Collaborative Research from the Center for Membrane Biosciences

poster abstractThe Center for Membrane Biosciences has been facilitating new research activities between the IUPUI School of Science and IU School of Medicine in the structure, biochemistry, and physiology of biological membranes. Results from two projects resulting from these collaborations are presented. Project 1: Ceramides are sphingolipids involved in the development of lung alveolar cell apoptosis (programmed death) and possibly in the clearance of apoptotic cells by alveolar macrophages. We use a combination of molecular and cellular methods to determine the effect of ceramides on the ability of alveolar macrophages to engulf apoptotic cells. Engulfment experiments of labeled apoptotic Jurkat cells were performed with rat alveolar macrophages (AM) obtained via bronchoalveolar lavage. AM were treated with various ceramide species and efferocytosis was quantified by flow cytometry. Using small-angle X-ray scattering and solid state 2H NMR we determined how ceramides (C6:0, C18:1) affect the molecular organization and the physical properties of model membranes. These studies can lead to a better understanding of the molecular mechanisms responsible for apoptotic cell clearance. If the clearance process is impaired, apoptotic cells may progress to secondary necrosis, resulting in release of harmful cellular contents and tissue inflammation. Project 2: Highly-photostable quantum dots (QD) conjugated to lipids or antibodies can be utilized to explore changes in compartmentalization of the plasma membrane due to hyperinsulinemia using wide field single molecule fluorescence microscopy. Protocols describing the bio-inertness and monovalent binding of QDs to antibodies are outlined, as well as use of confocal fluorescence correlation spectroscopy to determine colloidal stability of CdSe/ZnS QDs in aqueous solution. Tracking experiments on QD-conjugated to transferrin receptors in healthy and insulin-resistant adipocytes detect changes in membrane compartmentalization. The impact of chromium picolinate on receptor mobility was also investigated

Effects of Lipid Interactions on Model Vesicle Engulfment by Alveolar Macrophages

The engulfment function of macrophages relies on complex molecular interactions involving both lipids and proteins. In particular, the clearance of apoptotic bodies (efferocytosis) is enabled by externalization on the cell target of phosphatidylserine lipids, which activate receptors on macrophages, suggesting that (local) specific lipid-protein interactions are required at least for the initiation of efferocytosis. However, in addition to apoptotic cells, macrophages can engulf foreign bodies that vary substantially in size from a few nanometers to microns, suggesting that nonspecific interactions over a wide range of length scales could be relevant. Here, we use model lipid membranes (made of phosphatidylcholine, phosphatidylserine, and ceramide) and rat alveolar macrophages to show how lipid bilayer properties probed by small-angle x-ray scattering and solid-state 2H NMR correlate with engulfment rates measured by flow cytometry. We find that engulfment of protein-free model lipid vesicles is promoted by the presence of phosphatidylserine lipids but inhibited by ceramide, in accord with a previous study of apoptotic cells. We conclude that the roles of phosphatidylserine and ceramide in phagocytosis is based, at least in part, on lipid-mediated modification of membrane physical properties, including interactions at large length scales as well as local lipid ordering and possible domain formation

Silver-Containing Thin Films on Transparent Polymer Foils for Antimicrobial Applications

The increasing occurrence of infections caused by pathogens found on objects of everyday use requires a variety of solutions for active disinfection. Using active materials that do not require daily maintenance has a potential advantage for their acceptance. In this contribution, transparent films, with silver as the main antimicrobial agent and a total thickness of a few tens of nm, were deposited on flexible self-adhesive polymer foils used as screen protectors. TiO2 and SiO2 were used as transparent matrix to embed the Ag nanoparticles, ensuring also their mechanical protection and controlled growth. HiPIMS (High-Power Impulse Magnetron Sputtering) was used for the sputtering of the Ag target and fine control of the Ag amount in the layer, whereas TiO2 and SiO2 were sputtered in RF (Radio Frequency) mode. The thin film surface was investigated by AFM (Atomic Force Microscopy), providing information on the topography of the coatings and their preferential growth on the textured polymer foil. XRD (X-Ray Diffraction) revealed the presence of specific Ag peaks in an amorphous oxide matrix. UV-Vis-NIR (Ultraviolet-Visible-Near Infrared) spectroscopy revealed the presence of nanostructured Ag, characterized by preferential absorption in the 400 to 500 nm spectral range. The antimicrobial properties were assessed using an antimicrobial test with the Escherichia coli strain. The highest efficiency was observed for the Ag/SiO2 combination, in the concentration range of 104–105 CFU/mL