Single oral β-cryptoxanthin administration increases its serum concentration and enhances peripheral blood neutrophil function in Holstein cattle

We investigated the effect of oral administration of β-cryptoxanthin (β-CRX) on its serum concentration and peripheral neutrophil functions by the chemiluminescence (CL) response in Holstein cattle. A single oral administration of β-CRX was performed for serum β-CRX concentration (0, 0.05, 0.1, or 0.2 mg/kg body weight [BW]) and for peak CL response of peripheral neutrophils (0.2 mg/kg BW). The serum β-CRX concentration was peaked on 2 days after, similar to peak CL response on 3 days after β-CRX administration. Therefore, a single oral administration of β-CRX (0.2 mg/kg BW) induces higher serum concentration and concurrently enhances bactericidal ability of peripheral neutrophils in Holstein cattle.     

斑马鱼幼鱼嗜中性粒细胞对副溶血弧菌清除的动态变化

副溶血弧菌(Vibrio parahaemolyticus)是人-兽-鱼共患的致病菌,严重威胁食品安全和人类的健康。为了研究宿主嗜中性粒细胞清除细菌感染的动态变化过程,利用红色荧光蛋白标记的副溶血弧菌Vp57~(RFP)菌株感染受精后48 h(hours post fertilization,hpf)的斑马鱼(Danio rerio)幼鱼耳泡,建立局部感染模型,并以大肠杆菌(Escherichia coli)Ec01菌株为对照,系统比较了副溶血弧菌和大肠杆菌的半数致死剂量(median lethal dose,LD_(50))、存活曲线以及和嗜中性粒细胞相互作用的动态过程,RT-qPCR验证炎症相关基因il1b和il10在感染前后的表达量变化。结果显示副溶血弧菌和大肠杆菌感染斑马鱼幼鱼的LD_(50)分别为7.90×10~8 CFU/mL和1.14×10~(11) CFU/mL,与感染大肠杆菌相比,斑马鱼感染副溶血弧菌后招募嗜中性粒细胞的速度更快,且数量更多。RT-qPCR结果发现斑马鱼感染副溶血弧菌后在相同时间点能够激活更高的il1b和il10基因的表达,引起强烈的炎症反应。基于以上研究,我们建立的副溶血弧菌-斑马鱼幼鱼耳泡局部感染模型,为进一步研究嗜中性粒细胞清除副溶血弧菌感染的动态过程,以及深入揭示天然免疫应答机制提供了依据。     

β-羟丁酸抑制脂多糖诱导奶牛中性粒细胞核因子-κB信号通路的激活

高酮血症造成奶牛中性粒细胞先天免疫机能受到抑制,本研究探讨β-羟丁酸(BHBA)是否抑制脂多糖(LPS)诱导的奶牛中性粒细胞核因子-κB(NF-κB)信号通路的激活。分离健康奶牛中性粒细胞,采用LPS(100 ng/mL)和不同浓度(0.5、1.0、2.0和4.0 mmol/L)BHBA作用于中性粒细胞,收集细胞,应用实时荧光定量PCR(qRT-PCR)检测中性粒细胞中白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)和NF-κBp65 mRNA表达水平,Western blot检测NF-κBp65蛋白表达水平,比色法检测核因子-κB抑制物激酶β(IKKβ)激酶活性,酶联免疫吸附试验(ELISA)法检测促炎细胞因子TNF-α、IL-6和IL-1β的分泌量。结果表明：与对照组(不进行BHBA和LPS处理)相比较,LPS组(单独LPS处理)中IL-1β、IL-6、TNF-α和NF-κBp65 mRNA表达水平和NF-κBp65蛋白表达水平极显著增加(P <0.01),IKKβ激酶活性极显著增强(P<0.01),IL-1β和TNF-α的分泌...     

Neutrophil Phagocytosis Following Inoculation of Salmonella choleraesuis into Swine

Neutrophils are an important mediator of host defence, especially in early stages of infection. A major function of neutrophils is the uptake and killing of invading microbes. Little is known about the effect of neutrophil activity on the pathogenesis and development of the carrier state in swine following infection with Salmonella choleraesuis. A human whole-blood microassay using flow cytometry was modified to measure the effect of S. choleraesuis infection in vivo on the rate of ingestion, or rate of uptake, of homologous bacteria by porcine neutrophils. Pigs were inoculated intranasally with 5–8×10⁸ CFU S. choleraesuis and blood was collected in heparinized tubes at –5, 0, 1, 2, 3 and 4 days post inoculation (PI). Heat-killed S. choleraesuis were labelled with fluorescein isothiocyanate and incubated for various times with diluted whole blood. Red blood cells were lysed, external non-phagocytized bacteria were quenched with a commercially available lysing solution, and fluorescence from internalized bacteria labelled with fluorescein isothiocyanate was detected by flow cytometry. The rate of uptake by neutrophils did not increase until 2 days PI and then remained elevated to 4 days PI. The minimal uptake of S. choleraesuis early after exposure to these organisms may provide an opportunity for the pathogen to colonize and/or replicate to levels that facilitate establishment of a carrier state or clinical infection in swine.     

Intramammary infusion of an Enterococcus faecium SF68 preparation promoted the involution of drying off Holstein cows partly related to neutrophil‐associated matrix metalloproteinase 9

A problem for dairy cows following milk stasis is to cope with a high risk of intramammary infection and there is a need to initiate an extensive renewal of secretory modules in mammary glands so that milk production in next lactation may be optimized. We recently reported that ultrasonicated Enterococcus faecium SF68 (SF68) is compatible with cow mammary glands and an enhancer of innate immunity during the immediate post‐milk stasis period. The current study further examines the concomitant effect of ultrasonicated SF68 on mammary tissue remodeling. Four Holstein cows each received intramammary infusions of regular antibiotic dry‐cow formula (positive control) and two different doses of SF68 in different quarters. Analyses of individual quarter secretion samples showed faster neutrophil infiltration, earlier modifications in protein composition, including caseins and lactoferrins, as well as more prompt elevation of the specific unit of 92‐kDa matrix metalloproteinase 9 (MMP9) in SF68‐infused quarters compared to the positive controls. Intramammary infusion of ultrasonicated SF68 seems able to accelerate the regression of mammary synthetic capacity and potentiate the breakdown of glandular extracellular matrix, indicating a more efficient mammary gland involution. Correlation analyses imply that the ability of ultrasonicated SF68 to induce faster neutrophil chemotaxis and the associated MMP9 release is partly responsible.     

Demonstration of inflammatory cell population changes in rat lungs in response to intratracheal instillation of spring wheat dust using lung enzymatic digestion and centrifugal elutriation

The inhalation of grain dust by grain workers is responsible for a large number of pulmonary pathophysiologies. These problems may be acute or chronic and may be mediated by the chronic activation of the immune system. Constant inflammatory states in the lung may eventually lead to tissue damage and respiratory deficit. This study was designed to measure the changes in the relative number of inflammatory cells in peripheral blood, bronchoalveolar spaces, and lung interstitium that occur in response to intratracheally instilled airborne spring wheat dust in rats. It was found that 6h after instillation with dust, neutrophils were present in greater numbers in the blood and bronchoalveolar spaces than in lung interstitium. After 24h, there appeared to be a larger number of neutrophils in the lung interstitium in dust-instilled animals than in saline-instilled controls. These results indicate that intratracheal instillation of grain dust initiates an acute inflammatory reaction, and that there is an initial influx of neutrophils into the air spaces of the lung followed by transit of these cells into the lung interstitium.     

Effects of five oleanolic acid triterpenoid saponins from the rhizome of Anemone raddeana on stimulus-induced superoxide generation, phosphorylation of proteins and translocation of cytosolic compounds to cell membrane in human neutrophils

Five oleanolic acid triterpenoid saponins (OTS-1, 2, 3, 4 and 5) were isolated from the rhizome of Anemone raddeana. The effect of these triterpenoid saponins on stimulus-induced superoxide generation in human neutrophils was assayed by measuring the reduction of ferricytochrome c using a dual-beam spectrophotometer. The phosphorylation of neutrophil proteins, and translocation of p67(phox), p47(phox) and Rac to plasma membrane were investigated using specific monoclonal antibodies. The five oleanolic acid triterpenoid saponins used in this experiment suppressed N-formyl-methionyl-leucyl-phenylalanine (fMLP)-induced superoxide generation in a concentration-dependent manner. OTS-1, 2 and 4 suppressed phorbol 12-myristate 13-acetate (PMA)- and arachidonic acid (AA)-induced superoxide generation in a concentration-dependent manner, but OTS-3 and 5 showed no effect. fMLP- and PMA-induced tyrosyl or serine/threonine phosphorylation, and fMLP-, PMA- and AA-induced translocation of p67(phox), p47(phox) and Rac to plasma membrane were in parallel with the suppression of the stimulus-induced superoxide generation.