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Contribution of intestinal porcine organoids to the study of host-virus interactions of transmissible gastroenteritis virus
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International audience. Background and Objectives: Cellular culture (In Cellula) and/or animal experimentation (In Vivo) are standard methods for studying host-pathogen interactions. However, both methods have methodological or ethical limitations. Organoids, produced by the differentiation and self-organizationof stem cells, recapitulate the epithelial cellular diversity and structural organization of organs from which they are derived and, as such, represent an alternative to the In Cellula and In Vivo methods.The Viral Genetic and Biosafety Unit has implemented the production of porcine intestinal organoids (PIO) and developed a protocol for infection based on PIO’s monolayer to expose cell receptors to porcine transmissible gastroenteritis (TGEV) alpha-coronaviruses. Two TGEV strains were used forthe experiments: one belonging to the Purdue genogroup, adapted to cell culture, and the other belonging to the Miller genogroup, which is poorly adapted to cell culture.Material and Methods: To evaluate the advantages and limitations of organoids in the study of host-virus interactions, piglets, PIO and ST cells were infected with a comparable viral genomic load of 10E8 N gene copies of the virus. For each model, infection kinetics were performed, and cellular RNAs were harvested for RNA-seq analysis of host-virus interactions.Results: Comparison of the gene expression between the three models without any infection revealed Pearson’s correlation of 0,422 and 0,485 between PIO and piglet jejunum or ST cells, respectively, when this correlation was only 0,229 between piglets jejunum and ST cells. 56% of the genes expressed were common to all the three models. A total of 1103 genes were expressed in both the PIO and piglet jejunum. Gene ontology analysis of these genes revealed enrichment in the cellular components of the brush border, tight junctions, and apical/basal plasma membranes.The relative expression of the TGEV-N gene displayed differential dynamics depending on the strain. For the Miller strain, N viral gene expression decreased after 9 hpi, in contrast to the Purdue strain, for which N expression increased up to 24h. At the same MOI in organoids, 1000 timesmore of the viral N gene transcript was found after Purdue infection at 24h compared after Miller infection. In Vivo, a 3-log difference in viral N gene expression was observed in the piglet jejunum, in favor of the Miller strain.Conclusion: In conclusion, the Purdue strain maintained an increased ability to infect PIOs (as for ST cells) compared to the Miller strain, in contrast to the In Vivo model. Transcriptomic analysis is currently being performed for these three infected experimental models.
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