BEC) caused membrane hyperpolarization. •This hyperpolarization was due to the increased expression of Kir2.1 channels. •Hypoxia enhanced store-operated Ca{sup 2+} (SOC) entry via Kir2.1 up-regulation. •Expression levels of putative SOC channels were not affected by hypoxia. •Kir2.1 up-regulation is responsible for hypoxia-enhanced BEC proliferation.« less
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Sarkar, D; Singh, S K
2017-07-01
Thyroid hormones (THs) play an important role in maintaining the link between metabolism and reproduction and the altered THs status is associated with induction of oxidative stress in various organs like brain, heart, liver and testis. Further, reactive oxygen species play a pivotal role in regulation of glucose homeostasis in several organs, and glucose utilization by Leydig cells is essential for testosterone biosynthesis and thus is largely dependent on glucose transporter 8 (GLUT8). Glucose uptake by Sertoli cells is mediated through glucose transporter 3 (GLUT3) under the influence of THs to meet energy requirement of developing germ cells. THs also modulate level of gap junctional protein such as connexin 43 (Cx43), a potential regulator of cell proliferation and apoptosis in the seminiferous epithelium. Although the role of transient neonatal hypothyroidism in adult testis in terms of testosterone production is well documented, the effect of THs deficiency in early developmental period and its role in testicular glucose homeostasis and oxidative stress with reference to Cx43 in immature mice remain unknown. Therefore, the present study was conducted to evaluate the effect of neonatal hypothyroidism on testicular glucose homeostasis and oxidative stress at postnatal days (PND) 21 and 28 in relation to GLUT3, GLUT8 and Cx43. Hypothyroidism induced by 6-propyl-2-thiouracil (PTU) markedly decreased testicular glucose level with considerable reduction in expression level of GLUT3 and GLUT8. Likewise, lactate dehydrogenase (LDH) activity and intratesticular concentration of lactate were also decreased in hypothyroid mice. There was also a rise in germ cell apoptosis with increased expression of caspase-3 in PTU-treated mice. Further, neonatal hypothyroidism affected germ cell proliferation with decreased expression of proliferating cell nuclear antigen (PCNA) and Cx43. In conclusion, our results suggest that neonatal hypothyroidism alters testicular glucose homeostasis via increased oxidative stress in prepubertal mice, thereby affecting germ cell survival and proliferation. © 2017 American Society of Andrology and European Academy of Andrology.
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USDA-ARS?s Scientific Manuscript database
Short-chain fatty acids (SCFAs,), especially butyrate, affect cell differentiation, proliferation, and motility. Furthermore, butyrate induces cell cycle arrest and apoptosis through its inhibition on histone deacetylases (HDACs). Butyrate is a potent inducer of histone hyper-acetylation in cells a...
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Vitamin K2 improves proliferation and migration of bovine skeletal muscle cells in vitro.
Rønning, Sissel Beate; Pedersen, Mona Elisabeth; Berg, Ragnhild Stenberg; Kirkhus, Bente; Rødbotten, Rune
2018-01-01
Skeletal muscle function is highly dependent on the ability to regenerate, however, during ageing or disease, the proliferative capacity is reduced, leading to loss of muscle function. We have previously demonstrated the presence of vitamin K2 in bovine skeletal muscles, but whether vitamin K has a role in muscle regulation and function is unknown. In this study, we used primary bovine skeletal muscle cells, cultured in monolayers in vitro, to assess a potential effect of vitamin K2 (MK-4) during myogenesis of muscle cells. Cell viability experiments demonstrate that the amount of ATP produced by the cells was unchanged when MK-4 was added, indicating viable cells. Cytotoxicity analysis show that MK-4 reduced the lactate dehydrogenase (LDH) released into the media, suggesting that MK-4 was beneficial to the muscle cells. Cell migration, proliferation and differentiation was characterised after MK-4 incubation using wound scratch analysis, immunocytochemistry and real-time PCR analysis. Adding MK-4 to the cells led to an increased muscle proliferation, increased gene expression of the myogenic transcription factor myod as well as increased cell migration. In addition, we observed a reduction in the fusion index and relative gene expression of muscle differentiation markers, with fewer complex myotubes formed in MK-4 stimulated cells compared to control cells, indicating that the MK-4 plays a significant role during the early phases of muscle proliferation. Likewise, we see the same pattern for the relative gene expression of collagen 1A, showing increased gene expression in proliferating cells, and reduced expression in differentiating cells. Our results also suggest that MK-4 incubation affect low density lipoprotein receptor-related protein 1 (LRP1) and the low-density lipoprotein receptor (LDLR) with a peak in gene expression after 45 min of MK-4 incubation. Altogether, our experiments show that MK-4 has a positive effect on muscle cell migration and proliferation, which are two important steps during early myogenesis.
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Fei, Fei; Li, Xiaofei; Xu, Li; Li, Deyang; Zhang, Zhipei; Guo, Xu; Yang, Hushan; Chen, Zhinan; Xing, Jinliang
2014-12-01
It has been reported that CD147 and CD98 heavy chain (CD98hc) form a complex on the cell plasma membrane of several cancers; however, whether this complex exists in non-small cell lung cancer (NSCLC) cells and affects the prognosis of patients remains to be elucidated. The expression of CD147 and CD98hc was assessed in tissue samples from 241 NSCLC patients and NSCLC cell lines. The correlation between CD147 and CD98hc expression and their association with the prognosis of NSCLC patients were analyzed. We also evaluated the impact of CD147 and CD98hc on the growth of NSCLC cells as well as Akt phosphorylation. Both CD147 and CD98hc were significantly upregulated in NSCLC cells, and their expression levels were significantly correlated (p < 0.001). Immunoflurenece staining and co-immunoprecipitation demonstrated that CD147 and CD98hc could form a complex on NSCLC cells. Compared with NSCLC patients with CD147-/CD98hc-, those with CD147+/CD98hc+ exhibited a significantly poor overall survival (OS) with a hazard ratio (HR) of 1.92 (p = 0.010), and a significantly increased risk of recurrence with a HR of 1.97 (p = 0.004). Also, we demonstrated that the proliferation of lung cancer cell lines was significantly affected by knockdown and force-expression of the CD147-CD98hc complex. Western blot analysis indicated that the phosphorylation of Akt in NSCLC cells was significantly affected by knockdown and overexpression of either or both CD147 and CD98hc. Our findings indicate that the CD147-CD98hc complex significantly contributes to poor prognosis of NSCLC patients through promoting cell proliferation via the PI3K/Akt pathway.
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Islam, Md Soriful; Catherino, William H.; Protic, Olga; Janjusevic, Milijana; Gray, Peter Clarke; Giannubilo, Stefano Raffaele; Ciavattini, Andrea; Lamanna, Pasquale; Tranquilli, Andrea Luigi; Petraglia, Felice
2014-01-01
Context: Uterine leiomyomas are highly prevalent benign tumors of premenopausal women and the most common indication for hysterectomy. However, the exact etiology of this tumor is not fully understood. Objective: The objective of the study was to evaluate the role of activin-A and myostatin and their signaling pathways in human myometrial and leiomyoma cells. Design: This was a laboratory study. Setting: Myometrial and leiomyoma cells (primary and cell lines) were cultured in vitro. Patients: The study included premenopausal women who were admitted to the hospital for myomectomy or hysterectomy. Interventions: Primary myometrial and leiomyoma cells and/or cell lines were treated with activin-A (4 nM) and myostatin (4 nM) for different days of interval (to measure proliferation rate) or 30 minutes (to measure signaling molecules) or 48 hours to measure proliferating markers, extracellular matrix mRNA, and/or protein expression by real-time PCR, Western blot, and/or immunocytochemistry. Results: We found that activin-A and myostatin significantly reduce cell proliferation in primary myometrial cells but not in leiomyoma cells as measured by a CyQUANT cell proliferation assay kit. Reduced expression of proliferating cell nuclear antigen and Ki-67 were also observed in myometrial cells in response to activin-A and myostatin treatment. Activin-A also significantly increased mRNA expression of fibronectin, collagen1A1, and versican in primary leiomyoma cells. Finally, we found that activin-A and myostatin activate Smad-2/3 signaling but do not affect ERK or p38 signaling in both myometrial and leiomyoma cells. Conclusions: This study results suggest that activin-A and myostatin can exert antiproliferative and/or fibrotic effects on these cell types via Smad-2/3 signaling. PMID:24606069
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Kviecinski, M.R., E-mail: mrkviecinski@hotmail.com; Pedrosa, R.C., E-mail: rozangelapedrosa@gmail.com; Felipe, K.B., E-mail: kakabettega@yahoo.com.br
2012-05-04
Highlights: Black-Right-Pointing-Pointer The cytotoxicity of juglone is markedly increased by ascorbate. Black-Right-Pointing-Pointer T24 cell death by oxidative stress is necrosis-like. Black-Right-Pointing-Pointer Redox cycling by juglone/ascorbate inhibits cell proliferation. Black-Right-Pointing-Pointer Cellular migration is impaired by juglone/ascorbate. -- Abstract: The effects of juglone on T24 cells were assessed in the presence and absence of ascorbate. The EC{sub 50} value for juglone at 24 h decreased from 28.5 {mu}M to 6.3 {mu}M in the presence of ascorbate. In juglone-treated cells, ascorbate increased ROS formation (4-fold) and depleted GSH (65%). N-acetylcysteine or catalase restricted the juglone/ascorbate-mediated effects, highlighting the role of oxidative stress inmore » juglone cytotoxicity. Juglone alone or associated with ascorbate did not cause caspase-3 activation or PARP cleavage, suggesting necrosis-like cell death. DNA damage and the mild ER stress caused by juglone were both enhanced by ascorbate. In cells treated with juglone (1-5 {mu}M), a concentration-dependent decrease in cell proliferation was observed. Ascorbate did not impair cell proliferation but its association with juglone led to a clonogenic death state. The motility of ascorbate-treated cells was not affected. Juglone slightly restricted motility, but cells lost their ability to migrate most noticeably when treated with juglone plus ascorbate. We postulate that juglone kills cells by a necrosis-like mechanism inhibiting cell proliferation and the motility of T24 cells. These effects are enhanced in the presence of ascorbate.« less
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Kunzmann, Volker; Kimmel, Brigitte; Herrmann, Thomas; Einsele, Hermann; Wilhelm, Martin
2009-02-01
Tumour growth promotes the expansion of CD4(+) CD25(+) FoxP3(+) regulatory T cells (Tregs) which suppress various arms of immune responses and might therefore contribute to tumour immunosurveillance. In this study, we found an inverse correlation between circulating Treg frequencies and phosphoantigen-induced gammadelta T-cell proliferation in cancer patients, which prompted us to address the role of Tregs in controlling the gammadelta T-cell arm of innate immune responses. In vitro, human Treg-peripheral blood mononuclear cell (PBMC) co-cultures strongly inhibited phosphoantigen-induced proliferation of gammadelta T cells and depletion of Tregs restored the impaired phosphoantigen-induced gammadelta T-cell proliferation of cancer patients. Tregs did not suppress other effector functions of gammadelta T cells such as cytokine production or cytotoxicity. Our experiments indicate that Tregs do not mediate their suppressive activity via a cell-cell contact-dependent mechanism, but rather secrete a soluble non-proteinaceous factor, which is independent of known soluble factors interacting with amino acid depletion (e.g. arginase-diminished arginine and indolamine 2,3-dioxygenase-diminished tryptophan) or nitric oxide (NO) production. However, the proliferative activity of alphabeta T cells was not affected by this cell-cell contact-independent suppressive activity induced by Tregs. In conclusion, these findings indicate a potential new mechanism by which Tregs can specifically suppress gammadelta T cells and highlight the strategy of combining Treg inhibition with subsequent gammadelta T-cell activation to enhance gammadelta T cell-mediated immunotherapy.
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Parthymou, Anastasia; Lampropoulou, Evgenia; Mikelis, Constantinos; Drosou, Georgia; Papadimitriou, Evangelia
2008-01-01
Heparin affin regulatory peptide (HARP) or pleiotrophin seems to be involved in the progression of several tumors of diverse origin. In this study, we tried to determine the role of HARP in rat C6 glioma cells by using an antisense strategy for inhibition of HARP expression. Decrease of the expression of endogenous HARP in C6 cells (AS-C6 cells) significantly increased proliferation, migration, and anchorage-independent growth of cells. Implantation of AS-C6 cells onto chicken embryo chorioallantoic membranes resulted in a significant increase of tumor-induced angiogenesis compared with that induced by non-transfected or C6 cells transfected with the plasmid alone (PC-C6 cells). In the same line, conditioned medium from AS-C6 cells significantly increased endothelial cell proliferation, migration, and tube formation in vitro compared with the effect of conditioned medium from C6 or PC-C6 cells. Interestingly, vascular endothelial growth factor (VEGF) induced C6 cell proliferation and migration, and SU1496, a selective inhibitor of VEGF receptor 2 (VEGFR2), blocked increased glioma cell growth, migration, and angiogenicity observed in AS-C6 cell cultures. The above results seem to be due to a direct interaction between HARP and VEGF in the culture medium of C6 and PC-C6 cells, while AS-C6 cells secreted comparable amounts of VEGF that do not interact with HARP. Collectively, these data suggest that HARP negatively affects diverse biological activities in C6 glioma cells, mainly due to binding of HARP to VEGF, which may sequester secreted VEGF from signalling through VEGFR2.
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Tzeng, Wen-Yu; Wu, Hsin-Hua; Wang, Ching-Yi; Chen, Jin-Chung; Yu, Lung; Cherng, Chianfang G
2016-01-01
Sex differences in stress and coping responses have been frequently documented in aged people, while whether such differences in aged people may appear at the middle age are unknown. This study was undertaken to study the impact of acute stress and social interaction on early neurogenesis in the dentate gyrus (DG) and hippocampus-related memory in two sexes of middle-aged mice. The number of newly proliferated cells, neuroblasts in DG, the object recognition and location memory in 9-month-old male and female C57BL/6N mice were assessed under baseline conditions as well as following an acute stressor regimen and group housing. Three conspecific companions, serving as "the housing group," were used to model the social interaction throughout the stressor regimen. Males had lower numbers of newly proliferated cells and neuroblasts under baseline conditions as compared to females. The stressor regimen caused rapid decreases in the number of newly proliferated cells and neuroblasts in female DG but no obvious changes were observed in male DG. Group housing, regardless of companions' age, prevented the stress-induced decreases in the number of newly proliferated cells and neuroblasts in female DG. In contrast, the presence of young or age-matched companions potentiated the stress effect in males by decreasing the number of newly proliferated cells and neuroblasts. Finally, neither the stressor regimen nor group housing affected mouse performances in the object recognition and location memory in either sex. These findings, taken together, provide evidence to support a notion that middle-aged females appear to demonstrate more stress susceptibility on early neurogenesis in DG as compared to middle-aged males, although the hippocampus-related memory performances are comparable and not affected by stress in these males and females. Experiencing stress, middle-aged females are more prone to benefit from social interaction as compared to middle-aged males in this regard. We suggest, accordingly, that involving social interaction may afford a therapeutic advance in preventing stress-produced decreases in early neurogenesis in middle-aged females' DG.
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Li, Qian; Guo, Zhenhong; Xu, Xiongfei; Xia, Sheng; Cao, Xuetao
2008-10-01
The tissue microenvironment may affect the development and function of immune cells such as DC. Whether and how the pulmonary stromal microenvironment can affect the development and function of lung DC need to be investigated. Regulatory DC (DCreg) can regulate T-cell response. We wondered whether such regulatory DC exist in the lung and what is the effect of the pulmonary stromal microenvironment on the generation of DCreg. Here we demonstrate that murine pulmonary stromal cells can drive immature DC, which are regarded as being widely distributed in the lung, to proliferate and differentiate into a distinct subset of DCreg, which express high levels of CD11b but low levels of MHC class II (I-A), CD11c, secrete high amounts of IL-10, NO and prostaglandin E2 (PGE2) and suppress T-cell proliferation. The natural counterpart of DCreg in the lung with similar phenotype and regulatory function has been identified. Pulmonary stroma-derived TGF-beta is responsible for the differentiation of immature DC to DCreg, and DCreg-derived PGE2 contributes to their suppression of T-cell proliferation. Moreover, DCreg can induce the generation of CD4+CD25+Foxp3+ Treg. Importantly, infusion with DCreg attenuates T-cell-mediated eosinophilic airway inflammation in vivo. Therefore, the pulmonary microenvironment may drive the generation of DCreg, thus contributing to the maintenance of immune homoeostasis and the control of inflammation in the lung.
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Bravenboer, Nathalie
2016-01-01
During the initial stages of bone repair, proinflammatory cytokines are released within the injury site, quickly followed by a shift to anti-inflammatory cytokines. The effect of pro- and anti-inflammatory cytokines on osteogenic differentiation of mesenchymal stem cells is controversial. Here, we investigated the effect of the proinflammatory cytokines TNF-α, IL-6, IL-8, and IL-17F and the anti-inflammatory cytokine IL-4 on proliferation and osteogenic differentiation of human adipose stem cells (hASCs). hASCs were treated with TNF-α, IL-6, IL-8, IL-17F, or IL-4 (10 ng/mL) for 72 h mimicking bone repair. TNF-α reduced collagen type I gene expression but increased hASC proliferation and ALP activity. IL-6 also strongly enhanced ALP activity (18-fold), as well as bone nodule formation by hASCs. IL-8 did not affect proliferation or osteogenic gene expression but reduced bone nodule formation. IL-17F decreased hASC proliferation but enhanced ALP activity. IL-4 enhanced osteocalcin gene expression and ALP activity but reduced RUNX2 gene expression and bone nodule formation. In conclusion, all cytokines studied have both enhancing and reducing effects on osteogenic differentiation of hASCs, even when applied for 72 h only. Some cytokines, specifically IL-6, may be suitable to induce osteogenic differentiation of mesenchymal stem cells as a strategy for enhancing bone repair. PMID:27667999
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Eisenbeis, Janina; Peisker, Henrik; Backes, Christian S; Bur, Stephanie; Hölters, Sebastian; Thewes, Nicolas; Greiner, Markus; Junker, Christian; Schwarz, Eva C; Hoth, Markus; Junker, Kerstin; Preissner, Klaus T; Jacobs, Karin; Herrmann, Mathias; Bischoff, Markus
2017-02-01
Staphyloccocus aureus is a major human pathogen and a common cause for superficial and deep seated wound infections. The pathogen is equipped with a large arsenal of virulence factors, which facilitate attachment to various eukaryotic cell structures and modulate the host immune response. One of these factors is the extracellular adherence protein Eap, a member of the "secretable expanded repertoire adhesive molecules" (SERAM) protein family that possesses adhesive and immune modulatory properties. The secreted protein was previously shown to impair wound healing by interfering with host defense and neovascularization. However, its impact on keratinocyte proliferation and migration, two major steps in the re-epithelialization process of wounds, is not known. Here, we report that Eap affects the proliferation and migration capacities of keratinocytes by altering their morphology and adhesive properties. In particular, treatment of non-confluent HaCaT cell cultures with Eap resulted in cell morphology changes as well as a significant reduction in cell proliferation and migration. Eap-treated HaCaT cells changed their appearance from an oblong via a trapezoid to an astral-like shape, accompanied by decreases in cell volume and cell stiffness, and exhibited significantly increased cell adhesion. Eap had a similar influence on endothelial and cancer cells, indicative for a general effect of Eap on eukaryotic cell morphology and functions. Specifically, Eap was found to interfere with growth factor-stimulated activation of the mitogen-activated protein kinase (MAPK) pathway that is known to be responsible for cell shape modulation, induction of proliferation and migration of epithelial cells. Western blot analyses revealed that Eap blocked the phosphorylation of extracellular signal-regulated kinase 1 and 2 (Erk1/2) in keratinocyte growth factor (KGF)-stimulated HaCaT cells. Together, these data add another antagonistic mechanism of Eap in wound healing, whereby the bacterial protein interferes with keratinocyte migration and proliferation. Copyright © 2017 Elsevier GmbH. All rights reserved.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Tavakolinejad, Alireza; Rabbani, Mohsen, E-mail: m.rabbani@eng.ui.ac.ir; Janmaleki, Mohsen
2015-08-21
Alteration in specific inertial conditions can lead to changes in morphology, proliferation, mechanical properties and cytoskeleton of cells. In this report, the effects of hypergravity on morphology of Adipose-Derived Stem Cells (ADSCs) are indicated. ADSCs were repeatedly exposed to discontinuous hypergravity conditions of 10 g, 20 g, 40 g and 60 g by utilizing centrifuge (three times of 20 min exposure, with an interval of 40 min at 1 g). Cell morphology in terms of length, width and cell elongation index and cytoskeleton of actin filaments and microtubules were analyzed by image processing. Consistent changes observed in cell elongation index as morphological change. Moreover, cell proliferation wasmore » assessed and mechanical properties of cells in case of elastic modulus of cells were evaluated by Atomic Force Microscopy. Increase in proliferation and decrease in elastic modulus of cells are further results of this study. Staining ADSC was done to show changes in cytoskeleton of the cells associated to hypergravity condition specifically in microfilament and microtubule components. After exposing to hypergravity, significant changes were observed in microfilaments and microtubule density as components of cytoskeleton. It was concluded that there could be a relationship between changes in morphology and MFs as the main component of the cells. - Highlights: • Hypergravity (10 g, 20 g, 40 g and 60 g) affects on adipose derived stem cells (ADSCs). • ADSCs after exposure to the hypergravity are more slender. • The height of ADSCs increases in all test groups comparing their control group. • Hypergravity decreases ADSCs modulus of elasticity and cell actin fiber content. • Hypergravity enhances proliferation rate of ADSCs.« less
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Chu, Edward P F; Elso, Colleen M; Pollock, Abigail H; Alsayb, May A; Mackin, Leanne; Thomas, Helen E; Kay, Thomas W H; Silveira, Pablo A; Mansell, Ashley S; Gaus, Katharina; Brodnicki, Thomas C
2017-02-01
During immune cell activation, serine-derived lipids such as phosphatidylserine and sphingolipids contribute to the formation of protein signaling complexes within the plasma membrane. Altering lipid composition in the cell membrane can subsequently affect immune cell function and the development of autoimmune disease. Serine incorporator 1 (SERINC1) is a putative carrier protein that facilitates synthesis of serine-derived lipids. To determine if SERINC1 has a role in immune cell function and the development of autoimmunity, we characterized a mouse strain in which a retroviral insertion abolishes expression of the Serinc1 transcript. Expression analyses indicated that the Serinc1 transcript is readily detectable and expressed at relatively high levels in wildtype macrophages and lymphocytes. The ablation of Serinc1 expression in these immune cells, however, did not significantly alter serine-derived lipid composition or affect macrophage function and lymphocyte proliferation. Analyses of Serinc1-deficient mice also indicated that systemic ablation of Serinc1 expression did not affect viability, fertility or autoimmune disease susceptibility. These results suggest that Serinc1 is dispensable for certain immune cell functions and does not contribute to previously reported links between lipid composition in immune cells and autoimmunity. Copyright © 2016 Elsevier Ltd. All rights reserved.
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de Tomaso Portaz, Ana Clara; Caimi, Giselle Romero; Sánchez, Marcela; Chiappini, Florencia; Randi, Andrea S; Kleiman de Pisarev, Diana L; Alvarez, Laura
2015-10-02
Hexachlorobenzene (HCB) is a widespread environmental pollutant, and a liver tumor promoter in rodents. Depending on the particular cell lines studied, exposure to these compounds may lead to cell proliferation, terminal differentiation, or apoptosis. The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is involved in drug and xenobiotic metabolism. AhR can also modulate a variety of cellular and physiological processes that can affect cell proliferation and cell fate determination. The mechanisms by which AhR ligands, both exogenous and endogenous, affect these processes involve multiple interactions between AhR and other signaling pathways. In the present study, we examined the effect of HCB on cell proliferation and AhR expression, using an initiation-promotion hepatocarcinogenesis protocol in rat liver and in the human-derived hepatoma cell line, HepG2. Female Wistar rats were initiated with a single dose of 100 mg/kg of diethylnitrosamine (DEN) at the start of the experiment. Two weeks later, daily dosing of 100 mg/kg HCB was maintained for 10 weeks. Partial hepatectomy was performed 3 weeks after initiation. The number and area of glutathione S-transferase-P (GST-P)-positive foci, in the rat liver were used as biomarkers of liver precancerous lesions. Immunohistochemical staining showed an increase in proliferating cell nuclear antigen (PCNA)-positive cells, along with enhanced AhR protein expression in hepatocytes within GST-P-positive foci of (DEN HCB) group, when compared to DEN. In a similar manner, Western blot analysis demonstrated that HCB induced PCNA and AhR protein expression in HepG2 cells. Flow cytometry assay indicated that the cells were accumulated at S and G2/M phases of the cell cycle. HCB increased cyclin D1 protein levels and ERK1/2 phosphorylation in a dose-dependent manner. Treatment of cells with a selective MEK1 inhibitor, prevented HCB-stimulatory effect on PCNA and cyclinD1, indicating that these effects are mediated by ERK1/2. Pretreatment with an AhR antagonist, prevented HCB-induced PCNA protein levels, ERK1/2 phosphorylation and alterations in cell cycle distribution. These results demonstrate that HCB-induced HepG2 proliferation and cell cycle progression depend on ERK1/2 phosphorylation which is mediated by the AhR. Our results provide a clue to the molecular events involved in the mechanism of action of HCB-induced hepatocarcinogenesis. Copyright © 2015. Published by Elsevier Ireland Ltd.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Lo, Yuan-Hung; Ho, Po-Chun; Chen, Min-Shan
Highlights: Black-Right-Pointing-Pointer Proliferating Cell Nuclear Antigen (PCNA) is phosphorylated at Y114. Black-Right-Pointing-Pointer Phospho-Y114 of PCNA is not required for cell proliferation for normal growth. Black-Right-Pointing-Pointer MCE during adipogenesis is abolished in the lack of the phosphorylation. Black-Right-Pointing-Pointer Homozygous Y114F mice are resistant to high fat diet induced obesity. Black-Right-Pointing-Pointer Our results shed light on the interface between proliferation and differentiation. -- Abstract: Clonal proliferation is an obligatory component of adipogenesis. Although several cell cycle regulators are known to participate in the transition between pre-adipocyte proliferation and terminal adipocyte differentiation, how the core DNA synthesis machinery is coordinately regulated in adipogenesismore » remains elusive. PCNA (Proliferating Cell Nuclear Antigen) is an indispensable component for DNA synthesis during proliferation. Here we show that PCNA is subject to phosphorylation at the highly conserved tyrosine residue 114 (Y114). Replacing the Y114 residue with phenylalanine (Y114F), which is structurally similar to tyrosine but cannot be phosphorylated, does not affect normal animal development. However, when challenged with high fat diet, mice carrying homozygous Y114F alleles (PCNA{sup F/F}) are resistant to adipose tissue enlargement in comparison to wild-type (WT) mice. Mouse embryonic fibroblasts (MEFs) harboring WT or Y114F mutant PCNA proliferate at similar rates. However, when subjected to adipogenesis induction in culture, PCNA{sup F/F} MEFs are not able to re-enter the cell cycle and fail to form mature adipocytes, while WT MEFs undergo mitotic clonal expansion in response to the adipogenic stimulation, accompanied by enhanced Y114 phosphorylation of PCNA, and differentiate to mature adipocytes. Consistent with the function of Y114 phosphorylation in clonal proliferation in adipogenesis, fat tissues isolated from WT mice contain significantly more adipocytes than those isolated from PCNA{sup F/F} mice. This study identifies a critical role for PCNA in adipose tissue development, and for the first time identifies a role of the core DNA replication machinery at the interface between proliferation and differentiation.« less
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Koo, Bon Hyeock; Yi, Bong Gu; Wang, Wi Kwang; Ko, In Young; Hoe, Kwang Lae; Kwon, Young Guen; Won, Moo Ho; Kim, Young Myeong; Lim, Hyun Kyo; Ryoo, Sungwoo
2018-05-01
Vascular smooth muscle cell (VSMC) proliferation induced by native low-density lipoprotein (nLDL) stimulation is dependent on superoxide production from activated NADPH oxidase. The present study aimed to investigate whether the novel arginase inhibitor limonin could suppress nLDL-induced VSMC proliferation and to examine related mechanisms. Isolated VSMCs from rat aortas were treated with nLDL, and cell proliferation was measured by WST-1 and BrdU assays. NADPH oxidase activation was evaluated by lucigenin-induced chemiluminescence, and phosphorylation of protein kinase C (PKC) βII and extracellular signal-regulated kinase (ERK) 1/2 was determined by western blot analysis. Mitochondrial reactive oxygen species (ROS) generation was assessed using MitoSOX-red, and intracellular L-arginine concentrations were determined by high-performance liquid chromatography (HPLC) in the presence or absence of limonin. Limonin inhibited arginase I and II activity in the uncompetitive mode, and prevented nLDL-induced VSMC proliferation in a p21Waf1/Cip1-dependent manner without affecting arginase protein levels. Limonin blocked PKCβII phosphorylation, but not ERK1/2 phosphorylation, and translocation of p47phox to the membrane was decreased, as was superoxide production in nLDL-stimulated VSMCs. Moreover, mitochondrial ROS generation was increased by nLDL stimulation and blocked by preincubation with limonin. Mitochondrial ROS production was responsible for the phosphorylation of PKCβII. HPLC analysis showed that arginase inhibition with limonin increases intracellular L-arginine concentrations, but decreases polyamine concentrations. L-Arginine treatment prevented PKCβII phosphorylation without affecting ERK1/2 phosphorylation. Increased L-arginine levels following limonin-dependent arginase inhibition prohibited NADPH oxidase activation in a PKCβII-dependent manner, and blocked nLDL-stimulated VSMC proliferation. © Copyright: Yonsei University College of Medicine 2018.
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Wang, Wi-Kwang; Ko, In-Young; Hoe, Kwang-Lae; Kwon, Young-Guen; Won, Moo-Ho; Kim, Young-Myeong
2018-01-01
Purpose Vascular smooth muscle cell (VSMC) proliferation induced by native low-density lipoprotein (nLDL) stimulation is dependent on superoxide production from activated NADPH oxidase. The present study aimed to investigate whether the novel arginase inhibitor limonin could suppress nLDL-induced VSMC proliferation and to examine related mechanisms. Materials and Methods Isolated VSMCs from rat aortas were treated with nLDL, and cell proliferation was measured by WST-1 and BrdU assays. NADPH oxidase activation was evaluated by lucigenin-induced chemiluminescence, and phosphorylation of protein kinase C (PKC) βII and extracellular signal-regulated kinase (ERK) 1/2 was determined by western blot analysis. Mitochondrial reactive oxygen species (ROS) generation was assessed using MitoSOX-red, and intracellular L-arginine concentrations were determined by high-performance liquid chromatography (HPLC) in the presence or absence of limonin. Results Limonin inhibited arginase I and II activity in the uncompetitive mode, and prevented nLDL-induced VSMC proliferation in a p21Waf1/Cip1-dependent manner without affecting arginase protein levels. Limonin blocked PKCβII phosphorylation, but not ERK1/2 phosphorylation, and translocation of p47phox to the membrane was decreased, as was superoxide production in nLDL-stimulated VSMCs. Moreover, mitochondrial ROS generation was increased by nLDL stimulation and blocked by preincubation with limonin. Mitochondrial ROS production was responsible for the phosphorylation of PKCβII. HPLC analysis showed that arginase inhibition with limonin increases intracellular L-arginine concentrations, but decreases polyamine concentrations. L-Arginine treatment prevented PKCβII phosphorylation without affecting ERK1/2 phosphorylation. Conclusion Increased L-arginine levels following limonin-dependent arginase inhibition prohibited NADPH oxidase activation in a PKCβII-dependent manner, and blocked nLDL-stimulated VSMC proliferation. PMID:29611398
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A role for adult TLX-positive neural stem cells in learning and behaviour.
Zhang, Chun-Li; Zou, Yuhua; He, Weimin; Gage, Fred H; Evans, Ronald M
2008-02-21
Neurogenesis persists in the adult brain and can be regulated by a plethora of external stimuli, such as learning, memory, exercise, environment and stress. Although newly generated neurons are able to migrate and preferentially incorporate into the neural network, how these cells are molecularly regulated and whether they are required for any normal brain function are unresolved questions. The adult neural stem cell pool is composed of orphan nuclear receptor TLX-positive cells. Here, using genetic approaches in mice, we demonstrate that TLX (also called NR2E1) regulates adult neural stem cell proliferation in a cell-autonomous manner by controlling a defined genetic network implicated in cell proliferation and growth. Consequently, specific removal of TLX from the adult mouse brain through inducible recombination results in a significant reduction of stem cell proliferation and a marked decrement in spatial learning. In contrast, the resulting suppression of adult neurogenesis does not affect contextual fear conditioning, locomotion or diurnal rhythmic activities, indicating a more selective contribution of newly generated neurons to specific cognitive functions.
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An Emerging Allee Effect Is Critical for Tumor Initiation and Persistence
Böttger, Katrin; Hatzikirou, Haralambos; Voss-Böhme, Anja; Cavalcanti-Adam, Elisabetta Ada; Herrero, Miguel A.; Deutsch, Andreas
2015-01-01
Tumor cells develop different strategies to cope with changing microenvironmental conditions. A prominent example is the adaptive phenotypic switching between cell migration and proliferation. While it has been shown that the migration-proliferation plasticity influences tumor spread, it remains unclear how this particular phenotypic plasticity affects overall tumor growth, in particular initiation and persistence. To address this problem, we formulate and study a mathematical model of spatio-temporal tumor dynamics which incorporates the microenvironmental influence through a local cell density dependence. Our analysis reveals that two dynamic regimes can be distinguished. If cell motility is allowed to increase with local cell density, any tumor cell population will persist in time, irrespective of its initial size. On the contrary, if cell motility is assumed to decrease with respect to local cell density, any tumor population below a certain size threshold will eventually extinguish, a fact usually termed as Allee effect in ecology. These results suggest that strategies aimed at modulating migration are worth to be explored as alternatives to those mainly focused at keeping tumor proliferation under control. PMID:26335202
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Bertapelle, Carla; Polese, Gianluca; Di Cosmo, Anna
2017-06-01
Organisms showing a complex and centralized nervous system, such as teleosts, amphibians, reptiles, birds and mammals, and among invertebrates, crustaceans and insects, can adjust their behavior according to the environmental challenges. Proliferation, differentiation, migration, and axonal and dendritic development of newborn neurons take place in brain areas where structural plasticity, involved in learning, memory, and sensory stimuli integration, occurs. Octopus vulgaris has a complex and centralized nervous system, located between the eyes, with a hierarchical organization. It is considered the most "intelligent" invertebrate for its advanced cognitive capabilities, as learning and memory, and its sophisticated behaviors. The experimental data obtained by immunohistochemistry and western blot assay using proliferating cell nuclear antigen and poli (ADP-ribose) polymerase 1 as marker of cell proliferation and synaptogenesis, respectively, reviled cell proliferation in areas of brain involved in learning, memory, and sensory stimuli integration. Furthermore, we showed how enriched environmental conditions affect adult neurogenesis. © 2017 Wiley Periodicals, Inc.
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Kinetics of liver macrophages (Kupffer cells) in SIV-infected macaques
DOE Office of Scientific and Technical Information (OSTI.GOV)
Ahsan, Muhammad H.; Gill, Amy F.; Alvarez, Xavier
Since the liver drains antigens from the intestinal tract, and since the intestinal tract is a major site of viral replication, we examined the dynamics of liver macrophages (Kupffer cells) throughout SIV infection. Absolute numbers of Kupffer cells increased in the livers in acute infection, and in animals with AIDS. Significantly higher percentages of proliferating (BrdU+) Kupffer cells were detected in acute infection and in AIDS with similar trends in blood monocytes. Significantly higher percentages of apoptotic (AC3+) Kupffer cells were also found in acute and AIDS stages. However, productively infected cells were not detected in liver of 41/42 animalsmore » examined, despite abundant infected cells in gut and lymph nodes of all animals. Increased rates of Kupffer cell proliferation resulting in an increase in Kupffer cells without productive infection indicate SIV infection affects Kupffer cells, but the liver does not appear to be a major site of productive viral replication. - Highlights: • Kupffer cells increase in the liver of SIV-infected macaques. • Increased proliferation and apoptosis of Kupffer cells occurs in SIV infection. • Productively infected cells are rarely detected in the liver. • The liver is not a major site for SIV replication.« less
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Ginani, Fernanda; Soares, Diego Moura; Rabêlo, Luciana Maria; Rocha, Hugo Alexandre Oliveira; de Souza, Lélia Batista; Barboza, Carlos Augusto Galvão
2016-11-01
The aim of the present study was to evaluate the influence of a cryopreservation protocol on the proliferation and viability of stem cells from human exfoliated deciduous teeth (SHEDs). Cells from the pulp of three deciduous teeth were isolated and characterized to confirm their stem cell nature. In second passage, part of the cells were submitted to normal conditions of cell culture (Control group), while part of the cells were maintained in 10% DMSO diluted in foetal bovine serum and submitted to the following cryopreservation protocol: 2 h at 4 °C, 18 h at -20 °C and then at -80 °C for two intervals (30 days - Cryopreservation I; and 180 days Cryopreservation II). Cell proliferation and cell cycle were evaluated at intervals of 24, 48 and 72 h after plating, and apoptosis-related events were analyzed at 72 h. All groups exhibited an increase in the number of cells, and no significant differences between the cryopreserved and control groups were observed (p > .05). The distribution of cells in the cell cycle phases was consistent with cell proliferation, and the percentage of viable cells was higher than 99% in all groups, indicating that cell viability was not affected by the cryopreservation protocol throughout the experiment. The proposed cryopreservation protocol is adequate for the storage of SHED, permitting their use in future experimental studies.
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Effects and mechanisms of melatonin on the proliferation and neural differentiation of PC12 cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Liu, Yumei; Zhang, Ziqiang; Lv, Qiongxia
Melatonin, a lipophilic molecule that is mainly synthesized in the pineal gland, performs various neuroprotective functions. However, the detailed role and mechanisms of promoting neuronal differentiation remains limited. This study demonstrated that 10 μM melatonin led to significant increases in the proliferation and neurite outgrowth of PC12 cells. Increased expression of microtubule-associated protein 2 (MAP2, a neuron-specific protein) was also observed. However, luzindole (melatonin receptor antagonist) and PD98059 (MEK inhibitor) attenuated these increases. LY294002 (AKT inhibitor) inhibited melatonin-mediated proliferation in PC12 cells and did not affect melatonin-induced neural differentiation. The expression of p-ERK1/2/ERK1/2 was increased by melatonin treatment for 14 days in PC12 cells,more » whereas luzindole or PD98059 reduced the melatonin-induced increase. These results suggest that the activation of both the MEK/ERK and PI3K/AKT signaling pathways could potentially contribute to melatonin-mediated proliferation, but that only the MEK/ERK pathway participates in the melatonin-induced neural differentiation of PC12 cells. Altogether, our study demonstrates for the first time that melatonin may exert a positive effect on neural differentiation via melatonin receptor signalling and that the MEK/ERK1/2 signalling may act down stream from the melatonin pathway. - Highlights: • Melatonin improves the proliferation of PC12 cells. • Melatonin induces neural differentiation of PC12 cells. • Melatonin-mediated proliferation in PC12 cells relies on the ERK and AKT pathways. • Activation of ERK is essential for melatonin-induced neural differentiation of PC12.« less
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Tanaka, Satoshi; Sakaguchi, Minoru; Yoneyama, Hiroki
Histamine is involved in various physiological functions, including its neurotransmitter actions in the central nervous system and its action as a causative agent of inflammation, allergic reactions, and gastric acid secretions. Histamine expression and biosynthesis have been detected in breast cancer cells. It was recently suggested that the histamine H{sub 3} receptor (H{sub 3}R) plays a role in the proliferation of breast cancer cells. We recently developed the non-imidazole H{sub 3}R antagonist OUP-186 which exhibited a potent and selective human H{sub 3}R antagonistic activity as well as no activity against the human histamine H{sub 4} receptor (H{sub 4}R). In thismore » study, we compared the effects of OUP-186 on the proliferation of estrogen receptor negative (ER−) breast cancer cells (MDA-MB-231) and ER+ breast cancer cells (MCF7) to the effects of clobenpropit (potent imidazole-containing H{sub 3}R antagonist). OUP-186 and clobenpropit suppressed the proliferation of breast cancer cells. The IC{sub 50} values at 48 h for OUP-186 and clobenpropit were approximately 10 μM and 50 μM, respectively. Furthermore, OUP-186 potently induced cell death by activating caspase-3/7, whereas cell death was only slightly induced by clobenpropit. In addition, OUP-186 treatment blocked the proliferation increase triggered by 100 μM (R)-(-)-α-methylhistamine (H{sub 3}R agonist). The use of 4-methylhistamine (H{sub 4}R agonist) and JNJ10191584 (selective H{sub 4}R antagonist) did not affect breast cancer proliferation. These results indicate that OUP-186 potently suppresses proliferation and induces caspase-dependent apoptotic death in both ER+ and ER-breast cancer cells. - Highlights: • OUP-186, a histamine H{sub 3} receptor antagonist, effects breast cancer cell growth. • OUP-186 potently suppressed proliferation and induced caspase-dependent apoptosis. • OUP-186 may be an effective drug against ER+ and ER− breast cancers.« less
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The effect of nutritional status on myogenic satellite cell proliferation and differentiation.
Powell, D J; McFarland, D C; Cowieson, A J; Muir, W I; Velleman, S G
2013-08-01
Early posthatch satellite cell (SC) mitotic activity is a critical component of muscle development and growth. Satellite cells are stem cells that can be induced by nutrition to follow other cellular developmental pathways. The objective of the current study was to determine the effect of restricting protein synthesis on the proliferation and differentiation of SC, using variable concentrations of Met and Cys to modulate protein synthesis. Broiler pectoralis major SC were cultured and treated with 1 of 6 different Met/Cys concentrations: 60/192, 30/96 (control), 7.5/24, 3/9.6, 1/3.2, or 0/0 mg/L. The effect of Met/Cys concentration on SC proliferation and differentiation was measured, and myonuclear accretion was measured by counting the number of nuclei per myotube during differentiation. The 30/96 mg/L Met/Cys treatment resulted in the highest rate of proliferation compared with all other treatments by 72 h of proliferation (P < 0.05). Differentiation was measured with Met/Cys treatments only during proliferation and the cultures receiving normal differentiation medium (R/N), normal proliferation medium and differentiation medium with variable Met/Cys (N/R), or both proliferation and differentiation receiving variable Met/Cys treatments (R/R). Differentiation responded in a dose-dependent manner to Met/Cys concentration under all 3 of these treatment regimens, with a degree of recovery in the R/N regimen cells following reinstatement of the control medium. Reductions in both proliferation and differentiation were more pronounced as Met/Cys concentrations were further reduced, whereas increased differentiation was observed under the increased Met/Cys concentration treatment when applied during differentiation in the N/R and R/R regimens. The number of nuclei per myotube was significantly decreased in the severely Met/Cys restricted treatments (P < 0.05). These data demonstrate the sensitivity of pectoralis major SC to nutritional availability and the importance of optimal nutrition during both proliferation and differentiation for maximizing SC activity, which will affect subsequent muscle mass accretion.
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Patsoukis, Nikolaos; Brown, Julia; Petkova, Victoria; Liu, Fang; Li, Lequn; Boussiotis, Vassiliki A.
2017-01-01
The receptor programmed death 1 (PD-1) inhibits T cell proliferation and plays a critical role in suppressing self-reactive T cells, and it also compromises antiviral and antitumor responses. To determine how PD-1 signaling inhibits T cell proliferation, we used human CD4+ T cells to examine the effects of PD-1 signaling on the molecular control of the cell cycle. The ubiquitin ligase SCFSkp2 degrades p27kip1, an inhibitor of cyclin-dependent kinases (Cdks), and PD-1 blocked cell cycle progression through the G1 phase by suppressing transcription of SKP2, which encodes a component of this ubiquitin ligase. Thus, in T cells stimulated through PD-1, Cdks were not activated, and two critical Cdk substrates were not phosphorylated. Activation of PD-1 inhibited phosphorylation of the retinoblastoma gene product, which suppressed expression of E2F target genes. PD-1 also inhibited phosphorylation of the transcription factor Smad3, which increased its activity. These events induced additional inhibitory checkpoints in the cell cycle by increasing the abundance of the G1 phase inhibitor p15INK4 and repressing the Cdk-activating phosphatase Cdc25A. PD-1 suppressed SKP2 transcription by inhibiting phosphoinositide 3-kinase–Akt and Ras–mitogen-activated and extracellular signal–regulated kinase kinase (MEK)–extracellular signal–regulated kinase (ERK) signaling. Exposure of cells to the proliferation-promoting cytokine interleukin-2 restored activation of MEK-ERK signaling, but not Akt signaling, and only partially restored SKP2 expression. Thus, PD-1 blocks cell cycle progression and proliferation of T lymphocytes by affecting multiple regulators of the cell cycle. PMID:22740686
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Menon, Alessandra; Creo, Pasquale; Piccoli, Marco; Bergante, Sonia; Conforti, Erika; Banfi, Giuseppe; Randelli, Pietro; Anastasia, Luigi
2018-01-01
Adult stem cell-based therapeutic approaches for tissue regeneration have been proposed for several years. However, adult stem cells are usually limited in number and difficult to be expanded in vitro, and they usually tend to quickly lose their potency with passages, as they differentiate and become senescent. Culturing stem cells under reduced oxygen tensions (below 21%) has been proposed as a tool to increase cell proliferation, but many studies reported opposite effects. In particular, cell response to hypoxia seems to be very stem cell type specific. Nonetheless, it is clear that a major role in this process is played by the hypoxia inducible factor (HIF), the master regulator of cell response to oxygen deprivation, which affects cell metabolism and differentiation. Herein, we report that a chemical activation of HIF in human tendon stem cells reduces their proliferation and inhibits their differentiation in a reversible and dose-dependent manner. These results support the notion that hypoxia, by activating HIF, plays a crucial role in preserving stem cells in an undifferentiated state in the "hypoxic niches" present in the tissue in which they reside before migrating in more oxygenated areas to heal a damaged tissue.
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Changing the Properties of Multipotent Mesenchymal Stromal Cells by IFNγ Administration.
Petinati, N A; Kapranov, N M; Bigil'deev, A E; Popova, M D; Davydova, Yu O; Gal'tseva, I V; Drize, N I; Kuz'mina, L A; Parovichnikova, E N; Savchenko, V G
2017-06-01
We studied changes in the population of human multipotent mesenchymal stromal cells activated by IFNγ. The cells were cultured under standard conditions; IFNγ was added in various concentrations for 4 h or over 2 passages. It was shown that the total cell production significantly decreased after long-term culturing with IFNγ, but 4-h exposure did not affect this parameter. After 4-h culturing, the expression levels of IDO1, CSF1, and IL-6 increased by 300, 7, and 2.4 times, respectively, and this increase persisted 1 and 2 days after removal of IFNγ from the culture medium. The expression of class I and II MHC (HLA) on cell surface practically did not change immediately after exposure to IFNγ, but during further culturing, HLA-ABC (MHC I) and HLA-DR (MHC II) expression significantly increased, which abolished the immune privilege in these cells, the property allowing clinical use of allogenic multipotent mesenchymal stromal cells. Multipotent mesenchymal stromal cells can suppress proliferation of lymphocytes. The degree of this suppression depends on individual properties of multipotent mesenchymal stromal cell donor. Treatment with IFNγ did not significantly affect the intensity of inhibition of lymphocyte proliferation by these cells.
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Interleukin-15 regulates proliferation and self-renewal of adult neural stem cells
Gómez-Nicola, Diego; Valle-Argos, Beatriz; Pallas-Bazarra, Noemí; Nieto-Sampedro, Manuel
2011-01-01
The impact of inflammation is crucial for the regulation of the biology of neural stem cells (NSCs). Interleukin-15 (IL-15) appears as a likely candidate for regulating neurogenesis, based on its well-known mitogenic properties. We show here that NSCs of the subventricular zone (SVZ) express IL-15, which regulates NSC proliferation, as evidenced by the study of IL-15−/− mice and the effects of acute IL-15 administration, coupled to 5-bromo-2′-deoxyuridine/5-ethynyl-2′-deoxyuridine dual-pulse labeling. Moreover, IL-15 regulates NSC differentiation, its deficiency leading to an impaired generation of neuroblasts in the SVZ–rostral migratory stream axis, recoverable through the action of exogenous IL-15. IL-15 expressed in cultured NSCs is linked to self-renewal, proliferation, and differentiation. IL-15–/– NSCs presented deficient proliferation and self-renewal, as evidenced in proliferation and colony-forming assays and the analysis of cell cycle–regulatory proteins. Moreover, IL-15–deficient NSCs were more prone to differentiate than wild-type NSCs, not affecting the cell population balance. Lack of IL-15 led to a defective activation of the JAK/STAT and ERK pathways, key for the regulation of proliferation and differentiation of NSCs. The results show that IL-15 is a key regulator of neurogenesis in the adult and is essential to understanding diseases with an inflammatory component. PMID:21508317
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Xu, Yan; Hadjiargyrou, M; Rafailovich, Miriam; Mironava, Tatsiana
2017-07-11
Increasing production of nanomaterials requires fast and proper assessment of its potential toxicity. Therefore, there is a need to develop new assays that can be performed in vitro, be cost effective, and allow faster screening of engineered nanomaterials (ENMs). Herein, we report that titanium dioxide (TiO 2 ) nanoparticles (NPs) can induce damage to adipose derived stromal cells (ADSCs) at concentrations which are rated as safe by standard assays such as measuring proliferation, reactive oxygen species (ROS), and lactate dehydrogenase (LDH) levels. Specifically, we demonstrated that low concentrations of TiO 2 NPs, at which cellular LDH, ROS, or proliferation profiles were not affected, induced changes in the ADSCs secretory function and differentiation capability. These two functions are essential for ADSCs in wound healing, energy expenditure, and metabolism with serious health implications in vivo. We demonstrated that cytotoxicity assays based on specialized cell functions exhibit greater sensitivity and reveal damage induced by ENMs that was not otherwise detected by traditional ROS, LDH, and proliferation assays. For proper toxicological assessment of ENMs standard ROS, LDH, and proliferation assays should be combined with assays that investigate cellular functions relevant to the specific cell type.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Ai, Zhilong; Lu, Weiqi; Ton, Saixiong
2007-08-31
Gallbladder carcinoma (GBC), an aggressive and mostly lethal malignancy, is known to be resistant to a number of drug stimuli. Here, we demonstrated that arsenic trioxide inhibited the proliferation of gallbladder carcinoma in vivo and in vitro as well as the transcription of cell cycle-related protein Cyclin D1. And, Cyclin D1 overexpression inhibited the negative role of arsenic trioxide in cell cycle progression. We further explored the mechanisms by which arsenic trioxide affected Cyclin D1 transcription and found that the Sp1 transcription factor was down-regulated by arsenic trioxide, with a corresponding decrease in Cyclin D1 promoter activity. Taken together, thesemore » results suggested that arsenic trioxide inhibited gallbladder carcinoma cell proliferation via down-regulation of Cyclin D1 transcription in a Sp1-dependent manner, which provided a new mechanism of arsenic trioxide-involved cell proliferation and may have important therapeutic implications in gallbladder carcinoma patients.« less
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Mycophenolate mofetil attenuates pulmonary arterial hypertension in rats
DOE Office of Scientific and Technical Information (OSTI.GOV)
Suzuki, Chihiro; Takahashi, Masafumi; Morimoto, Hajime
Pulmonary arterial hypertension (PAH) is characterized by abnormal proliferation of smooth muscle cells (SMCs), leading to occlusion of pulmonary arterioles, right ventricular (RV) hypertrophy, and death. We investigated whether mycophenolate mofetil (MMF), a potent immunosuppresssant, prevents the development of monocrotaline (MCT)-induced PAH in rats. MMF effectively decreased RV systolic pressure and RV hypertrophy, and reduced the medial thickness of pulmonary arteries. MMF significantly inhibited the number of proliferating cell nuclear antigen (PCNA)-positive cells, infiltration of macrophages, and expression of P-selectin and interleukin-6 on the endothelium of pulmonary arteries. The infiltration of T cells and mast cells was not affected bymore » MMF. In vitro experiments revealed that mycophenolic acid (MPA), an active metabolite of MMF, dose-dependently inhibited proliferation of human pulmonary arterial SMCs. MMF attenuated the development of PAH through its anti-inflammatory and anti-proliferative properties. These findings provide new insight into the potential role of immunosuppressants in the treatment of PAH.« less
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Minaidou, Anna; Nicolaou, Paschalis; Christodoulou, Kyproula
2018-10-01
Deleterious variants in LRSAM1, a RING finger ubiquitin ligase which is also known as TSG101-associated ligase (TAL), have recently been associated with Charcot-Marie-Tooth disease type 2P (CMT2P). The mechanism by which mutant LRSAM1 contributes to the development of neuropathy is currently unclear. The aim of this study was to induce LRSAM1 deficiency in a neuronal cell model, observe its effect on cell growth and morphology and attempt to rescue the phenotype with ancestral and mutant LRSAM1 transfections. In this experimental study, we investigated the effect of LRSAM1 downregulation on neuroblastoma SH-SY5Y cells by siRNA technology where cells were transfected with siRNA against LRSAM1. The effects on the expression levels of TSG101, the only currently known LRSAM1 interacting molecule, were also examined. An equal dosage of ancestral or mutant LRSAM1 construct was transfected in LRSAM1-downregulated cells to investigate its effect on the phenotype of the cells and whether cell proliferation and morphology could be rescued. A significant reduction in TSG101 levels was observed with the downregulation of LRSAM1. In addition, LRSAM1 knockdown significantly decreased the growth rate of SH-SY5Y cells which is caused by a decrease in cell proliferation. An effect on cell morphology was also observed. Furthermore, we overexpressed the ancestral and the c.2047-1G>A mutant LRSAM1 in knocked down cells. Ancestral LRSAM1 recovered cell proliferation and partly the morphology, however, the c.2047-1G>A mutant did not recover cell proliferation and further aggravated the observed changes in cell morphology. Our findings suggest that depletion of LRSAM1 affects neuroblastoma cells growth and morphology and that overexpression of the c.2047-1G>A mutant form, unlike the ancestral LRSAM1, fails to rescue the phenotype. Copyright© by Royan Institute. All rights reserved.
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IGF-II and IGFBP-6 regulate cellular contractility and proliferation in Dupuytren's disease.
Raykha, Christina; Crawford, Justin; Gan, Bing Siang; Fu, Ping; Bach, Leon A; O'Gorman, David B
2013-10-01
Dupuytren's disease (DD) is a common and heritable fibrosis of the palmar fascia that typically manifests as permanent finger contractures. The molecular interactions that induce the development of hyper-contractile fibroblasts, or myofibroblasts, in DD are poorly understood. We have identified IGF2 and IGFBP6, encoding insulin-like growth factor (IGF)-II and IGF binding protein (IGFBP)-6 respectively, as reciprocally dysregulated genes and proteins in primary cells derived from contracture tissues (DD cells). Recombinant IGFBP-6 inhibited the proliferation of DD cells, patient-matched control (PF) cells and normal palmar fascia (CT) cells. Co-treatments with IGF-II, a high affinity IGFBP-6 ligand, were unable to rescue these effects. A non-IGF-II binding analog of IGFBP-6 also inhibited cellular proliferation, implicating IGF-II-independent roles for IGFBP-6 in this process. IGF-II enhanced the proliferation of CT cells, but not DD or PF cells, and significantly enhanced DD and PF cell contractility in stressed collagen lattices. While IGFBP-6 treatment did not affect cellular contractility, it abrogated the IGF-II-induced contractility of DD and PF cells in stressed collagen lattices. IGF-II also significantly increased the contraction of DD cells in relaxed lattices, however this effect was not evident in relaxed collagen lattices containing PF cells. The disparate effects of IGF-II on DD and PF cells in relaxed and stressed contraction models suggest that IGF-II can enhance lattice contractility through more than one mechanism. This is the first report to implicate IGFBP-6 as a suppressor of cellular proliferation and IGF-II as an inducer of cellular contractility in this connective tissue disease. Copyright © 2013 The Authors. Published by Elsevier B.V. All rights reserved.
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Aoyama, Yuki; Toriumi, Kazuya; Mouri, Akihiro; Hattori, Tomoya; Ueda, Eriko; Shimato, Akane; Sakakibara, Nami; Soh, Yuka; Mamiya, Takayoshi; Nagai, Taku; Kim, Hyoung-Chun; Hiramatsu, Masayuki; Nabeshima, Toshitaka; Yamada, Kiyofumi
2016-01-01
Cigarette smoking during pregnancy is associated with various disabilities in the offspring such as attention deficit/hyperactivity disorder, learning disabilities, and persistent anxiety. We have reported that nicotine exposure in female mice during pregnancy, in particular from embryonic day 14 (E14) to postnatal day 0 (P0), induces long-lasting behavioral deficits in offspring. However, the mechanism by which prenatal nicotine exposure (PNE) affects neurodevelopment, resulting in behavioral deficits, has remained unclear. Here, we report that PNE disrupted the proliferation of neuronal progenitors, leading to a decrease in the progenitor pool in the ventricular and subventricular zones. In addition, using a cumulative 5-bromo-2′-deoxyuridine labeling assay, we evaluated the rate of cell cycle progression causing the impairment of neuronal progenitor proliferation, and uncovered anomalous cell cycle kinetics in mice with PNE. Accordingly, the density of glutamatergic neurons in the medial prefrontal cortex (medial PFC) was reduced, implying glutamatergic dysregulation. Mice with PNE exhibited behavioral impairments in attentional function and behavioral flexibility in adulthood, and the deficits were ameliorated by microinjection of D-cycloserine into the PFC. Collectively, our findings suggest that PNE affects the proliferation and maturation of progenitor cells to glutamatergic neuron during neurodevelopment in the medial PFC, which may be associated with cognitive deficits in the offspring. PMID:26105135
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Zamorina, S A; Litvinova, L S; Yurova, K A; Khaziakhmatova, O G; Timganova, V P; Bochkova, M S; Khramtsov, P V; Rayev, M B
2018-01-01
The role of human chorionic gonadotropin (hCG) in the regulation of molecular genetics factors determining the functional activity of human naïve and memory T cells in vitro was studied. It was found that hCG (10 and 100IU/ml) inhibited CD28 and CD25 expression on the naïve T cells (CD45RA+) and CD25 expression on the memory T cells (CD45R0+). hCG didn't affect the CD71 proliferation marker expression in total. Nevertheless, hCG reduced the percentage of proliferating memory T cells with simultaneous suppression of CD71 expression on proliferating CD45R0+cells. In parallel, expression of U2af1l4, Gfi1, and hnRNPLL genes, which are Ptprc gene alternative splicing regulators was evaluated. It was established that hCG stimulated the expression of U2af1l4 and hnRNPLL genes, responsible for the assembly of CD45R0 in memory T cells, but reduced the expression of Gfi1 in these cells. In general, hCG promotes the differentiation of memory T cells by increasing of CD45R0 expression, but inhibits proliferation and CD25 expression which reflects their functional activity. Copyright © 2017 Elsevier B.V. All rights reserved.
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Attoff, K; Kertika, D; Lundqvist, J; Oredsson, S; Forsby, A
2016-09-01
Acrylamide is a well-known neurotoxic compound and people get exposed to the compound by food consumption and environmental pollutants. Since acrylamide crosses the placenta barrier, the fetus is also being exposed resulting in a risk for developmental neurotoxicity. In this study, the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y were used to study proliferation and differentiation as alerting indicators for developmental neurotoxicity. For both cell lines, acrylamide reduced the number of viable cells by reducing proliferation and inducing cell death in undifferentiated cells. Acrylamide concentrations starting at 10fM attenuated the differentiation process in SH-SY5Y cells by sustaining cell proliferation and neurite outgrowth was reduced at concentrations from 10pM. Acrylamide significantly reduced the number of neurons starting at 1μM and altered the ratio between the different phenotypes in differentiating C17.2 cell cultures. Ten micromolar of acrylamide also reduced the expression of the neuronal and astrocyte biomarkers. Although the neurotoxic concentrations in the femtomolar range seem to be specific for the SH-SY5Y cell line, the fact that micromolar concentrations of acrylamide seem to attenuate the differentiation process in both cell lines raises the interest to further investigations on the possible developmental neurotoxicity of acrylamide. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.
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Babkair, Hamzah; Yamazaki, Manabu; Uddin, Md Shihab; Maruyama, Satoshi; Abé, Tatsuya; Essa, Ahmed; Sumita, Yoshimasa; Ahsan, Md Shahidul; Swelam, Wael; Cheng, Jun; Saku, Takashi
2016-11-01
We reported that altered cell contact mediated by E-cadherin is an initial event in the pathogenesis of oral epithelial malignancies. To assess other effects of cell adhesion, we examined the expression levels of tight junction (TJ) molecules in oral carcinoma in situ (CIS) and squamous cell carcinoma (SCC). To identify changes in the expression of TJ molecules, we conducted an analysis of the immunohistochemical profiles of claudin-1 (CLDN-1) and zonula occludens-1 (ZO-1) in surgical specimens acquired from patients with oral SCC containing foci of epithelial dysplasia or from patients with CIS. We used immunofluorescence, Western blotting, reverse-transcription polymerase chain reaction, and RNA interference to evaluate the functions of CLDN-1 and ZO-1 in cultured oral SCC cells. TJ molecules were not detected in normal oral epithelial tissues but were expressed in SCC/CIS cells. ZO-1 was localized within the nucleus of proliferating cells. When CLDN-1 expression was inhibited by transfecting cells with specific small interference RNAs, SCC cells dissociated, and their ability to proliferate and invade Matrigel was inhibited. In contrast, although RNA interference-mediated inhibition of ZO-1 expression did not affect cell morphology, it inhibited cell proliferation and invasiveness. Our findings indicated that the detection of TJ molecules in the oral epithelia may serve as a marker for the malignant phenotype of cells in which CLDN-1 regulates proliferation and invasion. Copyright © 2016 Elsevier Inc. All rights reserved.
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Interdependence of cell growth and gene expression: origins and consequences.
Scott, Matthew; Gunderson, Carl W; Mateescu, Eduard M; Zhang, Zhongge; Hwa, Terence
2010-11-19
In bacteria, the rate of cell proliferation and the level of gene expression are intimately intertwined. Elucidating these relations is important both for understanding the physiological functions of endogenous genetic circuits and for designing robust synthetic systems. We describe a phenomenological study that reveals intrinsic constraints governing the allocation of resources toward protein synthesis and other aspects of cell growth. A theory incorporating these constraints can accurately predict how cell proliferation and gene expression affect one another, quantitatively accounting for the effect of translation-inhibiting antibiotics on gene expression and the effect of gratuitous protein expression on cell growth. The use of such empirical relations, analogous to phenomenological laws, may facilitate our understanding and manipulation of complex biological systems before underlying regulatory circuits are elucidated.
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A novel method for isolation of epithelial cells from ovine esophagus for tissue engineering.
Macheiner, Tanja; Kuess, Anna; Dye, Julian; Saxena, Amulya K
2014-01-01
The yield of a critical number of basal epithelial cells with high mitotic rates from native tissue is a challenge in the field of tissue engineering. There are many protocols that use enzymatic methods for isolation of epithelial cells with unsatisfactory results for tissue engineering. This study aimed to develop a protocol for isolating a sufficient number of epithelial cells with a high Proliferating Index from ovine esophagus for tissue engineering applications. Esophageal mucosa was pretreated with dispase-collagenase solution and plated on collagen-coated culture dishes. Distinction of the various types of epithelial cells and developmental stages was done with specific primary antibodies to Cytokeratins and to Proliferating Cell Nuclear Antigen (PCNA). Up to approximately 8100 epithelial cells/mm2 of mucosa tissue were found after one week of migration. Cytokeratin 14 (CK 14) was positive identified in cells even after 83 days. At the same time the Proliferating Index was 71%. Our protocol for isolation of basal epithelial cells was successful to yield sufficient numbers of cells predominantly with proliferative character and without noteworthy negative enzymatic affection. The results at this study offer the possibility of generation critical cell numbers for tissue engineering applications.
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A simple non-perturbing cell migration assay insensitive to proliferation effects.
Glenn, Honor L; Messner, Jacob; Meldrum, Deirdre R
2016-08-18
Migration is a fundamental cellular behavior that plays an indispensable role in development and homeostasis, but can also contribute to pathology such as cancer metastasis. Due to its relevance to many aspects of human health, the ability to accurately measure cell migration is of broad interest, and numerous approaches have been developed. One of the most commonly employed approaches, because of its simplicity and throughput, is the exclusion zone assay in which cells are allowed to migrate into an initially cell-free region. A major drawback of this assay is that it relies on simply counting cells in the exclusion zone and therefore cannot distinguish the effects of proliferation from migration. We report here a simple modification to the exclusion zone migration assay that exclusively measures cell migration and is not affected by proliferation. This approach makes use of a lineage-tracing vital stain that is retained through cell generations and effectively reads out migration relative to the original, parental cell population. This modification is simple, robust, non-perturbing, and inexpensive. We validate the method in a panel of cell lines under conditions that inhibit or promote migration and demonstrate its use in normal and cancer cell lines as well as primary cells.
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Roles of FGFR3 during morphogenesis of Meckel's cartilage and mandibular bones
Havens, Bruce A.; Velonis, Dimitris; Kronenberg, Mark S.; Lichtler, Alex C.; Oliver, Bonnie; Mina, Mina
2008-01-01
To address the functions of FGFR2 and FGFR3 signaling during mandibular skeletogenesis, we over-expressed in the developing chick mandible, replication-competent retroviruses carrying truncated FGFR2c or FGFR3c that function as dominant negative receptors (RCAS-dnFGFR2 and RCAS-dnFGFR3). Injection of RCAS-dnFGFR3 between HH15−20 led to reduced proliferation, increased apoptosis, and decreased differentiation of chondroblasts in Meckel's cartilage. These changes resulted in the formation of a hypoplastic mandibular process and truncated Meckel's cartilage. This treatment also affected the proliferation and survival of osteoprogenitor cells in osteogenic condensations, leading to the absence of five mandibular bones on the injected side. Injection of RCAS-dnFGFR2 between HH15−20 or RCAS-dnFGFR3 at HH26 did not affect the morphogenesis of Meckel's cartilage but resulted in truncations of the mandibular bones. RCAS-dnFGFR3 affected the proliferation and survival of the cells within the periosteum and osteoblasts. Together these results demonstrate that FGFR3 signaling is required for the elongation of Meckel's cartilage and FGFR2 and FGFR3 have roles during intramembranous ossification of mandibular bones. PMID:18339367
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Song, Yuanhui; Ju, Yang; Song, Guanbin; Morita, Yasuyuki
2013-01-01
Cell adhesion, migration, and proliferation are significantly affected by the surface topography of the substrates on which the cells are cultured. Alumina is one of the most popular implant materials used in orthopedics, but few data are available concerning the cellular responses of mesenchymal stem cells (MSCs) grown on nanoporous structures. MSCs were cultured on smooth alumina substrates and nanoporous alumina substrates to investigate the interaction between surface topographies of nanoporous alumina and cellular behavior. Nanoporous alumina substrates with pore sizes of 20 nm and 100 nm were used to evaluate the effect of pore size on MSCs as measured by proliferation, morphology, expression of integrin β1, and osteogenic differentiation. An MTT assay was used to measure cell viability of MSCs on different substrates, and determined that cell viability decreased with increasing pore size. Scanning electron microscopy was used to investigate the effect of pore size on cell morphology. Extremely elongated cells and prominent cell membrane protrusions were observed in cells cultured on alumina with the larger pore size. The expression of integrin β1 was enhanced in MSCs cultured on porous alumina, revealing that porous alumina substrates were more favorable for cell growth than smooth alumina substrates. Higher levels of osteoblastic differentiation markers such as alkaline phosphatase, osteocalcin, and mineralization were detected in cells cultured on alumina with 100 nm pores compared with cells cultured on alumina with either 20 nm pores or smooth alumina. This work demonstrates that cellular behavior is affected by variation in pore size, providing new insight into the potential application of this novel biocompatible material for the developing field of tissue engineering. PMID:23935364
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Physiological characterization of ocular melanosis-affected canine melanocytes.
Dawson-Baglien, Ethan M; Noland, Erica L; Sledge, Dodd G; Kiupel, Matti; Petersen-Jones, Simon M
2018-04-27
Cairn terriers with ocular melanosis (OM) accumulate large, heavily pigmented melanocytes in the anterior uvea. Darkly pigmented plaques develop within the sclera, leading us to hypothesize that OM uveal melanocytes may have an abnormal migratory capacity. Globes from OM-affected Cairn terriers and unaffected control eyes enucleated for reasons unrelated to this study were used for immunohistochemistry and to culture melanocytes for in vitro cell behavior assays. The scleral plaques of six dogs were immunolabeled for HMB-45, MelanA, PNL2, CD18, CD204, and Iba-1 and compared with the pigment cells accumulated within the irides. Cultured uveal melanocytes from OM-affected and control dogs were compared using conventional assays measuring cell proliferation, invasion capability, and melanin production. Melanocytes isolated from OM eyes had significantly elevated levels of per-cell melanin content and production compared to controls. The majority of pigmented cells in the scleral plaques were HMB45 positive indicating a melanocytic origin. Many were also CD18 positive. No differences were observed between cultured melanocytes from OM-affected and control uvea for standard in vitro proliferation or invasion assays. Pigmented cells which accumulate in the sclera of OM-affected Cairn terriers are predominantly melanocytes; however, in vitro assays of uveal melanocytes did not reveal differences in migratory behavior between OM and control cells. Migratory behavior of OM-melanocytes may be environment-dependent. We suggest that RNA sequencing and differential expression analysis would be a useful next step in understanding this disease. © 2018 American College of Veterinary Ophthalmologists.
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PKC{eta} is a negative regulator of AKT inhibiting the IGF-I induced proliferation
DOE Office of Scientific and Technical Information (OSTI.GOV)
Shahaf, Galit; Rotem-Dai, Noa; Koifman, Gabriela
2012-04-15
The PI3K-AKT pathway is frequently activated in human cancers, including breast cancer, and its activation appears to be critical for tumor maintenance. Some malignant cells are dependent on activated AKT for their survival; tumors exhibiting elevated AKT activity show sensitivity to its inhibition, providing an Achilles heel for their treatment. Here we show that the PKC{eta} isoform is a negative regulator of the AKT signaling pathway. The IGF-I induced phosphorylation on Ser473 of AKT was inhibited by the PKC{eta}-induced expression in MCF-7 breast adenocarcinoma cancer cells. This was further confirmed in shRNA PKC{eta}-knocked-down MCF-7 cells, demonstrating elevated phosphorylation on AKTmore » Ser473. While PKC{eta} exhibited negative regulation on AKT phosphorylation it did not alter the IGF-I induced ERK phosphorylation. However, it enhanced ERK phosphorylation when stimulated by PDGF. Moreover, its effects on IGF-I/AKT and PDGF/ERK pathways were in correlation with cell proliferation. We further show that both PKC{eta} and IGF-I confer protection against UV-induced apoptosis and cell death having additive effects. Although the protective effect of IGF-I involved activation of AKT, it was not affected by PKC{eta} expression, suggesting that PKC{eta} acts through a different route to increase cell survival. Hence, our studies show that PKC{eta} provides negative control on AKT pathway leading to reduced cell proliferation, and further suggest that its presence/absence in breast cancer cells will affect cell death, which could be of therapeutic value.« less
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Dunn, S.-J.; Osborne, J. M.; Appleton, P. L.; Näthke, I.
2016-01-01
Curative intervention is possible if colorectal cancer is identified early, underscoring the need to detect the earliest stages of malignant transformation. A candidate biomarker is the expanded proliferative zone observed in crypts before adenoma formation, also found in irradiated crypts. However, the underlying driving mechanism for this is not known. Wnt signaling is a key regulator of proliferation, and elevated Wnt signaling is implicated in cancer. Nonetheless, how cells differentiate Wnt signals of varying strengths is not understood. We use computational modeling to compare alternative hypotheses about how Wnt signaling and contact inhibition affect proliferation. Direct comparison of simulations with published experimental data revealed that the model that best reproduces proliferation patterns in normal crypts stipulates that proliferative fate and cell cycle duration are set by the Wnt stimulus experienced at birth. The model also showed that the broadened proliferation zone induced by tumorigenic radiation can be attributed to cells responding to lower Wnt concentrations and dividing at smaller volumes. Application of the model to data from irradiated crypts after an extended recovery period permitted deductions about the extent of the initial insult. Application of computational modeling to experimental data revealed how mechanisms that control cell dynamics are altered at the earliest stages of carcinogenesis. PMID:27053661
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Proliferation Rates of Bovine Primary Muscle Cells Relate to Liveweight and Carcase Weight in Cattle
Coles, Chantal A.; Wadeson, Jenny; Leyton, Carolina P.; Siddell, Jason P.; Greenwood, Paul L.; White, Jason D.; McDonagh, Matthew B.
2015-01-01
Muscling in cattle is largely influenced by genetic background, ultimately affecting beef yield and is of major interest to the beef industry. This investigation aimed to determine whether primary skeletal muscle cells isolated from different breeds of cattle with a varying genetic potential for muscling differ in their myogenic proliferative capacity. Primary skeletal muscle cells were isolated and cultured from the Longissimus muscle (LM) of 6 month old Angus, Hereford and Wagyu X Angus cattle. Cells were assessed for rate of proliferation and gene expression of PAX7, MYOD, MYF5, and MYOG. Proliferation rates were found to differ between breeds of cattle whereby myoblasts from Angus cattle were found to proliferate at a greater rate than those of Hereford and Wagyu X Angus during early stages of growth (5–20 hours in culture) in vitro (P < 0.05). The proliferation rates of myoblasts during early stages of culture in vitro were also found to be positively related to the liveweight and carcase weight of cattle (P < 0.05). Gene expression of MYF5 was also found to be significantly down-regulated in WagyuX compared with Angus cattle (P < 0.05). These findings suggest that early events during myogenesis are important for determining liveweight and caracase weights in cattle. PMID:25875203
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Sheng, Lingling; Mao, Xiyuan; Yu, Qingxiong; Yu, Dong
2017-01-01
Bone marrow-derived mesenchymal stem cell (BM-MSC) transplantation has been demonstrated to be an effective way of augmenting angiogenesis of ischemic tissue. The low oxygen conditions in ischemic tissue directly affect the biological behavior of engrafted cells. However, to date, the mechanism through which hypoxia regulates self-renewal, differentiation and paracrine function of BM-MSCs remains unclear. Clarification of this mechanism would be beneficial to the use of stem cell-based therapy. The PI3K/AKT pathway has been extensively investigated for its role in cell proliferation, cell transformation, paracrine function and angiogenesis. The present study aimed to analyze the role of PI3K/AKT pathway in hypoxia-induced proliferation of BM-MSCs and their differentiation into endothelial cells in vitro by the application of LY294002, a PI3K/AKT pathway inhibitor, with cells cultured in normoxia serving as a control. The results showed that rat BM-MSCs at passage 3 and 4 displayed only few phenotypical differences in the expression of surface antigens as detected by flow cytometry. When compared with the cells treated in normoxia, the proliferation of BM-MSCs in hypoxia was promoted, a greater number of cells expressed CD31 and a higher expression of vascular endothelial growth factor was observed after culture in hypoxic conditions. However, by inhibiting with LY294002, these changes induced by hypoxia were partly inhibited. In conclusion, the present study showed that the PI3K/AKT pathway served an important role in hypoxia-enhanced in vitro proliferation of BM-MSCs and their differentiation into endothelial cells and paracrine vascular endothelial growth factor. PMID:28123468
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Splicing factors PTBP1 and PTBP2 promote proliferation and migration of glioma cell lines
Cheung, Hannah C.; Hai, Tao; Zhu, Wen; Baggerly, Keith A.; Tsavachidis, Spiridon; Krahe, Ralf
2009-01-01
Polypyrimidine tract-binding protein 1 (PTBP1) is a multi-functional RNA-binding protein that is aberrantly overexpressed in glioma. PTBP1 and its brain-specific homologue polypyrimidine tract-binding protein 2 (PTBP2) regulate neural precursor cell differentiation. However, the overlapping and non-overlapping target transcripts involved in this process are still unclear. To determine why PTBP1 and not PTBP2 would promote glial cell-derived tumours, both PTBP1 and PTBP2 were knocked down in the human glioma cell lines U251 and LN229 to determine the role of these proteins in cell proliferation, migration, and adhesion. Surprisingly, removal of both PTBP1 and PTBP2 slowed cell proliferation, with the double knockdown having no additive effects. Decreased expression of both proteins individually and in combination inhibited cell migration and increased adhesion of cells to fibronectin and vitronectin. A global survey of differential exon expression was performed following PTBP1 knockdown in U251 cells using the Affymetrix Exon Array to identify PTBP1-specific splicing targets that enhance gliomagenesis. In the PTBP1 knockdown, previously determined targets were unaltered in their splicing patterns. A single gene, RTN4 (Nogo) had significantly enhanced inclusion of exon 3 when PTBP1 was removed. Overexpression of the splice isoform containing exon 3 decreased cell proliferation to a similar degree as the removal of PTBP1. These results provide the first evidence that RNA-binding proteins affect the invasive and rapid growth characteristics of glioma cell lines. Its actions on proliferation appear to be mediated, in part, through alternative splicing of RTN4. PMID:19506066
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Rallis, Charalampos; Codlin, Sandra; Bähler, Jürg
2013-08-01
Target of rapamycin complex 1 (TORC1) is implicated in growth control and aging from yeast to humans. Fission yeast is emerging as a popular model organism to study TOR signaling, although rapamycin has been thought to not affect cell growth in this organism. Here, we analyzed the effects of rapamycin and caffeine, singly and combined, on multiple cellular processes in fission yeast. The two drugs led to diverse and specific phenotypes that depended on TORC1 inhibition, including prolonged chronological lifespan, inhibition of global translation, inhibition of cell growth and division, and reprograming of global gene expression mimicking nitrogen starvation. Rapamycin and caffeine differentially affected these various TORC1-dependent processes. Combined drug treatment augmented most phenotypes and effectively blocked cell growth. Rapamycin showed a much more subtle effect on global translation than did caffeine, while both drugs were effective in prolonging chronological lifespan. Rapamycin and caffeine did not affect the lifespan via the pH of the growth media. Rapamycin prolonged the lifespan of nongrowing cells only when applied during the growth phase but not when applied after cells had stopped proliferation. The doses of rapamycin and caffeine strongly correlated with growth inhibition and with lifespan extension. This comprehensive analysis will inform future studies into TORC1 function and cellular aging in fission yeast and beyond. © 2013 The Authors. Aging Cell published by John Wiley & Sons Ltd and the Anatomical Society.
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Ruggiu, Alessandra; Ulivi, Valentina; Sanguineti, Francesca; Cancedda, Ranieri; Descalzi, Fiorella
2013-12-01
Platelet Lysate (PL) contains a cocktail of growth factors and cytokines, which actively participates in tissue repair and its clinical application has been broadly described. The aim of this study was to assess the regenerative potential of PL for bone repair. We demonstrated that PL stimulation induces a transient increase of the inflammatory response in quiescent human osteoblasts, via NF-kB activation, COX-2 induction, PGE2 production and secretion of pro-inflammatory cytokines. Furthermore, we showed that long-term PL stimulation enhances proliferation of actively replicating osteoblasts, without affecting their differentiation potential, along with changes of cell morphology, resulting in increased cell density at confluence. In confluent resting osteoblasts, PL treatment induced resumption of proliferation, change in cell morphology and increase of cell density at confluence. A burst of PL treatment (24-h) was sufficient to trigger such processes in both conditions. These results correlated with up-regulation of the proliferative and survival pathways ERKs and Akt and with cell cycle re-activation via induction of CyclinD1 and phosphorylation of Rb, following PL stimulation. Our findings demonstrate that PL treatment results in activation and expansion of resting osteoblasts, without affecting their differentiation potential. Therefore PL represents a good therapeutic candidate in regenerative medicine for bone repair. Copyright © 2013 Elsevier Ltd. All rights reserved.
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2013-01-01
Background Experimental clinical stem cell therapy has been used for more than a decade to alleviate the adverse aftermath of acute myocardial infarction (aMI). The post-infarcted myocardial microenvironment is characterized by cardiomyocyte death, caused by ischemia and inflammation. These conditions may negatively affect administered stem cells. As postnatal cardiomyocytes have a poor proliferation rate, while induction of proliferation seems even more rare. Thus stimulation of their proliferation rate is essential after aMI. In metaplastic disease, the pro-inflammatory cytokine interleukin-6 (IL-6) has been identified as potent mediators of the proliferation rate. We hypothesized that IL-6 could augment the proliferation rate of (slow-)dividing cardiomyocytes. Methods To mimic the behavior of therapeutic cells in the post-infarct cardiac microenvironment, human Adipose Derived Stromal Cells (ADSC) were cultured under hypoxic (2% O2) and pro-inflammatory conditions (IL-1β) for 24h. Serum-free conditioned medium from ADSC primed with hypoxia and/or IL-1β was added to rat neonatal cardiomyocytes and adult cardiomyocytes (HL-1) to assess paracrine-driven changes in cardiomyocyte proliferation rate and induction of myogenic signaling pathways. Results We demonstrate that ADSC enhance the proliferation rate of rat neonatal cardiomyocytes and adult HL-1 cardiomyocytes in a paracrine fashion. ADSC under hypoxia and inflammation in vitro had increased the interleukin-6 (IL-6) gene and protein expression. Similar to conditioned medium of ADSC, treatment of rat neonatal cardiomyocytes and HL-1 with recombinant IL-6 alone also stimulated their proliferation rate. This was corroborated by a strong decrease of cardiomyocyte proliferation after addition of IL-6 neutralizing antibody to conditioned medium of ADSC. The stimulatory effect of ADSC conditioned media or IL-6 was accomplished through activation of both Janus Kinase-Signal Transducer and Activator of Transcription (JAK/STAT) and Mitogen-Activated Protein (MAP) kinases (MAPK) mitogenic signaling pathways. Conclusion ADSC are promising therapeutic cells for cardiac stem cell therapy. The inflammatory and hypoxic host post-MI microenvironment enhances the regenerative potential of ADSC to promote the proliferation rate of cardiomyocytes. This was achieved in paracrine manner, which warrants the development of ADSC conditioned medium as an “of-the-shelf” product for treatment of post-myocardial infarction complications. PMID:23406316
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Kim, Hyun-Ju, E-mail: biohjk@knu.ac.kr; Yoon, Hye-Jin; Yoon, Kyung-Ae
Lipocalin-2 (LCN2) is a member of the lipocalin superfamily and plays a critical role in the regulation of various physiological processes, such as inflammation and obesity. In this study, we report that LCN2 negatively modulates the proliferation and differentiation of osteoclast precursors, resulting in impaired osteoclast formation. The overexpression of LCN2 in bone marrow-derived macrophages or the addition of recombinant LCN2 protein inhibits the formation of multinuclear osteoclasts. LCN2 suppresses macrophage colony-stimulating factor (M-CSF)-induced proliferation of osteoclast precursor cells without affecting their apoptotic cell death. Interestingly, LCN2 decreases the expression of the M-CSF receptor, c-Fms, and subsequently blocks its downstreammore » signaling cascades. In addition, LCN2 inhibits RANKL-induced osteoclast differentiation and attenuates the expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which are important modulators in osteoclastogenesis. Mechanistically, LCN2 inhibits NF-κB signaling pathways, as demonstrated by the suppression of IκBα phosphorylation, nuclear translocation of p65, and NF-κB transcriptional activity. Thus, LCN2 is an anti-osteoclastogenic molecule that exerts its effects by retarding the proliferation and differentiation of osteoclast lineage cells. - Highlights: • LCN2 expression is regulated during osteoclast development. • LCN2 suppresses M-CSF-mediated osteoclast precursor proliferation. • LCN2 inhibits RANKL-induced osteoclast differentiation.« less
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p53 Mutation suppresses adult neurogenesis in medaka fish (Oryzias latipes)
DOE Office of Scientific and Technical Information (OSTI.GOV)
Isoe, Yasuko; Okuyama, Teruhiro; Taniguchi, Yoshihito
2012-07-13
Highlights: Black-Right-Pointing-Pointer Progenitor migration is accompanied by an increase in their numbers in the adult brain. Black-Right-Pointing-Pointer p53 Mutation suppressed an increase in the number of the migrated progenitors. Black-Right-Pointing-Pointer The decreased progenitor number is not due to enhanced cell death. Black-Right-Pointing-Pointer p53 Mutation did not affect proliferation of stem cells. -- Abstract: Tumor suppressor p53 negatively regulates self-renewal of neural stem cells in the adult murine brain. Here, we report that the p53 null mutation in medaka fish (Oryzias latipes) suppressed neurogenesis in the telencephalon, independent of cell death. By using 5-bromo-29-deoxyuridine (BrdU) immunohistochemistry, we identified 18 proliferation zonesmore » in the brains of young medaka fish; in situ hybridization showed that p53 was expressed selectively in at least 12 proliferation zones. We also compared the number of BrdU-positive cells present in the whole telencephalon of wild-type (WT) and p53 mutant fish. Immediately after BrdU exposure, the number of BrdU-positive cells did not differ significantly between them. One week after BrdU-exposure, the BrdU-positive cells migrated from the proliferation zone, which was accompanied by an increased number in the WT brain. In contrast, no significant increase was observed in the p53 mutant brain. Terminal deoxynucleotidyl transferase (dUTP) nick end-labeling revealed that there was no significant difference in the number of apoptotic cells in the telencephalon of p53 mutant and WT medaka, suggesting that the decreased number of BrdU-positive cells in the mutant may be due to the suppression of proliferation rather than the enhancement of neural cell death. These results suggest that p53 positively regulates neurogenesis via cell proliferation.« less
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Olguín, Yusser; Campos, Cristian; Catalán, Javiera; Velásquez, Luís; Osorio, Fernando; Montenegro, Iván; Madrid, Alejandro; Acevedo, Cristian
2017-09-22
Advances in the generation of suitable thermosensitive hydrogels for the delivery of cells in neural tissue engineering demonstrate a delicate relationship between physical properties and capabilities to promote cell proliferation and differentiation. To improve the properties of these materials, it is possible to add liposomes for the controlled release of bioactive elements, which in turn can affect the physical and biological properties of the hydrogels. In the present investigation, different hydrogels based on Pluronic F127 have been formulated with the incorporation of chitosan and two types of liposomes of two different sizes. The rheological and thermal properties and their relation with the neurite proliferation and growth of the PC12 cell line were evaluated. Our results show that the incorporation of liposomes modifies the properties of the hydrogels dependent on the concentration of chitosan and the lipid type in the liposomes, which directly affect the capabilities of the hydrogels to promote the viability and differentiation of PC12 cells.
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Olguín, Yusser; Campos, Cristian; Catalán, Javiera; Velásquez, Luís; Osorio, Fernando; Montenegro, Iván; Madrid, Alejandro; Acevedo, Cristian
2017-01-01
Advances in the generation of suitable thermosensitive hydrogels for the delivery of cells in neural tissue engineering demonstrate a delicate relationship between physical properties and capabilities to promote cell proliferation and differentiation. To improve the properties of these materials, it is possible to add liposomes for the controlled release of bioactive elements, which in turn can affect the physical and biological properties of the hydrogels. In the present investigation, different hydrogels based on Pluronic F127 have been formulated with the incorporation of chitosan and two types of liposomes of two different sizes. The rheological and thermal properties and their relation with the neurite proliferation and growth of the PC12 cell line were evaluated. Our results show that the incorporation of liposomes modifies the properties of the hydrogels dependent on the concentration of chitosan and the lipid type in the liposomes, which directly affect the capabilities of the hydrogels to promote the viability and differentiation of PC12 cells. PMID:28937646
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Combinatorial growth of oxide nanoscaffolds and its influence in osteoblast cell adhesion
NASA Astrophysics Data System (ADS)
Acevedo-Morantes, Claudia Y.; Irizarry-Ortiz, Roberto A.; Caceres-Valencia, Pablo G.; Singh, Surinder P.; Ramirez-Vick, Jaime E.
2012-05-01
We report a novel method for high-throughput investigations on cell-material interactions based on metal oxide nanoscaffolds. These scaffolds possess a continuous gradient of various titanium alloys allowing the compositional and morphological variation that could substantially improve the formation of an osseointegrative interface with bone. The model nanoscaffold has been fabricated on commercially pure titanium (cp-Ti) substrate with a compositional gradients of tin (Sn), chromium (Cr), and niobium (Nb) deposited using a combinatorial approach followed by annealing to create native oxide surface. As an invitro test system, the human fetal osteoblastic cell line (hFOB 1.19) has been used. Cell-adhesion of hFOB 1.19 cells and the suitability of these alloys have been evaluated for cell-morphology, cell-number, and protein adsorption. Although, cell-morphology was not affected by surface composition, cell-proliferation rates varied significantly with surface metal oxide composition; with the Sn- and Nb-rich regions showing the highest proliferation rate and the Cr-rich regions presenting the lowest. The results suggest that Sn and Nb rich regions on surface seems to promote hFOB 1.19 cell proliferation and may therefore be considered as implant material candidates that deserve further analysis.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Konno, S.; Chiao, J.; Rossi, J.
1986-05-01
Addition of nicotine causes a dose- and time-dependent inhibition of cell growth in established human and murine cells. In the human promyelocytic HL-60 leukemic cells, 3 mM nicotine results in a 50% inhibition of cellular proliferation after 80 h. Nicotine was also found to affect the cell cycle distribution of HL-60 cells. Treatment with 4 mM nicotine for 20 h causes an increase in proportion of Gl-phase cells (from 49% to 57%) and a significant decrease in the proportion of S-phase cells (from 41% to 32%). These results suggest that nicotine causes cell arrest in the Gl-phase which may inmore » part account for its effects on cell growth. To determine whether nicotine has a primary effect on the uptake/transport of macromolecular precursors into cells, HL-60 cells were treated with 2-6 mM nicotine for 30 h/sub 3/ at the end of which time cells were labeled with (/sup 3/H)thymidine, (/sup 3/H)uridine, (/sup 14/C)lysine and (/sup 35/S)methionine, the trichloroacetic acid (TCA) soluble and insoluble radioactivities from each of the labeling conditions were determined. These studies show that nicotine primarily affect the synthesis of proteins.« less
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Strakova, Nicol; Ehrmann, Jiri; Dzubak, Petr; Bouchal, Jan; Kolar, Zdenek
2004-06-01
Glioblastoma multiforme is the most common malignant brain tumor in adults, and it is among the most lethal of all cancers. Recent studies have shown that ligand activation of peroxisome proliferator-activated receptor (PPAR)-gamma can induce differentiation and inhibit proliferation of several cancer cells. In this study, we have investigated whether one PPARgamma ligand in particular, ciglitazone, inhibits cell viability and, additionally, whether it affects the cell cycle and apoptosis of human glioblastoma cell lines T98G, U-87 MG, A172, and U-118 MG. All glioblastoma cell lines were found to express PPARgamma protein, and following treatment with ciglitazone, localization was unchanged. Ciglitazone inhibited viability in a dose-dependent manner in all four tested glioblastoma cells after 24 h of treatment. Analysis of the cell cycle showed arrest in the G(1) phase and partial block in G(2)/M phase of the cell cycle. Cyclin D1 and cyclin B expression was decreased. Phosphorylation of Rb protein dropped as well. We found that ciglitazone was followed by increased expression of p27(Kip1) and p21(Waf1/Cip1). It also led to apoptosis induction: bax expression in T98G was elevated. Expression of the antiapoptotic protein bcl-2 was reduced in U-118 MG and U-87 MG and showed a slight decrease in A172 cells. Flow cytometry confirmed the induction of apoptosis. Moreover, PPARgamma ligand decreased telomerase activity in U-87 MG and U-118 MG cell lines. Our results demonstrate that ciglitazone inhibits the viability of human glioblastoma cell lines via induction of apoptosis; as a result, this ligand may offer potential new therapy for the treatment of central nervous system neoplasms.
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Akt1 deficiency diminishes skeletal muscle hypertrophy by reducing satellite cell proliferation.
Moriya, Nobuki; Miyazaki, Mitsunori
2018-05-01
Skeletal muscle mass is determined by the net dynamic balance between protein synthesis and degradation. Although the Akt/mechanistic target of rapamycin (mTOR)-dependent pathway plays an important role in promoting protein synthesis and subsequent skeletal muscle hypertrophy, the precise molecular regulation of mTOR activity by the upstream protein kinase Akt is largely unknown. In addition, the activation of satellite cells has been indicated as a key regulator of muscle mass. However, the requirement of satellite cells for load-induced skeletal muscle hypertrophy is still under intense debate. In this study, female germline Akt1 knockout (KO) mice were used to examine whether Akt1 deficiency attenuates load-induced skeletal muscle hypertrophy through suppressing mTOR-dependent signaling and satellite cell proliferation. Akt1 KO mice showed a blunted hypertrophic response of skeletal muscle, with a diminished rate of satellite cell proliferation following mechanical overload. In contrast, Akt1 deficiency did not affect the load-induced activation of mTOR signaling and the subsequent enhanced rate of protein synthesis in skeletal muscle. These observations suggest that the load-induced activation of mTOR signaling occurs independently of Akt1 regulation and that Akt1 plays a critical role in regulating satellite cell proliferation during load-induced muscle hypertrophy.
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Citrus limonin and its glucoside inhibit colon adenocarcinoma cell proliferation through apoptosis.
Chidambara Murthy, Kotamballi N; Jayaprakasha, G K; Kumar, Vinod; Rathore, Keerti S; Patil, Bhimanagouda S
2011-03-23
The current study was an attempt to elucidate the mechanism of human colon cancer cell proliferation inhibition by limonin and limonin glucoside (LG) isolated from seeds of Citrus reticulata. The structures of purified compounds were confirmed by NMR and quantified using HPLC. These compounds of more than 95% purity were subjected to proliferation inhibition assay using human colon adenocarcinoma (SW480) cells. The IC50 value of 54.74 and 37.39 μM was observed for limonin and LG, respectively at 72 h. Following confirmation of proliferation inhibition, pattern of DNA fragmentation and activation of caspase-3 of the cells treated with limonoids suggest involvement of apoptosis. Furthermore, reduction in the transcription ratio of bcl2/bax and induction of cytochrome c release from mitochondria to cytosol with treatment of limonoids confirm the activation of intrinsic apoptosis pathway. The activity of Bax and Bcl2 was confirmed through analysis of mitochondrial membrane potential and intracellular calcium in the cells treated with limonin and LG; the net content of caspase-8 was not affected by limonoids. Results of the current study provide compelling evidence on the induction of mitochondria mediated intrinsic apoptosis by both limonin and LG in cultured SW480 cells for the first time.
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[Effects of ezrin silencing on pancreatic cancer cell line Panc-1].
Meng, Yun-xiao; Yu, Shuang-ni; Lu, Zhao-hui; Chen, Jie
2012-12-01
To explore the effects of ezrin silencing on pancreatic cancer cell line Panc-1. Pancreatic cancer cell line Panc-1 was transfected with ezrin silencing plasmid. The proliferation and the cell cycle status were determined by CCK-8 assay and flow cytometry analysis, respectively. Cellular membrane protrusions/microvilli formation were visualized by scanning election microscopy. Colony formation assay was used to determine the cell anchor-independent growth ability in vitro. Trans-filter migration and invasion assays were performed with 8 µm pore inserts in a 24-well BioCoat chamber with/without Matrigel. Ezrin silencing decreased cellular protrusions/microvilli formation, anchorage-independent growth, cell migration and invasion, but had no effects on cell proliferation in vitro and cell cycle, in pancreatic cancer cell line Panc-1. Ezrin expression affects the cellular protrusions/microvilli formation, anchorage-independent growth, cell migration and invasion in pancreatic cancer cell line Panc-1.
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Inhibition of cyclooxygenase (COX)-2 affects endothelial progenitor cell proliferation
DOE Office of Scientific and Technical Information (OSTI.GOV)
Colleselli, Daniela; Bijuklic, Klaudija; Mosheimer, Birgit A.
2006-09-10
Growing evidence indicates that inducible cyclooxygenase-2 (COX-2) is involved in the pathogenesis of inflammatory disorders and various types of cancer. Endothelial progenitor cells recruited from the bone marrow have been shown to be involved in the formation of new vessels in malignancies and discussed for being a key point in tumour progression and metastasis. However, until now, nothing is known about an interaction between COX and endothelial progenitor cells (EPC). Expression of COX-1 and COX-2 was detected by semiquantitative RT-PCR and Western blot. Proliferation kinetics, cell cycle distribution and rate of apoptosis were analysed by MTT test and FACS analysis.more » Further analyses revealed an implication of Akt phosphorylation and caspase-3 activation. Both COX-1 and COX-2 expression can be found in bone-marrow-derived endothelial progenitor cells in vitro. COX-2 inhibition leads to a significant reduction in proliferation of endothelial progenitor cells by an increase in apoptosis and cell cycle arrest. COX-2 inhibition leads further to an increased cleavage of caspase-3 protein and inversely to inhibition of Akt activation. Highly proliferating endothelial progenitor cells can be targeted by selective COX-2 inhibition in vitro. These results indicate that upcoming therapy strategies in cancer patients targeting COX-2 may be effective in inhibiting tumour vasculogenesis as well as angiogenic processes.« less
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APE/Ref-1 makes fine-tuning of CD40-induced B cell proliferation.
Merluzzi, Sonia; Gri, Giorgia; Gattei, Valter; Pagano, Michele; Pucillo, Carlo
2008-08-01
Apurinic/apyrimidinic endonuclease-1/Redox factor-1, a multifunctional DNA base excision repair and redox regulation enzyme, plays an important role in oxidative signalling, transcription factor regulation, and cell cycle control. Recently, we have demonstrated that following the triggering of CD40 on B cells, APE/Ref-1 translocates from the cytoplasm to the nucleus and regulates the activity of B cell-specific transcription factors. In the present paper we investigate whether APE/Ref-1 plays a role in controlling CD40-mediated B cell proliferation too. We demonstrate a concurrent increase in proliferation and decrease in apoptosis of primary mouse B cells activated by CD40 cross-linking and transfected with functional APE/Ref-1 antisense oligonucleotide. Moreover, we provide evidence that a redox-mediated signalling mechanism is involved in this process and we propose that APE/Ref-1, controlling the intracellular redox state, may also affect the cell cycle by inducing nucleus-cytoplasm redistribution of p21. Together, these findings suggest that APE/Ref-1 could act as a negative regulator in an adaptive response to elevated ROS levels following CD40 cross-linking. Considering the important role of ROS and APE/Ref-1 in CD40-mediated B cell proliferation, our data will contribute to understand the mechanisms of tumor escape and suggest APE/Ref-1 as a novel target for tumor therapeutic approaches.
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APE/Ref-1 makes fine-tuning of CD40-induced B cell proliferation
Merluzzi, Sonia; Gri, Giorgia; Gattei, Valter; Pagano, Michele; Pucillo, Carlo
2009-01-01
Apurinic/apyrimidinic endonuclease-1/Redox factor-1, a multifunctional DNA base excision repair and redox regulation enzyme, plays an important role in oxidative signalling, transcription factor regulation, and cell cycle control. Recently, we have demonstrated that following the triggering of CD40 on B cells, APE/Ref-1 translocates from the cytoplasm to the nucleus and regulates the activity of B cell-specific transcription factors. In the present paper we investigate whether APE/Ref-1 plays a role in controlling CD40-mediated B cell proliferation too. We demonstrate a concurrent increase in proliferation and decrease in apoptosis of primary mouse B cells activated by CD40 cross-linking and transfected with functional APE/Ref-1 antisense oligonucleotide. Moreover, we provide evidence that a redox-mediated signalling mechanism is involved in this process and we propose that APE/Ref-1, controlling the intracellular redox state, may also affect the cell cycle by inducing nucleus-cytoplasm redistribution of p21. Together, these findings suggest that APE/Ref-1 could act as a negative regulator in an adaptive response to elevated ROS levels following CD40 cross-linking. Considering the important role of ROS and APE/Ref-1 in CD40-mediated B cell proliferation, our data will contribute to understand the mechanisms of tumor escape and suggest APE/Ref-1 as a novel target for tumor therapeutic approaches. PMID:18617267
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Choi, Won Jin; Jung, Jongjin; Lee, Sujin; Chung, Yoon Jang; Yang, Cheol-Soo; Lee, Young Kuk; Lee, You-Seop; Park, Joung Kyu; Ko, Hyuk Wan; Lee, Jeong-O
2015-01-01
We demonstrate that ZnO films grown by atomic layer deposition (ALD) can be employed as a substrate to explore the effects of electrical conductivity on cell adhesion, proliferation, and morphogenesis. ZnO substrates with precisely tunable electrical conductivity were fabricated on glass substrates using ALD deposition. The electrical conductivity of the film increased linearly with increasing duration of the ZnO deposition cycle (thickness), whereas other physical characteristics, such as surface energy and roughness, tended to saturate at a certain value. Differences in conductivity dramatically affected the behavior of SF295 glioblastoma cells grown on ZnO films, with high conductivity (thick) ZnO films causing growth arrest and producing SF295 cell morphologies distinct from those cultured on insulating substrates. Based on simple electrostatic calculations, we propose that cells grown on highly conductive substrates may strongly adhere to the substrate without focal-adhesion complex formation, owing to the enhanced electrostatic interaction between cells and the substrate. Thus, the inactivation of focal adhesions leads to cell proliferation arrest. Taken together, the work presented here confirms that substrates with high conductivity disturb the cell-substrate interaction, producing cascading effects on cellular morphogenesis and disrupting proliferation, and suggests that ALD-grown ZnO offers a single-variable method for uniquely tailoring conductivity. PMID:25897486
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Balanced cell proliferation and expansion is essential for flowering stem growth control.
Ferjani, Ali; Hanai, Kenya; Gunji, Shizuka; Maeda, Saori; Sawa, Shinichiro; Tsukaya, Hirokazu
2015-01-01
The postembryonic development of aboveground plant organs relies on a continuous supply of cells from the shoot apical meristem. Previous studies of developmental regulation in leaves and flowers have revealed the crucial role of coordinated cell proliferation and differentiation during organogenesis. However, the importance of this coordination has not been examined in flowering stems. Very recently, we attempted to identify regulatory factors that maintain flowering stem integrity. We found that the increased cell number in clavata (clv) mutants and the decreased cell size in de-etiolated (det)3-1 resulted in flowering stems that were thicker and thinner, respectively, than in wild-type (WT) plants. Interestingly, in the cell proliferation- and cell expansion-defective double mutant clv det3-1, the flowering stems often exhibited severe cracking, resulting in exposure of their inner tissues. In this study, further quantification of the cellular phenotypes in the cotyledons and leaves revealed no differences between det3-1 and clv3 det3-1. Together, the above findings suggest that the clv3 mutation in a det3-1 background primarily affects flowering stems, while its effect on other organs is likely negligible. We propose that the coordination between cell proliferation and differentiation is not only important during leaf development, but also plays a role in the growth control of Arabidopsis flowering stems.
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Thompson, Christopher K.
2016-01-01
Thyroid hormone (TH) regulates many cellular events underlying perinatal brain development in vertebrates. Whether and how TH regulates brain development when neural circuits are first forming is less clear. Furthermore, although the molecular mechanisms that impose spatiotemporal constraints on TH action in the brain have been described, the effects of local TH signaling are poorly understood. We determined the effects of manipulating TH signaling on development of the optic tectum in stage 46–49 Xenopus laevis tadpoles. Global TH treatment caused large-scale morphological effects in tadpoles, including changes in brain morphology and increased tectal cell proliferation. Either increasing or decreasing endogenous TH signaling in tectum, by combining targeted DIO3 knockdown and methimazole, led to corresponding changes in tectal cell proliferation. Local increases in TH, accomplished by injecting suspensions of tri-iodothyronine (T3) in coconut oil into the midbrain ventricle or into the eye, selectively increased tectal or retinal cell proliferation, respectively. In vivo time-lapse imaging demonstrated that local TH first increased tectal progenitor cell proliferation, expanding the progenitor pool, and subsequently increased neuronal differentiation. Local T3 also dramatically increased dendritic arbor growth in neurons that had already reached a growth plateau. The time-lapse data indicate that the same cells are differentially sensitive to T3 at different time points. Finally, TH increased expression of genes pertaining to proliferation and neuronal differentiation. These experiments indicate that endogenous TH locally regulates neurogenesis at developmental stages relevant to circuit assembly by affecting cell proliferation and differentiation and by acting on neurons to increase dendritic arbor elaboration. SIGNIFICANCE STATEMENT Thyroid hormone (TH) is a critical regulator of perinatal brain development in vertebrates. Abnormal TH signaling in early pregnancy is associated with significant cognitive deficits in humans; however, it is difficult to probe the function of TH in early brain development in mammals because of the inaccessibility of the fetal brain in the uterine environment and the challenge of disambiguating maternal versus fetal contributions of TH. The external development of tadpoles allows manipulation and direct observation of the molecular and cellular mechanisms underlying TH's effects on brain development in ways not possible in mammals. We find that endogenous TH locally regulates neurogenesis at developmental stages relevant to circuit assembly by affecting neural progenitor cell proliferation and differentiation and by acting on neurons to enhance dendritic arbor elaboration. PMID:27707971
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Tossige-Gomes, Rosalina; Costa, Karine Beatriz; Ottone, Vinícius de Oliveira; Magalhães, Flávio de Castro; Amorim, Fabiano Trigueiro; Rocha-Vieira, Etel
2016-01-01
This study investigated whether an acute session of high-intensity interval training (HIIT) is sufficient to alter lymphocyte function and redox status. Sixteen young healthy men underwent a HIIT session on a cycloergometer, consisting of eight bouts of 1 min at 90-100% of peak power, with 75 seconds of active recovery at 30 W between bouts. Venous blood was collected before, immediately after, and 30 minutes after the HIIT session. In response to Staphylococcus aureus superantigen B (SEB) stimulation, lymphocyte proliferation decreased and the IL-2 concentration increased after the HIIT session. However, the HIIT session had no effect on lymphocyte proliferation or IL-2 response to phytohemagglutinin stimulation. The HIIT session also induced lymphocyte redox imbalance, characterized by an increase in the concentration of thiobarbituric acid reactive substances and a decrease in the activity of the antioxidant enzyme catalase. Lymphocyte viability was not affected by the HIIT session. The frequencies of CD25+ and CD69+ T helper and B lymphocytes in response to superantigen stimulation were lower after exercise, suggesting that superantigen-induced lymphocyte activation was reduced by HIIT. However, HIIT also led to a reduction in the frequency of CD4+ and CD19+ cells, so the frequencies of CD25+ and CD69+ cells within the CD4 and CD19 cell populations were not affected by HIIT. These data indicate that the reduced lymphocyte proliferation observed after HIIT is not due to reduced early lymphocyte activation by superantigen. Our findings show that an acute HIIT session promotes lymphocyte redox imbalance and reduces lymphocyte proliferation in response to superantigenic, but not to mitogenic stimulation. This observation cannot be explained by alteration of the early lymphocyte activation response to superantigen. The manner in which lymphocyte function modulation by an acute HIIT session can affect individual immunity and susceptibility to infection is important and requires further investigation.
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Tossige-Gomes, Rosalina; Costa, Karine Beatriz; Ottone, Vinícius de Oliveira; Magalhães, Flávio de Castro; Amorim, Fabiano Trigueiro; Rocha-Vieira, Etel
2016-01-01
This study investigated whether an acute session of high-intensity interval training (HIIT) is sufficient to alter lymphocyte function and redox status. Sixteen young healthy men underwent a HIIT session on a cycloergometer, consisting of eight bouts of 1 min at 90–100% of peak power, with 75 seconds of active recovery at 30 W between bouts. Venous blood was collected before, immediately after, and 30 minutes after the HIIT session. In response to Staphylococcus aureus superantigen B (SEB) stimulation, lymphocyte proliferation decreased and the IL-2 concentration increased after the HIIT session. However, the HIIT session had no effect on lymphocyte proliferation or IL-2 response to phytohemagglutinin stimulation. The HIIT session also induced lymphocyte redox imbalance, characterized by an increase in the concentration of thiobarbituric acid reactive substances and a decrease in the activity of the antioxidant enzyme catalase. Lymphocyte viability was not affected by the HIIT session. The frequencies of CD25+ and CD69+ T helper and B lymphocytes in response to superantigen stimulation were lower after exercise, suggesting that superantigen-induced lymphocyte activation was reduced by HIIT. However, HIIT also led to a reduction in the frequency of CD4+ and CD19+ cells, so the frequencies of CD25+ and CD69+ cells within the CD4 and CD19 cell populations were not affected by HIIT. These data indicate that the reduced lymphocyte proliferation observed after HIIT is not due to reduced early lymphocyte activation by superantigen. Our findings show that an acute HIIT session promotes lymphocyte redox imbalance and reduces lymphocyte proliferation in response to superantigenic, but not to mitogenic stimulation. This observation cannot be explained by alteration of the early lymphocyte activation response to superantigen. The manner in which lymphocyte function modulation by an acute HIIT session can affect individual immunity and susceptibility to infection is important and requires further investigation. PMID:27096389
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Tiemessen, Machteld M; Kunzmann, Steffen; Schmidt-Weber, Carsten B; Garssen, Johan; Bruijnzeel-Koomen, Carla A F M; Knol, Edward F; van Hoffen, Els
2003-12-01
Transforming growth factor (TGF)-beta has been demonstrated to play a key role in the regulation of the immune response, mainly by its suppressive function towards cells of the immune system. In humans, the effect of TGF-beta on antigen-specific established memory T cells has not been investigated yet. In this study antigen-specific CD4(+) T cell clones (TCC) were used to determine the effect of TGF-beta on antigen-specific proliferation, the activation status of the T cells and their cytokine production. This study demonstrates that TGF-beta is an adequate suppressor of antigen-specific T cell proliferation, by reducing the cell-cycle rate rather than induction of apoptosis. Addition of TGF-beta resulted in increased CD69 expression and decreased CD25 expression on T cells, indicating that TGF-beta is able to modulate the activation status of in vivo differentiated T cells. On the contrary, the antigen-specific cytokine production was not affected by TGF-beta. Although TGF-beta was suppressive towards the majority of the T cells, insensitivity of a few TCC towards TGF-beta was also observed. This could not be correlated to differential expression of TGF-beta signaling molecules such as Smad3, Smad7, SARA (Smad anchor for receptor activation) and Hgs (hepatocyte growth factor-regulated tyrosine kinase substrate). In summary, TGF-beta has a pronounced inhibitory effect on antigen-specific T cell proliferation without modulating their cytokine production.
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Li, Minglin; Ma, Huili; Yang, Lixin; Li, Peng
2016-01-01
Mangiferin, a flavonoid extracted from the mango tree, possesses anti-inflammatory, antibacterial, anti-herpes simplex and antitumor activity, and is able to affect immune function. The present study investigated the anticancer effects of mangiferin treatment on PC3 human prostate cancer cells, and the potential underlying mechanisms. In the present study, an MTT assay was used to analyze the proliferation of PC3 cells. Subsequently, flow cytometry and colorimetric assay kits were utilized to measure the PC3 cell apoptotic rate. The expression levels of B-cell lymphoma-2 (Bcl-2) and microRNA-182 (miR-182) were detected using western blot analysis and quantitative reverse transcription-polymerase chain reaction, respectively. Finally, miR-182 and anti-miR-182 were transfected into PC3 cells, which were used to investigate the effects of mangiferin. Mangiferin treatment reduced the proliferation of PC3 human prostate cancer cells in a concentration- and time-dependent manner. In addition, mangiferin was able to promote apoptosis and induce the caspase-3 activity of PC3 human prostate cancer cells. Mangiferin treatment was also able to significantly reduce Bcl-2 expression levels and enhance miR-182 expression in PC3 cells. Finally, it was observed that mangiferin inhibited proliferation and induced apoptosis in PC3 human prostate cancer cells, and this effect was correlated with downregulation of Bcl-2 and upregulation of miR-182.
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In vitro effects and mechanisms of lycopene in MCF-7 human breast cancer cells.
Peng, S J; Li, J; Zhou, Y; Tuo, M; Qin, X X; Yu, Q; Cheng, H; Li, Y M
2017-04-13
Breast cancer adversely affects the health status of women; therefore, the prevention and treatment of breast cancer is of critical importance. Lycopene is known to possess several biological effects such as removal of free radicals, alleviation of biological oxidative injury, and inhibition of tumor growth. In this study, we aimed to illustrate the effect of lycopene on tumor cell proliferation and modulation of cancer progression as well as its possible underlying mechanisms in human breast carcinoma cell line MCF-7 in vitro. MCF-7 cells were treated with different lycopene concentrations for 24, 48, and 72 h. Light field microscopy was used to observe cell morphology. MTT assay was used to determine the effect of lycopene on MCF-7 proliferation. Flow cytometry was employed to evaluate cell apoptosis. Real-time quantitative polymerase chain reaction was performed to detect the expression of p53 and Bax. Under microscopic examination, the untreated MCF-7 cells appeared to have a diamond or polygonal shape. Lycopene treatment resulted in cell shrinkage and breakage, whose severity increased in a dose and duration dependent manner. In addition, reduced cell proliferation and increased apoptosis (P < 0.05) were observed using MTT assay and flow cytometry, respectively. Moreover, lycopene could also upregulate the expression of p53 and Bax mRNAs in MCF-7 cells. In conclusion, lycopene inhibits proliferation and facilitates apoptosis of MCF-7 cells in vitro, possibly by regulating the expression of p53 and Bax.
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Vaca-González, J J; Gutiérrez, M L; Guevara, J M; Garzón-Alvarado, D A
2017-01-01
Articular cartilage is characterized by low cell density of only one cell type, chondrocytes, and has limited self-healing properties. When articular cartilage is affected by traumatic injuries, a therapeutic strategy such as autologous chondrocyte implantation is usually proposed for its treatment. This approach requires in vitro chondrocyte expansion to yield high cell number for cell transplantation. To improve the efficiency of this procedure, it is necessary to assess cell dynamics such as migration, proliferation and cell death during culture. Computational models such as cellular automata can be used to simulate cell dynamics in order to enhance the result of cell culture procedures. This methodology has been implemented for several cell types; however, an experimental validation is required for each one. For this reason, in this research a cellular automata model, based on random-walk theory, was devised in order to predict articular chondrocyte behavior in monolayer culture during cell expansion. Results demonstrated that the cellular automata model corresponded to cell dynamics and computed-accurate quantitative results. Moreover, it was possible to observe that cell dynamics depend on weighted probabilities derived from experimental data and cell behavior varies according to the cell culture period. Thus, depending on whether cells were just seeded or proliferated exponentially, culture time probabilities differed in percentages in the CA model. Furthermore, in the experimental assessment a decreased chondrocyte proliferation was observed along with increased passage number. This approach is expected to having other uses as in enhancing articular cartilage therapies based on tissue engineering and regenerative medicine.
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Ho, Y C; Wang, S
2010-11-24
Clinical and preclinical studies suggest that the hippocampus has a role in the pathophysiology of major depression. In the learned helplessness (LH) animal model of depression after inescapable shocks (ISs) animals that display LH behavior have reduced cell proliferation in the hippocampus; this effect can be reversed by antidepressant treatment. Using this model, we compared rats that displayed LH behavior and rats that did not show LH behavior (NoLH) after ISs to determine whether reduced hippocampal cell proliferation is associated with the manifestation of LH behavior or is a general response to stress. Specifically, we examined cell proliferation, neurogenesis, and synaptic function in dorsal and ventral hippocampus of LH and NoLH animals and control rats that were not shocked. The LH rats had showed reduced cell proliferation, neurogenesis, and synaptic transmission in the dorsal hippocampus, whereas no changes were seen in the ventral hippocampus. These changes were not observed in the NoLH animals. In a group of NoLH rats that received the same amount of electrical shock as the LH rats to control for the unequal shocks received in these two groups, we observed changes in Ki-67(+) cells associated with acute stress. We conclude that reduced hippocampal cell proliferation and neurogenesis are associated with the manifestation of LH behavior and that the dorsal hippocampus is the most affected area. Copyright © 2010 IBRO. Published by Elsevier Ltd. All rights reserved.
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Yuan, Ye; Yan, Gege; Gong, Rui; Zhang, Lai; Liu, Tianyi; Feng, Chao; Du, Weijie; Wang, Ying; Yang, Fan; Li, Yuan; Guo, Shuyuan; Ding, Fengzhi; Ma, Wenya; Idiiatullina, Elina; Pavlov, Valentin; Han, Zhenbo; Cai, Benzhi; Yang, Lei
2017-01-01
Blue light emitting diodes (LEDs) have been proven to affect the growth of several types of cells. The effects of blue LEDs have not been tested on bone marrow-derived mesenchymal stem cells (BMSCs), which are important for cell-based therapy in various medical fields. Therefore, the aim of this study was to determine the effects of blue LED on the proliferation, apoptosis and osteogenic differentiation of BMSCs. BMSCs were irradiated with a blue LED light at 470 nm for 1 min, 5 min, 10 min, 30 min and 60 min or not irradiated. Cell proliferation was measured by performing cell counting and EdU staining assays. Cell apoptosis was detected by TUNEL staining. Osteogenic differentiation was evaluated by ALP and ARS staining. DCFH-DA staining and γ-H2A.X immunostaining were used to measure intracellular levels of ROS production and DNA damage. Both cell counting and EdU staining assays showed that cell proliferation of BMSCs was significantly reduced upon blue LED irradiation. Furthermore, treatment of BMSCs with LED irradiation was followed by a remarkable increase in apoptosis, indicating that blue LED light induced toxic effects on BMSCs. Likewise, BMSC osteogenic differentiation was inhibited after exposure to blue LED irradiation. Further, blue LED irradiation was followed by the accumulation of ROS production and DNA damage. Taken together, our study demonstrated that blue LED light inhibited cell proliferation, inhibited osteogenic differentiation, and induced apoptosis in BMSCs, which are associated with increased ROS production and DNA damage. These findings may provide important insights for the application of LEDs in future BMSC-based therapies. © 2017 The Author(s). Published by S. Karger AG, Basel.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Lu Jiawei; Division of Molecular Medicine, Harbor-UCLA Medical Center, David Geffen School of Medicine, University of California Los Angeles, Torrance, CA 90502; Lu Zhenyu
The corneal endothelial cells form a boundary layer between anterior chamber and cornea. This single cell layer is important to maintain cornea transparency by eliciting net fluid transport into the anterior chamber. Injuries of the corneal endothelial layer in humans lead to corneal swelling and translucence. This hindrance is thought to be due to limited proliferative capacity of the endothelial layer. Fibroblast growth factor 2 (FGF-2) and transforming growth factor-beta 2 (TGF-{beta}2) are both found in aqueous humor, and these two cytokines promote and inhibit cell growth, respectively. The intracellular signaling mechanisms by which TGF-{beta}2 suppresses the mitogenic response tomore » FGF-2, however, remain unclear. We have addressed this question by investigating potential crosstalk between FGF-2-induced and TGF-{beta}2-regulated intracellular signaling events in cultured bovine corneal endothelial (BCE) cells. We found that TGF-{beta}2 and FGF-2 oppositely affect BCE cell proliferation and TGF-{beta}2 can override the stimulating effects of FGF-2 by increasing COX-2 expression in these cells. Consistent with these findings, overexpression of COX-2 significantly reduced FGF-2-induced cell proliferation whereas a COX-2 specific inhibitor NS398 reversed the effect of TGF-{beta}2 on FGF-2-induced cell proliferation. The COX-2 product prostaglandin E2 (PGE-2) blocks FGF-2-induced cell proliferation. Whereas FGF-2 stimulates cell proliferation by activating the AKT pathway, TGF-{beta}2 and PGE-2 both inhibit this pathway. In accordance with the effect of PGE-2, cAMP also inhibits FGF-2-induced AKT activation. These findings suggest that the mitogenic response to FGF-2 in vivo in the corneal endothelial layer may be inhibited by TGF-{beta}2-induced suppression of the PI3-kinase/AKT signaling pathway.« less
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Down-regulation of Wnt10a affects odontogenesis and proliferation in mesenchymal cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Liu, Yang, E-mail: Ly10160624@163.com; Han, Dong, E-mail: Donghan@bjmu.edu.cn; Wang, Lei, E-mail: wanglei_dentist@163.com
Highlights: •Down-regulation of Wnt10a in dental mesenchymal cells impairs odontogenesis of reassociated tooth germs. •Dspp is down- and up-regulated after Wnt10a-knockdown and overexpression in dental mesenchymal cells. •Down-regulation of Wnt10a inhibits proliferation of dental mesenchymal cells. -- Abstract: The WNT10a mutation has been found in patients with abnormal odontogenesis. In mice, Wnt10a expression is found in the tooth germ, but its role has not yet been elucidated. We aimed to investigate the role of Wnt10a in odontogenesis. Mesenchymal cells of the first mandibular molar germ at the bell stage were isolated, transfected with Wnt10a SiRNA or plasmid, and reassociated withmore » epithelial part of the molar germ. Scrambled SiRNA or empty vector was used in the control group. The reassociated tooth germs were transplanted into mice subrenal capsules. After gene modification, dental mesenchymal cells cultured in vitro were checked for cell proliferation and the expression of Dspp was examined. All 12 reassociated tooth germs in the control group resumed odontogenesis, while only 5 of 12 in the Wnt10a knockdown group developed into teeth. After Wnt10a knockdown, the mesenchymal cells cultured in vitro presented repressed proliferation. Wnt10a knockdown and overexpression led to both down- and up-regulation of Dspp. We conclude that the down-regulation of Wnt10a impairs odontogensis and cell proliferation, and that Wnt10a regulates Dspp expression in mesenchymal cells. These findings help to elucidate the mechanism of abnormal tooth development in patients with the WNT10A mutation.« less
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Control of Neural Daughter Cell Proliferation by Multi-level Notch/Su(H)/E(spl)-HLH Signaling
Bivik, Caroline; MacDonald, Ryan B.; Gunnar, Erika; Mazouni, Khalil; Schweisguth, Francois; Thor, Stefan
2016-01-01
The Notch pathway controls proliferation during development and in adulthood, and is frequently affected in many disorders. However, the genetic sensitivity and multi-layered transcriptional properties of the Notch pathway has made its molecular decoding challenging. Here, we address the complexity of Notch signaling with respect to proliferation, using the developing Drosophila CNS as model. We find that a Notch/Su(H)/E(spl)-HLH cascade specifically controls daughter, but not progenitor proliferation. Additionally, we find that different E(spl)-HLH genes are required in different neuroblast lineages. The Notch/Su(H)/E(spl)-HLH cascade alters daughter proliferation by regulating four key cell cycle factors: Cyclin E, String/Cdc25, E2f and Dacapo (mammalian p21CIP1/p27KIP1/p57Kip2). ChIP and DamID analysis of Su(H) and E(spl)-HLH indicates direct transcriptional regulation of the cell cycle genes, and of the Notch pathway itself. These results point to a multi-level signaling model and may help shed light on the dichotomous proliferative role of Notch signaling in many other systems. PMID:27070787
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Song, E Y; Han, S; Yang, B; Morris, G P; Bui, J D
2015-04-01
The proliferation rates of human T cells in vitro are affected by some factors such as initial T-cell number, dose of stimulating cells, and duration of culture. The transcription factor forkhead box P3 (FoxP3) has been used to identify regulatory T cells in humans and is thought to correlate with tolerance to allogeneic organ transplant. Thus, it is important to optimize conditions to expand FoxP3 cell proliferation to improve engraftment of allogeneic organ transplants. We studied proliferative responses and FoxP3 expression in divided T cells with the use of flow cytometric analysis of Ki-67 in culture of different concentrations of responding cells (6 × 10(6), 4 × 10(6), 2 × 10(6), 1 × 10(6), and 0.5 × 10(6)cells/mL), different types of stimulating cells (lymphocytes and low density cells), and different numbers of HLA mismatches. The proportion of CD3(+) cells, CD4(+)CD25(+) cells, and CD4(+)CD25(+)FoxP3(+) cells among mononuclear cells were highest at initial cell concentration of 2 × 10(6) responder cells/mL with lymphocytes as stimulators at day-5 mixed lymphocyte reaction (MLR). They were highest at a concentration of 4 × 10(6) responder cells/mL with low density cells as stimulators. The recovery (%), proportion of CD3(+) cells, CD4(+)CD25(+) cells, and CD4(+)CD25(+)FoxP3(+) cells with 2 HLA-DR incompatibility were significantly higher than those of 1 HLA-DR incompatibility at day-5 MLR. Initial cell concentration and HLA-DR incompatibility can affect the generation of FoxP3+ T cells in human MLR. These factors could be considered for efficient generation of Tregs for clinical trials in the future. Copyright © 2015 Elsevier Inc. All rights reserved.
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Toxicity of extracts from disposable chopsticks, toothpicks, and paper cups on L-929 cells.
Li, Juntao; Chen, Sifan; Li, Wenxue; Yang, Guangyu; Zhu, Wei
2015-04-01
To evaluate the toxicity of extracts from disposable chopsticks, toothpicks, and paper cups on L-929 cells. We followed national standards to prepare the extracts from disposable chopsticks, toothpicks, and paper cups used for the cell culture media, and the morphology of L-929 cells was observed with an optical microscope. The loss rate for adherent cells was evaluated with the trypan blue exclusion method, and cell proliferation was determined using the WST-1 assay. Compared with the control group, the cells cultured in media containing the extracts showed signs of apoptosis and necrosis after culturing for 4 or 7 days, and the loss rate for adherent cells was significantly increased (P < 0.05). An obvious decrease in cell viability was also observed (P < 0.05). The extracts from disposable chopsticks, toothpicks, and paper cups can affect the growth and proliferation of L-929 cells and are potentially toxic to humans.
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Kinetics of liver macrophages (Kupffer cells) in SIV-infected macaques
Ahsan, Muhammad H.; Gill, Amy F.; Alvarez, Xavier; Lackner, Andrew A.; Veazey, Ronald S.
2013-01-01
Since the liver drains antigens from the intestinal tract, and since the intestinal tract is a major site of viral replication, we examined the dynamics of liver macrophages (Kupffer cells) throughout SIV infection. Absolute numbers of Kupffer cells increased in the livers in acute infection, and in animals with AIDS. Significantly higher percentages of proliferating (BrdU+) Kupffer cells were detected in acute infection and in AIDS with similar trends in blood monocytes. Significantly higher percentages of apoptotic (AC3+) Kupffer cells were also found in acute and AIDS stages. However, productively infected cells were not detected in liver of 41/42 animals examined, despite abundant infected cells in gut and lymph nodes of all animals. Increased rates of Kupffer cell proliferation resulting in an increase in Kupffer cells without productive infection indicate SIV infection affects Kupffer cells, but the liver does not appear to be a major site of productive viral replication. PMID:24074569
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MiR-29b affects the secretion of PROG and promotes the proliferation of bovine corpus luteum cells
Zhang, Li-Qun; Sun, Xu-Lei; Luo, Dan; Fu, Yao; Gao, Yan; Zhang, Jia-Bao
2018-01-01
The regulatory role of miRNAs has been explored in ovarian cells, and their effects on gonadal development, apoptosis, ovulation, steroid production and corpus luteum (CL) development have been revealed. In this study, we analyzed the expression of miR-29b at different stages of bovine CL development and predicted the target genes of miR-29b. We confirmed that miR-29b reduces the expression of the oxytocin receptor (OXTR), affects progesterone (PROG) secretion and regulates the function of the CL. RT-PCR showed that the expression of miR-29b was significantly higher in functional CL phases than in the regressed CL phase. Immunohistochemistry showed that OXTR was expressed in both large and small CL cells and was mainly located in the cell membrane and cytoplasm of these cells. We analyzed the expression levels of OXTR and found that transfection with a miR-29b mimic decreased OXTR expression, but transfection with the inhibitor had a limited effect on the expression of the OXTR protein. At the same time, the secretion of PROG was significantly increased in the miR-29b mimic-transfected group. We also analyzed the effect of miR-29b on the apoptosis of CL cells. Finally, we found that miR-29b could promote the proliferation of bovine CL cells. In conclusion, we found that miR-29b reduces the expression of OXTR and can promote PROG secretion and the proliferation of CL cells via OXTR. PMID:29617446
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Eto, Masumi, E-mail: masumi.eto@jefferson.edu; Kirkbride, Jason A.; Chugh, Rishika
2013-04-26
Highlights: •Non-canonical roles of the myosin phosphatase inhibitor (CPI-17) were studied. •CPI-17 is localized in the nucleus of hyperplastic cancer and smooth muscle cells. •CPI-17 Ser12 phosphorylation may regulate the nuclear import. •CPI-17 regulates histone H3 phosphorylation and cell proliferation. •The nuclear CPI-17-PP1 axis plays a proliferative role in cells. -- Abstract: CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17 kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentratedmore » in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear import. CPI-17 phosphorylated at Ser12 was not localized at nuclei, suggesting a suppressive role of Ser12 phosphorylation in the nuclear import. Activated CPI-17 bound to all three isoforms of PP1 catalytic subunit in Panc1 nuclear extracts. CPI-17 knockdown in Panc1 resulted in dephosphorylation of histone H3 at Thr3, Ser10 and Thr11, whereas it had no effects on the phosphorylation of myosin light chain and merlin, the known targets of MLCP. In parallel, CPI-17 knockdown suppressed Panc1 proliferation. We propose that CPI-17 accumulated in the nucleus through the N-terminal tail targets multiple PP1 signaling pathways regulating cell proliferation.« less
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Role of Piezo Channels in Ultrasound-stimulated Dental Stem Cells.
Gao, Qianhua; Cooper, Paul R; Walmsley, A Damien; Scheven, Ben A
2017-07-01
Piezo1 and Piezo2 are mechanosensitive membrane ion channels. We hypothesized that Piezo proteins may play a role in transducing ultrasound-associated mechanical signals and activate downstream mitogen-activated protein kinase (MAPK) signaling processes in dental cells. In this study, the expression and role of Piezo channels were investigated in dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs) after treatment with low-intensity pulsed ultrasound (LIPUS). Cell proliferation was evaluated by bromodeoxyuridine incorporation. Western blots were used to analyze the proliferating cell nuclear antigen as well as the transcription factors c-fos and c-jun. Enzyme-linked immunosorbent assay and Western blotting were used to determine the activation of MAPK after LIPUS treatment. Ruthenium red (RR), a Piezo ion channel blocker, was applied to determine the functional role of Piezo proteins in LIPUS-stimulated cell proliferation and MAPK signaling. Western blotting showed the presence of Piezo1 and Piezo2 in both dental cell types. LIPUS treatment significantly increased the level of the Piezo proteins in DPSCs after 24 hours; however, no significant effects were observed in PDLSCs. Treatment with RR significantly inhibited LIPUS-stimulated DPSC proliferation but not PDLSC proliferation. Extracellular signal-related kinase (ERK) 1/2 MAPK was consistently activated in DPSCs over a 24-hour time period after LIPUS exposure, whereas phosphorylated c-Jun N-terminal kinase and p38 mitogen-activated protein kinase MAPK were mainly increased in PDLSCs. RR affected MAPK signaling in both dental cell types with its most prominent effects on ERK1/2/MAPK phosphorylation levels; the significant inhibition of LIPUS-induced stimulation of ERK1/2 activation in DPSCs by RR suggests that stimulation of DPSC proliferation by LIPUS involves Piezo-mediated regulation of ERK1/2 MAPK signaling. This study for the first time supports the role of Piezo ion channels in transducing the LIPUS response in dental stem cells. Copyright © 2017 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.
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Gil-Sanz, Cristina; Landeira, Bruna; Ramos, Cynthia; Costa, Marcos R; Müller, Ulrich
2014-08-06
Radial glial cells (RGCs) in the ventricular neuroepithelium of the dorsal telencephalon are the progenitor cells for neocortical projection neurons and astrocytes. Here we show that the adherens junction proteins afadin and CDH2 are critical for the control of cell proliferation in the dorsal telencephalon and for the formation of its normal laminar structure. Inactivation of afadin or CDH2 in the dorsal telencephalon leads to a phenotype resembling subcortical band heterotopia, also known as "double cortex," a brain malformation in which heterotopic gray matter is interposed between zones of white matter. Adherens junctions between RGCs are disrupted in the mutants, progenitor cells are widely dispersed throughout the developing neocortex, and their proliferation is dramatically increased. Major subtypes of neocortical projection neurons are generated, but their integration into cell layers is disrupted. Our findings suggest that defects in adherens junctions components in mice massively affects progenitor cell proliferation and leads to a double cortex-like phenotype. Copyright © 2014 the authors 0270-6474/14/3410475-13$15.00/0.
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lncRNA-HIT promotes cell proliferation of non-small cell lung cancer by association with E2F1.
Yu, L; Fang, F; Lu, S; Li, X; Yang, Y; Wang, Z
2017-05-01
Lung cancer is the leading cause of cancer-related death around the world. Long noncoding RNA (lncRNA) has pivotal roles in cancer occurrence and development. However, only a few lncRNAs have been functionally characterized. In the present study, we investigated the effects of lncRNA-HIT (HOXA transcript induced by TGFβ) expression on non-small cell lung cancer (NSCLC) cell phenotype with the gain-of-function and loss-of-function assays. We found that ectopic expression or knockdown of lncRNA-HIT markedly increased or decreased NSCLC cell proliferation, respectively. Moreover, we also showed that lncRNA-HIT interacted with E2F1 to regulate its target genes, such as Survivin, FOXM1, SKP2, NELL2 and DOK1. Collectively, our findings indicated that lncRNA-HIT affected the proliferation of NSCLC cells at least in part via regulating the occupancy of E2F1 in the promoter regions of its target genes. The lncRNA-HIT-E2F1 complex may be a potential target for NSCLC treatment.
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NASA Technical Reports Server (NTRS)
Ramesh, Govindarajan; Wu, Honglu
2012-01-01
Radiation affects several cellular and molecular processes including double strand breakage, modifications of sugar moieties and bases. In outer space, protons are the primary radiation source which poses a range of potential health risks to astronauts. On the other hand, the use of proton radiation for tumor radiation therapy is increasing as it largely spares healthy tissues while killing tumor tissues. Although radiation related research has been conducted extensively, the molecular toxicology and cellular mechanisms affected by proton radiation remain poorly understood. Therefore, in the present study, we irradiated rat epithelial cells (LE) with different doses of protons and investigated their effects on cell proliferation and cell death. Our data showed an inhibition of cell proliferation in proton irradiated cells with a significant dose dependent activation and repression of reactive oxygen species (ROS) and antioxidants, glutathione and superoxide dismutase respectively as compared to control cells. In addition, apoptotic related genes such as caspase-3 and -8 activities were induced in a dose dependent manner with corresponding increased levels of DNA fragmentation in proton irradiated cells than control cells. Together, our results show that proton radiation alters oxidant and antioxidant levels in the cells to activate apoptotic pathway for cell death.
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Fernandez-Rebollo, Eduardo; Mentrup, Birgit; Ebert, Regina; Franzen, Julia; Abagnale, Giulio; Sieben, Torsten; Ostrowska, Alina; Hoffmann, Per; Roux, Pierre-François; Rath, Björn; Goodhardt, Michele; Lemaitre, Jean-Marc; Bischof, Oliver; Jakob, Franz; Wagner, Wolfgang
2017-07-11
Culture medium of mesenchymal stromal cells (MSCs) is usually supplemented with either human platelet lysate (HPL) or fetal calf serum (FCS). Many studies have demonstrated that proliferation and cellular morphology are affected by these supplements - it is therefore important to determine if they favor outgrowth of different subpopulations and thereby impact on the heterogeneous composition of MSCs. We have isolated and expanded human bone marrow-derived MSCs in parallel with HPL or FCS and demonstrated that HPL significantly increases proliferation and leads to dramatic differences in cellular morphology. Remarkably, global DNA-methylation profiles did not reveal any significant differences. Even at the transcriptomic level, there were only moderate changes in pairwise comparison. Furthermore, the effects on proliferation, cytoskeletal organization, and focal adhesions were reversible by interchanging to opposite culture conditions. These results indicate that cultivation of MSCs with HPL or FCS has no systematic bias for specific cell types.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Ohashi, Kazuya, E-mail: asuno10k@yahoo.co.jp; Nagata, Yosuke, E-mail: cynagata@mail.ecc.u-tokyo.ac.jp; Wada, Eiji, E-mail: gacchu1@yahoo.co.jp
2015-05-01
Skeletal muscle stem cells named muscle satellite cells are normally quiescent but are activated in response to various stimuli, such as injury and overload. Activated satellite cells enter the cell cycle and proliferate to produce a large number of myogenic progenitor cells, and these cells then differentiate and fuse to form myofibers. Zinc is one of the essential elements in the human body, and has multiple roles, including cell growth and DNA synthesis. However, the role of zinc in myogenic cells is not well understood, and is the focus of this study. We first examined the effects of zinc onmore » differentiation of murine C2C12 myoblasts and found that zinc promoted proliferation, with an increased number of cells incorporating EdU, but inhibited differentiation with reduced myogenin expression and myotube formation. Furthermore, we used the C2C12 reserve cell model of myogenic quiescence to investigate the role of zinc on activation of myogenic cells. The number of reserve cells incorporating BrdU was increased by zinc in a dose dependent manner, with the number dramatically further increased using a combination of zinc and insulin. Akt and extracellular signal-regulated kinase (ERK) are downstream of insulin signaling, and both were phosphorylated after zinc treatment. The zinc/insulin combination-induced activation involved the phosphoinositide 3-kinase (PI3K)/Akt and ERK cascade. We conclude that zinc promotes activation and proliferation of myogenic cells, and this activation requires phosphorylation of PI3K/Akt and ERK as part of the signaling cascade. - Highlights: • Zinc has roles for promoting proliferation and inhibition differentiation of C2C12. • Zinc promotes activation of reserve cells. • Insulin and zinc synergize activation of reserve cells. • PI3K/Akt and ERK cascade affect zinc/insulin-mediated activation of reserve cells.« less
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Lauder, J M
1977-04-22
The effects of early hypo- and hyperthyroidism on the rates of cell acquisition and proliferation have been studied in the external granular layer (EGL) of the developing rat cerebellar cortex at 10 days of age using quantitative autoradiographic methods. Both altered thyroid states reduce the rate of cell acquisition in the EGL, but appear to do so for different reasons. Hyperthyroidism shortens the average length of the cell cycle by decreasing the duration of the pre-DNA synthetic phase (G1), indicating that excess thyroxine may exert a direct effect on the EGL. This action involves the early onset of neuronal differentiation (cessation of proliferation)46 which presumably leads to the observed decrease in the rate of cell acquisition (increased doubling time). Such differentiating cells do not, however, leave the proliferative zone or the EGL prematurely, resulting in a reduced labeling index, mitotic index, and growth fraction as non-dividing cells dilute the proliferating cell population. Hypothyroidism, on the other hand, leads to no significant change in the length of the cell cycle or in the mitotic index, but causes a decreased labeling index and growth fraction, as well as a reduced rate of cell acquisition (increased doubling time). No significant change in the amount of cell death in the EGL could be found to explain this apparent discrepancy between the rate of cell proliferation (cell cycle length) and cell acqusiition. The answer to this puzzle appears to lie in the mitotic index, which is not affected to the same extent as the labeling index, although it is also slightly reduced. If cells were to remain longer in mitosis, this could result in a decreased labeling index and growth fraction but nearly normal mitotic index and cell cycle length (as measured using the % labeled mitoses method), since those cells dropping out of the cycling population would be counted as mitoses...
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Ravindranath, T; Al-Ghoul, W; Namak, S; Fazal, N; Durazo-Arvizu, R; Choudhry, M; Sayeed, M M
2001-12-01
To evaluate the effect of burn injury with and without an Escherichia coliseptic complication on T-cell proliferation, interleukin-2 production, and Ca(2+) signaling responses in intestinal Peyer's patch and splenic T cells. Prospective, randomized, sham-controlled animal study. University medical center research laboratory. Adult male Sprague-Dawley rats. Rats were subjected to a 30% total body surface area, full skin thickness burn. Infection in rats was induced via intraperitoneal inoculation of E. coli, 10(9) colony forming units/kg, with or without a prior burn. Rat Peyer's patch and splenic T lymphocytes were isolated by using a nylon wool cell purification protocol. T-cell proliferation, interleukin-2 production, and Ca(2+) signaling responses were measured after stimulation of cells with the mitogen, concanavalin A. T-cell proliferation was determined by measuring incorporation of (3)H-thymidine into T-cell cultures. Interleukin-2 production by T-cell cultures was measured by using enzyme-linked immunosorbent assay. Intracellular T-cell Ca2(+ )concentration, [Ca(2+)](i), was measured by the use of Ca(2+)-specific fluorescent label, fura-2, and its fluorometric quantification. [Ca(2+)](i) was also evaluated by the use of digital video imaging of fura-2 loaded individual T cells. T-cell proliferation and interleukin-2 production were suppressed substantially in both Peyer's patch and splenic T cells 3 days after either the initial burn alone or burn followed by the E. coli inoculation at 24 hrs after the initial burn. There seemed to be no demonstrable additive effects of E. coli infection on the effects produced by burn injury alone. The T-cell proliferation and interleukin-2 production suppressions with burn or burn-plus-infection insults were correlated with attenuated Ca(2+) signaling. E. coli infection alone suppressed T-cell proliferation in Peyer's patch but not in splenic T cells at 2 days postbacterial inoculation; E. coli infection had no effect on Peyer's patch or splenic T cells at 1 day postinjury. On the other hand, burn injury alone caused a substantial T-cell proliferative suppression at 2 days postburn in both Peyer's patch and splenic cells and a significant suppression in T-cell proliferation on day 1 postburn in Peyer's patch but not in the spleen. An initial burn injury suppressed T-cell proliferation at a level that it would not be further affected by a subsequent infection even if the infection by itself has the potential of suppressing T-cell proliferation. An earlier onset of T-cell suppression in Peyer's patch cells than in the spleen with burn could be attributable to an initial hypoperfusion-related intestinal mucosal tissue injury. Overall, our study supports the concept that burn injury per se can significantly suppress T-cell mediated immunity and that the intestine is an early tissue site of such suppression.
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Sun, Mei-Na; Zhao, Xue-Jiao; Zhao, Han-Dong; Zhang, Wei-Guang; Li, Feng-Lan; Chen, Ming-Zi; Li, Hui; Li, Guangchao
2013-06-01
The aim of the present study was to investigate the effects of recombinant Escherichia coli (E. coli) Trx-jingzhaotoxin (JZTX)-III on cell growth in the mouse hepatocellular carcinoma (HCC) cell line Hepa1-6. The JZTX-III gene sequence was synthesized and cloned into the pET-32a(+) vector to construct the recombinant fusion protein Trx-JZTX-III, which was subsequently purified. Hepa1-6 cells were treated with 0 to 1,000-µg/ml concentrations of Trx-JZTX-III; this was demonstrated to affect cell viability, as determined by the 3-(4,5-dimethylthiazol‑2-yl)-2,5-diphenyltetra-zolium bromide (MTT) assay. The expression of the proliferating cell nuclear antigen (PCNA) protein was investigated using western blot analysis. A colony formation assay was used to determine Hepa1-6 cell proliferation, and the migration ability of cells was determined using a wound‑healing assay. Additionally, flow cytometry was employed to observe changes in the cell cycle. The MTT assay and quantification of PCNA expression indicated that recombinant E. coli Trx-JZTX-III significantly repressed the proliferation of Hepa1-6 cells. Colony formation and the migration of malignant cells was inhibited following treatment with recombinant E. coli Trx-JZTX-III. Flow cytometry showed that recombinant E. coli Trx-JZTX-III induced G0/G1 cell cycle arrest. In conclusion, recombinant E. coli Trx-JZTX-III functions as a tumor suppressor drug in mouse HCC and its underlying mechanism may involve the induction of G0/G1 cell cycle arrest.
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CD24 promotes the proliferation and inhibits the apoptosis of cervical cancer cells in vitro.
Pei, Zhen; Zhu, Guangchao; Huo, Xiaolei; Gao, Lu; Liao, Shan; He, Junyu; Long, Yuehua; Yi, Hong; Xiao, Songshu; Yi, Wei; Chen, Pan; Li, Xiaoling; Li, Guiyuan; Zhou, Yanhong
2016-03-01
The protein CD24 is a cell surface protein that appears to function as an adhesion molecule; its expression has been shown to correlate with prognosis in a variety of tumors. Herein, we investigated the possible role and mechanism of CD24 in cervical cancer. Our results showed that CD24 was overexpressed in cervical cancer tissues compared with that in the adjacent non‑cancerous tissues by qPCR, immunohistochemistry and western blotting technologies. To explore the possible mechanism of CD24 in cervical cancer, we elucidated the effect of CD24 on the proliferation and apoptosis of cervical cancer HeLa cells and found that a considerable increase in cell proliferation was observed in the HeLa cells with CD24 overexpession. The rate of cell apoptosis was decreased in the HeLa/CD24 cells compared with the HeLa or HeLa/vector cells. Cell apoptosis is closely related with a reduction in mitochondrial membrane potential (ΔΨm) and an increase in intracellular reactive oxygen species (ROS) and calcium ion (Ca2+) concentrations. Our results showed that overexpression of CD24 in the cervical cancer HeLa cells, led to an increase in ΔΨm and a decrease in intracellular ROS and Ca2+ concentrations. Furthermore, we found that CD24 was correlated with dysregulation of the MAPK signaling pathway in cervical cancer tissues in vitro. At the same time, we found that CD24 overexpression affected the expression of p38, JNK2 and c-Jun in vitro. In summary, our results suggest that CD24 is upregulated in cervical cancer tissues and plays its functions by affecting the MAPK signaling pathway in cervical cancer.
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Mutations in Alström protein impair terminal differentiation of cardiomyocytes.
Shenje, Lincoln T; Andersen, Peter; Halushka, Marc K; Lui, Cecillia; Fernandez, Laviel; Collin, Gayle B; Amat-Alarcon, Nuria; Meschino, Wendy; Cutz, Ernest; Chang, Kenneth; Yonescu, Raluca; Batista, Denise A S; Chen, Yan; Chelko, Stephen; Crosson, Jane E; Scheel, Janet; Vricella, Luca; Craig, Brian D; Marosy, Beth A; Mohr, David W; Hetrick, Kurt N; Romm, Jane M; Scott, Alan F; Valle, David; Naggert, Jürgen K; Kwon, Chulan; Doheny, Kimberly F; Judge, Daniel P
2014-03-04
Cardiomyocyte cell division and replication in mammals proceed through embryonic development and abruptly decline soon after birth. The process governing cardiomyocyte cell cycle arrest is poorly understood. Here we carry out whole-exome sequencing in an infant with evidence of persistent postnatal cardiomyocyte replication to determine the genetic risk factors. We identify compound heterozygous ALMS1 mutations in the proband, and confirm their presence in her affected sibling, one copy inherited from each heterozygous parent. Next, we recognize homozygous or compound heterozygous truncating mutations in ALMS1 in four other children with high levels of postnatal cardiomyocyte proliferation. Alms1 mRNA knockdown increases multiple markers of proliferation in cardiomyocytes, the percentage of cardiomyocytes in G2/M phases, and the number of cardiomyocytes by 10% in cultured cells. Homozygous Alms1-mutant mice have increased cardiomyocyte proliferation at 2 weeks postnatal compared with wild-type littermates. We conclude that deficiency of Alström protein impairs postnatal cardiomyocyte cell cycle arrest.
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Mutations in Alström Protein Impair Terminal Differentiation of Cardiomyocytes
Shenje, Lincoln T.; Andersen, Peter; Halushka, Marc K.; Lui, Cecillia; Fernandez, Laviel; Collin, Gayle B.; Amat-Alarcon, Nuria; Meschino, Wendy; Cutz, Ernest; Chang, Kenneth; Yonescu, Raluca; Batista, Denise A. S.; Chen, Yan; Chelko, Stephen; Crosson, Jane E.; Scheel, Janet; Vricella, Luca; Craig, Brian D.; Marosy, Beth A.; Mohr, David W.; Hetrick, Kurt N.; Romm, Jane M.; Scott, Alan F.; Valle, David; Naggert, Jürgen K.; Kwon, Chulan; Doheny, Kimberly F.; Judge, Daniel P.
2014-01-01
Cardiomyocyte cell division and replication in mammals proceed through embryonic development and abruptly decline soon after birth. The process governing cardiomyocyte cell cycle arrest is poorly understood. Here we carry out whole exome sequencing in an infant with evidence of persistent postnatal cardiomyocyte replication to determine the genetic risk factors. We identify compound heterozygous ALMS1 mutations in the proband, and confirm their presence in her affected sibling, one copy inherited from each heterozygous parent. Next, we recognise homozygous or compound heterozygous truncating mutations in ALMS1 in four other children with high levels of postnatal cardiomyocyte proliferation. Alms1 mRNA knockdown increases multiple markers of proliferation in cardiomyocytes, the percentage of cardiomyocytes in G2/M phases, and the number of cardiomyocytes by 10% in cultured cells. Homozygous Alms1-mutant mice have increased cardiomyocyte proliferation at two weeks postnatal compared to wild-type littermates. We conclude that deficiency of Alström protein impairs postnatal cardiomyocyte cell cycle arrest. PMID:24595103
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Zhang, Mingjun; Wang, Xiaolei; Li, Wanhu
Although a series of oncogenes and tumor suppressors were identified in the pathological development of gastric adenocarcinoma (GAC), the underlying molecule mechanism were still not fully understood. The current study explored the expression profile of miR-107 and miR-25 in GAC patients and their downstream regulative network. qRT-PCR analysis was performed to quantify the expression of these two miRNAs in serum samples from both patients and healthy controls. Dual luciferase assay was conducted to verify their putative bindings with LATS2. MTT assay, cell cycle assay and transwell assay were performed to explore how miR-107 and miR-25 regulate proliferation and invasion ofmore » gastric cancer cells. Findings of this study demonstrated that total miR-107 or miR-25 expression might be overexpressed in gastric cancer patients and they can simultaneously and synchronically regulate LATS2 expression, thereby affecting gastric cancer cell growth and invasion. Therefore, the miR-25/miR-107-LATS2 axis might play an important role in proliferation and invasion of the gastric cancer cells. - Highlights: • Total miR-107 and miR-25 expression is significantly increased in GAC patients. • Both miR-107 and miR-25 can promote proliferation and invasion of GAC cells. • Both miR-107 and miR-25 can target LATS2 and regulate its expression. • miR-107 and miR-25 regulate proliferation and invasion of GAC cells though LATS2.« less
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Colle, Dirleise; Farina, Marcelo; Ceccatelli, Sandra; Raciti, Marilena
2018-06-01
Pesticide exposure has been linked to the pathogenesis of neurodevelopmental and neurodegenerative disorders including autism spectrum disorders, attention deficit/hyperactivity, and Parkinson's disease (PD). Developmental exposure to pesticides, even at low concentrations not harmful for the adult brain, can lead to neuronal loss and functional deficits. It has been shown that prenatal or early postnatal exposure to the herbicide paraquat (PQ) and the fungicide maneb (MB), alone or in combination, causes permanent toxicity in the nigrostriatal dopamine system, supporting the idea that early exposure to these pesticides may contribute to the pathophysiology of PD. However, the mechanisms mediating PQ and MB developmental neurotoxicity are not yet understood. Therefore, we investigated the neurotoxic effect of low concentrations of PQ and MB in primary cultures of rat embryonic neural stem cells (NSCs), with particular focus on cell proliferation and oxidative stress. Exposure to PQ alone or in combination with MB (PQ + MB) led to a significant decrease in cell proliferation, while the cell death rate was not affected. Consistently, PQ + MB exposure altered the expression of major genes regulating the cell cycle, namely cyclin D1, cyclin D2, Rb1, and p19. Moreover, PQ and PQ + MB exposures increased the reactive oxygen species (ROS) production that could be neutralized upon N-acetylcysteine (NAC) treatment. Notably, in the presence of NAC, Rb1 expression was normalized and a normal cell proliferation pattern could be restored. These findings suggest that exposure to PQ + MB impairs NSCs proliferation by mechanisms involving alterations in the redox state.
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Conserva, Enrico; Menini, Maria; Ravera, Giambattista; Pera, Paolo
2013-08-01
The aim of this study was an in vitro comparison of osteoblast adhesion, proliferation and differentiation related to six dental implants with different surface characteristics, and to determine if the interaction between cells and implant is influenced by surface structure and chemical composition. Six types of implants were tested, presenting four different surface treatments: turned, sandblasted, acid-etched, anodized. The implant macro- and microstructure were analyzed using SEM, and the surface chemical composition was investigated using energy-dispersive X-ray analysis. SaOS-2 osteoblasts were used for the evaluation of cell adhesion and proliferation by SEM, and cell viability in contact with the various surfaces was determined using cytotoxicity MTT assays. Alkaline phosphatase (ALP) enzymatic activity in contact with the six surfaces was evaluated. Data relative to MTT assay and ALP activity were statistically analyzed using Kruskal-Wallis not parametric test and Nemenyi-Damico-Wolfe-Dunn post hoc test. All the implants tested supported cell adhesion, proliferation and differentiation, revealing neither organic contaminants nor cytotoxicity effects. The industrial treatments investigated changed the implant surface microscopic aspect and SaOS-2 cell morphology appeared to be influenced by the type of surface treatment at 6, 24, and 72 h of growth. SaOS-2 cells spread more rapidly on sandblasted surfaces. Turned surfaces showed the lowest cell proliferation at SEM observation. Sandblasted surfaces showed the greatest ALP activity values per cell, followed by turned surfaces (P < 0.05). On the base of this in vitro investigation, differently surfaced implants affected osteoblast morphology, adhesion, proliferation, and differentiation. Sandblasted surfaces promoted the most suitable osteoblast behavior. © 2012 John Wiley & Sons A/S.
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NASA Astrophysics Data System (ADS)
Žáková, Pavlína; Slepičková Kasálková, Nikola; Slepička, Petr; Kolská, Zdeňka; Karpíšková, Jana; Stibor, Ivan; Švorčík, Václav
2017-11-01
Various carbon nanostructures are widely researched as scaffolds for tissue engineering. We evaluated the surface properties and cell-substrate interactions of carbon nanoparticles functionalized with triethylenetetramine (CNPs) grafted polymer film. Two forms of polyethylene (HDPE, LDPE) were treated in an inert argon plasma discharge and, subsequently, grafted with CNPs. The surface properties were studied using multiple methods, including Raman spectroscopy, goniometry, atomic force microscopy, X-ray photoelectron spectroscopy and electrokinetic analysis. Cell-substrate interactions were determined in vitro by studying adhesion, proliferation and viability of vascular smooth muscle cells (VSMCs) from the aorta of a rat. Cell-substrate interactions on pristine and modified substrates were compared to standard tissue culture polystyrene. Our results show that CNPs affect surface morphology and wettability and therefore adhesion, proliferation and viability of cultured muscle cells.
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Keijzer, Richard; Liu, Jason; Deimling, Julie; Tibboel, Dick; Post, Martin
2000-01-01
Pulmonary hypoplasia associated with congenital diaphragmatic hernia (CDH) remains a major therapeutic problem. Moreover, the pathogenesis of pulmonary hypoplasia in case of CDH is controversial. In particular, little is known about early lung development in this anomaly. To investigate lung development separate from diaphragm development we used an in vitro modification of the 2,4-dichlorophenyl-p-nitrophenylether (Nitrofen) animal model for CDH. This enabled us to investigate the direct effects of Nitrofen on early lung development and branching morphogenesis in an organotypic explant system without the influence of impaired diaphragm development. Epithelial cell differentiation of the lung explants was assessed using surfactant protein-C and Clara cell secretory protein-10 mRNA expression as markers. Furthermore, cell proliferation and apoptosis were investigated. Our results indicate that Nitrofen negatively influences branching morphogenesis of the lung. Initial lung anlage formation is not affected. In addition, epithelial cell differentiation and cell proliferation are attenuated in lungs exposed to Nitrofen. These data indicate that Nitrofen interferes with early lung development before and separate from (aberrant) diaphragm development. Therefore, we postulate the dual-hit hypothesis, which explains pulmonary hypoplasia in CDH by two insults, one affecting both lungs before diaphragm development and one affecting the ipsilateral lung after defective diaphragm development. PMID:10751355
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Fetal programming in meat production.
Du, Min; Wang, Bo; Fu, Xing; Yang, Qiyuan; Zhu, Mei-Jun
2015-11-01
Nutrient fluctuations during the fetal stage affects fetal development, which has long-term impacts on the production efficiency and quality of meat. During the early development, a pool of mesenchymal progenitor cells proliferate and then diverge into either myogenic or adipogenic/fibrogenic lineages. Myogenic progenitor cells further develop into muscle fibers and satellite cells, while adipogenic/fibrogenic lineage cells develop into adipocytes, fibroblasts and resident fibro-adipogenic progenitor cells. Enhancing the proliferation and myogenic commitment of progenitor cells during fetal development enhances muscle growth and lean production in offspring. On the other hand, promoting the adipogenic differentiation of adipogenic/fibrogenic progenitor cells inside the muscle increases intramuscular adipocytes and reduces connective tissue, which improves meat marbling and tenderness. Available studies in mammalian livestock, including cattle, sheep and pigs, clearly show the link between maternal nutrition and the quantity and quality of meat production. Similarly, chicken muscle fibers develop before hatching and, thus, egg and yolk sizes and hatching temperature affect long-term growth performance and meat production of chicken. On the contrary, because fishes are able to generate new muscle fibers lifelong, the impact of early nutrition on fish growth performance is expected to be minor, which requires further studies. Copyright © 2015 Elsevier Ltd. All rights reserved.
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MicroRNA-944 Affects Cell Growth by Targeting EPHA7 in Non-Small Cell Lung Cancer.
Liu, Minxia; Zhou, Kecheng; Cao, Yi
2016-09-26
MicroRNAs (miRNAs) have critical roles in lung tumorigenesis and development. To determine aberrantly expressed miRNAs involved in non-small cell lung cancer (NSCLC) and investigate pathophysiological functions and mechanisms, we firstly carried out small RNA deep sequencing in NSCLC cell lines (EPLC-32M1, A549 and 801D) and a human immortalized cell line 16HBE, we then studied miRNA function by cell proliferation and apoptosis. cDNA microarray, luciferase reporter assay and miRNA transfection were used to investigate interaction between the miRNA and target gene. miR-944 was significantly down-regulated in NSCLC and had many putative targets. Moreover, the forced expression of miR-944 significantly inhibited the proliferation of NSCLC cells in vitro. By integrating mRNA expression data and miR-944-target prediction, we disclosed that EPHA7 was a potential target of miR-944, which was further verified by luciferase reporter assay and microRNA transfection. Our data indicated that miR-944 targets EPHA7 in NSCLC and regulates NSCLC cell proliferation, which may offer a new mechanism underlying the development and progression of NSCLC.
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Alfonso-Rodríguez, Camilo-Andrés; Garzón, Ingrid; Garrido-Gómez, Juan; Oliveira, Ana-Celeste-Ximenes; Martín-Piedra, Miguel-Ángel; Scionti, Giuseppe; Carriel, Víctor; Hernández-Cortés, Pedro; Campos, Antonio; Alaminos, Miguel
2014-01-01
Dupuytren's disease is a fibro-proliferative disease characterized by a disorder of the extracellular matrix (ECM) and high myofibroblast proliferation. However, studies failed to determine if the whole palm fascia is affected by the disease. The objective of this study was to analyze several components of the extracellular matrix of three types of tissues—Dupuytren's diseased contracture cords (DDC), palmar fascia clinically unaffected by Dupuytren's disease contracture (NPF), and normal forehand fascia (NFF). Histological analysis, quantification of cells recultured from each type of tissue, mRNA microarrays and immunohistochemistry for smooth muscle actin (SMA), fibrillar ECM components and non-fibrillar ECM components were carried out. The results showed that DDC samples had abundant fibrosis with reticular fibers and few elastic fibers, high cell proliferation and myofibroblasts, laminin and glycoproteins, whereas NFF did not show any of these findings. Interestingly, NPF tissues had more cells showing myofibroblasts differentiation and more collagen and reticular fibers, laminin and glycoproteins than NFF, although at lower level than DDC, with similar elastic fibers than DDC. Immunohistochemical expression of decorin was high in DDC, whereas versican was highly expressed NFF, with no differences for aggrecan. Cluster analysis revealed that the global expression profile of NPF was very similar to DDC, and reculturing methods showed that cells corresponding to DDC tissues proliferated more actively than NPF, and NPF more actively than NFF. All these results suggest that NPF tissues may be affected, and that a modification of the therapeutic approach used for the treatment of Dupuytren's disease should be considered. PMID:25379672
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Alfonso-Rodríguez, Camilo-Andrés; Garzón, Ingrid; Garrido-Gómez, Juan; Oliveira, Ana-Celeste-Ximenes; Martín-Piedra, Miguel-Ángel; Scionti, Giuseppe; Carriel, Víctor; Hernández-Cortés, Pedro; Campos, Antonio; Alaminos, Miguel
2014-01-01
Dupuytren's disease is a fibro-proliferative disease characterized by a disorder of the extracellular matrix (ECM) and high myofibroblast proliferation. However, studies failed to determine if the whole palm fascia is affected by the disease. The objective of this study was to analyze several components of the extracellular matrix of three types of tissues-Dupuytren's diseased contracture cords (DDC), palmar fascia clinically unaffected by Dupuytren's disease contracture (NPF), and normal forehand fascia (NFF). Histological analysis, quantification of cells recultured from each type of tissue, mRNA microarrays and immunohistochemistry for smooth muscle actin (SMA), fibrillar ECM components and non-fibrillar ECM components were carried out. The results showed that DDC samples had abundant fibrosis with reticular fibers and few elastic fibers, high cell proliferation and myofibroblasts, laminin and glycoproteins, whereas NFF did not show any of these findings. Interestingly, NPF tissues had more cells showing myofibroblasts differentiation and more collagen and reticular fibers, laminin and glycoproteins than NFF, although at lower level than DDC, with similar elastic fibers than DDC. Immunohistochemical expression of decorin was high in DDC, whereas versican was highly expressed NFF, with no differences for aggrecan. Cluster analysis revealed that the global expression profile of NPF was very similar to DDC, and reculturing methods showed that cells corresponding to DDC tissues proliferated more actively than NPF, and NPF more actively than NFF. All these results suggest that NPF tissues may be affected, and that a modification of the therapeutic approach used for the treatment of Dupuytren's disease should be considered.
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Morosetti, Roberta; Mirabella, Massimiliano; Gliubizzi, Carla; Broccolini, Aldobrando; Sancricca, Cristina; Pescatori, Mario; Gidaro, Teresa; Tasca, Giorgio; Frusciante, Roberto; Tonali, Pietro Attilio; Cossu, Giulio; Ricci, Enzo
2007-12-01
Facioscapulohumeral muscular dystrophy (FSHD) is the third most frequent inherited muscle disease. Because in FSHD patients the coexistence of affected and unaffected muscles is common, myoblasts expanded from unaffected FSHD muscles have been proposed as suitable tools for autologous cell transplantation. Mesoangioblasts are a new class of adult stem cells of mesodermal origin, potentially useful for the treatment of primitive myopathies of different etiology. Here, we report the isolation and characterization of mesoangioblasts from FSHD muscle biopsies and describe morphology, proliferation, and differentiation abilities of both mesoangioblasts and myoblasts derived from various affected and unaffected muscles of nine representative FSHD patients. We demonstrate that mesoangioblasts can be efficiently isolated from FSHD muscle biopsies and expanded to an amount of cells necessary to transplant into an adult patient. Proliferating mesoangioblasts from all muscles examined did not differ from controls in terms of morphology, phenotype, proliferation rate, or clonogenicity. However, their differentiation ability into skeletal muscle was variably impaired, and this defect correlated with the overall disease severity and the degree of histopathologic abnormalities of the muscle of origin. A remarkable differentiation defect was observed in mesoangioblasts from all mildly to severely affected FSHD muscles, whereas mesoangioblasts from morphologically normal muscles showed no myogenic differentiation block. Our study could open the way to cell therapy for FSHD patients to limit muscle damage in vivo through the use of autologous mesoangioblasts capable of reaching damaged muscles and engrafting into them, without requiring immune suppression or genetic correction in vitro. Disclosure of potential conflicts of interest is found at the end of this article.
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Aviello, Gabriella; Rowland, Ian; Gill, Christopher I; Acquaviva, Angela Maria; Capasso, Francesco; McCann, Mark; Capasso, Raffaele; Izzo, Angelo A; Borrelli, Francesca
2010-01-01
Abstract In recent years, the use of anthraquinone laxatives, in particular senna, has been associated with damage to the intestinal epithelial layer and an increased risk of developing colorectal cancer. In this study, we evaluated the cytotoxicity of rhein, the active metabolite of senna, on human colon adenocarcinoma cells (Caco-2) and its effect on cell proliferation. Cytotoxicity studies were performed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), neutral red (NR) and trans-epithelial electrical resistance (TEER) assays whereas 3H-thymidine incorporation and Western blot analysis were used to evaluate the effect of rhein on cell proliferation. Moreover, for genoprotection studies Comet assay and oxidative biomarkers measurement (malondialdehyde and reactive oxygen species) were used. Rhein (0.1–10 μg/ml) had no significant cytotoxic effect on proliferating and differentiated Caco-2 cells. Rhein (0.1 and 1 μg/ml) significantly reduced cell proliferation as well as mitogen-activated protein (MAP) kinase activation; by contrast, at high concentration (10 μg/ml) rhein significantly increased cell proliferation and extracellular-signal-related kinase (ERK) phosphorylation. Moreover, rhein (0.1–10 μg/ml): (i) did not adversely affect the integrity of tight junctions and hence epithelial barrier function; (ii) did not induce DNA damage, rather it was able to reduce H2O2-induced DNA damage and (iii) significantly inhibited the increase in malondialdehyde and reactive oxygen species (ROS) levels induced by H2O2/Fe2+. Rhein was devoid of cytotoxic and genotoxic effects in colon adenocarcinoma cells. Moreover, at concentrations present in the colon after a human therapeutic dosage of senna, rhein inhibited cell proliferation via a mechanism that seems to involve directly the MAP kinase pathway. Finally, rhein prevents the DNA damage probably via an anti-oxidant mechanism. PMID:19538468
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Domenis, Rossana; Cesselli, Daniela; Toffoletto, Barbara; Bourkoula, Evgenia; Caponnetto, Federica; Manini, Ivana; Beltrami, Antonio Paolo; Ius, Tamara; Skrap, Miran; Di Loreto, Carla
2017-01-01
A major contributing factor to glioma development and progression is its ability to evade the immune system. Nano-meter sized vesicles, exosomes, secreted by glioma-stem cells (GSC) can act as mediators of intercellular communication to promote tumor immune escape. Here, we investigated the immunomodulatory properties of GCS-derived exosomes on different peripheral immune cell populations. Healthy donor peripheral blood mononuclear cells (PBMCs) stimulated with anti-CD3, anti-CD28 and IL-2, were treated with GSC-derived exosomes. Phenotypic characterization, cell proliferation, Th1/Th2 cytokine secretion and intracellular cytokine production were analysed by distinguishing among effector T cells, regulatory T cells and monocytes. In unfractionated PBMCs, GSC-derived exosomes inhibited T cell activation (CD25 and CD69 expression), proliferation and Th1 cytokine production, and did not affect cell viability or regulatory T-cell suppression ability. Furthermore, exosomes were able to enhance proliferation of purified CD4+ T cells. In PBMCs culture, glioma-derived exosomes directly promoted IL-10 and arginase-1 production and downregulation of HLA-DR by unstimulated CD14+ monocytic cells, that displayed an immunophenotype resembling that of monocytic myeloid-derived suppressor cells (Mo-MDSCs). Importantly, the removal of CD14+ monocytic cell fraction from PBMCs restored T-cell proliferation. The same results were observed with exosomes purified from plasma of glioblastoma patients. Our results indicate that glioma-derived exosomes suppress T-cell immune response by acting on monocyte maturation rather than on direct interaction with T cells. Selective targeting of Mo-MDSC to treat glioma should be considered with regard to how immune cells allow the acquirement of effector functions and therefore counteracting tumor progression. PMID:28107450
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Domenis, Rossana; Cesselli, Daniela; Toffoletto, Barbara; Bourkoula, Evgenia; Caponnetto, Federica; Manini, Ivana; Beltrami, Antonio Paolo; Ius, Tamara; Skrap, Miran; Di Loreto, Carla; Gri, Giorgia
2017-01-01
A major contributing factor to glioma development and progression is its ability to evade the immune system. Nano-meter sized vesicles, exosomes, secreted by glioma-stem cells (GSC) can act as mediators of intercellular communication to promote tumor immune escape. Here, we investigated the immunomodulatory properties of GCS-derived exosomes on different peripheral immune cell populations. Healthy donor peripheral blood mononuclear cells (PBMCs) stimulated with anti-CD3, anti-CD28 and IL-2, were treated with GSC-derived exosomes. Phenotypic characterization, cell proliferation, Th1/Th2 cytokine secretion and intracellular cytokine production were analysed by distinguishing among effector T cells, regulatory T cells and monocytes. In unfractionated PBMCs, GSC-derived exosomes inhibited T cell activation (CD25 and CD69 expression), proliferation and Th1 cytokine production, and did not affect cell viability or regulatory T-cell suppression ability. Furthermore, exosomes were able to enhance proliferation of purified CD4+ T cells. In PBMCs culture, glioma-derived exosomes directly promoted IL-10 and arginase-1 production and downregulation of HLA-DR by unstimulated CD14+ monocytic cells, that displayed an immunophenotype resembling that of monocytic myeloid-derived suppressor cells (Mo-MDSCs). Importantly, the removal of CD14+ monocytic cell fraction from PBMCs restored T-cell proliferation. The same results were observed with exosomes purified from plasma of glioblastoma patients. Our results indicate that glioma-derived exosomes suppress T-cell immune response by acting on monocyte maturation rather than on direct interaction with T cells. Selective targeting of Mo-MDSC to treat glioma should be considered with regard to how immune cells allow the acquirement of effector functions and therefore counteracting tumor progression.
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Ding, Daofang; Wei, Songpu; Song, Yi; Li, Linghui; Du, Guoqing; Zhan, Hongsheng; Cao, Yuelong
2013-01-01
The purpose of this study was to investigate how Osthole affects glioma cell proliferation, apoptosis, invasion and migration. Rat glioma cells were treated with different concentrations of Osthole (0 µM, 25 µM, 50 µM, and 100 µM). Cell proliferation was assessed by measuring PCNA expression and CCK8 assay at different time points. Apoptosis was evaluated by measuring the expression of pro-apoptotic protein including Bax, Bcl2, PARP, and cleaved Caspase3, and of anti-apoptotic protein Survivin. Cell migration and invasion were assessed using different methods. Signaling pathways such as PI3K/Akt and MAPK, which are involved in the development of glioma cells, were also investigated in this study. Treatment with Osthole markedly inhibits glioma cell proliferation, as assessed by western blot with the PCNA antibody. Osthole also induces cell apoptosis by upregulating the expression of pro-apoptotic proteins, and by reducing the expression of anti-apoptotic factors. Moreover, C6 cell migration and invasion were efficiently inhibited in groups treated with Osthole, compared to the control group. Additionally, inhibition of PI3K/Akt and MAPK signaling pathway was also observed in C6 cells treated with Osthole. Our findings showed an anti-cancer effect of Osthole on glioma cells, including the proliferation inhibition, apoptosis induction, and migration/invasion inhibition. Further investigation in C6 glioma cells implicated the role of Osthole in essential pathways controlling glioma cell progression. Taken together, our data suggested that Osthole may have a potential application in glioma therapy. © 2014 S. Karger AG, Basel.
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Heim, Lisanne; Trump, Sonja; Mittler, Susanne; Sopel, Nina; Andreev, Katerina; Ferrazzi, Fulvia; Ekici, Arif B.; Rieker, Ralf; Springel, Rebekka; Assmann, Vera L.; Lechmann, Matthias; Koch, Sonja; Engelhardt, Marina; Trufa, Denis I.; Sirbu, Horia; Hartmann, Arndt; Finotto, Susetta
2017-01-01
ABSTRACT Genome-wide association studies (GWAS) associated Family with sequence similarity 13, member A (FAM13A) with non-small cell lung cancer (NSCLC) occurrence. Here, we found increased numbers of FAM13A protein expressing cells in the tumoral region of lung tissues from a cohort of patients with NSCLC. Moreover, FAM13A inversely correlated with CTLA4 but directly correlated with HIF1α levels in the control region of these patients. Consistently, FAM13A RhoGAP was found to be associated with T cell effector molecules like HIF1α and Tbet and was downregulated in immunosuppressive CD4+CD25+Foxp3+CTLA4+ T cells. TGFβ, a tumor suppressor factor, as well as siRNA to FAM13A, suppressed both isoforms of FAM13A and inhibited tumor cell proliferation. RNA-Seq analysis confirmed this finding. Moreover, siRNA to FAM13A induced TGFβ levels. Finally, in experimental tumor cell migration, FAM13A was induced and TGFβ accelerated this process by inducing cell migration, HIF1α, and the FAM13A RhoGAP isoform. Furthermore, siRNA to FAM13A inhibited tumor cell proliferation and induced cell migration without affecting HIF1α. In conclusion, FAM13A is involved in tumor cell proliferation and downstream of TGFβ and HIF1α, FAM13A RhoGAP is associated with Th1 gene expression and lung tumor cell migration. These findings identify FAM13A as key regulator of NSCLC growth and progression. PMID:28197372
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Salamon, Achim, E-mail: achim.salamon@med.uni-rostock.de; Jonitz-Heincke, Anika, E-mail: anika.jonitz@med.uni-rostock.de; Adam, Stefanie, E-mail: stefanie.adam@med.uni-rostock.de
Cartilaginous matrix-degenerative diseases like osteoarthritis (OA) are characterized by gradual cartilage erosion, and also by increased presence of cells with mesenchymal stem cell (MSC) character within the affected tissues. Moreover, primary chondrocytes long since are known to de-differentiate in vitro and to be chondrogenically re-differentiable. Since both findings appear to conflict with each other, we quantitatively assessed the mesenchymal differentiation potential of OA patient cartilage-derived cells (CDC) towards the osteogenic and adipogenic lineage in vitro and compared it to that of MSC isolated from adipose tissue (adMSC) of healthy donors. We analyzed expression of MSC markers CD29, CD44, CD105, andmore » CD166, and, following osteogenic and adipogenic induction in vitro, quantified their expression of osteogenic and adipogenic differentiation markers. Furthermore, CDC phenotype and proliferation were monitored. We found that CDC exhibit an MSC CD marker expression pattern similar to adMSC and a similar increase in proliferation rate during osteogenic differentiation. In contrast, the marked reduction of proliferation observed during adipogenic differentiation of adMSC was absent in CDC. Quantification of differentiation markers revealed a strong osteogenic differentiation potential for CDC, however almost no capacity for adipogenic differentiation. Since in the pathogenesis of OA, cartilage degeneration coincides with high bone turnover rates, the high osteogenic differentiation potential of OA patient-derived CDC may affect clinical therapeutic regimens aiming at autologous cartilage regeneration in these patients. - Highlights: • We analyze the mesenchymal differentiation capacity of cartilage-derived cells (CDC). • CDC express mesenchymal stem cell (MSC) markers CD29, CD44, CD105, and CD166. • CDC and MSC proliferation is reduced in adipogenesis and increased in osteogenesis. • Adipogenic differentiation is virtually absent in CDC, but strong in MSC. • Osteogenic differentiation is significantly stronger for CDC than for MSC.« less
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Creo, Pasquale; Bergante, Sonia; Conforti, Erika; Banfi, Giuseppe
2018-01-01
Adult stem cell-based therapeutic approaches for tissue regeneration have been proposed for several years. However, adult stem cells are usually limited in number and difficult to be expanded in vitro, and they usually tend to quickly lose their potency with passages, as they differentiate and become senescent. Culturing stem cells under reduced oxygen tensions (below 21%) has been proposed as a tool to increase cell proliferation, but many studies reported opposite effects. In particular, cell response to hypoxia seems to be very stem cell type specific. Nonetheless, it is clear that a major role in this process is played by the hypoxia inducible factor (HIF), the master regulator of cell response to oxygen deprivation, which affects cell metabolism and differentiation. Herein, we report that a chemical activation of HIF in human tendon stem cells reduces their proliferation and inhibits their differentiation in a reversible and dose-dependent manner. These results support the notion that hypoxia, by activating HIF, plays a crucial role in preserving stem cells in an undifferentiated state in the “hypoxic niches” present in the tissue in which they reside before migrating in more oxygenated areas to heal a damaged tissue. PMID:29713352
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Blanco-Calvo, Eduardo; Rivera, Patricia; Arrabal, Sergio; Vargas, Antonio; Pavón, Francisco Javier; Serrano, Antonia; Castilla-Ortega, Estela; Galeano, Pablo; Rubio, Leticia; Suárez, Juan; Rodriguez de Fonseca, Fernando
2014-01-01
Addiction to major drugs of abuse, such as cocaine, has recently been linked to alterations in adult neurogenesis in the hippocampus. The endogenous cannabinoid system modulates this proliferative response as demonstrated by the finding that pharmacological activation/blockade of cannabinoid CB1 and CB2 receptors not only modulates neurogenesis but also modulates cell death in the brain. In the present study, we evaluated whether the endogenous cannabinoid system affects cocaine-induced alterations in cell proliferation. To this end, we examined whether pharmacological blockade of either CB1 (Rimonabant, 3 mg/kg) or CB2 receptors (AM630, 3 mg/kg) would affect cell proliferation [the cells were labeled with 5-bromo-2′-deoxyuridine (BrdU)] in the subventricular zone (SVZ) of the lateral ventricle and the dentate subgranular zone (SGZ). Additionally, we measured cell apoptosis (as monitored by the expression of cleaved caspase-3) and glial activation [by analyzing the expression of glial fibrillary acidic protein (GFAP) and Iba-1] in the striatum and hippocampus during acute and repeated (4 days) cocaine administration (20 mg/kg). The results showed that acute cocaine exposure decreased the number of BrdU-immunoreactive (ir) cells in the SVZ and SGZ. In contrast, repeated cocaine exposure reduced the number of BrdU-ir cells only in the SVZ. Both acute and repeated cocaine exposure increased the number of cleaved caspase-3-, GFAP- and Iba1-ir cells in the hippocampus, and this effect was counteracted by AM630 or Rimonabant, which increased the number of BrdU-, GFAP-, and Iba1-ir cells in the hippocampus. These results indicate that the changes in neurogenic, apoptotic and gliotic processes that were produced by repeated cocaine administration were normalized by pharmacological blockade of CB1 and CB2. The restorative effects of cannabinoid receptor blockade on hippocampal cell proliferation were associated with the prevention of the induction of conditioned locomotion but not with the prevention of cocaine-induced sensitization. PMID:24409127
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Vitamin E, signalosomes and gene expression in T cells
USDA-ARS?s Scientific Manuscript database
CD4+T cells from aged humans or mice show significant reductions in IL-2 production upon activation. The resulting decreased proliferation is linked to higher risks of infection in the elderly. Several lines of evidence indicate that intrinsic defects preferentially affecting the naïve subset of CD4...
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Mi, Yuanyuan; Sun, Chuanyu; Wei, Bingbing; Sun, Feiyu; Guo, Yijun; Hu, Qingfeng; Ding, Weihong; Zhu, Lijie; Xia, Guowei
2018-01-01
Label-free quantitative proteomics has broad applications in the identification of differentially expressed proteins. Here, we applied this method to identify differentially expressed proteins (such as coatomer subunit beta 2 [COPB2]) and evaluated the functions and molecular mechanisms of these proteins in prostate cancer (PCA) cell proliferation. Proteins extracted from surgically resected PCA tissues and adjacent tissues of 3 patients were analyzed by label-free quantitative proteomics. The target protein was confirmed by bioinformatics and GEO dataset analyses. To investigate the role of the target protein in PCA, we used lentivirus-mediated small-interfering RNA (siRNA) to knockdown protein expression in the prostate carcinoma cell line, CWR22RV1 cells and assessed gene and protein expression by reverse transcription quantitative polymerase chain reaction and western blotting. CCK8 and colony formation assays were conducted to evaluate cell proliferation. Cell cycle distributions and apoptosis were assayed by flow cytometry. We selected the differentiation-related protein COPB2 as our target protein based on the results of label-free quantitative proteomics. High expression of COPB2 was found in PCA tissue and was related to poor overall survival based on a public dataset. Cell proliferation was significantly inhibited in COPB2-knockdown CWR22RV1 cells, as demonstrated by CCK8 and colony formation assays. Additionally, the apoptosis rate and percentage of cells in the G 1 phase were increased in COPB2-knockdown cells compared with those in control cells. CDK2, CDK4, and cyclin D1 were downregulated, whereas p21 Waf1/Cip1 and p27 Kip1 were upregulated, affecting the cell cycle signaling pathway. COPB2 significantly promoted CWR22RV1 cell proliferation through the cell cycle signaling pathway. Thus, silencing of COPB2 may have therapeutic applications in PCA. Copyright © 2017 Elsevier Inc. All rights reserved.
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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.
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N-acetylcysteine stimulates protein synthesis in enterocytes independently of glutathione synthesis.
Yi, Dan; Hou, Yongqing; Wang, Lei; Long, Minhui; Hu, Shengdi; Mei, Huimin; Yan, Liqiong; Hu, Chien-An Andy; Wu, Guoyao
2016-02-01
Dietary supplementation with N-acetylcysteine (NAC) has been reported to improve intestinal health and treat gastrointestinal diseases. However, the underlying mechanisms are not fully understood. According to previous reports, NAC was thought to exert its effect through glutathione synthesis. This study tested the hypothesis that NAC enhances enterocyte growth and protein synthesis independently of cellular glutathione synthesis. Intestinal porcine epithelial cells were cultured for 3 days in Dulbecco's modified Eagle medium containing 0 or 100 μM NAC. To determine a possible role for GSH (the reduced form of glutathione) in mediating the effect of NAC on cell growth and protein synthesis, additional experiments were conducted using culture medium containing 100 μM GSH, 100 μM GSH ethyl ester (GSHee), diethylmaleate (a GSH-depletion agent; 10 μM), or a GSH-synthesis inhibitor (buthionine sulfoximine, BSO; 20 μM). NAC increased cell proliferation, GSH concentration, and protein synthesis, while inhibiting proteolysis. GSHee enhanced cell proliferation and GSH concentration without affecting protein synthesis but inhibited proteolysis. Conversely, BSO or diethylmaleate reduced cell proliferation and GSH concentration without affecting protein synthesis, while promoting protein degradation. At the signaling level, NAC augmented the protein abundance of total mTOR, phosphorylated mTOR, and phosphorylated 70S6 kinase as well as mRNA levels for mTOR and p70S6 kinase in IPEC-1 cells. Collectively, these results indicate that NAC upregulates expression of mTOR signaling proteins to stimulate protein synthesis in enterocytes independently of GSH generation. Our findings provide a hitherto unrecognized biochemical mechanism for beneficial effects of NAC in intestinal cells.
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Cell Fusion as a Cause of Prostate Cancer Metastasis
2009-03-01
PC-3 cells? Does XRMV2 transform normal human cells? Does XRMV2 affect cell proliferation or viability? Is XRMV2 present in other prostate cancer...retroviral transduction. pathways regulated by tetraploidy in premalignant cells (Figure 1). In this experimental system, normal diploid human ...or findings contained in this report are those of the author(s) and should not be construed as an official Department of the Army position, policy
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Musa, Julian; Aynaud, Marie-Ming; Mirabeau, Olivier; Delattre, Olivier; Grünewald, Thomas GP
2017-01-01
Limitless cell proliferation, evasion from apoptosis, dedifferentiation, metastatic spread and therapy resistance: all these properties of a cancer cell contribute to its malignant phenotype and affect patient outcome. MYBL2 (alias B-Myb) is a transcription factor of the MYB transcription factor family and a physiological regulator of cell cycle progression, cell survival and cell differentiation. When deregulated in cancer cells, MYBL2 mediates the deregulation of these properties. In fact, MYBL2 is overexpressed and associated with poor patient outcome in numerous cancer entities. MYBL2 and players of its downstream transcriptional network can be used as prognostic and/or predictive biomarkers as well as potential therapeutic targets to offer less toxic and more specific anti-cancer therapies in future. In this review, we summarize current knowledge on the physiological roles of MYBL2 and highlight the impact of its deregulation on cancer initiation and progression. PMID:28640249
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The effect of simulated microgravity on hybridoma cells
NASA Astrophysics Data System (ADS)
Skok, Marina V.; Koval, Ludmila M.; Petrova, Yulia I.; Lykhmus, Olena Y.; Kolibo, Denis V.; Romanyuk, Svitlana I.; Yevdokimova, Nataliya Y.; Komisarenko, Sergiy V.
2005-04-01
The effect of clinostat-simulated microgravity on SP-2/0 and 1D6 hybridoma cells was studied. Clinorotation during 4-5 days at 1.5 rounds per minute decreased dramatically their proliferating capacity: the rotated cells divided less than once while control cells performed 4-5 divisions. They decreased the non-specific adhesion to tissue culture plastic, but increased the number of cell-to-cell contacts. Such phenomenological changes were accompanied with the alterations in pericellular glycosaminoglycans: decreased accumulation of hyaluronic acid and increased accumulation of chondroitin/dermatan-sulfate, as well as with the increase of cytoplasmic Ca concentration. Clinorotation resulted in hybridoma nicotinic receptor desensitization but not down-regulation. In contrast, both the quantity and quality (molecular isoforms, affinity and specificity) of the antibody produced by 1D6 hybridoma cells were not altered by clinorotation. It is concluded that simulated microgravity affected the proliferating and adhesive, but not biosynthetic properties of hybridoma cells.
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Tomasello, Laura; Mauceri, Rodolfo; Coppola, Antonina; Pitrone, Maria; Pizzo, Giuseppe; Campisi, Giuseppina; Pizzolanti, Giuseppe; Giordano, Carla
2017-08-01
Chronic periodontal disease is an infectious disease consisting of prolonged inflammation of the supporting tooth tissue and resulting in bone loss. Guided bone regeneration procedures have become common and safe treatments in dentistry, and in this context dental stem cells would represent the ideal solution as autologous cells. In this study, we verified the ability of dental pulp mesenchymal stem cells (DPSCs) and gingival mesenchymal stem cells (GMSCs) harvested from periodontally affected teeth to produce new mineralized bone tissue in vitro, and compared this to cells from healthy teeth. To characterize DPSCs and GMSCs, we assessed colony-forming assay, immunophenotyping, mesenchymal/stem cell phenotyping, stem gene profiling by means of flow cytometry, and quantitative polymerase chain reaction (qPCR). The effects of proinflammatory cytokines on mesenchymal stem cell (MSC) proliferation and differentiation potential were investigated. We also observed participation of several heat shock proteins (HSPs) and actin-depolymerizing factors (ADFs) during osteogenic differentiation. DPSCs and GMSCs were successfully isolated both from periodontally affected dental tissue and controls. Periodontally affected dental MSCs proliferated faster, and the inflamed environment did not affect MSC marker expressions. The calcium deposition was higher in periodontally affected MSCs than in the control group. Proinflammatory cytokines activate a cytoskeleton remodeling, interacting with HSPs including HSP90 and HSPA9, thioredoxin-1, and ADFs such as as profilin-1, cofilin-1, and vinculin that probably mediate the increased acquisition in the inflamed environment. Our findings provide evidence that periodontally affected dental tissue (both pulp and gingiva) can be used as a source of MSCs with intact stem cell properties. Moreover, we demonstrated that the osteogenic capability of DPSCs and GMSCs in the test group was not only preserved but increased by the overexpression of several proinflammatory cytokine-dependent chaperones and stress response proteins.
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MicroRNA-206: Effective Inhibition of Gastric Cancer Progression through the c-Met Pathway
Zheng, Zhiqiang; Yan, Dongsheng; Chen, Xiaoyan; Huang, He; Chen, Ke; Li, Guangjing; Zhou, Linglin; Zheng, Dandan; Tu, LiLi; Dong, Xiang Da
2015-01-01
MicroRNAs are endogenous short chain nucleotide RNAs that regulate gene function by direct binding of target mRNAs. In this study, we investigated the effects of microRNA-206 (miR-206) on the development of gastric cancer. miR-206 was first confirmed to be downregulated in gastric cancer specimens. Conversely, upregulation of c-Met was confirmed in tissue samples of human gastric cancer, with its level inversely correlated with miR-206 expression. Introduction of miR-206 inhibited cellular proliferation by inducing G1 cell cycle arrest, as well as migration and invasion. Moreover, important proliferation and/or migration related molecules such as c-Met, CDK4, p-Rb, p-Akt and p-ERK were confirmed to be downregulated by Western blot analysis. Targeting of c-Met also directly affected AGS cell proliferation, migration and invasion. In vivo, miR-206 expressing tumor cells also displayed growth delay in comparison to unaffected tumor cells. Our results demonstrated that miR-206 suppressed c-Met expression in gastric cancer and could function as a potent tumor suppressor in c-Met overexpressing tumors. Inhibition of miR-206 function could contribute to aberrant cell proliferation and migration, leading to gastric cancer development. PMID:26186594
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Yu, Sungsook; Cho, Hyun Hwa; Joo, Hye Joon; Bae, Yong Chan; Jung, Jin Sup
2008-10-01
Toll-like receptors (TLRs) sense microorganism components and are critical host mediators of inflammation during infection. Recently, TLRs have been reported to be involved in cell proliferation and differentiation. We previously reported that TLR agonists might affect proliferation and differentiation of human adipose tissue-derived mesenchymal stem cells (hASCs). In this study, we sought to determine whether TLR signaling is dependent on MyD88 in hASCs. The hASCs were downregulated using LV-GFP-miR-MyD88, a lentiviral construct inserted siRNA against human MyD88 that significantly inhibited cell proliferation. MyD88 downregulation reduced NF-kappaB activation and enhancement of osteogenic differentiation induced by peptidoglycan (PGN) more significantly than that induced by lipopolysaccharide (LPS). Although LPS- and PGN-induced cytokine secretions were decreased greatly by MyD88 downregulation, IFN-gamma-induced protein-10 (IP10) and IFNbeta expression were enhanced by LPS irrespective of the downregulation of MyD88. These results suggest that TLR signaling is mediated via MyD88-independent pathways as well as MyD88-dependent pathways in hASCs and that MyD88 contributes to the regulation of cell proliferation and differentiation in hASCs.
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Effects of corticosteroids and hyaluronic acid on torn rotator cuff tendons in vitro and in rats.
Nakamura, Hidehiro; Gotoh, Masafumi; Kanazawa, Tomonoshin; Ohta, Keisuke; Nakamura, Keiichirou; Honda, Hirokazu; Ohzono, Hiroki; Shimokobe, Hisao; Mitsui, Yasuhiro; Shirachi, Isao; Okawa, Takahiro; Higuchi, Fujio; Shirahama, Masahiro; Shiba, Naoto; Matsueda, Satoko
2015-10-01
Corticosteroids (CS) or hyaluronic acid (HA) is used in subacromial injection for the conservative treatment of rotator cuff tears (RCT); this study addresses the question of how CS and HA affect the tendon tissue and fibroblasts in vitro and in rats. Cell proliferation assays were performed in human tendon fibroblasts from RCT. Rats underwent surgery to create RCT, and the surgical sites were injected with CS or HA. The rotator cuff tendons were subjected to biomechanical testing, microscopic and immunohistochemical analysis of proliferating cell nuclear antigen (PCNA), and ultrastructural analysis. Cell proliferation was significantly decreased with CS in vitro (p < 0.05). Maximal load of CS-treated tendons was significantly decreased compared with that of HA-treated tendons (p < 0.05), as well as PCNA(+) cells at 2 weeks (p < 0.05). Ultrastructural observations of the CS-treated rats detected apoptosis of tendon fibroblasts 24 h after surgery. Histological and biomechanical data 4 weeks after surgery were not significant among the three groups. Unlike HA, CS caused cell death, and inhibition of the proliferation of tendon fibroblasts, leading to a delay of tendon healing involved and a subsequent decrease of biomechanical strength at the surgical site. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.
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Iskusnykh, Igor Y; Buddington, Randal K; Chizhikov, Victor V
2018-08-01
Preterm birth is a leading cause of long-term motor and cognitive deficits. Clinical studies suggest that some of these deficits result from disruption of cerebellar development, but the mechanisms that mediate cerebellar abnormalities in preterm infants are largely unknown. Furthermore, it remains unclear whether preterm birth and precocious exposure to the ex-utero environment directly disrupt cerebellar development or indirectly by increasing the probability of cerebellar injury, including that resulting from clinical interventions and protocols associated with the care of preterm infants. In this study, we analyzed the cerebellum of preterm pigs delivered via c-section at 91% term and raised for 10 days, until term-equivalent age. The pigs did not receive any treatments known or suspected to affect cerebellar development and had no evidence of brain damage. Term pigs sacrificed at birth were used as controls. Immunohistochemical analysis revealed that preterm birth did not affect either size or numbers of Purkinje cells or molecular layer interneurons at term-equivalent age. The number of granule cell precursors and Bergmann glial fibers, however, were reduced in preterm pigs. Preterm pigs had reduced proliferation but not differentiation of granule cells. qRT-PCR analysis of laser capture microdissected external granule cell layer showed that preterm pigs had a reduced expression of Ccnd1 (Cyclin D1), Ccnb1 (Cyclin B1), granule cell master regulatory transcription factor Atoh1, and signaling molecule Jag1. In vitro rescue experiments identified Jag1 as a central granule cell gene affected by preterm birth. Thus, preterm birth and precocious exposure to the ex-utero environment disrupt cerebellum by modulating expression of key cerebellar developmental genes, predominantly affecting development of granule precursors and Bergmann glia. Copyright © 2018 Elsevier Inc. All rights reserved.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Liu, Qingqing; Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qi-xiu Road, Nantong 226001, Jiangsu Province; Tao, Tao
As an essential post-translational modification, O-GlcNAcylation has been thought to be able to modulate various nuclear and cytoplasmic proteins and is emerging as a key regulator of multiple biological processes, such as transcription, cell growth, signal transduction, and cell motility. Recently, authoritative glycomics analyses have reported extensive crosstalk between O-GlcNAcylation and phosphorylation, which always dynamically interplay with each other and regulate signaling, transcription, and other cellular processes. Also, plentiful studies have shown close correlation between YB-1 phosphorylation and tumorigenesis. Therefore, our study aimed to determine whether YB-1 was O-GlcNAc modified and whether such modification could interact with its phosphorylation duringmore » the process of HCC development. Western blot and immunohistochemistry were firstly conducted to reveal obvious up-regulation of YB-1, OGT and O-GlcNAc modification in HCC tissues. What is more, not only YB-1 was identified to be O-GlcNAcylated but hyper-O-GlcNAcylation was demonstrated to facilitate HCC cell proliferation in a YB-1 dependent manner. Moreover, we detected four specific O-GlcNAc sites and confirmed T126A to be the most effective mutant in HCC cell proliferation via close O-GlcNAcylation-phosphorylation interaction. Even more interestingly, we discovered that T126A-induced HCC cell retardation and subdued transcriptional activity of YB-1 could be partially reversed by T126A/S102E mutant. From all above, it is not difficult to find that glycosylated-YB-1 mainly enhanced cell proliferation through congenerous actions with YB-1 phosphorylation and thus played indispensable roles in fine-tuning cell proliferation and procession of HCC. - Highlights: • YB-1 and OGT are associated with HCC prognosis. • YB-1 is O-GlcNAc modified in HCC. • Hyper-O-GlcNAcylation promotes HCC cell proliferation in dependent of YB-1. • The proliferating role of O-GlcNAcylation is based on Ser102 phosphorylation of YB-1.« less
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Protease-Activated Receptors and other G-Protein-Coupled Receptors: the Melanoma Connection
Rosero, Rebecca A.; Villares, Gabriel J.; Bar-Eli, Menashe
2016-01-01
The vast array of G-protein-coupled receptors (GPCRs) play crucial roles in both physiological and pathological processes, including vision, coagulation, inflammation, autophagy, and cell proliferation. GPCRs also affect processes that augment cell proliferation and metastases in many cancers including melanoma. Melanoma is the deadliest form of skin cancer, yet limited therapeutic modalities are available to patients with metastatic melanoma. Studies have found that both chemokine receptors and protease-activated receptors, both of which are GPCRs, are central to the metastatic melanoma phenotype and may serve as potential targets in novel therapies against melanoma and other cancers. PMID:27379162
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Protease-Activated Receptors and other G-Protein-Coupled Receptors: the Melanoma Connection.
Rosero, Rebecca A; Villares, Gabriel J; Bar-Eli, Menashe
2016-01-01
The vast array of G-protein-coupled receptors (GPCRs) play crucial roles in both physiological and pathological processes, including vision, coagulation, inflammation, autophagy, and cell proliferation. GPCRs also affect processes that augment cell proliferation and metastases in many cancers including melanoma. Melanoma is the deadliest form of skin cancer, yet limited therapeutic modalities are available to patients with metastatic melanoma. Studies have found that both chemokine receptors and protease-activated receptors, both of which are GPCRs, are central to the metastatic melanoma phenotype and may serve as potential targets in novel therapies against melanoma and other cancers.
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Nucleoplasmic calcium regulates cell proliferation through legumain.
Andrade, Viviane; Guerra, Mateus; Jardim, Camila; Melo, Flavia; Silva, Wamberto; Ortega, Jose M; Robert, Marie; Nathanson, Michael H; Leite, Fatima
2011-09-01
Nucleoplasmic Ca(2+) regulates cell growth in the liver, but the proteins through which this occurs are unknown. We used Rapid Subtraction Hybridization (RaSH) to subtract genes in SKHep1 liver cells expressing the Ca(2+) buffer protein parvalbumin (PV) targeted to the nucleus, from genes in cells expressing a mutated form of nuclear-targeted PV which has one of two Ca(2+)-binding sites inactivated. The subtraction permitted the selection of genes whose expression was affected by a small alteration in nuclear Ca(2+) concentration. The asparaginyl endopeptidase legumain (LGMN) was identified in this screening. When Ca(2+) was buffered in the nucleus of SKHep1 cells, LGMN mRNA was decreased by 97%, in part by a transcriptional mechanism, and decreased expression at the protein level was observed by immunoblot and immunofluorescence. Treatment with hepatocyte growth factor increased LGMN expression. Knockdown of LGMN by siRNA decreased proliferation of SKHep1 cells by ∼50% as measured both by BrdU uptake and mitotic index, although an inhibitor of LGMN activity did not affect BrdU incorporation. A significant reduction in the fraction of cells in G2/M phase was seen as well. This was associated with increases in the expression of cyclins A and E. Furthermore, LGMN expression was increased in hepatocellular carcinoma cells relative to normal hepatocytes in the same specimens. These findings suggest a new role for LGMN and provide evidence that nuclear Ca(2+) signals regulate cell proliferation in part through the modulation of LGMN expression. Increased expression of LGMN may be involved in liver carcinogenesis. Copyright © 2011 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.
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A simple non-perturbing cell migration assay insensitive to proliferation effects
Glenn, Honor L.; Messner, Jacob; Meldrum, Deirdre R.
2016-01-01
Migration is a fundamental cellular behavior that plays an indispensable role in development and homeostasis, but can also contribute to pathology such as cancer metastasis. Due to its relevance to many aspects of human health, the ability to accurately measure cell migration is of broad interest, and numerous approaches have been developed. One of the most commonly employed approaches, because of its simplicity and throughput, is the exclusion zone assay in which cells are allowed to migrate into an initially cell-free region. A major drawback of this assay is that it relies on simply counting cells in the exclusion zone and therefore cannot distinguish the effects of proliferation from migration. We report here a simple modification to the exclusion zone migration assay that exclusively measures cell migration and is not affected by proliferation. This approach makes use of a lineage-tracing vital stain that is retained through cell generations and effectively reads out migration relative to the original, parental cell population. This modification is simple, robust, non-perturbing, and inexpensive. We validate the method in a panel of cell lines under conditions that inhibit or promote migration and demonstrate its use in normal and cancer cell lines as well as primary cells. PMID:27535324
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HDAC inhibitors: modulating leukocyte differentiation, survival, proliferation and inflammation.
Sweet, Matthew J; Shakespear, Melanie R; Kamal, Nabilah A; Fairlie, David P
2012-01-01
Therapeutic effects of histone deacetylase (HDAC) inhibitors in cancer models were first linked to their ability to cause growth arrest and apoptosis of tumor cells. It is now clear that these agents also have pleiotropic effects on angiogenesis and the immune system, and some of these properties are likely to contribute to their anti-cancer activities. It is also emerging that inhibitors of specific HDACs affect the differentiation, survival and/or proliferation of distinct immune cell populations. This is true for innate immune cells such as macrophages, as well as cells of the acquired immune system, for example, T-regulatory cells. These effects may contribute to therapeutic profiles in some autoimmune and chronic inflammatory disease models. Here, we review our current understanding of how classical HDACs (HDACs 1-11) and their inhibitors impact on differentiation, survival and proliferation of distinct leukocyte populations, as well as the likely relevance of these effects to autoimmune and inflammatory disease processes. The ability of HDAC inhibitors to modulate leukocyte survival may have implications for the rationale of developing selective inhibitors as anti-inflammatory drugs.
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Fan, J Z; Yang, X; Bi, Z G
2015-07-01
We investigated whether 6-gingerol affects the maturation and proliferation of osteoblast-like MG63 cells in vitro. Osteoblast-like MG63 cells were treated with 6-gingerol under control conditions, and experimental inflammation was induced by tumor necrosis factor-α (TNF-α). Expression of different osteogenic markers and cytokines was analyzed by real-time PCR, Western blotting, and enzyme-linked immunosorbent assay. In addition, alkaline phosphatase (ALP) enzyme activity and biomineralization as markers for differentiation were measured. Treatment with 6-gingerol resulted in insignificant effects on the proliferation rate. 6-Gingerol induced the differentiation of osteoblast-like cells with increased transcription levels of osteogenic markers, upregulated ALP enzyme activity, and enhanced mineralized nodule formation. Stimulation with TNF-α led to enhanced interleukin-6 and nuclear factor-κB expression and downregulated markers of osteoblastic differentiation. 6-Gingerol reduced the degree of inflammation in TNF-α-treated MG-63 cells. In conclusion, 6-gingerol stimulated osteoblast differentiation in normal physiological and inflammatory settings, and therefore, 6-gingerol represents a promising agent for treating osteoporosis or bone inflammation.
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Fan, J.Z.; Yang, X.; Bi, Z.G.
2015-01-01
We investigated whether 6-gingerol affects the maturation and proliferation of osteoblast-like MG63 cells in vitro. Osteoblast-like MG63 cells were treated with 6-gingerol under control conditions, and experimental inflammation was induced by tumor necrosis factor-α (TNF-α). Expression of different osteogenic markers and cytokines was analyzed by real-time PCR, Western blotting, and enzyme-linked immunosorbent assay. In addition, alkaline phosphatase (ALP) enzyme activity and biomineralization as markers for differentiation were measured. Treatment with 6-gingerol resulted in insignificant effects on the proliferation rate. 6-Gingerol induced the differentiation of osteoblast-like cells with increased transcription levels of osteogenic markers, upregulated ALP enzyme activity, and enhanced mineralized nodule formation. Stimulation with TNF-α led to enhanced interleukin-6 and nuclear factor-κB expression and downregulated markers of osteoblastic differentiation. 6-Gingerol reduced the degree of inflammation in TNF-α-treated MG-63 cells. In conclusion, 6-gingerol stimulated osteoblast differentiation in normal physiological and inflammatory settings, and therefore, 6-gingerol represents a promising agent for treating osteoporosis or bone inflammation. PMID:25923459
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Choi, Yu-Jin; Choi, Yun-Sik
2016-02-01
Nonionizing radiation is emitted from electronic devices, such as smartphones. In this study, we intended to elucidate the effect of electromagnetic radiation from smartphones on spatial working memory and progenitor cell proliferation in the hippocampus. Both male and female mice were randomly separated into two groups (radiated and control) and the radiated group was exposed to electromagnetic radiation for 9 weeks and 11 weeks for male and female mice, respectively. Spatial working memory was examined with a Y maze, and proliferation of hippocampal progenitor cells were examined by 5-bromo-2'-deoxyuridine administration and immunohistochemical detection. When spatial working memory on a Y maze was examined in the 9(th) week, there was no significant difference in the spontaneous alternation score on the Y maze between the two groups. In addition, there was no significant difference in hippocampal progenitor cell proliferation. However, immunoreactivity to glial fibrillary acidic protein was increased in exposed animals. Next, to test the effect of recovery following chronic radiation exposure, the remaining female mice were further exposed to electromagnetic radiation for 2 more weeks (total 11 weeks), and spontaneous alternation was tested 4 weeks later. In this experiment, although there was no significant difference in the spontaneous alternation scores, the number of arm entry was significantly increased. These data indicate that although chronic electromagnetic radiation does not affect spatial working memory and hippocampal progenitor cell proliferation it can mediate astrocyte activation in the hippocampus and delayed hyperactivity-like behavior.
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Effects of leptin and adiponectin on proliferation and protein metabolism of porcine myoblasts.
Will, Katja; Kalbe, Claudia; Kuzinski, Judith; Lösel, Dorothea; Viergutz, Torsten; Palin, Marie-France; Rehfeldt, Charlotte
2012-08-01
The aim of this study was to show the abundance of leptin and adiponectin receptors (LEPR, ADIPOR1, ADIPOR2) and to determine the direct effects of leptin and adiponectin on the in vitro growth of porcine skeletal muscle cells. ADIPOR1 and ADIPOR2 were abundant at mRNA and protein level in proliferating and differentiating myoblast cultures derived from semimembranosus and semitendinosus muscles of newborn piglets, whereas LEPR expression was close to the detection limit. Adiponectin (10, 20, 40 μg/ml) attenuated the proliferation of porcine myoblasts, measured as [(3)H]-thymidine incorporation and real-time monitoring of the cells in response to 24- and 48-h exposure, in a dose-dependent manner. This effect resulted from suppressed basic fibroblast growth factor (bFGF)-mediated stimulation of DNA synthesis in serum-free medium (SFM) containing bFGF. No effects of leptin (5, 10, 20, 40, 80 ng/ml) on myoblast proliferation in SFM were detectable. Neither leptin nor adiponectin altered protein synthesis and degradation in differentiating porcine myoblasts cultured in SFM. The results on receptor abundance suggest that porcine skeletal muscle cells may be sensitive to adiponectin and leptin. However, except via inhibitory interaction of adiponectin with bFGF, these adipokines appear not to affect in vitro proliferation and protein metabolism of porcine muscle cells directly under serum-free culture conditions.
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Stadelmann, Britta; Merino, María C.; Persson, Lo; Svärd, Staffan G.
2012-01-01
In the field of infectious diseases the multifaceted amino acid arginine has reached special attention as substrate for the host´s production of the antimicrobial agent nitric oxide (NO). A variety of infectious organisms interfere with this part of the host immune response by reducing the availability of arginine. This prompted us to further investigate additional roles of arginine during pathogen infections. As a model we used the intestinal parasite Giardia intestinalis that actively consumes arginine as main energy source and secretes an arginine-consuming enzyme, arginine deiminase (ADI). Reduced intestinal epithelial cell (IEC) proliferation is a common theme during bacterial and viral intestinal infections, but it has never been connected to arginine-consumption. Our specific question was thereby, whether the arginine-consumption by Giardia leads to reduced IEC proliferation, in addition to NO reduction. In vitro cultivation of human IEC lines in arginine-free or arginine/citrulline-complemented medium, as well as in interaction with different G. intestinalis isolates, were used to study effects on host cell replication by MTT assay. IEC proliferation was further analyzed by DNA content analysis, polyamine measurements and expressional analysis of cell cycle regulatory genes. IEC proliferation was reduced upon arginine-withdrawal and also in an arginine-dependent manner upon interaction with G. intestinalis or addition of Giardia ADI. We show that arginine-withdrawal by intestinal pathogens leads to a halt in the cell cycle in IECs through reduced polyamine levels and upregulated cell cycle inhibitory genes. This is of importance with regards to intestinal tissue homeostasis that is affected through reduced cell proliferation. Thus, the slower epithelial cell turnover helps the pathogen to maintain a more stable niche for colonization. This study also shows why supplementation therapy of diarrhea patients with arginine/citrulline is helpful and that citrulline especially should gain further attention in future treatment strategies. PMID:23028934
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Rapacioli, Melina; Palma, Verónica; Flores, Vladimir
2016-01-01
The central nervous system areas displaying the highest structural and functional complexity correspond to the so called cortices, i.e., concentric alternating neuronal and fibrous layers. Corticogenesis, i.e., the development of the cortical organization, depends on the temporal-spatial organization of several developmental events: (a) the duration of the proliferative phase of the neuroepithelium, (b) the relative duration of symmetric (expansive) versus asymmetric (neuronogenic) sub phases, (c) the spatial organization of each kind of cell division, (e) the time of determination and cell cycle exit and (f) the time of onset of the post-mitotic neuronal migration and (g) the time of onset of the neuronal structural and functional differentiation. The first five events depend on molecular mechanisms that perform a fine tuning of the proliferative activity. Changes in any of them significantly influence the cortical size or volume (tangential expansion and radial thickness), morphology, architecture and also impact on neuritogenesis and synaptogenesis affecting the cortical wiring. This paper integrates information, obtained in several species, on the developmental roles of cell proliferation in the development of the optic tectum (OT) cortex, a multilayered associative area of the dorsal (alar) midbrain. The present review (1) compiles relevant information on the temporal and spatial organization of cell proliferation in different species (fish, amphibians, birds, and mammals), (2) revises the main molecular events involved in the isthmic organizer (IsO) determination and localization, (3) describes how the patterning installed by IsO is translated into spatially organized neural stem cell proliferation (i.e., by means of growth factors, receptors, transcription factors, signaling pathways, etc.) and (4) describes the morpho- and histogenetic effect of a spatially organized cell proliferation in the above mentioned species. A brief section on the OT evolution is also included. This section considers how the differential operation of cell proliferation could explain differences among species. PMID:27013978
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Chrysin and its phosphate ester inhibit cell proliferation and induce apoptosis in Hela cells.
Zhang, Ting; Chen, Xiaolan; Qu, Lingbo; Wu, Jinglan; Cui, Ran; Zhao, Yufen
2004-12-01
To improve the biological activities of chrysin (CR), we synthesize Diethyl Chysin-7-yl phosphate (CPE: C(19)H(19)O(7)P) and tetraethyl bis-phosphoric ester of chrysin (CP: C(23)H(28)O(10)P(2)) through a simplified Atheron-Todd reaction. The interactions of the CR and CPE with lysozyme were explored by electrospray ionization mass spectrometry (ESI) and fluorescence spectrometry method. Experimental results indicate that CPE could form the noncovalent compound with lysozyme, while the interaction of the CR with lysozyme was not detected. In addition, whether and how the compounds CPE and CP affect proliferation and apoptosis in human cervical cancer Hela cells were investigated. Moreover, the effects of CPE and CP in Hela cells were compared with that of the nonmodified CR compound. The Hela cells were co-cultured with CR, CP, and CPE as experimental groups, respectively, and corresponding control groups treated without CR, CP, and CPE. The proliferation and apoptosis were detected using MTT assay, HCl denatured-methyl green-pyronin staining, PCNA immunohistochemistry and TUNEL techniques. The cell growth IC(50), relative absorbance (RA), proliferating index (PI), PCNA-IR (immunoreactivity IR) integration value (IV), and apoptosis index (AI) were calculated and their correlation was analyzed in each group. The results show that all CR, CP, and CPE could inhibit proliferation and induce apoptosis in Hela cells. Moreover, the effects of CP and CPE were more potent than that of CR. The CP and CPE were proved to be a kind of stronger apoptosis inducers than nonphosphated CR. There was a negative correlation between proliferation and apoptosis. In conclusion, the CR, CP, and CPE could effectively inhibit growth by down-regulated expression of PCNA, and induce apoptosis in Hela cells. The efficiency of the modified CP and CPE preceded nonmodified CR compounds. The CP and CPE may be a new potential anti-cancer drug for therapy of human cervical carcinoma.
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Gandhapudi, Siva K.; Murapa, Patience; Threlkeld, Zachary D.; Ward, Martin; Sarge, Kevin D.; Snow, Charles; Woodward, Jerold G.
2013-01-01
Heat Shock Transcription Factor 1 (HSF1) is a major transcriptional regulator of the heat shock response in eukaryotic cells. HSF1 is also evoked in response to a variety of cellular stressors including elevated temperatures, oxidative stress, and other proteotoxic stressors. Previously, we demonstrated that HSF1 is activated in naive T cells at fever range temperatures (39.5°C) and is critical for in vitro T cell proliferation at fever temperatures. In this study, we demonstrated thatmurine HSF1 became activated to the DNA-binding form and trans-activated a large number of genes in lymphoid cells strictly as a consequence of receptor activation in the absence of apparent cellular stress. Microarray analysis comparing HSF1+/+ and HSF1−/− gene expression in T cells activated at 37°C revealed a diverse set of 323 genes significantly regulated by HSF1 in non-stressed T cells. In vivo proliferation studies revealed a significant impairment of HSF1−/− T cell expansion under conditions mimicking a robust immune response (staphylococcal enterotoxin B induced T cell activation). This proliferation defect due to loss of HSF1 is observed even under non-febrile temperatures. HSF1−/− T cells activated at fever temperatures show a dramatic reduction in cyclin E and cyclin A proteins during the cell cycle, although the transcription of these genes was not affected. Finally, B cell, and hematopoietic stem cell proliferation from HSF1−/− mice, but not HSF1+/+ mice were also attenuated under stressful conditions, indicating that HSF1 is critical for the cell cycle progression of lymphoid cells activated under stressful conditions. PMID:24043900
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Evaluation of role of Notch3 signaling pathway in human lung cancer cells.
Hassan, Wael Abdo; Yoshida, Ryoji; Kudoh, Shinji; Motooka, Yamato; Ito, Takaaki
2016-05-01
There is still a debate on the extent to which Notch3 signaling is involved in lung carcinogenesis and whether such function is dependent on cancer type or not. To evaluate Notch3 expression in different types of human lung cancer cells. Notch3 was detected in human lung cancer cell lines and in tissues. Then, small interfering RNA (siRNA) was used to down-regulate the expression of Notch3 in H69AR small cell lung carcinoma (SCLC) cells; two non-small cell lung carcinoma (NSCLC) cells; A549 adenocarcinoma (ADC); and H2170 squamous cell carcinoma (SCC). In addition, Notch3 intracellular domain (N3ICD) plasmid was transfected into H1688 human SCLC cells. We observed the effect of deregulating Notch3 signaling on the following cell properties: Notch-related proteins, cell morphology, adhesion, epithelial-mesenchymal transition (EMT), motility, proliferation and neuroendocrine (NE) features of SCLC. Notch3 is mainly expressed in NSCLC, and the expression of Notch1, Hes1 and Jagged1 is affected by Notch3. Notch3 has opposite functions in SCLC and NSCLC, being a tumor suppressor in the former and tumor promoting in the latter, in the context of cell adhesion, EMT and motility. Regarding cell proliferation, we found that inhibiting Notch3 in NSCLC decreases cell proliferation and induces apoptosis in NSCLC. Notch3 has no effect on cell proliferation or NE features of SCLC. Notch3 signaling in lung carcinoma is dependent on cell type. In SCLC, Notch3 behaves as a tumor suppressor pathway, while in NSCLC it acts as a tumor-promoting pathway.
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Bjørnebekk, Astrid; Mathé, Aleksander A; Gruber, Susanne H M; Brené, Stefan
2007-01-01
Genetic background influences the responsiveness to stress and plays a crucial role in the pathophysiology of depression. In an animal model of depression, Flinders Sensitive Line rats, and Sprague Dawley controls we analyzed if 7 weeks of social isolation of adult animals affect the number of newly proliferated cells in the dentate gyrus or mRNAs of Neuropeptide Y (NPY), the NPY-Y1 receptor, nociceptin, BDNF, and the serotonin 5HT1A and 5HT2A receptors, which are molecules involved in hippocampal plasticity. Since depressive illness more frequently affects women than men, and females seem to respond differently to stressful experiences than males, female rats were used in this study. Bromodeoxyuridine, which is a thymidin analogue that is incorporated into the DNA of newly formed cells, was administered during 9 days to even out the effects of hormonal fluctuations. Social isolation increased the number of newly proliferated Bromodeoxyuridine-immunoreactive cells in the Flinders Sensitive Line rats, whereas it had no impact on the number of cells in the Sprague Dawley strain. Group housed Sprague Dawley rats had a higher expression of BDNF, NPY, and the serotonin 5HT2A receptor mRNA than "depressed" Flinders Sensitive Line. Social isolation downregulated these molecules in Sprague Dawley but not in Flinders Sensitive Line rats thereby eliminating the differences between the two strains. We demonstrate strain and gender specific responses to stress induced regulation of factors important for hippocampal plasticity. (c) 2007 Wiley-Liss, Inc.
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Bracke, A; Schäfer, S; von Bohlen Und Halbach, V; Klempin, F; Bente, K; Bracke, K; Staar, D; van den Brandt, J; Harzsch, S; Bader, M; Wenzel, U O; Peters, J; von Bohlen Und Halbach, O
2018-02-23
The (pro)renin receptor [(P)RR], also known as ATP6AP2 [ATPase 6 accessory protein 2], is highly expressed in the brain. ATP6AP2 plays a role in early brain development, adult hippocampal neurogenesis and in cognitive functions. Lack of ATP6AP2 has deleterious effects, and mutations of ATP6AP2 in humans are associated with, e.g. X-linked intellectual disability. However, little is known about the effects of over-expression of ATP6AP2 in the adult brain. We hypothesized that mice over-expressing ATP6AP2 in the brain might exhibit altered neuroanatomical features and behavioural responses. To this end, we investigated heterozygous transgenic female mice and confirmed increased levels of ATP6AP2 in the brain. Our data show that over-expression of ATP6AP2 does not affect adult hippocampal neurogenesis, exercise-induced cell proliferation, or dendritic spine densities in the hippocampus. Only a reduced ventricular volume on the gross morphological level was found. However, ATP6AP2 over-expressing mice displayed altered exploratory behaviour with respect to the hole-board and novel object recognition tests. Moreover, primary adult hippocampal neural stem cells over-expressing ATP6AP2 exhibit a faster cell cycle progression and increased cell proliferation. Together, in contrast to the known deleterious effects of ATP6AP2 depletion, a moderate over-expression results in moderate behavioural changes and affects cell proliferation rate in vitro.
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Li, Xiaoxue; Wang, Siyang; Yang, Shuhong; Ying, Junjie; Yu, Hang; Yang, Chunlei; Liu, Yanyou; Wang, Yuhui; Cheng, Shuting; Xiao, Jing; Guo, Huiling; Jiang, Zhou; Wang, Zhengrong
2018-05-01
The circadian rhythm regulates numerous physiological activities, including sleep and wakefulness, behavior, immunity and metabolism. Previous studies have demonstrated that circadian rhythm disorder is associated with the occurrence of tumors. Responsible for regulating a number of functions, the Circadian locomotor output cycles kaput ( Clock ) gene is one of the core regulatory genes of circadian rhythm. The Clock gene has also been implicated in the occurrence and development of tumors in previously studies. The present study evaluated the role of the Clock gene in the proliferation and migration of mouse breast cancer 4T1 cells, and investigated its possible regulatory pathways and mechanisms. It was reported that downregulation of Clock facilitated the proliferation and migration of breast cancer cells. Further investigation revealed the involvement of IQ motif containing GTPase activating protein 1 (IQGAP1) protein expression in the Clock regulatory pathway, further influencing the expression of E-cadherin, a known proprietor of tumor cell migration and invasion. To the best of our knowledge, the present study is the first to report that Clock , acting through the regulation of the scaffolding protein IQGAP1, regulates the downstream expression of E-cadherin, thereby affecting tumor cell structure and motility. These results confirmed the role of Clock in breast cancer tumor etiology and provide insight regarding the molecular avenues of its regulatory nature, which may translate beyond breast cancer into other known functions of the gene.
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YAP regulates neuronal differentiation through Sonic hedgehog signaling pathway
DOE Office of Scientific and Technical Information (OSTI.GOV)
Lin, Yi-Ting; Ding, Jing-Ya; Li, Ming-Yang
2012-09-10
Tight regulation of cell numbers by controlling cell proliferation and apoptosis is important during development. Recently, the Hippo pathway has been shown to regulate tissue growth and organ size in Drosophila. In mammalian cells, it also affects cell proliferation and differentiation in various tissues, including the nervous system. Interplay of several signaling cascades, such as Notch, Wnt, and Sonic Hedgehog (Shh) pathways, control cell proliferation during neuronal differentiation. However, it remains unclear whether the Hippo pathway coordinates with other signaling cascades in regulating neuronal differentiation. Here, we used P19 cells, a mouse embryonic carcinoma cell line, as a model tomore » study roles of YAP, a core component of the Hippo pathway, in neuronal differentiation. P19 cells can be induced to differentiate into neurons by expressing a neural bHLH transcription factor gene Ascl1. Our results showed that YAP promoted cell proliferation and inhibited neuronal differentiation. Expression of Yap activated Shh but not Wnt or Notch signaling activity during neuronal differentiation. Furthermore, expression of Yap increased the expression of Patched homolog 1 (Ptch1), a downstream target of the Shh signaling. Knockdown of Gli2, a transcription factor of the Shh pathway, promoted neuronal differentiation even when Yap was over-expressed. We further demonstrated that over-expression of Yap inhibited neuronal differentiation in primary mouse cortical progenitors and Gli2 knockdown rescued the differentiation defect in Yap over-expressing cells. In conclusion, our study reveals that Shh signaling acts downstream of YAP in regulating neuronal differentiation. -- Highlights: Black-Right-Pointing-Pointer YAP promotes cell proliferation and inhibits neuronal differentiation in P19 cells. Black-Right-Pointing-Pointer YAP promotes Sonic hedgehog signaling activity during neuronal differentiation. Black-Right-Pointing-Pointer Knockdown of Gli2 rescues the Yap-overexpression phenotype in P19 cells. Black-Right-Pointing-Pointer Knockdown of Gli2 rescues the Yap-overexpression phenotype in cortical progenitors.« less
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CSF-1 Receptor-Dependent Colon Development, Homeostasis and Inflammatory Stress Response
Huynh, Duy; Akçora, Dilara; Malaterre, Jordane; Chan, Chee Kai; Dai, Xu-Ming; Bertoncello, Ivan; Stanley, E. Richard; Ramsay, Robert G.
2013-01-01
The colony stimulating factor-1 (CSF-1) receptor (CSF-1R) directly regulates the development of Paneth cells (PC) and influences proliferation and cell fate in the small intestine (SI). In the present study, we have examined the role of CSF-1 and the CSF-1R in the large intestine, which lacks PC, in the steady state and in response to acute inflammation induced by dextran sulfate sodium (DSS). As previously shown in mouse, immunohistochemical (IHC) analysis of CSF-1R expression showed that the receptor is baso-laterally expressed on epithelial cells of human colonic crypts, indicating that this expression pattern is shared between species. Colons from Csf1r null and Csf1op/op mice were isolated and sectioned for IHC identification of enterocytes, enteroendocrine cells, goblet cells and proliferating cells. Both Csf1r−/− and Csf1op/op mice were found to have colon defects in enterocytes and enteroendocrine cell fate, with excessive goblet cell staining and reduced cell proliferation. In addition, the gene expression profiles of the cell cycle genes, cyclinD1, c-myc, c-fos, and c-myb were suppressed in Csf1r−/− colonic crypt, compared with those of WT mice and the expression of the stem cell marker gene Lgr5 was markedly reduced. However, analysis of the proliferative responses of immortalized mouse colon epithelial cells (lines; Immorto-5 and YAMC) indicated that CSF-1R is not a major regulator of colonocyte proliferation and that its effects on proliferation are indirect. In an examination of the acute inflammatory response, Csf1r +/− male mice were protected from the adverse affects of DSS-induced colitis compared with WT mice, while Csf1r +/− female mice were significantly less protected. These data indicate that CSF-1R signaling plays an important role in colon homeostasis and stem cell gene expression but that the receptor exacerbates the response to inflammatory challenge in male mice. PMID:23451116
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Veeramani, S; Yuan, T-C; Lin, F-F; Lin, M-F
2009-01-01
p66Shc is shown to negatively regulate the life span in mice through reactive oxygen species (ROS) production. Recent reports, however, revealed that p66Shc protein level is significantly elevated in several human cancer tissues and growth-stimulated carcinoma cells, suggesting a mitogenic and carcinogenic role for p66Shc. In this communication, we demonstrate for the first time that p66Shc mediates androgenic growth signals in androgen-sensitive human prostate cancer cells through mitochondrial ROS production. Growth stimulation of prostate cancer cells with 5α-dihydrotestosterone (DHT) is accompanied by increased p66Shc level and ROS production, which is abolished by antioxidant treatments. However, antioxidant treatments do not affect the transcriptional activity of androgen receptor (AR) as observed by its inability to block DHT-induced prostate-specific antigen expression, an AR-dependent correlate of prostate cancer progression. Elevated expression of p66Shc by cDNA transfection increases the basal cell proliferation and, thus, reduces additional DHT-induced cell proliferation. Furthermore, DHT increases the translocation of p66Shc into mitochondria and its interaction with cytochrome c. Conversely, both redox-negative p66Shc mutant (W134F), which is deficient in cytochrome c interaction, and p66Shc small interfering RNA decrease DHT-induced cell proliferation. These results collectively reveal a novel role for p66Shc–ROS pathway in androgen-induced prostate cancer cell proliferation and, thus, may play a role in early prostate carcinogenesis. PMID:18504439
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Orciani, Monia; Lazzarini, Raffaella; Scartozzi, Mario; Bolletta, Elisa; Mattioli-Belmonte, Monica; Scalise, Alessandro; Di Benedetto, Giovanni; Di Primio, Roberto
2013-12-01
Breast implants are widely used and at times might cause inflammation as a foreign body, followed by fibrous capsule formation around the implant. In cancer, the inflamed stroma is essential for preservation of the tumor. Mesenchymal stem cells can be recruited to sites of inflammation, and their role in cancer development is debated. The authors assessed the effects of inflammation caused by breast implants' effects on tumor. Mesenchymal stem cells were isolated from the fibrous capsules of women who underwent a second operation after 1 year (presenting inflammation) or after 20 years (not presenting inflammation) since initial surgery. After characterization, cells were co-cultured with MCF7, a breast cancer cell line. The expression of genes involved in oncogenesis, proliferation, and epithelial-to-mesenchymal transition was investigated, followed by Western blot analyses. After co-culture with mesenchymal stem cells from the inflamed capsule, MCF7 induced a dose- and time-dependent increase in proliferation. Polymerase chain reaction analyses revealed a dysregulation of genes involved in oncogenesis, proliferation, and epithelial-to-mesenchymal transition. The subsequent evaluation by Western blot did not confirm these results, showing only a modest decrease in the expression of E-cadherin after co-culture with mesenchymal stem cells (both derived from inflamed or control capsules). These data indicate that inflammation caused by breast implants partially affects proliferation of MCF7 but does not influence key mechanisms of tumor development.
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Prosper, Boris W.; Marathe, Swanand; Husain, Basma F. A.; Kernie, Steven G.; Bartlett, Perry F.; Vaidya, Vidita A.
2014-01-01
Norepinephrine regulates latent neural stem cell activity and adult hippocampal neurogenesis, and has an important role in modulating hippocampal functions such as learning, memory and mood. Adult hippocampal neurogenesis is a multi-stage process, spanning from the activation and proliferation of hippocampal stem cells, to their differentiation into neurons. However, the stage-specific effects of noradrenergic receptors in regulating adult hippocampal neurogenesis remain poorly understood. In this study, we used transgenic Nestin-GFP mice and neurosphere assays to show that modulation of α2- and β-adrenergic receptor activity directly affects Nestin-GFP/GFAP-positive precursor cell population albeit in an opposing fashion. While selective stimulation of α2-adrenergic receptors decreases precursor cell activation, proliferation and immature neuron number, stimulation of β-adrenergic receptors activates the quiescent precursor pool and enhances their proliferation in the adult hippocampus. Furthermore, our data indicate no major role for α1-adrenergic receptors, as we did not observe any change in either the activation and proliferation of hippocampal precursors following selective stimulation or blockade of α1-adrenergic receptors. Taken together, our data suggest that under physiological as well as under conditions that lead to enhanced norepinephrine release, the balance between α2- and β-adrenergic receptor activity regulates precursor cell activity and hippocampal neurogenesis. PMID:24922313
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Pasternak, Taras; Asard, Han; Potters, Geert; Jansen, Marcel A K
2014-01-01
Glutathione (GSH) is an important scavenger of Reactive Oxygen Species (ROS), precursor of metal chelating phytochelatins, xenobiotic defence compound and regulator of cell proliferation. Homoglutathione (hGSH) is a GSH homologue that is present in several taxa in the family of Fabaceae. It is thought that hGSH performs many of the stress-defence roles typically ascribed to GSH, yet little is known about the potential involvement of hGSH in controlling cell proliferation. Here we show that hGSH/GSH ratios vary across organs and cells and that these changes in hGSH/GSH ratio occur during dedifferentiation and/or cell cycle activation events. The use of a GSH/hGSH biosynthesis inhibitor resulted in impaired cytokinesis in isolated protoplasts, showing the critical importance of these thiol-compounds for cell division. However, exposure of isolated protoplasts to exogenous GSH accelerated cytokinesis, while exogenous hGSH was found to inhibit the same process. We conclude that GSH and hGSH have distinct functional roles in cell cycle regulation in Medicago sativa L. GSH is associated with meristemic cells, and promotes cell cycle activation and induction of somatic embryogenesis, while hGSH is associated with differentiated cells and embryo proliferation. Copyright © 2013 Elsevier Masson SAS. All rights reserved.
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Panda, Dulal; Rathinasamy, K.; Santra, Manas K.; Wilson, Leslie
2005-01-01
The antifungal drug griseofulvin inhibits mitosis strongly in fungal cells and weakly in mammalian cells by affecting mitotic spindle microtubule (MT) function. Griseofulvin also blocks cell-cycle progression at G2/M and induces apoptosis in human tumor cell lines. Despite extensive study, the mechanism by which the drug inhibits mitosis in human cells remains unclear. Here, we analyzed the ability of griseofulvin to inhibit cell proliferation and mitosis and to affect MT polymerization and organization in HeLa cells together with its ability to affect MT polymerization and dynamic instability in vitro. Griseofulvin inhibited cell-cycle progression at prometaphase/anaphase of mitosis in parallel with its ability to inhibit cell proliferation. At its mitotic IC50 of 20 μM, spindles in blocked cells displayed nearly normal quantities of MTs and MT organization similar to spindles blocked by more powerful MT-targeted drugs. Similar to previously published data, we found that very high concentrations of griseofulvin (>100 μM) were required to inhibit MT polymerization in vitro. However, much lower drug concentrations (1–20 μM) strongly suppressed the dynamic instability behavior of the MTs. We suggest that the primary mechanism by which griseofulvin inhibits mitosis in human cells is by suppressing spindle MT dynamics in a manner qualitatively similar to that of much more powerful antimitotic drugs, including the vinca alkaloids and the taxanes. In view of griseofulvin's lack of significant toxicity in humans, we further suggest that it could be useful as an adjuvant in combination with more powerful drugs for the treatment of cancer. PMID:15985553
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Vinpocetine attenuates neointimal hyperplasia in diabetic rat carotid arteries after balloon injury.
Wang, Ke; Wen, Li; Peng, Wenhui; Li, Hailing; Zhuang, Jianhui; Lu, Yuyan; Liu, Baoxin; Li, Xiankai; Li, Weiming; Xu, Yawei
2014-01-01
Diabetes exacerbates abnormal vascular smooth muscle cell (VSMC) accumulation in response to arterial wall injury. Vinpocetine has been shown to improve vascular remolding; however, little is known about the direct effects of vinpocetine on vascular complications mediated by diabetes. The objective of this study was to determine the effects of vinpocetine on hyperglycemia-facilitated neointimal hyperplasia and explore its possible mechanism. Nondiabetic and diabetic rats were subjected to balloon injury of the carotid artery followed by 3-week treatment with either vinpocetine (10 mg/kg/day) or saline. Morphological analysis and proliferating cell nuclear antigen (PCNA) immunostaining were performed on day 21. Rat VSMCs proliferation was determined with 5-ethynyl-20-deoxyuridine cell proliferation assays. Chemokinesis was monitored with scratch assays, and production of reactive oxygen species (ROS) was assessed using a 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) flow cytometric assay. Apoptosis was detected by annexin V-FITC/PI flow cytometric assay. Cell signaling was assessed by immunblotting. Vinpocetine prevented intimal hyperplasia in carotid arteries in both normal (I/M ratio: 93.83 ± 26.45% versus 143.2 ± 38.18%, P<0.05) and diabetic animals (I/M ratio: 120.5 ± 42.55% versus 233.46 ± 33.98%, P<0.05) when compared to saline. The in vitro study demonstrated that vinpocetine significantly inhibited VSMCs proliferation and chemokinesis as well as ROS generation and apoptotic resistance, which was induced by high glucose (HG) treatment. Vinpocetine significantly abolished HG-induced phosphorylation of Akt and JNK1/2 without affecting their total levels. For downstream targets, HG-induced phosphorylation of IκBα was significantly inhibited by vinpocetine. Vinpocetine also attenuated HG-enhanced expression of PCNA, cyclin D1 and Bcl-2. Vinpocetine attenuated neointimal formation in diabetic rats and inhibited HG-induced VSMCs proliferation, chemokinesis and apoptotic resistance by preventing ROS activation and affecting MAPK, PI3K/Akt, and NF-κB signaling.
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Vinpocetine Attenuates Neointimal Hyperplasia in Diabetic Rat Carotid Arteries after Balloon Injury
Peng, Wenhui; Li, Hailing; Zhuang, Jianhui; Lu, Yuyan; Liu, Baoxin; Li, Xiankai; Li, Weiming; Xu, Yawei
2014-01-01
Background Diabetes exacerbates abnormal vascular smooth muscle cell (VSMC) accumulation in response to arterial wall injury. Vinpocetine has been shown to improve vascular remolding; however, little is known about the direct effects of vinpocetine on vascular complications mediated by diabetes. The objective of this study was to determine the effects of vinpocetine on hyperglycemia-facilitated neointimal hyperplasia and explore its possible mechanism. Materials and Methods Nondiabetic and diabetic rats were subjected to balloon injury of the carotid artery followed by 3-week treatment with either vinpocetine (10 mg/kg/day) or saline. Morphological analysis and proliferating cell nuclear antigen (PCNA) immunostaining were performed on day 21. Rat VSMCs proliferation was determined with 5-ethynyl-20-deoxyuridine cell proliferation assays. Chemokinesis was monitored with scratch assays, and production of reactive oxygen species (ROS) was assessed using a 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA) flow cytometric assay. Apoptosis was detected by annexin V-FITC/PI flow cytometric assay. Cell signaling was assessed by immunblotting. Results Vinpocetine prevented intimal hyperplasia in carotid arteries in both normal (I/M ratio: 93.83 ± 26.45% versus 143.2 ± 38.18%, P<0.05) and diabetic animals (I/M ratio: 120.5 ± 42.55% versus 233.46 ± 33.98%, P<0.05) when compared to saline. The in vitro study demonstrated that vinpocetine significantly inhibited VSMCs proliferation and chemokinesis as well as ROS generation and apoptotic resistance, which was induced by high glucose (HG) treatment. Vinpocetine significantly abolished HG-induced phosphorylation of Akt and JNK1/2 without affecting their total levels. For downstream targets, HG-induced phosphorylation of IκBα was significantly inhibited by vinpocetine. Vinpocetine also attenuated HG-enhanced expression of PCNA, cyclin D1 and Bcl-2. Conclusions Vinpocetine attenuated neointimal formation in diabetic rats and inhibited HG-induced VSMCs proliferation, chemokinesis and apoptotic resistance by preventing ROS activation and affecting MAPK, PI3K/Akt, and NF-κB signaling. PMID:24819198
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TAM receptors affect adult brain neurogenesis by negative regulation of microglial cell activation.
Ji, Rui; Tian, Shifu; Lu, Helen J; Lu, Qingjun; Zheng, Yan; Wang, Xiaomin; Ding, Jixiang; Li, Qiutang; Lu, Qingxian
2013-12-15
TAM tyrosine kinases play multiple functional roles, including regulation of the target genes important in homeostatic regulation of cytokine receptors or TLR-mediated signal transduction pathways. In this study, we show that TAM receptors affect adult hippocampal neurogenesis and loss of TAM receptors impairs hippocampal neurogenesis, largely attributed to exaggerated inflammatory responses by microglia characterized by increased MAPK and NF-κB activation and elevated production of proinflammatory cytokines that are detrimental to neuron stem cell proliferation and neuronal differentiation. Injection of LPS causes even more severe inhibition of BrdU incorporation in the Tyro3(-/-)Axl(-/-)Mertk(-/-) triple-knockout (TKO) brains, consistent with the LPS-elicited enhanced expression of proinflammatory mediators, for example, IL-1β, IL-6, TNF-α, and inducible NO synthase, and this effect is antagonized by coinjection of the anti-inflammatory drug indomethacin in wild-type but not TKO brains. Conditioned medium from TKO microglia cultures inhibits neuron stem cell proliferation and neuronal differentiation. IL-6 knockout in Axl(-/-)Mertk(-/-) double-knockout mice overcomes the inflammatory inhibition of neurogenesis, suggesting that IL-6 is a major downstream neurotoxic mediator under homeostatic regulation by TAM receptors in microglia. Additionally, autonomous trophic function of the TAM receptors on the proliferating neuronal progenitors may also promote progenitor differentiation into immature neurons.
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Liberio, Michelle S.; Sadowski, Martin C.; Soekmadji, Carolina; Davis, Rohan A.; Nelson, Colleen C.
2014-01-01
Weak cell-surface adhesion of cell lines to tissue culture surfaces is a common problem and presents technical limitations to the design of experiments. To overcome this problem, various surface coating protocols have been developed. However, a comparative and precise real-time measurement of their impact on cell behavior has not been conducted. The prostate cancer cell line LNCaP, derived from a patient lymph node metastasis, is a commonly used model system in prostate cancer research. However, the cells’ characteristically weak attachment to the surface of tissue culture vessels and cover slips has impeded their manipulation and analysis and use in high throughput screening. To improve the adherence of LNCaP cells to the culture surface, we compared different coating reagents (poly-l-lysine, poly-l-ornithine, collagen type IV, fibronectin, and laminin) and culturing conditions and analyzed their impact on cell proliferation, adhesion, morphology, mobility and gene expression using real-time technologies. The results showed that fibronectin, poly-l-lysine and poly-l-ornithine improved LNCaP cells adherence and provoked cell morphology alterations, such as increase of nuclear and cellular area. These coating reagents also induced a higher expression of F-actin and reduced cell mobility. In contrast, laminin and collagen type IV did not improve adherence but promoted cell aggregation and affected cell morphology. Cells cultured in the presence of laminin displayed higher mobility than control cells. All the coating conditions significantly affected cell viability; however, they did not affect the expression of androgen receptor-regulated genes. Our comparative findings provide important insight for the selection of the ideal coating reagent and culture conditions for the cancer cell lines with respect to their effect on proliferation rate, attachment, morphology, migration, transcriptional response and cellular cytoskeleton arrangement. PMID:25375165
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Kiss, Izabella; Unger, Christine; Huu, Chi Nguyen; Atanasov, Atanas Georgiev; Kramer, Nina; Chatruphonprasert, Waranya; Brenner, Stefan; McKinnon, Ruxandra; Peschel, Andrea; Vasas, Andrea; Lajter, Ildiko; Kain, Renate; Saiko, Philipp; Szekeres, Thomas; Kenner, Lukas; Hassler, Melanie R; Diaz, Rene; Frisch, Richard; Dirsch, Verena M; Jäger, Walter; de Martin, Rainer; Bochkov, Valery N; Passreiter, Claus M; Peter-Vörösmarty, Barbara; Mader, Robert M; Grusch, Michael; Dolznig, Helmut; Kopp, Brigitte; Zupko, Istvan; Hohmann, Judit; Krupitza, Georg
2015-01-28
An apolar extract of the traditional medicinal plant Neurolaena lobata inhibited the expression of the NPM/ALK chimera, which is causal for the majority of anaplastic large cell lymphomas (ALCLs). Therefore, an active principle of the extract, the furanoheliangolide sesquiterpene lactone lobatin B, was isolated and tested regarding the inhibition of ALCL expansion and tumour cell intravasation through the lymphendothelium. ALCL cell lines, HL-60 cells and PBMCs were treated with plant compounds and the ALK inhibitor TAE-684 to measure mitochondrial activity, proliferation and cell cycle progression and to correlate the results with protein- and mRNA-expression of selected gene products. Several endpoints indicative for cell death were analysed after lobatin B treatment. Tumour cell intravasation through lymphendothelial monolayers was measured and potential causal mechanisms were investigated analysing NF-κB- and cytochrome P450 activity, and 12(S)-HETE production. Lobatin B inhibited the expression of NPM/ALK, JunB and PDGF-Rβ, and attenuated proliferation of ALCL cells by arresting them in late M phase. Mitochondrial activity remained largely unaffected upon lobatin B treatment. Nevertheless, caspase 3 became activated in ALCL cells. Also HL-60 cell proliferation was attenuated whereas PBMCs of healthy donors were not affected by lobatin B. Additionally, tumour cell intravasation, which partly depends on NF-κB, was significantly suppressed by lobatin B most likely due to its NF-κB-inhibitory property. Lobatin B, which was isolated from a plant used in ethnomedicine, targets malignant cells by at least two properties: I) inhibition of NPM/ALK, thereby providing high specificity in combating this most prevalent fusion protein occurring in ALCL; II) inhibition of NF-κB, thereby not affecting normal cells with low constitutive NF-κB activity. This property also inhibits tumour cell intravasation into the lymphatic system and may provide an option to manage this early step of metastatic progression. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Kamanga-Sollo, E.; Pampusch, M.S.; White, M.E.
2005-11-15
We have previously shown that cultured porcine embryonic myogenic cells (PEMC) produce both insulin-like growth factor binding protein (IGFBP)-3 and IGFBP-5 and secrete these proteins into their media. Exogenously added recombinant porcine (rp) IGFBP-3 and rpIGFBP-5 act via IGF-dependent and IGF-independent mechanisms to suppress proliferation of PEMC cultures. Furthermore, immunoneutralization of endogenous IGFBP-3 and IGFBP-5 in the PEMC culture medium results in increased DNA synthesis rate suggesting that endogenous IGFBP-3 and IGFBP-5 suppress PEMC proliferation. TGF-{beta} superfamily members myostatin and TGF-{beta}{sub 1} have also been shown to suppress proliferation of myogenic cells, and treatment of cultured PEMC with either TGF-{beta}{submore » 1} or myostatin significantly (P < 0.01) increases levels of IGFBP-3 and -5 mRNA. We have previously shown that immunoneutralization of IGFBP-3 decreases the proliferation-suppressing activity of TGF-{beta}{sub 1} and myostatin. Here, we show that immunoneutralization of IGFBP-5 also significantly (P < 0.05) decreases the DNA synthesis-suppressing activity of these molecules. Simultaneous immunoneutralization of both IGFBP-3 and IGFBP-5 in TGF-{beta}{sub 1} or myostatin-treated PEMC cultures restores Long-R3-IGF-I-stimulated DNA synthesis rates to 90% of the levels observed in control cultures receiving no TGF-{beta}{sub 1} or myostatin treatment (P < 0.05). Even though immunoneutralization of IGFBP-3 and -5 increased DNA synthesis rates in TGF-{beta}{sub 1} or myostatin-treated PEMC cultures, phosphosmad2 levels in these cultures were not affected. These findings strongly suggest that IGFBP-3 and IGFBP-5 affect processes downstream from receptor-mediated Smad phosphorylation that facilitate the ability of TGF-{beta} and myostatin to suppress proliferation of PEMC.« less
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Petrangeli, E; Lenti, L; Buchetti, B; Chinzari, P; Sale, P; Salvatori, L; Ravenna, L; Lococo, E; Morgante, E; Russo, A; Frati, L; Di Silverio, F; Russo, M A
2009-04-01
The molecular mechanism by which the lipido-sterolic extract of Serenoa repens (LSESr, Permixon) affects prostate cells remains to be fully elucidated. In androgen-independent PC3 prostate cancer cells, the LSESr-induced effects on proliferation and apoptosis were evaluated by counting cells and using a FACScan cytofluorimeter. PC3 cells were stained with JC-1 dye to detect mitochondrial membrane potential. Cell membrane lipid composition was evaluated by thin layer chromatography and gas chromatographic analysis. Akt phosphorylation was analyzed by Western blotting and cellular ultrastructure through electron microscopy. LSESr (12.5 and 25 microg/ml) administration exerted a biphasic action by both inhibiting proliferation and stimulating apoptosis. After 1 h, it caused a marked reduction in the mitochondrial potential, decreased cholesterol content and modified phospholipid composition. A decrease in phosphatidylinositol-4,5-bisphosphate (PIP2) level was coupled with reduced Akt phosphorylation. After 24 h, all of these effects were restored to pre-treatment conditions; however, the saturated (SFA)/unsaturated fatty acid (UFA) ratio increased, mainly due to a significant decrease in omega 6 content. The reduction in cholesterol content could be responsible for both membrane raft disruption and redistribution of signaling complexes, allowing for a decrease of PIP2 levels, reduction of Akt phosphorylation and apoptosis induction. The decrease in omega 6 content appears to be responsible for the prolonged and more consistent increase in the apoptosis rate and inhibition of proliferation observed after 2-3 days of LSESr treatment. In conclusion, LSESr administration results in complex changes in cell membrane organization and fluidity of prostate cancer cells that have progressed to hormone-independent status. (c) 2008 Wiley-Liss, Inc.
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Effects of biaxial oscillatory shear stress on endothelial cell proliferation and morphology.
Chakraborty, Amlan; Chakraborty, Sutirtha; Jala, Venkatakrishna R; Haribabu, Bodduluri; Sharp, M Keith; Berson, R Eric
2012-03-01
Wall shear stress (WSS) on anchored cells affects their responses, including cell proliferation and morphology. In this study, the effects of the directionality of pulsatile WSS on endothelial cell proliferation and morphology were investigated for cells grown in a Petri dish orbiting on a shaker platform. Time and location dependent WSS was determined by computational fluid dynamics (CFD). At low orbital speed (50 rpm), WSS was shown to be uniform (0-1 dyne/cm(2)) across the bottom of the dish, while at higher orbital speed (100 and 150 rpm), WSS remained fairly uniform near the center and fluctuated significantly (0-9 dyne/cm(2)) near the side walls of the dish. Since WSS on the bottom of the dish is two-dimensional, a new directional oscillatory shear index (DOSI) was developed to quantify the directionality of oscillating shear. DOSI approached zero for biaxial oscillatory shear of equal magnitudes near the center and approached one for uniaxial pulsatile shear near the wall, where large tangential WSS dominated a much smaller radial component. Near the center (low DOSI), more, smaller and less elongated cells grew, whereas larger cells with greater elongation were observed in the more uniaxial oscillatory shear (high DOSI) near the periphery of the dish. Further, cells aligned with the direction of the largest component of shear but were randomly oriented in low magnitude biaxial shear. Statistical analyses of the individual and interacting effects of multiple factors (DOSI, shear magnitudes and orbital speeds) showed that DOSI significantly affected all the responses, indicating that directionality is an important determinant of cellular responses. Copyright © 2011 Wiley Periodicals, Inc.
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Low-grade myofibroblastic proliferations of the urinary bladder.
Alquati, Sara; Gira, Federica Alessandra; Bartoli, Veronica; Contini, Sandro; Corradi, Domenico
2013-08-01
Myofibroblastic proliferations of the urinary bladder, which share some similarities with nodular fasciitis, were first reported in 1980. Since then, they have had several designations, the most frequently used being inflammatory myofibroblastic tumor. Based on both histopathologic and prognostic grounds, some authors prefer the term pseudosarcomatous myofibroblastic proliferation, at least for some of the proliferations. These same scientists also assimilate the so-called postoperative spindle cell nodules with the pseudosarcomatous myofibroblastic proliferations. Little is known about these low-grade myofibroblastic proliferations. To review the literature about low-grade myofibroblastic proliferations occurring in the urinary bladder. Textbooks and literature review. We obtained most of the clinicopathologic peculiarities from a patient population composed of the most-relevant, previously reported cases. The low-grade myofibroblastic proliferations of the urinary bladder are rare lesions affecting males more often than they do females. The most-common signs and symptoms are hematuria and dysuria. Histopathologically, they are spindle cell proliferations in a loose myxoid stroma, even though compact proliferations or hypocellular fibrous patterns can be found. Immunohistochemistry is quite nonspecific, except for ALK-1 positivity (20%-89%). Fluorescence in situ hybridization has demonstrated clonal genetic aberrations involving the ALK gene in 50% to 60% of cases. After surgery, only 6% of patients experience local recurrence, without metastases or deaths from the disease. Malignant transformation has been reported exceptionally. These myofibroblastic proliferations are probably part of a continuum with, at one end, benign pseudosarcomatous proliferations and, at the opposite end, more-aggressive lesions. Because of the frequently indolent clinical course, aggressive treatment would be unjustified.
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Morgan, Peter J; Hübner, Rayk; Rolfs, Arndt; Frech, Moritz J
2013-09-15
Calcium signals affect many developmental processes, including proliferation, migration, survival, and apoptosis, processes that are of particular importance in stem cells intended for cell replacement therapies. The mechanisms underlying Ca(2+) signals, therefore, have a role in determining how stem cells respond to their environment, and how these responses might be controlled in vitro. In this study, we examined the spontaneous Ca(2+) activity in human neural progenitor cells during proliferation and differentiation. Pharmacological characterization indicates that in proliferating cells, most activity is the result of transient receptor potential (TRP) channels that are sensitive to Gd(3+) and La(3+), with the more subtype selective antagonist Ruthenium red also reducing activity, suggesting the involvement of transient receptor potential vanilloid (TRPV) channels. In differentiating cells, Gd(3+) and La(3+)-sensitive TRP channels also appear to underlie the spontaneous activity; however, no sub-type-specific antagonists had any effect. Protein levels of TRPV2 and TRPV3 decreased in differentiated cells, which is demonstrated by western blot. Thus, it appears that TRP channels represent the main route of Ca(2+) entry in human neural progenitor cells (hNPCs), but the responsible channel types are subject to substitution under differentiating conditions. The level of spontaneous activity could be increased and decreased by lowering and raising the extracellular K(+) concentration. Proliferating cells in low K(+) slowed the cell cycle, with a disproportionate increased percentage of cells in G1 phase and a reduction in S phase. Taken together, these results suggest a link between external K(+) concentration, spontaneous Ca(2+) transients, and cell cycle distribution, which is able to influence the fate of stem and progenitor cells.
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Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis
DOE Office of Scientific and Technical Information (OSTI.GOV)
Takahara, Kiyoshi; Ii, Masaaki, E-mail: masaii@art.osaka-med.ac.jp; Inamoto, Teruo
2014-04-18
Highlights: • AdSC transplantation exhibits inhibitory effect on tumor progressions of PCa cells. • AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway. • High expression of the TGF-β1 gene in AdSCs. - Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferationmore » of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa.« less
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Luteal function during the estrous cycle in arginine-treated ewes fed different planes of nutrition.
Bass, Casie S; Redmer, Dale A; Kaminski, Samantha L; Grazul-Bilska, Anna T
2017-03-01
Functions of corpus luteum (CL) are influenced by numerous factors including hormones, growth and angiogenic factors, nutritional plane and dietary supplements such as arginine (Arg), a semi-essential amino acid and precursor for proteins, polyamines and nitric oxide (NO). The aim of this study was to determine if Arg supplementation to ewes fed different planes of nutrition influences: (1) progesterone (P4) concentrations in serum and luteal tissue, (2) luteal vascularity, cell proliferation, endothelial NO synthase (eNOS) and receptor (R) soluble guanylate cyclase β protein and mRNA expression and (3) luteal mRNA expression for selected angiogenic factors during the estrous cycle. Ewes (n = 111) were categorized by weight and randomly assigned to one of three nutritional planes: maintenance control (C), overfed (2× C) and underfed (0.6× C) beginning 60 days prior to onset of estrus. After estrus synchronization, ewes from each nutritional plane were assigned randomly to one of two treatments: Arg or saline. Serum and CL were collected at the early, mid and late luteal phases. The results demonstrated that: (1) nutritional plane affected ovulation rates, luteal vascularity, cell proliferation and NOS3, GUCY1B3, vascular endothelial growth factor (VEGF) and VEGFR2 mRNA expression, (2) Arg affected luteal vascularity, cell proliferation and NOS3, GUCY1B3, VEGF and VEGFR2 mRNA expression and (3) luteal vascularity, cell proliferation and the VEGF and NO systems depend on the stage of the estrous cycle. These data indicate that plane of nutrition and/or Arg supplementation can alter vascularization and expression of selected angiogenic factors in luteal tissue during the estrous cycle in sheep. © 2017 Society for Reproduction and Fertility.
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Deng, Song-Bai; Jing, Xiao-Dong; Wei, Xiao-Ming; Du, Jian-Lin; Liu, Ya-Jie; Qin, Qin; She, Qiang
2017-04-29
Thyroid hormone has important functions in the development and physiological function of the heart. The aim of this study was to determine whether 3,5,3'-Triiodothyronine (T3) can promote the proliferation of epicardial progenitor cells (EPCs) and to investigate the potential underlying mechanism. Our results showed that T3 significantly promoted the proliferation of EPCs in a concentration- and time-dependent manner. The thyroid hormone nuclear receptor inhibitor bisphenol A (100 μmol/L) did not affect T3's ability to induce proliferation. Further studies showed that the mRNA expression levels of mitogen-activated protein kinase 1 (MAPK1), MAPK3, and Ki67 in EPCs in the T3 group (10 nmol/L) increased 2.9-, 3-, and 4.1-fold, respectively, compared with those in the control group (P < 0.05). In addition, the mRNA expression of the cell cycle protein cyclin D1 in the T3 group increased approximately 2-fold compared with the control group (P < 0.05), and there were more EPCs in the S phase of the cell cycle (20.6% vs. 12.0%, P < 0.05). The mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway inhibitor U0126 (10 μmol/L) significantly inhibited the ability of T3 to promote the proliferation of EPCs and to alter cell cycle progression. This study suggested that T3 significantly promotes the proliferation of EPCs, and this effect may be achieved through activation of the MAPK/ERK signaling pathway. Copyright © 2017 Elsevier Inc. All rights reserved.
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Can anesthetic-analgesic technique during primary cancer surgery affect recurrence or metastasis?
Byrne, Kathryn; Levins, Kirk J; Buggy, Donal J
2016-02-01
Mortality among cancer patients is more commonly due to the effects of metastasis and recurrence as opposed to the primary tumour. Various perioperative factors have been implicated in tumour growth, including anesthetic agents and analgesia techniques. In this narrative review, we integrate this information to present a summary of the best available evidence to guide the conduct of anesthesia for primary cancer surgery. We conducted a search of the PubMed database up to May 31, 2015 to identify relevant literature using the search terms "anesthesia and metastases", "anesthetic drugs and cancer", "volatile anesthetic agents and cancer", and "anesthetic technique and cancer". There is conflicting evidence regarding volatile agents; however, the majority of studies are in vitro, suggesting that these agents are associated with enhanced expression of tumourigenic markers as well as both proliferation and migration of cancer cells. Nitrous oxide has not been shown to have any effect on cancer recurrence. Local anesthetic agents may reduce the incidence of cancer recurrence through systemic anti-inflammatory action in addition to direct effects on the proliferation and migration of cancer cells. Nonsteroidal anti-inflammatory drugs affect cancer cells via inhibition of cyclooxygenase 2 (COX-2), which leads to reduced resistance of the cancer cell to apoptosis and reduced production of prostaglandins by cancer cells. Nonsteroidal anti-inflammatory drugs also suppress the cancer cell growth cycle through effects independent of COX-2 inhibition. Opioids have been shown to inhibit the function of natural killer cells and to stimulate cancer cell proliferation through effects on angiogenesis and tumour cell signalling pathways. Supplemental oxygen at the time of surgery has a proangiogenic effect on micrometastases, while the use of perioperative dexamethasone does not affect overall rates of cancer survival. Current laboratory research suggests that perioperative interventions may impact recurrence or metastasis through effects on cancer cell signalling, the immune response, or modulation of the neuroendocrine stress response. Further evidence is awaited from prospective randomized-controlled trials. Meanwhile, with limited data upon which to make strong recommendations, anesthesiologists should seek optimal anesthesia and analgesia for their patients based on individual risk-benefit analysis and best available evidence on outcomes other than cancer recurrence.
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Crucial role of vinexin for keratinocyte migration in vitro and epidermal wound healing in vivo
DOE Office of Scientific and Technical Information (OSTI.GOV)
Kioka, Noriyuki, E-mail: nkioka@kais.kyoto-u.ac.jp; Ito, Takuya; Yamashita, Hiroshi
2010-06-10
In the process of tissue injury and repair, epithelial cells rapidly migrate and form epithelial sheets. Vinexin is a cytoplasmic molecule of the integrin-containing cell adhesion complex localized at focal contacts in vitro. Here, we investigated the roles of vinexin in keratinocyte migration in vitro and wound healing in vivo. Vinexin knockdown using siRNA delayed migration of both HaCaT human keratinocytes and A431 epidermoid carcinoma cells in scratch assay but did not affect cell proliferation. Induction of cell migration by scratching the confluent monolayer culture of these cells activated both EGFR and ERK, and their inhibitors AG1478 and U0126 substantiallymore » suppressed scratch-induced keratinocyte migration. Vinexin knockdown in these cells inhibited the scratch-induced activation of EGFR, but not that of ERK, suggesting that vinexin promotes cell migration via activation of EGFR. We further generated vinexin (-/-) mice and isolated their keratinocytes. They similarly showed slow migration in scratch assay. Furthermore, vinexin (-/-) mice exhibited a delay in cutaneous wound healing in both the back skin and tail without affecting the proliferation of keratinocytes. Together, these results strongly suggest a crucial role of vinexin in keratinocyte migration in vitro and cutaneous wound healing in vivo.« less
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Zaldibar, B; Cancio, I; Soto, M; Marigómez, I
2007-11-01
Slugs, Arion ater (L), have been proposed as sentinel organisms to assess soil health. In slugs under the influence of pollutants, digestive cell loss and the concomitant increase of excretory cells of the digestive gland have been described. The aim of the present work was to determine up to what extent digestive cell loss affects biomarkers and whether the affectation is reversible after exposure to a mixture of metal and organic pollutants. Slugs were dosed with a mixture of cadmium and kerosene in the food for 27 days. Apart from chemical analyses, the volume density of black silver deposits (Vv(BSD)) after autometallography, and acyl-CoA oxidase (AOX) activity were used as biomarkers of exposure to metals and organic compounds, respectively. As effect biomarkers, changes in the volume density of the cell types that constitute the digestive gland epithelium were calculated. Proliferating cells were identified by means of bromodeoxyuridine (BrdU) immunohistochemistry. Results revealed that the mixture of pollutants provoked an increase in Vv(BSD) and AOX activity and a decrease in the number of digestive cells. These changes had no effect in the digestive gland accumulation capacity or in the effect and exposure biomarkers employed. BrdU-labelling showed that exposure to pollutants provoked an enhanced digestive cell proliferation.
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USDA-ARS?s Scientific Manuscript database
Butyrate affects cell proliferation, differentiation and motility. Butyrate inhibits histone deacetylase (HDAC) activities and induces cell cycle arrest and apoptosis. TP53 is one of the most active upstream regulators discovered by IPA in our RNA sequencing data set. The TP53 signaling pathway pl...
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Randelli, Pietro; Menon, Alessandra; Ragone, Vincenza; Creo, Pasquale; Bergante, Sonia; Randelli, Filippo; De Girolamo, Laura; Alfieri Montrasio, Umberto; Banfi, Giuseppe; Cabitza, Paolo; Tettamanti, Guido; Anastasia, Luigi
2016-01-01
Increasing the success rate of rotator cuff healing remains tremendous challenge. Among many approaches, the possibility of activating resident stem cells in situ, without the need to isolate them from biopsies, could represent valuable therapeutic strategy. Along this line, it has been recently demonstrated that lipoaspirate product, Lipogems, contains and produces growth-factors that may activate resident stem cells. In this study, human tendon stem cells (hTSCs) from the rotator cuff were cocultured in a transwell system with the Lipogems lipoaspirate product and compared to control untreated cells in terms of cell proliferation, morphology, stem cell marker and VEGF expression, and differentiation and migration capabilities. Results showed that the Lipogems product significantly increases the proliferation rate of hTSCs without altering their stemness and differentiation capability. Moreover, treated cells increase the expression of VEGF, which is crucial for the neovascularization of the tissue during the healing process. Overall, this study supports that directly activating hTSCs with the Lipogems lipoaspirate could represent a new practical therapeutic approach. In fact, obtaining a lipoaspirate is easier, safer, and more cost-effective than harvesting cells from tendon or bone marrow biopsies, expanding them in GMP facility and then reinjecting them in the patient. PMID:27057170
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BMI-1 Promotes Self-Renewal of Radio- and Temozolomide (TMZ)-Resistant Breast Cancer Cells.
Yan, Yanfang; Wang, Ying; Zhao, Pengxin; Ma, Weiyuan; Hu, Zhigang; Zhang, Kaili
2017-12-01
Breast cancer is a hormone-dependent malignancy and is the most prevalent cause of cancer-related mortality among females. Radiation therapy and chemotherapy are common treatments of breast cancer. However, tumor relapse and metastasis following therapy are major clinical challenges. The importance of B-lymphoma Moloney murine leukemia virus insertion region-1 (BMI-1) was implicated in cell proliferation, stem cell maintenance, and tumor initiation. We established radio- and temozolomide (TMZ)-resistant (IRC-R) MCF-7 and MDA-MB-231 cell lines to investigate the mechanism involved in therapeutic resistance. Cell proliferation and sphere number were dramatically elevated, and BMI-1 was remarkably upregulated, in IRC-R cells compared to parental cells. Silencing BMI-1 by RNA interference only affected the cell proliferation of IRC-R but not parental cells, suggesting the critical role of BMI-1 in radio- and TMZ resistance. We used a xenograft mice model to elucidate that BMI-1 was necessary in tumor development by assessing tumor volume and Ki67 expression. We found that Hedgehog (Hhg) signaling exerted synergized functions together with BMI-1, implicating the importance of BMI-1 in Hhg signaling. Downregulation of BMI-1 could be an effective strategy to suppress tumor growth, which supports the potential clinical use of targeting BMI-1 in breast cancer treatment.
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Randelli, Pietro; Menon, Alessandra; Ragone, Vincenza; Creo, Pasquale; Bergante, Sonia; Randelli, Filippo; De Girolamo, Laura; Alfieri Montrasio, Umberto; Banfi, Giuseppe; Cabitza, Paolo; Tettamanti, Guido; Anastasia, Luigi
2016-01-01
Increasing the success rate of rotator cuff healing remains tremendous challenge. Among many approaches, the possibility of activating resident stem cells in situ, without the need to isolate them from biopsies, could represent valuable therapeutic strategy. Along this line, it has been recently demonstrated that lipoaspirate product, Lipogems, contains and produces growth-factors that may activate resident stem cells. In this study, human tendon stem cells (hTSCs) from the rotator cuff were cocultured in a transwell system with the Lipogems lipoaspirate product and compared to control untreated cells in terms of cell proliferation, morphology, stem cell marker and VEGF expression, and differentiation and migration capabilities. Results showed that the Lipogems product significantly increases the proliferation rate of hTSCs without altering their stemness and differentiation capability. Moreover, treated cells increase the expression of VEGF, which is crucial for the neovascularization of the tissue during the healing process. Overall, this study supports that directly activating hTSCs with the Lipogems lipoaspirate could represent a new practical therapeutic approach. In fact, obtaining a lipoaspirate is easier, safer, and more cost-effective than harvesting cells from tendon or bone marrow biopsies, expanding them in GMP facility and then reinjecting them in the patient.
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Clonal analysis of human embryonic stem cell differentiation into teratomas.
Blum, Barak; Benvenisty, Nissim
2007-08-01
Differentiation of human embryonic stem cells (HESCs) can be studied in vivo through the induction of teratomas in immune-deficient mice. Cells within the teratomas differentiate into all three embryonic germ layers. However, the exact nature of the proliferation and differentiation of HESCs within the teratoma is not fully characterized, and it is not clear whether the differentiation is cell autonomous or affected by neighboring cells. Here, we establish a genetic approach to study the clonality of differentiation in teratomas using a mixture of HESC lines. We first demonstrate, by means of 5-bromo-2'-deoxyuridine incorporation, that cell proliferation occurs throughout the teratoma, and that there are no clusters of undifferentiated-proliferating cells. Using a combination of laser capture microdissection and DNA fingerprinting analysis, we show that different cell lines contribute mutually to the same distinctive tissue structures. Further support for the nonclonal differentiation within the teratoma was achieved by fluorescence in situ hybridization analysis of sex chromosomes. We therefore suggest that in vivo differentiation of HESCs is polyclonal and, thus, may not be cell autonomous, stressing the need for a three-dimensional growth in order to achieve complex differentiation of HESCs. Disclosure of potential conflicts of interest is found at the end of this article.
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Zhang, Yuanhao; Pan, Xiuwei; Shi, Zhen; Cai, Haibo; Gao, Yun; Zhang, Weian
2018-04-01
Stem cell factor (SCF) is considered as a commonly indispensable cytokine for proliferation of haematopoietic stem cells (HSCs), which is used in large dosages during ex vivo culture. The work presented here aimed to reduce the consumption of SCF by sustained release but still support cells proliferation and maintain the multipotency of HSCs. Stem cell factor was physically encapsulated within a hyaluronic acid/gelatin double network (HGDN) hydrogel to achieve a slow release rate. CD34 + cells were cultured within the SCF-loaded HGDN hydrogel for 14 days. The cell number, phenotype and functional capacity were investigated after culture. The HGDN hydrogels had desirable properties and encapsulated SCF kept being released for more than 6 days. SCF remained the native bioactivity, and the proliferation of HSCs within the SCF-loaded HGDN hydrogel was not affected, although the consumption of SCF was only a quarter in comparison with the conventional culture. Moreover, CD34 + cells harvested from the SCF-loaded HGDN hydrogels generated more multipotent colony-forming units (CFU-GEMM). The data suggested that the SCF-loaded HGDN hydrogel could support ex vivo culture of HSCs, thus providing a cost-effective culture protocol for HSCs. © 2017 John Wiley & Sons Ltd.
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Martínez-Cerdeño, Verónica; Camacho, Jasmin; Fox, Elizabeth; Miller, Elaine; Ariza, Jeanelle; Kienzle, Devon; Plank, Kaela; Noctor, Stephen C; Van de Water, Judy
2016-01-01
Autism spectrum disorders (ASDs) affect up to 1 in 68 children. Autism-specific autoantibodies directed against fetal brain proteins have been found exclusively in a subpopulation of mothers whose children were diagnosed with ASD or maternal autoantibody-related autism. We tested the impact of autoantibodies on brain development in mice by transferring human antigen-specific IgG directly into the cerebral ventricles of embryonic mice during cortical neurogenesis. We show that autoantibodies recognize radial glial cells during development. We also show that prenatal exposure to autism-specific maternal autoantibodies increased stem cell proliferation in the subventricular zone (SVZ) of the embryonic neocortex, increased adult brain size and weight, and increased the size of adult cortical neurons. We propose that prenatal exposure to autism-specific maternal autoantibodies directly affects radial glial cell development and presents a viable pathologic mechanism for the maternal autoantibody-related prenatal ASD risk factor. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.
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Yu, Ling; Shi, ZhuanZhuan; Gao, LiXia; Li, ChangMing
2015-09-01
In vitro cell-based analysis is strongly affected by material's surface chemical properties. The cell spreading, migration, and proliferation on a substrate surface are initiated and controlled by successful adhesion, particularly for anchor-dependent cells. Unfortunately, polydimethylsiloxane (PDMS), one of the most used polymeric materials for construction of microfluidic and miniaturized biomedical analytic devices, is not a cell-friendly surface because of its inherent hydrophobic property. Herein, a poly[glycidyl methacrylate-co-poly(ethylene glycol) methacrylate] (poly(GMA-co-pEGMA)) polymer brush was synthesized on a PDMS surface through a surface-initiated atom-transfer radical polymerization method. Contact angle and Fourier transform infrared characterization show that the poly (GMA-co-pEGMA) polymer brush functionalization can increase wettability of PDMS and introduce epoxy, hydroxyl, and ether groups into PDMS surface. In vitro cell growth assay demonstrates that cell adhesion and proliferation on poly(GMA-co-pEGMA) polymer brush-functionalized PDMS (poly(GMA-co-pEGMA)@PDMS) are better than on pristine PDMS. Additionally, immobilization of collagen type I (CI) and fibronectin (FN) on poly(GMA-co-pEGMA)@PDMS is better than direct coating of CI and FN on pristine PDMS to promote cell adhesion. Furthermore, increased intracellular reactive oxygen species and cell mitochondrial membrane depolarization, two indicators of cell oxidative stress, are observed from cells growing on pristine PDMS, but not from those on poly(GMA-co-pEGMA)@PDMS. Collectively, we demonstrate that poly(GMA-co-pEGMA) functionalization can enhance cell adhesion and proliferation on PDMS, and thus can be potentially used for microfluidic cell assay devices for cellular physiology study or drug screening. © 2015 Wiley Periodicals, Inc.
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Masaeli, Elahe; Morshed, Mohammad; Rasekhian, Parsa; Karbasi, Saeed; Karbalaie, Khadije; Karamali, Fereshte; Abedi, Daryoush; Razavi, Shahnaz; Jafarian-Dehkordi, Abbas; Nasr-Esfahani, Mohammad Hossein; Baharvand, Hossein
2012-07-01
A critical element in tissue engineering involves the fabrication of a three-dimensional scaffold. The scaffold provides a space for new tissue formation, supports cellular ingrowth, and proliferation and mimics many roles of the extracellular matrix. Poly(3-hydroxybutyrate) (PHB) is the most thoroughly investigated member of the polyhydroxyalkanoates (PHAs) family that has various degrees of biocompatibility and biodegradability for tissue engineering applications. In this study, we fabricated PHB scaffolds by utilizing electrospinning and salt-leaching procedures. The behavior of monkey epithelial kidney cells (Vero) and mouse mesenchymal stem cells (mMSCs) on these scaffolds was compared by the MTS assay and scanning electron microscopy. Additionally, this study investigated the mechanical and physical properties of these scaffolds by measuring tensile strength and modulus, dynamic contact angle and porosity. According to our results, the salt-leached scaffolds showed more wettability and permeability, but inferior mechanical properties when compared with nanofibrous scaffolds. In terms of cell response, salt-leached scaffolds showed enhanced Vero cell proliferation, whereas both scaffolds responded similarly in the case of mMSCs proliferation. In brief, nanofibrous scaffolds can be a better substrate for cell attachment and morphology. Copyright © 2012 Wiley Periodicals, Inc.
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Gregori, Josep; Méndez, Olga; Katsila, Theodora; Pujals, Mireia; Salvans, Cándida; Villarreal, Laura; Arribas, Joaquin; Tabernero, Josep; Sánchez, Alex; Villanueva, Josep
2014-07-15
Secretome profiling has become a methodology of choice for the identification of tumor biomarkers. We hypothesized that due to the dynamic nature of secretomes cellular perturbations could affect their composition but also change the global amount of protein secreted per cell. We confirmed our hypothesis by measuring the levels of secreted proteins taking into account the amount of proteome produced per cell. Then, we established a correlation between cell proliferation and protein secretion that explained the observed changes in global protein secretion. Next, we implemented a normalization correcting the statistical results of secretome studies by the global protein secretion of cells into a generalized linear model (GLM). The application of the normalization to two biological perturbations on tumor cells resulted in drastic changes in the list of statistically significant proteins. Furthermore, we found that known epithelial-to-mesenchymal transition (EMT) effectors were only statistically significant when the normalization was applied. Therefore, the normalization proposed here increases the sensitivity of statistical tests by increasing the number of true-positives. From an oncology perspective, the correlation between protein secretion and cellular proliferation suggests that slow-growing tumors could have high-protein secretion rates and consequently contribute strongly to tumor paracrine signaling.
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Xiong, Zekang; Lin, Hui; Zhao, Lei; Li, Zhiliang; Wang, Zhe; Peggrem, Shaun; Xia, Zhidao
2018-01-01
Background Link protein N-terminal peptide (LPP) in extracellular matrix (ECM) of cartilage could induce synthesis of proteoglycans and collagen type II in cartilaginous cells. Cartilage stem/progenitor cells (CSPCs), the endogenous stem cells in cartilage, are important in cartilage degeneration and regeneration. We hypothesized that LPP could be a stimulator for stem cell-based cartilage regeneration by affecting biological behaviors of CSPC. Methods CSPCs were isolated from rat knee cartilage. We evaluated the promoting effect of LPP on proliferation, migration, and chondrogenic differentiation of CSPCs. The chondrogenic differentiation-related genes and proteins were quantitated. Three-dimensional culture of CSPC was conducted in the presence of TGF-β3 or LPP, and the harvested pellets were analyzed to assess the function of LPP on cartilage regeneration. Results LPP stimulated the proliferation of CSPC and accelerated the site-directional migration. Higher expression of SOX9, collagen II, and aggrecan were demonstrated in CSPCs treated with LPP. The pellets treated with LPP showed more distinct characteristics of chondroid differentiation than those with TGF-β3. Conclusion LPP showed application prospect in cartilage regeneration medicine by stimulating proliferation, migration, and chondrogenic differentiation of cartilage stem/progenitor cells. PMID:29531532
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Ginani, Fernanda; Soares, Diego Moura; de Oliveira Rocha, Hugo Alexandre; de Souza, Lélia Batista; Barboza, Carlos Augusto Galvão
2018-01-01
The aim of this study was to evaluate the effect of low-level laser irradiation (LLLI) on the proliferation and viability of stem cells from human exfoliated deciduous teeth (SHED). Cells were irradiated or not (control) with an InGaAlP laser diode (660 nm, 30 mW, continuous action mode) using two different energy densities (0.5 J/cm 2 -16 s; 1.0 J/cm 2 -33 s). Irradiation was performed at 0 and 48 h, with the laser probe fixed at a distance of 0.5 cm from the cells. Cell proliferation was analyzed at 0, 24, 48, and 72 h by the Trypan blue exclusion method and MTT assay. Cell cycle and Ki67 expression were analyzed by flow cytometry. Apoptosis-related events were evaluated by expression of annexin V/PI and nuclear morphological changes by staining with DAPI. Differences between groups at each time were analyzed by the Kruskal-Wallis and Mann-Whitney tests, adopting a level of significance of 5% (p < 0.05). The results showed that an energy density of 1.0 J/cm 2 promoted an increase in cell proliferation at 48 and 72 h compared to the control and 0.5 J/cm 2 groups. Cell cycle analysis revealed a predominance of cells in the S and G2/M phases in the irradiated groups. This finding was confirmed by the increased expression of Ki67. Low positive staining for annexin V and PI was observed in all groups, and no nuclear changes were detected, indicating that cell viability was not affected by the energy densities tested. It can be concluded that the LLLI parameters used (660 nm, 30 mW, 1.0 J/cm 2 ) promote the proliferation of SHEDs and the maintenance of cell viability.
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Slattery, Eric L; Speck, Judith D; Warchol, Mark E
2009-09-01
The sensory hair cells of the cochlea and vestibular organs are essential for normal hearing and balance function. The mammalian ear possesses a very limited ability to regenerate hair cells and their loss can lead to permanent sensory impairment. In contrast, hair cells in the avian ear are quickly regenerated after acoustic trauma or ototoxic injury. The very different regenerative abilities of the avian vs. mammalian ear can be attributed to differences in injury-evoked expression of genes that either promote or inhibit the production of new hair cells. Gene expression is regulated both by the binding of cis-regulatory molecules to promoter regions as well as through structural modifications of chromatin (e.g., methylation and acetylation). This study examined effects of histone deacetylases (HDACs), whose main function is to modify histone acetylation, on the regulation of regenerative proliferation in the chick utricle. Cultures of regenerating utricles and dissociated cells from the utricular sensory epithelia were treated with the HDAC inhibitors valproic acid, trichostatin A, sodium butyrate, and MS-275. All of these molecules prevent the enzymatic removal of acetyl groups from histones, thus maintaining nuclear chromatin in a "relaxed" (open) configuration. Treatment with all inhibitors resulted in comparable decreases in supporting cell proliferation. We also observed that treatment with the HDAC1-, 2-, and 3-specific inhibitor MS-275 was sufficient to reduce proliferation and that two class I HDACs--HDAC1 and HDAC2--were expressed in the sensory epithelium of the utricle. These results suggest that inhibition of specific type I HDACs is sufficient to prevent cell cycle entry in supporting cells. Notably, treatment with HDAC inhibitors did not affect the differentiation of replacement hair cells. We conclude that histone deacetylation is a positive regulator of regenerative proliferation but is not critical for avian hair cell differentiation.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Kim, Yong Il; Ryu, Jae-Sung; Yeo, Jee Eun
2014-08-08
Highlights: • Continuous TGF-β1 overexpression in hSD-MSCs did not influence their phenotypes. • Retroviral-mediated transduction of TGFB1 in hSD-MSCs enhances cell proliferation. • TGF-β1 overexpression did not effect to adipo- or osteogenic potential of hSD-MSCs. • TGF-β1 overexpression in hSD-MSCs could stimulate and accelerate chondrogenesis. - Abstract: Transforming growth factor-beta (TGF-β) superfamily proteins play a critical role in proliferation, differentiation, and other functions of mesenchymal stem cells (MSCs). During chondrogenic differentiation of MSCs, TGF-β up-regulates chondrogenic gene expression by enhancing the expression of the transcription factor SRY (sex-determining region Y)-box9 (Sox9). In this study, we investigated the effect of continuousmore » TGF-β1 overexpression in human synovium-derived MSCs (hSD-MSCs) on immunophenotype, differentiation potential, and proliferation rate. hSD-MSCs were transduced with recombinant retroviruses (rRV) encoding TGF-β1. The results revealed that continuous overexpression of TGF-β1 did not affect their phenotype as evidenced by flow cytometry and reverse transcriptase PCR (RT-PCR). In addition, continuous TGF-β1 overexpression strongly enhanced cell proliferation of hSD-MSCs compared to the control groups. Also, induction of chondrogenesis was more effective in rRV-TGFB-transduced hSD-MSCs as shown by RT-PCR for chondrogenic markers, toluidine blue staining and glycosaminoglycan (GAG)/DNA ratio. Our data suggest that overexpression of TGF-β1 positively enhances the proliferation and chondrogenic potential of hSD-MSCs.« less
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ESR1 inhibits hCG-induced steroidogenesis and proliferation of progenitor Leydig cells in mice.
Oh, Yeong Seok; Koh, Il Kyoo; Choi, Bomi; Gye, Myung Chan
2017-03-07
Oestrogen is an important regulator in reproduction. To understand the role of oestrogen receptor 1 (ESR1) in Leydig cells, we investigated the expression of ESR1 in mouse Leydig cells during postnatal development and the effects of oestrogen on steroidogenesis and proliferation of progenitor Leydig cells (PLCs). In Leydig cells, the ESR1 expression was low at birth, increased until postnatal day 14 at which PLCs were predominant, and then decreased until adulthood. In foetal Leydig cells, ESR1 immunoreactivity increased from birth to postnatal day 14. These suggest that ESR1 is a potential biomarker of Leydig cell development. In PLCs, 17β-estradiol and the ESR1-selective agonist propylpyrazoletriol suppressed human chorionic gonadotropin (hCG)-induced progesterone production and steroidogenic gene expression. The ESR2-selective agonist diarylpropionitrile did not affect steroidogenesis. In PLCs from Esr1 knockout mice, hCG-stimulated steroidogenesis was not suppressed by 17β-estradiol, suggesting that oestrogen inhibits PLC steroidogenesis via ESR1. 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile decreased bromodeoxyuridine uptake in PLCs in the neonatal mice. In cultured PLCs, 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile reduced hCG-stimulated Ki67 and Pcna mRNA expression and the number of KI67-positive PLCs, suggesting that oestrogen inhibits PLC proliferation via both ESR1 and ESR2. In PLCs, ESR1 mediates the oestrogen-induced negative regulation of steroidogenesis and proliferation.
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ESR1 inhibits hCG-induced steroidogenesis and proliferation of progenitor Leydig cells in mice
Oh, Yeong Seok; Koh, Il Kyoo; Choi, Bomi; Gye, Myung Chan
2017-01-01
Oestrogen is an important regulator in reproduction. To understand the role of oestrogen receptor 1 (ESR1) in Leydig cells, we investigated the expression of ESR1 in mouse Leydig cells during postnatal development and the effects of oestrogen on steroidogenesis and proliferation of progenitor Leydig cells (PLCs). In Leydig cells, the ESR1 expression was low at birth, increased until postnatal day 14 at which PLCs were predominant, and then decreased until adulthood. In foetal Leydig cells, ESR1 immunoreactivity increased from birth to postnatal day 14. These suggest that ESR1 is a potential biomarker of Leydig cell development. In PLCs, 17β-estradiol and the ESR1-selective agonist propylpyrazoletriol suppressed human chorionic gonadotropin (hCG)-induced progesterone production and steroidogenic gene expression. The ESR2-selective agonist diarylpropionitrile did not affect steroidogenesis. In PLCs from Esr1 knockout mice, hCG-stimulated steroidogenesis was not suppressed by 17β-estradiol, suggesting that oestrogen inhibits PLC steroidogenesis via ESR1. 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile decreased bromodeoxyuridine uptake in PLCs in the neonatal mice. In cultured PLCs, 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile reduced hCG-stimulated Ki67 and Pcna mRNA expression and the number of KI67-positive PLCs, suggesting that oestrogen inhibits PLC proliferation via both ESR1 and ESR2. In PLCs, ESR1 mediates the oestrogen-induced negative regulation of steroidogenesis and proliferation. PMID:28266530
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Boku, Shuken, E-mail: shuboku@med.hokudai.ac.jp; Nakagawa, Shin; Takamura, Naoki
2013-05-17
Highlights: •GDNF has no effect on ADP proliferation and apoptosis. •GDNF increases ADP differentiation into astrocyte. •A specific inhibitor of STAT3 decreases the astrogliogenic effect of GDNF. •STAT3 knockdown by lentiviral shRNA vector also decreases the astrogliogenic effect of GDNF. •GDNF increases the phosphorylation of STAT3. -- Abstract: While the pro-neurogenic actions of antidepressants in the adult hippocampal dentate gyrus (DG) are thought to be one of the mechanisms through which antidepressants exert their therapeutic actions, antidepressants do not increase proliferation of neural precursor cells derived from the adult DG. Because previous studies showed that antidepressants increase the expression andmore » secretion of glial cell line-derived neurotrophic factor (GDNF) in C6 glioma cells derived from rat astrocytes and GDNF increases neurogenesis in adult DG in vivo, we investigated the effects of GDNF on the proliferation, differentiation and apoptosis of cultured neural precursor cells derived from the adult DG. Data showed that GDNF facilitated the differentiation of neural precursor cells into astrocytes but had no effect on their proliferation or apoptosis. Moreover, GDNF increased the phosphorylation of STAT3, and both a specific inhibitor of STAT3 and lentiviral shRNA for STAT3 decreased their differentiation into astrocytes. Taken together, our findings suggest that GDNF facilitates astrogliogenesis from neural precursor cells in adult DG through activating STAT3 and that this action might indirectly affect neurogenesis.« less
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Koning, Merel; Werker, Paul M N; van Luyn, Marja J A; Harmsen, Martin C
2011-07-01
Facial paralysis is a physically, psychologically, and socially disabling condition. Innovative treatment strategies based on regenerative medicine, in particular tissue engineering of skeletal muscle, are promising for treatment of patients with facial paralysis. The natural source for tissue-engineered muscle would be muscle stem cells, that is, human satellite cells (SC). In vivo, SC respond to hypoxic, ischemic muscle damage by activation, proliferation, differentiation to myotubes, and maturation to muscle fibers, while maintaining their reserve pool of SC. Therefore, our hypothesis is that hypoxia improves proliferation and differentiation of SC. During tissue engineering, a three-dimensional construct, or implanting SC in vivo, SC will encounter hypoxic environments. Thus, we set out to test our hypothesis on SC in vitro. During the first five passages, hypoxically cultured SC proliferated faster than their counterparts under normoxia. Moreover, also at higher passages, a switch from normoxia to hypoxia enhanced proliferation of SC. Hypoxia did not affect the expression of SC markers desmin and NCAM. However, the average surface expression per cell of NCAM was downregulated by hypoxia, and it also downregulated the gene expression of NCAM. The gene expression of the myogenic transcription factors PAX7, MYF5, and MYOD was upregulated by hypoxia. Moreover, gene expression of structural proteins α-sarcomeric actin, and myosins MYL1 and MYL3 was upregulated by hypoxia during differentiation. This indicates that hypoxia promotes a promyogenic shift in SC. Finally, Pax7 expression was not influenced by hypoxia and maintained in a subset of mononucleated cells, whereas these cells were devoid of structural muscle proteins. This suggests that during myogenesis in vitro, at least part of the SC adopt a quiescent, that is, reserve cells, phenotype. In conclusion, tissue engineering under hypoxic conditions would seem favorable in terms of myogenic proliferation, while maintaining the quiescent SC pool.
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Manavski, Yosif; Abel, Tobias; Hu, Junhao; Kleinlützum, Dina; Buchholz, Christian J.; Belz, Christina; Augustin, Hellmut G.; Dimmeler, Stefanie
2017-01-01
Endothelial cells (ECs) not only are important for oxygen delivery but also act as a paracrine source for signals that determine the balance between tissue regeneration and fibrosis. Here we show that genetic inactivation of flow-induced transcription factor Krüppel-like factor 2 (KLF2) in ECs results in reduced liver damage and augmentation of hepatocyte proliferation after chronic liver injury by treatment with carbon tetrachloride (CCl4). Serum levels of GLDH3 and ALT were significantly reduced in CCl4-treated EC-specific KLF2-deficient mice. In contrast, transgenic overexpression of KLF2 in liver sinusoidal ECs reduced hepatocyte proliferation. KLF2 induced activin A expression and secretion from endothelial cells in vitro and in vivo, which inhibited hepatocyte proliferation. However, loss or gain of KLF2 expression did not change capillary density and liver fibrosis, but significantly affected hepatocyte proliferation. Taken together, the data demonstrate that KLF2 induces an antiproliferative secretome, including activin A, which attenuates liver regeneration. PMID:28348240
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Su, Jingjing; Zhou, Houguang; Tao, Yinghong; Guo, Zhuangli; Zhang, Shuo; Zhang, Yu; Huang, Yanyan; Tang, Yuping; Hu, Renming; Dong, Qiang
2015-01-01
Cell cycle processes play a vital role in vascular endothelial proliferation and dysfunction. Cell division cycle protein 14 (Cdc14) is an important cell cycle regulatory phosphatase. Previous studies in budding yeast demonstrated that Cdc14 could trigger the inactivation of mitotic cyclin-dependent kinases (Cdks), which are required for mitotic exit and cytokinesis. However, the exact function of human Cdc14 (hCdc14) in cell cycle regulation during vascular diseases is yet to be elucidated. There are two HCdc14 homologs: hCdc14A and hCdc14B. In the current study, we investigated the potential role of hCdc14A in high glucose-, free fatty acids (FFAs)-, and hypoxia-induced injury in cultured human brain vascular endothelial cells (HBVECs). Data revealed that high glucose, FFA, and hypoxia down-regulated hCdc14A expression remarkably, and also affected the expression of other cell cycle-related proteins such as cyclin B, cyclin D, cyclin E, and p53. Furthermore, the combined addition of the three stimuli largely blocked cell cycle progression, decreased cell proliferation, and increased apoptosis. We also determined that hCdc14A was localized mainly to centrosomes during interphase and spindles during mitosis using confocal microscopy, and that it could affect the expression of other cycle-related proteins. More importantly, the overexpression of hCdc14A accelerated cell cycle progression, enhanced cell proliferation, and promoted neoplastic transformation, whereas the knockdown of hCdc14A using small interfering RNA produced the opposite effects. Therefore, these findings provide novel evidence that hCdc14A might be involved in cell cycle regulation in cultured HBVECs during high glucose-, FFA-, and hypoxia-induced injury. Copyright © 2014 Elsevier Inc. All rights reserved.
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Kinetics of liver macrophages (Kupffer cells) in SIV-infected macaques.
Ahsan, Muhammad H; Gill, Amy F; Alvarez, Xavier; Lackner, Andrew A; Veazey, Ronald S
2013-11-01
Since the liver drains antigens from the intestinal tract, and since the intestinal tract is a major site of viral replication, we examined the dynamics of liver macrophages (Kupffer cells) throughout SIV infection. Absolute numbers of Kupffer cells increased in the livers in acute infection, and in animals with AIDS. Significantly higher percentages of proliferating (BrdU+) Kupffer cells were detected in acute infection and in AIDS with similar trends in blood monocytes. Significantly higher percentages of apoptotic (AC3+) Kupffer cells were also found in acute and AIDS stages. However, productively infected cells were not detected in liver of 41/42 animals examined, despite abundant infected cells in gut and lymph nodes of all animals. Increased rates of Kupffer cell proliferation resulting in an increase in Kupffer cells without productive infection indicate SIV infection affects Kupffer cells, but the liver does not appear to be a major site of productive viral replication. © 2013 Elsevier Inc. All rights reserved.
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Szczepny, Anette; Hogarth, Cathryn A; Young, Julia; Loveland, Kate L
2009-02-01
The Hedgehog (Hh) signaling pathway affects fetal testis growth. Recently, we described the dynamic cellular production of Hh signaling pathway components in juvenile and adult rodent testes. The Hh signaling is understood to regulate cord formation in the fetal testis, but minimal knowledge exists regarding how Hh signaling impacts the postnatal testis. To investigate this, we employed hanging drop cultures, which are used routinely in embryoid body formation. This approach has the advantage of using small media volume, and we examined its suitability for short-term culture of both murine embryonic gonads and adult testis tubules. The effects of cyclopamine, a specific Hh signaling inhibitor, were examined following culture of Embryonic Day 11.5 urogenital ridges (as control) and adult seminiferous tubule fragments for 24-48 h using histological, cell proliferation, and gene expression analyses. Cultured embryonic testes displayed generally normal cord structure, anti-Müllerian hormone (Amh) expression, and cell proliferation; known Hh target gene expression (Gli1, osteopontin, official symbol Spp1, and Amh) was altered in response to cyclopamine. Cultured adult tubules exhibited some loss of seminiferous epithelium organization over 48 h. Spermatogonia continued to proliferate, however, and no significant loss of viability was noted overall. Addition of cyclopamine significantly affected levels of Gli1, Igfbp6, Ccnd2 (cyclin D2), Ccnb1 (cyclin B1), Spp1, Kit, and Amh mRNAs; these genes have been shown previously to be expressed in Sertoli and germ cells. These novel results identify Hh target genes in the testis and demonstrate this signaling pathway likely affects cell survival and differentiation in the context of normal adult testis.
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Szczepny, Anette; Hogarth, Cathryn A.; Young, Julia; Loveland, Kate L.
2008-01-01
The Hedgehog (Hh) signaling pathway affects fetal testis growth. Recently, we described the dynamic cellular production of Hh signaling pathway components in juvenile and adult rodent testes. The Hh signaling is understood to regulate cord formation in the fetal testis, but minimal knowledge exists regarding how Hh signaling impacts the postnatal testis. To investigate this, we employed hanging drop cultures, which are used routinely in embryoid body formation. This approach has the advantage of using small media volume, and we examined its suitability for short-term culture of both murine embryonic gonads and adult testis tubules. The effects of cyclopamine, a specific Hh signaling inhibitor, were examined following culture of Embryonic Day 11.5 urogenital ridges (as control) and adult seminiferous tubule fragments for 24–48 h using histological, cell proliferation, and gene expression analyses. Cultured embryonic testes displayed generally normal cord structure, anti-Müllerian hormone (Amh) expression, and cell proliferation; known Hh target gene expression (Gli1, osteopontin, official symbol Spp1, and Amh) was altered in response to cyclopamine. Cultured adult tubules exhibited some loss of seminiferous epithelium organization over 48 h. Spermatogonia continued to proliferate, however, and no significant loss of viability was noted overall. Addition of cyclopamine significantly affected levels of Gli1, Igfbp6, Ccnd2 (cyclin D2), Ccnb1 (cyclin B1), Spp1, Kit, and Amh mRNAs; these genes have been shown previously to be expressed in Sertoli and germ cells. These novel results identify Hh target genes in the testis and demonstrate this signaling pathway likely affects cell survival and differentiation in the context of normal adult testis. PMID:18843087
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Kay, Alasdair G; Long, Grace; Tyler, George; Stefan, Andrei; Broadfoot, Stephen J; Piccinini, Anna M; Middleton, Jim; Kehoe, Oksana
2017-12-21
We evaluated the therapeutic potential of mesenchymal stem cell-conditioned medium (CM-MSC) as an alternative to cell therapy in an antigen-induced model of arthritis (AIA). Disease severity and cartilage loss were evaluated by histopathological analysis of arthritic knee joints and immunostaining of aggrecan neoepitopes. Cell proliferation was assessed for activated and naïve CD4+ T cells from healthy mice following culture with CM-MSC or co-culture with MSCs. T cell polarization was analysed in CD4+ T cells isolated from spleens and lymph nodes of arthritic mice treated with CM-MSC or MSCs. CM-MSC treatment significantly reduced knee-joint swelling, histopathological signs of AIA, cartilage loss and suppressed TNFα induction. Proliferation of CD4+ cells from spleens of healthy mice was not affected by CM-MSC but reduced when cells were co-cultured with MSCs. In the presence of CM-MSC or MSCs, increases in IL-10 concentration were observed in culture medium. Finally, CD4+ T cells from arthritic mice treated with CM-MSC showed increases in FOXP3 and IL-4 expression and positively affected the Treg:Th17 balance in the tissue. CM-MSC treatment reduces cartilage damage and suppresses immune responses by reducing aggrecan cleavage, enhancing Treg function and adjusting the Treg:Th17 ratio. CM-MSC may provide an effective cell-free therapy for inflammatory arthritis.
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Rauner, Gat; Barash, Itamar
2014-10-15
The challenge in manipulating the proportion of somatic stem cells lies in having to override tissue homeostasis. Xanthosine infusion via the teat canal has been reported to augment the number of label-retaining cells in the mammary gland of 3-month-old bovine calves. To further delineate xanthosine׳s effect on defined stem cells in the mammary gland of heifers-which are candidates for increased prospective milk production following such manipulation-bovine mammary parenchymal tissue was transplanted and integrated into the cleared mammary fat pad of immunodeficient mice. Xanthosine administration for 14 days did not affect the number of label-retaining cells after 10- and 11-week chases. No change in stem cell proportion, analyzed according to CD49f and CD24 expression, was noted. Clone formation and propagation rate of cultured cells, as well as expression of stem cell markers, were also unaffected. In contrast, a latent 50% decrease in bovine mammary cell proliferation rate was observed 11 weeks after xanthosine administration. Tumor development in mice was also limited by xanthosine administration. These effects may have resulted from an initial decrease in expression of the rate-limiting enzyme in guanine synthesis, IMPDH. The data indicate that caution should be exerted when considering xanthosine for stem cell manipulation. Copyright © 2014 Elsevier Inc. All rights reserved.
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Cho, Sun-Mi; Lee, Eun-Ok; Kim, Sung-Hoon; Lee, Hyo-Jeong
2014-07-30
The essential oil of Pinus koraiensis (EOPK) is biologically active compound obtained from the leaves of P. koraiensis. The goal of this study was to investigate the anti-cancer mechanism of EOPK in HCT116 colorectal cancer cells. HCT116 cell proliferation was assessed by conducting crystal violet and BrdU assays. To assess the effects of EOPK on cell migration, we performed a wound-healing assay. Further, the contribution of PAK1 to EOPK-induced AKT and extracellular signal-regulated kinase (ERK) suppression was assessed by siRNA-mediated PAK1 knockdown. Changes to the expression and phosphorylation of PAK1 and its effectors were determined by western blotting, and changes to the actin cytoskeleton were determined by performing an immunofluorescence assay. EOPK significantly decreased HCT116 cell proliferation and migration, and induced G1 arrest without affecting normal cells. Additionally, EOPK suppressed the expression of PAK1, and decreased ERK and AKT phosphorylation in HCT116 cells. Finally, EOPK suppressed β-catenin, cyclin D1, and CDK4/6 expression. Our studies indicate that EOPK significantly reduced proliferation and migration of colorectal cancer cells. Furthermore, EOPK suppressed PAK1 expression in a dose-dependent manner, and this suppression of PAK1 led to inhibition of ERK, AKT, and β-catenin activities. Our findings suggest that EOPK exerts its anticancer activity via the inhibition of PAK1 expression, suggesting it may be a potent chemotherapeutic agent for colorectal cancer.
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Interferon-γ Reduces the Proliferation of Primed Human Renal Tubular Cells.
García-Sánchez, Omar; López-Novoa, José Miguel; López-Hernández, Francisco J
2014-01-01
Chronic kidney disease (CKD) is a progressive deterioration of the kidney function, which may eventually lead to renal failure and the need for dialysis or kidney transplant. Whether initiated in the glomeruli or the tubuli, CKD is characterized by progressive nephron loss, for which the process of tubular deletion is of key importance. Tubular deletion results from tubular epithelial cell death and defective repair, leading to scarring of the renal parenchyma. Several cytokines and signaling pathways, including transforming growth factor-β (TGF-β) and the Fas pathway, have been shown to participate in vivo in tubular cell death. However, there is some controversy about their mode of action, since a direct effect on normal tubular cells has not been demonstrated. We hypothesized that epithelial cells would require specific priming to become sensitive to TGF-β or Fas stimulation and that this priming would be brought about by specific mediators found in the pathological scenario. Herein we studied whether the combined effect of several stimuli known to take part in CKD progression, namely TGF-β, tumor necrosis factor-α, interferon-γ (IFN-γ), and Fas stimulation, on primed resistant human tubular cells caused cell death or reduced proliferation. We demonstrate that these cytokines have no synergistic effect on the proliferation or viability of human kidney (HK2) cells. We also demonstrate that IFN-γ, but not the other stimuli, reduces the proliferation of cycloheximide-primed HK2 cells without affecting their viability. Our results point at a potentially important role of IFN-γ in defective repair, leading to nephron loss during CKD.
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Interferon-γ Reduces the Proliferation of Primed Human Renal Tubular Cells
García-Sánchez, Omar; López-Novoa, José Miguel; López-Hernández, Francisco J.
2014-01-01
Background/Aims Chronic kidney disease (CKD) is a progressive deterioration of the kidney function, which may eventually lead to renal failure and the need for dialysis or kidney transplant. Whether initiated in the glomeruli or the tubuli, CKD is characterized by progressive nephron loss, for which the process of tubular deletion is of key importance. Tubular deletion results from tubular epithelial cell death and defective repair, leading to scarring of the renal parenchyma. Several cytokines and signaling pathways, including transforming growth factor-β (TGF-β) and the Fas pathway, have been shown to participate in vivo in tubular cell death. However, there is some controversy about their mode of action, since a direct effect on normal tubular cells has not been demonstrated. We hypothesized that epithelial cells would require specific priming to become sensitive to TGF-β or Fas stimulation and that this priming would be brought about by specific mediators found in the pathological scenario. Methods Herein we studied whether the combined effect of several stimuli known to take part in CKD progression, namely TGF-β, tumor necrosis factor-α, interferon-γ (IFN-γ), and Fas stimulation, on primed resistant human tubular cells caused cell death or reduced proliferation. Results We demonstrate that these cytokines have no synergistic effect on the proliferation or viability of human kidney (HK2) cells. We also demonstrate that IFN-γ, but not the other stimuli, reduces the proliferation of cycloheximide-primed HK2 cells without affecting their viability. Conclusion Our results point at a potentially important role of IFN-γ in defective repair, leading to nephron loss during CKD. PMID:24575118
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Zhu, DanHong; Sreekumar, Parameswaran G; Hinton, David R; Kannan, Ram
2010-03-31
Ceramide and its metabolic derivatives are important modulators of cellular apoptosis and proliferation. Dysregulation or imbalance of their metabolic pathways may promote the development of retinal degeneration. The aim of this study was to identify the expression and regulation of key enzymes of the ceramide pathway in retinal pigment epithelial (RPE) cells. RT-PCR was used to screen the enzymes involved in ceramide metabolism that are expressed in RPE. Over-expression of neutral sphingomyelinase-2 (SMPD3) or sphingosine kinase 1 (Sphk1) in ARPE-19 cells was achieved by transient transfection of SMPD3 or Sphk1 cDNA subcloned into an expression vector. The number of apoptotic or proliferating cells was determined using TUNEL and BrdU assays, respectively. Neutral sphingomyelinase-1, neutral sphingomyelinase-2, acidic ceramidase, ceramide kinase, SphK1 and Sphk2 were expressed in both ARPE-19 and early passage human fetal RPE (fRPE) cells, while alkaline ceramidase 2 was only expressed in fRPE cells. Over-expression of SMPD3 decreased RPE cell proliferation and increased cell apoptosis. The percentage of apoptotic cells increased proportionally with the amount of transfected SMPD3 DNA. Over-expression of SphK1 promoted cell proliferation and protected ARPE-19 cells from ceramide-induced apoptosis. The effect of C(2) ceramide on induction of apoptosis was evaluated in polarized vs. non-polarized RPE cultures; polarization of RPE was associated with much reduced apoptosis in response to ceramide. In conclusion, RPE cells possess the synthetic machinery for the production of ceramide, sphingosine, ceramide-1-phosphate (C1P), and sphingosine-1-phosphate (S1P). Over-expression of SMPD3 may increase cellular ceramide levels, leading to enhanced cell death and arrested cell proliferation. The selective induction of apoptosis in non-polarized RPE cultures by C(2) ceramide suggests that increased ceramide levels will preferentially affect non-polarized RPE, as are found in late age-related macular degeneration lesions, and may spare the normal RPE monolayer. SphK1 over-expression increased cellular S1P, which promoted cell proliferation and protected RPE from ceramide-induced apoptosis. Understanding the relationship between the metabolism of sphingolipids and their effects in RPE cell survival/death may help us to develop effective and efficient therapies for retinal degeneration. Copyright 2009 Elsevier Ltd. All rights reserved.
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Charif, N; Li, Y Y; Targa, L; Zhang, L; Ye, J S; Li, Y P; Stoltz, J F; Han, H Z; de Isla, N
2017-01-01
With their proliferation, differentiation into specific cell types, and secretion properties, mesenchymal stromal/stem cells (MSC) are very interesting tools to be used in regenerative medicine. Bone marrow (BM) was the first MSC source characterized. In the frame of autologous MSC therapy, it is important to detect donor's parameters affecting MSC potency. Age of the donors appears as one parameter that could greatly affect MSC properties. Moreover, in vitro cell expansion is needed to obtain the number of cells necessary for clinical developments. It will lead to in vitro cell aging that could modify cell properties. This review recapitulates several studies evaluating the effect of in vitro and in vivo MSC aging on cell properties.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Eke, Iris; Storch, Katja; Kaestner, Ina
Purpose: Cell invasion represents one of the major determinants that treatment has failed for patients suffering from glioblastoma. Contrary findings have been reported for cell migration upon exposure to ionizing radiation. Here, the migration and invasion capability of glioblastoma cells on and in collagen type I were evaluated upon irradiation with X-rays or carbon ions. Methods and Materials: Migration on and invasion in collagen type I were evaluated in four established human glioblastoma cell lines exposed to either X-rays or carbon ions. Furthermore, clonogenic radiation survival, proliferation (5-bromo-2-deoxyuridine positivity), DNA double-strand breaks ({gamma}H2AX/53BP1-positive foci), and expression of invasion-relevant proteins (eg,more » {beta}1 integrin, FAK, MMP2, and MMP9) were explored. Migration and invasion assays for primary glioblastoma cells also were carried out with X-ray irradiation. Results: Neither X-ray nor carbon ion irradiation affected glioblastoma cell migration and invasion, a finding similarly observed in primary glioblastoma cells. Intriguingly, irradiated cells migrated unhampered, despite DNA double-strand breaks and reduced proliferation. Clonogenic radiation survival was increased when cells had contact with extracellular matrix. Specific inhibition of the {beta}1 integrin or proliferation-associated signaling molecules revealed a critical function of JNK, PI3K, and p38 MAPK in glioblastoma cell invasion. Conclusions: These findings indicate that X-rays and carbon ion irradiation effectively reduce proliferation and clonogenic survival without modifying the migration and invasion ability of glioblastoma cells in a collagen type I environment. Addition of targeted agents against members of the MAPK and PI3K signaling axis to conventional chemoradiation therapy seems potentially useful to optimize glioblastoma therapy.« less
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Bi, H; Li, S; Qu, X; Wang, M; Bai, X; Xu, Z; Ao, X; Jia, Z; Jiang, X; Yang, Y; Wu, H
2015-01-01
Breast cancer that is accompanied by a high level of cyclin E expression usually exhibits poor prognosis and clinical outcome. Several factors are known to regulate the level of cyclin E during the cell cycle progression. The transcription factor DEC1 (also known as STRA13 and SHARP2) plays an important role in cell proliferation and apoptosis. Nevertheless, the mechanism of its role in cell proliferation is poorly understood. In this study, using the breast cancer cell lines MCF-7 and T47D, we showed that DEC1 could inhibit the cell cycle progression of breast cancer cells independently of its transcriptional activity. The cell cycle-dependent timing of DEC1 overexpression could affect the progression of the cell cycle through regulating the level of cyclin E protein. DEC1 stabilized cyclin E at the protein level by interacting with cyclin E. Overexpression of DEC1 repressed the interaction between cyclin E and its E3 ligase Fbw7α, consequently reducing the level of polyunbiquitinated cyclin E and increased the accumulation of non-ubiquitinated cyclin E. Furthermore, DEC1 also promoted the nuclear accumulation of Cdk2 and the formation of cyclin E/Cdk2 complex, as well as upregulating the activity of the cyclin E/Cdk2 complex, which inhibited the subsequent association of cyclin A with Cdk2. This had the effect of prolonging the S phase and suppressing the growth of breast cancers in a mouse xenograft model. These events probably constitute the essential steps in DEC1-regulated cell proliferation, thus opening up the possibility of a protein-based molecular strategy for eliminating cancer cells that manifest a high-level expression of cyclin E. PMID:26402517
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NASA Astrophysics Data System (ADS)
Picard, Frédéric; Wanatabe, Mitsuhiro; Schoonjans, Kristina; Lydon, John; O'Malley, Bert W.; Auwerx, Johan
2002-11-01
Gestational diabetes coincides with elevated circulating progesterone levels. We show that progesterone accelerates the progression of diabetes in female db/db mice. In contrast, RU486, an antagonist of the progesterone receptor (PR), reduces blood glucose levels in both female WT and db/db mice. Furthermore, female, but not male, PR-/- mice had lower fasting glycemia than PR+/+ mice and showed higher insulin levels on glucose injection. Pancreatic islets from female PR-/- mice were larger and secreted more insulin consequent to an increase in -cell mass due to an increase in -cell proliferation. These findings demonstrate an important role of progesterone signaling in insulin release and pancreatic function and suggest that it affects the susceptibility to diabetes.
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Tonic B-cell receptor signaling in diffuse large B-cell lymphoma.
Havranek, Ondrej; Xu, Jingda; Köhrer, Stefan; Wang, Zhiqiang; Becker, Lisa; Comer, Justin M; Henderson, Jared; Ma, Wencai; Man Chun Ma, John; Westin, Jason R; Ghosh, Dipanjan; Shinners, Nicholas; Sun, Luhong; Yi, Allen F; Karri, Anusha R; Burger, Jan A; Zal, Tomasz; Davis, R Eric
2017-08-24
We used clustered regularly interspaced short palindromic repeats/Cas9-mediated genomic modification to investigate B-cell receptor (BCR) signaling in cell lines of diffuse large B-cell lymphoma (DLBCL). Three manipulations that altered BCR genes without affecting surface BCR levels showed that BCR signaling differs between the germinal center B-cell (GCB) subtype, which is insensitive to Bruton tyrosine kinase inhibition by ibrutinib, and the activated B-cell (ABC) subtype. Replacing antigen-binding BCR regions had no effect on BCR signaling in GCB-DLBCL lines, reflecting this subtype's exclusive use of tonic BCR signaling. Conversely, Y188F mutation in the immunoreceptor tyrosine-based activation motif of CD79A inhibited tonic BCR signaling in GCB-DLBCL lines but did not affect their calcium flux after BCR cross-linking or the proliferation of otherwise-unmodified ABC-DLBCL lines. CD79A-GFP fusion showed BCR clustering or diffuse distribution, respectively, in lines of ABC and GCB subtypes. Tonic BCR signaling acts principally to activate AKT, and forced activation of AKT rescued GCB-DLBCL lines from knockout (KO) of the BCR or 2 mediators of tonic BCR signaling, SYK and CD19. The magnitude and importance of tonic BCR signaling to proliferation and size of GCB-DLBCL lines, shown by the effect of BCR KO, was highly variable; in contrast, pan-AKT KO was uniformly toxic. This discrepancy was explained by finding that BCR KO-induced changes in AKT activity (measured by gene expression, CXCR4 level, and a fluorescent reporter) correlated with changes in proliferation and with baseline BCR surface density. PTEN protein expression and BCR surface density may influence clinical response to therapeutic inhibition of tonic BCR signaling in DLBCL. © 2017 by The American Society of Hematology.
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Yamamura, Hideto; Suzuki, Yoshiaki; Yamamura, Hisao; Asai, Kiyofumi; Imaizumi, Yuji
2016-08-05
The blood-brain barrier (BBB) is mainly composed of brain capillary endothelial cells (BCECs), astrocytes and pericytes. Brain ischemia causes hypoxic encephalopathy and damages BBB. However, it remains still unclear how hypoxia affects BCECs. In the present study, t-BBEC117 cells, an immortalized bovine brain endothelial cell line, were cultured under hypoxic conditions at 4-5% oxygen for 72 h. This hypoxic stress caused hyperpolarization of resting membrane potential. Patch-clamp recordings revealed a marked increase in Ba(2+)-sensitive inward rectifier K(+) current in t-BBEC117 cells after hypoxic culture. Western blot and real-time PCR analyses showed that Kir2.1 expression was significantly up-regulated at protein level but not at mRNA level after the hypoxic culture. Ca(2+) imaging study revealed that the hypoxic stress enhanced store-operated Ca(2+) (SOC) entry, which was significantly reduced in the presence of 100 μM Ba(2+). On the other hand, the expression of SOC channels such as Orai1, Orai2, and transient receptor potential channels was not affected by hypoxic stress. MTT assay showed that the hypoxic stress significantly enhanced t-BBEC117 cell proliferation, which was inhibited by approximately 60% in the presence of 100 μM Ba(2+). We first show here that moderate cellular stress by cultivation under hypoxic conditions hyperpolarizes membrane potential via the up-regulation of functional Kir2.1 expression and presumably enhances Ca(2+) entry, resulting in the facilitation of BCEC proliferation. These findings suggest potential roles of Kir2.1 expression in functional changes of BCECs in BBB following ischemia. Copyright © 2016 Elsevier Inc. All rights reserved.
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Stroma provides an intestinal stem cell niche in the absence of epithelial Wnts.
Kabiri, Zahra; Greicius, Gediminas; Madan, Babita; Biechele, Steffen; Zhong, Zhendong; Zaribafzadeh, Hamed; Edison; Aliyev, Jamal; Wu, Yonghui; Bunte, Ralph; Williams, Bart O; Rossant, Janet; Virshup, David M
2014-06-01
Wnt/β-catenin signaling supports intestinal homeostasis by regulating proliferation in the crypt. Multiple Wnts are expressed in Paneth cells as well as other intestinal epithelial and stromal cells. Ex vivo, Wnts secreted by Paneth cells can support intestinal stem cells when Wnt signaling is enhanced with supplemental R-Spondin 1 (RSPO1). However, in vivo, the source of Wnts in the stem cell niche is less clear. Genetic ablation of Porcn, an endoplasmic reticulum resident O-acyltransferase that is essential for the secretion and activity of all vertebrate Wnts, confirmed the role of intestinal epithelial Wnts in ex vivo culture. Unexpectedly, mice lacking epithelial Wnt activity (Porcn(Del)/Villin-Cre mice) had normal intestinal proliferation and differentiation, as well as successful regeneration after radiation injury, indicating that epithelial Wnts are dispensable for these processes. Consistent with a key role for stroma in the crypt niche, intestinal stromal cells endogenously expressing Wnts and Rspo3 support the growth of Porcn(Del) organoids ex vivo without RSPO1 supplementation. Conversely, increasing pharmacologic PORCN inhibition, affecting both stroma and epithelium, reduced Lgr5 intestinal stem cells, inhibited recovery from radiation injury, and at the highest dose fully blocked intestinal proliferation. We conclude that epithelial Wnts are dispensable and that stromal production of Wnts can fully support normal murine intestinal homeostasis.
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Pefani, Dafni-Eleutheria; Dimaki, Maria; Spella, Magda; Karantzelis, Nickolas; Mitsiki, Eirini; Kyrousi, Christina; Symeonidou, Ioanna-Eleni; Perrakis, Anastassis; Taraviras, Stavros; Lygerou, Zoi
2011-01-01
Development and homeostasis of multicellular organisms relies on an intricate balance between cell proliferation and differentiation. Geminin regulates the cell cycle by directly binding and inhibiting the DNA replication licensing factor Cdt1. Geminin also interacts with transcriptional regulators of differentiation and chromatin remodelling factors, and its balanced interactions are implicated in proliferation-differentiation decisions during development. Here, we describe Idas (Idas being a cousin of the Gemini in Ancient Greek Mythology), a previously uncharacterised coiled-coil protein related to Geminin. We show that human Idas localizes to the nucleus, forms a complex with Geminin both in cells and in vitro through coiled-coil mediated interactions, and can change Geminin subcellular localization. Idas does not associate with Cdt1 and prevents Geminin from binding to Cdt1 in vitro. Idas depletion from cells affects cell cycle progression; cells accumulate in S phase and are unable to efficiently progress to mitosis. Idas protein levels decrease in anaphase, whereas its overexpression causes mitotic defects. During development, we show that Idas exhibits high level expression in the choroid plexus and the cortical hem of the mouse telencephalon. Our data highlight Idas as a novel Geminin binding partner, implicated in cell cycle progression, and a putative regulator of proliferation-differentiation decisions during development. PMID:21543332
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Zhang, Hengwei; Cao, Rui; Lee, Wai-Nang Paul; Deng, Caishu; Zhao, Yingchun; Lappe, Joan; Recker, Robert; Yen, Yun; Wang, Qi; Tsai, Ming-Ying; Go, Vay Liang; Xiao, Gary Guishan
2010-01-01
Oxythiamine (OT), a transketolase inhibitor, is known to inhibit pancreatic cancer cell proliferation. In this study, we investigated the effect of inhibition of the transketolase pathway on signaling pathways in MIA PaCa cancer cells using in-house proteomic techniques. We hypothesized that OT alter protein phosphorylation thus affecting cell cycle arrest and cell proliferation. MIA PaCa-2 cells were cultured in media containing an algal 15N amino acid mixture at 50% enrichment, with and without OT, to determine protein expression and synthesis. Analysis of cell lysates using two-dimensional gel electrophoresis matrix assisted laser desorption and ionization time-of-flight and time-of-flight mass spectrometry (2-DE-MALDI-TOF/TOF MS) identified 12 phosphor proteins that were significantly suppressed by OT treatment. Many of these proteins are involved in regulation of cycle activities and apoptosis. Among the proteins identified, expression of the phosphor heat shock protein 27 (Hsp27) was dramatically inhibited by OT treatment while the level of its total protein remained unchanged. Hsp27 expression and phoshporylation is known to be associated with drug resistance and cancer cell survival. The changes in phosphorylation of key proteins of cancer proliferation and survival suggest that protein phosphorylation is the confluence of the effects of OT on metabolic and signaling pathways. PMID:20035555
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Pefani, Dafni-Eleutheria; Dimaki, Maria; Spella, Magda; Karantzelis, Nickolas; Mitsiki, Eirini; Kyrousi, Christina; Symeonidou, Ioanna-Eleni; Perrakis, Anastassis; Taraviras, Stavros; Lygerou, Zoi
2011-07-01
Development and homeostasis of multicellular organisms relies on an intricate balance between cell proliferation and differentiation. Geminin regulates the cell cycle by directly binding and inhibiting the DNA replication licensing factor Cdt1. Geminin also interacts with transcriptional regulators of differentiation and chromatin remodelling factors, and its balanced interactions are implicated in proliferation-differentiation decisions during development. Here, we describe Idas (Idas being a cousin of the Gemini in Ancient Greek Mythology), a previously uncharacterised coiled-coil protein related to Geminin. We show that human Idas localizes to the nucleus, forms a complex with Geminin both in cells and in vitro through coiled-coil mediated interactions, and can change Geminin subcellular localization. Idas does not associate with Cdt1 and prevents Geminin from binding to Cdt1 in vitro. Idas depletion from cells affects cell cycle progression; cells accumulate in S phase and are unable to efficiently progress to mitosis. Idas protein levels decrease in anaphase, whereas its overexpression causes mitotic defects. During development, we show that Idas exhibits high level expression in the choroid plexus and the cortical hem of the mouse telencephalon. Our data highlight Idas as a novel Geminin binding partner, implicated in cell cycle progression, and a putative regulator of proliferation-differentiation decisions during development.
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Jenkins, A P; Thompson, R P
1992-01-01
This study investigated how substitution of long chain triglycerides for glucose in a mixed diet affects the overall small intestinal mucosal mass and the distribution of mucosal mass and cell proliferation along the small intestine. Four groups of eight female Wistar rats (180-200 g) were isocalorically fed mixed diets containing the essential fatty acid rich oil Efamol substituted for glucose at concentrations of 1.2%, 10%, 25%, and 50% total calories for 20 to 23 days. The small intestine was divided into three equal length segments and whole gut weights, mucosal weights, protein and DNA determined. Cell proliferation was estimated from the two hour accumulation of vincristine arrested metaphases in microdissected crypts at points 0%, 17%, 33%, 50%, 66%, and 100% small intestinal length. There were no differences between groups in parameters of overall small intestinal or distal segment mucosal mass. With increasing levels of fat, however, there was a significant trend for the mucosal mass of the proximal segment to fall and that of the middle segment to rise. The pattern of two hour metaphase accumulation reflected these changes. These regional changes in mucosal mass and cell proliferation may reflect differences in the sites of absorption of fat and glucose. PMID:1541418
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Reid, Katherine J; Lang, Kenneth; Froscio, Suzanne; Humpage, Andrew J; Young, Fiona M
2015-11-01
Undifferentiated mouse embryonic stem cell (mES) proliferation in vitro resembles aspects of in vivo pre-implantation embryonic development. mES were used to assess the embryo-toxicity of cylindrospermopsin (CYN), a water contaminant with an Australian Drinking Water Guideline (ADWG) of 1 μg/L. mES exposed to 0-1 μg/mL CYN for 24-168 h were subjected to an optimised crystal violet viability assay. mES exposed to retinoic acid ± 1 μg/L CYN differentiated into neural-like cells confirmed by morphological examination and RT-PCR for Oct4, Brachyury and Nestin. The CYN No Observed Effect Concentration (OEC) was 0.5 μg/mL, the Lowest OEC was 1 μg/mL (p < 0.001, n = 3), and the IC50 was 0.86 μg/mL after 24 h. The ADWG 1 μg/L CYN did not affect differentiation or proliferation after 72 h, but decreased proliferation after 168 h (p < 0.05). We conclude that higher algal bloom-associated CYN concentrations have the potential to impair in vivo pre-implantation development, and the mES crystal violet assay has broad application to screening environmental toxins. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Jenkins, A P; Thompson, R P
1992-02-01
This study investigated how substitution of long chain triglycerides for glucose in a mixed diet affects the overall small intestinal mucosal mass and the distribution of mucosal mass and cell proliferation along the small intestine. Four groups of eight female Wistar rats (180-200 g) were isocalorically fed mixed diets containing the essential fatty acid rich oil Efamol substituted for glucose at concentrations of 1.2%, 10%, 25%, and 50% total calories for 20 to 23 days. The small intestine was divided into three equal length segments and whole gut weights, mucosal weights, protein and DNA determined. Cell proliferation was estimated from the two hour accumulation of vincristine arrested metaphases in microdissected crypts at points 0%, 17%, 33%, 50%, 66%, and 100% small intestinal length. There were no differences between groups in parameters of overall small intestinal or distal segment mucosal mass. With increasing levels of fat, however, there was a significant trend for the mucosal mass of the proximal segment to fall and that of the middle segment to rise. The pattern of two hour metaphase accumulation reflected these changes. These regional changes in mucosal mass and cell proliferation may reflect differences in the sites of absorption of fat and glucose.
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Lim, Ji-Hey; McCullen, Seth D; Piedrahita, Jorge A; Loboa, Elizabeth G; Olby, Natasha J
2013-10-01
Application of sinusoidal electric fields (EFs) has been observed to affect cellular processes, including alignment, proliferation, and differentiation. In the present study, we applied low-frequency alternating current (AC) EFs to porcine neural progenitor cells (pNPCs) and investigated the effects on cell patterning, proliferation, and differentiation. pNPCs were grown directly on interdigitated electrodes (IDEs) localizing the EFs to a region accessible visually for fluorescence-based assays. Cultures of pNPCs were exposed to EFs (1 V/cm) of 1 Hz, 10 Hz, and 50 Hz for 3, 7, and 14 days and compared to control cultures. Immunocytochemistry was performed to evaluate the expression of neural markers. pNPCs grew uniformly with no evidence of alignment to the EFs and no change in cell numbers when compared with controls. Nestin expression was shown in all groups at 3 and 7 days, but not at 14 days. NG2 expression was low in all groups. Co-expression of glial fibrillary acidic protein (GFAP) and TUJ1 was significantly higher in the cultures exposed to 10- and 50-Hz EFs than the controls. In summary, sinusoidal AC EFs via IDEs did not alter the alignment and proliferation of pNPCs, but higher frequency stimulation appeared to delay differentiation into mature astrocytes.
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Makhija, Seema; Gupta, Renu; Toteja, Ravi
2015-08-01
Lithium is known to have profound biological effects of varying intensity in different life forms. In the present investigation, the effect of lithium was studied on the spirotrich ciliate Stylonychia lemnae. Lithium treatment brings about quantitative changes in the patterning of ciliary structures in S. lemnae. The dorsal surface of the affected cells develops supernumerary ciliary kineties due to excessive proliferation of the kinetosomes. The ventral surface on the other hand develops fewer than normal cirri formed from reduced numbers of ciliary primordia. The adoral zone of membranelles (AZM) fails to remodel properly as, in certain segments, membranelles become disarranged and misaligned. Lithium-induced changes are transitory as the normal pattern is restored during recovery after the cells are shifted to normal medium, suggesting non-genic regulation of cortical pattern. Lithium also affects the process of cell proliferation as the number of cells undergoing division is negligible as compared to reorganizing cells. The results point to the extremely complex and heterogeneous organization of the cellular cortex (plasma membrane and cytoskeleton) which is capable of exerting autonomous control over the phenotype and cortical pattern. Copyright © 2015 Elsevier GmbH. All rights reserved.
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Guiñazú, Natalia; Rena, Viviana; Genti-Raimondi, Susana; Rivero, Virginia; Magnarelli, Gladis
2012-04-01
Epidemiological data have associated environmental organophosphate insecticide (OP) exposure during pregnancy with fetal growth deficits. To better understand OP injury that may adversely affect pregnancy, we used the JEG-3 choriocarcinoma cell line, which provide a recognized in vitro model to study placental function. The effects of the OP phosmet (Pm) and chlorpyrifos (Cp) on JEG-3 cells viability, proliferation, cell cycle and inflammatory molecule production were evaluated. Both insecticides affected cellular viability in a concentration- and time-dependent manner, inducing apoptosis and decreasing [(3)H]-thymidine incorporation. However, only Pm reduced DNA synthesis independently of cellular death and decreased the cell percentage at the S-phase. Unlike apoptosis, TNFα production varied with the concentration tested, suggesting that other TNFα independent mechanisms might trigger cell death. No induction of the inflammatory molecule nitric oxide was detected. The mRNA levels of pro-inflammatory IL-6, IL-17 and the anti-inflammatory IL-13 cytokines were differentially modulated. These findings show that Pm and Cp generate a specific toxicity signature, altering cell viability and inducing an inflammatory cytokine profile, suggesting that trophoblasts may represent a possible target for OP adverse effects. Copyright © 2012 Elsevier Ltd. All rights reserved.
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Hsp47 mediates Cx43-dependent skeletal growth and patterning in the regenerating fin.
Bhadra, Joyita; Iovine, M Kathryn
2015-11-01
Skeletal morphogenesis describes how bones achieve their correct shape and size and appropriately position joints. We use the regenerating caudal fin of zebrafish to study this process. Our examination of the fin length mutant short fin (sof (b123)) has revealed that the gap junction protein Cx43 is involved in skeletal morphogenesis by promoting cell proliferation and inhibiting joint formation, thereby coordinating skeletal growth and patterning. Here we demonstrate that serpinh1b is molecularly and functionally downstream of cx43. The gene serpinh1b codes for a protein called Hsp47, a molecular chaperone responsible for proper folding of procollagen molecules. Knockdown of Hsp47 in regenerating fins recapitulates the sof (b123) phenotypes of reduced fin length, reduced segment length and reduced level of cell proliferation. Furthermore, Hsp47 knockdown affects the organization and localization of the collagen-based actinotrichia. Together, our findings reveal that serpinh1b acts in a cx43 dependent manner to regulate cell proliferation and joint formation. We conclude that disruption of the collagen-based extracellular matrix influences signaling events required for cell proliferation, as well as the patterning of skeletal precursor cells that influences segment length. Therefore, we suggest that Hsp47 function is necessary for skeletal growth and patterning during fin regeneration. Copyright © 2015 Elsevier B.V. All rights reserved.
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Synemin promotes AKT-dependent glioblastoma cell proliferation by antagonizing PP2A.
Pitre, Aaron; Davis, Nathan; Paul, Madhumita; Orr, A Wayne; Skalli, Omar
2012-04-01
The intermediate filament protein synemin is present in astrocyte progenitors and glioblastoma cells but not in mature astrocytes. Here we demonstrate a role for synemin in enhancing glioblastoma cell proliferation and clonogenic survival, as synemin RNA interference decreased both behaviors by inducing G1 arrest along with Rb hypophosphorylation and increased protein levels of the G1/S inhibitors p21(Cip1) and p27(Kip1). Akt involvement was demonstrated by decreased phosphorylation of its substrate, p21(Cip1), and reduced Akt catalytic activity and phosphorylation at essential activation sites. Synemin silencing, however, did not affect the activities of PDPK1 and mTOR complex 2, which directly phosphorylate Akt activation sites, but instead enhanced the activity of the major regulator of Akt dephosphorylation, protein phosphatase type 2A (PP2A). This was accompanied by changes in PP2A subcellular distribution resulting in increased physical interactions between PP2A and Akt, as shown by proximity ligation assays (PLAs). PLAs and immunoprecipitation experiments further revealed that synemin and PP2A form a protein complex. In addition, treatment of synemin-silenced cells with the PP2A inhibitor cantharidic acid resulted in proliferation and pAkt and pRb levels similar to those of controls. Collectively these results indicate that synemin positively regulates glioblastoma cell proliferation by helping sequester PP2A away from Akt, thereby favoring Akt activation.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Shim, Ki Shuk; Department of Neonatology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna; Rosner, Margit
Bach1 and Bach2 are evolutionarily related members of the BTB-basic region leucine zipper transcription factor family. We found that Bach2 downregulates cell proliferation of N1E-115 cells and negatively affects their potential to differentiate. Nuclear localization of the cyclin-dependent kinase inhibitor p21 is known to arrest cell cycle progression, and cytoplasmic p21 has been shown to promote neuronal differentiation of N1E-115 cells. We found that ectopic Bach2 causes upregulation of p21 expression in the nucleus and in the cytoplasm in undifferentiated N1E-115 cells. In differentiated cells, Bach2 specifically triggers upregulation of cytoplasmic p21. Our data suggest that Bach2 expression could representmore » a switch during the process of neuronal differentiation. Bach2 is not expressed in neuronal precursor cells. It would have negative effects on proliferation and differentiation of these cells. In differentiated neuronal cells Bach2 expression is upregulated, which could allow Bach2 to function as a gatekeeper of the differentiated status.« less
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Reconstructing the in vivo dynamics of hematopoietic stem cells from telomere length distributions
Werner, Benjamin; Beier, Fabian; Hummel, Sebastian; Balabanov, Stefan; Lassay, Lisa; Orlikowsky, Thorsten; Dingli, David; Brümmendorf, Tim H; Traulsen, Arne
2015-01-01
We investigate the in vivo patterns of stem cell divisions in the human hematopoietic system throughout life. In particular, we analyze the shape of telomere length distributions underlying stem cell behavior within individuals. Our mathematical model shows that these distributions contain a fingerprint of the progressive telomere loss and the fraction of symmetric cell proliferations. Our predictions are tested against measured telomere length distributions in humans across all ages, collected from lymphocyte and granulocyte sorted telomere length data of 356 healthy individuals, including 47 cord blood and 28 bone marrow samples. We find an increasing stem cell pool during childhood and adolescence and an approximately maintained stem cell population in adults. Furthermore, our method is able to detect individual differences from a single tissue sample, i.e. a single snapshot. Prospectively, this allows us to compare cell proliferation between individuals and identify abnormal stem cell dynamics, which affects the risk of stem cell related diseases. DOI: http://dx.doi.org/10.7554/eLife.08687.001 PMID:26468615
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Zhang, Haitao; Li, Ning; Wu, Jun; Su, Lijuan; Chen, Xiaoyi; Lin, Biyun; Luo, Hui
2013-10-15
Galangin, a flavonol derived from Alpinia officinarum Hance and used as food additives in southern China, induces apoptosis and autophagy to suppress the proliferation of HepG2 cells. In this study, we demonstrated that galangin induced autophagy by increasing the ratio of AMP/TAN in HepG2 cells. It stimulated the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) and LKB1, but inhibited the phosphorylation of AKT and mTOR. Inhibition of AMPK activation suppressed the dephosphorylation of mTOR to block galangin-induced autophagy. AMPK activation by galangin appeared to be independent of the LKB1 signaling pathway because the down-regulation of LKB1 by its siRNA failed to affect galangin-induced autophagy. Collectively, the findings demonstrated a novel mechanism of how galangin induces autophagy via activating AMPK in a LKB1- independent manner. The induction of autophagy can thus reflect the anti-proliferation effect of galangin in HCC cells. © 2013 Elsevier B.V. All rights reserved.
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Veno-occlusive disease in snow leopards (Panthera uncia) from zoological parks.
Munson, L; Worley, M B
1991-01-01
Livers from 54 snow leopards, 4 days to 23 years old, that had died in 23 US zoos, were evaluated histopathologically to determine if the hepatic fibrosis, which has been noted to be prevalent in this species, was due to chronic active hepatitis from hepadnaviral infection, Ito cell proliferation, or hemosiderosis. Forty-two of 54 snow leopards had subintimal vascular fibrosis with partial or total occlusion of central and sublobular veins (veno-occlusive disease) of unknown origin. All 21 leopards older than 5 years were affected. Four leopards had chronic active hepatitis, and 12 leopards had cholangiohepatitis; but these lesions were not connected anatomically to central and sublobular venous fibrosis. Hepatocellular and Kupffer cell siderosis and Ito cell proliferation were prevalent and often coexisted with perisinusoidal, central, and sublobular venous fibrosis; but fibrosis was present in leopards without siderosis or Ito cell proliferation. The pattern and prevalence of veno-occlusive disease in these leopards was similar to that reported in captive cheetah (Acinonyx jubatus), suggesting that a common extrinsic factor may cause the majority of hepatic disease in these large felid animals in captivity.
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Immunoexcitatory mechanisms in glioma proliferation, invasion and occasional metastasis
Blaylock, Russell L.
2013-01-01
There is increasing evidence of an interaction between inflammatory cytokines and glutamate receptors among a number of neurological diseases including traumatic brain injuries, neurodegenerative diseases and central nervous system (CNS) infections. A number of recent studies have now suggested a strong relation between inflammatory mechanisms and excitatory cascades and these may play a role in glioma invasiveness and proliferation. Chronic inflammation appears to be a major initiating mechanism in most human cancers, involving cell-signaling pathways, which are responsible for cell cycling, cancer cell migration, invasion, tumor aggressiveness, and angiogenesis. It is less well appreciated that glutamate receptors also play a significant role in both proliferation and especially glioma invasion. There is some evidence that sustained elevations in glutamate may play a role in initiating certain cancers and new studies demonstrate an interaction between inflammation and glutamate receptors that may enhance tumor invasion and metastasis by affecting a number of cell-signaling mechanisms. These mechanisms are discussed in this paper as well as novel treatment options for reducing immune-glutamate promotion of cancer growth and invasion. PMID:23493580
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Yang, Chenggang; Du, Wenfeng; Yang, Daogui
2016-11-01
(-)-Epigallocatechin-3-gallate (EGCG), the major polyphenol in green tea, could affect carcinogenesis and development of many cancers. However, the effects and underlying mechanisms of EGCG on gastric cancer remain unclear. We found that EGCG significantly inhibited proliferation and increased apoptosis of SGC-7901 cells in vitro. The decreased expressions of p-β-catenin(Ser552), p-GSK3β(S9) and β-catenin target genes were detected in SGC-7901 cells after treated by EGCG. XAV939 and β-catenin plasmid were further used to demonstrate the inhibition of EGCG on canonical Wnt/β-catenin signalling. Moreover, EGCG significantly inhibited gastric tumour growth in vivo by inhibiting Wnt/β-catenin signalling. Taken together, our findings establish that EGCG suppressed gastric cancer cell proliferation and demonstrate that this inhibitory effect is related to canonical Wnt/β-catenin signalling. This study raises a new insight into gastric cancer prevention and therapy, and provides evidence that green tea could be used as a nutraceutical beverage.
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NASA Astrophysics Data System (ADS)
Grzincic, E. M.; Yang, J. A.; Drnevich, J.; Falagan-Lotsch, P.; Murphy, C. J.
2015-01-01
Gold nanoparticles (Au NPs) are attractive for biomedical applications not only for their remarkable physical properties, but also for the ease of which their surface chemistry can be manipulated. Many applications involve functionalization of the Au NP surface in order to improve biocompatibility, attach targeting ligands or carry drugs. However, changes in cells exposed to Au NPs of different surface chemistries have been observed, and little is known about how Au NPs and their surface coatings may impact cellular gene expression. The gene expression of two model human cell lines, human dermal fibroblasts (HDF) and prostate cancer cells (PC3) was interrogated by microarray analysis of over 14 000 human genes. The cell lines were exposed to four differently functionalized Au NPs: citrate, poly(allylamine hydrochloride) (PAH), and lipid coatings combined with alkanethiols or PAH. Gene functional annotation categories and weighted gene correlation network analysis were used in order to connect gene expression changes to common cellular functions and to elucidate expression patterns between Au NP samples. Coated Au NPs affect genes implicated in proliferation, angiogenesis, and metabolism in HDF cells, and inflammation, angiogenesis, proliferation apoptosis regulation, survival and invasion in PC3 cells. Subtle changes in surface chemistry, such as the initial net charge, lability of the ligand, and underlying layers greatly influence the degree of expression change and the type of cellular pathway affected.Gold nanoparticles (Au NPs) are attractive for biomedical applications not only for their remarkable physical properties, but also for the ease of which their surface chemistry can be manipulated. Many applications involve functionalization of the Au NP surface in order to improve biocompatibility, attach targeting ligands or carry drugs. However, changes in cells exposed to Au NPs of different surface chemistries have been observed, and little is known about how Au NPs and their surface coatings may impact cellular gene expression. The gene expression of two model human cell lines, human dermal fibroblasts (HDF) and prostate cancer cells (PC3) was interrogated by microarray analysis of over 14 000 human genes. The cell lines were exposed to four differently functionalized Au NPs: citrate, poly(allylamine hydrochloride) (PAH), and lipid coatings combined with alkanethiols or PAH. Gene functional annotation categories and weighted gene correlation network analysis were used in order to connect gene expression changes to common cellular functions and to elucidate expression patterns between Au NP samples. Coated Au NPs affect genes implicated in proliferation, angiogenesis, and metabolism in HDF cells, and inflammation, angiogenesis, proliferation apoptosis regulation, survival and invasion in PC3 cells. Subtle changes in surface chemistry, such as the initial net charge, lability of the ligand, and underlying layers greatly influence the degree of expression change and the type of cellular pathway affected. Electronic supplementary information (ESI) available: UV-Vis spectra of Au NPs, the most significantly changed genes of HDF cells after Au NP incubation under GO accession number GO:0007049 ``cell cycle'', detailed information about the primer/probe sets used for RT-PCR validation of results. See DOI: 10.1039/c4nr05166a
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Wu, Ping; Liu, Siyuan; Su, Jianyu; Chen, Jianping; Li, Lin; Zhang, Runguang; Chen, Tianfeng
2017-10-18
Cancer cells are well known to require a constant supply of protein, lipid, RNA, and DNA via altered metabolism for accelerated cell proliferation. Targeting metabolic pathways is, therefore, a promising therapeutic strategy for cancers. Isoquercitrin (ISO) is widely distributed in dietary and medicinal plants and displays selective cytotoxicity to cancer cells, primarily by inducing apoptosis and cell cycle arrest. The aims of this study were to find out whether ISO could stabilize in a bladder-like acidic environment and inhibit bladder cancer cell proliferation by affecting their metabolism, and to investigate its molecular mechanism. In this study, the exposure of T24 bladder cancer cells to ISO (20-80 μM) decreased cell viability by causing ROS overproduction. This ROS change regulated the AMPK signaling pathway, and caused Caspase-dependent apoptosis as well as metabolism dysfunction. Metabolic alterations elevated metabolic pathway variation, which in turn destabilized lipid synthesis and altered anaerobic glycolysis. This linkage was proved by immunoblotting assay, and metabolomics as identified by UHPLC-QTOF-MS. Our findings provide comprehensive evidence that ISO influenced T24 bladder cancer cell metabolism, and that this process was mainly involved in activating the AMPK pathway. This study could lead to an understanding of how ISO suppresses bladder cancer cell growth, and whether the affected cancer metabolism is a common mechanism by which nutritional compounds suppress cancers.
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Choi, Yu-Jin; Choi, Yun-Sik
2015-01-01
Objectives Nonionizing radiation is emitted from electronic devices, such as smartphones. In this study, we intended to elucidate the effect of electromagnetic radiation from smartphones on spatial working memory and progenitor cell proliferation in the hippocampus. Methods Both male and female mice were randomly separated into two groups (radiated and control) and the radiated group was exposed to electromagnetic radiation for 9 weeks and 11 weeks for male and female mice, respectively. Spatial working memory was examined with a Y maze, and proliferation of hippocampal progenitor cells were examined by 5-bromo-2′-deoxyuridine administration and immunohistochemical detection. Results When spatial working memory on a Y maze was examined in the 9th week, there was no significant difference in the spontaneous alternation score on the Y maze between the two groups. In addition, there was no significant difference in hippocampal progenitor cell proliferation. However, immunoreactivity to glial fibrillary acidic protein was increased in exposed animals. Next, to test the effect of recovery following chronic radiation exposure, the remaining female mice were further exposed to electromagnetic radiation for 2 more weeks (total 11 weeks), and spontaneous alternation was tested 4 weeks later. In this experiment, although there was no significant difference in the spontaneous alternation scores, the number of arm entry was significantly increased. Conclusion These data indicate that although chronic electromagnetic radiation does not affect spatial working memory and hippocampal progenitor cell proliferation it can mediate astrocyte activation in the hippocampus and delayed hyperactivity-like behavior. PMID:26981337
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Seder, Christopher W; Hartojo, Wibisono; Lin, Lin; Silvers, Amy L; Wang, Zhuwen; Thomas, Dafydd G; Giordano, Thomas J; Chen, Guoan; Chang, Andrew C; Orringer, Mark B; Beer, David G
2009-01-01
Introduction The expression, mechanisms of regulation, and functional impact of INHBA (activin A) in lung adenocarcinoma (AD) have not been fully elucidated. Methods INHBA expression was examined in 96 lung samples (86 ADs, 10 normal lung) using oligonucleotide microarrays and 187 lung samples (164 ADs, 6 bronchioalveolar carcinomas, and 17 normal lung) using immunohistochemistry. The proliferation of AD cell lines H460 and SKLU1 was examined with WST-1 assays after treatment with recombinant activin A, follistatin, and INHBA-targeting small-interfering RNA. Cells were also treated with 5-aza-2′ deoxycytidine and trichostatin A to investigate the role of epigenetic regulation in INHBA expression. Results Primary ADs expressed 3.1 times more INHBA mRNA than normal lung. In stage I AD patients, high levels of primary tumor INHBA transcripts were associated with worse prognosis. Immunohistochemistry confirmed higher inhibin βA protein expression in ADs (78.7%) and bronchioalveolar carcinomas (66.7%) compared with normal lung (11.8%). H460 and SKLU1 demonstrated increased proliferation when treated with exogenous activin A and reduced proliferation when treated with follistatin or INHBA-targeting small-interfering RNA. INHBA mRNA expression in H460 cells was upregulated after treatment with trichostatin A and 5-aza-2′ deoxycytidine. Conclusions INHBA is overexpressed in AD relative to controls. Inhibin βA may promote cell proliferation, and its overexpression is associated with worse survival in stage I AD patients. In addition, overexpression of INHBA may be affected by promoter methylation and histone acetylation in a subset of lung ADs. PMID:19308293
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Everolimus selectively targets vemurafenib resistant BRAFV600E melanoma cells adapted to low pH.
Ruzzolini, Jessica; Peppicelli, Silvia; Andreucci, Elena; Bianchini, Francesca; Margheri, Francesca; Laurenzana, Anna; Fibbi, Gabriella; Pimpinelli, Nicola; Calorini, Lido
2017-11-01
Vemurafenib, a BRAF inhibitor, elicits in ∼80% of BRAF V600E -mutant melanoma patients a transient anti-tumor response which precedes the emergence of resistance. We tested whether an acidic tumor microenvironment may favor a BRAF inhibitor resistance. A375M6 BRAF V600E melanoma cells, either exposed for a short period or chronically adapted to an acidic medium, showed traits compatible with an epithelial-mesenchymal transition, reduced proliferation and high resistance to apoptosis. Both types of acidic cells treated with vemurafenib did not change their proliferation, distribution in cell cycle and level of p-AKT, in contrast to cells grown at standard pH, which showed reduced proliferation, cell cycle arrest and ERK/AKT inhibition. Even after treatment with trametinib (MEK inhibitor) acidic cell features did not change. Then, since both types of acidic cells exhibited high p-p70S6K, i.e. active mTOR signaling, we tested everolimus, an mTOR inhibitor, which was efficient in inducing apoptosis in acidic cells without affecting melanoma cells grown at standard pH. Our results indicate that an acidic microenvironment may cooperate in inducing a BRAF inhibitor resistance in melanoma cells and a combined therapy with everolimus could be used to overcome that resistance. Copyright © 2017 Elsevier B.V. All rights reserved.
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[Effects of methyl tertiary butyl ether on cell cycle and cell apoptosis].
Zhou, W; Huang, G; Zhang, H; Ye, S
2000-07-01
To explore the effects of the new gasoline additive, methyl tertiary butyl ether (MTBE) on cell cycle and cell apoptosis. Flow cytometry was used to evaluate the effect of MTBE (1, 2, 4 microl/ml, 24 h) on NIH/3T3 cell cycles; and the effect of MTBE on Hela cell apoptosis was evaluated by detecting cell survival using crystal violet staining. Flow cytometry showed that MTBE could change NIH/3T3 cell cycles, decrease the number of cells in S stage, and arrest cells at G(2) + M stage. The results suggested that MTBE could affect NIH/3T3 cell cycles and induce cell proliferation. This situation existed 48 hours after the treatment, and cell cycles came back normal 96 hours after the treatment. By detecting cell survival using crystal violet staining, we found that MTBE could inhibit the apoptosis of Hela cells which was induced by tumor necrosis factor (TNF)alpha and cycloheximide. MTBE's carcinogenicity to animals may relate to induction of cell proliferation and inhibition of cell apoptosis.
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Wu, Y; Wen, F; Gouk, S S; Lee, E H; Kuleshova, L
2015-01-01
The development of vitrification strategy for cell-biomaterial constructs, particularly biologically inspired nanoscale materials and hydrogels mimicking the in vivo environment is an active area. A cryopreservation strategy mimicking the in vivo environment for cell-hydrogel constructs may enhance cell proliferation and biological function. To demonstrate the efficacy of vitrification as a platform technology involving tissue engineering and human mesenchymal stem cells (hMSCs). Microcarriers made from alginate coated with chitosan and collagen are used. Conventional freezing and vitrification were compared. The vitrification strategy includes 10 min step-wise exposure to a vitrification solution (40% v/v EG, 0.6M sucrose) and immersion into liquid nitrogen. Confocal imaging of live/dead staining of hMSCs cultured on the surface of microcarriers demonstrated that vitrified cells had excellent appearance and prolonged spindle shape morphology. The proliferation ability of post-vitrified cells arbitrated to protein Ki-67 gene expression was not significantly different in comparison to untreated control, while that of post-freezing cells was almost lost. The ability of hMSCs cultured on the surface of microcarriers to proliferate has been not affected by vitrification and it was significantly better after vitrification than after conventional freezing during continuous culture. Collagen II related mRNA expression by 4 weeks post-vitrification and post-freezing showed that ability to differentiate into cartilage was sustained during vitrification and reduced during conventional freezing. No significant difference was found between control and vitrification groups only. Vitrification strategy coupled with advances in hMSC-expansion platform that completely preserves the ability of stem cells to proliferate and subsequently differentiate allows not only to reach a critical cell number, but also demonstrate prospects for effective utilization and transportation of cells with their support system, creating demand for novel biodegradable materials.
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Murillo-Carretero, Maribel; Geribaldi-Doldán, Noelia; Flores-Giubi, Eugenia; García-Bernal, Francisco; Navarro-Quiroz, Elkin A; Carrasco, Manuel; Macías-Sánchez, Antonio J; Herrero-Foncubierta, Pilar; Delgado-Ariza, Antonio; Verástegui, Cristina; Domínguez-Riscart, Jesús; Daoubi, Mourad; Hernández-Galán, Rosario; Castro, Carmen
2017-07-01
Pharmacological strategies aimed to facilitate neuronal renewal in the adult brain, by promoting endogenous neurogenesis, constitute promising therapeutic options for pathological or traumatic brain lesions. We have previously shown that non-tumour-promoting PKC-activating compounds (12-deoxyphorbols) promote adult neural progenitor cell (NPC) proliferation in vitro and in vivo, enhancing the endogenous neurogenic response of the brain to a traumatic injury. Here, we show for the first time that a diterpene with a lathyrane skeleton can also activate PKC and promote NPC proliferation. We isolated four lathyranes from the latex of Euphorbia plants and tested their effect on postnatal NPC proliferation, using neurosphere cultures. The bioactive lathyrane ELAC (3,12-di-O-acetyl-8-O-tigloilingol) was also injected into the ventricles of adult mice to analyse its effect on adult NPC proliferation in vivo. The lathyrane ELAC activated PKC and significantly increased postnatal NPC proliferation in vitro, particularly in synergy with FGF2. In addition ELAC stimulated proliferation of NPC, specifically affecting undifferentiated transit amplifying cells. The proliferative effect of ELAC was reversed by either the classical/novel PKC inhibitor Gö6850 or the classical PKC inhibitor Gö6976, suggesting that NPC proliferation is promoted in response to activation of classical PKCs, particularly PKCß. ELAC slightly increased the proportion of NPC expressing Sox2. The effects of ELAC disappeared upon acetylation of its C7-hydroxyl group. We propose lathyranes like ELAC as new drug candidates to modulate adult neurogenesis through PKC activation. Functional and structural comparisons between ELAC and phorboids are included. © 2017 The British Pharmacological Society.
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bFGF Regulates PI3-Kinase-Rac1-JNK Pathway and Promotes Fibroblast Migration in Wound Healing
Kanazawa, Shigeyuki; Fujiwara, Toshihiro; Matsuzaki, Shinsuke; Shingaki, Kenta; Taniguchi, Manabu; Miyata, Shingo; Tohyama, Masaya; Sakai, Yasuo; Yano, Kenji; Hosokawa, Ko; Kubo, Tateki
2010-01-01
Fibroblast proliferation and migration play important roles in wound healing. bFGF is known to promote both fibroblast proliferation and migration during the process of wound healing. However, the signal transduction of bFGF-induced fibroblast migration is still unclear, because bFGF can affect both proliferation and migration. Herein, we investigated the effect of bFGF on fibroblast migration regardless of its effect on fibroblast proliferation. We noticed involvement of the small GTPases of the Rho family, PI3-kinase, and JNK. bFGF activated RhoA, Rac1, PI3-kinase, and JNK in cultured fibroblasts. Inhibition of RhoA did not block bFGF-induced fibroblast migration, whereas inhibition of Rac1, PI3-kinase, or JNK blocked the fibroblast migration significantly. PI3-kinase-inhibited cells down-regulated the activities of Rac1 and JNK, and Rac1-inhibited cells down-regulated JNK activity, suggesting that PI3-kinase is upstream of Rac1 and that JNK is downstream of Rac1. Thus, we concluded that PI3-kinase, Rac1, and JNK were essential for bFGF-induced fibroblast migration, which is a novel pathway of bFGF-induced cell migration. PMID:20808927
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Mhanna, Rami; Kashyap, Aditya; Palazzolo, Gemma; Vallmajo-Martin, Queralt; Becher, Jana; Möller, Stephanie; Schnabelrauch, Matthias
2014-01-01
The loss of expression of chondrogenic markers during monolayer expansion remains a stumbling block for cell-based treatment of cartilage lesions. Here, we introduce sulfated alginate hydrogels as a cartilage biomimetic biomaterial that induces cell proliferation while maintaining the chondrogenic phenotype of encapsulated chondrocytes. Hydroxyl groups of alginate were converted to sulfates by incubation with sulfur trioxide–pyridine complex (SO3/pyridine), yielding a sulfated material cross-linkable with calcium chloride. Passage 3 bovine chondrocytes were encapsulated in alginate and alginate sulfate hydrogels for up to 35 days. Cell proliferation was five-fold higher in alginate sulfate compared with alginate (p=0.038). Blocking beta1 integrins in chondrocytes within alginate sulfate hydrogels significantly inhibited proliferation (p=0.002). Sulfated alginate increased the RhoA activity of chondrocytes compared with unmodified alginate, an increase that was blocked by β1 blocking antibodies (p=0.017). Expression and synthesis of type II collagen, type I collagen, and proteoglycan was not significantly affected by the encapsulation material evidenced by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry. Alginate sulfate constructs showed an opaque appearance in culture, whereas the unmodified alginate samples remained translucent. In conclusion, alginate sulfate provides a three dimensional microenvironment that promotes both chondrocyte proliferation and maintenance of the chondrogenic phenotype and represents an important advance for chondrocyte-based cartilage repair therapies providing a material in which cell expansion can be done in situ. PMID:24320935
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Rastegar, Hosein; Ahmadi Ashtiani, Hamidreza; Aghaei, Mahmoud; Ehsani, Amirohushang; Barikbin, Behrooz
2013-06-01
Recently, platelet-rich plasma (PRP) has attracted attention in various medical fields, including plastic surgery, treatment for problematic wounds, and dermatology. Specifically, PRP has been tested during hair transplantation to reduce swelling and pain and to increase hair density. We examined the effects of PRP and herbal extracts combination in order to identify potential stimulants of hair growth. PRP was prepared using the double-spin method and applied to dermal papilla cells (DPCs). MTT viability test and BrdU cell proliferation assay were used to study the effect of herbal extracts and PRP on proliferation of DPCs. To understand the mechanisms of herbal extracts and PRP involved in the regulation of hair growth, we evaluated signaling pathways and measured the expressions of ERK and Akt, by Western blot. Combination of herbal extracts and PRP was found to induce significant proliferation of human DPCs at concentrations ranging from 1.5% to 4.5%. The present study shows that herbal extracts and PRP affect the expressions of extracellular signal-regulated kinase (ERK) and Akt in DPCs. In this study, we have shown that combination of herbal extracts and PRP plays an active role in promoting the proliferation of human dermal papilla (DP) cells via the regulation of ERK and Akt proteins, and this may be applicable to the future development of herbal extracts and PRP combination therapeutics to enhance hair growth. © 2013 Wiley Periodicals, Inc.
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Dawson, R A; Goberdhan, N J; Freedlander, E; MacNeil, S
1996-03-01
The aim of this study was to investigate whether prior culture of cells on ECM proteins might positively influence the performance of keratinocytes when cells are transferred to a dermal in vitro wound bed model. Keratinocytes were cultured using a method for producing cultured epithelial autografts for severely burned patients (essentially using Green's medium, a mitogen-rich medium containing fetal calf serum, cholera toxin, EGF, insulin, transferrin and triiodothyronine). Cells were cultured either on irradiated 3T3 fibroblasts (as in the standard Rheinwald and Green technique) or, alternatively, on collagen I, collagen IV, matrigel, RGD, vitronectin or fibronectin. Under these conditions matrigel, collagen I and IV enhanced initial attachment, RGD, vitronectin, fibronectin and irradiated 3T3 fibroblasts did not. Proliferation of cells was positively influenced by matrigel, collagen I and IV and irradiated 3T3 fibroblasts; of these, however, only matrigel and 3T3 fibroblasts had sustained significant effects on keratinocyte proliferation over 4 days. Cells on fibronectin showed significantly reduced proliferation. An acellular non-viable dermis was then used to mimic the homograft allodermis onto which cultured epithelial autograft sheets are grafted clinically and cells cultured on the various ECM proteins for 96 h were transferred to this in vitro wound model. None of the substrates enhanced keratinocyte performance on this model. It was concluded that under these conditions some ECM proteins can significantly affect keratinocyte attachment and, to a lesser extent, proliferation but that the culture of keratinocytes on these ECM proteins does not appear to confer any lasting benefit to the attachment of these keratinocytes to an in vitro wound-bed model.
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Mattingly, Kathleen A.; Klinge, Carolyn M.
2011-01-01
Endothelial dysfunction precedes cardiovascular disease and is accompanied by mitochondrial dysfunction. Here we tested the hypothesis that diesel exhaust particulate extracts (DEPEs), prepared from a truck run at different speeds and engine loads, would inhibit genomic estrogen receptor activation of nuclear respiratory factor-1 (NRF-1) transcription in human umbilical vein endothelial cells (HUVECs). Additionally, we examined how DEPEs affect NRF-1 regulated TFAM expression and, in turn, Tfam-regulated mtDNA-encoded cytochrome c oxidase subunit I (COI, MTCO1) and NADH dehydrogenase subunit I (NDI) expression as well as cell proliferation and viability. We report that 17β-estradiol (E2), 4-hydroxytamoxifen (4-OHT), and raloxifene increased NRF-1 transcription in HUVECs in an ER-dependent manner. DEPEs inhibited NRF-1 transcription and this suppression was not ablated by concomitant treatment with E2, 4-OHT, or raloxifene, indicating that the effect was not due to inhibition of ER activity. While E2 increased HUVEC proliferation and viability, DEPEs inhibited viability but not proliferation. Resveratrol increased NRF-1 transcription in an ER-dependent manner in HUVECs, and ablated DEPE inhibition of basal NRF-1 expression. Given that NRF-1 is a key nuclear transcription factor regulating genes involved in mitochondrial activity and biogenesis, these data suggest that DEPEs may adversely affect mitochondrial function leading to endothelial dysfunction and resveratrol may block these effects. PMID:22105178
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Intrauterine Growth Restriction Alters Mouse Intestinal Architecture during Development.
Fung, Camille M; White, Jessica R; Brown, Ashley S; Gong, Huiyu; Weitkamp, Jörn-Hendrik; Frey, Mark R; McElroy, Steven J
2016-01-01
Infants with intrauterine growth restriction (IUGR) are at increased risk for neonatal and lifelong morbidities affecting multiple organ systems including the intestinal tract. The underlying mechanisms for the risk to the intestine remain poorly understood. In this study, we tested the hypothesis that IUGR affects the development of goblet and Paneth cell lineages, thus compromising the innate immunity and barrier functions of the epithelium. Using a mouse model of maternal thromboxane A2-analog infusion to elicit maternal hypertension and resultant IUGR, we tested whether IUGR alters ileal maturation and specifically disrupts mucus-producing goblet and antimicrobial-secreting Paneth cell development. We measured body weights, ileal weights and ileal lengths from birth to postnatal day (P) 56. We also determined the abundance of goblet and Paneth cells and their mRNA products, localization of cellular tight junctions, cell proliferation, and apoptosis to interrogate cellular homeostasis. Comparison of the murine findings with human IUGR ileum allowed us to verify observed changes in the mouse were relevant to clinical IUGR. At P14 IUGR mice had decreased ileal lengths, fewer goblet and Paneth cells, reductions in Paneth cell specific mRNAs, and decreased cell proliferation. These findings positively correlated with severity of IUGR. Furthermore, the decrease in murine Paneth cells was also seen in human IUGR ileum. IUGR disrupts the normal trajectory of ileal development, particularly affecting the composition and secretory products of the epithelial surface of the intestine. We speculate that this abnormal intestinal development may constitute an inherent "first hit", rendering IUGR intestine susceptible to further injury, infection, or inflammation.
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Wang, Yuan-Hsi; Twu, Yuh-Ching; Wang, Chung-Kwe; Lin, Fu-Zhen; Lee, Chun-Ya; Liao, Yi-Jen
2018-06-05
Liver fibrosis is the first step toward the progression to cirrhosis, portal hypertension, and hepatocellular carcinoma. A high-cholesterol diet is associated with liver fibrosis via the accumulation of free cholesterol in hepatic stellate cells (HSCs). Niemann-Pick type C2 (NPC2) plays an important role in the regulation of intracellular free cholesterol homeostasis via direct binding with free cholesterol. Previously, we reported that NPC2 was downregulated in liver cirrhosis tissues. Loss of NPC2 enhanced the accumulation of free cholesterol in HSCs and made them more susceptible to transforming growth factor (TGF)-β1. In this study, we showed that knockdown of NPC2 resulted in marked increases in platelet-derived growth factor BB (PDGF-BB)-induced HSC proliferation through enhanced extracellular signal-regulated kinases (ERK), p38, c-Jun N-terminal kinases (JNK), and protein kinase B (AKT) phosphorylation. In contrast, NPC2 overexpression decreased PDGF-BB-induced cell proliferation by inhibiting p38, JNK, and AKT phosphorylation. Although NPC2 expression did not affect caspase-related apoptosis, the autophagy marker light chain 3β (LC3B) was decreased in NPC2 knockdown, and free cholesterol accumulated in the HSCs. The mitochondrial respiration functions (such as oxygen consumption rate, ATP production, and maximal respiratory capacity) were decreased in NPC2 knockdown, and free cholesterol accumulated in the HSCs, while NPC2-overexpressed cells remained normal. In addition, NPC2 expression did not affect the susceptibility of HSCs to lipopolysaccharides (LPS), and U18666A treatment induced free cholesterol accumulation, which enhanced LPS-induced Toll-like receptor 4 (TLR4), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 phosphorylation, interleukin (IL)-1 and IL-6 expression. Our study demonstrated that NPC2-mediated free cholesterol homeostasis controls HSC proliferation and mitochondrial function.
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Théon, A P; Lecouteur, R A; Carr, E A; Griffey, S M
2000-03-01
To assess the influence of tumor cell proliferation and sex-hormone receptors on the efficacy of megavoltage irradiation for dogs with incompletely resected meningiomas. Longitudinal clinical trial. 20 dogs with incompletely resected intracranial meningiomas. Dogs were treated with 48 Gy of radiation administered 3 times per week on an alternate-day schedule of 4 Gy/fraction for 4 weeks, using bilateral parallel-opposed fields. Tumor proliferative fraction measured by immunohistochemical detection of proliferating cell nuclear antigen (PFPCNA index) ranged from 10 to 42% (median, 24%). Progesterone receptor immunoreactivity was detected in 70% of tumors. Estrogen receptor immunoreactivity was not detected. An inverse correlation was found between detection of progesterone receptors and the PFPCNA index. The overall 2-year progression-free survival (PFS) rate was 68%. The only prognostic factor that significantly affected PFS rate was the PFPCNA index. The 2-year PFS was 42% for tumors with a high PFPCNA index (value > or = 24%) and 91% for tumors with a low PFPCNA index (value < 24%). Tumors with a high PFPCNA index were 9.1 times as likely to recur as were tumors with a low PFPCNA index. This study confirms the value of irradiation for dogs with incompletely resected meningiomas. Prognostic value of the PFPCNA index suggests-that duration of treatment and interval from surgery to start of irradiation may affect outcome. Loss of progesterone receptors in some tumors may be responsible for an increase in PFPCNA index and may indirectly affect prognosis after radiation therapy.
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Tong, Jie; Sun, Lijun; Zhu, Bin; Fan, Yun; Ma, Xingfeng; Yu, Liyin; Zhang, Jianbao
2017-10-01
Pulsed electromagnetic fields (PEMF) can be used to treat bone-related diseases, but the underlying mechanism remains unclear, especially the process by which PEMFs initiate biological effects. In this study, we demonstrated the effects of PEMF on proliferation and differentiation of osteoblasts using the model of calcium transients induced by high extracellular calcium. Our results showed that PEMF can increase both the percentage of responding cells and amplitude of intracellular calcium transients induced by high extracellular calcium stimulation. Compared with corresponding extracellular calcium levels, PEMF stimulation increased proliferation and differentiation of osteoblasts and related gene expressions, such as insulin-like growth factor 1 (IGF-1), alkaline phosphatase (ALP), runt-related transcription factor 2 (Runx2), and osteocalcin (OCN), which can be completely abolished by BAPTA-AM. Moreover, PEMF did not affect proliferation and differentiation of osteoblasts if no intracellular calcium transient was present in osteoblasts during PEMF exposure. Our results revealed that PEMF affects osteoblast proliferation and differentiation through enhanced intracellular calcium transients, which provided a cue to treat bone-related diseases with PEMF. Bioelectromagnetics. 38:541-549, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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Marsch-Martinez, Nayelli; Greco, Raffaella; Becker, Jörg D; Dixit, Shital; Bergervoet, Jan H W; Karaba, Aarati; de Folter, Stefan; Pereira, Andy
2006-12-01
The BOLITA (BOL) gene, an AP2/ERF transcription factor, was characterized with the help of an activation tag mutant and overexpression lines in Arabidopsis and tobacco. The leaf size of plants overexpressing BOL was smaller than wild type plants due to a reduction in both cell size and cell number. Moreover, severe overexpressors showed ectopic callus formation in roots. Accordingly, global gene expression analysis using the overexpression mutant reflected the alterations in cell proliferation, differentiation and growth through expression changes in RBR, CYCD, and TCP genes, as well as genes involved in cell expansion (i.e. expansins and the actin remodeling factor ADF5). Furthermore, the expression of hormone signaling (i.e. auxin and cytokinin), biosynthesis (i.e. ethylene and jasmonic acid) and regulatory genes was found to be perturbed in bol-D mutant leaves.
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Metabolic Features of Multiple Myeloma.
El Arfani, Chaima; De Veirman, Kim; Maes, Ken; De Bruyne, Elke; Menu, Eline
2018-04-14
Cancer is known for its cellular changes contributing to tumour growth and cell proliferation. As part of these changes, metabolic rearrangements are identified in several cancers, including multiple myeloma (MM), which is a condition whereby malignant plasma cells accumulate in the bone marrow (BM). These metabolic changes consist of generation, inhibition and accumulation of metabolites and metabolic shifts in MM cells. Changes in the BM micro-environment could be the reason for such adjustments. Enhancement of glycolysis and glutaminolysis is found in MM cells compared to healthy cells. Metabolites and enzymes can be upregulated or downregulated and play a crucial role in drug resistance. Therefore, this review will focus on changes in glucose and glutamine metabolism linked with the emergence of drug resistance. Moreover, metabolites do not only affect other metabolic components to benefit cancer development; they also interfere with transcription factors involved in proliferation and apoptotic regulation.
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VEGF promotes tumorigenesis and angiogenesis of human glioblastoma stem cells
DOE Office of Scientific and Technical Information (OSTI.GOV)
Oka, Naoki; Soeda, Akio; Inagaki, Akihito
2007-08-31
There is increasing evidence for the presence of cancer stem cells (CSCs) in malignant brain tumors, and these CSCs may play a pivotal role in tumor initiation, growth, and recurrence. Vascular endothelial growth factor (VEGF) promotes the proliferation of vascular endothelial cells (VECs) and the neurogenesis of neural stem cells. Using CSCs derived from human glioblastomas and a retrovirus expressing VEGF, we examined the effects of VEGF on the properties of CSCs in vitro and in vivo. Although VEGF did not affect the property of CSCs in vitro, the injection of mouse brains with VEGF-expressing CSCs led to the massivemore » expansion of vascular-rich GBM, tumor-associated hemorrhage, and high morbidity, suggesting that VEGF promoted tumorigenesis via angiogenesis. These results revealed that VEGF induced the proliferation of VEC in the vascular-rich tumor environment, the so-called stem cell niche.« less
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Perucca, Paola; Mocchi, Roberto; Guardamagna, Isabella; Bassi, Elisabetta; Sommatis, Sabrina; Nardo, Tiziana; Prosperi, Ennio; Stivala, Lucia Anna; Cazzalini, Ornella
2018-06-01
In mammalian cells, Nucleotide Excision Repair (NER) plays a role in removing DNA damage induced by UV radiation. In Global Genome-NER subpathway, DDB2 protein forms a complex with DDB1 (UV-DDB), recognizing photolesions. During DNA repair, DDB2 interacts directly with PCNA through a conserved region in N-terminal tail and this interaction is important for DDB2 degradation. In this work, we sought to investigate the role of DDB2-PCNA association in DNA repair and cell proliferation after UV-induced DNA damage. To this end, stable clones expressing DDB2 Wt and DDB2 PCNA- were used. We have found that cells expressing a mutant DDB2 show inefficient photolesions removal, and a concomitant lack of binding to damaged DNA in vitro. Unexpected cellular behaviour after DNA damage, such as UV-resistance, increased cell growth and motility were found in DDB2 PCNA- stable cell clones, in which the most significant defects in cell cycle checkpoint were observed, suggesting a role in the new cellular phenotype. Based on these findings, we propose that DDB2-PCNA interaction may contribute to a correct DNA damage response for maintaining genome integrity. Copyright © 2018 Elsevier B.V. All rights reserved.
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Gandolfi, Maria Giovanna; Ciapetti, Gabriela; Taddei, Paola; Perut, Francesca; Tinti, Anna; Cardoso, Marcio Vivan; Van Meerbeek, Bart; Prati, Carlo
2010-10-01
The effect of ageing in phosphate-containing solution of bioactive calcium-silicate cements on the chemistry, morphology and topography of the surface, as well as on in vitro human marrow stromal cells viability and proliferation was investigated. A calcium-silicate cement (wTC) mainly based on dicalcium-silicate and tricalcium-silicate was prepared. Alpha-TCP was added to wTC to obtain wTC-TCP. Bismuth oxide was inserted in wTC to prepare a radiopaque cement (wTC-Bi). A commercial calcium-silicate cement (ProRoot MTA) was tested as control. Cement disks were aged in DPBS for 5 h ('fresh samples'), 14 and 28 days, and analyzed by ESEM/EDX, SEM/EDX, ATR-FTIR, micro-Raman techniques and scanning white-light interferometry. Proliferation, LDH release, ALP activity and collagen production of human marrow stromal cells (MSC) seeded for 1-28 days on the cements were evaluated. Fresh samples exposed a surface mainly composed of calcium-silicate hydrates CSH (from the hydration of belite and alite), calcium hydroxide, calcium carbonate, and ettringite. Apatite nano-spherulites rapidly precipitated on cement surfaces within 5 h. On wTC-TCP the Ca-P deposits appeared thicker than on the other cements. Aged cements showed an irregular porous calcium-phosphate (Ca-P) coating, formed by aggregated apatite spherulites with interspersed calcite crystals. All the experimental cements exerted no acute toxicity in the cell assay system and allowed cell growth. Using biochemical results, the scores were: fresh cements>aged cements for cell proliferation and ALP activity (except for wTC-Bi), whereas fresh cements
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Metabolite profiling of microfluidic cell culture conditions for droplet based screening.
Bjork, Sara M; Sjostrom, Staffan L; Andersson-Svahn, Helene; Joensson, Haakan N
2015-07-01
We investigate the impact of droplet culture conditions on cell metabolic state by determining key metabolite concentrations in S. cerevisiae cultures in different microfluidic droplet culture formats. Control of culture conditions is critical for single cell/clone screening in droplets, such as directed evolution of yeast, as cell metabolic state directly affects production yields from cell factories. Here, we analyze glucose, pyruvate, ethanol, and glycerol, central metabolites in yeast glucose dissimilation to establish culture formats for screening of respiring as well as fermenting yeast. Metabolite profiling provides a more nuanced estimate of cell state compared to proliferation studies alone. We show that the choice of droplet incubation format impacts cell proliferation and metabolite production. The standard syringe incubation of droplets exhibited metabolite profiles similar to oxygen limited cultures, whereas the metabolite profiles of cells cultured in the alternative wide tube droplet incubation format resemble those from aerobic culture. Furthermore, we demonstrate retained droplet stability and size in the new better oxygenated droplet incubation format.
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Raimondo, Stefania; Naselli, Flores; Fontana, Simona; Monteleone, Francesca; Lo Dico, Alessia; Saieva, Laura; Zito, Giovanni; Flugy, Anna; Manno, Mauro; Di Bella, Maria Antonietta; De Leo, Giacomo; Alessandro, Riccardo
2015-01-01
Nanosized vesicles are considered key players in cell to cell communication, thus influencing physiological and pathological processes, including cancer. Nanovesicles have also been found in edible-plants and have shown therapeutic activity in inflammatory bowel diseases; however information on their role in affecting cancer progression is missing. Our study identify for the first time a fraction of vesicles from lemon juice (Citrus limon L.), obtained as a result of different ultracentrifugation, with density ranging from 1,15 to 1,19 g/ml and specific proteomic profile. By using an in vitro approach, we show that isolated nanovesicles inhibit cancer cell proliferation in different tumor cell lines, by activating a TRAIL-mediated apoptotic cell death. Furthermore, we demonstrate that lemon nanovesicles suppress CML tumor growth in vivo by specifically reaching tumor site and by activating TRAIL-mediated apoptotic cell processes. Overall, this study suggests the possible use of plant-edible nanovesicles as a feasible approach in cancer treatment. PMID:26098775
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Zhu, Wenjun; Tan, Yuanyuan; Qiu, Qihong; Li, Xiting; Huang, Zixian; Fu, Yun; Liang, Min
2013-12-01
Periodontal ligament stem cells (PDLSCs) can be used in periodontal regeneration. Tumour necrosis factor-alpha (TNF-α) participates in the regulation of cell proliferation, apoptosis, differentiation, and migration. However, whether TNF-α can affect the biological features of PDLSCs is still unclear. The objective of this study was to illustrate the biological effects (proliferation, apoptosis, osteogenesis and migration) of TNF-α on human CD146 positive periodontal ligament cells (CD146+PLDCs) and CD146 negative periodontal ligament cells (CD146-PDLCs). CD146±PDLCs were isolated from human PDLCs and analyzed using a fluorescence-activated cell sorter. The biological effects of TNF-α on CD146±PDLCs were evaluated by CCK-8 assay (proliferation), DAPI staining (apoptosis), alizarin red staining and alkaline phosphatase activities assay (osteogenesis), and wounding assay and transwell assay (migration). CD146+PDLCs, which expressed MSC surface markers CD105, CD90, CD73, CD44, and Stro-1, showed higher proliferative and osteogenic potential than CD146-PDLCs. TNF-α at a dose of 2.5ng/ml was found to enhance both proliferation and osteogenesis in CD146+PDLCs. At 5ng/ml, TNF-α promoted proliferation, osteogenesis, and apoptosis in CD146+PDLCs and enhanced osteogenesis in CD146-PDLCs. At 10ng/ml, TNF-α only aggravated apoptosis in CD146+PDLCs. The migratory ability of both CD146+PDLCs and CD146-PDLCs was not altered by TNF-α. CD146+PDLCs were subpopulation of MSC. It showed greater proliferative and osteogenic potential than CD146-PDLCs. At low concentration, TNF-α was beneficial to CD146+PDLCs on proliferation and osteogenesis, and at high concentration it was detrimental. CD146-PDLCs were found to be less sensitive to TNF-α. Copyright © 2013 Elsevier Ltd. All rights reserved.
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FGF signaling supports Drosophila fertility by regulating development of ovarian muscle tissues
Irizarry, Jihyun; Stathopoulos, Angelike
2015-01-01
The thisbe (ths) gene encodes a Drosophila fibroblast growth factor (FGF), and mutant females are viable but sterile suggesting a link between FGF signaling and fertility. Ovaries exhibit abnormal morphology including lack of epithelial sheaths, muscle tissues that surround ovarioles. Here we investigated how FGF influences Drosophila ovary morphogenesis and identified several roles. Heartless (Htl) FGF receptor was found expressed within somatic cells at the larval and pupal stages, and phenotypes were uncovered using RNAi. Differentiation of terminal filament cells was affected, but this effect did not alter ovariole number. In addition, proliferation of epithelial sheath progenitors, the apical cells, was decreased in both htl and ths mutants, while ectopic expression of the Ths ligand led to these cells’ over-proliferation suggesting that FGF signaling supports ovarian muscle sheath formation by controlling apical cell number in the developing gonad. Additionally, live imaging of adult ovaries was used to show that htl RNAi mutants, hypomorphic mutants in which epithelial sheaths are present, exhibit abnormal muscle contractions. Collectively, our results demonstrate that proper formation of ovarian muscle tissues is regulated by FGF signaling in the larval and pupal stages through control of apical cell proliferation and is required to support fertility. PMID:25958090
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Potassium Channels in Regulation of Vascular Smooth Muscle Contraction and Growth
Jackson, William F.
2017-01-01
Potassium channels importantly contribute to the regulation of vascular smooth muscle (VSM) contraction and growth. They are the dominant ion conductance of the VSM cell membrane and importantly determine and regulate membrane potential. Membrane potential, in turn, regulates the open-state probability of voltage-gated Ca2+ channels (VGCC), Ca2+ influx through VGCC, intracellular Ca2+ and VSM contraction. Membrane potential also affects release of Ca2+ from internal stores and the Ca2+ sensitivity of the contractile machinery such that K+ channels participate in all aspects of regulation of VSM contraction. Potassium channels also regulate proliferation of VSM cells through membrane potential-dependent and membrane potential-independent mechanisms. Vascular smooth muscle cells express multiple isoforms of at least five classes of K+ channels contribute to the regulation of contraction and cell proliferation (growth). This review will examine the structure, expression and function of large-conductance, Ca2+-activated K+ (BKCa) channels, intermediate-conductance Ca2+-activated K+ (KCa3.1) channels, multiple isoforms of voltage-gated K+ (KV) channels, ATP-sensitive K+ (KATP) channels, and inward-rectifier K+ (KIR) channels in both contractile and proliferating VSM cells. PMID:28212804
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Okamoto, Michio; Tanaka, Hiroyuki, E-mail: tanahiro-osk@umin.ac.jp; Okada, Kiyoshi
2014-01-17
Highlights: •Methylcobalamin activated the Erk1/2 signaling pathway in C2C12 cells. •Methylcobalamin promoted the proliferation and migration in C2C12 cells. •C2C12 cell apoptosis during differentiation was inhibited by methylcobalamin. -- Abstract: Methylcobalamin (MeCbl) is a vitamin B12 analog that has some positive effects on peripheral nervous disorders. Although some previous studies revealed the effects of MeCbl on neurons, its effect on the muscle, which is the final target of motoneuron axons, remains to be elucidated. This study aimed to determine the effect of MeCbl on the muscle. We found that MeCbl promoted the proliferation and migration of C2C12 myoblasts in vitromore » and that these effects are mediated by the Erk1/2 signaling pathway without affecting the activity of the Akt signaling pathway. We also demonstrated that MeCbl inhibits C2C12 cell apoptosis during differentiation. Our results suggest that MeCbl has beneficial effects on the muscle in vitro. MeCbl administration may provide a novel therapeutic approach for muscle injury or degenerating muscle after denervation.« less
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García, Celina; Nuñez-Anita, Rosa Elvira; Thebault, Stéphanie; Arredondo Zamarripa, David; Jeziorsky, Michael C; Martínez de la Escalera, Gonzalo; Clapp, Carmen
2014-03-01
Endothelial nitric oxide synthase (eNOS)-derived nitric oxide is a major vasorelaxing factor and a mediator of vasopermeability and angiogenesis. Vasoinhibins, a family of antiangiogenic prolactin fragments that include 16 K prolactin, block most eNOS-mediated vascular effects. Vasoinhibins activate protein phosphatase 2A, causing eNOS inactivation through dephosphorylation of eNOS at serine residue 1179 in bovine endothelial cells and thereby blocking vascular permeability. In this study, we examined whether human eNOS phosphorylation at S1177 (analogous to bovine S1179) influences other actions of vasoinhibins. Bovine umbilical vein endothelial cells were stably transfected with human wild-type eNOS (WT) or with phospho-mimetic (S1177D) or non-phosphorylatable (S1177A) eNOS mutants. Vasoinhibins inhibited the increases in eNOS activity, migration, and proliferation following the overexpression of WT eNOS but did not affect these responses in cells expressing S1177D and S1177A eNOS mutants. We conclude that eNOS inhibition by dephosphorylation of S1177 is fundamental for the inhibition of endothelial cell migration and proliferation by vasoinhibins.
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Arumuggam, Niroshaathevi; Melong, Nicole; Too, Catherine Kl; Berman, Jason N; Rupasinghe, Hp Vasantha
2017-01-01
The overall clinical outcome in T-cell acute lymphoblastic leukemia (T-ALL) can be improved by minimizing risk for treatment failure using effective pharmacological adjuvants. Phloridzin (PZ), a flavonoid precursor found in apple peels, was acylated with docosahexaenoic acid (DHA) yielding a novel ester known as phloridzin docosahexaenoate (PZ-DHA). Here, we have studied the cytotoxic effects of PZ-DHA on human leukemia cells using in vitro and in vivo models. The inhibitory effects of PZ-DHA were tested on human Jurkat T-ALL cells in comparison to K562 chronic myeloid leukemia (CML) cells and non-malignant murine T-cells. PZ-DHA, not PZ or DHA alone, reduced cell viability and ATP levels, increased intracellular LDH release, and caused extensive morphological alterations in both Jurkat and K562 cells. PZ-DHA also inhibited cell proliferation, and selectively induced apoptosis in Jurkat and K562 cells while sparing normal murine T-cells. The cytotoxic effects of PZ-DHA on Jurkat cells were associated with caspase activation, DNA fragmentation, and selective down-regulation of STAT3 phosphorylation. PZ-DHA significantly inhibited Jurkat cell proliferation in zebrafish larvae; however, the proliferation of K562 cells was not affected in vivo . We propose that PZ-DHA-induced cytotoxic response is selective towards T-ALL in the presence of a tumor-stromal microenvironment. Prospective studies evaluating the combinatorial effects of PZ-DHA with conventional chemotherapy for T-ALL are underway.
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Arumuggam, Niroshaathevi; Melong, Nicole; Too, Catherine KL; Berman, Jason N; Rupasinghe, HP Vasantha
2017-01-01
The overall clinical outcome in T-cell acute lymphoblastic leukemia (T-ALL) can be improved by minimizing risk for treatment failure using effective pharmacological adjuvants. Phloridzin (PZ), a flavonoid precursor found in apple peels, was acylated with docosahexaenoic acid (DHA) yielding a novel ester known as phloridzin docosahexaenoate (PZ-DHA). Here, we have studied the cytotoxic effects of PZ-DHA on human leukemia cells using in vitro and in vivo models. The inhibitory effects of PZ-DHA were tested on human Jurkat T-ALL cells in comparison to K562 chronic myeloid leukemia (CML) cells and non-malignant murine T-cells. PZ-DHA, not PZ or DHA alone, reduced cell viability and ATP levels, increased intracellular LDH release, and caused extensive morphological alterations in both Jurkat and K562 cells. PZ-DHA also inhibited cell proliferation, and selectively induced apoptosis in Jurkat and K562 cells while sparing normal murine T-cells. The cytotoxic effects of PZ-DHA on Jurkat cells were associated with caspase activation, DNA fragmentation, and selective down-regulation of STAT3 phosphorylation. PZ-DHA significantly inhibited Jurkat cell proliferation in zebrafish larvae; however, the proliferation of K562 cells was not affected in vivo. We propose that PZ-DHA-induced cytotoxic response is selective towards T-ALL in the presence of a tumor-stromal microenvironment. Prospective studies evaluating the combinatorial effects of PZ-DHA with conventional chemotherapy for T-ALL are underway. PMID:29312799