摘要
[目的]探讨L-型电压依赖性钙通道在NFAT1调节软骨细胞基质代谢中的作用。[方法]Hulth法制作兔骨关节炎(KOA)模型。使用q RT-PCR法观察对照组和KOA组兔软骨细胞5种NFAT亚型的m RNA表达情况,然后使用L-型钙通道阻断剂硝苯地平,检测软骨细胞NFAT各亚型和基质代谢各标记物的改变,以及膜片钳技术观察KOA细胞膜电位的变化。[结果]与对照组相比,KOA组软骨细胞5种NFAT的m RNA表达变化并不一致。KOA组软骨细胞NFAT1明显升高,是对照组的2.42倍(P<0.01),NFAT2显著降低,是对照组的0.34倍(P<0.01),而NFAT3、NFAT4和NFAT5的m RNA表达没有改变。与单纯DMSO培养基对照组相比,10μmol/L硝苯地平对KOA软骨细胞的活力没有影响,但完全阻断了KOA软骨细胞Cav1的m RNA表达,增加了软骨细胞内基质成分II型胶原(7.46倍,P<0.01)、聚集蛋白聚糖(4.98倍,P<0.05)和转化生长因子-β的m RNA表达(4.37倍,P<0.01),降低了分解酶基质金属蛋白酶-1(0.17倍,P<0.01),含血小板反应蛋白基序的解聚蛋白样金属蛋白酶-4(ADAMTS)(0.29倍,P<0.01)和ADAMTS-5的表达(0.10倍,P<0.01),与对照组的差异均具有统计学意义。同时,硝苯地平显著降低了NFAT1的表达,是对照组的0.20倍(P<0.05),但对NFAT2的m RNA表达没有影响。此外,硝苯地平使KOA组细胞膜电位显著增加(P<0.01)。[结论]L-型钙通道通过上调NFAT1.的m RNA表达参与调控KOA软骨细胞基质代谢。
[Objective] To explore the possible role of L-type calcium channel in regulation of matrix metabolism of chondrocytes via nuclear factor of activated T cell 1 （NFAT1） in rabbit knee osteoarthritis （KOA） model. [Methods] KOA model in rabbits was surgically established by Hulth method. The mRNA expression of five members of NFATs in normal and KOA chondrocytes were detected with qRT-PCR technique. After L-type voltage-gated calcium channel blocker, nifedipine （10μM） was added, the mRNA expressions of various subtypes of NFAT and matrix metabolism markers in KOA chon- drocytes were further measured. Additionally, the changes of membrane potential in KOA chondrocytes were detected by patch clamp technique. [Results] The mRNA expressions of NFAT1-NFAT5 could be detected in the normal and KOA chondro- cytes with incongruent variations. Compared with the normal cells, the NFAT1 expression in the KOA chondrocytes obviously increased （2.42 folds, P〈0.01）, while the NFAT2 reduced significantly （0.34 folds, P〈0.01）, but there were on significant differences in the mRNA expressions of NFAT3, NFAT4 and NFAT5 between the two groups. Compared with solvent control, 10μM nifedipine had no significantly impact on viability of the KOA chondrocytes in vitro, however completely blocked the mRNA expressions of calcium channel Cavl. In addition, nifedipine significantly increased the mRNA expressions of chondrocyte matrix components Col II （7.46 folds, P〈0.01）, Aggrecan （4.98 folds, P〈0.05） and TGF-β （4.37folds, P〈0.01）, and conversely, significantly decreased the mRNA expressions of MMP- 1 （0.17 folds, P〈0.01）, ADAMTS-4 （0.29fold, P〈0.01） andADAMTS-5 （0.10 folds, P〈0.01） .Concomitantly, nifedipine remarkably reduced the mRNA expressions of NFAT1 （0.20 folds, P〈0.01 ）, but had no effect on the expressions of NFAT2. Moreover, the KOA chondrocytes had a more hyperpolarized membrane potential in the nifedipine group than the solvent control group （P〈0.01） . [Conclusion] The role of L-type calcium channel on matrix metabolism of KOA chondrocytes is through upgrading the mRNA expression of NFAT 1.
出处
《中国矫形外科杂志》
CAS
CSCD
北大核心
2017年第13期1225-1230,共6页
Orthopedic Journal of China
基金
河北省科技计划项目(16277785D)
河北省高等学校科学技术研究项目(ZD20131059)
国家级大学生创新创业训练计划项目(2016100890014)
