Background: Some male survivors of childhood cancer are in distress from azoospermia.
Spermatogonial stem cells (SSC) isolation and purification are really important. Testicular organoids and 3D scaffolds are possible applications in treatment of male infertility. 3D nanofibers scaffolds play important role in cell culture, due to these scaffolds provide a microenvironment similar to extracellular matrix for proliferation and self-renewal of cells. A micro fabricated chip made of polydimethylsiloxane (PDMS) and having a set of micro-channels etched or molded into it, can serve as a culture vessel for cells and can easily accommodate medium few or Laboratory of Proteomics.Objective: The present study aimed to evaluate efficiency of 3D microenvironment containing the chitosan-alginate -Graphene oxide nanocomposites and microfluidic culture system for improve cell culture and proliferation of spermatogonial cells.Materials and Methods: Spermatogonial cells were cultured and divided into 5 culture groups: 1/ Control (culture in basic media), 2/ SSC culture in CA/GO scaffold, 3/ SSC culture on
CA/GO scaffold in microfluidic system, 4/ cells culture on GO nanocomposites and 5/ the cells culture on alginate-chitosan. The identity of the cultured cells was confirmed by flow cytometry (ckit and GFRa1). The Scaffolds were investigated by SEM to observe surface topography and the morphology. Cytotoxicity of scaffold was assayed at 24h, 72h and one week after seeding using MTT assay. The stem cells related markers for SSCs (Id4 ,GFRa1 and PLZF) were detected on all experimental groups by qRT-PCR.Results: These results showed that SSCs can easily attach and proliferate on. The CA/GO scaffolds were biocompatible, as evidenced by the MTT assay.
Spermatogonial stem cells that were seeded onto the surface of the scaffold exhibited good proliferation. The quantity of proliferations marker significantly increased compared with control group. The qRT-PCR results confirmed that GO-based/Alginate-chitosan scaffold may provide an ideal environment for SSC proliferations.Conclusion: We conclude that SSCs culture in the group CA/GO/ MF have potential use for SSCs proliferation in vitro. This three-dimensional scaffold is applicable for culturing and encapsulation of spermatogonial stem cells.