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

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, I_KCa3.1 ) blockers and corresponding gene (KCNN4)-expression knockdown. Depletion of Ca²⁺_i induced membrane-potential (V_mem ) depolarization, while store-operated Ca²⁺ entry (SOCE) hyperpolarized V_mem in both cell types. The hyperpolarizing SOCE effect was substantially reduced by I_KCa3.1 or SOCE blockade (TRAM-34, 2-APB), and I_KCa3.1 blockade (TRAM-34) or KCNN4-knockdown decreased the Ca²⁺ entry resulting from SOCE. I_KCa3.1 suppression reduced c-Kit⁺ eCPC and BM-MSC proliferation, while significantly altering the profile of cyclin expression. I_KCa3.1 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 I_KCa3.1 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.