猪德尔塔冠状病毒感染对初生仔猪小肠杯状细胞数量及Hes1和MUC2表达的影响

doi:10.11843/j.issn.0366-6964.2021.03.021

摘要/Abstract

摘要： 旨在探讨猪德尔塔冠状病毒（PDCoV）感染仔猪对小肠杯状细胞（goblet cell，GC）的影响，将6头未吃初乳的初生仔猪静养2 d后随机分为感染组（n=3）和对照组（n=3），感染组仔猪口服接种5 mL 1×10⁵ TCID₅₀·mL^-1 PDCoV-CHN-HG-2017毒株，对照组仔猪口服5 mL DMEM培养基。感染组仔猪分别在口服接种后22、39和70 h出现明显腹泻、脱水和嗜睡等临床症状，及时实施安乐死并采集小肠组织样品；对应时间点分别选取对照组1头仔猪实施安乐死并采集小肠组织样品。采用HE染色、PAS染色和AB染色观察小肠组织病理学变化和GC数量变化。采用荧光定量PCR和ELISA分别检测初生仔猪小肠组织中发状分裂相关增强子1（Hes1）和黏蛋白2（MUC2）的转录和含量的变化。结果显示，感染组仔猪与对照组仔猪相比小肠各段黏膜结构均有不同程度损伤，绒毛长度降低且差异显著（P<0.05），VH：CD的比值降低且差异显著（P<0.05）。PAS染色与AB染色结果一致，小肠各段GC数量均有不同程度减少且差异显著（P<0.05、P<0.01或P<0.001）。感染组仔猪与对照组仔猪相比小肠各段Hes1 mRNA的转录量和含量均升高，在空肠和回肠中Hes1 mRNA转录量差异显著（P<0.01或P<0.05），在十二指肠和回肠中Hes1含量差异显著（P<0.01）。感染组仔猪与对照组仔猪相比小肠各段MUC2 mRNA的转录量和含量均降低，在十二指肠和回肠中差异显著（P<0.05、P<0.01或P<0.001）。结果表明，PDCoV感染初生仔猪后引起小肠GC数量明显减少。PDCoV感染仔猪可能通过激活肠道Notch信号通路下游靶基因Hes1表达来阻碍小肠中GC形成和分泌，导致GC数量减少和MUC2表达降低。

关键词: 猪德尔塔冠状病毒, 仔猪, 杯状细胞, 小肠, 发状分裂相关增强子1, 黏蛋白2

Abstract: The aim of this study was to investigate the effect of porcine deltacoronavirus(PDCoV) infected neonatal piglet on small intestinal goblet cells (GC). Six neonatal piglets without colostrum were randomly divided into infection group (n=3) and control group (n=3). After acclimation for 48 h, the infected group was inoculated orally with a PDCoV-CHN-HG-2017 preparation with a titer of 1×10⁵TCID₅₀·mL^-1 (5 mL per pig), while the mock-infected group was inoculated with 5 mL of maintenance medium. Clinical signs were monitored hourly. The infected piglets developed typical symptoms, such as diarrhea, dehydration and lethargy, from 22 to 70 hours post inoculation (hpi). Piglets were euthanized at 22, 39 and 70 hpi after the onset of clinical symptoms. At the same time point, one piglet was treated in the control group. HE staining, PAS staining and AB staining were used to observe the histopathological changes and the number of GC in the small intestine of piglets. Fluorescent quantitative PCR and ELISA were used to detect the transcriptional quantity and content of mucoprotein-2 (MUC2) and hairy and enhancer of split-1 (Hes1) in small intestine of neonatal piglets. The results showed that piglets in the infected group had different degree of damage to the mucosal structure of small intestine compared with piglets in the control group, the villus length were significantly decreased (P<0.05), the ratio of VH:CD significantly decreased (P<0.05). Compared with the control group, the number of GC in each segment of small intestine of infected piglets was significantly reduced. The results of PAS staining and AB staining were consistent. Compared with the control group, the transcription and content of Hes1 mRNA in each segment of the small intestine of piglets in the infected group were all increased, and there were significant differences in the transcription of Hes1 in jejunum and ileum (P<0.01 or P<0.05), and there were significant differences in the content of Hes1 mRNA in the duodenum and ileum (P<0.01). Compared with the control group, the transcription and content of MUC2 mRNA in each segment of the small intestine of infected piglets were reduced, and the differences were significant in the duodenum and ileum (P<0.05, P<0.01 or P<0.001). In summary, the number of small intestinal GC was significantly reduced after PDCoV infection in neonatal piglets. PDCoV infection in piglets may inhibit the formation and secretion of GC in the small intestine by activating the Notch signaling pathway in the intestine to enhance the expression of target gene Hes1, resulting in a decrease in the number of GC and the expression of MUC2.

Key words: PDCoV, piglet, goblet cell, small intestine, Hes1, MUC2

中图分类号:

S858.285.3

梁继翔, 焦哲, 严治山, 李旸, 李栋祺, 刘晓丽, 谷长勤, 胡薛英, 程国富, 张万坡. 猪德尔塔冠状病毒感染对初生仔猪小肠杯状细胞数量及Hes1和MUC2表达的影响[J]. 畜牧兽医学报, 2021, 52(3): 772-781.

LIANG Jixiang, JIAO Zhe, YAN Zhishan, LI Yang, LI Dongqi, LIU Xiaoli, GU Changqin, HU Xueying, CHENG Guofu, ZHANG Wanpo. Effect of Porcine Deltacoronavirus Infection on the Number of Intestinal Goblet Cells and the Expression of Hes1 and MUC2 in the Small Intestine of Neonatal Piglets[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(3): 772-781.

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