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. 2016 May;4(2):175.
doi: 10.4172/2375-4508.1000175. Epub 2016 Mar 18.

Cryorecovery of Mouse Sperm by Different IVF Methods Using MBCD and GSH

Ming-Wen Li  1 , Olivia C Glass  1 , Jasmin Zarrabi  1 , Lisa N Baker  1 , K C Kent Lloyd  1
Affiliations

Cryorecovery of Mouse Sperm by Different IVF Methods Using MBCD and GSH

Ming-Wen Li et al. J Fertili In Vitro. 2016 May.

Abstract

Different protocols incorporating methyl-β-cyclodextrin (MBCD) and reduced glutathione (GSH) have been reported to improve IVF recovery of cryopreserved mouse sperm on a C57BL/6 (J and N) genetic background. However, it is not clear which IVF protocol is most appropriate when using the various methods to cryorecover sperm with different sperm quality and sample volumes. Therefore, in the present study we correlated sperm motility with fertilization rate and compared the efficiency of different IVF methods using various sperm samples so as to establish general guidelines for mouse sperm cryorecovery by IVF. High linear correlation between sperm fertilization rate and progressive motility was found, R2 was 0.9623 and 0.9993 for pre-freezing and post-thaw progressive motility, respectively. High amounts of cryoprotective agent (CPA) were observed to impair both sperm capacitation and fertilization. Moreover, the presence of a large number of immotile sperm in the sperm-oocyte co-incubation drop was found to reduce IVF success which could be partially reversed by supplementation using monothioglycerol (MTG) during centrifugation. It was concluded that the efficiency of IVF using cryorecovered mouse sperm in media containing MBCD and GSH can be predicted from sperm progressive motility. High concentrations of CPA and immotile sperm should be mitigated prior to IVF. The optimum IVF method should be selected based on sperm sample volume and sperm parameters.

Keywords: Cryopreservation; IVF; Mouse; Sperm.

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Figures

Figure 1
Figure 1
Correlation between post-thaw sperm progressive motility and fertilization rate of MBCD-GSH IVF, where n represents the number of IVF procedures performed for each mean progressive motility.
Figure 2
Figure 2
Linear regression between the mean of sperm progressive motility (pre-freeze and post-thaw) and the mean of fertilization rates of MBCD-GSH IVF. Ypre = pre-freeze progressive motility; Ypost = post-thaw progressive motility; X = IVF rate.
Figure 3
Figure 3
Comparisons of cryorecovery IVF rates of different IVF methods using wildtype C57BL/6N sperm. Each IVF procedure was repeated 3 times. Compared with the other 3 IVF methods, the IVF rate of the “IVF in MBCD” method was significantly lower (P<0.05).
Figure 4
Figure 4
Effect of high CPA concentration on IVF rates of MBCD-GSH IVF and Rescue MBCD-GSH methods (n=4, P<0.05).
Figure 5
Figure 5
Comparison of IVF rates of Rescue MBCD-GSH IVF and Rescue IVF in MBCD using sperm samples with high concentration and low motility (n=4, P<0.05).
Figure 6
Figure 6
Comparison of IVF rates of Rescue IVF in MBCD using sperm centrifuged in the presence (RVF+MTG) or absence of MTG (RVF only). n=3, P<0.05.
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