Research

Protein kinase C promotes restoration of calcium homeostasis to platelet activating factor-stimulated human neutrophils by inhibition of phospholipase C

Gregory R Tintinger¹ , Annette J Theron² , Helen C Steel² , Riana Cockeran² , Lynette Pretorius² and Ronald Anderson²

¹  Department of Internal Medicine, Faculty of Health Sciences, University of Pretoria, South Africa

²  Medical Research Council Unit for Inflammation and Immunity, Department of Immunology, School of Medicine, Faculty of Health Sciences, University of Pretoria and Tshwane Academic Division of the National Health Laboratory Service, Pretoria, South Africa

author email corresponding author email

Journal of Inflammation 2009, 6:29doi:10.1186/1476-9255-6-29

Published:	30 October 2009

Abstract

Background

The role of protein kinase C (PKC) in regulating the activity of phospholipase C (PLC) in neutrophils activated with the chemoattractant, platelet-activating factor (PAF, 20 and 200 nM), was probed in the current study using the selective PKC inhibitors, GF10903X (0.5 - 1 μM) and staurosporine (400 nM).

Methods

Alterations in cytosolic Ca²⁺, Ca²⁺ influx, inositol triphosphate (IP₃), and leukotriene B₄ production were measured using spectrofluorimetric, radiometric and competitive binding radioreceptor and immunoassay procedures, respectively.

Results

Activation of the cells with PAF was accompanied by an abrupt increase in cytosolic Ca²⁺ followed by a gradual decline towards basal levels. Pretreatment of neutrophils with the PKC inhibitors significantly increased IP₃ production with associated enhanced Ca²⁺ release from storage vesicles, prolongation of the peak cytosolic Ca²⁺ transients, delayed clearance and exaggerated reuptake of the cation, and markedly increased synthesis of LTB₄. The alterations in Ca²⁺ fluxes observed with the PKC inhibitors were significantly attenuated by U73122, a PLC inhibitor, as well as by cyclic AMP-mediated upregulation of the Ca²⁺-resequestering endomembrane ATPase.

Taken together, these observations are compatible with a mechanism whereby PKC negatively modulates the activity of PLC, with consequent suppression of IP₃ production and down-regulation of Ca²⁺ mediated pro-inflammatory responses of PAF-activated neutrophils.

Conclusion

Although generally considered to initiate and/or amplify intracellular signalling cascades which activate and sustain the pro-inflammatory activities of neutrophils and other cell types, the findings of the current study have identified a potentially important physiological, anti-inflammatory function for PKC, at least in neutrophils.