Sperm enter glands of preovulatory bovine endometrial explants and initiate inflammation.

We previously reported that sperm binding to cultured monolayers of bovine uterine epithelial cells induces an acute inflammatory response involving the Toll-like receptor (TLR2) signaling pathway. This response serves to clear the uterus of sperm and thereby prepares the endometrium for implantation. The endometrium is lined by surface epithelial cells; however, epithelial cells also line uterine glands. To investigate the source of the immune response, we used an explant model. Explants of bovine endometrium were incubated with bull sperm illuminated by JC1 fluorescent labeling in their mitochondria. The sperm glided over the surface epithelium until they encountered and entered uterine glands, where they remained. Scanning electron microscopy of explants revealed polymorphonuclear neutrophils (PMNs) in uterine glands along with sperm. In the absence of sperm, PMNs were not seen in glands. The incubation of sperm with explants resulted in an acute inflammatory response, seen as the upregulation of mRNA expression of IL8, TNFA, IL1B, PGES and TLR2 in whole explants, as well as increased TNFA protein expression in uterine glands. TLR1/2 antagonist reduced sperm numbers in the glands and inhibited the increase of TNFA. Our observations suggest that uterine glands serve as a site where sperm interact with the uterine epithelium to trigger the innate immune response to clear excess sperm from the uterus.

A proinflammatory response of bovine endometrial epithelial cells to active sperm in vitro.

In the cow, cryopreserved semen is inseminated into the uterus, and most of sperm are removed by backflow and phagocytes. Nevertheless, the mechanism responsible for sperm phagocytosis is unclear. Here, we used cultured bovine uterine epithelial cells (BUECs) to investigate the uterine response to sperm and the mechanism that activates polymorphonuclear neutrophils (PMNs). BUEC monolayers were co-cultured with different numbers of washed sperm obtained from cryopreserved semen (104 , 105 , and 106 sperm/ml) for 3 hr. Sperm dose-dependently up-regulated IL8 (Interleukin 8). Sperm at 106 /ml increased mRNA expression of TNFA (Tumor necrosis factor alpha), IL1B (Interleukin 1B), NFKB2 (Nuclear factor kappa B2), and C3 (Complement factor 3), as well as PGES (Prostaglandin E synthase) expression and PGE2 release. Live sperm, but not dead sperm, attached to BUECs, and dead sperm did not induce an acute inflammatory response. Time-dependent effects were evaluated by co-culture of 106 /ml washed sperm with BUECs for 0, 1, 3, and 6 hr. The number of detached sperm increased gradually toward 6 hr. Maximum mRNA expression of IL8, TNFA, IL1B, and NFKB2 was induced at 3 hr, while C3 continued to increase toward 6 hr. Sperm did not stimulate mRNA expression of anti-inflammatory cytokines TGFB1 (Transforming growth factor beta 1) or IL10 (Interleukin 10). Medium conditioned by sperm co-incubated with BUECs stimulated PMNs phagocytosis of sperm in vitro. Fresh media supplemented with low levels of IL1B, TNFA, and PGE2 up-regulated sperm phagocytosis by PMNs as well. In conclusion, our findings strongly suggest that the active sperm attach to BUECs and trigger uterine local innate immunity with induction of a pro-inflammatory response that enhances sperm phagocytosis by PMNs.

Involvement of IL-23 in enteropathic arthritis patients with inflammatory bowel disease: preliminary results.

The role of interleukin (IL)-23 in the pathogenesis of inflammatory bowel disease (IBD) remains unclear. The aim of this work was to study the serum level of IL-23 in IBD with and without arthritis and determine its relation to the subsets and clinical features of the disease. Thirty-seven patients with IBD including 11 with arthritis were included in the study with a mean age of 30.86 ± 4.66 years. Twenty healthy subjects served as control. Seronegative spondyloarthropathy was present in 11 (29.73 %) of the IBD patients; Crohn's disease (CD) was present in 23 and 14 had ulcerative colitis (UC). Serum level of IL-23 was measured in all patients and control by ELISA. IL-23 was significantly higher in IBD patients (46.24 ± 27.19 pg/ml) compared to control (24.1 ± 2.31 pg/ml) (p < 0.0001) being higher in CD patients (52.57 ± 32.78 pg/ml) compared to those with UC (35.86 ± 6.41 pg/ml) (p = 0.026). Furthermore, it was significantly higher in those with peripheral and/or axial arthritis (67.73 ± 40.85 pg/ml) compared to patients without (37.15 ± 10.37 pg/ml) (p = 0.03). There was a tendency to a higher level in males (49.15 ± 30.97 pg/ml) compared to females (38.4 ± 9.54 pg/ml). Serum IL-23 is increased in IBD especially those with CD associated with arthritis and sacroiliitis. The IL-23 could be added to the biomarkers of development of arthritis in IBD patients. These results also confirm the findings of previous studies on the critical role played by IL-23 in the pathogenesis of IBD making it an important new therapeutic target for these patients.