Complement component C1q induces endothelial cell adhesion and spreading through a docking/signaling partnership of C1q receptors and integrins.
The interaction of C1q with endothelial cells elicits a multiplicity of biologic responses. Although these specific responses are thought to be mediated by the interaction of C1q with proteins of the endothelial cell surface, the molecular identity of the participant(s) has not been clearly defined. In this study, we examined the role of two C1q-binding proteins, cC1q-R/CR and gC1q-R/p33, on C1q-mediated adhesion and spreading of human dermal microvascular endothelial cells (HDMVECs). A specific and dose-dependent adhesion and spreading was observed when HDMVECs were cultured in microtiter plate wells coated with concentrations of C1q ranging from 0 to 50 microg/ml. The extent of adhesion and spreading was similar to the adhesion seen on collagen-coated wells. Furthermore, the effect of C1q was mimicked by either polyclonal anti-cC1q-R or mAb 60.11, but not with isotype- and species-matched control IgG. More importantly, however, a 100% inhibition of spreading but not adhesion to C1q-coated wells was observed when HDMVECs were cultured in the presence of 30 mM of the peptide GRRGDSP but not GRRGESP. Furthermore, while anti-beta1 integrin antibody blocked adhesion and spreading, antialpha5 integrin only blocked spreading. Since earlier studies have shown that zinc induces the exposure of hydrophobic sites in the C-terminus of gC1q-R including the putative high-molecular weight kininogen (HK)-binding site corresponding to residues 204-218, we also examined the effect of zinc on antibody binding to cell surface gC1q-R. Flow cytometric data show that the binding of mAb 74.5.2, which recognizes residues 204-218, is greatly enhanced when endothelial cells were incubated in the presence of 50 microM zinc. In summary, our data show that: (a) C1q-mediated endothelial cell adhesion and spreading requires the cooperation of both C1q receptors and 1 integrins, and possibly other membrane-spanning molecules, and (b) zinc can induce the exposure of hydrophobic sites in the C-terminal domain of gC1q-R allowing a more efficient binding of mAb 74.5.2 and HK.
Ghebrehiwet
Berhane
B
Department of Medicine, Health Sciences Center, Division of Rheumatology, State University of New York, Stony Brook, NY 11794, USA. berhane@mail.som.sunysb.edu
Feng
Xiaodong
X
Kumar
Rajeev
R
Peerschke
Ellinor I B
EI
D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001710:Biotin / Q000378:metabolism; D002196:Capillaries / Q000166:cytology / Q000187:drug effects; D002448:Cell Adhesion / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D015922:Complement C1q / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D019012:Integrin beta1 / Q000187:drug effects / Q000378:metabolism; D016023:Integrins / Q000378:metabolism*; D007704:Kininogens / Q000378:metabolism; D008856:Microscopy, Fluorescence; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000378:metabolism; D011951:Receptors, Complement / Q000187:drug effects* / Q000378:metabolism*; D015032:Zinc / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D019012:Integrin beta1; D016023:Integrins; D007704:Kininogens; D011956:Receptors, Cell Surface; D011951:Receptors, Complement; D001710:Biotin; D015922:Complement C1q; D015032:Zinc