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

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.