Role of the Nitric Oxide System in Persistent Breeding-Induced Endometritis in the Mare

ROLE OF THE NITRIC OXIDE SYSTEM IN PERSISTENT BREEDING-INDUCED ENDOMETRITIS IN THE MARE Firdous Ahmad Khan Advisor: University of Guelph, 2016 Dr. Tracey S. Chenier Co-advisor: Dr. Elizabeth L. Scholtz Persistent breeding-induced endometritis (PBIE) is a major cause of infertility in the mare. Delayed uterine clearance, attributed mostly to impaired myometrial contractility, is recognized as an important factor in the development of PBIE. Nitric oxide (NO) may play a role in determining susceptibility of mares to PBIE through an inhibitory effect on uterine contractility. The purpose of this research was to investigate the role of the nitric oxide system in the pathogenesis of PBIE in mares. First, a randomized controlled experiment was used to evaluate the effect of NO on spontaneous uterine contractility. Using an in-vitro muscle contractility model, uterine tissue strips from euthanized mares were treated with increasing concentrations of S-nitroso-N-acetylpenicillamine (an NO donor) and N-acetyl-D-penicillamine (vehicle and timematched control) at regular intervals. Results indicated a dose-dependent inhibitory effect of NO on spontaneous uterine contractility. Secondly, differences in endometrial nitric oxide synthase (NOS) activity between PBIE-susceptible and resistant mares, and the effect of a specific inducible nitric oxide synthase (iNOS) inhibitor on endometrial NOS activity, were evaluated. Mares susceptible or resistant to PBIE were selected based on preset criteria and the results of an intrauterine challenge with killed spermatozoa during estrus. Endometrial biopsy samples were cultured in L-arginine supplemented minimum essential medium with or without a specific iNOS inhibitor. The medium and the endometrial tissue were collected after 24 hours of culture to determine endometrial NOS activity. Susceptible mares had significantly greater endometrial NOS activity than resistant mares. The iNOS inhibitor treatment resulted in a significant reduction in endometrial NOS activity. Coupled with the previously reported presence of increased NO levels in the uteri of susceptible mares, the dose-dependent inhibition of spontaneous uterine contractility by NO and the greater endometrial NOS activity in susceptible mares in the studies described in this thesis suggest that the nitric oxide system may play an important role in the development of PBIE in mares. The reduction of endometrial NOS activity by the iNOS inhibitor treatment provides a basis for clinical testing of NOS inhibitors as preventative or therapeutic options for PBIE in mares. PUBLICATIONS ARISING FROM THIS RESEARCH Khan FA, Scholtz EL, Chenier TS. The nitric oxide system in equine reproduction: current status and future directions. J Equine Vet Sci 2015;35:481–7. Khan FA, Chenier TS, Murrant CL, Foster RA, Hewson J, Scholtz, EL. Dose-dependent inhibition of uterine contractility by nitric oxide: a potential mechanism underlying persistent breeding-induced endometritis in the mare. Theriogenology 2017;90:59–64. Khan FA, Chenier TS, Foster RA, Hewson J, Scholtz, EL. Endometrial nitric oxide synthase activity in mares susceptible or resistant to persistent breeding-induced endometritis and the effect of a specific iNOS inhibitor. Theriogenology (submitted for publication). Khan FA, Chenier TS, Murrant CL, Foster RA, Hewson J, Scholtz EL. Nitric oxide’s dosedependent inhibition of uterine contractility: a potential mechanism underlying persistent breeding-induced endometritis in the mare. Clin Theriogenol 2016;8:307 (Abstract). Khan FA, Murrant CL, Chenier TS, Hewson J, Foster RA, Scholtz EL. The effect of nitric oxide on in-vitro contractility of the ovine uterus: a dose response study. Proc 43 rd Annual Southern Ontario Reproductive Biology Meeting, McMaster University, Hamilton, Canada, 2015 (Abstract P27).