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Uptake of aggregated low density lipoproteins by vascular smooth muscle cells
Wednesday 12 October 1994: Poster Abstracts Lipoproteins
222
similar size to IDL and LDL (fraction II), and large HDL (fraction III). Accurate quantitation of apo E in these fractions has been difficult, however, since fractions do not coincide with classical ultracentrifugal density cuts and a large proportion of lipoproteinbound apo E is removed by ultracentrifugation. In the present study, we have determined the plasma lipoprotein distribution of apo E in normolipidemic (group A, n = 6), hypercholesterolemic (type IIa, group B, II = 8), mixed hyperlipidemic (type IIb, group C, n = 8) and hypertriglyceridemic (type IV, group D, n =7) subjects after an overnight fast. Plasma lipoproteins were separated by fast protein liquid chromatography (FPLC) on a Superose 6 column and apo E was determined by a polyclonal antibody ELISA assay. The total plasma apo E concentrations were 3.6 f 0.3, 4.6 f 0.3, 7.7 + 1.1, and 7.4 & 0.8 mg/dl (mean f SE) for groups A-D respectively. In the normolipidemic subjects, 6.8, 40.1 and 53.1% of total plasma apo E was contained within fractions I, II and III. A greater proportion of plasma apo E was found in TRL (6.8, 11.7, 33.8, 49.9%) and a smaller proportion in HDL (53.1, 33.1, 22.1, 15.3%) as TRL triglyceride increased (0.55 f 0.08, 0.91 f 0.18, 2.73 kO.28. 4.89 i0.61 mmol/l) for zrouos A-D. respectively, Importantly, remnant lipoproteins (fra&on II) were responsible for transporting a significant proportion of plasma apo E in all subjects (40.1, 55.1, 44.1, 34.9%), representing mean plasma ‘remnant’ apo E concentrations of 1.5 f 0.2, 2.6 + 0.2, 3.4 + 0.5 and 2.6 + 0.3 mg/dl, respectively. We conclude that irrespective of a patient’s plasma tiglyceride concentration, a significant proportion of plasma apo E is associated with remnant-sized lipoproteins. Elevated LDL may contribute to increased requirement for beparin in patients with acute myocardial infarction g&ski 1, Sbnecka A, Flasi6ski J, Wasilewska A, Winnicka A, Kurek P, Dmochowska W, Inst. of Maritime and Tropical Med.,
1
Gdynia, Poland
LDL are believed to interfere with heparin and antithrombin III (AT III) activity, so that an increased dose of heparin may be necessary in patients with unstable angina or acute MI and a high plasma LDL concentration. The aim of our study was to compare the dose of heparin required to maintain the activated partial thromboplastin time (APTT) ratio within the range 2.0-2.5 in angina-AM1 patients with normal (A) or high (B) plasma HDL-C, to compare AT III activity in groups A and B, and to correlate the required dose of heparin with plasma LDL-C and AT III activity. 18 patients (A) with LDL-C 149 +_20 mg/dl (i.e. normal, <160 mg/dl) required 29 000 * 3900 units of heparin/24 h; their AT III activity was 12.9 +_1.9 ng/ml (normal range O-15 ng/ml). For group B (LDL-C 224 + 30 ng/dl) the heparin need was 37 500 + 5400 units/24 h (P < 0.05) and AT III 11.9 f 1.7 ng/ml (difference ns). The only significant correlation found (r = 0.659, P < 0.02) was between LDL-C and heparin requirement in group B. Con&sions Patients with high LDL-C require an increased dose of heparin if a desirable APTT ratio is to be preserved; LDL C correlated directly with this dose; and normo- and hypercholesterolemic patients did not differ in AT III activity.
taken up and metabolized to retinoic acid in bone marrow-derived cells. Retinoic acid is a ligand to nuclear receptors and has an important role in the regulation of many genes. Chylomicron remnants were made from mesenteric lymph of rats given [‘HIretin through a duodenal catheter. Retinoids were determined by HPLC and scintillation counting of elution fiattions. At 4°C the cell-associated radioactivity reached a maximum of 2.6% of added dose/mg cell protein after 1 h of incubation. During further incubation for 3 h at 37”C, 80% of the retinyl ester was hydrolyzed to retinol and about 25% of the [3H]retinol was secreted to the medium during the same period. The hydrolysis of retinyl esters was partly inhibited by monensin, chloroquine and brefeldlin A. The amount of [3H]retinol taken up by the cells during 5 h of incubation at 37°C was increased 4-fold by adding 1 ,@ml lipoprotein lipase and decreased by 80% by inhibiting proteoglycan synthesis. By incubating the cells 5 and 24 h at 37°C we were able to detect a small amount of all-trans [3H]retinoic acid. More polar metabolites, not yet identified, were also detected in the cells. In the medium about 50% of the radioactivity was found as polar metabolites after 24 h. (1621 LDL subclass patterns and Syndrome X elements Hartwich J, Baczydska E, Zdzienicka A, Pajqk A, Goldsztajn P, .I .I s, Dept. of Clin. Biochem., Sch. of Publ. Health, Colleg&m Meicum, Jagiellonian Krakhw, Koperniko 15a, Poland
Univ. of Cracow,
31-501
Heterogeneity of LDL particles can be described by two distinct patterns denoted A and B. Pattern A is characterized by a predominance of large LDL, and pattern B by a predominance of small, dense LDL. Pattern B of LDL is associated with increased risk of CHD and with an atherogenic lipoprotein profile. Since dyslipidemia is one of the signs of Syndrome X we asked which of the LDL subclass patterns may be related to this phenomenon. Observations were carried out on the participants of the PolMONICA Cracow Study. 139 non-diabetic men 45-68 years old were divided into subgroups with LDL subclass pattern A (70%) or LDL subclass pattern B (30%). The LDL subclass pattern was photographed after density gradient ultracentrifugation of stained plasma and quantified by densitometric scanning of the slides. Fasting glucose and insulin, SUMGLUC, SUMINS (endogenous insulin response during oral glucose tolerance test) were significantly higher in subjects with LDL subclass pattern B. The frequencies of Syndrome X symptoms according to LDL subclass pattern (percent of subjects in subgroup) were as follows: hyperinsulinemia in 60% of the patients in group B vs 41% in group A, hypertension 48.7% vs 27.8%. obesity 57% vs 35% and impaired glucose tolerance 33% vs 16.5% respectively. In patients with LDL subclass pattern B we observed higher plasma apo B and triglyceride concentrations, whereas significantly lower were HDL-C concentration, plasma apo A-I and LDL free cholesterol concentrations. LDL patternB is, therefore, associated with the abnormalities described as Syndrome X. [1631 WE,
Uptake of aggregated low density lipoproteins by vascular smooth muscle cells Brown SA, Klemp KF, Guyton JR, Dept. of Med., Box
3510, Duke Univ. Med. Center, Durham, NC 27710, USA
Retinoic acid is formed when retinyl esters (vitamin A) are given to J 774 macrophages in chylomicron remnants SE, Halvorsen B, Nenseter MS, Blomhoff R, Nomm KR, Inst. of Nutrition, Rex Sch. of Med., Univ. qf Oslo, PO Box 1046,
(161)
Blind&,
0316, Oslo, Norway
The objective of this study was to show that vitamin A given physiologically as retinyl esters in chylomicron remnants can be
When human LDL are aggregated by vortexing, particle fusion and phase separation of lipid domains occurs, producing lipid droplets and vesicles. The resulting lipid structures bear a strong resemblance to extracellular lipid deposits in human atherosclerosis (Guyton et al. J Lipid Res 1991; 32: 953-962). Macmphages take up aggregated LDL (agLDL) by phagocytosis (Khoo et al Arteriosclerosis 1988; 8: 348-358), but the interaction of agLDL with vascular smooth muscle cells (SMC) is not known.
Atherosclerosis X, Montreal, October I994
Wednesday 12 October 1994: Poster Abstracts HDL and reverse cholesterol transport
AgLDL in concentrations of lO-5O,~g/ml were applied to cultures of 4th to 10th passage swine aortic smooth muscle cells. To facilitate interaction with cells, agLDL were briefly homogenized. Witbin 2 days, agLDL were observed to cluster on the surface of SMC. By 3 or more days, agLDL were internalized in large lysosomes, as confirmed by fluorescent labeling with diI and identification of lysosomes with acridine orange. Electron microscopy with the use of the osmium-tannic acid-paraphenylenediamine procedure to preserve lipid structure and of acid phosphatase to identify lysosomes likewise showed large lysosomes (up to 16 per cell profile, producing a ‘foamy’ appearance). However, cytoplasmic lipid droplets were not evident by electron microscopy or by Nile red fluorescent staining. lz51labeling of LDL prior to aggregation allowed the demonstration of protein degradation after the application of agLDL to SMC. However, degradation rates were always higher for native LDL than for agLDL. I*?-agLDL degradation was partially inhibited by lO,ug/ml cytochalasin B, but because of its slow toxicity this inhibitor of phagocytosis could not be applied to the cells for sufficient time to follow agLDL ingestion fully. The results suggest a novel mechanism for uptake of agLDL by SMC, which may be associated with Iysosomal dysfunction. The lipid profile of healthy individuals and coronary patients in Israel. The Betibrate Infarction Prevention @UP) Study and the Israel MONICA Study w, Graff E, Benderly M, Reicher-Reiss H, Behar S, for the BIP Study Group, Neufeld Cardiac Res. Inst., Sheba Med.
1
Center, Tel-Hashomer, Israel
During 1991-1992, we compared the lipid profiles of healthy volunteers (324 M, 494 F) in the Israel MONICA project with those of age-matched (45-74 years) patients (323 M, 449 F) with established coronary disease (6 months to 5 years after acute myocardial infarction and/or with stable angina pectoris), randomly selected from 7600 screenees for the BIP study, a trial of bezafibrate for raising HDL in persons with hypoalphalipoproteinemia (HDL-C < 45 mg/dl). The results were as follows. MONICA Men Agea 55.1+7.7 212.Ok47.8 TCb LDL-Cb 144.3~47.6 HDL-Cb 38.9k9.7 HDL-C%C 19.0?6.2 143.6k75.4 TGb
Women
BIP Men
Women
53.0+7.1 218.9rt40.9 147.6+49.5 49.1k12.2 23.Ok6.8 116.6kl47.6
55.3k5.4 226.Ok41.3 156.1-c34.1 35.7k8.6 16.3k4.8 174.2+107
53.0+4.1 235.8zt45.5 159.8+40.0 42.7i11.4 17.8+6.0 169.h88.0
223
High TC, LDL-C and TG and low HDL-C are well accepted as risk factors for CHD. In the coronary (BIP) patients the values are within the normal range or moderately outranged, with the exception of HDL-C% which proves a valuable marker for atherogenicity. Differences in lipid profiles between these groups of healthy persons and coronary patients, though statistically significant, were minor. Other risk factors such as coagulation disorders and other lipoproteins as well as hereditary factors and lifestyles could account for the atherogenic process in individuals and on the population level. Intra-family correlation of plasma fibrinogen levels and their relation to lipid and lipoprotein values. The Tel Aviv-Heidelberg Three Generation Offspring Study BrunnerQ, Graff E, Hoting 1, Blettner M, Wahrendorf J, Schettler G, Inst. of Physiol. Hygiene, Edith Wolfson Hosp., Holon 58100, 11651
Israel; Geomed. Res. Unit, Heidelberg Acad. for the Humanities and Sciences, Germany
In the ongoing Three Generation Offspring Study more than 1200 men and women in the age range 5-95 years from about 400 families were examined. Plasma fibrinogen levels (F) were determined in more than 600 individuals. The pedigrees contain up to three vertical generations. In tbe analyses the members of the second generation are included as children of the first generation and also as parents of the third generation. In healthy men and women F gradually increases with age (in age-specific subgroups ~30 years, 30-64 years and 165 years), F strongly positively correlated with TC, LDL-C, TG, apo B, Lp(a) and body mass index and strongly negatively correlated with apo A-I, HDL-C and its subfractions HDL2C and HDL3-C. Familial correlations of F were calculated for all pairs of relatives (mother-father, mother-daughter, etc.). Equal weight was given to pedigrees. The correlations between cousins were higher than the marital correlations although the cousins do not share the same environment. This indicates a considerable genetic determination of plasma fibrinogen levels.
aIn years. bin mg/dl. ‘HDL-C as a percentage of TC. P < 0.001 for all differences BIP vs MONICA.
HDL AND REVERSE
CHOLESTEROL
pG-J
Cholesterol emux from different subcellular cholesterol pools in HepG2 cells Svmdov D, Fidge N, Baker Med. Res. Inst., Commercial Road,
.
Prahran, Vie., 3181, Australia
Cholesterol efflux is a key element of reverse cholesterol transport which may be rate-limited by the step involving translocation of cholesterol from intracelluiar compartments to the plasma membrane (PM). Since we have recently demonstrated that liver cells also donate cholesterol to the plasma pool independent of lipoprotein biosynthesis, we have addressed this question by investigating cholesterol efflux from different cellular pools using human hepatoma cells HepG2. Cholesterol in the PM was labeled by incubation of cells with
TRANSPORT
[14C]cholesterol which had been incorporated into phosphatidylcholine micelles or HDL. Cholesterol in the lysosomes and endoplasmic reticulum (ER) was labeled by incubation of cells with [t4C]cholesterol which had been incorporated into LDL, or [14C]acetate, respectively. Localization of [14C]cholesterol in the particular membrane was validated by cell fractionation. [‘4C]Cholesterol associated with the intracellular compartments was esterified twice as effectively as that from PM. Efflux of [‘4C]cholesterol born the PM into human serum after 3 h incubation was double that from lysosomes and eight times that from the ER. Extension of the incubation time from 3 to 6 h markedly diminished the difference in cholesterol efIlux from different membranes. Further incubation up to 12 h abolished the different responses. In contrast, cell-free preparations of membranes ob-
Atherosclerosis X, Montreal, October 1994
222
similar size to IDL and LDL (fraction II), and large HDL (fraction III). Accurate quantitation of apo E in these fractions has been difficult, however, since fractions do not coincide with classical ultracentrifugal density cuts and a large proportion of lipoproteinbound apo E is removed by ultracentrifugation. In the present study, we have determined the plasma lipoprotein distribution of apo E in normolipidemic (group A, n = 6), hypercholesterolemic (type IIa, group B, II = 8), mixed hyperlipidemic (type IIb, group C, n = 8) and hypertriglyceridemic (type IV, group D, n =7) subjects after an overnight fast. Plasma lipoproteins were separated by fast protein liquid chromatography (FPLC) on a Superose 6 column and apo E was determined by a polyclonal antibody ELISA assay. The total plasma apo E concentrations were 3.6 f 0.3, 4.6 f 0.3, 7.7 + 1.1, and 7.4 & 0.8 mg/dl (mean f SE) for groups A-D respectively. In the normolipidemic subjects, 6.8, 40.1 and 53.1% of total plasma apo E was contained within fractions I, II and III. A greater proportion of plasma apo E was found in TRL (6.8, 11.7, 33.8, 49.9%) and a smaller proportion in HDL (53.1, 33.1, 22.1, 15.3%) as TRL triglyceride increased (0.55 f 0.08, 0.91 f 0.18, 2.73 kO.28. 4.89 i0.61 mmol/l) for zrouos A-D. respectively, Importantly, remnant lipoproteins (fra&on II) were responsible for transporting a significant proportion of plasma apo E in all subjects (40.1, 55.1, 44.1, 34.9%), representing mean plasma ‘remnant’ apo E concentrations of 1.5 f 0.2, 2.6 + 0.2, 3.4 + 0.5 and 2.6 + 0.3 mg/dl, respectively. We conclude that irrespective of a patient’s plasma tiglyceride concentration, a significant proportion of plasma apo E is associated with remnant-sized lipoproteins. Elevated LDL may contribute to increased requirement for beparin in patients with acute myocardial infarction g&ski 1, Sbnecka A, Flasi6ski J, Wasilewska A, Winnicka A, Kurek P, Dmochowska W, Inst. of Maritime and Tropical Med.,
1
Gdynia, Poland
LDL are believed to interfere with heparin and antithrombin III (AT III) activity, so that an increased dose of heparin may be necessary in patients with unstable angina or acute MI and a high plasma LDL concentration. The aim of our study was to compare the dose of heparin required to maintain the activated partial thromboplastin time (APTT) ratio within the range 2.0-2.5 in angina-AM1 patients with normal (A) or high (B) plasma HDL-C, to compare AT III activity in groups A and B, and to correlate the required dose of heparin with plasma LDL-C and AT III activity. 18 patients (A) with LDL-C 149 +_20 mg/dl (i.e. normal, <160 mg/dl) required 29 000 * 3900 units of heparin/24 h; their AT III activity was 12.9 +_1.9 ng/ml (normal range O-15 ng/ml). For group B (LDL-C 224 + 30 ng/dl) the heparin need was 37 500 + 5400 units/24 h (P < 0.05) and AT III 11.9 f 1.7 ng/ml (difference ns). The only significant correlation found (r = 0.659, P < 0.02) was between LDL-C and heparin requirement in group B. Con&sions Patients with high LDL-C require an increased dose of heparin if a desirable APTT ratio is to be preserved; LDL C correlated directly with this dose; and normo- and hypercholesterolemic patients did not differ in AT III activity.
taken up and metabolized to retinoic acid in bone marrow-derived cells. Retinoic acid is a ligand to nuclear receptors and has an important role in the regulation of many genes. Chylomicron remnants were made from mesenteric lymph of rats given [‘HIretin through a duodenal catheter. Retinoids were determined by HPLC and scintillation counting of elution fiattions. At 4°C the cell-associated radioactivity reached a maximum of 2.6% of added dose/mg cell protein after 1 h of incubation. During further incubation for 3 h at 37”C, 80% of the retinyl ester was hydrolyzed to retinol and about 25% of the [3H]retinol was secreted to the medium during the same period. The hydrolysis of retinyl esters was partly inhibited by monensin, chloroquine and brefeldlin A. The amount of [3H]retinol taken up by the cells during 5 h of incubation at 37°C was increased 4-fold by adding 1 ,@ml lipoprotein lipase and decreased by 80% by inhibiting proteoglycan synthesis. By incubating the cells 5 and 24 h at 37°C we were able to detect a small amount of all-trans [3H]retinoic acid. More polar metabolites, not yet identified, were also detected in the cells. In the medium about 50% of the radioactivity was found as polar metabolites after 24 h. (1621 LDL subclass patterns and Syndrome X elements Hartwich J, Baczydska E, Zdzienicka A, Pajqk A, Goldsztajn P, .I .I s, Dept. of Clin. Biochem., Sch. of Publ. Health, Colleg&m Meicum, Jagiellonian Krakhw, Koperniko 15a, Poland
Univ. of Cracow,
31-501
Heterogeneity of LDL particles can be described by two distinct patterns denoted A and B. Pattern A is characterized by a predominance of large LDL, and pattern B by a predominance of small, dense LDL. Pattern B of LDL is associated with increased risk of CHD and with an atherogenic lipoprotein profile. Since dyslipidemia is one of the signs of Syndrome X we asked which of the LDL subclass patterns may be related to this phenomenon. Observations were carried out on the participants of the PolMONICA Cracow Study. 139 non-diabetic men 45-68 years old were divided into subgroups with LDL subclass pattern A (70%) or LDL subclass pattern B (30%). The LDL subclass pattern was photographed after density gradient ultracentrifugation of stained plasma and quantified by densitometric scanning of the slides. Fasting glucose and insulin, SUMGLUC, SUMINS (endogenous insulin response during oral glucose tolerance test) were significantly higher in subjects with LDL subclass pattern B. The frequencies of Syndrome X symptoms according to LDL subclass pattern (percent of subjects in subgroup) were as follows: hyperinsulinemia in 60% of the patients in group B vs 41% in group A, hypertension 48.7% vs 27.8%. obesity 57% vs 35% and impaired glucose tolerance 33% vs 16.5% respectively. In patients with LDL subclass pattern B we observed higher plasma apo B and triglyceride concentrations, whereas significantly lower were HDL-C concentration, plasma apo A-I and LDL free cholesterol concentrations. LDL patternB is, therefore, associated with the abnormalities described as Syndrome X. [1631 WE,
Uptake of aggregated low density lipoproteins by vascular smooth muscle cells Brown SA, Klemp KF, Guyton JR, Dept. of Med., Box
3510, Duke Univ. Med. Center, Durham, NC 27710, USA
Retinoic acid is formed when retinyl esters (vitamin A) are given to J 774 macrophages in chylomicron remnants SE, Halvorsen B, Nenseter MS, Blomhoff R, Nomm KR, Inst. of Nutrition, Rex Sch. of Med., Univ. qf Oslo, PO Box 1046,
(161)
Blind&,
0316, Oslo, Norway
The objective of this study was to show that vitamin A given physiologically as retinyl esters in chylomicron remnants can be
When human LDL are aggregated by vortexing, particle fusion and phase separation of lipid domains occurs, producing lipid droplets and vesicles. The resulting lipid structures bear a strong resemblance to extracellular lipid deposits in human atherosclerosis (Guyton et al. J Lipid Res 1991; 32: 953-962). Macmphages take up aggregated LDL (agLDL) by phagocytosis (Khoo et al Arteriosclerosis 1988; 8: 348-358), but the interaction of agLDL with vascular smooth muscle cells (SMC) is not known.
Atherosclerosis X, Montreal, October I994
Wednesday 12 October 1994: Poster Abstracts HDL and reverse cholesterol transport
AgLDL in concentrations of lO-5O,~g/ml were applied to cultures of 4th to 10th passage swine aortic smooth muscle cells. To facilitate interaction with cells, agLDL were briefly homogenized. Witbin 2 days, agLDL were observed to cluster on the surface of SMC. By 3 or more days, agLDL were internalized in large lysosomes, as confirmed by fluorescent labeling with diI and identification of lysosomes with acridine orange. Electron microscopy with the use of the osmium-tannic acid-paraphenylenediamine procedure to preserve lipid structure and of acid phosphatase to identify lysosomes likewise showed large lysosomes (up to 16 per cell profile, producing a ‘foamy’ appearance). However, cytoplasmic lipid droplets were not evident by electron microscopy or by Nile red fluorescent staining. lz51labeling of LDL prior to aggregation allowed the demonstration of protein degradation after the application of agLDL to SMC. However, degradation rates were always higher for native LDL than for agLDL. I*?-agLDL degradation was partially inhibited by lO,ug/ml cytochalasin B, but because of its slow toxicity this inhibitor of phagocytosis could not be applied to the cells for sufficient time to follow agLDL ingestion fully. The results suggest a novel mechanism for uptake of agLDL by SMC, which may be associated with Iysosomal dysfunction. The lipid profile of healthy individuals and coronary patients in Israel. The Betibrate Infarction Prevention @UP) Study and the Israel MONICA Study w, Graff E, Benderly M, Reicher-Reiss H, Behar S, for the BIP Study Group, Neufeld Cardiac Res. Inst., Sheba Med.
1
Center, Tel-Hashomer, Israel
During 1991-1992, we compared the lipid profiles of healthy volunteers (324 M, 494 F) in the Israel MONICA project with those of age-matched (45-74 years) patients (323 M, 449 F) with established coronary disease (6 months to 5 years after acute myocardial infarction and/or with stable angina pectoris), randomly selected from 7600 screenees for the BIP study, a trial of bezafibrate for raising HDL in persons with hypoalphalipoproteinemia (HDL-C < 45 mg/dl). The results were as follows. MONICA Men Agea 55.1+7.7 212.Ok47.8 TCb LDL-Cb 144.3~47.6 HDL-Cb 38.9k9.7 HDL-C%C 19.0?6.2 143.6k75.4 TGb
Women
BIP Men
Women
53.0+7.1 218.9rt40.9 147.6+49.5 49.1k12.2 23.Ok6.8 116.6kl47.6
55.3k5.4 226.Ok41.3 156.1-c34.1 35.7k8.6 16.3k4.8 174.2+107
53.0+4.1 235.8zt45.5 159.8+40.0 42.7i11.4 17.8+6.0 169.h88.0
223
High TC, LDL-C and TG and low HDL-C are well accepted as risk factors for CHD. In the coronary (BIP) patients the values are within the normal range or moderately outranged, with the exception of HDL-C% which proves a valuable marker for atherogenicity. Differences in lipid profiles between these groups of healthy persons and coronary patients, though statistically significant, were minor. Other risk factors such as coagulation disorders and other lipoproteins as well as hereditary factors and lifestyles could account for the atherogenic process in individuals and on the population level. Intra-family correlation of plasma fibrinogen levels and their relation to lipid and lipoprotein values. The Tel Aviv-Heidelberg Three Generation Offspring Study BrunnerQ, Graff E, Hoting 1, Blettner M, Wahrendorf J, Schettler G, Inst. of Physiol. Hygiene, Edith Wolfson Hosp., Holon 58100, 11651
Israel; Geomed. Res. Unit, Heidelberg Acad. for the Humanities and Sciences, Germany
In the ongoing Three Generation Offspring Study more than 1200 men and women in the age range 5-95 years from about 400 families were examined. Plasma fibrinogen levels (F) were determined in more than 600 individuals. The pedigrees contain up to three vertical generations. In tbe analyses the members of the second generation are included as children of the first generation and also as parents of the third generation. In healthy men and women F gradually increases with age (in age-specific subgroups ~30 years, 30-64 years and 165 years), F strongly positively correlated with TC, LDL-C, TG, apo B, Lp(a) and body mass index and strongly negatively correlated with apo A-I, HDL-C and its subfractions HDL2C and HDL3-C. Familial correlations of F were calculated for all pairs of relatives (mother-father, mother-daughter, etc.). Equal weight was given to pedigrees. The correlations between cousins were higher than the marital correlations although the cousins do not share the same environment. This indicates a considerable genetic determination of plasma fibrinogen levels.
aIn years. bin mg/dl. ‘HDL-C as a percentage of TC. P < 0.001 for all differences BIP vs MONICA.
HDL AND REVERSE
CHOLESTEROL
pG-J
Cholesterol emux from different subcellular cholesterol pools in HepG2 cells Svmdov D, Fidge N, Baker Med. Res. Inst., Commercial Road,
.
Prahran, Vie., 3181, Australia
Cholesterol efflux is a key element of reverse cholesterol transport which may be rate-limited by the step involving translocation of cholesterol from intracelluiar compartments to the plasma membrane (PM). Since we have recently demonstrated that liver cells also donate cholesterol to the plasma pool independent of lipoprotein biosynthesis, we have addressed this question by investigating cholesterol efflux from different cellular pools using human hepatoma cells HepG2. Cholesterol in the PM was labeled by incubation of cells with
TRANSPORT
[14C]cholesterol which had been incorporated into phosphatidylcholine micelles or HDL. Cholesterol in the lysosomes and endoplasmic reticulum (ER) was labeled by incubation of cells with [t4C]cholesterol which had been incorporated into LDL, or [14C]acetate, respectively. Localization of [14C]cholesterol in the particular membrane was validated by cell fractionation. [‘4C]Cholesterol associated with the intracellular compartments was esterified twice as effectively as that from PM. Efflux of [‘4C]cholesterol born the PM into human serum after 3 h incubation was double that from lysosomes and eight times that from the ER. Extension of the incubation time from 3 to 6 h markedly diminished the difference in cholesterol efIlux from different membranes. Further incubation up to 12 h abolished the different responses. In contrast, cell-free preparations of membranes ob-
Atherosclerosis X, Montreal, October 1994