Minna L. Hannuksela

Relation of polymorphisms in the cholesteryl ester transfer protein gene to transfer protein activity and plasma lipoprotein levels in alcohol drinkers

We investigated the interaction between genetic and environmental factors in the regulation of plasma HDL cholesterol concentration by determining TaqI and EcoN I restriction fragment length polymorphisms at the cholesteryl ester transfer protein (CETP) gene locus in 93 male alcohol drinkers and 82 control men. The highest plasma CETP activity and the lowest HDL cholesterol concentration were in the control subjects who were homozygous for the presence of the TaqI B restriction site (genotype 1-1). The lowest CETP activity and the highest HDL cholesterol among the control subjects were in those with genotype 2-2. These associations were, however, evident only in the non-smokers (P = 0.03 for CETP activity and P = 0.05 for HDL cholesterol). The non-smoking control subjects with genotype 1-1 had 19% higher CETP activity and 16% lower HDL cholesterol than those with genotype 2-2 (mean +/- S.D., 113 +/- 25 nmol/h/ml and 1.16 +/- 0.30 mmol/l vs. 95 +/- 16 nmol/h/ml and 1.38 +/- 0.34 mmol/l, respectively), and CETP activity and HDL cholesterol were negatively correlated (r = -0.280, P = 0.03, n = 59). The alcohol drinkers had 30% lower CETP activity (P < 0.001) and 48% higher HDL cholesterol (P < 0.001) than the controls. CETP activity was not affected by the TaqI B genotype in the alcohol drinkers. The lowest HDL cholesterol was in subjects with genotype 1-1 (1.68 +/- 0.60 mmol/l), but those with genotype 2-2 had lower HDL cholesterol than those with genotype 1-2 (1.78 +/- 0.59 and 1.93 +/- 0.66 mmol/l, respectively). The data of the alcohol drinkers fitted better with the quadratic regression model than with the linear one, suggesting a trend towards a curved relationship between the TaqI B genotype and HDL cholesterol in both the non-smoking and smoking alcohol drinkers. Total, LDL or VLDL cholesterol, total or VLDL triglycerides did not differ between the TaqI B genotypes either in the alcohol drinkers or the controls. Lipid and lipoprotein levels and CETP activities were likewise similar in the TaqI A and EcoN I polymorphisms. Our data indicate that CETP TaqI B polymorphism is related to plasma CETP activity and HDL cholesterol concentration in non-smoking men, but these associations are affected by smoking and alcohol drinking.

Biochemical markers of alcoholism.

Abstract Alcohol and alcohol-related diseases have become a major cause of death in Western countries. The most sensitive and specific of the commonly used biomarkers of alcohol intake are carbohydrate-deficient transferrin (CDT), and the combination of γ-glutamyltransferase (GGT) and CDT. Other widely used laboratory markers are GGT, mean corpuscular volume of erythrocytes and the ratio of aspartate aminotransferase to alanine aminotransferase. Blood ethanol levels reveal recent alcohol use. However, more specific and sensitive biomarkers to improve the detection of excessive alcohol use at an early stage are needed. New biomarkers, not yet used in routine clinical work, include phosphatidylethanol, fatty acid ethyl esters, ethyl glucuronide, sialic acid, and acetaldehyde adducts. Clin Chem Lab Med 2007;45:953–61.

Benefits and risks of sauna bathing

Although sauna bathing causes various acute, transient cardiovascular and hormonal changes, it is well tolerated by most healthy adults and children. Sauna bathing does not influence fertility and is safe during the uncomplicated pregnancies of healthy women. Some studies have suggested that long-term sauna bathing may help lower blood pressure in patients with hypertension and improve the left ventricular ejection fraction in patients with chronic congestive heart failure, but additional data are needed to confirm these findings. The transient improvements in pulmonary function that occur in the sauna may provide some relief to patients with asthma and chronic bronchitis. Sauna bathing may also alleviate pain and improve joint mobility in patients with rheumatic disease. Although sauna bathing does not cause drying of the skin-and may even benefit patients with psoriasis-sweating may increase itching in patients with atopic dermatitis. Contraindications to sauna bathing include unstable angina pectoris, recent myocardial infarction, and severe aortic stenosis. Sauna bathing is safe, however, for most people with coronary heart disease with stable angina pectoris or old myocardial infarction. Very few acute myocardial infarctions and sudden deaths occur in saunas, but alcohol consumption during sauna bathing increases the risk of hypotension, arrhythmia, and sudden death, and should be avoided.

Increased high-density lipoprotein cholesterol concentration in alcoholics is related to low cholesteryl ester transfer protein activity

Abstract. Cholesteryl ester transfer protein (CETP) facilitates the transfer of cholesteryl esters from HDL to apo B‐containing lipoproteins. Since alcoholics have high HDL cholesterol and low LDL cholesterol levels, a defect in cholesteryl ester transfer could be responsible for the alcohol‐induced alteration in cholesterol distribution between lipoproteins. To test this hypothesis, we compared CETP activity in plasma from 30 alcoholics without severe liver damage and 16 control subjects.

Effect of alcohol on lipids and lipoproteins in relation to atherosclerosis

Several studies indicate that light-to-moderate alcohol consumption is associated with a low prevalence of coronary heart disease. An increase in high-density lipoprotein (HDL) cholesterol is associated with alcohol intake and appears to account for approximately half of alcohols cardioprotective effect. In addition to changes in the concentration and composition of lipoproteins, alcohol consumption may alter the activities of plasma proteins and enzymes involved in lipoprotein metabolism: cholesteryl ester transfer protein, phospholipid transfer protein, lecithin: cholesterol acyltransferase, lipoprotein lipase, hepatic lipase, paraoxonase-1 and phospholipases. Alcohol intake also results in modifications of lipoprotein particles: low sialic acid content in apolipoprotein components of lipoprotein particles (e.g., HDL apo E and apo J) and acetaldehyde modification of apolipoproteins. In addition, “abnormal” lipids, phosphatidylethanol, and fatty acid ethyl esters formed in the presence of ethanol are associated with lipoproteins in plasma. The effects of lipoproteins on the vascular wall cells (endothelial cells, smooth muscle cells, and monocyte/macrophages) may be modulated by ethanol and the alterations further enhanced by modified lipids. The present review discusses the effects of alcohol on lipoproteins in cholesterol transport, as well as the novel effects of lipoproteins on vascular wall cells.

A novel Bayesian approach to quantify clinical variables and to determine their spectroscopic counterparts in 1H NMR metabonomic data

BackgroundA key challenge in metabonomics is to uncover quantitative associations between multidimensional spectroscopic data and biochemical measures used for disease risk assessment and diagnostics. Here we focus on clinically relevant estimation of lipoprotein lipids by 1H NMR spectroscopy of serum.ResultsA Bayesian methodology, with a biochemical motivation, is presented for a real 1H NMR metabonomics data set of 75 serum samples. Lipoprotein lipid concentrations were independently obtained for these samples via ultracentrifugation and specific biochemical assays. The Bayesian models were constructed by Markov chain Monte Carlo (MCMC) and they showed remarkably good quantitative performance, the predictive R-values being 0.985 for the very low density lipoprotein triglycerides (VLDL-TG), 0.787 for the intermediate, 0.943 for the low, and 0.933 for the high density lipoprotein cholesterol (IDL-C, LDL-C and HDL-C, respectively). The modelling produced a kernel-based reformulation of the data, the parameters of which coincided with the well-known biochemical characteristics of the 1H NMR spectra; particularly for VLDL-TG and HDL-C the Bayesian methodology was able to clearly identify the most characteristic resonances within the heavily overlapping information in the spectra. For IDL-C and LDL-C the resulting model kernels were more complex than those for VLDL-TG and HDL-C, probably reflecting the severe overlap of the IDL and LDL resonances in the 1H NMR spectra.ConclusionThe systematic use of Bayesian MCMC analysis is computationally demanding. Nevertheless, the combination of high-quality quantification and the biochemical rationale of the resulting models is expected to be useful in the field of metabonomics.

Altered Transfer of Cholesteryl Esters and Phospholipids in Plasma From Alcohol Abusers

The net mass transfer (NMT) of cholesteryl esters (CEs), triglycerides (TGs), and phospholipids (PLs) between lipoproteins was measured after incubation of fresh plasma for up to 2 hours from 18 male alcohol abusers and 17 male volunteer control subjects. In alcohol abusers the mean value of CE NMT was 3.7 nmol.mL-1.h-1 from apolipoprotein B-containing lipoproteins (apoB-containing lipoproteins) to HDL and in control subjects 8.7 nmol.mL-1.h-1 from HDL to apoB-containing lipoproteins. The NMT of PL was higher in alcohol abusers than in control subjects (35.0 vs 11.6 nmol.mL-1.h-1 from apoB-containing lipoproteins to HDL, respectively), and plasma PL transfer protein (TP) activity was 33% higher (P < .05) in alcohol abusers than in control subjects. The lack of correlation between the NMTs and CETP and PLTP activities suggests that the NMT could more closely reflect the role of lipoprotein properties in reverse cholesterol transport in vivo, whereas in vitro activities reflect the total capacity of transfer but not its direction. The rate of CE NMT from HDL to apoB-containing lipoproteins was dependent on the VLDL TG concentration. Moreover, at low VLDL TG levels, the increased HDL cholesterol concentration in alcohol abusers reversed the direction of CE NMT. This situation could be reconstructed in the plasma of control subjects by adding autologous HDL or VLDL to mimic the lipoprotein profiles of the alcohol abusers. Addition of VLDL enhanced the CE NMT from HDL to apoB-containing lipoproteins, whereas addition of HDL had an opposite effect, and at higher HDL levels, even reversed the direction of CE NMT. In conclusion, the NMT of CE and PL in alcohol abusers differs from that in control subjects. The concentrations of HDL and VLDL seem to be the major determinants of the direction of CE NMT in alcohol abusers.

Estimation of VLDL, IDL, LDL, HDL2, apoA-I, and apoB from the Friedewald inputs—apoB and IDL, but not LDL, are associated with mortality in type 1 diabetes

Background. There is an unmet need for a straightforward and cost-effective assessment of multiple lipoprotein risk factors for vascular diseases. Aims. 1) To study the relation of various lipoprotein lipid and apolipoprotein (apo) measures on the Friedewald inputs, i.e. plasma triglycerides (TG), cholesterol (TC), and high-density lipoprotein cholesterol (HDL-C). 2) To build up regression models for the appropriate measures based solely on the Friedewald inputs. Methods. Data were available for 1,775 plasma samples, from which very-low-density lipoprotein (VLDL), intermediate-density lipoprotein (IDL), low-density lipoprotein (LDL), and HDL were also isolated by ultracentrifugation. For HDL2-C and apolipoproteins, 343 and 247 samples were available, respectively. Results. Accurate models were obtained for VLDL-TG (cross-validation r=0.98), LDL-C (r=0.91), HDL2-C (r=0.92), apoA-I (r=0.92), and apoB (r=0.95). A semi-quantitative model was obtained for IDL-C (r=0.78). Due to the anticipated role of IDL-C in atherosclerosis, it was still kept within the accepted models and pursued further. The associations of the estimates with premature deaths were studied in 4,084 patients with type 1 diabetes. The associations of IDL-C and LDL-C were markedly different, the best predictors of mortality being apoB, apoB to apoA-I ratio, and IDL-C. Conclusions. The new models allow identification of clinically relevant lipoprotein profiles with no added cost to the conventional Friedewald formula.

Immunological detection of in vitro formed phosphatidylethanol--an alcohol biomarker--with monoclonal antibodies.

BACKGROUND Phosphatidylethanol (PEth) is a promising new marker for detecting long-term alcohol abuse with excellent sensitivity and specificity. Current methods are based on the high performance liquid chromatography-mass spectrometric method and therefore require high levels of expertise and expensive instrumentation. This study was designed to generate PEth-specific monoclonal antibodies for PEth immunoassay development. METHODS C57/BL6 mice were immunized with PEth in 3 different carriers, mouse serum albumin, mouse high-density lipoproteins, and human low-density lipoprotein (LDL). Mouse splenocytes were fused with a mouse myeloma cell line using the hybridoma technique. Mouse IgM-producing cell lines were selected by limiting dilutions. Binding characteristics of the anti-PEth antibodies were studied using luminometric immunoassays and sequence analysis of the variable region mRNA sequences of the antibodies. Produced antibodies were purified by chromatographic methods. PEth was detected with these antibodies in fluorescence immunoassay and flow cytometric analysis. RESULTS We generated monoclonal cell lines (2B1 and 2E9) that produce IgM antibodies binding specifically to PEth but not to structurally or chemically similar phospholipids, such as phosphatidylcholine, phosphatidic acid, and cardiolipin. We show here that these anti-PEth antibodies can be used to detect PEth in a fluorescent PEth assay and FACS analysis of human red blood cell samples spiked with PEth. CONCLUSIONS The present study shows that PEth-specific monoclonal antibodies can be generated using traditional hybridoma technique. Immunogenicity of PEth was enhanced using human LDL as an immunization carrier. The generated monoclonal anti-PEth antibodies, 2B1 and 2E9 bind to PEth in fluid phase and in biological membranes.

HDL2 of Heavy Alcohol Drinkers Enhances Cholesterol Efflux From Raw Macrophages via Phospholipid‐Rich HDL2b Particles

BACKGROUND Alcohol consumption is associated with increased serum high density lipoprotein (HDL) cholesterol levels and a decreased risk for the development of atherosclerosis. However, the effects of heavy alcohol intake on reverse cholesterol transport, one of the key anti-atherogenic processes related to HDL, are poorly known. METHODS The ability of total HDL as well as HDL(2) and HDL(3) subclasses to promote cholesterol efflux from (3)H-cholesterol-labeled RAW 264.7 macrophages was studied among 6 heavy alcohol drinkers and 6 controls. Distribution of HDL subclasses was analyzed by 4 to 30% native gradient gels. Serum phospholipid transfer protein (PLTP) and cholesteryl ester transfer protein (CETP) activities were analyzed among several other biochemical measures. RESULTS Cholesterol efflux to HDL(2) of heavy drinkers was 22% (p = 0.025) higher relative to controls. The increase in HDL(2) phospholipids, with a concomitant 2-fold (p = 0.055) increase in large HDL(2b) particles, was associated with enhanced cholesterol efflux to HDL(2). Interestingly, the cholesterol efflux to HDL(3) did not differ between the 2 study groups. These findings may be partially explained by a decreased CETP activity (-26%, p = 0.037) and an increased PLTP activity (39%, p = 0.045) in heavy drinkers. CONCLUSIONS The increased cholesterol efflux potential of HDL(2) is most likely an anti-atherogenic feature linked to heavy alcohol consumption. The cholesterol efflux and HDL phospholipids also associated strongly within the whole study group (r(s) = 0.910, p <or= 0.01) suggesting a common pathway of enhanced cholesterol efflux via enlarged phospholipid-rich HDL particles.