Normal DNA polymorphism at the low density lipoprotein receptor (LDLR) locus associated with serum cholesterol level

A restriction fragment length polymorphism (RFLP) at the low density lipoprotein receptor (LDLR) locus detectable with the restriction enzyme PvuII exhibits association with total serum cholesterol level. People who are homozygous for absence of the PvuII restriction site have a significantly higher total cholesterol level than heterozygotes (the number of homozygotes for presence of the restriction site was too small to permit meaningful comparison). This difference is significant at the 2% level. Thus, this study of sex- and age-adjusted cholesterol levels in a sample of healthy people yields additional evidence and sustains our previous proposal that normal alleles at the LDLR locus contribute to the population variation in total cholesterol levels. Absence of the PvuII site appears to confer an odds ratio of approximately 2.7 for having a cholesterol level in the top quartile of the population distribution.     

Genetics of the low density lipoprotein receptor:

Fibroblast low density lipoprotein (LDL) plasma membrane receptor activity, measured as 125I-LDL association (plasma membrane binding plus intracellular accumulation) and degradation was determined in cell strains from 14 monozygotic (MZ) and 21 like-sexed dizygotic (DZ) normolipidemic twin pairs. The twins were between 57 and 62 years old and had liver apart for an average of 38 years (range 0-60). The intrapair differences were significantly smaller in MZ than in DZ twin pairs in fibroblast 125I-LDL association as well as degradation assays (P less than 0.05). These findings suggest a genetic influence on normal variation in LDL receptor activity in vitro. In two MZ pairs discordant for psoriasis, the psoriatic twin had markedly lower LDL receptor activity than the cotwin.     

Associations of usual sleep duration with serum lipid and lipoprotein levels

STUDY OBJECTIVES: We examined the individual association between sleep duration and a high serum triglyceride, low HDL cholesterol, or high LDL cholesterol level. DESIGN AND SETTING: The present study analyzed data from the National Health and Nutrition Survey that was conducted in November 2003 by the Japanese Ministry of Health, Labour and Welfare. This survey was conducted on residents in the districts selected randomly from all over Japan. PARTICIPANTS: The subjects included in the statistical analysis were 1,666 men and 2,329 women aged 20 years or older. INTERVENTION: N/A. MEASUREMENTS AND RESULTS: Among women, both short and long sleep durations are associated with a high serum triglyceride level or a low HDL cholesterol level. Compared with women sleeping 6 to 7 h, the relative risk of a high triglyceride level among women sleeping <5 h was 1.51 (95% CI, 0.96-2.35), and among women sleeping > or =8 h was 1.45 (95% CI, 1.00-2.11); the relative risk of a low HDL cholesterol level among women sleeping <5 h was 5.85 (95% CI, 2.29-14.94), and among women sleeping > or =8 h was 4.27 (95% CI, 1.88-9.72). On the other hand, it was observed that the risk of a high LDL cholesterol level was lower among men sleeping > or =8 h. These analyses were adjusted for the following items: age, blood pressure, body mass index, plasma glucose level, smoking habit, alcohol consumption, dietary habits, psychological stress, and taking cholesterol-lowering medications. CONCLUSIONS: Usual sleep duration is closely associated with serum lipid and lipoprotein levels.     

Genetics of the low density lipoprotein receptor:

Fibroblast association (plasma membrane binding plus intracellular accumulation) and degradation of radioiodinated low density lipoprotein (125I-LDL) index plasma membrane LDL receptor activity. Cultured fibroblasts from 23 subjects affected with familial hypercholesterolemia (HC) and from 95 subjects without HC (non-HCs) were tested for 125I-LDL association and degradation. Both LDL receptor activity indices were twice as high in non-HC and HC heterozygous cell strains. This is compatible with a major gene effect on LDL receptor activity. However, a considerable overlap between non-HC and HC heterozygous values was found in the 125I-LDL association assay [median (range) 970 (330-2500), and 450 (250-490), respectively] and in the degradation assay [median (range) 810 (280-2020), and 470 (160-790), respectively]. The values are expressed as ng 125I-LDL X mg cell protein-1 X 4.5 h-1. These great overlaps in the LDL receptor activity indices support the view that the influence of LDL receptor activity on the HC phenotype may be smaller than believed previously. Furthermore, for the diagnosis of HC, these LDL receptor activity assays are far more expensive and have less sensitivity and specificity than simple serum cholesterol determination. The LDL receptor-dependent 125I-LDL association values for the HC heterozygous individuals clustered into four groups. Family data supported the hypothesis that this variation could be due to four different LDL receptor variants, each coded for by different alleles at the LDL receptor locus. If confirmed, this finding may have implications for the understanding of the variable expression of HC and also of the genetic impact on lipoprotein metabolism and susceptibility to atherosclerosis in non-HCs.     

Lp(a) lipoprotein enters cultured fibroblasts independently of the plasma membrane low density lipoprotein receptor

Lp(a) lipoprotein shares the apoB antigen with low density lipoprotein (LDL). The Lp(a) antigen is unique for Lp(a) lipoprotein. Fibroblast association (i.e. plasma membrane binding plus intracellular accumulation), plasma membrane binding, intracellular accumulation and degradation of 125I-Lp(a) lipoprotein were studied in strains from subjects with or without autosomal dominant hypercholesterolemia (HC). Subjects without HC (non-HCs) have cell surface receptors for low density lipoprotein (LDL receptors). On the average, HC heterozygotes have half-normal LDL receptor activity and "receptor-negative" HC homozygous cell strains lack functional receptors. Fibroblast processing of 125I-Lp(a) lipoprotein was compared to fibroblast processing of 125I-LDL. LDL receptor-dependent processing of 125I-LDL was saturated at about 50 microgram apo 125I-LDL.ml-1 in non-HC fibroblasts. 125I-Lp(a) lipoprotein was, however, largely processed independently of receptor mechanisms by non-HC cells (highest concentration examined 150 microgram apo 125I-Lp(a) lipoprotein . ml-1). Lp(a) lipoprotein did not interfere with 125I-LDL for fibroblast association, but inhibited 125I-LDL degradation. The interference with 125I-LDL degradation was time dependent. Only slightly higher 125I-Lp(a) lipoprotein processing values were found in non-HC and HC heterozygous strains than in "receptor-negative" HC homozygous strains. However, non-HC cells had more than tenfold higher 125I-LDL processing values than "receptor-negative" HC homozygous cells.     

内源性高甘油三酯血症患者血浆VLDL、LDL及HDL对血凝的影响

目的：研究内源性高甘油三酯血症(HTG)患血浆极低密度脂蛋白(VLDL)、低密度脂蛋白(LDL)及高密度脂蛋白(HDL)是否发生了氧化修饰及其对血凝的影响。方法：对2l例内源性高甘油三酯血症患与2l例年龄性别相近的正常人的血脂、脂质过氧化物进行了分析。用一次性密度梯度超速离心法分离血浆VLDL、LDL及HDL，测定这三种脂蛋白的234nm光吸收、相对电泳迁移率(REM)和硫代巴比妥酸反应物质(TBARS)，分别将这三种脂蛋白加入由正常人新鲜混合血浆构成的反应系统中，按试剂盒分别测定凝血酶原时间(PT)及活化部分凝血酶原时间(APIT)。结果：内源性HTG患血浆TG含量平均升高2．73倍，HDLC下降l.7l倍，同时LPO升高1．22倍;HTG组VLDL、LDL及HDL的REM、234nm光吸收值、TBARS含量均较对照组显增加(P＜0．01)，表明内源性HTG患血浆VLDL、LDL及LDL均发生了氧化修饰生成Ox—VLDL、Ox-LDL.PT及APTT在分别加入HTG组的VLDL、LDL及HDL后均比加入相应正常组脂蛋白明显缩短(P均＜0．05)。相关分析表明，HTG组血浆VLDL及HDL相对电泳迁移率(REM)与PT呈负相关(P＜0．01)。结论：HTG患血浆VLDL、LDL及HDL发生了氧化修饰，并使PT及APTT明显缩短。     

Gene-gene interaction between the low density lipoprotein receptor and apolipoprotein E loci affects lipid levels

A restriction fragment length polymorphism (RFLP) at the low density lipoprotein receptor (LDLR) locus, detectable with the restriction enzyme PvuII has been studied in a second series of Norwegian subjects, believed to be representative of the general population. The results confirm the association of normal alleles at the LDLR locus with differences in age- and sex-corrected total and LDL cholesterol. A gene identified as one of the alleles in this RFLP appears to eliminate the effect of the apolipoprotein E4 (apoE4) gene on cholesterol (or its allele must be present for the apoE4 effect on lipid level to manifest itself). The findings in this series substantiate previous indications that normal alleles are of importance in the control of LDLR activity and that normal LDLR alleles contribute to the population variation in cholesterol. Finally, they confirm that an interaction between LDLR and apoE genes contributes to the population variation in total and LDL cholesterol.     

Normal genetic variation at the low density lipoprotein receptor (LDLR) locus influences cholesterol levels in children

The population of Czechoslovakia is at high risk of premature atherosclerosis. Normal DNA polymorphism at the low density lipoprotein receptor (LDLR) locus detectable with the restriction enzyme PvuII was analyzed in Czech children with a high or a low concentration of total serum cholesterol. The PvuII restriction site was found significantly more often in the low cholesterol group than in the high cholesterol group. Thus, normal genetic variation at the LDLR locus contributes to the population variation in cholesterol in children in the population studied.     

家族性高胆固醇血症家系低密度脂蛋白受体基因剪接突变的研究

目的　检测中国汉族家族性高胆固醇血症 (familial hypercholesterolemia,FH)大家系低密度脂蛋白受体 (low density lipoprotein receptor,L DL R)基因突变 ,探讨 FH发病的分子机理。方法　首先采用聚合酶链反应 -限制性片段长度多态性 (polymerase chain reaction- restriction fragment lengthpolymorphism,PCR- RFL P)技术检测载脂蛋白 B1 0 0 (apo B1 0 0 )基因 Q35 0 0 R突变 ,排除家族性 apo B1 0 0 缺陷症 ,再采用 PCR扩增结合核苷酸序列分析检测 1例临床诊断为 FH纯合子患儿及其家系成员 L DL R基因启动子和全部 18个外显子片段 ,结果与 Gen Bank公布的该基因正常序列对比找出突变 ,并在家系其他成员中证实该突变。结果　该患儿 L DL R基因第 3内含子剪接供体处存在 IN 5′GT→AT纯合剪接突变 ,并且在家系中得到证实 ,一级和二级亲属中各发现 2例相同位点和相同形式的杂合子 ,其基因型表现为野生型和突变型杂合现象。同时未检测出患儿及其父母 apo B1 0 0 Q35 0 0 R突变。结论　发现 L DL R基因第 3内含子 G→ A纯合剪接突变 ,可能是该 FH家系发病的分子基础 ;检测该突变对临床干预和遗传指导有参考价值。     

Low density lipoprotein receptor activity in cultured skin fibroblasts from octa- and nonagenarians

Low density lipoprotein (LDL) receptor activity determined as association or degradation of 125I-LDL at 37 degrees C, was measured in cultured skin fibroblasts from 48 elderly, independent subjects (84-95 years old). Data concerning these subjects were compared with those from a reference group of 27 younger, healthy subjects (5-64 years old) and 23 heterozygotes for familial hypercholesterolemia (9-67 years old). The median (10th-90th percentile) association and degradation value for the older subjects were 294 (172-535) ng 125I-LDL/mg protein/6h and 191 (99-376) ng 125I-LDL/mg protein/6h, respectively. These values did not differ significantly from the corresponding values in the normal subjects (296 (60-491) ng 125I-LDL/mg protein/6h and 171 (99-275) ng 125I-LDL/mg protein/6h, respectively). Thus, maximal LDL receptor activity in fibroblasts from the older subjects seems to be comparable to that of younger, healthy controls. However, the elderly subjects had markedly higher values for total serum cholesterol and LDL cholesterol than the normal controls. There was a significant increase in LDL receptor activity with age in the normal controls. Such an increase was not found in heterozygotes for familial hypercholesterolemia.     

Effect of lipid lowering diet on low density lipoprotein receptor activity in freshly isolated peripheral blood mononuclear cells

The effect of lipid lowering diet on low density lipoprotein (LDL) receptor activity has been studied in freshly isolated peripheral blood mononuclear cells (PBMCs) from 16 hypercholesterolemic male subjects during a three weeks' dietary intervention trial. The participants were randomized to an intervention group or to a control group. The subjects in the intervention group had a non-significantly larger increase in LDL receptor activity, determined as degradation of 125I-LDL at 37 degrees C, than the control group (49.7 +/- 18.1 and 35.4 +/- 21.2 ng/mg, respectively). Irrespective of assignment to the intervention group or control group, the eight subjects with the largest reduction in total serum cholesterol had significantly larger absolute and percentage increases in LDL receptor activity than the eight subjects with the smallest reduction in total serum cholesterol (p less than 0.05). Thus, it appears that the cholesterol reduction that can be achieved in humans by dietary intervention results in a small increase in LDL receptor activity in freshly isolated PBMCs.     

The effect of vigorous physical activity at work on serum lipids with a special reference to serum high-density lipoprotein cholesterol

Serum lipid concentrations of lumberjacks whose occupational physical activity is most vigorous were compared with those of electricians. The lumberjacks had significantly higher serum HDL-cholesterol and significantly lower triglyceride and free fatty acid concentrations hut there were no significant differences in total cholesterol levels. The favourable effect of vigorous physical activity at work on lipid metabolism is, however, epidemiologically not seen, obviously due to negative risk factors in lumberjacks' life.     

Interaction between low density lipoprotein receptor (LDLR) and apolipoprotein E (apoE) alleles contributes to normal variation in lipid level

Subjects drawn from a population-based register were studied with respect to lipid level association. The association of isoforms of apolipoprotein E (apoE) with lipid level in the general population was found to be limited to people with one particular genotype at the low density lipoprotein receptor (LDLR) locus. The results presented in this paper suggest that functional LDLR variants enhance or limit the effect of isoforms of apoE. The association between apoE4 and serum total and LDL cholesterol level may be mediated through the LDL (apoB100, apoE) receptor to a greater extent than previously thought.     

Characterization of low density lipoprotein receptor (LDLR) gene mutations in Albania

Introduction

Familial hypercholesterolaemia (FH) is a clinical syndrome characterised by elevated serum total cholesterol (TCHOL) levels due to an increase in low-density lipoprotein (LDL) cholesterol, by tendon xanthomata and clinical manifestations of ischaemic heart disease in early life. Typically, it results from mutations in the low-density lipoprotein receptor (LDLR) gene. So far, more than 800 mutations have been reported for the LDLR gene and account for FH. The nature of LDLR gene mutations varies among different ethnicities. Until now no mutations of LDLR have been reported in the Albanian population.

Material and methods

We assessed the contribution of the LDLR gene mutations as causes of FH in an Albanian population. Fifty probands with a clinical diagnosis of FH were included. We analysed all the exons and the promoter of the LDLR gene by using restriction isotyping or direct sequencing.

Results

Twenty-one patients were heterozygous for the 1646G>A mutation (FH Genoa) in exon 11 and 9 patients were heterozygous for the 81T>C mutation in exon 2 of the LDLR gene.

Conclusions

This report describes two LDLR gene mutations accounting for FH in Albania (1646G>A, 81T>C).     

Reduced plasma concentrations of total, low density lipoprotein and high density lipoprotein cholesterol in patients with Gaucher type I disease

Plasma lipid and serum apoprotein concentrations were determined in twenty-nine individuals with Gaucher type I disease. Plasma total cholesterol, low density lipoprotein (LDL) cholesterol and high density lipoprotein (HDL) cholesterol were all significantly reduced in the patients with Gaucher disease compared to a group of matched control subjects. Total, LDL and HDL cholesterol were lower in males than in females with Gaucher disease. These sex differences appeared to be inversely correlated with the severity of disease manifestations which were greater in the males. Serum levels of apoprotein-B and apoprotein-AI, the major structural apoproteins of LDL and HDL, respectively, were decreased in the subjects with Gaucher disease. Thus, the reductions in LDL and HDL cholesterol were associated with reduced numbers of lipoprotein particles in plasma. In contrast, apoprotein-E, a protein which is secreted by several tissues, including activated macrophages and which may mediate hepatic catabolism of lipoproteins, was elevated in the patients. Since macrophages may also catabolize lipoproteins, Gaucher disease may serve as a model for the effect of activated macrophages upon human lipoprotein metabolism.
This work was supported by the following grants: USPHS HL 23077, HD 07105, HL 25752, CA 31656 and RR-71 from the National Institutes of Health; 1–273 and 1–578 from the March of Dimes Birth Defects Foundation; grant from the New York Heart Assocation. Dr. Ginsberg is a recipient of a Research Career Development Award HL 00949 and is an Irma T. Hirschl Career Scientist. Dr. Grabowski is a recipient of a Clinical Investigator Award HD 00386.     

低密度脂蛋白受体相关蛋白5基因的基因组结构

目的 确定2低密度脂蛋白受体相关蛋白5（low density lipoprotein receptor related protein,5 LRP5）的基因组结构。方法 以LRP5基因的cDNA序列为线索，采用计算机杂交方法首先通过对比分析该基因的cDNA序列和基因组序列，初步确定LRP5基因的基因组结构，按分析得到的基因组结构设计引物，扩增并测定外显子序列和外显子内含子接头序列，确定该基因的基因组结构。结果 LRP5基因的基因组DNA全长为131.6kb，含4有23个外显子和22个内含子；在编码序列中检测到3个单核苷酸多态，即位于第2外显子的A459G，位于第10外显子的C2220T和位于第21外显子的G4416C；LRP5基因含有4个已知的短串联重复序列，即D11S1917，D11S4087，D11S1337和D11S4178，它们分别位于该基因的5'端和第1，4，13内含子内。结论 LRP5基因的基因组结构的确定，为分析该基因突变和功能研究奠定了基础。     

A 9.6 kilobase deletion in the low density lipoprotein receptor gene in Norwegian familial hypercholesterolemia subjects

Haplotype analysis of the low density lipoprotein receptor (LDLR) gene was performed in Norwegian subjects heterozygous for familial hypercholesterolemia (FH). Southern blot analysis of genomic DNA, using an exon 18 specific probe and the restriction enzyme NcoI, showed that two out of 57 unrelated FH subjects had an abnormal 3.6 kb band. Further analyses revealed that this abnormal band was due to a 9.6 kb deletion that included exons 16 and 17. The 5' deletion breakpoint was after 245 bp of intron 15, and the 3' deletion breakpoint was in exon 18 after nucleotide 3390 of cDNA. Thus, both the membrane-spanning and cytoplasmatic domains of the receptor had been deleted. A polymerase chain reaction (PCR) method was developed to identify this deletion among other Norwegian FH subjects. As a result of this screening one additional subject was found out of 124 subjects screened. Thus, three out of 181 (1.7%) unrelated Norwegian FH subject possessed this deletion. The deletion was found on the same haplotype in the three unrelated subjects, suggesting a common mutagenic event. The deletion is identical to a deletion (FH-Helsinki) that is very common among Finnish FH subjects. However, it is not yet known whether the mutations evolved separately in the two countries.     

动脉粥样硬化性脑梗塞与低密度脂蛋白受体基因NcoI、AvaⅡ多态性的关系

目的 探讨动脉粥样硬化性脑梗死（atherosclerotic cerebral infarction,ACI)与低密度脂蛋白受体（low density lipoprotein receptor,LDL-R)基因NcoI、AvaⅡ多态性的关系。方法 用聚合酶链反应技术检测113名辽宁藉汉族健康人和77例ACI患者的LD－R基因NcoI、AvaⅡ多态性及血脂、载脂蛋白的含量。结果 LDL－R基因NcoI、AvaⅡ等位基因频率健康人N^ 为0．667、A^ 为0．230；ACI组N^ 为0．662、A^ 为0．125。A^-A^-与N^ N^ 联合存在时ACI的发病相对风险率（RR）为5．56（P＜0．001），引起血清TG、TC、LDL－C、LP（a)升高的相对风险率依次为4．29、7．67、9．33、3．09（P＜0．05）。结论 LDL－R基因A^-A^-与N^ N^ 联合存在影响血脂、脂蛋白的含量，与ACI密切相关。