Sperm protein reactive with antisperm antibody is immunoexpressed in equine primordial, primary, secondary, and tertiary follicles

Sperm protein reactive with antisperm antibody (SPRASA) is present in all stages of ovarian follicles in humans, mice, cows, dogs, and cats, 1,2 and is localized to granulosa cells and ooplasm.1,2  Objective was to determine if equine ovarian follicles expressed SPRASA. We hypothesized that SPRASA immune expression would be present in all stages of equine ovarian follicle, and localize to granulosa cells and ooplasm. Routine immunohistochemistry was performed on formalin-fixed paraffinembedded ovarian sections (4 µm) from 3 adult grade Quarter Horse mares. Briefly, sections were deparaffinized and rehydrated before antigen retrieval was accomplished by heat activation in a sodium citrate buffer. Endogenous peroxidase activity was blocked using 3% hydrogen peroxide and nonspecific binding was blocked using serum-free protein block (Dako #X0909, Carpinteria, CA). Polyclonal antiSPRASA primary antibody was applied to slides at a 1:200 dilution. Universal negative rabbit antibody (Dako #S3022) was applied to adjacent sections to serve as negative control. Horseradish peroxidase-conjugated antirabbit polymer secondary antibody (Immuno BiosCience, #IH-8064-OSU-15, Mukilteo, WA) was applied undiluted to all slides. Peroxidase activity was detected using Nova Red Kit (Vector Laboratories Inc, #SK4800, Burlingame, CA). Slides were counterstained with hematoxylin, dehydrated, and cover slipped. Digital images were captured at 10 and 40 x magnifications (QImaging #QIC-F-M-12-C). SPRASA expressed in pregranulosa cells of primordial, and in granulosa cells of primary, secondary, and tertiary follicles in equine ovaries. SPRASA was not expressed in equine ooplasm. There was no positive staining in negative controls. In US, wild horse and burro populations have drastically exceeded the carrying capacity of public lands where they are managed. With the exception of surgery (removal of ovaries), current methods for sterilizing wild female horses and burros (e.g. porcine zona pellucida or GnRH immunization) are temporary reversible solutions. Immunization against SPRASA may prove to be a permanent nonsurgical sterilant method because it targets all follicles including primordial follicles.

Keywords: Antisperm antibodies, contraceptive, horse, immunohistochemistry, oocyte, SPACA3

References

1. Chiu WW, Erikson EK, Sole CA, et al: SPRASA, a novel sperm protein involved in immune-mediated infertility. Hum Reprod 2004;9:243-249.
2. Wagner A, Holland OJ, Tong M, et al: The role of SPRASA in female fertility. Reprod Sci 2015;22:452-461.

This manuscript was originally published in the journal Clinical Theriogenology Vol 12(3) Sept 2020.  Clinical Theriogenology is the official journal of the Society for Theriogenology (SFT) and the American College of Theriogenologists (ACT).  This content has been reproduced on the IVIS website with the explicit permission of the SFT/ACT.