Deng Zhaoju, Shahid Muhammad, Zhang Limei, Gao Jian, Gu Xiaolong, Zhang Shiyao, Zou Jiaqi, Fanning Séamus, Han Bo
Department of Clinical Veterinary Medicine, College of Veterinary Medicine, China Agricultural University, Yuan Ming Yuan West Road No. 2, Haidian District, Beijing, 100193, People's Republic of China.
UCD-Centre for Food Safety, School of Public Health, Physiotherapy and Population Science, University College Dublin, Belfield, Dublin 4, Ireland.
Mycopathologia. 2016 Dec;181(11-12):823-832. doi: 10.1007/s11046-016-0053-0. Epub 2016 Aug 24.
Prototheca zopfii is an important bovine mastitis pathogen, which could result in severe mammary infection. However, the innate immune response in bovine mastitis associated with P. zopfii was not clear. Therefore, the aim of this study is to investigate in vitro innate immune responses implicated by P. zopfii. Bovine mammary epithelial cells (bMECs) were infected with 5.0 × 10 cells/ml P. zopfii genotypes I and II independently, and the mRNA expression of TLR-2, TLR-4, TNF-α, IL-1β, IL-8, NOD-1, NOD-2 and β-defensin-5 was determined by real-time polymerase chain reaction (RT-PCR) over a time course of 1, 3, 6, 12 and 24 h. The detection of the NF-κB p65 protein in nucleus and cytoplasm of infected bMECs over the same time course was evaluated. Results showed that P. zopfii genotype II has ability to up-regulate the expression of TLR-2, TLR-4, TNF-α, IL-1β, IL-8, NOD-1, NOD-2 and β-defensin-5 'more strongly than genotype I. Western blot results showed that when bMECs were challenged by P. zopfii genotype II, the expression of NF-κB p65 protein in the nucleus was up-regulated, while in cytoplasm it appeared to be repressed, which indicated that bMECs partly regulate the innate immune responses and inflammation by the NF-κB signaling pathway while being infected by P. zopfii genotype II. It was concluded that adhesion of genotype II was stronger than genotype I, and therefore the genotype II regulatory ability is more robust than that of the genotype I, which causes inflammation of bovine mammary tissue.
威克海姆原藻是一种重要的牛乳腺炎病原体，可导致严重的乳腺感染。然而，与威克海姆原藻相关的牛乳腺炎中的固有免疫反应尚不清楚。因此，本研究的目的是调查威克海姆原藻引发的体外固有免疫反应。将牛乳腺上皮细胞（bMECs）分别用5.0×10个细胞/毫升的威克海姆原藻基因型I和II独立感染，并通过实时聚合酶链反应（RT-PCR）在1、3、6、12和24小时的时间进程中测定TLR-2、TLR-4、TNF-α、IL-1β、IL-8、NOD-1、NOD-2和β-防御素-5的mRNA表达。评估了在相同时间进程中感染的bMECs细胞核和细胞质中NF-κB p65蛋白的检测情况。结果表明，威克海姆原藻基因型II比基因型I更能强烈上调TLR-2、TLR-4、TNF-α、IL-1β、IL-8、NOD-1、NOD-2和β-防御素-5的表达。蛋白质印迹结果表明，当bMECs受到威克海姆原藻基因型II攻击时，细胞核中NF-κB p65蛋白的表达上调，而细胞质中的表达似乎受到抑制，这表明bMECs在被威克海姆原藻基因型II感染时通过NF-κB信号通路部分调节固有免疫反应和炎症。得出结论：基因型II的黏附力比基因型I更强，因此基因型II的调节能力比基因型I更强，这会导致牛乳腺组织发炎。
