Centre for Cellular and Molecular Biology, Hyderabad, India.
University Medical Centre, Johannes Gutenberg University, Mainz, Germany.
Transgenic Res. 2019 Dec;28(5-6):573-587. doi: 10.1007/s11248-019-00174-x. Epub 2019 Oct 10.
EchAMP, the tenth most abundant transcript expressed in the mammary gland of echidna, has in vitro broad-spectrum antibacterial effects. However, the effects of EchAMP on mastitis, a condition where inflammation is triggered following mammary gland infection, has not been investigated. To investigate the impact of EchAMP against mastitis, EchAMP transgenic mice were generated. In antibacterial assays, the whey fractions of milk from transgenic mice significantly reduced growth of Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosa compared with whey fractions from wildtype mice. Furthermore, a mastitis model created by infecting mammary gland with these four bacterial strains displayed a significant reduction in bacterial load in transgenic mice injected with S. aureus and B. subtilis. On further confirmation, histomorphologic analysis showed absence of necrosis and cell infiltration in the mammary glands of transgenic mice. To understand the role of EchAMP against inflammation, we employed an LPS-injected mastitis mouse model. LPS is known to induce phopshorylation of NF-κB and MAPK pathways, which in turn activate downstream proinflammatory signaling mediators, to promote inflammation. In LPS-treated EchAMP transgenic mice, phosphorylation levels of NF-κB, p38 and ERK1/2 were significantly downregulated. Furthermore, in mammary gland of transgenic mice, there was a significant downregulation of mRNA levels of proinflammatory cytokines, namely TNF-α, IL-6 and IL-1β. Taken together, these data suggest that EchAMP has an antiinflammatory response and is effective against S. aureus and B. subtilis. We suggest that EchAMP may be a potential prophylactic protein against mastitis in dairy animals by expressing this gene in their mammary gland.
棘皮动物乳腺中表达丰度第十的 EchAMP 在体外具有广谱抗菌作用。然而，EchAMP 对乳腺炎（乳腺感染引发炎症的一种疾病）的影响尚未得到研究。为了研究 EchAMP 对乳腺炎的影响，我们生成了 EchAMP 转基因小鼠。在抗菌测定中，与野生型小鼠的乳清部分相比，转基因小鼠的乳清部分显著降低了金黄色葡萄球菌、枯草芽孢杆菌、大肠杆菌和铜绿假单胞菌的生长。此外，用这四种细菌感染乳腺创建的乳腺炎模型显示，用金黄色葡萄球菌和枯草芽孢杆菌注射的转基因小鼠的细菌负荷显著降低。进一步确认后，组织形态学分析显示转基因小鼠的乳腺中没有坏死和细胞浸润。为了了解 EchAMP 对抗炎症的作用，我们使用了 LPS 注射乳腺炎小鼠模型。LPS 已知可诱导 NF-κB 和 MAPK 通路的磷酸化，进而激活下游的促炎信号转导介质，以促进炎症。在 LPS 处理的 EchAMP 转基因小鼠中，NF-κB、p38 和 ERK1/2 的磷酸化水平显著下调。此外，在转基因小鼠的乳腺中，促炎细胞因子 TNF-α、IL-6 和 IL-1β 的 mRNA 水平也显著下调。总之，这些数据表明 EchAMP 具有抗炎反应，对金黄色葡萄球菌和枯草芽孢杆菌有效。我们建议，通过在乳腺中表达该基因，EchAMP 可能成为预防奶牛乳腺炎的潜在预防性蛋白。
