RESUMEN
Interferon-τ (IFN-τ) participates in the establishment of endometrial receptivity in ruminants. However, the precise mechanisms by which IFN-τ establishes bovine endometrial receptivity remain largely unknown. Interferon regulatory factor 1 (IRF1) is a classical interferon-stimulated gene (ISG) induced by type I interferon, including IFN-τ. Leukemia inhibitory factor receptor (LIFR) is a transmembrane receptor for leukemia inhibitory factor (LIF), which is a key factor in regulating embryo implantation in mammals. This study aimed to investigate the roles of IRF1 and LIFR in the regulation of bovine endometrial receptivity by IFN-τ. In vivo, we found IRF1 and LIFR were upregulated in the bovine endometrial luminal epithelium on Day 18 of pregnancy compared to Day 18 of the estrous cycle. In vitro, IFN-τ could upregulate IRF1, LIFR, and endometrial receptivity markers (LIF, HOXA10, ITGAV, and ITGB3) expression, downregulate E-cadherin expression and reduce the quantity of microvilli of bovine endometrial epithelial cells (bEECs). Overexpression of IRF1 had similar effects to IFN-τ on endometrial receptivity, and interference of LIFR could block these effects, suggesting the positive effects of IRF1 on endometrial receptivity were mediated by LIFR. Dual luciferase reporter assay verified that IRF1 could transactivate LIFR transcription by binding to its promoter. In conclusion, IFN-τ can induce IRF1 expression in bovine endometrial epithelial cells, and IRF1 upregulates LIFR expression by binding to LIFR promoter, contributing to the enhancement of bovine endometrial receptivity.
Asunto(s)
Implantación del Embrión , Endometrio , Factor 1 Regulador del Interferón , Interferón Tipo I , Animales , Femenino , Bovinos , Endometrio/metabolismo , Endometrio/inmunología , Factor 1 Regulador del Interferón/metabolismo , Factor 1 Regulador del Interferón/genética , Implantación del Embrión/inmunología , Interferón Tipo I/metabolismo , Embarazo , Receptores OSM-LIF/metabolismo , Proteínas Gestacionales/metabolismo , Proteínas Gestacionales/genética , Activación Transcripcional , Células Cultivadas , Células Epiteliales/metabolismo , Células Epiteliales/inmunologíaRESUMEN
Endometritis is a severe postpartum inflammatory disease that puts cows' reproductive health at risk and causes the dairy industry to suffer significant financial losses. The present study aimed to investigate the regulatory role of miR26a in LPSinduced bovine endometrial epithelial cells (bEECs) and the implication for endometritis. Here, we found inflammatory cell infiltration and destruction of endometrial structure in cow uterus, and dramatic increase in myeloperoxidase (MPO) activity and upregulation of pro-inflammatory cytokines (IL-1ß, TNF-α, and IL-6) in endometritis. Meanwhile, miR-26a was down-regulated, but MAP3K8 was increased in the uterine tissue of endometritis. Similarly, the expression of miR-26a was significantly decreased in LPS-stimulated bEECs, while MAP3K8 was risen. In addition, we further verified that MAP3K8 was a target of miR-26a by dual-luciferase reporter assay. Under LPS stress, over-expressing miR-26a markedly decreased MAP3K8 expression levels, along with the reduced expression of inflammatory factors, such as IL-1ß, TNF-α and IL-6, whereas this effect was countered by the inhibition of miR-26a. Furthermore, we demonstrated that miR-26a overexpression prevented the MAPK pathway from being activated by targeting MAP3K8. Then we carried out experiments in LPS-stimulated mice uterus to expound that MAP3K8 was essential in endometritis development, which further confirmed the reliability of the above results. In conclusion, overexpression of miR-26a effectively inhibited the expression of MAP3K8 in LPS-induced bEECs and thereby partially suppressed the activation of MAPK signaling pathway. miR-26a and MAP3K8 may be a promising biomarker and therapeutic target for dairy cow endometritis.
Asunto(s)
Endometritis , Quinasas Quinasa Quinasa PAM , MicroARNs , Animales , Bovinos , Femenino , Ratones , Endometritis/tratamiento farmacológico , Endometritis/veterinaria , Células Epiteliales/metabolismo , Interleucina-6/metabolismo , Lipopolisacáridos , Quinasas Quinasa Quinasa PAM/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Reproducibilidad de los Resultados , Transducción de Señal , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Bovine endometritis is a serious hazard to husbandry, so it is necessary to know the mechanism of endometritis. In past research, we found microRNAs (miRNAs) might be regulators in inflammation, including miR-196b, but the mechanism of miR-196b in bovine endometritis was unknown. Therefore, we tended to find out what role miR-196b would play in bovine endometritis. As a result, we found miR-196b up-regulated in the endometritis tissue and the high concentration lipopolysaccharide (LPS)-stimulated bovine endometrial epithelial (BEND) cell line, but down-regulated in the low concentration. And, over-expression of miR-196b inhibited the expressions of some inflammatory factors, such as tumor necrosis factor-alpha (TNF-α), interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and neuroblastoma RAS (NRAS)/nuclear factor (NF)-κB pathway proteins. Furthermore, the dual-luciferase reporter assay and NRAS knockdown confirmed that miR-196b inhibited activation of the downstream pathway by directly targeting NRAS. In conclusion, we provide evidence that miR-196b alleviates LPS-induced inflammatory injury by targeting NRAS.
Asunto(s)
Endometritis , MicroARNs , Neuroblastoma , Animales , Bovinos , Células Epiteliales/metabolismo , Femenino , GTP Fosfohidrolasas/metabolismo , Humanos , Interleucina-6/metabolismo , Lipopolisacáridos/farmacología , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , FN-kappa B/metabolismo , Neuroblastoma/metabolismo , Neuroblastoma/patología , Transducción de Señal/genéticaRESUMEN
AIMS: Patients with acute kidney injury (AKI) have higher mortality, and sepsis is among its main causes. MicroRNAs (miRNAs) are essential for regulating kidney function and could have curative potential. This study explored the possibility to treat AKI with miR-125a-5p and reveal the possible mechanism. MATERIALS AND METHODS: LPS-induced mouse model and LPS-induced RAW264.7 cell model of AKI were established and treated with miR-125a-5p mimics or inhibitors. Serum creatinine and blood urea were measured to evaluate kidney function. The pathological changes of kidney tissues were detected by H&E and PAS staining technique, and the infiltration of macrophages were observed by immunohistochemistry. RAW264.7 cell viability, TRAF6 and cytokines expressions under LPS stimulation were measured. The role and therapeutic potential of miR-125a-5p were verified in vivo and in vitro after given miR-125a-5p mimics or inhibitors. KEY FINDINGS: LPS-induced mice had increasing serum creatinine and urea, and evident pathological changes, including severe tubular dilatation and macrophages infiltration. TRAF6 expression in the kidney was significantly higher, while miR-125a-5p expression was suppressed. MiR-125a-5p targeted TRAF6, and its overexpression deactivated NF-κB signaling pathway, reducing downstream TNF-α, IL-1ß and IL-6 expressions. MiR-125a-5p mimics rescued LPS-induced kidney damage and suppressed pro-inflammatory cytokines expression through inhibiting TRAF6/NF-κB axis. SIGNIFICANCE: We highlighted that miR-125a-5p could inhibit LPS-induced acute inflammation in the kidney through targeting TRAF6/NF-κB axis. These results might contribute to the development of molecular therapy in AKI.
Asunto(s)
Lesión Renal Aguda/patología , Regulación de la Expresión Génica , Lipopolisacáridos/toxicidad , MicroARNs/genética , Factor 6 Asociado a Receptor de TNF/antagonistas & inhibidores , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/genética , Lesión Renal Aguda/metabolismo , Animales , Masculino , Ratones , Ratones Endogámicos BALB C , Células RAW 264.7 , Factor 6 Asociado a Receptor de TNF/genética , Factor 6 Asociado a Receptor de TNF/metabolismoRESUMEN
Solving the reproductive barriers of dairy cows has become one of the most critical factors determining the dairy industry's development. Clinically, inflammation disease like endometritis is the most crucial cause in reducing dairy production's financial viability. MiR-193 family can induce cell apoptosis and differentiation has been reported in various diseases. LGR4 plays a vital role in reproductive system development and immune system regulation, and it is closely related to animal reproductive function and cytokine regulation. In this study, we observed a negative relationship between miR-193a-3p and LGR4 expression level in both inflammatory tissues and cells. The expression level of miR-193a-3p and LGR4 in bovine endometrial epithelial cells (BENDs) is regulated by lipopolysaccharide (LPS) stimulation time and dose-dependent. Subsequently, miR-193a-3p mimics and inhibitors were used to explore its functions in the inflammation response process, finding that overexpression of miR-193a-3p markedly increases the expression level of pro-inflammatory cytokines induced by LPS, such as IL-1ß, IL-6 and TNF-α, while the group in which transfected inhibitor is on the contrary. Of note, immunofluorescence and western blot results showed that miR-193a-3p enhanced LPS-induced NF-κB p65 phosphorylation through targeting LGR4, whereas inhibiting miR-193a-3p could suppress the activation of NF-κB pathway significantly. In conclusion, our study firstly reported the mechanism by which miR-193a-3p targets LGR4 to elevate the inflammatory response in bovine endometrium injury, thereby implying that knockdown miR-193a-3p may lay the theoretical and practical basis for drug development of alleviating endometritis in dairy cows.
Asunto(s)
Endometritis/veterinaria , Endometrio/inmunología , MicroARNs/metabolismo , Receptores Acoplados a Proteínas G/genética , Animales , Bovinos , Línea Celular , Endometritis/genética , Endometritis/inmunología , Endometrio/patología , Células Epiteliales/inmunología , Células Epiteliales/patología , Femenino , Regulación de la Expresión Génica/inmunología , Técnicas de Silenciamiento del Gen , Lipopolisacáridos/inmunología , MicroARNs/genéticaRESUMEN
Endometritis is a common postpartum inflammatory disease that endangers the reproductive health of humans and animals. Emerging evidence shows that microRNA is a new type of therapeutic molecule that plays a vital role in many diseases; however, its mechanism of action in lipopolysaccharide (LPS)-induced endometritis is still unclear. This study aims to investigate the regulatory role of miR-211 in the innate immune response involved in endometritis, and to evaluate its potential therapeutic value. Here, we found that the expression of miR-211 in bovine endometrial epithelial cells (bEECs) stimulated by lipopolysaccharide (LPS) was significantly reduced. Importantly, overexpression of miR-211 can significantly reduce the production of pro-inflammatory cytokines (IL-1ß , IL-6 and TNF-α). In addition, we proved that TAB1 is the target gene of miR-211. MiR-211 inhibits TAB1 protein expression by binding to the 3'-UTR of TAB1 mRNA. Subsequently, we verified that the overexpression of miR-211 inhibited the activation of NF-κB p65 by targeting the TAB1-mediated pathway. Therefore, miR-211 has anti-inflammatory effects and mediates the negative regulation of the NF-κB signaling pathway in LPS-induced endometritis by targeting TAB1.