Juhani Kahri

Cholesterol efflux from Fu5AH hepatoma cells induced by plasma of subjects with or without coronary artery disease and non-insulin-dependent diabetes: importance of LpA-I:A-II particles and phospholipid transfer protein

We measured the capacity of human plasma to induce cholesterol efflux from Fu5AH rat hepatoma cells in four groups of men with or without non-insulin-dependent diabetes mellitus (NIDDM) and coronary artery disease (CAD). Plasma from men with both NIDDM and CAD (n = 47) had the lowest efflux capacity (17.3 +/- 3.6%) whereas healthy control subjects with neither diabetes nor CAD (n = 25) had the highest capacity (19.8 +/- 3.4%). The groups with CAD but no diabetes (n = 44) and with NIDDM but no CAD (n = 35) had intermediate efflux values (18.5 +/- 3.8 and 18.5 +/- 3.9%, respectively). In a 2 x 2 factorial ANOVA, the differences were significant with respect to the presence of CAD (P = 0.038) and NIDDM (P = 0.041), with no interaction between the factors. The concentration of HDL particles containing apolipoprotein (apo) A-I but no apo A-II (LpA-I) was not related to efflux capacity in univariate or multivariate analyses. A multivariate regression analysis showed that when controlled for the presence of NIDDM and CAD, the concentration of particles containing both apo A-I and apo A-II (LpA-I:A-II) and plasma phospholipid transfer protein activity were both positively, independently, and significantly (P < 0.001) related to cholesterol efflux capacity.

Regulation of low-density lipoprotein particle size distribution in NIDDM and coronary disease: importance of serum triglycerides

SummaryAn increase of low-density lipoprotein triglycerides (LDL-Tg) was found to be an independent coronary artery disease (CAD) risk factor for non-insulin-dependent diabetic (NIDDM) patients in a recent prospective study. We examined the composition and size of LDL particles in 50 NIDDM men with angiographically verified CAD (NIDDM+ CAD+) and in 50 NIDDM men without CAD (NIDDM+ CAD−) as compared to 50 non-diabetic men with CAD (NIDDM− CAD+) and 31 non-diabetic men without CAD (NIDDM− CAD−). The groups had similar ranges of age and BMI LDL particle size was determined by gradient gel electrophoresis, and LDL was isolated by sequential ultracentrifugation for compositional analyses. Serum Tg was increased in NIDDM patients as compared to non-diabetic subjects (p<0.05), and in patients with CAD as compared to subjects without the disease (p<0.05). LDL cholesterol was lower in NIDDM patients than in non-diabetic subjects (p<0.001). Mean diameter of LDL particles was less than 255 å, but closely comparable in all groups. The presence of NIDDM was associated with increases of Tg and protein but lowering of free cholesterol in LDL (p<0.005 for all). In multivariate regression analyses neither NIDDM nor CAD were associated with LDL particle size, but serum Tg was the major determinant of LDL size in both NIDDM and non-diabetic subjects (p<0.001). When the patients were divided into quartiles according to fasting serum Tg levels, the LDL particle size and free cholesterol content decreased, but Tg and protein contents of LDL particles increased from the lowest to the highest Tg quartile (analysis of variance p<0.001 for all). When the subjects were categorized into two groups according to the median of VLDL-Tg (1.10 mmol/l) LDL size was associated with VLDL-Tg in the high but not in the low VLDL-Tg group. We conclude that in NIDDM patients with or without CAD serum Tg is the major determinant of the properties of LDL particles. The clinical implication is that in NIDDM serum Tg should be as low as possible to prevent atherogenic changes in LDL.

Dual Metabolic Defects Are Required to Produce Hypertriglyceridemia in Obese Subjects

Objective— Obesity increases the risk of cardiovascular disease and premature death. However, not all obese subjects develop the metabolic abnormalities associated with obesity. The aim of this study was to clarify the mechanisms that induce dyslipidemia in obese subjects. Methods and Results— Stable isotope tracers were used to elucidate the pathophysiology of the dyslipidemia in hypertriglyceridemic (n=14) and normotriglyceridemic (n=14) obese men (with comparable body mass index and visceral fat volume) and in normotriglyceridemic nonobese men (n=10). Liver fat was determined using proton magnetic resonance spectroscopy, and subcutaneous abdominal and visceral fat were measured by magnetic resonance imaging. Serum triglycerides in obese subjects were increased by the combination of increased secretion and severely impaired clearance of triglyceride-rich very-low-density lipoprotein1 particles. Furthermore, increased liver and subcutaneous abdominal fat were linked to increased secretion of very-low-density lipoprotein 1 particles, whereas increased plasma levels of apolipoprotein C-III were associated with impaired clearance in obese hypertriglyceridemic subjects. Conclusion— Dual metabolic defects are required to produce hypertriglyceridemia in obese subjects with similar levels of visceral adiposity. The results emphasize the clinical importance of assessing hypertriglyceridemic waist in obese subjects to identify subjects at high cardiometabolic risk.

HDLs Containing Apolipoproteins A-I and A-II (LpA-I:A-II) as Markers of Coronary Artery Disease in Men With Non–Insulin- Dependent Diabetes Mellitus

BACKGROUND Abnormalities in HDL and an increased risk of coronary artery disease (CAD) coexist in non-insulin-dependent diabetes mellitus (NIDDM). HDLs can be separated by their apolipoprotein (apo) content into particles containing apoA-I but not apoA-II (LpA-I) and those containing both apoA-I and apoA-II (LpA-I:A-II). The LpA-I particles have been suggested to be more effective in conferring protection against CAD than the LpA-I:A-II particles. However, data are sparse, and no studies have defined the role of these two classes of particles in NIDDM. METHODS AND RESULTS LpA-I and LpA-I:A-II particles were quantified by a differential electroimmunoassay in four groups of men with similar age and body mass index (BMI) distributions. Group 1 consisted of 50 patients with NIDDM and angiographically verified CAD; group 2, 50 men with CAD but no diabetes; group 3, 50 men with NIDDM but no CAD; and group 4, 31 healthy men. Serum apoA-I and apoA-II concentrations were measured by immunoturbidimetry, and HDL2 and HDL3 were separated by ultracentrifugation. Concentrations of LpA-I:A-II particles in group 1 were 13.8%, 18.3%, and 26.9% lower than in groups 2 through 4, respectively. In a two-by-two factorial ANOVA, adjusted for age and BMI, the differences were significant for both CAD (P < .001) and NIDDM (P < .001), with no interaction between the factors. These results were confirmed by comparable differences in the serum concentrations of apoA-I and apoA-II. LpA-I particles were related to the presence or absence of CAD (P = .013), but the difference was lost in a multivariate analysis. A low HDL3 cholesterol concentration characterized both CAD (P = .002) and NIDDM (P = .024). HDL2 cholesterol differed significantly with regard to the presence of NIDDM (P = .033) but only borderline with respect to CAD (P = .073). CONCLUSIONS ApoA-II-containing lipoproteins and HDL3 cholesterol are powerful markers of CAD in men with NIDDM.

High density lipoprotein subfractions, apolipoprotein A‐I containing lipoproteins, lipoprotein (a), and cholesteryl ester transfer protein activity in alcoholic women before and after ethanol withdrawal

Abstract. We studied 11 female alcoholics before and after ethanol withdrawal of 2 weeks and 10 healthy normolipidaemic, nonalcoholic women of similar age. In alcoholic women the HDL2 mass was increased by 63% (P<0.01) on admission and normalized (P<0.01) during abstention. The concentrations of HDL3 cholesterol and its mass remained unchanged throughout the study. Consistently with the fall of HDL2 gradient gel electrophoresis analyses also demonstrated decrease of the cholesterol concentration of HDL2b and HDL2a (P<0.05) during alcohol withdrawal. On admission the apo A‐II concentration was increased by 48% (P < 0.01) and it was normalized (P< 0.001) during abstention. Among apo A‐I containing lipoproteins the most prominent change occurred in Lp A‐1: A‐11, which fell by 32% (P<0.01) during 1 weeks alcohol withdrawal. During abstention the lipoprotein (a) concentration increased in 10 out of 11 women. In patients cholesteryl ester transfer (CETP) activity increased by 35% (P<0.01) during 1 week of ethanol withdrawal. On admission post‐heparin plasma lipoprotein (LPL) and hepatic lipase activities were increased by 25% (P = NS); during 1 weeks abstention they both returned to the control level (P < 0.05–< 0.01). In conclusion, chronic alcoholic women display multiple changes of lipoprotein metabolism which are rapidly reversed during abstinence. In contrast to alcoholic men, studied previously by us using the same study design and methods, there was no significant elevation of HDL3 cholesterol and apo A‐I. The data suggest that alcohol interferes with several regulatory steps of HDL metabolism which are partly gender dependent.

ApoA-IHelsinki (Lys107→0) Associated With Reduced HDL Cholesterol and LpA-I:A-II Deficiency

A Finnish kindred with premature coronary heart disease and decreased HDL cholesterol levels was identified as having an apoA-I variant, apoA-I (Lys107-->0), caused by a 3-bp deletion of nucleotides 1396 through 1398 in exon 4 of the apoA-I gene. These subjects (n = 10) were heterozygous for this mutation. The mean serum HDL cholesterol concentration (26.7 +/- 9.7 mg/dL) of affected family members was 36%, lower than that of unaffected family members (P < .05). Mean serum apoA-I and apoA-II concentrations in heterozygotes were reduced by 18% and 22%, respectively, compared with normal family members (P < .05). In heterozygotes the mean concentration of lipoprotein containing both apoA-I and apoA-II (LpA-I:A-II) was 31% lower than in those with normal apoA-I (P < .001), while the mean level of lipoproteins containing apoA-I without apoA-II was similar in the two groups. HDL density-gradient ultracentrifugation showed a lack of HDL2 and small dense HDL3 in heterozygotes compared with unaffected family members. The HDL particle size distribution, as analyzed by nondenaturing gradient gel electrophoresis of heterozygotes, revealed one major peak at 8.0 to 9.7 nm, a minor peak at 7.8 to 8.5 nm, and an absence of HDL2b and HDL2a peaks. These latter peaks were observed in unaffected family members. Serum levels of LDL cholesterol, triglycerides, VLDL, IDL, and LDL subclasses were similar in the two groups. However, in heterozygotes the cholesterol-to-triglyceride ratios in VLDL2, LDL1, LDL3, HDL2b, HDL2a, and HDL3a were 8% to 54% lower than in unaffected family members (P < .05). Cholesteryl ester transfer protein activity in heterozygotes was reduced by 25% compared with unaffected family members (P < .05), while the plasma lecithin:cholesterol acyltransferase (LCAT) activity did not differ between heterozygotes and unaffected family members. The ability of isolated variant apoA-I to serve as a cofactor for LCAT in vitro did not differ from that of normal apoA-I. Our data are consistent with the concept that a low HDL cholesterol level in subjects heterozygous for the apoA-IHelsinki mutation (Lys107-->0) having normal LCAT activity is a consequence of decreased concentration of LpA-I:A-II particles and of a smaller size and reduced cholesterol content of HDL particles.

Regulation of Apolipoprotein A-I–Containing Lipoproteins in IDDM

In IDDM patients, serum high-density lipoprotein cholesterol concentrations have been reported to be normal or elevated. The spectrum of high-density lipoprotein particles is highly heterogeneous, but no data are available on the subpopulations of high-density lipoprotein in IDDM. We, therefore, studied the spectrum of high-density lipoprotein particles in 86 IDDM patients (51 men and 35 women) 37 ± 10 yr of age and in 74 sex-, age-, and body mass index-matched healthy nondiabetic subjects. The concentrations of high-density lipoprotein and HDL2 cholesterol were higher in the IDDM group than in the control subjects (P < 0.01). The apoA-I-to-apoA-II ratio was higher in the IDDM patients than in the nondiabetic subjects (P < 0.001) because of an increased concentration of LpA-I particles (61 ± 17 vs. 53 ± 15, P < 0.01). LpA-I particles correlated positively with high-density lipoprotein and HDL2 cholesterol in the two groups. Postheparin plasma lipoprotein lipase activity was significantly higher in the IDDM group than in the control group (P < 0.001), whereas postheparin plasma hepatic lipase activities were similar in both groups. Plasma cholesteryl ester transfer protein activity was estimated in an in vitro isotopic assay using exogenous labeled donor (low-density) and acceptor (high-density) lipoproteins in the absence of native lipoproteins. We observed no difference in cholesteryl ester transfer protein activity between the groups, and no significant correlations existed between cholesteryl ester transfer protein activity and high-density lipoprotein subpopulations. A positive correlation existed between HDL2 cholesterol and lipoprotein lipase-to-hepatic lipase ratio in IDDM patients (r = 0.35, P = 0.001) and in nondiabetic subjects (r = 0.55, P < 0.001). A positive correlation existed between LpA-I particles and lipoprotein lipase-to-hepatic lipase ratio in both groups (r = 0.34, P < 0.01; r = 0.38, P < 0.01, respectively) suggesting that lipolytic enzymes participate in the regulation of the metabolism of LpA-I particles. In conclusion, the elevation of high-density lipoprotein cholesterol in IDDM patients is mainly caused by a rise of LpA-I particles, which are suggested to have a key role in reverse cholesterol transport.

Effect of gemfibrozil on high density lipoprotein subspecies in non-insulin dependent diabetes mellitus. Relations to lipolytic enzymes and to the cholesteryl ester transfer protein activity

Twenty patients (18 men, 2 women) with non-insulin dependent diabetes mellitus (NIDDM) were randomized to receive either gemfibrozil 1200 mg daily or placebo for 3 months in a double-blind study. The effect of gemfibrozil on plasma HDL subfraction distribution was studied with sequential and density gradient ultracentrifugation and in gradient gel electrophoresis. The concentrations of apo A-I, apo A-II, Lp A-I and Lp A-I:A-II particles were measured. Postheparin plasma lipoprotein lipase (LPL) and hepatic lipase (HL) activities and plasma cholesteryl ester transfer protein (CETP) activities were also determined. Gemfibrozil increased the concentration of HDL cholesterol (P < 0.01), which was due to the rise of HDL3 cholesterol (+16%), while in the placebo group these values remained unchanged. Gemfibrozil increased the concentrations of apo A-I(+12.6%, NS), apo A-II (+28.2%, P < 0.01) and Lp A-I:A-II particles (+21.6%, P < 0.06) but there were no changes in the placebo group. Neither gemfibrozil nor placebo had any effect on the concentration of Lp A-I particles. As determined by density-gradient ultracentrifugation, gemfibrozil increased the concentration of cholesterol in the most dense HDL fractions (mean density 1.193 g/ml, +22%, P < 0.05 and mean density 1.158 g/ml, +19.3%, P < 0.05). In gradient gel electrophoresis, the gemfibrozil-induced elevations of the cholesterol and protein were most pronounced in the HDL3a (8.8-8.2 nm) region. Gemfibrozil increased LPL and HL activities by 14.7% (P < 0.05) and by 18.8% (P < 0.01), respectively, while in the placebo group LPL and HL activities remained unchanged. Plasma CETP activity was also increased during gemfibrozil treatment while in the placebo group it remained unchanged. We conclude that gemfibrozil causes multiple changes in plasma HDL metabolism. The gemfibrozil-induced elevation of HDL3 and dense HDL subpopulations may reflect the concerted action of LPL, HL and CETP on plasma HDL metabolism.

Plasma Cholesteryl Ester Transfer Protein and Its Relationship to Plasma Lipoproteins and Apolipoprotein A-I-Containing Lipoproteins in IDDM Patients With Microalbuminuria and Clinical Nephropathy

OBJECTIVE To study the distribution of high-density lipoprotein (HDL) subclasses in insulindependent diabetes mellitus (IDDM) patients with nephropathy and factors involved in the regulation of HDL, including plasma cholesteryl ester transfer protein (CETP) and postheparin plasma lipoprotein lipase (LPL) and hepatic lipase (HL) activities. RESEARCH DESIGN AND METHODS Participants included 52 microalbuminuric IDDM patients (with a urinary albumin excretion rate [UAER] of 20–200 μg/min), 37 macroalbuminuric IDDM patients (UAER >200 μg/min), and 64 normoalbuminuric IDDM patients (UAER <20 μg/min). Groups were matched for age, body mass index, duration of diabetes, and glycemic control (HbA1). RESULTS Median concentrations of HDL and HDL2 cholesterol were 11.6 (P = 0.01) and 22.7% (P = 0.01) less in microalbuminuric patients and 5.1 and 15.5% less in macroalbuminuric patients compared with normoalbuminuric patients. No significant differences were observed in the concentrations of apoA-I, apoA-II (apolipoprotein) or LpA-I or LpA-I:A-II (lipoprotein) particles between the groups. HDL cholesterol: apoA-I + apoA-II ratio was significantly lower in micro- (19.7 ± 4.2 (± SD); P < 0.01) and macroalbuminuric patients (20.0 ± 3.7, P < 0.05) than in normoalbuminuric patients (22.1 ± 4.4). Postheparin plasma LPL:IIL ratio was lower in microalbuminuric patients compared with normoalbuminuric patients (1.65 vs. 1.05 [median], P < 0.01). Plasma CETP activity was higher in the macroalbuminuric patients than in micro- (P < 0.05) and normoalbuminuric patients (P < 0.05) but did not correlate with HDL, HDL2, or HDL, cholesterol. LPLHL ratio correlated positively with HDL cholesterol (r = 0.372, P < 0.001), HDL2 cholesterol (r = 0.413, P < 0.001) and with LpA-I particles (r = 0.355, P < 0.001) but not with LpA-I:A-II particles (r = –0.065, NS). CONCLUSIONS IDDM patients with micro- and macroalbuminuria show only trivial changes in concentrations of different HDL parameters, which cannot explain the excess risk of coronary heart disease in these patients. Data also indicate that elevation of CETP activity in IDDM patients with nephropathy is probably not responsible for the lowering of HDL cholesterol.

Impact of postprandial lipaemia on low-density lipoprotein (LDL) size and oxidized LDL in patients with coronary artery disease

Background  Remnant lipoprotein particles (RLPs) and oxidative stress are components of postprandial state. We investigated the concentrations of triglyceride‐rich lipoproteins (TRLs), RLPs, low‐density lipoprotein (LDL) size, and oxidized LDL (oxLDL) during alimentary lipaemia, and evaluated whether changes among these variables could be associated with the severity and extent of coronary artery disease (CAD).