Neurotrophins and the anti-inflammatory agents interleukin-4 (IL-4), IL-10, IL-11 and transforming growth factor-β1 (TGF-β1) down-regulate T cell costimulatory molecules B7 and CD40 on cultured rat microglia

Abstract

Microglia are essential for T cell activation in the CNS. Since T cell activation requires costimulation by B7 and/or CD40, we examined the regulation by cytokines of B7-1, B7-2 and CD40 mRNA expression in cultured rat microglia in serum-free medium. All three ligands are expressed constitutively, but are profoundly up-regulated by granulocyte-macrophage colony-stimulating factor (GM-CSF). By contrast, interferon-γ raises only B7-2 and CD40 mRNA, and the B7-2 increase is inhibited by IL-10. IL-4, transforming growth factor-β1, and nerve growth factor (NGF) repress GM-CSF-induced B7-2 and CD40, but not B7-1. NGF also down-regulates its own high-affinity trkA receptor. IL-11, unrecognized for its effect on antigen presentation, represses GM-CSF-induced B7-2.

Introduction

Brain microglia are salient mediators of the CNS immune response to a variety of insults (for review, see Merrill and Jonakait, 1995; Kreutzberg, 1996). T cell activation is often an essential component of that response. Because they resemble other professional antigen presenting cells (APCs) with respect to morphology, FcR expression, complement receptor-mediated phagocytosis and cell surface marker profile (Lowe et al., 1989; Ulvestad et al., 1994), microglia are likely to perform the task of antigen presentation and, hence, T cell activation in brain. We have, therefore, sought to examine the role played by microglia in effecting it.

The antigen-presenting ability of microglia has been addressed for a decade in terms of expression of major histocompatibility complex (MHC) type II molecules (Frei et al., 1987; Loughlin et al., 1993; Bo et al., 1994). However, the activation of T cells requires two membrane-bound signals from APCs (for review, see Lenschow et al., 1996): binding of the MHC class II (with its processed antigen) to the T cell receptor as well as engagement of costimulatory molecules on APCs with their corresponding receptors on T cells. Since failure of costimulatory ligand interaction results in negative, anergizing signals for T cell activation, the presence of MHC class II on microglia is not a sufficient indication of antigen-presenting ability. Hence, the potential role of microglia in antigen presentation needs to include detection of costimulatory molecule expression in microglia.

Candidate costimulators on APCs include B7 (B7-1 and/or B7-2) and/or CD40 (for review, see Foy et al., 1996; Lenschow et al., 1996). In the nervous system, both may be important. Both B7 and CD40 may play a role in the progress of multiple sclerosis and its experimental surrogate, experimental autoimmune encephalomyelitis (EAE) (Miller et al., 1995; Racke et al., 1995; Windhagen et al., 1995; Gerritse et al., 1996; Grewal et al., 1996; Issazadeh et al., 1998). Therefore, an understanding of the regulation of microglial costimulatory molecules is essential for a complete understanding of autoimmune diseases and other CNS lesions.

Several groups have begun to study both the constitutive and cytokine-inducible expression of B7 and CD40 on human and mouse microglia (Satoh et al., 1995; Becher and Antel, 1996; Dangond et al., 1997; Menendez Iglesias et al., 1997; Carson et al., 1998; Havenith et al., 1998). However, regulation of costimulatory molecules in isolated rat microglia has not been addressed; no study has performed this analysis in the absence of serum constituents; and very few studies have assessed microglial regulation of both B7 and CD40 simultaneously. We have systematically investigated cytokine regulation of B7-1, B7-2, and CD40 mRNA expression in microglia derived from neonatal rat cerebral cortices and cultured in serum-free medium utilizing semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) analysis and immunohistochemistry.

Because the activation of T cells often results in damaging inflammation, the role of anti-inflammatory regulators is of particular concern. In our study, we have focused on several molecules with anti-inflammatory properties including interleukin-4 (IL-4; Suzumura et al., 1994), IL-10 (Fiorentino et al., 1989), IL-11 (Trepicchio et al., 1996), and transforming growth factor-beta (TGF-β) (Kehrl et al., 1986). The recent suggestion that members of the neurotrophin family of growth factors and their receptors are expressed by microglia (Condorelli et al., 1995; Elkabes et al., 1996; Hart et al., 1997; Heese et al., 1998) and that they can serve an anti-inflammatory function (Neumann et al., 1998) has caused us to investigate their role in the regulation of costimulatory molecules. We find that the neurotrophins may have an important role in the down-regulation of CD40 and B7-2, but have a less important role for B7-1.