Western Society of Periodontics

Laboratory Studies

Volume Number 4, 1995

Polyamines found in gingival fluid inhibit chemotaxis by human polymorphonuclear leukocytes in vitro

This in vitro study was designed to test the hypothesis that polyamines such as putrescine or spermidine found at inflammatory sites would not enhance the regulation of PMN chemotaxis and phagocytosis. Whole blood was collected from healthy, medication-free human volunteers. PMNs were isolated, washed, and resuspended. Chemotaxis was assayed with a modified Boyden chamber using 3 ~m nitrocellulose filters. Putrescine or spermidine was added, and migration was expressed as a chemotactic index, defined as the sum of the products of the number of PMNs present at each 10 um interval times the distance traveled in microns. Phagocytosis of fluorescein-labeled Micrococcus Iysodeithcus was assayed, the bacteria were opsonized, and ingested bacteria were quantified with a fluorescence spectrometer. In addition, PMN filamentous actin (F-action), PMN adhesion, and FMet-Leu-Phe receptor expression were assayed. The influence of the polyamines on chemotactic index, adhesion, and phagocytosis was analyzed statistically.

Putrescine inhibited chemotaxis in a dose dependent manner; spermidine was more effective. Spermidine had no significant effect on the time course of actin polymerization (nor did putrescine). This test was used to determine whether the observed inhibition of chemotaxis might be related to polyamine-induced changes in actin polymerization. Although spermidine significantly inhibited adhesion at a concentration of 0.5 mM, it had no significant effect at 1 mM or below 0.5 mM. Putrescine had no apparent effect on phagocytosis over the range of 0.1 to 1 mM, whereas spermidine produced modest enhancement of phagocytosis at concentrations above 0.1 mM. This was not significant, however.

Putrescine and spermidine significantly inhibited chemotaxis to fMet-Leu-Phe and C5a (cytokine). This inhibition, however, was not strongly related to any effect polyamines have on PMN adhesion, actin polymerization, or formyl peptide receptor expression. Phagocytosis of opsonized bacteria was not significantly impacted by the polyamines. Therefore, at concentrations similar to those found in gingival fluid, polyamines could, under the findings in this study, potentially inhibit the migration of PMNs to diseased pockets without impairing their ability to phagocytize invading bacteria. [D.S.S.]