Abstract 1641: Altered Hemoglobin is Associated with High Density Lipoproteins in Sera from Mice Fed Atherogenic/Hyperlipidemic Diets
Objective: Recent studies in both mice and humans suggest that the anti- or pro-inflammatory nature of HDL may be a sensitive predictor of risk for CHD events. We used ProteinChip technology coupled with surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) on serum and high density lipoproteins from mice fed atherogenic/hyperlipidemic diets, and identified an eight-protein core signature that serves as a serum biomarker panel for identifying pro-inflammatory HDL in mice. The objective of the present study was to characterize two of the potential biomarker peaks (m/z 14,900 and m/z 15,600) most dramatically associated with pro-inflammatory HDL
Methods and Results: Using micro-liquid chromatography-tandem mass spectrometry, we identified the SELDI peaks representing m/z 14,900 and m/z 15,600, as mouse hemoglobin alpha chain (Hb-Alpha, 14.9 kDa) and mouse hemoglobin beta chain (Hb-beta, 15.6 kDa), respectively. Western blot analysis confirmed the differential association of Hb with pro-inflammatory HDL when compared to normal HDL. Biochemical characterization of Hb associated with HDL further showed that the Hb associated with pro-inflammatory HDL possess distinct physical and chemical properties including reduced pI (pI 4.0 & pI 7.0 vs. pI 7.5 or higher for RBC Hb), and association with high molecular weight complexes found in fractions containing HDL. The main form of hemoglobin found associated with HDL was oxyhemoglobin (oxyHb).
Conclusion: Hb with distinct physical and chemical properties associates with pro-inflammatory HDL in animal models of atherosclerosis. Based on the pro-oxidant nature of oxyHb, our data suggest that Hb may contribute to the pro-inflammatory nature of HDL under atherogenic conditions. If these data can be extended to humans, HDL-associated Hb, may serve as a novel biomarker for atherosclerosis.