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Lipoprotein lipase inhibits hepatitis C virus (HCV) infection by blocking virus cell entry.

Maillard P, Walic M, Meuleman P, Roohvand F, Huby T, Le Goff W, Leroux-Roels G, Pécheur EI, Budkowska A - PLoS ONE (2011)

Bottom Line: The effect of LPL depended on its enzymatic activity.These analyses demonstrated the internalization of virus particles into hepatoma cells and their presence in intracellular vesicles and associated with lipid droplets.HCV-associated lipoproteins may therefore be a promising target for the development of new therapeutic approaches.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur, Unité Hépacivirus et Immunité Innée, Département de Virologie, Paris, France.

ABSTRACT
A distinctive feature of HCV is that its life cycle depends on lipoprotein metabolism. Viral morphogenesis and secretion follow the very low-density lipoprotein (VLDL) biogenesis pathway and, consequently, infectious HCV in the serum is associated with triglyceride-rich lipoproteins (TRL). Lipoprotein lipase (LPL) hydrolyzes TRL within chylomicrons and VLDL but, independently of its catalytic activity, it has a bridging activity, mediating the hepatic uptake of chylomicrons and VLDL remnants. We previously showed that exogenously added LPL increases HCV binding to hepatoma cells by acting as a bridge between virus-associated lipoproteins and cell surface heparan sulfate, while simultaneously decreasing infection levels. We show here that LPL efficiently inhibits cell infection with two HCV strains produced in hepatoma cells or in primary human hepatocytes transplanted into uPA-SCID mice with fully functional human ApoB-lipoprotein profiles. Viruses produced in vitro or in vivo were separated on iodixanol gradients into low and higher density populations, and the infection of Huh 7.5 cells by both virus populations was inhibited by LPL. The effect of LPL depended on its enzymatic activity. However, the lipase inhibitor tetrahydrolipstatin restored only a minor part of HCV infectivity, suggesting an important role of the LPL bridging function in the inhibition of infection. We followed HCV cell entry by immunoelectron microscopy with anti-envelope and anti-core antibodies. These analyses demonstrated the internalization of virus particles into hepatoma cells and their presence in intracellular vesicles and associated with lipid droplets. In the presence of LPL, HCV was retained at the cell surface. We conclude that LPL efficiently inhibits HCV infection by acting on TRL associated with HCV particles through mechanisms involving its lipolytic function, but mostly its bridging function. These mechanisms lead to immobilization of the virus at the cell surface. HCV-associated lipoproteins may therefore be a promising target for the development of new therapeutic approaches.

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The inhibitory effect of LPL on HCVcc infection is only partly related to its catalytic activity.(A) Huh7.5 cells were pre-incubated with 1 µg/ml LPL at 4°C in the presence or absence of 50 µg/ml THL before infection with JFH1. The infected cells were grown for 24 h and HCV RNA was then extracted and quantified by RT-qPCR. (B) THL does not influence HCV replication. Huh7.5 cells were pre-incubated with indicated concentrations of THL before cell infection with JFH-1, as for experiments with LPL. THL was maintained in the medium for 24 h post infection. HCV RNA was then extracted and quantified by RT-qPCR. Results are expressed as a percent of RNA as compared with control cells infected in the absence of LPL and THL.
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pone-0026637-g005: The inhibitory effect of LPL on HCVcc infection is only partly related to its catalytic activity.(A) Huh7.5 cells were pre-incubated with 1 µg/ml LPL at 4°C in the presence or absence of 50 µg/ml THL before infection with JFH1. The infected cells were grown for 24 h and HCV RNA was then extracted and quantified by RT-qPCR. (B) THL does not influence HCV replication. Huh7.5 cells were pre-incubated with indicated concentrations of THL before cell infection with JFH-1, as for experiments with LPL. THL was maintained in the medium for 24 h post infection. HCV RNA was then extracted and quantified by RT-qPCR. Results are expressed as a percent of RNA as compared with control cells infected in the absence of LPL and THL.

Mentions: LPL has two physiological roles, either of which may affect virus infectivity. Its enzymatic activity might induce changes in the lipid composition and structure of the virus particles, whereas its “bridging” function might play a role in the inhibitory process, either binding the virus tightly to the cell surface or directing it to the abortive infection pathway. We investigated which of these LPL functions was determinant for the inhibition of HCV infection, by performing experiments with THL, blocking the catalytic activity of LPL. The inhibitory effect of LPL on HCVcc infection was only partially abrogated by THL (Figure 5A). In the absence of THL, LPL inhibited viral infection by two orders of magnitude, whereas, in the presence of THL, LPL also decreased infection levels, but only by one order of magnitude. We confirmed that THL alone had no influence on viral replication capacity, by measuring intracellular HCV RNA levels, 24 h after infection (Figure 5B).


Lipoprotein lipase inhibits hepatitis C virus (HCV) infection by blocking virus cell entry.

Maillard P, Walic M, Meuleman P, Roohvand F, Huby T, Le Goff W, Leroux-Roels G, Pécheur EI, Budkowska A - PLoS ONE (2011)

The inhibitory effect of LPL on HCVcc infection is only partly related to its catalytic activity.(A) Huh7.5 cells were pre-incubated with 1 µg/ml LPL at 4°C in the presence or absence of 50 µg/ml THL before infection with JFH1. The infected cells were grown for 24 h and HCV RNA was then extracted and quantified by RT-qPCR. (B) THL does not influence HCV replication. Huh7.5 cells were pre-incubated with indicated concentrations of THL before cell infection with JFH-1, as for experiments with LPL. THL was maintained in the medium for 24 h post infection. HCV RNA was then extracted and quantified by RT-qPCR. Results are expressed as a percent of RNA as compared with control cells infected in the absence of LPL and THL.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3198807&req=5

pone-0026637-g005: The inhibitory effect of LPL on HCVcc infection is only partly related to its catalytic activity.(A) Huh7.5 cells were pre-incubated with 1 µg/ml LPL at 4°C in the presence or absence of 50 µg/ml THL before infection with JFH1. The infected cells were grown for 24 h and HCV RNA was then extracted and quantified by RT-qPCR. (B) THL does not influence HCV replication. Huh7.5 cells were pre-incubated with indicated concentrations of THL before cell infection with JFH-1, as for experiments with LPL. THL was maintained in the medium for 24 h post infection. HCV RNA was then extracted and quantified by RT-qPCR. Results are expressed as a percent of RNA as compared with control cells infected in the absence of LPL and THL.
Mentions: LPL has two physiological roles, either of which may affect virus infectivity. Its enzymatic activity might induce changes in the lipid composition and structure of the virus particles, whereas its “bridging” function might play a role in the inhibitory process, either binding the virus tightly to the cell surface or directing it to the abortive infection pathway. We investigated which of these LPL functions was determinant for the inhibition of HCV infection, by performing experiments with THL, blocking the catalytic activity of LPL. The inhibitory effect of LPL on HCVcc infection was only partially abrogated by THL (Figure 5A). In the absence of THL, LPL inhibited viral infection by two orders of magnitude, whereas, in the presence of THL, LPL also decreased infection levels, but only by one order of magnitude. We confirmed that THL alone had no influence on viral replication capacity, by measuring intracellular HCV RNA levels, 24 h after infection (Figure 5B).

Bottom Line: The effect of LPL depended on its enzymatic activity.These analyses demonstrated the internalization of virus particles into hepatoma cells and their presence in intracellular vesicles and associated with lipid droplets.HCV-associated lipoproteins may therefore be a promising target for the development of new therapeutic approaches.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur, Unité Hépacivirus et Immunité Innée, Département de Virologie, Paris, France.

ABSTRACT
A distinctive feature of HCV is that its life cycle depends on lipoprotein metabolism. Viral morphogenesis and secretion follow the very low-density lipoprotein (VLDL) biogenesis pathway and, consequently, infectious HCV in the serum is associated with triglyceride-rich lipoproteins (TRL). Lipoprotein lipase (LPL) hydrolyzes TRL within chylomicrons and VLDL but, independently of its catalytic activity, it has a bridging activity, mediating the hepatic uptake of chylomicrons and VLDL remnants. We previously showed that exogenously added LPL increases HCV binding to hepatoma cells by acting as a bridge between virus-associated lipoproteins and cell surface heparan sulfate, while simultaneously decreasing infection levels. We show here that LPL efficiently inhibits cell infection with two HCV strains produced in hepatoma cells or in primary human hepatocytes transplanted into uPA-SCID mice with fully functional human ApoB-lipoprotein profiles. Viruses produced in vitro or in vivo were separated on iodixanol gradients into low and higher density populations, and the infection of Huh 7.5 cells by both virus populations was inhibited by LPL. The effect of LPL depended on its enzymatic activity. However, the lipase inhibitor tetrahydrolipstatin restored only a minor part of HCV infectivity, suggesting an important role of the LPL bridging function in the inhibition of infection. We followed HCV cell entry by immunoelectron microscopy with anti-envelope and anti-core antibodies. These analyses demonstrated the internalization of virus particles into hepatoma cells and their presence in intracellular vesicles and associated with lipid droplets. In the presence of LPL, HCV was retained at the cell surface. We conclude that LPL efficiently inhibits HCV infection by acting on TRL associated with HCV particles through mechanisms involving its lipolytic function, but mostly its bridging function. These mechanisms lead to immobilization of the virus at the cell surface. HCV-associated lipoproteins may therefore be a promising target for the development of new therapeutic approaches.

Show MeSH
Related in: MedlinePlus