Novel roles for voltage-gated T-type Ca2+ and ClC-2 channels in phagocytosis and angiogenic factor balance identified in human iPSC-derived RPE

Abstract

Human-induced pluripotent stem cell (hiPSC)-derived retinal pigment epithelium (RPE) is a powerful tool for pathophysiological studies and preclinical therapeutic screening, as well as a source for clinical cell transplantation. Thus, it must be validated for maturity and functionality to ensure correct data readouts and clinical safety. Previous studies have validated hiPSC-derived RPE as morphologically characteristic of the tissue in the human eye. However, information concerning the expression and functionality of ion channels is still limited. We screened hiPSC-derived RPE for the polarized expression of a panel of L-type (Ca_V 1.1, Ca_V 1.3) and T-type (Ca_V 3.1, Ca_V 3.3) Ca²⁺ channels, K⁺ channels (Maxi-K, Kir4.1, Kir7.1), and the Cl^- channel ClC-2 known to be expressed in native RPE. We also tested the roles of these channels in key RPE functions using specific inhibitors. In addition to confirming the native expression profiles and function of certain channels, such as L-type Ca²⁺ channels, we show for the first time that T-type Ca²⁺ channels play a role in both phagocytosis and vascular endothelial growth factor (VEGF) secretion. Moreover, we demonstrate that Maxi-K and Kir7.1 channels are involved in the polarized secretion of VEGF and pigment epithelium-derived factor (PEDF). Furthermore, we show a novel localization for ClC-2 channel on the apical side of hiPSC-derived RPE, with an overexpression at the level of fluid-filled domes, and demonstrate that it plays an important role in phagocytosis, as well as VEGF and PEDF secretion. Taken together, hiPSC-derived RPE is a powerful model for advancing fundamental knowledge of RPE functions.

Keywords: Ca2+ and K+ channels; ClC-2 chloride channel; iPSC-derived RPE; phagocytosis; secretion.