激活素A抑制人晶状体上皮细胞增殖的实验研究
发布时间：2018-07-29 18:30
【摘要】：目的： 探讨激活素A(activinA, ACTA)对人晶状体上皮细胞株SRA01/04(human lens epithelial cell,HLEC SRA01/04)增殖的抑制作用及作用机制的研究,及其与时间—效应的关系。 方法： (1)细胞培养:HLECs用10%FBS的LDMEM培养液培养,并收集状态良好的对数生长期细胞进行下列实验; (2)四甲基偶氮唑盐(methyl thiazolyl tetrazolium, MTT)比色法：检测10ng/mlACTA作用于HLECs 24h、48h、72h后细胞生长抑制率; (3)流式细胞术(flow cytometrc analysis, FCM):检测10ng/ml的ACTA作用于HLECs 24h、48h、72h后早期细胞凋亡率; (4) DAPI免疫荧光染色(4,6—diamidino—2—phenylindole dihydro-chloride, DAPI):观察lOng/ml的ACTA作用于HLECs 48h后的形态学改变; (5)免疫荧光：观察lOng/ml的ACTA作用于HLECs 24h、48h、72h后,检测caspase-3蛋白的表达量。实验结果均行统计学分析。 结果： (1)MTT:10ng/mlACTA溶液作用于HLECs 24h、48h、72h后其抑制率分别为：10.02%、10.91%、18.11%,说明ACTA对HLECs有明显的抑制作用,并随作用时间增加而增加,差异有统计学意义(P均0.05)。 (2)FCM:ACTA作用于HLECs 24h、48h、72h后,早期细胞凋亡率分别为2.3%、6.2%、9.4%,而空白对照组的早期细胞凋亡率为1.4%,说明ACTA对HLECs有诱导凋亡的作用,各组间差异有统计学意义(p0.05),并呈时间一效应关系。 (3)DAPI:ACTA作用于HLECs 48h后细胞变小变圆、细胞膜发生皱缩、染色质浓缩、染色质断裂、聚集、核碎裂等凋亡的形态学改变。对照组细胞形态规则,细胞核饱满,荧光染色均一。 (4)免疫荧光:ACTA作用于HLECs 24h、48h、72h后,caspase-3蛋白表达随时间增加而增加。差异有统计学意义(p0.05)。 结论： 激活素A对人晶状体上皮细胞的增殖有抑制作用及诱导凋亡的作用,并呈时间—效应关系,因此可望成为防治后发性白内障的有效药物。
[Abstract]:Aim: to investigate the inhibitory effect of activin A (activinA, ACTA) on the proliferation of human lens epithelial cell line SRA01/04 (human lens epithelial cell line HLEC SRA01/04 and its relationship with time-effect. Methods: (1) the cells were cultured in 10 S LDMEM medium. The following experiments were carried out: (2) tetramethylazolium (methyl thiazolyl tetrazolium, MTT) colorimetry: to detect the inhibition rate of cell growth by 10ng/mlACTA on HLECs 24 h, 48 h and 72 h; (3) flow cytometry. (4) DAPI immunofluorescence staining (46-diamidino-2-phenylindole dihydro-chloride (DAPI): the morphological changes of lOng/ml ACTA on HLECs 48 h were observed. (5) Immunofluorescence: the effects of ACTA of lOng/ml on HLECs for 24 h and 48 h after 72 h were observed. The expression of caspase-3 protein was detected. The experimental results were analyzed statistically. Results: (1) after treated with MTT:10ng/mlACTA solution for 24 h or 48 h or 72 h, the inhibition rates of ACTA were 10.02 and 10.91, respectively, indicating that ACTA had obvious inhibitory effect on HLECs and increased with the increase of time. The difference was statistically significant (P0. 05). (2). After treated with FCM:ACTA for 24 h or 48 h or 72 h, the rate of early apoptosis was 2. 3% and 6. 2%, respectively, while that of the blank control group was 1. 4%, indicating that ACTA could induce apoptosis of HLECs. There were significant differences among the groups (p0. 05). (3) after DAPI:ACTA was treated with HLECs for 48 h, the cells became smaller and rounded, the cell membrane shrank, chromatin condensed, chromatin breaks and aggregates. Morphological changes of apoptosis such as nuclear fragmentation. In the control group, the cell morphology was regular, the nucleus was full and the fluorescence staining was uniform. (4) the expression of caspase-3 protein increased with the increase of time after the treatment of HLECs 24 h, 48 h and 72 h with immunofluorescence: ACTA. The difference was statistically significant (p0.05). Conclusion: Activin A can inhibit the proliferation of human lens epithelial cells and induce apoptosis in a time-dependent manner, so it is expected to be an effective drug for the prevention and treatment of post-cataract.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R776.1
本文编号：2153615
[Abstract]:Aim: to investigate the inhibitory effect of activin A (activinA, ACTA) on the proliferation of human lens epithelial cell line SRA01/04 (human lens epithelial cell line HLEC SRA01/04 and its relationship with time-effect. Methods: (1) the cells were cultured in 10 S LDMEM medium. The following experiments were carried out: (2) tetramethylazolium (methyl thiazolyl tetrazolium, MTT) colorimetry: to detect the inhibition rate of cell growth by 10ng/mlACTA on HLECs 24 h, 48 h and 72 h; (3) flow cytometry. (4) DAPI immunofluorescence staining (46-diamidino-2-phenylindole dihydro-chloride (DAPI): the morphological changes of lOng/ml ACTA on HLECs 48 h were observed. (5) Immunofluorescence: the effects of ACTA of lOng/ml on HLECs for 24 h and 48 h after 72 h were observed. The expression of caspase-3 protein was detected. The experimental results were analyzed statistically. Results: (1) after treated with MTT:10ng/mlACTA solution for 24 h or 48 h or 72 h, the inhibition rates of ACTA were 10.02 and 10.91, respectively, indicating that ACTA had obvious inhibitory effect on HLECs and increased with the increase of time. The difference was statistically significant (P0. 05). (2). After treated with FCM:ACTA for 24 h or 48 h or 72 h, the rate of early apoptosis was 2. 3% and 6. 2%, respectively, while that of the blank control group was 1. 4%, indicating that ACTA could induce apoptosis of HLECs. There were significant differences among the groups (p0. 05). (3) after DAPI:ACTA was treated with HLECs for 48 h, the cells became smaller and rounded, the cell membrane shrank, chromatin condensed, chromatin breaks and aggregates. Morphological changes of apoptosis such as nuclear fragmentation. In the control group, the cell morphology was regular, the nucleus was full and the fluorescence staining was uniform. (4) the expression of caspase-3 protein increased with the increase of time after the treatment of HLECs 24 h, 48 h and 72 h with immunofluorescence: ACTA. The difference was statistically significant (p0.05). Conclusion: Activin A can inhibit the proliferation of human lens epithelial cells and induce apoptosis in a time-dependent manner, so it is expected to be an effective drug for the prevention and treatment of post-cataract.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R776.1
【引证文献】
相关硕士学位论文 前1条
1 赵瑞苓;外源表达人激活要素受体ⅡA对人晶状体上皮细胞增殖的影响[D];吉林大学;2012年
，本文编号：2153615
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