Mitochondrial Function Determines the Viability and Osteogenic Potency of Human Mesenchymal Stem Cells

Abstract

Advanced therapies medicinal products (ATMPs) have introduced innovative cell-based products. However, the regulatory demands for characterization of ATMPs are currently unable to adequately address the safety of such products. As recent studies have emphasized the role of mitochondria in the osteogenic differentiation of human mesenchymal stem cells (hMSCs), we have studied in detail the viability and osteogenic differentiation potency of the hMSCs intended for use as ATMPs based on analyses of the mitochondrial inner membrane potential (DeltaPsi(m)). Flow cytometric measurement of 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide (JC-1), propidium iodide fluorescence, and AnnexinV was employed to determine DeltaPsi(m), plasma membrane integrity, and organization of phosphatidylserine in plasma membrane, respectively, in cultured hMSCs. Apoptosis was induced by incubating cells at critical concentration (20 muM) of menadione. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was used as an indicator for cell proliferation and alkaline phosphatase activity and calcium deposition as indicators of osteogenic differentiation. Based on JC-1 fluorescence, cell morphology, organization of phosphatidylserine, and plasma membrane integrity, we could sort cells into four categories that represented different cell quality. A strong correlation between JC-1 and osteogenic differentiation was demonstrated for the first time and thus this analytical tool is suitable not only to determine cell viability but also to predict osteogenic differentiation of hMSC.

Mitochondrial membrane potential and reactive oxygen species in cancer stem cells

“…The Dwm is related to apoptosis due to the fact that dissipation of Dwm is a critical event in apoptosis process [11]. It is convincing that the high Dwm cell population possesses a stronger resistance to apoptotic inducers than low Dwm cell population in cultured hMSCs (human mesenchymal stem cells) [32]. The stronger apoptotic resistance of CSCs compared to the non-CSCs in human oral squamous cell carcinoma cell line Ho-1-N-1 was also related to Dwm [33].…”

Strategies for Isolating and Enriching Cancer Stem Cells: Well Begun Is Half Done

“…It is thought that mitochondrial function and integrity may affect SC viability, the proliferative and differentiation potential of these cells, and their lifespan [111]. Heerdt et al found that the mitochondrial membrane potential (Dcm) is associated with a cell's tumorigenicity.…”

Mitochondrial Function and Energy Metabolism in Umbilical Cord Blood- and Bone Marrow-Derived Mesenchymal Stem Cells

“…Mitochondrial inner membrane potential (DC m ) specific dye 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide ( JC-1; Sigma-Aldrich) was used as earlier described [30]. Cells were suspended in PBS and incubated with 1 mM JC-1 for 20 min at room temperature.…”

“…Recently, more attention has been focused on mitochondria and cellular bioenergetics of stem cells, which has revealed their unique metabolic status [27][28][29]. It has been shown that mitochondrial activity or dormancy plays a major role in maintaining the undifferentiated state, whereas the proper activation and function of mitochondria is essential during the differentiation processes [29][30][31]. In addition, mitochondrial reactive oxygen species (ROS) regulate the differentiation of several stem cell types; and recently, ROS have been shown to inhibit adhesion of hMSCs to the site of injury [32][33][34].…”