1 Stimulation of chemotaxis of human polymorphonuclear leucocytes (PMNs) with the chemoattractive peptide fMLP (N-formyl-Met-Leu-Phe) is paralleled by profound morphological and metabolic alterations like changes of intracellular pH (pH i ) and cell shape. The present study was performed to investigate the interrelation of cell volume (CV) regulatory ion transport, pH i and migration of fMLP stimulated PMNs. 2 Addition of fMLP to PMNs stimulated directed migration in Boyden chamber assays and was accompanied by rapid initial intracellular acidi®cation and cell swelling. 3 Inhibition of the Na + /H + exchanger suppressed fMLP stimulated cell migration, accelerated the intracellular acidi®cation and inhibited the fMLP-induced cell swelling. 4Step omission of extracellular Na + caused intracellular acidi®cation, which was accelerated by subsequent addition of gastric H + /K + ATPase inhibitor SCH 28080, or by omission of extracellular K + ions. In addition Na + removal caused cell swelling, which was further enhanced by fMLP. 5 H + /K + ATPase inhibitors omeprazole and SCH 28080 inhibited stimulated migration and blunted the fMLP-induced increase in CV. 6 Increasing extracellular osmolarity by addition of mannitol to the extracellular solution caused cell shrinkage followed by regulatory volume increase, partially due to activation of the Na + /H + exchanger. In fMLP-stimulated cells the CV increase was counteracted by simultaneous addition of mannitol. Under these conditions the fMLP stimulated migration was inhibited. 7 The antibacterial activity of PMNs was not modi®ed by Hoe 694 or omeprazole. 8 Western analysis with a monoclonal anti gastric H + /K + ATPase b-subunit antibody detected a glycosylated 35 kD core protein in lysates of mouse and human gastric mucosa as well as in human PMNs. 9 The results indicate that fMLP leads to cell swelling of PMNs due to activation of the Na + /H + exchanger and a K + -dependent H + -extruding mechanism, presumably an H + /K + ATPase. Inhibition of these ion transporters suppresses the increase in CV and precludes PMNs from stimulated migration.

Pravastatin, a hydrophilic inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, has been reported to beneficially affect atherogenesis, plaque stability, and transient myocardial ischemia in significant coronary artery disease by influencing lipid metabolism and by intracellular signaling via mevalonate pathway products other than cholesterol. Leukocytes are implicated to play a pathophysiological role in these events. We were interested in finding out whether pravastatin could affect transendothelial migration (TEM), chemotaxis, and respiratory burst activity of the neutrophil ex vivo. In addition, effects on monocyte and T-lymphocyte chemotaxis were tested. For TEM assays, monolayers of human umbilical vein endothelial cells (HUVECs) were grown to confluence on polycarbonate filters bearing 5-microns pores in Transwell (Costar) culture plate inserts. Chemotaxis experiments were performed using modified Boyden chambers with cellulose nitrate micropore filters. Respiratory burst activity was measured fluorometrically. Treatment of neutrophils and monocytes with pravastatin at 2 to 200 mumol/L and 10 to 1000 mumol/L, respectively, significantly decreased chemotaxis triggered by fMet-Leu-Phe. This effect was abolished in the presence of mevalonic acid (500 mumol/L); no effect of pravastatin was seen on T-lymphocyte chemotaxis triggered by interleukin-8. Preincubation of neutrophils with pravastatin (200 mumol/L) also resulted in a significant reduction in the number of neutrophils that transmigrated a tumor necrosis factor-stimulated or lipopolysaccharide-stimulated HUVEC monolayer. At none of the concentrations tested (2 pmol/L to 200 mumol/L) did pravastatin affect neutrophil respiratory burst activity. We conclude that pravastatin may alter monocyte chemotaxis and neutrophil-endothelial interactions in migratory responses at concentrations obtained in vivo with cholesterol-lowering doses.

Secretoneurin is a newly discovered 33-amino-acid peptide derived from secretogranin II (chromogranin C) that is found in sensory afferent C-fibers. We show here that secretoneurin triggers the selective migration of human monocytes in vitro and in vivo. Combinations of secretoneurin with the sensory neuropeptides, substance P or somatostatin, synergistically stimulate such migration. The attraction of monocytes represents the first established function of secretoneurin as a sensory neuropeptide.

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