The Effects of Chemotherapy with Bleomycin, Etoposide, and Cis-Platinum (BEP) on Rat Sperm Chromatin Remodeling, Fecundity and Testicular Gene Expression in the Progeny1

Abstract

During spermiogenesis, histones are replaced first by transition proteins and then by protamines, resulting in a very condensed sperm DNA structure that is absolutely critical for normal sperm function. We have demonstrated previously that, despite a 9-wk recovery period, mature sperm from rats treated for 9 wk with bleomycin, etoposide, and cis-platinum (BEP), the drugs used to treat testicular cancer, have reduced levels of protamine 1 and a concomitant upregulation of specific histones, highlighting a problem in histone eviction. Here, we demonstrate that regulators of histone removal are increased in elongating spermatids following recovery; however, Ac-H4 and gamma H2AX histones remain elevated in elongating spermatids or caudal epididymal spermatozoa 9 wk post-BEP treatment. This indicates that chromatin remodelers and effector proteins that respond to histone removal cues may be a target of BEP treatment. A decrease in the expression of SMARCE1 in elongating spermatids may explain the persistent retention of histones in cauda epididymal sperm 9 wk after the cessation of BEP treatment. Remarkably, proteins implicated in the translational control and posttranslational processing of protamine 1 are also significantly elevated 9 wk post-BEP treatment, suggesting that histone eviction may dictate the DNA availability for protamine binding. Males mated to control females 9 wk after BEP treatment have reduced litter sizes; moreover, the profile of gene expression in the developing testes of their pups is altered. Altering the proportion of histones to protamine in mature spermatozoa has an adverse impact on male fecundity, with modifications to epigenetic marks potentially threatening normal progeny development.

Paternal exposure to testis cancer chemotherapeutics alters sperm fertilizing capacity and affects gene expression in the eight-cell stage rat embryo

“…Interestingly, similar to our results (Table 1), gene expression at the eight-cell stage was mostly upregulated in embryos derived from benzo[a]pyrene-exposed fathers compared with controls (Brevik et al, 2012). Furthermore, they strengthen our hypothesis that the more open chromatin structure of spermatozoa following a prolonged cessation of BEP treatment may lead to dysregulated gene expression in the developing embryo (Maselli et al, 2013). Furthermore, they strengthen our hypothesis that the more open chromatin structure of spermatozoa following a prolonged cessation of BEP treatment may lead to dysregulated gene expression in the developing embryo (Maselli et al, 2013).…”

“…Together, these results indicate that the eight-cell stage may represent a sensitive window in embryo development for paternally mediated chemically induced changes in gene expression. Furthermore, they strengthen our hypothesis that the more open chromatin structure of spermatozoa following a prolonged cessation of BEP treatment may lead to dysregulated gene expression in the developing embryo (Maselli et al, 2013).…”

“…In animal studies, treatment with BEP causes DNA damage, a disruption of spermatogenesis leading to a decrease in sperm count and spermatozoa with abnormal morphology and chromatin structure (Bieber et al, 2006;Delbes et al, 2007;Maselli et al, 2012). In addition, an increase in the early post-natal mortality of pups sired by BEP-treated males is observed (Bieber et al, 2006;Maselli et al, 2013). As with BEP, treatment with cyclophosphamide, another chemotherapeutic drug, induces DNA damage in spermatozoa (Barton et al, 2003;Codrington et al, 2004); paternal exposure leads to a perturbed epigenetic reprogramming in the pre-implantation rat embryo (Barton et al, 2005), while triggering a DNA damage response at various stages of early embryogenesis (Barton et al, 2007;Grenier et al, 2012).…”

Effects of Four Chemotherapeutic Agents, Bleomycin, Etoposide, Cisplatin, and Cyclophosphamide, on DNA Damage and Telomeres in a Mouse Spermatogonial Cell Line1

“…In the Terc gene knockout mouse, severe male sterility, caused by telomere shortening, was observed after the sixth generation [43], but progressive defects in germ cells, with consequences on embryo development, were detected in the second generation [6]. Treatment of male rats with bleomycin, etoposide, and cisplatin induced adverse effects on progeny during early embryonic development [44]. In cancer survivors, sperm DNA damage may persist 2 yr posttreatment with chemotherapeutic agents [1,45].…”

Genetic Effect of Chemotherapy Exposure in Children of Testicular Cancer Survivors

“…In addition, non-genetic effects, such as epigenetic changes(34, 35) or transcription-based alternative splicing not observable with whole genome sequencing, may identify transmissible mutagenic effects in humans and warrant further evaluation.…”