Tryptase and chymase: comparison of extraction and release in two dog mastocytoma lines.
Mast cell secretory granules contain unique tryptic and chymotryptic serine proteases that differ between species and tissues. Direct comparison of these proteases in single-cell types has been hindered by the difficulty of obtaining adequate numbers of pure mast cells. In this study, we were able to compare tryptic and chymotryptic enzyme activity in cells of presumed monoclonal origin, using two stable lines ('BR' and 'G') of dog mastocytomas. The gel-filtration profiles, inhibitor susceptibilities and substrate preferences of tryptic and chymotryptic mastocytoma protease activities established their close resemblance to the tryptases and chymases of human and rodent mast cells. Striking heterogeneity was observed in the amounts and solubilities of the tryptic and chymotryptic activity in the two different mastocytoma cell lines. Incubation of cells from both lines with calcium ionophore A23187 caused non-cytotoxic release of protease activity. In contrast to chymase from rat connective tissue mast cells, protease activity that was insoluble after extraction at low ionic strength became soluble following ionophore-stimulated release. Neither tryptic nor chymotryptic activity was activated during degranulation, suggesting the absence of inactive precursors. Cells of the 'BR' line released both tryptic and chymotryptic activity in parallel with the granule marker histamine; cells of the 'G' line released a much smaller proportion of tryptic activity than of either chymotryptic activity or histamine. These differences in release of granule constituents from cells of common origin could be explained by developmental variations in the production of performed mediators or by differential regulation of preformed mediator release. We conclude that the differences in protease content, solubility and release in these mastocytoma lines are useful in evaluating the potential pathophysiological significance of the contribution of proteases to mast cell heterogeneity.
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D053818:Chymases; D002918:Chymotrypsin / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000201:enzymology; D004285:Dogs; D008407:Mast Cells / Q000201:enzymology; D012515:Mast-Cell Sarcoma / Q000201:enzymology*; D010447:Peptide Hydrolases / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000378:metabolism*; D013379:Substrate Specificity; D014357:Trypsin / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000201:enzymology
D000001:Calcimycin; D010447:Peptide Hydrolases; C023073:tosylarginine methyl ester hydrolase; D012697:Serine Endopeptidases; D002918:Chymotrypsin; D053818:Chymases; D014357:Trypsin