Effects of cyclophosphamide exposure of male rats on progeny outcome and site of action on male germ cell nuclei in the testis and epididymis

Abstract

Cyclophosphamide is a bifunctional alkylating agent. Administration of low doses of cyclophosphamide to male rats produces severe effects on the outcome of their progeny. The studies in this thesis show that one week treatment with cyclophosphamide can produce about 30% post-implantation loss. This indicates that post-testicular spermatozoa are sensitive to alkylating agents. Previous studies have shown that six weeks treatment caused more than 95% post-implantation loss. The effects of cyclophosphamide treatment on rat spermatozoal nuclei in the testis and epididymis were also addressed in this thesis. Rat cauda epididymal spermatozoa collected after one week of cyclophosphamide treatment showed no change in decondensation pattern in vitro; while samples obtained from rats treated for six weeks showed a slowed chromatin elongation and dispersion pattern. Iodoacetamide binding revealed that there was about a 30% decrease in the amount of free sulfhydryls in spermatozoal nuclei from six weeks treated samples, while there was no detectable change in one week treated samples. DNA single strand breakage was detected in one week treated samples only in the presence of proteinase K in the lysis solution, while significant amounts of both DNA single strand breakage and DNA cross-links were detected in six weeks treated samples. Using a DNA synthesis in vitro system, it was found that six weeks treatment decreased spermatozoal template function, while one week treatment did not result in any significant difference in DNA template function when compared with the control. It is proposed that as the mele germ cells undergo the chromatin transition processes, the chromatin is in an open dynamic status, which expresses many vulnerable sites of the genome to aklylating agent. Since there is no DNA repair system, the alkylation of the genome is cumulative and causes severe damage to the germ cell nuclei; this would lead to early embryo death. Furthermore it is hypothesized tha