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Macrophage-specific apoE gene repair reduces diet-induced hyperlipidemia and atherosclerosis in hypomorphic Apoe mice.

Gaudreault N, Kumar N, Olivas VR, Eberlé D, Rapp JH, Raffai RL - PLoS ONE (2012)

Bottom Line: Although the liver is the major source of plasma apoE, extra-hepatic sources of apoE, including from macrophages, account for up to 10% of plasma apoE levels.This difference in atherosclerosis lesions size was proportional to the observed reduction in plasma cholesterol.Macrophage-derived apoE raises plasma apoE levels in response to diet-induced hyperlipidemia and by such reduces atherosclerosis proportionally to the extent to which it lowers plasma cholesterol levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, University of California San Francisco, VA Medical Center, San Francisco, California, United States of America.

ABSTRACT

Background: Apolipoprotein (apo) E is best known for its ability to lower plasma cholesterol and protect against atherosclerosis. Although the liver is the major source of plasma apoE, extra-hepatic sources of apoE, including from macrophages, account for up to 10% of plasma apoE levels. This study examined the contribution of macrophage-derived apoE expression levels in diet-induced hyperlipidemia and atherosclerosis.

Methodology/principal findings: Hypomorphic apoE (Apoe(h/h)) mice expressing wildtype mouse apoE at ∼2-5% of physiological levels in all tissues were derived by gene targeting in embryonic stem cells. Cre-mediated gene repair of the Apoe(h/h) allele in Apoe(h/h)LysM-Cre mice raised apoE expression levels by 26 fold in freshly isolated peritoneal macrophages, restoring it to 37% of levels seen in wildtype mice. Chow-fed Apoe(h/h)LysM-Cre and Apoe(h/h) mice displayed similar plasma apoE and cholesterol levels (55.53±2.90 mg/dl versus 62.70±2.77 mg/dl, n = 12). When fed a high-cholesterol diet (HCD) for 16 weeks, Apoe(h/h)LysM-Cre mice displayed a 3-fold increase in plasma apoE and a concomitant 32% decrease in plasma cholesterol when compared to Apoe(h/h) mice (602.20±22.30 mg/dl versus 888.80±24.99 mg/dl, n = 7). On HCD, Apoe(h/h)LysM-Cre mice showed increased apoE immunoreactivity in lesional macrophages and liver-associated Kupffer cells but not hepatocytes. In addition, Apoe(h/h)LysM-Cre mice developed 35% less atherosclerotic lesions in the aortic root than Apoe(h/h) mice (167×10(3)±16×10(3) µm(2) versus 259×10(3)±56×10(3) µm(2), n = 7). This difference in atherosclerosis lesions size was proportional to the observed reduction in plasma cholesterol.

Conclusions/significance: Macrophage-derived apoE raises plasma apoE levels in response to diet-induced hyperlipidemia and by such reduces atherosclerosis proportionally to the extent to which it lowers plasma cholesterol levels.

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Related in: MedlinePlus

Conditional repair of the hypomorphic Apoeh/h allele.Hypomorphic Apoe gene expression and gene repair strategies (A). Relative Apoe gene expression levels in peritoneal macrophages collected from both groups of mice (n = 4; B). ApoE secreted by peritoneal macrophages in the presence and absence of LXR agonist pre-treatment was quantified (n = 3; C) from Western Blot (D), mean±sem, **p<0.01, ***p<0.001; control is media without macrophages.
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pone-0035816-g001: Conditional repair of the hypomorphic Apoeh/h allele.Hypomorphic Apoe gene expression and gene repair strategies (A). Relative Apoe gene expression levels in peritoneal macrophages collected from both groups of mice (n = 4; B). ApoE secreted by peritoneal macrophages in the presence and absence of LXR agonist pre-treatment was quantified (n = 3; C) from Western Blot (D), mean±sem, **p<0.01, ***p<0.001; control is media without macrophages.

Mentions: The insertion of a neomycin (neo) resistance cassette flanked by loxP sites into intron 3 of the WT Apoe allele resulted in the attenuated expression of apoE as previously described in the apoE4-like Arg-61 Apoeh/h strain of mice [28]. The hypomorphic effect observed in Apoeh/h mice presumably arises from an aberrant mRNA splicing between Apoe exon 3 and the neo cassette (Fig. 1a). Removal of the neomycin cassette in Arg-61 Apoeh/h mice by Cre-mediated gene repair permanently restores normal plasma apoE levels in plasma [28]. In this study, Apoeh/h mice were bred to LysM-Cre transgenic mice that restricts the expression of Cre recombinase to the myeloid cell lineage, including mature macrophages [29].


Macrophage-specific apoE gene repair reduces diet-induced hyperlipidemia and atherosclerosis in hypomorphic Apoe mice.

Gaudreault N, Kumar N, Olivas VR, Eberlé D, Rapp JH, Raffai RL - PLoS ONE (2012)

Conditional repair of the hypomorphic Apoeh/h allele.Hypomorphic Apoe gene expression and gene repair strategies (A). Relative Apoe gene expression levels in peritoneal macrophages collected from both groups of mice (n = 4; B). ApoE secreted by peritoneal macrophages in the presence and absence of LXR agonist pre-treatment was quantified (n = 3; C) from Western Blot (D), mean±sem, **p<0.01, ***p<0.001; control is media without macrophages.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3351426&req=5

pone-0035816-g001: Conditional repair of the hypomorphic Apoeh/h allele.Hypomorphic Apoe gene expression and gene repair strategies (A). Relative Apoe gene expression levels in peritoneal macrophages collected from both groups of mice (n = 4; B). ApoE secreted by peritoneal macrophages in the presence and absence of LXR agonist pre-treatment was quantified (n = 3; C) from Western Blot (D), mean±sem, **p<0.01, ***p<0.001; control is media without macrophages.
Mentions: The insertion of a neomycin (neo) resistance cassette flanked by loxP sites into intron 3 of the WT Apoe allele resulted in the attenuated expression of apoE as previously described in the apoE4-like Arg-61 Apoeh/h strain of mice [28]. The hypomorphic effect observed in Apoeh/h mice presumably arises from an aberrant mRNA splicing between Apoe exon 3 and the neo cassette (Fig. 1a). Removal of the neomycin cassette in Arg-61 Apoeh/h mice by Cre-mediated gene repair permanently restores normal plasma apoE levels in plasma [28]. In this study, Apoeh/h mice were bred to LysM-Cre transgenic mice that restricts the expression of Cre recombinase to the myeloid cell lineage, including mature macrophages [29].

Bottom Line: Although the liver is the major source of plasma apoE, extra-hepatic sources of apoE, including from macrophages, account for up to 10% of plasma apoE levels.This difference in atherosclerosis lesions size was proportional to the observed reduction in plasma cholesterol.Macrophage-derived apoE raises plasma apoE levels in response to diet-induced hyperlipidemia and by such reduces atherosclerosis proportionally to the extent to which it lowers plasma cholesterol levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, University of California San Francisco, VA Medical Center, San Francisco, California, United States of America.

ABSTRACT

Background: Apolipoprotein (apo) E is best known for its ability to lower plasma cholesterol and protect against atherosclerosis. Although the liver is the major source of plasma apoE, extra-hepatic sources of apoE, including from macrophages, account for up to 10% of plasma apoE levels. This study examined the contribution of macrophage-derived apoE expression levels in diet-induced hyperlipidemia and atherosclerosis.

Methodology/principal findings: Hypomorphic apoE (Apoe(h/h)) mice expressing wildtype mouse apoE at ∼2-5% of physiological levels in all tissues were derived by gene targeting in embryonic stem cells. Cre-mediated gene repair of the Apoe(h/h) allele in Apoe(h/h)LysM-Cre mice raised apoE expression levels by 26 fold in freshly isolated peritoneal macrophages, restoring it to 37% of levels seen in wildtype mice. Chow-fed Apoe(h/h)LysM-Cre and Apoe(h/h) mice displayed similar plasma apoE and cholesterol levels (55.53±2.90 mg/dl versus 62.70±2.77 mg/dl, n = 12). When fed a high-cholesterol diet (HCD) for 16 weeks, Apoe(h/h)LysM-Cre mice displayed a 3-fold increase in plasma apoE and a concomitant 32% decrease in plasma cholesterol when compared to Apoe(h/h) mice (602.20±22.30 mg/dl versus 888.80±24.99 mg/dl, n = 7). On HCD, Apoe(h/h)LysM-Cre mice showed increased apoE immunoreactivity in lesional macrophages and liver-associated Kupffer cells but not hepatocytes. In addition, Apoe(h/h)LysM-Cre mice developed 35% less atherosclerotic lesions in the aortic root than Apoe(h/h) mice (167×10(3)±16×10(3) µm(2) versus 259×10(3)±56×10(3) µm(2), n = 7). This difference in atherosclerosis lesions size was proportional to the observed reduction in plasma cholesterol.

Conclusions/significance: Macrophage-derived apoE raises plasma apoE levels in response to diet-induced hyperlipidemia and by such reduces atherosclerosis proportionally to the extent to which it lowers plasma cholesterol levels.

Show MeSH
Related in: MedlinePlus