[Frontiers in Bioscience 8, a40-47, January 1, 2003]

[Frontiers in Bioscience 8, a40-47, January 1, 2003]

MORAXELLA CATARRHALIS INDUCES MAST CELL ACTIVATION AND NUCLEAR FACTOR KAPPAB-DEPENDENT CYTOKINE SYNTHESIS

G. Krishnaswamy ^1,4, R. Martin ¹, E. Walker^1,4, C. Li ², F. Hossler³, K. Hall ¹ and D. S. Chi ¹

1 The Departments of Internal Medicine, ²Surgery and ¹Anatomy and Cell Biology, East Tennessee State University, Johnson City, Tennessee and ⁴The James H. Quillen V.A. Medical Center, Mountain Home, Tennessee

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and methods: 3.1. Mast cell culture and activation; 3.2. Preparation of bacterial suspensions; 3.3. ELISA for cytokines; 3.4. EMSA for nuclear factor kappaB; 3.5. Scanning electron microscopy
4. Results: 4.1. Binding of bacteria to mast cells; 4.2. Bacterial stimulation of cytokine production; 4.3. Direct bacterial contact is necessary for mast cell activation; 4.4. Role of nuclear factor kappaB in mast cell activation
5. Discussion
6. Acknowledgment
7. References

1. ABSTRACT

Human mast cells are often found perivascularly and at mucosal sites and may play crucial roles in the inflammatory response. Recent studies have suggested a prominent role for mast cells in host defense. In this study, we analyzed the effects of a common airway pathogen, Moraxella catarrhalis and a commensal bacterium, Neiserria cinerea, on activation of human mast cells. Human mast cell leukemia cells (HMC-1) were activated with either phorbol myristate acetate (PMA) and calcium ionophore or with varying concentrations of heat-killed suspensions of bacteria. Supernatants were assayed for the cytokines interleukin-4 (IL-4), granulocyte macrophage colony stimulating factor (GM-CSF), IL-6, IL-8, IL-13 and monocyte chemotactic protein-1 (MCP-1). Nuclear proteins were isolated and assayed by electrophoretic mobility shift assay (EMSA) for nuclear factor kappaB (NF-kB) nuclear binding activity. In some experiments, NF-kB inhibitor, Bay-11 was added to determine functional significance. Both M. catarrhalis and N. cinerea induced mast cell activation and selective secretion of two key inflammatory cytokines, IL-6 and MCP-1. This was accompanied by NF-kB activation. Neither spun bacterial supernatants nor bacterial lipopolysaccharide induced cytokine secretion, suggesting need for direct bacterial contact with mast cells. Scanning electron microscopy revealed active aggregation of bacteria over mast cell surfaces. The NF-kB inhibitor, Bay-11, inhibited expression of MCP-1. These findings suggest the possibility of direct interactions between human mast cells and common bacteria and provide evidence for a novel role for human mast cells in innate immunity.