The role of complement activation in autoimmune liver disease

IntroductionThe complement system, an essential part of the innate immune system, is involved in various autoimmune diseases. Activation of the complement system by autoantibodies results in immune activation and tissue damage. At the moment little is known about the role of the complement system in autoimmune liver disease, including primary biliary cholangitis (PBC), primary sclerosing cholangitis (PSC) and autoimmune hepatitis (AIH). Since inhibition of the complement system is currently being tested in several autoimmune diseases as a therapeutic option, its role in autoimmune liver disease requires further clarification.MethodsA review of the literature was performed on studies investigating complement activation in PBC, PSC and AIH. Since data on AIH were lacking immunohistochemical staining for IgG, C1q, C3d, C4d and C5b9 was performed on liver tissue of nine AIH patients, two healthy controls and one positive control (acute liver failure caused by paracetamol intoxication).ResultsImmunohistochemical analysis in AIH revealed increased production of C3 and C4 by hepatocytes. Despite a strong staining for IgG in the immune infiltrate in AIH, C3d, C4d and C5b9 deposition was only present in one AIH patient and the deposition was restricted to the interface between portal tracts and liver parenchyma. No deposition was found in all other AIH patients or healthy controls.Literature review showed raised plasma C3 and C4 levels in AIH, PBC and PSC patients compared to healthy controls. For PBC and PSC no complement depositions at the bile ducts were reported.Conclusion and discussionAlthough complement is involved in various autoimmune diseases, the role of complement in autoimmune liver disease seems limited. Therefore it is unlikely that complement inhibition will become a novel treatment option for these diseases.     

Autoantibodies against complement C1q correlate with the thyroid function in patients with autoimmune thyroid disease

Autoantibodies against complement C1q (anti-C1q) have been well described in patients with systemic lupus erythematosus, where they correlate with the occurrence of severe lupus nephritis. However, data on anti-C1q in organ-specific autoimmune diseases are scarce. In order to determine the prevalence of anti-C1q in patients with autoimmune thyroid disorders (AITD) and a possible association with thyroid function, we measured prospectively anti-C1q in 23 patients with Graves' disease (GD) and 52 patients with Hashimoto's thyroiditis (HT). Anti-C1q levels were correlated with parameters of thyroid function and autoantibodies against thyroperoxidase, thyroglobulin and thyroid stimulating hormone (TSH) receptor. Twenty-one patients with multi-nodular goitre and 72 normal blood donors served as controls. We found elevated concentrations of anti-C1q more frequently in patients with AITD than in controls: seven of 23 (30%) patients with GD and 11 of 52 (21%) patients with HT, compared with one of 21 (5%) patients with multi-nodular goitre and six of 72 (8%) normal controls. Anti-C1q levels did not correlate with thyroid autoantibodies. However, in GD absolute levels of anti-C1q correlated negatively with TSH and positively with free thyroxine (FT4) and triiodothyronine (FT3). In contrast, in HT, anti-C1q correlated positively with TSH levels. No correlation between TSH and thyroid autoantibodies was found. In conclusion, we found an increased prevalence of anti-C1q in patients with AITD and their levels correlated with the thyroid function in both GD and HT. This correlation seems to be independent of thyroid autoantibodies. Therefore, anti-C1q might point to a pathogenic mechanism involved in the development of AITD that is independent of classical thyroid autoantibodies.     

Viral regulators of complement activation: Structure,function and evolution

The complement system surveillance in the host is effective in controlling viral propagation. Consequently, to subvert this effector mechanism, viruses have developed a series of adaptations. One among these is encoding mimics of host regulators of complement activation (RCA) which help viruses to avoid being labeled as ‘foreign’ and protect them from complement-mediated neutralization and complement-enhanced antiviral adaptive immunity. In this review, we provide an overview on the structure, function and evolution of viral RCA proteins.     

Terminal complement complexes and C1/C1 inhibitor complexes in autoimmune thyroid disease.

The potential role of complement activation and the membrane attack complex in the pathogenesis of Graves' disease and Hashimoto's thyroiditis has been investigated by measuring serum concentrations of the C1r-C1s-C1 inhibitor complex (C1/C1-inh) and the terminal complement complex (TCC), and by studying the binding to thyroid tissue of monoclonal and polyclonal antibodies against TCC neoantigens. Serum C1/C1-inh and TCC concentrations were significantly increased in 29 patients with untreated Graves' disease compared with 47 healthy subjects (P less than 0.001 for both), and decreased significantly after carbimazole treatment in 18 of these patients for whom post-treatment samples were available (P less than 0.01 and P less than 0.02, respectively). The serum TCC concentration, but not that of C1/C1-inh, was also significantly increased in 15 patients with Hashimoto's thyroiditis compared with the 47 healthy subjects (P less than 0.001). TCCs were identified by immunohistochemical staining around the thyroid follicles in thyroidectomy specimens from patients with Graves' disease (six out of six) and Hashimoto's thyroiditis (two out of two); normal thyroid tissue from two subjects showed no staining. These results suggest a role for complement, in particular the membrane attack complex in the pathogenesis of autoimmune thyroid disease.     

The complement system in systemic autoimmune disease

Complement is part of the innate immune system. Its major function is recognition and elimination of pathogens via direct killing and/or stimulation of phagocytosis. Activation of the complement system is, however, also involved in the pathogenesis of the systemic autoimmune diseases. Activation via the classical pathway has long been recognized in immune complex-mediated diseases such as cryoglobulinemic vasculitis and systemic lupus erythematosus (SLE). In SLE, the role of complement is somewhat paradoxical. It is involved in autoantibody-initiated tissue damage on the one hand, but, on the other hand, it appears to have protective features as hereditary deficiencies of classical pathway components are associated with an increased risk for SLE. There is increasing evidence that the alternative pathway of complement, even more than the classical pathway, is involved in many systemic autoimmune diseases. This is true for IgA-dominant Henoch Schönlein Purpura, in which additional activation of the lectin pathway contributes to more severe disease. In anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis the complement system was considered not to be involved since immunoglobulin deposition is generally absent in the lesions. However, recent studies, both in human and animal models, demonstrated complement activation via the alternative pathway as a major pathogenic mechanism. Insight into the role of the various pathways of complement in the systemic autoimmune diseases including the vasculitides opens up new ways of treatment by blocking effector pathways of complement. This has been demonstrated for monoclonal antibodies to C5 or C5a in experimental anti-phospholipid antibody syndrome and ANCA-associated vasculitis.     

The role of complement in autoimmune renal disease

The predominance of renal involvement in autoimmune diseases can most likely be assigned to the specialised function of the kidneys filtrating over 120 ml plasma per minute. Complement activation by autoantibodies directed against planted antigens or antigens already present in renal tissue in the subendothelial and mesangial regions provoke an inflammatory response ultimately resulting in renal damage. New data also suggest complement involvement in the pathogenesis of renal disease caused by subepithelial immune complex deposition. On the other hand complement itself can also be a target of an autoimmune responses causing renal damage as seen in SLE. The results on intervention of complement activation in clinical practise are awaited.     

Gm phenotypes in autoimmune thyroid disease

The Gm phenotype Gm f,b or Gm f,n,b was found in all forty patients with Graves' disease studied, contrasted with thirty-five out of forty controls and twenty out of thirty-one patients with thyroiditis. The difference between the two groups with autoimmune thyroid disease was significant. These results suggest that thyroid stimulating antibodies may be allotypically restricted.     

Cell-mediated immunity in autoimmune thyroid disease

The initiation of an autoimmune response requires the establishment of an appropriate microenvironment. This, in turn, involves several requirements, including antigen expression on the membrane surface of the target cells, class II antigen expression on the antigen-presenting cell or target cell, a relative systemic or local increase in the helper/inducer subset of T cells, and/or a relative decrease in the suppressor subset of T cells. All of these conditions have been described in the thyroid gland. Appropriate cellular interactions result in the appearance of activated T cells and the generation of cytotoxic T cells. The pathologic alterations may be produced by the local production of antibody and subsequent formation of immune complexes, by direct lymphocyte damage, or by lymphokine production. Autoimmune thyroid disease remains, to our minds, the most instructive paradigm of the organ-specific autoimmune endocrinopathies.     

The role of Fas-mediated apoptosis in thyroid autoimmune disease.

Apoptosis is a carefully regulated mechanism of cell death that differs from necrosis and plays an important role in normal tissue development and homeostasis, as well as disease processes. Apoptosis also plays an important role in autoimmunity. Defective apoptosis can cause systemic autoimmunity by allowing the survival of autoreactive lymphocytes. It may also be involved in the pathogenesis of organ-specific autoimmune diseases, such as Hashimoto's thyroiditis, through altered target organ susceptibility. Apoptosis signaling pathways can be initiated through activation of death receptors. One of these pathways employs the death receptor Fas and its ligand (FasL). Fas expression and death pathway signaling have been demonstrated in the thyroid, but there is controversy surrounding the expression of FasL and its role in thyroid autoimmunity. A number of proteins, including FAP-1, Bcl-2 and I-FLICE may regulate the Fas pathway in the thyroid and provide potential mechanisms for modifying the pathogenesis of autoimmune thyroid disease.     

Antigen-specific T cell recognition of affinity-purified and recombinant thyroid peroxidase in autoimmune thyroid disease.

The T cell proliferative responses of peripheral blood lymphocytes from 20 patients with autoimmune thyroid disease (AITD) and 20 healthy controls were analysed to immunoaffinity-purified thyroid peroxidase (TPO) and recombinant antigen preparations generated in Escherichia coli as glutathione-s-transferase fusion proteins. The epitope specificity of the T cell response was investigated using a selection of eight discrete recombinant fragments encompassing the whole of the extracellular region of the TPO molecule. Significant differences in the proliferative responses between patients and controls were observed to the full length, affinity-purified TPO molecule (P less than 0.002) as well as to the recombinant fragments R1c (residues 145-250) (P less than 0.001) and R2b (residues 457-589) (P less than 0.001) suggesting the presence of at least two distinct T cell determinants on this autoantigen. One of these T cell epitopes, localized within the region R1c, has not previously been identified by studies with synthetic peptides.     

T, B and K cells in autoimmune thyroid disease.

The K-cell cytotoxic activity of peripheral blood lymphoid cells from 104 patients with autoimmune thyroid disease and from age and sex matched control subjects was measured using chicken erythrocytes as target cells. Patients with Hashimoto thyroiditis,primary hypothyroidism and thyrotoxicosis who were either newly diagnosed and untreated or had received therapy for less than or equal to 1 year showed a significant increase in K-cell cytotoxic activity. Patients who had received treatment for greater than 1 year and less than or equal to 5 years showed no such comparable increase in cytotoxic activity. Within the group of patients with untreated thyrotoxicosis it was found that K-cell cytotoxic activity was related to both goitre size and serum antibody titre. Thus patients with little or no goitre showed a highly significant elevation of cytotoxic activity whereas patients with moderate to large goitres gave values within the normal range. Similarly patients with no detectable serum thyroid autoantibodies showed high K-cell activity while patients with positive antibody titres did not. It was also shown that neither the absolute number nor the proportion of circulating T and B lymphocytes in patients with autoimmune thyroid disease as assessed by the sheep red cell rosette method and by indirect immunofluorescence was significantly different from that observed in the normal control population. No correlation was found between peripheral blood K-cell cytotoxic activity and the percentage of circulating null cells, i.e. 100-(percentage T + percentage B) in either patients or control subjects.     

T cell responses to synthetic thyroid peroxidase peptides in autoimmune thyroid disease.

Sixteen peptides, representing four different extracellular regions of thyroid peroxidase (TPO) predicted to contain a high proportion of potential T cell epitopes, were synthesized to investigate which parts of this autoantigen may be targets for the T cell response in thyroid autoimmunity. Compared with 25 controls, peripheral blood T cells from 23-37% of 30 patients with Graves' disease or autoimmune hypothyroidism were stimulated significantly by three peptides, representing amino acids 415-432, 439-457 and 463-481 of the TPO sequence; T cells from individual patients were also stimulated by several other peptides. These results indicate that the T cell response to TPO is directed against several epitopes which may be recognized by different patients.     

Anti-perinuclear and rheumatoid factor in different forms of autoimmune thyroid disease.

Anti-perinuclear factors (APF), IgM-, IgG- and IgA-rheumatoid factors have been detected in six, five, one and three out of 20 patients with primary myxoedema, respectively. We were unable to find APF in 22 patients with Hashimoto's thyroiditis nor in 20 patients with Graves' thyrotoxicosis. APF appears to be a marker for non-organ specific autoimmune diseases.     

Expression of CD40 and apoptosis related molecules in autoimmune thyroid diseases.

Apoptosis is responsible for the loss of thyrocytes in autoimmune thyroiditis. Recent investigations into the pathogenesis of apoptosis have revealed that the important roles of suicide molecules expression on both thyrocytes and cytotoxic T-lymphocytes. To study the mechanism of thyrocyte loss in various forms of thyroiditis, we evaluated in situ expression patterns of CD40, Fas, and Fas-L on thyrocytes and infiltrating inflammatory cells by immunohistochemical staining of thyroid samples obtained from 49 patients (Graves' disease, n=10: Hashimoto's thyroiditis, n=14; nonspecific lymphocytic thyroiditis, n=11; subacute granulomatous thyroiditis, n=11; normal, n=3). The role of cytotoxic T-lymphocytes was also evaluated by analyzing the expression of granzyme B along with their phenotypic characteristics. CD40 was not expressed on thyrocytes of normal controls while they showed a diffuse expression of Fas and a scattered focal expression of Fas-L. The plump thyrocytes proximal to the inflammatory infiltrates showed more intense expressions of these three molecules in various forms of thyroiditis and a close correlation was found between CD40 and Fas-L expression on thyrocytes. Unlike Fas, which was expressed on infiltrating lymphocytes in all groups, Fas-L was not expressed on infiltrating lymphocytes, except those in subacute granulomatous thyroiditis. Granzyme B expressing activated cytotoxic T-lymphocytes occupied a negligible proportion of CD8+ T-lymphocytes in various forms of thyroiditis, and no difference was found in terms of their proportions according to the type of thyroiditis. These results show the acquisition of CD40, Fas and Fas-L molecules on thyrocytes proximal to inflammatory cell aggregates and the negligible expression of granzyme B and Fas-L on the infiltrating lymphocytes, and suggest that Fas and Fas-L mediated apoptosis of thyrocytes (fratricide) may be more important than T cell-mediated cytotoxicity in various forms of thyroiditis.     

The immunoregulatory disturbance in autoimmune thyroid disease

There is now considerable evidence for a genetically induced antigen-specific defect in suppressor T lymphocytes as the basis for AITD, derived from several laboratories and via different types of experimental techniques. In addition, there is now evidence for additive effects on reducing generalized suppressor T lymphocyte numbers and/or function by environmental factors as well as hyperthyroidism itself, and these effects would be superimposed upon the organ-specific defect. Such effects on generalized suppressor T lymphocyte numbers may act as precipitating and self-perpetuating factors, in Graves' disease at least. Presentation of the antigen by the thyroid cell via HLA-DR expression on its membrane does occur as a result of interferon gamma production by T lymphocytes. This in turn appears to be secondary to the initial specific immune assault and is not a primary inductive step. However, it may be important as an amplifying intermediate factor but cannot perpetuate the process in the absence of the underlying immune disorder. There is, however, no evidence for an underlying antigenic abnormality or stimulus in human autoimmune thyroid disease, and the initiating event would appear to be due to perturbation of the generalized immune system superimposed on the organ-specific abnormality. Variations in the serological and clinical expression of AITD would appear to depend on the severity of the original organ-specific disturbance in suppressor T lymphocyte function, plus the added factor of environmental influences playing upon generalized suppressor T lymphocyte function and numbers. Remissions in Graves' disease brought about by antithyroid drugs may well be via their effect on modulating the thyroid cell activity; this will then reduce thyrocyte-immunocyte signalling, allowing remission to occur in those patients with a partial organ-specific defect in suppressor T lymphocytes.