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目的 观察抗氧化剂N 乙酰 L 半胱氨酸 (NAC)和过氧化氢酶 (catalase ,CAT)对晶状体上皮细胞凋亡的抑制作用及对凋亡关键蛋白酶caspase 3酶活性的影响。方法 Sprayue Dawley(SD)大鼠 84只分成 4组 ,每组 2 1只。取晶状体于MEM培养液中培养 ,过氧化氢 (H2 O2 )组加入终浓度为 2mmol/L的H2 O2 ,对照组不加H2 O2 ,H2 O2 +NAC组及H2 O2 +CAT组除加入 2mmol/LH2 O2 外再分别加入 10 0 μmol/LNAC和 9× 10 5U/LCAT。培养 2 4h后观察晶状体透明度改变 ,透射电镜下观察细胞超微结构改变 ,流式细胞AnnexinV PI双染色法检测细胞凋亡率 ,并用Westernblot法分析caspase 3相对分子量为 2 0 0 0 0的活性亚单位的表达。结果 2mmol/LH2 O2 作用 2 4h后可引起晶状体明显混浊 ,透射电镜下晶状体上皮细胞出现典型的凋亡形态学改变 ,细胞凋亡率上升为 (31 2± 3 3) % ,并伴随caspase 3酶活性增高 ;分别加入抗氧化剂NAC和CAT后 ,晶状体混浊度明显减轻 ,细胞凋亡率下降为(2 0 9± 3 2 ) %和 (15 0± 2 4 ) % (P <0 0 1) ,caspase 3酶活性均降低。结论NAC和CAT可有效抑制氧化损伤引起的晶状体混浊及上皮细胞凋亡 ,其抑制作用可能与降低caspase 3酶活性有关。
Objective To observe the inhibitory effects of antioxidants NAC and catalase (CAT) on the apoptosis of lens epithelial cells and the effect on the activity of caspase 3, a key protease involved in apoptosis. Methods Eighty-four Sprayue Dawley rats were divided into 4 groups with 21 rats in each group. The cells were cultured in MEM medium, hydrogen peroxide (H2O2) group was added to the final concentration of 2mmol / L H2O2, the control group without H2O2, H2O2 + NAC group and H2O2 + CAT group addition of 2mmol / LH 2 O 2 were then added to 10 0 μmol / LNAC and 9 × 10 5 U / LCAT. After cultured for 24 hours, the changes of the transparency of the lens were observed. The changes of ultrastructure were observed under transmission electron microscope. The apoptosis rate was detected by flow cytometry with Annexin V PI double staining. The relative molecular weight of caspase 3 was analyzed by Western blot Unit of expression. Results 2 mmol / LH 2 O 2 could cause lens opacification after 24 hours. The morphological changes of lens epithelial cells were observed under transmission electron microscope. The rate of apoptosis increased to (31 2 ± 3 3)%, accompanied with caspase 3 After the addition of antioxidants NAC and CAT respectively, the lens opacity was significantly reduced and the apoptosis rate was decreased to (209 ± 2)% and (15 0 ± 2 4)% (P 0 01), respectively. caspase 3 enzyme activity are reduced. Conclusion NAC and CAT can effectively inhibit lens opacification and epithelial cell apoptosis induced by oxidative damage. The inhibitory effect may be related to the decrease of caspase 3 activity.