Leucocyte migration and amount of COX-2 protein in the susceptible mares endometrium treated with platelet-rich plasma

The Post-Mating Induced Endometritis (PMIE) is characterized by the exacerbated influx of polymorphonuclear cells (PMNs) into the uterus and inability to eliminate intrauterine fluid after breeding, resulting in an unfavourable environment to embryo growth. The platelet rich-plasma (PRP) is an inexpensive and accessible product that has been used in the treatment of localized inflammations. Thus, the present study aimed to determine if the intrauterine infusion of PRP could modulate the uterine inflammatory response in susceptible mares. A total of 13 mares classified as susceptible to PMIE based on presence of intrauterine fluid 48 hours after artificial insemination (AI), more than 20% of polymorphonuclear cells (PMNs) 24 hours after AI and historic of less than 30% of embryo recovery rates were used for this study. The mares were inseminated with fresh semen in three consecutive cycles in a cross-over study design. PRP was prepared by single centrifugation protocol and just samples with a minimum concentration of 250.000 platelets/μL were used for treatments. The cycles were classified as control cycle (C): no pharmacological interference; PreT: 20 mL of PRP was infused 24 hours before AI; PostT: 20 mL of PRP was infused four hours after AI. Follicular dynamic was monitored daily by transrectal ultrasound and when a >35 mm follicle and endometrial edema at least grade 2 (min. 0 and max. 4) were detected, the ovulation induction was performed with deslorelin acetate (1mg, IM). AI was performed 24 hours after ovulation induction with the semen deposition (800 x 106 total sperm) in the uterine body. The percentage of PMNs in uterine cytology (CIT) was observed before and 24 hours after AI, using bright-field microscopy; in the biopsy (HIP) samples five fields were count and the average number of PMNs obtained, samples with an average of >17 PMNs cells were positive for endometritis; the number of COX-2 positive cells was evaluated by immunohistochemistry using a score system (0 – negative to COX-2; 1 – mild presence of positive cells; 2 – moderate presence of positive cells; 3 – severe presence of positive cells). The results were evaluated by Kruskal-Wallis test followed by Tukey-Krammer test, considering a 5% significance level. It was possible to observe that PMNs number decrease (P < 0.05) in CIT in both treated cycles (PreT & PostT) after AI (%, C – 46.6±6.3; PreT – 13.3±3.1; PostT - 18±5.2). The number of positive mares to endometritis in the HIP, also decrease (P < 0.05) in both treated cycles (Positive: C – 69%; PreT – 15%; PostT – 23%). Intensely (P < 0.05) positive COX-2 labelling was observed in the control (Score 2 – 30.7%; 3 – 69.2%) cycle compared to the PreT (1 – 15.4%; 2 – 84%) and PostT (1 – 30.7%; 2 – 53.8; 3 – 15.4) cycles. In conclusion, PRP is benefit to susceptible mares downregulating the PMNs influx into the uterine lumen and the amount of COX-2 protein in the endometrium after breed, becoming an option as a complementary treatment in these mares.