Sperm mRNA Transcripts Are Indicators of Sub-Chronic Low Dose Testicular Injury in the Fischer 344 Rat

Abstract

Current human reproductive risk assessment methods rely on semen and serum hormone analyses, which are not easily comparable to the histopathological endpoints and mating studies used in animal testing. Because of these limitations, there is a need to develop universal evaluations that reliably reflect male reproductive function. We hypothesized that toxicant-induced testicular injury can be detected in sperm using mRNA transcripts as indicators of insult. To test this, we exposed adult male Fischer 344 rats to low doses of model testicular toxicants and classically characterized the testicular injury while simultaneously evaluating sperm mRNA transcripts from the same animals. Overall, this study aimed to: 1) identify sperm transcripts altered after exposure to the model testicular toxicant, 2,5-hexanedione (HD) using microarrays; 2) expand on the HD-induced transcript changes in a comprehensive time course experiment using qRT-PCR arrays; and 3) test these injury indicators after exposure to another model testicular toxicant, carbendazim (CBZ). Microarray analysis of HD-treated adult Fischer 344 rats identified 128 altered sperm mRNA transcripts when compared to control using linear models of microarray analysis (q<0.05). All transcript alterations disappeared after 3 months of post-exposure recovery. In the time course experiment, time-dependent alterations were observed for 12 candidate transcripts selected from the microarray data based upon fold change and biological relevance, and 8 of these transcripts remained significantly altered after the 3-month recovery period (p<0.05). In the last experiment, 8 candidate transcripts changed after exposure to CBZ (p<0.05). The two testicular toxicants produced distinct molecular signatures with only 4 overlapping transcripts between them, each occurring in opposite directions. Overall, these results suggest that sperm mRNA transcripts are indicators of low dose toxicant-induced testicular injury in the rat.

Biomarkers of chemotherapy-induced testicular damage

“…While the mRNA content of sperm is low relative to many human or animal tissues, it is sufficient to detect exposure-related changes. Moreover, a panel of sperm mRNA transcripts has been developed that can detect and predict low-level exposures to Sertoli cell toxicants in rats (59). A similar approach may be used to identify the effects of chemotherapy-induced testicular or sperm damage at the molecular level.…”

Small RNA sequencing of cryopreserved semen from single bull revealed altered miRNAs and piRNAs expression between High- and Low-motile sperm populations

“…Furthermore, sperm transcripts retain information of the past events of spermatogenesis and probably contribute to egg fertilization and development. Comparisons between sperm from fertile and infertile males in different species indicate that sperm transcripts may have diagnostic value, and suggest a relationship between sperm transcripts composition and proper sperm functions [8, 14–17]. …”

Assessing Male Reproductive Toxicity during Drug Development

“…Of the sperm biomarkers, the human sperm membrane protein SP22 was shown to decline after exposure to both epidydimal and testicular toxicants in rat and ram models, changes in human sperm gene expression or mRNA transcript content [24,[42][43][44]. DNA methylation profile during human spermatogenesis were correlated with impair of sperm quality (such as concentration, morphology and motility); and another specific group of sperm mRNA transcripts was shown to predict low level exposures to Sertoli cell toxicants in the rat [45][46][47].…”