Repair of injured soft and hard tissues in horses can benefit greatly from the use of regenerative therapies with mesenchymal stem cells (MSC). Vitamin-C and platelet-rich-plasma had been used for in vitro differentiation of MSC. This study was aimed to evaluate the effect of vitamin-C, platelet-rich-plasma and their combination on the in vitro differentiation of adipose horse MSC. We isolated MSC from horse fat and differentiated them in vitro into osteogenic and chondrogenic lineages, as demonstrated by specific staining and RT-qPCR of selected genes. Combining vitamin-C and plasma-rich-platelet positively affected the ability of MSC to differentiate in vitro into mesodermal lineages during 14 days of culture; this effect was not as marked when differentiation was attempted for 21 days. This provides valuable information on the effect of combined use of these molecules in regenerative therapies and their potential application along stem cells for lesions of musculoskeletal tissue in sport horses.