The ILDs are a heterogeneous group of disorders, the most common of which is CFA. The latter is an inflammatory condition of the lungs that results in scarring, pulmonary failure and death. The aetiology of the disease is unknown, but the pathogenesis may involve an immunological reaction to unidentified antigens in the lung, resulting in tissue damage. Recent reports suggest a higher prevalence of CFA than was previously documented (13.2–20.2/100,000 population) and a rising mortality rate. The prognosis is universally poor, with 50% of patients dying within 5 years. Although approximately 30% of patients may live for long periods (>10 years), morbidity is significant and quality of life in long-term survivors is poor [1].

Diagnosis is traditionally based on an open lung biopsy, but more recently high-resolution computed tomography has been used. It has been shown that CFA is associated with the production of circulating IgG autoantibodies to antigen(s) that are associated with alveolar lining cells, and there is evidence for a local humoral immune response associated with B-lymphocyte aggregates in the lungs [2,3,4]. The presence of large numbers of B lymphocytes in open lung biopsies may be associated with a poor prognosis, but it should be noted that B cells are not a prominent feature of this disease. Although serum levels of anticytokeratin 8 antibody are increased, it is not clear whether they are involved in the pathological process or are epiphenomenal [5]. Antinuclear antibodies have long been recognized as a feature of CFA [6]. Antitopoisomerase II antibodies have also been found in approximately 37% of sera from patients with CFA [7,8]. In addition, antibodies to poly (ADP-ribose) polymerase have been identified in up to 25% of patients [9].

Serological identification of antigens by recombinant expression cloning can readily be applied to define antigens at the molecular level. We applied the technique using serum from a patient with the canonical features of CFA.

It is evident that the humoral immune system plays some role in the pathogenesis of CFA. It is less clear whether this role is causative or merely epiphenomenal. Although circulating autoantibodies to lung protein(s) have been mooted as the pathogenic mechanism, relatively few antigens have actually been characterized and none of these is accepted as diagnostic for the disease. An autoantibody reactive to cytokeratin 8 was defined by western immunoblotting and by enzyme-linked immunosorbent assay [5]. Similar data support a role for antibodies that are reactive with cytokeratin 19 in the process of lung injury in pulmonary fibrosis [13]. The majority of autoantibodies appear directed against antigens that are not specifically localized to lung tissue, for example antibodies to DNA topoisomerase IIα, a nuclear antigen that is constitutively expressed by dividing cells. These antibodies are prevalent in the sera of some patients, and analysis of the specific patterns of reactivity with various deletion recombinants revealed the existence of multiple epitopes in the protein [14]. Interestingly, the authors of that study noted a correlation between the type of epitope and the disease duration, supporting the hypothesis that the autoantibody response is an antigen-driven process.

Not many of the antigens associated with CFA actually show a tissue-specific pattern of expression that is limited to the lung. This is surprising because the disease is explicitly manifest as a pulmonary lesion. Lungs may be targeted simply because their high blood flow means that they are more likely to trap immune complexes. Alternatively, the majority of antibodies may be epiphenomenal and associated with tissue damage caused by the primary lesion. Two of the antigens described here show strong association with epithelial cells, although they are clearly expressed in a wider range of tissues. The third antigen, ATS, is expressed in all tissues at high levels. This high level of expression is evident from the finding of six independent clones encoding ATS in the library that we probed. The mRNA source of this library originated from a lung tumour cell line. We chose this library because of its complexity and because we knew that it contained large inserts. In addition, western blots using patients' sera strongly suggested that there was no bias in the target tissue for the dominant antigens. The SAGE analysis, through its virtual northern programme, showed that each of the antigens that we cloned are widely expressed.

Each of the reactivities that we defined is associated with T-cell recognition of the antigen. This follows simply because we used an isotype-specific second antibody, and class switching to IgG is a T-cell-dependent event. It seems likely then that the spectrum of antigen reactivity in the serological response is dictated by the availability of autoreactive T cells. How these arise, particularly to highly expressed ubiquitous proteins such as ATS, and why subsets of antinuclear antibodies to ubiquitous determinants define particular syndromes remain unresolved issues.

The antisynthetase syndrome is such a syndrome. It typically associates polymyositis and dermatomyositis with complicating ILD, Raynaud's phenomenon and frequently polyarthritis. Autoantibodies against aminoacyl-tRNA synthetases (antisynthetases) define the syndrome. Patients can present with ILD and antisynthetase antibodies, but no clinically evident myositis. Previous studies have shown that anti-PL-12 antibodies (anti-ATS) are the most common in this group, but anti-Jo-1 (antihistidyl-tRNA synthetase), anti-EJ (antiglycyl-tRNA synthetase) and anti-OJ (anti-isoleucyl-tRNA synthetase) also occur [15,16]. Patients usually exhibit reactivity to only one of the synthetases, but different subsets have occasionally been recorded together in the same patient [17]. Of interest, autoantibodies to synthetases have been found in mice with graft-versus-host disease [18], suggesting that they may be a secondary consequence of tissue damage and cell death.

The serum samples were obtained from 3 months to 4 years after the onset of disease in the patients with CFA, and 1–5 years after presentation in the sarcoidosis patients. The SLE patient had been diagnosed with the disease for 3 months. Immunosuppressive therapy, including oral prednisolone, was being taken by five of the CFA patients and by four of the sarcoidosis patients. The index patient was on no treatment when the initial sample was obtained, but he was on immunosuppressive therapy when the later samples were obtained and was improving physiologically. The SLE patient was receiving plaquenil only. Taken together, it seems unlikely that therapy was associated with the negative results because not all patients were on steroids and because steroids made no difference to the results in the index case.

CFA can involve an inappropriate immunological reaction in the lung interstitia. It is not clear what promotes and maintains this reactivity. We anticipated that the targets of the humoral immune response might give us clues to the originating insult. We demonstrated by western blot that there were likely to be common as well as unique antigenic specificities in the patient population studied. However, by molecularly cloning and identifying the antigens that were recognized by our index patient, we were able to demonstrate that they were not recognized by any other patient. CFA may represent the end stage of a number of different disease processes, some of which may be autoantibody mediated. It is also possible that autoantibodies are epiphenomenal to CFA, representing a secondary feature of tissue damage induced by some other mechanism.