MicroRNA-143 is a critical regulator of cell cycle activity in stem cells with co-overexpression of Akt and angiopoietin-1 via transcriptional regulation of Erk5/cyclin D1 signaling

We report that simultaneous expression of Akt and angiopoietin-1 (Ang-1) transgenes supported mitogenesis in stem cells with a critical role for microRNA-143 (miR-143) downstream of FoxO1 transcription factor. Mesenchymal stem cells (MSC) from young male rats were transduced with Ad-vectors encoding for Akt ( Akt MSC) and Ang-1 ( Ang-1 MSC) transgenes for their individual or simultaneous overexpression ( AA MSC; > 5-fold gene level and > 4-fold Akt and Ang-1 protein expression in AA MSC vs. Ad-Empty transduced MSC; Emp MSC). AA MSC had higher phosphorylation of FoxO1, which activated Erk5, a distinct mitogen-induced MAPK that drove transcriptional activation of cyclin D1 and Cdk4. Flow cytometry showed > 10% higher S-phase cell population that was confirmed by BrdU assay (15%) and immunohistology for Ki67 (11%) in AA MSC using Emp MSC as controls. miR array supported by real-time PCR showed induction of miR-143 in AA MSC (4.73-fold vs.. Emp MSC). Luciferase assay indicated a dependent relationship between miR-143 and Erk5 in AA MSC. FoxO1-specific siRNA upregulated miR-143, whereas inhibition of miR-143 did not change FoxO1 activation. However, miR-143 inhibition repressed phosphorylation of Erk5 and abrogated cyclin D1 with concomitant reduction in cells entering cell cycle. During in vivo studies, male GFP+ AA MSC transplanted into wild-type female infarcted rat hearts showed significantly higher number of Ki67 expressing cells (p < 0.05 vs. Emp MSC) 7 days after engraftment (n = 4 animals/group). In conclusion, co-overexpression of Akt and Ang-1 in MSC activated cell cycle progression by upregulation of miR-143 and stimulation of FoxO1 and Erk5 signaling.