Erythrocyte Membrane Biophysical Changes Mediated by Pooled Immunoglobulin G and Hematin: Electrokinetic and Lipid Peroxidation Studies

Abstract

Pooled Immunoglobulin G (IgG), hematin and the membrane-disruptive amphipathic peptide melittin have received attention as powerful biomacromolecules for biomedical and pharmacology applications. Their action on surface properties, oxidation status and epifluorescence properties measured in vitro provide useful information about the functional activity of upper biomacromolecules in erythrocytes in vivo. The hemolysis of erythrocyte membranes, as well as changes in hematocrit and the morphology of erythrocytes, was investigated here via fluorescence microscopy using FITC-concanavalin A binding to cells. The effect of melittin on the membrane capacitance and resistance of model lipid bilayers was probed via electrochemical impedance spectroscopy. Lipid bilayer capacitance was higher in the presence of 0.10 g/L melittin compared to that in the control, which is likely related to bilayer thinning and alterations of the dielectric permittivity of melittin-treated membranes. The biomolecule interactions with red blood cells were probed in physiological media in which the surface of erythrocyte membranes was negatively charged. Surface parameters of erythrocytes are reported upon IgG/hematin and IgG/melittin treatment. Pooled IgG in the presence of melittin, preincubated IgG/hematin preparations promoted a significant decrease in the electrokinetic potential of erythrocytes (Rh-positive). A malondialdehyde (MDA) assay revealed a high rate of lipid peroxidation in erythrocytes treated with IgG/hematin or IgG/melittin preparations. This finding might be a result of pooled IgG interactions with the hematin molecule and the subsequent conformational changes in the protein molecule altering the electrokinetic properties of the erythrocyte membrane related to the Rh group type of erythrocytes. The pooled IgG and hematin are reported to have important consequences for the biophysical understanding of the immunopathological mechanisms of inflammatory, autoimmune and antibody-mediated pathological processes.