Clinical expression of familial hypercholesterolemia in clusters of mutations of the LDL receptor gene that cause a receptor-defective or receptor-negative phenotype.

Bertolini S, Cantafora A, Averna M, Cortese C, Motti C, Martini S, Pes G, Postiglione A, Stefanutti C, Blotta I, Pisciotta L, Rolleri M, Langheim S, Ghisellini M, Rabbone I, Calandra S.

Department of Internal Medicine, University of Genoa, Italy.

Seventy-one mutations of the low density lipoprotein (LDL) receptor gene were identified in 282 unrelated Italian familial hypercholesterolemia (FH) heterozygotes. By extending genotype analysis to families of the index cases, we identified 12 mutation clusters and localized them in specific areas of Italy. To evaluate the impact of these mutations on the clinical expression of FH, the clusters were separated into 2 groups: receptor-defective and receptor-negative, according to the LDL receptor defect caused by each mutation. These 2 groups were comparable in terms of the patients' age, sex distribution, body mass index, arterial hypertension, and smoking status. In receptor-negative subjects, LDL cholesterol was higher (+18%) and high density lipoprotein cholesterol lower (-5%) than the values found in receptor-defective subjects. The prevalence of tendon xanthomas and coronary artery disease (CAD) was 2-fold higher in receptor-negative subjects. In patients >30 years of age in both groups, the presence of CAD was related to age, arterial hypertension, previous smoking, and LDL cholesterol level. Independent contributors to CAD in the receptor-defective subjects were male sex, arterial hypertension, and LDL cholesterol level; in the receptor-negative subjects, the first 2 variables were strong predictors of CAD, whereas the LDL cholesterol level had a lower impact than in receptor-defective subjects. Overall, in receptor-negative subjects, the risk of CAD was 2.6-fold that of receptor-defective subjects. Wide interindividual variability in LDL cholesterol levels was found in each cluster. Apolipoprotein E genotype analysis showed a lowering effect of the epsilon2 allele and a raising effect of the epsilon4 allele on the LDL cholesterol level in both groups; however, the apolipoprotein E genotype accounted for only 4% of the variation in LDL cholesterol. Haplotype analysis showed that all families of the major clusters shared the same intragenic haplotype cosegregating with the mutation, thus suggesting the presence of common ancestors.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10978268&dopt=Abstract cholesterol




[Differences in clinical presentation between subjects with a phenotype of familial hypercholesterolemia determined by defects in the LDL-receptor and defects in Apo B-100]

[Article in Spanish]

Garcia-Alvarez I, Castillo S, Mozas P, Tejedor D, Reyes G, Artieda M, Cenarro A, Alonso R, Mata P, Pocovi M, Civeira F; Grupo de Estudio de la Hipercolesterolemia Familiar.

Departamentos de Medicina y Psiquiatria. Hospital Universitario Miguel Servet. Zaragoza. Espana. garalgar excite.com

INTRODUCTION AND OBJECTIVES: Familial hypercholesterolemia and familial defective Apo B-100 are phenotypically indistinguishable. At present they can be distinguished by genetic analysis. PATIENTS AND METHODc We compared the clinical features of 13 subjects with familial defective Apo B-100 and 39 subjects with familial hypercholesterolemia. We used data from first degree relatives to compare morbidity and mortality between the two groups. RESULTS: We found statistically significant differences in total cholesterol and LDL cholesterol, which were lower in the familial defective Apo B-100 group (TC = 357 37.3 mg/dl vs 415 79.7 mg/dl and LDLc = 270 34.2 mg/dl vs 355 72.4 mg/dl). We found no differences in xanthomas, corneal arcus, smoking status, vascular events, blood pressure, BMI or waist/hip ratio. There were no differences between the two groups in the proportions of patients with cardiovascular disease or patients who died. We found statistically significant differences between the groups (p = 0.023) in the mean age at first vascular event (familial hypercholesterolemia and first degree relatives: 45.3 19.9 years; familial defective Apo B-100 and first degree relatives: 51.5 20.8 years). CONCLUSIONS: We conclude that familial defective Apo B-100 results in clinically milder hypercholesterolemia than familial hypercholesterolemia, and that discerning between them could be helpful to stratify the risk in persons with hereditary hypercholesterolemia.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12892621&dopt=Abstract cholesterol




Analysis of unsaturated compounds by Ag+ coordination ionspray mass spectrometry: studies of the formation of the Ag+/lipid complex.

Seal JR, Havrilla CM, Porter NA, Hachey DL.

Department of Chemistry, the Center in Molecular Toxicology, Vanderbilt University, Nashville, Tennessee 37235, USA.

Coordination ionspray mass spectrometry (CIS-MS) is a useful tool in the detection and identification of cholesterol ester and phospholipid hydroperoxides and diacyl peroxides. Extensive studies of a series of cholesterol esters using CIS-MS revealed the following: (1) Cholesterol esters with equal number of double bonds as the internal standard showed a linear relative response in the mass spectrometer while compounds with non-equal numbers of double bonds gave a nonlinear relative response. (2) Complex adducts containing cholesterol ester, silver ion, AgF, AgBF(4), and 2-propanoxide form when silver is in molar excess of cholesterol esters, reducing the [M + Ag](+) signal. (3) In a mixture of cholesterol esters where silver is limiting, Ch22:6 and Ch20:4 bind to silver at the expense of Ch18:2 and have a higher signal in the mass spectrometer. (4) In a mixture of cholesterol esters where silver concentration is twofold greater than total cholesterol ester concentration, Ch22:6 and Ch20:4 form large complex adducts more frequently than Ch18:2 and have a lower signal in the mass spectrometer.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12892911&dopt=Abstract cholesterol




Inhibition of cholesterol absorption associated with a PPAR alpha-dependent increase in ABC binding cassette transporter A1 in mice.

Knight BL, Patel DD, Humphreys SM, Wiggins D, Gibbons GF.

Lipoprotein Group, MRC Clinical Sciences Centre, Faculty of Medicine, Imperial College, Hammersmith Hospital, London W12 ONN, UK. brian.knight csc.mrc.ac.uk

Dietary supplementation with the peroxisome proliferator-activated receptor alpha (PPAR alpha) ligand WY 14,643 gave rise to a 4- to 5-fold increase in the expression of mRNA for the ATP binding cassette transporter A1 (ABCA1) in the intestine of normal mice. There was no effect in the intestine of PPAR alpha-null mice. Consumption of a high-cholesterol diet also increased intestinal ABCA1 expression. The effects of WY 14,643 and the high-cholesterol diet were not additive. WY 14,643 feeding reduced intestinal absorption of cholesterol in the normal mice, irrespective of the dietary cholesterol concentration, and this resulted in lower diet-derived cholesterol and cholesteryl ester concentrations in plasma and liver. At each concentration of dietary cholesterol, there was a similar significant inverse correlation between intestinal ABCA1 mRNA content and the amount of cholesterol absorbed. The fibrate-induced changes in the intestines of the normal mice were accompanied by an increased concentration of the mRNA encoding the sterol-regulatory element binding protein-1c gene (SREBP-1c), a known target gene for the oxysterol receptor liver X receptor alpha (LXR alpha). There was a correlation between intestinal ABCA1 mRNA and SREBP-1c mRNA contents, but not between SREBP-1c mRNA content and cholesterol absorption. These results suggest that PPAR alpha influences cholesterol absorption through modulating ABCA1 activity in the intestine by a mechanism involving LXR alpha.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12897186&dopt=Abstract cholesterol




Serum cholesterol predictive equations in product development.

Pedersen JI, Kirkhus B, Muller H.

Institute for Nutrition Research, University of Oslo, P.O.B. 1046, Blindern, N-0316 Oslo, Norway. j.i.pedersen basalmed.uio.no

The aim of the study was to incorporate trans fatty acids into predictive equations for serum cholesterol and compare their effects with the effects of the individual saturated fatty acids 12:0, 14:0 and 16:0. We have introduced trans fatty acids from partially hydrogenated soybean oil (TransV) and fish oil (TransF) into previously published equations by constrained regression analysis. Prior knowledge about the signs and ordering of existing regression coefficients were incorporated into the regression modelling by adding lower and upper bounds to the coefficients. Oleic acid (18:1) and polyunsaturated fatty acids (18:2, 18:3) were not sufficiently varied in the studies and the respective regression coefficients therefore set equal to those found by Yu et al. (Am J Clin Nutr 1995;61:1129-39). Stearic acid (18:0) considered to be neutral was not included in the equations. The regression analyses were based on results from four controlled dietary studies with a total of 95 participants and including 10 diets differing in fatty acid composition. The analyses resulted in the following equations where the change in cholesterol is expressed in mmol/L and the change in intake of fatty acids is expressed in E%: Delta Total cholesterol = 0.01 delta(12:0) + 0.12 Delta(14:0) + 0.057 delta(16:0) + 0.039 delta(TransF) + 0.031 delta(TransV)- 0.0044 delta(18:1) - 0.017 delta(18:2, 18:3) and deltaLDL cholesterol = 0.01 delta(12:0) + 0.071 delta(14:0) + 0.047 delta(16:0) + 0.043 delta(TransF) + 0.025 delta(TransV) - 0.0044 delta(18:1) - 0.017 delta(18:2, 18:3). The test set used for validation consisted of 22 data points from seven recently published dietary studies. The equation for total cholesterol showed good prediction ability with a correlation coefficient of 0.981 between observed and predicted values. The equation has been used to reformulate margarines into "trans free" products all with more favourable effects on serum cholesterol than previous products. Also a cholesterol reducing margarine has been produced. When tested against butter in an open clinical trial among subjects with mild hypercholesterolemia the observed cholesterol-lowering effect of this margarine corresponded reasonably well with the predicted (0.77 vs. 0.64 mmol/L). We conclude that the equation has practical applicability and can be used to formulate and nutritionally optimise fat products as well as to evaluate already existing products on the market.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12915327&dopt=Abstract cholesterol




Pancreatic triglyceride lipase deficiency minimally affects dietary fat absorption but dramatically decreases dietary cholesterol absorption in mice.

Huggins KW, Camarota LM, Howles PN, Hui DY.

Department of Pathology and Laboratory Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio 45267, USA.

This study generated pancreatic triglyceride lipase (PTL)-null mice to test the hypothesis that PTL-mediated hydrolysis of dietary triglyceride is necessary for efficient dietary cholesterol absorption. The PTL-/- mice grew normally and displayed similar body weight as their PTL+/+ littermates. Plasma lipid levels between animals of various PTL genotypes were similar when they were maintained on either a basal low fat diet or a western-type high fat/high cholesterol diet. Although the lack of a functional PTL delayed fat absorption during the initial hour of feeding a bolus load of olive oil containing [3H]triolein and [14C]cholesterol, the rate of [3H]triolein absorption was similar between PTL+/+ and PTL-/- mice after the initial 1-h period. Importantly, comparison of fecal fat content revealed similar overall fat absorption efficiency between PTL+/+ and PTL-/- mice. In contrast, the PTL-/- mice displayed significant decrease in both the rate and the amount of cholesterol absorbed after a single meal. The plasma appearance of [14C]cholesterol was found to be 75% lower (p < 0.0005) in PTL-/- mice compared with PTL+/+ mice after 4 h. The total amount of [14C]cholesterol excreted in the feces was 45% higher (p < 0.0004) in PTL-/- mice compared with PTL+/+ mice over a 24-h period. These results indicate that the delayed fat digestion due to PTL deficiency results in a significant reduction in cholesterol absorption, although other enzymes in the digestive tract may compensate for the lack of PTL in PTL-/- mice in fat digestion and absorption.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12915407&dopt=Abstract cholesterol




Cyclodextrins differentially mobilize free and esterified cholesterol from primary human foam cell macrophages.

Liu SM, Cogny A, Kockx M, Dean RT, Gaus K, Jessup W, Kritharides L.

Heart Research Institute, Camperdown, Sydney New South Wales, Australia.

Human monocyte-derived foam cell macrophages (HMFCs) are resistant to cholesterol efflux mediated by physiological acceptors. The role of the plasma membrane in regulating depletion of free cholesterol (FC) and of cholesteryl ester (CE) was investigated using cyclodextrins (CDs). HMFCs were incubated in media containing CDs (1.0 mg/ml, approximately 0.7 mM) with low [hydroxypropyl-beta-CD (HP-CD)] or high [trimethyl-beta-CD (TM-CD)] affinity for cholesterol in the presence or absence of phospholipid vesicles (PLVs). Low-affinity HP-CD caused minimal cholesterol efflux on its own, but HP-CD+ PLV depleted cell FC and CE to 54.5 +/- 6.7% of control by 24 h. TM-CD depleted FC at least as well as HP-CD+PLV but without depleting CE, even when combined with PLV. This was not explained by acceptor saturation, instability of PLV vesicles, de novo cholesterol synthesis, kinetically distinct cholesterol pools, or inhibition of CE hydrolysis. TM-CD did, however, deplete CE when lower concentrations of TM-CD were combined with PLV and when acetyl-CoA cholesteryl acyltransferase was inhibited. TM-CD caused much greater depletion of plasma membrane cholesterol than HP-CD without depleting plasma membrane sphingomyelin. It is concluded that differential depletion of plasma membrane cholesterol pools regulates cholesterol efflux and CE clearance in human macrophages.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12671029&dopt=Abstract cholesterol




Plasma markers of cholesterol homeostasis and apolipoprotein B-100 kinetics in the metabolic syndrome.

Chan DC, Watts GF, Barrett PH, O'Neill FH, Thompson GR.

School of Medicine and Pharmacology, University of Western Australia, Perth, Western Australia.

OBJECTIVE: The metabolic syndrome is characterized by defective hepatic apolipoprotein B-100 (apoB) metabolism. Hepato-intestinal cholesterol metabolism may contribute to this abnormality. RESEARCH METHODS AND PROCEDURES: We examined the association of cholesterol absorption and synthesis with the kinetics of apoB in 35 obese subjects with the metabolic syndrome. Plasma ratios of campesterol and lathosterol to cholesterol were used to estimate cholesterol absorption and synthesis, respectively. Very-low-density lipoprotein (VLDL), intermediate-density lipoprotein (IDL), and low-density lipoprotein apoB kinetics were studied using stable isotopy and mass spectrometry. Kinetic parameters were derived using multicompartmental modeling. RESULTS: Compared with controls, the obese subjects had significantly lower plasma ratios of campesterol, but higher plasma ratios of lathosterol (p < 0.05 in both). This was associated with elevated VLDL-apoB secretion rate (p < 0.05) and delayed fractional catabolism of IDL and low-density lipoprotein-apoB (p < 0.01). In the obese group, plasma ratios of campesterol correlated inversely with VLDL-apoB secretion (r = -0.359, p < 0.05), VLDL-apoB (r = -0.513, p < 0.01) and IDL-apoB (r = -0.511, p < 0.01) pool size, and plasma lathosterol ratio (r = -0.366, p < 0.05). Subjects with low cholesterol absorption had significantly higher VLDL-apoB secretion, VLDL-apoB and IDL-apoB pool size, and plasma lathosterol ratio (p < 0.05 in both) than those with high cholesterol absorption. DISCUSSION: Subjects with the metabolic syndrome have oversecretion of VLDL-apoB and decreased catabolism of apoB-containing particles and low absorption and high synthesis rates of cholesterol. These changes in cholesterol homeostasis may contribute to the kinetic defects in apoB metabolism in the metabolic syndrome.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12690090&dopt=Abstract cholesterol




Single repeat deletion in ApoA-I blocks cholesterol esterification and results in rapid catabolism of delta6 and wild-type ApoA-I in transgenic mice.

Sorci-Thomas MG, Thomas M, Curtiss L, Landrum M.

Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157, USA. msthomas wfubmc.edu

The deletion mutation Delta6 apolipoprotein A-I lacks residues 143-164 or repeat 6 in the mature apoA-I protein. In vitro studies show this mutation dramatically reduces the rate of lecithin:cholesterol acyltransferase (LCAT) catalyzed cholesterol esterification. The present study was initiated to investigate the effect of this mutation on in vivo high density lipoprotein (HDL) cholesterol esterification and metabolism. Transgenic mice expressing human Delta6 apoA-I (TgDelta6 +/+) were created and then crossed with apoA-I knockout mice (-/-) to generate mice expressing only human Delta6 apoA-I (TgDelta6 -/-). Human Delta6 apoA-I was associated with homogeneous sized alpha-HDL, when wild-type mouse apoA-I was present (in TgDelta6 +/+ and +/- mice). However, in the absence of endogenous mouse apoA-I, Delta6 apoA-I was found exclusively in cholesterol ester-poor HDL, and lipid-free HDL fractions. This observation coincides with the 6-fold lower cholesterol ester mass in TgDelta6 -/- mouse plasma compared with control. Structural studies show that despite the structural perturbation of a domain extending from repeat 5 to repeat 8 (137-178), Delta6 apoA-I binds to spherical unilamellar vesicles with only 2-fold less binding affinity. In summary, these data show a domain corresponding to apoA-I repeat 6 is responsible for providing an essential conformation for LCAT catalyzed generation of cholesterol esters. Deletion of apoA-I repeat 6 not only blocks normal levels of cholesterol esterification but also exerts a dominant inhibition on the ability of wild-type apoA-I to activate LCAT in vivo.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10766851&dopt=Abstract cholesterol