Inheritance of cholesterol metabolism of probands with high or low cholesterol absorption

Heredity of cholesterol absorption and synthesis was studied in siblings of hypercholesterolemic probands with low and high serum cholestanol to cholesterol ratio (assumed to indicate low and high absorption of cholesterol, respectively). Cholesterol synthesis was assayed with sterol balance technique and measuring serum cholesterol precursor to cholesterol ratios (synthesis markers of cholesterol), and cholesterol absorption with measuring dietary cholesterol absorption percentage and serum plant sterol and cholestanol to cholesterol ratios (absorption markers of cholesterol). In the siblings of the low absorption families, cholesterol absorption percentage and ratios of absorption markers were significantly lower, and cholesterol and bile acid synthesis, cholesterol turnover, fecal steroids and ratios of synthesis markers significantly higher than in the siblings of the high absorption families. The ratios of absorption and synthesis markers were inversely interrelated, and they were correlated with cholesterol absorption and synthesis in the siblings. In addition, low absorption was associated with high body mass index, low HDL cholesterol, and serum sex hormone binding globulin levels, suggesting that low absorption was associated with metabolic syndrome. Intrafamily correlations were significant for serum synthesis markers, cholestanol, triglycerides, and blood glucose level.In conclusion, cholesterol absorption efficiency and synthesis are partly inherited phenomena, and they can be predicted by the ratios of non-cholesterol sterols to cholesterol in serum. Heredity of cholesterol absorption and synthesis was studied in siblings of hypercholesterolemic probands with low and high serum cholestanol to cholesterol ratio (assumed to indicate low and high absorption of cholesterol, respectively). Cholesterol synthesis was assayed with sterol balance technique and measuring serum cholesterol precursor to cholesterol ratios (synthesis markers of cholesterol), and cholesterol absorption with measuring dietary cholesterol absorption percentage and serum plant sterol and cholestanol to cholesterol ratios (absorption markers of cholesterol). In the siblings of the low absorption families, cholesterol absorption percentage and ratios of absorption markers were significantly lower, and cholesterol and bile acid synthesis, cholesterol turnover, fecal steroids and ratios of synthesis markers significantly higher than in the siblings of the high absorption families. The ratios of absorption and synthesis markers were inversely interrelated, and they were correlated with cholesterol absorption and synthesis in the siblings. In addition, low absorption was associated with high body mass index, low HDL cholesterol, and serum sex hormone binding globulin levels, suggesting that low absorption was associated with metabolic syndrome. Intrafamily correlations were significant for serum synthesis markers, cholestanol, triglycerides, and blood glucose level. In conclusion, cholesterol absorption efficiency and synthesis are partly inherited phenomena, and they can be predicted by the ratios of non-cholesterol sterols to cholesterol in serum. Owing to small absorption of dietary plant sterols, serum contains low concentrations of these sterols, mainly campesterol and sitosterol. In addition, serum also contains a low concentration of cholestanol, a 5α derivative of cholesterol, which is partly synthesized in the body, but is also present in diet in very small amounts. These three sterols are transported by serum lipoproteins, mainly by LDL like cholesterol. The ratio of the three sterols to cholesterol correlates positively to absorption efficiency and negatively to synthesis of cholesterol (1Miettinen T.A. Tilvis R.S. Kesäniemi Y.A. Serum plant sterols and cholesterol precursors reflect absorption and synthesis in volunteers of a randomly selected male population.Am. J. Epidemiol. 1990; 131: 20-31Google Scholar, 2Miettinen T.A. Tilvis R.S. Kesäniemi Y.A. Serum cholestanol and plant sterol levels in relation to cholesterol metabolism in middle-aged men.Metabolism. 1989; 38: 136-140Google Scholar). Accordingly, their increased serum ratio points to a high absorption percentage of cholesterol and low cholesterol synthesis. Furthermore, low cholesterol synthesis decreases the release of cholesterol precursor sterols, most likely mainly from the liver to circulation, such that the ratios of squalene, methyl sterols, cholestenol, desmosterol, and lathosterol to cholesterol are also reduced (1Miettinen T.A. Tilvis R.S. Kesäniemi Y.A. Serum plant sterols and cholesterol precursors reflect absorption and synthesis in volunteers of a randomly selected male population.Am. J. Epidemiol. 1990; 131: 20-31Google Scholar). In general, the ratios of the absorption markers, and synthesis markers of cholesterol are negatively related to each other (3Miettinen T.A. Strandberg T.E. Gylling H. for the Finnish Investigators of the Scandinavian Simvastatin Survival Study GroupNoncholesterol sterols and cholesterol lowering by long-term simvastatin treatment in coronary patients. Relation to basal serum cholestanol.Arterioscler. Thromb. Vasc. Biol. 2000; 20: 1340-1346Google Scholar). Accordingly, a selection of subjects from a population with high ratios of the absorption markers predicts high absorption and low synthesis of cholesterol in these cases (1Miettinen T.A. Tilvis R.S. Kesäniemi Y.A. Serum plant sterols and cholesterol precursors reflect absorption and synthesis in volunteers of a randomly selected male population.Am. J. Epidemiol. 1990; 131: 20-31Google Scholar). On the other hand, the higher the ratios of the synthesis markers, e.g., lathosterol, the higher is cholesterol synthesis. In addition, baseline synthesis rate of cholesterol predicts to some extent lowering of cholesterol to statins, inhibitors of cholesterol synthesis. For instance, in the highest cholestanol quartile of the Finnish subgroup to the Scandinavian Simvastatin Survival Study (4S), serum cholesterol lowering was slightly but significantly lower than in the lowest quartile with high cholesterol synthesis (3Miettinen T.A. Strandberg T.E. Gylling H. for the Finnish Investigators of the Scandinavian Simvastatin Survival Study GroupNoncholesterol sterols and cholesterol lowering by long-term simvastatin treatment in coronary patients. Relation to basal serum cholestanol.Arterioscler. Thromb. Vasc. Biol. 2000; 20: 1340-1346Google Scholar). From among the non-cholesterol sterols, methyl sterol ratios to cholesterol exhibited significant intrapair correlations in serum of monozygotic but not dizygotic twins (4Kesäniemi, Y. A. Koskenvuo M. Vuoristo M. Miettinen T.A. Biliary lipid composition in monozygotic and dizygotic pairs of twins.Gut. 1989; 30: 1750-1756Google Scholar). However, concentrations of demethylated precursor sterols in plasma of a large number of families and twins showed associations, e.g., with apoE polymorphism (5Kempen H.J.M. Glatz J.F.C. Leuven J.A.G. van der Voort H.A. Katan M.B. Serum lathosterol concentration is an indicator of whole-body cholesterol synthesis in humans.J. Lipid Res. 1988; 29: 1149-1155Google Scholar), but no familial studies have been performed. Our routine non-cholesterol sterol analyses have revealed several families with high serum plant sterol ratios associated with low ratios of synthesis markers, suggesting heredity of variables of cholesterol metabolism. To this end, this study was planned to show the possible association of cholesterol absorption and synthesis in siblings of probands with high versus low ratios of serum absorption marker sterols for cholesterol, suggesting high versus low cholesterol absorption. For this pupose, the first-degree relatives of those probands were studied for serum precursor sterols, cholesterol absorption percentage, and cholesterol synthesis with the sterol balance technique. From a large number (n = 330) of coronary subjects with baseline measurement of serum non-cholesterol sterols, six and seven subjects with respective lowest and highest ratios of serum cholestanol to cholesterol were selected as probands. Definition for low and high absorption families was performed by the serum cholestanol to cholesterol ratios, such that the respective ratios were 98 ± 5 (mean ± SE) and 162 ± 12 102 mmol/mol of cholesterol with no overlapping between the groups. Probands' sisters and brothers were questioned by mail about their willingness to participate in the study. Of the 75 siblings 37, 14 from the low and 23 from the high absorption families, responded positively and were sent an invitation letter to participate in the study. None of them took lipid-lowering drugs or other products known to affect specifically cholesterol metabolism. A blood sample was drawn after an overnight fast, and a careful clinical examination was performed, and personal and family histories were recorded. The participants were advised to keep their normal habitual diet and possible drugs unchanged. They were asked to keep food diary for the next week, and consume a capsule (contained chromic oxide, C14-labeled cholesterol and tritiated sitostanol) three times a day with each major meal. During the last three days of the week, the participants were asked to provide stool samples to test tubes and bring them to laboratory, and give another blood sample. Owing to varying age and willingness, cholesterol absorption could be performed only in ten participants.The proband group had been studied earlier, and most of them had been given statin treatment by doctors at the Outpatient Department after the baseline blood samples, such that absorption and fecal studies could not be performed. All subjects volunteered to the study, the protocol of which was accepted by the Ethics Committee of the hospital. Table 1 shows the demographic data of the probands with low and high cholesterol absorption and of their respective siblings. Dietary diaries were calculated for different variables using a recent national method for their computerized analysis (6Knuts L-R. Rastas M. Haapala P. Micro-Nutrica, version 1.0. National Pensions Institute, Helsinki1991Google Scholar). Serum lipid analysis for total, HDL cholesterol (HDL-C) and LDL-C, total triglycerides, and phospholipids were made with the routine commercial kits (Boehringer Mannheim, Germany; Roche, Switzerland; Wako Chemicals, Germany). Ultracentrifugation was performed in the sibling's samples according to ealier methods (7Lipid Research Clinics programManual of laboratory operations, lipid research clinic program. Lipid Research Clinics Program, DHEW publication no. NIH/75–628. National Institutes of Health, Washington DC1974: 51-59Google Scholar) separating VLDL, IDL, LDL, and HDL followed by measurement of total, free, and esterified cholesterol, phospholipids, and triglycerides with commercial kits. In addition, blood glucose (hexokinase method; Roche, Switzerland), fasting serum insulin (radioimmunoassay; Pharmacia & Upjohn, Sweden), and serum sex hormone binding globulin (SHBG), an indicator of insulin resistance (8Haffner S.M. Sex hormone-binding protein, hyperinsulinemia, insulin resistance and noninsulin-dependent diabetes.Horm. Res. 1996; 45: 233-237Google Scholar) (fluoroimmunoassay, Wallac, Finland), were quantified with routine commercial kits. Plasma total homocysteine level was assayed with high-pressure liquid chromatography. Serum cholesterol, squalene, and non-cholesterol sterols mere measured by gas-liquid chromatography (GLC) on a 50 m long capillary column (9Miettinen T.A. Koivisto P. Non-cholesterol sterols and bile acid production in hypercholesterolaemic patients with ileal bypass.in: Paumgartner G. Stiehl A. Gerok W. Bile Acids and Cholesterol in Health and Disease. MTP Press, Lancaster, PA.1983: 183-187Google Scholar) from non-saponifiable material in serum. Each GLC run separates cholesterol, squalene, cholestenol, desmosterol, lathosterol (three sterols reflecting cholesterol synthesis), cholestanol, campesterol, sitosterol, and avenasterols (four sterols reflecting cholesterol absorption). The squalene and non-cholesterol sterol levels were standardized by cholesterol of the same GLC run, because this procedure eliminates differences caused by variable lipoprotein contents transporting cholesterol and non-cholesterol sterols. Thus, the values are expressed in terms of 102 × mmol/mol of cholesterol and expressed in the text as ratio.TABLE 1Demographic data, variables of glucose metabolism, and plasma total homocysteine level of study populationProbandsSiblingsVariablesLow AbsorbersHigh AbsorbersAllLow AbsorbersHigh AbsorbersAllNumber of patients6713142337Age, years58 ± 358 ± 158 ± 358 ± 252 ± 3aSignificantly different from probands.54 ± 3aSignificantly different from probands.Weight, kg82 ± 578 ± 380 ± 375 ± 468 ± 2aSignificantly different from probands.70 ± 2aSignificantly different from probands.BMI, kg/m229 ± 125 ± 1bSignificantly different from low absorbers.27 ± 125 ± 1aSignificantly different from probands.24 ± 124 ± 1aSignificantly different from probands.Blood glucose, mmol/l5.4 ± 0.64.4 ± 0.24.9 ± 0.35.2 ± 0.24.8 ± 0.15.0 ± 0.1Serum insulin, pmol/l——8.3 ± 1.86.6 ± 0.76.4 ± 0.56.5 ± 0.4Serum SHBG, nmol/l——37 ± 552 ± 889 ± 15bSignificantly different from low absorbers. 75 ± 10aSignificantly different from probands.Plasma total homocysteine, μmol/l———9.2 ± 0.611.5 ± 1.110.6 ± 0.7Mean ± SE. BMI, body mass index; SHBG, sex hormone binding globulin.a Significantly different from probands.b Significantly different from low absorbers. Open table in a new tab Mean ± SE. BMI, body mass index; SHBG, sex hormone binding globulin. Cholesterol absorption percentage and fecal sterols of both cholesterol and plant sterol origin, and bile acids were measured from a pooled sample of the three daily stool samples by earlier methods (10Crouse J.R. Grundy S.M. Evaluation of a continuous isotope feeding method for measurement of cholesterol absorption in man.J. Lipid Res. 1978; 19: 967-971Google Scholar, 11Miettinen T.A. Gas-liquid chromatographic determination of fecal neutral sterols using a capillary column.Clin. Chim. Acta. 1982; 124: 245-248Google Scholar, 12Grundy S.M. Ahrens Jr., E.H. Miettinen T.A. Quantitative isolation and gas-liquid chromatographic analysis of total fecal bile acids.J. Lipid Res. 1965; 6: 397-410Google Scholar, 13Miettinen T.A. Ahrens Jr., E.H. Grundy S.M. Quantitative isolation and gas-liquid chromatographic analysis of total dietary and fecal neutral steroids.J. Lipid Res. 1965; 6: 411-424Google Scholar). Mean daily intestinal cholesterol pool was calculated dividing fecal neutral sterols by (1-fractional absorption of cholesterol). Biliary cholesterol secretion was the difference between the intestinal pool and daily dietary cholesterol. Fractional absorption multiplied by the intestinal pool, biliary secretion, or dietary cholesterol revealed the absorption of total, biliary, and dietary cholesterol, respectively. Cholesterol synthesis was calculated by the sterol balance technique subtracting dietary cholesterol intake from the fecal sum of neutral sterols of cholesterol origin plus bile acids. Cholesterol turnover was cholesterol synthesis plus absorbed dietary cholesterol. All values are expressed in terms of mg/day. Means and SEs were calculated and the differences between the four groups were analyzed by ANOVA and Student's two-sided t-test. Correlation coefficients were calculated by Pearson's product-moment correlation. P < 0.05 was considered statistically significant. Table 1 shows the demographic values for probands and siblings in the low and high absorption groups. The siblings were 8 years younger than the probands, but the difference was significant only in the high absorbing families. Body mass index was higher in the low absorbing probands, and higher overall in probands than in siblings. Both diastolic and systolic blood pressure values were higher in the probands than siblings, but the low and high absorbers did not differ from each other. Blood glucose and serum insulin values were similar in the subgroups, even though SHBG values were high in the siblings, especially in the high absorbing ones. Plasma concentration of total homocysteine did not differ between the groups. Table 2 shows the major lipids of probands and siblings and ultracentrifugation analysis of the siblings. It can be seen that serum total and LDL-C levels did not differ between the low and high absorbers, even though they were higher in the probands, but HDL-C level was significantly lower in the low than high absorbers. However, no consistent difference in triglycerides was seen between low and high absorbers, even after ultracentrifugation (data not shown), such that they only tended to be higher in the low than high absorbers, especially in VLDL and IDL. In contrast to cholesterol, serum phopholipids were lower in the low than high absorbers due to difference in HDL. Esterification percentage of cholesterol was significantly lower only in HDL fraction of the low than high absorbers.TABLE 2Serum and lipoprotein lipids of study populationProbandsSiblingsVariablesLow Absorbers (n = 6)High Absorbers (n = 7)All (n = Absorbers (n = Absorbers (n = (n = cholesterol ± ± ± ± different from ± ± different from analyses from serum. The other lipoprotein lipid values are from ± ± ± ± different from ± different from different from low ± different from triglycerides, ± ± ± ± ± ± analyses from serum. The other lipoprotein lipid values are from ± ± ± ± different from ± different from ± different from ± ± ± ± ± different from low ± ± ± ± ± ± different from low ± phospholipids, ± ± different from low ± ± ± ± ± 14 ± 14 ± ± 578 ± ± ± ± different from low ± ± analyses from serum. The other lipoprotein lipid values are from Significantly different from Significantly different from low absorbers. Open table in a new tab Mean ± SE. The ratio of serum squalene only tended to in siblings as with probands, but the differences were not significant. The levels were not by the absorption squalene and non-cholesterol sterol to cholesterol ratios mmol/mol of in study AbsorbersHigh ± ± ± ± ± ± ± ± different from low ± ± ± 1bSignificantly different from low ± ± ± different from low ± ± different from ± different from low ± ± ± different from low ± ± ± different from different from low ± ± ± different from low ± ± different from ± different from low ± ± ± different from low ± ± different from ± different from different from low ± different from ± ± different from low ± ± ± ± ± ± ± ± ± ± ± Significantly different from probands.b Significantly different from low absorbers. Open table in a new tab Mean ± SE. The ratios of serum cholestenol, desmosterol, and lathosterol were significantly higher in the low than high absorbers of both the probands and siblings, respectively. The low absorbing siblings tended to have lower ratios than their probands for in the high absorbing siblings the lathosterol ratio was higher than in the respective probands. In to the ratio of serum cholestanol, those of campesterol and sitosterol, but not of were higher in the high than low absorbers in both the probands for and siblings, respectively. In the low absorbers, the ratios of serum cholestanol and campesterol were lower in the probands than siblings, this difference significant only for serum campesterol in the high absorbers. In general, as in the probands, the ratios of the absorption markers were negatively related to those of the synthesis markers, but in the siblings their respective ratios to serum cholesterol, for instance, were = for P < and = for P < Intrafamily correlations were significant for the ratios of = = lathosterol = triglycerides = and glucose = but not for squalene, total or or body mass index, and only tended to be significant for plant sterol ratios = for but was that for cholestanol = Cholesterol absorption efficiency ± ± was significantly higher in the high than low absorbers, a the ratios of the cholesterol absorption sterols in serum fecal of cholesterol as neutral sterols of cholesterol origin and as bile acids was significantly increased in the low absorbers, in increased cholesterol synthesis also bile acid synthesis from cholesterol), as with the high absorbers. the low cholesterol absorption efficiency in the low absorbers, their dietary intake of cholesterol was lower than in the high absorbers, but the respective dietary intake of plant sterols was also Owing to a high biliary secretion of cholesterol in the low absorbers, their cholesterol absorption was the of the high absorbers low absorption of dietary absorption and metabolism in siblings of low and high Absorbers (n = Absorbers (n = ± ± different from high cholesterol, ± ± different from high ± ± different from high ± ± different from high ± ± different from high ± ± different from high ± 14 ± different from high synthesis, ± ± different from high turnover, ± ± different from high cholesterol, ± ± ± ± different from high ± ± ± ± different from high ± ± ± Significantly different from high absorbers. Open table in a new tab Mean ± SE. Cholesterol absorption efficiency was positively related to the ratios of the serum absorption markers < 0.05 to P < that the higher the serum plant sterol and cholestanol ratios, the higher is cholesterol absorption efficiency Cholesterol synthesis was positively related to the ratios of the synthesis markers < to P < and negatively to the absorption markers < and cholesterol absorption efficiency < Thus, the higher the ratios of the synthesis markers the higher is synthesis of cholesterol, and the lower is cholesterol absorption in both groups. It is to small that the correlations were significant in the high or low absorbing coefficients of cholesterol synthesis and absorption with ratios of serum markers of cholesterol absorption and synthesis mmol/mol of in siblings of high and low absorption absorption different for low absorbers. different for low absorbers. different for low different for low different for high different for low absorbers. different for high absorbers. different for high different for high different for low different for high different for low absorbers. coefficients (n = between squalene and P < Significantly different for high Significantly different for low absorbers. Open table in a new tab All coefficients (n = between squalene and P < The present show that cholesterol efficiency and cholesterol synthesis are inherited earlier that the high serum cholestanol, campesterol, and ratios to serum cholesterol low serum precursor sterol indicate high cholesterol absorption efficiency and low cholesterol synthesis was to be The on variables of cholesterol metabolism in these coronary families are not only by but also some It to be in a normal selection of subjects absorption using high versus low ratios of serum absorption markers low high synthesis of cholesterol also indicate of families with respective high versus low absorption low high of cholesterol, as in the present hypercholesterolemic coronary However, differences in synthesis and absorption of cholesterol, serum total and LDL-C levels were similar e.g., in the that synthesis of LDL LDL-C level. The of of total and LDL-C be by the small number of lipid association is in family studies A. G. and of serum low and high lipoprotein cholesterol and serum and Scholar). On the other hand, lower HDL-C level and a of higher serum and body in low with high absorbers earlier in a large number of coronary patients T.A. Gylling H. Strandberg serum cholestanol as of coronary in subgroup of Scandinavian simvastatin Scholar), and the in metabolic and J. Scholar). In addition, low serum SHBG levels the of metabolic in the low absorbers. The intake of cholesterol, and plant sterols, the three dietary variables with cholesterol metabolism, were different in the groups. Dietary intake due to different body and the intake was in terms of of body dietary intake in the low than high absorbers absorption and could have cholesterol synthesis. It is to that the of synthesis by in Table could be a of about of dietary cholesterol absorption. low absorbers were of plant sterols, which could have cholesterol absorption increased cholesterol synthesis. for body plant sterol intake similar in the suggesting that to different cholesterol absorption was It been earlier that in subjects with the at baseline low cholesterol absorption efficiency P. Gylling H. Miettinen T.A. a new of the metabolic low cholesterol J. 2000; Scholar), suggesting in with the present that low cholesterol absorption efficiency to be associated with insulin be the absorption or synthesis of cholesterol, for the of cholesterol metabolism, that cholesterol absorption is by H. J. P. of dietary cholesterol in caused by in 2000; Scholar) the of H. of a in the of dietary cholesterol Scholar), which at plant and also cholesterol absorption. It been J. Lipid Res. that in the plasma concentrations of sterols is and polymorphism in to in the plasma plant sterol The study was by a from the Finnish Research The of and is