Menopausal hormone therapy and risk of sarcoidosis: a population-based nested case-control study in Sweden.

Sarcoidosis incidence peaks in women between 50 and 60 years old, which coincides with menopause, suggesting that certain sex hormones, mainly estrogen, may play a role in disease development. We investigated whether menopausal hormone therapy (MHT) was associated with sarcoidosis risk in women and whether the risk varied by treatment type. We performed a nested case-control study (2007-2020) including incident sarcoidosis cases from the Swedish National Patient Register (n = 2593) and matched (1:10) to general population controls (n = 20,003) on birth year, county, and living in Sweden at the time of sarcoidosis diagnosis. Dispensations of MHT were obtained from the Swedish Prescribed Drug Register before sarcoidosis diagnosis/matching. Adjusted odds ratios (aOR) of sarcoidosis were estimated using conditional logistic regression. Ever MHT use was associated with a 25% higher risk of sarcoidosis compared with never use (aOR 1.25, 95% CI 1.13-1.38). When MHT type and route of administration were considered together, systemic estrogen was associated with the highest risk of sarcoidosis (aOR 1.51, 95% CI 1.23-1.85), followed by local estrogen (aOR 1.25, 95% CI 1.11-1.42), while systemic estrogen-progestogen combined was associated with the lowest risk compared to never users (aOR 1.12, 95% CI 0.96-1.31). The aOR of sarcoidosis did not differ greatly by duration of MHT use. Our findings suggest that a history of MHT use is associated with increased risk of sarcoidosis, with women receiving estrogen administered systemically having the highest risk.

From Karl Wurm and Guy Scadding's staging to 18F-FDG PET/CT scan phenotyping and far beyond: perspective in the evading history of phenotyping in sarcoidosis.

Sarcoidosis is an inflammatory granulomatous disease of unknown etiology involving any organ or tissue along with any combination of active sites, even the most silent ones clinically. The unpredictable nature of the sites involved in sarcoidosis dictates the highly variable natural history of the disease and the necessity to cluster cases at diagnosis based on clinical and/or imaging common characteristics in an attempt to classify patients based on their more homogeneous phenotypes, possibly with similar clinical behavior, prognosis, outcome, and therefore with therapeutic requirements. In the course of the disease's history, this attempt relates to the availability of a means of detection of the sites involved, from the Karl Wurm and Guy Scadding's chest x-ray staging through the ACCESS, the WASOG Sarcoidosis Organ Assessment Instruments, and the GenPhenReSa study to the 18F-FDG PET/CT scan phenotyping and far beyond to new technologies and/or the current "omics." The hybrid molecular imaging of the 18F-FDG PET/CT scan, by unveiling the glucose metabolism of inflammatory cells, can identify high sensitivity inflammatory active granulomas, the hallmark of sarcoidosis-even in clinically and physiologically silent sites-and, as recently shown, is successful in identifying an unexpected ordered stratification into four phenotypes: (I) hilar-mediastinal nodal, (II) lungs and hilar-mediastinal nodal, (III) an extended nodal supraclavicular, thoracic, abdominal, inguinal, and (IV) all the above in addition to systemic organs and tissues, which is therefore the ideal phenotyping instrument. During the "omics era," studies could provide significant, distinct, and exclusive insights into sarcoidosis phenotypes linking clinical, laboratory, imaging, and histologic characteristics with molecular signatures. In this context, the personalization of treatment for sarcoidosis patients might have reached its goal.

Modifiable lifestyle risk factors for sarcoidosis: a nested case-control study.

Objective: We aimed to investigate whether obesity, tobacco use, alcohol consumption and physical inactivity are associated with sarcoidosis risk. Methods: We conducted a matched case-control study nested within the Northern Sweden Health and Disease Study. Incident sarcoidosis cases (n=165) were identified via medical records and matched to controls (n=660) on sub-cohort, sex, birth and questionnaire date (1:4). Data on lifestyle factors were obtained through questionnaires, and physical measurements of height, weight and waist were collected prior to sarcoidosis diagnosis. Conditional logistic regression estimated adjusted odds ratios with 95% confidence intervals (aOR; 95% CI). Results: Compared with never-smoking, current smoking was associated with lower sarcoidosis odds (aOR 0.48; 95% CI 0.32-0.71), and former smoking with higher odds (aOR 1.33; 95% CI 0.98-1.81). Snus use was not associated with sarcoidosis. There was an increased odds of sarcoidosis associated with obesity (aOR 1.34; 95% CI 0.94-1.92) but not with overweight (aOR 0.99; 95% CI 0.76-1.30). Compared with those who were physically inactive, those who were active had a 25% higher odds of sarcoidosis (aOR 1.25; 95% CI 0.91-1.72). No association was found with moderate alcohol consumption (aOR 0.95; 95% CI 0.56-1.62). All results were similar when cases diagnosed within 5 years after exposure assessment were excluded, except the aOR for former smoking decreased to 1.1. Conclusion: We observed a reduced sarcoidosis risk associated with smoking, which cannot be fully explained by early symptoms of sarcoidosis influencing smoking habits. Results indicate an increased risk associated with obesity, but not overweight, and being physically active.

Immune mapping of human tuberculosis and sarcoidosis lung granulomas.

Tuberculosis (TB) and sarcoidosis are both granulomatous diseases. Here, we compared the immunological microenvironments of granulomas from TB and sarcoidosis patients using in situ sequencing (ISS) transcriptomic analysis and multiplexed immunolabeling of tissue sections. TB lesions consisted of large necrotic and cellular granulomas, whereas "multifocal" granulomas with macrophages or epitheloid cell core and a T-cell rim were observed in sarcoidosis samples. The necrotic core in TB lesions was surrounded by macrophages and encircled by a dense T-cell layer. Within the T-cell layer, compact B-cell aggregates were observed in most TB samples. These B-cell clusters were vascularized and could contain defined B-/T-cell and macrophage-rich areas. The ISS of 40-60 immune transcripts revealed the enriched expression of transcripts involved in homing or migration to lymph nodes, which formed networks at single-cell distances in lymphoid areas of the TB lesions. Instead, myeloid-annotated regions were enriched in CD68, CD14, ITGAM, ITGAX, and CD4 mRNA. CXCL8 and IL1B mRNA were observed in granulocytic areas in which M. tuberculosis was also detected. In line with ISS data indicating tertiary lymphoid structures, immune labeling of TB sections expressed markers of high endothelial venules, follicular dendritic cells, follicular helper T cells, and lymph-node homing receptors on T cells. Neither ISS nor immunolabeling showed evidence of tertiary lymphoid aggregates in sarcoidosis samples. Together, our finding suggests that despite their heterogeneity, the formation of tertiary immune structures is a common feature in granulomas from TB patients.

Reproductive and hormonal risk factors for sarcoidosis: a nested case-control study.

BACKGROUND: Sarcoidosis incidence peaks in females around the fifth decade of life, which coincides with menopause, suggesting hormonal factors play a role in disease development. We investigated whether longer exposure to reproductive and hormonal factors is associated with reduced sarcoidosis risk. METHODS: We conducted a matched case-control study nested within the Mammography Screening Project. Incident sarcoidosis cases were identified via medical records and matched to controls on birth and questionnaire date (1:4). Information on hormonal factors was obtained through questionnaires prior to sarcoidosis diagnosis. Multilevel modelling was used to estimate adjusted odds ratios with 95% credible intervals (OR; 95% CI). RESULTS: In total, 32 sarcoidosis cases and 124 controls were included. Higher sarcoidosis odds were associated with older age at menarche (OR 1.19: 95% CI 0.92-1.55), natural menopause versus non-natural (OR 1.53: 95% CI 0.80-2.93), later age at first pregnancy (OR 1.11: 95% CI 0.76-1.63) and ever hormone replacement therapy (HRT) use (OR 1.40: 95% CI 0.76-2.59). Lower odds were associated with older age at menopause (OR 0.90: 95% CI 0.52-1.55), longer duration of oral contraceptive use (OR 0.70: 95% CI 0.45-1.07), longer duration of HRT use (OR 0.61: 95% CI 0.22-1.70), ever local estrogen therapy (LET) use (OR 0.83: 95% CI 0.34-2.04) and longer duration of LET use (OR 0.78: 95% CI 0.21-2.81). However, the CIs could not rule out null associations. CONCLUSION: Given the inconsistency and modest magnitude in our estimates, and that the 95% credible intervals included one, it still remains unclear whether longer estrogen exposure is associated with reduced sarcoidosis risk.

Pulmonary and blood dendritic cells from sarcoidosis patients more potently induce IFNγ-producing Th1 cells compared with monocytes.

Sarcoidosis is a systemic inflammatory disease mainly affecting the lungs. The hallmark of sarcoidosis are granulomas that are surrounded by activated T cells, likely targeting the disease-inducing antigen. IFNγ-producing Th1 and Th17.1 T cells are elevated in sarcoidosis and associate with disease progression. Monocytes and dendritic cells (DCs) are antigen-presenting cells (APCs) and required for T cell activation. Several subsets of monocytes and DCs with different functions were identified in sarcoidosis. However, to what extent different monocyte and DC subsets can support activation and skewing of T cells in sarcoidosis is still unclear. In this study, we performed a transcriptional and functional side-by-side comparison of sorted monocytes and DCs from matched blood and bronchoalveolar lavage (BAL) fluid of sarcoidosis patients. Transcriptomic analysis of all subsets showed upregulation of genes related to T cell activation and antigen presentation in DCs compared with monocytes. Allogeneic T cell proliferation was higher after coculture with monocytes and DCs from blood compared with BAL and DCs induced more T cell proliferation compared with monocytes. After coculture, proliferating T cells showed high expression of the transcription factor Tbet and IFNγ production. We also identified Tbet and RORγt coexpressing T cells that mainly produced IFNγ. Our data show that DCs rather than monocytes from sarcoidosis patients have the ability to activate and polarize T cells towards Th1 and Th17.1 cells. This study provides a useful in vitro tool to better understand the contribution of monocytes and DCs to T cell activation and immunopathology in sarcoidosis.

Effects of infliximab on lung and circulating natural killer cells, CD56+ T cells and B cells in sarcoidosis.

BACKGROUND: Tumour necrosis factor α (TNF-α) is pivotal in sarcoid granuloma formation, and inhibitors of TNF-α offer an attractive third-line treatment option in sarcoidosis. The sarcoid inflammation is characterised by an exaggerated T helper 1 response, and evidence indicates a contribution of dysregulated and/or deficient NK (natural killer) cells, CD56+ T cells and B cells. OBJECTIVES: Insight into how TNF-α inhibitors influence these cells may provide more information on inflammatory mechanisms in sarcoidosis and improve understanding of such treatment. We therefore evaluated treatment effects of the TNF-α inhibitor infliximab on lung and peripheral blood (PB) NK, CD56+ T cells and B cells. METHODS: Fifteen patients were assessed with PB samples, spirometry and CT scan, and 11 of them also underwent bronchoalveolar lavage (BAL) close to start of infliximab treatment. These investigations were repeated after 6 months of treatment. RESULTS: Twelve out of 15 patients disclosed a clinical improvement at follow-up. Median percentage of BAL fluid (BALF) CD56+ T cells increased while a decrease was seen in PB (p<0.05 and 0.005, respectively). No significant changes were observed for NK cells. There was a trend towards increased median percentage of PB B cells (p=0.07), and a negative correlation was observed between PB and BALF B cells after treatment (p<0.05). CONCLUSION: In conclusion, 6 months of infliximab treatment in patients with sarcoidosis, of whom the majority benefited from the treatment, influenced immune cells in the lung and circulation differently, highlighting the importance of investigating several compartments concomitantly when evaluating treatment effects on the inflammatory activity.

Bronchoalveolar lavage fluid cell subsets associate with the disease course in Löfgren's and non-Löfgren's sarcoidosis patients.

BACKGROUND: Sarcoidosis is a multisystem granulomatous inflammatory disorder, that predominantly involves the lungs. Patients with Löfgren's syndrome (LS) are characterized by acute onset and usually have the HLA-DRB1*03 (DR3positive) allele and a good prognosis. Non-LS patients are usually DR3negative and are more likely to develop chronic disease. The study aimed to identify bronchoalveolar lavage fluid (BALF) cells that could associate with disease severity (reduced pulmonary function tests (PFTs), advanced chest radiographs, need for treatment) and/or chronicity (duration >2 years) in newly diagnosed LS and non-LS patients, respectively. METHODS: We retrospectively included data from 955 non-LS patients, 477 LS patients, and 295 healthy controls (HC) in this study. Intra-group comparison of patients with resolving versus chronic disease was performed in LS and non-LS, respectively. Non-LS patients were divided into two subgroups according to the binary BALF cell concentrations for intra-group comparison (i.e. higher or lower than the 95th percentile of the BALF cells references in healthy individuals). RESULTS: LS patients with a non-resolving disease course had higher BALF lymphocytes, neutrophils, and eosinophils than LS with a favourable outcome. In non-LS subjects increased BALF of the same cells and in addition also of basophils and mast cells were more likely associated with more severe disease course. CONCLUSION: Increased BALF cells display prognostic significance in sarcoidosis. Certain BALF profiles should promote the clinician to monitor these patients more closely as they may associate non-resolving disease, in turn, resulting in future irreversible functional impairment.

Arrhythmias in Cardiac Sarcoidosis Bench to Bedside: A Case-Based Review.

Cardiac sarcoidosis is a component of an often multiorgan granulomatous disease of still uncertain cause. It is being recognized with increasing frequency, mainly as the result of heightened awareness and new diagnostic tests, specifically cardiac magnetic resonance imaging and 18F-fluorodeoxyglucose positron emission tomography scans. The purpose of this case-based review is to highlight the potentially life-saving importance of making the early diagnosis of cardiac sarcoidosis using these new tools and to provide a framework for the optimal care of patients with this disease. We will review disease mechanisms as currently understood, associated arrhythmias including conduction abnormalities, and atrial and ventricular tachyarrhythmias, guideline-directed diagnostic criteria, screening of patients with extracardiac sarcoidosis, and the use of pacemakers and defibrillators in this setting. Treatment options, including those related to heart failure, and those which may help clarify disease mechanisms are included.

Monocytes in sarcoidosis are potent tumour necrosis factor producers and predict disease outcome.

BACKGROUND: Pulmonary sarcoidosis is an inflammatory disease characterised by granuloma formation and heterogeneous clinical outcome. Tumour necrosis factor (TNF) is a pro-inflammatory cytokine contributing to granuloma formation and high levels of TNF have been shown to associate with progressive disease. Mononuclear phagocytes (MNPs) are potent producers of TNF and highly responsive to inflammation. In sarcoidosis, alveolar macrophages have been well studied. However, MNPs also include monocytes/monocyte-derived cells and dendritic cells, which are poorly studied in sarcoidosis, despite their central role in inflammation. OBJECTIVE: To determine the role of pulmonary monocyte-derived cells and dendritic cells during sarcoidosis. METHODS: We performed in-depth phenotypic, functional and transcriptomic analysis of MNP subsets from blood and bronchoalveolar lavage (BAL) fluid from 108 sarcoidosis patients and 30 healthy controls. We followed the clinical development of patients and assessed how the repertoire and function of MNP subsets at diagnosis correlated with 2-year disease outcome. RESULTS: Monocytes/monocyte-derived cells were increased in blood and BAL of sarcoidosis patients compared to healthy controls. Interestingly, high frequencies of blood intermediate monocytes at time of diagnosis associated with chronic disease development. RNA sequencing analysis showed highly inflammatory MNPs in BAL of sarcoidosis patients. Furthermore, frequencies of BAL monocytes/monocyte-derived cells producing TNF without exogenous stimulation at time of diagnosis increased in patients that were followed longitudinally. In contrast to alveolar macrophages, the frequency of TNF-producing BAL monocytes/monocyte-derived cells at time of diagnosis was highest in sarcoidosis patients that developed progressive disease. CONCLUSION: Our data show that pulmonary monocytes/monocyte-derived cells are highly inflammatory and can be used as a predictor of disease outcome in sarcoidosis patients.

Methylome and transcriptome signature of bronchoalveolar cells from multiple sclerosis patients in relation to smoking.

BACKGROUND: Despite compelling evidence that cigarette smoking impacts the risk of developing multiple sclerosis (MS), little is known about smoking-associated changes in the primary exposed lung cells of patients. OBJECTIVES: We aimed to examine molecular changes occurring in bronchoalveolar lavage (BAL) cells from MS patients in relation to smoking and in comparison to healthy controls (HCs). METHODS: We profiled DNA methylation in BAL cells from female MS (n = 17) and HC (n = 22) individuals, using Illumina Infinium EPIC and performed RNA-sequencing in non-smokers. RESULTS: The most prominent changes were found in relation to smoking, with 1376 CpG sites (adjusted P < 0.05) differing between MS smokers and non-smokers. Approximately 30% of the affected genes overlapped with smoking-associated changes in HC, leading to a strong common smoking signature in both MS and HC after gene ontology analysis. Smoking in MS patients resulted in additional discrete changes related to neuronal processes. Methylome and transcriptome analyses in non-smokers suggest that BAL cells from MS patients display very subtle (not reaching adjusted P < 0.05) but concordant changes in genes connected to reduced transcriptional/translational processes and enhanced cellular motility. CONCLUSIONS: Our study provides insights into the impact of smoking on lung inflammation and immunopathogenesis of MS.

Sarcoidosis exosomes stimulate monocytes to produce pro-inflammatory cytokines and CCL2.

Pulmonary sarcoidosis has unknown etiology, a difficult diagnostic procedure and no curative treatment. Extracellular vesicles including exosomes are nano-sized entities released from all cell types. Previous studies of exosomes from bronchoalveolar lavage fluid (BALF) of sarcoidosis patients have revealed pro-inflammatory components and abilities, but cell sources and mechanisms have not been identified. In the current study, we found that BALF exosomes from sarcoidosis patients, but not from healthy individuals, induced a dose-dependent elevation of intracellular IL-1ß in monocytes. Analyses of supernatants showed that patient exosomes also induced release of IL-1ß, IL-6 and TNF from both PBMCs and enriched monocytes, suggesting that the observed effect is direct on monocytes. The potently chemotactic chemokine CCL2 was induced by exosomes from a subgroup of patients, and in a blocking assay the exosome-induced CCL2 was reduced for 13 out of 19 patients by the asthma drug Montelukast, a cysteinyl leukotriene receptor antagonist. Further, reactive oxygen species generation by PBMCs was induced to a higher degree by patient exosomes compared to healthy exosomes. These findings add to an emerging picture of exosomes as mediators and disseminators of inflammation, and open for further investigations of the link between CCL2 and exosomal leukotrienes in sarcoidosis.

Maternal and infant outcomes in sarcoidosis pregnancy: a Swedish population-based cohort study of first births.

BACKGROUND: It is unclear whether sarcoidosis, a multisystem inflammatory disease, is associated with adverse pregnancy outcomes. We aimed to assess the risk of adverse maternal and infant outcomes in sarcoidosis pregnancies, focused on first births. METHODS: Using a population-based cohort study design and Swedish national registers (2002-2013), we identified 182 singleton first pregnancies in the Medical Birth Register with at least two maternal ICD-coded sarcoidosis visits prior to pregnancy in the National Patient Register. Modified Poisson regression models estimated relative risks (RR) of adverse outcomes in sarcoidosis pregnancies compared to the general population adjusted for maternal age at delivery, calendar year and educational level. Some models were additionally adjusted for maternal body mass index and smoking status. RESULTS: The prevalence of pre-existing diabetes and hypertension was higher in mothers with sarcoidosis than those without sarcoidosis. Mothers with sarcoidosis had an increased risk of preeclampsia/eclampsia (RR 1.6; 95%CI 1.0, 2.6) and cesarean delivery (RR 1.3; 95%CI 1.0, 1.6). There were < 5 stillbirths and cases of infection and no cases of placental abruption, venous thromboembolism, cardiac arrest or maternal death. Newborns of first-time mothers with sarcoidosis had a 70% increased risk of preterm birth (RR 1.7; 95%CI 1.1, 2.5). There was an increased risk of birth defects (RR 1.6; 95%CI 0.9, 2.8) the majority of which were non-cardiac. CONCLUSIONS: Sarcoidosis is associated with increased risks for preeclampsia/eclampsia, cesarean delivery, preterm birth and some birth defects. Awareness of these conditions may prevent possible pregnancy complications in mothers with sarcoidosis and their newborns.

Looking into the future of sarcoidosis: what is next for treatment?

PURPOSE OF REVIEW: Sarcoidosis is a complex granulomatous disease of unknown cause. Corticosteroids and immune suppressants are often given long term in chronic disease, which may result in substantial toxicity. Validated strategies for selecting patients at risk for disease progression, who might benefit from early and targeted treatment, are lacking. Consequently, the unmet need for new treatment options in sarcoidosis is high. In this review, we critically discuss potential therapeutic targets and ongoing clinical trials in sarcoidosis. RECENT FINDINGS: Despite the heterogeneous clinical manifestations and the lack of a reliable animal model, our knowledge and understanding of the pathogenesis of sarcoidosis has improved in recent years, which has resulted in the identification of several potential therapeutic strategies. They include the inhibition of cytokines involved in maturation of macrophages, activation of dendritic cells, and maturation and activation of pathogenic T-lymphocytes. The inflammasome and the autophagy are additional areas for future research. Antifibrotic therapy might also be a reasonable choice in selected patients, although the best treatment strategy in progressive fibrotic sarcoidosis remains undetermined. SUMMARY: In this article, we review novel approaches to sarcoidosis treatment and potential therapeutic targets.

Lung CD4+ Vα2.3+ T-cells in sarcoidosis cohorts with Löfgren's syndrome.

BACKGROUND: Sarcoidosis is diagnosed by a combination of typical clinical and radiological findings together with biopsy proof of non-caseating epithelioid cell granulomas in affected tissues and/or the cell distribution in bronchoalveolar lavage fluid (BALF). We aimed at investigating the usefulness of measuring the proportion of T-cell receptor (TCR) CD4+ Vα2.3+ T-cells in BALF as an additive marker to CD4/CD8-ratio to confirm the diagnosis. METHODS: From a register consisting of 749 sarcoidosis patients [Löfgren's syndrome (LS) n = 274, non-LS n = 475] with information on Vα2.3+ T-cells, an expansion of CD4+ Vα2.3+ T-cells (CD4+ Vα2.3+ T cells > 10.5% in BALF) was seen in 268 (36%). Controls were healthy volunteers (n = 69) and patients with other pulmonary conditions (n = 39), investigated because of suspicion of sarcoidosis. RESULTS: A proportion of CD4+ Vα2.3+ T-cells in BALF > 10.5% was highly specific for sarcoidosis, with a specificity of 97% and with a sensitivity of 36% (p < 0.0001). Receiver operating characteristic (ROC) curves show that testing for CD4+ Vα2.3+ T-cells in BALF was a more useable test in individuals with LS [area under the curve (AUC) 0.82, p < 0.0001] compared to the whole patient group (AUC 0.64, p < 0.0001). CONCLUSION: In this study, we show that an increased proportion of CD4+ Vα2.3+ T-cells in BALF is highly specific for sarcoidosis. This suggests that this T-cell subset could be used as an additional tool to the CD4/CD8-ratio to support the sarcoidosis diagnosis, particularly in patients with LS but also in patients with non-LS.

Proinflammatory Histidyl-Transfer RNA Synthetase-Specific CD4+ T Cells in the Blood and Lungs of Patients With Idiopathic Inflammatory Myopathies.

OBJECTIVE: Autoantibodies targeting histidyl-transfer RNA synthetase (HisRS; anti-Jo-1) are common in the idiopathic inflammatory myopathies (IIMs) and antisynthetase syndrome. This study was undertaken to investigate immunity against HisRS in the blood and lungs of patients with IIM/antisynthetase syndrome. METHODS: Bronchoalveolar lavage (BAL) fluid, BAL fluid cells, and peripheral blood mononuclear cells (PBMCs) from patients with IIM/antisynthetase syndrome (n = 24) were stimulated with full-length HisRS protein or a HisRS-derived peptide (HisRS11-23 ). BAL fluid and PBMCs from patients with sarcoidosis (n = 7) and healthy subjects (n = 12) were included as controls. The CD4+ T cell response was determined according to levels of CD40L up-regulation and cytokine expression using flow cytometry. Anti-Jo-1 autoantibody responses in the serum and BAL fluid were assessed by enzyme-linked immunosorbent assay. Lung biopsy samples from patients with IIM/antisynthetase syndrome (n = 14) were investigated by immunohistochemistry. RESULTS: In BAL fluid, CD4+ T cells from 3 of 4 patients with IIM/antisynthetase syndrome responded to stimulation with HisRS protein, as measured by the median fold change in CD40L expresssion in stimulated cells compared to unstimulated cells (median fold change 3.6, interquartile range [IQR] 2.7-14.7), and 2 of 3 patients with IIM/antisynthetase syndrome had the highest responses to HisRS11-23 (median fold change 88, IQR 27-149). In PBMCs, CD4+ T cells from 14 of 18 patients with IIM/antisynthetase syndrome responded to HisRS protein (median fold change 7.38, IQR 2.69-31.86; P < 0.001), whereas a HisRS11-23 response was present in 11 of 14 patients with IIM/antisynthetase syndrome (median fold change 3.4, IQR 1.87-10.9; P < 0.001). In the control group, there was a HisRS11-23 response in 3 of 7 patients with sarcoidosis (median fold change 2.09, IQR 1.45-3.29) and in 5 of 12 healthy controls (median fold change 2, IQR 1.89-2.42). CD4+ T cells from patients with IIM/antisynthetase syndrome displayed a pronounced Th1 phenotype in the BAL fluid when compared to the PBMCs (P < 0.001), producing high amounts of interferon-γ and interleukin-2 following stimulation. Anti-Jo-1 autoantibodies were detected in BAL fluid and germinal center (GC)-like structures were seen in the lung biopsy samples from patients with IIM/antisynthetase syndrome. CONCLUSION: The results of this study demonstrate a pronounced presence of HisRS-reactive CD4+ T cells in PBMCs and BAL fluid cells from patients with IIM/antisynthetase syndrome as compared to patients with sarcoidosis and healthy controls. These findings, combined with the presence of anti-Jo-1 autoantibodies in BAL fluid and GC-like structures in the lungs, suggest that immune activation against HisRS might take place within the lungs of patients with IIM/antisynthetase syndrome.

A Gene-Environment Interaction Between Smoking and Gene polymorphisms Provides a High Risk of Two Subgroups of Sarcoidosis.

The influence and effect of cigarette smoking in sarcoidosis is unclear. Here, we evaluated gene-environment interaction between multiple genetic variants including HLA genes and smoking in sarcoidosis defined by two clinical phenotypes, Löfgren's syndrome (LS) and patients without Löfgren's syndrome (non-LS). To quantify smoking effects in sarcoidosis, we performed a gene-environment interaction study in a Swedish population-based case-control study consisting of 3,713 individuals. Cases and controls were classified according to their cigarette smoking status and genotypes by Immunochip platform. Gene-smoking interactions were quantified by an additive interaction model using a logistic regression adjusted by sex, age and first two principal components. The estimated attributable proportion (AP) was used to quantify the interaction effect. Assessment of smoking effects with inclusion of genetic information revealed 53 (in LS) and 34 (in non-LS) SNP-smoking additive interactions at false discovery rate (FDR) below 5%. The lead signals interacting with smoking were rs12132140 (AP = 0.56, 95% CI = 0.22-0.90), p = 1.28e-03) in FCRL1 for LS and rs61780312 (AP = 0.62, 95% CI = 0.28-0.90), p = 3e-04) in IL23R for non-LS. We further identified 16 genomic loci (in LS) and 13 (in non-LS) that interact with cigarette smoking. These findings suggest that sarcoidosis risk is modulated by smoking due to genetic susceptibility. Therefore, patients having certain gene variants, are at a higher risk for the disease. Consideration of individual's genetic predisposition is crucial to quantify effects of smoking in sarcoidosis.

Sarcoidosis.

Sarcoidosis is an inflammatory disorder of unknown cause that is characterized by granuloma formation in affected organs, most often in the lungs. Patients frequently suffer from cough, shortness of breath, chest pain and pronounced fatigue and are at risk of developing lung fibrosis or irreversible damage to other organs. The disease develops in genetically predisposed individuals with exposure to an as-yet unknown antigen. Genetic factors affect not only the risk of developing sarcoidosis but also the disease course, which is highly variable and difficult to predict. The typical T cell accumulation, local T cell immune response and granuloma formation in the lungs indicate that the inflammatory response in sarcoidosis is induced by specific antigens, possibly including self-antigens, which is consistent with an autoimmune involvement. Diagnosis can be challenging for clinicians because of the potential for almost any organ to be affected. As the aetiology of sarcoidosis is unknown, no specific treatment and no pathognomic markers exist. Thus, improved biomarkers to determine disease activity and to identify patients at risk of developing fibrosis are needed. Corticosteroids still constitute the first-line treatment, but new treatment strategies, including those targeting quality-of-life issues, are being evaluated and should yield appropriate, personalized and more effective treatments.

Tobacco smoking induces changes in true DNA methylation, hydroxymethylation and gene expression in bronchoalveolar lavage cells.

BACKGROUND: While smoking is known to associate with development of multiple diseases, the underlying mechanisms are still poorly understood. Tobacco smoking can modify the chemical integrity of DNA leading to changes in transcriptional activity, partly through an altered epigenetic state. We aimed to investigate the impact of smoking on lung cells collected from bronchoalveolar lavage (BAL). METHODS: We profiled changes in DNA methylation (5mC) and its oxidised form hydroxymethylation (5hmC) using conventional bisulphite (BS) treatment and oxidative bisulphite treatment with Illumina Infinium MethylationEPIC BeadChip, and examined gene expression by RNA-seq in healthy smokers. FINDINGS: We identified 1667 total 5mC + 5hmC, 1756 5mC and 67 5hmC differentially methylated positions (DMPs) between smokers and non-smokers (FDR-adjusted P <.05, absolute Δß >0.15). Both 5mC DMPs and to a lesser extent 5mC + 5hmC were predominantly hypomethylated. In contrast, almost all 5hmC DMPs were hypermethylated, supporting the hypothesis that smoking-associated oxidative stress can lead to DNA demethylation, via the established sequential oxidation of which 5hmC is the first step. While we confirmed differential methylation of previously reported smoking-associated 5mC + 5hmC CpGs using former generations of BeadChips in alveolar macrophages, the large majority of identified DMPs, 5mC + 5hmC (1639/1667), 5mC (1738/1756), and 5hmC (67/67), have not been previously reported. Most of these novel smoking-associating sites are specific to the EPIC BeadChip and, interestingly, many of them are associated to FANTOM5 enhancers. Transcriptional changes affecting 633 transcripts were consistent with DNA methylation profiles and converged to alteration of genes involved in migration, signalling and inflammatory response of immune cells. INTERPRETATION: Collectively, these findings suggest that tobacco smoke exposure epigenetically modifies BAL cells, possibly involving a continuous active demethylation and subsequent increased activity of inflammatory processes in the lungs. FUND: The study was supported by the Swedish Research Council, the Swedish Heart-Lung Foundation, the Stockholm County Council (ALF), the King Gustav's and Queen Victoria's Freemasons' Foundation, Knut and Alice Wallenberg Foundation, Neuro Sweden, and the Swedish MS foundation.