Neutrophil invasion of collagen gels: Is collagenolytic activity involved?

246

Cell Biology

International

Reports,

Vol. 11, No. 3, March

1987

Neutroohil invasion of CollaaenGels,IsCollaaenolvticActivitv Involved? K.V.Crocket,J.M. Lack&, Departmentof Cell Biology, University of Glasgow,612 803

During an inflammatory response neutrophil leucocytes (PIIN) invade, and move through, a collagen-rich ECM,Normal humanPMNcontain a specific collagenase,25-3Og of which can be released in response to damagedtissue and immune complexes I1 I. There is some evidence that proteases are important for local invasion from primary tumours and at secondary sites /2,31, and it is pertinent to ask whether invading PMN engagein proteolytic activity. Gels made from reconstituted collagen provide a model for normal connective tissue, and using PMNfrom rabbit peritoneal exudate and collagen radio-labelled with tritium it is possible to look for the appearanceof soluble collagen fragments producedby cell-released collagenase. Thin layers of collagen were used to maximise the area accessible to cells, which were stimulated with optimum concentrations of f-MLP. Aliquots for counting were taken from the medium abovethe gels after a 2 h incubation at 37’C, with or without cells. Results from 4 assays using cells showed no increase in counts. Wells containing collagenase at concentrations from 0.1 to 10.0 pg/ ml, in contrast (3 assays), showed significantly increased counts. At the cell numbers used, and assumingcomparableintracellular levels of collagenase,we might expect about 20 ug of collagenaseto be released into the medium.As we found no increase in counts with cells, yet an increase in counts using much lower levels of collagenase,we conclude that in this invasive assay system, PMNare not engagedin any significant proteolytic activity. L11.Dhlsson& Olsson(1977). Stand J Haematol19, 145-152. 121.Matzner ds/ (1985). J Clin Invest 76, 1306-1313 131.Levine d J/ (1984). CollagenRel Res4.49-61,