Enhanced proliferation and differentiation of Oct4- and Sox2-overexpressing human adipose tissue mesenchymal stem cells

Abstract

Mesenchymal stem cells (MSCs) are attractive candidates for clinical repair or regeneration of damaged tissues. Oct4 and Sox2, which are essential transcription factors for pluripotency and self-renewal, are naturally expressed in MSCs at low levels in early passages, and their levels gradually decrease as the passage number increases. Therefore, to improve MSC proliferation and stemness, we introduced human Oct4 and Sox2 for conferring higher expansion and differentiation capabilities. The Oct4-IRES-Sox2 vector was transfected into human adipose tissue MSCs (ATMSCs) by liposomal transfection and used directly. Oct4 and Sox2 were successfully transfected into ATMSCs, and we confirmed maintenance of MSC surface markers without alterations in both red fluorescent protein (RFP) (control) and Oct4/Sox2-ATMSCs. Enhanced proliferative activity of Oct4/Sox2-ATMSCs was shown by WST-1 assay, and this result was further confirmed by cell counting using trypan blue exclusion for a long period. In addition, FACs cell cycle analysis showed that there was a reduction in the fraction of Oct4/Sox2-ATMSCs in G1 with a concomitant increase in the fraction of cells in S, compared with RFP-ATMSCs. Increased levels of cyclin D1 were also seen in Oct4/Sox2-ATMSCs, indicating acceleration in the transition of cells from G1 to S phase. Furthermore, Oct4/Sox2-overexpressing ATMSCs showed higher differentiation abilities for adipocytes or osteoblasts than controls. The markers of adipogenic or osteogenic differentiation were also upregulated by Oct4/Sox2 overexpression. The improvement in cell proliferation and differentiation using Oct4/Sox2 expression in ATMSCs may be a useful method for expanding the population and increasing the stemness of ATMSCs. Researchers in Korea have shown that forced expression of two genes stimulates growth in adult stem cells, increasing their potential therapeutic applications. Mesenchymal stem cells (MSCs), a type of adult stem cell, lack the developmental flexibility of embryonic stem cells but can potentially repair tissues such as bone or cartilage and are easily harvested from fat tissue. Hee-Woo Lee and Hwa-Young Youn of Seoul National University have shown that forcing MSCs to express the Sox2 and Oct4 genes increases their viability and capacity to multiply. These two genes help lock MSCs in their developmentally immature state, while maintaining their capacity to differentiate into fat or bone cells. This treatment could greatly boost the clinical usefulness of these cells, which normally lose their stem cell qualities over time when cultured.