Metabolism of Apolipoprotein B in Members of a Family With Accelerated Atherosclerosis: Influence of Apolipoprotein E-3/E-2 Pattern Steven M. Haffner,
Rampratap
S. Kushwaha,
and William
R. Hazard
Familial combined hyperlipidemia (FCHL) appears to be the most common, simply inherited hyperlipidemia strongly associated with coronary heart disease. In the family examined in this study, two of the siblings who met diagnostic criteria for FCHL had extensive clinical atherosclerosis before age 30, unusually premature for this form of hyperfipidemia. Lipoproteins and low-density lipoprotein (LDL) apolipoprotein (ape) B metabolism were characterized in these siblings in an attempt to gain insight into the cause of the rapid atherosclerosis in the two siblings so affected. LDL apo B production rates were very high in all three siblings (25 to 30 mg/kg/d), consistent with FCHL. p-Very-low-density lipoprotein-p ((%VLDL) was present in the plasma of both siblings with accelerated atherosclerosis. The isoapolipoprotein E pattern in both of these siblings was E-3/E-2. In the third sibling, who was free of premature clinical atherosclerosis and lacked plasma @VLDL, the pattern was E-3/E-3. Thus, the heterozygote apo E-3/E-2 pattern may be related to the accumulation of p-VLDL in persons with a very high apo B production rate. The abnormal accumulation of (3-VLDL may be one of the possible explanations for the rapid, premature atherosclerosis in the two siblings with FCHL in this kindred. Both male members in this kindred also had low levels of high-density lipoproteins, and thus may have had an additional risk of developing atherosclerosis due to this lipoprotein abnormality as well. Copyright 0 1992 by W.B. Saunders Company
M
EMBERS OF A KINDRED who variously have elevated very-low-density lipoproteins (VLDL), lowdensity lipoproteins (LDL), or both, fulfill the criteria for the diagnosis of familial combined hyperlipidemia (FCHL).’ Moreover, the lipoprotein pattern of an individual with this disorder may characteristically change over time.’ FCHL may occur in as many as one in 50 to 100 persons’J in the general population and is the most common genetic hyperlipidemia associated with premature coronary heart disease (affecting as many as lo%, with average age at first myocardial infarction approximately 50 years).‘.’ Metabolic studies have shown an increased apolipoprotein (apo) B production in persons with this disorder.6 In the present study, plasma lipoproteins, LDL apo B metabolism, and isoapolipoprotein E patterns were examined in three affected siblings from a family with FCHL to gain insight into the mechanism of the extraordinarily accelerated atherosclerosis in the proband and his brother (onset before age 30). The results suggest an important role of the isoapolipoprotein E pattern and p-VLDL in the genesis of their premature vascular disease. MATERIALS
AND METHODS
Study Subjects
studies. All subjects had at least two lipoprotein lipid measurements and two apo E isoelectric-focusing patterns determined. Values presented in Table 1 reflect lipoproteins at the time of admission to the Clinical Research Center. None of these subjects had tendinous xanthomas, which are characteristic of familial hypercholesterolemia (FH). During their stay at the Clinical Research Center, they did not take drugs affecting lipid metabolism. Informed written consent was obtained from all subjects, and the study protocol was approved by the Human Subjects Review Committee of the University of Washington. Experimental Protocol LDL apo B turnover studies were conducted in each subject. Blood was drawn from the subjects for isolation and iodination of LDL 1 week before apo B turnover studies. The turnover study was begun by simultaneously injecting autologous ‘zI-labeled LDL after an overnight fast. All of the subjects ingested a saturated solution of potassium iodide to block the uptake of iodide by the thyroid (750 mg/d, starting 3 days before and continuing throughout the study). Iodinated lipoproteins were passed through a millipore filter (Millipore, Bedford, MA), and pyrogen was tested in rabbits before its injection into study subjects, who had been fasting for 12 to 14 hours. Subjects were maintained on an isocaloric fat-free liquid formula diet for the first 60 hours of the study, as described earlier.’ Thereafter, they were given a solidfood isocalorie diet containing 40% calories as fat, given in three meals daily. Serial blood samples (10 mL) were obtained in tubes
Three young adult siblings from a single kindred (Fig 1, subjects IV-l, IV-2, and IV-3) with various elevations of VLDL and LDL were referred to the University of Washington Northwest Lipid Research Clinic for evaluation of their hyperlipidemia (Fig 1 gives the health history and lipid levels of various family members). Two male members of the family had extensive atherosclerosis and underwent coronary bypass surgery at very early ages. Table 1 describes the clinical characteristics of these subjects. Subject IV-2, propositus, had a myocardial infarction at age 25 and a three-vessel coronary bypass graft at age 27. Subject IV-3, his younger brother, had a four-vessel bypass at age 27. Subject IV-2 never smoked, and subject IV-3 smoked less than 10 cigarettes per day for 8 years before his bypass surgery. Subject IV-l, their older sister, smoked one pack of cigarettes per day for 20 years. She was free of clinical atherosclerosis at the time of study. All these siblings had normal glucose tolerance and blood pressure levels. None had received any treatment for hyperlipidemia within 6 months of these metabolic Metabolism, Vol41, No 3 (March), 1992: pp 241-245
From the Deparhnent of Medicine, University of Texas Health Science Center, San Antonio, TX; the Depaltment of Physiology and Medicine, Southwest Foundation for Biomedical Research, San Antonio, TX,, and the Department of Internal Medicine, Bowman Gray School of Medicine, Wake Forest Universi~, Weston-Salem, NC. Supporled by National Institutes of Health Grants No. RR-137 (CRC), HL 22607, HL 23474, and a grantfrom R.J. Reynolds to the University of Washington. Most of this work was conducted at the University of Washington Clinical Research Center, Seattle, WA. Address reprint requests to Rampratap S. Zbshwaha, PhD, Department of Physiology and Medicine, Southwest Foundation for Biomedical Research, San Antonio, TX 78228-0147. Copyright 0 1992 by U! B. Saunders Company 0026-0495/92/4103-G@3$03.OOlO 241
HAFFNER, KUSHWAHA, AND HAZZARD
242
MI 4s
Characterization of Labeled LDL
OLD AGE
Labeled lipoproteins were characterized by the radioactivity associated in the trichloroacetic acid-precipitable fraction, free iodide, lipid, and apo B.‘.” Most of the radioactivity ( > 93%) in LDL was in the trichloroacetic acid-precipitable fraction. Only a very small amount (
Rampratap
S. Kushwaha,
and William
R. Hazard
Familial combined hyperlipidemia (FCHL) appears to be the most common, simply inherited hyperlipidemia strongly associated with coronary heart disease. In the family examined in this study, two of the siblings who met diagnostic criteria for FCHL had extensive clinical atherosclerosis before age 30, unusually premature for this form of hyperfipidemia. Lipoproteins and low-density lipoprotein (LDL) apolipoprotein (ape) B metabolism were characterized in these siblings in an attempt to gain insight into the cause of the rapid atherosclerosis in the two siblings so affected. LDL apo B production rates were very high in all three siblings (25 to 30 mg/kg/d), consistent with FCHL. p-Very-low-density lipoprotein-p ((%VLDL) was present in the plasma of both siblings with accelerated atherosclerosis. The isoapolipoprotein E pattern in both of these siblings was E-3/E-2. In the third sibling, who was free of premature clinical atherosclerosis and lacked plasma @VLDL, the pattern was E-3/E-3. Thus, the heterozygote apo E-3/E-2 pattern may be related to the accumulation of p-VLDL in persons with a very high apo B production rate. The abnormal accumulation of (3-VLDL may be one of the possible explanations for the rapid, premature atherosclerosis in the two siblings with FCHL in this kindred. Both male members in this kindred also had low levels of high-density lipoproteins, and thus may have had an additional risk of developing atherosclerosis due to this lipoprotein abnormality as well. Copyright 0 1992 by W.B. Saunders Company
M
EMBERS OF A KINDRED who variously have elevated very-low-density lipoproteins (VLDL), lowdensity lipoproteins (LDL), or both, fulfill the criteria for the diagnosis of familial combined hyperlipidemia (FCHL).’ Moreover, the lipoprotein pattern of an individual with this disorder may characteristically change over time.’ FCHL may occur in as many as one in 50 to 100 persons’J in the general population and is the most common genetic hyperlipidemia associated with premature coronary heart disease (affecting as many as lo%, with average age at first myocardial infarction approximately 50 years).‘.’ Metabolic studies have shown an increased apolipoprotein (apo) B production in persons with this disorder.6 In the present study, plasma lipoproteins, LDL apo B metabolism, and isoapolipoprotein E patterns were examined in three affected siblings from a family with FCHL to gain insight into the mechanism of the extraordinarily accelerated atherosclerosis in the proband and his brother (onset before age 30). The results suggest an important role of the isoapolipoprotein E pattern and p-VLDL in the genesis of their premature vascular disease. MATERIALS
AND METHODS
Study Subjects
studies. All subjects had at least two lipoprotein lipid measurements and two apo E isoelectric-focusing patterns determined. Values presented in Table 1 reflect lipoproteins at the time of admission to the Clinical Research Center. None of these subjects had tendinous xanthomas, which are characteristic of familial hypercholesterolemia (FH). During their stay at the Clinical Research Center, they did not take drugs affecting lipid metabolism. Informed written consent was obtained from all subjects, and the study protocol was approved by the Human Subjects Review Committee of the University of Washington. Experimental Protocol LDL apo B turnover studies were conducted in each subject. Blood was drawn from the subjects for isolation and iodination of LDL 1 week before apo B turnover studies. The turnover study was begun by simultaneously injecting autologous ‘zI-labeled LDL after an overnight fast. All of the subjects ingested a saturated solution of potassium iodide to block the uptake of iodide by the thyroid (750 mg/d, starting 3 days before and continuing throughout the study). Iodinated lipoproteins were passed through a millipore filter (Millipore, Bedford, MA), and pyrogen was tested in rabbits before its injection into study subjects, who had been fasting for 12 to 14 hours. Subjects were maintained on an isocaloric fat-free liquid formula diet for the first 60 hours of the study, as described earlier.’ Thereafter, they were given a solidfood isocalorie diet containing 40% calories as fat, given in three meals daily. Serial blood samples (10 mL) were obtained in tubes
Three young adult siblings from a single kindred (Fig 1, subjects IV-l, IV-2, and IV-3) with various elevations of VLDL and LDL were referred to the University of Washington Northwest Lipid Research Clinic for evaluation of their hyperlipidemia (Fig 1 gives the health history and lipid levels of various family members). Two male members of the family had extensive atherosclerosis and underwent coronary bypass surgery at very early ages. Table 1 describes the clinical characteristics of these subjects. Subject IV-2, propositus, had a myocardial infarction at age 25 and a three-vessel coronary bypass graft at age 27. Subject IV-3, his younger brother, had a four-vessel bypass at age 27. Subject IV-2 never smoked, and subject IV-3 smoked less than 10 cigarettes per day for 8 years before his bypass surgery. Subject IV-l, their older sister, smoked one pack of cigarettes per day for 20 years. She was free of clinical atherosclerosis at the time of study. All these siblings had normal glucose tolerance and blood pressure levels. None had received any treatment for hyperlipidemia within 6 months of these metabolic Metabolism, Vol41, No 3 (March), 1992: pp 241-245
From the Deparhnent of Medicine, University of Texas Health Science Center, San Antonio, TX; the Depaltment of Physiology and Medicine, Southwest Foundation for Biomedical Research, San Antonio, TX,, and the Department of Internal Medicine, Bowman Gray School of Medicine, Wake Forest Universi~, Weston-Salem, NC. Supporled by National Institutes of Health Grants No. RR-137 (CRC), HL 22607, HL 23474, and a grantfrom R.J. Reynolds to the University of Washington. Most of this work was conducted at the University of Washington Clinical Research Center, Seattle, WA. Address reprint requests to Rampratap S. Zbshwaha, PhD, Department of Physiology and Medicine, Southwest Foundation for Biomedical Research, San Antonio, TX 78228-0147. Copyright 0 1992 by U! B. Saunders Company 0026-0495/92/4103-G@3$03.OOlO 241
HAFFNER, KUSHWAHA, AND HAZZARD
242
MI 4s
Characterization of Labeled LDL
OLD AGE
Labeled lipoproteins were characterized by the radioactivity associated in the trichloroacetic acid-precipitable fraction, free iodide, lipid, and apo B.‘.” Most of the radioactivity ( > 93%) in LDL was in the trichloroacetic acid-precipitable fraction. Only a very small amount (
