Elevation of protein kinase Cα stimulates osteogenic differentiation of mesenchymal stem cells through the TAT-mediated protein transduction system.
Mesenchymal stem cells (MSCs) can differentiate toward various lineages, including the osteogenic lineage, and thus hold great potential for clinic purposes. By using pharmacological inhibitors, protein kinase C (PKC) signaling has been shown to either negatively or positively regulate differentiation of bone, however, due to the low transfection efficiency in MSCs, the role of individual PKC isoforms is still not fully understood. In this study, we established a TAT peptide-mediated transduction system that efficiently delivered PKCα proteins into MSCs in a non-invasive fashion. The increased PKCα protein levels significantly promoted osteogenic differentiation in the murine mesenchymal C3H10T1/2 cells and in primary MSCs from both human and mouse, as demonstrated by the enhanced activity of the osteoblast marker, alkaline phosphatase, and the enhanced expression of the key transcription factor runx2. Mineralization is an important functional indication for bone differentiation. Our results further showed that PKCα promoted expression of the important osteocalcin gene and the accumulation of calcium minerals. Taken together, this study provides direct evidence showing that PKCα positively regulates osteogenic differentiation and demonstrates that the TAT peptide-mediated method enables functional study of specific PKC isoforms in MSCs without using viral infection. This may promote the application of PKCs in therapeutic treatment.
a Institute of Biopharmaceutical Sciences, National Yang-Ming University, No. 155, Sec. 2, Linong St., Taipei 112, Taiwan.
Wei-Kai
WK
Hua
Ya-Huei
YH
Shiau
Oscar K
OK
Lee
Wey-Jinq
WJ
Lin
D000469:Alkaline Phosphatase / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002113:Calcification, Physiologic; D002454:Cell Differentiation; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D015696:Gene Products, tat / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D010012:Osteogenesis / Q000235:genetics; D051571:Protein Kinase C-alpha / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D014161:Transduction, Genetic / Q000379:methods*
D015415:Biomarkers; D050659:Core Binding Factor Alpha 1 Subunit; D015696:Gene Products, tat; C493709:RUNX2 protein, human; D015675:Osteocalcin; D051571:Protein Kinase C-alpha; D000469:Alkaline Phosphatase