Integrated genomic profiling of endometrial carcinoma associates aggressive tumors with indicators of PI3 kinase activation.

Although 75% of endometrial cancers are treated at an early stage, 15% to 20% of these recur. We performed an integrated analysis of genome-wide expression and copy-number data for primary endometrial carcinomas with extensive clinical and histopathological data to detect features predictive of recurrent disease. Unsupervised analysis of the expression data distinguished 2 major clusters with strikingly different phenotypes, including significant differences in disease-free survival. To identify possible mechanisms for these differences, we performed a global genomic survey of amplifications, deletions, and loss of heterozygosity, which identified 11 significantly amplified and 13 significantly deleted regions. Amplifications of 3q26.32 harboring the oncogene PIK3CA were associated with poor prognosis and segregated with the aggressive transcriptional cluster. Moreover, samples with PIK3CA amplification carried signatures associated with in vitro activation of PI3 kinase (PI3K), a signature that was shared by aggressive tumors without PIK3CA amplification. Tumors with loss of PTEN expression or PIK3CA overexpression that did not have PIK3CA amplification also shared the PI3K activation signature, high protein expression of the PI3K pathway member STMN1, and an aggressive phenotype in test and validation datasets. However, mutations of PTEN or PIK3CA were not associated with the same expression profile or aggressive phenotype. STMN1 expression had independent prognostic value. The results affirm the utility of systematic characterization of the cancer genome in clinically annotated specimens and suggest the particular importance of the PI3K pathway in patients who have aggressive endometrial cancer.

Cardiolipin content of wild type and mutant yeasts in relation to mitochondrial function and development.

The phospholipid composition of various strains of the yeast, Saccharomyces cerevisiae, and several of their derived mitochondrial mutants grown under conditions designed to induce variations in the complement of mitochondrial membranes has been examined. Wild type and petite (cytoplasmic respiratory deficient) yeasts were fractionated into various subcellular fractions, which were monitored by electron microscopy and analyzed for cytochrome oxidase (in wild type) and phospholipid composition. 90% or more of the phospholipid, cardiolipin was found in the mitochondrial membranes of wild type and petite yeast. Cardiolipin content differed markedly under various growth conditions. Stationary yeast grown in glucose had better developed mitochondria and more cardiolipin than repressed log phase yeast. Aerobic yeast contained more cardiolipin than anaerobic yeast. Respiration-deficient cytoplasmic mitochondrial mutants, both suppressive and neutral, contained less cardiolipin than corresponding wild types. A chromosomal mutant lacking respiratory function had normal cardiolipin content. Log phase cells grown in galactose and lactate, which do not readily repress the development of mitochondrial membranes, contained as much cardiolipin as stationary phase cells grown in glucose. Cytoplasmic mitochondrial mutants respond to changes in the glucose concentration of the growth medium by variations in their cardiolipin content in the same way as wild type yeast does under similar growth conditions. It is concluded that cardiolipin content of yeast is correlated with, and is a good indicator of, the state of development of mitochondrial membrane.

Distinct mutational signatures characterize concurrent loss of polymerase proofreading and mismatch repair.

Fidelity of DNA replication is maintained using polymerase proofreading and the mismatch repair pathway. Tumors with loss of function of either mechanism have elevated mutation rates with characteristic mutational signatures. Here we report that tumors with concurrent loss of both polymerase proofreading and mismatch repair function have mutational patterns that are not a simple sum of the signatures of the individual alterations, but correspond to distinct, previously unexplained signatures: COSMIC database signatures 14 and 20. We then demonstrate that in all five cases in which the chronological order of events could be determined, polymerase epsilon proofreading alterations precede the defect in mismatch repair. Overall, we illustrate that multiple distinct mutational signatures can result from different combinations of a smaller number of mutational processes (of either damage or repair), which can influence the interpretation and discovery of mutational signatures.

Whole-exome sequencing of cell-free DNA and circulating tumor cells in multiple myeloma.

Liquid biopsies including circulating tumor cells (CTCs) and cell-free DNA (cfDNA) have enabled minimally invasive characterization of many cancers, but are rarely analyzed together. Understanding the detectability and genomic concordance of CTCs and cfDNA may inform their use in guiding cancer precision medicine. Here, we report the detectability of cfDNA and CTCs in blood samples from 107 and 56 patients with multiple myeloma (MM), respectively. Using ultra-low pass whole-genome sequencing, we find both tumor fractions correlate with disease progression. Applying whole-exome sequencing (WES) to cfDNA, CTCs, and matched tumor biopsies, we find concordance in clonal somatic mutations (~99%) and copy number alterations (~81%) between liquid and tumor biopsies. Importantly, analyzing CTCs and cfDNA together enables cross-validation of mutations, uncovers mutations exclusive to either CTCs or cfDNA, and allows blood-based tumor profiling in a greater fraction of patients. Our study demonstrates the utility of analyzing both CTCs and cfDNA in MM.

Effect of Reye's syndrome serum on isolated chinchilla liver mitochondria.

A general impairment of liver mitochondrial enzymes is central to Reye's syndrome (RS). The respiration of isolated liver mitochondria was measured after the addition of concentrated normal serum or RS serum derived from 12 patients. RS serum stimulates oxygen consumption in isolated rat liver mitochondria. This effect is due to the oxidation of uric acid by peroxisomes contaminating the preparation and a stimulation of mitochondrial respiration (1.05 +/- 0.14 nmol of O2/min X mg of protein; control 0.30 +/- 0.08 nmol O2/min X mg). The stimulation of respiration occurs in the presence of all respiratory substrates, is dependent on the amount of serum added, and represents an uncoupling of oxidative phosphorylation. RS serum reduces ATP formation by 15-76%. The uncoupling effect correlates with the amount of free fatty acid in the serum sample and resembles the effect induced by the addition of a dicarboxylic fatty acid. Dicarboxylic fatty acids, especially long-chain dicarboxylic acids, impair ATP formation. Dicarboxylic acids were found in the serum of all RS patients and comprised as much as 54% of the total serum free fatty acids. 90% of the serum dicarboxylic acids were of 16-18 carbon lengths. The amount of dicarboxylic acids in the RS serum corresponded directly with the reduction in ATP formation by the RS serum. This demonstrates that dicarboxylic acids occur in RS and may be important in the general impairment of mitochondrial function in RS and other disorders where they are present.

Induction of omega-oxidation of monocarboxylic acids in rats by acetylsalicylic acid.

The accumulation of dicarboxylic acids, particularly long chain, is a prominent feature of Reye's syndrome and diseases of peroxisomal metabolism. We assessed the omega-oxidation of a spectrum of fatty acids in rats and asked whether pretreatment of rats with aspirin, which is known to predispose children to Reye's syndrome, would affect omega-oxidation of long chain fatty acids. We found that aspirin increased liver free fatty acids and increased the capacity for omega-oxidation three- to sevenfold. Omega-oxidation of long chain substrate was stimulated to a greater degree than medium chain substrate and was apparent within one day of treatment, at serum aspirin concentrations below the therapeutic range in humans. The apparent Km for lauric acid was 0.9 microM and 12 microM for palmitate. We also found a difference in the storage stability of activity toward medium and long chain substrate. Saturating concentrations of palmitate had no effect on the formation of dodecanedioic acid, whereas laurate decreased but never eliminated the omega-oxidation of palmitate. 97% of the total laurate omega-oxidative activity recovered was found in the microsomes, but 32% of palmitate omega-oxidative activity was present in the cytosol. These results demonstrate that aspirin is a potent stimulator of omega-oxidation and suggest that there may be multiple enzymes for omega-oxidation with overlapping substrate specificity.

The metabolism of proinsulin and insulin by the liver.

The removal of bovine proinsulin by the isolated perfused rat liver has been studied and the results compared with the removal of insulin. At high concentrations of insulin (> 180 ng/ml) the removal process was saturated and the t(1/2) varied between 35 and 56 min. With low initial insulin levels the disappearance followed first-order kinetics, the mean regression coefficient being - 0.022, t(1/2) 13.8 min, and the hepatic extraction 4.0 ml/min. The results with proinsulin were in striking contrast to these findings. At both high and low concentrations the hepatic removal of proinsulin was considerably slower, averaging 10-15 times less than that of insulin. Specific immunoassay techniques and gel filtration of samples taken from perfusions to which both labeled and unlabeled proinsulin had been added did not show conversion to either insulin or the C-peptide. Bovine and rat (131)I-labeled proinsulins were degraded more slowly than bovine insulin-(131)I by bovine and rat liver homogenates. Both proinsulin and insulin inhibited the degradation of insulin-(131)I, equimolar quantities of proinsulin being 2-5 times less effective than insulin. These results indicate significant differences in the capacity of the liver to remove and degrade insulin and proinsulin. The low hepatic extraction of proinsulin may account for its prolonged half-life in vivo and contribute to its relatively high plasma concentration in the fasting state. Furthermore this finding will have to be taken into account in the interpretation of changes in the proinsulin:insulin ratios in peripheral blood in a variety of metabolich situations.

Alteration of protein synthesis and induction of specific protein phosphorylation by hyperthermia.

Confluent cultures of the mouse cell line clone 1D were subjected to 1-hr hyperthermic treatments. Temperatures were increased from the control level of 37 degrees to values ranging from 38 to 45 degrees. Protein synthesis patterns were determined in fluorograms of sodium dodecyl sulfate-polyacrylamide gels labeled with [3H]leucine. Although incorporation into most proteins was either repressed or decreased by the treatment, several proteins showed an increased label of were apparently induced de novo. Among the induced proteins was a prominent band, probably a doublet, with an estimated molecular weight of 70,000 to 69,000. Crude cell lysates made from 37 degrees, 41 degrees, and 45 degrees-treated cells were tested for kinase activity at 30 degrees by a 10-min incubation with adenosine [gamma-32P]triphosphate. Several specific proteins exhibited increased phosphorylation, while phosphorylation of other proteins decreased. The most significant increase in phosphorylation was shown by a protein with molecular weight of about 37,000. We suggest that heat treatment induces or activates one or more specific phosphokinase(s) with the ability to phosphorylate proteins with approximate molecular weights of 37,000, 36,000, 23,000, and 16,000.

DNA microarrays identification of primary and secondary target genes regulated by p53.

The transcriptional program regulated by the tumor suppressor p53 was analysed using oligonucleotide microarrays. A human lung cancer cell line that expresses the temperature sensitive murine p53 was utilized to quantitate mRNA levels of various genes at different time points after shifting the temperature to 32 degrees C. Inhibition of protein synthesis by cycloheximide (CHX) was used to distinguish between primary and secondary target genes regulated by p53. In the absence of CHX, 259 and 125 genes were up or down-regulated respectively; only 38 and 24 of these genes were up and down-regulated by p53 also in the presence of CHX and are considered primary targets in this cell line. Cluster analysis of these data using the super paramagnetic clustering (SPC) algorithm demonstrate that the primary genes can be distinguished as a single cluster among a large pool of p53 regulated genes. This procedure identified additional genes that co-cluster with the primary targets and can also be classified as such genes. In addition to cell cycle (e.g. p21, TGF-beta, Cyclin E) and apoptosis (e.g. Fas, Bak, IAP) related genes, the primary targets of p53 include genes involved in many aspects of cell function, including cell adhesion (e.g. Thymosin, Smoothelin), signaling (e.g. H-Ras, Diacylglycerol kinase), transcription (e.g. ATF3, LISCH7), neuronal growth (e.g. Ninjurin, NSCL2) and DNA repair (e.g. BTG2, DDB2). The results suggest that p53 activates concerted opposing signals and exerts its effect through a diverse network of transcriptional changes that collectively alter the cell phenotype in response to stress.

Position-specific inhibition of yeast mitochondrial transcription by a poly(T) sequence.

The 3' flanking nucleotide(s) of the octanucleotide promoter sequence regulates transcriptional efficiency of some mitochondrial genes in Saccharomyces cerevisiae. To understand this regulation the in vitro transcriptional activity of various synthetic mitochondrial promoters carrying different 3' flanking sequences was examined. The results presented here demonstrate that consecutive thymidine residues, but no other polynucleotides or secondary structure, in the promoter-proximal non-transcribed DNA strand inhibited mitochondrial transcription. The location and the number of T residues in the cluster as well as the concentration of UTP in the transcription reaction are the important factors determining this transcriptional inhibition. For example, a pair of thymidine nucleotides at positions +2 and +3 is sufficient for inactivation of mitochondrial transcription, whereas more than three consecutive thymidine nucleotides beyond these positions are required for inhibition of mitochondrial transcription. However, a cluster of six to 12 thymidine residues beyond position +11, a point where mtRNA polymerase has been shown to form a stable transcription complex, did not interfere with mitochondrial transcription. Interestingly, at low UTP concentration the mtRNA polymerase generates a large quantity of aborted initiation products on a template carrying promoter-proximal poly(T) sequence probably due to the inability of the polymerase to clear this promoter. On the other hand at high UTP concentration the same mtRNA polymerase on the same mitochondrial promoter produces a higher level of productive initiation complex. These observations suggest that the mechanism of poly(T) inhibition of mitochondrial transcription is a UTP-limited transcriptional attenuation at the promoter site, which might occur under specific physiological conditions (i.e. glucose repression-derepression, switching of aerobic-anaerobic conditions).

Effect of immune deficiency on lipoproteins and atherosclerosis in male apolipoprotein E-deficient mice.

To determine whether T cells and B cells influence lipid metabolism and atherosclerosis, we crossed apolipoprotein E-deficient (apoE degrees ) mice with recombination activating gene 2-deficient (RAG2 degrees ) mice. Total plasma cholesterol levels were approximately 20% higher in male apoE degrees mice compared with the apoE degrees RAG2 degrees mice at 8 weeks of age, and plasma triglyceride levels were 2.5-fold higher in the apoE degrees mice even when plasma cholesterol levels were similar. Male mice with plasma cholesterol levels between 400 and 600 mg/dL at 8 weeks of age were euthanized at 27 and 40 weeks of age. The aortic root lesion area in the apoE degrees RAG2 degrees mice, compared with that in the immune-competent apoE degrees mice, was 81% and 57% smaller at 27 and 40 weeks of age, respectively. In contrast, there was no difference in the size of the brachiocephalic trunk lesions. Similar results were obtained with mice euthanized at 40 weeks of age that had 8-week cholesterol levels between 300 and 399 mg/dL. In apoE degrees RAG2 degrees mice, aortic root atherosclerosis was more profoundly suppressed at lower cholesterol levels. Thus, T and B cells and their products differentially influence the development of atherosclerosis at different sites. We also demonstrate a profound effect of the immune system on plasma lipid homeostasis.

Whole-genome sequencing reveals activation-induced cytidine deaminase signatures during indolent chronic lymphocytic leukaemia evolution.

Patients with chromosome 13q deletion or normal cytogenetics represent the majority of chronic lymphocytic leukaemia (CLL) cases, yet have relatively few driver mutations. To better understand their genomic landscape, here we perform whole-genome sequencing on a cohort of patients enriched with these cytogenetic characteristics. Mutations in known CLL drivers are seen in only 33% of this cohort, and associated with normal cytogenetics and unmutated IGHV. The most commonly mutated gene in our cohort, IGLL5, shows a mutational pattern suggestive of activation-induced cytidine deaminase (AID) activity. Unsupervised analysis of mutational signatures demonstrates the activities of canonical AID (c-AID), leading to clustered mutations near active transcriptional start sites; non-canonical AID (nc-AID), leading to genome-wide non-clustered mutations, and an ageing signature responsible for most mutations. Using mutation clonality to infer time of onset, we find that while ageing and c-AID activities are ongoing, nc-AID-associated mutations likely occur earlier in tumour evolution.

Neurons of the human frontal cortex display apolipoprotein E immunoreactivity: implications for Alzheimer's disease.

Apolipoprotein E (apoE) is a plasma protein that regulates lipid transport and cholesterol homeostasis. In humans, apoE occurs as 3 major isoforms (apoE2, E3, and E4). Genetic evidence demonstrates an overrepresentation of the apoE epsilon 4 allele in Alzheimer's disease (AD). While apoE immunoreactivity (IR) is associated with the amyloid plaques and neurofibrillary tangles of AD, few studies have characterized the localization of apoE in normal human brains. We examined the distribution of apoE in the cerebral cortex of normal aged individuals and compared the results to clinically diagnosed and pathologically confirmed AD cases. In addition, we characterized the apoE IR in brains from high plaque non-demented (HPND) cases. We observed consistent and widespread apoE staining in cortical neurons from normal and HPND individuals. This finding was confirmed by double immunostaining which colocalized apoE with microtubule-associated protein-2, as well as low density lipoprotein receptor-related protein, an apoE receptor found on neurons. In contrast, AD brains displayed apoE IR in plaques and neurofibrillary tangles with little neuronal staining. These data clearly establish the presence of apoE in normal neurons, supporting an intracellular role for apoE. Moreover, the results suggest that this function of apoE is disrupted in AD, where apoE staining of neurons was drastically reduced.

The effect of thyroid hormone on in vitro rat liver mitochondrial RNA synthesis.

Liver mitochondrial preparations from normal, thyroidectomized, and triiodothyronine-treated thyroidectomized rats were assayed for in vitro RNA synthetic activity. Thyroidectomized rat mitochondrial preparations incorporated UTP into RNA at 70% the rate of normal control preparations. Mitochondrial preparations from triiodothyronine-treated thyroidectomized rats incorporated UTP at rates 35%-45% greater than those of sham-injected thyroidectomized rats. These differences were statistically significant and could not be attributed to inequalities in mitochondrial sampling, dilution of labeled precursor specific activity, nucleotide substrate concentrations, or differences in ribonuclease activities.

Loss of atheroprotective effect of estradiol in immunodeficient mice.

Estradiol significantly decreases fatty streak formation in the aortic root of chow-fed apolipoprotein E-deficient mice. In contrast, immunodeficient mice with homozygous disruption at the recombinase activating gene 2 loci present fatty streak development that is insensitive to estradiol. Lymphocytes thus appear to be required for development of the atheroprotective effect of estradiol in this mouse model.

Suppression of plasma testosterone leads to an increase in serum total and high density lipoprotein cholesterol and apoproteins A-I and B.

Men have lower high density lipoprotein (HDL) and higher low density lipoprotein (LDL) levels than women. To dynamically evaluate the role of endogenous testosterone on the lipoprotein profile, eight normal men received a long-acting gonadotropin releasing hormone analog (LHRHA) for 10 weeks by SC injection. Plasma testosterone levels were acutely lowered below 1 ng/ml after 4 weeks of LHRHA treatment and remained depressed at this level for the duration of administration of the analog. There were prompt increases in total cholesterol [baseline vs. peak (milligrams per dl) mean +/- SEM, 177 +/- 18 vs. 208 +/- 22; P less than 0.005], apoprotein B (apo B; 69 +/- 12 vs. 97 +/- 13; P less than 0.05), HDL-cholesterol (23 +/- 2 vs. 33 +/- 2; P less than 0.005), and apo A-I (80 +/- 7 vs. 112 +/- 5; P less than 0.005), but not in apo A-II (40 +/- 3 vs. 40 +/- 4; P = NS) levels. The peaks occurred after 10 weeks of treatment and were followed by a fall in these values after discontinuing LHRHA. These changes were largely prevented in a second study (six men) in which LHRHA was administered together with im testosterone enanthate, which was given every 2 weeks. These results show that suppression of endogenous testosterone leads to increases in HDL and LDL, demonstrating that testosterone has an important effect on lipoprotein metabolism and plays a key role in defining the lipoprotein profile in men.

Fasting hypertriglyceridemia in noninsulin-dependent diabetes mellitus is an important predictor of postprandial lipid and lipoprotein abnormalities.

Postprandial lipoprotein metabolism may be important in atherogenesis and has not been studied in detail in noninsulin-dependent diabetes mellitus (NIDDM). We used the vitamin A fat-loading test to label triglyceride-rich lipoprotein particles of intestinal origin after ingestion of a high fat mixed meal containing 60 g fat/m2 and 60,000 U vitamin A/m2 in 12 untreated NIDDM subjects with normotriglyceridemia (NTG; triglycerides, less than 1.7 mmol/L), 7 untreated NIDDM subjects with moderate hypertriglyceridemia (HTG; triglycerides, 1.7-4.7 mmol/L), and 8 age- and weight-matched normotriglyceridemic nondiabetic controls. The postprandial triglyceride increment was greater in NIDDM with HTG (P = 0.0001) and correlated strongly in all groups with the fasting triglyceride concentration (r = 0.83; P = 0.0001). Retinyl palmitate measured in whole plasma, an Sf greater than 1000 chylomicron fraction, and an Sf less than 1000 nonchylomicron fraction was also significantly greater in NIDDM with HTG, but did not differ significantly between NIDDM with NTG and controls. In NIDDM with HTG, chylomicrons appeared to be cleared at a slower rate, as evidenced by the significantly later intersection of the chylomicron and nonchylomicron retinyl palmitate response curves (13.7 h in HTG NIDDM vs. 8.5 h in NTG NIDDM vs. 7.3 h in controls; P less than 0.01). Although fasting FFA levels were similar in all three groups, the HTG diabetic subjects had a late postprandial surge in FFAs that lasted for up to 14 h. The postprandial FFA elevation in all groups correlated with the fasting triglyceride concentration (r = 0.57; P less than 0.002) and postprandial triglyceride increment (r = 0.80; P = 0.0001). The fasting core triglyceride content of the HDL particles in NIDDM with HTG was significantly elevated compared to those in NIDDM with NTG and controls (21.0% vs. 14.0% vs. 14.1% respectively; P less than 0.05), and this increased proportionately in all groups after the meal at the expense of cholesteryl ester, the increase correlating with total plasma postprandial triglyceride increment (r = 0.51; P less than 0.01). We conclude that moderate fasting hypertriglyceridemia in NIDDM is predictive of a constellation of postprandial changes in lipids and lipoproteins that may potentiate the already unfavorable atherogenic fasting lipid profile in these subjects.

Role of basal triglyceride and high density lipoprotein in determination of postprandial lipid and lipoprotein responses.

The present study reports on the interaction between basal triglyceride and high density lipoprotein (HDL) cholesterol in determining the magnitude of postprandial triglyceridemia. The vitamin A fat-loading test was used to label intestinally derived triglyceride-rich particles after a high fat meal in 18 subjects with low HDL cholesterol and 6 control subjects who had normal fasting triglyceride and HDL cholesterol levels. The patients with low HDL cholesterol were divided into 2 groups on the basis of their basal triglyceride concentrations; 11 had normal triglyceride levels, and 7 had elevated serum triglycerides (HTG). In the HTG-low HDL group, the incremental area under the triglyceride curve was significantly greater (P less than 0.0003) than that in the other 2 groups, between whom no significant differences in triglyceride response were observed. Retinyl palmitate levels measured in whole plasma, an Sf greater than 1000 chylomicron fraction, and an Sf less than 1000 nonchylomicron fraction were also significantly greater in low HDL subjects with HTG, while the concentrations in low HDL subjects with normal triglyceride levels and control subjects were similar. Although basal HDL cholesterol levels in all study subjects were negatively correlated with the area under the incremental triglyceride curve (r = -0.42; P less than 0.05), this correlation was weak, in contrast to the correlation between fasting triglyceride levels and incremental triglyceride area (r = 0.56; P less than 0.005). Furthermore, basal HDL cholesterol levels did not correlate with the area under the chylomicron or nonchylomicron curves, whereas basal triglyceride levels were significantly correlated (P = 0.0001) with both of these variables. The HDL particles of both low HDL groups had a significantly higher proportion of triglyceride compared to the HDL particles in the control subjects. In conclusion, 1) fasting triglyceride levels are a more powerful indicator of the postprandial lipid response than basal HDL cholesterol in subjects with low HDL cholesterol levels; 2) patients with low HDL cholesterol levels do not preferentially accumulate chylomicron remnants after a meal unless they have coexisting hypertriglyceridemia; and 3) abnormalities in the levels of triglyceride-rich particles post-prandially are unlikely to be responsible for the increased incidence of atherosclerosis in low HDL patients who are normotriglyceridemic.

Postprandial lipoprotein metabolism in normal and obese subjects: comparison after the vitamin A fat-loading test.

Abnormalities in fasting lipid and lipoprotein levels are known to occur in obesity and other hyperinsulinemic states. However, postprandial lipoprotein metabolism has not been studied systematically in obese subjects using sensitive techniques to distinguish between triglyceride-rich lipoprotein particles derived from the intestine and the liver. In the present study the vitamin A fat-loading test was used to label intestinally derived triglyceride-rich lipoprotein particles in the postprandial state. Lipid parameters in seven normolipidemic obese subjects [body mass index, 43.7 +/- 2.81 kg/m2 (mean +/- SEM)] were compared to those in eight matched normal weight controls (body mass index, 23.6 +/- 0.72 kg/m2) during the 24-h period following ingestion of a mixed meal with a high fat content to which vitamin A had been added. Although subjects were selected for normal fasting lipid levels, in the obese group fasting triglycerides were significantly higher (1.35 +/- 0.12 vs. 0.68 +/- 0.08 mmol/L; P less than 0.0005) and high density lipoprotein (HDL) cholesterol was lower (0.94 +/- 0.08 vs. 1.35 +/- 0.11 mmol/L; P less than 0.01). The obese subjects had a greater postprandial triglyceride response to the test meal (P less than 0.05). The cumulative increment in total plasma triglycerides was 3.35-fold greater in obese than control subjects, while that of retinyl ester was only 1.63-fold greater, suggesting that a significant portion of the postprandial triglyceride response is due to endogenous hepatic lipoproteins. Postprandial plasma triglyceride and retinyl ester increment correlated with basal triglycerides (r = 0.72; P less than 0.005 and r = 0.57; P less than 0.03, respectively) and negatively with fasting HDL (r = -0.51; P less than 0.05 and r = -0.60; P less than 0.02, respectively). In the obese, the HDL triglyceride content increased maximally 4 h postprandially (4.1% to 6.1%; P less than 0.005) and phospholipid at 12 h (25.8% to 28.7%; P less than 0.05), with lower cholesteryl ester (21.1% to 17.5%; P less than 0.002) at 8 h, reflecting exchange of surface and core lipids with triglyceride-rich particles after the meal. In obese and control subjects the magnitude of HDL triglyceride enrichment after the meal correlated positively with the postprandial triglyceride increment (r = 0.74; P less than 0.007) and negatively with the fasting HDL cholesterol concentration (r = -0.80; P = 0.002). We conclude that even normolipidemic obese subjects have greater postprandial lipemia and triglyceride enrichment of HDL after ingestion of a high fat meal.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of feeding fish oil on the properties of lipoproteins isolated from rhesus monkeys consuming an atherogenic diet.

This study examined plasma lipids and lipoproteins of rhesus monkeys fed fish oil incorporated into a highly atherogenic diet containing saturated fat and cholesterol. The animals were fed diets containing 2% cholesterol and either 25% coconut oil (group I), 25% fish oil/coconut oil (1:1; group II), or 25% fish oil/coconut oil (3:1; group III) for 12 months (n = 8/group). Adding menhaden fish oil to the diet increased plasma eicosapentaenoic acid and docosahexaenoic acid and decreased plasma linoleic acid in animals fed the fish oil containing diets. Plasma concentrations of all lipoprotein fractions were decreased in the fish oil groups. VLDL isolated from group I animals exhibited beta-mobility on agarose gels but the VLDL from groups II and III animals did not. The group I VLDL was more highly enriched in cholesteryl ester than was VLDL from groups II and III. Group I LDL had a small but significant increase in cholesteryl ester content compared to group III LDL. No differences in HDL composition were observed in the 3 groups. At least 6 times less apo E was recovered in VLDL, IDL, and LDL from group III animals than from group I animals. Assuming 1 molecule of apo B per lipoprotein particle, there were 50% fewer VLDL, IDL, and LDL particles in group III than in group I animals. Group III also had significantly lower molar ratios of apo E/apo B in VLDL, IDL, and LDL than did group I animals. When VLDL from all 3 groups were incubated with J774 macrophages at equal protein concentrations, only the VLDL from the group I animals stimulated cholesterol esterification. Thus, introducing fish oil into an atherogenic diet reduced the number of VLDL, IDL and LDL particles in plasma by as much as 50%, reduced the cholesteryl ester content of the circulating lipoprotein, and reduced the ability of the VLDL to stimulate cholesterol esterification in macrophages.