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Gunther Marsche, PhD:

Impact of inflammation and oxidative stress on lipoprotein metabolism

Otto Loewi Research Centre, Division of Pharmacology, Medical University of Graz, Universitätsplatz 4, A-8010 Graz;
phone: +43-316-380 4513, fax: +43-316-380 9645, ✉ e-mail
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Keywords:

Inflammation, high-density lipoprotein, endothelium, gestational diabetes, leukocytes, cell adhesion, placenta, umbilical cord blood

Research interest:

A central research goal of our laboratory is to understand effects of inflammation and oxidant stress on lipoprotein metabolism. There is clear epidemiological evidence that plasma levels of high-density lipoprotein (HDL)-cholesterol are inverse and independent predictors of cardiovascular disease risk. However, we and others have shown that inflammation markedly modifies composition and function of HDL, affecting putative atheroprotective properties of HDL, including reverse cholesterol transport capability, as well as anti-oxidant and anti-inflammatory activities (1, 2). We are interested in understanding the molecular basis underlying inflammation-induced loss of protective activities of HDL since those qualitative alterations of HDL appear to be directly linked to increased cardiovascular complications and mortality. We study disease-specific changes in the proteome and lipidome of HDL, assess post-translational modifications of HDL, examine metrics of functional properties of HDL and several enzyme activities involved in lipoprotein maturation and metabolism (1). The role of oxidative stress in several inflammatory diseases is also a major research area. Protein oxidation induced by free radicals and peroxidases have shown to be of importance in many inflammatory- and oxidative stress-related diseases. Increased oxidative stress, a feature of inflammatory pregnancy disorders, including gestational diabetes and pre-eclampsia, promotes diverse alterations in the composition and metabolism of HDL (3). This is of particular importance, given that HDL represents the major cholesterol carrying lipoprotein class in human cord blood, while in maternal serum cholesterol is mainly carried by low-density lipoproteins. Previous findings suggested that HDL isolated from cord blood differs from maternally derived HDL with respect to its proteome, size and function (3). Mass and activity of the HDL-associated anti-oxidant enzyme paraoxonase are 5-fold lower in cord blood, accompanied by very low anti-oxidant capacity of fetal HDL. These results suggest that HDL is a major target for oxidation during fetal development, and dysfunctional HDL might promote fatty streak formation which is observed in fetal aortas under inflammatory conditions.

Illustrations:

Fig. 1: Oxidative modifications of HDL-associated apoA-I, apoE and albumin in fetal sera (5 pregnancies complicated by severe pre-eclampsia) and 5 fetal sera of healthy controls.
Sera were derivatized with dinitrophenylhydrazine, subjected to SDS  PAGE and transferred to poly(vinylidene  difluoride) membranes. HDL apolipo­proteins and albumin-associated carbonyls were detected using an anti-DNP antibody.

References:

  1. Birner-Gruenberger R, Schittmayer M, Holzer M, Marsche G: Understanding high-density lipoprotein function in disease: recent advances in proteomics unravel the complexity of its composition and biology. Prog Lipid Res, 2014; 56:36–46.
  2. Marsche G, Saemann MD, Heinemann A, Holzer M: Inflammation alters HDL composition and function: implications for HDL-raising therapies. Pharmacol Ther, 2013; 137(3):341–351.
  3. Sreckovic I, Birner-Gruenberger R, Besenboeck C, Miljkovic M, Stojakovic T, Scharnagl H, Marsche G, Lang U, Kotur-Stevuljevic J, Jelic-Ivanovic Z, Desoye G, Wadsack C: Gestational diabetes mellitus modulates neonatal high-density lipoprotein composition and its functional heterogeneity. Biochim Biophys Acta, 2014; 1841(11):1619–1627.

Collaborations within the DP-iDP:

  • G. Desoye will assist the students in detailed analysis of endothelial cell function such as proliferation/cell cycle assays and train students in the isolation of placental endothelial cells.
  • C. Wadsack will train the students in placental perfusion and to isolate fetal HDL.
  • M. Gauster will train the students in immunhistochemical analysis of placental tissue.
  • Á. Heinemann will train the students in isolating leukocytes from cord blood and assays of leukocyte-endothelial interaction, such as adhesion under flow and transendothelial migration.
  • M. van Poppel will introduce students to biostatistics and the biology of angiogenesis.

Collaborating research groups where PhD students could perform their research stay abroad:

  • Regine Heller (University of Jena, Germany) has a broad expertise in the investigation of key signaling molecules and their interaction in relation to endothelial functions such as vasorelaxation, migration or permeability.
  • Uwe Tietge (University of Groningen, Netherlands) will teach the students how to assess in vivo reverse cholesterol transport.
  • Volker Adams (University of Leipzig, Germany) is a cardiologist investigating impaired HDL function in obese children and patients with chronic heart failure. He will advise the students in exploring HDL-mediated endothelial repair and HDL induced stimulation of endothelial nitric oxide synthase.

Know-how and infrastructure of the research group:

Gunther Marsche’s laboratory has expertise in the field of lipoprotein metabolism and inflammatory vascular biology. Several techniques are routinely being used to study loss of function of post-translational modified lipoproteins and function / integrity of endothelial cells and macrophages. His laboratory uses a wide range of techniques spanning biophysical chemistry, molecular biology, cell biology and mass spectrometry to monitor the protein and lipid composition of HDL in the setting of cardiovascular disease, chronic kidney disease, liver failure, diabetes and psoriasis. His group is experienced in isolating and characterizing lipids / lipoproteins, in radioactive and non-radioactive labeling of lipids / lipoproteins, in cell culture and flow cytometry. Gunther Marsche’s research projects rely on chemical as well as analytical methods to identify specific reaction products and post-translational modifications to elaborate pathways responsible for disease, mass-spectrometric quantification of 3-chlorotyrosine (specific marker for myeloperoxidase-catalyzed chlorination at sites of inflammation), carbamyllysine, nitrotyrosine, carboxymethyllysine (major advanced glycation end product) and in analyses of HDL-associated anti-inflammatory enzymes (lecithin-cholesterol acyltransferase, platelet-activating factor acetylhydrolase, paroxonase-1). All required equipment is available at the institute, including animal and cell culture facilities, radionuclide laboratory, flow cytometryand a microscopy system to study cell-to-cell interaction under flow conditions. Mass spectrometry analyses are performed at the Centre for Medical Research and the Institute of Medical and Chemical Laboratory Diagnostics, both located at the MUG.

Scientific concepts and techniques that students will learn in this laboratory:

DP-iDP students will receive insights in lipid metabolism and the role of lipids in health and disease. The students will learn how to isolate lipoproteins either by ultracentrifugation or immune precipitation. The students will be trained to characterize and quantitate specific lipid subclasses and apolipoproteins by mass spectroscopy (GC-MS, LC-MS). They will be trained in radioactive and non-radioactive labeling of lipids / lipoproteins. The students will learn how to isolate platelets and leukocytes from peripheral blood. Further techniques that the students will acquire include flow cytometry to determine the expression of receptors and adhesion molecules, cell adhesion assays under flow, cholesterol efflux studies, endothelial regenerative capacity, Western blotting, multiplex ELISA to measure cytokine release, immunofluorescence microscopy, and assays to detect reactive oxygen species, phagocytosis, degranulation, and neutrophil–endothelial adhesion under flow.

28.05.2018 DP-iDP Doctoral Programme “Inflammatory Disorders in Pregnancy”