Acute hypoxic increases in labile zinc in cultured pulmonary endothelial cells are associated with activation of PKC‐epsilon

We have recently demonstrated that acute hypoxia increases labile zinc in an NO‐dependent fashion in cultured pulmonary endothelial cells. Although metallothionein appeared to be the source of labile zinc under these conditions, the cellular targets for this new pool of zinc remain unclear. We hypothesized that PKC‐ ε, a zinc dependent enzyme that others have shown is critical in hypoxic pulmonary vasoconstriction, may be one such target. Microspectrofluorometric analysis of cultured lung endothelial cells from several species incubated with the zinc sensitive fluorophore, Fluozin3, and exposed to low oxygen (P O2 =24mmHg) revealed an acute increase in labile (i.e. TPEN chelatable) [Zn 2+ ] I that was sensitive to L‐NAME (and partially reversible with supplemental L‐arginine). Exposure of rat pulmonary artery endothelial cells (RPACE) to exogenous zinc (1–10 μM) or hypoxia was associated with increases in PKC‐ε activity as determined by translocation of immunoreactive PKC‐ε from the cytosol to the plasma membrane and enhanced kinase activity associated with fractions enriched in PKC‐ε by immunoprecipitation. The hypoxic (and zinc) dependent increases in PKC‐ε was sensitive to TPEN suggesting that the new compartment of chelatable zinc in response to hypoxia in pulmonary endothelial cells is associated with zinc dependent activation of a critical signaling molecule in vascular response to low oxygen. Future studies are designed to examine the contribution of this pathway to vasomotor responses to low oxygen. This research was supported in part by NIH HL 65697 (BRP) and Parker B. Francis FNDT.