Quantification of apolipoprotein B in grossly normal human aorta

Area of Science:

• Biochemistry
• Cardiovascular Biology
• Lipid Metabolism

Background:

• Apolipoprotein B (apoB) is the primary protein component of human plasma low-density lipoproteins (LDL) and very-low-density lipoproteins (VLDL).
• Understanding apoB distribution in the arterial wall is crucial for cardiovascular research.

Purpose of the Study:

• To develop a quantitative electroimmunodiffusion (EID) assay for apolipoprotein B (apoB).
• To measure the concentration and localization of apoB within the intima of normal human aortas.

Main Methods:

• Development and validation of a quantitative electroimmunodiffusion (EID) assay for apoB, establishing specificity, sensitivity (30-200 ng), and reproducibility (11%).
• Analysis of buffer-soluble apoB in supernatant fractions from homogenates of normal human aortic intima.
• Differential ultracentrifugation to separate lipoprotein fractions (VLDL and LDL) from aortic homogenates.
• Electron microscopy to characterize particles within the LDL density fraction.

Main Results:

• The developed EID assay demonstrated high specificity, sensitivity, and reproducibility for apoB quantification.
• Intimal homogenates from normal aortas contained significant amounts of apoB (mean 5.42 ± 3.95 µg/mg dry weight), primarily in the supernatant fraction.
• No apoB was detected in the adjacent tunica media.
• Differential ultracentrifugation revealed that most intimal apoB was associated with the LDL density fraction, with negligible amounts in the VLDL fraction.
• Electron microscopy confirmed the presence of LDL-sized particles (200-250 Å diameter) in the LDL fraction.

Conclusions:

• A reliable quantitative assay for apoB has been established.
• Grossly normal human aortas contain substantial quantities of intact LDL, primarily within the intimal layer.
• The findings suggest that LDL infiltration into the intima may be a feature even in the absence of overt atherosclerotic lesions.