Biomedical subjects

H B Brewer

Publications and source records attributed to H B Brewer.

At least 55 records · Page 3Linked to original sources

PubMed1997

Overexpression of human lecithin:cholesterol acyltransferase in cholesterol-fed rabbits: LDL metabolism and HDL metabolism are affected in a gene dose-dependent manner.

Lecithin:cholesterol acyltransferase (LCAT) is an enzyme well known for its involvement in the intravascular metabolism of high density lipoproteins; however, its role in the regulation of apolipoprotein (apo) B-containing lipoproteins remains elusive. The present study was designed to investigate the metabolic mechanisms responsible for the differential lipoprotein response observed between cholesterol-fed hLCAT transgenic and control rabbits. 131I-labeled HDL apoA-I and 125I-labeled LDL kinetics were assessed in age- and sex-matched groups of rabbits with high (HE), low (LE), or no hLCAT expression after 6 weeks on a 0.3% cholesterol diet. In HE, the mean total cholesterol concentration on this diet, mg/dl (230 +/- 50), was not significantly different from that of either LE (313 +/- 46) or controls (332 +/- 52) due to the elevated level of HDL-C observed in HE (127 +/- 19), as compared with both LE (100 +/- 33) and controls (31 +/- 4). In contrast, the mean nonHDL-C concentration for HE (103 +/- 33) was much lower than that for either LE (213 +/- 39) or controls (301 +/- 55). FPLC analysis of plasma confirmed that HDL was the predominant lipoprotein class in HE on the cholesterol diet, whereas cholesteryl ester-rich, apoB-containing lipoproteins characterized the plasma of LE and, most notably, of controls. In vivo kinetic experiments demonstrated that the differences in HDL levels noted between the three groups were attributable to distinctive rates of apoA-I catabolism, with the mean fractional catabolic rate (FCR, d-1) of apoA-I slowest in HE (0.282 +/- 0.03), followed by LE (0.340 +/- 0.01) and controls (0.496 +/- 0.04). A similar, but opposite, pattern was observed for nonHDL-C levels and LDL metabolism (h-1), such that HE had the lowest nonHDL-C levels with the fastest rate of clearance (0.131 +/- 0.027), followed by LE (0.057 +/- 0.009) and controls (0.031 +/- 0.001). Strong correlations were noted between LCAT activity and both apoA-I (r= -0.868, P < 0.01) and LDL (r = 0.670, P = 0.06) FCR, indicating that LCAT activity played a major role in the mediation of lipoprotein metabolism. In summary, these data are the first to show that LCAT overexpression can regulate both LDL and HDL metabolism in cholesterol-fed rabbits and provide a potential explanation for the prevention of diet-induced atherosclerosis observed in our previous study.

Animals↗

PubMed1996

Overexpression of lecithin:cholesterol acyltransferase in transgenic rabbits prevents diet-induced atherosclerosis.

Lecithin:cholesterol acyltransferase (LCAT) is a key plasma enzyme in cholesterol and high density lipoprotein (HDL) metabolism. Transgenic rabbits overexpressing human LCAT had 15-fold greater plasma LCAT activity that nontransgenic control rabbits. This degree of overexpression was associated with a 6.7-fold increase in the plasma HDL cholesterol concentration in LCAT transgenic rabbits. On a 0.3% cholesterol diet, the HDL cholesterol concentrations increased from 24 +/- 1 to 39 +/- 3 mg/dl in nontransgenic control rabbits (n = 10; P < 0.05) and increased from 161 +/- 5 to 200 +/- 21 mg/dl (P < 0.001) in the LCAT transgenic rabbits (n = 9). Although the baseline non-HDL concentrations of control (4 +/- 3 mg/dl) and transgenic rabbits (18 +/- 4 mg/dl) were similar, the cholesterol-rich diet raised the non-HDL cholesterol concentrations, reflecting the atherogenic very low density, intermediate density, and low density lipoprotein particles observed by gel filtration chromatography. The non-HDL cholesterol rose to 509 +/- 57 mg/dl in controls compared with only 196 +/- 14 mg/dl in the LCAT transgenic rabbits (P < 0.005). The differences in the plasma lipoprotein response to a cholesterol-rich diet observed in the transgenic rabbits paralleled the susceptibility to developing aortic atherosclerosis. Compared with nontransgenic controls, LCAT transgenic rabbits were protected from diet-induced atherosclerosis with significant reductions determined by both quantitative planimetry (-86%; P < 0.003) and quantitative immunohistochemistry (-93%; P < 0.009). Our results establish the importance of LCAT in the metabolism of both HDL and apolipoprotein B-containing lipoprotein particles with cholesterol feeding and the response to diet-induced atherosclerosis. In addition, these findings identify LCAT as a new target for therapy to prevent atherosclerosis.

Animals↗

PubMed1996

Hyperalphalipoproteinemia in human lecithin cholesterol acyltransferase transgenic rabbits. In vivo apolipoprotein A-I catabolism is delayed in a gene dose-dependent manner.

Lecithin cholesterol acyltransferase (LCAT) is an enzyme involved in the intravascular metabolism of high density lipoproteins (HDLs). Overexpression of human LCAT (hLCAT) in transgenic rabbits leads to gene dose-dependent increases of total and HDL cholesterol concentrations. To elucidate the mechanisms responsible for this effect, 131I-HDL apoA-I kinetics were assessed in age- and sex-matched groups of rabbits (n=3 each) with high, low, or no hLCAT expression. Mean total and HDL cholesterol concentrations (mg/dl), respectively, were 162+/-18 and 121+/-12 for high expressors (HE), 55+/-6 and 55+/-10 for low expressors (LE), and 29+/-2 and 28+/-4 for controls. Fast protein liquid chromatography analysis of plasma revealed that the HDL of both HE and LE were cholesteryl ester and phospholipid enriched, as compared with controls, with the greatest differences noted between HE and controls. These compositional changes resulted in an incremental shift in apparent HDL particle size which correlated directly with the level of hLCAT expression, such that HE had the largest HDL particles and controls the smallest. In vivo kinetic experiments demonstrated that the fractional catabolic rate(FCR, d(-1)) of apoA-I was slowest in HE (0.328+/-0.03) followed by LE (0.408+/-0.01) and, lastly, by controls (0.528+/-0.04). ApoA-I FCR was inversely associated with HDL cholesterol level (r=-0.851,P<0.01) and hLCAT activity (r=-0.816, P<0.01). These data indicate that fractional catabolic rate is the predominant mechanism by which hLCAT overexpression differentially modulates HDL concentrations in this animal model. We hypothesize that LCAT-induced changes in HDL composition and size ultimately reduce apoA-I catabolism by altering apoA-I conformation and/or HDL particle regeneration.

Animals↗

PubMed1996

Lecithin:cholesterol acyltransferase overexpression generates hyperalpha-lipoproteinemia and a nonatherogenic lipoprotein pattern in transgenic rabbits.

Cholesterol esterification within plasma lipoprotein particles is catalyzed by lecithin:cholesterol acyltransferase (LCAT). The impact of the overexpression of this enzyme on plasma concentrations of the different plasma lipoproteins in an animal model expressing cholesteryl ester transfer protein was evaluated by generating rabbits expressing human LCAT. A 6.2-kilobase human genomic DNA construct was injected into the pronuclei of rabbit embryos. Of the 1002 embryos that were injected, 3 founder rabbits were characterized that expressed the human LCAT gene. As in mice and humans, the principal sites of mRNA expression in these rabbits is in the liver and brain, indicating that the regulatory elements required for tissue-specific expression among these species are similar. The alpha-LCAT activity correlated with the number of copies of LCAT that integrated into the rabbit DNA. Compared with controls, the high expressor LCAT-transgenic rabbits total and high density lipoprotein (HDL) cholesterol concentrations were increased 1.5-2.5-fold with a 3.1-fold increase in the plasma cholesterol esterification rate. Analysis of the plasma lipoproteins by fast protein liquid chromatography indicates that these changes reflected an increased concentration of apolipoprotein E-enriched, HDL1-sized particles, whereas atherogenic apolipoprotein B particles disappeared from the plasma. The concentrations of plasma HDL cholesterol were highly correlated with both human LCAT mass (r = 0.93; p = 0.001) and the log LCAT activity (r = 0.94; p < 0.001) in the transgenic rabbits. These results indicate that overexpression of LCAT in the presence of cholesteryl ester transfer protein leads to both hyperalpha-lipoproteinemia and reduced concentrations of atherogenic lipoproteins.

Animals↗

PubMed1996

Hepatic lipase gene therapy in hepatic lipase-deficient mice. Adenovirus-mediated replacement of a lipolytic enzyme to the vascular endothelium.

Hepatic lipase (HL) is an endothelial-bound lipolytic enzyme which functions as a phospholipase as well as a triacylglycerol hydrolase and is necessary for the metabolism of IDL and HDL. To evaluate the feasibility of replacing an enzyme whose in vivo physiologic function depends on its localization on the vascular endothelium, we have infused recombinant replication-deficient adenovirus vectors expressing either human HL (HL-rAdV; n = 7) or luciferase cDNA (Lucif-rAdV; n = 4) into HL-deficient mice with pretreatment plasma cholesterol, phospholipid, and HDL cholesterol values of 176 +/- 9, 314 +/- 12, and 129 +/- 9, respectively. After infusion of HL-rAdV, HL could be detected in the postheparin plasma of HL-deficient mice by immunoblotting and postheparin plasma HL activities were 25,700 +/- 4,810 and 1,510 +/- 688 nmol/min/ml on days 5 and 15, respectively. Unlike the mouse HL, 97% of the newly synthesized human HL was heparin releasable, indicating that the human enzyme was virtually totally bound to the mouse vascular endothelium. Infusion of HL-rAdV in HL-deficient mice was associated with a 50-80% decrease in total cholesterol, triglyceride, phospholipids, cholesteryl ester, and HDL cholesterol (P < 0.001) as well as normalization of the plasma fast protein liquid chromatography lipoprotein profile by day 8. These studies demonstrate successful expression and delivery of a lipolytic enzyme to the vascular endothelium for ultimate correction of the HL gene defect in HL-deficient mice and indicate that recombinant adenovirus vectors may be useful in the replacement of endothelial-bound lipolytic enzymes in human lipolytic deficiency states.

Adenoviridae↗

PubMed1996

Adenoviral delivery of low-density lipoprotein receptors to hyperlipidemic rabbits: receptor expression modulates high-density lipoproteins.

Plasma concentrations of low-density lipoproteins (LDLs) and high-density lipoproteins (HDLs) are inversely related in several dyslipoproteinemias. To elucidate the interactions between these lipoproteins, we used a recombinant adenovirus (hLDLR-rAdV) to express human LDL receptors (hLDLRs) in LDL receptor-deficient rabbits. hLDLR-rAdV administration resulted in hepatocyte expression and a reduction of total, intermediate-density lipoprotein (IDL), and LDL cholesterol. In addition, we found that hLDLR-rAdV treatment induced (1) increased very-low-density lipoprotein (VLDL) cholesterol, (2) increased VLDL, IDL and LDL triglycerides, (3) decreased alpha- and pre-beta-migrating apolipoprotein E (apo E) and decreased pre-beta-migrating apo A-I at 2 to 4 days posttreatment, and (4) increased total plasma apo A-I and pre-beta-migrating apo A-I beginning 8 to 10 days posttreatment. Virtually all plasma apo A-I was present on alpha- and pre-beta-HDL. Pre-beta-HDL particles with size and electrophoretic properties consistent with nascent HDL demonstrated the greatest relative apo A-I enrichment following hLDLR-rAdV treatment. In summary, enhanced expression of hepatocyte LDLRs by hLDLR-rAdV treatment markedly altered apo A-I-containing lipoproteins and IDL and LDL. The use of recombinant viruses to express physiologically relevant genes in intact animals, analogous to transfection of cells in culture, provides a new strategy for the evaluation of effects of specific gene products on metabolic systems in vivo.

Adenoviridae↗

PubMed1996

Homozygous familial hypobetalipoproteinemia. Increased LDL catabolism in hypobetalipoproteinemia due to a truncated apolipoprotein B species, apo B-87Padova.

Mutations on the apolipoprotein (apo) B gene that interfere with the full-length translation of the apoB molecule are associated with familial hypobetalipoproteinemia (FHBL), a disease characterized by the reduction of plasma apoB and LDL cholesterol. In this report, we describe an FHBL kindred carrying a unique truncated apoB form, apoB-87Padova. Sequence analysis of amplified genomic DNA identified a single G deletion at nucleotide 12032, which shifts the translation reading frame and causes a termination at amino acid 3978. Two homozygous subjects and seven heterozygous relatives were studied. Although homozygous individuals had only trace amounts of LDL, they were virtually free from the symptoms typical of homozygous FHBL subjects. We investigated the in vivo turnover of radiolabeled normal apoB-100 LDL and apoB-87 LDL in one homozygous patient and two normal control subjects. ApoB-87 LDL showed a similar metabolism in all three subjects, with a fractional catabolic rate more than double that of normal LDL. The rate of entry of apoB-87 in the LDL compartment was also markedly decreased compared with normal apoB-100. The increased in vivo catabolism of apoB-87 LDL was paralleled in vitro by a 2.5-fold increased ability of these particles to inhibit the uptake and degradation of normal apoB-100 LDL by normal human cultured fibroblasts. These results indicate that apoB-87 LDL has an enhanced ability to interact with the LDL receptor, the increased apoB catabolism contributes to the hypobetalipoproteinemia and may explain the mild expression of the disease in the two homozygous individuals.

Adult↗

PubMed1996

Exon 10 skipping caused by intron 10 splice donor site mutation in cholesteryl ester transfer protein gene results in abnormal downstream splice site selection.

Cholesteryl ester transfer protein (CETP) deficiency is the most common cause of hyperalphalipoproteinemia in Japan. However, the genetic basis of this disorder has not been fully characterized. We have studied a 49-year-old Japanese male presenting with total cholesterol, HDL-cholesterol, and apolipoprotein A-I levels of 300, 236, and 233 mg/dl, respectively, and total absence of CETP activity and mass in plasma. Sequence analysis of the patient's CETP gene revealed that the splice donor consensus GT was substituted by GG in intron 10 (intron 10 splice defect) and by AT in intron 14 (intron 14 splice defect). Restriction digestion of PCR-amplified DNA using NdeI and MaeIII established that the patient was a compound heterozygote for both gene defects. Sequencing of cDNA amplified by RT-PCR from the patient's monocyte-derived macrophage RNA demonstrated abnormal splicing with deletion of exon 10 as well as alternative splicing at a native AG site located 31 nucleotides 5' of the normal splice acceptor in intron 13. Thus, the intron 10 splice defect results in exon 10 skipping and the insertion of a 31 bp fragment between exon 13 and exon 14, which contains an in frame stop codon. The presence of abnormally spliced mRNA was further confirmed by amplification of patient cDNA using CETP specific primers. Abnormal splicing of exon 14 as a result of the intron 14 splice defect was not detected, indicating potential unstable CETP mRNA derived from that mutation. These findings demonstrate that a novel splice site mutation in intron 10 of the CETP gene results in the skipping of exon 10, as well as disruption of downstream splicing at intron 13 identifying a novel mechanism leading to CETP deficiency.

Base Sequence↗

PubMed1996

ApoA-II kinetics in humans using endogenous labeling with stable isotopes: slower turnover of apoA-II compared with the exogenous radiotracer method.

ApoA-II is a major apolipoprotein constituent of high density lipoproteins (HDL) and may play an important role in lipoprotein metabolism and predisposition to atherosclerosis. Previous radiotracer kinetic studies have suggested that the metabolism of apoA-II in humans may be different than the metabolism of apoA-I, the major HDL apolipoprotein. In the present study, we have used an endogenous labeling technique using stable isotopically labeled amino acids to study apoA-II metabolism and compared the results to those obtained by a simultaneous exogenous radiotracer labeling method. Seven subjects with HDL cholesterol levels ranging from 9 to 93 mg/dl and apoA-II levels from 13 to 60 mg/dl were investigated in this study. [13C6]phenylalanine and 131I-labeled apoA-II were simultaneously administered as a primed-constant infusion and a bolus injection, respectively. In the endogenous labeling study, plateau tracer/tracee ratios of VLDL apoB-100 were used as estimates for the precursor pool tracer/tracee ratios for apoA-II synthesis. Residence times of apoA-II using these two independent methods were found to be highly correlated (r = 0.973, P < 0.0002). These results indicate that the endogenous labeling of apoA-II using stable isotopically labeled amino acids is a reasonable alternative to the conventional exogenous radiotracer labeling method for the investigation of apoA-II turnover. However, under the conditions of our experimental design and modeling strategy, the apoA-II residence times as determined by endogenous labeling were significantly longer (mean 5.33 days) than by exogenous radiotracer (mean 4.65 days). This suggests that apoA-II turnover may be even slower than believed based on radiotracer studies, and further supports the concept that HDL containing apoA-II are metabolized differently than HDL without apoA-II.

Apolipoprotein A-I↗

PubMed1995

In vitro expression of structural defects in the lecithin-cholesterol acyltransferase gene.

Classic LCAT deficiency (CLD) and fish eye disease (FED) are two clinically distinct syndromes, associated with defects in the lecithin-cholesterol acyltransferase (LCAT) gene resulting in total (CLD) or partial (FED) enzyme deficiency. In order to investigate the underlying molecular mechanisms that lead to different phenotypic expression in CLD and FED, LCAT mutants associated with either CLD (LCAT147, LCAT156, and LCAT228) or FED (LCAT10, LCAT123, LCAT158, LCAT293, LCAT300, and LCAT347) were expressed in vitro in human embryonic kidney 293 cells and characterized with respect to LCAT expression and enzyme activity. Evaluation of mutant LCAT gene transcription by Northern blot analysis demonstrated LCAT mRNA of normal size and concentration. Although all constructs gave rise to similar intracellular LCAT mass, the amount of enzyme present in the media for LCAT147, LCAT156, and LCAT300 was reduced to less than 10% of normal, suggesting that these mutations disrupted LCAT secretion. Western blot analysis of cell culture media containing wild type or mutant LCAT demonstrated the presence of a single normal-sized band of 67 kDa. The ability of the different enzymes to esterify free cholesterol in high density lipoprotein-like proteoliposomes (alpha-LCAT-specific activity) was reduced to less than 5% of normal for CLD mutants LCAT147 and LCAT228 and FED mutants LCAT10, LCAT123, LCAT293, and LCAT347, whereas that of LCAT156, LCAT158, and LCAT300 ranged from 45 to 110% of control. Although most FED mutant LCAT enzymes retained the ability to esterify free cholesterol present in alpha- and beta-lipoproteins of heat-inactivated plasma, esterification was undetectable in all CLD mutants (LCAT147, LCAT156, and LCAT228). In contrast, all mutant enzymes retained the ability to hydrolyze the water soluble, short-chained fatty acid substrate p-nitrophenolbutyrate. In summary, our studies establish the functional significance of nine LCAT gene defects associated with either FED or CLD. Characterization of the expressed LCAT mutants identified multiple, overlapping functional abnormalities that include defects in secretion and/or disruption of enzymic activity. All nine LCAT mutants retained the ability to hydrolyze the water-soluble PNPB substrate, indicating intact hydrolytic function. Based on these studies we propose that mutations in LCAT residues 147, 156, 228 (CLD) and 10, 123, 158, 293, 300, and 347 (FED) do not disrupt the functional domain mediating LCAT phospholipase activity, but alter structural domains involved in lipid binding or transesterification.

Blotting, Northern↗

PubMed1995

Carboxyl-terminal domain truncation alters apolipoprotein A-I in vivo catabolism.

Apolipoprotein A-I (apoA-I), the major protein of high density lipoproteins, facilitates reverse cholesterol transport from peripheral tissue to liver. To determine the structural motifs important for modulating the in vivo catabolism of human apoA-I (h-apoA-I), we generated carboxyl-terminal truncation mutants at residues 201 (apoA-I201), 217 (apoA-I217), and 226 (apoA-I226) by site-directed mutagenesis. ApoA-I was expressed in Escherichia coli as a fusion protein with the maltose binding protein, which was removed by factor Xa cleavage. The in vivo kinetic analysis of the radioiodinated apoA-I in normolipemic rabbits revealed a markedly increased rate of catabolism for the truncated forms of apoA-I. The fractional catabolic rates (FCR) of 9.10 +/- 1.28/day (+/- S.D.) for apoA-I201, 6.34 +/- 0.81/day for apoA-I217, and 4.42 +/- 0.51/day for apoA-I226 were much faster than the FCR of recombinant intact apoA-I (r-apoA-I, 0.93 +/- 0.07/day) and h-apoA-I (0.91 +/- 0.34/day). All the truncated forms of apoA-I were associated with very high density lipoproteins, whereas the intact recombinant apoA-I (r-apoA-I) and h-apoA-I associated with HDL2 and HDL3. Gel filtration chromatography revealed that in contrast to r-apoA-I, the mutant apoA-I201 associated with a phospholipid-rich rabbit apoA-I containing particle. Analysis by agarose gel electrophoresis demonstrated that the same mutant migrated in the pre-beta position, but not within the alpha position as did r-apoA-I. These results indicate that the carboxyl-terminal region (residue 227-243) of apoA-I is critical in modulating the association of apoA-I with lipoproteins and in vivo metabolism of apoA-I.

Animals↗

PubMed1995

Lipids.

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Cardiovascular Diseases↗

PubMed1995

Characterization of high-density apolipoprotein particles A-I and A-I:A-II isolated from humans with cholesteryl ester transfer protein deficiency.

The cholesteryl ester transfer protein (CETP) plays an important role in metabolism of high-density lipoprotein and reverse cholesterol transport in humans. The two major classes of high-density lipoprotein particles are those containing apolipoprotein A-I (LpA-I) and those containing both apoA-I and apoA-II (LpA-I:A-II). We isolated and characterized the apoA-I-containing lipoprotein particles from three subjects with homozygous CETP deficiency (CETP-D) and compared the results with those from normolipidemic control subjects. Plasma concentrations of apoA-I in both LpA-I and LpA-I:A-II were significantly elevated in CETP-D subjects. Both LpA-I and LpA-I:A-II from these subjects were larger and contained more cholesteryl ester per particle than control particles. In CETP-D, subpopulations of LpA-I and LpA-I:A-II with an unusually large size (Stokes diameters 13.8 nm and 12.6 nm, respectively) not detected in normal subjects were isolated. The molar ratio of apoA-I to apoA-II in LpA-I:A-II isolated from CETP-D subjects was higher (mean 2.4) than those of controls (mean 1.4). ApoE was primarily associated with LpA-I:A-II in CETP-D subjects. A subclass of LpA-I with pre-beta migration on agarose electrophoresis was increased in CETP-D subjects. Both LpA-I and LpA-I:A-II from CETP-D subjects bound with higher affinity but less capacity to HepG2 cells compared with control particles, and were internalized to a lesser extent than control particles. These data suggest that the absence of CETP in humans significantly affects the plasma concentration, size, composition, and cellular interaction of both major classes of apoA-I-containing lipoprotein particles.

Adult↗

PubMed1995

Lipoproteins containing apolipoprotein B isolated from patients with abetalipoproteinemia and homozygous hypobetalipoproteinemia: identification and characterization.

Abetalipoproteinemia (ABL) and homozygous hypobetalipoproteinemia (HBL) are inherited disorders which are classically characterized by progressive retinal and spinocerebellar disease, fat-soluble vitamin deficiency, and absence of apolipoprotein (apo) B from the plasma. Using immunoaffinity chromatography with an anti-apo B antiserum, we isolated apo B-containing lipoprotein (LpB) particles from the plasma of 4 ABL and 2 HBL patients. The LpB particles were characterized and compared with low density lipoprotein (LDL) and LpB isolated from normal plasma. The ABL/HBL LpB particles were similar in size and charge to normal LpB particles but were relatively enriched in several other apolipoproteins. They contained alpha-tocopherol in a ratio to cholesterol that was proportionately much higher than the very low ratio of alpha-tocopherol to cholesterol in plasma. They bound saturably to fibroblasts and were internalized and degraded similarly to LDL. Hence, the molecular defects in ABL and HBL permit the secretion of a very small number of apo B-containing lipoproteins which may be important for transport of alpha-tocopherol to peripheral tissues.

Abetalipoproteinemia↗

PubMed1995

A pilot study of ex vivo gene therapy for homozygous familial hypercholesterolaemia.

The outcome of the first pilot study of liver-directed gene therapy is reported here. Five patients with homozygous familial hypercholesterolaemia (FH) ranging in age from 7 to 41 years were enrolled; each patient tolerated the procedure well without significant complications. Transgene expression was detected in a limited number of hepatocytes of liver tissue harvested four months after gene transfer from all five patients. Significant and prolonged reductions in low density lipoprotein (LDL) cholesterol were demonstrated in three of five patients; in vivo LDL catabolism was increased 53% following gene therapy in a receptor negative patient, who realized a reduction in serum LDL equal to approximately 150 mg dl-1. This study demonstrates the feasibility of engrafting limited numbers of retrovirus-transduced hepatocytes without morbidity and achieving persistent gene expression lasting at least four months after gene therapy. The variable metabolic responses observed following low-level genetic reconstitution in the five patients studied precludes a broader application of liver-directed gene therapy without modifications that consistently effect substantially greater gene transfer.

Adult↗

PubMed1995

Lipoprotein (a) in patients with hyperlipidaemia.

Lipoprotein (a) [Lp(a)] is an atherogenic lipoprotein which is similar in structure to, but metabolically distinct from, LDL. Factors modulating plasma Lp(a) concentrations are poorly understood. We hypothesized that patients with hyperlipidaemia have elevated Lp(a) levels and determined the phenotype, concentration and distribution of Lp(a) in a group of hyperlipidaemic patients (n = 107) compared with a control group (n = 128). Lp(a) concentrations were significantly increased in the hyperlipidaemic patients (mean, 34 +/- 4 mg dL-1; median, 19 mg dL-1) as compared with the controls (20 +/- 3 mg dL-1; 9 mg dL-1) (P < 0.01). Interestingly, after dividing the patients into one group with elevated cholesterol (> 200 mg dL-1) (n = 44) and another group with elevated triglycerides (> 200 mg dL-1) (n = 51) we found that Lp(a) concentrations were 2.3-fold higher in the high cholesterol patients (mean, 45 +/- 5; median, 41 mg dL-1) compared to the high triglyceride subjects (20 +/- 4; 8 mg dL-1) (P < 0.01). Furthermore, a negative correlation between triglyceride and Lp(a) plasma concentrations was found in patients exhibiting triglyceride levels > 300 mg dL-1 (r = -0.41, P = 0.04, n = 36) and with triglycerides > 400 mg dL-1 (r = -0.52, P = 0.03, n = 17). These data indicate that plasma Lp(a) concentrations are elevated in hyperlipidaemia if the patients have high cholesterol levels, whereas Lp(a) is normal to low in patients with elevated triglycerides.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1995

Apolipoprotein A-II production rate is a major factor regulating the distribution of apolipoprotein A-I among HDL subclasses LpA-I and LpA-I:A-II in normolipidemic humans.

HDLs are heterogeneous in their apolipoprotein composition. Apolipoprotein (apo) A-I and apoA-II are the major proteins found in HDL and form the two major HDL subclasses: those that contain only apoA-I (LpA-I) and those that contain both apoA-I and apoA-II (LpA-I:A-II). Substantial evidence indicates that these two subclasses differ in their in vivo metabolism and effect on atherosclerosis, with LpA-I the more specifically protective subfraction against atherosclerosis. The purpose of this study was to investigate the effect of apoA-I and apoA-II production and catabolism on plasma LpA-I and LpA-I:A-II levels. Fifty normolipidemic subjects (those with HDL cholesterol levels in the top and bottom tenth percentiles were excluded) underwent kinetic studies with radiolabeled apoA-I and apoA-II, and the kinetic parameters of apoA-I and apoA-II were correlated with LpA-I and LpA-I:A-II levels. ApoA-I levels were strongly correlated with apoA-I residence times and less strongly correlated with apoA-I production rates. In contrast, apoA-II levels were correlated only with apoA-II production rates and not with apoA-II residence times. Levels of apoA-I in LpA-I were correlated with apoA-I residence times, whereas levels of apoA-I in LpA-I:A-II were correlated primarily with apoA-II production rates. The fraction of apoA-I in LpA-I was highly inversely correlated with apoA-II production rate (r = -.67, P < .001). In multiple regression analysis, apoA-II production rate was the most significant independent variable determining percent apoA-I in LpA-I among all the kinetic parameters.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1995

Dominant expression of type III hyperlipoproteinemia. Pathophysiological insights derived from the structural and kinetic characteristics of ApoE-1 (Lys146-->Glu).

Type III hyperlipoproteinemia is characterized by delayed chylomicron and VLDL remnant catabolism and is associated with homozygosity for the apoE-2 allele. We have identified a kindred in which heterozygosity for an apoE mutant, apoE-1 (Lys146-->Glu), is dominantly associated with the expression of type III hyperlipoproteinemia. DNA sequence analysis of the mutant apoE gene revealed a single-point mutation that resulted in the substitution of glutamic acid (GAG) for lysine (AAG) at residue 146 in the proposed receptor-binding domain of apoE. The pathophysiological effect of this mutation was investigated in vivo by kinetic studies in the patient and six normal subjects, and in vitro by binding studies of apoE-1 (Lys146-->Glu) to LDL receptors on human fibroblasts and to heparin. The kinetic studies revealed that apoE-1 (Lys146-->Glu) was catabolized significantly slower than apoE-3 in normals (P < 0.005). In the proband, the plasma residence times of both apoEs were substantially longer and the production rate of total apoE was about two times higher than in the control subjects. ApoE-1 (Lys146-->Glu) was defective in interacting with LDL receptors, and its ability to displace LDL in an in vitro assay was reduced to 7.7% compared with apoE-3. The affinity of apoE-1 (Lys146-->Glu) to heparin was also markedly reduced compared with both apoE-2 (Arg158-->Cys) and apoE-3. These abnormal in vitro binding characteristics and the altered in vivo metabolism of apoE-1 (Lys146-->Glu) are proposed to result in the functional dominance of this mutation in the affected kindred.

Adult↗