Molecular basis of complement activation in ischemic myocardium: identification of specific molecules of mitochondrial origin that bind human C1q and fix complement.
Mitochondria may be a source of molecules that activate complement during ischemic injury to myocardium, providing therewith a stimulus for infiltration of polymorphonuclear leukocytes. To identify specific molecules that activate the classical complement pathway, detergent lysates of canine cardiac mitochondria were fractionated by polyacrylamide gel electrophoresis and transferred electrophoretically to nitrocellulose paper (NCP). The NCP replicas of the gels were incubated with isolated C1q and fresh sera as a source of complement, washed briefly, and overlaid with sensitized sheep erythrocytes (RBC) in agarose. A cluster of four to six molecules between 45 and 53 kDa as well as four others, 34, 30, 26, and 23 kDa, consumed complement thereby preventing complement-mediated lysis of sensitized sheep RBC in the agarose overlay. Additional molecules reactive with C1 were identified by their ability to bind isolated human C1q and to serve as assembly sites for later acting complement components. Sites of localization of complement were demonstrated by incubating NCP replicas of fractionated mitochondria with antisera specific for C1q, C3, C5, and C9, followed by peroxidase-conjugated anti-immunoglobulin and substrate. A total of 12 C1q binding molecules ranging in size from 67 kDa to 23 kDa, which can fix later acting complement components, were identified. At least two of these reacted with antisera prepared against canine cardiac lymph collected in the first 3-4 hours after a 45-minute coronary artery occlusion. These studies present direct evidence that specific molecules, released from subcellular fractions of myocardial cells rich in mitochondria, can activate the complement cascade.
Kagiyama
A
A
Laboratory for Immunology Research, Veterans Administration Medical Center, Houston, TX 77211.
Savage
H E
HE
Michael
L H
LH
Hanson
G
G
Entman
M L
ML
Rossen
R D
RD
D000818:Animals; D001665:Binding Sites; D003166:Complement Activating Enzymes / Q000032:analysis / Q000276:immunology*; D003167:Complement Activation; D003172:Complement C1 / Q000032:analysis / Q000276:immunology*; D015922:Complement C1q; D003168:Complement Fixation Tests / Q000379:methods; D003171:Complement Pathway, Classical; D003327:Coronary Disease / Q000276:immunology*; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006168:Guinea Pigs; D006801:Humans; D008196:Lymph / Q000276:immunology; D008929:Mitochondria, Heart / Q000032:analysis / Q000276:immunology*; D008970:Molecular Weight; D009124:Muscle Proteins / Q000032:analysis / Q000276:immunology
D003172:Complement C1; D009124:Muscle Proteins; D015922:Complement C1q; D003166:Complement Activating Enzymes