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Serological analysis may provide relevant information on endometritis diagnostics. Therefore, mares scheduled for AI with refrigerated semen, at the time of heat signs, underwent blood and uterine fluid samplings using a swab, uterine lavage for culture analysis, and treatment with human chorionic gonadotropin to induce ovulation. After 24–28 h, the mares were inseminated and, if positive at the culture test, treated with antibiotics chosen based on the susceptibility test. Uterine cells obtained by swabs were used for cytological examination with both classical and fluorescence techniques. Blood serum and uterine fluid samples were analyzed for assessing parameters related to redox balance, inflammation, and protease regulator potential. In blood serum, total antioxidant capacity, measured as the ferric reducing ability of plasma (FRAP), was significantly lower in cytologically endometritis-positive than -negative mares. In the uterine fluid, total thiol levels (TTL), nitric oxide metabolites (NOx), protease activity and total protein content varied significantly between groups. Although the cytological examination was more capable of discriminating between endometritis-positive and -negative mares in relation to the parameters examined, no statistically significant differences emerged in terms of pregnancy rate in relation to cytological and culture diagnosis as well as in mares diagnosed as positive and negative for endometritis.

The etiopathogenesis of endometritis in mares and cows differs significantly; this could depend on a different sensitivity and reactivity of the uterus but also on endocrine and rearing factors and different stress sources. In both species, microorganisms and the immune system play a primary role in the generation of this pathology. Microbiological and cytological tests support clinical examination and significantly improve diagnostic accuracy. For both species, during the inflammation, immune cells invade the endometrium and release bioactive substances to contrast primary or secondary pathogen contamination. These molecules are traceable to cytokines, chemokines, and prostaglandins as well as reactive oxygen and nitrogen species (ROS and RNS), collectively known as RONS. The RONS-mediated oxidation causes morphological and functional alterations of macromolecules, such as proteins, lipids, and nucleic acids, with the consequent production of derivative compounds capable of playing harmful effects. These bioactive molecules and by-products, which have recently become increasingly popular as diagnostic biomarkers, enter the bloodstream, influencing the functionality of organs and tissues. This review has collected and compared information obtained in cows and mares related to the diagnostic potential of these biomarkers that are assessed by using different methods in samples from either blood plasma or uterine fluid.

By exploiting the PMN property to produce high quantities of oxygen peroxide to neutralize pathogens, the oxygen peroxide content of uterine cells was measured to diagnose endometritis. After preliminary in vitro studies in which endometrial cells from slaughtered mares were mixed with leukocytes from peripheral blood, endometrial samples were collected by uterine flushing from mares before insemination. Staining endometrial cells with H2DCF-DA was combined with hydroethidine to normalize the fluorescence intensity with the cellular content of the sample. Stained cell smears were assumed as the gold standard of endometritis, and based on this assay, the samples were considered positive (C+) and negative (C−) for endometritis. The amount and the turbidity of fluid recovered by uterine flushing were significantly (p < 0.01) higher in C+ than in C−. Moreover, the oxygen peroxide content of the endometrial cells was significantly higher in the C+ than in the C− group (6.31 ± 1.92 vs. 3.12 ± 1.26, p = 0.001). Using the value of 4.4 as the cutoff level of this fluorescence cytology assay, it was found that only one C− sample exceeded the cutoff level (false positives = 7.7%) while three C+ samples showed values below the cutoff level (false negative = 11.5%).