RESUMO
Pigeons are natural intermediate host of Neospora caninum (N. caninum). In comparison to ruminants, N. caninum causes milder clinical symptoms and less financial loss to pigeons. Natural infectious rates and high prevalence of N. caninum in pigeons, and death cases of N. caninum-infected pigeons under experimental conditions have been reported, but the detailed pathological characteristics and congenital immunological responses of pigeons-infected with N. caninum remain not well described. In this study, pigeons were infected intraperitoneally with 107 N. caninum tachyzoites. N. caninum in tissues was detected by qPCR. Pathological changes of tissues were examined by hematoxylin-eosin staining. Blood smears were prepared for counting eosinophils changes in blood. Heterophil extracellular traps (HETs) in vivo and in vitro were quantified by Pico Green. N. caninum-induced HETs structures were observed by immunofluorescence staining. The model of pigeons-infected with N. caninum was successfully established. Lung and duodenum were the main target organs of pigeons-infected with N. caninum. N. caninum caused hemorrhage, edema and inflammatory cell infiltration in liver, pulmonary congestion and hemorrhage, organizational destruction in lung, and shorter villi or even disappear in duodenum. N. caninum also increased the number of eosinophils in blood of pigeons. Moreover, N. caninum-induced HETs release in the congenital immunological system of pigeons were first demonstrated, and the HETs structures were consisted of DNA as the skeleton and modified with citH3 and elastase. N. caninum-induced HETs release was related with NADPH oxidase, TLR 2 and 4, ERK1/2 and p38 MAPK signaling pathways, and glycolysis. In summary, it is the first report on the detailed pathological characteristics and congenital immunological responses of pigeons-infected with N. caninum, which may provide theoretical basis for the prevention and control of Neosporosis in pigeons.
Assuntos
Coccidiose , Armadilhas Extracelulares , Neospora , Animais , Coccidiose/veterinária , Columbidae , NeutrófilosRESUMO
Toxoplasma gondii is a zoonotic parasite with a global distribution. Heterophil extracellular traps (HETs) are a novel innate immune mechanism of chickens against pathogens, but whether T. gondii can induce HETs release in chickens has not been reported. The effects of T. gondii on heterophils viability were assessed by using Cell Counting Kit-8. T. gondii-induced HETs were observed and analysed by the immunofluorescence method. T. gondii-induced reactive oxygen species (ROS) was determined by the DCFH-DA method. The mechanisms underlying T. gondii-triggered HETs were investigated by inhibitors and fluorescence microplate reader. T. gondii did not significantly affect heterophils viability at a 1:1 ratio within 1 h. It was demonstrated for the first time that T. gondii could induce HETs release in chicken, and the structure of HETs was comprised of DNA, elastase and citrullinated histone 3 (citH3). T. gondii increased ROS production in a dose-dependent manner. Inhibitors of NADPH oxidase, extracellular signal-regulated kinase 1/2 (ERK1/2 ) and P38 signalling pathways, glycolysis and autophagy significantly decreased the release of T. gondii-induced HETs. Taken together, T. gondii can induce HETs release in chickens, and ROS, NADPH oxidase, ERK1/2 and P38 signalling pathways, glycolysis and autophagy participate in the process of HETs release, which provides new insights into the innate immune mechanism of chickens against T. gondii infection.
Assuntos
Armadilhas Extracelulares , Toxoplasma , Animais , Galinhas , NADPH Oxidases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Autofagia , GlicóliseRESUMO
Toxoplasma gondii (T. gondii) exhibits a significantly high prevalence of infection in goats, leading to adverse consequences such as abortion and stillbirth in ewes, thereby posing a substantial challenge to the goat farming industry. Neutrophil extracellular traps (NETs) have been shown to capture T. gondii in goats; however, the precise mechanisms underlying NET release in goats remain poorly understood. Therefore, the aim of our research was to elucidate the involved mechanism. We assessed the cytotoxicity of T. gondii on neutrophils using CCK-8 assay, visualized the structure of T. gondii-induced goat NETs through immunofluorescence, quantified ROS release during T. gondii-induced NET formation using fluorescence microplate analysis, and employed inhibitors targeting TLR 2, TLR4, NADPH oxidase, ERK1/2, and P38 MAPK signaling pathways as well as glycolysis to dissect the mechanisms underlying T. gondii-induced NET release. Within 1 h, T. gondii did not exhibit significant cytotoxicity towards neutrophils in our findings. The formation of typical NET structures induced by T. gondii involved DNA, citrullinated histone 3 (citH3), and neutrophil elastase (NE). Additionally, T. gondii significantly stimulated the release of NETs in goats. The process was accompanied by the production of reactive oxygen species (ROS) mediated through NADPH oxidase, p38, and ERK1/2 signaling pathways. Inhibition of these pathways resulted in a decrease in NET release. Moreover, inhibition of TLR 2, TLR4, and glycolysis also led to a reduction in T. gondii-induced NET release. Overall, our study demonstrates that T. gondii can induce characteristic NET structures and elucidates the involvement of various mechanisms including TLR2/TLR4 signaling pathway activation, NADPH oxidase activity modulation via ROS production regulation through p38 MAPK and ERK1/2 signaling pathways, and glycolysis regulation during the innate immune response against T. gondii infection in goats.
Assuntos
Armadilhas Extracelulares , Toxoplasma , Animais , Feminino , Ovinos , Sistema de Sinalização das MAP Quinases , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , Espécies Reativas de Oxigênio/metabolismo , Cabras , Neutrófilos , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , NADPH Oxidases/metabolismoRESUMO
Artificial insemination has been a predominant technique employed in goat husbandry for breeding purposes. Subsequent to artificial insemination, sperm can elicit inflammation in the reproductive tract, resulting in substantial the accumulation of neutrophils. Recognized as foreign entities, sperm may become entrapped within neutrophil extracellular traps (NETs) released by neutrophils, thereby exploiting their properties of pathogen elimination. Deoxyribonuclease I (DNase I), which is known for disintegrating NETs and causing loss of function, has been utilized to ameliorate liver and brain damage resulting from NETs, as well as to enhance sperm quality. This study investigated the mechanism of sperm-induced NETs and further explored the impact of DNase I on NETs. Sperm quality was evaluated using optical microscopy, while the structure of NETs was observed through immunofluorescence staining. The formation mechanism of NETs was examined using inhibitors and PicoGreen. The findings revealed that sperm induced the formation of NETs, a process regulated by glycolysis, NADPH oxidase, ERK1/2, and p38 signaling pathways. The composition of NETs encompassed DNA, citrullinated histone H3 (citH3), and elastase (NE). DNase I protects sperm by degrading NETs, thereby concurrently preserving the integrity of plasma membrane and motility of sperm. In summary, the release of sperm-induced NETs leads to its damage, but this detrimental effect is counteracted by DNase I through degradation of NETs. These observations provide novel insights into reproductive immunity in goats.
Assuntos
Armadilhas Extracelulares , Masculino , Animais , Armadilhas Extracelulares/metabolismo , Cabras , Sêmen , Neutrófilos , Espermatozoides , Desoxirribonuclease I/metabolismo , Desoxirribonuclease I/farmacologiaRESUMO
Gliotoxin (GT) belongs to the epipolythiodioxopiperazine (ETP) family, which is considered a crucial virulence determinant among the secondary metabolites produced by Aspergillus fumigatus. The metabolites are commonly found in food and feed, contributing to the invasion and immune escape of Aspergillus fumigatus, thereby posing a significant threat to the health of livestock, poultry, and humans. Heterophil extracellular traps (HETs), a novel form of innate immune defense, have been documented in the chicken's innate immune systems for capturing and eliminating invading microbes. However, the effects and mechanisms of GT on the production of duck HETs in vitro remain unknown. In this study, we first confirmed the presence of HETs in duck innate immune systems and further investigated the molecular mechanism underlying GT-induced HETs release. Our results demonstrate that GT can trigger typical release of HETs in duck. The structures of GT-induced HETs structures were characterized by DNA decoration, citrullinated histones 3, and elastase. Furthermore, NADPH oxidase, glycolysis, ERK1/2 and p38 signaling pathway were found to regulate GT-induced HETs. In summary, our findings reveal that gliotoxin activates HETs release in the early innate immune system of duck while providing new insights into the immunotoxicity of GT towards ducks.
Assuntos
Patos , Gliotoxina , Imunidade Inata , Animais , Imunidade Inata/efeitos dos fármacos , Armadilhas Extracelulares/efeitos dos fármacos , Imunotoxinas/toxicidadeRESUMO
Aspergillus fumigatus causes aspergillosis with high morbidity and mortality in the duck industry. As a vital virulence factor produced by A. fumigatus, gliotoxin (GT) is widely present in food and feed, threatening duck industry and human health. Quercetin is a polyphenol flavonoid compound from natural plants with anti-inflammatory and antioxidant functions. However, the effects of quercetin on ducklings with GT poisoning are unknown. The model of ducklings with GT poisoning was established, and the protective effects and molecular mechanisms of quercetin on ducklings with GT poisoning were investigated. Ducklings were divided into control, GT, and quercetin groups. A model of GT (2.5 mg/kg) poisoning in ducklings was successfully established. Quercetin protected GT-induced liver and kidney functions and alleviated GT-induced alveolar wall thickening in lungs, cell fragmentation, and inflammatory cell infiltration in liver and kidney. Quercetin decreased malondialdehyde (MDA) and increased superoxide dismutase (SOD) and catalase (CAT) after GT treatment. Quercetin significantly reduced GT-induced mRNA expression levels of inflammatory factors. Furthermore, quercetin increased GT-reduced heterophil extracellular traps (HETs) in serum. These results indicated that quercetin protected ducklings against GT poisoning by inhibiting oxidative stress, inflammation and increasing HETs release, which confirms the potential applicability of quercetin in treating GT-induced duckling poisoning.
Assuntos
Armadilhas Extracelulares , Gliotoxina , Animais , Humanos , Quercetina/farmacologia , Patos , Gliotoxina/farmacologia , Estresse Oxidativo , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Antioxidantes/farmacologiaRESUMO
Ketosis is a metabolic disease of dairy cows in the perinatal period, ß-hydroxybutyrate (ß-HB) is the main component of ketosis. High levels of ß-HB can trigger oxidative stress and inflammatory response in dairy cows, leading to decreased milk yield and multiple postpartum diseases. Forsythin (FOR), the major constituent of the herbal medicine Forsythia, has anti-inflammatory, anti-oxidant, and antiviral effects. FOR was demonstrated to have an antioxidant effect on PC12 cells. However, the effects of FOR on ß-HB-stimulated bovine macrophages (BMs) has not been reported. Thus, the aim of the present study was to investigate the effects of FOR on ß-HB-stimulated BMs. Firstly, the CCK8 test confirmed that FOR (50, 100, 200 µg/mL) has no effect on BMs activity, and we selected these concentrations for subsequent experiments. Secondly, through detecting the oxidation indexes ROS, MDA and antioxidant indexes CAT and SOD, we confirmed the antioxidant effect of FOR on BMs. Next, qRT-PCR confirmed that FOR dramatically reduced the mRNA levels of IL-1ß and IL-6. Furthermore, the western blotting confirmed that FOR observably down-regulated ß-HB-stimulated phosphorylation of p38, ERK and Akt and up-regulated expression of Nrf2, and HO-1. Above results suggested that FOR plays antioxidant effects on ß-HB-induced BMs through p38, ERK and PI3K/Akt, Nrf2 and HO-1 signaling pathways. Therefore, we speculated that FOR may be a potential medicine to alleviate ß-HB-induced inflammatory response and provide a preliminary reference for the research and development of FOR.