Anti-Fx1A produces complement-dependent cytotoxicity of glomerular epithelial cells.
Glomerular injury in passive Heymann nephritis (PHN) in rats is mediated by the C5b-9 membrane attack complex (MAC) and is associated with morphologic changes in glomerular visceral epithelial cells (GEC). We determined if the nephritogenic antibody of PHN (gamma 1 sheep anti-Fx1A IgG) directs insertion of the MAC into GEC plasma membranes with consequent cytotoxicity. Antibody-sensitized GEC were exposed to various sera serving as sources of complement. Loss of cell viability was determined by trypan blue uptake and/or by release of cellular lactate dehydrogenase (LDH). Incubation of antibody-sensitized primary and passaged GEC in fresh human serum (FHS) resulted in sigmoidal relationships between cytotoxicity and complement dose (r = 0.97 and 0.94, respectively) such that cytolysis approached 100% with FHS (10% vol/vol). Cytotoxicity was not evident if C8-deficient (C8D) plasma was substituted for FHS, but was restored in a dose-dependent manner by reconstitution with purified rat C8. Sublytic injury was demonstrated by wide separation between simultaneous release curves of cell-incorporated biscarboxyethyl carboxyfluorescein (BCECF; mol wt approximately equal to 520) and LDH at limiting doses of complement (at 2% FHS, BCECF release was 51.1 +/- 0.6% of maximum vs. 3.2 +/- 1.3% for LDH; N = 3) and by blebbing of the plasma membrane on electron microscopy. Thus, the pathogenic antibody of PHN produces complement-mediated sublytic as well as lytic cytotoxicity of GEC.
Quigg
R J
RJ
Department of Medicine, University Hospital, Boston, Massachusetts.
Cybulsky
A V
AV
Jacobs
J B
JB
Salant
D J
DJ
D000818:Animals; D000920:Antibody-Dependent Cell Cytotoxicity; D002478:Cells, Cultured; D003165:Complement System Proteins / Q000276:immunology*; D004847:Epithelial Cells; D004848:Epithelium / Q000276:immunology / Q000648:ultrastructure; D007678:Kidney Glomerulus / Q000276:immunology* / Q000648:ultrastructure; D007687:Kidney Tubules, Proximal / Q000276:immunology*; D008297:Male; D008854:Microscopy, Electron; D008855:Microscopy, Electron, Scanning; D008871:Microvilli / Q000276:immunology; D051381:Rats; D011919:Rats, Inbred Strains
D003165:Complement System Proteins