The effect of platelet-rich plasma on endometrial pro-inflammatory cytokines in susceptible mares following semen deposition

Abstract

Mares susceptible to persistent mating-induced endometritis (PMIE) are a source of substantial economic inefficiency. Recent studies report a significant post insemination up-regulation of endometrial anti-inflammatory cytokines IL-10 and IL-Ra in normal versus susceptible mares, as well as significantly increased and prolonged expression of IL-1b, IL-6, IL-8, and IL1Ra in susceptible mares following intrauterine inoculation with E.coli [1]. Furthermore, nitric oxide is suspected to play a significant role in uterine clearance, because increased endometrial expression of inducible nitric oxide synthetase (iNOS) has been reported in susceptible mares following insemination [2]. This project was developed to investigate the expression of inflammatory biomarkers of uterine biopsies obtained from susceptible mares following intrauterine PRP (platelet-rich plasma) infusion. Platelets contain alpha-granules that, upon activation, secrete bioactive molecules with regenerative capacity in the form of growth factors, cytokines, and chemokines. Barrenmares with a history of PMIE (n 1⁄4 9) underwent daily ultrasonographic examination of the reproductive tract per rectum while in estrus. All mares underwent an untreated (no PRP) cycle followed by a treated (PRP) cycle. When a dominant follicle >35 mm in diameter and evidence of endometrial edema were detected, 1.0 mg deslorelin acetate (SucromateR Bioniche Animal Health Pullman, WA) was administered intramuscularly. On the treatment cycle, 180 ml of citrated whole blood was obtained for PRP processing in a specialized blood fraction separating centrifuge system (Angel Cytomedix, Inc. Gaithersburg, MD). The resultant PRP was brought to a final volume of 10 ml with plateletpoor plasma and infused into the uterus. Mares were inseminated with >200 million motile sperm 24-36 hours following deslorelin administration. An endometrial biopsy obtained the following day was preserved in RNAlaterR (Sigma Inc., St Louis, MO) for reverse transcription polymerase chain reaction (RT–PCR) analysis [1] for mRNA expression of inflammatory biomarkers IL-1b, IL1-Ra, TNFa, IL-6, IL-8, IL-10, and iNOS. Data were analyzed using SAS (SAS Institute, Cory NC) with Wilcoxon signed-rank test (non-parametric). Comparisons were made between pretreatment and post-treatment groups. Significance was set to P < 0.05. In susceptible mares treated with an intrauterine infusion of PRP, the mRNA expression of IL-1b, IL-6, IL-8 and iNOS was significantly down-regulated (P < 0.05) when compared with untreated mares. All other cytokines were similar in treated and untreated mares. Therefore, following semen deposition, PRP influences the mRNA expression of endometrial pro-inflammatory cytokines and iNOS in mares susceptible to PMIE. References