Extracellular acidosis suppresses endothelial function by inhibiting store-operated Ca2+ entry via non-selective...

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 Ca(2+) concentration, and we therefore examined the effects of extracellular and intracellular acidosis on Ca(2+) responses and EDRF production in cultured porcine aortic endothelial cells.

Cytosolic pH (pH(i)) and Ca(2+) were measured using fluorescent dyes, BCECM/AM (pH-indicator) and fura-2/AM (Ca(2+)-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(1alpha) 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 micromol/L)-induced Ca(2+) 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 pH(i) 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 Ca(2+) responses and NO production were reversible. Propionate changed pH(i) from 7.3 to 6.9, without altering pH(o) (7.4). Intracellular acidosis had no effect on BK- and TG-induced Ca(2+) responses or NO production.

These results indicate that extracellular, but not intracellular, acidosis causes endothelial dysfunction by inhibiting store-operated Ca(2+) entry, so helping to clarify the vascular pathophysiology of conditions such as ischaemia, hypoxia, acidosis, and ischaemia-reperfusion.