Inhibition of neutrophil chemotaxis by protease inhibitors. Differential effect of inhibitors of serine and thiol proteases.

Previous studies have demonstrated that neutrophils possess an active serine protease(s) which may be involved in the process of chemotaxis but the precise identity of this enzyme(s) remains to be determined. In this study fourteen different protease inhibitors were tested over a wide concentration range for their ability to inhibit unstimulated neutrophil movement and chemotaxis to C5a, fMLP and IL-8. Pretreatment of neutrophils with aspartyl or metallo-protease inhibitors had no effect on either chemotaxis or random cell movement. The thiol protease inhibitors E-64 and cystatin, as well as the thiol/serine inhibitors antipain and leupeptin, diminished only C5a-induced chemotaxis. Pretreatment of neutrophils with the serine protease inhibitors PMSF or 3,4-DCI significantly reduced chemotaxis to C5a, fMLP and IL-8. The inhibitor of trypsin-like serine proteases, TLCK, and the neutrophil elastase inhibitor MeO-Suc-AAPV-CMK had no inhibitory effect on cell movement. However, two different inhibitors of chymotrypsin-like serine proteases, TPCK and chymostatin, significantly inhibited movement to any chemoattractant. These results suggest that an active chymotrypsin-like serine protease is essential for neutrophils to respond to chemotactic stimuli.

A comparison of large-scale Sf9 insect cell growth and protein production: stirred vessel vs. airlift.

A simple and relatively inexpensive stirred-vessel system for the large-scale growth of insect cells (Sf9) is described. Sf9 cell growth in a stirred-vessel fermentor and an airlift fermentor were compared on the basis of maximum cell density and average population doubling time. Also, both fermentor systems were compared with respect to the large-scale production of a recombinant human protein (protein kinase C-eta). No significant differences in Sf9 cell growth or protein expression levels were apparent between the two fermentor systems. However, large differences in cost and scale-up of each system are discussed with respect to the large-scale production of recombinant proteins.