EXPRESSÃO GÊNICA DE CITOCINAS NO ENDOMÉTRIO BOVINO PRÉ-SENSIBILIZADO COM ANTÍGENOS MATERNOS E PATERNOS E NA INFLAMAÇÃO SUBCLÍNICA

Abstract

Uterine infections soon after delivery and embryonic losses related to failures in the immunotolerance to the concept at the beginning of gestation are important causes of low reproductive efficiency in bovine breeding. The aim of this study was to compare the expression of the CCL5, IL-8, IL-6 and IL-1β genes in the endometrial mucosa with and without inflammation in the caruncular and intercaruncular regions using endometrial biopsy and cytobrush collection methods. Twenty uteri were used in the experiment after evaluation of the percentage of polymorphonuclear cells (PMNs) in endometrial cytology, being ten without PMNs, seven with PMNs ranging from 11 to 19% (soft endometrial inflammation) and three with PMNs between 75 and 81% (severe endometrial inflammation). These samples were collected, its RNAs were extracted, and then the reverse transcription and the qPCR reaction were performed for the quantification of the gene expression. No difference was observed in the gene expression between the caruncular and intercaruncular regions and the expression was dependent on the degree of inflammation. Soft inflammations did not present significant differences in comparison to uninflamed uteri. On the other hand, severe inflammation presented difference in comparison to the control group, independent of the collection method, biopsy or cytobrush. The influence of the endometrial collection technique in the gene expression was observed only in the severe inflammation, and the best results were observed in samples collected by cytobrush. This study had also the objective to understand the mechanisms of uterine immunotolerance to the concept by analyzing the IL-1β, IL-2, IL-6, IL-17, IFNG and LIF gene expression during the early gestation. Forty-five cows synchronized to estrus were submitted to intrauterine immunological induction, administered in D0 (estrus day), with four treatments: semen, PBMCs, semen and PBMCs and control (without induction). In vivo uterine biopsies were collected at three different times: the first at D0 (no immunological induction), the second at day seven (D7) and the final one at day 14 (D14) after immunological stimulation. After immunological induction, IL-1β mRNA expression was significantly reduced in all sensitized groups when compared to control. The IL-2 gene presented greater expression only in semen induction. It was likewise observed that IL-1β and IL-2 gene expression varied significantly between D0, D7 and D14 of the estrous cycle. The IL-6 gene did not show expression variations from the immunological inductions nor during the first 14 days of the estrous cycle. The results of the IL-17, IFNG and LIF genes were inconclusive. It was concluded that the identification of uterine inflammation through gene expression is only possible in severe cases, and the best results are observed in samples collected by cytobrush. In addition, the IL-1β and IL-2 gene expression can be modulated by immune-induction, suggesting that immunological sensitization may contribute to the success of gestation.