Comparative Effect of Hypoxia and Ascorbic Acid for Cardiomyocyte Differentiation of WhartonʼS Jelly MesenchymalStem Cells

Background: ۵-azacytidine is a major classical factor for myogenicinduction, but this drug is toxic. Many studies investigatethe efficacy of the other alternative factors for cardiomyocytedifferentiation. The present study aims to survey the effect ofvarious cocktails, containing hypoxia (Hypo) and ascorbic acid(Asc) for cardiomyocyte differentiation of whartonʼs jelly mesenchymalstem cells (WJMSCs).Materials and Methods: WJMSCs were isolated by explantculture. They were characterized by osteogenic and adipogenicdifferentiation and flow cytometry of mesenchymal markers.Heart conditioned medium (HCM) was collected from neonatalrat heart. ۸ cocktail of ۵-azacytidine (۵-aza) and HCM wereused including: ۱) [HCM], ۲) [HCM + Asc], ۳) [HCM + Hypo],۴) [HCM + Asc + Hypo], ۵) [۵-aza], ۶) [۵-aza + Asc], ۷) [۵-aza +Hypo], ۸) [۵-aza + Asc + Hypo]. Then, the expression of α-actinand connexin۴۳ was investigated by immunofluorescence. Theequivalent groups were compared; (۱ ~ ۵), (۲ ~ ۶), (۳ ~ ۷) and(۴ ~ ۸).Results: The cells were positive for mesenchymal markers(CD۴۴, CD۴۵, CD۷۳, CD۹۰ and CD۱۰۵) and negative forCD۳۴. They demonstrated adipogenic and osteogenic differentiation.In [HCM + Asc], α-actin had higher level than [۵-Aza+ Asc]. The comparison of [Hypo + Aza] and [Hypo + HCM]showed that cardiac markers had higher expression level ingroup containing ۵-Aza, while the comparison of [۵-aza + Asc+ Hypo] and [HCM + Asc + Hypo] showed that connexin۴۳and α-actin had the most expression level in group containingHCM.Conclusion: As, connexin۴۳ and α-actin had the most expressionlevel in group ۴ and ۶. It means that the cocktail of “Asc+ Hypo” in combination with ۵-Aza or HCM can be the bestcombination for cardiomyocyte differentiation. We can concludethat when hypoxia is combined with ascorbic acid, it canaccomplish better cardiomyocyte differentiation than absenceof ascorbic acid.

کلیدواژه ها:

Cardiomyocyte Differentiation ، Wharton's Jelly Mesenchymal Stem Cells ، Heart Conditioned Medium