乌司他丁对缺氧诱导PC12细胞神经损伤的保护作用研究

Protective effect of ulinastatin against hypoxia-induced injury in PC12 nerve cells

摘要:

目的 探讨乌司他丁对缺氧诱导PC12细胞神经损伤的作用及分子机制。方法 PC12细胞分为对照组、缺氧组、缺氧+乌司他丁低、中、高剂量组、缺氧+miR-NC组、缺氧+miR-190组、缺氧+乌司他丁+anti-miR-NC组、缺氧+乌司他丁+anti-miR-190组。检测培养液中乳酸脱氢酶（LDH）漏出率及细胞中超氧化物歧化酶（SOD）活性、丙二醛（MDA）含量；流式细胞术检测细胞凋亡；蛋白质印迹法（Western blotting）检测蛋白表达；实时荧光定量聚合酶链反应（qRT-PCR）检测miR-190表达水平。结果 与缺氧组比较，缺氧+乌司他丁低、中、高剂量组PC12细胞中LDH漏出率降低，SOD活性升高，MDA含量降低，细胞凋亡率降低，Bcl-2相对表达量升高，Bax相对表达量降低，miR-190相对表达量升高，且呈剂量依赖性（均P<0.05）。miR-190过表达后，LDH漏出率降低，SOD活性升高，MDA含量降低，细胞凋亡率降低，Bcl-2相对表达量升高，Bax相对表达量降低（均P<0.05）。抑制miR-190表达能逆转乌司他丁对缺氧诱导的PC12细胞损伤的作用。结论 乌司他丁可能通过上调miR-190表达对缺氧诱导PC12细胞神经损伤起保护作用。

Abstract:

Objective To investigate the effect of ulinastatin on hypoxia-induced injury in PC12 nerve cells and its molecular mechanism.Methods PC12 cells were divided into control group, hypoxia group, hypoxia+low/middle/high-dose ulinastatin groups, hypoxia+miR-NC group, hypoxia+miR-190 group, hypoxia+ulinastatin+anti-miR-NC group, and hypoxia+ulinastatin+anti-miR-190 group. Related kits were used to measure the leakage rate of lactate dehydrogenase (LDH) in the culture medium and the activity of superoxide dismutase (SOD) and the content of malondialdehyde (MDA) in cells; flow cytometry was used to measure cell apoptosis; Western blotting was used to measure protein expression; quantitative real-time PCR was used to measure the expression of miR-190.Results Compared with the hypoxia group, the hypoxia+low/middle/high-dose ulinastatin groups had significant reductions in LDH leakage rate, MDA content, and cell apoptosis rate and a significant increase in SOD activity, as well as significant increases in the relative expression of Bcl-2 and miR-190 and a significant reduction in the relative expression of Bax, in a dose-dependent manner (all P<0.05). After miR-190 overexpression, there was a significant reduction in LDH leakage rate, a significant increase in SOD activity, significant reductions in MDA content and cell apoptosis rate, a significant increase in the relative expression of Bcl-2, and a significant reduction in the relative expression of Bax (all P<0.05). Inhibition of the expression of miR-190 reversed the effect of ulinastatin on hypoxia-induced injury of PC12 cells.Conclusions Ulinastatin can exert a protective effect against hypoxia-induced injury of PC12 nerve cells possibly by upregulating the expression of miR-190.