大肠杆菌K88攻毒对幼龄仔猪肠道离子载体和水通道蛋白表达的影响

畜牧编辑

　　大肠杆菌K88攻毒对幼龄仔猪肠道离子载体和水通道蛋白表达的影响
　　C. Zhu, J. L. Ye, J. Yang, K. M. Yang, Z.Chen, R. Liang, X. J. Wu, L. Wang and Z. Y. Jiang

　　本试验研究大肠杆菌K88攻毒在体内或者体外实验中是否会对水和离子转运载体基因表达产生影响。

　　选用36头公猪（4日龄）随机分为对照组和攻毒组，每个处理6个重复，每个重复3头猪。所有猪只饲喂相同日粮17天。在第15天，攻毒组猪只接受K88攻毒（血清型O149:K91:K88ac，剂量为1×108 cfu/头），而对照组接受同剂量的磷酸盐缓冲液处理。攻毒72小时后（第18天），每个重复挑选一头猪屠宰，收集空肠、回肠和结肠样品。

　　攻毒组回肠和结肠中囊性纤维化跨膜传导调节因子(CFTR)的蛋白丰度和mRNA的表达显著提高（P＜0.05）。此外，攻毒还提高了回肠和结肠中钠钾氯共转运蛋白1（NKCC1）的mRNA表达（P＜0.05），而在空肠中还额外提高了钠钾氯共转运蛋白1蛋白的表达（P＜0.05）。除此之外还通过猪小肠上皮细胞系（IPEC-J2）研究K88对水通道蛋白和离子转运载体的影响及可能的作用机理。在6孔平板上培养上皮细胞（每孔1.17×106个细胞），用50:1的K88攻毒，处理3小时。攻毒显著降低了上皮细胞中水通道蛋白3（AQP3）、AQP11和钠氢交换蛋白3（NHE3）mRNA的表达（P＜0.05）。蛋白质免疫印迹法（western blotting）的进一步分析表明K88攻毒降低了上皮细胞AQP3、AQP9、AQP11蛋白的表达（P＜0.05）。此外，攻毒还降低了蛋白激酶A和环磷酸腺苷效应元件结合蛋白（CREB）的磷酸化水平（P＜0.05）。

　　试验结果表明K88攻毒可能通过调节环磷酸腺苷-蛋白激酶A信号通路导致肠道离子转运载体和水通道蛋白的表达差异。本研究结果一定程度上从另一方面揭示了幼龄仔猪腹泻控制中离子平衡的重要性。

　　Differential expression of intestinal ion transporters and water channel aquaporins in young piglets challenged with enterotoxigenic Escherichia coli K88

　　C. Zhu, J. L. Ye, J. Yang, K. M. Yang, Z. Chen, R. Liang, X. J. Wu, L. Wang and Z. Y. Jiang

　　The study was to determine whether the expression of genes involved in intestinal water and ion transport would be affected by enterotoxigenic Escherichia coli (ETEC) K88 both in vitro and invivo. First, 36 male piglets (4 d old) were randomly allotted to either the control or the ETEC K88 group. Each group had 6 replicates with 3 piglets per replicate. All piglets were fed with the same diets for 17 d. On d 15, piglets in the ETEC K88 group were challenged with ETEC K88 (serotype O149:K91:K88ac) at 1 × 108 cfu per pig, whereas those in the control group received the same volume of sterile PBS. After being challenged with ETEC K88 for 72 h (d 18), 1 piglet from each replicate was selected for slaughter to collect samples from the jejunum, ileum, and colon. The mRNA expression and protein abundance of cystic fibrosis transmembrane conductance regulator (CFTR) in the ileum and colon were increased compared with that in the control group (P < 0.05). Furthermore, the mRNA expression of Na-K-Cl cotransporter 1 (NKCC1) in the ileum and colon was increased by ETEC K88 challenge (P < 0.05), whereas in the jejunum, both its mRNA and protein expression were increased by ETEC K88 treatment (P < 0.05). Additionally, an established porcine intestinal epithelial cell line (IPEC-J2) was used to investigate the effect and possible mechanism of ETEC K88 on expression of water channel aquaporins (AQP) and ion transporters. Cells (1.17 × 106 per well) were grown in 6-well plates and treated with ETEC K88 at a multiplicity of infection of 50:1 for 3 h. The mRNA expression of AQP3, AQP11, and Na+/H+ exchanger 3 (NHE3) in IPEC-J2 cells was reduced after ETEC K88 treatment (P < 0.05). Further analyses using western blotting also demonstrated that ETEC K88 decreased the protein expression of AQP3, AQP9, and AQP11 in IPEC-J2 cells (P < 0.05). Moreover, the phosphorylation levels of protein kinase A (PKA) and cyclic adenosine monophosphate (cAMP)-response element binding protein (CREB) were decreased by ETEC K88 challenge (P < 0.05). The results indicate that ETEC K88 challenge induced differential expression of intestinal ion transporters and AQP in young piglets, probably by regulation of the cAMP–PKA signaling pathway. This study might provide new insights about the importance of fluid homeostasis in control of ETEC-induced diarrhea in young piglets。

山中的漫游者

这个实验证明了大肠杆菌是腹泻的主要因素，