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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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LPL inhibits cell infection by the JFH-1 and J6/JFH-1 strains produced in vitro and in vivo in a chimeric uPA-SCID mouse model.The HCVcc strains JFH-1 (A) and J6/JFH-1 (B) were produced in the Huh7.5 hepatoma cell line. Cells were incubated with (or without) LPL for 30 min at 4°C and then with virus preparations for 2 h at 37°C to allow infection. RNA was extracted from cells 24 h post infection and HCV RNA was quantified by RT-qPCR. The data obtained were normalized with respect to levels of GADPH. The mJFH-1 (A) and mJ6/JFH-1 (B) correspond to HCVcc strains produced in chimeric uPA-SCID mice into which we transplanted human hepatocytes. Serum samples collected from infected mice were pooled and their capacity to infect Huh7.5 cells was assessed in the presence and absence of LPL, as outlined above. Cells infected in the absence (black bar) and in the presence of LPL (gray bar). The data are expressed as the amount of HCV RNA detected in cells infected in the presence of LPL as compared with the amount of HCV RNA in cells infected in the absence of LPL, expressed as a percentage.
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pone-0026637-g001: LPL inhibits cell infection by the JFH-1 and J6/JFH-1 strains produced in vitro and in vivo in a chimeric uPA-SCID mouse model.The HCVcc strains JFH-1 (A) and J6/JFH-1 (B) were produced in the Huh7.5 hepatoma cell line. Cells were incubated with (or without) LPL for 30 min at 4°C and then with virus preparations for 2 h at 37°C to allow infection. RNA was extracted from cells 24 h post infection and HCV RNA was quantified by RT-qPCR. The data obtained were normalized with respect to levels of GADPH. The mJFH-1 (A) and mJ6/JFH-1 (B) correspond to HCVcc strains produced in chimeric uPA-SCID mice into which we transplanted human hepatocytes. Serum samples collected from infected mice were pooled and their capacity to infect Huh7.5 cells was assessed in the presence and absence of LPL, as outlined above. Cells infected in the absence (black bar) and in the presence of LPL (gray bar). The data are expressed as the amount of HCV RNA detected in cells infected in the presence of LPL as compared with the amount of HCV RNA in cells infected in the absence of LPL, expressed as a percentage.

Mentions: We investigated the effect of LPL on cell infection with these two HCVcc strains, by incubating Huh7.5 cells with bovine LPL for 30 min at 4°C before adding the virus, and determining intracellular HCV RNA levels 24 h after infection, as previously described [32]. The infection of cells with the two virus strains was inhibited by more than 90% in the presence of LPL (Figure 1A). This inhibition was also observed 48 and 72 h after infection. Thus, the addition of LPL to Huh7.5 cells as an exogenous ligand inhibited cell infection with various HCVcc strains.


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)

LPL inhibits cell infection by the JFH-1 and J6/JFH-1 strains produced in vitro and in vivo in a chimeric uPA-SCID mouse model.The HCVcc strains JFH-1 (A) and J6/JFH-1 (B) were produced in the Huh7.5 hepatoma cell line. Cells were incubated with (or without) LPL for 30 min at 4°C and then with virus preparations for 2 h at 37°C to allow infection. RNA was extracted from cells 24 h post infection and HCV RNA was quantified by RT-qPCR. The data obtained were normalized with respect to levels of GADPH. The mJFH-1 (A) and mJ6/JFH-1 (B) correspond to HCVcc strains produced in chimeric uPA-SCID mice into which we transplanted human hepatocytes. Serum samples collected from infected mice were pooled and their capacity to infect Huh7.5 cells was assessed in the presence and absence of LPL, as outlined above. Cells infected in the absence (black bar) and in the presence of LPL (gray bar). The data are expressed as the amount of HCV RNA detected in cells infected in the presence of LPL as compared with the amount of HCV RNA in cells infected in the absence of LPL, expressed as a percentage.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0026637-g001: LPL inhibits cell infection by the JFH-1 and J6/JFH-1 strains produced in vitro and in vivo in a chimeric uPA-SCID mouse model.The HCVcc strains JFH-1 (A) and J6/JFH-1 (B) were produced in the Huh7.5 hepatoma cell line. Cells were incubated with (or without) LPL for 30 min at 4°C and then with virus preparations for 2 h at 37°C to allow infection. RNA was extracted from cells 24 h post infection and HCV RNA was quantified by RT-qPCR. The data obtained were normalized with respect to levels of GADPH. The mJFH-1 (A) and mJ6/JFH-1 (B) correspond to HCVcc strains produced in chimeric uPA-SCID mice into which we transplanted human hepatocytes. Serum samples collected from infected mice were pooled and their capacity to infect Huh7.5 cells was assessed in the presence and absence of LPL, as outlined above. Cells infected in the absence (black bar) and in the presence of LPL (gray bar). The data are expressed as the amount of HCV RNA detected in cells infected in the presence of LPL as compared with the amount of HCV RNA in cells infected in the absence of LPL, expressed as a percentage.
Mentions: We investigated the effect of LPL on cell infection with these two HCVcc strains, by incubating Huh7.5 cells with bovine LPL for 30 min at 4°C before adding the virus, and determining intracellular HCV RNA levels 24 h after infection, as previously described [32]. The infection of cells with the two virus strains was inhibited by more than 90% in the presence of LPL (Figure 1A). This inhibition was also observed 48 and 72 h after infection. Thus, the addition of LPL to Huh7.5 cells as an exogenous ligand inhibited cell infection with various HCVcc strains.

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