Membrane Lesions in Immune Lysis

Abstract

A B S T R A C TIt is known that there are 100 A-wide circular structures associated with the erythrocyte membrane in immune lysis. To determine whether these structures were functional holes extending through the membrane, freeze-etch electron microscopy was carried out. Sheep erythrocytcs incubatcd with either rabbit complement or rabbit antibody (anti-sheep erythrocyte antibody) did not hemolyzc and did not reveal any abnormalitics in freezectch or negative-stain clcctron microscopy. Erythrocytes incubated with both complement and antibody revealed rings on the extracellular surface (etch face) of the cell membrane. o Allowing for the 30 A-thick Pt/C replica, the dimensions of the surface rings were similar to those seen by negative staining. The ring's central depression was level with the plane of the membrane; some rings were closed circles, othcrs were crescent shaped. The cleavage face of the cxtraccllular leaflet revealed globule aggregates, each aggregate appearing to be composed of about four fused globules. The cleavage face of the cytoplasmic leaflet was normal. When immune lysis was carried out in the presence of ferritin, fcrritin was subsequently detected in all lysed erythrocytes. If ferritin was added after immune lysis was complete, only 15 % of the cells were pcrmcatcd by fcrritin, indicating that transien[ openings exist in the cell membrane during immune lysis. No abnormal structures were detected when C6-deficicnt rabbit serum was used as a source of complement. It is concluded that antibody and complement produce surface rings, prelytic leakage of K +, colloid osmotic swelling, membrane disruption, and membrane resealing; the surface rings persist after these events.

Membrane attack complex of complement: generation of high-affinity phospholipid binding sites by fusion of five hydrophilic plasma proteins.

“…It is emphasized that model c proposes a reorientation of boundary lipid, which may be envisioned as depicted schematically in the figure or in a different manner provided it results in a perturbation of the normal bilayer structure of the membrane. Model c is consistent with the body of available data: (i) the initial complement produced channel is small (25,30), (ii) the cytoplasmic leaflet of membranes attacked by the MAC exhibits no morphological alterations (31), and (iii) the MAC (28) and lipid (32) tend to detach from liposomes (33)(34)(35) and bacterial cell walls (36), probably in the form of protein-phospholipid complexes.…”

Role of Complement Components on Cells and in Cell Interactions

“…This insertional theory has been called the 'doughnut theory' by M a y e r [1973]. Freeze-etching electron micrographs support the theory [Iles et al, 1973].…”

Assembly of complement components C5b-8 and C5b-9 on lipid bilayer membranes: visualization by freeze-etch electron microscopy

“…Early support for the doughnut hypothesis came from negative-stain electron micrographs (EM) of complement-lysed erythrocytes, which show accumulations of stain in dark circular areas of 9-13-nm diameter. Freeze-etch EM of the same system also reveals doughnut-like projections on the external membrane surface and proteinaceous stubs projecting from the hydrophobic fracture face of the outer monolayer (lies et al, 1973;Bhakdi et al, 1974;Tranum-Jensen & Bhakdi, 1983).More recent support for the doughnut hypothesis comes from the finding that under certain conditions, e.g., high temperature or the presence of heavy metals or exogenous proteases, purified complement component C9 forms a tubular structure composed of 12-18 C9 protomers (Podack & Tschopp, 1982;Tschopp et al, 1984). The tube has an annulus at one end such that the external dimensions are =21 x 15 X 15 nm (DiScipio & Hugh, 1985).…”