| 邓慧琳,杨婧,郭俐.α?硫辛酸对白细胞介素 1β诱导的软骨细胞损伤的保护作用[J].安徽医药,2021,25(1):57-63. |
| α?硫辛酸对白细胞介素 1β诱导的软骨细胞损伤的保护作用 |
| Induction of IL?1β by α?lipoic acid protective effect of chondrocyte injury |
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| DOI：10.3969/j.issn.1009?6469.2021.01.015. |
| 中文关键词: 硫辛酸 微小 RNA?877 基因， myc N?myc下游调节基因 2 软骨细胞 肿瘤坏死因子 α 白细胞介素类 细胞凋亡 大鼠， Sprague?Dawley |
| 英文关键词: Thioctic acid miR?877 Genes,myc NDRG2 Chondrocytes Tumor necrosis factor?alpha Interleukins Apoptosis Rats,Sprague?Dawley |
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| 中文摘要: |
| 目的探讨 α?硫辛酸(ALA)调控微小 RNA?877(miR?877)/N?myc下游调节基因 2(NDRG2)分子轴对白细胞介素 1β(IL ?1β)诱导的软骨细胞损伤的影响及其机制。方法本研究于 2019年 1月至 2019年 7月在南充市中心医院风湿免疫科实验室完成。分离培养 SD大鼠膝关节软骨细胞，按照随机数字表法分为对照(Con)组、 IL?1β组、 IL?1β+ALA?低剂量(L)组、 IL?1β+ ALA?中剂量(M)组、 IL?1β+ALA?高剂量(H)组、 IL?1β+miRNA抑制物阴性对照(anti?miR?NC)组、 IL?1β+miR?877抑制物(anti? miR?877)组、 IL?1β+空载体质粒(pcDNA)组和 IL?1β+NDRG2过表达质粒(pcDNA?NDRG2)组、 IL?1β+ALA+miRNA模拟物阴性对照(miR?NC)组和 IL?1β+ALA+miR?877组。流式细胞术检测细胞凋亡，酶联免疫吸附测定(ELISA)试剂盒检测白细胞介素 ?6(IL?6)、肿瘤坏死因子 α(TNF?α)、干扰素 γ(IFN?γ)的分泌量，蛋白质印记(western blot)法检测 NDRG2蛋白的表达水平，实时定量 PCR(qRT?PCR)检测 miR?877和 NDRG2 mRNA的表达水平。双荧光素酶报告基因实验和 western blot检测 miR?877和 NDRG2的靶向调控关系。结果与 Con组比较， IL?1β组软骨细胞 IL?6［(3.27±0.31)比(1.06±0.09)ng/L］、 TNF?α、IFN?γ的分泌增加， miR?877［(2.65±0.24)比(1.02±0.09)］表达升高， NDRG2［(0.21±0.03)比(0.69±0.06)］表达降低，细胞凋亡率［(0.69±0.06)比(6.24±0.63)%］升高(均 P＜0.05)；与 IL?1β组比较， IL?1β+ALA?L组、 IL?1β+ALA?M组、 IL?1β+ALA?H组软骨细胞 IL?6［(2.91± 0.25)(2.33±0.23)，(1.65±0.16)比(3.27±0.31)ng/L］、 TNF?α、IFN?γ的分泌降低， miR?877［(2.21±0.19)(1.96±0.17)，(1.43± 0.14)比(，2.65±0.24)］的表达降低， NDRG2［(0.34±0.03)(0.46±0.04)(0.58±0.05)比(0.21±0.03)］表达，升高，细胞凋亡率［(20.13±2.12)(16.32±1.63)(11.04±1.15)比(27.62±2.43)，］%降低，差异有，统计学意义(P＜0.05)；与 IL?1β+anti?miR?NC组比较， IL?1β+anti?mi，R?877组软骨细，胞 IL?6［(1.38±0.14)比(3.34±0.33)ng/L］、 TNF?α、IFN?γ的分泌降低，细胞凋亡率［(12.54±1.25)比(27.14±2.73)］%降低，差异有统计学意义(P＜0.05)；与 IL?1β+pcDNA组比较， IL?1β+pcDNA?NDRG2组软骨细胞 IL?6［(1.43±0.15)比(3.47±0.34)ng/L］、 TNF?α、IFN?γ的分泌降低，细胞凋亡率［(13.55±1.35)比(28.14±2.83)］%降低，差异有统计学意义(P＜0.05)；与 IL?1β+ALA+miR?NC组相比， IL?1β+ALA+miR?877组软骨细胞 IL?6［(3.01±0.29)比(1.42±0.13)ng/L］、 TNF?α、IFN?γ的分泌升高，细胞凋亡率［(25.47±2.53)比(11.68±1.87)］%增加，差异有统计学意义(P＜0.05)。结论 α?硫辛酸能够减轻 IL? 1β诱导的软骨细胞损伤，其机制与调控 miR?877/NDRG2通路有关。 |
| 英文摘要: |
| Objective To investigate the effect of α?lipoic acid(ALA)on the interleukin 1β(IL?1β)?induced chondrocyte inju? ry induced by regulating microRNA?877(miR?877)/Downstream regulatory gene 2 of N?myc(NDRG2)molecular axis and its mechanism.Methods The experiment was completed in the Laboratory of Rheumatology and Immunology of Nanchong CentralHospital from January 2019 to July 2019.SD rats were purchased from Hunan Slack Jingda Experimental Animal Co.，Ltd.Isolated and cultured chondrocytes in vitro were divided into control(Con)group，IL?1β group，IL?1β+ALA low dose?(L)group，IL?1β+ ALA medium dose(M)group，IL?1β+ALA high dose(H)group.IL?1β+negative control of miRNA inhibitor(anti?miR?NC)group， IL?1β+miR?877 inhibitor(anti?miR?877)group，IL?1β+ empty vector plasmid(pcDNA)group and IL?1β+NDRG2 overexpression plasmid(pcDNA?NDRG2)group，IL?1β+ALA+ negative control of miRNA mimics(miR?NC)group and IL?1β+ALA+miR?877 group according to the random number table method.Apoptosis was detected by flow cytometry，the secretion of interleukin 6(IL? 6)，tumor necrosis factor α(TNF?α)and interferon?γ(IFN?γ)was detected by enzyme?linked immunosorbent assay(ELISA)kit， the expression NDRG2 protein were detected by Western blot，and the expression levels of NDRG2 mRNA and miR?877 were de?tected by Real?time quantitative PCR(qRT?PCR).The dual luciferase reporter gene assay and western blot were used to detect thetargeted regulatoryrelationship between miR?877 and NDRG2.Results Compared with the Con group，the secretion of IL?6(3.27±0.31 vs. 1.06±0.09)ng/L，TNF?α and IFN?γ in chondrocytes of IL?1β group increased，the expression of miR?877(2.65±0.24 vs.1.02±0.09)increased，and the expression of NDRG2(0.21±0.03 vs. 0.69±0.06)decreased，the apoptotic rate(0.69±0.06 vs. 6.24±0.63)% increased，the difference was statistically significant(P＜0.05)； Compared withIL?1β group，the secretion of IL?6(2.91±0.25，2.33±0.23，1.65±0.16 vs. 3.27±0.31)ng/L，TNF?αand IFN?γ in chondrocytes of IL?1β+ALA?L group，IL?1β+ALA?M group，IL? 1β+ALA?H group decreased，the expression of miR?877(2.21±0.19，1.96±0.17，1.43±0.14 vs. 2.65±0.24)decreased，the expression of NDRG2(0.34±0.03，0.46±0.04，0.58±0.05 vs. 0.21±0.03)increased，and the apoptosis rate(20.13±2.12，16.32±1.63，11.04±1.15 vs. 27.62±2.43)% decreased，the difference was statistically significant(P＜0.05).Compared with IL?1β+anti?miR?NC group， the secretion of IL?6(1.38±0.14 vs. 3.34±0.33)ng/L，TNF?α and IFN?γ in chondrocytes of IL?1β+anti?miR?877 group decreased， and the apoptosis(12.54±1.25 vs. 27.14±2.73)% rate decreased，the difference was statistically significant(P＜0.05).Compared with IL?1β+pcDNA group，the secretion of IL?6(1.43±0.15 vs. 3.47±0.34)ng/L，TNF?α and IFN?γ in chondrocytes of IL?1β+pcD? NA?NDRG2 group decreased，and the apoptosis rate(13.55±1.35 vs. 28.14±2.83)% decreased，the difference was statistically sig? nificant(P＜0.05).Compared with IL?1β+ALA+miR?NC group，the secretion of IL?6(3.01±0.29 vs. 1.42±0.13)ng/L，TNF?α and IFN?γ in chondrocytes of IL?1β+ALA+miR ?877 group increased，and the apoptosis rate(25.47±2.53 vs. 11.68±1.87)% increased， the difference was statistically significant(P＜0.05).Conclusion α?lipoic acid could attenuate IL?1β?induced chondrocyte injury， and its mechanism is related to the regulation of miR?877/NDRG2 pathway. |
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