THE CONTROL OF NEUTROPHIL CHEMOTAXIS BY INHIBITORS OF CATHEPSIN-G ANDCHYMOTRYPSIN

Neutrophil chemotaxis plays an important role in the inflammatory resp onse and when excessive or persistent may augment tissue damage, The e ffects of inhibitors indicated the involvement of one or more serine p roteinases in human neutrophil migration and shape change in response to a chemoattractant. Monospecific antibodies, chloromethylketone inhi bitors, and reactive-site mutants of alpha(1)-antitrypsin and alpha(1) -antichymotrypsin were used to probe the specificity of the proteinase s involved in chemotaxis, Antibodies specific for cathepsin G inhibite d chemotaxis, Moreover, rapid inhibitors of cathepsin G and alpha-chym otrypsin suppressed neutrophil chemotaxis to the chemoattractants N-fo rmyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) and zymosan-activated serum in multiple blind well assays and to fMLP in migration assays u nder agarose, The concentrations of antichymotrypsin mutants that redu ced chemotaxis by 50% would inactivate free cathepsin G with a half-li fe of 1.5-3 s, whereas the concentrations of chloromethylketones requi red to produce a similar inhibition of chemotaxis would inactivate cat hepsin G with a half life of 345 s. These data suggest different modes of action for these two classes of inhibitors, Indeed the chloromethy lketone inhibitors of cathepsin G (Z-Gly-Leu-Phe-CMK) and to a lesser extent of chymotrypsin (Cbz-Gly-Gly-Phe-CMK) mediated their effect by preventing a shape change in the purified neutrophils exposed to fMLP, Antichymotrypsin did not affect shape change in response to fMLP even at concentrations that were able to reduce neutrophil chemotaxis by 5 0%, These results support the involvement of cell surface proteinases in the control of cell migration and show that antichymotrypsin and ch loromethylketones have differing modes of action, This opens the possi bility for the rational design of anti-inflammatory agents targeted at neutrophil membrane enzymes.