O-211 The DNA double-strand break repair genes expression exhibits significant changes in the postnatal mouse testes from early to aged terms

Abstract

Abstract Study question How does the DNA double-strand break (DSB) repair genes, Rad51, Rpa70, Ku80, and Xrcc4, expression change in the postnatal mouse testes from early to aged terms. Summary answer The Rad51, Rpa70, and Ku80 genes expression decreased in the postnatal mouse testes from early to aged terms. What is known already The DSB levels occurring in the spermatogenic cells during spermatogenesis increase during biological aging in men. DSBs can be repaired by specific repair mechanisms including homologous recombination (HR) or non-homologous end joining (NHEJ) pathways. While RAD51 and RPA70 are the main components of HR process, KU80 and XRCC4 play important roles in NHEJ pathway. As is known, gH2AX is a commonly used biomarker for determining the DSBs. Study design, size, duration The potential reasons of DSB levels in the spermatogenic cells during aging is not fully addressed yet. In this study, we aimed to analyze the expression of the Rad51, Rpa70, Ku80 and Xrcc4 genes at mRNA and protein levels in the postnatal mouse testes from early to aged terms. Participants/materials, setting, methods We comprised five groups based on the testicular histology, consisting of early (1- and 2-week-old), young (3- and 4-week-old), adult (5- and 6-week-old), late-adult (16-, 18- and 20-week-old), and aged (48-, 50- and 52-week-old). DSB repair genes expression at mRNA and protein levels were determined using qRT-PCR and immunohistochemistry techniques, respectively. The data were evaluated by using one-way ANOVA and Tukey post hoc test. P<0.05 was considered statistically significant. Main results and the role of chance The Rad51, Rpa70, Ku80, and Xrcc4 mRNA levels significantly decreased in the aged group when compared to the young, adult and late-adult groups (P<0.05). gH2AX was intensively localized in the nucleus of primary spermatocytes of postnatal testes, and its levels either in the total or seminiferous tubules or germinal epithelial cells involving primary spermatocytes, round spermatids, elongating spermatids, elongated spermatids, and Sertoli Cells were higher in the aged group than the remained groups (P<0.05). The DSB repair proteins were detected in the spermatogenic cells, in which pachytene spermatocytes showed stronger intensity. The levels of RAD51 and RP70 proteins implicating in HR pathway were lower in the seminiferous tubules of the aged group when compared to the adult and late-adult groups (P<0.05). Moreover, the total and seminiferous tubules analysis revealed that KU80 levels decreased in the aged group in comparison to the remaining groups except for early group, as observed for the spermatogonia, primary spermatocytes and round spermatids of the aged group (P<0.05). Although there were no significant differences found in the total and seminiferous tubule analysis for XRCC4 protein, its levels decreased in the round spermatids and elongating spermatids of the aged group compared to the adult and late-adult groups. Limitations, reasons for caution The limitation of this study is that we did not isolate spermatogenic cell types from aged mice to compare adult ones. Wider implications of the findings The increase of DSB in the spermatogenic cells of aged mice may derive from reduced levels of RAD51, RPA70 and KU80 proteins playing roles in DSB repair. Further researches are required to determine the molecular mechanisms resulting in decrease of these protein levels. Trial registration number not applicable