Catabolism of hypochlorite-modified low density lipoprotein (LDL) in vivo: insights from small animal PET studies

Ziel/Aim:

Oxidative modification of LDL apolipoprotein (apo) B-100 by myeloperoxidase-generated hypochlorite (HOCL) is regarded as a crucial event in atherogenesis. Recently, HOCl-modified LDL (OCl-LDL) have been shown to be present in human atherosclerotic lesions. On the other hand, data concerning the role of circulating OCl-LDL in the development of atherosclerosis are scarce. One reason for this is the shortage of methods for direct assessment of metabolism of oxidized LDL in vivo. We report an improved methodology for radiolabeling of both native LDL (nLDL) and OCl-LDL with the positron-emitter fluorine-18 (F-18) by N-succinimidyl-4-(F-18)fluorobenzoate (F-18-SFB) and the use of F-18-fluorobenzoylated LDL particles in dynamic PET studies in rats.

Methodik/Methods:

As a model, nLDL were modified in vitro by 3 mM NaOCl. For radiolabeling, pools of chemically and biochemically well characterized human nLDL and OCl-LDL were used. Module-assisted synthesis of F-18-SFB resulted in radiochemical yields of 36±2% (corrected for decay) and purites of >95%.

Ergebnisse/Results:

LDL labeling with F-18-SFB resulted in radiochemical yield of 30±10% (nLDL; corrected for decay) and 10±5% (OCl-LDL), respectively, with specific radioactivity of 50-400 GBq/µmol. Radiolabeling of native and modified LDL using F-18-SFB caused neither additional oxidative structural modifications of LDL lipids and proteins nor alteration of their biological activity and functionality in vitro, respectively. The method was further evaluated with respect to the uptake of F-18-fluorobenzoylated native and modified LDL, respectively, in various human cells. Biodistribution studies in rats revealed high in vivo stability for the fluorine-18-fluorobenzoylated LDL. The metabolic fate of F-18-fluorobenzoylated nLDL and OCl-LDL particles in vivo was delineated by dynamic PET studies using a dedicated small animal positron emission tomograph. Dynamic PET data demonstrated a significantly enhanced catabolism of OCl-LDL when compared with nLDL. The in vivo distribution and kinetics of both native and modified LDL correlated well with the anatomical localization of LDL receptors and scavenger receptors.

Schlussfolgerungen/Conclusions:

F-18-SFB-labeling of LDL and the use of small animal PET provide a valuable tool to discriminate the kinetics and the metabolic fate of both native and oxidized LDL in animal models in vivo.