Upregulation of hepatic LDL transport by n-3 fatty acids in LDL receptor knockout mice

We determined the effects of dietary n-6 and n-3 polyunsaturated fatty acids (PUFA) on parameters of plasma lipoprotein and hepatic lipid metabolism in LDL receptor (LDLr) knockout mice. Dietary n-3 PUFA decreased the rate of appearance and increased the hepatic clearance of IDL/LDL resulting in a marked decrease in the plasma concentration of these particles. Dietary n-3 PUFA increased the hepatic clearance of IDL/LDL through a mechanism that appears to involve apolipoprotein (apo)E but is independent of the LDLr, the LDLr related protein (LRP), the scavenger receptor B1, and the VLDLr. The decreased rate of appearance of IDL/VLDL in the plasma of animals fed n-3 PUFA could be attributed to a marked decrease in the plasma concentration of precursor VLDL. Decreased plasma VLDL concentrations were due in part to decreased hepatic secretion of VLDL triglyceride and cholesteryl esters, which in turn was associated with decreased concentrations of these lipids in liver. Decreased hepatic triglyceride concentrations in animals fed n-3 PUFA were due in part to suppression of fatty acid synthesis as a result of a decrease in sterol regulatory element binding protein-1 (SREBP-1) expression and processing. In conclusion, these studies indicate that n-3 PUFA can markedly decrease the plasma concentration of apoB-containing lipoproteins and enhance hepatic LDL clearance through a mechanism that does not involve the LDLr pathway or LRP.     

The role of hepatic Surf4 in lipoprotein metabolism and the development of atherosclerosis in apoE−/− mice

Elevated plasma levels of low-density lipoprotein-C (LDL-C) increase the risk of atherosclerotic cardiovascular disease. Circulating LDL is derived from very low-density lipoprotein (VLDL) metabolism and cleared by LDL receptor (LDLR). We have previously demonstrated that cargo receptor Surfeit 4 (Surf4) mediates VLDL secretion. Inhibition of hepatic Surf4 impairs VLDL secretion, significantly reduces plasma LDL-C levels, and markedly mitigates the development of atherosclerosis in LDLR knockout (Ldlr^?/?) mice. Here, we investigated the role of Surf4 in lipoprotein metabolism and the development of atherosclerosis in another commonly used mouse model of atherosclerosis, apolipoprotein E knockout (apoE^?/?) mice. Adeno-associated viral shRNA was used to silence Surf4 expression mainly in the liver of apoE^?/? mice. In apoE^?/? mice fed a regular chow diet, knockdown of Surf4 expression significantly reduced triglyceride secretion and plasma levels of non-HDL cholesterol and triglycerides without causing hepatic lipid accumulation or liver damage. When Surf4 was knocked down in apoE^?/? mice fed the Western-type diet, we observed a significant reduction in plasma levels of non-HDL cholesterol, but not triglycerides. Knockdown of Surf4 did not increase hepatic cholesterol and triglyceride levels or cause liver damage, but significantly diminished atherosclerosis lesions. Therefore, our findings indicate the potential of hepatic Surf4 inhibition as a novel therapeutic strategy to reduce the risk of atherosclerotic cardiovascular disease.     

Stimulation of the in vivo production of very low density lipoproteins by apolipoprotein E is independent of the presence of the low density lipoprotein receptor

Apolipoprotein (apo) E stimulates the secretion of very low density lipoproteins (VLDLs) by an as yet unknown mechanism. Recently, a working mechanism for apoE was proposed (Twisk, J., Gillian-Daniel, D. L., Tebon, A., Wang, L., Barrett, P. H., and Attie, A. D. (2000) J. Clin. Invest. 105, 521-532) in which apoE prevents the inhibitory action of the low density lipoprotein receptor (LDLr) by binding to it. We have first tested whether this newly described effect of the LDLr on VLDL secretion, obtained in vitro, is also observed in vivo. In LDLr knockout mice (LDLr-/-), the production of VLDL triglycerides and apoB was 30% higher than that in controls. Also the ratio of apoB100:apoB48 secretion was increased in the LDLr-/- mice. The composition of nascent VLDL was similar in both strains. To test whether the action of apoE depends on the presence of the LDLr, VLDL production was measured in LDLr-/- and apoE-/- LDLr-/- mice. Deletion of apoE on a LDLr-/- background still caused a 50% decrease of VLDL triglycerides and apoB production. The composition of nascent VLDL was again similar for both strains. We conclude that the effect of apoE on hepatic VLDL production is independent of the presence of the LDLr.     

Oligomeric proanthocyanidins mitigate cholesterol and cholic acid diet–induced hepatic dysfunction in male Sprague Dawley rats

Dysregulated synthesis of hepatic cholesterol is a critical determinant of atherosclerosis. The combination of cholesterol and cholic acid (CC) diet supplementation to animal models is associated with hepatic dysfunction‐mediated atherosclerosis. The current study was designed to investigate the hepatic cholesterol–lowering effects of oligomeric proanthocyanidins (OPC) in CC diet fed rats. CC diet–induced group exhibited significant increase in the hepatic lipid profile, activities of 3‐hydroxy‐3‐methylglutaryl coenzyme A (HMG‐CoA) reductase (HMGR), PON‐1, LCAT, LPL, and LPO levels, and messenger RNA expression of HMGR, low‐density lipoprotein receptor (LDLr), and HNF‐4α. Administration of OPC (100 mg/kg/bwt) resulted in the significant reduction of lipid profile and HMGR levels, with concomitant increase in the levels of cholesterol‐regulating enzymes and upregulated expression of LDLr and HNF‐4α, which was similar to atorvastatin. Molecular docking studies also revealed that proanthocyanidins had a strong binding affinity to HMGR, similar to atorvastatin. Our findings suggest that OPC regulate the impaired cholesterol metabolism–associated atherosclerosis through hepatic cholesterol–lowering effect.     

A decreased expression of angiopoietin-like 3 is protective against atherosclerosis in apoE-deficient mice

KK/Snk mice (previously KK/San) possessing a recessive mutation (hypl) of the angiopoietin-like 3 (Angptl3) gene homozygously exhibit a marked reduction of VLDL due to the decreased Angptl3 expression. Recently, we proposed that Angptl3 is a new class of lipid metabolism modulator regulating VLDL triglyceride (TG) levels through the inhibition of lipoprotein lipase (LPL) activity. In this study, to elucidate the role of Angptl3 in atherogenesis, we investigated the effects of hypl mutation against hyperlipidemia and atherosclerosis in apolipoprotein E knockout (apoEKO) mice. ApoEKO mice with hypl mutation (apoEKO-hypl) exhibited a significant reduction of VLDL TG, VLDL cholesterol, and plasma apoB levels compared with apoEKO mice. Hepatic VLDL TG secretion was comparable between both apoE-deficient mice. Turnover studies revealed that the clearance of both [3H]TG-labeled and 125I-labeled VLDL was significantly enhanced in apoEKO-hypl mice. Postprandial plasma TG levels also decreased in apoEKO-hypl mice. Both LPL and hepatic lipase activities in the postheparin plasma increased significantly in apoEKO-hypl mice, explaining the enhanced lipid metabolism. Furthermore, apoEKO-hypl mice developed 3-fold smaller atherogenic lesions in the aortic sinus compared with apoEKO mice. Taken together, the reduction of Angptl3 expression is protective against hyperlipidemia and atherosclerosis, even in the absence of apoE, owing to the enhanced catabolism and clearance of TG-rich lipoproteins.     

Dietary supplementation with Pluronic L-81 modifies hepatic secretion of very low density lipoproteins in the rat

Supplementation of high fat/cholesterol-enriched diets with polyoxypropylene-polyoxyethylene copolymers containing 90% hydrophobic constituents has been found to impair enteric secretion of chylomicrons, lower plasma levels of very low density (VLDL) and low density (LDL) lipoprotein cholesterol and prevent diet-induced hypercholesterolemia and atherosclerosis. These agents are known to be absorbed from the gastrointestinal tract and excreted in bile. In order to determine whether dietary supplementation with this group of hydrophobic poloxalenes influences hepatic secretion of triglyceride-rich lipoproteins, groups of rats were maintained for 21-34 days on either standard chow, semisynthetic diet containing 10.0% safflower oil/1.0% cholesterol, or each of the above diets supplemented with the hydrophobic poloxalene Pluronic L-81. At the end of the feeding period, newly secreted hepatic VLDL were isolated from 2-hr recirculating liver perfusates, quantitated, and characterized. Compared to perfusions in chow-fed rats, perfusion experiments in rats fed the high fat/cholesterol-enriched semisynthetic diet revealed a 3.1-fold increased net hepatic VLDL secretion rate; enrichment of secretory VLDL in cholesteryl esters and in C18:2 core lipid fatty acids; and a shift in the size distribution of secretory VLDL towards larger particles. When the 0.5% Pluronic L-81 was included in the high fat/cholesterol-enriched semisynthetic diet, the net hepatic VLDL secretion rate fell significantly and the physicochemical properties of secretory VLDL in these rats were found to resemble those of chow-fed animals. Supplementation of the chow diet with L-81 resulted in a significant fall in the net hepatic VLDL secretion rate from that observed in rats fed chow alone. Compared to rats fed chow alone, perfusate VLDL from rats fed each of the other experimental diets contained markedly lower amounts of both apoB molecular weight variants, as analyzed by gradient gel electrophoresis and densitometric gel scanning. Since previous studies have demonstrated that VLDL are the major cholesterol transport lipoproteins following fat/cholesterol feeding; a precursor-product relationship exists between fat/cholesterol-induced hepatic VLDL and plasma VLDL; such particles are capable of delivering cholesterol to the arterial wall; and dietary supplementation with hydrophobic poloxalenes prevents both the increase in plasma VLDL-cholesterol and diet-induced atherosclerosis, it is possible that dietary supplementation with hydrophobic poloxalenes may influence the atherogenic process through direct and/or indirect effects on hepatic VLDL transport.     

Hepatic insulin resistance is sufficient to produce dyslipidemia and susceptibility to atherosclerosis

Insulin resistance plays a central role in the development of the metabolic syndrome, but how it relates to cardiovascular disease remains controversial. Liver insulin receptor knockout (LIRKO) mice have pure hepatic insulin resistance. On a standard chow diet, LIRKO mice have a proatherogenic lipoprotein profile with reduced high-density lipoprotein (HDL) cholesterol and very low-density lipoprotein (VLDL) particles that are markedly enriched in cholesterol. This is due to increased secretion and decreased clearance of apolipoprotein B-containing lipoproteins, coupled with decreased triglyceride secretion secondary to increased expression of Pgc-1 beta (Ppargc-1b), which promotes VLDL secretion, but decreased expression of Srebp-1c (Srebf1), Srebp-2 (Srebf2), and their targets, the lipogenic enzymes and the LDL receptor. Within 12 weeks on an atherogenic diet, LIRKO mice show marked hypercholesterolemia, and 100% of LIRKO mice, but 0% of controls, develop severe atherosclerosis. Thus, insulin resistance at the level of the liver is sufficient to produce the dyslipidemia and increased risk of atherosclerosis associated with the metabolic syndrome.     

The Liver-Selective Thyromimetic T-0681 Influences Reverse Cholesterol Transport and Atherosclerosis Development in Mice

Background

Liver-selective thyromimetics have been reported to efficiently reduce plasma cholesterol through the hepatic induction of both, the low-density lipoprotein receptor (LDLr) and the high-density lipoprotein (HDL) receptor; the scavenger receptor class B type I (SR-BI). Here, we investigated the effect of the thyromimetic T-0681 on reverse cholesterol transport (RCT) and atherosclerosis, and studied the underlying mechanisms using different mouse models, including mice lacking LDLr, SR-BI, and apoE, as well as CETP transgenic mice.

Methodology/Principal Findings

T-0681 treatment promoted bile acid production and biliary sterol secretion consistently in the majority of the studied mouse models, which was associated with a marked reduction of plasma cholesterol. Using an assay of macrophage RCT in mice, we found T-0681 to significantly increase fecal excretion of macrophage-derived neutral and acidic sterols. No positive effect on RCT was found in CETP transgenic mice, most likely due to the observed decrease in plasma CETP mass. Studies in SR-BI KO and LDLr KO mice suggested hepatic LDLr to be necessary for the action of T-0681 on lipid metabolism, as the compound did not have any influence on plasma cholesterol levels in mice lacking this receptor. Finally, prolonged treatment with T-0681 reduced the development of atherosclerosis by 60% in apoE KOs on Western type diet. In contrast, at an earlier time-point T-0681 slightly increased small fatty streak lesions, in part due to an impaired macrophage cholesterol efflux capacity, when compared to controls.

Conclusions/Significance

The present results show that liver-selective thyromimetics can promote RCT and that such compounds may protect from atherosclerosis partly through induction of bile acid metabolism and biliary sterol secretion. On-going clinical trials will show whether selective thyromimetics do prevent atherosclerosis also in humans.     

Antisense oligonucleotide reduction of apoB-ameliorated atherosclerosis in LDL receptor-deficient mice

Chronic elevations of plasma apolipoprotein B (apoB) are strongly associated with cardiovascular disease. We have previously demonstrated that inhibition of hepatic apoB mRNA using antisense oligonucleotides (ASO) results in reductions of apoB, VLDL, and LDL in several preclinical animal models and humans. In this study, we evaluated the anti-atherogenic effects of a murine-specific apoB ASO (ISIS 147764) in hypercholesterolemic LDLr deficient (LDLr(-/-)) mice. ISIS 147764 was administered weekly at 25-100 mg/kg for 10-12 weeks and produced dose-dependent reductions of hepatic apoB mRNA and plasma LDL by 60-90%. No effects on these parameters were seen in mice receiving control ASOs. ApoB ASO treatment also produced dose-dependent reductions of aortic en face and sinus atherosclerosis from 50-90%, with high-dose treatment displaying less disease than the saline-treated, chow-fed LDLr(-/-) mice. No changes in intestinal cholesterol absorption were seen with apoB ASO treatment, suggesting that the cholesterol-lowering pharmacology of 147764 was primarily due to inhibition of hepatic apoB synthesis and secretion. In summary, ASO-mediated suppression of apoB mRNA expression profoundly reduced plasma lipids and atherogenesis in LDLr(-/-) mice, leading to the hypothesis that apoB inhibition in humans with impaired LDLr activity may produce similar effects.     

Apolipoprotein A-IV regulates chylomicron metabolism-mechanism and function

Dietary fat is an important mediator of atherosclerosis and obesity. Despite its importance in mediating metabolic disease, there is still much unknown about dietary fat absorption in the intestine and especially the detailed biological roles of intestinal apolipoproteins involved in that process. We were specifically interested in determining the physiological role of the intestinal apolipoprotein A-IV (A-IV) using A-IV knockout (KO) mice. A-IV is stimulated by fat absorption in the intestine and is secreted on nascent chylomicrons into intestinal lymph. We found that A-IV KO mice had reduced plasma triglyceride (TG) and cholesterol levels and that this hypolipidemia persisted on a high-fat diet. A-IV KO did not cause abnormal intestinal lipid absorption, food intake, or adiposity. Additionally, A-IV KO did not cause abnormal liver TG and cholesterol metabolism, as assessed by measuring hepatic lipid content, lipogenic and cholesterol synthetic gene expression, and in vivo VLDL secretion. Instead, A-IV KO resulted in the secretion of larger chylomicrons from the intestine into the lymph, and those chylomicrons were cleared from the plasma more slowly than wild-type chylomicrons. These data suggest that A-IV has a previously unknown role in mediating the metabolism of chylomicrons, and therefore may be important in regulating plasma lipid metabolism.     

Disruption of the sterol carrier protein 2 gene in mice impairs biliary lipid and hepatic cholesterol metabolism.

Hepatic up-regulation of sterol carrier protein 2 (Scp2) in mice promotes hypersecretion of cholesterol into bile and gallstone formation in response to a lithogenic diet. We hypothesized that Scp2 deficiency may alter biliary lipid secretion and hepatic cholesterol metabolism. Male gallstone-susceptible C57BL/6 and C57BL/6(Scp2(-/-)) knockout mice were fed a standard chow or lithogenic diet. Hepatic biles were collected to determine biliary lipid secretion rates, bile flow, and bile salt pool size. Plasma lipoprotein distribution was investigated, and gene expression of cytosolic lipid-binding proteins, lipoprotein receptors, hepatic regulatory enzymes, and intestinal cholesterol absorption was measured. Compared with chow-fed wild-type animals, C57BL/6(Scp2(-/-)) mice had higher bile flow and lower bile salt secretion rates, decreased hepatic apolipoprotein expression, increased hepatic cholesterol synthesis, and up-regulation of liver fatty acid-binding protein. In addition, the bile salt pool size was reduced and intestinal cholesterol absorption was unaltered in C57BL/6(Scp2(-/-)) mice. When C57BL/6(Scp2(-/-)) mice were challenged with a lithogenic diet, a smaller increase of hepatic free cholesterol failed to suppress cholesterol synthesis and biliary cholesterol secretion increased to a much smaller extent than phospholipid and bile salt secretion. Scp2 deficiency did not prevent gallstone formation and may be compensated in part by hepatic up-regulation of liver fatty acid-binding protein. These results support a role of Scp2 in hepatic cholesterol metabolism, biliary lipid secretion, and intracellular cholesterol distribution.     

Effect of macrophage-derived apolipoprotein E on hyperlipidemia and atherosclerosis of LDLR-deficient mice

LDL receptor-deficient (LDLR(-/-)) mice fed a Western diet exhibit severe hyperlipidemia and develop significant atherosclerosis. Apolipoprotein E (apoE) is a multifunctional protein synthesized by hepatocytes and macrophages. We sought to determine effect of macrophage apoE deficiency on severe hyperlipidemia and atherosclerosis. Female LDLR(-/-) mice were lethally irradiated and reconstituted with bone marrow from either apoE(-/-) or apoE(+/+) mice. Four weeks after transplantation, recipient mice were fed a Western diet for 8 weeks. Reconstitution of LDLR(-/-) mice with apoE(-/-) bone marrow resulted in a slight reduction in plasma apoE levels and a dramatic reduction in accumulation of apoE and apoB in the aortic wall. Plasma lipid levels were unaffected when mice had mild hyperlipidemia on a chow diet, whereas IDL/LDL cholesterol levels were significantly reduced when mice developed severe hyperlipidemia on the Western diet. The hepatic VLDL production rate of mice on the Western diet was decreased by 46% as determined by injection of Triton WR1339 to block VLDL clearance. Atherosclerotic lesions in the proximal aorta were significantly reduced, partially due to reduction in plasma total cholesterol levels (r=0.56; P<0.0001). Thus, macrophage apoE-deficiency alleviates severe hyperlipidemia by slowing hepatic VLDL production and consequently reduces atherosclerosis in LDLR(-/-) mice.     

Influence of low-density lipoprotein (LDL) receptor on lipid composition, inflammation and parasitism during Toxoplasma gondii infection

Intracellular replication of Toxoplasma gondii requires cholesterol uptake by host cell low-density lipoprotein receptor (LDLr), a critical element in atherosclerosis. We evaluated host parasitism, inflammatory responses and development of atherosclerosis in LDLr knockout (LDLr(-/-)) and their controls C57BL/6 mice infected with T. gondii. Our results show that T. gondii cysts were reduced in LDLr(-/-) mice when compared to C57BL/6 mice. However, in presence of hypercholesterolemic diet, parasite growth in LDLr(-/-) mice was similar to that seen in infected C57BL/6 mice. In presence of a hypercholesterolemic diet, T. gondii infection leads to a 60% reduction of serum triacylglycerol, total and atherogenic lipoprotein cholesterol. When aortic valve lesion was analyzed, infected mice showed a reduction of atherosclerotic lesion area as well as CD36 expression. MCP-1, SRA-I, SRA-II, ICAM-1 and VCAM-1 mRNA expression was kept similar between infected and control groups. Thus, despite the intense inflammatory process, the drastic reduction in serum lipids seems to limit the development of atherosclerosis in LDLr(-/-) mice infected with T. gondii. In conclusion, our results indicate that T. gondii employs host LDLr to acquire cholesterol and favor its growth. However, in the presence of hypercholesterolemia, T. gondii parasites are able to acquire cholesterol-rich lipoproteins through an alternative host receptor, and overcome LDLr deficiency, favoring host parasitism and impairing lipid loading of foam cells.     

The effect of dietary sphingolipids on plasma sphingomyelin metabolism and atherosclerosis

Sphingomyelin (SM) plays a very important role in cell membrane formation and plasma lipoprotein metabolism. All these functions may have an impact on atherosclerotic development. To investigate the relationship between SM metabolism and atherosclerosis, we utilized a sphingolipid-rich diet to feed LDL receptor gene knockout (LDLr KO) mice and studied lipid metabolism and atherosclerosis in the mice. After 3 months of a sphingolipid-rich diet, we found a significant increase in SM, cholesterol, and SM/phosphatidylcholine (PC) ratio (50%, P<0.001; 62%, P<0.01; and 45%, P<0.01, respectively), compared to chow fed diet. HDL-lipids were not significantly altered. Non-HDL-SM, non-HDL-C, and non-HDL-SM/non-HDL-PC ratio were significantly increased (115%, P<0.001; 106%, P<0.001; and 106%, P<0.01, respectively). FPLC confirmed the results. SDS-PAGE showed an increase of apoB48 and apoB100, but no changes of apoAI. Moreover, we found that an SM-rich diet significantly increased atherosclerotic lesion area in both root assay and en face assay, compared to chow diet (58,210+/-15,300 microm(2) vs. 9670+/-2370 microm(2), P<0.001; 5.9+/-3.1% vs. 1.1+/-0.9%, P<0.001). These results indicate that the enrichment of sphingolipids in diet has proatherogenic properties.     

Absence of VLDL secretion does not affect alpha-tocopherol content in peripheral tissues

alpha-Tocopherol is a lipid-soluble antioxidant that helps to prevent oxidative damage to cellular lipids. alpha-Tocopherol is absorbed by the intestine and is taken up and retained by the liver; it is widely presumed that alpha-tocopherol is then delivered to peripheral tissues by the secretion of VLDL. To determine whether VLDL secretion is truly important for the delivery of alpha-tocopherol to peripheral tissues, we examined alpha-tocopherol metabolism in mice that lack microsomal triglyceride transfer protein (Mttp) expression in the liver and therefore cannot secrete VLDL (Mttp(Delta/Delta) mice). Mttp(Delta/Delta) mice have low plasma lipid levels and increased stores of lipids in the liver. Similarly, alpha-tocopherol levels in the plasma were lower in Mttp(Delta/Delta) mice than in controls, whereas hepatic alpha-tocopherol stores were higher. However, alpha-tocopherol levels in the peripheral tissues of Mttp(Delta/Delta) mice were nearly identical to those of control mice, suggesting that VLDL secretion is not critical for the delivery of alpha-tocopherol to peripheral tissues. When fed a diet containing deuterated alpha-tocopherol, Mttp(Delta/Delta) and control mice had similar incorporation of deuterated alpha-tocopherol into plasma and various peripheral tissues. We conclude that the absence of VLDL secretion has little effect on the stores of alpha-tocopherol in peripheral tissues, at least in the mouse.     

Defective VLDL metabolism and severe atherosclerosis in mice expressing human apolipoprotein E isoforms but lacking the LDL receptor

Differences in affinity of human apolipoprotein E (apoE) isoforms for the low density lipoprotein receptor (LDLR) are thought to result in the differences in lipid metabolism observed in humans with different APOE genotypes. Mice expressing three common human apoE isoforms, E2, E3, and E4, in place of endogenous mouse apoE were used to investigate the relative roles of apoE isoforms in LDLR- and non-LDLR-mediated very low density lipoprotein (VLDL) clearance. While both VLDL particles isolated from mice expressing apoE3 and apoE4 bound to mouse LDLR with affinity and Bmax similar to VLDL containing mouse apoE, VLDL with apoE2 bound with only half the Bmax. In the absence of the LDLR, all lines of mice expressing human apoE showed dramatic increases in VLDL cholesterol and triglycerides (TG) compared to LDLR knockout mice expressing mouse apoE. The mechanism of the hyperlipidemia in mice expressing human apoE isoforms is due to impairment of non-LDL-receptor-mediated VLDL clearance. This results in the severe atherosclerosis observed in mice expressing human apoE but lacking the LDLR, even when fed normal chow diet. Our data show that defects in LDLR independent pathway(s) are a potential factor that trigger hyperlipoproteinemia when the LDLR pathway is perturbed, as in E2/2 mice.     

Loss of resistin ameliorates hyperlipidemia and hepatic steatosis in leptin-deficient mice

Resistin has been linked to components of the metabolic syndrome, including obesity, insulin resistance, and hyperlipidemia. We hypothesized that resistin deficiency would reverse hyperlipidemia in genetic obesity. C57Bl/6J mice lacking resistin [resistin knockout (RKO)] had similar body weight and fat as wild-type mice when fed standard rodent chow or a high-fat diet. Nonetheless, hepatic steatosis, serum cholesterol, and very low-density lipoprotein (VLDL) secretion were decreased in diet-induced obese RKO mice. Resistin deficiency exacerbated obesity in ob/ob mice, but hepatic steatosis was drastically attenuated. Moreover, the levels of triglycerides, cholesterol, insulin, and glucose were reduced in ob/ob-RKO mice. The antisteatotic effect of resistin deficiency was related to reductions in the expression of genes involved in hepatic lipogenesis and VLDL export. Together, these results demonstrate a crucial role of resistin in promoting hepatic steatosis and hyperlipidemia in obese mice.     

12/15-Lipoxygenase Is Required for the Early Onset of High Fat Diet-Induced Adipose Tissue Inflammation and Insulin Resistance in Mice

Background

Recent understanding that insulin resistance is an inflammatory condition necessitates searching for genes that regulate inflammation in insulin sensitive tissues. 12/15-lipoxygenase (12/15LO) regulates the expression of proinflammatory cytokines and chemokines and is implicated in the early development of diet-induced atherosclerosis. Thus, we tested the hypothesis that 12/15LO is involved in the onset of high fat diet (HFD)-induced insulin resistance.

Methodology/Principal Findings

Cells over-expressing 12/15LO secreted two potent chemokines, MCP-1 and osteopontin, implicated in the development of insulin resistance. We assessed adipose tissue inflammation and whole body insulin resistance in wild type (WT) and 12/15LO knockout (KO) mice after 2–4 weeks on HFD. In adipose tissue from WT mice, HFD resulted in recruitment of CD11b⁺, F4/80⁺ macrophages and elevated protein levels of the inflammatory markers IL-1β, IL-6, IL-10, IL-12, IFNγ, Cxcl1 and TNFα. Remarkably, adipose tissue from HFD-fed 12/15LO KO mice was not infiltrated by macrophages and did not display any increase in the inflammatory markers compared to adipose tissue from normal chow-fed mice. WT mice developed severe whole body (hepatic and skeletal muscle) insulin resistance after HFD, as measured by hyperinsulinemic euglycemic clamp. In contrast, 12/15LO KO mice exhibited no HFD-induced change in insulin-stimulated glucose disposal rate or hepatic glucose output during clamp studies. Insulin-stimulated Akt phosphorylation in muscle tissue from HFD-fed mice was significantly greater in 12/15LO KO mice than in WT mice.

Conclusions

These results demonstrate that 12/15LO mediates early stages of adipose tissue inflammation and whole body insulin resistance induced by high fat feeding.     

LOX-1 deletion and macrophage trafficking in atherosclerosis

Background

Atherosclerosis is associated with macrophage accumulation. LOX-1 has been shown to induce macrophage attachment, and its deletion (LOX-1 knockout, KO) reduces atherosclerosis in LDLr KO mice fed a high cholesterol diet. We examined differences in macrophage trafficking in age-matched wild type, LOX-1 KO, LDLr KO, and LDLr/LOX-1 double KO mice.

Methods

Sections of aortas of mice fed high cholesterol diet were collected at weeks 0, 4, 8, 12 and 19 and analyzed by immunohistochemistry and flow cytometry.

Results

In the LDLr KO mice aorta, CD68 positivity (macrophage accumulation) increased over time up to 12 weeks, and then the accumulation fell modestly but significantly. The periaortal fat and adventitia showed more CD68 positivity than the media and intima. This pattern was also evident in the non-atherosclerotic areas. Importantly, LOX-1 KO and LDLr–LOX-1 double KO mice showed diminished CD68 positivity in comparison to wild type and LDLR KO mice, respectively. Further, macrophages from LOX-1 KO mice revealed a marked reduction in migration (vs. macrophages from wild type mice) in in vitro migration assay.

Conclusions

LOX-1 deletion translates into reduction in macrophage trafficking in the aorta of LDLr KO mice. Most of the macrophage trafficking appears in the subadventitial regions.