Retinoic acid (RA) is known to induce osteogenic differentiation in many cell types. Mesenchymal stem cells are being used as promising tool for regenerative therapy. The objective of this study was to investigate the potential of retinoic acid as an inducer of osteogenic differentiation in human umbilical cord derived mesenchymal stem cells (hUMSCs) and its subsequent use in bone tissue engineering for the repair and regeneration of craniofacial bony defects with special emphasis on cleft-palate.
The hUMSCs obtained from Sciencell (Carlsbad, CA) were cultured in complete medium containing low glucose Dulbecco’s modified eagle medium with 10% of Fetal Bovine serum and 1% antibiotic and antimycotic solution at 37°C in 5% CO2. Cells with 70-80% confluency, were treated with retinoic acid (0.5, 1 and 2µM) in the presence of complete medium along with ascorbic acid and β glycerophsophate. Cells grown in similar culture conditions in absence of retinoic acid were considered as control. Cell proliferation was measured at 1 day, 2 day and 3 day intervals. On day 7, the expression of osteogenic marker genes was measured by RT-PCR. The activity of alkaline phosphatase (ALP) enzyme was determined using colorimetric assay. Matrix mineralization was assessed on day 21, by alizarin red staining.
Cell proliferation was significantly decreased in dose dependent manner in the cells exposed to retinoic acid when compared to control. The ALP gene expression was significantly down regulated by RA in a dose dependent manner. The ALP activity measured on days 7 and 14 also showed a significant decrease in the cells treated with retinoic acid. hUMSCs did not show matrix mineralization.
The results of this study suggested that under the above experimental conditions RA may not support osteogenic differentiation of hUMSCs. (Support NSUHPD).
Keywords: Bone, Bone repair, Cell culture, Cleft lip-palate and Human
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