In vitro transplantation of spermatogonial stem cells isolated from human frozen\u2013thawed testis tissue can induce spermatogenesis under 3-dimensional tissue culture conditions

Mahdi Mohaqiq,Zohreh Mazaheri,Mansoureh Movahedin,Naser Amirjannati

doi:10.1186/s40659-019-0223-x

Abstract

BackgroundSperm production is one of the most complex biological processes in the body. In vitro production of sperm is one of the most important goals of researches in the field of male infertility treatment, which is very important in male cancer patients treated with gonadotoxic methods and drugs. In this study, we examine the progression of spermatogenesis after transplantation of spermatogonial stem cells under conditions of testicular tissue culture.ResultsTesticular tissue samples from azoospermic patients were obtained and then these were freeze–thawed. Spermatogonial stem cells were isolated by two enzymatic digestion steps and the identification of these cells was confirmed by detecting the PLZF protein. These cells, after being labeled with DiI, were transplanted in azoospermia adult mice model. The host testes were placed on agarose gel as tissue culture system. After 8 weeks, histomorphometric, immunohistochemical and molecular studies were performed. The results of histomorphometric studies showed that the mean number of spermatogonial cells, spermatocytes and spermatids in the experimental group was significantly more than the control group (without transplantation) (P < 0.05) and most of the cells responded positively to the detection of DiI. Immunohistochemical studies in host testes fragments in the experimental group express the PLZF, SCP3 and ACRBP proteins in spermatogonial cells, spermatocyte and spermatozoa, respectively, which confirmed the human nature of these cells. Also, in molecular studies of PLZF, Tekt1 and TP1, the results indicated that the genes were positive in the test group, while not in the control group.ConclusionThese results suggest that the slow freezing of SSCs can support the induction of spermatogenesis to produce haploid cells under the 3-dimensional testicular tissue culture.

Highlights

Sperm production is one of the most complex biological processes in the body
Sato et al [13] put the immature mouse testis tissue that contains only gonocytes and spermatogonial cells precursor under tissue culture conditions on agarose gel and, that leads to full progression of spermatogenesis
The identification of the isolated spermatogonial stem cells (SSCs) was confirmed by tracing the PLZF protein in the colonies derived from the culture cell suspensions obtained (Fig. 1)

Summary

Introduction

Sperm production is one of the most complex biological processes in the body. In vitro production of sperm is one of the most important goals of researches in the field of male infertility treatment, which is very important in male cancer patients treated with gonadotoxic methods and drugs. Sato et al [13] put the immature mouse testis tissue that contains only gonocytes and spermatogonial cells precursor under tissue culture conditions on agarose gel and, that leads to full progression of spermatogenesis. They obtained yielded adult mature sperm that can be fertilized by microinjection. They placed immature mouse testicular tissue after freezing and thawing under tissue culture conditions and reported complete progress in spermatogenesis. Until now, the progress of the spermatogenesis process has not been studied by human SSCs in mouse testis niche

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Discussion

Conclusion