槲皮素通过下调MALAT1抑制膝骨关节炎小鼠软骨损伤及软骨细胞凋亡的相关机制  

Mechanism of quercetin inhibiting cartilage injury and chondrocyte apoptosis in mice with knee osteoarthritis by down-regulating MALAT1

作　　者：李伟[1] 黄耀凯 王洪林[1] 汪礼军[1] 黄珍谷 陈兴华[1] 唐鹏[1] LI Wei;HUANG Yao-kai;WANG Hong-lin;WANG Li-jun;HUANG Zhen-gu;CHEN Xing-hua;TANG Peng(Department of Orthopedics,Dazu Hospital Affiliated to Chongqing Medical University,Chongqing 402360,China)

机构地区：[1]重庆医科大学附属大足医院骨科,重庆402360

出　　处：《中华骨质疏松和骨矿盐疾病杂志》2023年第1期33-40,共8页Chinese Journal Of Osteoporosis And Bone Mineral Research

摘　　要：目的探究槲皮素(quercetin,QUE)通过下调人肺腺癌转移相关转录本1(metastasis-associated lung adenocarcinoma transcript 1,MALAT1)抑制膝骨关节炎(knee osteoarthritis,KOA)小鼠软骨损伤及软骨细胞凋亡的相关机制。方法将雄性C57BL/6小鼠分为假手术组、OA组、OA+QUE组,每组各10只。采用内侧半月板失稳术(destabilization of medial meniscus,DMM)诱导膝OA模型,术后5周,小鼠膝关节腔内注射100μL 0.9%(质量分数)氯化钠注射液或QUE(8μmol/L),每周2次,共8周。术后13周,处死小鼠并采集膝关节,分别进行病理观察、Western blot或反转录定量聚合酶链反应检测。从健康雄性C57BL/6小鼠的膝关节软骨中分离软骨细胞,进行原代培养,分为空白组、白细胞介素-1β(interleukin-1β,IL-1β)组、QUE组、IL-1β+QUE组。IL-1β组用小鼠重组IL-1β处理24 h;QUE组用2~256μmol/L QUE作用48 h;IL-1β+QUE组先用IL-1β处理24 h,然后再用4、8、16μmol/L QUE作用;空白组不做任何特殊处理。噻唑兰(methyl thiazolyl tetrazolium,MTT)试剂检测细胞活力。结果与假手术组相比,OA组小鼠OARSI评分(4.50±1.08分vs.0.40±0.52分)、MALAT1 mRNA(1.47±0.06 vs.1.00±0.04)和核因子κB(nuclear transcription factor-κB,NF-κB)mRNA(1.71±0.08 vs.1.00±0.03)以及白细胞介素6(interleukin 6,IL-6)、基质金属蛋白酶13(matrix metalloproteinase 13,MMP-13)、caspase-3蛋白表达水平显著升高,同时miR-9表达(0.78±0.03 vs.1.00±0.03)降低(P<0.05);相反地,OA+QUE组小鼠OARSI评分改善至2.40±1.26分,同时MALAT1、miR-9、NF-κB、IL-6、MMP-13和caspase-3表达较OA组被逆转(P<0.05)。miR-9存在与MALAT1和NF-κB结合的靶向互补序列,而且QUE可抑制MALAT1启动子的转录活性和相对mRNA表达水平。对于细胞实验,与空白组相比,IL-1β组miR-9表达降低(0.45±0.04 vs.1.00±0.03),NF-κB表达(1.52±0.08 vs.0.99±0.02)升高(P<0.05);而IL-1β+QUE组miR-9和NF-κB表达水平则以剂量依赖性方式较IL-1β组被反转(P<0.05)。结�Objective To explore the mechanism of quercetin(QUE)inhibiting cartilage injury and chondrocyte apoptosis in knee osteoarthritis(KOA)mice by down-regulating human metastasis assoiated lung adenocarcinoma transcript 1(MALAT1).Methods Male C57BL/6 mice were divided into sham-operation group,OA group,and OA+QUE group,with 10 mice in each group.The knee OA model was induced by destabilization of medial meniscus(DMM).Five weeks after operation,mice were injected with 100μL of normal saline or QUE(8μmol/L),twice a week for 8 weeks.At 13 weeks after operation,the mice were killed and the knee joints were collected for pathological observation,Western blot and reverse transcription quantitative polymerase chain reaction.Chondrocytes were isolated from the knee cartilage of healthy male C57BL/6 mice and cultured in primary culture.They were divided into blank group and interleukin-1β(IL-1β)group,QUE group,IL-1β+QUE group.IL-1βgroup were treated with mice recombinant IL-1βfor 24 h.QUE group were treated with 2-256μmol/L QUE for 48 h.IL-1β+QUE group was treated with IL-1βfor 24 h and then 4,8,16μmol/L QUE.The blank group did not undergo any special treatment.MTT assay was used to detect cell viability.Results Compared with the sham surgery group,the OA group mice had OARSI scores(4.50±1.08 vs.0.40±0.52),MALAT1 mRNA(1.47±0.06 vs.1.00±0.04),and MALAT1 mRNA(1.47±0.06 vs.1.00±0.04),nuclear transcription factorκB(NF-κB)mRNA(1.71±0.08 vs.1.00±0.03),interleukin 6(IL-6),matrix metalloproteinase 13(MMP-13),and caspase-3 protein expression levels of mice in the OA group significantly increased;meanwhile,the expression of miR-9 decreased(0.78±0.03 vs.1.00±0.03)(P<0.05).On the contrary,the OARSI score of OA+QUE group mice improved to 2.40±1.26,while MALAT1,miR-9,NF-κB.The expression of IL-6,MMP-13,and caspase-3 was reversed compared to the OA group(P<0.05).The presence of miR-9 in MALAT1 and NF-κB binding targets complementary sequences,and QUE can inhibit the transcriptional activity and relative mRNA express

关 键 词：槲皮素 膝骨关节炎 人肺腺癌转移相关转录本1 软骨细胞 炎性反应 

分 类 号：R681[医药卫生—骨科学]