Quercetin regulates Fas expression and induces apoptosis in hepatocellular carcinoma HepG2 cells

AIM: To investigate the role of 1, 4, 5- trisphosphate inositol (IP3) and Fas gene expression in apoptosis of HepG2 cells induced by quercetin. METHODS: HepG2 cells were treated with quercetin at different concentrations (including 20, 40, 60, 80 μmol/ L) for 72 h and treated with 60 μmol/ L quercetin for 6 h, 12 h, 24 h, 48 h and 72 h. IP3, Fas mRNA, Fas protein and apoptosis rate were assayed by IP3 - ［3H］ Birtrak assay, RT-PCR, Western blotting and flow cytometry, respectively. RESULTS: When HepG2 cells were incubated with different concentrations of quercetin for 72 h, the IP3 content was lower than those in control. Fas mRNA expression, Fas protein expression and the apoptosis rate were higher than those in control. When HepG2 cells were incubated with quercetin for 6 h, 12 h, 24 h, 48 h, 72 h, the IP3 contents were lower than those in control incubated with 60 μmol/L quercetin for 12 h. Fas mRNA expression was higher than that in control incubated with 60 μmol/L quercetin for 12 h . Fas protein expression was higher than that in control. The apoptosis rate was significantly higher than that in control incubated with 60 μmol/L quercetin for 24 h (P<0.01). CONCLUSION: Quercetin induces apoptosis of HepG2 cells by reducing IP3 production and upregulating Fas gene expression.     

Fas/FasL通路对镉激活PC12细胞MAPK通路的影响

旨在研究Fas/FasL通路对镉激活PC12细胞MAPK通路的影响,用10 μmol L^-1醋酸镉（CdAc₂）分别处理插入非特异性序列PC12细胞与Fas基因沉默PC12细胞12 h;用10 μmol·L^-1CdAc₂与40 μmol·L^-1 Z-IETD-FMK （caspase-8特异性抑制剂）单独或联合处理PC12细胞12 h。通过Western blot检测Fas/FasL通路相关蛋白Fas、FasL、Fas相关死亡域蛋白（FADD）、Cleaved caspase-8、死亡结构域相关蛋白（Daxx）、凋亡信号调节激酶1（ASK1）的表达与MAPK通路相关蛋白ERK1/2、JNK1/2、p-ERK1/2、p-JNK1/2的表达;Hoechst33258荧光染色检测细胞凋亡形态学变化,流式细胞术检测细胞凋亡率。结果显示,Fas shRNA慢病毒极显著抑制镉引起的PC12细胞Fas/FasL通路相关蛋白表达量和凋亡率的升高（P<0.01）,缓解镉引起的PC12细胞凋亡形态学变化;Fas shRNA与Z-IETD-FMK均能够极显著抑制镉引起的PC12细胞MAPK通路相关蛋白ERK1/2与JNK1/2磷酸化水平升高（P<0.01）。综上表明,Fas/FasL通路调控MAPK通路参与镉致PC12细胞凋亡。     

氯氨酮对大鼠缺血再灌注心肌细胞凋亡和Fas/FasL蛋白表达的影响

目的:探讨氯胺酮对大鼠实验性心肌缺血再灌注时心肌细胞凋亡与Fas及Fas蛋白配体(FasLi-gand,FasL)表达的影响,并分析心肌组织病理学损伤程度。方法:以穿线结扎或松扎左冠状动脉制备大鼠心肌缺血再灌注模型。32只大鼠随机平均分成假手术组(假手术4.5 h)、缺血再灌注组(缺血30min、再灌注4 h)、低剂量氯胺酮再灌注组(缺血30 min、再灌注4 h)及高剂量氯胺酮再灌注组(缺血30 min、再灌注4 h)。以缺口末端标记法检测心肌细胞凋亡的变化,S-P免疫组化法分别检测Fas与FasL蛋白水平变化,做病理组织切片检查心肌损伤情况。结果:心肌缺血再灌注前心肌细胞凋亡指数、Fas蛋白阳性染色指数与炎性细胞FasL蛋白阳性染色指数分别为4.25±2.04、1.06±0.25和0,心肌缺血再灌注后心肌细胞凋亡指数、Fas蛋白阳性染色指数和炎性细胞FasL蛋白阳性染色指数分别为12.58±1.35,11.05±3.02和8.12±3.54,低剂量氯氨酮作用组心肌细胞凋亡指数及Fas蛋白阳性染色指数与炎性细胞FasL蛋白阳性染色指数分别为7.36±1.30,7.61±3.41和3.69±3.13;心肌缺血再灌注后心肌组织呈大小不一的灶性坏死,坏死周围有炎性细胞浸润,氯胺酮作用后坏死减轻,低剂量氯胺酮作用更明显。结论:心肌缺血再灌注时心肌细胞凋亡、Fas基因的蛋白与炎性细胞的FasL蛋白表达量均增加,氯氨酮可减少心肌凋亡,减少细胞Fas和FasL蛋白阳性表达,从而减轻心肌损伤,且低剂量氯氨酮作用更明显。     

睾丸免疫豁免研究进展

睾丸免疫豁免对精子发生具有重要的保护作用.文章对免疫豁免这一概念和作用进行了概述,论述了睾丸巨噬细胞、树状突细胞作用和Leydig细胞分泌的雄激素在免疫豁免中的作用机制,Sertoli细胞中Fas/FasL系统在睾丸免疫豁免中的作用,并阐述了睾丸免疫豁免在临床实践中的应用,提出了下一步需要研究的问题.     

Effects of the grub extract on apoptosis of MCF-7 human breast cancer cell line

AIM: To investigate the apoptotic pathway of MCF-7 breast cancer induced by the grub extract in vitro.METHODS: MTT assay was used to determine the effect of the grub extract on proliferation of MCF-7 human breast cancer cell line and cell toxicity. Morphological changes of the apoptosis in cancer cells were observed by HE staining through invert microscope, light microscope, AO/EB double fluorescent staining under fluorescent microscope. FCM was used to assay the change of apoptotic rate. The expression of Bcl-2, Fas, caspase-9, caspase-3 in apoptotic pathway was detected with immunocytochemical method before and after exposure to the grub extract, and the effect of that on apoptotic pathway was explored.RESULTS: (1) The MTT test showed that the growth of MCF-7 human breast cancer cell line was significantly inhibited by the grub extract in dose and time dependent manners. The inhibitory rate in exposure group was significantly different from that in control group (P<0.01). (2) Morphological changes of apoptosis including nuclear condensation, fragment and apoptosis body formation were observed by invert microscope. (3) The MCF-7 human breast cancer cells in experimental group by HE staining showed nuclear condensation and blue-black, cytoplasm slight red, nuclear chromatin condensation and fragment shape, apoptosis body formations. (4) Apoptosis in the experimental group was observed by AO/EB double fluorescent staining under fluorescent microscope. (5) FCM assay indicated that apoptotic rate increased significantly in time dependent manner in experimental group. (6) The expression of Bcl-2 was down-regulated, while that of Fas, caspase-3, caspase-9 was up-regulated, compared with control group (P<0.01).CONCLUSION: (1) The proliferation of MCF-7 human breast cancer cell line can be inhibited significantly by the grub extract in vitro. (2)The mechanism of effect of the grub extract on MCF-7 human breast cancer cell line might be mediated by down-regulation of Bcl-2 and up-regulation of Fas, caspase-3, caspase-9. This type of apoptosis starting and performing is through death receptor pathway and mitochondrial pathway.     

Effect of CoQ10 on apoptosis and proliferation of cultured mouse microvascular endothelial cells and it's mechanism

AIM: To study the inhibitory effect of CoQ₁₀ on the apoptosis of microvascular endothelial cells and it's probable mechanism. METHODS: Using serum pharmacology method and cytoflowmetery, the effects of CoQ₁₀ at different concentrations on apoptosis and proliferation in cultured mouse brain microvascular endothelial cells (bEnd.3) were investigated. The expression of Fas protein and Bcl-2 protein were observed with immunocytochemical method (ABC). RESULTS: The cell apoptosis was inhibited significantly in CoQ₁₀ groups (50 μL and 25 μL) in cultured bEnd.3 cells. The results of immunocytochemical staining showed that the expressions of Fas protein was inhibited and Bcl-2 protein was stimulated significantly in CoQ10 group with above concentration. But there was no significant change in cell proliferation. CONCLUSIONS: CoQ₁₀ may inhibit apoptosis of microvascular endothelial cells (bEnd.3) via up-regulation of Bcl-2 and down-regulation of Fas. Authors suggest that this is one of the protection mechanisms of CoQ₁₀ from dysfunction of microvascular endothelial cells.     

Association of Fas promoter-670 polymorphism with systemic lupus erythematosus in southern Chinese

AIM: To investigate whether Fas promoter-670 polymorphism is associated with systemic lupus erythematosus(SLE) in Southern Chinese. METHODS: 103 SLE patients and 110 controls were studied. Fas promoter -670 polymorphism was typed by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). RESULTS: No statistically significant differences were found when Fas promoter -670 genotype and allele frequencies were compared between the SLE and the controls. Similarly, no significant differences were seen between the male and female SLE and the controls, the SLE with lupus nephritis (LN) and the controls, the SLE with LN and the SLE without LN. CONCLUSION: Fas promoter -670 polymorphism does not appear to be associated with susceptibility to SLE in Southern Chinese.     

三氧化二砷对鸡马立克氏病肿瘤细胞凋亡的影响

研究了三氧化二砷（As2O3）对诱导鸡马立克氏病（MD）肿瘤细胞株MDCC—MSB1细胞凋亡相关基因Fas、Caspase-3 mRNA表达及Caspase-3活性的影响，进而探讨了AS2O3诱导MD肿瘤细胞株凋亡的机制。以体外培养MDCC-MSB1为研究对象，经终浓度分别为0、2、4和8μmol／L的As2O3作用48h后，采用荧光显微镜观察细胞的形态学变化，DNA Ladder法检测细胞凋亡情况，RT-PCR方法检测Fas、Caspase-3 mRNA的表达水平，试剂盒比色法检测Caspase-3活性的变化。结果表明，AS2O3能引起典型的凋亡形态学变化，并出现典型的DNA Ladder，同时Fas、Caspase-3 mRNA的表达水平和Caspase-3活性均增加，组间差异均极显著（P〈0．01），呈现剂量依赖性。证实As2O3诱导鸡MD肿瘤细胞株MDCC-MSB1细胞凋亡是通过增加Fas、Caspase-3 mRNA的表达和提高Caspase-3活性来实现的。     

豆豉对早期动脉粥样硬化大鼠主动脉平滑肌细胞凋亡的影响

观察豆豉对早期动脉粥样硬化大鼠主动脉平滑肌细胞凋亡及相关凋亡基因突变型p53和Fas蛋白表达的调节作用,以探讨其机制.采用高脂饲料喂养法建立大鼠早期动脉粥样硬化模型,同时连续10周灌胃给予豆豉提取物进行干预.末次给药后处死动物,光镜下观察血管平滑肌细胞的形态,透射电镜观察超微结构,采用流式细胞术检测细胞凋亡率以及凋亡相关基因突变型p53和Fas蛋白的表达.模型对照组主动脉平滑肌细胞的凋亡率及增值指数(FI)明显高于正常对照组(P＜0.05),053蛋白表达上调;豆豉干预组(20 g生药·kg-1,dO g生药·kg-1)大鼠主动脉平滑肌细胞的凋亡率明显高于模型对照组(P＜0.05),增殖指数明显低于模型对照组(P＜0.05),Fas蛋白表达上调(P＜0.05),p53蛋白表达下调(P＜0.05),光镜及透射电镜可见主动脉损伤明显较模型对照组改善.豆豉可调节早期动脉粥样硬化大鼠血管平滑肌细胞凋亡与增殖的平衡,其机制可能与调节突变型p53和Fas蛋白的表达有关.     

Effect of Fas/FasL on late reperfusion of acute myocardial infarction and the potential oxidative stress mechanism in canine

AIM: To discuss the effect of Fas/FasL on the late reperfusion of acute myocardial infarction (AMI) and the potential oxidative stress mechanism. METHODS:Eighteen anesthetized dogs were randomly divided into three groups: late reperfusion group (n=6): ligated the coronary for 6 h, followed by reperfusion for 6 h; permanent ischemia group (n=6): after pericardium were opened for 6 h, ligated the coronary for 6 h, and did not reperfuse; control group (n=6): did not ligate the coronary but operation last for 12 h. Infarction brim myocardial Fas/FasL was detected by immunohistochemistry. Apoptosis index (AI) was detected by TUNEL. SOD and GR activity and MDA content were detected by colorimetry. RESULTS:The expression of Fas/FasL and apoptosis index were significantly higher in permanent ischemia group and late reperfusion group than those in control group (P<0.05, P<0.01), and the difference between them was also significant (P<0.05). SOD and GR activities were lower in permanent ischemia group and late reperfusion group than those in control group (P<0.05, P<0.01). The MDA contents in permanent ischemia group and late reperfusion group were higher than that in control group (P<0.05). CONCLUSION:The late reperfusion of AMI promotes the expression of Fas/FasL and myocardial apoptosis, and it may be due to oxidative stress mechanism.