Related Experiment Videos
Decay-accelerating factor regulates complement-mediated damage in the human atherosclerotic wall
F Niculescu1, H G Rus, R Vlaicu
1Medical Clinic No. 1, Cluj-Napoca, Roumania.
Insights
Decay-accelerating factor (DAF) protects arterial walls from complement damage. However, DAF is insufficient to prevent complement activation in human atherosclerotic arteries.
Area of Science:
- Immunology
- Vascular Biology
- Biochemistry
Background:
- Decay-accelerating factor (DAF) is a membrane protein that inhibits complement pathways.
- The role of DAF in the human atherosclerotic arterial wall is not fully understood.
Purpose of the Study:
- To investigate the localization and function of DAF in human atherosclerotic arteries.
- To determine if DAF protects against complement-mediated damage in the arterial wall.
Main Methods:
- Immunohistochemistry and immunogold labeling to localize DAF in arterial tissue.
- Western blotting to analyze DAF molecular weight.
- Enzyme digestion and cell-based assays to assess DAF's protective role against complement-mediated lysis.
Main Results:
- DAF was found in the cells and connective tissue matrix of the arterial wall, with higher concentrations in fibrous plaques and intimal thickenings.
- DAF co-localized with C5b-9 complement complexes on arterial cells and debris.
- Blocking DAF significantly increased complement-mediated cell lysis from 10-15% to 60-70%.
Conclusions:
- DAF plays a protective role against autologous complement activation in the arterial wall.
- DAF's protective effect is insufficient to completely prevent complement activation in human atherosclerosis.
Abstract:
Decay-accelerating factor (DAF) is an intrinsic membrane inhibitor that regulates the activity of C3 and C5 convertases of the classical and alternative complement pathways. Using two monoclonal antibodies, IC6 and IA10, DAF was localized by immunohistochemistry using streptavidin-biotin-peroxidase complex or silver-intensified immunogold techniques in aortic, iliac and femoral samples obtained at surgery and autopsy from 32 patients. DAF was localized on the cells and in the connective tissue matrix of the arterial wall. Fibrous plaques and intimal thickenings presented larger amounts than fatty streaks, intimae and normal areas. By Western blotting analysis, DAF extracted from the arterial wall had a molecular weight of about 67 kDa. Using a double-labeling technique, DAF and C5b-9 complexes were co-localized on nucleated cells and on cell debris. The cells isolated after enzyme digestion of the arterial wall were tested for the protective role of DAF to complement-mediated damage. When DAF of the sensitized cells was blocked by monoclonal antibodies, complement-mediated cell lysis was enhanced from 10-15% to 60-70%. The effect of anti-DAF antibodies was dose-dependent. DAF blocking in the absence of antibodies used for sensitization led to a lysis under 10%. These data suggest a protective role of DAF against autologous complement activation, however insufficient to prevent complement activation in the human atherosclerotic wall.