Establishment and characterization of human theca stem cells and their differentiation into theca progenitor cells

: In this study, we have characterized the human theca stem cells (hTSCs) and their differentiation into human theca progenitor cells (hTPCs). hTSCs were isolated from the theca layer of small antral follicles (3-5 mm in size). Alkaline phosphatase activity, cell cycle status, and cell surface markers were evaluated in hTSCs. The differentiation potential of these cells was investigated via differentiation of hTSCs into adipocyte-, osteocyte-, and chondrocyte-like cells. The cells also differentiated into hTPCs. The hTSCs were morphologically similar to human fibroblast cells (hFCs). Some of the cells were positive for alkaline phosphatase activity. The expression of OCT4 in hTSCs was significantly higher than that of human bone marrow mesenchymal stem cells (hBMSCs) and hFCs. To determine the type of OCT4 (isoform A or B), RT PCR was performed. The data showed that OCT-4A was expressed in hBMSCs and hTSCs but immunofluorescence analyses using the OCT-4A-specific and OCT4 antibodies did not show OCT-4A protein. In addition, cell cycle status showed that the number of hTSCs in the S phase was significantly higher than that of hFCs. CD29, CD44, CD73, CD90, and CD105 were present in hTSCs. Osteogenic, adipogenic, and chondrogenic differentiation was confirmed by cytochemical staining and lineage-specific transcripts. Our results showed that specific Dulbecco modified Eagle medium F12 culture medium results in the presence of hTPC markers. hTPCs displayed lipid droplets, appropriate gene expression, and secreted dehydroepiandrosterone and estradiol. hTSCs have the ability to differentiate into mesenchymal lineages and hTPCs. This study may provide a novel in vitro model for further investigation of theca cell maturation and differentiation.