南方医科大学学报 ›› 2015, Vol. 35 ›› Issue (07): 960-.

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哇巴因对大鼠血管平滑肌细胞内钙离子浓度的影响

张明娟,张美程,张超英,杨军,朱参战,段宗明   

  • 出版日期:2015-07-20 发布日期:2015-07-20

Effect of ouabain on intracellular Ca2+ concentration in rat vascular smooth muscle cells in
vitro

  • Online:2015-07-20 Published:2015-07-20

摘要: 目的探讨哇巴因对大鼠胸主动脉血管平滑肌细胞(vascular smooth muscle cells, VSMCs)内Ca2+浓度的影响。方法用组织
块贴壁法进行大鼠胸主动脉VSMCs原代培养,用免疫组织化学方法进行细胞鉴定,采用放射自显影术检测哇巴因对大鼠胸主动
脉VSMCs的结合能力,采用Fluo 3-AM(钙离子荧光探针)法研究哇巴因在短时间内对VSMCs细胞内Ca2+浓度的影响,探讨不同
浓度的钠泵α2亚单位截断性片段的拮抗作用。结果(1)放射自显影术检测结果显示,哇巴因与VSMCs间有一定的结合能力。
在0.1~100 nmol/L浓度范围,随着哇巴因浓度的增加,VSMCs与哇巴因的结合能力增加;(2)Fluo 3-AM检测VSMCs内Ca2+浓度
结果显示:不同浓度的哇巴因在短时间内能够引起VSMCs内Ca2+浓度的变化。在0~3200 nmol/L浓度范围,细胞内Ca2+离子浓
度会随着哇巴因浓度的增加呈现出逐渐升高的趋势;(3)不同浓度的钠泵α2亚单位截断性片段可以拮抗哇巴因引起的VSMCs内
Ca2+浓度的升高作用。结论哇巴因引起的细胞内高钙是高哇巴因高血压发生的细胞学基础,钠泵α2亚单位截断性片段可以拮
抗哇巴因引起的VSMCs内Ca2+浓度的升高作用,为进一步探讨哇巴因的作用机制结高血压的治疗提供了实验依据。

Abstract: Objective To explore the effect of ouabain on intracellular Ca2 + concentration ([Ca2 + ]i) in thoracic aorta vascular
smooth muscle cells (VSMCs) in vitro. Methods Primary SD rat thoracic aorta VSMCs were cultured by tissue adherent method
and identified by immunochemistry. The binding ability between ouabain and VSMCs was detected by autoradiography, and
fluo 3-AM (a Ca2 + fluorescent probe) was employed to investigate whether ouabain affected VSMCs within a short period of
time. The effect of a truncated fragment of the sodium pump α2 subunit was assayed in antagonizing the effect of ouabain on
[Ca2+]i in the VSMCs. Results Within the concentration range of 0.1-100 nmol/L, ouabain was found to dose-dependently bind
to the VSMCs. Different concentrations of ouabain (0-3200 nmol/L) caused a transient, dose-dependent increase in [Ca2 + ]i in
the VSMCs, which was antagonized by the application of the truncated fragment of sodium pump α2 subunit. Conclusions
Elevations in [Ca2+]i in the VSMCs can be the cytological basis of high ouabain-induced hypertension. The truncated fragment
of the sodium pump α2 subunit can antagonize ouabain-induced increase of [Ca2 + ]i in the VSMCs, which provides a clue for
understanding the pathogenesis of and devising a therapeutic strategy for high ouabain-induced hypertension.