Extracellular acidosis suppresses endothelial function by inhibiting store-operated Ca2+ entry via non-selective cation channels
Extracellular acidosis suppresses endothelial function by inhibiting store-operated Ca2+ entry via non-selective cation channels
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DOI：
10.1093/cvr/cvp105
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发表时间：
2009-07-01
影响因子：
10.8
通讯作者：
Watanabe, Hiroshi
中科院分区：
文献类型：
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作者：
Asai, Masayoshi;Takeuchi, Kazuhiko;Watanabe, Hiroshi
目的缺氧、缺血和外源性化学物质可引起细胞外和细胞内酸中毒，但目前尚不清楚哪种类型的酸中毒会影响内皮细胞功能。内皮源性舒张因子（EDRF）的合成和释放与细胞内Ca 2+浓度的增加有关，因此我们研究了细胞外和细胞内酸中毒对培养的猪主动脉内皮细胞中Ca 2+反应和EDRF产生的影响。方法和结果使用荧光染料测量细胞内pH（pH（i））和Ca 2+，BCECM/AM（pH指示剂）和fura-2/AM（Ca 2+指示剂）。EDRF、一氧化氮（NO）和前列腺素I-2（PGI（2））分别采用DAF-FM/DA（NO指示染料）荧光测定法和6-酮PGF（1 α）酶免疫测定法进行评估。使用滴定至pH 6.4、6.9和7.4的HEPES缓冲液改变细胞外pH（pH（o）），并使用丙酸盐（20 mmol/L）引起细胞内酸中毒。细胞外酸中毒强烈抑制缓激肽（BK，10 nmol/L）和毒胡萝卜素（TG，1 μ mol/L）诱导的Ca 2+反应的30%和23%，在pH（o）6.9和80%和97%，分别在pH（o）6.4。在检查期间，在pH（o）7.4、6.9和6.4下，三组之间的pHi无显著差异。细胞外酸中毒也抑制BK刺激的PGI（2）的产生，在pH（o）6.9时抑制55%，在pH（o）6.4时抑制77%，在pH（o）6.9时抑制38%，在pH（o）6.4时抑制91%。细胞外酸中毒对Ca ~（2+）反应和NO产生的抑制作用是可逆的。丙酸盐将pHi从7.3改变为6.9，而不改变pH（o）（7.4）。细胞内酸中毒没有影响BK和TG诱导的Ca 2+反应或NO production.Conclusion这些结果表明，细胞外，而不是细胞内，酸中毒导致内皮功能障碍，通过抑制存储操作的Ca 2+进入，因此有助于澄清血管病理生理条件，如缺血，缺氧，酸中毒，缺血再灌注。
Aims Hypoxia, ischaemia, and exogenous chemicals can induce extracellular and intracellular acidosis, but it is not clear which of these types of acidosis affects endothelial cell function. The synthesis and release of endothelium-derived relaxing factors (EDRFs) are linked to an increase in cytosolic Ca2+ concentration, and we therefore examined the effects of extracellular and intracellular acidosis on Ca2+ responses and EDRF production in cultured porcine aortic endothelial cells.Methods and results Cytosolic pH (pH(i)) and Ca2+ were measured using fluorescent dyes, BCECM/AM (pH-indicator) and fura-2/AM (Ca2+-indicator), respectively. EDRFs, nitric oxide (NO) and prostaglandin I-2 (PGI(2)) were assessed using DAF-FM/DA (NO-indicator dye) fluorometry and 6-keto PGF(1 alpha) enzyme immunoassay, respectively. HEPES buffers titrated to pH 6.4, 6.9, and 7.4 were used to alter extracellular pH (pH(o)), and propionate (20 mmol/L) was applied to cause intracellular acidosis. Extracellular acidosis strongly suppressed bradykinin (BK, 10 nmol/L)- and thapsigargin (TG, 1 mu mol/L)-induced Ca2+ responses by 30 and 23% at pH(o) 6.9, and by 80 and 97% at pH(o) 6.4, respectively. During the examinations, there were no significant differences in pHi among the three groups at pH(o) 7.4, 6.9, and 6.4. Extracellular acidosis also inhibited BK-stimulated PGI(2) production by 55% at pH(o) 6.9 and by 77% at pH(o) 6.4, and NO production by 38% at pH(o) 6.9 and by 91% at pH(o) 6.4. The suppressive effects of extracellular acidosis on Ca2+ responses and NO production were reversible. Propionate changed pHi from 7.3 to 6.9, without altering pH(o) (7.4). Intracellular acidosis had no effect on BK- and TG-induced Ca2+ responses or NO production.Conclusion These results indicate that extracellular, but not intracellular, acidosis causes endothelial dysfunction by inhibiting store-operated Ca2+ entry, so helping to clarify the vascular pathophysiology of conditions such as ischaemia, hypoxia, acidosis, and ischaemia-reperfusion.