Regulation of TNF-α Receptor Expression on Human Neutrophils by Various Cell Activating Factors

Abstract

Since tumor necrosis factor-α (TNF-α) has been implicated in a wide range of inflammatory states, we sought to examine the regulation of TNF-α receptors on human neutrophils (PMNs). We were particularly interested in the relationship of TNF-α binding characteristics to various known neutrophil-priming stimuli such as temperature, N -formylmethionyl-leucyl-phenylanine (fMLP), phorbol myristate acetate (PMA), C5a des Arg, and human recombinant C5a (HrC5a). Competition binding studies indicated that there was no significant change in receptor number or affinity due to the temperature used for neutrophil isolation. PMNs isolated at 4 or 25°C had a single class of receptors with a K d of 83.1 ± 3.3 vs 90.0 ±8.3 pmole/liter and a similar number of receptors, 1575 ± 99 vs 1876 ± 279 receptors/cell. However, when either group of isolated PMNs was further incubated in buffer at 37°C for 15 min, a significant increase in K d (4-37°C, 147.9 ± 23.7 pmole/liter; 25-37°C, 165.2 ± 29.2 pmole/liter) and a decrease in receptor number (4-37°C, 1116 ± 86 receptors/cell; 25-37°C, 957 ± 71 receptors/cell) were seen. To determine the effect of other activating substances such as fMLP, PMA, C5a des Arg, HrC5a, 4°C isolated PMNs were used. Pretreatment of PMNs (15 min at 37°C) by all of these agents down-regulated TNF-α receptors by decreasing receptor number and by decreasing receptor affinity. In the course of these studies, we identified a substantial disparity in the chemotactic deactivation and TNF-α receptor down-regulation of ED 50's for native C5a des Arg compared to recombinant C5a. These results would suggest that both loss of receptors and a change in affinity of the TNF-α receptor could be a means of modulating TNF-α response in neutrophils. We speculate that activation and release of the TNF-binding portion of the receptor from a proposed trimer receptor complex explains the affinity loss.