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R J Havel

Publications and source records attributed to R J Havel.

At least 19 recordsLinked to original sources
PubMed1998

Lamellar lipoproteins uniquely contribute to hyperlipidemia in mice doubly deficient in apolipoprotein E and hepatic lipase.

Remnants of triglyceride-rich lipoproteins containing apolipoprotein (apo) B-48 accumulate in apo E-deficient mice, causing pronounced hypercholesterolemia. Mice doubly deficient in apo E and hepatic lipase have more pronounced hypercholesterolemia, even though remnants do not accumulate appreciably in mice deficient in hepatic lipase alone. Here we show that the doubly deficient mice manifest a unique lamellar hyperlipoproteinemia, characterized by vesicular particles 600 A-1,300 A in diameter. As seen by negative-staining electron microscopy, these lipoproteins also contain an electron-lucent region adjacent to the vesicle wall, similar to the core of typical lipoproteins. Correlative chemical analysis indicates that the vesicle wall is composed of a 1:1 molar mixture of cholesterol and phospholipids, whereas the electron-lucent region appears to be composed of cholesteryl esters (about 12% of the particle mass). Like the spherical lipoproteins of doubly deficient mice, the vesicular particles contain apo B-48, but they are particularly rich in apo A-IV. We propose that cholesteryl esters are removed from spherical lipoproteins of these mice by scavenger receptor B1, leaving behind polar lipid-rich particles that fuse to form vesicular lipoproteins. Hepatic lipase may prevent such vesicular lipoproteins from accumulating in apo E-deficient mice by hydrolyzing phosphatidyl choline as scavenger receptor B1 removes the cholesteryl esters and by gradual endocytosis of lipoproteins bound to hepatic lipase on the surface of hepatocytes.

Animals↗
PubMed1998

Rationale for use of non-high-density lipoprotein cholesterol rather than low-density lipoprotein cholesterol as a tool for lipoprotein cholesterol screening and assessment of risk and therapy.

The plasma level of low-density lipoprotein (LDL) cholesterol is the "gold standard" for estimating the lipoprotein-related risk for complications of atherosclerotic vascular disease. LDL cholesterol concentrations are commonly estimated by the Friedewald formula that requires only the measurement (after overnight fasting) of plasma cholesterol and triglycerides along with high-density lipoprotein (HDL) cholesterol. This value, however, is not in fact a true estimate of LDL cholesterol but rather of LDL cholesterol along with variable, usually smaller, amounts of intermediate-density lipoprotein (IDL) cholesterol and lipoprotein(a). Estimation of LDL cholesterol levels by the Friedewald formula becomes progressively less accurate as plasma triglyceride concentrations increase, and the formula is generally considered inapplicable when triglyceride levels exceed 400 mg/dL. We believe that a very simple measurement-non-HDL cholesterol (serum cholesterol minus HDL cholesterol)-has considerable potential as a screening tool for identifying dyslipoproteinemias, for risk assessment, and for assessing the results of hypolipidemic therapy. Unlike the estimation of LDL cholesterol levels by the Friedewald formula, the estimation of non-HDL cholesterol concentrations requires no assumptions about the relation of very-low-density (VLDL) cholesterol levels to plasma triglyceride concentrations. This method includes all of the cholesterol present in lipoprotein particles now considered to be potentially atherogenic [VLDL, IDL, LDL, and lipoprotein(a)]. This article provides examples of the utility of non-HDL cholesterol concentrations in clinical medicine.

Apolipoproteins B↗
PubMed1998

Receptor and non-receptor mediated uptake of chylomicron remnants by the liver.

The uptake of chylomicron remnants by rodent liver is mediated by proteins residing on the microvillous surface of hepatocytes and occurs in two steps. First, initial removal of the remnants from the blood occurs through binding to the low density lipoprotein (LDL) receptor via apo E and to hepatic lipase via polar lipids and proteins on the remnant surface. Second, chylomicron remnants are taken up into the cell mainly by the LDL receptor and follow the classical receptor-mediated pathway of endocytosis. The LDL receptor-related protein (LRP), which binds weakly to chylomicron remnants via apo E, does not appear to have a significant role in the initial removal process. The remnant particles can, however, be enriched with proteoglycan-bound apo E present on hepatocytic microvilli, which increases their affinity for LRP to the extent that they are subject to endocytosis by this receptor, particularly when the LDL receptor is deficient or down-regulated. Hepatic lipase can also mediate binding of remnants to LRP, for which it has high affinity. Lipolysis of remnant lipids by hepatic lipase may promote but is not required for interaction of remnants with the endocytic receptors. Proteoglycan-bound hepatic lipase may also mediate endocytosis of chylomicron remnants independent of apo E, so that hepatic catabolism of these particles is not completely dependent upon this apoprotein. Available data from experiments in vivo thus indicate redundancy of both steps of hepatic uptake of chylomicron remnants, consistent with the centrality of this process in nutrient delivery.

Animals↗
PubMed1998

Metabolism of lipoproteins containing apolipoprotein B in hepatic lipase-deficient mice.

Mice lacking hepatic lipase have been reported to express mild hyperlipidemia characterized by increased concentrations of large high density lipoproteins, but normal concentrations of lipoproteins containing apolipoprotein B. Whereas hepatic lipase has been implicated in the clearance and processing of chylomicron remnants in rats, no such defect was found in these mice. We have further characterized the abnormal lipoprotein phenotype in young hepatic lipase-deficient mice and have found more pronounced elevations of high density lipoproteins associated in particular with a 5-fold increase in plasma concentrations of apolipoprotein E. In addition, there was a reduction in the concentration of low density lipoproteins containing apolipoprotein B-100 and B-48 relative to precursor lipoproteins of lower density and a pronounced deficiency of apolipoprotein B-containing low density lipoproteins with density exceeding 1.029 g/mL. Conversion of radiolabeled rabbit intermediate density lipoproteins to low density lipoproteins was reduced by 6-fold as compared with wild-type mice. Although clearance of cholesteryl ester-labeled chylomicrons from the blood was unimpaired in the deficient mice, that of chylomicron remnants was reduced. Furthermore, endocytosis of chylomicron cholesteryl esters into liver cells occurred more rapidly than in wild-type mice. The unimpaired hepatic clearance of injected chylomicron particles in hepatic lipase-deficient mice may be the result of greater acquisition of apoE from high density lipoproteins during remnant formation. These studies thus demonstrate a critical role for mouse hepatic lipase in the formation of small, dense low density lipoproteins, as well as participation in the normal clearance and processing of chylomicron remnants.

Animals↗
PubMed1998

Evaluation of an immunoseparation method for quantitative measurement of remnant-like particle-cholesterol in serum and plasma.

Substantial evidence indicates that triglyceride-rich lipoprotein remnants are atherogenic. Additional research has, however, been limited by available methods for separation and quantification of remnants. We have evaluated an immunoseparation assay developed to measure cholesterol in remnant-like particles (RLP-C). This method uses monoclonal antibodies to human apolipoproteins B-100 and A-I to remove most of the apolipoprotein B-100-containing lipoproteins (namely LDL and nascent VLDL) and apolipoprotein A-I-containing lipoproteins (namely chylomicrons and HDL), leaving behind a fraction of triglyceride-rich lipoproteins, including chylomicron and VLDL remnants, both of which are enriched in apolipoprotein E. Cholesterol in the unbound fraction is measured with a sensitive enzymatic assay. The RLP-C concentration was highly correlated with total triglyceride-rich lipoproteins (sum of VLDL-cholesterol and IDL-cholesterol) separated by ultracentrifugation and by polyacrylamide gel electrophoresis (r = 0.86 and 0.76, respectively). The within-run and run-to-run imprecision (CV) of the assay was approximately 6% and 10%, respectively. The assay was not affected by hemoglobin up to 5000 mg/L (500 mg/dL), bilirubin up to 342 mmol/L (20 mg/dL), glucose up to 67 mmol/L (1200 mg/dL), or ascorbic acid up to 170 mmol/L (3.0 mg/dL). In 726 subjects (men, n = 364; women, n = 362) in the US, the 75th percentiles of RLP-C concentration were 0.17 mmol/L (6.6 mg/dL) and 0.23 mmol/L (8.8 mg/dL) in sera obtained after overnight fasting or randomly, respectively. A group of 151 patients from nine US centers and one Canadian center with coronary artery atherosclerosis established by angiography had higher median RLP-C concentrations than 302 gender- and age-matched controls (P <0.05). We conclude that the RLP-C assay compares favorably to ultracentrifugation and electrophoresis and provides a convenient and economical approach to measure triglyceride-rich lipoprotein remnants in routine clinical laboratories.

Adolescent↗
PubMed1997

Assessment of postprandial lipemia: nutritional influences.

Recent reports have contributed new information on the influence of the size and spacing of fat-containing meals on processes related to fat digestion and absorption, lymphatic transport, and postprandial responses of plasma lipoproteins. The postabsorptive concentrations of plasma triglycerides and common polymorphisms of apolipoprotein B and apolipoprotein E exert important influences on postprandial lipoprotein responses. Attention to these characteristics of individuals and groups can facilitate interpretation of responses to meals containing different fatty acids and varying amounts of cholesterol. Additional evidence is accumulating that points to disease related abnormalities of postprandial lipoprotein metabolism related to clearance mechanisms for triglyceride-rich lipoproteins and processes related to reverse cholesterol transport.

Apolipoproteins B↗
PubMed1996

Calmodulin binds to the basolateral targeting signal of the polymeric immunoglobulin receptor.

We have identified a major calmodulin (CaM)-binding protein in rat liver endosomes using 125I-CaM overlays from two-dimensional protein blots. Immunostaining of blots demonstrates that this protein is the polymeric immunoglobulin receptor (pIgR). We further investigated the interaction between pIgR and CaM using Madin-Darby canine kidney cells stably expressing cloned wild-type and mutant pIgR. We found that detergent-solubilized pIgR binds to CaM-agarose in a Ca(2+)-dependent fashion, and binding is inhibited by the addition of excess free CaM or the CaM antagonist W-13 (N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide), suggesting that pIgR binding to CaM is specific. Furthermore, pIgR is the most prominent 35S-labeled CaM-binding protein in the detergent phase of Triton X-114-solubilized, metabolically labeled pIgR-expressing Madin-Darby canine kidney cells. CaM can be chemically cross-linked to both solubilized and membrane-associated pIgR, suggesting that binding can occur while the pIgR is in intact membranes. The CaM binding site is located in the membrane-proximal 17-amino acid segment of the pIgR cytoplasmic tail. This region of pIgR constitutes an autonomous basolateral targeting signal. However, binding of CaM to various pIgR mutants suggests that CaM binding is not necessary for basolateral targeting. We suggest that CaM may be involved in regulation of pIgR transcytosis and/or signaling by pIgR.

Amino Acid Sequence↗
PubMed1996

Identification, characterization, cloning, and expression of apolipoprotein C-IV, a novel sialoglycoprotein of rabbit plasma lipoproteins.

We have identified and characterized a novel proline- and arginine-rich protein component of lipoproteins, present in up to five sialylated isoforms, in rabbit blood plasma. The pI of the desialylated protein is 5.7. Based upon its N-terminal sequence, a complete cDNA sequence of 555 nucleotides was cloned from rabbit liver. The synthesized protein is predicted to contain 124 amino acids, including a typical signal peptide of 27 residues. The mature protein of 97 amino acids, designated apolipoprotein C-IV, is associated with the lipoproteins of blood plasma, primarily very low density and high density lipoproteins. It contains two potential amphipathic helices characteristic of plasma apolipoproteins and forms discoidal micelles with phosphatidylcholine. Northern analysis shows a single 0.6-kilobase apolipoprotein C-IV mRNA, detected only in the liver, and Southern analysis suggests a single copy gene. Sialylated apolipoprotein C-IV is secreted from transfected mammalian cells. Nucleotide sequence comparisons demonstrate a strong homology to portions of the upstream regions of the mouse and human apolipoprotein C2 genes, within each of which a distinct gene has recently been identified. The nucleotide sequences and the predicted amino acid sequences, as well as corresponding cDNA sequences in the rat and monkey, indicate that the apolipoprotein C4 gene has been highly conserved during mammalian evolution.

Amino Acid Sequence↗
PubMed1996

Prolonged postprandial responses of lipids and apolipoproteins in triglyceride-rich lipoproteins of individuals expressing an apolipoprotein epsilon 4 allele.

The postprandial responses of apo B48, B100, E and lipids in triglyceride-rich lipoproteins (TRL) to a meal containing one-third of daily energy (39% fat calories) were compared in normolipidemic young men with apo E3/3 and apo E4/3 phenotypes. After the two groups consumed a diet rich in polyunsaturated fat for 15-29 d, their postabsorptive concentrations of TRL triglycerides, apo B48, and apo B100 were virtually identical, but their postprandial responses differed. In both groups, TRL apo B48 increased at 3 h but returned to postabsorptive values at 6 h only in the apo E3/3 group; in the apo E4/3 group the concentration of apo B48 at 6 h was 80% higher than postabsorptive values. TRL apo B100 also increased at 3 h in the two groups and fell to post-absorptive values at 6 h in the apo E3/3 group but remained 51% higher than postabsorptive concentrations in the apo E4/3 group; this response was closely coupled to that of TRL cholesterol and apo E. These observations suggest that clearance of intestinal and hepatogenous TRL remnants is impaired in young men with an apo E4/3 phenotype.

Adult↗
PubMed1996

The polymeric immunoglobulin receptor is the major calmodulin-binding protein in an endosome fraction from rat liver enriched in recycling receptors.

Rat liver endosomes contain one major high-affinity calmodulin-binding protein (CaMBP) that now has been identified as the polymeric immunoglobulin receptor (pIgR). In isolated endosomes pIgR was enriched in the receptor-recycling compartment (RRC); lesser enrichment was found in 'early' endosome (CURL) and much less in 'late' endosome fractions (multivesicular bodies, MVB). The distribution of the major CaMBP, shown by Western blotting or by overlay with I125-calmodulin in the isolated fractions, was consistent with rapid accumulation of I125-immunoglobulin A (IgA) in RRC and CURL after intravenous injection into rats. The receptor was also found in sinusoidal plasma membranes but not in cell fractions containing apical (bile canalicular) or lateral plasma membrane domains of the hepatocyte. The interaction of pIgR with calmodulin was shown by direct binding assays and by affinity chromatography. Thus, calmodulin is the first cytoplasmic protein shown to interact with the pIgR. We postulate that calmodulin regulates pIgA trafficking in rat liver. In addition, the receptor recycling fraction emerges as an endosomal subcompartment involved in pIgA transport via pIgR.

Adult↗
PubMed1996

Sepsis increases endocytosis of endotoxin into hepatocytes.

BACKGROUND: Chylomicrons bind endotoxins and accelerate their clearance from plasma to the liver. This results in reduced mortality from septic shock in a rodent model. We hypothesized that the clearance of the LPS-chylomicron (LPS-CM) complex by hepatocytes is due to receptor-mediated endocytosis and that sepsis up-regulates this process. METHODS: Three groups of Sprague-Dawley rats; (1) control; (2) pretreated with 10 micrograms/kg LPS 24 hours before treatment; and (3) pretreated with 17-alpha-ethinyl estradiol (EE, 5 mg/kg subcutaneously for 3 days), were infused with labeled I125-LPS alone or with I125-LPS bound to chylomicron. Livers were removed 2.5, 15, and 30 minutes after LPS injection, and hepatic endosomes were isolated from the liver homogenates by serial ultracentrifugation in sucrose gradients. RESULTS: The injection of I125-LPS-CM complexes resulted in higher levels of endosomal I125-LPS in all groups, as compared with I125-LPS alone. In addition, the endosomal uptake of I125-LPS was markedly increased by both LPS and EE pretreatments. CONCLUSIONS: These data strongly suggest a primary role for receptor-mediated endocytosis in the increased clearance of LPS when bound to chylomicron. In addition, exposure to LPS appears to increase the accumulation of LPS in endosomes by a mechanism similar to that of EE, which is known to up-regulate receptor-mediated lipoprotein uptake. This endogenous pathway for the catabolism of endotoxins may provide a teleological explanation for the hypertriglyceridemia observed during sepsis.

Animals↗