Downregulation of XBP1s aggravates lipopolysaccharide-induced inflammation by promoting NF-κB and NLRP3 pathways' activation in goat endometrial epithelial cells.

After delivery, bacterial contamination and uterine tissue degeneration in animals can lead to the development of uterine diseases, such as endometritis, accompanied by endoplasmic reticulum stress (ERS). Increasing evidence suggests that spliced X-box binding protein 1 (XBP1s), a critical component of ERS, is involved in several pathological processes in various organisms. However, the specific molecular mechanisms by which XBP1s mediates the inflammatory response in goat endometrial epithelial cells (gEECs) remain largely unknown. In the present study, XBP1s protein was induced into the nucleus in the lipopolysaccharide (LPS, 5 µg/mL)-induced inflammatory response of gEECs. Lipopolysaccharide-induced expression and nucleation of XBP1s were reduced by the inhibition of Toll-like receptor 4 (TLR4) using TAK-242 (1 µM; a TLR4 inhibitor). Expression and nucleation of XBP1s were similarly reduced when the activity of inositol-requiring enzyme 1α (IRE1α) was inhibited using 4µ8C (10 µM; an IRE1α inhibitor). In addition, inhibition of IRE1a increased IL-1ß, TNF-α, and IL-8 levels and secretion of IL-6 induced by LPS. Notably, phosphorylation of nuclear factor kappa-B (NF-κB) P65 protein and expression of NOD-like receptor thermal protein domain associated protein 3 (NLRP3) were similarly increased. Furthermore, knockdown of XBP1s in gEECs consistently promoted NF-κB P65 protein phosphorylation, NLRP3 protein expression, and inflammatory cytokine secretion. In summary, the current results suggest that in the LPS-induced inflammatory response in gEECs, LPS generates intracellular signaling cascades in gEECs via TLR4, which may promote XBP1s protein expression and nucleation by activating IRE1a. However, downregulation of XBP1s expression exacerbates inflammation by promoting activation of the NF-κB and NLRP3 inflammatory vesicle pathways. These results will potentially contribute to the treatment and prevention of endometritis in ruminants.

UFM1 inhibits the activation of the pyroptosis in LPS-induced goat endometritis.

Infertility, abortion, and stillbirth caused by endometritis are the main factors affecting fertility in ruminants. Lipopolysaccharide (LPS)-mediated inflammation is the main cause of endometritis. Toll-like receptor 4 (TLR4) pathway and pyroptosis play an important role in the inflammation, but the underlying mechanism is still unclear. Previous studies have reported that UFMylation, a ubiquitin-like post-translational modifier, plays an important regulatory role in inflammation via the TLR4 pathway; however, its relationship with pyroptosis is still unclear. Our result showed that LPS induced inflammation by activating the TLR4 pathway and pyroptosis in goat endometrial epithelial cells (gEECs). We also found that TAK-242,a specific inhibitor of the TLR4 pathway, inhibited the pyroptosis pathway. In addition, with an increased LPS treatment time, ubiquitin-folding modifier factor 1 (UFM1) conjugated proteins were highly expressed in gEECs. Moreover, overexpression of UFM1 inhibited LPS-induced activation of the TLR4 pathway and pyroptosis, whereas si-UFM1 produced contrasting results. After inhibiting the TLR4 pathway, si-UFM1 could not upregulate the expression of nod-like receptor family protein 3 (NLRP3), cleavage caspase-1, or cleavage gasdermin D (GSDMD). In conclusion, these results suggest that UFM1 inhibits pyroptosis activation in LPS-induced gEECs through the TLR4 pathway.

Molecular detection of Hepatozoon canis in dogs and ticks in Shaanxi province, China.

Hepatozoon canis, transmitted by Rhipicephalus sanguineus, is a tick-borne pathogen and causes canine hepatozoonosis. Until now, only limited previous studies were conducted on the molecular detection and characterization of Hepatozoon sp. in dogs in China. Blood samples were collected from 93 sick dogs that were clinically diagnosed as babesiosis but tested negative for Babesia, and 103 apparently healthy dogs, as well as their infesting ticks in Xi'an and Hanzhong cities, Shaanxi province of China. PCR amplifying partial 18S rRNA gene was used to detect the DNA of Hepatozoon sp. Genetic and phylogenetic analysis were performed to determine the Hepatozoon species. Our results demonstrated that H. canis was identified from the sick dogs and the infested ticks in Hanzhong, with no significant differences of prevalence between both genders and ages. No positive blood or tick samples were found in Xi'an. Moreover, all the 18S rRNA gene sequences recovered from both dogs and the infested ticks showed a high genetic similarity with each other, and also presented a close relationship with other known sequences in and outside China. In conclusion, H. canis was identified in babesiosis-suspected dogs and ticks infesting them in Shaanxi, China, although the association between clinical signs and H. canis need further study.