Characterization of the lipid envelope of exosome encapsulated HEV particles protected from the immune response

Chapuy‐Regaud, Sabine; Dubois, Martine; Plisson‐Chastang, Célia; Bonnefois, Tiffany; Lhomme, Sébastien; Bertrand‐Michel, Justine; You, Bruno; Simoneau, Steve; Gleizes, Pierre‐Emmanuel; Flan, Benoît; Abravanel, Florence; Izopet, Jacques

doi:10.1016/j.biochi.2017.05.003

Cited by 105 publications

(101 citation statements)

References 68 publications

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“…As they are not polarized, these cells must be lysed to recover non-enveloped particles (intracellular HEV). In agreement with previous studies 28, [33][34][35][36] we found that the quasi-envelope rendered extracellular HEV less infectious (Supplementary Fig. 5), but protected the virus from neutralization by anti-HEV capsid ORF2 protein antibodies (Fig.…”

Section: Resultssupporting

confidence: 93%

Stem cell-derived polarized hepatocytes

Thi

Belote

et al. 2020

Nat Commun

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Human stem cell-derived hepatocyte-like cells (HLCs) offer an attractive platform to study liver biology. Despite their numerous advantages, HLCs lack critical in vivo characteristics, including cell polarity. Here, we report a stem cell differentiation protocol that uses transwell filters to generate columnar polarized HLCs with clearly defined basolateral and apical membranes separated by tight junctions. We show that polarized HLCs secrete cargo directionally: Albumin, urea, and lipoproteins are secreted basolaterally, whereas bile acids are secreted apically. Further, we show that enterically transmitted hepatitis E virus (HEV) progeny particles are secreted basolaterally as quasi-enveloped particles and apically as naked virions, recapitulating essential steps of the natural infectious cycle in vivo. We also provide proof-of-concept that polarized HLCs can be used for pharmacokinetic and drug-drug interaction studies. This novel system provides a powerful tool to study hepatocyte biology, disease mechanisms, genetic variation, and drug metabolism in a more physiologically relevant setting.

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Section: Resultssupporting

confidence: 93%

Stem cell-derived polarized hepatocytes

Thi

Belote

et al. 2020

Nat Commun

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“…Extracellular vesicles in the first four columns of Table 1 show high PS levels (138,143). Higher concentrations of PS on the outer leaflet of the lipid bilayer would promote uptake by other cells like macrophages (144,145).…”

Section: Lipid Rafts and Ev Lipidsmentioning

confidence: 99%

The ins and outs of lipid rafts: functions in intracellular cholesterol homeostasis, microparticles, and cell membranes

Ouweneel

Thomas

Sorci‐Thomas

2020

Journal of Lipid Research

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Cellular membranes are not homogenous mixtures of proteins; rather, they are segregated into microdomains on the basis of preferential association between specific lipids and proteins. These microdomains, called lipid rafts, are well known for their role in receptor signaling on the plasma membrane (PM) and are essential to such cellular functions as signal transduction and spatial organization of the PM. A number of disease states, including atherosclerosis and other cardiovascular disorders, may be caused by dysfunctional maintenance of lipid rafts. Lipid rafts do not occur only in the PM but also have been found in intracellular membranes and extracellular vesicles (EVs). Here, we focus on discussing newly discovered functions of lipid rafts and microdomains in intracellular membranes, including lipid and protein trafficking from the ER, Golgi bodies, and endosomes to the PM, and we examine lipid raft involvement in the production and composition of EVs. Because lipid rafts are small and transient, visualization remains challenging. Future work with advanced techniques will continue to expand our knowledge about the roles of lipid rafts in cellular functioning.

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“…HEV has been cultured in several hepatocarcinoma cell systems (14,(20)(21)(22)(23). The HEV secreted from cells is lipid associated (eHEV) (14,15,23,24). This process involves the interaction of ORF3 with the ESCRT (endosome sorting complex required for transport)associated protein TSG101 (25).…”

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confidence: 99%

Vectorial Release of Hepatitis E Virus in Polarized Human Hepatocytes

Capelli

Marion

Dubois

et al. 2019

J Virol

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Hepatitis E virus (HEV) is a common cause of acute viral hepatitis worldwide. Most HEV infections are asymptomatic, but immunocompromised patients infected with HEV genotype 3 (HEV3), HEV4, or HEV7 may develop chronic infections. The HEV particles in stools are naked (nHEV), while those in the serum and culture supernatants (eHEV) are associated with lipids. Hepatocytes are polarized epithelial cells that have basolateral (oriented toward the blood) and apical (oriented toward the bile) exosomal pathways. We isolated a subclone, F2, from the human hepatocarcinoma cell line HepG2/C3A that grew as a polarized monolayer culture and had better HEV production than HepG2/C3A cells. F2 cells cultured on semipermeable collagen inserts and infected basolaterally with nHEV3 released 94.6% of virus particles apically, those infected with eHEV3 released 96.8% apically, and eHEV1-infected cells released 99.3% apically. Transcytosis was not involved. Density gradient centrifugation and NP-40 treatment showed that HEV particles released both apically and basolaterally were lipid associated. The apically released HEV3 and HEV1 particles were six and nine times more infectious than those released basolaterally, respectively. Confocal microscopy indicated that the open reading frame 2 (ORF2) capsid protein colocalized apically with ORF3 virus protein, the apical marker DPP4, and the recycling endosome GTPase Rab27a. The amounts of soluble glycosylated ORF2 secreted apically and basolaterally were similar. These polarized-hepatocyte data suggest that infectious HEV particles are mainly released into bile, while the small fraction released into blood could spread HEV throughout the host. IMPORTANCE Hepatitis E virus (HEV) in stools is naked, while that in culture supernatants and patients’ blood is lipid associated. Its life cycle in hepatocytes, polarized cells with a basolateral side communicating with blood and an apical side connected with bile, is incompletely understood. We have developed a polarized hepatocyte model and used the cells to analyze the supernatants bathing the apical and basolateral sides and HEV subcellular distribution. HEV particles from both sides were lipid associated, and most infectious HEV particles left the cell via its apical side. Similar amounts of the open reading frame 2 (ORF2) soluble capsid protein were secreted from both sides of the hepatocytes. This model mimicking physiological conditions should help clarify the HEV cell cycle in polarized hepatocytes.

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Characterization of the lipid envelope of exosome encapsulated HEV particles protected from the immune response

Cited by 105 publications

References 68 publications

Stem cell-derived polarized hepatocytes

Stem cell-derived polarized hepatocytes

The ins and outs of lipid rafts: functions in intracellular cholesterol homeostasis, microparticles, and cell membranes

Vectorial Release of Hepatitis E Virus in Polarized Human Hepatocytes

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