Internalization and Transportation of Endothelial Cell Surface K_Ca2.3 and K_Ca3.1 in Normal Pregnancy and Preeclampsia

Altered redox state modulates the expression levels of endothelial K_Ca2.3 and K_Ca3.1 (K_Cas) in normal pregnancy (NP) and preeclampsia (PE), thereby regulating vascular contractility. The mechanisms underlying K_Cas endocytosis and transportation remain unknown. We investigated the regulation of K_Cas expression in plasma membrane (PM) during NP and PE. Cultured human uterine artery endothelial cells were incubated in serum from normal nonpregnant women and women with NP or PE, or in oxidized LDL-, or lysophosphatidylcholine- (LPC-) containing a medium for 24 hours. NP serum elevated PM levels of K_Cas and reduced caveolin-1 and clathrin levels. PE serum, oxidized LDL, or LPC reduced PM levels of K_Cas and elevated caveolin-1, clathrin, Rab5c, and early endosome antigen-1 (EEA1) levels. Reduced K_Cas levels by PE serum or LPC were reversed by inhibition of caveolin-1, clathrin, or EEA1. Catalase and glutathione peroxidase 1 (GPX1) knockdown elevated PM-localized K_Cas levels and reduced caveolin-1 and clathrin levels. Elevated K_Ca2.3 levels upon catalase and GPX1 knockdown were reversed by PEG-catalase treatment. An H₂O₂ donor reduced clathrin and Rab5c. In contrast, elevated clathrin, caveolin-1, or colocalization of caveolin-1 with K_Ca3.1 by PE serum or LPC was reversed by NADPH oxidase inhibitors or antioxidants. A superoxide donor xanthine+xanthine oxidase elevated caveolin-1 or Rab5c levels. We concluded that K_Cas are endocytosed in a caveola- or a clathrin-dependent manner and transported in a Rab5c- and EEA1-dependent manner during pregnancy. The endocytosis and transportation processes may slow down via H₂O₂-mediated pathways in NP and may be accelerated via superoxide-mediated pathways in PE.