J.J.P. Kastelein

Cholesteryl ester transfer protein TaqIB variant, high-density lipoprotein cholesterol levels, cardiovascular risk, and efficacy of pravastatin treatment : individual patient meta-analysis of 13,677 subjects

Background—Several studies have reported that the cholesteryl ester transfer protein (CETP) TaqIB gene polymorphism is associated with HDL cholesterol (HDL-C) levels and the risk of coronary artery disease (CAD), but the results are inconsistent. In addition, an interaction has been implicated between this genetic variant and pravastatin treatment, but this has not been confirmed. Methods and Results—A meta-analysis was performed on individual patient data from 7 large, population-based studies (each >500 individuals) and 3 randomized, placebo-controlled, pravastatin trials. Linear and logistic regression models were used to assess the relation between TaqIB genotype and HDL-C levels and CAD risk. After adjustment for study, age, sex, smoking, body mass index (BMI), diabetes, LDL-C, use of alcohol, and prevalence of CAD, TaqIB genotype exhibited a highly significant association with HDL-C levels, such that B2B2 individuals had 0.11 mmol/L (0.10 to 0.12, P<0.0001) higher HDL-C levels than did B1B1 individuals. Second, after adjustment for study, sex, age, smoking, BMI, diabetes, systolic blood pressure, LDL-C, and use of alcohol, TaqIB genotype was significantly associated with the risk of CAD (odds ratio=0.78 [0.66 to 0.93]) in B2B2 individuals compared with B1B1 individuals (P for linearity=0.008). Additional adjustment for HDL-C levels rendered a loss of statistical significance (P=0.4). Last, no pharmacogenetic interaction between TaqIB genotype and pravastatin treatment could be demonstrated. Conclusions—The CETP TaqIB variant is firmly associated with HDL-C plasma levels and as a result, with the risk of CAD. Importantly, this CETP variant does not influence the response to pravastatin therapy.

Short-term efficacy and safety of pravastatin in 72 children with familial hypercholesterolemia

The safety, tolerability, and efficacy of a 12-wk treatment with pravastatin, 5, 10, and 20 mg/d, was evaluated in 72 children with heterozygous familial hypercholesterolemia (FH) in a double-blind, randomized, and placebo-controlled study. The results show that pravastatin was well tolerated and that adverse events were mild and equally distributed among the three treatment groups. Plasma total and LDL cholesterol levels were significantly reduced in all pravastatin treatment groups, in comparison with the control group; -24.6% (-28.1 to 21.0) and -32.9% (-37.0 to -28.6), for mean change and 95% confidence interval, respectively. In four children plasma LDL cholesterol levels were reduced within normal limits for sex and age. HDL cholesterol increased in the pravastatin 20-mg group, +10.8% (+3.4 to +18.8), whereas plasma apo B100 and very LDL (VLDL) cholesterol levels were reduced within all pravastatin-treated groups -26.8% (-13.2 to -21.7) and-24.5% (-35.0 to -12.3). These data show that short-term pravastatin treatment of children with FH is safe and effective, although long-term dose titration studies with 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors need to be performed, to reduce plasma LDL cholesterol levels below a predefined level. The results of these studies have to be awaited before new treatment strategies are to be considered in these children.

The 20210 G→ A mutation in the 3'-untranslated region of the prothrombin gene and the risk for arterial thrombotic disease

A sequence variation in the 3′‐untranslated region of the prothrombin (PT) gene (20210 G → A) was recently claimed to be associated with elevated plasma prothrombin levels and an increased risk for venous and arterial thrombosis. We examined the prevalence of the 20210 A allele in the prothrombin gene in 400 healthy controls and in 263 patients with proven premature atherosclerotic disease. In addition, we measured prothrombin, prothrombin fragment 1 + 2, thrombin–antithrombin (TAT) complex and D‐dimer levels in plasma from carrier and non‐carrier patients. The frequency of the variant allele was 1% in the control subjects and 2.7% in the patient group, yielding a relative risk (RR) for the 20210 A allele of 2.7 (95% CI 0.8–9.4). All heterozygotes in the patient group were found to have had a myocardial infarction (MI), yielding a RR for MI of 4.2 (95% CI 1.2–14.6). Plasma prothrombin levels in carriers (126 ± 10) were higher than in non‐carriers (103 ± 1, P = 0.02). The levels of TAT complexes (16 ± 9 v 6 ± 1 μg/ml, P = 0.02) as well as of prothrombin fragment 1 + 2 (1.5 ± 0.3 v 1.0 ± 0.1 nmol/l, P = 0.02) were also elevated in carriers of the mutation. Our findings suggest that the 20210 G → A mutation in the prothrombin gene is a genetic risk factor for MI. In addition, our data provide evidence for an association of the mutation with excessive thrombin generation, which may contribute to the understanding of its role in venous and arterial disease.

Cholesteryl ester transfer protein concentration is associated with progression of atherosclerosis and response to pravastatin in men with coronary artery disease (REGRESS)

Background  The TaqIB polymorphism in the cholesteryl ester transfer protein (CETP) gene is associated with HDL‐C, progression of coronary artery disease (CAD) and response to pravastatin treatment in men with angiographically proven CAD (REGRESS). We hypothesized that differences in CETP concentration could explain these associations and now investigated whether CETP concentration is an independent determinant of these parameters.

Tumor necrosis factor-α inhibition protects against endotoxin-induced endothelial glycocalyx perturbation

OBJECTIVE Inflammatory stimuli profoundly increase the vulnerability of the vessel wall to atherogenesis. The endothelial glycocalyx, a layer of glycosaminoglycans and proteoglycans covering the luminal side of the vasculature, has recently emerged as an orchestrator of vascular homeostasis. In the present study, we investigated whether endotoxin-induced inflammatory reactions lead to a decrease of endothelial glycocalyx thickness in humans and whether tumor necrosis factor-alpha (TNFalpha) plays a role in this process. DESIGN, SUBJECTS AND INTERVENTION Healthy male volunteers received low-dose endotoxin (1ng/kg) intravenously, with (n=8) or without (n=13) pre-treatment with the soluble TNFalpha receptor etanercept. Endothelial glycocalyx thickness and related parameters were determined after endotoxin challenge. RESULTS Endotoxin resulted in a profound reduction in microvascular glycocalyx thickness (from 0.60+/-0.1 to 0.30+/-0.1microm, p<0.01). Concomitantly, plasma levels of the principal glycocalyx constituent hyaluronan (62+/-18 to 85+/-24ng/mL, p<0.05), monocyte activation and coagulation activation increased (F1+2; 0.3+/-0.1 to 2.8+/-1.5nmol/L, p<0.05 and d-dimer; from 0.2+/-0.1 to 0.4+/-0.1mg/L, p<0.05 compared to baseline). Inhibition of TNFalpha by etanercept attenuated loss of microvascular glycocalyx thickness (0.54+/-0.1 to 0.35+/-0.1mum, p<0.05). Changes in hyaluronan (58+/-13 to 46+/-10ng/mL, p<0.05) and coagulation activation were also attenuated (F1+2; 0.3+/-0.1 to 2.1+/-0.9nmol/L and d-dimer; from 0.2+/-0.1 to 0.3+/-0.1mg/L, p<0.05 compared to baseline). CONCLUSIONS These data suggest that inflammatory activity, in part mediated by TNFalpha, leads to perturbation of the endothelial glycocalyx in humans. This may contribute to the vascular vulnerability induced by inflammation.

An Intronic Mutation in a Lariat Branchpoint Sequence Is a Direct Cause of an Inherited Human Disorder (Fish-Eye Disease)

The first step in the splicing of an intron from nuclear precursors of mRNA results in the formation of a lariat structure. A distinct intronic nucleotide sequence, known as the branchpoint region, plays a central role in this process. We here describe a point mutation in such a sequence. Three sisters were shown to suffer from fish-eye disease (FED), a disorder which is caused by mutations in the gene coding for lecithin:cholesterol acyltransferase (LCAT). Sequencing of the LCAT gene of all three probands revealed compound heterozygosity for a missense mutation in exon 4 which is reported to underlie the FED phenotype, and a point mutation located in intron 4 (IVS4:T-22C). By performing in vitro expression of LCAT minigenes and reverse transcriptase PCR on mRNA isolated from leukocytes of the patient, this gene defect was shown to cause a null allele as the result of complete intron retention. In conclusion, we demonstrated that a point mutation in a lariat branchpoint consensus sequence causes a null allele in a patient with FED. In addition, our finding illustrates the importance of this sequence for normal human mRNA processing. Finally, this report provides a widely applicable strategy which ensures fast and effective screening for intronic defects that underlie differential gene expression.

The effect of a common methylenetetrahydrofolate reductase mutation on levels of homocysteine, folate, vitamin B12 and on the risk of premature atherosclerosis.

An increased total plasma homocysteine level is an established risk factor for atherosclerotic vascular disease. The plasma level of homocysteine is influenced by both environmental and genetic factors. An important genetic determinant of plasma homocysteine is a common amino acid dimorphism (Ala222Val) in the methylenetetrahydrofolate reductase (MTHFR) gene. Individuals homozygous for the Val allele have significantly higher homocysteine levels than those with an Ala/Val or Ala/Ala genotype. Moreover, the Val/Val genotype has been claimed to be a strong genetic risk factor for atherosclerosis. The aim of the present study is: (1) to determine the risk associated with the MTHFR dimorphism by comparing the genotype distribution in patients with premature atherosclerosis with that in a group of healthy controls; and (2) to investigate the relationship between the MTHFR genotype and parameters of homocysteine metabolism. The patient group consisted of 257 consecutive referred individuals with angiographically proven premature ( <50 years of age) arterial disease (coronary, and/or peripheral vascular disease). A total of 272 healthy hospital workers without a history of vascular disease were selected as a control group. The MTHFR-genotype was determined by PCR and gel-electrophoresis. A methionine-loading test was performed on 245 patients, and, in addition to homocysteine, levels of folate and vitamin B12 were measured. We found a strong correlation between MTHFR genotype and plasma homocysteine levels both before and after methionine loading. In addition, the MTHFR genotype seems important for the inverse relationship between homocysteine and folate and vitamin B12 levels. Lastly, the MTHFR genotype distribution was not different between patient and control groups. MTHFR genotype is a strong determinant of plasma homocysteine levels. Moreover, the plasma level of folate, which by itself influences homocysteine levels, is also dependent on the MTHFR genotype. In Val/Val genotypes, low levels of both folate and B12 lead to a relatively large increase in homocysteine levels. Nevertheless, the MTHFR genotype does not increase the risk for premature coronary artery disease.

Sustained changes in lipid profile and macrophage migration inhibitory factor levels after anti-tumour necrosis factor therapy in rheumatoid arthritis

Background: Macrophage migration inhibitory factor (MIF) has recently emerged as an important cytokine possibly linking rheumatoid arthritis (RA) and atherogenesis. Because atherogenesis is accelerated in RA this study was conducted to investigate whether anti-tumour necrosis factor (TNF) therapy could lead to sustained downregulation of systemic MIF levels and improvement in lipid profiles. Methods: Fifty RA patients with active disease (disease activity score in 28 joints (DAS28) ⩾3.2), who started adalimumab therapy at 40 mg every other week, were included. At baseline, weeks 16 and 52 serum levels of MIF and lipids were assessed. In addition, the DAS28 and serum C-reactive protein (CRP) levels and erythrocyte sedimentation rate (ESR) were determined. Results: After 16 weeks of adalimumab therapy, both DAS28 and MIF levels were significantly decreased (p<0.001 and p = 0.020, respectively). This was sustained up to week 52 (p<0.001 and p = 0.012, respectively). CRP levels and ESR were significantly reduced after 16 and 52 weeks of adalimumab therapy (p<0.001). High-density lipoprotein cholesterol levels increased at week 16 (p<0.001), but returned to baseline at week 52. Apolipoprotein (apo) A-I levels increased at week 16 (p<0.001) and remained stable (p = 0.005). This resulted in an improved apo B/A-I ratio. Conclusions: The results underline the sustained downregulation of MIF as a potential new mechanism by which anti-TNF therapy might reduce vascular inflammation, and as such perhaps cardiovascular morbidity in RA patients. This hypothesis is supported by an improved apo B/A-I ratio as well as reduced CRP levels in these patients.

The future of best practice

Evidence from epidemiological and clinical studies continues to improve our understanding of the pathogenesis of coronary heart disease (CHD). However, despite major advances in the development of diagnostic methods and effective treatment, CHD remains the leading cause of mortality in the Western world. That cholesterol lowering is of major importance in lowering the risk of developing coronary artery disease (CAD) is now an accepted principle in medicine. Most patients with CAD will require drug therapy to achieve target lipid levels. Subgroup analyses of data from the landmark statin trials show that this benefit is seen in all patient groups: male and female, older and younger patients, diabetics and non-diabetics, and patients with and without myocardial revascularization. Recent evidence suggests that the extent to which cholesterol is lowered is also important and that the attainment of reduced low-density lipoprotein cholesterol levels is associated with greater reductions in the risk of cardiovascular events. Unfortunately, data from studies to date do not provide a definitive answer to the question of whether there is a benefit in lowering cholesterol to very low levels.

Treatment of primary hypercholesterolaemia. Short-term efficacy and safety of increasing doses of simvastatin and pravastatin: a double-blind comparative study

Abstract. Objectives. To compare the efficacy and safety of increasing doses (0, 10, 20 and 40 mg day−1, each dose for 6 weeks) of the inhibitors of 3‐hydroxy‐3‐methylglutaryl coenzyme A reductase, simvastatin and pravastatin, in the treatment of primary hypercholesterolaemia.