Calcium-dependent potassium channels control proliferation of cardiac progenitor cells and bone marrow-derived mesenchymal stem cells.

ABSTRACT

KEY POINTS Ex vivo proliferated c-Kit+ endogenous cardiac progenitor cells (eCPCs) obtained from mouse and human cardiac tissues have been reported to express a wide range of functional ion channels. In contrast to previous reports in cultured c-Kit+ eCPCs, we found that ion currents were minimal in freshly isolated cells. However, inclusion of free Ca2+ intracellularly revealed a prominent inwardly rectifying current identified as the intermediate conductance Ca2+ -activated K+ current (KCa3.1) Electrical function of both c-Kit+ eCPCs and bone marrow-derived mesenchymal stem cells is critically governed by KCa3.1 calcium-dependent potassium channels. Ca2+ -induced increases in KCa3.1 conductance are necessary to optimize membrane potential during Ca2+ entry. Membrane hyperpolarization due to KCa3.1 activation maintains the driving force for Ca2+ entry that activates stem cell proliferation. Cardiac disease downregulates KCa3.1 channels in resident cardiac progenitor cells. Alterations in KCa3.1 may have pathophysiological and therapeutic significance in regenerative medicine. ABSTRACT Endogenous c-Kit+ cardiac progenitor cells (eCPCs) and bone marrow (BM)-derived mesenchymal stem cells (MSCs) are being developed for cardiac regenerative therapy, but a better understanding of their physiology is needed. Here, we addressed the unknown functional role of ion channels in freshly isolated eCPCs and expanded BM-MSCs using patch-clamp, microfluorometry and confocal microscopy. Isolated c-Kit+ eCPCs were purified from dog hearts by immunomagnetic selection. Ion currents were barely detectable in freshly isolated c-Kit+ eCPCs with buffering of intracellular calcium (Ca2+i ). Under conditions allowing free intracellular Ca2+ , freshly isolated c-Kit+ eCPCs and ex vivo proliferated BM-MSCs showed prominent voltage-independent conductances that were sensitive to intermediate-conductance K+ -channel (KCa3.1 current, IKCa3.1 ) blockers and corresponding gene (KCNN4)-expression knockdown. Depletion of Ca2+i induced membrane-potential (Vmem ) depolarization, while store-operated Ca2+ entry (SOCE) hyperpolarized Vmem in both cell types. The hyperpolarizing SOCE effect was substantially reduced by IKCa3.1 or SOCE blockade (TRAM-34, 2-APB), and IKCa3.1 blockade (TRAM-34) or KCNN4-knockdown decreased the Ca2+ entry resulting from SOCE. IKCa3.1 suppression reduced c-Kit+ eCPC and BM-MSC proliferation, while significantly altering the profile of cyclin expression. IKCa3.1 was reduced in c-Kit+ eCPCs isolated from dogs with congestive heart failure (CHF), along with corresponding KCNN4 mRNA. Under perforated-patch conditions to maintain physiological [Ca2+ ]i , c-Kit+ eCPCs from CHF dogs had less negative resting membrane potentials (-58 ± 7 mV) versus c-Kit+ eCPCs from control dogs (-73 ± 3 mV, P < 0.05), along with slower proliferation. Our study suggests that Ca2+ -induced increases in IKCa3.1 are necessary to optimize membrane potential during the Ca2+ entry that activates progenitor cell proliferation, and that alterations in KCa3.1 may have pathophysiological and therapeutic significance in regenerative medicine.