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Impaired LDL receptor-related protein 1 translocation correlates with improved dyslipidemia and atherosclerosis in apoE-deficient mice.

Gordts PL, Bartelt A, Nilsson SK, Annaert W, Christoffersen C, Nielsen LB, Heeren J, Roebroek AJ - PLoS ONE (2012)

Bottom Line: Postprandial lipoprotein improvement was explained by increased hepatic clearance of triglyceride-rich remnant lipoproteins and accompanied by a compensatory 1.6-fold upregulation of LDLR expression in hepatocytes.These findings demonstrate that the NPxYxxL motif in LRP1 is important for insulin-mediated translocation and slow perinuclear endosomal recycling.These LRP1 impairments correlated with reduced atherogenesis and cholesterol levels in apoE-deficient mice, likely via compensatory LDLR upregulation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Experimental Mouse Genetics, Center for Human Genetics, KU Leuven, Leuven, Belgium.

ABSTRACT

Objective: Determination of the in vivo significance of LDL receptor-related protein 1 (LRP1) dysfunction on lipid metabolism and atherosclerosis development in absence of its main ligand apoE.

Methods and results: LRP1 knock-in mice carrying an inactivating mutation in the NPxYxxL motif were crossed with apoE-deficient mice. In the absence of apoE, relative to LRP1 wild-type animals, LRP1 mutated mice showed an increased clearance of postprandial lipids despite a compromised LRP1 endocytosis rate and inefficient insulin-mediated translocation of the receptor to the plasma membrane, likely due to inefficient slow recycling of the mutated receptor. Postprandial lipoprotein improvement was explained by increased hepatic clearance of triglyceride-rich remnant lipoproteins and accompanied by a compensatory 1.6-fold upregulation of LDLR expression in hepatocytes. One year-old apoE-deficient mice having the dysfunctional LRP1 revealed a 3-fold decrease in spontaneous atherosclerosis development and a 2-fold reduction in LDL-cholesterol levels.

Conclusion: These findings demonstrate that the NPxYxxL motif in LRP1 is important for insulin-mediated translocation and slow perinuclear endosomal recycling. These LRP1 impairments correlated with reduced atherogenesis and cholesterol levels in apoE-deficient mice, likely via compensatory LDLR upregulation.

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Compensatory LDLR up-regulation associated with an increased chylomicron remnant clearance.A–C, Internalization of 125I-CR (A) and 125I-CR-K1 (B) in primary hepatocytes and LDLR immunofluorescence staining in primary hepatocytes (bars are 20 µm) (C). D, Immunoblot analyses of hepatocytes for LDLR and β-actin protein levels after a 16 h incubation period with either 10% FBS or 10% Lipoprotein Deficient FBS (LPDS). E, Immunoblot analysis of microsomal liver extracts for LDLR and β-actin protein levels (n  = 6 per genotype). ApoE−/− (□) and apoE−/−LRP1n2/n2 (▪) mice, data are mean±SEM. *P<0.05, **P<0.005.
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pone-0038330-g003: Compensatory LDLR up-regulation associated with an increased chylomicron remnant clearance.A–C, Internalization of 125I-CR (A) and 125I-CR-K1 (B) in primary hepatocytes and LDLR immunofluorescence staining in primary hepatocytes (bars are 20 µm) (C). D, Immunoblot analyses of hepatocytes for LDLR and β-actin protein levels after a 16 h incubation period with either 10% FBS or 10% Lipoprotein Deficient FBS (LPDS). E, Immunoblot analysis of microsomal liver extracts for LDLR and β-actin protein levels (n  = 6 per genotype). ApoE−/− (□) and apoE−/−LRP1n2/n2 (▪) mice, data are mean±SEM. *P<0.05, **P<0.005.

Mentions: In order to identify whether the increased postprandial lipid accumulation in the liver was due to an increased uptake of TRLs into hepatocytes, 125I-radiolabeled CR (Figure 3A) and CR supplemented with vitamin K1 (CR-K1) (Figure 3B) were evaluated in vitro for their uptake by primary hepatocytes. Uptake of radiolabeled CR and CR-K1 was respectively significantly 1.25-fold and 1.5-fold increased in hepatocytes isolated from the apoE−/−LRP1n2/n2 mice compared to the apoE−/− control cells. A possible compensatory mechanism for CR clearance might be upregulated, as it was previously shown that the LRP1 mutation negatively affects the internalization property of the receptor [7] and that apoE-deficiency negatively affects LRP1 mediated clearance of CR-K1 in osteoblasts [33]. It is known that there exists a certain degree of functional redundancy between LDLR and LRP1 at the level of the liver [6]. Both via immunofluorescence (Figure 3C) and immunoblotting (Figure 3D–E) a significant increase in the expression levels of LDLR in hepatocytes (Figure 3C–D) and liver extracts (showing a 1.6-fold increase) (Figure 3E) could be observed between apoE−/−LRP1n2/n2 and apoE−/− mice. These results suggest that LRP1 NPxYxxL-motif inactivation leads to a compensatory up-regulation of the LDLR, which could be responsible for improved CR clearance via hepatocytes.


Impaired LDL receptor-related protein 1 translocation correlates with improved dyslipidemia and atherosclerosis in apoE-deficient mice.

Gordts PL, Bartelt A, Nilsson SK, Annaert W, Christoffersen C, Nielsen LB, Heeren J, Roebroek AJ - PLoS ONE (2012)

Compensatory LDLR up-regulation associated with an increased chylomicron remnant clearance.A–C, Internalization of 125I-CR (A) and 125I-CR-K1 (B) in primary hepatocytes and LDLR immunofluorescence staining in primary hepatocytes (bars are 20 µm) (C). D, Immunoblot analyses of hepatocytes for LDLR and β-actin protein levels after a 16 h incubation period with either 10% FBS or 10% Lipoprotein Deficient FBS (LPDS). E, Immunoblot analysis of microsomal liver extracts for LDLR and β-actin protein levels (n  = 6 per genotype). ApoE−/− (□) and apoE−/−LRP1n2/n2 (▪) mice, data are mean±SEM. *P<0.05, **P<0.005.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3368875&req=5

pone-0038330-g003: Compensatory LDLR up-regulation associated with an increased chylomicron remnant clearance.A–C, Internalization of 125I-CR (A) and 125I-CR-K1 (B) in primary hepatocytes and LDLR immunofluorescence staining in primary hepatocytes (bars are 20 µm) (C). D, Immunoblot analyses of hepatocytes for LDLR and β-actin protein levels after a 16 h incubation period with either 10% FBS or 10% Lipoprotein Deficient FBS (LPDS). E, Immunoblot analysis of microsomal liver extracts for LDLR and β-actin protein levels (n  = 6 per genotype). ApoE−/− (□) and apoE−/−LRP1n2/n2 (▪) mice, data are mean±SEM. *P<0.05, **P<0.005.
Mentions: In order to identify whether the increased postprandial lipid accumulation in the liver was due to an increased uptake of TRLs into hepatocytes, 125I-radiolabeled CR (Figure 3A) and CR supplemented with vitamin K1 (CR-K1) (Figure 3B) were evaluated in vitro for their uptake by primary hepatocytes. Uptake of radiolabeled CR and CR-K1 was respectively significantly 1.25-fold and 1.5-fold increased in hepatocytes isolated from the apoE−/−LRP1n2/n2 mice compared to the apoE−/− control cells. A possible compensatory mechanism for CR clearance might be upregulated, as it was previously shown that the LRP1 mutation negatively affects the internalization property of the receptor [7] and that apoE-deficiency negatively affects LRP1 mediated clearance of CR-K1 in osteoblasts [33]. It is known that there exists a certain degree of functional redundancy between LDLR and LRP1 at the level of the liver [6]. Both via immunofluorescence (Figure 3C) and immunoblotting (Figure 3D–E) a significant increase in the expression levels of LDLR in hepatocytes (Figure 3C–D) and liver extracts (showing a 1.6-fold increase) (Figure 3E) could be observed between apoE−/−LRP1n2/n2 and apoE−/− mice. These results suggest that LRP1 NPxYxxL-motif inactivation leads to a compensatory up-regulation of the LDLR, which could be responsible for improved CR clearance via hepatocytes.

Bottom Line: Postprandial lipoprotein improvement was explained by increased hepatic clearance of triglyceride-rich remnant lipoproteins and accompanied by a compensatory 1.6-fold upregulation of LDLR expression in hepatocytes.These findings demonstrate that the NPxYxxL motif in LRP1 is important for insulin-mediated translocation and slow perinuclear endosomal recycling.These LRP1 impairments correlated with reduced atherogenesis and cholesterol levels in apoE-deficient mice, likely via compensatory LDLR upregulation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Experimental Mouse Genetics, Center for Human Genetics, KU Leuven, Leuven, Belgium.

ABSTRACT

Objective: Determination of the in vivo significance of LDL receptor-related protein 1 (LRP1) dysfunction on lipid metabolism and atherosclerosis development in absence of its main ligand apoE.

Methods and results: LRP1 knock-in mice carrying an inactivating mutation in the NPxYxxL motif were crossed with apoE-deficient mice. In the absence of apoE, relative to LRP1 wild-type animals, LRP1 mutated mice showed an increased clearance of postprandial lipids despite a compromised LRP1 endocytosis rate and inefficient insulin-mediated translocation of the receptor to the plasma membrane, likely due to inefficient slow recycling of the mutated receptor. Postprandial lipoprotein improvement was explained by increased hepatic clearance of triglyceride-rich remnant lipoproteins and accompanied by a compensatory 1.6-fold upregulation of LDLR expression in hepatocytes. One year-old apoE-deficient mice having the dysfunctional LRP1 revealed a 3-fold decrease in spontaneous atherosclerosis development and a 2-fold reduction in LDL-cholesterol levels.

Conclusion: These findings demonstrate that the NPxYxxL motif in LRP1 is important for insulin-mediated translocation and slow perinuclear endosomal recycling. These LRP1 impairments correlated with reduced atherogenesis and cholesterol levels in apoE-deficient mice, likely via compensatory LDLR upregulation.

Show MeSH
Related in: MedlinePlus