Modulation of Endothelin-1-Induced Cytosolic Free Calcium Mobilization and Mitogen-Activated Protein Kinase Activation by Erythropoietin in Vascular Smooth Muscle Cells

Abstract

Background: It has been reported that human recombinant erythropoietin (rHuEPO) modulates the sensitivity of the cardiovascular system to vasoconstrictors. We investigated whether rHuEPO has modulative effects on the endothelin-1 (ET-1)-induced elevation of cytosolic free calcium concentration ([Ca²⁺]_i) and mitogen-activated protein (MAP) kinase activation in vascular smooth muscle cells (VSMC). Methods: [Ca²⁺]_i was measured by fura-2/AM, and MAP kinase activation was analyzed by Western blotting. Results: Exposure of VSMC to rHuEPO prior to stimulation with ET-1 enhanced both basal and ET-1-induced elevation of [Ca²⁺]_i in a dose-dependent manner in the presence of extracellular Ca²⁺. The synergistic effect was also retained in the absence of extracellular Ca²⁺ after exposure to rHuEPO. However, the effect was diminished in the presence of extracellular Ca²⁺ combined with the intracellular Ca²⁺ release inhibitor TMB-8, PKC inhibitor, or PKC depletion. Exposure to rHuEPO also had a synergistic effect on the activation of MAP kinase induced by ET-1; however, this effect was diminished in the presence of the Ca²⁺ chelator BAPTA-AM. Conclusion: The results suggest that rHuEPO has synergistic effects on ET-1-induced [Ca²⁺]_i mobilization, particularly on intracellular Ca²⁺ release, and MAP kinase activation in VSMC.