Abstract
The pulmonary leukostasis and lung dysfunction associated with intravascular complement activation results from C5a-mediated granulocyte (GR) aggregation, a phenomenon which can be reproduced in vitro using standard nephelometric techniques. To produce a more subjective measure of the extent and rate of GR aggregation responses we added a digital integrator to the system. The validity of this approach was substantiated by the close correlation between the aggregating and chemotactic activities of C5a andN-formyl-metionine-leucine-phenylalanine. Use of this technique enabled us to define the dose-response relationship of the aggregation produced by the cationophore A23187 and the inhibitory effect of tetracaine on divalent cationdependent aggregation responses. The aggregation produced by these three stimuli does not result primarily from simple cross-linking of surface charges because, unlike the passive cell-cell association produced by the cation poly-l-lysine, it is not inhibited by anionic poly-l-glutamic acid. The importance of microtubules as regulators of GR adhesiveness was substantiated by the inhibitory effects of colchicine (but not lumicolchicine) on aggregation in this system. These data suggest that this integration of light transmission increments is a useful adjunct to this basic technique, whether used as a bioassay for chemotactic stimuli or as a model to study the many factors which regulate GR adhesiveness.
Similar content being viewed by others
References
Craddock, P. R., J. Fehr, A. P. Dalmasso, K. L. Brigham, andH. S. Jacob. 1977. Hemodialysis leukopenia. Pulmonary vascular leukostasis resulting from complement activation by dialyzer cellophane membranes.J. Clin. Invest. 59:879.
Craddock, P. R., J. Fehr, K. L. Brigham, R. S. Dronenberg, andH. S. Jacob. 1977. Complement and leukocyte-mediated pulmonary dysfunction in hemodialysis.N. Engl. J. Med. 296:769.
Craddock, P. R., D. Hammerschmidt, J. G. White, A. P. Dalmasso, andH. S. Jacob. 1977. Complement (C5a)-induced granulocyte aggregation in vitro. A possible mechanism of complement-mediated leukostasis and leukopenia.J. Clin. Invest. 60:260.
Hammerschmidt, D. E., J. G. White, P. R. Craddock, andH. S. Jacob. 1979. Cortocosteroids inhibit complement-induced granulocyte aggregation. A possible mechanism for their efficacy in shock states.J. Clin. Invest. 63:798.
Sacks, T., C. F. Moldow, P. R. Craddock, T. K. Bowers, andH. S. Jacob. 1978. Oxygen radicals mediate endothelial damage by complement-stimulated granulocytes. An in vitro model of immune vascular damage.J. Clin. Invest. 61:1161.
MacGregor, R., P. Spagnuolo, andA. Lentner. 1974. Inhibition of granulocyte adherence by ethanol, prednisone, and aspirin measured with an assay system.N. Engl. J. Med. 291:642.
O'flaherty, J. T., D. L. Kreutzer, andP. A. Ward. 1977. Neutrophil aggregation and swelling induced by chemotactic agents.J. Immunol. 119:232–239.
Boucek, M. M., andR. Synderman. 1976. Calcium influx requirement for human neutrophil chemotaxis: Inhibition by lanthanum chloride.Science 193:905.
Craddock, P. R., Y. Yawata, L. Van Santen, S. Gilberstadt, S. Silvis, andH. S. Jacob. 1974. Acquired phagocyte dysfunction. A complication of the hypophospotemic of parenthal hyperalimentation.N. Engl. J. Med. 290:1403.
Zigmond, S. H., andJ. G. Hirsch. 1973. Leukocyte locomotion and chemotaxis: New methods for evaluation and demonstration of cell-derived chemotactic factor.J. Exp. Med. 137:187.
Craddock, P. R., J. G. White, andH. S. Jacob. 1978. The potentiation of complement (C5a)-induced granulocyte aggregation by cytochalasin B.J. Lab. Clin. Med. 91:490.
Smith, A. L., I. Rosenberg, D. R. Avfrill, F. R. Moxon, T. Stossel, andD. H. Smith. 1974. Brain polymorphonuclear leukocyte quantitation by peroxidase assay.Infect. Immun. 10:356.
Hoffstein, S., I. M. Goldstein, andG. Weissmann. 1977. Role of microtubule assembly in lysosomal enzyme secretion from human polymorphonuclear leukocytes. A reevaluation.J. Cell Biol. 73:242.
Beutler, E. 1975. Red Cell Metabolism.In A Manual of Biochemical Methods, second ed. Grune and Stratton, New York. 63.
Wilson, L., andM. Griedkin. 1966. The biochemical events of mitoses. 1. Synthesis and properties of colchicine labeled with tritium in its acetyl moiety.Biochemistry. 5:2463.
Boxer, L. A., J. M. Allen, A. M. Watanabe, H. R. Besch, andR. L. Baehner. 1978. Role of microtubules on granulocyte adherence.Blood 51:1045.
White, J. G. 1968. Fine structural alterations induced in platelets by adenosine diphosphate.Blood 31:604.
Gosset, W. S. 1908. The probable error of the mean.Biometrika. 6:1.
Craddock, P. R., D. E. Hammerschmidt, C. R. Moldow, O. Yamada, andH. S. Jacob. 1979. Granulocyte aggregation as a manifestation of membrane interactions with complement: Possible role in leukocyte margination, microvascular occlusion, and endothelial damage.Sem. Hematol. 16:140.
Craddock, P. R., R. P. Hebbel, J. G. White, andH. S. Jacob. 1977. Complementinduced Ca2++-mediated granulocyte aggregation; a previously unsuspected response of phagocytes.Clin. Res. 25:474A.
O'Flaherty, J. T., H. J. Showell, E. L. Becker, andP. A. Ward. 1978. Substances which aggregate neutrophils. Mechanism of action.Am. J. Pathol. 92:155.
Seeman, P. 1972. The membrane actions of anesthetics and tranquilizers.Pharmacol. Rev. 24:583.
Suarez-Kurtz, G., andF. J. R. Baumgartten. 1973. Blockade of quinine-induced contracture and45Ca efflux by procaine and tetracaine in frog sartorius muscle.J. Pharmacol. Exp. Ther. 186:562.
Papahadjopoulos, D. 1972. Studies on the mechanism of action of local anesthetics with phospholipid model membranes.Biochem. Biophys. Acta 265:169.
Gallin, J. I. andA. S. Rosenthal. 1974. The regulatory role of divalent cations in human granulocyte chemotaxis; Evidence for an association between calcium exchanges and microtubule assembly.J. Cell Biol. 62:594.
Hoffstein, S., I. M. Goldstein, andG. Weissmann. 1977. Role of microtubule assembly in lysosomal enzyme secretion from human polymorphonuclear leukocytes. A re-evaluationJ. Cell Biol. 73:242.
Oliver, J. M., T. E. Ukena, andR. D. Berlin. 1974, Effects of phagocytes and colchicine on the distribution of lectin-binding sites on the cell surface.Proc. Natl. Acad. Sci. U.S.A. 71:395.
Weisdorf, D. J., D. E. Hammerschmidt, H. S. Jacob, andP. R. Craddock. 1979. The roles of endogenous protrases, circulating granulocytes, and the spleen in the clearance of activated C5 components: Possible relevance to the shock lung syndrome (ARDS).Clin. Res. 27:649A.
Fletcher, J. R., andP. W. Ramwell, 1978. Lidocaine or indomethacin improves survival in baboon endotoxic shock.J. Surg. Res. 24:154.
Author information
Authors and Affiliations
Additional information
This work was supported by grants CA15627, HL11880, and HLO6316 from the National Institutes of Health. Dr. Craddock is the recipient of Research Career Development Award HLO479 also from the National Institutes of Health.
Rights and permissions
About this article
Cite this article
Craddock, P.R., White, J.G., Weisdorf, D.J. et al. Digital integration of granulocyte aggregation responses. Inflammation 4, 381–395 (1980). https://doi.org/10.1007/BF00916049
Issue Date:
DOI: https://doi.org/10.1007/BF00916049