Pulmonary autoimmunity as a feature of autoimmune polyendocrine syndrome type 1 and identification of KCNRG as a bronchial autoantigen

Abstract

Patients with autoimmune polyendocrine syndrome type 1 (APS-1) suffer from multiple organ-specific autoimmunity with autoantibodies against target tissue-specific autoantigens. Endocrine and nonendocrine organs such as skin, hair follicles, and liver are targeted by the immune system. Despite sporadic observations of pulmonary symptoms among APS-1 patients, an autoimmune mechanism for pulmonary involvement has not been elucidated. We report here on a subset of APS-1 patients with respiratory symptoms. Eight patients with pulmonary involvement were identified. Severe airway obstruction was found in 4 patients, leading to death in 2. Immunoscreening of a cDNA library using serum samples from a patient with APS-1 and obstructive respiratory symptoms identified a putative potassium channel regulator (KCNRG) as a pulmonary autoantigen. Reactivity to recombinant KCNRG was assessed in 110 APS-1 patients by using immunoprecipitation. Autoantibodies to KCNRG were present in 7 of the 8 patients with respiratory symptoms, but in only 1 of 102 APS-1 patients without respiratory symptoms. Expression of KCNRG messenger RNA and protein was found to be predominantly restricted to the epithelial cells of terminal bronchioles. Autoantibodies to KCNRG, a protein mainly expressed in bronchial epithelium, are strongly associated with pulmonary involvement in APS-1. These findings may facilitate the recognition, diagnosis, characterization, and understanding of the pulmonary manifestations of APS-1.

Fig. 1.

Radiological and histological aspects of pulmonary involvement in APS-1 patients. (A) Pulmonary CT scan in patient 1 at the age of 11 years: peribronchial ground glass opacities. (B) Pulmonary CT scan in patient 1 at the age of 16 years, on immunosuppressive treatment with mycophenloate mofetil; the peribronchiolar abnormalities are improved. (C and D) Histological appearance of lung biopsy in patient 1 at the age of 11 years [10× magnification (C) and 40× magnification (D)]: peribronchiolar lymphoid infiltrate. (E) Pulmonary CT scan in patient 3 showing established bronchiectasis. (F) Lung biopsy on patient 3 showing bronchiolitis obliterans organizing pneumonia. (G and H) Histological appearance of lung biopsy in patient 4, at the age of 35 years, 2 years before his death from chronic respiratory failure [10× magnification (G) and 40× magnification (H)] showing severe peribronchiolar infiltrate.

Fig. 2.

Autoantibody reactivity to KCNRG and tissue expression of KCNRG messenger RNA. (A) Comparison of KCNRG autoantibody titers in sera from APS-1 patients, patients with different pulmonary disorders or other autoimmune disorders, and healthy blood donors. The assay for detection of autoantibodies is described in Materials and Methods. The dashed line indicates a cut of value of 0.41, which is the upper level of normal, defined as the mean results obtained for the healthy blood donors + 3 SD. (B) Expression of KCNRG mRNA in adult human tissues as measured by quantitative PCR, demonstrating that the expression of KCNRG is mainly restricted to pulmonary tissue. Please note the noncontinuous y axis.

Fig. 3.

Distribution of KCNRG protein in bovine lung. (A–C). Immunohistochemistry on 4-μm paraffin-embedded sections, using affinity-purified anti-KCNRG rabbit antiserum. (A) Background staining without the primary antibody. (B) Antiserum used in dilution 1:1000. (C) Antiserum in dilution 1:1,000 preabsorbed with 20 nmol of the peptide used for the immunization. (D–K) Immunofluorescence on 6-μm cryosections of lung, using patient and control serum samples. The blue background is from DAPI that stains nucleoli. FITC-conjugated goat anti-human IgG secondary antibody (green) was used. (D) Background staining when no primary serum is used. (E–G) Staining with 3 APS-1 patients' sera with KCNRG reactivity. (H and I) Staining with 2 APS-1 sera without KCNRG reactivity. (J and K) Staining with sera from 2 healthy blood donors.