Riollet C, Rainard P, Poutrel B
Laboratoire de Pathologie Infectieuse et Immunologie, INRA, Centre de Tours, Nouzilly, France.
Inflamm Res. 2000 Sep;49(9):486-96. doi: 10.1007/s000110050621.
To examine changes in inflammatory mediators, lymphocyte subpopulations and neutrophil activation that occur during an immune-mediated recruitment of neutrophils in the mammary gland.
11 clinically healthy cows.
5 cows received 2 subcutaneous injections of 30 microg of alpha-toxin of Staphylococcus aureus, two months apart. Three months after the last immunization, 5 microg of alpha-toxin were injected, via the teat end, in one randomly selected quarter of the 5 immunized cows and of the 6 unimmunized cows (control group).
Blood and milk samples were collected at several times during 4 days post-challenge. Blood and milk cells were purified to be stained with specific mAbs and analysed by flow cytometry, or to be used for cytokine RT-PCR. Bovine serum albumin, haptoglobin, cytokines and C5a were also analysed in milk or plasma samples using radial immunodiffusion assay or ELISA.
Large amounts of cells (> 1 million/ml of milk) were recruited in the quarters of the immunized cows, whereas no recruitment occurred in the control group. In blood of immunized animals, haptoglobin was present and expression of surface adhesion molecules on neutrophils was modified whereas no change was observed concerning the lymphocyte subpopulations. On milk-derived neutrophils, the expression of CD11b and CD18 was upregulated compared to blood, in contrast to CD62L that was downregulated. The CD8+ cells were recruited as soon as 12 h post-challenge, in contrast to the CD4+ cells, 96 h post-challenge. No IL-1beta, TNF-alpha, IL-8 and C5a were detected using ELISA. mRNA of IL-1alpha, IL-1beta, IL-6, TNF-alpha, IL-8 and IL-12 were found in almost all the samples.
The immunization triggered an early and massive neutrophil recruitment from the blood into the milk compartment as well as the recruitment of a cytotoxic/suppressor lyphocyte population during the early acute phase response. These results could help to devise new vaccinal strategies to fight against staphylococcal mastitis.
研究乳腺免疫介导的中性粒细胞募集过程中炎症介质、淋巴细胞亚群和中性粒细胞活化的变化。
11头临床健康奶牛。
5头奶牛皮下注射2次，每次30微克金黄色葡萄球菌α毒素，间隔两个月。最后一次免疫后三个月，在5头免疫奶牛和6头未免疫奶牛（对照组）的一个随机选择的乳区经乳头端注射5微克α毒素。
攻毒后4天内多次采集血液和乳汁样本。血液和乳汁细胞经纯化后用特异性单克隆抗体染色，通过流式细胞术分析，或用于细胞因子逆转录聚合酶链反应。牛奶或血浆样本中的牛血清白蛋白、触珠蛋白、细胞因子和C5a也使用放射免疫扩散试验或酶联免疫吸附测定进行分析。
免疫奶牛的乳区募集了大量细胞（>100万个/毫升乳汁），而对照组未发生募集。在免疫动物的血液中，存在触珠蛋白，中性粒细胞表面粘附分子的表达发生改变，而淋巴细胞亚群未观察到变化。与血液相比，乳汁来源的中性粒细胞上CD11b和CD18的表达上调，而CD62L下调。与CD4+细胞在攻毒后96小时被募集不同，CD8+细胞在攻毒后12小时即被募集。使用酶联免疫吸附测定未检测到白细胞介素-1β、肿瘤坏死因子-α、白细胞介素-8和C5a。几乎所有样本中都发现了白细胞介素-1α、白细胞介素-1β、白细胞介素-6、肿瘤坏死因子-α、白细胞介素-8和白细胞介素-12的信使核糖核酸。
免疫引发了早期大量中性粒细胞从血液募集到乳腺区，以及在早期急性期反应中细胞毒性/抑制性淋巴细胞群体的募集。这些结果有助于设计新的疫苗策略来对抗葡萄球菌性乳腺炎。
