Journal Logo


Effect of a Reduced-Fat Diet With or Without Pravastatin on Glucose Tolerance and Insulin Sensitivity in Patients with Primary Hypercholesterolemia

Galvan, Alfredo; Natali, Andrea; Baldi, Simona; Frascerra, Silvia; Sampietro, Tiziana; Galetta, Fabio; Seghieri, Giuseppe; Ferrannini, Ele

Author Information
Journal of Cardiovascular Pharmacology: October 1996 - Volume 28 - Issue 4 - p 595-602
  • Free


The notion that high serum total cholesterol and low high-density-lipoprotein(HDL)-cholesterol concentrations are major risk factors for the development of atherosclerosis is so well established (1-3) that the prevention and treatment of hypercholesterolemic states are public health priorities in developed countries (1,4). Intervention studies have confirmed that reduction in total cholesterol concentrations leads to a reduction in cardiovascular morbidity and mortality (8-12). Low-cholesterol diets and pharmacological treatment constitute the cornerstones of the management of hypercholesterolemia.

Cardiovascular risk factors tend to cluster in the same person. Diabetes mellitus, essential hypertension, and dyslipidemia are frequently associated with one another. These conditions are characterized by insulin resistance and hyperinsulinemia. Indeed, in three large epidemiological studies, investigators have concluded that hyperinsulinemia per se is an independent risk factor for coronary heart disease (5-7). Therefore, any dietary or pharmacological intervention in the primary or secondary prevention of atherogenic conditions should assess the effect on the entire risk factor profile.

The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors effectively decrease low density lipoprotein (LDL)-cholesterol levels (8), but information on the effect of this class of drugs on glucose tolerance is scarce (13) and no studies in which sophisticated metabolic techniques are used are available. Therefore, we compared the effects of a reduced-fat diet alone with those of a reduced-fat diet combined with pravastatin on some of the major cardiovascular risk factors [glucose tolerance, insulin sensitivity, plasma lipids, lipoprotein profile, and blood pressure (BP)] in a selected group of hypercholesterolemic patients.


Study subjects

Twenty patients with primary familial hypercholesterolemia (of the heterozygous type) were selected from 45 consecutive patients referred to the Lipid Clinic. The diagnosis was based on the presence of high serum total cholesterol levels (>6.48 mM) in the probands and a well-documented family history of hypercholesterolemia. Secondary causes of hypercholesterolemia were excluded by clinical and laboratory assessment. Ten normocholesterolemic healthy subjects without family history of hypercholesterolemia served as the control group (Table 1). Inclusion criteria were (a) age 30-60 years, (b) body mass index <26 kg m-2, (c) fasting plasma glucose levels <5.7 mM and normal glucose tolerance by National Diabetes Data Group criteria (14); (d) arterial BP <160/90 mm Hg, and (e) plasma triglyceride levels <2.90 mM. All patients were free of intercurrent acute or chronic illness (as judged by clinical and laboratory workup), were nonsmokers, and were not receiving medications. Lipid-lowering treatment (probucol or fibrate) was discontinued at least 12 weeks before study enrollment. Therapy with any HMG-CoA reductase inhibitors was discontinued at least 4 weeks before study enrollment. The protocol was reviewed and approved by the Institutional Review Board of the National Research Council Institute of Clinical Physiology. Purpose, nature, and risks involved in the study were explained to all patients before their consent to participate was obtained.

Study design

In the hypercholesterolemic patients, glucose tolerance and insulin sensitivity were assessed twice, i.e., at baseline and at the end of an 8-week period during which placebo or oral pravastatin (40 mg once daily) was administered in a randomized, double-blind design. Healthy subjects were studied only at baseline. Throughout the observation period, a low-cholesterol (200 mg/day), reduced-fat (30% fat, 17% protein, 53% carbohydrate), isocaloric (as judged by indirect calorimetry) diet was prescribed to all the patients by a dietitian. Every 2 weeks, body weight, BP, diet and treatment compliance, and possible untoward effects of treatment were accurately checked. Patients were encouraged to maintain body weight throughout the study period. Blood samples were drawn at baseline and every 4 weeks for lipid and lipoprotein analysis. An ECG and ophthalmoscopic examination were performed at baseline and at every 4 weeks of treatment.

Oral glucose tolerance test (OGTT)

After at least 3 days of a 250-g carbohydrate diet and after an overnight (10-12 h) fast, glucose tolerance was assessed by a 3-h 75-g OGTT. At 8:00 a.m., a polyethylene 18-guage catheter was inserted in an antecubital vein for blood sampling. At baseline and at 30-min intervals during the OGTT, blood samples were obtained for substrate and hormone determinations. BP and heart rate (HR) were measured at baseline and at 30-min intervals during the entire study.

Insulin sensitivity test

At 9:00 a.m. on a different day, insulin sensitivity was measured by the euglycemic insulin clamp technique (15) at an insulin infusion rate of 40 mU min-1/m2 body surface area. Before the start of the study and at 20-min intervals during the clamp, arterialized blood samples were obtained for the measurement of substrate and hormone concentrations. BP and HR were measured at baseline and at 20-min intervals during the test.

Indirect calorimetry

During the insulin clamp, substrate oxidation and energy expenditure were measured by indirect calorimetry with a computerized, continuous open-circuit system with a canopy (Metabolic Measurement Cart Horizon, Sensor Medics, Anaheim, CA, U.S.A.), as previously described (16). Urine volumes were measured, and urine samples were taken at the end of the basal period and again at the end of the 2-h clamp for the measurement of nonprotein urinary nitrogen excretion [Kjeldahl method (17)]. We estimated protein oxidation from the urinary nonprotein nitrogen excretion rate by multiplying the latter value by 6.25, after correction for urea pool changes (18). Net rates of whole-body carbohydrate and fat oxidation were estimated from gas-exchange measurements according to equations described in detail previously (16). Nonoxidative glucose disposal (which encompasses anaerobic glycolysis, glycogen synthesis, and de novo lipid synthesis from glucose-derived carbons) was calculated as the difference between whole-body glucose disposal and total carbohydrate oxidation.

Hepatic glucose production (HGP)

Under fasting and clamp conditions, HGP was measured with the tracer technique according to the prime-constant infusion protocol as originally suggested by Norwich (19). A primed (22 μCi) constant (0.174 μCi min-1) infusion of [6-3H]glucose (Dupont NEN, Milan, Italy) was started. After 100 min of tracer infusion, three blood samples were drawn in 20 min for the determination of plasma [6-3H]glucose specific activity (SA). Under the steady-state conditions present in the fasting state, the basal glucose appearance rate (Ra) is calculated as the ratio between the tracer infusion rate and the basal glucose SA. During insulin infusion, the estimation of change in glucose Ra is optimized by minimizing the changes in plasma glucose SA (20). To approximate a clamp of glucose SA, we halved the basal tracer glucose infusion rate every 15 min after the start of insulin infusion until 45 min into the clamp, when it was stopped. At the same time, the exogenous glucose solution used to clamp plasma glucose concentrations was enriched with 150 μCi [6-3H]glucose. This level of enrichment was chosen on the basis of previous experiments to match the steady-state plasma glucose SA prevailing during the basal state. Non-steady-state glucose Ras were calculated from the isotopic data by a two-compartment model with an ad hoc computer program (21). HGP was calculated as the difference between Ra and the exogenous glucose infusion rate.

Analytical procedures

Blood samples were kept in an ice bath after being drawn and then spun at 4°C. The plasma was aliquoted and stored at -20°C until assay. Plasma glucose concentration was immediately assayed by the glucose oxidase method (Beckman Glucose Analyzer, Beckman Instruments, Fullerton, CA, U.S.A.). Plasma insulin was measured by radioimmunoassay (RIA: INSKIT, Sorin, Saluggia, Italy). Blood levels of lactate, pyruvate, β-hydroxybutyrate, alanine, and free fatty acids (FFA) were assayed by spectrophotometry. Tracer glucose concentrations in plasma were measured in duplicate by the Somogyi method (21). We processed diluted aliquots of the tracer infusates in the same way as the plasma samples to determine the tracer infusion rate precisely.

Serum cholesterol, triglycerides, and HDL-cholesterol were determined as described previously (22-24). Concentrations of apolipoproteins in serum were determined by single radial immunodiffusion (25) on an agarose gel plate, with the gel containing 30 ml/L goat antiserum against purified ApoA-1, AII, B, CIII, and E. (Daiichi Pure Chemicals, Tokyo, Japan). Lp(a) was measured with the apo (a) RIA kit from Pharmacia (Uppsala, Sweden); between-run coefficients of variation were 6.8% at 45 mg/L and 9.7% at 93 mg/L. Plasma LDL-cholesterol was calculated by the Friedewald formula (26).

Data analysis

Insulin-mediated glucose disposal rates (GDR) were averaged over the second hour of the clamp, during which near-steady-state conditions prevailed. Areas under time-concentration curves (AUC) were calculated by trapezoidal integration. Results are mean ± SEM. The statistical significance of differences between the group means was tested by analysis of variance (ANOVA). Two-way ANOVA with repeated measures over time was used to test for the effect of time and treatment simultaneously (placebo vs. pravastatin).


Clinical data

Eleven hypercholesterolemic patients were assigned to pravastatin treatment and 9 were assigned to placebo administration. Clinical characteristics and the baseline serum levels of total and LDL-cholesterol were similar in the placebo and pravastatin groups (Table 1). Weight loss at the end of the study was negligible in both hypercholesterolemic groups [from 67.7 ± 2.8 to 67.4 ± 2.8 kg in the pravastatin group (p = NS), and from 71.7 ± 3.7 to 70.5 ± 3.7 in the placebo group (p = NS)]. Baseline mean BP values were similar across study groups (91 ± 3, 94 ± 3, and 90 ± 3 mm Hg, in pravastatin, placebo and controls, respectively). After treatment, no significant changes in BP were observed in either group of hypercholesterolemic patients. Treatment compliance was 100% (as estimated by tablet count). In only 1 patient (assigned to pravastatin) was an asymptomatic, transient increase in serum creatine phosphokinase levels observed. No ophthalmic or hepatic abnormalities were noted during treatment.

Lipid parameters

As summarized in Table 2, there were no significant differences in baseline serum lipid concentrations between the placebo and treatment groups. Diet alone was associated with significant decrements in total and LDL-cholesterol and triglyceride levels. As indicated by the presence of significant statistical interactions, pravastatin administration was associated with a greater decrease in LDL-cholesterol and with an increase, rather than a decrease, in HDL-cholesterol concentrations. Percentagewise, the effect of pravastatin on LDL-cholesterol was twice as great as that of diet alone, whereas its effect on HDL-cholesterol was similar but opposite to that of diet alone. As a consequence, the basal LDL:HDL ratio, which was 60% higher in patients than controls (p = 0.02), remained unchanged in the placebo group, but decreased significantly in the pravastatin group to a mean value that was no longer different from that of the controls (Fig. 1). The observed changes in HDL-cholesterol levels were entirely due to the HDL3 subfraction, with no significant changes in the HDL2 subfraction.

At baseline, Apo B and Apo E levels were significantly higher in patients than in controls, with no difference between the placebo and pravastatin groups. Apo A, C2, C3, and Lp(a) were similar across study groups. After the 8-week period of diet treatment, a significant decrement in Apo B, C2, C3, and E, but not in Apo A or Lp(a) levels was observed, with no significant independent effect of pravastatin (Table 3).

Metabolic parameters

Baseline values for fasting HGP and glucose, insulin, and FFA AUCs in response to oral glucose were similar in controls and patient groups. Both diet alone and pravastatin were associated with similar slight improvements in glucose tolerance (AUC = 1.36 ± 0.08 and 1.32 ± 0.05 mM/180 min, respectively, at 8 weeks), which fell short of being statistically different from baseline. Likewise, 8 weeks of pravastatin or placebo induced similar reductions (≈16%) in insulin AUC (Fig. 2) and FFA AUC (data now shown), none of which reached statistical significance. During the clamp, similar steady-state plasma insulin concentrations (≈500 pM) were achieved in the three groups. The time course of HGP during the clamp (Fig. 3) shows that in both the placebo and the pravastatin group suppression of HGP by euglycemic hyperinsulinemia was slightly but not significantly more effective after 8 weeks of treatment than at baseline. At baseline, insulin sensitivity was similar in all three groups (GDR = 33.8 ± 1.7, 27.2 ± 2.2, and 32.2 ± 3.3 μmol · min-1 · kg-1 in the placebo, pravastatin, and control groups, respectively). After treatment, GDR was similar within the pravastatin and placebo groups (Fig. 3).

Calorimetric parameters

Fasting and insulin-stimulated rates of carbohydrate and lipid oxidation, nonoxidative glucose disposal, and energy expenditure are shown in Table 4. These parameters were all similar in the two groups at baseline, were all affected by insulin in the same manner, and did not change in response to placebo or pravastatin treatment.


Our group of nonobese, nondiabetic, normotensive patients with moderate hypercholesterolemia received an isocaloric diet with a reduced fat and low cholesterol content, similar to that recommended by the National Cholesterol Education Program as step 2 diet (<30% of calories from total fat, intake of saturated fat <10%, and cholesterol <200 mg/day) (1). Detailed dietary advice and biweekly monitoring resulted in weight maintenance for the 8-week period of follow-up, allowing evaluation of the metabolic effects of changes in diet composition-with or without the addition of pravastatin-independent of the body weight changes frequently observed (and slightly underreported) in clinical trials of lipid-lowering drugs including dietary intervention (27-30).

In our series, diet alone was associated with a significant decrease in serum LDL-cholesterol and triglycerides and a slight decrease in HDL-cholesterol concentrations, accompanied by parallel reductions in Apo B, C2, C3, and E levels. The addition of pravastatin led to a significantly greater reduction in LDL-cholesterol and an 8% increase in total HDL-cholesterol concentrations. Accordingly, the LDL:HDL cholesterol ratio remained almost unchanged throughout the 8-week period in the placebo-diet group, whereas it was restored to normal in the pravastatin-diet group (Fig. 1)

Reduced-fat diets effectively reduce total and LDL cholesterol levels, but concern has been raised over the reduction of HDL-cholesterol and the increase in triglyceride levels associated with the isocaloric replacement of fat with carbohydrate. Coulston and co-workers (31) reported that a low-fat diet (21% as fat, 60% as carbohydrate) produced marked hypertriglyceridemia and hyperinsulinemia in the short term. Our results do not support this concern, as plasma triglycerides concentrations decreased consistently (≈20%) in response to the reduced-fat diet independent of pravastatin administration and in the absence of weight loss. Furthermore, plasma insulin and glucose concentrations were, if anything, lower after diet intervention as compared with baseline levels (Fig. 2). One possible explanation for the discrepant effects of low-fat diets is the degree of change in the fat content of the diet. Lichtenstein and colleagues (27), reported that in response to a reduced fat (≈30%) isocaloric diet (similar to that used in the present study) triglyceride concentrations remained relatively stable while LDL- and HDL- cholesterol concentrations decreased. Moreover, Mattson and Grundy reported that when the fat content is further reduced to 15% of total calorie intake, a marked increase in triglycerides together with a decrease in HDL-cholesterol concentrations occurs (32). Therefore, the hypertriglyceridemic effect of reduced-fat diets appears to be related to the fat content of the diet. Clearly, an abrupt increase in the carbohydrate content of the diet (31) can enhance very LDL (VLDL)-triglyceride synthesis through stimulation of insulin release. Results of other trials in which moderate reductions in fat content were used are in concordance with the current results (28,32). Geographical differences in dietary habits may also play a role. Our patients consumed the Mediterranean diet, enriched with monounsaturated fat as olive oil (33), which has been associated with a low coronary heart disease risk (34). Our dietary intervention consisted of a reduction in total fat from an estimated 40% to 30% and conservation of the characteristic monounsaturated fat preponderance. In most trials in which a hypertriglyceridemic effect of low-fat diet has been reported, more drastic quantitative changes in fat content (from 40 to 20-25%), as well as changes in fat composition [replacing saturated with poly- and monounsaturated fats (28,32)], have been implemented. The degree to which fat quality may affect triglycerides metabolism, alone or in interaction with total fat content, remains to be determined.

The beneficial effects of low-fat diets on serum lipid profile have been suggested to be achieved only when dietary modification leads to weight loss (27). In the current study, in which body weight remained stable, the reduction in LDL-cholesterol and triglycerides levels was counterbalanced by a decrease in HDL-cholesterol, such that the LDL:HDL ratio remained unchanged. Normalization of the ratio was attained only when pravastatin was associated. We therefore confirm what has been suggested previously (35,36): that the reduction in HDL-cholesterol concentrations associated with low- or reduced-cholesterol, low-fat diets can be attenuated or fully prevented by the concomitant use of an HMGCoA reductase inhibitor.

Higher serum Apo B levels are prevalent in patients with coronary heart disease (38), and reduction of Apo B is an independent factor associated with regression of documented coronary artery disease (39). The ratio of Apo B:Apo A1, an atherogenic risk index, tended to be higher (p = 0.06) in the hypercholesterolemic groups (1.46 ± 0.20 and 1.33 ± 0.11, placebo and pravastatin groups, versus 0.92 ± 0.11 in controls). After treatment, this ratio decreased in the pravastatin group (to 1.16 ± 0.14, p = 0.08) while remaining unmodified in the placebo group (1.31 ± 0.18, p = 0.89).

An LDL subclass pattern characterized by the predominance of small, dense LDL particles (pattern B) has been associated with an increased risk of coronary heart disease (40,41). As the size of the particle diminishes, the protein:lipid ratio and the density increase (42). In our series, the calculated protein:lipid ratio (Apo B:LDL-cholesterol) was 0.65 ± 0.03, 0.66 ± 0.03, and 0.60 ± 0.04 in the pravastatin, placebo, and control groups, respectively (p = NS). After treatment, the ratio increased to 0.76 ± 0.08 with pravastatin and to 0.73 ± 0.03 with placebo (p < 0.01 vs. baseline, p = NS for the difference between the two groups), thus adding evidence to the notion that low-fat diets can increase the density of LDL-cholesterol particles (43). In accord with previous evidence obtained in hypercholesterolemic patients (39), in our study pravastatin therapy itself did not adversely change LDL subclass pattern.

A clinically relevant issue is whether lipid-lowering treatment with an HMGCoA reductase inhibitor has any influence on carbohydrate metabolism. Other widely used antilipemic agents, such as nicotinic acid, can worsen glucose tolerance (44). As previously shown by us (45) and other researchers (46), isolated hypercholesterolemia is not associated with insulin resistance. Therefore, a further improvement in insulin sensitivity would be unexpected, but other metabolic changes may occur. We explored tolerance to oral glucose and the endogenous plasma insulin response to glucose and the ability of insulin to inhibit hepatic glucose production and lipolysis and to promote peripheral glucose uptake. For each of these responses at the end of the 8-week period, the pravastatin and placebo groups alike showed slight improvement, with no evidence of an added effect of pravastatin. As did other researchers using other HMG-CoA reductase inhibitors (47), we noted that pravastatin does not modify BP. Our results extend to nondiabetic hypercholesterolemic patients previous findings in diabetic individuals, in whom long-term treatment with pravastatin did not change fasting plasma glucose or glycohemoglobin levels or the insulin response to oral glucose (13).

In nondiabetic hypercholesterolemic patients, pravastatin therapy plus a reduced-fat isocaloric diet, in addition to improving the lipid and lipoprotein profile, is at least neutral (or marginally beneficial) with regard to glucose tolerance, insulin action, and energy metabolism. Whether the overall effects of pravastatin on carbohydrate metabolism are class- or drug-specific remains to be determined.

Acknowledgment: This work was supported in part by a grant from Bristol-Myers-Squibb.

FIG. 1.
FIG. 1.:
Changes in the low density:high density lipoprotein (LDL:HDL)-cholesterol ratio after 8 weeks of diet + placebo or diet + pravastatin treatment in hypercholesterolemia patients as compared with healthy controls.
FIG. 2.
FIG. 2.:
Plasma glucose (top) and insulin concentrations (bottom) in the fasting state and after oral glucose administration in the diet + placebo and diet + pravastatin group at baseline (lines) and after 8 weeks of treatment (dotted lines).
FIG. 3.
FIG. 3.:
Hepatic glucose production and glucose disposal rate during the insulin clamp in the diet + placebo and diet + pravastatin groups at baseline (solid lines, shaded columns) and after 8 weeks of treatment (dotted lines, solid columns).


1. Summary of the Second Report of the National Cholesterol Education Program (NCEP) Expert Panel On Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults. Adult treatment panel II. JAMA 1993;269:3015-23.
2. Verschuren WM, Jacobs DR, Blomberg PM, et al. Serum total cholesterol and long term coronary heart disease mortality in different cultures. Twenty-five-year follow up of the seven countries study. JAMA 1995;274:131-6.
3. Gotto AM Jr, LaRosa JC, Hunninghake D. The cholesterol facts. A summary of the evidence relating dietary fats, serum cholesterol, and coronary heart disease. Circulation 1990;81:1721-33.
4. Study Group of the European Atherosclerosis Society. Strategies for the prevention of coronary heart disease: a policy statement of the European Atherosclerosis Society. Eur Heart J 1987;8:7-88.
5. Pyöräla K, Savolainen E, Kaukola S, Haapakoski J. Plasma insulin as coronary heart disease risk factor: relationship to other risk factors and predictive value during 9 ½-year follow-up of the Helsinki Policemen Study Population. Acta Med Scand 1985;70:38-52.
6. Ducimetiere P, Eschwege E, Papoz L, Richard JL, Claude JR, Rosselin G. Relationship of plasma insulin levels to the incidence of myocardial infarction and coronary heart disease mortality in a middle-aged population. Diabetologia 1980;19:205-10.
7. Welborn TA, Wearne K. Coronary heart disease incidence and cardiovascular mortality in Busselton with reference to glucose and insulin concentrations. Diabetes Care 1979;2:154-60.
8. Stein EA. Drugs and alternative therapies for hyperlipidemia. Atherosclerosis 1994;108(suppl):S105-16.
9. Lipid Research Clinics Program. The Lipid Research Clinics Coronary Primary Intervention Trials results. II. The relationship in incidence of coronary heart disease to cholesterol lowering. JAMA 1984;251:351-74.
10. Loren P. The Oslo Diet Heart Study: eleven-year report. Circulation 1970;42:935-42.
11. Frick MH, Elo O, Happa K. Helsinki Heart Study: primary prevention with gemfibrozil in middle-aged men with dyslipidemia. N Engl J Med 1987;317:1237-45.
12. Scandinavian Simvastatin Survival Study Group. Randomized trial of cholesterol lowering in 4444 patients with coronary heart disease: the Scandinavian Simvastatin Survival Study (4S). Lancet 1994;344:1383-9.
13. Yoshino G, Kazumi T, Iwai M, et al. Long-term treatment of hypercholesterolemic non-insulin dependent diabetics (NIDDM) with pravastatin (CS-514). Atherosclerosis 1989;75:67-72.
14. National Diabetes Data Group. Classification and diagnosis of diabetes mellitus and other categories of glucose intolerance. Diabetes 1979;28:1039-57.
15. DeFronzo RA, Tobin JD, Andres R. Glucose clamp technique: a method for quantifying insulin secretion and resistance. Am J Physiol 1979;237:E214-233.
16. Ferrannini E. The theoretical bases of indirect calorimetry: a review. Metabolism 1988;37:287-301.
17. Hawk PB, Oser BL, Summerson WH. Practical physiological chemistry, 12th ed. Toronto: Blakiston, 1947:814-22.
18. Tappy L, Owen OE, Boden G. Effect of hyperinsulinemia on urea pool size and substrate oxidation rates. Diabetes 1988;37:1212-6.
19. Norwich KH. Measuring rates of appearance in systems which are not in steady state. Can J Physiol Pharmacol 1973;51:105-21.
20. Butler PC, Caumo A, Zerman A, O'Brien PC, Cobelli C, Rizza RA. Methods for assessment of the rate of onset and offset of insulin action during nonsteady state in humans. Am J Physiol 1993;264:E548-60.
21. Mari A. Estimation of the rate of appearance in the non-steady state with a two-compartment model. Am J Physiol 1992;263:E400-15.
22. Roeschlau P, Bernt T, Gruber W. Enzymatic determination of total cholesterol in serum. Z Klin Chem Klin Biochem 1974;12:226-32.
23. Wahlefeld AW. Triglycerides determination after enzymatic hydrolysis. In: Bergmeyer HU, ed. Methods of enzymatic analysis. San Diego: Academic Press, 1974:1831-5.
24. Enzi B, Baroni L, Piccoli A, Baiocchi MR, Fellin R. Cholesterol determination in HDL, HDL2 and HDL3 fractions after polyanion precipitation: a comparison between chemical extractive and totally enzymatic procedures. Clin Chim Acta 1985;146:81-6.
25. Mancini G, Carbonara AO, Heremans JF. Immunochemical quantitation of antigens by single radial immunodiffusion. Immunochemistry 1965;2:235-54.
26. De Long DM, De Long ER, Wood PD, Lippel K, Rifkind BM. A comparison of methods for the estimation of low and very low density lipoprotein cholesterol: the Lipid Research Clinic Prevalence Study. JAMA 1986;256:2372-7.
27. Lichtenstein AH, Ausman LM, Carrasco W, Jenner JL, Ordovas JM, Schaefer EJ. Short-term consumption of a reduced-fat diet beneficially affects plasma lipid concentrations only when accompanied by weight loss. Arterioscl Thromb 1994;14:1751-60.
28. Wiklund O, Angelin B, Bergman M, et al. Pravastatin and gemfibrozil alone and in combination for the treatment of hypercholesterolemia. Am J Med 1993;94:13-20.
29. Pravastatin Multicenter Study Group II. Comparative efficacy and safety of pravastatin and cholestyramine alone and combined in patients with hypercholesterolemia. Arch Intern Med 1993;153:1321-9.
30. Brussaard JH, Dallinga-Thie G, Groot PHE, Katan MB. Effects of amount and type of dietary fat on serum lipids, lipoproteins and apolipoproteins in man: a controlled 8-week trial. Atherosclerosis 1982;42:205-19.
31. Coulston AM, Liu GC, Reaven GM. Plasma glucose, insulin and lipid responses to high carbohydrate reduced-fat diets in normal humans. Metabolism 1983;32:52-6.
32. Mattsson FH, Grundy S. Comparison of effects of dietary saturated, monounsaturated and polyunsaturated fatty acids on plasma lipids and lipoproteins in man. J Lipid Res 1985;26:194-202.
33. Mensink RP, Katan MB. Effect of a diet enriched with monounsaturated or polyunsaturated fatty acids on levels of low-density and high-density lipoprotein cholesterol in healthy women and men. N Engl J Med 1989;321:436-41.
34. Kromhout D, Keys A, Aravanis C. Food consumption patterns in the 1960s in seven countries. Am J Clin Nutr 1989;49:889-94.
35. Verschuren WM, Jacobs DR, Blomberg PM, et al. Serum total cholesterol and long term coronary heart disease mortality in different cultures. Twenty-five-year follow up of the seven countries study. JAMA 1995;274:131-6.
36. Hunninghake DB, Stein EA, Dujovne CA, et al. The efficacy of intensive dietary therapy alone or combined with lovastatin in outpatients with hypercholesterolemia. N Engl J Med 1993;328:1213-9.
37. Clifton PM, Wight MB, Nestel PJ. Is fat restriction needed with an HMGCoA reductase inhibitor treatment? Atherosclerosis 1992;93:59-70.
38. Parra H, Fievet C, Boniface B. Lipoproteins, apolipoproteins, and coronary heart disease assessed by coronary arteriography. In: De Genes JL, Polovski J, Paoletti R, eds. Latent dyslipoproteinemias and atherosclerosis. New York: Raven Press, 1984:187-97.
39. Brown G, Albers JJ, Fisher LD, et al. Regression of coronary artery disease as a result of intensive lipid lowering therapy in men with high levels of apolipoprotein B. N Engl J Med 1990;323:1289-98.
40. Franceschini G, Cassiontti M, Vecchio G, et al. Pravastatin effectively lowers LDL cholesterol in familial combined hyperlipidemia without changing LDL subclass pattern. Arterioscler Thromb 1994;14:1569-75.
41. Austin MA, King MC, Vranizan KM, Krauss RM. Atherogenic lipoprotein phenotype: a proposed marker for coronary heart disease risk. Circulation 1990;82:495-506.
42. Rajman I, Maxwell S, Cramb R, Kendall M. Particle size: the key to the atherogenic lipoprotein? Q J Med 1994;87:709-20.
43. Dreon DM, Fernstrom HA, Miller B, Krauss R. Low density subclass patterns and lipoprotein response to a reduced fat diet in men. FASEB J 1994;8:121-6.
44. Garg A, Grundy SM. Nicotinic acid as therapy for dyslipidemia in non-insulin dependent diabetes mellitus. JAMA 1990;72:3-26.
45. Quiñones Galvan A, Santoro D, et al. Insulin sensitivity in familial hypercholesterolemia. Metabolism 1993;42:1359-64.
46. Paolisso G, Ferrannini E, D'Amore A, Volpe C, Varricchio M, D'Onofrio F. Effects of physiological plasma insulin levels on turnover parameters in familial hypercholesterolemia. Atherosclerosis 1993;101:111-5.
47. D'Agostino RB, Kannel WB, Stepanians MN, D'Agostino L. Efficacy and tolerability of lovastatin in hypercholesterolemia in patients with systemic hypertension. Am J Cardiol 1993;71:82-7.

Reduced-fat diet; 3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors; Insulin sensitivity; Glucose tolerance; Hypercholesterolemia

© Lippincott-Raven Publishers