Testicular Cells Derived Conditioned Medium Supports Germ Cell Differentiation of Human Embryonic Stem Cells

Abstract

Objective: There are ethical and technical challenges in studying human germ cell development. Therefore, the aim ofthe study is in vitro differentiation of human embryonic stem cells (hESCs), as pluripotent cells, to the germ cells whichis a valuable tool for studying molecular and cellular aspects of gametogenesis and understanding causes of infertility.Materials and Methods: In this experimental study, two different complete media [Dulbecco’s Modified Eagle Medium(DMEM)+20% fetal bovine serum (FBS) and embryoid bodies (EBs) medium; KOSR/HES without basic fibroblastgrowth factor (bFGF)] were used in the both of test groups using testicular cells derived conditioned medium (TCCM)and control groups spontaneously differentiated (SD). Thereby, EBs from hESCs (Yazd2; 46XY) were cultured indifferent conditions EB medium; EB medium and conditioned EB medium; EB medium, DMEM, and FBS withoutconditioning; EB medium, conditioned DMEM, and FBS medium. EBs were collected after 4, 7, and 14 days and theirgene expression profiles were assessed and compared to hESCs, as day 0, using IF and relative reverse transcriptionquantitative polymerase chain reaction (RT-qPCR).Results: An increase in the gametogenesis gene expression level in TCCM groups was showed in comparison withSD groups. Additionally, immunostaining of differentiated cells in all groups showed in vitro gametogenesis (IVG).Conclusion: Our findings showed that human TCCM could be used as a natural niche for in vitro male and femalegerm cell development. However, further studies are needed to define the factors and metabolites within the humanTCCM.

Keywords

• conditioned medium
• differentiation
• germ cells
• human embryonic stem cells