Articular cartilage (AC) is a thin tissue that lines moving joints. Once injured, AC has very limited ability to regenerate due to lack of blood vessels and sensory neurons, leading to osteoarthritis (OA), the most common joint disease in the US. Available treatments, including knee replacement and microfracture surgeries, are insufficient for restoring native function. Current research is focusing on the potential of adipose derived stem cells (ASCs) to undergo chondrogenic differentiation for OA therapy. Multiple exogenous factors, including vitamins and proteins, can enhance ASC chondrogenesis. ASCs exhibit higher chondrogenic potential when supplemented with TGF-b3 (T), dexamethasone (D) and ascorbic acid (A) compared to a negative control. In this study, individual effects of T, D, and A, binary combinations, DA, DT and TA, and a combination of the three factors, DTA, on ASC chondrogenesis has been studied in micromass cultures as a function of time. An additional experimental set to test the temporal effect of growth factor supplementation was conducted. In this set, cultures were supplemented with DT was for the first three days, DTA from the third day till the seventh, and D till day 25. We are testing the hypothesis that this temporal addition will enhance chondrogenesis better than individual and other combinations. ASC differentiation and chondrogenesis are being evaluated by comparing mRNA gene expression of chondrogenic, hypertrophic, angiogenic, fibrogenic and osteogenic markers, colorimetric assays of collagen (Col) and Glycosaminoglycan (GAG), and upregulation of chondrocyte surface markers as characterized by flow cytometry. Expression of the chondrogenic indices, i.e. Col II, Col XI, SOX9, ACAN and Lubricin, are combined with the normalized colorimetric analysis results to calculate a chondrogenic index to identify the optimal growth factor supplementation regime. Our current findings indicate that D containing media cause an increase in total GAG and Col contents compared to other treatments.

Enhancing chondrogenesis of adipose derived stem cells through temporal supplementation of dexamethasone, TGF-b3, ascorbic acid and their combinations

2019 Spring National Meeting

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