Spermatogenic differentiation of spermatogonial stem cells on three-dimensional silk nanofiber scaffold

Abstract Background Nano-fibrous scaffolds provide a three-dimensional matrix that guides sufficient orientation of seeded cells similar to natural niche. In this research, we designed silk scaffold improve the differention mouse spermatogonial stem spermatogenic cell lines. Spermatogonial were collected from neonatal (2–6 days) testes ( n =60) using two steps mechanical and enzymatic method. Cells on cultured in Dulbecco’s modified Eagle’s medium, supplemented with 15 % fetal bovine serum 1000 units/ml leukemia inhibitory factor, incubated at 32°C humidified atmosphere 5% CO 2 air. SEM technique was done for confirmation seeding cells. study major groups (i.e., 2D 3D culture groups) 30 mice each. Isolated testicular each group absence or presence scaffold. For induction differentiation, exposed 1 μM 50 ng/ml BMP-4. The specific genes, e.g.; VASA, DAZL, PLZF, Piwil2, assessed via real-time PCR immunocytochemistry techniques. P values less than 0.05 assumed significant. All experiments performed least three times. Results analysis confirmed homogeneity fabricated average diameter 450 nm nanofibers fibers. Silk induces attachment SSCs comparison monolayer group. Spermatogonia colonies observed gradually after week culture. Electrospun supports differentiation Dates showed expression meiotic markers, Piwil2 as related had significant upregulation cell-seeded compared control < 0.05). Immunocytochemistry founding approved markers; DAZL PLZF higher experiment Conclusion It is concluded vitro.