Intestine-specific expression of Apobec-1 rescues apolipoprotein B RNA editing and alters chylomicron production in Apobec1−/− mice

Intestinal apolipoprotein B (apoB) mRNA undergoes C-to-U editing, mediated by the catalytic deaminase apobec-1, which results in translation of apoB48. Apobec1^−/− mice produce only apoB100 and secrete larger chylomicron particles than those observed in wild-type (WT) mice. Here we show that transgenic rescue of intestinal apobec-1 expression (Apobec1^Int/O) restores C-to-U RNA editing of apoB mRNA in vivo, including the canonical site at position 6666 and also at approximately 20 other newly identified downstream sites present in WT mice. The small intestine of Apobec1^Int/O mice produces only apoB48, and the liver produces only apoB100. Serum chylomicron particles were smaller in Apobec1^Int/O mice compared with those from Apobec1^−/− mice, and the predominant fraction of serum apoB48 in Apobec1^Int/O mice migrated in lipoproteins smaller than chylomicrons, even when these mice were fed a high-fat diet. Because apoB48 arises exclusively from the intestine in Apobec1^Int/O mice and intestinal apoB48 synthesis and secretion rates were comparable to WT mice, we were able to infer the major sites of origin of serum apoB48 in WT mice. Our findings imply that less than 25% of serum apoB48 in WT mice arises from the intestine, with the majority originating from the liver.     

Loss of the nutrient receptor Tas1R3 reduces atherosclerotic plaque accumulation and hepatic steatosis in ApoE−/− mice

Journal of Physiology and Biochemistry - The taste receptor type I (Tas1R) family consists of three G protein-coupled receptors (T1R1, T1R2, and T1R3) that form heterodimers recognizing sweet...     

Deletion of tumor necrosis factor-α receptor type 1 exacerbates insulin resistance and hepatic steatosis in aromatase knockout mice

The relevance of estrogen functions in lipid metabolism has been suggested in patients with estrogen-signaling deficiencies. Their importance was further implied by studies in estrogen-deficient mice (ArKO mice), which progressively developed hepatic steatosis. As circulating tumor necrosis factor (TNF)-α levels are known to positively correlate with disturbances in lipid metabolism, we investigated the impact of the loss of TNF-α signaling on carbohydrate and lipid metabolism in ArKO mice. Histological examinations of the livers of mice at 5 months of age revealed that ArKO male mice lacking the TNF-α receptor type 1 (TNFR1) gene (ArKO/TNFR1KO) or both the TNFR 1 and 2 genes (ArKO/TNFR1&2KO) developed more severe hepatic steatosis than ArKO or ArKO/TNFR2KO mice. Serum analyses demonstrated a clear increase in cholesterol and insulin levels in the ArKO/TNFR1KO mice compared with the ArKO mice. Glucose- and insulin-tolerance tests further revealed exacerbation of the systemic insulin resistant phenotype in the ArKO/TNFR1KO mice. Hepatic expression of lipogenic genes including fatty-acid synthase and stearoyl-Coenzyme A desaturase 1 were more markedly upregulated in the ArKO/TNFR1KO mice than the ArKO mice. These findings indicate that under estrogen-deficient physiological conditions, hepatic lipid metabolism would benefit from TNF-α mediated signaling via TNFR1.     

TNF-α impairs endothelial function in adipose tissue resistance arteries of mice with diet-induced obesity

We tested the hypothesis that high fat (HF) feeding results in endothelial dysfunction in resistance arteries of epididymal white adipose tissue (eWAT) and is mediated by adipose tissue inflammation. When compared with normal chow (NC)-fed mice (n = 17), HF-fed male B6D2F1 mice were glucose intolerant and insulin resistant as assessed by glucose tolerance test (area under the curve; HF, 18,174 ± 1,889 vs. NC, 15,814 ± 666 mg·dl(-1)·min(-1); P < 0.05) and the homeostatic model assessment (HF, 64.1 ± 4.3 vs. NC, 85.7 ± 6.4; P = 0.05). HF diet-induced metabolic dysfunction was concomitant with a proinflammatory eWAT phenotype characterized by greater macrophage infiltration (HF, 3.9 ± 0.8 vs. NC, 0.8 ± 0.4%; P = 0.01) and TNF-α (HF, 22.6 ± 4.3 vs. NC, 11.4 ± 2.5 pg/dl; P < 0.05) and was associated with resistance artery dysfunction, evidenced by impaired endothelium-dependent dilation (EDD) (maximal dilation; HF, 49.2 ± 10.7 vs. NC, 92.4 ± 1.4%; P < 0.01). Inhibition of nitric oxide (NO) synthase by N(ω)-nitro-l-arginine methyl ester (l-NAME) reduced dilation in NC (28.9 ± 6.3%; P < 0.01)- and tended to reduce dilation in HF (29.8 ± 9.9%; P = 0.07)-fed mice, eliminating the differences in eWAT artery EDD between NC- and HF-fed mice, indicative of reduced NO bioavailability in eWAT resistance arteries after HF feeding. In vitro treatment of excised eWAT arteries with recombinant TNF-α (rTNF) impaired EDD (P < 0.01) in NC (59.7 ± 10.9%)- but not HF (59.0 ± 9.3%)-fed mice. l-NAME reduced EDD in rTNF-treated arteries from both NC (21.9 ± 6.4%)- and HF (29.1 ± 9.2%)-fed mice (both P < 0.01). In vitro treatment of arteries with a neutralizing antibody against TNF-α (abTNF) improved EDD in HF (88.2 ± 4.6%; P = 0.05)-fed mice but was without effect on maximal dilation in NC (89.0 ± 5.1%)-fed mice. l-NAME reduced EDD in abTNF-treated arteries from both NC (25.4 ± 7.5%)- and HF (27.1 ± 16.8%)-fed mice (both P < 0.01). These results demonstrate that inflammation in the visceral adipose tissue resulting from diet-induced obesity impairs endothelial function and NO bioavailability in the associated resistance arteries. This dysfunction may have important implications for adipose tissue blood flow and appropriate tissue function.     

iPLA2β deficiency attenuates obesity and hepatic steatosis in ob/ob mice through hepatic fatty-acyl phospholipid remodeling

PLA2G6 or GVIA calcium-independent PLA2 (iPLA2β) is identified as one of the NAFLD modifier genes in humans, and thought to be a target for NAFLD therapy. iPLA2β is known to play a house-keeping role in phospholipid metabolism and remodeling. However, its role in NAFLD pathogenesis has not been supported by results obtained from high-fat feeding of iPLA2β–null (PKO) mice. Unlike livers of human NAFLD and genetically obese rodents, fatty liver induced by high-fat diet is not associated with depletion of hepatic phospholipids. We therefore tested whether iPLA2β could regulate obesity and hepatic steatosis in leptin-deficient mice by cross-breeding PKO with ob/ob mice to generate ob/ob–PKO mice. Here we observed an improvement in ob/ob–PKO mice with significant reduction in serum enzymes, lipids, glucose, insulin as well as improved glucose tolerance, and reduction in islet hyperplasia. The improvement in hepatic steatosis measured by liver triglycerides, fatty acids and cholesterol esters was associated with decreased expression of PPARγ and de novo lipogenesis genes, and the reversal of β-oxidation gene expression. Notably, ob/ob livers contained depleted levels of lysophospholipids and phospholipids, and iPLA2β deficiency in ob/ob–PKO livers lowers the former, but replenished the latter particularly phosphatidylethanolamine (PE) and phosphatidylcholine (PC) that contained arachidonic (AA) and docosahexaenoic (DHA) acids. Compared with WT livers, PKO livers also contained increased PE and PC containing AA and DHA. Thus, iPLA2β deficiency protected against obesity and ob/ob fatty liver which was associated with hepatic fatty-acyl phospholipid remodeling. Our results support the deleterious role of iPLA2β in severe obesity associated NAFLD.     

Reduced VLDL clearance in Apoe−/−Npc1−/− mice is associated with increased Pcsk9 and Idol expression and decreased hepatic LDL-receptor levels

Niemann-Pick type C1 (NPC1) promotes the transport of LDL receptor (LDL-R)-derived cholesterol from late endosomes/lysosomes to other cellular compartments. NPC1-deficient cells showed impaired regulation of liver_X receptor (LXR) and sterol regulatory element-binding protein (SREBP) target genes. We observed that Apoe^−/−Npc1^−/− mice displayed a marked increase in total plasma cholesterol mainly due to increased VLDL, reflecting decreased clearance. Although nuclear SREBP-2 and Ldlr mRNA levels were increased in Apoe^−/−Npc1^−/− liver, LDL-R protein levels were decreased in association with marked induction of proprotein convertase subtilisin/kexin type 9 (Pcsk9) and inducible degrader of the LDL-R (Idol), both known to promote proteolytic degradation of LDL-R. While Pcsk9 is known to be an SREBP-2 target, marked upregulation of IDOL in Apoe^−/−Npc1^−/− liver was unexpected. However, several other LXR target genes also increased in Apoe^−/−Npc1^−/− liver, suggesting increased synthesis of endogenous LXR ligands secondary to activation of sterol biosynthesis. In conclusion, we demonstrate that NPC1 deficiency has a major impact on VLDL metabolism in Apoe^−/− mice through modulation of hepatic LDL-R protein levels. In contrast to modest induction of hepatic IDOL with synthetic LXR ligands, a striking upregulation of IDOL in Apoe^−/−Npc1^−/− mice could indicate a role of endogenous LXR ligands in regulation of hepatic IDOL.     

Reversal of high-fat diet-induced hepatic steatosis by n-3 LCPUFA: role of PPAR-α and SREBP-1c

Nonalcoholic fatty liver disease is characterized by an abnormal accumulation of triacylglycerides in the liver in absence of significant alcohol consumption. Under these conditions, it has been observed an impaired bioavailability of hepatic n-3 long-chain polyunsaturated fatty acids (LCPUFAs). The aim of this study was to test the reversion of the prosteatotic and proinflammatory effects of high-fat diet (HFD) in the mouse liver by changing to normocaloric diet and n-3 LCPUFA supplementation. Male C57BL/6J mice were given either control diet (CD) or HFD for 12 weeks. Control and HFD groups were divided into subgroups that continue with CD or subjected to CD plus n-3 LCPUFA for 8 additional weeks. After this time, blood and liver samples were taken and metabolic, morphologic, oxidative stress, inflammatory and signaling parameters were analyzed. The dietary change from HFD to a normocaloric diet with n-3 LCPUFA supplementation significantly reduced insulin resistance and liver steatosis when compared to switching HFD to normocaloric diet alone. In addition, HFD-induced increases in adiposity, adipocyte enlargement and liver oxidative stress and inflammatory cytokine expression were suppressed by n-3 LCPUFA to control values. Importantly, n-3 LCPUFA supplementation abolish HFD-induced enhancement in hepatic SREBP-1c/PPAR-α ratios, suggesting a change in the metabolic status of the liver from a lipogenic condition to one favoring fatty acid oxidation and steatosis attenuation. These findings may provide the rational basis for the use of normocaloric diets supplemented with n-3 LCPUFA in patients with liver steatosis.     

Characterization of intestinal inflammation and identification of related gene expression changes in mdr1a−/− mice

Multidrug resistance targeted mutation (mdr1a (-/-) ) mice spontaneously develop intestinal inflammation. The aim of this study was to further characterize the intestinal inflammation in mdr1a (-/-) mice. Intestinal samples were collected to measure inflammation and gene expression changes over time. The first signs of inflammation occurred around 16 weeks of age and most mdr1a (-/-) mice developed inflammation between 16 and 27 weeks of age. The total histological injury score was the highest in the colon. The inflammatory lesions were transmural and discontinuous, revealing similarities to human inflammatory bowel diseases (IBD). Genes involved in inflammatory response pathways were up-regulated whereas genes involved in biotransformation and transport were down-regulated in colonic epithelial cell scrapings of inflamed mdra1 (-/-) mice at 25 weeks of age compared to non-inflamed FVB mice. These results show overlap to human IBD and strengthen the use of this in vivo model to study human IBD. The anti-inflammatory regenerating islet-derived genes were expressed at a lower level during inflammation initiation in non-inflamed colonic epithelial cell scrapings of mdr1a (-/-) mice at 12 weeks of age. This result suggests that an insufficiently suppressed immune response could be crucial to the initiation and development of intestinal inflammation in mdr1a (-/-) mice.     

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.     

Gene expression changes in the secondary palate and mandible of Prdm16 −/− mice

Loss of Prdm16 expression in the mouse leads to a complete cleft of the secondary palate. We have now determined changes in gene expression in the secondary palates of Prdm16 ^?/? fetuses in an attempt to reveal the mechanism(s) leading to the failure of palate closure in these mice. Defined pathway-based polymerase chain reaction arrays were used to analyze the expression of genes associated with the extracellular matrix and the transforming growth factor-β and bone morphogenetic protein signaling networks, perturbations of which can lead to palatal clefting. Loss of Prdm16 expression in the secondary palate leads to alterations in numerous genes within these groups, many of which have been linked to chondrogenesis and osteogenesis. The expression of several genes linked to bone development was significantly changed in the developing secondary palate. Analysis of gene expression in the mandibles of Prdm16 ^?/? fetuses revealed similar alterations in the same gene set. These data suggest that one function of Prdm16 is the regulation of genes that play a role in the differentiation of mesenchymal cells into chondro-/osteocytes.     

Diacylglycerol acyltransferase 2 acts upstream of diacylglycerol acyltransferase 1 and utilizes nascent diglycerides and de novo synthesized fatty acids in HepG2 cells

The two diacylglycerol acyltransferases, DGAT1 and DGAT2, are known to have non-redundant functions, in spite of catalysing the same reaction and being present in the same cell types. The basis for this distinctiveness, which is reflected in the very different phenotypes of Dgat1(-/-) and Dgat2(-/-) mice, has not been resolved. Using selective inhibitors of human DGAT1 and DGAT2 on HepG2 cells and gene silencing, we show that, although DGAT2 activity accounts for a modest fraction (相似文献   

Rag-dependent and Rag-independent mechanisms of Notch1 rearrangement in thymic lymphomas of Atm−/− and scid mice

The pathways of thymic lymphomagenesis are classified as Rag-dependent or -independent according to their dependence on recombination-activating gene (Rag1/2) proteins. The role of the two-lymphoma pathways in oncogene rearrangements and the connection between lymphoma pathways and rearrangement mechanisms, however, remain obscure. We compared the incidence and latency of thymic lymphomas, and associated rearrangements of the representative oncogene Notch1 among Rag2^?/?, ataxia telangiectasia mutated (Atm)^?/?, and severe combined immune deficiency (scid) mice combined with Rag2 deficiency. Contrary to expectations, Rag2^?/? mice were prone to thymic lymphoma development, suggesting the existence of a Rag2-independent lymphoma pathway in Rag2^?/? mice. The lymphoma incidence in Rag2^?/?Atm^?/? mice was lower than that in Atm^?/? mice, but higher than that in Rag2^?/? mice, indicating that Atm^?/? mice develop lymphomas through both pathways. Scid mice developed lymphomas with an incidence and latency similar to Rag2^?/?scid mice, suggesting that Rag2-mediated V(D)J recombination-driven events are not necessarily required for lymphomagenesis in scid mice. Notch1 rearrangement mechanisms were classified as Rag2-dependent or Rag2-independent based on the presence of recombination signal-like sequences at rearranged sites. In Rag2^?/? lymphomas, Notch1 must be rearranged independently of Rag2 function, implying that Rag2^?/? mice are susceptible to lymphomagenesis due to the presence of other rearrangement mechanisms. The results in Atm^?/? mice suggest that Notch1 was rearranged through both lymphoma pathways. In scid mice, the frequency of Rag2-mediated rearrangements was relatively low compared with that in wild-type mice, suggesting that the Rag2-independent lymphoma pathway prevails in the development of thymic lymphomas in scid mice. Thus, two rearrangement mechanisms underlie the lymphoma pathways and constitute the mechanistic bases for lymphomagenesis, thereby providing the molecular criteria for distinguishing between Rag2-dependent and Rag2-independent lymphoma pathways.     

Hepatic bile acid metabolism and expression of cytochrome P450 and related enzymes are altered in Bsep −/− mice

The bile salt export pump (BSEP/Bsep; gene symbol ABCB11/Abcb11) translocates bile salts across the hepatocyte canalicular membrane into bile in humans and mice. In humans, mutations in the ABCB11 gene cause a severe childhood liver disease known as progressive familial intrahepatic cholestasis type 2. Targeted inactivation of mouse Bsep produces milder persistent cholestasis due to detoxification of bile acids through hydroxylation and alternative transport pathways. The purpose of the present study was to determine whether functional expression of hepatic cytochrome P450 (CYP) and microsomal epoxide hydrolase (mEH) is altered by Bsep inactivation in mice and whether bile acids regulate CYP and mEH expression in Bsep ^?/? mice. CYP expression was determined by measuring protein levels of Cyp2b, Cyp2c and Cyp3a enzymes and CYP-mediated activities including lithocholic acid hydroxylation, testosterone hydroxylation and alkoxyresorufin O-dealkylation in hepatic microsomes prepared from female and male Bsep ^?/? mice fed a normal or cholic acid (CA)-enriched diet. The results indicated that hepatic lithocholic acid hydroxylation was catalyzed by Cyp3a/Cyp3a11 enzymes in Bsep ^?/? mice and that 3-ketocholanoic acid and murideoxycholic acid were major metabolites. CA feeding of Bsep ^?/? mice increased hepatic Cyp3a11 protein levels and Cyp3a11-mediated testosterone 2β-, 6β-, and 15β-hydroxylation activities, increased Cyp2b10 protein levels and Cyp2b10-mediated benzyloxyresorufin O-debenzylation activity, and elevated Cyp2c29 and mEH protein levels. We propose that bile acids upregulate expression of hepatic Cyp3a11, Cyp2b10, Cyp2c29 and mEH in Bsep ^?/? mice and that Cyp3a11 and multidrug resistance-1 P-glycoproteins (Mdr1a/1b) are vital components of two distinct pathways utilized by mouse hepatocytes to expel bile acids.