Mitochondrial function and energy metabolism in umbilical cord blood- and bone marrow-derived mesenchymal stem cells.

Human mesenchymal stem cells (hMSCs) are an attractive choice for a variety of cellular therapies. hMSCs can be isolated from many different tissues and possess unique mitochondrial properties that can be used to determine their differentiation potential. Mitochondrial properties may possibly be used as a quality measure of hMSC-based products. Accordingly, the present work focuses on the mitochondrial function of hMSCs from umbilical cord blood (UCBMSC) cells and bone marrow cells from donors younger than 18 years of age (BMMSC <18) and those more than 50 years of age (BMMSC >50). Changes of ultrastructure and energy metabolism during osteogenic differentiation in all hMSC types were studied in detail. Results show that despite similar surface antigen characteristics, the UCBMSCs had smaller cell surface area and possessed more abundant rough endoplasmic reticulum than BMMSC >50. BMMSC <18 were morphologically more UCBMSC-like. UCBMSC showed dramatically higher mitochondrial-to-cytoplasm area ratio and elevated superoxide and manganese superoxide dismutase (MnSOD) levels as compared with BMMSC >50 and BMMSC <18. All hMSCs types showed changes indicative of mitochondrial activation after 2 weeks of osteogenic differentiation, and the increase in mitochondrial-to-cytoplasm area ratio appears to be one of the first steps in the differentiation process. However, BMMSC >50 showed a lower level of mitochondrial maturation and differentiation capacity. UCBMSCs and BMMSCs also showed a different pattern of exocytosed proteins and glycoproteoglycansins. These results indicate that hMSCs with similar cell surface antigen expression have different mitochondrial and functional properties, suggesting different maturation levels and other significant biological variations of the hMSCs. Therefore, it appears that mitochondrial analysis presents useful characterization criteria for hMSCs intended for clinical use.

Mitochondrial function determines the viability and osteogenic potency of human mesenchymal stem cells.

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.