Prostaglandin E1 protects hepatocytes against endoplasmic reticulum stress-induced apoptosis via protein kinase A-dependent induction of glucose-regulated protein 78 expression

AIM To investigate the protective effect of prostaglandin E1(PGE1) against endoplasmic reticulum(ER) stressinduced hepatocyte apoptosis, and to explore its underlying mechanisms.METHODS Thapsigargin(TG) was used to induce ER stress in the human hepatic cell line L02 and hepatocarcinomaderived cell line Hep G2. To evaluate the effects of PGE1 on TG-induced apoptosis, PGE1 was used an hour prior to TG treatment. Activation of unfolded protein response signaling pathways were detected by western blotting and quantitative real-time RTPCR. Apoptotic index and cell viability of L02 cells and Hep G2 cells were determined with flow cytometry and MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2 H-tetrazolium] assay. RESULTS Pretreatment with 1 μmol/L PGE1 protected against TG-induced apoptosis in both L02 cells and Hep G2 cells. PGE1 enhanced the TG-induced expression of C/EBP homologous protein(CHOP), glucose-regulated protein(GRP) 78 and spliced X box-binding protein 1 at 6 h. However, it attenuated their expressions after 24 h. PGE1 alone induced protein and m RNA expressions of GRP78; PGE1 also induced protein expression of DNA damage-inducible gene 34 and inhibited the expressions of phospho-PKR-like ER kinase, phosphoeukaryotic initiation factor 2α and CHOP. Treatment with protein kinase A(PKA)-inhibitor H89 or KT5720 blocked PGE1-induced up-regulation of GRP78. Further, the cytoprotective effect of PGE1 on hepatocytes was not observed after blockade of GRP78 expression by H89 or small interfering RNA specifically targeted against human GRP78.CONCLUSION Our study demonstrates that PGE1 protects against ER stress-induced hepatocyte apoptosis via PKA pathwaydependent induction of GRP78 expression.     

Experimental study on the apoptosis of cervical cancer Hela cells induced by juglone through c-Jun N-terminal kinase/c-Jun pathway

Objective: To study the regulatory effect and molecular mechanism of juglone on apoptosis of cervical cancer Hela cells. Methods: Cervical cancer Hela cells were cultured and treated with different dosages of juglone(10, 20, and 40 μmol/L, respectively) and c-Jun N-terminal kinase(JNK) inhibitor SP600125(10, 20, and 40 μmol/L, respectively). Then cellular proliferative activity and the expression of JNK/c-Jun pathway molecule and apoptotic molecule in the cells were detected. Results: After 6, 12, 18 and 24 h of treatment, the value for proliferative activity of cells treated with juglone was significantly lower than that of control group(P0.05), and the anti-proliferative effect was more significant as the treatment period and juglone dosage increased(P0.05). The protein expressions of Bax, Cyt C, Fas, Fas L, Caspase-3, p-JNK and p-c-Jun in cells treated with juglone were significantly higher than those of control group(P0.05), and the protein expressions of Bax, Cyt C, Fas, Fas L, Caspase-3, p-JNK and p-c-Jun increased more remarkably as the juglone dosage increased(P0.05). In cells treated with 40 μmol/L juglone and SP600125, the protein expressions of Bax, Cyt C, Fas, Fas L and Caspase-3 were significantly lower than those of cells treated with 40 μmol/L juglone(P0.05), and the protein expressions of Bax, Cyt C, Fas, Fas L and Caspase-3 reduced more remarkably as the SP600125 dosage increased(P0.05). Conclusion: Juglone can increase the expression of apoptotic molecules in mitochondrial pathway and death receptor pathway by activating JNK/c-Jun pathway, thus inducing apoptosis of cervical cancer cells.     

Salubrinal protects against tunicamycin and hypoxia induced cardiomyocyte apoptosis via the PERK-eIF2α signaling pathway

Objectives This study examined the protective effect of salubrinal and the mechanism underlying this protection on tunicamycin (TM)- and hypoxia-induced apoptosis in rat cardiomyocytes. Methods Neonatal rat cardiomyocytes were cultured from the ventricles of 1-day-old Wistar rats. Cells were exposed to different concentrations of salubrinal (10, 20, and 40 μmol/L) for 30 minutes followed by TM treatment or hypoxia for 36 hours. Apoptosis was measured by a multiparameter HCS (high content screening) apoptosis assay, TUNEL assay and flow cytometry. The phosphorylation of eukaryotic translation initiation factor 2 subunit alpha (p-eIF2α) and the expression of cleaved caspase-12 were determined by western blotting. C/EBP homologous protein (CHOP) was detected by immunocytochemistry. Results HCS, TUNEL assays and flow cytometry showed that salubrinal protected against apoptosis induced by TM or hypoxia. Western blotting showed that salubrinal protected cardiomyocytes against apoptosis by inducing eIF2α phosphorylation and down-regulating the expression of the endoplasmic reticulum stress-mediated apoptotic proteins, CHOP and cleaved caspase-12. Conclusions Our study suggests that salubrinal protects rat cardiomyocytes against TM- or hypoxia-associated apoptosis via a mechanism involving the inhibition of ER stress-mediated apoptosis.     

Synergistic effect of cell differential agent-Ⅱ and arsenic trioxide on induction of cell cycle arrest and apoptosis in hepatoma cells

AIM: To illustrate the possible role of cell differential agent-Ⅱ (CDA-Ⅱ) in the apoptosis of hepatoma cells induced byarsenic trioxide (As2O3).METHODS: Hepatoma cell lines BEL-7402 and HepG2 weretreated with As2O3 together with CDA-Ⅱ. Cell survivingfraction was determined by MTT assay; morphologicalchanges were observed by immunofluorescence staining ofHoechst 33 258; and cell cycle and the apoptosis index weredetermined by flow cytometry (FCM).RESULTS: Cytotoxity of CDA-Ⅱ was low. Nevertheless, CDA-Ⅱ could strongly potentiate arsenic trioxide-inducedapoptosis. At 1.0 g/L CDA-Ⅱ, IC50 of As2O3 in hepatoma celllines was reduced from 5.0 μmol/L to 1.0 μmol/L (P＜0.01).The potentiation of apoptosis was dependent on the dosageof CDA-Ⅱ. FCM indicated that in hepatoma, cell growth wasinhibited by CDA-Ⅱ at lower concentrations (＜2.0 g/L)primarily by arresting at S and G2 phase, and at higherconcentrations (＞2.0 g/L) apoptotic cell and cell cyclearresting at G1 phaseincreased proportionally. Thecombination of two drugs led to much higher apoptotic rates,as compared with the either drug used alone.CONCLUSION: CDA-Ⅱ can strongly potentiate As2O3-induced apoptosis in hepatoma cells, and two drugs canproduce a significant synergic effect.     

Effects of epidermal growth factor and its signal transduction inhibitors on apoptosis in human colorectal cancer cells

AIM:The study investigated if EGF signaling inhibitors,EGFantibody and tyrphostin 51 (a tyrosine kinase inhibitor),mediated the action of EGF on apoptosis and the expressionof EGF receptors and p21 (a cyclin-dependent kinaseinhibitor) of human colorectal cancer cells.METHODS:Human colorectal adenocarcinoma cells(SW480) were incubated with 0.6mL/L dimethyl sulfoxide(DNSO,the control group),225ng/mL (37.5nmol/L) EGFin 0.6mL/L DNSO,225ng/mL EGF 2.5μg/mL (17nmol/L)EGF antibody in 0.6mL/L DMSO,or 225ng/mL EGF 215ng/mL (0.8μmol/L) tyrphostin 51 in 0.6mL/L DMSO.RESULTS:After 48h incubation,the levels of EGF in mediumsignificantly increased (P<0.05) in the EGF-treated groups.The numbers of apoptotic cells were significantly fewer(P<0.05) in the EGF EGF antibody and EGF tyrphostin51 groups than those in the control and EGF groups after12 h treatments.The expression of phosphorylated EGFreceptors in the EGF,EGF EGF antibody,and EGF tyrphostin 51 groups was 176.8%,62.4%,and 138.1% ofthe control group,respectively.The expression of p21protein in the EGF,EGF EGF antibody,and EGF tyrphostin51 groups was 115.7%,4.8%,and 61.5% of the controlgroup,respectively.CONCLUSION:The data suggest that EGF antibody andtyrphostin 51 can inhibit the action of EGF on apoptosis inhuman colorectal cancer cells through down-regulation ofEGF receptor and p21 expression.     

Role of mitochondrial dysfunction in hydrogen peroxide-induced apoptosis of intestinal epithelial cells

AIM: To study the role of mitochondrial dysfunction in hydrogen peroxide-induced apoptosis of intestinal epithelial cells.METHODS: Hydrogen peroxide-induced apoptosis of human intestinal epithelial cell line SW-480 was established. Cell apoptosis was determined by Annexin-V and PI doublestained flow cytometry and DNA gel electrophoresis.Morphological changes were examined with light and electron microscopy. For other observations, mitochondrial function,cytochrome c release, mitochondrial translocation and membrane potential were determined simultaneously.RESULTS: Percentage of apoptotic cells induced with 400μmol/L hydrogen peroxide increased significantly at 1 h or 3 h after stimulation and recovered rapidly. Meanwhile percentage of apoptotic cells induced with 4 mmol/L hydrogen peroxide increased with time. In accordance with these changes, we observed decreased mitochondrial function in 400μmol/L H2O2-stimualted cells at 1 h or 3 h and in 4 mmol/L H2O2-stimualted cells at times examined.Correspondingly, swelling cristae and vacuole-like mitochondria were noted. Release of cytochrome c,decreased mitochondrial membrane potential and mitochondrial translocation were also found to be the early signs of apoptosis.CONCLUSION: Dysfunctional mitochondria play a role in the apoptosis of SW-480 cell line inline induced by hydrogen peroxide.     

Schistosoma japonicum attenuates dextran sodium sulfateinduced colitis in mice via reduction of endoplasmic reticulum stress

AIM To elucidate the impact of Schistosoma(S.) japonicum infection on inflammatory bowel disease by studying the effects of exposure to S. japonicum cercariae on dextran sodium sulfate(DSS)-induced colitis. METHODS Infection was percutaneously established with 20 ± 2 cercariae of S. japonicum, and colitis was induced by administration of 3% DSS at 4 wk post infection. Weight change, colon length, histological score(HS) and disease activity index(DAI) were evaluated. Inflammatory cytokines, such as IL-2, IL-10 and IFN-γ, were tested by a cytometric bead array and real-time quantitative polymerase chain reaction(RT-PCR). Protein and m RNA levels of IRE1α, IRE1β, GRP78, CHOP, P65, P-P65, P-IκBα and IκBα in colon tissues were examined by Western blot and RT-PCR, respectively. Terminal deoxynucleotidyl transferase-mediated d UTP nick-end labeling positive cells, cleaved-caspase 3 expression and Bcl2/Bax were investigated to assess the apoptosis in colon tissues.RESULTS Mice infected with S. japonicum cercariae were less susceptible to DSS. Mice infected with S. japonicum cercariae and treated with DSS showed decreased weight loss, longer colon, and lower HS and DAI compared with mice treated with DSS alone. A substantial decrease in Th1/Th2/Th17 response was observed after infection with S. japonicum. Endoplasmic reticulum(ER) stress and the nuclear factor-kappa B(NF-κB) pathway were reduced in mice infected with S. japonicum cercariae and treated with DSS, along with ameliorated celluar apoptosis, in contrast to mice treated with DSS alone. CONCLUSION Exposure to S. japonicum attenuated inflammatory response in a DSS-induced colitis model. In addition to the Th1/Th2/Th17 pathway and NF-κB pathway, ER stress was shown to be involved in mitigating inflammation and decreasing apoptosis. Thus, ER stress is a new aspect in elucidating the relationship between helminth infection and inflammatory bowel disease(IBD), which may offer new therapeutic methods for IBD.     

Isoflavone genistein protects high glucose-induced human aortic endothelial cell apoptosis through estrogen receptor-mediated pathway

Objective The aim of this study was to determine if isoflavone genistien has protective effects against high glucose-induced cell apoptosis in human aortic endlthelial cells,and investigate the possible mechanism for this protection.Methods Human aortic endothelial cells subjected to normal (5mmol/L) or high glucose (25mmol/L) were treated with genistein at 0,50,100nmol/L.Parallel experiments were performed with 100nM 17b-estradiol,and also in the presence and absence of the pure anti-estrogen ICI-182,780 (100nmol/L).The effects on cell apoptotic DNA fragmentation were determined using cell death ELISA,and the effects on cellular proliferation were determined using tritiated thymidine incorporation assay.Estrogen receptor expression was detected by Taqman quantitative PCR.Results Genistein at 100nmol/L significantly reduced high glucose-induced DNA fragmentation,and reversed cell DNA synthesis inhibition (P＜0.001) after 24 hours' incubation.The effect of genistein was completely blocked by ICI-182,780administration.Estrogen receptor beta,but not alpha was found to be expressed in these cells.Conclusion Isoflavone genistein shows protection against high glucose-induced cell damage through estrogen receptor beta,reducing apoptotic DNA damage and protecting from the inhibition of cell proliferation.     

Low intensity ultrasound-induced apoptosis in human gastric carcinoma cells

AIM： To investigate the low intensity ultrasound （US）- induced apoptosis in human gastric carcinoma cells and its potential mechanism and to suggest a new therapeutic approach to gastric carcinoma. METHODS： Human SGC-7901 gastric carcinoma cells were cultured in vitro and irradiated by low intensity US for 10 min at different intensities with different incubation times after irradiation. Morphologic changes were examined under microscope with trypan blue staining and then the percentage of early apoptotic cells was detected by flow cytometry （FCM） with double staining of fiuorescein isothiocyanate （FITC）- Annexin V/propidium iodide （PI）. Two-dimensional electrophoresis （2DE） was used to get the protein profile and some proteins differently expressed after US irradiation were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS）. Functional analysis was performed to investigate the mechanism of US-induced cell apoptosis. RESULTS： The percentage of apoptotic cells increased about 10% after US irradiation （12.0 W/cm^2, 12 h culture）, The percentage of early apoptosis and secondary necrosis in the US-irradiated cells increased with the increased US intensity. Moreover, apoptotic cells increased with the increased culture time after US irradiation and reached its maximum at about 12 h.Several new proteins appeared after US irradiation and were up or down regulated more than 2 times. Some heat shock proteins （HSPs） were found to be associated with the signal process simulating the apoptosis of cells. CONCLUSION： Low intensity US could induce apoptosis in human gastric carcinoma cells. US-induced apoptosis is related to US intensity/culture time. US-induced apoptosis may be caspases-dependent and endoplasmic reticulum （ER） stress-triggered apoptosis may also contribute to it. Proteomic experimental system is useful in finding the protein alteration in carcinoma cells after US irradiation, helping to develop a new cancer ther     

Effects of Angiotensin Ⅱ on Expression of the Gap Junction Channel Protein Connexin 43 in Neonatal Rat Ventricular Myocytes

To study the effects of angiotensin Ⅱ,as a mediator of cardiac hypertrophy,on expression of connexin 43 (Cx43) in cultured neonatal rat ventricular myocytes and correlation of expression of Cx43 and cardiomyocyte hypertrophy.Methods Cardiomyocytes were isolated from newborn SD rats.Angiotensin Ⅱ was added into the media to induce myocyte hypertrophy.Cultures were exposed to 10 ～6 mol/L angiotensin Ⅱ for 72 h,Cx43 expression was characterized by RT-PCR and Immunofluorescence methods.Results Immunofluorescence analysis revealed decreased Cx43 immunoreactivity in cells treated for 72 h with angiotensin Ⅱ.RT-PCR analysis demonstrated there was an obvious decrease of Cx43 mRNA level in cells exposed to angiotensin Ⅱ for 72 h.The changes of expression of connexin 43 were related to its entrance into S phase of the cell cycle.Cultured neonatal rat cardiomyocytes were exposed for 72 h to increase concentrations of angiotensin Ⅱ ( 1.0 × 10-9 ～ 1.0 × 10-6mol/L),resulting in significantly decreased Cx43 expression.Conclusions Angiotensin Ⅱ leads to a concentration-dependent decrease in Cx43 protein in cultured neonatal rat ventricular myocytes by decreasing Cx43 mRNA synthesis.Signal transduction pathways activated by angiotensin Ⅱ under pathophysiologic conditions of cardiac hypertrophy could initiate remodeling of gap junctions.     

Inhibition effect of captopril and losartan on expression of matrix metalloproteinase-1,-9 induced by angiotensin Ⅱ in rat vascular smooth muscle cells

Objective To investigate the effect of angiotensin converting enzyme inhibitor (ACEI) captopril and angiotensin Ⅱ receptor antagonist losartan on the mRNA expression of matrix metalloproteinase (MMP)-1 and MMP-9 in vascular smooth muscle cells induced by angiotensin Ⅱ (Ang Ⅱ ).Methods Male Wistar rats' thoracic aortic vascular smooth muscle cells were cultured in vitro.The cultured cells were divided in to control group,Ang Ⅱ group,captopril group,losartan group,and captopril plus losartan group.Cells in all groups were collected at the culture end-point.MMP-1 and MMP-9 mRNA expressions were detected by RT-PCR method in the collected specimens,and the effects of Ang Ⅱ on MMP-1 and MMP-9 mRNA expression and the intervention effects of captopril and losartan were observed in different Ang Ⅱ concentrations and different action times to vascular smooth muscle cells.Results ( 1 ) MMP-1 mRNA expression gradually increased along with the increments of Ang Ⅱ concentration and the action time (P＜0.05),and the most significant concentration was 10-6 mol/L (P＜0.01).(2)Captopril (5 × 10-6 mol/L) and losartan (5 × 10-6mol/L) inhibited the action of AngⅡ (P＜0.05,P＜0.01).MMP-9 mRNA expression was 0.47±0.03 ,0.86 ± 0.04,0.94±0.14 and 1.12±0.19 vs.0.10±0.04 (P＜0.05,P＜0.01) respectively when Ang Ⅱ concentration was 10-7 ,10-6 ,10-5 and 10-4 mol/L respectively.Captopril (5 × 10-6mol/L) and losartan (5 × 10-6 mol/I) significantly inhibited the MMP-9 mRNA expression which was stimulated by Ang Ⅱ (P＜0.05,P＜0.01),especially in captopril plus losartan group.The MMP-9 mRNA expression increased with the prolonging of stimulating time of Ang Ⅱ,MMP-9 mRNA expression was earlier than that of MMP-1.Conclusions AngⅡ increases the expression of MMP-1 and MMP-9 of vascular smooth muscle cells in a dose-and time-dependence manner.Captopril and losartan inhibit the MMP-1 and MMP-9 mRNA expression of vascular smooth muscle cells induced by Ang Ⅱ ,and the inhibition is the strongest when losartan was combined with captopril.The inhibitive effects is positively correlated to action time.     

Inhibition effect of captopril and losartan on expression of matrix metalloproteinase-1,-9 induced by angiotensin Ⅱ in rat vascular smooth muscle cells

Objective To investigate the effect of angiotensin converting enzyme inhibitor (ACEI) captopril and angiotensin Ⅱ receptor antagonist losartan on the mRNA expression of matrix metalloproteinase (MMP)-1 and MMP-9 in vascular smooth muscle cells induced by angiotensin Ⅱ (Ang Ⅱ ).Methods Male Wistar rats' thoracic aortic vascular smooth muscle cells were cultured in vitro.The cultured cells were divided in to control group,Ang Ⅱ group,captopril group,losartan group,and captopril plus losartan group.Cells in all groups were collected at the culture end-point.MMP-1 and MMP-9 mRNA expressions were detected by RT-PCR method in the collected specimens,and the effects of Ang Ⅱ on MMP-1 and MMP-9 mRNA expression and the intervention effects of captopril and losartan were observed in different Ang Ⅱ concentrations and different action times to vascular smooth muscle cells.Results ( 1 ) MMP-1 mRNA expression gradually increased along with the increments of Ang Ⅱ concentration and the action time (P＜0.05),and the most significant concentration was 10-6 mol/L (P＜0.01).(2)Captopril (5 × 10-6 mol/L) and losartan (5 × 10-6mol/L) inhibited the action of AngⅡ (P＜0.05,P＜0.01).MMP-9 mRNA expression was 0.47±0.03 ,0.86 ± 0.04,0.94±0.14 and 1.12±0.19 vs.0.10±0.04 (P＜0.05,P＜0.01) respectively when Ang Ⅱ concentration was 10-7 ,10-6 ,10-5 and 10-4 mol/L respectively.Captopril (5 × 10-6mol/L) and losartan (5 × 10-6 mol/I) significantly inhibited the MMP-9 mRNA expression which was stimulated by Ang Ⅱ (P＜0.05,P＜0.01),especially in captopril plus losartan group.The MMP-9 mRNA expression increased with the prolonging of stimulating time of Ang Ⅱ,MMP-9 mRNA expression was earlier than that of MMP-1.Conclusions AngⅡ increases the expression of MMP-1 and MMP-9 of vascular smooth muscle cells in a dose-and time-dependence manner.Captopril and losartan inhibit the MMP-1 and MMP-9 mRNA expression of vascular smooth muscle cells induced by Ang Ⅱ ,and the inhibition is the strongest when losartan was combined with captopril.The inhibitive effects is positively correlated to action time.     

Inhibition effect of captopril and losartan on expression of matrix metalloproteinase-1,-9 induced by angiotensin Ⅱ in rat vascular smooth muscle cells

Objective To investigate the effect of angiotensin converting enzyme inhibitor (ACEI) captopril and angiotensin Ⅱ receptor antagonist losartan on the mRNA expression of matrix metalloproteinase (MMP)-1 and MMP-9 in vascular smooth muscle cells induced by angiotensin Ⅱ (Ang Ⅱ ).Methods Male Wistar rats' thoracic aortic vascular smooth muscle cells were cultured in vitro.The cultured cells were divided in to control group,Ang Ⅱ group,captopril group,losartan group,and captopril plus losartan group.Cells in all groups were collected at the culture end-point.MMP-1 and MMP-9 mRNA expressions were detected by RT-PCR method in the collected specimens,and the effects of Ang Ⅱ on MMP-1 and MMP-9 mRNA expression and the intervention effects of captopril and losartan were observed in different Ang Ⅱ concentrations and different action times to vascular smooth muscle cells.Results ( 1 ) MMP-1 mRNA expression gradually increased along with the increments of Ang Ⅱ concentration and the action time (P＜0.05),and the most significant concentration was 10-6 mol/L (P＜0.01).(2)Captopril (5 × 10-6 mol/L) and losartan (5 × 10-6mol/L) inhibited the action of AngⅡ (P＜0.05,P＜0.01).MMP-9 mRNA expression was 0.47±0.03 ,0.86 ± 0.04,0.94±0.14 and 1.12±0.19 vs.0.10±0.04 (P＜0.05,P＜0.01) respectively when Ang Ⅱ concentration was 10-7 ,10-6 ,10-5 and 10-4 mol/L respectively.Captopril (5 × 10-6mol/L) and losartan (5 × 10-6 mol/I) significantly inhibited the MMP-9 mRNA expression which was stimulated by Ang Ⅱ (P＜0.05,P＜0.01),especially in captopril plus losartan group.The MMP-9 mRNA expression increased with the prolonging of stimulating time of Ang Ⅱ,MMP-9 mRNA expression was earlier than that of MMP-1.Conclusions AngⅡ increases the expression of MMP-1 and MMP-9 of vascular smooth muscle cells in a dose-and time-dependence manner.Captopril and losartan inhibit the MMP-1 and MMP-9 mRNA expression of vascular smooth muscle cells induced by Ang Ⅱ ,and the inhibition is the strongest when losartan was combined with captopril.The inhibitive effects is positively correlated to action time.     

Transfection of apoptosis related gene Fas ligand in human hepatocellular carcinoma cells and its significance in apoptosis

AIM: To evaluate the expression of apoptosis related gene Fas ligand (FasL) in human hepatocellular carcinoma (HCC) cells HepG2 and its significance in apoptosis. METHODS: Levels of soluble Fas ligand (sFasL) in a group of patients with hepatitis B virus (HBV)-induced chronic hepatitis, HBV-positive liver cirrhosis and HCC were evaluated. In a further study, the recombinant eukaryotic expression plasmid pcDNA3.1hisB-FasL was transfected into HCC cells HepG2 by lipofection, and then soluble FasL was examined in the supernatant of culture cells by EIA, FasL expression in HepG2 cells was detected by immuohistochemistry. After being stained by annexin V and propidium iodine, cells were passed through a flow cytometer and examined by a fluorescence microscope and a laser scanning microscope. RESULTS: The sFasL levels were significantly lower in patients with HCC when compared to the patients with hepatitis or liver cirrhosis. In comparison with untransfected cells, the soluble FasL could be detected in the supernatant of transfected cells. FasL was expressed on the membranes and cytoplasm of transfected cells. The apoptotic cell rate was 36.30% in transfected cells, and was 11.53% in untransfected cells. Moreover, the different stage of apoptotic cells could be distinguished by annexin V and propidium iodine staining. CONCLUSION: Fas ligand is an apoptotic pathway of HCC cells.     

Apoptosis of human primary gastric carcinoma cells induced by genistein

AIM:To investigate the apoptosis in primary gastric cancercells induced by genistein,and the relationship betweenthis apoptosis and expression of bcl-2 and bax.METHODS:MTT assay was used to determine the cell growthinhibitory rate in vitro.Transmission electron microscope andTUNEL staining were used to quantitatively and qualitativelydetect the apoptosis of primary gastric cancer cells beforeand after genistein treatment.Immunohistochemical stainingand RT-PCR were used to detect the expression of apoptosis-associated genes bcl-2 and bax.RESULTS:Genistein inhibited the growth of primary gastriccancer cells in dose-and time-dependent manner.Genisteininduced primary gastric cancer cells to undergo apoptosiswith typically apoptotic characteristics.TUNEL assay showedthat after the treatment of primary gastric cancer cells withgenistein for 24 to 96h,the apoptotic rates of primary gastriccancer cells increased time-dependently.Immunohistochemicalstaining showed that after the treatment of primary gastriccancer cells with genistein for 24 to 96h,the positivity ratesof Bcl-2 proteins were apparently reduced with time andthe positivity rates of Bax proteins were apparently increasedwith time.After exposed to genistein at 20 μmol/L for 24,48,72 and 96 respectively,the density of bcl-2 mRNAdecreased progressively and the density of bax mRNAincreased progressively with elongation of time.CONCLUSION:Genistein is able to induce the apoptosis inprimary gastric cancer cells.This apoptosis may be mediatedby down-regulating the apoptosis- associated bcl-2 geneand up-regulating the expression of apoptosis-associatedbax gene.     

Role of C-terminal Src kinase in angiotensin Ⅱ-induced cytoskeletal rearrangement in glomerular podocytes

<正>Objective To investigate the effects of angiotensinⅡ(AngⅡ)on the expression of C-terminal Src kinase(Csk)in AngⅡ-infused rat model and cultured podocytes,and to explore the role of Csk in AngⅡ-induced cytoskeletal rearrangement of podocytes.Methods Twenty-four Wista rats were randomly subjected to normal sa-