孙静磊;陈康;秦志明;常延超;武文婷;王晓峰;师晨霞;

• 1:河北医科大学
• 2:河北医科大学第三医院关节外科

摘要(Abstract)：

[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。

关键词(KeyWords)： 膝骨关节炎;基质代谢;L-型钙通道;钙活化T细胞核因子(NFATs);膜电位

Abstract:

Keywords:

基金项目(Foundation): 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)

作者(Author): 孙静磊;陈康;秦志明;常延超;武文婷;王晓峰;师晨霞;

Email:

DOI:

参考文献(References)：

• [1]Sitara D,Aliprantis AO.Transcriptional regulation of bone and joint remodeling by NFAT[J].Immunol Rev,2010,233(1):286-300.
• [2]Jahr H,Matta C,Mobasheri A.Physicochemical and biomechanical stimuli in cell-based articular cartilage repair[J].Curr Rheumatol Rep,2015,17(3):22.
• [3]Matta C,Zákány R,Mobasheri A.Voltage-dependent calcium channels in chondrocytes:roles in health and disease[J].Curr Rheumatol Rep,2015,17(7):43.
• [4]王晓峰,武文婷,常延超,等.骨关节炎软骨细胞电压依赖性钙通道α1亚单位表达的改变[J].中华实验外科杂志,2016,33(7):1727-1730.
• [5]Ranger AM,Gerstenfeld LC,Wang J,et al.The nuclear factor of activated T cells(NFAT)transcription factor NFATp(NFATc2)is a repressor of chondrogenesis[J].J Exp Med,2000,191(1):9-22.
• [6]Thirunavukkarasu K,Pei Y,Moore TL,et al.Regulation of the human ADAMTS-4 promoter by transcription factors and cytokines[J].Biochem Biophys Res Commun,2006,345(2):197-204.
• [7]Yaykasli KO,Oohashi T,Hirohata S,et al.ADAMTS9 activation by interleukin 1 beta via NFATc1 in OUMS-27 chondrosarcoma cells and in human chondrocytes[J].Mol Cell Biochem,2009,323:69-79.
• [8]Sun R,Zhang B,Chen L,et al.Role of nuclear factor of activated T cells 1 in the pathogenesis of osteoarthritis[J].Exp Ther Med,2014,7(1):195-198.
• [9]Wang J,Gardner BM,Lu Q,et al.Transcription factor Nfat1 deficiency causes osteoarthritis through dysfunction of adult articular chondrocytes[J].J Pathol,2009,219(2):163-172.
• [10]Greenblatt MB,Ritter SY,Wright J,et al.NFATc1 and NFATc2 repress spontaneous osteoarthritis[J].Proc Natl Acad Sci U S A,2013,110(49):19914-19919.
• [11]Tomita M,Reinhold MI,Molkentin JD,et al.Calcineurin and NFAT4 induce chondrogenesis[J].J Biol Chem,2002,277(44):42214-42218.
• [12]Tardif G,Pelletier JP,Fahmi H,et al.NFAT3 and TGF-β/SMAD3regulate the expression of mi R-140 in osteoarthritis[J].Arthritis Res Ther,2013,15(6):R197.
• [13]Tomida T,Hirose K,Takizawa A,et al.NFAT functions as a working memory of Ca2+signals in decoding Ca2+oscillation[J].EMBO J,2003,22(15):3825-3832.
• [14]Lin SS,Tzeng BH,Lee KR,et al.Cav3.2 T-type calcium channel is required for the NFAT-dependent Sox9 expression in tracheal cartilage[J].Proc Natl Acad Sci U S A,2014,111(19):1990-1998.
• [15]Nguyen C,Lieberherr M,Bordat C,et al.Intracellular calcium oscillations in articular chondrocytes induced by basic calcium phosphate crystals lead to cartilage degradation[J].Osteoarthritis Car-tilage,2012,20(11):1399-1408.
• [16]Boileau C,Martel-Pelletier J,Brunet J,et al.PD-0200347,an alpha2delta ligand of the voltage gated calcium channel,inhibits in vivo activation of the Erk1/2 pathway in osteoarthritic chondrocytes:a PKCalpha dependent effect[J].Ann Rheum Dis,2006,65(5):573-580.
• [17]Navedo MF,Hell JW.AKAP5 keeps L-type channels and NFAT on their toes[J].Cell Rep,2014,7(5):1341-1342.
• [18]Hagenfeld D,Borkenhagen B,Schulz T,et al.Hyaluronan export through plasma membranes depends on concurrent K+efflux by K(ir)channels[J].PLo S One,2012,7(6):e39096.
• [19]Suzuki Y,Ohya S,Yamamura H.A new splice variant of large conductance Ca2+-activated K+(BK)channelαsubunit alters human chondrocyte function[J].J Biol Chem,2016,291(46):24247-24260.
• [20]Lewis R,Barrett-Jolley R.Changes in membrane receptors and ion channels as potential biomarkers for osteoarthritis[J].Front Physiol,2015,6(12):357.