We report the engineering and characterization of paraoxonase-3 knockout mice (Pon3KO). The mice were generally healthy but exhibited quantitative alterations in bile acid metabolism and a 37% increased body weight compared to the wild-type mice on a high fat diet. PON3 was enriched in the mitochondria-associated membrane fraction of hepatocytes. PON3 deficiency resulted in impaired mitochondrial respiration, increased mitochondrial superoxide levels, and increased hepatic expression of inflammatory genes. PON3 deficiency did not influence atherosclerosis development on an apolipoprotein E null hyperlipidemic background, but it did lead to a significant 60% increase in atherosclerotic lesion size in Pon3KO mice on the C57BL/6J background when fed a cholate-cholesterol diet. On the diet, the Pon3KO had significantly increased plasma intermediate-density lipoprotein/LDL cholesterol and bile acid levels. They also exhibited significantly elevated levels of hepatotoxicity markers in circulation, a 58% increase in gallstone weight, a 40% increase in hepatic cholesterol level, and increased mortality. Furthermore, Pon3KO mice exhibited decreased hepatic bile acid synthesis and decreased bile acid levels in the small intestine compared with wild-type mice. Our study suggests a role for PON3 in the metabolism of lipid and bile acid as well as protection against atherosclerosis, gallstone disease, and obesity.

To identify genetic and environmental factors contributing to the pathogenesis of nonalcoholic fatty liver disease, we examined liver steatosis and related clinical and molecular traits in more than 100 unique inbred mouse strains, which were fed a diet rich in fat and carbohydrates. A > 30-fold variation in hepatic TG accumulation was observed among the strains. Genome-wide association studies revealed three loci associated with hepatic TG accumulation. Utilizing transcriptomic data from the liver and adipose tissue, we identified several high-confidence candidate genes for hepatic steatosis, including Gde1, a glycerophosphodiester phosphodiesterase not previously implicated in triglyceride metabolism. We confirmed the role of Gde1 by in vivo hepatic over-expression and shRNA knockdown studies. We hypothesize that Gde1 expression increases TG production by contributing to the production of glycerol-3-phosphate. Our multi-level data, including transcript levels, metabolite levels, and gut microbiota composition, provide a framework for understanding genetic and environmental interactions underlying hepatic steatosis.

Insulin resistance (IR) is a complex trait with multiple genetic and environmental components. Confounded by large differences between the sexes, environment, and disease pathology, the genetic basis of IR has been difficult to dissect. Here we examine IR and related traits in a diverse population of more than 100 unique male and female inbred mouse strains after feeding a diet rich in fat and refined carbohydrates. Our results show dramatic variation in IR among strains of mice and widespread differences between sexes that are dependent on genotype. We uncover more than 15 genome-wide significant loci and validate a gene, Agpat5, associated with IR. We also integrate plasma metabolite levels and global gene expression from liver and adipose tissue to identify metabolite quantitative trait loci (mQTL) and expression QTL (eQTL), respectively. Our results provide a resource for analysis of interactions between diet, sex, and genetic background in IR.

Autocrine VEGF is necessary for endothelial survival, although the cellular mechanisms supporting this function are unknown. Here, we show that – even after full differentiation and maturation – continuous expression of VEGF by endothelial cells is needed to sustain vascular integrity and cellular viability. Depletion of VEGF from the endothelium results in mitochondria fragmentation and suppression ofglucosemetabolism,leadingtoincreasedautophagythatcontributes to cell death. Gene-expression profiling showed that endothelial VEGF contributes to the regulation of cell cycle and mitochondrial gene clusters, as well as several – but not all – targets of the transcription factor FOXO1. Indeed, VEGF-deficient endothelium in vitro and in vivo showed increased levels of FOXO1 protein in the nucleus and cytoplasm. Silencing of FOXO1 in VEGF-depleted cells reversed expression profiles of several of the gene clusters that were de-regulated in VEGF knockdown, and rescued both cell death and autophagy phenotypes. Our data suggest that endothelial VEGF maintains vascular homeostasis through regulation of FOXO1 levels, thereby ensuring physiological metabolism and endothelial cell survival.

The CCAAT/enhancer binding proteins (C/EBPs) are transcription factors involved in hematopoietic cell development and induction of several inflammatory mediators. C/EBPε is expressed only in myeloid cells including monocytes/macrophages. Atherosclerosis is an inflammatory disorder of the vascular wall and circulating immune cells such as monocytes/macrophages. Mice deficient in the low density lipoprotein (LDL) receptor (Ldlr−/−) fed on a high cholesterol diet (HCD) show elevated blood cholesterol levels and are widely used as models to study human atherosclerosis. In this study, we generated Ldlr and Cebpe double-knockout (llee) mice and compared their atherogenic phenotypes to Ldlr single deficient (llEE) mice after HCD. Macrophages from llee mice have reduced lipid uptake by foam cells and impaired phagokinetic motility in vitro compared to macrophages from llEE mice. Also, compared to llEE mice, llee mice have alterations of lipid metabolism, and reduced atheroma and obesity, particularly the males. Peritoneal macrophages of llee male mice have reduced mRNA expression of FABP4, a fatty acid binding protein implicated in atherosclerosis. Overall, our study suggests that the myeloid specific factor C/EBPε is involved in systemic lipid metabolism and that silencing of C/EBPε could decrease the development of atherosclerosis.

HomeCirculation ResearchVol. 112, No. 11Response to Pomozi et al's Research Commentary Free AccessLetterPDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessLetterPDF/EPUBResponse to Pomozi et al's Research Commentary Lisa J. Martin Edward Lau Harpreet Singh Laurent Vergnes Elizabeth J. Tarling, Margarete Mehrabian, Imran Mungrue, Sheila Xiao and Diana Shih Lawrence Castellani Peipei Ping Karen Reue Enrico Stefani Thomas A. Drake Kristina Bostrom Aldons J. Lusis Lisa J. MartinLisa J. Martin Department of Medicine Edward LauEdward Lau Departments of Medicine and Physiology Harpreet SinghHarpreet Singh Department of Anesthesiology Laurent VergnesLaurent Vergnes Department of Human Genetics Elizabeth J. TarlingElizabeth J. Tarling , Margarete MehrabianMargarete Mehrabian , Imran MungrueImran Mungrue , Sheila XiaoSheila Xiao and Diana ShihDiana Shih Lawrence CastellaniLawrence Castellani Department of Medicine Peipei PingPeipei Ping Departments of Medicine and Physiology Karen ReueKaren Reue Department of Human GeneticsMolecular Biology Institute Enrico StefaniEnrico Stefani Department of Anesthesiology Thomas A. DrakeThomas A. Drake Department of Pathology and Laboratory Medicine Kristina BostromKristina Bostrom Department of Medicine Aldons J. LusisAldons J. Lusis Departments of Medicine, Human Genetics, and Microbiology, Immunology and Molecular GeneticsDavid Geffen School of Medicine University of CaliforniaLos Angeles, CA Originally published26 Apr 2013https://doi.org/10.1161/CIRCRESAHA.113.301666Circulation Research. 2013;112:e152–e153Other version(s) of this articleYou are viewing the most recent version of this article. Previous versions: January 1, 2013: Previous Version 1 To the EditorIn the course of studying the effects of ABCC6 deficiency in mice, we observed an enrichment of mitochondrial gene expression signatures. In subsequent studies, we found that ABCC6 null mice exhibited abnormal mitochondrial morphology and functional mitochondrial deficiencies.1 We then carried out subcellular fractionation studies, indicating that ABCC6 colocalized with markers of the mitochondria-associated membranes in mouse liver and kidney.1 Our results differed from Le Saux et al2 from the Varadi group, who had concluded in a recent publication that ABCC6 resided in the plasma membrane. To test whether ABCC6 was localized in plasma membrane, we performed cell surface protein biotin labeling experiments, which were negative for ABCC6.1See Research Commentary, p e148Pomozi et al3 of the Varadi group have now challenged our conclusions based, as in their previous report,2 on immunofluorescent imaging of frozen liver sections and cells in culture showing peripheral cellular localization of antibody binding. They argue that cell disruption and subcellular fractionation in our study1 may have resulted in artifactual associations of membrane proteins. However, this seems improbable, given that the plasma membrane markers that we examined did not fractionate with ABCC6, and mitochondria-associated membranes constitute a very small fraction of the total membranes.1 Cellular fractionation techniques have been used almost universally to provide definitive evidence of subcellular localization.4,5 Pomozi et al3 also argue that ABCC6, lacks sufficient amine groups on the extracellular surface to allow biotin labeling. This possibility cannot be excluded, although the manufacturer of the surface protein biotin labeling assay (Thermo Pierce) indicates that it is sensitive to even a single amine group, which would be present in the N terminus or in one of the predicted available lysines.In our studies, we used the N-terminal binding (S-20) antibody from the same commercial supplier (Santa Cruz Biotechnology) as reported by Pomozi et al.3 In our hands, this antibody exhibited significant nonspecific binding in ABCC6 null mouse tissue sections (not shown) and in Western blots.1 We note that in their publication,2 the methods state that the S-20 antibody is made in rabbit and blocking was performed using goat serum. In fact, the antibody is made in goat and blocking with goat serum would produce nonspecific labeling (per Santa Cruz Biotechnology). An advantage of our results obtained by subcellular fractionation compared with immunofluorescence of tissue sections is that, in the former, the protein is detected on Western blots after separation by gel electrophoresis, thus allowing better separation from cross-reacting proteins. A lack of accompanying Western data in the current challenge, in our opinion, significantly weakens the ability to ascertain the specificity of the signal.In conclusion, we recognize that localization of proteins can be challenging and is highly dependent on the quality of the antibody reagents. Although we cannot exclude the possibility that some of the ABCC6 protein resides on the plasma membrane, low-resolution imaging studies of frozen liver sections and cells in culture do not provide convincing evidence against the localization of ABCC6 in the mitochondria-associated membranes.Lisa J. MartinDepartment of MedicineEdward LauDepartments of Medicine and PhysiologyHarpreet SinghDepartment of AnesthesiologyLaurent VergnesDepartment of Human GeneticsElizabeth J. TarlingMargarete MehrabianImran MungrueSheila XiaoDiana ShihLawrence CastellaniDepartment of MedicinePeipei PingDepartments of Medicine and PhysiologyKaren ReueDepartment of Human GeneticsMolecular Biology InstituteEnrico StefaniDepartment of AnesthesiologyThomas A. DrakeDepartment of Pathology and Laboratory MedicineKristina BostromDepartment of MedicineAldons J. LusisDepartments of Medicine, Human Genetics, andMicrobiology, Immunology and Molecular GeneticsDavid Geffen School of Medicine University of CaliforniaLos Angeles, CASources of FundingThis work was supported by funding from the NIH (HL30568, S10RR026744, HL088640, and HHSN268201000035C) and American Heart Association (11SDG7230059, 11POST7300060, and 12PRE11610024).DisclosuresNone.FootnotesLetters to the Editor will be published, if suitable, as space permits. They should not exceed 1000 words (typed double-spaced) in length and may be subject to editing or abridgment.References1. Martin LJ, Lau E, Singh H, et al. ABCC6 localizes to the mitochondria-associated membrane.Circ Res. 2012; 111:516–520.LinkGoogle Scholar2. Le Saux O, Fülöp K, Yamaguchi Y, Iliás A, Szabó Z, Brampton CN, Pomozi V, Huszár K, Arányi T, Váradi A. Expression and in vivo rescue of human ABCC6 disease-causing mutants in mouse liver.PLoS One. 2011; 6:e24738.CrossrefMedlineGoogle Scholar3. Pomozi V, Le Saux O, Brampton CN, Apana ALY, Iliás A, Szeri F, Martin L, Monostory K, Paku S, Sarkadi B, Szakács G, Varadi A. ABCC6 is a basolateral plasma membrane protein.Circ Res. 2013; 112. DOI: 10.1161/CIRCRESAHA.111.300194LinkGoogle Scholar4. Huber LA, Pfaller K, Vietor I. Organelle proteomics: implications for subcellular fractionation in proteomics.Circ Res. 2003; 92:962–968.LinkGoogle Scholar5. Rockstroh M, Müller S, Jende C, Kerzhner A, von Bergen M, Tomm JM. Cell fractionation: an important tool for compartment proteomics.JIntegrated OMICS. 2011; 1:135–143.Google Scholar Previous Back to top Next FiguresReferencesRelatedDetailsCited By Borst P, Váradi A and van de Wetering K (2019) PXE, a Mysterious Inborn Error Clarified, Trends in Biochemical Sciences, 10.1016/j.tibs.2018.10.005, 44:2, (125-140), Online publication date: 1-Feb-2019. Ferré M, Reynier P, Chevrollier A, Prunier-Mirebeau D, Lefthériotis G, Henrion D, Bonneau D, Procaccio V and Martin L (2013) Is ABCC6 a genuine mitochondrial protein?, BMC Research Notes, 10.1186/1756-0500-6-427, 6:1, Online publication date: 1-Dec-2013. May 24, 2013Vol 112, Issue 11 Advertisement Article InformationMetrics © 2013 American Heart Association, Inc.https://doi.org/10.1161/CIRCRESAHA.113.301666PMID: 23625949 Originally publishedApril 26, 2013 PDF download Advertisement SubjectsVascular Biology

Obesity is a highly heritable disease driven by complex interactions between genetic and environmental factors. Human genome-wide association studies (GWAS) have identified a number of loci contributing to obesity; however, a major limitation of these studies is the inability to assess environmental interactions common to obesity. Using a systems genetics approach, we measured obesity traits, global gene expression, and gut microbiota composition in response to a high-fat/high-sucrose (HF/HS) diet of more than 100 inbred strains of mice. Here we show that HF/HS feeding promotes robust, strain-specific changes in obesity that are not accounted for by food intake and provide evidence for a genetically determined set point for obesity. GWAS analysis identified 11 genome-wide significant loci associated with obesity traits, several of which overlap with loci identified in human studies. We also show strong relationships between genotype and gut microbiota plasticity during HF/HS feeding and identify gut microbial phylotypes associated with obesity.

BACKGROUND:The human 9p21.3 chromosome locus has been shown to be an independent risk factor for atherosclerosis in multiple large-scale genome-wide association studies, but the underlying mechanism remains unknown. We set out to investigate the potential role of the 9p21.3 locus neighboring genes, including Mtap, the 2 isoforms of Cdkn2a, p16Ink4a and p19Arf, and Cdkn2b, in atherosclerosis using knockout mice models.METHODS AND RESULTS:Gene-targeted mice for neighboring genes, including Mtap, Cdkn2a, p19Arf, and Cdkn2b, were each bred to mice carrying the human APO*E3 Leiden transgene that sensitizes the mice for atherosclerotic lesions through elevated plasma cholesterol. We found that the mice heterozygous for Mtap developed larger lesions compared with wild-type mice (49623±21650 versus 18899±9604 μm(2) per section [mean±SD]; P=0.01), with morphology similar to that of wild-type mice. The Mtap heterozygous mice demonstrated changes in metabolic and methylation profiles and CD4(+) cell counts. The Cdkn2a knockout mice had smaller lesions compared with wild-type and heterozygous mice, and there were no significant differences in lesion size in p19Arf and Cdkn2b mutants compared with wild type. We observed extensive, tissue-specific compensatory regulation of the Cdkn2a and Cdkn2b genes among the various knockout mice, making the effects on atherosclerosis difficult to interpret.CONCLUSIONS:Mtap plays a protective role against atherosclerosis, whereas Cdkn2a appears to be modestly proatherogenic. However, no relation was found between the 9p21 genotype and the transcription of 9p21 neighboring genes in primary human aortic vascular cells in vitro. There is extensive compensatory regulation in the highly conserved 9p21 orthologous region in mice.

The liver X receptor (LXR) signaling pathway is an important modulator of atherosclerosis, but the relative importance of the two LXRs in atheroprotection is incompletely understood. We show here that LXRα, the dominant LXR isotype expressed in liver, plays a particularly important role in whole-body sterol homeostasis. In the context of the ApoE−/− background, deletion of LXRα, but not LXRβ, led to prominent increases in atherosclerosis and peripheral cholesterol accumulation. However, combined loss of LXRα and LXRβ on the ApoE−/− background led to an even more severe cholesterol accumulation phenotype compared to LXRα−/−ApoE−/− mice, indicating that LXRβ does contribute to reverse cholesterol transport (RCT) but that this contribution is quantitatively less important than that of LXRα. Unexpectedly, macrophages did not appear to underlie the differential phenotype of LXRα−/−ApoE−/− and LXRβ−/−ApoE−/− mice, as in vitro assays revealed no difference in the efficiency of cholesterol efflux from isolated macrophages. By contrast, in vivo assays of RCT using exogenously labeled macrophages revealed a marked defect in fecal sterol efflux in LXRα−/−ApoE−/− mice. Mechanistically, this defect was linked to a specific requirement for LXRα−/− in the expression of hepatic LXR target genes involved in sterol transport and metabolism. These studies reveal a previously unrecognized requirement for hepatic LXRα for optimal reverse cholesterol transport in mice.

Rationale: Mutations of the orphan transporter ABCC6 (ATP-binding cassette, subfamily C, member 6) cause the connective tissue disorder pseudoxanthoma elasticum. ABCC6 was thought to be located on the plasma membrane of liver and kidney cells. Objective: Mouse systems genetics and bioinformatics suggested that ABCC6 deficiency affects mitochondrial gene expression. We therefore tested whether ABCC6 associates with mitochondria. Methods and Results: We found ABCC6 in crude mitochondrial fractions and subsequently pinpointed its localization to the purified mitochondria-associated membrane fraction. Cell-surface biotinylation in hepatocytes confirmed that ABCC6 is intracellular. Abcc6-knockout mice demonstrated mitochondrial abnormalities and decreased respiration reserve capacity. Conclusions: Our finding that ABCC6 localizes to the mitochondria-associated membrane has implications for its mechanism of action in normal and diseased states.

Inbred strains of mice are strikingly different in susceptibility to obesity-driven diabetes. For instance, deficiency in leptin receptor (db/db) leads to hyperphagia and obesity in both C57BL/6 and DBA/2 mice, but only on the DBA/2 background do the mice develop beta-cell loss leading to severe diabetes, while C57BL/6 mice are relatively resistant. To further investigate the genetic factors predisposing to diabetes, we have studied leptin receptor-deficient offspring of an F2 cross between C57BL/6J (db/+) males and DBA/2J females. The results show that the genetics of diabetes susceptibility are enormously complex and a number of quantitative trait loci (QTL) contributing to diabetes-related traits were identified, notably on chromosomes 4, 6, 7, 9, 10, 11, 12, and 19. The Chr. 4 locus is likely due to a disruption of the Zfp69 gene in C57BL/6J mice. To identify candidate genes and to model coexpression networks, we performed global expression array analysis in livers of the F2 mice. Expression QTL (eQTL) were identified and used to prioritize candidate genes at clinical trait QTL. In several cases, clusters of eQTLs colocalized with clinical trait QTLs, suggesting a common genetic basis. We constructed coexpression networks for both 5 and 12 wk old mice and identified several modules significantly associated with clinical traits. One module in 12 wk old mice was associated with several measures of hepatic fat content as well as with other lipid- and diabetes-related traits. These results add to the understanding of the complex genetic interactions contributing to obesity-induced diabetes.

Epidemiological studies show that high HDL-cholesterol (HDLc) decreases the risk of cardiovascular disease. To map genes controlling lipid metabolism, particularly HDLc levels, we screened the plasma lipids of 36 AcB/BcA RC mouse strains subjected to either a normal or a high-fat/cholesterol diet. Strains BcA68 and AcB65 showed deviant HDLc plasma levels compared with the parental A/J and C57BL/6J strains; they were thus selected to generate informative F2 crosses. Linkage analyses in the AcB65 strain identified a locus on chromosome 4 ( Hdlq78) responsible for high post-high fat diet HDLc levels. This locus has been previously associated at genome-wide significance to two regions in the human genome. A second linkage analysis in strain BcA68 identified linkage in the vicinity of a gene cluster known to control HDLc levels. Sequence analysis of these candidates identified a de novo, loss-of-function mutation in the ApoA1 gene of BcA68 that prematurely truncates the ApoA1 protein. The possibility of dissecting the specific effects of this new ApoA1 deficiency in the context of isogenic controls makes the BcA68 mouse a valuable new tool.

Objective— To test the hypothesis that NF-E2–related factor 2 (Nrf2) expression plays an antiatherogenic role by its vascular antioxidant and anti-inflammatory properties. Methods and Results— Nrf2 is an important transcription factor that regulates the expression of phase 2 detoxifying enzymes and antioxidant genes. Its expression in vascular cells appears to be an important factor in the protection against vascular oxidative stress and inflammation. We developed Nrf2 heterozygous (HET) and homozygous knockout (KO) mice on an apolipoprotein (apo) E–null background by sequential breeding, resulting in Nrf2 −/− , apoE −/− (KO), Nrf2 −/+ , apoE −/− (HET) and Nrf2 +/+ , and apoE −/− wild-type littermates. KO mice exhibited decreased levels of antioxidant genes with evidence of increased reactive oxygen species generation compared with wild-type controls. Surprisingly, KO males exhibited 47% and 53% reductions in the degree of aortic atherosclerosis compared with HET or wild-type littermates, respectively. Decreased atherosclerosis in KO mice correlated with lower plasma total cholesterol in a sex-dependent manner. KO mice also had a decreased hepatic cholesterol content and a lower expression of lipogenic genes, suggesting that hepatic lipogenesis could be reduced. In addition, KO mice exhibited atherosclerotic plaques characterized by a lesser macrophage component and decreased foam cell formation in an in vitro lipid-loading assay. This was associated with a lower rate of cholesterol influx, mediated in part by decreased expression of the scavenger receptor CD36. Conclusion— Nrf2 expression unexpectedly promotes atherosclerotic lesion formation in a sex-dependent manner, most likely by a combination of systemic metabolic and local vascular effects.

We report a systems genetic analysis of high density lipoprotein (HDL) levels in an F2 intercross between inbred strains CAST/EiJ and C57BL/6J. We previously showed that there are dramatic differences in HDL metabolism in a cross between these strains, and we now report co-expression network analysis of HDL that integrates global expression data from liver and adipose with relevant metabolic traits. Using data from a total of 293 F2 intercross mice, we constructed weighted gene co-expression networks and identified modules (subnetworks) associated with HDL and clinical traits. These were examined for genes implicated in HDL levels based on large human genome-wide associations studies (GWAS) and examined with respect to conservation between tissue and sexes in a total of 9 data sets. We identify genes that are consistently ranked high by association with HDL across the 9 data sets. We focus in particular on two genes, Wfdc2 and Hdac3, that are located in close proximity to HDL QTL peaks where causal testing indicates that they may affect HDL. Our results provide a rich resource for studies of complex metabolic interactions involving HDL. This article is part of a Special Issue entitled Advances in High Density Lipoprotein Formation and Metabolism: A Tribute to John F. Oram (1945-2010).

Systems genetics relies on common genetic variants to elucidate biologic networks contributing to complex disease-related phenotypes. Mice are ideal model organisms for such approaches, but linkage analysis has been only modestly successful due to low mapping resolution. Association analysis in mice has the potential of much better resolution, but it is confounded by population structure and inadequate power to map traits that explain less than 10% of the variance, typical of mouse quantitative trait loci (QTL). We report a novel strategy for association mapping that combines classic inbred strains for mapping resolution and recombinant inbred strains for mapping power. Using a mixed model algorithm to correct for population structure, we validate the approach by mapping over 2500 cis -expression QTL with a resolution an order of magnitude narrower than traditional QTL analysis. We also report the fine mapping of metabolic traits such as plasma lipids. This resource, termed the Hybrid Mouse Diversity Panel, makes possible the integration of multiple data sets and should prove useful for systems-based approaches to complex traits and studies of gene-by-environment interactions.

Background— We previously mapped a quantitative trait locus on chromosome 15 in mice contributing to high-density lipoprotein cholesterol and triglyceride levels and now report the identification of the underlying gene. Methods and Results— We first fine-mapped the locus by studying a series of congenic strains derived from the parental strains BALB/cJ and MRL/MpJ. Analysis of gene expression and sequencing followed by transgenic complementation led to the identification of zinc fingers and homeoboxes 2 (Zhx2), a transcription factor previously implicated in the developmental regulation of α-fetoprotein. Reduced expression of the protein in BALB/cJ mice resulted in altered hepatic transcript levels for several genes involved in lipoprotein metabolism. Most notably, the Zhx2 mutation resulted in a failure to suppress expression of lipoprotein lipase, a gene normally silenced in the adult liver, and this was normalized in BALB/cJ mice carrying the Zhx2 transgene. Conclusions— We identified the gene underlying the chromosome 15 quantitative trait locus, and our results show that Zhx2 functions as a novel developmental regulator of key genes influencing lipoprotein metabolism.

INTRODUCTION:The purpose of this study was to evaluate the effects of L-4F, an apolipoprotein A-1 mimetic peptide, alone or with pravastatin, in apoE-/-Fas-/-C57BL/6 mice that spontaneously develop immunoglobulin G (IgG) autoantibodies, glomerulonephritis, osteopenia, and atherosclerotic lesions on a normal chow diet.METHODS:Female mice, starting at eight to nine weeks of age, were treated for 27 weeks with 1) pravastatin, 2) L-4F, 3) L-4F plus pravastatin, or 4) vehicle control, followed by disease phenotype assessment.RESULTS:In preliminary studies, dysfunctional, proinflammatory high-density lipoproteins (piHDL) were decreased six hours after a single L-4F, but not scrambled L-4F, injection in eight- to nine-week old mice. After 35 weeks, L-4F-treated mice, in the absence/presence of pravastatin, had significantly smaller lymph nodes and glomerular tufts (PL, LP<0.05), lower serum levels of IgG antibodies to double stranded DNA (dsDNA) (PL<0.05) and oxidized phospholipids (oxPLs) (PL, LP<0.005), and elevated total and vertebral bone mineral density (PL, LP<0.01) compared to vehicle controls. Although all treatment groups presented larger aortic root lesions compared to vehicle controls, enlarged atheromas in combination treatment mice had significantly less infiltrated CD68+ macrophages (PLP<0.01), significantly increased mean alpha-actin stained area (PLP<0.05), and significantly lower levels of circulating markers for atherosclerosis progression, CCL19 (PL, LP<0.0005) and VCAM-1 (PL<0.0002).CONCLUSIONS:L-4F treatment, alone or with pravastatin, significantly reduced IgG anti-dsDNA and IgG anti-oxPLs, proteinuria, glomerulonephritis, and osteopenia in a murine lupus model of accelerated atherosclerosis. Despite enlarged aortic lesions, increased smooth muscle content, decreased macrophage infiltration, and decreased pro-atherogenic chemokines in L-4F plus pravastatin treated mice suggest protective mechanisms not only on lupus-like disease, but also on potential plaque remodeling in a murine model of systemic lupus erythematosus (SLE) and accelerated atherosclerosis.

Objective— The risk of atherosclerosis in the setting of chylomicronemia has been a topic of debate. In this study, we examined susceptibility to atherosclerosis in Gpihbp1 -deficient mice ( Gpihbp1 −/− ), which manifest severe chylomicronemia as a result of defective lipolysis. Methods and Results— Gpihbp1 −/− mice on a chow diet have plasma triglyceride and cholesterol levels of 2812±209 and 319±27 mg/dL, respectively. Even though nearly all of the lipids were contained in large lipoproteins (50 to 135 nm), the mice developed progressive aortic atherosclerosis. In other experiments, we found that both Gpihbp1 -deficient “apo-B48–only” mice and Gpihbp1 -deficient “apo-B100–only” mice manifest severe chylomicronemia. Thus, GPIHBP1 is required for the processing of both apo-B48– and apo-B100–containing lipoproteins. Conclusions— Chylomicronemia causes atherosclerosis in mice. Also, we found that GPIHBP1 is required for the lipolytic processing of both apo-B48– and apo-B100–containing lipoproteins.

OBJECTIVE To identify metabolic derangements contributing to diabetes susceptibility in the leptin receptor–deficient obese C57BLKS/J-db/db (BKS-db) mouse strain. RESEARCH DESIGN AND METHODS Young BKS-db mice were used to identify metabolic pathways contributing to the development of diabetes. Using the diabetes-resistant B6-db strain as a comparison, in vivo and in vitro approaches were applied to identify metabolic and molecular differences between the two strains. RESULTS Despite higher plasma insulin levels, BKS-db mice exhibit lower lipogenic gene expression, rate of lipogenesis, hepatic triglyceride and glycogen content, and impaired insulin suppression of gluconeogenic genes. Hepatic insulin receptor substrate (IRS)-1 and IRS-2 expression and insulin-stimulated Akt-phosphorylation are decreased in BKS-db primary hepatocytes. Hyperinsulinemic-euglycemic clamp studies indicate that in contrast to hepatic insulin resistance, skeletal muscle is more insulin sensitive in BKS-db than in B6-db mice. We also demonstrate that elevated plasma triglyceride levels in BKS-db mice are associated with reduced triglyceride clearance due to lower lipase activities. CONCLUSIONS Our study demonstrates the presence of metabolic derangements in BKS-db before the onset of β-cell failure and identifies early hepatic insulin resistance as a component of the BKS-db phenotype. We propose that defects in hepatic insulin signaling contribute to the development of diabetes in the BKS-db mouse strain.