高血压大鼠动脉平滑肌细胞钙超载与腺苷三磷酸酶

Calcium Overload and Adenosinetriphosphatase in Artery Smooth Muscle Cells in Spontaneously Hypertensive Rats

目的探讨自发性高血压大鼠(SHR)动脉血管平滑肌细胞钙超载与腺苷三磷酸酶(AT-Pase)的关系。方法组织块种植法培养SHR和Wistar-Kyoto(WKY)大鼠胸主动脉平滑肌细胞,应用流式细胞术、生化酶学方法和逆转录聚合酶链反应(RT-PCR)技术等检测SHR和WKY大鼠动脉平滑肌细胞内游离钙浓度([Ca2+]i)、细胞膜Ca2+-ATPase和Na+-K+-ATPase活性及其mRNA表达水平。结果SHR动脉平滑肌细胞[Ca2+]i浓度高于WKY大鼠[(307.7±32.1vs118.9±21.4)nmol/L,P<0.01)],Ca2+-ATPase和Na+-K+-ATPase活性低于WKY大鼠(1.3±0.2vs2.6±0.3;3.1±0.5vs4.9±0.5,P均<0.01),细胞质膜Ca2+-ATPase(plasmamembraneCa2+-ATPase,PMCA)亚型1和Na+-K+-ATPaseα1亚单位mRNA表达低于WKY大鼠(0.231±0.007vs0.403±0.021;0.253±0.028vs0.634±0.033,P均<0.01)。结论SHR动脉平滑肌细胞出现钙超载,其机制可能部分与细胞膜PMCA和Na+-K+-ATPase活性降低有关,两种ATPase功能降低可能是其基因转录水平下调的结果。

Objective To investigate the relationship of calcium overload and adenosine triphosphatase （ ATPase ） in artery smooth muscle cells （ ASMCs） isolated from spontaneously hypertensive rats （SHR）. Methods The thoracic ASMCs isolated from SHR and Wistar-Kyoto （ WKY ） rats were cultured. Intracellular free Ca^2＋ concentration （ [Ca^2＋ ]i ） was detected by flow cytometry. The activity of Ca^2＋ -ATPase and Na^＋-K^＋-ATPase was measured by biochemistry and enzymology. Reverse transcriptase-polymerase chain reaction （RT-PCR） assay was employed to determine the levels of plasma membrane Ca^2＋-ATPase isoform 1 （ PMCA1 ） and Na^＋-K^＋-ATPasea1-subunit mRNA. Results [Ca2＋ -]i in ASMCs of SHR was higher than those from WKY （307.7±32. 1 vs 118.9±21.4 nmol/L, P 〈0.01）. The activities of Ca^2＋-ATPase and Na^＋-K^＋-ATPase in ASMCs from SHR were lower than those from WKY （1.3±0.2 vs 2. 6±0.3; 3. 1±0.5 vs 4.9±0. 5,P〈0.01, respectively）. Meanwhile, the levels of PMCA1 and Na^＋-K^＋-ATPase a1-subunit mRNA were significantly decreased in SHR ASMCs compared with WKY （PMCA1 mRNA 0. 231±0. 007 vs 0. 403±0. 021;Na^＋ -K^＋ ATPase a1-subunit mRNA 0. 253±0. 028 vs 0. 634±0. 033, P〈0.01）. Conclusion Calcium overload in ASMCs of SHR may be partially ascribed to the decreased activities of PMCA and Na^＋-K^＋-ATPase.