PGG抑制人晶状体上皮细胞生长及其作用分子

发布时间：2018-09-18 19:33

【摘要】：目的:五没食子酰基葡萄糖(1,2,3,4,6-Pentagalloylglucose,PGG)是一种天然多酚类化合物,它广泛存在于中草药植物中,五倍子、牡丹皮和白芍等植物含量较高。研究表明,PGG具有抗炎、抗肿瘤及抗糖尿病等作用。后发性白内障是白内障囊外摘除术后最常见的并发症,目前认为术中残留的晶状体上皮细胞的增殖是引起后发性白内障的主要原因。本研究拟探讨PGG对人晶状体上皮细胞增殖的抑制作用及其作用机制。方法:(1)PGG的浓度设为0、25、50、75μmol/L,作用SRA细胞的时间为24、48、72 h,采用MTT法检测PGG对人晶状体上皮细胞增殖的影响,显微镜观察细胞形态变化。(2)PGG作用SRA细胞48 h后,采用Annexin VFITC/PI双染,流式细胞技术分析PGG对人晶状体上皮细胞凋亡率的影响。(3)PGG作用SRA细胞48 h后,采用PI单染,流式细胞技术分析PGG对人晶状体上皮细胞周期的影响。(4)PGG作用SRA细胞48 h后,采用Western blotting检测PGG对人晶状体上皮细胞NF-κB p65、phosphoNF-κB p65、phospho-stat3和cyclin D1蛋白表达的影响。结果:(1)PGG孵育后,贴壁的SRA细胞数量明显减少,细胞变圆、变小,细胞增殖受到明显的抑制,且具有时间和剂量依赖性。在同一时间段,药物浓度越大,PGG对SRA细胞增殖的抑制率越大,其中72 h最明显。72 h时,药物浓度为25μmol/L时,抑制率为(30.75±2.63)%,药物浓度为50μmol/L时,抑制率为(39.69±0.67)%,药物浓度为75μmol/L时,抑制率达到(47.84±0.98)%,以对照组(药物浓度为0)增殖抑制率为0做参照,不同药物浓度组抑制率明显增加,差异有统计学意义(F=1842.49,P0.01)。同一药物浓度时,PGG作用时间越长,SRA细胞的增殖抑制率越大。药物浓度为75μmol/L作用24 h时,SRA细胞的增殖抑制率为(11.14±1.29)%,作用48 h时,抑制率为(30.42±2.42)%,72 h时,抑制率高达(47.84±0.98)%,抑制率明显增加,差异有统计学意义(F=716.26,P0.01)。(2)与对照组比较,PGG处理的SRA细胞早期凋亡率和晚期凋亡率均增加,随PGG浓度的增加细胞凋亡率明显增加,差异均有统计学意义(P0.01)。(3)与对照组比较,随着PGG药物浓度的增加,S期细胞所占百分率增加,药物浓度为75μmol/L时,S期细胞百分率为(44.28±1.33)%,药物浓度为50μmol/L时,S期细胞百分率为(37.67±1.29)%,与对照组(28.58±1.27)%比较,S期细胞所占百分率明显增加,差异有统计学意义(F=77.11,P0.01)。(4)PGG浓度增加,PGG孵育的SRA细胞的NF-κB p65的表达无明显改变,但phospho-NF-κB p65表达上调,phospho-stat3与cyclin D1表达下调。结论:PGG可抑制晶状体上皮细胞的增殖,促进晶状体上皮细胞的凋亡,阻滞晶状体上皮细胞于S期。PGG的这些作用可能与其上调蛋白phospho-NF-κB p65,降低phospho-Stat3、cyclin D1的表达有关。因此,PGG有望成为防治后发性白内障的有效药物。
[Abstract]:Objective: PGG is a kind of natural polyphenols. It is widely used in Chinese herbal medicine plants such as Galla chinensis peony and Paeonia lanceolata. The results showed that PGG had anti-inflammatory, anti-tumor and anti-diabetes effects. Posterior cataract is the most common complication after extracapsular cataract extraction. The aim of this study was to investigate the inhibitory effect of PGG on the proliferation of human lens epithelial cells and its mechanism. Methods: (1) the effect of PGG on the proliferation of human lens epithelial cells was detected by MTT assay, and the morphological changes of SRA cells were observed by microscope. (2) SRA cells treated with PGG for 48 h were treated with Annexin VFITC/PI double staining. Flow cytometry was used to analyze the effect of PGG on apoptosis rate of human lens epithelial cells. (3) SRA cells were treated with PGG for 48 h, then PI staining was used. Flow cytometry was used to analyze the effect of PGG on the cell cycle of human lens epithelial cells. (4) SRA cells were treated with PGG for 48 h. The effects of PGG on the expression of phosphoNF- 魏 B p65 phospho-stat3 and cyclin D1 protein in human lens epithelial cells were detected by Western blotting. Results: (1) after PGG incubation, the number of adherent SRA cells decreased significantly, the cells became round and smaller, and the proliferation of SRA cells was significantly inhibited in a time-and dose-dependent manner. At the same time, the inhibition rate of SRA cell proliferation was increased with the increase of drug concentration. The inhibition rate was (30.75 卤2.63) at 72 h, (30.75 卤2.63) at 25 渭 mol/L, (39.69 卤0.67) at 50 渭 mol/L and (39.69 卤0.67) at 75 渭 mol/L. The inhibition rate was (47.84 卤0.98) and the control group (drug concentration was 0). The inhibition rate of different drug concentration groups was significantly increased (FF1842.49, P0.01). At the same drug concentration, the proliferation inhibition rate of SRA cells increased with the time of treatment with PGG. The inhibitory rate of proliferation was (11.14 卤1.29) at the concentration of 75 渭 mol/L for 24 h, and the inhibitory rate was (30.42 卤2.42) and (47.84 卤0.98) at 48 h. Compared with the control group, the early and late apoptosis rates of SRA cells treated with SRA increased significantly with the increase of PGG concentration, and the difference was statistically significant (P0.01). (3) compared with the control group (P0.01). (3), the difference was statistically significant (FF716.26 P0.01). (2), and the apoptosis rate of SRA cells increased significantly with the increase of PGG concentration (P0.01). (3). With the increase of PGG concentration, the percentage of S phase cells was (44.28 卤1.33) at 75 渭 mol/L and (37.67 卤1.29) at 50 渭 mol/L. Compared with the control group (28.58 卤1.27), the percentage of S phase cells increased significantly. There was no significant difference in the expression of NF- 魏 B p65 in SRA cells incubated with SRA, but the expression of phospho-NF- 魏 B p65 up-regulated phospho-stat3 and down-regulated the expression of cyclin D1. Conclusion WPGG can inhibit the proliferation of lens epithelial cells and promote the apoptosis of lens epithelial cells. These effects may be related to the up-regulation of phospho-NF- 魏 B p65 and the decrease of phospho-Stat3,cyclin D1 expression. Therefore, PGG is expected to be an effective drug for the prevention and treatment of post-cataract.
【学位授予单位】：桂林医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R776.1

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