Evaluation of conditioned medium from Sertoli cells as a potential inducer of in vitro germ cell differentiation of bovine fetal mesenchymal stem cells (MSCs).

Abstract

Due to their self-renewal and differentiation potentials, mesenchymal stem cells (MSCs) may be induced into germ cells (GC) differentiation under in vitro conditions. In veterinary medicine, this technology could provide an alternative method to artificial insemination, as well as potentially useful for the conservation of endangered species. Previous studies have reported the use of SCs and MSCs co-culture systems, as well as SCs conditioned medium (SCCM) to induce GC differentiation of human and murine embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). The objective of this study was to evaluate the effect of SCCM as an inducer of in vitro GC differentiation of MSCs derived from fetal bovine adipose tissue (AT-MSCs). SCCM was collected from bovine SC cultures generated from adult bull testis. The effect of SCCM on MSCs was analyzed using quantitative PCR (Q-PCR) and flow cytometry. CD73 mRNA levels were decreased (P<0.05) in AT-MSC/SCCM at day 14 of culture compared to control. CD90 and CD105 gene expression were detected during the 21 days of culture; however, relative expression levels were not different (P>0.05) between treated and controls cells. DAZL gene expression was detected on day 21 of culture, as well as a proportion of AT-MSC positive for DAZL at day 21 of culture. OCT4, PIWIL2 and DAZL gene expressions were detected from day 0, 7 and 21 of culture, respectively, as well as a proportion of cells positive for each marker were detected at day 21 of culture. However, similar gene and protein expression levels (P>0,05) were detected between AT-MSCs/SCCM and control cultures. DMC1 gene expression levels were detected from day 7 of culture, and expression levels were not different (P>0,05) between treatment and control cells. Expression patterns of MSC, pluripotent, GC and meiotic markers indicate that SCCM did not induce GC differentiation of AT-MSCs.