M. Castro Cabezas

Apo B versus cholesterol in estimating cardiovascular risk and in guiding therapy: report of the thirty-person/ten-country panel.

There is abundant evidence that the risk of atherosclerotic vascular disease is directly related to plasma cholesterol levels. Accordingly, all of the national and transnational screening and therapeutic guidelines are based on total or LDL cholesterol. This presumes that cholesterol is the most important lipoprotein‐related proatherogenic risk variable. On the contrary, risk appears to be more directly related to the number of circulating atherogenic particles that contact and enter the arterial wall than to the measured concentration of cholesterol in these lipoprotein fractions. Each of the atherogenic lipoprotein particles contains a single molecule of apolipoprotein (apo) B and therefore the concentration of apo B provides a direct measure of the number of circulating atherogenic lipoproteins. Evidence from fundamental, epidemiological and clinical trial studies indicates that apo B is superior to any of the cholesterol indices to recognize those at increased risk of vascular disease and to judge the adequacy of lipid‐lowering therapy. On the basis of this evidence, we believe that apo B should be included in all guidelines as an indicator of cardiovascular risk. In addition, the present target adopted by the Canadian guideline groups of an apo B <90 mg dL−1 in high‐risk patients should be reassessed in the light of the new clinical trial results and a new ultra‐low target of <80 mg dL−1 be considered. The evidence also indicates that the apo B/apo A‐I ratio is superior to any of the conventional cholesterol ratios in patients without symptomatic vascular disease or diabetes to evaluate the lipoprotein‐related risk of vascular disease.

Impaired fatty acid metabolism in familial combined hyperlipidemia. A mechanism associating hepatic apolipoprotein B overproduction and insulin resistance.

To establish whether insulin resistance and/or postprandial fatty acid metabolism might contribute to familial combined hyperlipidemia (FCH) we have examined parameters of insulin resistance and lipid metabolism in six FCH kindreds. Probands and relatives (n = 56) were divided into three tertiles on the basis of fasting plasma triglycerides (TG). Individuals in the highest tertile (TG > 2.5 mM; n = 14) were older and had increased body mass index, systolic blood pressure, and fasting plasma insulin concentrations compared with individuals in the lowest tertile (n = 24). The former also presented with decreased HDL cholesterol and increased total plasma cholesterol, HDL-TG, and apoprotein B, E, and CIII concentrations. Insulin concentrations were positively correlated with plasma apo B, apo CIII, apo E, and TG, and inversely with HDL cholesterol. Fasting nonesterified fatty acids (NEFA) were elevated in FCH subjects compared to six unrelated controls and five subjects with familial hypertriglyceridemia. Prolonged and exaggerated postprandial plasma NEFA concentrations were found in five hypertriglyceridemic FCH probands. In FCH the X2 minor allele of the AI-CIII-AIV gene cluster was associated with increased fasting plasma TG, apo CIII, apo AI, and NEFA concentrations and decreased postheparin lipolytic activities. The clustering of risk factors associated with insulin resistance in FCH indicates a common metabolic basis for the FCH phenotype and the syndrome of insulin resistance probably mediated by an impaired fatty acid metabolism.

Triglyceride-rich lipoproteins in non-insulin-dependent diabetes mellitus: post-prandial metabolism and relation to premature atherosclerosis.

Non‐insulin‐dependent diabetes mellitus is frequently associated with premature atherosclerosis. Abnormalities in lipid and lipoprotein metabolism contribute to the increased risk of coronary heart disease. One of the most common lipid abnormalities in non‐insulin‐dependent diabetes mellitus is hypertriglyceridaemia. In the present paper, the authors review the metabolism of triglyceride‐rich lipoproteins, with special emphasis on the post‐prandial state. Several studies have demonstrated that levels of atherogenic post‐prandial lipoproteins are increased in patients with non‐insulin‐dependent diabetes mellitus. An increased supply of glucose and free fatty acids contributes to overproduction of very low‐density lipoproteins, increasing the burden of triglyceride‐rich lipoproteins on the common lipolytic pathway at the level of lipoprotein lipase. Low lipoprotein lipase activity and increased amounts of lipolysis‐inhibiting free fatty acids further impair lipolysis of post‐prandial lipoproteins. The clearance of atherogenic remnants is also delayed in non‐insulin‐dependent diabetes mellitus. There is evidence that a relative hepatic removal defect exists, secondary to impaired remnant‐receptor interaction and increased competition with very low density lipoprotein remnants. Correction of the increased post‐prandial lipaemia in non‐insulin‐dependent diabetes mellitus is advisable, as it may contribute to attenuation of the risk on premature atherosclerosis. When dietary measures and hypoglycaemic agents have failed to achieve acceptable lipid levels, lipid‐lowering drugs should be advised. Fibric acids and hydroxymethylglutaryl coenzyme A (HMG CoA) reductase inhibitors are the drugs of choice.

Novel aspects of postprandial lipemia in relation to atherosclerosis

Postprandial hyperlipidemia is considered to be a substantial risk factor for atherosclerosis. Interestingly, this concept has never been supported by randomized clinical trials. The difficulty lies in the fact that most interventions aimed to reduce postprandial lipemia, will also affect LDL-C levels. The atherogenic mechanisms of postprandial lipids and lipoproteins can be divided into direct lipoprotein-mediated and indirect effects; the latter, in part, by inducing an inflammatory state. Elevations in postprandial triglycerides (TG) have been related to the increased expression of postprandial leukocyte activation markers, up-regulation of pro-inflammatory genes in endothelial cells and involvement of the complement system. This set of events is part of the postprandial inflammatory response, which is one of the recently identified potential pro-atherogenic mechanisms of postprandial lipemia. Especially, complement component 3 levels show a close correlation with postprandial lipemia and are also important determinants of the metabolic syndrome. In clinical practice, fasting TG are frequently used as reflections of postprandial lipemia due to the close correlation between the two. The use of serial capillary measurements in an out-of-hospital situation is an alternative for oral fat loading tests. Daylong TG profiles reflect postprandial lipemia and are increased in conditions like the metabolic syndrome, type 2 diabetes and atherosclerosis. Studies are needed to elucidate the role of postprandial inflammation in atherogenesis and to find new methods in order to reduce selectively the postprandial inflammatory response. Future studies are needed to find new methods in order to reduce selectively the postprandial inflammatory response.

Postprandial inflammation and endothelial dysfuction

Postprandial hyperlipidaemia is a common metabolic disturbance in atherosclerosis. During the postprandial phase, chylomicrons and their remnants can penetrate the intact endothelium and cause foam cell formation. These particles are highly atherogenic after modification. People in the Western world are non-fasting for most of the day, which consequently leads to a continuous challenge of the endothelium by atherogenic lipoproteins and their remnants. Furthermore, atherosclerosis is considered a low-grade chronic inflammatory disease. Many studies have shown that the process of atherogenesis in part starts with the interaction between the activated leucocytes and activated endothelium. Postprandial lipoproteins can activate leucocytes in the blood and up-regulate the expression of leucocyte adhesion molecules on the endothelium, facilitating adhesion and migration of inflammatory cells into the subendothelial space. Another inflammatory process associated with postprandial lipaemia is the activation of the complement system. Its central component C3 has been associated with obesity, coronary sclerosis, the metabolic syndrome and fasting and postprandial TAGs (triacylglycerols). Moreover, chylomicrons are the strongest stimulators of adipocyte C3 production via activation of the alternative complement cascade. A postprandial C3 increment has been shown in healthy subjects and in patients with CAD (coronary artery disease) and with FCHL (familial combined hyperlipidaemia). Postprandial lipaemia has been related to TAG and free fatty acid metabolism. All of these mechanisms provide an alternative explanation for the atherogenicity of the postprandial period.

Diurnal triglyceride profiles: a novel approach to study triglyceride changes

Fasting plasma triglycerides (TG) show a high intra-individual variability, and therefore, repeated measurements and alternative methodology are necessary when studying TG metabolism. In search for novel approaches to study TG changes, we evaluated the feasibility of determining ambulatory capillary TG. In addition, well-known characteristics (e.g. gender differences) of TG metabolism in healthy subjects were determined. In 18 subjects with a wide range of fasting plasma TG, the results of standardised oral fat loading tests (50 g m(-2)) were compared to their diurnal capillary TG profiles, measured on 3 different days, six times each day in an out-patient clinic setting. The diurnal TG-profile was calculated as area under the capillary TG curve (TGc-AUC) and as incremental area (dTGc-AUC). Clearance of plasma TG after the acute oral fat load correlated well with the diurnal TGc-AUC (r=0.77; P<0.01). In addition, hypertriglyceridemic subjects (plasma TG >2.0 mmol l(-1)) had a higher diurnal triglyceridemia (49.83+/-15.37 h mmol l(-1)) as well as a higher response of plasma TG to the oral fat load (42.10+/-15.37 h mmol l(-1)), than the subjects with normal fasting plasma TG (29.83+/-11.75 h mmol l(-1) (P<0.05) and 20.75+/-5.89 h mmol l(-1) (P<0.01), respectively). In an observational study, 106 volunteers (54 females and 52 males) measured capillary triglycerides. Food intake was recorded and fasting blood was drawn once at the start of the study. Body composition was assessed by anthropometric parameters and body-impedance. Repeated measurements of diurnal triglyceridemia tended to be less variable than fasting capillary triglycerides (mean coefficients of variation 15.1% (range: 0.60-45.9%) and 24.9% (range: 1.44-72.7%), respectively; P=0.09) for the whole group and in males (18.6% (0.60-45.9%) and 24.0% (1.4-58.2%), respectively; P=0.07). The mean diurnal TGc-AUC and dTGc-AUC were lower in females (16.50+/-4.85 and 1.82+/-3.46 h mmol l(-1), respectively) than in males (23.44+/-6.50 and 6.93+/-4.67 h mmol l(-1); P<0.001 for each). The total daily energy intake was lower in females (8911+/-1905 kJ) than in males (11042+/-2604 kJ, P<0.001) because of a lower intake of all nutrients. In females, estrogen status determined significantly the capillary TG profiles. Stepwise multiple regression analysis for females and males, with TGc-AUC as the dependent variable, showed that the best predictors were fasting capillary TG, gender, systolic blood pressure and mean daily energy intake, explaining 72% of the variation. Incremental triglyceridemia was best described by gender, mean daily protein intake and systolic blood pressure, explaining 42% of the variation. Diurnal capillary TG profiles may be used to estimate the total daily load of potential atherogenic particles to which individuals are subjected during the day without the need for metabolic ward studies.

Lipoprotein lipase gene mutations D9N and N291S in four pedigrees with familial combined hyperlipidaemia

Abstract. The role of the lipoprotein lipase (LPL) gene in familial combined hyperlipidaemia (FCH) is unclear at present. We screened a group of 28 probands with familial combined hyperlipidaemia and a group of 91 population controls for two LPL gene mutations. D9N and N291S. LPL‐D9N was found in two probands and one normolipidaemic population control. LPL‐N291S was found in four probands and four population controls. Subsequently, two pedigrees from probands with the D9N mutation and two pedigrees from probands with the N291S mutation were studied, representing a total of 24 subjects. Both LPL gene mutations were associated with a significant effect on plasma lipids and apolipoproteins. Presence of the D9N mutation (n = 7) was associated with hypertriglyceridaemia [2.69± 1.43 (SD) mmol L‐1] and reduced plasma high‐density lipoprotein cholesterol (HDL‐C) concentrations (0.92± 0.21 mmol L‐1) compared with 11 non‐carriers (triglyceride 1.75± 0.64 mmol L‐1; HDL‐C 1.23± 0.30 mmol L‐1, P= 0.03 and P= 0.025 respectively). LPL‐D9N carriers had higher diastolic blood pressures than non‐carriers. LPL‐N291S carriers (n= 6) showed significantly higher (26%) apo B plasma concentrations (174± 26 mg dL‐1) than non‐carriers (138± 26 mg dL‐1; P= 0.023), with normal post‐heparin plasma LPL activities. Linkage analysis revealed no significant relationship between the D9N or N291S LPL gene mutations and the FCH phenotype (hypertriglyceridaemia, hypercholesterolaemia or increased apo B concentrations). It is concluded that the LPL gene did not represent the major single gene causing familial combined hyperlipidaemia in the four pedigrees studied, but that the LPL‐D9N and LPL‐N291S mutations had significant additional effects on lipid and apolipoprotein phenotype.

A proposal to redefine familial combined hyperlipidaemia -- third workshop on FCHL held in Barcelona from 3 to 5 May 2001, during the scientific sessions of the European Society for Clinical Investigation.

Familial combined hyperlipidaemia (FCHL) was described in 1973 by three separate groups as a common familial disorder characterized by multiple lipoprotein phenotypes and an increased risk of premature coronary artery disease [1– 3]. No metabolic explanation was offered for the variable lipid phenotypes and opinion differed as to whether this was likely to be a monogenic or polygenic disorder. In 1986, the first FCHL workshop was held in Seattle and at that meeting an elevated plasma apolipoprotein B (apoB) was added to the list of characteristics. However, it was not made an essential feature, nor was any change to the fundamental approach to phenotypic classification suggested, notwithstanding that complexity of diagnosis severely limits clinical application and comparison of research results [4]. In 1998, investigators met in Helsinki for the second FCHL workshop and heard of the newest efforts to identify the genetic and metabolic bases for FCHL. All of the analyses were presented within the context of the original diagnostic approach. At the most recent meeting of the European Society for Clinical Investigation in Barcelona, the third workshop on FCHL was organized by Dr J. Ribalta (Reus, Spain) and Dr M. Castro Cabezas (Utrecht, the Netherlands) to reconsider this most common, but least well characterized, familial atherogenic dyslipoproteinaemia. Our objective became to search for the most important pathophysiological features. From the outset, as outlined by Professor M-R. Taskinen (Helsinki, Finland) , the two most well-documented features are increased very low-density lipoprotein (VLDL) secretion and impaired clearance of postprandial lipoproteins [5,6]. The increased VLDL2 secretion results in hypertriglyceridaemia and an elevated plasma apoB. Long residence time of VLDL1 particles favour the formation of small dense low-density lipoprotein (LDL). Based on this, we considered the hypothesis that the phenotype of FCHL might not be multiple but unitary – namely, hypertriglyceridaemic (hyperTg) hyperapoB. If so, FCHL phenotype could be defined more simply and consistently as follows. The phenotype of hyperTg hyperapoB would have to be present in more than one family member and at least one individual in the family must have premature symptomatic coronary artery disease. Other genetic disorders and secondary causes of dyslipidaemia, including type 1 and type 2 diabetes would, of course, have to be excluded [4,5]. It was emphasized that such a change only represents an evolution in diagnosis based on the advances in knowledge and technology that have occurred since the disorder was Mike Rosenbloom Laboratory for Cardiovascular Research, McGill University Health Centre, McGill University, Montreal, Quebec, Canada (A. D. Sniderman); Departments of Internal Medicine and Endocrinology, University Medical Centre, Utrecht, the Netherlands (M. Castro Cabezas, D. W. Erkelens); Unitat de Recerca de Lípids i Arteriosclerosi, Facultat de Medicina, Hospital Universitari de Sant Joan, Universitat Rovira i Virgili, Reus, Spain (J. Ribalta, L. Masana); Servicio de Endocrinología, Departamento de Medicina, Universidad de Valencia, Valencia, Spain (R. Carmena, J. T. Real); Departments of Medicine and Endocrinology, Academic Hospital, 6202 AZ Maastricht, the Netherlands (T. W. A. de Bruin); Department General Internal Medicine 541, UMC Nijmegen, the Netherlands (J. de Graaf); Division of Cardiovascular Genetics, Department of Medicine, Royal Free and University College Medical School, London WC1E 6JJ, UK (S. E. Humphries, P. J. Talmud); Department of Medicine, University of Helsinki, Helsinki, Finland (M. R. Taskinen).

Delayed chylomicron remnant clearance in subjects with heterozygous familial hypercholesterolaemia

Castro Cabezas M, De Bruin TWA, Westerveld HE, Meijer E, Erkelens DW (Departments of Internal Medicine and Endocrinology, University Hospital Utrecht and University Hospital Maastricht, The Netherlands). Delayed chylomicron remnant clearance in subjects with heterozygous familial hypercholesterolaemia. J Intern Med 1998; 244: 299–307.

Gender differences in diurnal triglyceridemia in lean and overweight subjects

AIMS: Increased fasting and postprandial triglyceridemia is one of the cardiovascular risk factors for patients with insulin resistance. Since triglyceride (TG) metabolism largely depends on gender, we have investigated diurnal TG changes in patients with and without overweight, focusing on gender differences.METHODS: Twenty-two males and 22 females with overweight (mean body mass index (BMI) 28.0±2.3 kg/m2) measured capillary TG concentrations at six fixed time points on three different days. Diurnal TG profiles were calculated as area under the capillary TG curves (TGc-AUCs). The control group consisted of 24 males and 21 females who were not overweight (mean BMI 22.4±1.5 kg/m2). Biochemical and anthropometric parameters associated with insulin resistance were measured.RESULTS: Lean males and lean females had comparable fasting insulin levels (6.9±2.6 and 8.1±4.7 mU/l, respectively), but females had a more favorable fasting lipoprotein profile when compared to males. Diurnal TG profiles were lower in lean females than in lean males (16.9±4.3 vs 20.3±5.7 mMh, respectively, P<0.05). Overweight males and females had comparable fasting insulin levels (10.3±3.4 and 12.1±4.9 mU/l, respectively), which were higher than in lean subjects. Overweight females also had a more favorable fasting lipoprotein profile compared to overweight males. Diurnal TG profiles were similar in overweight females and overweight males (31.1±15.6 and 32.9±13.2 mMh, respectively). Stepwise multiple regression analysis showed that in both males and females, waist circumference was the strongest determinant of diurnal TG profiles when fasting TG concentrations were excluded from the model (R 2=0.49 for males and R 2=0.33 for females). These results suggest that overweight resulted in a ‘male diurnal TG profile’ in females due to abdominal fat accumulation.CONCLUSION: Insulin resistance in overweight subjects partly mitigates the gender differences of fasting and postprandial TG metabolism. The significant positive association between diurnal triglyceridemia and waist circumference supports the view that especially abdominal fat associated with insulin resistance enhances postprandial lipemia.