Pirnia A, Parivar K, Hemadi M, Yaghmaei P, Gholami M
Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.
Fertility and Infertility Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.
Andrologia. 2017 Jun;49(5). doi: 10.1111/and.12650. Epub 2016 Jul 29.
This study investigated the effect of spermatogonial stem cell encapsulated in alginate hydrogel during cryopreservation, as cells were protected against damage during cryopreservation within the hydrogel. Spermatogonial stem cells were isolated from the testes of Balb/c mice pups (6 days old), purified in laminin-coated dishes and CD90.1 microbeads, encapsulated in alginate hydrogel and then cryopreserved. After thawing, cell viability and Spermatogonial stem cell (SSC) colony diameter were evaluated. After RNA was isolated and cDNA was synthesised, the expression of stemness genes was considered using RT real-time PCR. Finally, spermatogonial stem cells labelled with BrdU were transplanted to busulfan azoospermic mouse models. Lin28a and Sall4 genes were significantly upregulated after cryopreservation in alginate hydrogel. However, cell viability was significantly decreased. The diameter of colonies consisting of spermatogonial stem cells freeze-thawed in alginate microbeads showed no significant difference with fresh spermatogonial stem cells and the control group. The injection of freeze-thawed spermatogonial stem cells encapsulated in alginate hydrogel resulted in spermatogenesis recovery. Alginate mimics the extracellular matrices (ECM) for spermatogonial stem cells; therefore, it can support stemness potential during the cell cryopreservation process and restart spermatogenesis after transplantation.
本研究调查了封装在海藻酸盐水凝胶中的精原干细胞在冷冻保存期间的效果，因为细胞在水凝胶内的冷冻保存过程中受到保护而免受损伤。从6日龄的Balb/c小鼠幼崽的睾丸中分离出精原干细胞，在层粘连蛋白包被的培养皿中并用CD90.1微珠纯化，封装在海藻酸盐水凝胶中，然后进行冷冻保存。解冻后，评估细胞活力和精原干细胞（SSC）集落直径。分离RNA并合成cDNA后，使用RT实时PCR检测干性基因的表达。最后，将用BrdU标记的精原干细胞移植到白消安诱导的无精子症小鼠模型中。在海藻酸盐水凝胶中冷冻保存后，Lin28a和Sall4基因显著上调。然而，细胞活力显著下降。在海藻酸盐微珠中冻融的由精原干细胞组成的集落直径与新鲜精原干细胞及对照组相比无显著差异。注射封装在海藻酸盐水凝胶中的冻融精原干细胞可使精子发生恢复。海藻酸盐模拟精原干细胞的细胞外基质（ECM）；因此，它可以在细胞冷冻保存过程中支持干性潜能，并在移植后重启精子发生。
