Blocking VLDL secretion causes hepatic steatosis but does not affect peripheral lipid stores or insulin sensitivity in mice

The liver secretes triglyceride-rich VLDLs, and the triglycerides in these particles are taken up by peripheral tissues, mainly heart, skeletal muscle, and adipose tissue. Blocking hepatic VLDL secretion interferes with the delivery of liver-derived triglycerides to peripheral tissues and results in an accumulation of triglycerides in the liver. However, it is unclear how interfering with hepatic triglyceride secretion affects adiposity, muscle triglyceride stores, and insulin sensitivity. To explore these issues, we examined mice that cannot secrete VLDL [due to the absence of microsomal triglyceride transfer protein (Mttp) in the liver]. These mice exhibit markedly reduced levels of apolipoprotein B-100 in the plasma, along with reduced levels of triglycerides in the plasma. Despite the low plasma triglyceride levels, triglyceride levels in skeletal muscle were unaffected. Adiposity and adipose tissue triglyceride synthesis rates were also normal, and body weight curves were unaffected. Even though the blockade of VLDL secretion caused hepatic steatosis accompanied by increased ceramides and diacylglycerols in the liver, the mice exhibited normal glucose tolerance and were sensitive to insulin at the whole-body level, as judged by hyperinsulinemic euglycemic clamp studies. Normal hepatic glucose production and insulin signaling were also maintained in the fatty liver induced by Mttp deletion. Thus, blocking VLDL secretion causes hepatic steatosis without insulin resistance, and there is little effect on muscle triglyceride stores or adiposity.     

The low density lipoprotein receptor prevents secretion of dense apoB100-containing lipoproteins from the liver

The assembly and secretion of very low density lipoproteins (VLDL) require microsomal triglyceride transfer protein (MTP). Recent evidence also suggests a role for the low density lipoprotein (LDL) receptor in this process. However, the relative importance of MTP in the two steps of VLDL assembly and the specific role of the LDL receptor still remain unclear. To further investigate the role of MTP and the LDL receptor in VLDL assembly, we bred mice harboring "floxed" Mttp alleles (Mttpflox/flox) and a Cre transgene on a low-density lipoprotein receptor-deficient background to generate mice with double deficiency in the liver (Ldlr-/- MttpDelta/Delta). In contrast to the plasma of Ldlr+/+ MttpDelta/Delta mice, the plasma of Ldlr-/- MttpDelta/Delta mice contained apoB100. Accordingly, Ldlr-/- MttpDelta/Delta but not Ldlr+/+ MttpDelta/Delta hepatocytes secreted apoB100-containing lipoprotein particles. The secreted lipoproteins were of LDL and HDL sizes but no VLDL-sized lipoproteins could be detected. These findings indicate that hepatic LDL receptors function as "gatekeepers" targeting dense apoB100-containing lipoproteins for degradation. In addition, these results suggest that very low levels of MTP are insufficient to mediate the second step but sufficient for the first step of VLDL assembly.     

Alpha-tocopherol is secreted from rat liver in very low density lipoproteins

Three separate studies were carried out to test the hypothesis that rat liver secretes vitamin E (alpha-tocopherol) within very low density lipoproteins (VLDL). i) When the clearance of plasma chylomicrons (CM) and VLDL was blocked by the administration of Triton WR-1339, alpha-tocopherol concentrations increased linearly with time in both classes of triacylglycerol-rich lipoproteins, although accumulation rates within VLDL exceeded those within CM. For fasted rats, appearance of alpha-tocopherol in VLDL persisted at slightly reduced rates. alpha-Tocopherol and triglycerides in the VLDL fraction responded to Triton WR-1339 administration by coordinate increases. In contrast to the situation in serum, alpha-tocopherol concentrations decreased in the liver following injection of Triton. ii) In order to inhibit the secretion of hepatic lipoproteins containing apolipoprotein B (apoB), rats were fed a diet containing orotic acid. This resulted in a reduction of apoB and alpha-tocopherol concentrations in serum and VLDL, whereas the vitamin E content of liver was increased. iii) In primary cultures of hepatocytes, alpha-tocopherol was secreted into the culture media predominantly within VLDL. We, therefore, conclude that the liver secretes alpha-tocopherol within VLDL and in this way contributes to the maintenance of serum vitamin E concentrations.     

ApoB100 is required for increased VLDL-triglyceride secretion by microsomal triglyceride transfer protein in ob/ob mice

Microsomal triglyceride transfer protein (Mttp) is a key player in the assembly and secretion of hepatic very low density lipoproteins (VLDL). Here we determined the effects of Mttp overexpression on hepatic triglyceride (TG) and VLDL secretion in leptin-deficient (ob/ob) mice, specifically in relation to apolipoproteinB (apoB) isoforms. We crossed Apobec1(-/-) mice with congenic ob/ob mice to generate apoB100-only ob/ob mice (A-ob/ob). The obesity phenotype in both genotypes was similar, but A-ob/ob mice had greater hepatic TG content. Administration of recombinant adenovirus expressing murine Mttp cDNA (Ad-mMTP) increased hepatic Mttp content and activity and increased hepatic VLDL-TG secretion in A-ob/ob mice. However, despite equivalent overexpression of Mttp, there was no change in VLDL-TG secretion in ob/ob mice in a wild-type Apobec1 background. Metabolic labeling studies in primary hepatocytes from A-ob/ob mice demonstrated that Ad-mMTP increased triglyceride secretion without changing the synthesis and secretion of apoB100, suggesting greater incorporation of TG into existing VLDL particles rather than increased particle number. Ad-mMTP administration failed to increase hepatic VLDL secretion in lean Apobec1(-/-) mice or controls. By contrast, VLDL secretion increased and hepatic TG content decreased following Ad-mMTP administration to human APOB transgenic mice crossed into the Apobec1(-/-) line. These findings demonstrate that Ad-mMTP increases murine hepatic VLDL-TG secretion only in the apoB100 background, and even then only in situations with either increased hepatic TG accumulation or increased apoB100 expression.     

A study of hyperlipidaemia induced by ascites tumours.

In mice four stages of hyperlipidaemia induced by Ehrlich ascites tumour could be distinguished. Hyperlipidaemia is characterized mainly by increased serum VLDL content accompanied by high triglyceride concentration. The only exception was the regressive stage III where the serum lipid level (VLDL) has temporarily decreased. From the results obtained with the simultaneous examination of the liver, mesenteric fat tissue, tumour cell and ascites plasma lipids, it may be concluded that the endogeneous fat mobilization induced by tumour cells via increased VLDL synthesis and secretion of liver will lead to hyperlipidaemia and to the total depletion of the lipid stores. Rapid and reversible fall in lipid level following the withdrawal of ascites fluid in tumorous animals demonstrated clearly the direct effect of the tumour cells on lipid-lipoprotein metabolism of the host organism.     

Hypolipidemic effect of intravenous pluronic L-81 in fasted rats treated with Triton WR-1339: possible inhibition of hepatic lipoprotein secretion

Pluronic L-81 (L-81), a non-ionic hydrophobic surfactant, is a powerful inhibitor of the secretion of lipid-transporting chylomicrons from intestinal epithelial cells to lymph. Since the other major organ that secretes lipoproteins into the circulation is the liver, whose principal lipid secretory product is very low density lipoprotein (VLDL), we tested the hypothesis that L-81 will also inhibit hepatic lipid secretion. Rats were fasted so that they had little lipid input from the intestine. We then administered Triton WR-1339 (tyloxapol) intravenously to block peripheral utilization of VLDL, causing plasma lipids to rise rapidly. Some animals were also given L-81 intravenously to test whether the L-81 would retard the tyloxapol-induced rise in plasma lipids. Administration of tyloxapol alone (250 mg/kg) increased plasma triglyceride, phospholipid and cholesterol concentrations considerably. Simultaneous administration of a small dose of L-81 (6 mg/kg) markedly reduced the rise in plasma triglyceride, particularly in the first hour (by 45%). L-81 also diminished the rise in plasma phospholipid and cholesterol, but to a lesser extent (30%). In the fasting rat, most of the plasma triglyceride is in VLDL; therefore, L-81 probably acts by decreasing the secretion of hepatic VLDL. Thus, Pluronic L-81 may be a useful tool for examining the secretion and metabolism of hepatic lipoproteins, in particular, VLDL.     

Apolipoprotein B and triacylglycerol secretion in human triacylglycerol hydrolase transgenic mice

Apolipoprotein B (apoB)-containing lipoproteins play a critical role in whole body lipid homeostasis and the pathogenesis of atherosclerosis. The assembly of hepatic apoB-containing lipoproteins, VLDL, is governed by the availability of lipids, including triacylglycerol (TG). The majority of TG associated with VLDL is derived from the hepatic cytoplasmic lipid stores by a process involving lipolysis followed by reesterification. Microsomal triacylglycerol hydrolase (TGH) has been demonstrated to play a role in the lipolysis/reesterification process. To evaluate the potential regulatory role of TGH in hepatic VLDL assembly, we developed inducible transgenic mice expressing a human TGH minigene under the control of the mouse metallothionein promoter. Induction of human TGH by zinc resulted in liver-specific expression of the enzyme associated with 3- to 4-fold increases in lipolytic activity that could be attenuated with a TGH-specific inhibitor. Augmented TGH activity led to increased secretion of newly synthesized apoB and plasma TG levels. These results suggest that increased hepatic expression of TGH leads to a more proatherogenic plasma lipid and apoB profile.     

Secretion of alpha-tocopherol from cultured rat hepatocytes

Primary cultures of rat hepatocytes and rat liver perfusions were used to study hepatic secretion of alpha-tocopherol. The secretion of alpha-tocopherol from hepatocytes in culture was linear with time for 4 h. Ultracentrifugation of the medium revealed that 89.4 +/- 2.1% of alpha-tocopherol secreted during 4 h incubation was associated with the very-low density lipoprotein fraction (VLDL, d less than 1.006 g/ml). Oleic acid had no significant effect on the secretory rate of alpha-tocopherol, whereas eicosapentaenoic acid reduced the amount of alpha-tocopherol secreted to 48.4 +/- 12.7% of the control value after 20 h incubation (P less than 0.01). Monensin, a known inhibitor of VLDL secretion, reduced the secretion of alpha-tocopherol to 14.1 +/- 4.3% of the control value (P less than 0.02). Colchicine and chloroquine inhibited the secretion of alpha-tocopherol in the same order of magnitude as monensin. Hepatic perfusion after intravenous injection of in vivo labeled alpha-[3H]tocopherol lymph, showed that about 75% of the secreted radioactivity was in the VLDL fraction. From these results we conclude that most alpha-tocopherol is secreted from the liver associated with nascent VLDL in rats.     

Hepatic CTP:phosphocholine cytidylyltransferase-alpha is a critical predictor of plasma high density lipoprotein and very low density lipoprotein

CTP:phosphocholine cytidylyltransferase (CT) is the key regulatory enzyme in the CDP-choline pathway for the biosynthesis of phosphatidylcholine (PC). We previously generated a mouse in which the hepatic CTalpha gene was specifically inactivated by the cre/loxP procedure. In CTalpha knock-out mice, plasma high density lipoprotein (HDL) and very low density lipoprotein (VLDL) levels were markedly lower than in wild type mice (Jacobs, R. L., Devlin, C., Tabas, I., and Vance, D. E. (2004) J. Biol. Chem. 279, 47402-47410.) To investigate the mechanism(s) responsible for the decrease in plasma lipoprotein levels, we isolated primary hepatocytes from knock-out and wild type mice. ABCA1 expression was reduced in knock-out hepatocytes and apoAI-dependent cholesterol, and PC efflux was impaired. When knock-out hepatocytes were infected with an adenovirus expressing CTalpha, apoAI-dependent PC efflux returned partially, whereas cholesterol efflux and ABCA1 levels were not restored to normal levels. Adenoviral expression of CTalpha did not increase VLDL secretion in knock-out hepatocytes, even though cellular PC levels returned to normal. However, in vivo adenoviral delivery of CTalpha normalized plasma HDL and VLDL levels in knock-out mice. The observations demonstrate that hepatic PC biosynthesis is a key player in maintaining plasma VLDL and HDL, and further underscores the importance of the liver in HDL formation.     

Mechanism of hypertriglyceridemia in CTP:phosphoethanolamine cytidylyltransferase-deficient mice

Phosphatidylethanolamine is an important inner-leaflet phospholipid, and CTP:phosphoethanolamine cytidylyltransferase-Pcyt2 acts as the main regulator of the de novo phosphatidylethanolamine synthesis from ethanolamine and diacylglycerol. Complete deletion of the mouse Pcyt2 gene is embryonic lethal, and the single-allele deficiency leads to development of the metabolic syndrome phenotype, including liver steatosis, hypertriglyceridemia, obesity, and insulin resistance. This study aimed to specifically elucidate the mechanisms of hypertriglyceridemia in Pcyt2 heterozygous mice (Pcyt2(+/-)). Evidence here shows that unlike 8 week-old mice, 32 week- and 42 week-old Pcyt2(+/-) mice experience increased VLDL secretion and liver microsomal triglyceride transfer protein activity. Older Pcyt2(+/-) mice also demonstrate increased levels of postprandial plasma TAGs, increased stimulation of genes responsible for intestinal lipid absorption, transport and chylomicron secretion, and dramatically elevated plasma Angptl4, apoB-100, and apoB-48 content. In addition, plasma HL and LPL activities and TAG clearance following a lipid challenge were significantly reduced in Pcyt2(+/-) mice relative to control littermates. Collectively, these results establish that the hypertriglyceridemia that accompanies Pcyt2 deficiency is the result of multiple metabolic adaptations, including elevated hepatic and intestinal lipoprotein secretion and stimulated expression and/or activity of genes involved in lipid absorption and transport and lipoprotein assembly, together with reduced plasma TAG clearance and utilization with peripheral tissues.     

Carbenoxolone treatment attenuates symptoms of metabolic syndrome and atherogenesis in obese, hyperlipidemic mice

Glucocorticoids, which are well established to regulate body fat mass distribution, adipocyte lipolysis, hepatic gluconeogenesis, and hepatocyte VLDL secretion, are speculated to play a role in the pathology of metabolic syndrome. Recent focus has been on the activity of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), which is capable of regenerating, and thus amplifying, glucocorticoids in key metabolic tissues such as liver and adipose tissue. To determine the effects of global 11beta-HSD1 inhibition on metabolic syndrome risk factors, we subcutaneously injected "Western"-type diet-fed hyperlipidemic mice displaying moderate or severe obesity [LDL receptor (LDLR)-deficient (LDLR(-/-)) mice and mice derived from heterozygous agouti (A(y)/a) and homozygous LDLR(-/-) breeding pairs (A(y)/a;LDLR(-/-) mice)] with the nonselective 11beta-HSD inhibitor carbenoxolone for 4 wk. Body composition throughout the study, end-point fasting plasma, and extent of hepatic steatosis and atherosclerosis were assessed. This route of treatment led to detection of high levels of carbenoxolone in liver and fat and resulted in decreased weight gain due to reduced body fat mass in both mouse models. However, only A(y)/a;LDLR(-/-) mice showed an effect of 11beta-HSD1 inhibition on fasting insulin and plasma lipids, coincident with a reduction in VLDL due to mildly increased VLDL clearance and dramatically decreased hepatic triglyceride production. A(y)/a;LDLR(-/-) mice also showed a greater effect of the drug on reducing atherosclerotic lesion formation. These findings indicate that subcutaneous injection of an 11beta-HSD1 inhibitor allows for the targeting of the enzyme in not only liver, but also adipose tissue, and attenuates many metabolic syndrome risk factors, with more pronounced effects in cases of severe obesity and hyperlipidemia.     

Metabolic homeostasis in mice with disrupted Clock gene expression in peripheral tissues

The role of peripheral vs. central circadian rhythms and Clock in the maintenance of metabolic homeostasis and with aging was examined by using Clock(Delta19)+MEL mice. These have preserved suprachiasmatic nucleus and pineal gland rhythmicity but arrhythmic Clock gene expression in the liver and skeletal muscle. Clock(Delta19)+MEL mice showed fasting hypoglycemia in young-adult males, fasting hyperglycemia in older females, and substantially impaired glucose tolerance overall. Clock(Delta19)+MEL mice had substantially reduced plasma insulin and plasma insulin/glucose nocturnally in males and during a glucose tolerance test in females, suggesting impaired insulin secretion. Clock(Delta19)+MEL mice had reduced hepatic expression and loss of rhythmicity of gck, pfkfb3, and pepck mRNA, which is likely to impair glycolysis and gluconeogenesis. Clock(Delta19)+MEL mice also had reduced glut4 mRNA in skeletal muscle, and this may contribute to poor glucose tolerance. Whole body insulin tolerance was enhanced in Clock(Delta19)+MEL mice, however, suggesting enhanced insulin sensitivity. These responses occurred although the Clock(Delta19) mutation did not cause obesity and reduced plasma free fatty acids while increasing plasma adiponectin. These studies on clock-gene disruption in peripheral tissues and metabolic homeostasis provide compelling evidence of a relationship between circadian rhythms and the glucose/insulin and adipoinsular axes. It is, however, premature to declare that clock-gene disruption causes the full metabolic syndrome.     

Burn and smoke inhalation injury in sheep depletes vitamin E: kinetic studies using deuterated tocopherols

To test the hypothesis that burn and smoke injury will deplete tissue alpha-tocopherol and cause its faster plasma disappearance, deuterium-labeled vitamin E was administered to sheep exposed to both surface skin burn and smoke insufflation, which cause injuries similar to those of human victims of fire accidents. Two different protocols were used: (1) deuterated vitamin E was administered orally with food at time 0 (just before injury) or (2) the labeled vitamin E was administered orally with food the day before injury. The animals, which had been operatively prepared seven days before, were anesthetized and then received both 40% body surface area third-degree burn and 48 breaths of cotton smoke or sham injuries. All were resuscitated with Ringer's lactate solution (4 ml/kg/% BSA burn/24 h) and mechanically ventilated. Blood samples were collected at various times after vitamin E dosing. In both studies the depletion of plasma alpha-tocopherol was faster in the injured sheep. The sheep given deuterated vitamin E 24 h before injury had similar maximum alpha-tocopherol concentrations at similar times. The exponential rates of alpha-tocopherol disappearance were 1.5 times greater and half-lives were 12 h shorter (p < 0.05) in the injured sheep. In separate studies, various tissues were obtained from sheep that were sacrificed from 4 to 48 h after injury. The liver alpha-tocopherol concentrations in sheep killed at various times after injury seem to show a linear decrease at a rate of 0.1 nmol alpha-tocopherol/g liver per hour, suggesting that the liver is supplying alpha-tocopherol to maintain the plasma and lung alpha-tocopherol concentrations, but that this injury is so severe the liver is unable to maintain lung alpha-tocopherol concentrations. These findings suggest that alpha-tocopherol should be administered to burn patients to prevent vitamin E depletion and to protect against oxidative stress from burn injury.     

Hormone-sensitive lipase deficiency in mice changes the plasma lipid profile by affecting the tissue-specific expression pattern of lipoprotein lipase in adipose tissue and muscle.

Hormone-sensitive lipase (HSL) is believed to play an important role in the mobilization of fatty acids from triglycerides (TG), diglycerides, and cholesteryl esters in various tissues. Because HSL-mediated lipolysis of TG in adipose tissue (AT) directly feeds non-esterified fatty acids (NEFA) into the vascular system, the enzyme is expected to affect many metabolic processes including the metabolism of plasma lipids and lipoproteins. In the present study we examined these metabolic changes in induced mutant mouse lines that lack HSL expression (HSL-ko mice). During fasting, when HSL is normally strongly induced in AT, HSL-ko animals exhibited markedly decreased plasma concentrations of NEFA (-40%) and TG (-63%), whereas total cholesterol and HDL cholesterol levels were increased (+34%). Except for the increased HDL cholesterol concentrations, these differences were not observed in fed animals, in which HSL activity is generally low. Decreased plasma TG levels in fasted HSL-ko mice were mainly caused by decreased hepatic very low density lipid lipoprotein (VLDL) synthesis as a result of decreased NEFA transport from the periphery to the liver. Reduced NEFA transport was also indicated by a depletion of hepatic TG stores (-90%) and strongly decreased ketone body concentrations in plasma (-80%). Decreased plasma NEFA and TG levels in fasted HSL-ko mice were associated with increased fractional catabolic rates of VLDL-TG and an induction of the tissue-specific lipoprotein lipase (LPL) activity in cardiac muscle, skeletal muscle, and white AT. In brown AT, LPL activity was decreased. Both increased VLDL fractional catabolic rates and increased LPL activity in muscle were unable to provide the heart with sufficient NEFA, which led to decreased tissue TG levels in cardiac muscle. Our results demonstrate that HSL deficiency markedly affects the metabolism of TG-rich lipoproteins by the coordinate down-regulation of VLDL synthesis and up-regulation of LPL in muscle and white adipose tissue. These changes result in an "anti-atherogenic" lipoprotein profile.     

Nascent VLDL from liver perfusions of cynomolgus monkeys are preferentially enriched in RRR- compared with SRR-alpha-tocopherol: studies using deuterated tocopherols

The transport and secretion of vitamin E in lipoproteins have been studied in cynomolgus monkeys fed tocopherols labeled with different amounts of deuterium. The animals were fed a single dose of vitamin E containing 60 mumol of each 2R,4'R,8'R-alpha-(5,7-(C2H3)2)tocopheryl acetate (d6-RRR-alpha-tocopheryl acetate; alpha-tocopherol with natural stereochemistry), 2S,4'R,8'R-alpha-5-(C2H3)tocopheryl acetate (d3-SRR-alpha-tocopheryl acetate; alpha-tocopherol with unnatural stereochemistry), and 2R,4'R,8'R-gamma-(3,4-2H)tocopherol (d2-RRR-gamma-tocopherol; gamma-tocopherol with natural stereochemistry). Chylomicrons, as well as the other plasma lipoproteins, contained equal concentrations of all three tocopherols at the earliest time points after feeding suggesting that all three tocopherols were absorbed equally. At later times plasma lipoproteins became preferentially enriched in d6-RRR-alpha-tocopherol. This is likely to be due to hepatic secretion of VLDL (very low density lipoproteins) and other lipoproteins, which were enriched in d6-RRR-alpha-tocopherol, as demonstrated in the lipoproteins isolated from perfused livers that had been obtained 24 h following the administration of the deuterated tocopherols. Taken together these data demonstrate that the liver, not the intestine, is the likely site of discrimination between tocopherol isomers and that the liver secretes nascent lipoproteins preferentially enriched in d6-RRR-alpha-tocopherol.     

Regulation of hepatic secretion of very low density lipoprotein by dietary cholesterol.

Male rats were fed a cholesterol-free diet or the same diet supplemented with either 0.05, 0.1, 0.25, 0.5, 1, or 2% C for 21 days to investigate the effects of cholesterol on secretion of very low density lipoprotein (VLDL). Cholesterol feeding increased plasma and hepatic concentrations of triglyceride (TG) and cholesteryl esters (CE) in a dose-dependent manner. Plasma VLDL and low density lipoprotein (LDL) lipids were elevated by cholesterol feeding, while the high density lipoprotein (HDL) lipids were reduced. The secretion of the VLDL by perfused livers from these cholesterol-fed rats was examined to establish the relationship between the accumulation of lipids in the liver and the concurrent hyperlipemia. Liver perfusions were carried out for 4 h with a medium containing bovine serum albumin (3% w/v), glucose (0.1% w/v), bovine erythrocytes (30% v/v), and a 10-mCi 3H2O initial pulse. Oleic acid was infused to maintain a concentration of 0.6 mM. Hepatic secretion of VLDL-TG, PL (phospholipid), free cholesterol (FC), and CE increased in proportion to dietary cholesterol and was maximal at 0.5% cholesterol in these experiments in which TG synthesis was stimulated by oleic acid. Secretion of VLDL protein and apoB by the perfused liver was also increased. The molar ratios of surface (sum of PL and cholesterol) to core (sum of TG and CE) lipid components of the secreted VLDL, regardless of cholesterol feeding, were the same, as were the mean diameters of the secreted particles. The molar ratios of surface to core lipid of VLDL isolated from the plasma also were not affected by cholesterol feeding. During perfusion with oleic acid of livers from the rats fed the higher levels of cholesterol, the hepatic concentration of CE decreased, while the level of TG was not changed. We conclude that the hypercholesterolemia and hypertriglyceridemia that occur in vivo from cholesterol feeding, concurrent with accumulation of CE and TG in the liver, must result, in part, from increased hepatic secretion of all VLDL lipids and apoB. The VLDL particles produced by the liver of the cholesterol-fed rat are assembled without modification of the surface lipid ratios (PL/FC), but contain a greater proportion of cholesteryl esters compared to triglyceride in the core, because of the stimulated transport of CE from the expanded pool in the liver.(ABSTRACT TRUNCATED AT 400 WORDS)     

Compensatory increase in hepatic lipogenesis in mice with conditional intestine-specific Mttp deficiency

Microsomal TG transfer protein (MTTP) is required for the assembly and secretion of TG (TG)-rich lipoproteins from both enterocytes and hepatocytes. Liver-specific deletion of Mttp produced a dramatic reduction in plasma very low density lipoprotein-TG and virtually eliminated apolipoprotein B100 (apoB100) secretion yet caused only modest reductions in plasma apoB48 and apoB48 secretion from primary hepatocytes. These observations prompted us to examine the phenotype following intestine-specific Mttp deletion because murine, like human enterocytes, secrete virtually exclusively apoB48. We generated mice with conditional Mttp deletion in villus enterocytes (Mttp-IKO), using a tamoxifen-inducible, intestine-specific Cre transgene. Villus enterocytes from chow-fed Mttp-IKO mice contained large cytoplasmic TG droplets and no chylomicron-sized particles within the secretory pathway. Chow-fed, Mttp-IKO mice manifested steatorrhea, growth arrest, and decreased cholesterol absorption, features that collectively recapitulate the phenotype associated with abetalipoproteinemia. Chylomicron secretion was reduced dramatically in vivo, in conjunction with an approximately 80% decrease in apoB48 secretion from primary enterocytes. Additionally, although plasma and hepatic cholesterol and TG content were decreased, Mttp-IKO mice demonstrated a paradoxical increase in both hepatic lipogenesis and very low density lipoprotein secretion. These findings establish distinctive features for MTTP involvement in intestinal chylomicron assembly and secretion and suggest that hepatic lipogenesis undergoes compensatory induction in the face of defective intestinal TG secretion.     

Transbilayer distribution of alpha-tocopherol and lipid asymmetry in nerve tissue membranes

The alpha-tocopherol content and fatty acid composition of lipids in various types of nervous tissue membranes were studied. The transbilayer distribution of alpha-tocopherol and polyunsaturated fatty acids in liposomes and plasma membranes of synaptosomes was examined. It was shown that both phosphatidylethanolamine and phosphatidylserine are localized predominantly in the inner monolayer and they contain the bulk of polyenoic fatty acid residues. alpha-Tocopherol incorporated into liposomes from synaptosome plasma membrane lipids and present in synaptosome plasma membranes is also predominantly localized in the inner monolayers. No asymmetrical distribution of incorporated alpha-tocopherol was observed in liposomes prepared from a single phospholipid, e.g., dioleoylphosphatidylcholine.     

Hepatic SH2B1 and SH2B2 Regulate Liver Lipid Metabolism and VLDL Secretion in Mice

SH2B1 is an SH2 and PH domain-containing adaptor protein. Genetic deletion of SH2B1 results in obesity, type 2 diabetes, and fatty liver diseases in mice. Mutations in SH2B1 are linked to obesity in humans. SH2B1 in the brain controls energy balance and body weight at least in part by enhancing leptin sensitivity in the hypothalamus. SH2B1 in peripheral tissues also regulates glucose and lipid metabolism, presumably by enhancing insulin sensitivity in peripheral metabolically-active tissues. However, the function of SH2B1 in individual peripheral tissues is unknown. Here we generated and metabolically characterized hepatocyte-specific SH2B1 knockout (HKO) mice. Blood glucose and plasma insulin levels, glucose tolerance, and insulin tolerance were similar between HKO, albumin-Cre, and SH2B1^f/f mice fed either a normal chow diet or a high fat diet (HFD). Adult-onset deletion of SH2B1 in the liver either alone or in combination with whole body SH2B2 knockout also did not exacerbate HFD-induced insulin resistance and glucose intolerance. Adult-onset, but not embryonic, deletion of SH2B1 in the liver attenuated HFD-induced hepatic steatosis. In agreement, adult-onset deletion of hepatic SH2B1 decreased the expression of diacylglycerol acyltransferase-2 (DGAT2) and increased the expression of adipose triglyceride lipase (ATGL). Furthermore, deletion of liver SH2B1 in SH2B2 null mice attenuated very low-density lipoprotein (VLDL) secretion. These data indicate that hepatic SH2B1 is not required for the maintenance of normal insulin sensitivity and glucose metabolism; however, it regulates liver triacylglycerol synthesis, lipolysis, and VLDL secretion.