灵芝提取物通过抑制miR-143-3p保护Aβ25-35处理PC12细胞损伤的机制研究  

Mechanism of Ganoderma lucidum extract protecting Aβ25-35 from PC12 cell injury by inhibiting miR-143-3p

作　　者：尹志鹏 吴玲玲 陈光东 YIN Zhi-peng;WU Ling-ling;CHEN Guang-dong(Department of Psychiatry,Wenzhou Seventh People's Hospital,Wenzhou 325000,Chin)

机构地区：[1]温州市第七人民医院精神科,325000

出　　处：《临床神经病学杂志》2023年第3期202-207,共6页Journal of Clinical Neurology

基　　金：温州市科技局课题(Y20210507)。

摘　　要：目的探究灵芝提取物通过抑制miR-143-3p对β-淀粉样蛋白(Aβ25-35)诱导的PC12细胞损伤的影响。方法使用20μmol/L的Aβ25-35处理PC12细胞用于建立体外Alzheimer’s病模型,细胞分为空白组(未经Aβ25-35处理细胞)、模型组(经过Aβ25-35处理细胞24 h)、低剂量组(2.5 mg/L灵芝提取物和Aβ25-35处理细胞24 h)、中剂量组(5.0 mg/L灵芝提取物和Aβ25-35处理细胞24 h)、高剂量组(10 mg/L灵芝提取物和Aβ25-35处理细胞24 h)、miR-NC+高剂量组(转染miR-NC后,使用10 mg/L灵芝提取物和Aβ25-35处理细胞24 h)、miR-143-3p+高剂量组(转染miR-143-3p后,使用10 mg/L灵芝提取物和Aβ25-35处理细胞24 h)。用噻唑蓝检测细胞增殖活性;流式细胞术检测细胞凋亡;Western Blotting法检测剪切的含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved-caspase-3)、Bcl-2相关X蛋白(Bax蛋白)的表达;ELISA检测丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性、过氧化氢酶(CAT)活性、Aβ、TNF-α、IL-6水平;实时定量RT-PCR(qRT-PCR)检测miR-143-3p表达水平。结果与空白组相比,模型组细胞活性、SOD活性、CAT活性显著降低,凋亡率、Cleaved-caspase-3蛋白表达、Bax蛋白表达、MDA含量、Aβ、TNF-α、IL-6水平以及miR-143-3p表达水平显著增加(均P<0.05)。与模型组相比,低剂量组、中剂量组、高剂量组细胞活性、SOD活性、CAT活性显著增加,凋亡率、Cleaved-caspase-3蛋白表达、Bax蛋白表达、MDA含量、Aβ、TNF-α、IL-6水平以及miR-143-3p表达水平显著降低(均P<0.05)。与miR-NC+高剂量组相比,miR-143-3p+高剂量组miR-143-3p表达水平、Cleaved-caspase-3、Bax蛋白表达、凋亡率、MDA含量、Aβ、TNF-α、IL-6水平显著增加,细胞活性、SOD活性、CAT活性显著降低(均P<0.05)。结论灵芝提取物可能通过下调miR-143-3p减弱Aβ25-35诱导的神经细胞凋亡、氧化应激和炎症反应。Objective To explore the effect of Ganoderma lucidum extract on PC12 cell damage induced byβ-amyloid protein(Aβ25-35)by inhibiting miR-143-3p.Methods PC12 cells were treated with 20μmol/L AB25-3s to establish an in vitro Alzheimer's disease model.The cells were divided into blank group(cells not treated with Aβ25-35),model group(cells treated with Aβ25-35 for 24 h),Low-dose group(cells were treated with 2.5 mg/L Ganoderma lucidum extract and Aβ25-3s for 24 h),medium-dose group(cells were treated with 5.0 mg/L Ganoderma lucidum extract and Aβ25-3 for 24 h),high-dose group(cells were treated with 10 mg/L Ganoderma lucidum extract and Aβ25-3s for 24 h),miR-NC+high-dose group(after transfection of miR-NC,cells were treated with 10 mg/L Ganoderma lucidum extract and Aβ25-35 for 24 h),miR-143-3p+high-dose group(after transfection of miR-143-3p,cells were treated with 10 mg/L Ganoderma lucidum extract and Aβ253s for 24 h).Cell proliferation activity was detected by thiazolyl blue;cell apoptosis was detected by flow cytometry;Western blotting was used to detect the expression of cleaved cysteine-containing aspartate protease 3(Cleaved-caspase-3),Bcl-2-related X protein(Bax protein);ELISA was used to detect malondialdehyde(MDA)content,superoxide dismutase(SOD)activity,catalase(CAT)activity,Aβ,TNF-α,IL-6 levels;quantitative real-time RT-PCR(qRT-PCR)was used to detect miR-143-3p expression levels.Results Compared with the blank group,the cell activity,SOD activity and CAT activity in the model group were significantly decreased,and the apoptosis rate,Cleaved-caspase-3 protein,Bax protein,MDA content,Aβ,TNF-α,IL-6 levels,and miR-143-3p expression level were significantly increased(all P<0.05).Compared with the model group,the cell activity,SOD activity,and CAT activity in the low-dose group,middle-dose group,and high-dose group were significantly increased,apoptosis rate,Cleaved-caspase-3 protein expression,Bax protein expression,MDA content,Aβ,TNF-α,IL-6 levels,and miR-143-3p expression levels were significa

关 键 词：灵芝提取物 miR-143-3p Alzheimer’s病 神经细胞 

分 类 号：R749.16[医药卫生—神经病学与精神病学]