Activation of the alternative pathway of complement by calcium-loaded erythrocytes resulting from loss of membrane phospholipid asymmetry.
The aminophospholipids phosphatidylserine (PS) and phosphatidylethanolamine (PE) are exposed on the outer membrane leaflet of deoxygenated and irreversibly sickled erythrocytes and senescent normal cells. PS exposure on erythrocytes results in the expression of procoagulant activity for the conversion of prothrombin to thrombin. Because liposomes or vesicles composed of aminophospholipids can activate the alternative pathway of complement, the possibility that increased exposure of PS and PE on intact erythrocytes would also make them capable of activating the alternative pathway was examined. Loss of normal membrane phospholipid asymmetry was induced by incubation of erythrocytes with calcium (Ca2+) and the calcium ionophore A23187. PS exposure on 60% of erythrocytes was confirmed by binding of fluorescein isothiocyanate-conjugated annexin V. Expression of procoagulant activity, measured with the Russell's viper venom clotting assay, was significantly increased on the Ca2+/A23187-treated erythrocytes. In addition, the erythrocytes became capable of activating the alternative pathway of complement, as judged by an increase in cell-bound C3b after incubation with serum and a decrease in alternative pathway hemolytic activity of the serum. The effect could be reversed by incubation of the Ca2+/A23187-treated erythrocytes under conditions that induced recovery of normal membrane phospholipid asymmetry. In contrast, tetrathionate-treated erythrocytes showed no increase in binding of annexin V and no procoagulant activity and failed to activate the alternative pathway of complement. These findings demonstrate that loss of phospholipid asymmetry in erythrocytes not only results in expression of procoagulant activity but also renders the cells capable of activating the alternative pathway of complement.
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Children's Hospital Oakland Research Institute, California 94609, USA.
Mitsuyoshi
J
J
D017304:Annexin A5 / Q000378:metabolism; D001777:Blood Coagulation / Q000502:physiology; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000097:blood*; D003179:Complement C3b / Q000378:metabolism; D003170:Complement Pathway, Alternative; D004910:Erythrocyte Membrane / Q000737:chemistry / Q000502:physiology*; D004912:Erythrocytes / Q000737:chemistry / Q000502:physiology*; D005434:Flow Cytometry; D006801:Humans; D007118:Immunoassay; D007476:Ionophores / Q000494:pharmacology; D008081:Liposomes / Q000378:metabolism; D008563:Membrane Lipids / Q000502:physiology*; D010714:Phosphatidylethanolamines / Q000502:physiology; D010718:Phosphatidylserines / Q000502:physiology; D010743:Phospholipids / Q000502:physiology; D011485:Protein Binding; D011951:Receptors, Complement / Q000378:metabolism; D013776:Tetrathionic Acid / Q000494:pharmacology
D017304:Annexin A5; D007476:Ionophores; D008081:Liposomes; D008563:Membrane Lipids; D010714:Phosphatidylethanolamines; D010718:Phosphatidylserines; D010743:Phospholipids; D011951:Receptors, Complement; D000001:Calcimycin; D003179:Complement C3b; D013776:Tetrathionic Acid; D002118:Calcium