Arterioscler Thromb Vasc Biol 1991 Hill 290 7

7/29/2019 Arterioscler Thromb Vasc Biol 1991 Hill 290 7

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290

Genetic and Environmental Factors Affectingthe Incidence of Coronary Artery Disease inHeterozygous Familial HypercholesterolemiaJ.S. Hill, M.R. Hayden, J. Frohlich, and P.H. Pritchard

This study explores the influence of selected genetic and environmental factors on the clinicalexpression of heterozygous familial hypercholesterolemia (FH). A detailed examination of thephysical and biochemical features of FHwas performed in a large cohort of 208 females a nd 156males. Females with FH had higher levels of total, low density lipoprotein (LDL), and highdensity lipoprotein (HDL) cholesterol when compared with ma les, although the concentrationof HDL cholesterol was significantly lower for both sexes when compared with normals. Thereported incide nce of coronary artery dise ase (CAD) was 31% for men a nd 13% for wom en,which was lower when compared with figures from previous studies. The average age of onsetof coronary symptoms was delayed in females, with a mean age of 55 years compared with 48years for males (/?


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Hill et al Heterozygous Familial Hypercholesterolemia 291other factor that determines diversity in the clinicalexpression of FH. The genetic locus for plasma apo Ehas three common alleles (e2, e3, e4), which encode forthe three major apo E isoforms found in plasma, E2,E3 , and E4, respectively, resulting in the expression ofsix apo E phenotypes.8 Several studies have estab-lished that in a normal population, apo E4 and apo E2have opposite effects on the concentrations of totalcholesterol (TC) and LDL-C.9.10 In addition, it hasbeen shown that the incidence of the e2 and e4 allelesis greater in patients with ischemic heart disease.11Although some previous reports have attempted tostudy the effects of coinheritance of these differentapo E alleles on the phenotypic expression of FH, noclear pattern has been established.12"15

The current clinical and biochemical features ofheterozygous FH have been based on a number ofdifferent population studies.16"22 However, many ofthese investigations differ with respect to size, selec-tion criteria, and genetic background, making itdifficult to definitively describe the characteristics ofFH. In the present study, we have used specificselection criteria to perform a detailed examinationof the physical and biochemical features of a largecohort of individuals with heterozygous FH. A com-parative analysis was made between males and fe-males and between those with and without CADthrough the investigation of lipids, lipoproteins, andapos as well as other potential risk factors, includingthe polymorphism of apo E. This study has allowedus to identify those factors that, in addition to TClevels, are associated with the development of pre-mature coronary atherosclerosis in heterozygous FH.

MethodsSubjects

A total of 364 patients from 283 families withheterozygous FH were identified among a populationin the University Hospital Lipid Clinic. The ethnicorigin of this cohort was very diverse, with at least 38different countries represented. FH was diagnosed ifsubjects satisfied both of the following criteria: 1) alevel of LDL-C greater than the 95th percentilecorrected for both age and sex and 2) tendon xan-thomas in the patient or a first-degree relative.The criteria for CAD were the presence of angina(history of typical exercise-associated chest pain) ormyocardial infarction (MI, proven by electrocardio-gram and/or serum enzyme changes) or angiograph-ically proven disease or a history of coronary bypasssurgery. The frequency for each of these findings isindicated in Table 3, where more than one of theCAD indicators may occur in a single patient. Smok-ing was defined in both former and current smokerswho had a history of smoking of at least 5 pack-years,where 1 pack-year is equivalent to smoking 1 pack/day for 1 year. Hypertension was indicated if clini-cally documented, even if patients were currently onantihypertensive medication. The body mass index(BMI) was calculated as body weight in kilograms

divided by the sq uare of the height in meters. Plasmasamples from 125 unrelated FH subjects from thelarger patient population were randomly chosen forapo E phenorype determination. In addition, speci-mens from a randomly selected normal population of203 subjects aged 18-78 years living in the Vancouverarea were analyzed for lipids and apo E phenotype.Laboratory Methods

Venous blood was collected from all subjects afteran overnight fast of 12-16 hours. The EDTA/plasmawas separated from cells by low-speed centrifugation(l,20Qt> 20 minutes) and was analyzed immediatelyor frozen at 70C. TC and triglycerides (TGs) weremeasured by established enzymatic techniques.23-24High density lipoprotein cholesterol (HDL-C) wasdetermined as the amount of cholesterol remainingafter precipitation of apo B-containing lipoproteinswith heparin/MnQ 2.25 LDL-C was calculated fromthe formula [TC-(HDL-C)]-(TG/2.2), where allvalues were measured in millimoles per liter. Plasmaapo A-I and apo B were measured by rate nephelom-etry using a Beckman Immunochemistry System,Beckman Instruments, Ontario, Canada.26 Apo Ephenotypes were determined as previously de-scribed27 in plasma that was neuraminidase treated,delipidated, and focused in vertical polyacrylamideminigels. After immuno blotting, apo E was visualizedwith a polyclonal goat anti-apo E antibody followedby a protein G-peroxidase conjugate.Data AnalysisStatistical analysis was performed using data ob-tained on the patient's first visit to the lipid clinic toensure that the lipid values represented were obtainedbefore treatment. Patients with hypothyroidism orpoorly controlled diabetes and those on medicationaffecting lipoprotein metabolism were excluded fromanalysis. The significance of difference between twomeans was determined by Student's t test. The statis-tical significance of the differences in proportion be-tween two groups was determined by the x2 t e s t(employing Yates' correction for continuity).

ResultsAge and Sex DistributionA total of 364 patients (208 females and 156 males)with FH were identified from 283 families. The agedistribution of this popu lation is shown in Figure 1. Th emean age for males was 40.317 years and 45.417years for women. This age difference was statisticallysignificant (/?


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292 Arteriosclerosis and Throm bosis Vo l 11, No 2 March/April 199130-,43 -40-35 -30-23-20-13-10-3-030-|43-40-33 -30 -23 -20 -15-10-

5-0

0- t 10-1* SO-tt 30-90 40-40 SO-M M- M 70-71

0-1 10-it 10-M 30-3i 40-M SO-WAge in Years

7O-7t

FEMALES(n-208)

FIGURE 1. Bar graphs showing age distribution(years) of familial hypercholesterolemia patients

for each sex (No. of patients/age group).

MALES

(n-136)

normal population. In addition, the levels of HDL-Cand apo A-I were consistently lower in all FHpatients when compared with normals (/?


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Hill et al Heterozygous Famil ial Hyperch oieste rolemi a 293TABLE 2. Mean Values for LJpids, LJpoproteins, and Apoproteins for Each Age Division in Familial Hyperchoiesterolemia

VariableTotal cholesterol (mmol/1)LDL cholesterol (mmol/1)HDL cholesterol (mmol/1)Triglycerides (mmol/1)Apoprotein B (g/1)Apoprotein A-I (g/1)

Total cholesterol (mmol/1)LDL cholesterol (mmol/1)HDL cholesterol (mmol/1)Triglycerides (mmol/1)Apoprotein B (g/1)Apoprotein A-I (g/1)

0-9(" = 12)

7.24 1.55.661.41.140.20.990.41.150.31.220.1

0-9(/i =9)

7.650.96.11 0.81.090.21.070.21.280.21.190.1

10-19(=9)

7.63 1.55.771.41.160.20.75 0.31.090.21300.2

10-19(=8)

8i61.46.85 1.21.060.21.410.71.530.41.240.2

Male age division (yr) (n = 156)20-29(" = 19)

8.541.36.90Ol.ll0.21.200.71.300.41.160.1

30-39(n=28)

8.49+1.36.391.61.090.21.88+1.21350.41.24+0.2

40-49(n=35)

8.81 + 1.66.971.61.040.31.840.81J7O.71.1803

50-59("=32)8.63 1.26.881.21.050.21.570.71.500J1.250.2

Female age division (yr) (n=208)20-29( = 18)

8.571.46.85 1.41.22031.040.41.480.51.250.3

30-39(n=44)8.96+1.97.01 + 1.81.20+0.31.240.71.480_51.280.3

40-49(n=30)9.00U6.91 + 1.41.41 0.41.29+1.01.61 0.41.40 0.3

50-59("=45)

9.212.06.961.71.280.41.61 0.71.520.31.440J

60-69(" = 19)

8.841.76.861.41.150.21.45 0.71.380.41.340.2

60-69("=45)9.48l-57.331.61.240.31.860.91.590.41.410.3

70-79("=2)

6.91 0.15.390.40.920.11.43 0.41.41 0.11.31 0.03

70-79(=9)

9.39 2.06.902.01.420.32.201.01.47 0.41.390.3

Values are given as meanSD except for the male age group of 70-79 years, for which data are expressed as meanVi the range.

declined after age 50. In contrast, females generallyshowed a steady increase in the concentration of TGswith age. Between the ages of 30 and 49, men hadsignificantly higher TG levels than did women,1.85 + 1.0 mmol/1 compared with 1.260.9 mmol/1(/>


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294 Arteriosclerosis and Thromb osis Vo l 11 , No 2 March/April 199140-i

o 30-1oU. 20-

10-1o

0-1

MALES

FEMALES

6 10 20 30 40 50 6 0 7 0Age in Years

FIGURE 2. Line plot showing cumulative requency(%) ofcoronary artery disease in male ( ) an d female () familialhypercholesterolemia patients as a function of ag e (years).incidence of tendon xanthomas was greater in fe-males (89%) compared with males (77%) Q?


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Hill et al Heterozygous Familial Hypercholesterolemia 295TABLE 5. Assessment of Effects of Smoking, Hypertension, High Density Lipoprotein Cholesterol Levels, and BodyMass Index on Coronary Artery Disease in Familial Hypercholesterolemia

VariablenSmokingHypertensionHDL cholesterol inlower quartileHDL cholesterol inupper quartileBody mass index

CAD-68

26 (38%)7 (10%)

11 (16%)23 (34%)24.753.0

MalesCAD+

4733 (70%)'

3(6%)18 (38%)t6(13%)t

25.152.8

CAD-147

58 (39%)18(12%)34 (23%)38(26%)24.104.0

FemalesCAD +

269 (35%)9 (35%)t9 (35%)5 (19%)

25.313.5CAD, coronary artery disease; HDL, high density lipoprotein.Significance of difference between CA D+ and C A D - group s for males and females is */?


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296 Arteriosclerosis and Thrombosis Vol 11 , No 2 March/April 1991TABLE 7. Lipid, LJpoprotein, and Apoprotein Levels in Familial Hrpercbolesterolemia Patients With and WithoutApolipoprotein E4 and E2VariablenAge(yr)Total cholesterol (mmol/1)LDL cholesterol (mmol/1)HDL cholesterol (mmol/1)Triglycerides (mmol/1)Apoprotein B (g/l)Apoprotein A-I (g/l)

E 4 -77

45.6179.321.97.161.91.130.31.75+0.91.530.51.280.3

E4+48

47.7148.81 + 1.76.541.71.18+0.31.79+1.21.460.51.310.2

E2-111

46.6169.07 1.76.97 1.71.16+0.31.640.91.500.51.280.3

E 2 +14

44.5129.56+2.56.522.41.080.22.701.5*1.470.41.35 0.2

LDL, low density lipoprotein; HDL, high density lipoprotein.Values are given as mea nSD. Significance of difference between the + and - groups for each apolipoprotein E

isoform is */?


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Hill et al Heterozygous Fam ilial Hypercholesterolemia 297males and females is related to differences betweenthe impact of known risk factors and the incidence ofCAD. Even in the presence of a genetic mutationcausing overt hypercholesterolemia, the expressionof this disease is markedly affected by gender, lipidprofile, smoking, and hypertension. In our study, onlya small number of females with FH had symptoms ofCAD; however, the risk of developing CAD in fe-males was significantly increased in the smaller frac-tion of patients who had hypertension or elevatedTGs. In contrast, males had a higher general fre-quency of disease but were at a much greater risk ifthey had lower HDL-C values and a history ofsmoking. In fact, the presence of CAD was almostalways associated with one or more of these riskfactors. Considering these observations, it may benecessary to reevaluate the conditions of treatmentfor the general FH population. From this investiga-tion, it is apparent that an individualized assessmentto consider the differences between males and fe-males, including the effects of these risk factors,should be emphasized when treating and predictingthe development of CAD in patients with FH.

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bolic Basis of Inherited Disease, 6th ed. New York, McGraw-Hill Book Co, 1989, pp 1195-12502. Kwiterovich PO Jr , Fredrickson DS, Levy R I: Familial hyper-cholesterolemia (one form of familial type II hyperlipopro-teinemia): A study of its biochemical, genetic and clinicalrepresentation in childhood. J Clin Invest 1974^3:1237-1249

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24. Buccolo G, David H: Quantitative determination of serumtriglycerides by the use of enzymes. Clin Chem 1973;19:476-48225. Warnick GR, Albers JJ: Heparin-Mn 2+ quantitation of highdensity lipoprotein cholesterol: An ultrafiltration procedurefor lipemic samples. Clin Chem 1978;24:900-90426. Wieland H, Cremer P, Seidel D: Determination of apolipo-protein B by kinetic (rate) nephelometry. / Lipid Res 1982;23:893-90227. Hill JS, Pritchard PH: Improved phenotyping of apolipopro-tein E: Application to population frequency distribution. ClinChem 1990;36:1871-187428. Streja D, Steiner G, Kwiterovich PO Jr: Plasma high-densitylipoproteins and ischemic heart disease. Ann Intern Med1978;89:871-88029. Das HK, McPherson J, Brans GAP, Karathanasis SK, BreslowJL: Isolation, characterization and m apping to chromosome 19of the human apolipoprotein E gene. J Biol Chem 1985;260:6240-624730. Lusis AJ, Heinzmann C, Sparkes RS, Scott J, Knott TJ, GellerR, Sparkes MC, Mohandas R: Regional mapping of humanchromosome 19: Organization of genes of plasma lipid trans-port (apo cl, -c2, and -E and LDLR) and the genes c3, PEPDand GPI. Proc NatlAcad Sd USA 1986;83:3929-393331. Assmann G, Schmitz G, Menzel JH, Schultz H: Apolipopro-tein E polymorphism and hyperlipidemia. Clin Chem 1984;30:641-64332. Utermann G, Kindermann I, Kaffarnik H, Steinmetz A: Apo-lipoprotein E phenotypes and hyperlipidemia. Hum Genet1984;65:232-236

KEY WORDS familial hypercholesterolemia coronary arterydisease plasma lipids smoking apolipoprotein E hypertension

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Arterioscler Thromb Vasc Biol 1991 Hill 290 7