哈獸研馮力團隊首次建立冠狀病毒PEDV體外感染豬腸類器官模型

近日，中國農業科學院哈爾濱獸醫研究所馮力研究員領銜的豬消化道傳染病創新團隊在豬流行性腹瀉病毒（PEDV) 體外感染腸類器官模型的研究上取得新進展，相關研究成果以「Porcine Intestinal Enteroids: a New Model for Studying Enteric Coronavirus PEDV Infection and the Host Innate Response」為題，發表在病毒學專業期刊《病毒學雜誌（Journal of Virology）》上。該研究成功建立了一個可以模擬體內PEDV由腸道幹細胞分化成的體外腸類器官感染的模型，為解析豬腸道PEDV致病性和免疫應答的相關研究奠定了基礎。該模型不僅可以用於PEDV的相關研究，還可以用於其他豬腸道病毒的研究。解決了PEDV缺少合適的體外感染模型的瓶頸問題。

豬流行性腹瀉病毒（Porcine Epidemic Diarrhea Virus，PEDV）屬於α冠狀病毒屬，是引起仔豬腹瀉的最主要病原， 每年給養殖業造成重大經濟損失。腸道上皮是由多種細胞組成的絨毛和隱窩複雜結構，而當前常用的PEDV體外感染模型都是非豬源的綠猴腎細胞（ Vero 細胞） 或者單一的豬小腸上皮細胞（ IPEC-J2) ，不能夠很好地模擬體內情況。由於缺乏很好的體外豬腸道模型，因此PEDV感染致病機制的相關研究仍然不是很清楚。

在本研究中，科研人員分別從十二指腸、空腸和迴腸分離腸隱窩幹細胞並體外培養分化成豬小腸類器官細胞。並且發現腸隱窩幹細胞分化的小腸類器官能夠在體外很好地重現小腸上皮細胞結構的複雜性並且能夠感染PEDV。 進一步證實PEDV可以感染多種類型的腸上皮細胞，包括腸道細胞、幹細胞和杯狀細胞。而且和體內感染結果一致，迴腸類器官比結腸類器官對PEDV更敏感，體現腸段的感染差異性。此外，IFN-λ比IFN-α更有效地誘導抗病毒反應和抑制PEDV在迴腸類器官中的感染。該研究中，建立了一種新的可模擬體內的體外類器官細胞模型，用於探索PEDV的致病機制及宿主和各種豬腸道病毒之間相互作用的體外感染模型。

ABSTRACT

Porcine epidemic diarrhea virus (PEDV), a member of the group of alphacoronaviruses, is the pathogen of a highly contagious gastrointestinal swine disease. The elucidation of the events associated with the intestinal epithelial response to PEDV infection has been limited by the absence of good in vitroporcine intestinal models that recapitulate the multicellular complexity of the gastrointestinal tract. Here, we generated swine enteroids from the intestinal crypt stem cells of the duodenum, jejunum, or ileum, and found that the generated enteroids are able to satisfactorily recapitulate the complicated intestinal epithelium in vivo and are susceptible to infection by PEDV. PEDV infected multiple types of cells including enterocytes, stem cells, and goblet cells, and exhibited segmental infection discrepancies compared with ileal enteroids and colonoids, and this finding was verified in vivo. Moreover, the clinical isolate PEDV-JMS propagated better in ileal enteroids than the cell-adapted PEDV CV777, and PEDV infection suppressed IFN production early during the infection course. IFN-lambda elicited a potent antiviral response and inhibited PEDV in enteroids more efficiently than IFN-α. Therefore, swine enteroids provide a novel in vitro model for exploring the pathogenesis of PEDV and for the in vitro study of the interplay between a host and a variety of swine enteric viruses.

IMPORTANCE

PEDV is a highly contagious enteric coronavirus that causes significant economic losses, and the lack of a good in vitro model system is a major roadblock to an in-depth understanding of PEDV pathogenesis. Here, we generated a porcine intestinal enteroid model for PEDV infection. Utilizing porcine intestinal enteroids, we demonstrated that PEDV infects multiple lineages of the intestinal epithelium and preferably infects ileal enteroids over colonoids and that enteroids prefer to respond to IFN-lambda 1 over IFN-α. These events recapitulate the events that occur in vivo. This study constitutes the first use of a primary intestinal enteroid model to investigate the susceptibility of porcine enteroids to PEDV and to determine the antiviral response following infection. Our study provides important insights into the events associated with PEDV infection of the porcine intestine and provides a valuable in vitro model for studying not only PEDV but also other swine enteric viruses.

來源：哈獸研

本期編輯：Tony

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