Related Experiment Video
Updated: Aug 15, 2026
Germ Cell Transplantation and Testis Tissue Xenografting in Mice
Published on: February 6, 2012
Dose-response of RAG2-/-/gammac-/- mice to busulfan in preparation for spermatogonial transplantation
A E Moisan1, R A Foster, K J Betteridge
1Department of Biomedical Sciences, University of Guelph, Guelph, Ontario N1G 2W1, Canada.
Abstract:
Practical applications of spermatogonial transplantation require good rates of colonization by the donor cells. Recipient testes are usually depleted of competing endogenous spermatogonia by administration of 32-44 mg busulfan kg(-1) body weight before transplantation. However, it is not clear that this is the optimum dose, especially for immunodeficient mice. In the present study, the response of adult RAG2(-/-)/gamma(c)(-/-) (RAG2) male mice to treatment with 10-50 mg busulfan kg(-1) body weight was determined in terms of mortality rates, testicular masses and histology, and colonization of seminiferous tubules by transplanted spermatogonia. Mortality increased from 0 to 50% at doses between 20 mg busulfan kg(-1) and 40 mg busulfan kg(-1), whereas the maximum effects on testicular mass and histology were observed at 20 mg busulfan kg(-1). Colonization of testes by genetically marked spermatogonia after treatment of mice with 20 mg busulfan kg(-1) was equivalent to rates previously reported in recipients treated with 32-44 mg busulfan kg(-1). Thus, 20 mg busulfan kg(-1) appears to be the optimum dose for preparing RAG2 mice for spermatogonial transplantation. However, because the steepness of the dose-response curves indicates that direct administration of busulfan is not ideal for this purpose, 15 mg busulfan kg(-1) was administered to pregnant females at various times between day 10.5 and day 16.5 of gestation to determine whether this would deplete the number of germ cells in male offspring. Although there were large variations in testicular mass and histology, no mortality was observed and administration of busulfan at day 10.5 or 12.5 after mating delayed initiation of spermatogenesis, indicating that prenatal administration of busulfan combined with neonatal transplantation might be an effective method for further increasing rates of colonization by donor spermatogonia.
Insights
Optimizing busulfan dosage for spermatogonial transplantation in RAG2 mice is crucial. A dose of 20 mg/kg is found to be optimal for adult mice, while prenatal exposure shows promise for future applications.
Area of Science:
- Reproductive Biology
- Developmental Biology
- Transplantation Immunology
Background:
- Spermatogonial transplantation is vital for practical applications in fertility research.
- Effective colonization by donor cells requires depletion of endogenous spermatogonia in recipient testes.
- Current busulfan doses (32-44 mg/kg) for depletion may not be optimal, especially in immunodeficient models.
Purpose of the Study:
- To determine the optimal busulfan dose for depleting endogenous spermatogonia in RAG2 male mice.
- To evaluate the effects of varying busulfan doses on mortality, testicular parameters, and donor cell colonization.
- To investigate prenatal busulfan administration as an alternative depletion strategy.
Main Methods:
- Adult RAG2 male mice were treated with busulfan at doses ranging from 10-50 mg/kg.
- Evaluated mortality rates, testicular mass, histology, and colonization by transplanted spermatogonia.
- Administered 15 mg/kg busulfan to pregnant females at different gestational days (10.5-16.5) to assess effects on male offspring.
Main Results:
- A busulfan dose of 20 mg/kg maximized testicular depletion and histology effects with minimal mortality.
- Spermatogonial colonization in testes treated with 20 mg/kg busulfan was comparable to higher doses.
- Prenatal busulfan exposure (days 10.5-12.5) in dams resulted in germ cell depletion in male offspring without mortality, delaying spermatogenesis initiation.
Conclusions:
- 20 mg/kg busulfan is the optimal dose for preparing adult RAG2 mice for spermatogonial transplantation.
- Direct administration of busulfan presents dose-response challenges; prenatal administration offers a potentially safer alternative.
- Prenatal busulfan exposure followed by neonatal transplantation may enhance donor spermatogonial colonization rates.
More Related Videos
11:25Busulfan as a Myelosuppressive Agent for Generating Stable High-level Bone Marrow Chimerism in Mice
Published on: April 1, 2015
06:08Assessment of Chimeric Antigen Receptor T Cell-Associated Toxicities Using an Acute Lymphoblastic Leukemia Patient-Derived Xenograft Mouse Model
Published on: February 10, 2023