Regeneratie van de spermatogenese na transplantatie van testiculair weefsel, testiculaire cellen en beenmerg naar de testes van steriele muizen

Student thesis: Master's Thesis

Abstract

Infertility is a common problem in our society. Gonadotoxic treatments like chemo- and radiotherapy are an important cause of male infertility. Because this therapy is toxic for fast dividing cells and thus also for spermatogonia, sterility is known as a long-term side effect of this therapy. Adult men can cryopreserve their sperm but little boys do not have this opportunity because spermatogenesis has not started yet. Their only hope to restore fertility are the spermatogonial stem cells.
Two techniques, in which these stem cells are transplanted to the testis, were compared with each other. In one technique, the spermatogonial stem cell transplantation (SSCT), the stem cells were injected as a cell suspension into the tubules of an infertile testis. The second method is tissue grafting, where a prepubertal tissue fragment is transplanted to the testis. The aim of both techniques is to restore spermatogenesis from the transplanted stem cells. Both techniques are evaluated in a GFP mouse model. Donor cells from a GFP+ mouse are transplanted in sterile GFP- mouse.
The cell suspension was injected in the tubules through the efferent duct or directly through the tunica albuginea. Green fluorescent colonies were visible four months after transplantation with both injection procedures. The largest amount of donor derived spermatogenesis was present after tissue transplantation, followed by ductal injection. When the cell suspension was injected through the tunica very little and small colonies were observed. The presence of donor derived spermatogenesis was confirmed by anti-GFP staining.
In a second phase the impact of cryopreservation on testicular tissue was evaluated by comparing tissue transplantation of fresh tissue with frozen-thawed pieces. Cryopreservation didn't cause a significant difference in the amount of donorspermatogenesis present after transplantation. All stages of spermatogenesis were found in fresh and frozen-thawed pieces. Donor origin of spermatogenesis was confirmed by anti-GFP staining. The morphology of the tubules and spermatogenesis was preserved during transplantation.
In a third phase the potential of bone marrow stem cells to colonize the seminiferous tubules and give rise to spermatogenesis was evaluated. The mononuclear cells, containing the stem cells, are injected through the ductus or directly through the tunica. A comparison was made with SSCT. Bone marrow transplantation did not give rise to fluorescent colonies. A few GFP+ cells were observed after anti-GFP staining in the interstitium and some tubules.
Stem cells have the potential to colonize the seminiferous tubules as well after SSCT (ductal and blind injection) as after tissue transplantation. The largest amount of donor spermatogenesis occurred after tissue transplantation. Ductal injection of the cell suspension gave significantly more colonies than blind injection. After bone marrow transplantation only a few GFP+ cells were observed.
Date of Award27 Jun 2007
Original languageDutch
SupervisorHerman Tournaye (Promotor) & Ellen Goossens (Co-promotor)

Keywords

  • transplantatie

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