tetano
Editor, Senior Moderator
PLoS One
. 2024 Jan 25;19(1):e0292977.
doi: 10.1371/journal.pone.0292977. eCollection 2024. A polarized cell system amenable to subcellular resolution imaging of influenza virus infection
Jean-Baptiste Brault[SUP] 1 [/SUP], Catherine Thouvenot[SUP] 2 [/SUP], Magda Cannata Serio[SUP] 1 [/SUP], Sylvain Paisant[SUP] 1 [/SUP], Julien Fernandes[SUP] 3 [/SUP], David Gény[SUP] 4 [/SUP], Lydia Danglot[SUP] 4 5 [/SUP], Adeline Mallet[SUP] 2 [/SUP], Nadia Naffakh[SUP] 1 [/SUP]
Affiliations
The life cycle of influenza A viruses (IAV), and notably intracellular trafficking of the viral genome, depends on multiple interactions with the cellular cytoskeleton and endomembrane system. A limitation of the conventional cellular models used for mechanistic study and subcellular imaging of IAV infection is that they are cultured in two dimensions (2D) under non-polarizing conditions, and therefore they do not recapitulate the intracellular organization of the polarized respiratory epithelial cells naturally targeted by IAVs. To overcome this limitation, we developed an IAV-infection assay in a 3D cell culture system which allows imaging along the baso-lateral axis of polarized cells, with subcellular resolution. Here we describe a protocol to grow polarized monolayers of Caco2-TC7 cells on static Cytodex-3 microcarrier beads, infect them with IAV, and subsequently perform immunostaining and confocal imaging, or electron microscopy, on polarized IAV-infected cells. This method can be extended to other pathogens that infect human polarized epithelial cells.
. 2024 Jan 25;19(1):e0292977.
doi: 10.1371/journal.pone.0292977. eCollection 2024. A polarized cell system amenable to subcellular resolution imaging of influenza virus infection
Jean-Baptiste Brault[SUP] 1 [/SUP], Catherine Thouvenot[SUP] 2 [/SUP], Magda Cannata Serio[SUP] 1 [/SUP], Sylvain Paisant[SUP] 1 [/SUP], Julien Fernandes[SUP] 3 [/SUP], David Gény[SUP] 4 [/SUP], Lydia Danglot[SUP] 4 5 [/SUP], Adeline Mallet[SUP] 2 [/SUP], Nadia Naffakh[SUP] 1 [/SUP]
Affiliations
- PMID: 38271396
- PMCID: PMC10810476
- DOI: 10.1371/journal.pone.0292977
The life cycle of influenza A viruses (IAV), and notably intracellular trafficking of the viral genome, depends on multiple interactions with the cellular cytoskeleton and endomembrane system. A limitation of the conventional cellular models used for mechanistic study and subcellular imaging of IAV infection is that they are cultured in two dimensions (2D) under non-polarizing conditions, and therefore they do not recapitulate the intracellular organization of the polarized respiratory epithelial cells naturally targeted by IAVs. To overcome this limitation, we developed an IAV-infection assay in a 3D cell culture system which allows imaging along the baso-lateral axis of polarized cells, with subcellular resolution. Here we describe a protocol to grow polarized monolayers of Caco2-TC7 cells on static Cytodex-3 microcarrier beads, infect them with IAV, and subsequently perform immunostaining and confocal imaging, or electron microscopy, on polarized IAV-infected cells. This method can be extended to other pathogens that infect human polarized epithelial cells.