TRPV4影响软骨细胞退变的初步研究
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作者Author单位AddressE-Mail
沈雪 SHEN Xue 上海中医药大学附属曙光医院, 上海 201203 Shuguang Hospital Affiliated to Shanghai University of Traditiontal Chinese Medicine, Shanghai 201203, China  
张琥 ZHANG Hu 上海中医药大学附属曙光医院, 上海 201203 Shuguang Hospital Affiliated to Shanghai University of Traditiontal Chinese Medicine, Shanghai 201203, China  
陈德塔 CHEN De-ta 上海中医药大学附属市中医医院, 上海 200071 Hospital of Traditional Chinese Medicine Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China  
曹月龙 CAO Yue-long 上海中医药大学附属曙光医院, 上海 201203 Shuguang Hospital Affiliated to Shanghai University of Traditiontal Chinese Medicine, Shanghai 201203, China ningtcm@126.com 
期刊信息:《中国骨伤》2023年,第36卷,第10期,第990-995页
DOI:10.12200/j.issn.1003-0034.2023.10.015
基金项目:国家自然科学基金资助项目(编号:81373665);国家自然科学基金资助项目(编号:81973874);上海市慢性筋骨病临床医学研究中心(编号:20MC1920600)
中文摘要:

目的:探讨并验证瞬时受体电位香草素受体4(transient receptor potential vanilloid 4,TRPV4)影响软骨细胞的退变。

方法:取新生SD大鼠软骨细胞,甲苯胺蓝染色和阿利新蓝染色鉴定;通过白细胞介素1β(interleukin-1β,IL-1β)构建体外软骨细胞炎症模型,采用TRPV4抑制剂处理炎症条件下软骨细胞,经荧光定量PCR(real-time PCR,PCR)法检测软骨细胞中TRPV4、基质金属肽酶-13(matrix metallopeptidase 13,MMP-13)、血小板反应蛋白解整合素金属肽酶-5(A aisintegrin and metalloproteinase with thrombospondin 5,ADAMTS-5)、一氧化氮合酶 2(nitric oxide synthase 2,NOS2)、Ⅱ型胶原α1(collagen,typeⅡ,alpha 1,Col2α1)和软骨蛋白聚糖(aggrecan,Acan)mRNA;通过TRPV4不同浓度过表达质粒处理原代软骨细胞,筛选最佳过表达剂量,并经RT-PCR法检测最佳TRPV4过表达剂量下软骨细胞中TRPV4、MMP-13、ADAMTS-5、NOS2、Col2α1、Acan mRNA表达情况。

结果:甲苯胺蓝染色和阿利新蓝染色鉴定提取细胞为原代软骨细胞。RT-PCR显示,经TRPV4抑制剂处理后炎症条件下软骨细胞中TRPV4、MMP-13、ADAMTS-5、NOS2 mRNA的表达明显下降(P<0.05),Col2α1 mRNA、Acan mRNA的表达升高(P<0.05);经0.8 ng·μl-1 TRPV4过表达质粒处理的软骨细胞中Col2α1、Acan mRNA表达情况明显下降(P<0.05),NOS2 mRNA的表达升高(P<0.05),MMP-13、ADAMTS-5mRNA表达差异无统计学意义(P>0.05)。

结论:抑制TRPV4的表达能下调软骨细胞退变相关基因表达。
【关键词】膝骨关节炎  软骨退变  TRPV4
 
Preliminary study of TRPV4 affects chondrocyte degeneration
ABSTRACT  

Objective To explore and verify that transient receptor potential vanilloid 4(TRPV4) affects chondrocyte degeneration.

Method Neonatal SD rats were selected,primary chondrocytes were extracted,and identified by toluidine blue staining and alcian blue staining;an in vitro chondrocyte inflammation model was constructed by IL-1β,and TRPV4 inhibitor was used to treat chondrocytes under inflammatory conditions,and the chondrocytes were treated by RT-PCR method was used to detect matrix metallopeptidase 13(MMP-13),a disintegrin and metalloproteinase with thrombospondin 5,(ADAMTS-5)、nitric oxide synthase 2(NOS2)、Collagen,type II alpha 1(Col2α1)and aggrecan (Acan) mRNA in chondrocytes; primary chondrocytes were treated with different concentrations of TRPV4 overexpression plasmid,and the optimal overexpression dose was screened. The mRNA expressions of TRPV4,MMP-13,ADAMTS-5,NOS2,Col2α1 and Acan in chondrocytes under the optimal TRPV4 overexpression dose were detected.

Results Toluidine blue staining and Alcian blue staining identified the extracted cells as primary chondrocytes;RT-PCR showed that TRPV4,MMP-13,ADAMTS-5,NOS2 mRNA in chondrocytes treated with TRPV4 inhibitor under inflammatory conditions. The expression of Col2α1 mRNA was significantly decreased (P<0.05),and the expression of Col2α1 mRNA was increased (P<0.05). Although there was no significant difference in the expression of Acan mRNA,the overall trend was also increasing. The expression of Col2α1 and Acan mRNA in chondrocytes was significantly decreased (P<0.05),and the expression of NOS2 mRNA was increased(P<0.05),but there was no significant difference in MMP-13 and ADAMTS-5 (P>0.05).

Conclusion Inhibiting the expression of TRPV4 can down-regulate the expression of genes related to chondrocyte degeneration.
KEY WORDS  Knee osteoarthritis  Cartilage degeneration  TRPV4
 
引用本文,请按以下格式著录参考文献:
中文格式:沈雪,张琥,陈德塔,曹月龙.TRPV4影响软骨细胞退变的初步研究[J].中国骨伤,2023,36(10):990~995
英文格式:SHEN Xue,ZHANG Hu,CHEN De-ta,CAO Yue-long.Preliminary study of TRPV4 affects chondrocyte degeneration[J].zhongguo gu shang / China J Orthop Trauma ,2023,36(10):990~995
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