Increasing cytosolic Ca²⁺ levels restore cell proliferation and stem cell potency in aged MSCs

Aging is an inescapable complex physiological but extendable process, and all cells, including stem cells, are altered over time. Diverse mechanism(s) could modulate stem cell number, their proliferation rate, and promote tissue repair during aging that leads to longevity. However, the factors that could restore aging stem cell potency and would lead to healthy aging are not fully identified. Here we show that maintaining cytosolic Ca²⁺ levels was essential for modulating stem cells function in aged mesenchymal stem cells (MSCs). Increasing external Ca²⁺ induced spindle shape stem cell morphology and maintained stem cell surface marker expression in aged bone marrow-derived MSCs. Similarly, stem cell survival and proliferation of aged MSCs was dependent on cytosolic Ca²⁺ levels. Importantly, Ca²⁺ entry potentiated cell cycle progression, and stem cell potential was increased in cells incubated with higher external Ca²⁺. Moreover, blocking Ca²⁺ entry using SKF 96365, decreased stem cell survival and its proliferation but, treatment with 2-APB did not significantly affected cell proliferation, rather only modulated cell viability. Evaluation of Ca²⁺ entry channels, showed that TRPC1/Orai1/Orai3 and their regulator STIM1 was essential for MSCs proliferation/viability as gene silencing of Orai1/Orai3/TRPC1/STIM1 significantly inhibited stem cell viability. Finally, MSCs isolated from aged mice that were subjected to higher Ca²⁺ levels, were able to rescue age-induced loss of MSCs function. Together these results suggest that Ca²⁺ entry is essential for preventing the loss of aged stem cell function and supplementing Ca²⁺ not only restored their proliferative potential but, allowed them to develop into younger stem cell lineages that could be critical for regenerative medicine.