The diffusion rate of lipids in the cell membrane is reduced by a factor of 5-100 from that in artificial bilayers. This slowing mechanism has puzzled cell biologists for the last 25 yr. Here we address this issue by studying the movement of unsaturated phospholipids in rat kidney fibroblasts at the single molecule level at the temporal resolution of 25 micros. The cell membrane was found to be compartmentalized: phospholipids are confined within 230-nm-diameter (phi) compartments for 11 ms on average before hopping to adjacent compartments. These 230-nm compartments exist within greater 750-nm-phi compartments where these phospholipids are confined for 0.33 s on average. The diffusion rate within 230-nm compartments is 5.4 microm2/s, which is nearly as fast as that in large unilamellar vesicles, indicating that the diffusion in the cell membrane is reduced not because diffusion per se is slow, but because the cell membrane is compartmentalized with regard to lateral diffusion of phospholipids. Such compartmentalization depends on the actin-based membrane skeleton, but not on the extracellular matrix, extracellular domains of membrane proteins, or cholesterol-enriched rafts. We propose that various transmembrane proteins anchored to the actin-based membrane skeleton meshwork act as rows of pickets that temporarily confine phospholipids.

BACKGROUND

Metabolic risk conferred by adiposity may be due to associated risk factor clustering.

OBJECTIVE

The objective of this study was to assess risk for diabetes or cardiovascular disease (CVD) stratified by body mass index (BMI) and the presence or absence of metabolic syndrome (MetS) or insulin resistance (IR).

METHODS

This was a community-based, longitudinal study of 2902 people (55% women, mean age 53 yr) without diabetes or CVD in 1989-1992 followed for up to 11 yr. We categorized subjects by normal weight (BMI < 25 kg/m(2)), overweight (25-29.9 kg/m(2)), or obese (>30 kg/m(2)) and by the National Cholesterol Education Program's Adult Treatment Panel MetS or the top quartile of homeostasis model IR. We used proportional hazard models to estimate risk relative to normal weight and no MetS or IR.

METHODS

Incident type 2 diabetes (treatment or fasting glucose>> or = 7 mmol/liter, 141 events) or CVD (myocardial infarction, stroke, or claudication, 252 events) were measured.

RESULTS

Among 1056 normal-weight subjects, 7% had MetS and a risk factor-adjusted relative risk for diabetes of 3.97 (95% confidence interval, 1.35-11.6) and for CVD of 3.01 (1.68-5.41). Among 638 obese subjects, 37% did not have MetS or significantly increased risk. Obese subjects with MetS had an adjusted relative risk for diabetes of 10.3 (5.44-19.5) and for CVD of 2.13 (1.43-3.18). Results were similar in analyses of BMI-IR categories.

CONCLUSIONS

People with normal weight and MetS or IR or with obesity but no MetS or IR were not uncommon in our sample. Risk factor clustering or IR appear to confer much of the risk for diabetes or CVD commonly associated with elevated BMI.

Chronic inflammation is a key feature of vascular disease states such as atherosclerosis. Multiple clinical studies have shown that a class of medications termed statins lower cardiovascular morbidity and mortality. Originally developed to lower serum cholesterol, increasing evidence suggests that these medications have potent anti-inflammatory effects that contribute to their beneficial effects in patients. Here, we discuss the clinical and experimental evidence underlying the anti-inflammatory effects of these agents.

Recent investigations of atherosclerosis have focused on inflammation, providing new insight into mechanisms of disease. Inflammatory cytokines involved in vascular inflammation stimulate the generation of endothelial adhesion molecules, proteases, and other mediators, which may enter the circulation in soluble form. These primary cytokines also induce production of the messenger cytokine interleukin-6, which stimulates the liver to increase production of acute-phase reactants such as C-reactive protein. In addition, platelets and adipose tissue can generate inflammatory mediators relevant to atherothrombosis. Despite the irreplaceable utility of plasma lipid profiles in assessment of atherosclerotic risk, these profiles provide an incomplete picture. Indeed, many cardiovascular events occur in individuals with plasma cholesterol concentrations below the National Cholesterol Education Program thresholds of 200 mg/dL for total cholesterol and 130 mg/dL for low-density lipoprotein (LDL) cholesterol. The concept of the involvement of inflammation in atherosclerosis has spurred the discovery and adoption of inflammatory biomarkers for cardiovascular risk prediction. C-reactive protein is currently the best validated inflammatory biomarker; in addition, soluble CD40 ligand, adiponectin, interleukin 18, and matrix metalloproteinase 9 may provide additional information for cardiovascular risk stratification and prediction. This review retraces the biology of atherothrombosis and the evidence supporting the role of inflammatory biomarkers in predicting primary cardiovascular events in this biologic context.

BACKGROUND

We report the first large renal biopsy-based clinicopathologic study on obesity-related glomerulopathy.

METHODS

Obesity was defined as body mass index (BMI>> 30 kg/m2. Obesity-related glomerulopathy (ORG) was defined morphologically as focal segmental glomerulosclerosis and glomerulomegaly (O-FSGS; N = 57) or glomerulomegaly alone (O-GM; N = 14).

RESULTS

Review of 6818 native renal biopsies received from 1986 to 2000 revealed a progressive increase in biopsy incidence of ORG from 0.2% in 1986-1990 to 2.0% in 1996-2000 (P = 0.0001). Mean BMI in ORG was 41.7 (range 30.9 to 62.7). Indications for renal biopsy included proteinuria (N = 40) or proteinuria and renal insufficiency (N = 31). Seventy-one patients with ORG were compared to 50 patients with idiopathic FSGS (I-FSGS). Patients with ORG were older (mean 42.9 vs. 32.6 years, P < 0.001) and more often Caucasian (75% vs. 52%; P = 0.003). ORG patients had a lower incidence of nephrotic range proteinuria (48% vs. 66%; P = 0.007) and nephrotic syndrome (5.6% vs. 54%; P < 0.001), with higher serum albumin (3.9 vs. 2.9 g/dL; P < 0.001), lower serum cholesterol (229 vs. 335 mg/dL; P < 0.001), and less edema (35% vs. 68%; P = 0.003). On renal biopsy, patients with ORG had fewer lesions of segmental sclerosis (10 vs. 39%; P < 0.001), more glomerulomegaly (100% vs. 10%; P < 0.001), and less extensive foot process effacement (40 vs. 75%; P < 0.001). Glomerular diameter in ORG (mean 226 mu) was significantly larger than age- and sex-matched normal controls (mean 168 mu; P < 0.001). Follow-up was available in 56 ORG patients (mean 27 months) and 50 idiopathic FSGS controls (mean 38 months). A total of 75% of ORG patients received angiotensin-converting enzyme (ACE) inhibition or A2 blockade while 78% of the I-FSGS patients received immunosuppressive therapy. ORG patients had less frequent doubling of serum creatinine (14.3% vs. 50%; P < 0.001) and progression to ESRD (3.6% vs. 42%; P < 0.001). On multivariate analysis, presenting serum creatinine and severity of proteinuria were the only predictors of poor outcome in ORG.

CONCLUSIONS

ORG is distinct from idiopathic FSGS, with a lower incidence of nephrotic syndrome, more indolent course, consistent presence of glomerulomegaly, and milder foot process fusion. The ten-fold increase in incidence over 15 years suggests a newly emerging epidemic. Heightened physician awareness of this entity is needed to ensure accurate diagnosis and appropriate therapy.

The concentrations of bile salt, lecithin, and cholesterol were determined on each of 66 samples of gall bladder bile from patients with cholesterol gallstones and 25 samples of normal gall bladder bile. When these three constituents were plotted simultaneously on triangular coordinates, a complete separation of the normal and "abnormal" bile was achieved. This separation was the result of an increase in the quantity of cholesterol relative to the amounts of bile salts and lecithin contained in the bile from patients with cholesterol gallstones. An in vitro model system was constructed (on triangular coordinates) that allows prediction of the maximum amount of cholesterol that can be solubilized in solutions containing varying proportions of bile salt and lecithin. When the bile data were compared with the solubility of cholesterol derived from the model system, normal biles were found to be less than saturated with cholesterol, whereas biles from patients with cholesterol gallstones were saturated and in some cases contained insoluble cholesterol in the form of microcrystals. It is suggested that the physical state of bile (i.e., the presence or absence of insoluble cholesterol) is determined by the relative concentrations of bile salt, lecithin, and cholesterol, and the other biliary constituents do not appear to significantly effect the solubility of cholesterol in bile.

BACKGROUND

Reduced intake of saturated fat is widely recommended for prevention of cardiovascular disease. The type of macronutrient that should replace saturated fat remains uncertain.

OBJECTIVE

To compare the effects of 3 healthful diets, each with reduced saturated fat intake, on blood pressure and serum lipids.

METHODS

Randomized, 3-period, crossover feeding study (April 2003 to June 2005) conducted in Baltimore, Md, and Boston, Mass. Participants were 164 adults with prehypertension or stage 1 hypertension. Each feeding period lasted 6 weeks and body weight was kept constant.

METHODS

A diet rich in carbohydrates; a diet rich in protein, about half from plant sources; and a diet rich in unsaturated fat, predominantly monounsaturated fat.

METHODS

Systolic blood pressure and low-density lipoprotein cholesterol.

RESULTS

Blood pressure, low-density lipoprotein cholesterol, and estimated coronary heart disease risk were lower on each diet compared with baseline. Compared with the carbohydrate diet, the protein diet further decreased mean systolic blood pressure by 1.4 mm Hg (P = .002) and by 3.5 mm Hg (P = .006) among those with hypertension and decreased low-density lipoprotein cholesterol by 3.3 mg/dL (0.09 mmol/L; P = .01), high-density lipoprotein cholesterol by 1.3 mg/dL (0.03 mmol/L; P = .02), and triglycerides by 15.7 mg/dL (0.18 mmol/L; P<.001). Compared with the carbohydrate diet, the unsaturated fat diet decreased systolic blood pressure by 1.3 mm Hg (P = .005) and by 2.9 mm Hg among those with hypertension (P = .02), had no significant effect on low-density lipoprotein cholesterol, increased high-density lipoprotein cholesterol by 1.1 mg/dL (0.03 mmol/L; P = .03), and lowered triglycerides by 9.6 mg/dL (0.11 mmol/L; P = .02). Compared with the carbohydrate diet, estimated 10-year coronary heart disease risk was lower and similar on the protein and unsaturated fat diets.

CONCLUSIONS

In the setting of a healthful diet, partial substitution of carbohydrate with either protein or monounsaturated fat can further lower blood pressure, improve lipid levels, and reduce estimated cardiovascular risk. Clinical Trials Registration ClinicalTrials.gov Identifier: NCT00051350.

Sterol regulatory element 1 (SRE-1), a decamer (5'-ATC-ACCCCAC-3') flanking the low density lipoprotein (LDL) receptor gene, activates transcription in sterol-depleted cells and is silenced by sterols. We report the cDNA cloning of human SREBP-1, a protein that binds SRE-1, activates transcription, and thereby mediates the final regulatory step in LDL metabolism. SREBP-1 contains a basic-helix-loop-helix-leucine zipper (bHLH-ZIP) motif, but it differs from other bHLH-ZIP proteins in its larger size (1147 amino acids) and target sequence. Instead of an inverted repeat (CANNTG), the target for all known bHLH-ZIP proteins, SRE-1 contains a direct repeat of CAC. Overexpression of SREBP-1 activates transcription of reporter genes containing SRE-1 in the absence (15-fold) and presence (90-fold) of sterols, abolishing sterol regulation. We suggest that SREBP-1 is regulated by an unknown factor that is overwhelmed when SREBP-1 is overexpressed. Understanding the regulation of SREBP-1 may be crucial for understanding the control of plasma cholesterol in humans.

Cellular cholesterol levels reflect a balance between uptake, efflux, and endogenous synthesis. Here we show that the sterol-responsive nuclear liver X receptor (LXR) helps maintain cholesterol homeostasis, not only through promotion of cholesterol efflux but also through suppression of low-density lipoprotein (LDL) uptake. LXR inhibits the LDL receptor (LDLR) pathway through transcriptional induction of Idol (inducible degrader of the LDLR), an E3 ubiquitin ligase that triggers ubiquitination of the LDLR on its cytoplasmic domain, thereby targeting it for degradation. LXR ligand reduces, whereas LXR knockout increases, LDLR protein levels in vivo in a tissue-selective manner. Idol knockdown in hepatocytes increases LDLR protein levels and promotes LDL uptake. Conversely, adenovirus-mediated expression of Idol in mouse liver promotes LDLR degradation and elevates plasma LDL levels. The LXR-Idol-LDLR axis defines a complementary pathway to sterol response element-binding proteins for sterol regulation of cholesterol uptake.

Twenty healthy adult humans had serum samples drawn on four occasions within a 24-hr period: after a 12 hr overnight fast, 4-5 hr after a high fat breakfast, at midafternoon, and the next morning after another 12 hr fast. Nonfasting samples had 22% to 29% higher mean concentrations (p less than 0.05) than did fasting samples for polychlorinated biphenyls (PCBs, 4.81 vs 3.74 ng/g serum wt), hexachlorobenzene (HCB, 0.163 vs 0.134 ng/g serum wt), and p,p'-dichlorodiphenyl-dichloroethylene (p,p'-DDE, 6.74 vs 5.37 ng/g serum wt) measured by electron capture gas liquid chromatography. Total serum lipids were estimated from measurements of total cholesterol, free cholesterol, triglycerides, and phospholipids and were 20% higher in nonfasting samples than in fasting samples (7.05 g/L vs 5.86 g/L). When PCBs, HCB, and p,p'-DDE concentrations were corrected by total serum lipids, results from fasting and non-fasting samples were not statistically different. Because of the differences in these chlorinated hydrocarbon concentrations observed with different sample collection regimens, meaningful comparison of analytical results requires standardizing collection procedures or correcting by total serum lipid levels.

PPARalpha is a nuclear receptor that regulates liver and skeletal muscle lipid metabolism as well as glucose homeostasis. Acting as a molecular sensor of endogenous fatty acids (FAs) and their derivatives, this ligand-activated transcription factor regulates the expression of genes encoding enzymes and transport proteins controlling lipid homeostasis, thereby stimulating FA oxidation and improving lipoprotein metabolism. PPARalpha also exerts pleiotropic antiinflammatory and antiproliferative effects and prevents the proatherogenic effects of cholesterol accumulation in macrophages by stimulating cholesterol efflux. Cellular and animal models of PPARalpha help explain the clinical actions of fibrates, synthetic PPARalpha agonists used to treat dyslipidemia and reduce cardiovascular disease and its complications in patients with the metabolic syndrome. Although these preclinical studies cannot predict all of the effects of PPARalpha in humans, recent findings have revealed potential adverse effects of PPARalpha action, underlining the need for further study. This Review will focus on the mechanisms of action of PPARalpha in metabolic diseases and their associated vascular pathologies.

BACKGROUND

Statins lower high-sensitivity C-reactive protein (hsCRP) and cholesterol concentrations, and hypothesis generating analyses suggest that clinical outcomes improve in patients given statins who achieve hsCRP concentrations less than 2 mg/L in addition to LDL cholesterol less than 1.8 mmol/L (<70 mg/dL). However, the benefit of lowering both LDL cholesterol and hsCRP after the start of statin therapy is controversial. We prospectively tested this hypothesis.

METHODS

In an analysis of 15 548 initially healthy men and women participating in the JUPITER trial (87% of full cohort), we prospectively assessed the effects of rosuvastatin 20 mg versus placebo on rates of non-fatal myocardial infarction, non-fatal stroke, admission for unstable angina, arterial revascularisation, or cardiovascular death (prespecified endpoints) during a maximum follow-up of 5 years (median 1.9 years), according to on-treatment concentrations of LDL cholesterol >>/=1.8 mmol/L or <1.8 mmol/L) and hsCRP >>/=2 mg/L or <2 mg/L). We included all events occurring after randomisation. This trial is registered with ClinicalTrials.gov, number NCT00239681.

RESULTS

Compared with placebo, participants allocated to rosuvastatin who achieved LDL cholesterol less than 1.8 mmol/L had a 55% reduction in vascular events (event rate 1.11 vs 0.51 per 100 person-years; hazard ratio [HR] 0.45, 95% CI 0.34-0.60, p<0.0001), and those achieving hsCRP less than 2 mg/L a 62% reduction (event rate 0.42 per 100 person-years; HR 0.38, 95% CI 0.26-0.56, p<0.0001). Although LDL cholesterol and hsCRP reductions were only weakly correlated in individual patients (r values <0.15), we recorded a 65% reduction in vascular events in participants allocated to rosuvastatin who achieved both LDL cholesterol less than 1.8 mmol/L and hsCRP less than 2 mg/L (event rate 0.38 per 100 person-years; adjusted HR 0.35, 95% CI 0.23-0.54), versus a 33% reduction in those who achieved one or neither target (event rate 0.74 per 100 person-years; HR 0.67, 95% CI 0.52-0.87) (p across treatment groups <0.0001). In participants who achieved LDL cholesterol less than 1.8 mmol/L and hsCRP less than 1 mg/L, we noted a 79% reduction (event rate 0.24 per 100 person-years; HR 0.21, 95% CI 0.09-0.52). Achieved hsCRP concentrations were predictive of event rates irrespective of the lipid endpoint used, including the apolipoprotein B to apolipoprotein AI ratio.

CONCLUSIONS

For people choosing to start pharmacological prophylaxis, reduction in both LDL cholesterol and hsCRP are indicators of successful treatment with rosuvastatin.

Taking into account a continuous exposure in regression models by using categorization, when non-linear dose-response associations are expected, have been widely criticized. As one alternative, restricted cubic spline (RCS) functions are powerful tools (i) to characterize a dose-response association between a continuous exposure and an outcome, (ii) to visually and/or statistically check the assumption of linearity of the association, and (iii) to minimize residual confounding when adjusting for a continuous exposure. Because their implementation with SAS® software is limited, we developed and present here an SAS macro that (i) creates an RCS function of continuous exposures, (ii) displays graphs showing the dose-response association with 95 per cent confidence interval between one main continuous exposure and an outcome when performing linear, logistic, or Cox models, as well as linear and logistic-generalized estimating equations, and (iii) provides statistical tests for overall and non-linear associations. We illustrate the SAS macro using the third National Health and Nutrition Examination Survey data to investigate adjusted dose-response associations (with different models) between calcium intake and bone mineral density (linear regression), folate intake and hyperhomocysteinemia (logistic regression), and serum high-density lipoprotein cholesterol and cardiovascular mortality (Cox model).

Little is known regarding the long-term effects of caloric restriction (CR) on the risk for atherosclerosis. We evaluated the effect of CR on risk factors for atherosclerosis in individuals who are restricting food intake to slow aging. We studied 18 individuals who had been on CR for an average of 6 years and 18 age-matched healthy individuals on typical American diets. We measured serum lipids and lipoproteins, fasting plasma glucose and insulin, blood pressure (BP), high-sensitivity C-reactive protein (CRP), platelet-derived growth factor AB (PDGF-AB), body composition, and carotid artery intima-media thickness (IMT). The CR group were leaner than the comparison group (body mass index, 19.6 +/- 1.9 vs. 25.9 +/- 3.2 kg/m(2); percent body fat, 8.7 +/- 7% vs. 24 +/- 8%). Serum total cholesterol (Tchol), low-density lipoprotein cholesterol, ratio of Tchol to high-density lipoprotein cholesterol (HDL-C), triglycerides, fasting glucose, fasting insulin, CRP, PDFG-AB, and systolic and diastolic BP were all markedly lower, whereas HDL-C was higher, in the CR than in the American diet group. Medical records indicated that the CR group had serum lipid-lipoprotein and BP levels in the usual range for individuals on typical American diets, and similar to those of the comparison group, before they began CR. Carotid artery IMT was approximately 40% less in the CR group than in the comparison group. Based on a range of risk factors, it appears that long-term CR has a powerful protective effect against atherosclerosis. This interpretation is supported by the finding of a low carotid artery IMT.

HMG-CoA (3-hydroxy-3-methylglutaryl-coenzyme A) reductase (HMGR) catalyzes the committed step in cholesterol biosynthesis. Statins are HMGR inhibitors with inhibition constant values in the nanomolar range that effectively lower serum cholesterol levels and are widely prescribed in the treatment of hypercholesterolemia. We have determined structures of the catalytic portion of human HMGR complexed with six different statins. The statins occupy a portion of the binding site of HMG-CoA, thus blocking access of this substrate to the active site. Near the carboxyl terminus of HMGR, several catalytically relevant residues are disordered in the enzyme-statin complexes. If these residues were not flexible, they would sterically hinder statin binding.

Tissue factor (TF) circulates in plasma, largely on monocyte/macrophage-derived microvesicles that can bind activated platelets through a mechanism involving P-selectin glycoprotein ligand-1 (PSGL-1) on the microvesicles and P-selectin on the platelets. We found these microvesicles to be selectively enriched in both TF and PSGL-1, and deficient in CD45, suggesting that they arise from distinct membrane microdomains. We investigated the possibility that microvesicles arise from cholesterol-rich lipid rafts and found that both TF and PSGL-1, but not CD45, localize to lipid rafts in blood monocytes and in the monocytic cell line THP-1. Consistent with a raft origin of TF-bearing microvesicles, their shedding was significantly reduced with depletion of membrane cholesterol. We also evaluated the interaction between TF-bearing microvesicles and platelets. Microvesicles bound only activated platelets, and required PSGL-1 to do so. The microvesicles not only bound the activated platelets, they fused with them, transferring both proteins and lipid to the platelet membrane. Fusion was blocked by either annexin V or an antibody to PSGL-1 and had an important functional consequence: increasing the proteolytic activity of the TF-VIIa complex. These findings suggest a mechanism by which all of the membrane-bound reactions of the coagulation system can be localized to the surface of activated platelets.

BACKGROUND

Higher levels of serum phosphate are associated with adverse cardiovascular outcomes, especially in the setting of overt hyperphosphatemia. Given the biological importance of phosphorus, it is plausible that higher levels of serum phosphate within the normal range may also be associated with adverse outcomes.

RESULTS

We performed a post hoc analysis of data from the Cholesterol And Recurrent Events (CARE) study. Baseline serum phosphate levels were measured in 4127 fasting participants who were randomized to receive pravastatin 40 mg daily or placebo and followed up for a median of 59.7 months. We used Cox proportional-hazards models to examine the association between serum phosphate and adverse clinical outcomes after adjustment for potential confounders. During nearly 60 months of follow-up, 375 participants died. A significant association was noted between baseline serum phosphate level and the age-, race-, and sex-adjusted risk of all-cause death (hazard ratio per 1 mg/dL, 1.27; 95% confidence interval, 1.02 to 1.58). After categorization based on baseline phosphate level (<2.5, 2.5 to 3.4, 3.5 to 3.9, and>> or =4 mg/dL) and further adjustment, a graded independent relation between phosphate and death was observed (P for trend=0.03). For instance, participants with serum phosphate>> or =3.5 mg/dL had an adjusted hazard ratio for death of 1.27 (95% confidence interval, 1.02 to 1.59) compared with those with serum phosphate of <3.5 mg/dL. Higher levels of serum phosphate were also associated with increased risk of new heart failure, myocardial infarction, and the composite of coronary death or nonfatal myocardial infarction, but not the risk of stroke.

CONCLUSIONS

We found a graded independent relation between higher levels of serum phosphate and the risk of death and cardiovascular events in people with prior myocardial infarction, most of whom had serum phosphate levels within the normal range. Given the ready availability and low cost of serum phosphate assays, this finding may prove clinically useful.

The primary circulating form of vitamin D, 25-hydroxy-vitamin D [25(OH)D], is associated with multiple medical outcomes, including rickets, osteoporosis, multiple sclerosis and cancer. In a genome-wide association study (GWAS) of 4501 persons of European ancestry drawn from five cohorts, we identified single-nucleotide polymorphisms (SNPs) in the gene encoding group-specific component (vitamin D binding) protein, GC, on chromosome 4q12-13 that were associated with 25(OH)D concentrations: rs2282679 (P=2.0x10(-30)), in linkage disequilibrium (LD) with rs7041, a non-synonymous SNP (D432E; P=4.1x10(-22)) and rs1155563 (P=3.8x10(-25)). Suggestive signals for association with 25(OH)D were also observed for SNPs in or near three other genes involved in vitamin D synthesis or activation: rs3829251 on chromosome 11q13.4 in NADSYN1 [encoding nicotinamide adenine dinucleotide (NAD) synthetase; P=8.8x10(-7)], which was in high LD with rs1790349, located in DHCR7, the gene encoding 7-dehydrocholesterol reductase that synthesizes cholesterol from 7-dehydrocholesterol; rs6599638 in the region harboring the open-reading frame 88 (C10orf88) on chromosome 10q26.13 in the vicinity of ACADSB (acyl-Coenzyme A dehydrogenase), involved in cholesterol and vitamin D synthesis (P=3.3x10(-7)); and rs2060793 on chromosome 11p15.2 in CYP2R1 (cytochrome P450, family 2, subfamily R, polypeptide 1, encoding a key C-25 hydroxylase that converts vitamin D3 to an active vitamin D receptor ligand; P=1.4x10(-5)). We genotyped SNPs in these four regions in 2221 additional samples and confirmed strong genome-wide significant associations with 25(OH)D through meta-analysis with the GWAS data for GC (P=1.8x10(-49)), NADSYN1/DHCR7 (P=3.4x10(-9)) and CYP2R1 (P=2.9x10(-17)), but not C10orf88 (P=2.4x10(-5)).

Exosomes are 60-100-nm membrane vesicles that are secreted into the extracellular milieu as a consequence of multivesicular body fusion with the plasma membrane. Here we determined the protein and lipid compositions of highly purified human B cell-derived exosomes. Mass spectrometric analysis indicated the abundant presence of major histocompatibility complex (MHC) class I and class II, heat shock cognate 70, heat shock protein 90, integrin alpha 4, CD45, moesin, tubulin (alpha and beta), actin, G(i)alpha(2), and a multitude of other proteins. An alpha 4-integrin may direct B cell-derived exosomes to follicular dendritic cells, which were described previously as potential target cells. Clathrin, heat shock cognate 70, and heat shock protein 90 may be involved in protein sorting at multivesicular bodies. Exosomes were also enriched in cholesterol, sphingomyelin, and ganglioside GM3, lipids that are typically enriched in detergent-resistant membranes. Most exosome-associated proteins, including MHC class II and tetraspanins, were insoluble in 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid (CHAPS)-containing buffers. Multivesicular body-linked MHC class II was also resistant to CHAPS whereas plasma membrane-associated MHC class II was solubilized readily. Together, these data suggest that recruitment of membrane proteins from the limiting membranes into the internal vesicles of multivesicular bodies may involve their incorporation into tetraspanin-containing detergent-resistant membrane domains.

Simian virus 40 (SV40) utilizes endocytosis through caveolae for infectious entry into host cells. We found that after binding to caveolae, virus particles induced transient breakdown of actin stress fibers. Actin was then recruited to virus-loaded caveolae as actin patches that served as sites for actin "tail" formation. Dynamin II was also transiently recruited. These events depended on the presence of cholesterol and on the activation of tyrosine kinases that phosphorylated proteins in caveolae. They were necessary for formation of caveolae-derived endocytic vesicles and for infection of the cell. Thus, caveolar endocytosis is ligand-triggered and involves extensive rearrangement of the actin cytoskeleton.

BACKGROUND

Lowering low-density lipoprotein cholesterol (LDL-C) is known to reduce risk of recurrent coronary heart disease in middle-aged men. However, this effect has been uncertain in elderly people and women.

OBJECTIVE

To estimate the risk reduction of coronary heart disease and total mortality associated with statin drug treatment, particularly in elderly individuals and women.

METHODS

Trials published in English-language journals were retrieved by searching MEDLINE (1966-December 1998), bibliographies, and authors' reference files.

METHODS

Studies in which participants were randomized to statin or control treatment for at least 4 years and clinical disease or death was the primary outcome were included in the meta-analysis (5 of 182 initially identified).

METHODS

Information on sample size, study drug duration, type and dosage of statin drug, participant characteristics at baseline, reduction in lipids during intervention, and outcomes was abstracted independently by 2 authors (J.H. and S.V.) using a standardized protocol. Disagreements were resolved by consensus.

RESULTS

Data from the 5 trials, with 30 817 participants, were included in this meta-analysis. The mean duration of treatment was 5.4 years. Stati n drug treatment was associated with a20% reduction in total cholesterol, 28% reduction in LDL-C, 13% reduction in triglycerides, and 5% increase in high-density lipoprotein cholesterol. Overall, statin drug treatment reduced risk 31 % in major coronary events (95% confidence interval [CI], 26%-36%) and 21 % in all-cause mortality (95% CI, 14%-28%). The risk reduction in major coronary events was similar between women (29%; 95% Cl, 13 %-42 %) and men (31 %; 95% CI, 26%-35%), and between persons aged at least 65 years (32%; 95% CI, 23%-39%) and persons younger than 65 years (31 %; 95% CI, 24%-36%).

CONCLUSIONS

Our meta-analysis indicates that reduction in LDL-C associated with statin drug treatment decreases the risk of coronary heart disease and all-cause mortality. The risk reduction was similar for men and women and for elderly and middle-aged persons.

Farnesoid X receptor (FXR) plays an important role in maintaining bile acid and cholesterol homeostasis. Here we demonstrate that FXR also regulates glucose metabolism. Activation of FXR by the synthetic agonist GW4064 or hepatic overexpression of constitutively active FXR by adenovirus-mediated gene transfer significantly lowered blood glucose levels in both diabetic db/db and wild-type mice. Consistent with these data, FXR null mice exhibited glucose intolerance and insulin insensitivity. We further demonstrate that activation of FXR in db/db mice repressed hepatic gluconeogenic genes and increased hepatic glycogen synthesis and glycogen content by a mechanism that involves enhanced insulin sensitivity. In view of its central roles in coordinating regulation of both glucose and lipid metabolism, we propose that FXR agonists are promising therapeutic agents for treatment of diabetes mellitus.

BACKGROUND

Evolocumab, a monoclonal antibody that inhibits proprotein convertase subtilisin-kexin type 9 (PCSK9), significantly reduced low-density lipoprotein (LDL) cholesterol levels in short-term studies. We conducted two extension studies to obtain longer-term data.

METHODS

In two open-label, randomized trials, we enrolled 4465 patients who had completed 1 of 12 phase 2 or 3 studies ("parent trials") of evolocumab. Regardless of study-group assignments in the parent trials, eligible patients were randomly assigned in a 2:1 ratio to receive either evolocumab (140 mg every 2 weeks or 420 mg monthly) plus standard therapy or standard therapy alone. Patients were followed for a median of 11.1 months with assessment of lipid levels, safety, and (as a prespecified exploratory analysis) adjudicated cardiovascular events including death, myocardial infarction, unstable angina, coronary revascularization, stroke, transient ischemic attack, and heart failure. Data from the two trials were combined.

RESULTS

As compared with standard therapy alone, evolocumab reduced the level of LDL cholesterol by 61%, from a median of 120 mg per deciliter to 48 mg per deciliter (P<0.001). Most adverse events occurred with similar frequency in the two groups, although neurocognitive events were reported more frequently in the evolocumab group. The risk of adverse events, including neurocognitive events, did not vary significantly according to the achieved level of LDL cholesterol. The rate of cardiovascular events at 1 year was reduced from 2.18% in the standard-therapy group to 0.95% in the evolocumab group (hazard ratio in the evolocumab group, 0.47; 95% confidence interval, 0.28 to 0.78; P=0.003).

CONCLUSIONS

During approximately 1 year of therapy, the use of evolocumab plus standard therapy, as compared with standard therapy alone, significantly reduced LDL cholesterol levels and reduced the incidence of cardiovascular events in a prespecified but exploratory analysis. (Funded by Amgen; OSLER-1 and OSLER-2 ClinicalTrials.gov numbers, NCT01439880 and NCT01854918.).

A thermodynamic and a microscopic interaction model are proposed to describe the phase equilibria in the phosphatidylcholine-cholesterol system. The model calculations allow for a solid phase with conformationally ordered acyl chains and liquid phases with conformationally ordered as well as disordered chains. The resulting phase diagram is in excellent agreement with the experimental phase diagram for dipalmitoylphosphatidylcholine bilayers with cholesterol as determined by a recent NMR and calorimetry study. It is thus demonstrated that the phase behaviour of phosphatidylcholine-cholesterol mixtures can be rationalized using only a few basic assumptions: (i) Cholesterol interacts favourably with phosphatidylcholine chains in an extended conformation, (ii) the main transition of pure phosphatidylcholine bilayers takes place in terms of translational variables as well acyl-chain conformational variables, and (iii) cholesterol disturbs the translational order in the crystalline (gel) state of phosphatidylcholine. These results suggest that the occurrence of specific phosphatidylcholine-cholesterol complexes is not implied by the experimental thermodynamic data.