Dedicated to understanding the pathways for the generation of dysfunctional HDL and the roles of dysfunctional HDL in the pathogenesis of atherosclerosis, diabetes, and other inflammatory diseases in humans.
HDL – the good form of cholesterol – removes cholesterol from macrophages, which plays a key role in protecting humans from atherosclerosis. However, not all forms of HDL are cardioprotective. The research in my lab focuses on the impact of oxidative modifications and diabetes on the anti-atherogenic effects of HDL.
One major area of interest is myeloperoxidase (MPO), a heme enzyme secreted by macrophages. MPO is a source of oxidative stress in the human artery wall. We demonstrated that tyrosine 192 was the predominant site of both chlorination and nitration of apoA-I by MPO system. Importantly, when apoA-I is oxidized by MPO, its ability to promote cellular cholesterol efflux by the membrane-associated ATP binding cassette transporter A1 (ABCA1) pathway is diminished. To evaluate the functional significance of such damage, we generated a series of mutated forms of apoA-I, and then determined whether those mutant proteins were resistant to oxidative inactivation. These studies showed that tyrosine chlorination in concert with methionine oxidation impairs reverse cholesterol transport by apoA-I. Therefore, our studies strongly indicate that MPO represents a potential pathway for the generation of dysfunctional HDL.
Another potential mechanism for generating dysfunctional HDL involves covalent modification of apoA-I by reactive carbonyls, which have been implicated in atherogenesis and diabetic vascular disease. Indeed, modification of apoA-I by malondialdehyde (MDA) or acrolein also markedly impaired the lipoprotein’s ability to promote cellular cholesterol efflux by the ABCA1 pathway. Tandem mass spectrometric analyses revealed that these reactive carbonyls target specific Lys residues in the C-terminus of apoA-I. Importantly, immunochemical analyses showed that levels of MDA-protein adducts are elevated in HDL isolated from human atherosclerotic lesions. Also, apoA-I co-localized with acrolein adducts in such lesions. Thus, lipid peroxidation products might specifically modify HDL in vivo. Our observations support the hypotheses that reactive carbonyls might generate dysfunctional HDL in humans.
A major focus of the research in my lab is the development of LC-MS and LC-MS/MS methods for quantitative analysis of protein biomarkers in biological samples and in translational studies. We developed a label free quantitation approach to identify and quantify the oxidation products when proteins are exposed to various oxidative systems. The levels of oxidation products in HDL from in vivo samples are extremely low. In order to detect and quantify the oxidation products in HDL isolated from human plasma and lesions, we developed an LC-MS/MS approach using isotope-dilution selective reaction monitoring (SRM) or parallel reaction monitoring (PRM). Shotgun proteomics is a powerful approach for detection of proteins in biological samples, while targeted proteomics analysis based on SRM or PRM with isotope dilution provides precise relative quantification of proteins in complex mixtures. Alterations of HDL protein composition may impair its cardioprotective functions. In a recent study, we used both shotgun and targeted isotope dilution proteomics analyses to investigate alterations in the HDL proteome of end-stage renal disease (ESRD) patients undergoing hemodialysis. Our observations indicate that ESRD markedly remodels the HDL proteome and support the proposal that the protein cargo of HDL can serve as novel biomarkers for ESRD status.
Our lab uses cell biology together with tandem mass spectrometry to pinpoint specific amino acid residues in HDL that are damaged by reactive intermediates. Our long term goal is to understand the role of oxidative reactions and dysfunctional HDL in the pathogenesis of atherosclerosis, diabetes and other inflammatory diseases in humans.
Baohai Shao, PhD
Research Associate ProfessorBaohai Shao received his PhD from the Zhejiang University, China in 2002. After completing postdoctoral training in the laboratory of Dr. Jay Heinecke, he joined the Diabetes and Obesity Center of Excellence (now Diabetes Institute) in the Department of Medicine at the University of Washington.
UW Medicine Diabetes Institute
750 Republican Street, Box 358062
Seattle, WA 98109
Baohai Shao: (206) 616-8360
Laboratory: (206) 616-4524
Fax: (206) 543-3567
Baoshai Shao: firstname.lastname@example.org
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