Optimal dose of busulfan for depleting testicular germ cells of recipient mice before spermatogonial transplantation

Wang, Dezhi; Zhou, Xinhua; Yuan, Yuexing; Zheng, Xiaodong

doi:10.1038/aja.2009.67

Cited by 50 publications

(40 citation statements)

References 26 publications

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“…This is typically performed by using either chemical compounds like the alkylating agent busulfan or alternatively testis irradiation. However, protocols need to be species optimized, which requires further research [97,98] . SSC transplantation could also be used for gene delivering purposes in order to increase the frequency of certain genes in animal populations.…”

mentioning

confidence: 99%

Spermatogonial stem cells: Current biotechnological advances in reproduction and regenerative medicine

Aponte

2015

WJSC

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mentioning

confidence: 99%

Spermatogonial stem cells: Current biotechnological advances in reproduction and regenerative medicine

Aponte

2015

WJSC

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“…Meanwhile, a mortality rate of 30% was observed in the 40 mg/kg group. This was lower than the 50% mortality rate defined as the lethal dose rate for tree shrews (27). All tree shrews injected with 45 mg/kg busulfan succumbed, suggesting that the high busulfan dose exerted acute toxic effects.…”

Section: Discussionmentioning

confidence: 87%

Depletion of endogenous germ cells in tree shrews in preparation for spermatogonial transplantation

Liu

Guo

Yan

et al. 2017

Experimental and Therapeutic Medicine

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Abstract. To achieve successful spermatogonial transplantation, endogenous germ cells must be depleted in recipient animals to allow donor germ cells to colonize efficiently. Busulfan is commonly used for the depletion of endogenous germ cells in recipient males. However, the optimal dose of busulfan is species-specific, and the optimal dose in tree shrews is yet to be determined. The current study aimed to determine the optimal dose of busulfan for effective suppression of endogenous spermatogenesis in tree shrews. Different doses (15,20,25, 30, 35, 40 and 45 mg/kg) of busulfan were injected into tree shrews intraperitoneally. Survival rates of the different treatment groups were calculated at 2 weeks and body weights were measured at 4, 6, 8, 10 and 28 weeks post-busulfan treatment. The testes were also removed and weighed at 4, 6, 8, 10 and 28 weeks post-treatment, and the cross and longitude diameters of the testes and diameters of the seminiferous tubules were measured and histologically evaluated. It was observed that there were no significant differences in the survival rates between the 15-35 mg/kg treatment groups and the control group (P>0.05), while the survival rate of the 40 mg/kg treatment group significantly decreased relative to the control group (P<0.05) and the survival rate of the 45 mg/kg treatment group was 0% (P<0.05 vs. control). In addition, the weight and diameters of the testes, diameters of the seminiferous tubules and proportion of normal type tubules in the 40 mg/kg group significantly decreased over 4-10 weeks relative to the control group (P<0.05), though gradually recovered with time. At 28 weeks, the recovery was significant relative to 4 weeks (P<0.05). Similarly, histological analysis indicted that recovery of abnormal tubules was delayed in the 40 mg/kg group relative to the lower dose groups as the 40 mg/kg dose tree shrews had more tubules with no spermatogenesis compared with the lower dose group at the same time points. These data indicate that a busulfan dose of 40 mg/kg is optimal for the depletion of endogenous germ cells in tree shrews. This dose led to maximum suppression of endogenous spermatogenesis while maintaining an acceptable survival rate of >50% of the lethal dose of busulfan for tree shrews.

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“…busulfan), pesticides and ionizing radiation have been described to have this effect. Busulfan is of some interest as it is one of the most potent gonadotoxicants known and is routinely used to sterilize mice (41). Unfortunately, busulfan’s chemical structure did not allow for linkage to FSHβ (33-53), and it is too toxic when administered alone in terms of bone marrow and neurological toxicity to be considered for sterilizing larger captive and wild animals (42-44).…”

Section: Discussionmentioning

confidence: 99%

“…It must be noted, however, that this approach to screening compounds in vitro has a limitation in that the behavior of Sertoli cells in vitro may different from their behavior in vivo . Sertoli cells in culture can proliferate (41), and therefore may be more sensitive to the effects of melphalan compared to Sertoli cells in vivo , which are non-replicating. With these caveats, future work directed at developing novel compounds for male sterility could use this system to screen candidate molecules prior to in vivo testing.…”

Section: Discussionmentioning

confidence: 99%

Melphalan, alone or conjugated to an FSH-β peptide, kills murine testicular cells in vitro and transiently suppresses murine spermatogenesis in vivo

Amory¹,

Hong²,

Yu³

et al. 2014

Theriogenology

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New approaches to sterilizing male animals are needed to control captive and wild animal populations. We sought to develop a non-surgical method of permanent sterilization for male animals by administering the gonadotoxicant melphalan conjugated to peptides derived from the β-chain of follicle stimulating hormone (FSHβ). We hypothesized that conjugating melphalan to FSHβ peptides would magnify the gonadotoxic effects of melphalan while minimizing systemic toxicity. The ability of conjugates of melphalan and FSHβ peptides to kill murine testicular cells was first tested in vitro in a three-dimensional testicular cell co-culture system. In this system, melphalan caused considerable cell death as measured both by increases in LDH concentrations in the culture supernatant and direct visualization of the cultures. Of the conjugates tested, melphalan conjugated to a 20 amino acid peptide derived from human FSHβ consisting of amino acids 33-53 (FSHβ (33-53)-melphalan) was very potent, with cell cytotoxicity and LDH release roughly one-half that of melphalan. The effects of melphalan and FSHβ (33-53)-melphalan on spermatogenesis were then tested in vivo in mature C56Bl/6 male mice. Four weeks after intraperitoneal injection, all mice treated with either FSHβ (33-53)-melphalan or melphalan had ~75% reductions in testicular spermatid counts compared with control animals. Testicular histology revealed significant reduction in mature spermatids and spermatocytes in most tubules. However, 12 weeks after the injection, testicular spermatid counts and histology were similar to controls, except in one animal receiving FSHβ (33-53)-melphalan that had no apparent spermatogenesis. We conclude that melphalan and FSHβ (33-53)-melphalan are potent gonadotoxicants in male mice resulting in marked suppression of spermatogenesis 4 weeks after a single intraperitoneal injection. However, this effect is transient in most mice as spermatogenesis is similar to control animals 12 weeks after drug administration. Melphalan or FSHβ (33-53)-melphalan may be useful for the temporary control of fertility in male animals, but additional research will be needed to develop a single dose method of permanent sterilization for male animals.

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Optimal dose of busulfan for depleting testicular germ cells of recipient mice before spermatogonial transplantation

Cited by 50 publications

References 26 publications

Spermatogonial stem cells: Current biotechnological advances in reproduction and regenerative medicine

Spermatogonial stem cells: Current biotechnological advances in reproduction and regenerative medicine

Depletion of endogenous germ cells in tree shrews in preparation for spermatogonial transplantation

Melphalan, alone or conjugated to an FSH-β peptide, kills murine testicular cells in vitro and transiently suppresses murine spermatogenesis in vivo

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