Potential of human testicular endosialin+ cells to differentiate towards LCs lineages. (A) Schematic of the experimental procedure used to induce LCs lineage differentiation. (B) After 14 days of differentiation in DIM, endosialin+ cells expressed the LCs lineage-specific markers LHCGR, SF-1, STAR and HSD3β, as shown by immunofluorescence (n = 3). The nuclei were counterstained with DAPI. NC stained in the absence of the primary antibody. The scale bar denotes 50 μm. (C) DIFF were examined by quantitative RT–PCR analysis of the expression of the indicated markers. The expression levels of each gene were compared with those in UNDIFF and human testis tissue. The data are expressed as the mean ± SEM (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001, ns = no significance. (D) Testosterone production was analysed by ELISA during LCs lineage induction in DIM. (E) Testosterone production of DIFF stimulated by LH increased in a concentration-dependent manner, as analysed by ELISA. The data are expressed as the mean ± SEM (n = 3). DIFF, differentiated endosialin+ cells; DIM, differentiation-inducing medium; HSD3β, hydroxysteroid dehydrogenase 3β; LHCGR, LH receptor; SF-1, steroidogenic factor 1; STAR, steroidogenic acute regulatory protein; Testis, human testis tissues; UNDIFF, undifferentiated endosialin+ cells.