Effect of Oxygen on Cardiac Differentiation in Mouse iPS Cells: Role of Hypoxia Inducible Factor-1 and Wnt/Beta-Catenin Signaling

Abstract

BackgroundDisturbances in oxygen levels have been found to impair cardiac organogenesis. It is known that stem cells and differentiating cells may respond variably to hypoxic conditions, whereby hypoxia may enhance stem cell pluripotency, while differentiation of multiple cell types can be restricted or enhanced under hypoxia. Here we examined whether HIF-1alpha modulated Wnt signaling affected differentiation of iPS cells into beating cardiomyocytes.ObjectiveWe investigated whether transient and sustained hypoxia affects differentiation of cardiomyocytes derived from murine induced pluripotent stem (iPS) cells, assessed the involvement of HIF-1alpha (hypoxia-inducible factor-1alpha) and the canonical Wnt pathway in this process.MethodsEmbryoid bodies (EBs) derived from iPS cells were differentiated into cardiomyocytes and were exposed either to 24 h normoxia or transient hypoxia followed by a further 13 days of normoxic culture.ResultsAt 14 days of differentiation, 59±2% of normoxic EBs were beating, whilst transient hypoxia abolished beating at 14 days and EBs appeared immature. Hypoxia induced a significant increase in Brachyury and islet-1 mRNA expression, together with reduced troponin C expression. Collectively, these data suggest that transient and sustained hypoxia inhibits maturation of differentiating cardiomyocytes. Compared to normoxia, hypoxia increased HIF-1alpha, Wnt target and ligand genes in EBs, as well as accumulation of HIF-1alpha and beta-catenin in nuclear protein extracts, suggesting involvement of the Wnt/beta-catenin pathway.ConclusionHypoxia impairs cardiomyocyte differentiation and activates Wnt signaling in undifferentiated iPS cells. Taken together the study suggests that oxygenation levels play a critical role in cardiomyocyte differentiation and suggest that hypoxia may play a role in early cardiogenesis.

Systems analysis reveals down-regulation of a network of pro-survival miRNAs drives the apoptotic response in dilated cardiomyopathy

“…For example, we demonstrated here that, in the presence of an antagomiR blocking mir-499, expression of APC was significantly increased (more than 2-fold), presumably causing a depletion in β-catenin 50 . Alterations in β-catenin-dependent Wnt signaling have been demonstrated under certain cardiac conditions, such as hypoxic conditions during differentiation of mouse pluripotent stem cells to beating cardiomyocytes, hypoxia activated Wnt signaling and impaired cardiomyocyte differentiation 75 . Furthermore, Wnt has been shown to enhance apoptosis in cardiomyoctes 76 suggesting a plausible mechanism for therapeutic intervention.…”

Selection of human induced pluripotent stem cells lines optimization of cardiomyocytes differentiation in an integrated suspension microcarrier bioreactor

“…The oxygen levels were maintained at 30% DO saturation to support the hiPSC growth during the expansion phase and early differentiation phase as shown in a previous publication [47]. Reports showed that hypoxia activated the expression of hypoxia inducible factor (HIF-1), which may indirectly enhance cardiomyocyte differentiation by the activation of a number of growth factors (such as VEGF) that have a synergistic effect on mesodermal cardiogenesis [18,43,49,51]. Further work is required, however, to optimize DO level during the different phases of the bioprocess system in order to improve the overall efficiency of CM generation.…”

Higher cardiogenic potential of iPSCs derived from cardiac versus skin stromal cells

“…On the other hand, the HIF-1 signaling pathway, known to reduce the number and the maturation of differentiating cardiomyocytes, is specifically targeted by miRs during the reprogramming process of SStCs. Of note, HIF-1 signaling appears to impair cardiogenic differentiation by reducing the number and the maturation of differentiating cardiomyocytes, which might provide a possible explanation of the lower cardiac differentiation capabilities observed in S-iPSCs (47). Finally, among the commonly targeted pathways, the PI3K/Akt pathway can be involved in the molecular mechanisms regulating cardiomyogenesis specifically by suppressing the GSK-3β activity and maintaining the Wnt/beta-catenin activity (48).…”