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Clinical Chemistry 50: 1189-1200, 2004. First published April 23, 2004; 10.1373/clinchem.2004.032763
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(Clinical Chemistry. 2004;50:1189-1200.)
© 2004 American Association for Clinical Chemistry, Inc.

Lipids, Lipoproteins, and Cardiovascular Risk Factors

Sex and Age Differences in Lipoprotein Subclasses Measured by Nuclear Magnetic Resonance Spectroscopy: The Framingham Study

David S. Freedman1,a, James D. Otvos2, Elias J. Jeyarajah2, Irina Shalaurova2, L. Adrienne Cupples3, Helen Parise4, Ralph B. D’Agostino4, Peter W.F. Wilson4 and Ernst J. Schaefer5

1 Division of Nutrition and Physical Activity, CDC, Atlanta, GA. 2 LipoScience, Inc, Raleigh, NC. 3 Boston University School of Public Health, Boston, MA. 4 Boston University School of Medicine, Boston, MA. 5 Jean Mayer-US Department of Agriculture Human Nutrition Research Center on Aging at Tufts University, Boston, MA.

aAddress correspondence to this author at: CDC K26, 4770 Buford Hwy, Atlanta, GA 30341-3717. Fax 770-488-6027; e-mail DFreedman{at}cdc.gov.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Background: The sex differential in coronary heart disease (CHD) risk, which is not explained by male/female differences in lipid and lipoprotein concentrations, narrows with age. We examined whether this differential CHD risk might, in part, be attributable to the sizes of lipoprotein particles or concentrations of lipoprotein subclasses.

Methods: We analyzed frozen plasma samples from 1574 men and 1692 women from exam cycle 4 (1988–1990) of the Framingham Offspring Study. Nuclear magnetic resonance (NMR) spectroscopy was used to determine the subclass concentrations and mean sizes of VLDL, LDL, and HDL particles. Concentrations of lipids and apolipoproteins were measured by standard chemical methods.

Results: In addition to the expected sex differences in concentrations of triglycerides, LDL-cholesterol, and HDL-cholesterol, women also had a lower-risk subclass profile consisting of larger LDL (0.4 nm) and HDL (0.5 nm) particles. The sex difference was most pronounced for HDL, with women having a twofold higher (8 vs 4 µmol/L) concentration of large HDL particles than men. Furthermore, similar to the narrowing of the sex difference in CHD risk with age, the observed male/female difference in HDL particle size also decreased with age. Although lipoprotein particle sizes were highly correlated with lipid and lipoprotein concentrations, the sex differences in the mean sizes of lipoprotein particles persisted (P <0.001) even after adjustment for lipid and lipoprotein concentrations.

Conclusions: Women have a less atherogenic subclass profile than men, even after accounting for differences in lipid concentrations.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
The sex differential in coronary heart disease (CHD)1 narrows with age, but the death rate among women never exceeds that of men (1)(2). Concentrations of lipids and lipoproteins may partly explain this sex difference, and compared with similarly aged men, premenopausal women have lower LDL-cholesterol and triglyceride (TG) concentrations and higher HDL-cholesterol concentrations (3). Several of these male/female differences in lipid concentrations are reduced (or reversed) at older ages, but because the sex differential in CHD risk exists even at comparable lipid and lipoprotein concentrations (2), other factors need to be considered.

For example, a predominance of small, dense LDL subclass particles (pattern B) is more prevalent among men than among women (4)(5)(6)(7)(8). A small LDL particle size, as characterized by gradient gel electrophoresis (GGE), is associated with an increased CHD risk, but the association is not independent of concentrations of total cholesterol (TC), HDL-cholesterol, and TGs (9)(10)(11). It is possible, however, that the characterization of LDL particles on the basis of LDL subclass distribution or phenotype is not optimal. Small LDL concentration as determined by GGE has been found to be independently associated with CHD risk (12). Independent associations with CHD (or the extent of coronary artery disease) have also been found in studies in which density gradient ultracentrifugation (13)(14) or nuclear magnetic resonance (NMR) spectroscopy (15)(16)(17)(18)(19)(20) were used to characterize LDL subclass heterogeneity in a quantitative, rather than qualitative, manner.

The metabolic reactions responsible for high concentrations of small LDL particles produce abnormalities in VLDL and HDL subclasses that may also contribute to CHD risk. For example, insulin resistance is associated with overproduction of large VLDL particles (21), and independent of the TG concentration, concentrations of large VLDL are associated with coronary artery disease (15) and coronary calcium (19). Differing associations across HDL subclasses have also been noted, with a protective effect most consistently observed for the large (e.g., HDL2b) subfraction(s) (15)(18)(22)(23)(24).

Because the traditional methods of lipoprotein subfractionation are laborious and time-consuming, the possible contribution of lipoprotein subclasses to the sex differential in CHD has not been studied extensively. In the current study, we used a rapid and efficient NMR spectroscopic method of analysis to examine sex and age differences in VLDL, LDL, and HDL subclasses in a large, population-based sample.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
sample
The individuals included in this study were participants in exam cycle 4 (1987–1990) of the Framingham Offspring Study, and all procedures were approved by the appropriate Institutional Review Board with participants giving informed consent. NMR lipoprotein determinations were performed among 3523 persons for whom a frozen plasma specimen was available.

Individuals were excluded from the analyses for any the following reasons: (a) reported use of cholesterol-lowering drugs (n = 136); (b) a missing chemical lipid or lipoprotein determinations (n = 60); or (c) a fasting TG concentration ≥4 g/L (n = 72). Hypertriglyceridemic individuals were excluded because LDL-cholesterol concentrations could not be estimated (25). (Freezing may also alter NMR-measured VLDL subclasses of persons with hypertriglyceridemia.) These exclusions gave 3266 individuals for the current analyses.

chemical analyses of lipids and lipoproteins
As described previously (26), blood was drawn from participants after a 12-h fast into tubes containing EDTA (1.5 g/L) and centrifuged at 1000g for 20 min at 4 °C to obtain plasma. TC, TG, and HDL-cholesterol concentrations were determined enzymatically with an Abbott Diagnostics ABA-200. LDL-cholesterol was estimated, among those with a TG concentration <4.0 g/L, as TC – HDL-cholesterol – TG/5 (25). Plasma concentrations of apolipoprotein A-1 (apoA-I) were measured by noncompetitive ELISA with the use of affinity-purified polyclonal antibodies (27), and apolipoprotein B (apoB) was measured with a commercial immunoturbidimetric assay (Incstar Corp.) (26).

nmr analysis of lipoprotein subclasses
Plasma samples stored at –70 °C for 6–10 years were analyzed by a commercially available proton NMR spectroscopic assay (LipoScience) at North Carolina State University. In brief, this method uses the characteristic signals broadcast by lipoprotein subclasses of different size as the basis of their quantification (28)(29). Each subclass signal emanates from the total number of terminal methyl groups on the lipids contained within the particle core (cholesterol ester and TGs, each contributing three methyl groups) and in the surface shell (phospholipid and unesterified cholesterol, each contributing two methyl groups). The number of methyl groups contained within a particle depends, to a close approximation, only on the diameter of the particle and is not affected by differences in lipid composition arising from variations in (a) the relative amounts of cholesterol ester and TGs in the particle core, (b) the degree of unsaturation of the lipid fatty acyl chains, or (c) phospholipid composition. The methyl NMR signal emitted by each subclass therefore provides a direct measure of the concentration of that subclass.

NMR spectra of each plasma specimen (0.4 mL) were acquired in duplicate by an automated 360 MHz lipoprotein analyzer, and the lipid methyl signal envelope was decomposed computationally to give the amplitudes of the contributing signals of 16 subclasses [1 for chylomicrons; 6 for VLDL; 1 for intermediate-density lipoprotein (IDL); 3 for LDL; and 5 for HDL]. Conversion factors relating signal amplitudes to subclass concentrations expressed in particle concentration units or in lipid mass concentration units (cholesterol or TGs) were then applied. These factors were derived from NMR and chemical analyses performed on a set of purified subclass standards of defined size, which were isolated from a diverse group of normo- and dyslipidemic individuals by a combination of ultracentrifugation and agarose gel- filtration chromatography. Particle concentrations (nmol/L for VLDL and LDL; µmol/L for HDL) were calculated for each subclass standard by measuring the total concentration of core lipid (cholesterol ester plus TGs) and dividing the volume occupied by these lipids by the core volume per particle calculated from the diameter of that particle (30). From lipid measurements performed on each subclass standard, conversion factors were also generated to give lipid mass concentration estimates of the VLDL subclasses (g/L TGs) and LDL and HDL subclasses (g/L cholesterol).

We emphasize that NMR-derived lipid values are estimates obtained from direct measurement of the lipoprotein particles carrying the lipids, not from an actual lipid measurement. Because the calculation of these lipid concentrations assumes that each subclass particle contains the same amount of cholesterol and TGs as the corresponding isolated subclass standard, they therefore represent the lipid values for a person with lipoproteins of "normal" composition.

The 16 measured subclasses were grouped for analysis into the following categories: 3 VLDL subclasses [large, >60 nm (including chylomicrons); intermediate, 35–60 nm; small, 27–35 nm], IDL (23–27 nm), 3 LDL subclasses (large, 21.3–23 nm; intermediate, 19.8–21.2 nm; small, 18.3–19.7 nm), and 3 HDL subclasses (large, 8.8–13 nm; intermediate, 8.2–8.8 nm; small, 7.3–8.2 nm). The IDL and LDL subclass diameters, which were ~5 nm smaller than those estimated by GGE, were consistent with both electron microscopy (31) and LDL lipid compositional data (32). Weighted average VLDL, LDL, and HDL particle sizes (nm diameter) were computed as the sum of the diameter of each subclass multiplied by its relative mass percentage as estimated from the amplitude of its methyl NMR signal.

LDL and HDL subclass distributions determined by GGE and NMR are highly correlated (16)(33). Replicate analyses of plasma pools indicated that NMR subclass measurements are reproducible, with CV <3% for NMR-derived concentrations of VLDL, LDL, and HDL; <4% for VLDL size; and <1% for LDL and HDL size (29).

statistical methods
Male/female differences in lipoprotein subclass concentrations were examined, and linear regression was used to determine whether these sex differences were independent of TG, LDL-cholesterol, and HDL-cholesterol concentrations. Although concentrations of several NMR subclasses were skewed toward higher values, the use of robust regression techniques (34) and ordinary (least-squares) regression (with and without square-root transformations) yielded similar results. Spearman rank correlations were used to examine the relationship of NMR-derived lipoprotein concentrations and particle sizes with chemically determined lipoprotein concentrations, and age trends were examined graphically using lowess, a robust smoothing technique based on weighted linear regression. Possible sex differences in the relationship of age to NMR-determined subclass concentrations were assessed by including terms for both sex x age and sex x age2 in regression models.

Several of these analyses were performed for NMR lipoprotein concentrations expressed in both particle concentration and lipid mass concentration units. Lipoprotein lipid mass concentrations are strongly influenced by the largest subclass particles (because larger particles contain more lipid than smaller particles), whereas lipoprotein particle concentrations are influenced by the smallest subclass particles.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
There was no sex difference in the mean TC concentration, but men had higher concentrations of TGs, LDL-cholesterol, and apoB, and women had higher concentrations of HDL-cholesterol and apoA-I (Table 1 ). Sex differences in lipoprotein particle concentrations generally paralleled these findings. Men had higher concentrations of (total) VLDL particles and a larger mean VLDL particle size because of higher concentrations of large and intermediate VLDL subclasses. Men also had a higher (total) LDL particle concentration because of higher concentrations of small and intermediate LDL subclasses, but women had higher concentrations of the large LDL subclass. These distributional differences led to men having a 0.4-nm smaller mean LDL particle size and a threefold higher (34% vs 11%) prevalence of the small (≤20.5 nm) LDL phenotype. Interestingly, there was no sex difference in the concentration of (total) HDL particles despite the 0.12 g/L higher mean HDL-cholesterol concentration for women. Men had higher concentrations of small and intermediate HDL particles, but the concentration of cholesterol-rich large HDL particles was twofold higher (8 vs 4 µmol/L) among women.


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  		Table 1. Values for selected characteristics, by sex.3

The Spearman (rank) correlations between various NMR determinations and chemically measured lipid and lipoprotein concentrations are shown in Table 2 . As expected, TG concentrations were highly correlated with lipid mass concentrations of VLDL (r ~ 0.9) and with the size of VLDL (r = 0.54–0.71), LDL (r = –0.30 to –0.55), and HDL (r ~ –0.5) particles. HDL-cholesterol concentrations were strongly associated with HDL lipid mass (r ~ 0.9) and the sizes of HDL (r ~ 0.7) and LDL (r = 0.39–0.57) particles. Strong associations were also observed between apoB and LDL particle concentrations (r = 0.86–0.88) and between Friedewald- and NMR-derived LDL-cholesterol concentrations (r = 0.83).


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  		Table 2. Relationship of chemically determined lipid and lipoprotein concentrations with NMR-determined particle concentration, lipid mass, and size.

Because of these intercorrelations, we examined whether the sex differences in particle sizes existed at comparable concentrations of TGs and HDL-cholesterol (Fig. 1 ). The overall sex difference in VLDL particle size was reduced by 50% (from 4 nm to 2 nm; Fig. 1 , upper left panel) when comparisons were made at similar TG concentrations; furthermore, at HDL-cholesterol concentrations <0.35 g/L, the mean VLDL particle size was slightly larger among women than among men (Fig. 1 , upper right panel). In contrast, women had larger LDL (Fig. 1 , middle panels) and HDL (Fig. 1 , bottom panels) particles than did men at almost all concentrations of TGs and HDL-cholesterol. Many of these associations were markedly nonlinear. For example, the mean HDL particle size decreased by ~0.6 nm between TG concentrations of 0.5 and 2.0 g/L, but there was no further decrease as the TG concentration approached 4.0 g/L.



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  		Figure 1. Relationship of VLDL (top), LDL (middle), and HDL (bottom) particle sizes with chemically determined concentrations of TG (left panels) and HDL-cholesterol (right panels).

Associations were estimated, within each sex, by lowess. Black dashed line, men; solid gray line, women.

We then examined whether the observed sex differences in the NMR determinations were independent of TG, LDL-cholesterol, and HDL-cholesterol concentrations in regression analyses (Table 1Up , last column). Adjusting for concentrations of these lipids (considered jointly) eliminated the sex differences in apoB and apoA-I concentrations. However, even at equivalent lipid concentrations, the VLDL particle size remained 2 nm larger among men, whereas the sizes of LDL (0.1 nm) and HDL (0.2 nm) particles remained larger among women (P <0.001 for all comparisons). Furthermore, concentrations of small LDL and small HDL remained higher among men, whereas women had higher adjusted concentrations of large LDL and large HDL.

Tables 3 through 5 present various percentiles of VLDL, LDL, and HDL particle sizes and subclass concentrations (in both particle and lipid mass concentration units), along with correlations with age. As seen in Table 3 , TG concentrations were more strongly associated with age among women than men (r = 0.34 vs 0.17), and comparable differences were seen for each subclass. For example, whereas the median concentration of small VLDL particles was 7 nmol/L higher among men than women before age 45 years, among older individuals (≥55 years), the median concentration was 4 nmol/L higher among women.


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  		Table 3. Selected percentiles (10th, 50th, and 90th) of VLDL particle and lipid mass concentrations, by sex and age.1


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  		Table 4. Selected percentiles of LDL particle and lipid mass concentrations, by sex and age.1


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  		Table 5. Selected percentiles of HDL particle and lipid mass concentrations, by sex and age.1

As seen in Table 4Up and Fig. 2 , LDL-cholesterol and LDL particle concentrations were also more strongly associated with age among women (r = 0.35–0.36) than among men (r = 0.03–0.12). After 55 years of age, the mean LDL-cholesterol concentration was higher among women than men, but the concentration of LDL particles remained higher among men until age 70 (Fig. 2 ). These analyses also indicated that the male/female crossover in LDL-cholesterol concentrations was attributable to large and intermediate particles. With increasing age, the female excess in concentrations of large LDL particles increased from ~0 to 200 nmol/L, whereas the male excess in intermediate LDL decreased. In contrast, the male excess in concentrations of small LDL particles increased between the ages of 25 and 50 years. LDL size was weakly (inversely) associated with age among both men and women, but the mean LDL particle size was 0.3–0.4 nm smaller among men than among women at all ages >35 years.



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  		Figure 2. Relationship of LDL-cholesterol, LDL size, LDL particle concentration, and LDL subclasses with age.

Within each sex, separate lines were constructed by use of lowess. Black dashed line, men; solid gray line, women.

Although HDL-cholesterol concentrations were weakly associated with age (r = –0.07 for men; –0.03 for women), we observed stronger associations with concentrations of the HDL subclasses (Table 5Up and Fig. 3 ). Among women, but not men, age was inversely associated with concentrations of large HDL but positively associated with concentrations of small and intermediate HDL. These differing associations led to the mean HDL size decreasing with age among women (r = –0.10) but increasing with age among men (r = 0.13).



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  		Figure 3. Relationship of HDL-cholesterol, HDL particle size, and HDL subclasses with age.

Within each sex, separate lines were constructed by use of lowess. Black dashed line, men; solid gray line, women.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
In this large, population-based study, we examined whether male/female differences in the sizes of lipoprotein particles and concentrations of lipoprotein subclasses can, at least in part, account for the sex difference in CHD risk. We found that men had smaller LDL and HDL particles, but larger VLDL particles, than did women. Furthermore, despite the strong associations between lipoprotein particle sizes and TG and HDL-cholesterol concentrations, the sex differences in particle sizes persisted even when comparisons were made at equivalent lipid concentrations. We also found that the sex difference in HDL particle size decreased with age, a trend similar to that for the male/female difference in CHD, and that the increased LDL-cholesterol concentration of older women is attributable to high concentrations of intermediate and large LDL, rather than small LDL.

Of the various characteristics that we examined, CHD risk has been most extensively studied in relation to LDL subclasses and particle size. Although persons with a relatively high proportion of small, dense LDL particles are at increased risk for CHD, this risk appears to be attributable to the adverse concentrations of TGs and HDL that are frequently seen among these persons (9)(10)(11). [In the current study, TG and HDL-cholesterol concentrations could statistically account for 33% (women) to 44% (men) of the variability in LDL particle size.] Other characteristics of LDL particles, however, such as the amount of cholesterol in small LDL particles (12), the LDL particle concentration (16)(17) (which largely reflects concentrations of small and intermediate LDL), and concentrations of small LDL (13)(14)(15)(18)(19), appear to provide additional, independent information on CHD risk, the extent of coronary artery disease, and coronary calcium.

Men have smaller LDL particles than women (4)(5)(6)(7)(8)(35), and we found that 34% of men but only 11% of women had a mean LDL particle size ≤20.5 nm. This sex difference is similar to that observed for a GGE-measured LDL peak particle diameter of <25.5 nm (pattern B) (5). Also in agreement with our findings, most investigators have found little change in LDL particle size with age (6)(36). However, we found that the sex difference in concentrations of (total) LDL particles narrowed with age, a trend that parallels the narrowing of the sex differential in CHD risk with age (1)(2). It is also possible that the increase in concentrations of small LDL among men (but not women) before age 50 years may be associated with the higher CHD risk seen among middle-aged (vs younger) men.

Several cross-sectional studies have also shown that quantification of HDL subclasses by GGE, two-dimensional gel electrophoresis, or NMR may provide information on CHD risk beyond that conveyed by the HDL-cholesterol concentration. Compared with controls, individuals with CHD or occlusive disease have frequently been found to have low concentrations of the larger HDL subclasses (HDL2b and lipoprotein A-I {alpha}1 and pre-{alpha}) along with high concentrations of small HDL, yielding a smaller mean HDL particle size (15)(18)(22)(23)(24). In contrast, quantification of the HDL2 and HDL3 subclasses does not improve the prediction of CHD (37). Whereas women had larger HDL particles than men, we found that this sex differential narrowed by ~50% with increasing age. Age-related changes in HDL subclasses have been reported by other investigators, with concentrations of HDL3b increasing after menopause (38) and concentrations of small HDL increasing during puberty among boys (39). There is also some evidence that HDL-cholesterol concentrations are more strongly (inversely) associated with CHD risk among women than men (40), and this may, in part, be attributable to the larger mean HDL particle size of women.

Although the sex difference in CHD risk is independent of several risk factors (2), males with diabetes are at only slightly greater risk for CHD than are females with diabetes (40)(41). Dyslipidemia among individuals with type 2 diabetes is characterized not only by high TG concentrations and low HDL-cholesterol concentrations, but also by increases in the size of VLDL particles and decreases in the size of LDL and HDL particles (41)(42)(43). These differences parallel the male/female differences observed in the current study. In addition, it has been found that (a) insulin resistance is associated with smaller HDL particles (41), (b) diabetes is associated with a greater reduction in LDL particle size among women than among men (6)(8)(36)(41), and (c) the mean VLDL particle size among individuals with type 1 diabetes is larger among women than among men (44). These sex difference in lipoprotein subclasses could possibly account for the greatly increased CHD risk of females with diabetes.

Increased plasma TG concentrations, carried mainly in large VLDL particles, are necessary, but not sufficient to promote the cycle of lipid exchange and lipolysis that leads to small LDL and HDL subclasses (7)(42). The process begins with a reaction catalyzed by cholesterol ester transfer protein, in which TGs from the core of (mainly large) VLDL exchange one-for-one with cholesterol ester molecules in LDL and HDL particles. When LDL and HDL particles are depleted in cholesterol ester and enriched in TGs, they become a substrate for hepatic lipase (HL), which produces smaller, denser particles. Therefore, the concentrations and/or activities of these enzymes can potentially influence the distributions of LDL and HDL subclasses at any given plasma TG concentration, and HL may be a determinant of sex- and diabetes-related differences in these subclasses. Normolipidemic men have approximately twice the HL activity of women (7)(45), and HL activity is significantly increased among individuals with diabetes (42). Although there are exceptions (46), HL activity has generally been found to be inversely correlated, independently of the TG concentration, with the sizes of LDL (47) and HDL(33) particles. Our results showing that women have larger LDL and HDL particle sizes than men, even after adjustment for lipid and lipoprotein concentrations, are consistent with these previous findings.

From an analytical perspective, the NMR spectroscopic method of lipoprotein subclass analysis used in this study has several advantages over the classic techniques of electrophoresis and ultracentrifugation in terms of ease and speed of measurement. By avoiding the need for physical fractionation of lipoprotein subspecies, which requires several hours to days of effort and achieves only partial resolution of the subspecies, the NMR process requires only minutes and is completely automated (29). NMR also eliminates the sources of analytical variability inherent in separation procedures. Perhaps most relevant to the assessment of CHD risk is that the NMR method provides a direct measure of lipoprotein subclass particle concentrations. Rather than basing quantification on the amount of cholesterol contained within a population of lipoprotein particles or the relative degree of lipid/protein staining of separated particles, NMR uses the characteristic signals broadcast by lipoprotein subclasses of different sizes (48). The measured amplitudes of these subclass NMR signals are directly proportional to the number of particles emitting the signal, even when the amount of lipid or protein per particle varies from person to person. As a result, NMR-derived lipoprotein concentrations may therefore differ from those measured by traditional methods.

Despite the differences between NMR and other methods used to quantify lipoprotein subspecies, studies of split samples have shown good agreement between LDL and HDL particle sizes measured by NMR and GGE (16)(33). In addition, many of the sex differences in lipoprotein particle sizes and subclasses that we observed, as well as the associations with concentrations of lipids and lipoproteins, are similar to those reported by others. For example, the overall correlations that we observed between TG and LDL size in the current study (r = –0.30 for women to –0.55 for men) are similar to those reported by other investigators (35)(49)(50), as is the stronger association among men (7)(48). Moreover, results from nested case–control studies of frozen specimens indicate that NMR-derived lipoprotein particle concentrations can improve the prediction of CHD risk beyond that obtained with lipid and apolipoprotein measurements (15)(16)(17)(18)(19)(20).

There are, however, various limitations of our study that should be considered. Although the sex differences that we observed are consistent with male/female differences in CHD risk, we did not have information on subsequent CHD and therefore cannot assess whether controlling for the lipoprotein subclass differences would have reduced the male excess in CHD. In addition, there is measurement variability in the NMR determinations, with CV <4% for VLDL size, <1% for LDL and HDL sizes, and <3% for total VLDL, LDL, and HDL concentrations (29). However, assuming that measurement reliability does not vary by sex and age, the magnitudes of the associations that we observed would have been biased toward 0. Another limitation is that, at the present time, NMR lipoprotein testing is performed in only a single reference laboratory in Raleigh, NC. However, a more robust and reliable NMR analyzer is under development to enable general laboratory usage.

In conclusion, our results indicate that, compared with men, women have a less atherogenic lipoprotein subclass profile that consists of smaller-sized VLDL and larger-sized LDL and HDL. This pattern persists after controlling for differences in plasma lipid and lipoprotein concentrations. In addition, our data provide reference values for men and women of various ages for the many lipoprotein values measured by NMR spectroscopic analysis.


   Acknowledgments
 
This study was supported by NIH Grant HL-43230.


   Footnotes
 
1 Nonstandard abbreviations: CHD, coronary heart disease; TG, triglyceride; GGE, gradient gel electrophoresis; TC, total cholesterol; NMR, nuclear magnetic resonance; apoA-I, apolipoprotein A-I; apoB, apolipoprotein B; IDL, intermediate-density lipoprotein; and HL, hepatic lipase.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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