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Objective To investigate the effect of highly expressed β-catenin on the proliferative activity of and expressions of two apoptosis-related genes Bcl-2 and Bax by human skin fibroblasts induced by hydrogen peroxide (H2O2).Methods Normal human skin fibroblasts (HSFs) from child foreskin were divided into three groups:empty vector group transfected with the empty vector pcDNA3.1,H2O2 group transfected with the empty vector pcDNA3.1 followed by treatment with H2O2 (150 μ mol/L) for 2 hours,β-catenin group transfected with pcDNA3.1-β-catenin followed by treatment with H2O2 (150 μ mol/L) for 2 hours.Subsequently,methyl thiazolyl tetrazolium (MTT) assay was conducted to estimate proliferative activity of fibroblasts,flow cytometry to detect cell apoptosis,and reverse transcription (RT)-PCR and Western blot were performed to measure the mRNA and protein expressions of Bcl-2 and Bax respectively.The relative expression levels of genes were expressed as the ratios between the targets and GAPDH.Results Significant differences were found between the empty vector group,H2O2 group and β-catenin group in cellular proliferative activity (expressed as absorbance value at 570 nm:0.792 ± 0.012 vs.0.462 ± 0.012 vs.0.521 ± 0.015,P＜ 0.01) and apoptosis rate (3.407％ ± 0.217％ vs.24.555％ ± 1.793％ vs.15.360％ ± 0.755％,P＜ 0.01).Both mRNA and protein expression levels of Bcl-2 were significantly lower in the H2O2 group (0.333 ± 0.003 and 0.336 ± 0.004 respectively) than in the empty vector group (0.507 ± 0.013 and 0.514 ± 0.021,respectively,both P ＜ 0.01) and β-catenin group (0.404 ± 0.006 and 0.411 ± 0.005,respectively,both P ＜ 0.01).Increased expression levels of Bax mRNA and protein were observed in the H2O2 group compared with the empty vector group and β-catenin group (mRNA:0.451 ± 0.002 vs.0.303 ± 0.005 and 0.339 ± 0.012,protein:0.460 ± 0.008 vs.0.320 ± 0.013 and 0.346 ± 0.013,all P＜ 0.01).Conclusion High expression of β-catenin can raise proliferative activity of aging HSFs.
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Objective To explore the relationship between mitochondrial DNA microsatellite instability(mtMSI) and expression of Bcl 2 and Bax mRNA in gastric cancer and its precancerous lesions. Methods mtMSI and expressions of Bcl 2 and Bax mRNA were detected with PCR SSCP and RT PCR. Results Expressions of Bcl 2 mRNA in intestinal metaplasia (IM,53 3%) and dysplasia (Dys, 70%) were significantly higher than that in normal control(10%, P
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Objective To investigate the effects of gross saponins from Tribulus terrestris L (GSTT) on myocardial apoptosis, and its related gene bcl-2 and bax expression after hypoxia/reoxygenation in cultured myocardial cells of neonatal rat. Methods After myocardial hypoxia/ reoxygenation model was established by culturing primary myocardial cell of neonatal rat in vivo. The myocardial cells were divided into four groups, the sham-operated group, model group, large- and low-dose of GSTT groups. The apoptotic rate of myocardial cells was determined by flow cytometry, and the expression of bcl-2 and bax was detected by using immuno-histochemical method. Results Both large- and low- dose of GSTT could significantly decrease the apoptotic rate (P
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Objective To study the role of 1,4,5-trisphosphate inositol(IP3) and bax gene expression in inhibiting transplanted hepatocellular carcinoma of nude mice by quercetin.Methods After animals with hepatocellular carcinoma were treated with quercetin 1mg/kg/d(ip)for 3 weeks,the volume and weight of tumor was measured,and IP3,Bax mRNA,and Bax protein were assayed by IP3- Birtrak assay,RT-PCR,and Western blotting,respectively;and compard with coutrol group.Results The tumor volume and weight of animals treated with quercetin were lower than those of control[(15.8?10.1) mm3 vs.(52.3?26.5 mm3;(44.8?10.4) mg vs.(91.3?31.4) mg],IP3 content was lower than that of control[(15.9?2.8)pmol/mg protein vs(35.3?6.6)pmol/mg protein],Bax mRNA expression was not significantly different between the 2 groups[RI which was the gray degree multiply area of bax / the gray degree multiply area of ?-actin(0.64?0.12) vs.(0.56?0.15)],Bax mRNA expression was higher in group treated with quercetin than that of control[(3.16?0.95) vs.(1.37?0.48)].Conclusions Quercetin can inhibit growth of transplanted hepatocellular carcinoma of nude mice by reducing IP3 production and bax protein expression.
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AIM:To evaluate the antitumor effect of caffeic acid Ge on U14 tumor bearing mice.METHODS:The tumor inhibitory ratios of caffeic acid Ge on the growth of U14 in mice was observed.Apoptosis morphological transformation of U14 cells induced by caffeic acid Ge was detected by electronic scan microscope and MG-P staining.Alteration of cell cycle was analyzed by flow cytometry.Apoptosis-related protein levels of Bax and Bcl-2 were determined by immunity histochemistry technology.MTT assay was applied to study the antitumor activities of caffeic acid Ge in U14 cell lines in vitro.RESULTS:Tumor inhibitory rates in caffeic-acid Ge groups were 38.50%,47.17% and 64.02%(from low dose to high dose)(P
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AIM:To investigate the protective effects of sodium ferulate(SF)on apoptosis in cultured hippocampal neurons induced by sodium nitroprusside(SNP),and the effect of SF on expression of bcl-2 and bax.METHODS:The primary cultured hippocampal neurons were exposed to 50 ?mol SNP,a nitric oxide-donor,for 24 h after pretreatment with different concentrations of SF(10-160 ?mol/mL)for 6 h.Then neuronal viability was tested by MTT assay.Fluorescent staining with Hoechst 33258 and agarose gel electrophoresis was used to analyze apoptosis.The expressions of bcl-2,bax mRNA and protein were tested by RT-PCR and Western blotting.RESULTS:Pretreatment with SF(10-160 ?mol/L)for 6 h increased the survival rate of neurons.SF prevented the neuronal nuclei from shrinkage,condensation and cleavage and blocked neuronal nuclear DNA fragmentation induced by SNP.SF also increased the expressions of bcl-2 mRNA and Bcl-2 protein and decreased the expressions of bax mRNA and Bax protein.CONCLUSION:SF prevents the cultured hippocampal neurons against SNP neurotoxicity.The mechanism of protection is related to the increase in Bcl-2 level and the decrease in Bax level.As a result,the ratio of Bcl-2/Bax is changed.
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Objective To explore the roles of Bcl-2, Bax, Bcl-xl proteins in the carcinogenesis of basal cell carcinoma(BCC). Methods Forty-six BCC cases were divided into 2 groups (21 in non-aggressive BCC group and 26 in aggressive BCC group) according to their pathological features. Bcl-2, Bax and Bcl-xl were evaluated in formalin fixed, paraffin embedded specimens by immunohistochemical methods and analyzed by image-analysis methods to obtain average optical density (AOD) of each protein for each case. The AOD data of 3 proteins and Bcl-2/Bax ratios were then compared between the 2 BCC groups. Results The level of Bcl-2 expression and Bcl-2/Bax ratio of non-aggressive BCC group were higher than those of aggressive group,(P=0.005 and 0.044, respectively). For the expression level of Bcl-xl and Bax, there was no statistically difference between the two groups (P=0.097 and 0.979, respectively). Conclusion Bcl-2, Bax,Bcl-xl proteins might participate in the carcinogenesis and development of BCC.
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AIM:To study the senescence of human umbilical vein endothelial cells(HUVECs) and Bcl-2,Bax gene expression associated with apoptosis induced by angiotensinⅡ(AngⅡ).METHODS:HUVECs were cultured in vitro and the cell viability was observed by methyl thiazolyl tetrazolium(MTT).HUVECs were intervened by AngⅡ and valsartan(AngⅡ type 1 receptor blocking) and divided into 3 groups:the control group,AngⅡ group(stimulated with AngⅡ10-6mol/L for 48 h),valsartan group(valsartan was added to cells 1 h before 10-6mol/L AngⅡ treatment).?-gal staining aod cell cycle analysis were used to identify the cell aging status.Morphologic changes and percentage of apoptosis were assayed with Hoechst33258 under fluorescent microscope.The expressions of Bcl-2 and Bax,and the apoptosis-associated genes were detected by immunocytochemical staining,RT-PCR and Western blotting.RESULTS:The cell viability by AngⅡ-induced cells was(81.9%?4.1)%,the positive cell number of ?-gal staining was significantly higher in AngⅡ-induced cells(80.10%?6.81)% than that in the control cells.The cell cycle was at G0-G1(91.36%?6.45)%,the apoptotic cells significantly increased(31.84?2.86)% under fluorescent microscope.In valsartan group,Bcl-2 mRNA and protein expression increased markedly(P
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AIM:To investigate the effect of L-arginine(L-Arg) on expression of bcl-2,bax mRNA during pulmonary ischemia and reperfusion injury(PIRI) in rabbits.METHODS: Single lung ischemia and reperfusion animal model was used in vivo.The rabbits were randomly divided into three groups: sham operated group(sham,n=12),ischemia-reperfusion group(I/R,n=12) and I/R+ L-arginine group(L-Arg,n=12).Changes of several parameters,which included apoptotic index(AI),wet to dry ratio of lung tissue weight(W/D) and index of quantitative assessment of histologic lung injury(IQA),were measured at 300 min after reperfusion in lung tissue.Meanwhile the location and expression of bcl-2,bax mRNA as well as the ratio of bcl-2 mRNA/bax mRNA were observed.The lung tissue was prepared for light microscopic and electron microscopic observation at 60,180 and 300 min after reperfusion.RESULTS: As compared with I/R group,in intima and extima of small pulmonary artery,alveoli,and bronchiole epithelia,the expression of bcl-2 mRNA and the ratio of bcl-2 mRNA/bax mRNA were increased,and the expression of bax mRNA was decreased in L-Arg treatment group.The values of AI,W/D and IQA showed significantly lower than that in I/R group at 180 minutes after reperfusion in lung tissue(P
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AIM: To explore the effects of homocysteine on the apoptosis in PC12 cells and the relationship between the apoptosis and the expression of the bcl-2 as well as bax gene. METHODS: Cell viability was determined by MTT assay. Cell apoptosis was assessed by phase-contrast microscope, fluorescence microscopy and flow cytometry (FCM). The expression of bcl-2 and bax mRNA was measured by semiquantitative reverse transcription polymerse chain reaction (RT-PCR). RESULTS: Treatment of PC12 cells with Hcy plus 10 ?mol/L copper for 12 h, in the range of 0.125, 0.25, 0.5, 1.0 mmol/L, caused a great decrease in cell viability: 81%, 79%, 69%, 57%, and induced typical morphological changes of apoptosis in a dose-dependent manner. The apoptosis ratios were respectively 8.00%, 10.37%, 17.26% and 20.19%, respectively. The expression of bcl-2 was significantly decreased (from 0.517 to 0.198) whereas bax was significantly increased (from 0.302 to 0.619). CONCLUSION: Homocysteine plus copper may induce apoptosis in PC12 cells through the down-regulation of bcl-2 and the up-regulation of bax gene expression.
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AIM: To investigate gene expression of bax, bcl-2 and p53 in fetal skin at different gestational ages and children skin in order to explore their potentially biological significance. METHODS: Apoptosis in skin specimens was determined by terminal deoxynucleotidy transferase mediated dUTP-biotin nick-end labeling technique (TUNEL). Gene expressions of bax, bcl-2 and p53 in skin at different developmental stages was examined with reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Along with fetal growth and development, the incidence rate of apoptosis was increased progressively in skin. In skin from early gestational fetus, bcl-2 was strongly expressed. This gene expression was progressively decreased with increment in gestational age. In children skin, the mRNA content of this gene was significantly reduced compared with fetal skin (P
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Objective To investigate the effect of preoperative regional intra-arterial chemotherapy on apoptosis and apoptosis-regulating genes in advanced breast cancer.Methods 56 patients were divided into 2 groups:Preoperative regional intra-arterial chemotherapy group(treatment group,28 cases)and non-preoperative chemotherapy group(control group,28 cases).Apoptotic cells were examined by TUNEL and bcl-2 expression,and bax was detected by immunohistochemical techniques.Results Apoptotic index(AI)of breast cancer in the treatment group was 8.74%,while that of the control group was 4.65%.The bcl-2 expression in treatment group was 0.68?0.06,while in control group was 2.24?0.36.Expression of bax in treatment group was 0.72?0.06,while in control group was 0.38?0.04(P
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Objective To investigate the relationship between the expression of Fas, FasL, Bcl-2 and Bax in transitional cell carcinoma of bladder and its clinical significance. Methods Immunohistochemistry method was used to detect the expression of Fas, FasL, Bcl-2 and Bax in 40 specimens of transitional cell carcinoma of bladder and 24 specimens of normal bladder mucosa tissue. Results The positive expression of Fas was detected both in transitional cell carcinonma of bladder and in normal bladder mucosa tissue, while FasL, Bcl-2 and Bax only in transitional cell carcinonma of bladder. The expression of Fas was significantly lower in careinoma of high-grade malignancy than in carcinoma of low-grade malignancy (P