Effect of transforming growth factor beta 1 on chondrogenic differentiation of cultured equine mesenchymal stem cells

Objective-To determine the morphologic and phenotypic effects of transformi ng growth factor beta 1 (TGF-beta 1) on cultured equine mesenchymal stem ce lls (MSC) and articular chondrocytes. Sample Population-Bone marrow aspirates and articular cartilage samples fro m a 2-year-old and two 8-month-old horses. Procedure-After initial isolation and culture, MSC and chondrocytes were cu ltured in Ham's F-12 medium supplemented with TGF-beta 1 at a concentration of 0, 1, 5, or 10 ng/ml. Medium was exchanged on day 2, and cells were har vested on day 4. Medium was assayed for proteoglycan (PG) content. Total RN A was isolated from cell cultures, and expression of aggrecan, decrin, coll agen type-I, and collagen type-II mRNA was assessed by means of Northern bl ot analyses. Cell cultures were stained with H&E or toluidine blue and exam ined histologically. Additional cultures were examined after immunohistoche mical staining for type-I and -II collagen. Results-MSC cultures exposed to TGF-beta 1 had an increased cellular densit y with cell layering and nodule formation that was most pronounced in cultu res treated with 5 ng of TGF-beta 1/ml. Expression of collagen type-ii mRNA in MSC cultures exposed to 5 ng of TGF-beta 1/ml was 1.7 times expression in control cultures, and expression of collagen type-I mRNA was 2.8 times e xpression in control cultures. Treatment of MSC with TGF-beta 1 led to dose -related increases in area and intensity of type-II collagen immunoreaction . Conclusion-Results suggest that TGF-beta 1 enhances chondrogenic differenti ation of bone marrow-derived MSC in a dose-dependent manner.