Mannose binding lectin deficiency and triglyceride-rich lipoprotein metabolism in normolipidemic subjects.

Mannose binding lectin (MBL) is one of the three initiators of complement activation and is therefore closely linked to inflammation. MBL deficiency has been associated with the generation of atherosclerosis. Since atherosclerosis, the complement system and postprandial lipemia are linked to inflammation, we studied postprandial lipoprotein metabolism in MBL deficiency. An observational study was carried out in 107 volunteers (21% MBL deficient). Classical cardiovascular risk factors were not different between subjects with and without MBL deficiency. Oral fat loading tests in 8 MBL deficient and 14 MBL sufficient subjects showed similar postprandial triglyceride, free fatty acid, hydroxybutyric acid and complement component 3 concentrations. MBL deficient subjects had 2.4 times lower postprandial Sf>400 (chylomicron)-apoB48 concentrations, but in contrast a 2-3.5 times increased Sf 60-400 (VLDL1-TG) and Sf 60-400-apoB100 response. MBL activity was inversely related to the postprandial Sf 60-400-TG increase. Despite lower postprandial Sf>400-apoB48 concentrations, MBL deficient subjects show an accumulation of Sf 60-400 lipoproteins.

Comparison of different methods to investigate postprandial lipaemia.

Postprandial hyperlipidaemia has been associated with coronary artery disease (CAD). We investigated which of the generally used methods to test postprandial lipaemia differentiated best between patients with premature CAD (50+/-4 years, n=20) and healthy controls. Furthermore, the effects of rosuvastatin 40 mg/day on postprandial parameters were assessed. Standardised oral fat-loading tests (OFLT) and ambulant self-measurements of daylong capillary triglycerides (TGc) were performed. Total responses of individual lipoproteins, plasma TG (TGp) and remnant-like particle cholesterol (RLP-C) were estimated as area under the curve (AUC). Most AUCs were highest in untreated patients and reached control levels after rosuvastatin. From all AUCs, RLP-C-AUC was best associated to TGp-AUC in untreated patients and controls (adjusted R2=0.84, beta=0.92, p.

Effects of rosuvastatin on postprandial leukocytes in mildly hyperlipidemic patients with premature coronary sclerosis.

We investigated whether pro-inflammatory aspects of the postprandial phase can be modulated by rosuvastatin in premature coronary artery disease (CAD) patients. Herefore standardized 8 h oral fat loading tests were performed off-treatment and after rosuvastatin 40 mg/d in 20 male CAD patients (50 +/- 4 years). The expression of leukocyte activation markers CD11a, CD11b, CD62L and CD66b was studied using flowcytometry. Migration of isolated neutrophils towards chemoattractants was determined in a fluorescence-based assay. Rosuvastatin did not affect baseline leukocyte counts nor the postprandial neutrophil increment (maximum mean increase +10% pre- and +14% post-treatment, P < 0.01 for each). Rosuvastatin reduced baseline platelets (from 266 +/- 78 to 225 +/- 74 x 10(9) cells/L, P < 0.001) and blunted the postprandial platelet count change (maximum mean increase +6%, P = 0.01, and 0%, respectively). The baseline expression of CD11a, CD11b and CD62L increased on most types of leukocytes by rosuvastatin, whereas the postprandial responses were unaffected. Pretreatment, postprandial neutrophil migration increased dose-dependently, but there were no postprandial changes after rosuvastatin. The latter effect was unrelated to changes in lipoprotein concentrations. In conclusion, in CAD patients postprandial pro-inflammatory and pro-coagulant changes can be modified by rosuvastatin. These apparently lipid-lowering independent effects may render protection against atherosclerosis.

Daylong triglyceridaemia in healthy Mediterranean and northern European subjects.

BACKGROUND: A Mediterranean eating pattern and diet enriched in monounsaturated fatty acids may result in a favourable daylong lipid profile. METHODS: 19 Spanish males (aged 32 +/- 8 years) and 28 females (34 +/- 8 years) were matched to Dutch subjects on the basis of fasting capillary triglycerides (TGc), gender and age. TGc were self-measured at six fixed time points over three days. Daylong TGc profiles were calculated as areas under the curve (TGc-AUC). RESULTS: Anthropometric parameters and fasting plasma lipids were comparable between Spanish participants and Dutch subjects. Insulin sensitivity (expressed as HOMA) was highest in the Dutch females (1.41 +/- 1.09 vs. 2.09 +/- 1.23 in the Spanish females, p < 0.05). Daylong TGc values were not different between Spanish and Dutch participants. Male Spanish subjects showed the largest daylong TGc increase after lunch, while in the Dutch males, the largest TGc increase was seen after dinner. Total daytime dietary energy and total fat intake were comparable when analysed by gender. However, the Spanish participants had a higher intake of monounsaturated and polyunsaturated fatty acids as percentage of energy. CONCLUSION: There are no major differences in daylong triglyceridaemia between Dutch and Spanish subjects, despite different eating habits and a diet enriched in monounsaturated and polyunsaturated fat in the latter.

Activation of leukocytes by postprandial lipemia in healthy volunteers.

Activation of leukocytes is obligatory for inflammation and atherogenesis by adhering to the endothelium via specific ligands. Although in vitro studies have shown that triglycerides (TG) can activate leukocytes, it is unknown whether this occurs in vivo. Using flowcytometry, we studied the expression of leukocyte activation markers CD11A, CD11B, CD62L (all involved in endothelium adhesion) and CD66B (a neutrophil degranulation marker) during a 6 h fat challenge (50 g/m2) and a water test in 10 healthy males (52 +/- 3 years). After fat, neutrophil counts were increased between t=1 and t =6 h, with a maximum at t=3 h (+32% versus t=0, P <0.05), while they remained unchanged after water. Both tests showed gradual lymphocyte count increments. The expression of activation markers on lymphocytes was low and showed comparable responses after both tests. After fat, a significant increase up to a maximum at t=6 h was seen for CD11B on monocytes and on neutrophils for CD11B, CD62L and CD66B. Postprandial activation of monocytes and neutrophils was higher after fat than after water. The maximal postprandial TG increment was significantly related to the increase of CD11B on monocytes (Pearson's R=0.64, P <0.05). In conclusion, postprandially there is a TG-specific increase of neutrophil counts and increased activation of monocytes and neutrophils. These results are suggestive of a pro-inflammatory situation that may correspond with increased adhesive capacity of these cells contributing to the inflammatory component of atherosclerosis.

Increased expression of activation markers on monocytes and neutrophils in type 2 diabetes.

BACKGROUND: Activation of leukocytes is obligatory for adherence to the endothelium and atherogenesis. Since leukocyte activation by triglycerides (TG) and glucose has been described in vitro, we hypothesised higher leukocyte activation in patients with type 2 diabetes. METHODS: Using flow cytometry, we studied the expression of the leukocyte activation markers CD11A, CD11B, CD62L and CD66B in 15 patients with type 2 diabetes without clinical evidence of atherosclerosis (55+/-7 years) and in 15 healthy controls (53+/-2 years). All patients were on oral antidiabetic treatment (glyHb 6.3+/-0.9%) and not taking statins or anti-inflammatory drugs. RESULTS: In comparison with controls, the patients had a higher waist circumference (1.08+/-0.09 vs 0.94+/-0.11 m, p<0.005) and higher fasting glucose (8.4+/-2.3 vs 5.3+/-0.7 mM, p<0.005), whereas fasting plasma lipids were not statistically different. The leukocyte count was higher in the patients (6.55+/-1.55 vs 5.07+/-1.10 x 10(9) cells/l, p<0.005) due to higher neutrophils and lymphocytes (+34% and +24%, p<0.05 for each). CD11B on monocytes and CD11B and CD66B on neutrophils were higher in the patients (+30%, +52% and +43%, P<0.05 for each). Fasting glucose, waist circumference, body mass index and systolic blood pressure were positively associated with the leukocyte and neutrophil count. The expressions of CD11B and CD66B on monocytes and neutrophils were strongly positively interrelated, but unrelated to TG and glucose. CONCLUSION: In patients with type 2 diabetes, the expression of activation markers on monocytes and neutrophils is enhanced and not correlated to fasting glucose or TG. These results suggest a proinflammatory situation in type 2 diabetes and most likely represent increased adhesive capacity of neutrophils and monocytes to the endothelium.

Postprandial leukocyte increase in healthy subjects.

Atherosclerosis is an inflammatory disorder involving leukocytes and lipids. To study the relationship between leukocytes and lipids in vivo, leukocyte changes were determined in 14 healthy males (age, 23 +/- 3 years; body mass index [BMI], 21.9 +/- 1.5 kg/m(2)) after an 8-hour oral fat load (50 g/m(2)) and after water. The postprandial triglyceride (TG) increment after fat was paralleled by a leukocyte increment, due to an increase in neutrophils in the first 2 hours (142% +/- 69% higher than baseline, P =.04). Neutrophil counts did not return to baseline at the end of the test. Water ingestion did not induce significant neutrophil changes. Blood lymphocytes increased gradually in both tests (142% +/- 30% higher than baseline, P <.001 after fat, and 128% +/- 36%, P =.02 after water). The total leukocyte increment after fat ingestion was related to the postprandial TG increase (Spearman's r = 0.73, P =.003). An early postprandial, lipid-specific, neutrophil increment is a new characteristic of the postprandial phase. Future studies will elucidate the role of postprandial leukocyte changes in the pathogenesis of atherosclerosis.

Postprandial recruitment of neutrophils may contribute to endothelial dysfunction.

Atherosclerosis is a low-grade inflammatory disease involving leukocytes, lipids, and glucose leading to endothelial dysfunction. Since activation of neutrophils by triglycerides and glucose has been described in vitro, we hypothesized that the postprandial phase is an inflammatory state affecting leukocytes, possibly contributing to endothelial dysfunction. We measured postprandial blood leukocyte counts, cytokines, hydroperoxides (HPOs), and flow-mediated vasodilation (FMD) in eight healthy males (age 23 +/- 2 years) after a FAT (50 g/m2) and GLUCOSE challenge (37.5 g/m2), a combination of both (MIXED test), and after WATER. All tests, except WATER, resulted in significantly impaired FMD (10% reduction) between t = 1 h and t = 3 h, accompanied by a significant increase of neutrophils (59% after FAT and 28% after GLUCOSE and MIXED), total plasma HPOs (15 to 31% increase), and plasma interleukin-8 (IL-8) (50-130% increase). WATER did not affect FMD, neutrophils, HPOs, or IL-8. Lymphocytes increased gradually in all tests (40-70% increase at t = 10 h compared with t = 0; P < 0.005), paralleling a gradual 3- to 5-fold interleukin-6 increase. Monocyte and erythrocyte counts did not change in any test. In conclusion, the neutrophil increment during postprandial lipemia and glycemia with concomitant IL-8 and HPO increases may contribute to endothelial dysfunction. Lymphocyte increment is a nonspecific diurnal process. Postprandial intravascular inflammatory changes may be relevant for the pathogenesis of atherosclerosis.

Insulin resistance and vessel endothelial function.

IRS is a complex disease consisting of a clustering of metabolic disorders, of which hyperglycaemia, hyper-insulinaemia and dyslipidaemia are the most important. Endothelial dysfunction plays an important role in the pathogenesis of atherosclerosis. The effects of hyperinsulinaemia seem to depend on lipidaemia and glycaemia. Hyperglycaemia and hyperlipidaemia have detrimental effects on endothelial function in the fasting as well as the postprandial states. In both situations, the generation of ROS and vasoactive molecules plays a major role in interfering with the atheroprotective endothelium-dependent NO system. Treatment of IRS in regard to endothelial function should be focused initially on lifestyle improvement, such as stopping smoking and eating a balanced diet containing antioxidant vitamins, folic-acid, L-arginine and long-chain omega-3 unsaturated FA. Strict glucose control has shown to improve endothelial function and decrease microvascular complications. However, macrovascular complications, in line with endothelial functional improvement, have so far been reduced only when treatment was focused on other characteristics of the IRS syndrome, in particular dyslipidaemia. Other relevant treatments include ACE inhibitors and thiazolidinediones, and probably tetrahydrobiopterin and folic acid supplementation. Future studies should address the effects of therapeutic neovascularization on endothelial dysfunction.

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.

Diurnal triglyceride profiles in healthy normolipidemic male subjects are associated to insulin sensitivity, body composition and diet.

BACKGROUND: Elevated fasting and postprandial triglycerides (TG) are established risk factors for Coronary Heart Disease (CHD). Usually, fasting plasma TG are measured, although TG are mainly produced in a postprandial state. Our objective was to investigate diurnal TG profiles using serial capillary TG measurements, in normolipidemic healthy males. MATERIALS AND METHODS: Forty-eight, non-obese, non-smoking males (range: 20-55 years, mean age: 32 +/- 12 years), measured diurnal capillary TG, at six fixed timepoints during the day on three different days and recorded their food intake. Insulin sensitivity was estimated by HOMA. Diurnal capillary TG profiles were calculated as integrated area under the mean capillary TG curve (TGc-AUC). RESULTS: All subjects had normal fasting plasma TG and cholesterol. The average TGc-AUC was 23.6 +/- 6.7 mmol h L-1. Significant correlations with TGc-AUC were: fasting insulin (r = 0. 40, P < 0.005), HOMA (r = 0.32, P < 0.05), relative fat mass (r = 0. 31, P < 0.05), dietary protein-(r = 0.31, P < 0.05) and saturated fat intake (r = 0.30, P < 0.05). Age was not associated to diurnal triglyceridemia. After subdividing the group into quartiles on the base of TGc-AUC, differences were found between the highest (n = 12) and lowest quartile (n = 12) for: fasting capillary TG, fasting insulin, HOMA and systolic blood pressure. Fasting plasma TG and dietary intake were not different. CONCLUSION: Diurnal TG profiles in healthy normolipidemic males are not age-dependent, but are associated to insulin sensitivity, fat mass and diet. Diurnal capillary TG profiles may be a valuable additional tool in estimating a risk profile for CHD since significant differences in diurnal TG are not always reflected by elevated fasting plasma TG.

[Diurnal triglyceride profiles in 30 young healthy men as a function of diet, fasting triglyceride levels, body composition and insulin sensitivity]. / Triglyceridedagprofielen bij 30 jonge gezonde mannen als functie van voeding, nuchtere triglycerideconcentraties, lichaamssamenstelling en insulinegevoeligheid.

OBJECTIVE: To study predictors of diurnal capillary triglyceride (TG-c) profiles in healthy males. DESIGN: Observational, cross-sectional. SETTING: University Hospital Utrecht, Department of Internal Medicine, the Netherlands. METHOD: In 30 healthy males (20-34 years) TG-c was measured during three days, six times a day. Fasting blood was collected at inclusion. Body composition and HOMA ratio as insulin sensitivity index were determined. TG-c profiles were calculated as integrated area under the mean TG-c curve (TG-AUC). RESULTS: All subjects had normal fasting plasma and capillary TG and cholesterol concentrations. Diurnal TG-c values were higher than fasting values. The average TG-AUC was 24.6 +/- 6.7 mmol/l over 14 hrs. Variables associated with TG-AUC were: fasting TG-c, relative fat mass, total protein and saturated fat intake. After correction for fasting TG-c only diet and fat mass were correlated with TG-AUC. The relative fat mass was positively correlated with the HOMA ratio and fasting insulin concentrations, suggesting that decreased insulin sensitivity accompanied increased body fat. CONCLUSION: Triglyceride profiles provide information about the total diurnal TG load. The best determinant of diurnal triglyceride changes was fasting triglycerides. However, diet, body composition and insulin sensitivity are also important. Future investigations should address the question whether triglyceride profiles may be used to estimate more accurately the individual risk profile for coronary heart disease.