Depolarization alters phenotype, maintains plasticity of pre-differentiated mesenchymal stem cells

Abstract

Although adult stem cell transplantation has been implemented as a therapy for tissue repair, it is limited by the availability of functional adult stem cells. A potential approach to generate stem and progenitor cells may be to modulate the differentiated status of somatic cells. Therefore, there is a need for a better understanding of how the differentiated phenotype of mature cells is regulated. We hypothesize that bioelectric signaling plays an important role in the maintenance of the differentiated state, as it is a functional regulator of the differentiation process in various cells and tissues. In this study, we asked whether the mature phenotype of osteoblasts and adipocytes derived from human mesenchymal stem cells (hMSCs) could be altered by modulation of their membrane potential. hMSC-derived osteoblasts and adipocytes were depolarized by treatment with ouabain, a Na + /K + ATPase inhibitor, or by treatment with high concentrations of extracellular K + . To characterize the effect of voltage modulation on the differentiated state, the depolarized cells were evaluated for (1) the loss of differentiation markers; (2) the up-regulation of stemness markers and stem properties; and (3) differences in gene expression profiles in response to voltage modulation. hMSC-derived osteoblasts and adipocytes exhibited significant down-regulation of bone and fat tissue markers in response to depolarization, despite the presence of differentiation-inducing soluble factors, suggesting that bioelectric signaling overrides biochemical signaling in the maintenance of cell state. Suppression of the osteoblast or adipocyte phenotype was not accompanied by upregulation of genes associated with the stem state. Thus, depolarization does not activate the stem cell genetic signature and, therefore, does not induce a full reprogramming event. However, after transdifferentiating the depolarized cells to evaluate for multi-lineage potential, depolarized osteoblasts demonstrated improved ability to achieve correct adipocyte morphology compared with nondepolarized osteoblasts. The present study thus demonstrates that depolarization reduces the differentiated phenotype of hMSC-derived cells and improves their transdifferentiation capacity, but does not restore a stem-like genetic profile. Through global transcript profiling of depolarized osteoblasts, we identified pathways that may mediate the effects of voltage signaling on cell state, which will require a detailed mechanistic inquiry in future studies.

Assessment of Enrichment of Human Mesenchymal Stem Cells Based on Plasma and Mitochondrial Membrane Potentials

“…[47][48][49][50][51][52][53][54][55] In terms of stem cell state, undifferentiated stem cells with greater proliferative capacity (than their differentiated progeny) have been suggested to be associated with a depolarized plasma membrane, while terminally differentiated and quiescent cells tend to be hyperpolarized. 53 This relationship may be true for hMSCs as well; modulation of plasma membrane potential via pharmaceuticals or changing the ionic milieu has been shown to be able to override biochemical signaling to prevent hMSC differentiation, 47 or suppress their differentiated state, 56 even in the presence of the most potent chemical mediators known to drive differentiation down certain lineages. In addition, mitochondrial membrane potential may also be involved in differentiation and immunomodulatory functions of hMSCs.…”

Measurement of ATP-Induced Membrane Potential Changes in IVD Cells

“…The combined actions of ion channels and transporters establish membrane potentials by regulating intracellular and extracellular ionic concentrations 10, 11 . Increasing evidence suggests that membrane potentials are not only involved in the cellular processes of proliferation, migration and differentiation 12–14 , but are also related to complex processes such as control of wound healing 15, 16 , regeneration 17, 18 , left-right patterning 19 and development 15 . Thus, it is suggested that membrane potentials are a fundamental control mechanism of cell functions 10 .…”