In Vivo Ca(V)3 Channel Inhibition Promotes Maturation of Glucose-Dependent Ca(2+) Signaling in Human iPSC-Islets

Ca(V)3 channels are ontogenetically downregulated with the maturation of certain electrically excitable cells, including pancreatic β cells. Abnormally exaggerated Ca(V)3 channels drive the dedifferentiation of mature β cells. This led us to question whether excessive Ca(V)3 channels, retained mistakenly in engineered human-induced pluripotent stem cell-derived islet (hiPSC-islet) cells, act as an obstacle to hiPSC-islet maturation. We addressed this question by using the anterior chamber of the eye (ACE) of immunodeficient mice as a site for recapitulation of in vivo hiPSC-islet maturation in combination with intravitreal drug infusion, intravital microimaging, measurements of cytoplasmic-free Ca(2+) concentration ([Ca(2+)](i)) and patch clamp analysis. We observed that the ACE is well suited for recapitulation, observation and intervention of hiPSC-islet maturation. Intriguingly, intraocular hiPSC-islet grafts, retrieved intact following intravitreal infusion of the Ca(V)3 channel blocker NNC55-0396, exhibited decreased basal [Ca(2+)](i) levels and increased glucose-stimulated [Ca(2+)](i) responses. Insulin-expressing cells of these islet grafts indeed expressed the NNC55-0396 target Ca(V)3 channels. Intraocular hiPSC-islets underwent satisfactory engraftment, vascularization and light scattering without being influenced by the intravitreally infused NNC55-0396. These data demonstrate that inhibiting Ca(V)3 channels facilitates the maturation of glucose-activated Ca(2+) signaling in hiPSC-islets, supporting the notion that excessive Ca(V)3 channels as a developmental error impede the maturation of engineered hiPSC-islet insulin-expressing cells.