P172 - Immortalization approach dependent variation in proliferation and differentiation of infrapatellar fat pad derived stem cells
Cellular immortalization using human telomerase (TERT) or SV40 large T (LT) antigen facilitates stem cell-based research. In this study, we hypothesized that TERT and SV40 transduced infrapatellar fat pad derived stem cells (IPFSCs) would perform differently in proliferation and differentiation capacity which could also be modulated by decellularized extracellular matrix (dECM).
Methods and Materials
TERT and SV40LT lentiviruses were used to immortalize IPFSCs. We compared their differences in cell proliferation, cell surface markers, stemness gene expression, and chondrogenic and adipogenic differentiation potential. High passage (P12) IPFSCs were expanded on either tissue culture plastic (TCP) alone or dECMs deposited by un-transduced cells, GFP control vector-transduced cells, TERT or SV40LT lentivirus transduced cells from either a low (P5) or high (P12) passage. Proliferation and bi-differentiation capacities were evaluated. Premature pellets were also evaluated in nude rats by implanting into either joints or subcutaneous pockets for 3 weeks.
Compared with TERT immortalized cells, SV40LT immortalized cells exhibited a superior proliferative capacity. TERT immortalized IPFSCs exhibited a significant increase in chondrogenesis but a decrease in adipogenesis, which is contrary to the response of SV40LT immortalized cells. IPFSCs plated on dECM deposited by TERT-immortalized cells grew faster and showed higher chondrogenic differentiation potential. Interestingly, dECM deposited by SV40LT immortalized cells could reverse P12 IPFSCs’ differentiation preference, by promoting chondrogenic differentiation and decreasing adipogenic differentiation. Compared with subcutaneous implantation, growing pellets in joints decreased vascularization.
Cellular immortalization approach affects stem cell proliferation capacity and differentiation preference, which can also be modulated by dECM rejuvenation. Despite the variation of IPFSCs’ response to TERT and SV40LT immortalization, both approaches can be fine-tuned to regulate adult stem cells’ fate.