MiR-199及TNF-α在人退变椎间盘髓核组织中的表达及生物学作用  

Expression levels of miR-199 and TNF-αin the human degenerated intervertebral disc nucleus pulposus tissue and their biological effects

作　　者：王伟[1] 郭召[2] 杨利新 乔昱皓 张利 范伟业 WANG Wei;GUO Zhao;YANG Lixin(Fourth Department of Orthopedics,Xiangjiang Branch,the Third Hospital of Hebei Medical University,Hebei,Shijiazhuang 050053,China;不详)

机构地区：[1]河北医科大学第三医院湘江院区骨四科,石家庄市050053 [2]河北省保定市河北大学附属医院

出　　处：《河北医药》2025年第2期212-217,共6页Hebei Medical Journal

基　　金：河北省自然科学基金精准医学联合基金项目(编号:H2021206139)。

摘　　要：目的探讨退变的人椎间盘髓核组织中miR-199及肿瘤坏死因子-α(TNF-α)的表达变化及生物学作用。方法收集人退变椎间盘及正常对照髓核组织标本各5例,高通量测序筛查2组样本中表达差异明显的微小RNA,ELISA方法检测TNF-α水平变化及RT-qPCR检测miR-199表达量变化;人髓核细胞株经体外培养后应用TNF-α(20 ng/mL)诱导建立体外细胞凋亡模型;miR-control及miR mimics-199转染至髓核细胞,然后应用TNF-α刺激,探讨miR-199的调控作用。结果与正常髓核组织比较miR-199在退变髓核组织中表达降低,TNF-α表达明显升高(P<0.05)。随着TNF-α诱导时间的延长miR-199表达及髓核细胞增殖下降(P<0.05),而乳酸脱氢酶(LDH)升高(P<0.05)。与正常组比较,TNF-α组细胞增殖能力下降(P<0.05),细胞凋亡率升高(P<0.05),凋亡相关蛋白(Bax、Bcl-2、caspase-3及cleaved caspase-3)表达水平升高(P<0.05);与TNF-α组比较,TNF-α+miR-199mimics组则逆转上述现象,且LDH升高。结论在退变髓核组织中miR-199表达降低,TNF-α表达升高;髓核细胞凋亡可由TNF-α诱导产生,且呈时间依赖性;TNF-α诱导的髓核细胞凋亡可通过上调miR-199表达而逆转,提示miR-199参与椎间盘退变的发病过程,可作为椎间盘退变的早期诊治标志物及治疗靶点。Objective To investigate the expression levels of miR-199 and tumor necrosis factor-α(TNF-α)in human degenerated intervertebral disc nucleus pulposus tissue and their biological effects.Methods Five tissue samples of degenerated intervertebral discs nucleus pulposus and 5 tissue samples of normal intervertebral discs nucleus pulposus form patients were collected.Differentially expressed microRNA(miRNA)between the two groups was screened by high-throughput sequencing.Changes in TNF-αlevel between groups were detected by enzyme-linked immunosorbent assay(ELISA),and miR-199 level was detected by reverse transcription-quantitative polymerase chain reaction(RT-qPCR).In vitro nucleus pulposus cell lines were cultured and induced with TNF-α(20ng/mL)to establish an in vitro apoptosis model.Nucleus pulposus cells were transfected with miR-control and miR-199 mimics,followed by TNF-αinduction,thus exploring the regulatory effect of miR-199.Results Compared with normal nucleus pulposus tissue,miR-199 was significantly decreased in degenerated nucleus pulposus tissue,and TNF-αwas significantly upregulated(P<0.05).With the prolongation of TNF-αinduction,the expression of miR-199 and the cell proliferation ability of nucleus pulposus cells decreased significantly(P<0.05),and Lactate dehydrogenase(LDH)increased significantly(P<0.05).Compared with the normal group,nucleus pulposus cells induced with TNF-αshowed significantly lower viability(P<0.05),and the expression of apoptosis-related proteins(Bax,Bcl-2,caspase-3 and cleaved caspase-3)were significantly increased(P<0.05).While compared with the TNF-αgroup,the TNF-α+miR-199 mimics group reversed the experimental changes mentioned above,and LDH increased.Conclusion Down-regulated miR-199 and up-regulated TNF-αare seen in the nucleus pulposus tissue of degenerative intervertebral disc.Apoptosis of nucleus pulposus cells can be induced by TNF-αin a time-dependent manner,and TNF-α-induced apoptosis of nucleus pulposus cells can be reversed by overexpression of miR-199

关 键 词：椎间盘退变 微小RNA TNF-Α 凋亡 

分 类 号：R34[医药卫生—基础医学]