Inhibition by protease inhibitors of chemotaxis induced by elastin-derived peptides.
We studied the effect of two inhibitors of human neutrophil proteases on neutrophil chemotaxis induced by the hexapeptide Val-Gly-Val-Ala-Pro-Gly (VGVAPG), a recurring sequence in the elastin molecule. The inhibitors were tosyl-Phe chloromethyl ketone (TFCK) and N-methoxysuccinyl-Ala-Ala-Pro-Val chloromethyl ketone (MAAPVCK). We assayed chemotactic activity by the double-membrane technique in a modified Boyden chamber, after incubating the cells for 1 hr with varying concentrations of inhibitor. We observed a concentration-dependent inhibitory effect. We also measured the potency of the two chloromethyl ketones as protease inhibitors. The more potent protease inhibitor, MAAPVCK, was also the more effective in inhibiting VGVAPG-induced chemotaxis; its inhibitor dissociation constant was KI = 28 nM with elastase and KI = 33 nM with cathepsin G. For TFCK the corresponding KI values were 37 microM and 200 microM. The incubating concentration required to lower chemotaxis by half its uninhibited value was C0.5 = 0.64 microM for MAAPVCK, compared to C0.5 = 3.4 microM for TFCK. A third peptide, triglycinate (gly3), which did not inhibit the proteolytic activity of either elastase or cathepsin G, did not inhibit chemotaxis. Chemotaxis induced by formyl Met-Leu-Phe (fMLP) was weakly inhibited by both chloromethyl ketones with TFCK being somewhat more effective than MAAPVCK. We concluded that inhibition of VGVAPG-induced chemotaxis is in part specific, receptor mediated. We suggest that proteolytic inhibitors protect the extracellular matrix from degradation by inhibiting chemotaxis. Comparing the inhibitor concentrations required to half proteolytic activity with the concentration required to half chemotactic activity, we further suggest that the two functions may be of comparable significance.
Sarfati
I
I
Department of Surgery, Long Island Jewish Medical Center, New Hyde Park, New York 11042, USA.
Lopes
D
D
Murphy
E A
EA
Rao
S
S
Cohen
J R
JR
D000595:Amino Acid Sequence; D002633:Chemotaxis / Q000187:drug effects*; D004549:Elastin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D008969:Molecular Sequence Data; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000235:genetics / Q000494:pharmacology*; D011480:Protease Inhibitors / Q000494:pharmacology*
D010446:Peptide Fragments; D011480:Protease Inhibitors; D009240:N-Formylmethionine Leucyl-Phenylalanine; D004549:Elastin