Regeneration of Leydig cells in ectopically autografted adult mouse testes

Abstract

Ectopic autografting of testis tissue is a promising approach for studying testicular development, male germline preservation and restoration of male fertility. In this study, we examined the fate of various testicular cells in adult mouse testes following ectopic autografting at 1, 2, 4 and 8 weeks post grafting. Histological examination showed no evidence of re-establishment of spermatogenesis in autografts, and progressive degeneration of seminiferous tubules was detected. Expression of germ cell-specific proteins such as POU5F1, DAZL, TNP1, TNP2, PRM1 and PRM2 revealed that, although proliferating and differentiating spermatogenic germ cells such as spermatogonia, spermatocytes and spermatids could survive in autografts until 4 weeks, only terminally differentiated germ cells such as sperm persisted in autografts until 8 weeks. The presence of Sertoli and peritubular myoid cells, as indicated by expression of WT1 and ACTA2 proteins, respectively, was evident in the autografts until 8 weeks. Interestingly, seminal vesicle weight and serum testosterone level were restored in autografted mice by 8 weeks post grafting. The expression of Leydig cell-specific proteins such as CYP11A1, HSD3B2 and LHCGR showed revival of Leydig cell (LC) populations in autografts over time since grafting. Elevated expression of PDGFRA, LIF, DHH and NEFH in autografts indicated de novo regeneration of LC populations. Autografted adult testis can be used as a model for investigating Leydig cell regeneration, steroidogenesis and regulation of the intrinsic factors involved in Leydig cell development. The success of this rodent model can have therapeutic applications for adult human males undergoing sterilizing cancer therapy.

Dysregulation of angiogenesis-specific signalling in adult testis results in xenograft degeneration

“…The inability of xenografts from adult testis to produce sufficient testosterone could be due to lack of mature Leydig cells. These results contradict previous findings where restoration of seminal vesicle weight in mice 26, 27 and, serum testosterone level in rats 28 following adult testis autografting has been reported. This inconsistency could be due to failure or delay in the establishment of angiogenesis in xenografted adult testis leading to loss of Leydig cells 29 .…”

“…It is likely that spermatozoa being terminally differentiated cells are rather resistant to degeneration and are able to survive in degenerated tubules. We had observed similar findings in autografted adult mice testis 27 and another report also confirmed similar finding 16 . Testis from immature donors could establish spermatogenesis following ectopic xenografting and meiosis was induced as indicated by the presence of pachytene-stage spermatocytes as the most advanced germ cells (Fig.…”

“…Interestingly, PCNA-stained cells were observed in the peritubular region and interstitial space in adult testis grafts. These PCNA-stained cells possibly represent stem or foetal Leydig cells as reported previously 27 .

Figure 2Proliferating cell nuclear antigen (PCNA) immunostaining ( A ) In donor tissue from 10-wk-old rat (T0). Black arrows indicate PCNA-positive spermatogonia, red arrows indicate PCNA-positive Sertoli cells and, arrow heads indicate PCNA-positive spermatocytes.

…”

Wistar rats immature testicular tissue vitrification and heterotopic grafting

“…Heterotopic transplantation showed signs of testicular structure disintegration and extensive seminiferous tubules degeneration, except for one sample where it was possible to identify intact seminiferous tubules. Obtained data corroborates with Makala et al . (2015) results, showing that when autologously transplanting testicular fragments (5mm) on the back of adult mice, there was an extensive seminiferous tubule degeneration 4 weeks after grafting and no intact seminiferous tubule present after 8 weeks.…”

Characterization of the testicular regeneration potential in premature cockerels

“…The testis is not only responsible for spermatogenesis, but also for sex hormone production in animals. Several attempts using zebrafish, mice, and pigs for the characterization of testis regeneration revealed limited spermatogenic or steroidogenic functions, except in newborn animals [ 6 , 7 , 8 ]. Similarly, studies by Song and Silversides (2007) [ 9 ] reported that hetero-transplanted testicular slices from a newly hatched chick into a recipient chick’s abdomen were able to grow and produce functional sperm that generated donor-derived progeny, which suggested that chicken testes have a strong regenerative potential.…”