Long-Term Culture of Mouse Male Germline Stem Cells Under Serum-or Feeder-Free Conditions1

TLDR

Spermatogonial stem cells are the sole germline stem cells that transmit genetic information, can be cultured long‑term with serum and feeder cells, yet the factors governing their self‑renewal remain poorly understood. The study aimed to develop serum‑ and feeder‑free culture systems for spermatogonial stem cells to facilitate investigation of their regulatory mechanisms. The authors tested culture conditions lacking serum or feeder cells, using serum‑free media on mouse embryonic fibroblasts and feeder‑free laminin‑coated plates. Spermatogonial stem cells expanded in serum‑free MEFs or feeder‑free laminin plates, but not when both serum and feeder were absent; the laminin‑cultured cells grew exponentially for at least six months and yielded normal, fertile progeny after transplantation.

Abstract

Spermatogonial stem cells are the only stem cells in the body that transmit genetic information to the next generation. These cells can be cultured for extended periods in the presence of serum and feeder cells. However, little is known about factors that regulate self-renewal division of spermatogonial stem cells. In this investigation we examined the possibility of establishing culture systems for spermatogonial stem cells that lack serum or a feeder cell layer. Spermatogonial stem cells could expand in serum-free conditions on mouse embryonic fibroblasts (MEFs), or were successfully cultivated without feeder cells on a laminin-coated plate. However, they could not expand when both serum and feeder cells were absent. Although the cells cultured on laminin differed phenotypically from those on feeder cells, they grew exponentially for at least 6 mo, and produced normal, fertile progeny following transplantation into infertile mouse testis. This culture system will provide a new opportunity for understanding the regulatory mechanism that governs spermatogonial stem cells.