© copyright 2009 by the american association for clinical chemistry agreement between fasting and...

© Copyright 2009 by the American Association for Clinical Chemistry

Agreement Between Fasting and Postprandial LDL Cholesterol Measured with 3 Methods in Patients with Type 2 Diabetes Mellitus

S.S. Lund, M. Petersen, M. Frandsen, U.M. Smidt, H.-H. Parving, A.A. Vaag, and T. Jensen

February 2011

http://www.clinchem.org/cgi/content/full/57/2/298


Journal ClubJournal Club


Introduction:Introduction:

The use of fasting or nonfasting lipid measurements is debated.

LDL cholesterol (LDL-C) is considered the primary target of lipid lowering therapy.(Adult Treatment Panel III (ATP-III), Circulation 2002).

International guidelines recommend using fasting samples to assess LDL-C (e.g., ATP III).

However, the use of nonfasting (postprandial) samples is more convenient.


Introduction (cont.):Introduction (cont.):

The relation between fasting and nonfasting LDL-C:

At the population level, nonfasting LDL-C is typically lower than fasting LDL-C as measured by the mean nonfasting-fasting difference.

However, at the individual level (i.e., for patient

management), the extent of agreement between nonfasting and fasting LDL-C is probably a more relevant measure.



Assessment of LDL-C.The LDL-C reference method, -quantification (BQ), is

cumbersome – it involves ultracentrifugation.

In daily practice, LDL-C is usually calculated by the Friedewald equation (FE), or measured by a direct assay (DA).

Unlike DA, the use of FE is recommended only in fasting samples and if triglycerides (TG) are <4.52 mM (400 mg/dL).

Contrary to recommendations, many routine clinics and research studies use FE in nonfasting samples.



Patients with type-2 diabetes (T2DM):In the postprandial state, T2DM patients are often

characterized by elevations of TG.

The elevated postprandial TG could interact with or influence the measurement of LDL-C.

Study aim:To evaluate the agreement between fasting and

postprandial LDL-C measured with 3 methods in patients with T2DM.


Questions:Questions:

How can fasting and postprandial concentrations of LDL-C be investigated?

Which LDL-C methods should be included in the investigation?


Material and methods:Material and methods:

Study protocol:

After an overnight fast of minimum 10 h, 74 T2DM patients were served a standard fat-rich breakfast meal.

The meal consisted of typical western foods: Bread, butter, cheese, milk, jam, and sausage (3515 kJ, 54% fat (50 g total fat),13% protein, 33% carbohydrates). Any medications were taken with the meal. Only drinking water was allowed postprandially.

Plasma was sampled at time 0 (fasting) as well as 1.5, 3.0, 4.5 and 6 h (postprandially).

Data are presented for 66 patients with complete measurements.


Material and methods:Material and methods: Study protocol (cont.):Measurements:

Total cholesterol (TC) and TG: enzymatic methods (Roche).

HDL cholesterol (HDL-C): Homogeneous assay (HDL-C Plus, Roche).

LDL-C:

1) Modified BQ (MBQ): Ultracentrifugation for 18 h at 105.000g. The bottom fraction (d>1.006 g/mL) containing the LDL and HDL particles was brought to volume and analyzed for TC and HDL-C. LDL-C was calculated as bottom fraction TC – HDL-C.

2) FE: LDL-C (mM) = TC – HDL-C – TG/2.2 (use TG/5 if in mg/dL) in samples with TG <4.52 mM (400 mg/dL).

3) DA: Homogeneous assay (LDL-C Plus, Roche), attempts to inhibit the cholesterol reactivity in non-LDL particles. This enables enzymatic measurement of LDL-C.


Questions:Questions:

How should agreement between fasting and postprandial LDL-C be evaluated?

What is a clinically relevant difference in LDL-C?


Statistics:Statistics:Agreement was evaluated according to the Bland-Altman method (Lancet 1986):

The ‘limits of agreement’ (LOA) given by the mean 1.96 x standard deviation (i.e., mean and 95% LOA) of the postprandial-fasting differences was calculated and plotted.

Acceptable agreement was concluded if the mean and 95% LOA of the postprandial-fasting difference in LDL-C was within 0.20 mM (7.7 mg/dL). This was considered to be a clinically relevant difference.(Law, BMJ 2003; Linsel-Nitschke, PlosOne 2008).


Statistics (cont.):Statistics (cont.):

Interpretation of the Bland-Altman method:The LOA is not a 95% confidence interval (CI) for the mean difference. It is a

prediction interval within which 95% of the individual data points, e.g., postprandial-fasting LDL-C differences, can be expected to lie.

About half of data points, differences, will typically lie between the mean and either of the lower or higher 95% LOA, respectively.

The 95% LOA can also be interpreted as the estimated ‘maximum’ disagreement between two methods of measurement, e.g., fasting and postprandial LDL-C.

The Bland-Altman method provides optimal clinical information, that for the population, the mean difference, and that for the individual, the LOA.

© Copyright 2009 by the American Association for Clinical Chemistry© Copyright 2009 by the American Association for Clinical Chemistry

Results:Results:

Table 1. Patient characteristics.Data are median (range) or numbers (%).*Calculated by FE.To convert mM to mg/dL multiply by 38.67 for cholesterol and by 88.57 for TG.

Gender (men/women) 55/11

Age (years) 61.7 (44.4; 81.2)

Known duration of diabetes (years) 5 (0; 29)

Body mass index (kg/m2) 24.6 (18.8; 28.2)

HemoglobinA1c (%) 7.7 (5.2; 10.7)

Glucose-lowering therapy,

(Diet only/oral agents/insulin) 25/41/0

Fasting TC (mM) 4.70 (2.80; 6.60)

Fasting LDL-C (mM)* 2.85 (1.50; 4.90)

Fasting HDL-C (mM) 1.14 (0.56; 1.93)

Fasting TG (mM) 1.27 (0.40; 3.40)

Ongoing statin treatment 17 (26)

Known cardiovascular disease 15 (23)


Figure 1. Fasting and postprandial LDL-C concentrations in T2DM patients measured by a modified quantification, the Friedewald equation and a direct assay.Data are meanSE. For all methods, mean LDL-C concentrations at all postprandial times were significantly lower, typically by about 0.2 to 0.4 mM, than time 0 (fasting), p<0.005 versus fasting for all.Data are also shown for TC, HDL-C and TG (meanSE, or geomean) (*p<0.05 versus fasting for these).


Figure 2. Bland-Altman plots of postprandial and fasting LDL-C measured by 3 methods in T2DM patients. Difference: Postprandial minus fasting LDL-C; Average: Mean of postprandial and fasting LDL-C. Solid line: The identity line. Dashed black lines: Mean difference with 95% LOA. Dashed grey lines: 95% CI. Grey area: The window of acceptable agreement of 0.2 mM. Acceptable agreement was unmet for all LDL-C methods at all times (e.g., LOA: –0 .52 to –0.89 mM at 4.5 h; data not shown at 3.0 or 6.0h).


Misclassification according to LDL-C <2.6 Misclassification according to LDL-C <2.6 mM (ATP-III) using postprandial LDL-C:mM (ATP-III) using postprandial LDL-C:

Table 2. Data are percentages and number of patients. For each method, the reference is the fasting LDL-C for that method. All: The 66 patient-group. Statin users: The 17 statin users. Misclassified patients: Those with fasting LDL-C >= 2.6 mM and at least one postprandial LDL-C <2.6. A: Misclassified patients (n1) among patients with fasting LDL-C >= 2.6 mM (n2).B: Misclassified patients (n1) among patients with at least one postprandial LDL-C <2.6 mM (n2).

In patients with fasting LDL-C >= 2.6 mM (columns A), 10-38%, and up to 63% of statin users, were misclassified into lower ATP-III risk categories using postprandial instead of fasting LDL-C.

In patients with postprandial LDL-C <2.6 mM (columns B), 24-50%, and up to 36% of statin users, were similarly misclassified.


Question:Question:

Do statin treatment or high postprandial TG influence the difference between postprandial and fasting LDL-C?


Figure 3. Fasting and postprandial LDL-C in T2DM patients partitioned by statin treatment or TG. Data are meanSE. TG partition was based on at least 1 postprandial TG concentration >2.08 mM (the median postprandial TG concentration). †p<0.05 for the change from time 0. *p<0.05 between groups for the change from time 0. §p<0.05 between groups for the absolute LDL-C concentrations. Unlike statin use, postprandial TG >2.08 mM meant greater postprandial decrease in LDL-C for all methods, most pronounced for FE.


Summary:Summary:For 3 LDL-C methods the mean LDL-C decreased by 0.2-0.4 mM postprandially in T2DM patients.

Postprandial and fasting LDL-C disagreed for all methods with 95% LOA (estimated ‘maximum’ disagreement) ranging from –0.52 to –0.89 mM at 4.5 h.That is, for FE, about half of patients may disagree of from ~0.4 mM (the mean) to ~0.9 mM (the LOA) lower postprandial than fasting LDL-C.

Using postprandial LDL-C, FE misclassified 38% and two-thirds of statin users into lower ATP-III risk categories.

Unlike statin use, postprandial TG >2.08 mM had greater disagreement between fasting and postprandial LDL-C for all methods.


Authors’ conclusions:Authors’ conclusions:

In T2DM patients, postprandial LDL-C concentrations might differ substantially from fasting concentrations, with postprandial LDL-C concentrations usually being lower.

The findings support that, irrespective of the method, postprandial LDL-C should not be used for assessing CVD risk.

© copyright 2009 by the american association for clinical chemistry agreement between fasting and...

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