Uptake Of Remnants Research Articles

P32 EVALUATION OF SYSTEMIC INFLAMMATION IN FAMILIAL HYPERCHOLESTEROLEMIA BY BIOMARKER PROFILING ON BIOCHIP ARRAYS: IMPACT OF LDL-APHERESIS

The extent and kinetics of postprandial changes are highly variable and are modulated by numerous factors including diet, lifestyle conditions, genetic background and pathological conditions. This review focuses on dietary factors affecting postprandial chylomicron and lipoprotein metabolism in humans. The first key step in the process is fat digestion within the stomach and small intestine which can be modulated by fat and dietary fiber amounts and types.The second key step is intestinal absorption per se. The uptake and secretion of absorbed nutrients in chylomicrons is affected by numerous factors, including nutrients themselves. The amount of dietary triglycerides as well as the nature of fatty acids ingested from habitual diet or present in a test meal modulates the extent of chylomicron and plasma triglyceride levels postprandially. The amount of dietary cholesterol also modulates the extent of postprandial chylomicron and plasma triglycerides. Other nutrients can alter the postprandial occurrence of chylomicrons such as proteins, alcohol, carbohydrates or fibers. Addition of glucose and more markedly fructose, to a fat test meal increases the accumulation of chylomicrons, as does a high-glycemic index meal. In contrast, some sources of dietary fibers such as oat bran noticeably decrease the occurrence of chylomicron postprandially. Overall, changing the habitual dietary pattern, for instance from a Western-type diet to Mediterranean-type one, can in turn alter the postprandial response of the subjects. The last step in postprandial lipid metabolism is clearance from plasma through combined lipolysis processes and tissue uptake of chylomicron remnants. Both the processes can also be modulated by the nutrients.Because most day-time is usually spent in a postprandial state, the link between dietary patterns and cardiovascular health or risk is clearly mediated by the secretion and clearance rates of chylomicrons in the circulation.

Hepatic SR-BI, not endothelial lipase, expression determines biliary cholesterol secretion in mice

High density lipoprotein cholesterol is thought to represent a preferred source of sterols secreted into bile following hepatic uptake by scavenger receptor class B type I (SR-BI). The present study aimed to determine the metabolic effects of an endothelial lipase (EL)-mediated stimulation of HDL cholesterol uptake on liver lipid metabolism and biliary cholesterol secretion in wild-type, SR-BI knockout, and SR-BI overexpressing mice. In each model, injection of an EL expressing adenovirus decreased plasma HDL cholesterol (P < 0.001) whereas hepatic cholesterol content increased (P < 0.05), translating into decreased expression of sterol-regulatory element binding protein 2 (SREBP2) and its target genes HMG-CoA reductase and LDL receptor (each P < 0.01). Biliary cholesterol secretion was dependent on hepatic SR-BI expression, being decreased in SR-BI knockouts (P < 0.001) and increased following hepatic SR-BI overexpression (P < 0.001). However, in each model, biliary secretion of cholesterol, bile acids, and phospholipids as well as fecal bile acid and neutral sterol content, remained unchanged in response to EL overexpression. Importantly, hepatic ABCG5/G8 expression did not correlate with biliary cholesterol secretion rates under these conditions. These results demonstrate that an acute decrease of plasma HDL cholesterol levels by overexpressing EL increases hepatic cholesterol content but leaves biliary sterol secretion unaltered. Instead, biliary cholesterol secretion rates are related to the hepatic expression level of SR-BI. These data stress the importance of SR-BI for biliary cholesterol secretion and might have relevance for concepts of reverse cholesterol transport.

Clinical Implications of Diffuse Hepatic Uptake Observed in Postablative and Post-Therapeutic I-131 Scans

Diffuse hepatic uptake of I-131 either on diagnostic or post-therapeutic scans is a usual finding in patients with differentiated thyroid carcinoma. The aim of this study was to evaluate the frequency and clinical significance of diffuse hepatic uptake of radioiodine on post-therapeutic (PT) and postablative (PA) whole-body scans.A total of 720 PA and 172 PT I-131 scans in a total of 732 patients with differentiated thyroid carcinoma were retrospectively reviewed. Residual thyroid tissue and diffuse liver uptake of I-131 were classified from 0 to 4. The correlation between the liver and thyroid remnant uptake score, dose of radioiodine, serum thyroglobuline (Tg), liver function test levels (ALT, AST), liver ultrasonography, presence of metastatic foci, and recurrent disease were examined.Diffuse hepatic uptake was observed in 701 of 722 (94.2%) PA and 162 of 172 (97%) PT whole body I-131 scans. Hepatic radioiodine uptake was positively correlated with the dose of administered I-131 and increased levels of serum AST and ALT. Liver uptake scores of patients with hepatosteatosis were significantly higher than all study groups. However, no evidence of a relationship between diffuse visualization of liver and serum thyroglobulin levels, uptake score of thyroid remnants, presence of local or distant metastatic foci on I-131 scan, and recurrence rate could be demonstrated.The lack of correlation between hepatic radioactive iodine (RAI) uptake and Tg levels, functioning metastatic tissue or thyroid remnants suggests that this finding may be related to factors other than thyroid tissue. The positive correlation between administered RAI dose, hepatic enzymes and hepatosteatosis support the conclusion that diffuse hepatic RAI uptake may be related to different mechanisms as well.

Incidence of radiation thyroiditis and thyroid remnant ablation success rates following 1110 MBq (30 mCi) and 3700 MBq (100 mCi) post‐surgical 131I ablation therapy for differentiated thyroid carcinoma

To evaluate the relationships between thyroid remnant (131)I uptake, radiation thyroiditis and remnant ablation success rate between lower (1110 MBq) and higher (3700 MBq) initial ablative (131)I dose for post-surgical ablation therapy for differentiated thyroid cancer. Patients having post-surgical administration of 1110 MBq (68 patients) or 3700 MBq (115 patients) (131)I were retrospectively reviewed. Thyroid remnant (131)I uptake on a 48 h post-administration scan was correlated with neck symptoms experienced. Patients were classified as having insignificant, mild or severe thyroiditis based on symptoms. Absent thyroid bed (131)I uptake on a follow-up 74 MBq (131)I study was considered successful ablation. 183 patients were included. Median (131)I remnant uptake was 37 MBq. 21% (39/183) of patients developed thyroiditis. Incidence and severity of thyroiditis increased with increasing remnant (131)I activity (P <or= 0.0001). Risk of thyroiditis increased by 64% for every 37 MBq increase in remnant (131)I uptake. OR = 1.64 [1.36-1.99]. Severe thyoriditis was only seen with remnant (131)I uptake > 73 MBq. For patients treated with 1110 MBq and 3700 MBq, incidence of thyroiditis was 12% and 27% (P = 0.02) and remnant ablation success rate was 76% and 84% (P = NS), respectively. Occurrence of thyroiditis did not correlate with successful ablation. Incidence and severity of radiation thyroiditis following (131)I remnant ablation therapy is directly related to thyroid remnant (131)I uptake. As 1110 MBq (131)I is associated with a significantly lower frequency of thyroiditis but similar remnant ablation rate to 3700 MBq, it warrants consideration for thyroid remnant ablation particularly in patients with low risk disease.

The hepatic uptake of VLDL in lrp−ldlr−/−vldlr−/− mice is regulated by LPL activity and involves proteoglycans and SR-BI

LPL activity plays an important role in preceding the VLDL remnant clearance via the three major apolipoprotein E (apoE)-recognizing receptors: the LDL receptor (LDLr), LDL receptor-related protein (LRP), and VLDL receptor (VLDLr). The aim of this study was to determine whether LPL activity is also important for VLDL remnant clearance irrespective of these receptors and to determine the mechanisms involved in the hepatic remnant uptake. Administration of an adenovirus expressing LPL (AdLPL) into lrp(-)ldlr(-/-)vldlr(-/-) mice reduced both VLDL-triglyceride (TG) and VLDL-total cholesterol (TC) levels. Conversely, inhibition of LPL by AdAPOC1 increased plasma VLDL-TG and VLDL-TC levels. Metabolic studies with radiolabeled VLDL-like emulsion particles showed that the clearance and hepatic association of their remnants positively correlated with LPL activity. This hepatic association was independent of the bridging function of LPL and HL, since heparin did not reduce the liver association. In vitro studies demonstrated that VLDL-like emulsion particles avidly bound to the cell surface of primary hepatocytes from lrp(-)ldlr(-/-)vldlr(-/-) mice, followed by slow internalization, and involved heparin-releaseable cell surface proteins as well as scavenger receptor class B type I (SR-BI). Collectively, we conclude that hepatic VLDL remnant uptake in the absence of the three classical apoE-recognizing receptors is regulated by LPL activity and involves heparan sulfate proteoglycans and SR-BI.

Macronutrient intake and modulation on chylomicron production and clearance

The extent and kinetics of postprandial changes are highly variable and are modulated by numerous factors including diet, lifestyle conditions, genetic background and pathological conditions. This review focuses on dietary factors affecting postprandial chylomicron and lipoprotein metabolism in humans. The first key step in the process is fat digestion within the stomach and small intestine which can be modulated by fat and dietary fiber amounts and types. The second key step is intestinal absorption per se. The uptake and secretion of absorbed nutrients in chylomicrons is affected by numerous factors, including nutrients themselves. The amount of dietary triglycerides as well as the nature of fatty acids ingested from habitual diet or present in a test meal modulates the extent of chylomicron and plasma triglyceride levels postprandially. The amount of dietary cholesterol also modulates the extent of postprandial chylomicron and plasma triglycerides. Other nutrients can alter the postprandial occurrence of chylomicrons such as proteins, alcohol, carbohydrates or fibers. Addition of glucose and more markedly fructose, to a fat test meal increases the accumulation of chylomicrons, as does a high-glycemic index meal. In contrast, some sources of dietary fibers such as oat bran noticeably decrease the occurrence of chylomicron postprandially. Overall, changing the habitual dietary pattern, for instance from a Western-type diet to Mediterranean-type one, can in turn alter the postprandial response of the subjects. The last step in postprandial lipid metabolism is clearance from plasma through combined lipolysis processes and tissue uptake of chylomicron remnants. Both the processes can also be modulated by the nutrients. Because most day-time is usually spent in a postprandial state, the link between dietary patterns and cardiovascular health or risk is clearly mediated by the secretion and clearance rates of chylomicrons in the circulation.

An ABC of apolipoprotein C-III: a clinically useful new cardiovascular risk factor?

Hypertriglyceridaemia, commonly found in subjects with obesity and type 2 diabetes mellitus, is associated with increased risk of coronary heart disease (CHD). Apolipoprotein C-III (apoC-III) plays an important role in regulating the metabolism of triglyceride-rich lipoproteins (TRLs) and may provide a new approach to assessing hypertriglyceridaemia. We review the role of apoC-III in regulating TRL metabolism and address the potential importance of apoC-III in clinical practice. Hypertriglyceridaemia is chiefly a consequence of alterations in the kinetics of TRLs, including overproduction and delayed clearance of very-low density lipoprotein (VLDL). ApoC-III is an inhibitor of lipoprotein lipase and of TRLs remnant uptake by hepatic lipoprotein receptors. Elevated apoC-III, usually resulting from hepatic overproduction of VLDL apoC-III, may cause accumulation of plasma TRLs leading to hypertriglyceridaemia. The results from recent observational studies demonstrate that apoC-III is a strong predictor of risk for CHD, but this chiefly relates to apoC-III in apoB-containing lipoproteins. Lifestyle and pharmacological intervention can correct hypertriglyceridaemia by a mechanism of action that regulates apoC-III transport. Targeting apoC-III metabolism may therefore be an important, new therapeutic approach to managing dyslipidaemia and CHD risk in obesity, insulin resistance and type 2 diabetes mellitus. However, further work is required to establish the practical aspects of measuring apoC-III in routine laboratory service and the precise therapeutic targets for serum total apoC-III and/or apoC-III in apoB-containing lipoproteins. While showing much promise as a potentially useful cardiovascular risk factor, apoC-III is not yet ready for prime time use in clinical practice.

Dietary sphingolipids lower plasma cholesterol and triacylglycerol and prevent liver steatosis

AbstractThe prevalence of dyslipidemia and obesity resulting from excess energy intake and physical inactivity is increasing. The liver plays a pivotal role in the systemic lipid homeostasis. Effective, natural dietary interventions that lower plasma lipids and promote liver health are needed. APOE*3Leiden mice were fed a Western‐type diet, supplemented with different sphingolipids, to determine their effect on plasma lipids and liver steatosis. Hepatic lipid levels and lipid‐related gene expression were also determined. Dietary sphingolipids dose‐dependently lowered both plasma cholesterol (C) and triglycerides (TG) in APOE*3Leiden mice. 1% Phytosphingosine (PS) reduced C and TG by 57 and 58%, respectively. PS (a) decreased the absorption of dietary C and free fatty acid but did not affect the intestinal TG lipolysis, (b) increased hepatic VLDL‐TG production whereas plasma lipolysis was not affected; and (c) increased the hepatic uptake of VLDL remnants. Hepatic mRNA levels indicated enhanced hepatic lipid synthesis and VLDL and LDL uptake. Livers of PS (1%) fed mice were lighter (−22%), less pale, and contained less cholesteryl ester (−61%) and TG (−56%). Furthermore, markers for liver inflammation (SAA) and liver damage (ALAT) were decreased by 74% and 79%, respectively in PS‐fed mice. Sphingolipids lower plasma C and TG and protect the liver from fat‐ and cholesterol‐induced steatosis. In a preliminary small double‐blind cross‐over study with six middle‐aged slightly overweight male volunteers the daily supplementation of one gram of PS to the diet resulted in a −9.8% (p = 0.0074) and – 13.2% (p = 0.0002) reduction of total C and LDL‐C, respectively. The C/HDL‐C ratio was not significantly affected (p = 0.0571). Due to the relatively low pre‐study levels of TG in the human volunteers, and the individual variability of TG levels, the TG lowering in humans was not significant in this first small study, but per individual there was a clear trend in TG lowering.

Lipoprotein lipase-facilitated uptake of LDL is mediated by the LDL receptor

LPL mediates the uptake of lipoproteins into different cell types independent of its catalytic activity. The mechanism of this process and its physiological relevance are not clear. Taking into account the importance of the endothelial barrier for lipoprotein uptake, in vitro studies with primary aortic endothelial cells from wild-type and low density lipoprotein receptor (LDLR)-deficient (LDLR(-/-)) mice were performed. Addition of LPL almost doubled the uptake of LDL into wild-type cells. However, there was virtually no LPL-mediated change of LDL uptake into LDLR(-/-) cells. Upregulation of LDLR by lipoprotein-deficient serum/lovastatin in wild-type cells resulted in a 7-fold increase of LPL-mediated LDL uptake. Uptake of chylomicron remnants was not affected by LDLR expression. In proteoglycan-deficient cells, LPL did not increase the uptake of lipoproteins. The physiological relevance of this pathway was studied in mice that were both LDLR(-/-) and transgenic for catalytically inactive LPL in muscle. In the presence of LDLR, inactive LPL reduced LDL cholesterol significantly (13-24%). In the absence of LDLR, LDL cholesterol was not affected by transgenic LPL. Metabolic studies showed that in the presence of LDLR, LPL increased the muscular uptake of LDL by 77%. In the absence of LDLR, transgenic LPL did not augment LDL uptake. Chylomicron uptake was not affected by the LDLR genotype. We conclude that LPL-mediated cellular uptake of LDL, but not of chylomicrons, is dependent on the presence of both LDLR and proteoglycans.

Atherogenic remnant lipoproteins: role for proteoglycans in trapping, transferring, and internalizing

Unraveling the mechanisms controlling remnant lipoprotein clearance is important, as these lipoproteins are highly atherogenic. The most critical molecule in this process is apoE, which mediates high-affinity binding of remnant lipoproteins to members of the LDL receptor (LDLR) family and cell-surface heparan sulfate proteoglycans (HSPGs), which have been shown to play major independent as well as cooperative roles in remnant lipoprotein clearance. While all the players may have been identified, our understanding of how they interact and function together continues to evolve. In this issue of the JCI, MacArthur et al. (see the related article beginning on page 153) demonstrated that HSPGs under normal physiological conditions are critically important in the clearance of remnant lipoproteins, independent of LDLR family members. The complexity of VLDL and chylomicron remnant clearance was exemplified by the studies of Jones et al., also in this issue (see the related article beginning on page 165). Despite defective clearance of LDL in mice with a deficiency in the adaptor protein controlling internalization of the LDLR, called autosomal recessive hypercholesterolemia (ARH), remnant lipoprotein clearance was not grossly abnormal. A likely explanation is that the abnormal LDLRs bind the remnants and then transfer them to another acceptor for internalization. While the studies clearly demonstrate that the LDLR-related protein 1 is not involved and suggest a role for an additional unidentified receptor, it remains a possibility that HSPGs are responsible for remnant uptake by hepatocytes in the presence of defective LDLR internalization.

Dietary sphingolipids lower plasma cholesterol and triacylglycerol and prevent liver steatosis in APOE*3Leiden mice1–3

The prevalence of dyslipidemia and obesity resulting from excess energy intake and physical inactivity is increasing. The liver plays a pivotal role in systemic lipid homeostasis. Effective, natural dietary interventions that lower plasma lipids and promote liver health are needed. Our goal was to determine the effect of dietary sphingolipids on plasma lipids and liver steatosis. APOE*3Leiden mice were fed a Western-type diet supplemented with different sphingolipids. Body cholesterol and triacylglycerol metabolism as well as hepatic lipid concentrations and lipid-related gene expression were determined. Dietary sphingolipids dose-dependently lowered both plasma cholesterol and triacylglycerol in APOE*3Leiden mice; 1% phytosphingosine (PS) reduced plasma cholesterol and triacylglycerol by 57% and 58%, respectively. PS decreased the absorption of dietary cholesterol and free fatty acids by 50% and 40%, respectively, whereas intestinal triacylglycerol lipolysis was not affected. PS increased hepatic VLDL-triacylglycerol production by 20%, whereas plasma lipolysis was not affected. PS increased the hepatic uptake of VLDL remnants by 60%. Hepatic messenger RNA concentrations indicated enhanced hepatic lipid synthesis and VLDL and LDL uptake. The net result of these changes was a strong decrease in plasma cholesterol and triacylglycerol. The livers of 1% PS-fed mice were less pale, 22% lighter, and contained 61% less cholesteryl ester and 56% less triacylglycerol than livers of control mice. Furthermore, markers of liver inflammation (serum amyloid A) and liver damage (alanine aminotransferase) decreased by 74% and 79%, respectively, in PS-fed mice. Sphingolipids lower plasma cholesterol and triacylglycerol and protect the liver from fat- and cholesterol-induced steatosis.

LDL composition in E2/2 subjects and LDL distribution by Apo E genotype in type 1 diabetes

Apo E plays an important role in chylomicron and VLDL remnant processing, uptake or conversion to LDL. The type of lipoprotein that isolates in the LDL density of E2/2 subjects was investigated and the effect of the apo E isoforms on LDL mass was determined in all genotypes in a large group of Type 1 diabetics. Analysis of the LDL composition of E2/2 homozygotes ( n = 6) compared to subjects with the common E3/3 isoform ( n = 6) demonstrated an enrichment in apo E, unesterified cholesterol, phospholipid and triglyceride relative to apo B in E2/2 subjects, more typical of a dense IDL remnant than of LDL. Although diabetics were studied, these findings are considered to reflect those of the general population. Comparison of the lipoprotein distribution of homozygous and heterozygous subjects revealed that, as genotype changed from E4/4 ( n = 22) to E3/4 ( n = 262), E3/3 ( n = 710) = E2/4 ( n = 30), E2/3 ( n = 151), E2/2 ( n = 6), LDL cholesterol decreased significantly in a stepwise manner. The decrease was not in a specific subgroup of LDL. In conclusion, for E2/2 subjects, lipoproteins isolated in the LDL density range appear to be composed mainly of dense IDL remnants and some Lp(a). The apo E isoform also has a significant effect on LDL concentration in both homozygotes and heterozygotes.

An investigation by electron microscopy of chylomicron remnant uptake by human monocyte-derived macrophages

Human monocyte-derived macrophages (HMM) internalise proatherogenic chylomicron remnants via several high affinity receptor pathways. However, the endocytic ultrastructures responsible for the uptake of chylomicron remnants by macrophages have not previously been described. In this study, we have utilised transmission electron microscopy together with colloidal gold-labelling of chylomicron remnants to investigate the pathways involved in macrophage uptake of chylomicron remnants. We found that macrophages internalise chylomicron remnants via surface-connected compartments of up to 600 nm as well as non-clathrin coated pits. Chylomicron remnants were found to be distributed internally in a number of endocytic vesicles including early cysternal endosomes, spherical late endosomes and tubular vesicular compartments. Uptake of chylomicron remnants by HMM via phagocytosis or macropinocytosis was excluded based on the observations that lipoproteins were not found in phagolysosomes nor modified by inhibitors of these two processes, respectively. The latter observation contrasts with previous reports of chylomicron remnant internalisation by macrophages of other species.

SR-BI-mediated selective lipid uptake segregates apoA-I and apoA-II catabolism

The HDL receptor scavenger receptor class B type I (SR-BI) binds HDL and mediates the selective uptake of cholesteryl ester. We previously showed that remnants, produced when human HDL(2) is catabolized in mice overexpressing SR-BI, become incrementally smaller, ultimately consisting of small alpha-migrating particles, distinct from pre-beta HDL. When mixed with mouse plasma, some remnant particles rapidly increase in size by associating with HDL without the mediation of cholesteryl ester transfer protein, LCAT, or phospholipid transfer protein. Here, we show that processing of HDL(2) by SR-BI-overexpressing mice resulted in the preferential loss of apolipoprotein A-II (apoA-II). Short-term processing generated two distinct, small alpha-migrating particles. One particle (8.0 nm diameter) contained apoA-I and apoA-II; the other particle (7.7 nm diameter) contained only apoA-I. With extensive SR-BI processing, only the 7.7 nm particle remained. Only the 8.0 nm remnants were able to associate with HDL. Compared with HDL(2), this remnant was more readily taken up by the liver than by the kidney. We conclude that SR-BI-generated HDL remnants consist of particles with or without apoA-II and that only those containing apoA-II associate with HDL in an enzyme-independent manner. Extensive SR-BI processing generates small apoA-II-depleted particles unable to reassociate with HDL and readily taken up by the liver. This represents a pathway by which apoA-I and apoA-II catabolism are segregated.

Apolipoprotein A-V Deficiency Results in Marked Hypertriglyceridemia Attributable to Decreased Lipolysis of Triglyceride-Rich Lipoproteins and Removal of Their Remnants

ApoAV, a newly discovered apoprotein, affects plasma triglyceride level. To determine how this occurs, we studied triglyceride-rich lipoprotein (TRL) metabolism in mice deficient in apoAV. No significant difference in triglyceride production rate was found between apoa5(-/-) mice and controls. The presence or absence of apoAV affected TRL catabolism. After the injection of 14C-palmitate and 3H-cholesterol labeled chylomicrons and (125)I-labeled chylomicron remnants, the disappearance of 14C, 3H, and (125)I was significantly slower in apoa5(-/-) mice relative to controls. This was because of diminished lipolysis of TRL and the reduced rate of uptake of their remnants in apoa5(-/-) mice. Observed elevated cholesterol level was caused by increased high-density lipoprotein (HDL) cholesterol in apoa5(-/-) mice. VLDL from apoa5(-/-) mice were poor substrate for lipoprotein lipase, and did not bind to the low-density lipoprotein (LDL) receptor as well as normal very-low-density lipoprotein (VLDL). LDL receptor levels were slightly elevated in apoa5(-/-) mice consistent with lower remnant uptake rates. These alterations may be the result of the lower apoE-to-apoC ratio found in VLDL isolated from apoa5(-/-) mice. These results support the hypothesis that the absence of apoAV slows lipolysis of TRL and the removal of their remnants by regulating their apoproteins content after secretion.

Apolipoprotein E (APOE) and warfarin dosing in an Italian population

Sirs, Despite large interindividual differences in dose requirement and a narrow therapeutic range, warfarin is still the most widely prescribed anticoagulant for thromboembolic therapy. However, the problems with dose titration, especially during the initial phase of treatment, lead to a high risk of either severe bleeding or the failure to prevent thromboembolism [1]. A working knowledge of warfarin interactive pathways and of the human genome will facilitate the determination of genetic factors that cause variable response to this drug. Warfarin acts through the inhibition of vitamin K reductase in the vitamin K cycle, thereby preventing the activation of clotting factors II, VII, IX and X and proteins C, S and Z [2, 3]. However, vitamin K1 can be reduced through another pathway, and a high intake of vitamin K1 may reverse the anticoagulant effect of warfarin [3–5]. Vitamin K1 is absorbed from the small intestine together with dietary fat, transported by chylomicrons in the blood and subsequently cleared by the liver through an apolipoprotein E (APOE) receptorspecific uptake [3, 6]. The major isoforms of apolipoprotein E are encoded by the three APOE alleles E2, E3 and E4 [7]. Individuals carrying the APOE*E4 allele have a more rapid uptake of chylomicron remnants following a meal than those that do not [8], and they also have lower serum concentrations of vitamin K1 following an overnight fast [6, 9, 10]. These findings suggest that carriers of APOE*E4 have a more rapid uptake of vitamin K1, leading to an enhanced availability of reduced vitamin K for the activation of clotting factors and proteins in the liver. Warfarin is administered as a racemate of Rand Senantiomers [11]. Cytochrome P450 2C9 (CYP2C9) is the main enzyme responsible for the metabolism of the more active S-warfarin [12], and genetic variability in CYP2C9 has a significant effect on warfarin dose requirement [13]. We have recently published a study on warfarin-treated Swedish patients showing that CYP2C9*1/*1 extensive metabolisers that are homozygous for APOE*E4 require higher warfarin doses than patients carrying other APOE alleles [14]. This study was repeated using 145 Italian patients, most of whom had participated in a previously published study on warfarin pharmacokinetics [15]. The local Ethics Committee (Comitato Etico dell’Azienda Ospedaliera di Padova) gave permission for the APOE genotyping. The warfarin patients had stable prothrombin time international normalised ratios (PT INR), and no interfering drugs were allowed during the study. Two APOE single nucleotide polymorphisms (SNPs) at nucleotide positions 334 and 472 were investigated using TaqMan (Applied Biosystems, Foster City, Calif.), and the ABI PRISM 7000 Sequence Detection System (Applied Biosystems) [14]. Twenty patients carried one APOE*E4 allele (E2/E4, n=1 and E3/E4, n=19), 105 were homozygous for E3 and 20 were heterozygous E2/E3. No one was homozygous for APOE*E4. There were no significant differences in age, gender, bodyweight or PT INR among the APOE genotype groups. We found that the weekly warfarin dose requirement did not vary between patients with different APOE genotypes (analysis of variance: p=0.5937; Fig. 1a). To test the contribution of other important factors, we combined the POE genotype with the CYP2C9 genotype, age, bodyweight and PT INR in a multiple regression model and found that CYP2C9 H. Kohnke AE M. G. Scordo AE M. Wadelius (&) Department of Medical Sciences, Clinical Pharmacology, University Hospital, Uppsala, Sweden E-mail: mia.wadelius@medsci.uu.se Tel.: +46-18-6114945

Spillover of Dietary Fatty Acids and Use of Serum Nonesterified Fatty Acids for the Synthesis of VLDL-Triacylglycerol Under Two Different Feeding Regimens

The present study quantified dietary fatty acid flux in healthy men (n = 6) who were fed a liquid formula through a duodenal feeding tube (continuous feeding group) or who consumed the same formula in meals (meal feeding group). A triacylglycerol (TAG) stable isotope was added to the formula to determine the entry of dietary fatty acids into the serum and its clearance to the liver and resecretion into serum via VLDL. The contribution of dietary fatty acids to serum nonesterified fatty acids (NEFAs) was higher with meal feeding (24.4 +/- 2.6%) compared with continuous feeding (10.8 +/- 2.9%, P < 0.01) and, when multiplied by the NEFA concentration, resulted in similar absolute fatty acid spillover. Diet-derived NEFAs subsequently represented 10.6 +/- 1.2% and 4.7 +/- 1.3% of hepatic VLDL-TAG (meal feeding vs. continuous feeding, respectively, P = 0.004). Chylomicron remnant uptake by the liver contributed 9.3 +/- 1.9% of fatty acids to hepatic VLDL-TAG synthesis with meal feeding compared with continuous feeding (4.4 +/- 0.8%, P < 0.03). These data suggest that the extent of dietary fatty acid recycling via serum NEFAs and VLDL-TAG is determined by the rate of delivery of dietary fat to the intestine. The inefficient removal of dietary fat from the circulation may maintain VLDL-TAG production but may also result in prolonged postprandial lipemia.

Interplay Between Apolipoprotein E and Scavenger Receptor Class B Type I Controls Coronary Atherosclerosis and Lifespan in the Mouse

The mouse has become a standard model for the study of atherosclerosis induced by extreme dyslipidemia in genetically modified strains such as the apolipoprotein E (apoE)–deficient or the LDL-receptor–deficient mouse. Although the process of plaque formation appears to repeat the fundamental steps of the human disease, it has been notoriously difficult to reproduce in the mouse the clinical consequences of atherosclerosis, such as myocardial infarction and stroke, that so commonly in patients signal plaque rupture or total lumen occlusion.1 The apoE-deficient mouse is a particularly interesting model because of its susceptibility to atherosclerotic lesions spanning from the aortic sinus to the abdominal aorta and involving the brachiocephalic and carotid arteries as well.2,3 It has recently been reported that the brachiocephalic plaque in the apoE-deficient mouse is prone to rupture on feeding a high-fat diet for periods as short as 8 weeks,4 but even in this setting of exaggerated hypercholesterolemia, the aortic sinus and coronary tree do not appear to be susceptible to plaque rupture. Scavenger receptor class B type I (SR-BI) is a scavenger-type receptor expressed by many tissues and accepted by consensus to represent the docking receptor for HDL in the liver. SR-BI-deficient mice have raised HDL levels and increased atherosclerosis susceptibility on fat feeding.5 ApoE and SR-BI are both expressed, among other cells, by the hepatocyte and the macrophage. ApoE is an efficient ligand for receptor-mediated lipoprotein removal by the liver, but it is also a strong acceptor of cellular cholesterol. As such, it may control the rate of foam cell formation in the vessel wall. Similarly, SR-BI channels cholesterol in and out of the cell, and its abundant expression by the macrophage may regulate cholesterol homeostasis in arterial foam cells. Although apoE and SR-BI play crucial roles in lipoprotein metabolism and have been …

Triglyceride-rich lipoprotein metabolism in unique VLDL receptor, LDL receptor, and LRP triple-deficient mice

The very low density lipoprotein receptor (VLDLR), low density lipoprotein receptor (LDLR), and low density lipoprotein receptor-related protein (LRP) are the three main apolipoprotein E-recognizing endocytic receptors involved in the clearance of triglyceride (TG)-rich lipoproteins from plasma. Whereas LDLR deficiency in mice results in the accumulation of plasma LDL-sized lipoproteins, VLDLR or LRP deficiency alone only minimally affects plasma lipoproteins. To investigate the combined effect of the absence of these receptors on TG-rich lipoprotein levels, we have generated unique VLDLR, LDLR, and LRP triple-deficient mice. Compared with wild-type mice, these mice markedly accumulated plasma lipids and lipases. These mice did not show aggravated hyperlipidemia compared with LDLR and LRP double-deficient mice, but plasma TG was increased after high-fat diet feeding. In addition, these mice showed a severely decreased postprandial TG clearance typical of VLDLR-deficient (VLDLR-/-) mice. Collectively, although VLDLR deficiency in LRP- and LDLR-/- mice does not aggravate hyperlipidemia, these triple-deficient mice represent a unique model of markedly delayed TG clearance on a hyperlipidemic background.

Endotoxin promotes preferential periportal upregulation of VLDL secretion in the rat liver

Zonation affects liver parenchymal cell function and metabolism as well as nonparenchymal cell activation, but whether VLDL production is zonated has yet to be elucidated. Infection induces enhanced VLDL secretion by the liver. Ex vivo studies were undertaken to examine the liver heterogeneity for VLDL formation and secretion and their in vivo response to endotoxin. Highly pure periportal (PP) and perivenous (PV) hepatocytes were isolated from fasted lipopolysaccharide-treated, fasted, and fed rats. They were used to assess their capacity to release VLDL-apolipoprotein B (apoB) and lipid classes in relation to de novo lipid synthesis and the expression of genes crucial to VLDL production. Despite the common superior ability of PP hepatocytes for lipid release and zonal differences in lipid synthesis, zonated secretion of VLDL particles was observed in septic but not in normal fed or fasted livers. The endotoxin-induced apoB secretion was more accentuated in PP hepatocytes; this was accompanied by a preferential PP increase in apoB and microsomal triglyceride transfer protein mRNA levels, whereas lipogenesis indicators were, if anything, similarly modified in hepatocytes of either acinar origin. We conclude that PP and PV hepatocytes exhibited similar capabilities for VLDL formation/secretion in normal conditions; however, the endotoxic pressure did zonate periportally.