Effect of scrotal insulation on clusterin-positive cells in ram semen and their relationship to semen quality

Scrotal contents of 2 rams were insulated for 96 hours and the fraction (as a percentage) of clusterin-positive cells (CPCs) and its relationship to s emen quality was investigated. Semen collection was started 18 days before insulation and was terminated on day 78 and day 63 after insulation in anim als 1 and 2, respectively. Sperm clusterin was localized by immunostaining with anti-bovine clusterin antibody (anti-bCAb) and fluorescein isothiocyan ate-conjugated immunoglobulin G. Scrotal insulation led to deterioration of semen quality and increased the percentage of CPCs in both rams. Two types of sperm reactivity were observed: an extensive, intensive staining patter n (ESP); and a localized, less-intensive staining pattern (LSP). The percen tage of ESP-CPCs began to increase from day 6 and reached 88.8% and 100% on day 15 after insulation in animals 1 and 2, respectively. The increase in CPCs coincided with the presence of a high percentage of teratoid forms (88 .3%) in semen from animal 1, and detached heads (81.4%) in semen from anima l 2. After normal semen production was restored on day 60 in animal 1, the percentages of ESP-CPCs and LSP-CPCs returned to preinsulation rates, where as only the ESP-CPCs returned to normal in animal 2. A negative relationshi p was observed between ESP-CPCs and total sperm/ejaculate (r = -.62), motil ity (r = -.78), viability (r = -.68), and filtration rate (r = -.71) in sem en from animal 1. Conversely, a positive relationship was seen between ESP- CPCs and total abnormal spermatozoa (r =.82). Similar results were obtained in semen from animal 2. CPCs were nearly completely absent in glass wool-S ephadex (GWS)-filtered semen, suggesting a role for clusterin in the proces s of trapping abnormal spermatozoa in the GWS filters. We conclude that abe rrant spermatogenesis induced by scrotal insulation increases the percentag e of CPCs in ram semen. We suggest that the percentage of CPCs in ram semen could be a useful marker in poor-quality ejaculates.