Three-dimensional Huh7 cell culture system for the study of Hepatitis C virus infection
In order to elucidate how Hepatitis C Virus (HCV) interacts with polarized hepatocytes in vivo and how HCV-induced alterations in cellular function contribute to HCV-associated liver disease, a more physiologically relevant hepatocyte culture model is needed. As such, NASA-engineered three-dimensional (3-D) rotating wall vessel (RWV) bioreactors were used in effort to promote differentiation of HCV-permissive Huh7 hepatoma cells.
Results: When cultured in the RWV, Huh7 cells became morphologically and transcriptionally distinct from more standard Huh7 two-dimensional (2-D) monolayers.
Specifically, RWV-cultured Huh7 cells formed complex, multilayered 3-D aggregates in which Phase I and Phase II xenobiotic drug metabolism genes, as well as hepatocyte-specific transcripts (HNF4-alpha, Albumin, TTR and alpha-1AT), were upregulated compared to 2-D cultured Huh7 cells. Immunofluorescence analysis revealed that these HCV-permissive 3-D cultured Huh7 cells were more polarized than their 2D counterparts with the expression of HCV receptors, cell adhesion and tight junction markers (CD81, scavenger receptor class B member 1, claudin-1, occludin, ZO-1, beta-Catenin and E-Cadherin) significantly increased and exhibiting apical, lateral and/or basolateral localization.
Conclusions: These findings show that when cultured in 3-D, Huh7 cells acquire a more differentiated hepatocyte-like phenotype.
Importantly, we show that these 3D cultures are highly permissive for HCV infection, thus providing an opportunity to study HCV entry and the effects of HCV infection on host cell function in a more physiologically relevant cell culture system.
Author: Bruno SainzVeronica TenCateSusan Uprichard
Credits/Source: Virology Journal 2009, 6:103
Published on: 2009-07-15
Copyright by the authors listed above - made available via BioMedCentral (Open Access). Please
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