PS6-4. Localization of Toll-like Receptor 4 in the Reproductive Tract of Boars Prior to and Post Puberty Attainment.

Abstract Hormones, such as Testosterone and Dihydrotestosterone (DHT), play essential roles in male sexual development. In both rodents and humans, the expression of toll-like receptors (TLRs) in various tissues has been shown to be regulated by these hormones. Preliminary results also indicate potential TLR expression in the acrosomal region of sperm cells. Therefore, the objective of the current study was to immunolocalize TLR4 and peanut agglutinin (PNA) in pre- and post-pubertal boar testes and epididymal tissues. Intact reproductive tracts of crossbred boars (n = 4 pre-pubertal and n = 7 post-pubertal) were harvested and tissues were fixed in 4% paraformaldehyde prior to paraffin embedding and sectioning. Blood was also collected and processed for plasma before testosterone and DHT concentrations were quantified using a competitive ELISA and chemiluminescence via the Texas A&M Veterinary Medical Diagnostic Laboratory, respectively. Concentrations of testosterone and DHT were analyzed by PROC GLM in SAS 9.4 with fixed effect of puberty status and results were reported as least square means ± SEM. The concentration of testosterone differed between pre- and post-pubertal boars (0.96 ± 0.36 vs. 3.01 ± 0.26 ng/mL, respectively; P < 0.0001); thus, indicating that blood testosterone concentration rises in post-pubertal as compared to pre-pubertal boars as anticipated. There were no differences in DHT concentrations between pre- and post-pubertal boars (9.35 ± 0.52 vs. 8.92 ± 0.42 ng/mL, respectively; P = 0.53). Immunoreactive TLR4 was detected within the interstitial space; however, it was not detected in seminiferous tubules of pre-pubertal boar. Similarly, TLR4 was detected in the interstitial tissue, while remaining undetectable in the seminiferous tubules of post-pubertal boars. In pre-pubertal boar epididymal tissues, TLR4 was localized to the luminal epithelium. TLR4 was present throughout luminal epithelium of caput, corpus, and cauda epididymis of post-pubertal boars, as well as the vascular endothelium. This pattern was also observed in post-pubertal boar epididymal tissue, with majority of localization at the apical portion of the epididymal epithelium. Slight colocalization of TLR4 with PNA was observed on the acrosomal region of sperm cells within the lumen of the cauda epididymal duct. Based on these findings, TLR4 is present in specific regions of reproductive tissues in both pre- and post-pubertal boars. However, immunolocalization of TLR4 was more intensive in post-pubertal boar testes and epididymal tissues compared to pre-pubertal tissues. The observed differences in TLR expression could be associated with circulating steroid hormone concentrations. In addition, slight colocalization of TLR4 with PNA in the cauda epididymis suggests a potential role for TLR4 in sperm maturation or acrosomal function. These results indicate a possible regulatory relationship between steroid hormones and TLR expression in the male reproductive tract, warranting further investigation.