Comparison of the Effect of Laminin and Matrigel on the Colonization of Frozen-thawed Spermatogonial Stem Cells in Vitro

Background and Objective: Spermatogonial stem cells (SSCs) are the only adult stem cells that serve as the origin of spermatogenesis, possessing both proliferation and self-renewal capabilities, as well as the ability to transmit genetic information to the next generation. These cells play a significant role in reproductive medicine, the treatment of various types of infertility, and cancers. The proliferation of this cell line in vitro is a prerequisite for all related studies. The aim of this study was to investigate the in-vitro effects of the extracellular matrix of laminin and matrigel on the colonization of frozen-thawed sheep spermatogonia stem cells. Materials and Methods: Spermatogonial stem cells were isolated from the testes of lambs and purified using a differentiation removal method. After freezing and thawing, the cells were cultured for 10 days in three experimental groups: the control group (simple cell culture in basal medium: DMEM containing 1% antibiotics and 5% fetal bovine serum), treatment 1 (cultured on laminin), and treatment 2 (cultured on matrigel). The number and area of colonies were measured on days 4, 7, and 10 in each group. Specific markers for spermatogonial cells (PLZF, ITGA6, PGP9.5, VASA) were also assessed using immunocytochemistry. Results: The number of colonies in the laminin group (118.92±6.2) was significantly higher than that in the control (78.08±3.8) and matrigel groups (95.00±4.3) (P≤0.05). The colony area was significantly higher in the laminin (1.37±0.30 mm2) and matrigel groups (1.97±0.38 mm2) than in the control group (0.59±0.08 mm2) (P≤0.05). Conclusion: The extracellular matrix of laminin can provide a suitable microenvironment for cryopreserved Ovine SSCs in an in vitro culture system.