Effect of cyclic nucleotides on PGI2 generation of cultured human vascular endothelial cells with references to Ca++ kinetics.

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Effects of cyclic nucleotides on PGI2 generaton were investigated with references to Ca++ kinetics using cultured human vascular endothelial cell. Endothelial cells were isolated and cultured by the modified method of Jaffe et al. 6-keto PGF1α and cyclic nucleotides concentration were measured by RIA. Ca++ kinetics or cytosolic free Ca++ concentration ([Ca++]i) was measured using 45Ca or a new fluorescence indicator, Fura 2. Ca ionophore A23187 increased 45Ca uptake and release, or [Ca++]i, and it also enhanced PGI2 generation. The pretreatment of the cells with a Ca++ immobilizer, TMB-8 decreased all of them. Dibutyryl cAMP or 8-bromoc GMP increased intracellular cAMP or cGMP concentration, respectively. While dibutyryl cAMP increased 45Ca uptake or release, and decreased [Ca++]i, 8-bromo cGMP decreased 45Ca uptake, 45Ca release, and [Ca++]i. And administration of both these cyclic nucleotides analogues decreased PGI2 generation. Through these results it was concluded that: 1) PGI2 generation of cultured human vascular endothelial cells was brought about by the increase of [Ca++]i via the increase of Ca++ uptake or the succeeded enhancement of the release of Ca++ from the storage sites. 2) The direct evidence was shown that PGI2 generation was suppressed by the decrease of [Ca++]i via the increase of intracellular cyclic nucleotides concentration.
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