The Study Of Effect Of Reduced Glutathione On In Vitro Fertilization Of Fresh And Frozen/Thawed C57BL/6J Mouse Sperm

Sperm cryopreservation and in vitro fertilization(IVF)are useful for the effective reproduction and storage of genomic resources derived from genetically engineered mice.However,sperm cryopreservation of C57BL/6 mice,the most commonly used genetic background for genetically engineered mice,considerably reduces its fertility.In the present study,in contrast to the fertilization medium without reduced glutathione(GSH),we examined the effect of different concentration of GSH added to the fertilization medium on the fertilization rate,the rate of blastocyst development and the birth rate of fresh and frozen/thawed C57BL/6J mouse sperm.We found that adding reduced glutathione to the fertilization medium increased the fertilization rate,but did not adversely affect embryo development in vitro or in vivo.We used this technique with 5 strains of genetically engineered mice having a C57BL/6J background in order to verify the influence of reduced glutathione.In order to understand the mechanism of the enhancement of fertilization mediated by GSH,an attempt to determine the effects of amino acid constituents of GSH on the fertilization rate,the rate of blastocyst development and the birth rate of frozen/thawed C57BL/6J mouse sperm was made.Meanwhile,we also observed the effect of GSH and its amino acid constituents on the amount of free thiols in the zona pellucida.Results showed that the GSH-stimulated fertilization was associated with an increase in the number of free thiols in the zona pellucida.Thus,we demonstrated that adding GSH to fertilization medium enhanced the fertility of fresh and frozen/thawed C57BL/6J mouse sperm and that the enhancement was achieved by the interaction between the free thiols of GSH and the disulfide bonds.