Expression of matrix metalloproteinase-9 in mononuclear cells of hyperhomocysteinaemic subjects.

Atherosclerotic plaque instability and rupture requires extracellular matrix modification, a complex process regulated by matrix metalloproteinases (MMPs). We hypothesized that homocysteine's atherogenic effects may involve MMP-mediated mechanisms. Our results showed the following: (i) Compared with healthy control subjects (n = 9), patients with hyperhomocysteinaemia (n = 9) had elevated mRNA levels of MMP-9 and tissue inhibitors of metalloproteinases-1 (TIMP-1) in freshly isolated peripheral blood mononuclear cells (PBMCs), which were positively correlated with homocysteine and negatively correlated with folate and vitamin B12 levels. (ii) Peripheral blood mononuclear cells obtained from these patients released markedly enhanced the amount of MMP-9 upon oxidized LDL (oxLDL) stimulation, whereas no such enhancing effect was seen in cells from healthy controls. (iii) During folic acid 6 weeks' treatment, normalization of homocysteine levels was accompanied by a significant reduction in mRNA levels of MMP-9 and TIMP-1 in PBMCs, as well as a marked reduction in oxLDL-stimulated release of MMP enzyme activity. These novel findings may suggest that homocysteine exerts its atherogenic effect in part by elevating levels and activity of MMPs, which in turn may enhance matrix degradation, potentially promoting atherogenesis and plaque instability. Moreover, our findings suggest that folic acid supplementation may down-regulate these inappropriate responses in these patients.

Effect of folic acid treatment on endothelium-dependent vasodilation and nitric oxide-derived end products in hyperhomocysteinemic subjects.

PURPOSE: An elevated plasma homocysteine concentration is an independent risk factor for cardiovascular diseases. In this study, we tested the hypothesis that hyperhomocysteinemia induces endothelial dysfunction mediated, at least in part, through nitric oxide-dependent mechanisms and that folic acid supplementation improves endothelial function in hyperhomocysteinemic subjects. SUBJECTS AND METHODS: Endothelial function was evaluated in healthy controls and hyperhomocysteinemic subjects by measuring plasma levels of the nitric oxide-derived end products nitrite and nitrate and by assessing vasodilatory responses in the skin microcirculation and forearm vasculature. In the subjects with hyperhomocysteinemia, these measurements were repeated after 6 weeks and 12 months of folic acid supplementation. RESULTS: Compared with healthy controls, hyperhomocysteinemic subjects had significantly lower median plasma levels of nitric oxide-derived end products (12.1 microM [range 4.4 to 41.8] versus 24.6 microM [13.6 to 53.2]; P <0.001), a significantly lower endothelium-dependent vasodilatory response to acetylcholine (P <0.01), hyperemic response in the microcirculation (P <0.01), and total forearm blood flow during reactive hyperemia (P = 0.01). There was no significant difference in the endothelium-independent response. Folic acid treatment for 12 months increased the plasma level of nitric oxide-derived end products by 121% (95% confidence interval [CI], 72% to 170%), the vasodilatory response to acetylcholine by 124% (95% CI, 36% to 212%), and the ischemia-mediated hyperemic responses in the microcirculation by 60% (95% CI, 25% to 96%) and in the forearm vasculature by 47% (95% CI, 21% to 73%). CONCLUSIONS: Homocysteine appears to induce its atherogenic effect, at least in part, by depressing endothelial function, possibly through nitric oxide-dependent mechanisms. This effect can be reversed by folic acid supplementation.

Serum cholesterol response to replacing butter with a new trans-free margarine in hypercholesterolemic subjects.

UNLABELLED: Margarine leads to lower total and LDL cholesterol (LDL-C) levels than butter but may contain trans fatty acids that increase atherogenic lipids. A food company has used data concerning the cholesterolemic effects of individual fatty acids, including trans fatty acids, to develop a commercially available and virtually trans-free margarine. OBJECTIVE: The effect of this novel margarine on serum lipids and lipoproteins was compared with that of butter in free-living, hypercholesterolemic subjects. DESIGN AND SETTING: A two-period, outpatient cross-over trial at a university hospital lipid clinic. SUBJECTS: The study involved 77 subjects, and was completed by 53 men and 19 women aged 35-65 years with total serum cholesterol levels of between 6.0 and 7.9 mmol/L. INTERVENTION: Two 23-day regimens, separated by a 4-week washout period, included individualised dietary prescriptions supplemented with butter or margarine designed to provide 15% of total dietary energy. RESULTS: In comparison with butter, margarine intake lowered total and LDL-C levels by respectively 11.1% (99% CI: 8.1-14.1) and 11.3% (99% CI: 7.6-15.1). The reduction in LDL-C was < 3% in nearly one-fifth of the subjects despite appropriate changes in serum triglyceride fatty acids. Of the tested clinical and demographic variables, only the percentage of energy obtained from saturated fat during the margarine intake period was associated with dietary responsiveness (explaining 12% of the variation; p < 0.01). CONCLUSION: Our results suggest that a margarine designed to meet nutritional recommendations for hypercholesterolemia is more efficacious than butter in reducing atherogenic lipid levels in hypercholesterolemic subjects.

Effect of Norwegian fish powder on risk factors for coronary heart disease among hypercholesterolemic individuals.

BACKGROUND AND AIM: Numerous studies suggest an association between high intake of fatty fish and reduced risk of coronary heart disease. Very long-chain omega-3 fatty acids are thought to be responsible for the benefits observed, though other fatty fish components may act in concert with them. Norwegian fish powder is a dry herring product that contains essential amino acids, marine omega-3 fatty acids, vitamins and minerals. The aim of the present study was to determine whether it has beneficial effects on risk factors for coronary heart disease in man. METHODS AND RESULTS: A single center, randomized, double-blind, parallel-treatment study was carried out for 12 weeks. Subjects with primary hypercholesterolemia were randomly allocated to 10 g fish powder or placebo (20 tablets/day). Participants were instructed to follow National Cholesterol Education Program (NCEP) Step I Diet during a 4-week diet run-in phase and during the study. Concentrations of lipids, lipoproteins, hemostatic variables and endothelial cell markers were determined before and after supplementation. Our data showed that the fish powder supplement was well tolerated. A significant decrease and increase respectively were observed in plasma alpha-linolenic acid (p = 0.03) and docosahexaenoic acid (DHA) (p = 0.03). Concentrations of lipids, lipoproteins, homocysteine, factor VII, fibrinogen, tissue plasminogen activator (tPA), plasminogen activator inhibitor (PAI)-1, soluble intercellular adhesion molecule (ICAM)-1, P-selectin and interleukin (IL)-8 were not beneficially affected. CONCLUSIONS: Fish powder supplementation does not seem an effective approach to improve risk factors for coronary heart disease in hypercholesterolemic subjects following the NCEP Step I Diet.

Effects of high doses of simvastatin and atorvastatin on high-density lipoprotein cholesterol and apolipoprotein A-I.

A randomized, blinded, multicenter clinical trial was performed comparing low- and high-dose simvastatin (40 and 80 mg) with comparable doses of atorvastatin (20 and 40 mg) for effects on plasma concentrations of lipoproteins and apolipoprotein A-I over 12 weeks in 842 patients with elevated low-density lipoprotein cholesterol. The 2 agents reduced low-density lipoprotein cholesterol and triglycerides to a comparable degree, but simvastatin raised high-density lipoprotein cholesterol and apolipoprotein A-I more than atorvastatin, suggesting differences in metabolic effects of the 2 agents on plasma lipids and lipoproteins.

Phenotypic consequences of a deletion of exons 2 and 3 of the LDL receptor gene.

Screening for structural alterations of the low density lipoprotein (LDL) receptor gene by Southern blot analysis revealed an abnormal band pattern in one subject with a clinical diagnosis of homozygous familial hypercholesterolemia (FH). The molecular defect was further characterized by polymerase chain reaction and cDNA sequencing. These analyses identified a 4.8 kb in-frame deletion of exons 2 and 3, where exon 1 was spliced to exon 4. This deletion is expected to produce a receptor that has lost the two first cysteine-rich repeats of the ligand-binding domain. Previously published data of in vitro site-directed mutagenesis has shown that binding of LDL to such a receptor is reduced to 70% of normal. A mild phenotype in our FH homozygote is consistent with that observation. In contrast, heterozygotes carrying this deletion have a relatively more severe phenotype that is comparable to that of heterozygotes carrying a null-allele. A severe phenotype was also found in a compound heterozygote carrying this deletion. Possible mechanisms for this phenotypic variability are discussed.-Rødningen, O. K., S. Tonstad, J. D. Medh, D. A. Chappell, L. Ose, and T. P. Leren. Phenotypic consequences of a deletion of exons 2 and 3 of the LDL receptor gene.

The C677T mutation in the methylenetetrahydrofolate reductase gene predisposes to hyperhomocysteinemia in children with familial hypercholesterolemia treated with cholestyramine.

In children with familial hypercholesterolemia, heterozygosity and homozygosity for the C677T mutation in the methylenetetrahydrofolate reductase gene was associated with low serum folate and increased susceptibility to elevation of plasma total homocysteine during cholestyramine treatment. Because of the independent relationship between elevated plasma total homocysteine and cardiovascular disease, folate supplementation may be prudent in these children.

[Diagnosis and treatment of severe hyperlipidemia]. / Diagnose og behandling av svaer hyperlipidemi.

Though severe hyperlipidaemia (total cholesterol level > or = 13 mmol/l in this study) is uncommon, it is important to make a precise diagnosis. We examined 57 patients with isolated severe hypercholesterolaemia. Of these, four were homozygotes for familial hypercholesterolaemia, 48 were heterozygotes for familial hypercholesterolacmia and one had sitosterolemia. The heterozygotes carried 15 different LDL receptor mutations, with no one mutation predominating. When the diagnosis is made, relatives should be given the opportunity to be tested. Combined severe hyperlipidaemia is usually due to a secondary cause, at our clinic, the most common cause is diabetes mellitus. The underlying disease should be treated first. However, many patients will require additional lipid-lowering drugs because the underlying disease may be associated with an increased risk of cardiovascular disease. With the exception of fish oil capsules, drugs that reduce serum triglyceride levels substantially are not registered in Norway at present.

Effect of hormone replacement therapy on the susceptibility of low-density lipoprotein to oxidation among postmenopausal hypercholesterolaemic women.

The effect of sequential combined hormone replacement therapy on the susceptibility of low-density lipoprotein to oxidative modification was investigated in a double-blind, randomized placebo-controlled study. Hypercholesterolaemic, postmenopausal women were supplemented with 17 beta-oestradiol and norethisterone acetate (n = 13 subjects) or placebo capsules (n = 15 subjects) for 12 weeks. They were instructed to follow the American Heart Association step one diet. Low-density lipoprotein, isolated before and after treatment, was subjected to copper-catalysed lipid peroxidation. There were no significant differences between low-density lipoprotein from the hormone replacement therapy and placebo groups, as assessed by measuring the lag time for formation of conjugated dienes, the rate of formation and the amount of conjugated dienes formed, the amount of lipid peroxides generated, and the relative electrophoretic mobility at baseline and after treatment. Dietary records showed that the subjects were consuming similar amounts of fat and vitamins. No major differences were found in the fatty acid pattern of low-density lipoprotein from the two groups. In conclusion, the results indicated that hormone replacement therapy with 17 beta-oestradiol sequentially combined with norethisterone acetate in non-smoking, hypercholesterolaemic, postmenopausal women has no protective effect on the susceptibility of low-density lipoprotein to copper-catalysed modification in vitro.

Results of intensive long-term treatment of familial hypercholesterolemia.

Fifty-seven patients with familial hypercholesterolemia (FH) with mean age of 48 years (range 30 to 69), participated in a follow-up examination 5.5 years after the completion of a 1-year trial with lovastatin, cholestyramine, probucol, or omega-3 fatty acids. The goals were to record quality of life, compliance to treatment, adverse effects, and clinical outcome. The quality of life was similar to that in a Norwegian reference population. The factors causing most distress to patients were keeping a diet low in saturated fats, taking medication, and fear of death. The medication was mostly prescribed in maximum dosages. At follow-up, the reduction in total cholesterol was 36% (p < 0.05), low-density lipoprotein (LDL) cholesterol 38% (p < 0.05), triglycerides 20% (p < 0.05) compared with being on diet therapy only. High-density lipoprotein (HDL) cholesterol increased 8% (p < 0.05). Intake of saturated and monounsaturated fat increased 1.5% and 1.7% (p < 0.05), respectively; polyunsaturated fat was unchanged. Three patients experienced myocardial infarction, of whom 2 died and 1 developed angina pectoris. Before the start of lovastatin treatment, 27 coronary events occurred per 1,000 patient-years in this group compared with 12 events per 1,000 patient-years thereafter. Of 28 patients reporting adverse events, 4 discontinued lovastatin and 3 discontinued cholestyramine. Several practical and psychological difficulties were associated with FH. Long-term intensive lipid-lowering therapy was possible in FH outpatients without loss of effect and with good compliance to therapy. Intensive therapy, today is, however, not sufficient for many FH patients to reach a therapeutic goal of LDL cholesterol < 4.0 mmol/L. More potent lipid-lowering agents are needed.

Efficacy and safety of cholestyramine therapy in peripubertal and prepubertal children with familial hypercholesterolemia.

OBJECTIVE: To determine the efficacy and safety of cholestyramine therapy in young children with familial hypercholesterolemia. SUBJECTS: Boys aged 6 to 11 years (n = 57) and girls aged 6 to 10 years (n = 39) with familial hypercholesterolemia. DESIGN: After 1 year of a low-fat, low-cholesterol diet, children with low-density lipoprotein (LDL) cholesterol levels > or = 4.9 mmol/L (190 mg/di) or < or = 4.1 mmol/L (160 mg/dl) in the presence of familial premature cardiovascular disease were randomly assigned to a double-blind comparison of 8 gm cholestyramine (n = 36) and placebo (n = 36) for 1 year. OUTCOME MEASURES: The primary efficacy and safety outcomes were serum LDL cholesterol levels and height velocity, respectively. Secondary safety outcomes were erythrocyte folate, total plasma homocysteine, serum fat-soluble vitamins, and side effects. RESULTS: Twenty-two subjects in the cholestyramine group and 26 in the placebo group completed the 1-year study. Most withdrawals from the study were related to unpalatability of the study drug or placebo. The LDL cholesterol levels changed by -16.9% (95% confidence interval, -10.8% to -22.9%) in the cholestyramine group compared with 1.4% (95% confidence interval, -4.4% to 7.2%) in the placebo group. Mean height velocity standard deviation scores during 1 year for the children in the cholestyramine and the placebo groups who had not started puberty were 0.24 +/- 1.14 and 0.11 +/- 0.68, respectively (not significant). In the cholestyramine group, mean levels of 25-hydroxyvitamin D decreased. One girl had low folate and elevated homocysteine levels, and there was one case of intestinal obstruction caused by adhesions. CONCLUSIONS: Significant reductions in LDL cholesterol are achievable during treatment with cholestyramine in about half of eligible children. Growth is not adversely affected. Folate deficiency may occur, even with a low dose of cholestyramine, and vitamin D supplements should be considered. Caution should possibly be exercised in starting cholestyramine therapy within 3 months of abdominal surgery in children.

Low dose colestipol in adolescents with familial hypercholesterolaemia.

The effects of orange flavoured colestipol granules, 10 g/day, in 37 boys and 29 girls aged 10-16 years with familial hypercholesterolaemia were examined first in an eight week double blind, placebo controlled protocol, then in open treatment for 44-52 weeks. All patients were on a low fat diet. Low density lipoprotein cholesterol levels were reduced by 19.5% by colestipol v 1.0% by placebo. Levels of serum folate, vitamin E, and carotenoids were reduced in the colestipol group, but not the vitamin E/cholesterol and carotenoid/cholesterol ratios or serum concentrations of vitamins A and D. After one year of colestipol, two thirds of the participants remained in the study, of whom half took > or = 80% of the prescribed dose. Those who took > or = 80% of the dose had a greater decrease in serum 25-hydroxyvitamin D levels than those who took < 80%. No adverse effects on weight gain or linear growth velocity were observed. Although low dose colestipol effectively reduces low density lipoprotein cholesterol levels, only a minority of adolescents adhered to the new formulation for one year. Folate and possibly vitamin D supplementation is recommended.

[Familial hypercholesterolemia in children. A management program. Procedures for diagnosis and treatment in our pediatric departments]. / Familiaer hyperkolesterolemi hos barn. Handlingsprogram prosedyrer til diagnostisering og behandling ved våre barneavdelinger. Gruppen som har utarbeidet handlingsprogrammet er nedsatt av Norsk barnelegeforening, 1995.

A Norwegian programme for treatment and selective screening of familial hypercholesterolaemia has been developed which takes into account family history and levels of hypercholesterolaemia. The programme includes recommendations on when and whom to screen for familial hypercholesterolaemia. With regard to treatment, special emphasis is placed on diet. The working group suggests that small lipid clinics with a dietitian should be established in some paediatric departments.

No effect of beta-carotene supplementation on the susceptibility of low density lipoprotein to in vitro oxidation among hypercholesterolaemic, postmenopausal women.

The effect of beta-carotene on the susceptibility of low density lipoprotein (LDL) to oxidative modification was investigated in a double-blind, randomized placebo-controlled study. Hypercholesterolaemic, postmenopausal women were given 30 mg beta-carotene per day (n = 15 subjects) or placebo capsules (n = 15 subjects) for 10 weeks. They were instructed to follow the American Heart Association Step One diet. LDL, isolated before and after treatment was subjected to copper-catalysed lipid peroxidation. There were no significant differences between LDL from the beta-carotene and placebo groups, as assessed by measuring the lag time for formation of conjugated dienes; the rate of formation and the amount of conjugated dienes formed; the amount of lipid peroxides generated; and the relative electrophoretic mobility, at baseline and after treatment. Dietary records showed that the subjects were consuming similar amounts and types of fat. No significant differences were found in the lipid composition and fatty acid pattern of LDL from the two groups. In conclusion, the results indicated that supplementation with beta-carotene in non-smoking, hypercholesterolaemic, postmenopausal women had no protective effect on the susceptibility of LDL to copper-catalysed modification in vitro.

[Diet therapy of hypercholesterolemia. Can the diet of persons who already follow a cholesterol-lowering diet be further improved by guidelines given by a clinical nutritionist?]. / Kostbehandling ved hyperkolesterolemi. Kan kostholdet til personer som allerede følger et kolesterolsenkende kosthold, forbedres ytterligere etter veiledning av klinisk ernaeringsfysiolog?

The objective of the study was to investigate whether expert advice from a clinical nutritionist improves the diet of patients who have previously been instructed by health professionals to follow a lipid-lowering diet. We investigated dietary composition before and after dietary intervention by a clinical nutritionist in 46 individuals, aged 33 to 65 years, with primary hypercholesterolemia. After intervention, there was a 25% reduction in the intake of saturated fat and of cholesterol (p < 0.0001) and an 65% elevation of the P/S-ratio (p < 0.05). Percentage of energy from fat remained unchanged. The diet of patients who are already following a lipid-lowering diet can be improved by expert advice from a clinical nutritionist, and they easily adjust to new habits.

[Iron status in postmenopausal women]. / Jernstatus hos postmenopausale kvinner.

Iron intake was determined in 42 postmenopausal women from food records over two three-day periods, with weighing of the food. The average daily intake was 9.4 mg (95% confidence interval 8.8-9.9). In 70% of the women the intake was lower than the nationally recommended 10 mg per day. Serum-ferritin was measured to detect possible iron deficiency. The average serum-ferritin was 60 micrograms/l (95% confidence interval 47-72). Only two women were advised to take iron supplements. One of these women had a serum-ferritin below 12 micrograms/l, which is regarded as empty iron store. A low iron intake in postmenopausal women is not an indication to prescribe iron supplements. Iron deficiency can be assessed only by serum-ferritin measurements.

The effect of the gastrointestinal lipase inhibitor, orlistat, on serum lipids and lipoproteins in patients with primary hyperlipidaemia.

The effect of orlistat, a nonabsorbed inhibitor of gastric and pancreatic lipases, was examined in patients with primary hyperlipidaemia (serum cholesterol > or = 6.2 mmol.l-1 and triglycerides < or = 5.0 mmol.l-1) not responsive to dietary change alone. In a multicentre, randomised, double-blind study, 103 men and 70 women received 30, 90, 180, or 360 mg or orlistat or placebo for 8 weeks. Total and low-density lipoprotein cholesterol levels were reduced by 4% and 5% with 30 mg orlistat, by 7% and 8% with 90 mg orlistat, by 7% and 7% with 180 mg orlistat and by 11% and 10% with 360 mg orlistat compared to placebo. High density lipoprotein cholesterol levels significantly decreased in the 360 mg orlistat group. Triglyceride levels significantly increased in the placebo group but not in the drug groups. Body weight decreased by 1.2 kg with 360 mg orlistat, despite a weight maintenance diet. Decreases in vitamin E and D levels occurred, although both vitamins remained within the normal range. Adverse effects from the gastrointestinal tract were frequent, but led to discontinuation of therapy in only seven patients. Orlistat is a new therapeutic drug for the treatment of hyperlipidaemia that may be particularly useful among overweight patients. Its potential place in therapy will await long-term studies. Vitamin supplementation should be considered during treatment.

[Familial hypercholesterolemia--intensive diet therapy combined with drug therapy]. / Familiaer hyperkolesterolemi--intensiv kostbehandling i kombinasjon med medikamentell behandling.

The aim of the investigation was twofold: to study the effect of lovastatin, a potent inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, alone and in combination with other lipid lowering drugs in an open 48 week single centre study, and to study if lipid lowering drugs influence adherence to diet in adult patients with familial hypercholesterolemia. Lovastatin monotherapy (80 mg daily) for 12 weeks reduced serum cholesterol, LDL-cholesterol and triglycerides levels by 36%, 44% and 24% respectively. HDL-cholesterol level was increased by 12%. The addition of 16 g cholestyramine daily further increased the reduction of total cholesterol and LDL-cholesterol levels by 17% and 24% respectively. Addition of 1 g probucol daily decreased total cholesterol, LDL-cholesterol and HDL-cholesterol levels by 9%, 5% and 27% respectively. Addition of omega-3-fatty acids (3.6 g daily) reduced total cholesterol, LDL-cholesterol and triglycerides levels by 10%, 12% and 20% respectively. Administration of potent lipid lowering agents did not influence adherence to a diet with a mean daily fat energy of 21% (CI: 20-22), cholesterol of 177 mg (CI: 157-196) and P/S ratio of 0.75 (CI: 0.66-0.84). A significant increase in liver enzymes was recorded in only one patient. One patient was withdrawn from the study because of myositis.

Dietary n-6 fatty acids inhibit the incorporation of dietary n-3 fatty acids in thrombocyte and serum phospholipids in humans: a controlled dietetic study.

The effect of a high dietary intake of n-6 fatty acids (36 g daily) vs a low intake (4-6 g daily) on the incorporation of fatty acids from a dietary supplementation of n-3 fatty acids (6 g daily) was studied for 8 weeks in 15 healthy, normolipaemic volunteers. The importance of a high (43.6) vs a low (20.6) energy percentage from fat was also investigated in the participants on a low n-6 intake. Fatty acid analyses of serum and thrombocyte phospholipids showed a marked increase in docosahexaenoic acid (22:6 (n-3), DHA) and especially eicosapentaenoic acid (20:5 (n-3), EPA) in both the high and low n-6 groups after 14 days, but the changes were significantly greater in the low n-6 diet groups. Changes of the ratio between EPA and arachidonic acid (20:4 (n-6), AA) in phospholipids followed an identical pattern in serum and thrombocytes. This indicates that thrombocytes are influenced by the fatty acid composition in serum. The results showed that incorporation of n-3 fatty acids in phospholipids was reduced by a high intake of dietary n-6 fatty acids in the cells and lipid fractions studied. The observed effect of dietary n-6 fatty acids was independent of the energy percentage provided by dietary fat. In order to obtain an optimal effect of n-3 supplementation, the intake of linoleic acid has to be considered and kept at a low level. The serum content of cholesterol was unaffected, but the concentration of triacylglycerol was reduced during the supplementation period.