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Effects of complement activation products on the synthesis of decay accelerating factor and membrane cofactor protein
F G Cosio1, T Shibata, B H Rovin
1Department of Internal Medicine, Nephrology, Ohio State University, Columbus.
Insights
Complement activation stimulates decay accelerating factor (DAF) synthesis via C3 and C5a, with sustained increases following full complement cascade activation. Membrane cofactor protein (MCP) levels showed minimal changes.
Area of Science:
- Immunology
- Nephrology
Background:
- Decay accelerating factor (DAF) and membrane cofactor protein (MCP) are complement regulatory proteins.
- DAF and MCP are expressed on human glomerular cells, including human mesangial cells (HMC).
- Previous work showed terminal complement activation upregulates DAF on glomerular cells.
Purpose of the Study:
- Investigate mechanisms of complement-induced DAF synthesis in HMC.
- Evaluate complement activation's effect on MCP synthesis and expression in HMC.
Main Methods:
- Cultured human mesangial cells (HMC).
- Stimulation with complement activators (immune complexes, activated C3, C5a).
- Measurement of DAF and MCP mRNA and protein levels (including FACS analysis).
- Use of cycloheximide (protein synthesis inhibitor).
Main Results:
- Complement activation increased DAF mRNA via C3 deposition and C5a generation.
- DAF mRNA upregulation by C3/C5a was transient (<4 hr); full cascade activation sustained it (>8 hr).
- Cycloheximide alone upregulated DAF mRNA; it did not affect complement's effect on DAF mRNA.
- Complement activation did not upregulate MCP mRNA or protein synthesis, but slightly increased MCP protein levels by FACS.
Conclusions:
- Complement activation upregulates DAF mRNA through multiple pathways in HMC.
- Sustained DAF protein increase requires full complement cascade activation.
- MCP expression is less responsive to complement activation compared to DAF.
Abstract:
We previously demonstrated that activation of terminal complement components (C8 and/or C9) increases the synthesis and expression of decay accelerating factor (DAF) on human glomerular cells. DAF is a cell membrane-associated complement regulatory protein that inhibits complement activation on cell surfaces. In the present studies we evaluated, first, the mechanisms by which complement activation stimulates DAF synthesis, and second the effect of complement activation on the synthesis. and expression of membrane cofactor protein (MCP), another complement regulatory protein, by human mesangial cells (HMC) in culture. Complement activation by immune complexes resulted in increased DAF mRNA levels by at least two mechanisms: deposition of activated C3 on HMC and generation of soluble complement activation products, specifically C5a. The increase in DAF mRNA levels induced by activated C3 or C5a was short lived (less than 4 hr). In contrast, the up-regulation of DAF mRNA levels induced by activation of the complete complement cascade persisted for at least eight hours. The effect of complement activation on DAF mRNA levels was not affected by cycloheximide, a protein synthesis inhibitor. However, cycloheximide alone resulted in a significant up-regulation of DAF mRNA levels on HMC. In contrast to those findings complement activation did not cause an up-regulation of MCP mRNA, nor an increase in the synthesis of this protein. However, by FACS, complement produced a small but significant increase of MCP protein levels on HMC. In conclusion, both MCP and DAF are present on HMC. Several activated complement components are capable of increasing DAF mRNA levels, but DAF protein levels increase only after activation of the whole complement cascade.(ABSTRACT TRUNCATED AT 250 WORDS)