Osteoporosis: the current status of mesenchymal stem cell-based therapy.

Osteoporosis, or bone loss, is a progressive, systemic skeletal disease that affects millions of people worldwide. Osteoporosis is generally age related, and it is underdiagnosed because it remains asymptomatic for several years until the development of fractures that confine daily life activities, particularly in elderly people. Most patients with osteoporotic fractures become bedridden and are in a life-threatening state. The consequences of fracture can be devastating, leading to substantial morbidity and mortality of the patients. The normal physiologic process of bone remodeling involves a balance between bone resorption and bone formation during early adulthood. In osteoporosis, this process becomes imbalanced, resulting in gradual losses of bone mass and density due to enhanced bone resorption and/or inadequate bone formation. Several growth factors underlying age-related osteoporosis and their signaling pathways have been identified, such as osteoprotegerin (OPG)/receptor activator of nuclear factor B (RANK)/RANK ligand (RANKL), bone morphogenetic protein (BMP), wingless-type MMTV integration site family (Wnt) proteins and signaling through parathyroid hormone receptors. In addition, the pathogenesis of osteoporosis has been connected to genetics. The current treatment of osteoporosis predominantly consists of antiresorptive and anabolic agents; however, the serious adverse effects of using these drugs are of concern. Cell-based replacement therapy via the use of mesenchymal stem cells (MSCs) may become one of the strategies for osteoporosis treatment in the future.

Cardiomyocyte differentiation of perinatally­derived mesenchymal stem cells.

Coronary heart disease is major cause of mortality worldwide and several risk factors have been shown to play a role in its pathogenesis, including smoking, obesity, hypertension and hypercholesterolemia. A number of therapeutic methods have been developed to improve the quality of patients' lives, including stem cell therapy using mesenchymal stem cells (MSCs). Perinatal sources, including the placenta (PL) and umbilical cord (UC), are rich sources of MSCs and have been identified as a potential source of cells for therapeutic use. Their role in cardiogenic differentiation is also of contemporary medical interest. The present study demonstrated the induced differentiation of MSCs obtained from the UC, PL and Wharton's jelly (WJ) into cardiomyocytes, using 10 µM 5­azacytidine. The characteristics of the MSCs from each source were studied and their morphology was compared. An immunofluorescence analysis for the cardiac­specific markers, GATA4 and Troponin T (TnT), was performed and tested positive in all sources. The expression of the cardiac­specific genes, Nkx2.5, α­cardiac actin and TnT, was analyzed by real­time RT­PCR and presented as fold change increases. The expression of each of the markers was observed to be higher in the 5­azacytidine­treated MSCs. The differences in expression among the sources of treated MSCs was as follows: TnT had the highest level of expression in the bone marrow (BM) MSCs; α­cardiac actin had the highest level of expression in the PLMSCs; and all the genes were expressed at significantly high levels in the WJMSCs compared with the control group. The present study showed the ability of alternative perinatally­derived MSCs to differentiate into cardiomyocyte­like cells and how this affects the therapeutic use of these cells.

Cardiogenic and myogenic gene expression in mesenchymal stem cells after 5-azacytidine treatment.

OBJECTIVE: 5-Azacytidine is a hypomethylating agent that is used for the treatment of myelodysplastic syndrome. This histone modifier is widely employed and plays a nonspecific role in influencing the differentiation capability of stem cells. The ability of bone marrow mesenchymal stem cells to differentiate into cardiomyocyte- and myocyte-like cells after exposure to 3 different doses of 5-azacytidine has been evaluated and compared. The aim of the study was to optimize the effective dose of 5-azacytidine for promoting the cardiomyocyte and myocyte differentiation capabilities of human mesenchymal stem cells (MSCs). MATERIALS AND METHODS: Human bone marrow aspirations were collected from healthy donors. MSCs were used for the study of mesodermal differentiation. MSCs were cultured to promote osteoblast differentiation and adipocyte differentiation. The evaluation of osteogenic or adipogenic properties was then performed through immunocytochemical staining. BMMSCs were trypsinized into single-cell suspensions and then prepared for flow cytometric analysis. The MSCs were treated with 5, 10, or 15 µM 5-azacytidine for 24 h and then cultured for 3 weeks. Total RNA was extracted from untreated and 5-azacytidine-treated cells. Troponin T and GATA4 antibodies were used as cardiogenic markers, whereas myogenin and MyoD antibodies were used as myocyte markers. RESULTS: The morphology and growth rate of MSCs that were treated with any of the 3 doses of 5-azacytidine were similar to the morphology and growth rate of control MSCs. An immunofluorescence analysis examining the expression of the cardiac-specific markers GATA4 and troponin T and the skeletal muscle-specific markers MyoD and myogenin revealed that cells treated with 15 µM 5-azacytidine were strongly positive for these markers. Real-time RT-PCR results were examined; these amplifications indicated that there were higher expression levels of cardiac- and skeletal muscle-specific mRNAs in MSCs treated with 15 µm 5-azacytidine than in MSCs that had either been treated with lower doses of 5-azacytidine or left untreated. CONCLUSION: MSCs treated with 5-azacytidine demonstrated the capacity to differentiate into both cardiomyocytes and skeletal myocytes, and 15 µM 5-azacytidine could be the optimal dose of this drug. Other promoting factors should be examined to investigate the possibility of promoting the differentiation of MSCs into specific cell types. CONFLICT OF INTEREST: None declared.