IgG4-related pulmonary disease: the protean impersonator?

IgG4-related disease is an immune-mediated fibro-inflammatory disease, characterised by distinct pathological features. An increasing number of clinical phenotypes are described, from single-organ disease to a multisystem disorder, which can present to a variety of different specialities. Recognition is key; its protean manifestations can mimic other inflammatory diseases, infection and malignancy. Here, we present three cases to highlight the importance of being familiar with this condition in its various forms.

The role of biologics in treatment of connective tissue disease-associated interstitial lung disease.

With an increased understanding of the molecular pathways of inflammation and autoimmunity, the development of targeted biological agents has revolutionized the management of connective tissue diseases (CTDs). There has been an explosion in the development of these drugs in the last decade, targeting diseases in diverse fields including: allergic disorders, oncology, neuroinflammatory disorders, inflammatory bowel disease, macular degeneration and CTDs. In this last field, commonly applied biologics fall into two categories: cytokine inhibitors and lymphocyte-targeted therapies. The former group includes the antitumour necrosis factor alpha (TNF-α), anti-interleukin (IL)-6 receptor monoclonal antibodies and IL-1 receptor antagonists, whilst the latter encompasses the anti-CD20, B-cell depleting, monoclonal antibody (mAb), Rituximab and the anti-T-cell activation agent, Abatacept. This review will examine our developing experience in the use of these agents in the treatment of CTD-related interstitial lung diseases, with a particular focus on B-cell depletion.

BCLAF1 is a radiation-induced H2AX-interacting partner involved in γH2AX-mediated regulation of apoptosis and DNA repair.

H2AX, a histone H2A variant, has a key role in the cellular response to DNA double-strand breaks (DSBs). H2AX senses DSBs through rapid serine 139 phosphorylation, concurrently leading to the formation of phospho-(γ)H2AX foci with various proteins. However, in the cells with different sensitivity to ionizing radiation (IR)-induced DSBs, still incomplete are those specific proteins selectively recruited by γH2AX to decide different cell fates. Because the abundance of γH2AX indicates the extent of DSBs, we first identified IR-induced dose-dependent H2AX-interacting partners and found that Bcl-2-associated transcription factor 1 (BCLAF1/Btf) showed enhanced association with γH2AX only under high-dose radiation. In acutely irradiated cells, BCLAF1 promoted apoptosis of irreparable cells through disturbing p21-mediated inhibition of Caspase/cyclin E-dependent, mitochondrial-mediated pathways. Meanwhile, BCLAF1 co-localized with γH2AX foci in nuclei and stabilized the Ku70/DNA-PKcs complex therein, facilitating non-homologous end joining (NHEJ)-based DSB repair in surviving cells. In tumor cells, BCLAF1 was intrinsically suppressed, leading to formation of anti-apoptotic Ku70-Bax complexes and disruption of Ku70/DNA-PKcs complexes, all of which contribute to tumor-associated apoptotic resistance and cell survival with defective NHEJ DNA repair. For the first time, our studies reveal that, based on the extent of DNA damage, BCLAF1 is involved in the γH2AX-mediated regulation of apoptosis and DNA repair, and is a γH2AX-interacting tumor suppressor.

Autoantibodies to a 140-kd protein in juvenile dermatomyositis are associated with calcinosis.

OBJECTIVE: The identification of novel autoantibodies in juvenile dermatomyositis (DM) may have etiologic and clinical implications. The aim of this study was to describe autoantibodies to a 140-kd protein in children recruited to the Juvenile DM National Registry and Repository for UK and Ireland. METHODS: Clinical data and sera were collected from children with juvenile myositis. Sera that recognized a 140-kd protein by immunoprecipitation were identified. The identity of the p140 autoantigen was investigated by immunoprecipitation/immunodepletion, using commercial monoclonal antibodies to NXP-2, reference anti-p140, and anti-p155/140, the other autoantibody recently described in juvenile DM. DNA samples from 100 Caucasian children with myositis were genotyped for HLA class II haplotype associations and compared with those from 864 randomly selected UK Caucasian control subjects. RESULTS: Sera from 37 (23%) of 162 patients with juvenile myositis were positive for anti-p140 autoantibodies, which were detected exclusively in patients with juvenile DM and not in patients with juvenile DM-overlap syndrome or control subjects. No anti-p140 antibody-positive patients were positive for other recognized autoantibodies. Immunodepletion suggested that the identity of p140 was consistent with NXP-2 (the previously identified MJ autoantigen). In children with anti-p140 antibodies, the association with calcinosis was significant compared with the rest of the cohort (corrected P < 0.005, odds ratio 7.0, 95% confidence interval 3.0-16.1). The clinical features of patients with anti-p140 autoantibodies were different from those of children with anti-p155/140 autoantibodies. The presence of HLA-DRB1*08 was a possible risk factor for anti-p140 autoantibody positivity. CONCLUSION: This study has established that anti-p140 autoantibodies represent a major autoantibody subset in juvenile DM. This specificity may identify a further immunogenetic and clinical phenotype within the juvenile myositis spectrum that includes an association with calcinosis.

Clinical and human leucocyte antigen class II haplotype associations of autoantibodies to small ubiquitin-like modifier enzyme, a dermatomyositis-specific autoantigen target, in UK Caucasian adult-onset myositis.

OBJECTIVES: Autoantibodies to a novel autoantigen small ubiquitin-like modifier activating enzyme (SAE) associated with dermatomyositis (DM) have previously been identified. The aim of this study was to establish the frequency of anti-SAE autoantibodies in a UK myositis cohort and investigate clinicoimmunogenetic associations. METHODS: Clinical data and sera were studied from 266 patients recruited to the Adult Onset Myositis Immunogenetic Collaboration. Myositis sera, control sera including 250 patients with other connective tissue diseases and 50 healthy participants were screened using radio-immunoprecipitation. Immunodepletion was performed on all sera immunoprecipitating 40 and 90 kDa bands to confirm the presence of anti-SAE. DNA from 202 patients with myositis was genotyped for human leucocyte antigen (HLA)-DRB1 and DQB1; DQA1 data were inferred. RESULTS: Out of 266 patients with myositis, 11 (4%) were positive for anti-SAE, which was found exclusively in DM with a frequency of 8%. Patients with anti-SAE had a high frequency of cutaneous lesions including heliotrope (82%) and Gottron rash (82%). Of the 11, 9 (82%) had systemic features and 7 of 9 (78%) developed dysphagia. Of those nine, seven (78%) presented with skin disease before myositis onset. All patients with anti-SAE possessed at least one copy of HLA-DQB1*03. HLA-DRB1*04-DQA1*03-DQB1*03 was a significant risk factor in anti-SAE positive versus patients who were anti-SAE negative (haplotype frequency 18% vs 6%, p<0.001, OR 5.7, 95% CI 1.9 to 17.3). CONCLUSIONS: Anti-SAE is a myositis-specific autoantibody that identifies a subset of patients with adult DM. The majority of patients with anti-SAE presented with cutaneous disease and progressed to myositis with systemic features including dysphagia. This novel autoantibody has a strong association with the HLA-DRB1*04-DQA1*03-DQB1*03 haplotype.

The protein tyrosine phosphatase N22 gene is associated with juvenile and adult idiopathic inflammatory myopathy independent of the HLA 8.1 haplotype in British Caucasian patients.

OBJECTIVE: To examine single-nucleotide polymorphisms (SNPs) of the protein tyrosine phosphatase N22 gene (PTPN22) and to study the relationship between PTPN22 and the HLA region in patients with idiopathic inflammatory myopathies (IIMs). METHODS: PTPN22 SNPs were assessed in a large, cross-sectional, case-control study from the UK involving patients with adult or juvenile IIM, comprising patients with polymyositis (PM) (n=114), dermatomyositis (DM) (n=102), myositis associated with another connective tissue disease (myositis-CTD overlap syndrome) (n=64), or juvenile DM (n=101), in comparison with 748 control subjects. Seventeen PTPN22 SNPs were genotyped using the Sequenom MassArray iPLEX platform. Serotyping for myositis-specific/myositis-associated autoantibodies (MSAs/MAAs) was performed by radioimmunoprecipitation. RESULTS: A significant association was noted between the R620W variant (rs2476601) and IIM (corrected P [Pcorr]=0.0009 versus controls), and specifically with the clinical subgroup of PM (Pcorr=0.003 versus controls). A weaker association was noted with juvenile DM (Pcorr=0.009 versus controls). No significant associations were noted after stratification by serologic subgroups. The association with the R620W variant was independent of alleles forming the HLA 8.1 haplotype. No other PTPN22 SNPs were associated with IIM. The PTPN22 haplotype containing the R620W T allele was the only haplotype significantly associated with IIM. CONCLUSION: The R620W variant is a significant risk factor for IIM, independent of the HLA 8.1 haplotype. Unlike that in the HLA region, risk is not increased in individuals possessing MSAs/MAAs. These results are further evidence that the PTPN22 gene confers autoimmune susceptibility.

Clinical associations of autoantibodies to a p155/140 kDa doublet protein in juvenile dermatomyositis.

OBJECTIVES: Myositis-specific autoantibodies (MSAs) may define homogeneous clinical subsets of adult patients with dermatomyositis (DM). Recently, there have been descriptions of novel autoantibodies in DM. This study was conducted to establish the clinical significance of anti-p155/140 autoantibodies in juvenile DM (JDM). METHODS: The first 116 children recruited to the JDM National Registry and Repository (UK and Ireland) were studied. Comprehensive clinical features were recorded and sera screened for anti-p155/140 autoantibodies using radio-immunoprecipitation. Sera from adults with DM (n = 20), PM (n = 25), SSc (n = 150), SLE (n = 40) and healthy subjects (n = 50) were used for comparison. Immunodepletion experiments were used to establish whether the p155/140 kDa targets recognized by JDM sera were the same as adult DM sera. RESULTS: Twenty-seven out of 116 (23%) JDM cases were positive for anti-p155/140 in comparison with 6/20 (30%) adults with DM. Immunodepletion confirmed that the 155/140 kDa proteins recognized by JDM and adult DM sera were the same targets. All other adult control sera were negative for anti-p155/140 autoantibodies. There was a higher frequency of males in the anti-p155/140-positive JDM group (P = 0.02). JDM patients with anti-p155/140 autoantibodies had significantly more cutaneous involvement including Gottron's papules (P = 0.003), ulceration (P = 0.005) and oedema (P = 0.013). The distribution of skin lesions was more extensive particularly periorbitally (P = 0.014) and over the small (P < 0.001) and large joints (P = 0.003). CONCLUSIONS: Anti-p155/140 autoantibodies are clinically significant in JDM and may define a clinical subset in terms of disease severity and outcome. The same autoantigen target is detected in adult DM patients.

Microvascular responses following digital thermal hyperaemia and iontophoresis measured by laser Doppler imaging in idiopathic inflammatory myopathy.

OBJECTIVE: To measure microvascular function using laser Doppler imaging following digital hyperaemia and iontophoresis of vasoactive substances in patients with idiopathic inflammatory myopathy (IIM). METHODS: Fifteen patients with IIM including eight patients with dermatomyositis (DM) and seven patients with polymyositis (PM) were studied. Fifteen age-matched normal controls were also recruited. Scanning red laser Doppler imaging (LDI) was performed after resting the subject in a temperature controlled room at 23 degrees C for 20 min. An initial LDI scan was performed to assess baseline blood flow. Digital microvascular responses were quantified following a maximum hyperaemic response (MHR) and iontophoresis with endothelial dependent acetylcholine (Ach) and endothelial independent sodium nitroprusside (SNP). Maximum vasodilation following iontophoresis was expressed as a percentage of the MHR. RESULTS: All subjects were age matched, and the duration of disease was similar between the IIM patients. There was no significant difference in baseline blood flow when comparing the three study groups. There was no significant difference in MHR or SNP/MHR when comparing DM or PM with controls. However, Ach/MHR was significantly lower in both the DM and PM group compared with controls (both P < 0.01). There was no significant difference in any of the microvascular responses when comparing patients with DM directly with PM. CONCLUSION: This is the first study to evaluate microvascular responses using LDI in patients with IIM. We have demonstrated that patients with DM have abnormal endothelial dependent mediated vasodilation and the same abnormality is present in patients with PM.

Maximum blood flow and microvascular regulatory responses in systemic sclerosis.

Objectives. To investigate microvascular function using laser Doppler imaging (LDI) following response to hyperaemia, neurovascular reflex and iontophoresis in systemic sclerosis (SSc) in comparison with primary Raynaud's phenomenon (PRP) and age-matched controls. A secondary aim was to evaluate if SSc patients with a higher Medsger vascular score have lower endothelial responses. Methods. Twenty patients with SSc, 10 PRP and 17 controls were studied. Patients with SSc were scored using the vascular component of the Medsger severity scale. A baseline LDI scan was performed on the dorsal aspect of both hands. Digital responses were quantified following maximum hyperaemic response (MHR), contralateral vasoconstrictor response (CLVc) and iontophoresis with acetylcholine (Ach)-endothelial dependent and sodium nitroprusside (SNP)-endothelial independent. Mean blood flow was quantified over a standard region of interest. Results. MHR was lower in SSc patients compared with controls (P < 0.001). A similar trend was seen when comparing SSc with PRP although this did not reach significance (P = 0.07). CLVc and Ach/MHR were lower in SSc vs PRP (P < 0.05) and controls (P < 0.001). No difference was observed in MHR, CLVc and Ach/MHR between PRP and controls. Overall, SNP/MHR was similar in all the three groups. SSc patients with a higher Medsger vascular score had lower endothelial-dependent (P < 0.01) and independent (P < 0.05) responses. Conclusion. SSc patients have abnormal microvascular regulatory responses compared with PRP and controls. This study also suggests that the degree of endothelial dysfunction may be related to the degree of peripheral vascular involvement.

Anti-synthetase syndrome: a new autoantibody to phenylalanyl transfer RNA synthetase (anti-Zo) associated with polymyositis and interstitial pneumonia.

OBJECTIVE: Autoantibodies directed against the aminoacyl tRNA synthetases are associated with myositis, arthritis, Raynaud's phenomenon, mechanic's hands, fever and interstitial pneumonia, clinically referred to as the anti-synthetase syndrome (ASS). The aim of this study was to characterize the autoantibody profile in a patient with clinical features of ASS whose routine diagnostic testing was negative for the previously identified anti-synthetase autoantibodies. METHODS: Serum from a patient presenting with interstitial pneumonia followed by proximal myopathy, Raynaud's phenomenon and arthrlagia was analysed for autoantigen specificity by routine methods including indirect immunofluorescence, immunodiffusion, ELISA and immunoblotting. The autoantibody specificity was further analysed by RNA and protein immunoprecipitation. Novel autoantigens found on protein immunoprecipitation were further characterized using a proteomic approach, combining immunoprecipitation, SDS-PAGE and MALDI-TOF mass spectrometry. RESULTS: Diagnostic testing on the patient's serum was negative by ELISA and immunodiffusion. Indirect immunofluorescence using Hep-2 cells was ANA negative, although a strong cytoplasmic speckle was seen. Immunoblotting with the patient serum displayed an unknown positive band at approximately 60 kDa. Protein immunoprecipitation revealed the presence of two proteins with molecular weights of approximately 60 and 70 kDa, and RNA immunoprecipitation revealed the presence of a band corresponding to a tRNA synthetase. Using a combination of immunoprecipitation and mass spectrometry, the novel immunoprecipitation targets were identified as phenylalanyl tRNA synthetase alpha and beta chains. CONCLUSIONS: We report the identification of previously uncharacterized autoantibodies to phenylalanyl tRNA synthetase, entitled anti-Zo. This is the eighth anti-synthetase autoantibody in a patient with anti-synthetase syndrome.

Is cerebrospinal fluid spectrophotometry useful in CT scan-negative suspected subarachnoid haemorrage?

Missed cerebral aneurysms in CT-negative patients can have serious implications. We set out to determine the usefulness of cerebrospinal fluid (CSF) spectrophotometry and the individual significance of CSF oxyhaemoglobin, bilirubin and methaemoglobin in 463 CT scan-negative patients with suspected subarachnoid haemorrhage (SAH) and normal neurological examination. CSF spectrophotometry resulted in the diagnosis of an intracranial aneurysm in 2% (9/463) of patients with CT-negative suspected SAH. No aneurysms were found in patients in whom spectrophotometry was negative for haem pigments. Less than 1% of patients with oxyhaemoglobin alone had aneurysms diagnosed, whilst 21% of patients with bilirubin had an aneurysm. CSF spectrophotometry is an important investigation in patients with CT-negative suspected SAH, particularly where clinical suspicion is strong. Patients positive for bilirubin are likely to provide a high yield of aneurysmal bleed and should undergo angiography. Patients with oxyhaemoglobin alone in whom SAH is strongly suspected may benefit from angiography. Based on a small number of patients, we recommend that patients with methaemoglobin should also be investigated. Patients with negative spectrophotometry are unlikely to benefit from further investigation.

Use of the titanium(IV) chelate of salicylic acid for a selective absorptiometric determination of titanium.

Titanium(IV) forms a 1:1 chelate with salicylic acid in 40% aqueous ethanolic solution. The chelate is extractable into chloroform. A selective extraction method for the estimation of titanium in the presence of a variety of commonly interfering species is described. The addition of pyridine is found to increase the sensitivity and selectivity and makes it unnecessary to use alcohol in the extraction. The extracted species is a pyridine adduct of the titanium-salicylic acid chelate.