Effect of cryopreservation on DNA normality of mice epididymal sperms following in vitro preparation with pentoxifylline and Glycyrrhiza glabra

Abstract

Background:Many studies conducted on the pentoxifylline (PX) and Glycyrrhiza glabra (G.glabra) as motility I stimulants showed positive effect on sperm preparation in vitro improving the progressive forward movement. However, it was not known the impact of these stimulants on the nature of | genetic material, especially after cryopreservation.Objective:The experiment was designed to find out any harmful effect of a medium containing a mixture of PX and aqueous extract of G.glabra on the sperm DNA before and after cryopreservation.Materials and Methods:Fifty mature fertile male mice 812-weeks old were used. Epididymal sperms were obtained from caudal region and prepared in vitro by direct activation technique using four media namely;PX alone, G.glabra alone, a mixture of PX with G.glabra and Ham's F-12 (as control medium). Certain sperm function characteristics were examined as well as evaluation of DNA integrity | by acridine orange test and comet assay before and after cryopreservation.Results:The results showed a highly significant (P<0,001) improvement in the sperm forward movement, with a highly significant (P0.001) decrease in DNA abnormality using acridine orange test | and comet assay, following in vitro activation by the four media compared to before activation. Following in vitro activation and cryopreservation, the results of epididymal sperm showed a highly significant (P<0.001) decrease in DNA abnormality by using PX medium alone, G.glabra medium alone and the mixture of PX and G.glabra compared to control medium and the results before cryopreservation.Conclusion:According to the results of present study, it has been found no toxic effect of medium containing PX and/or G.glabra on the epididymal sperms DNA after activation in vitro and cryopreservation using acridine orange test and comet assay. The best result obtained when using a medium containing a mixture of PX+ G.glabra together. These data could be applied for preparation and activation of epididymal and testicular sperms obtained from obstructive azoospermic men.