title: 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.
abstract: 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.
authors.affiliation: Department of Pathology, University of Cincinnati, Cincinnati, OH, USA.
authors.forename: Vien Khach
authors.initials: VK
authors.lastname: Lai
authors.forename: Muhammad
authors.initials: M
authors.lastname: Ashraf
authors.forename: Shujia
authors.initials: S
authors.lastname: Jiang
authors.forename: Khawaja
authors.initials: K
authors.lastname: Haider
mesh_terms: D042683:Angiopoietin-1 / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002453:Cell Cycle / Q000235:genetics / Q000502:physiology*; D002478:Cells, Cultured; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D048054:Mitogen-Activated Protein Kinase 7 / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011916:Rats, Inbred F344; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D042683:Angiopoietin-1; D035683:MicroRNAs; D019938:Cyclin D1; D051057:Proto-Oncogene Proteins c-akt; D048054:Mitogen-Activated Protein Kinase 7