摘要
目的探讨miR-214-3p对IL-1β诱导的软骨细胞自噬和凋亡的影响及其可能的机制。方法采用10 ng/mL的IL-1β诱导大鼠关节软骨细胞24 h,构建骨关节炎(osteoarthritis,OA)细胞模型,将细胞分为对照组、模型组、miR-NC组、miR-214-3p mimics组、740Y-P(PI3K/AKT通路激活剂)+miR-214-3p mimics组和LY294002(PI3K/AKT通路抑制剂)+miR-214-3p mimics组。通过CCK8检测细胞活力、流式细胞术检测细胞凋亡、透射电镜观察细胞中的自噬小体、Western blot检测凋亡(Bcl-2、Bax)、自噬(LC3II、Beclin-1)和PI3K/AKT通路(AKT、p-AKT)相关蛋白的表达。结果与对照组相比,模型组细胞活力降低(P<0.01),细胞凋亡率升高(P<0.01),自噬小体明显减少,细胞中Bcl-2、LC3II和Beclin-1蛋白表达水平降低(P<0.01),Bax和p-AKT/AKT水平升高(P<0.01);与miR-NC组相比,miR-214-3p mimics组细胞活力升高(P<0.01),细胞凋亡率降低(P<0.01),自噬小体明显增多,细胞中Bcl-2、LC3II和Beclin-1蛋白表达水平升高(P<0.01),Bax和p-AKT/AKT水平降低(P<0.01);与miR-214-3p mimics单独作用相比,联合740Y-P能够逆转miR-214-3p mimics的上述作用,而联合LY294002能够进一步加重miR-214-3p mimics的上述作用。结论miR-214-3p能够促进OA细胞增殖和自噬,并抑制细胞凋亡,其机制可能与抑制PI3K/AKT通路的激活有关。
Objective To investigate the effects of miR-214-3p on chondrocyte autophagy and apoptosis induced by IL-1βand its possible mechanism.Methods Rat articular chondrocytes were induced by 10 ng/mL IL-1βfor 24 h,to establish osteoarthritis(OA)cell model.The cells were divided into control group,model group,miR-NC group,miR-214-3p mimics group,740Y-P(PI3K/AKT pathway activator)+miR-214-3p mimics group,and LY294002(PI3K/AKT pathway inhibitor)+miR-214-3p mimics group.Cell viability was detected with CCK8.Cell apoptosis was detected with flow cytometry.Autophagosomes in cells were observed using transmission electron microscopy.Expressions of apoptosis(Bcl-2,Bax),autophagy(LC3II,Beclin-1),and PI3K/AKT pathway(AKT,p-AKT)related proteins were detected using Western blotting.Results Compared to those in the control group,the cell viability in the model group decreased(P<0.01),the apoptosis rate increased(P<0.01),autophagosomes decreased,the expression levels of Bcl-2,LC3II,and Beclin-1 proteins decreased(P<0.01),and Bax and p-AKT/AKT levels increased(P<0.01).Compared to those in the miR-NC group,cell viability in miR-214-3p mimics group increased(P<0.01),the apoptosis rate decreased(P<0.01),autophagosomes increased,the expression levels of Bcl-2,LC3II and Beclin-1 proteins increased(P<0.01),and Bax and p-AKT/AKT levels decreased(P<0.01).Compared to those in the effects of miR-214-3p mimics alone,the combination of 740Y-P reversed the above effects of miR-214-3p mimics,while the combination of LY294002 further aggravated the above effects of miR-214-3p mimics.Conclusion miR-214-3p promotes OA cell proliferation and autophagy,and inhibits apoptosis,which may be related to the inhibition of the activation of PI3K/AKT pathway.
作者
付亚静
雷豫
张莉莹
刘超
汪伟
FU Yajing;LEI Yu;ZHANG Liying;LIU Chao;WANG Wei(Department of Anesthesiology,People’s Hospital of Wuhan University,Wuhan 430000,China;Department of Spinal Orthopedics,the Second Affiliated Hospital of Guizhou University of Traditional Chinese Medicine,Guiyang 550003,China;School of Acupuncture and Bone Injury,Hubei University of Chinese Medicine,Wuhan 430065,China;Department of Orthopedics and Traumatology,the Affiliated Hospital of Hubei University of Chinese Medicine,Hubei Provincial Hospital of Traditional Chinese Medicine,Hubei Provincial Institute of Traditional Chinese Medicine,Wuhan 430060,China)
出处
《中国骨质疏松杂志》
CAS
CSCD
北大核心
2024年第11期1561-1567,共7页
Chinese Journal of Osteoporosis
基金
湖北省自然科学基金(2021CFB217)。
