The Role of Endogenous Tumor Necrosis Factor Alpha (TNF-a) on the Osteogenic Potential of Mesenchymal Stem Cells Derived from Adipose Tissue

Abstract

Cell-based therapy has been used as a promising strategy for bone regeneration purposes, an approach of interest in the filed of maxillofacial and orthopedics surgeries. Among the cell types, the mesenchymal stem cells derived from adipose tissue (AT-MSC) emerge as a good alternative once they are easily harvested with lower morbidity, compared to the cells obtained from bone marrow (BM-MSC). However, the AT-MSC have been assigned to a lower osteogenic potential. Previous results showed the impairment on osteoblastic differentiation due to the presence of the tumor necrosis factor alpha (TNF-±) released by adipocytes. Thus, we assume that TNF-± produced and released by AT-MSC have an inhibitory effect on osteoblast differentiation. In this context, we propose to evaluate the role of endogenous TNF-± in the osteogenic potential of AT-MSC. AT-MSC will be harvested from Wistar rats and temporal and gene expression of TNF-± and its receptors (TNFR1A and TNFR1B) will be evaluated. Then, the AT-MSC will be transfected with shRNA clones for TNF-± receptors alone or associated and evaluated for osteoblastic differentiation by: 1) alkaline phosphatase (ALP), RUNX2, osteopontin (OPN) and osteocalcin (OC) gene expression; 2) ALP activity and 3) production of mineralized extracellular matrix. As control of osteoblastic differentiation, BM-MSC obtained from the same animals will be used. To evaluate the effect of TNF-± silencing on bone repair, the selected shRNA will be transfected to CTM-TAs and will be injected into bone defects created in rat calvaria defects. The bone repair will be evaluated after 4 weeks using computerized microtomography and histological analysis. The results of this study may contribute to the understanding of the intracellular mechanisms involved in the differentiation and activity of osteoblastic cells derived from AT-MSC and contribute to the application and development of new treatment strategies involving cell-based therapy on bone regeneration. (AU)