Production of IL-1ß, IL-6 and CXCL8 by endometrium of crossbred heifers stimulated with various pathogen-associated molecular patterns.

Uterine bacterial infections are common during the post-partum period of dairy herds and, apparently, incidences in crossbred cattle are less than in Holsteins. The aims of this study were (I) to evaluate production of interleukin 1-ß (IL-1ß), interleukin-6 (IL-6) and chemokine CXCL8 using endometrial explants from Bos indicus crossbred heifers at diestrous, stimulated by various pathogen-associated molecular patterns (PAMP), and (II) assess production of these cytokines by lipopolysaccharide-stimulated endometrial explants from heifers when samples were collected at different stages of estrous cycle. In the first experiment, endometrial explants from heifers at diestrous were stimulated by ten-fold serial dilutions of lipopolysaccharide (LPS), triacylated lipopeptide (PAM3) or peptidoglycan (PGN). In the second experiment, endometrial explants collected at different stages of the estrous cycle were treated with LPS. Concentrations of IL-1ß, IL-6 and CXCL8 were quantified in supernatant. There was a marked (P < 0.05) production of IL-1ß, IL-6, and CXCL8 in response to LPS treatment. There was also production of IL-1ß (P < 0.05) in response to PGN treatment. Explant samples collected at different stages of the estrous cycle responded to LPS treatment with production of IL-1ß and IL-6, but with no differences (P > 0.05) between stages of estrous cycle. In conclusion, endometrial samples of crossbred Zebu-based heifers collected during diestrous produced IL-1ß, IL-6 and CXCL8 in response to LPS and IL-1ß in response to PGN. The cytokine production in response to LPS, however, was not affected by the stage of the estrous cycle in Bos indicus crossbred heifers.

Transcription of pattern recognition receptors and abortive agents induced chemokines in the bovine pregnant uterus.

Pattern recognition receptors (PRRs) are important components of the innate immune system whose ligands are specific pathogen associated molecular patterns (PAMPs). Considering the scarcity of studies on transcription of PRRs in the pregnant uterus of cows, and its response to PAMPs and microorganisms that cause abortion in cattle, this study aimed to characterize the transcription of TLR1-10, NOD1, NOD2 and MD2 in bovine uterus throughout gestation and to investigate the sensitivity of different uterine tissues at third trimester of pregnancy to purified TLR ligands or heat-killed Brucella abortus, Salmonella enterica serotype Dublin (S. Dublin), Listeria monocytogenes, and Aspergillus fumigatus, by assessing chemokine transcription. RNA extracted from endometrium, placentome and intercotiledonary region of cows at the first (n=6), second (n=6), and third (n=6) trimesters of pregnancy were subjected to real time RT-PCR. After stimulation of endometrium and intercotiledonary regions with purified TLR ligands or heat-killed microorganisms, gene transcription was assessed by real time RT-PCR. In the placentome, there was no significant variation in TLRs transcription throughout the three trimesters of pregnancy. In the endometrium, there was significant variation in TLR4 and TLR5 transcription during the three stages of gestation; i.e. TLR4 transcription was higher during the third trimester, whereas TLR5 transcription was higher during the last two trimesters. In the intercotiledonary region, there was significant variation in transcription of TLR1/6, TLR7, and TLR8, which were more strongly expressed during the first trimester of pregnancy. At the third trimester of gestation, significant transcription of CXCL6 and CXCL8 was detected mostly in endometrial tissues in response to purified TLR4 and TLR2 ligands. Transcription of these chemokines was induced in the endometrium and intercotiledonary region at the third trimester of pregnancy when stimulated with heat-killed B. abortus or S. Dublin. Therefore, this study demonstrates that some PRRs are expressed in the uterus during pregnancy, which coincides with its ability to respond to stimulation with TLRs ligands as well as heat-killed organisms known to cause abortion in cattle.

Postpartum toll-like receptors and ß-defensin 5 mRNA levels in the endometrium of Holstein cows.

Toll-like receptors (TLRs) and ß-defensins are important components of the innate immune system. This study aimed to evaluate endometrial mRNA levels of TLRs (1/6, 2, 4, and 5) and ß-defensin 5 in Holstein cows by quantitative real time RT-PCR. Uterine biopsies were performed from 6 to 12 h after parturition, and cows were divided into two groups: (i) cows with placental retention and clinical signs of uterine infection until 45 days postpartum (n=10) or (ii) cows with normal puerperium (n=10). All cows had detectable levels of TLRs and ß-defensin 5 mRNAs, but these levels did not differ between groups (P>0.05). Levels of TLR4 mRNA had a positive and significant correlation with the time required for uterine involution in both groups.