A role for autoantibodies in enhancement of pro-inflammatory cytokine responses to a self-antigen, thyroid peroxidase

Abstract

The role of thyroid peroxidase (TPO) antibodies (TPOAbs) in the pathogenesis of autoimmune thyroid disease is unclear. We selected sera with a high concentration of TPOAbs from eleven patients with Hashimoto's thyroiditis (HT), ten healthy monozygotic co-twins to HT patients, and twelve healthy individuals with no familiar disposition to AITD, and mixed each serum with normal mononuclear cells (MNCs). Following challenge with TPO, the MNCs' production of the pro-inflammatory cytokines TNF-α, IL-6 and IFN-γ, and the anti-inflammatory cytokine IL-10, correlated with the TPOAb content of the serum present in the culture (p = 0.0002–0.05). Enrichment of foetal calf serum-containing media with IgG with a high content of TPOAbs enhanced the TPO-elicited production of TNF-α, IL-6 and IFN-γ by normal MNCs in a dose- and Fcγ-receptor dependent manner (p < 0.0002–0.05). The data indicate that TPO-induced release of pro-inflammatory cytokines from phagocytic cells and T-cell responses to TPO are promoted by TPOAbs.

Introduction

While autoantibodies have a clearly defined role in the pathogenesis of certain autoimmune diseases, including Graves' disease (GD) and Myasthenia Gravis [1], the role of antibodies in autoimmune diseases known to be T-cell mediated, e.g. Hashimoto's thyroiditis (HT), remains obscure. Nonetheless, their presence often precedes clinically overt disease [1]. Examples are antibodies to thyroglobulin (TG) and thyroid peroxidase (TPO), which are predictive of the development of autoimmune thyroid disease (AITD), comprising HT and GD. Virtually all patients with active HT are seropositive for TPO antibodies (TPOAbs), and 80% of the patients are positive for TGAbs [2], [3], and it has been a matter of controversy whether they play a direct role in the pathogenesis, or occur merely as a by-product of T-cell mediated destruction of thyroid cells [4]. TPOAbs and TGAbs are present in low concentrations in the circulation of healthy individuals [5], [6], but the presence of these autoantibodies at elevated concentrations is predictive for the development of clinically overt disease [7].

Unlike TG, TPO is expressed on the surface of thyroid cells, rendering the cells susceptible to TPOAb-mediated complement-dependent cytotoxicity (CDC) [8], [9], [10], [11], [12], [13] and antibody-dependent cellular cytotoxicity (ADCC) [13], [14], [15], [16]. The primary effector cells appear to be monocytes, triggered via binding of TPOAbs to Fcγ-receptor I (FcγRI, CD64) or FcγRIII (CD16) [13].Thus, TPOAbs of the IgG1 subclass, that bind to FcγRs (reviewed in [17]), mediate thyroid cell damage, whereas non-FcγR-binding IgG4 antibodies, with the same specificity, do not [18]. Others, however, have failed to find any correlation between the TPOAb IgG subclass and lysis of thyroid cells [19].

Despite the possible involvement of TPOAbs in thyroid cell damage, HT is thought to be caused primarily by T-cell mediated inflammatory processes that lead to the death of thyroid cells and, consequently, to hypothyroidism [20]. We have previously proposed that activation of pathogenic T cells may be promoted by autoantibodies, by virtue of the capability of the antibodies to promote uptake of self-antigen by antigen-presenting cells (APCs) via their FcγRs and, in the case of complement-containing autoantibody/autoantigen complexes, via complement receptors [5], [21]. In support of this notion, macrophages from immune guinea pigs are capable of presenting low concentrations of antigen to specific T cells, while macrophages from naïve animals are effective only at high antigen concentrations, unless incubated with sera from immunised animals [22]. Moreover, specific antibodies to hepatitis B surface antigen [23], trinitrophenyl-keyhole limpet hemocyanin [24], β-galactosidase [25] and a self-antigen, the acetylcholine receptor [26], [27], enhance T-cell responses to the respective antigens. In addition, the activity of diabetogenic OVA-reactive CD8+ T cells in transgenic mice expressing ovalbumin (OVA) as “self” in the thymus and by the β cells of the pancreas is greatly enhanced by administration of anti-OVA IgG [28]. While a role for IgE in directing the binding of TPO to B cells via the low affinity FcεR (CD23) has been demonstrated [29], it is unclear whether IgG autoantibodies, generated spontaneously in autoimmune disease or in healthy conditions, facilitate such uptake of TPO by antigen-presenting cells.

In the present study, we have examined to which extent serum TPOAbs affect the cytokine responses of normal mononuclear cells (MNCs) to stimulation with TPO. The cells were incubated with TPO in the presence of sera from HT patients, and sera from two groups of healthy, euthyroid individuals: a group with no familiar disposition to autoimmune thyroid disease (AITD), and – to allow for any influence of polymorphic serum proteins on the cytokine responses – a group of healthy monozygotic (MZ) co-twins to patients with HT. Our data indicate that IgG autoantibodies, regardless of their origin, promote cytokine responses to TPO through enhancement of TPO uptake by antigen-presenting MNCs bearing Fcγ-receptors (FcγRs).