Heat Stress Induced Testicular Impairment is Related to Orchitis and Complement Activation in Rongchang Boars

Background Heat stress is posing as a tremendous threat to pig industry, especially to the thermos-sensitive gonads of boars. Testis serves as an immune-privileged organ to keep its spermatogenesis undisturbed, while immunity is thermo-sensitive, especially for macrophages who present as the most abundant testicular immune cell. Our study aims to deeply unveil the underlying immune responses and assess its consequences to semen quality under heat stress, seeking for understanding and solutions to obstruction in boar reproductivity under heat stress. Methods We conducted the experiment using 26 8-week-old Rongchang male pigs who were assigned into thermal neutral pair-feed group and heat stress group for a 3-week experiment with the last two weeks as heat stress period. During heat stress period, pigs of heat stress group were subjected to 14-day 35 ± 1℃ heat stress, while pigs of thermal neutral pair-feed group were kept at 26 ± 1℃. Pigs were sampled at the end of heat stress period to obtain gonads tissue for assessing and measurement. Results Our findings confirmed heat stress reactions by elevated respiration rate ( P  < 0.05) and expression of heat shock proteins 60 ( P  < 0.05) and heat shock proteins 90 (P  < 0.05). Tendencies to reduce sperm motility ( P  = 0.06) and progressive sperms (P = 0.08) under HS were observed as well as significant reduction in average straight-line velocity ( P  < 0.05) and total abnormality were also recorded ( P  < 0.05). Visualized fibrosis, caspase-3 expression and accumulation of tumor necrosis factor-α ( P  < 0.05) and Interleukin-1β ( P  < 0.05) along with elevated macrophage composition ( P  < 0.05) characterized the orchitis under heat stress. Accordingly, single cell RNA sequencing revealed fluctuation in engulfing and inflammatory signals in testicular macrophages, especially promotion in complement cascades by CD163 + macropahegs, resulting in final membrane attack complex assembling ( P  < 0.05). Linear regression further exhibited a negative correlation between the membrane attack complex and sperm motility ( P  < 0.05) a long with near-negative correlation between the membrane attack complex and progressive motility (P = 0.08) or velocity straight line (P = 0.06). Conclusions Our findings highlighted the relationship between heat stress, the onset of orchitis, and the activation of the complement system, all of which caused obstruction in semen quality of boars.