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Complement-opsonized IgG antibody/dsDNA immune complexes bind to CR1 clusters on isolated human erythrocytes
R P Taylor1, F Pocanic, C Reist
1Department of Biochemistry, University of Virginia School of Medicine, Charlottesville 22908.
Insights
Immune adherence (IA) involves complement (C3b)-opsonized immune complexes (IC) binding to erythrocyte complement receptor 1 (CR1). CR1 clusters on red blood cells (RBCs) dictate IC binding, with aggregated complexes potentially aiding transfer to phagocytic cells.
Area of Science:
- Immunology
- Cell Biology
- Hematology
Background:
- Immune adherence (IA) is a mechanism where complement-opsonized immune complexes (IC) bind to erythrocytes (RBCs) via the complement receptor 1 (CR1).
- The distribution and binding characteristics of these complexes on RBCs are crucial for understanding immune complex clearance.
Purpose of the Study:
- To investigate the distribution of complement (C3b)-opsonized antibody/dsDNA immune complexes (IC) bound to human erythrocytes (RBCs) via immune adherence (IA).
- To correlate IC binding with the number and distribution of CR1 receptors on RBCs.
Main Methods:
- Utilized fluorescence microscopy to visualize IC and CR1 distribution.
- Employed quantitative flow cytometry (FACS) analyses to assess CR1 numbers and IC binding.
- Conducted radioimmunoassays to quantify IC binding.
Main Results:
- RBCs showed significant heterogeneity in IC binding, with some binding no complexes.
- Bound IC localized to CR1 clusters on RBCs, correlating strongly with CR1 number per cell.
- Fewer fluorescent spots (aggregated IC) were observed per RBC than the total number of bound IC, suggesting aggregation.
Conclusions:
- The number of CR1 clusters capable of binding IC is proportional to the total CR1 count per RBC.
- Aggregated immune complexes on RBCs may facilitate their transfer to mononuclear phagocytic cells.
- Efficient IA requires specific geometric alignment between C3b on IC and multiple CR1 molecules within a cluster.
Abstract:
We used fluorescence microscopy, quantitative FACS analyses, and radioimmunoassays to examine the distribution of complement (C3b)-opsonized antibody/dsDNA immune complexes (IC) bound to normal human erythrocytes (RBCs) via immune adherence (IA). IC were detected with fluorescent anti-human IgG, and the RBC IA-receptor (CR1) was revealed with monoclonal antibodies (Mabs) to CR1 and fluorescent anti-mouse IgG. Under saturating conditions RBCs exhibit a large heterogeneity in binding; a significant fraction binds no IC. The positions of bound IC coincide with CR1 clusters on RBCs, as predicted by several investigators. FACS experiments indicate an excellent correlation between CR1 number and IC binding within an RBC population. The number of CR1 clusters able to bind IC is proportional to the number of CR1 per RBC. However, IC (fluorescent spots) detected per RBC (on average less than 10) are less than the average number of IC bound (20-30). This suggests that each fluorescent spot represents a small number of aggregated IC bound to a CR1 cluster. These "patches" of aggregated complexes may facilitate transfer of RBC-bound IC to cells of the mononuclear phagocytic system. Finally, not all the CR1 clusters bind IC, suggesting that proper geometric alignment of multiple C3b per IC with several CR1 in the cluster is required for IA.