Phenotypes of organ involvement in sarcoidosis.

Sarcoidosis is a highly variable, systemic granulomatous disease of hitherto unknown aetiology. The GenPhenReSa (Genotype-Phenotype Relationship in Sarcoidosis) project represents a European multicentre study to investigate the influence of genotype on disease phenotypes in sarcoidosis.The baseline phenotype module of GenPhenReSa comprised 2163 Caucasian patients with sarcoidosis who were phenotyped at 31 study centres according to a standardised protocol.From this module, we found that patients with acute onset were mainly female, young and of Scadding type I or II. Female patients showed a significantly higher frequency of eye and skin involvement, and complained more of fatigue. Based on multidimensional correspondence analysis and subsequent cluster analysis, patients could be clearly stratified into five distinct, yet undescribed, subgroups according to predominant organ involvement: 1) abdominal organ involvement, 2) ocular-cardiac-cutaneous-central nervous system disease involvement, 3) musculoskeletal-cutaneous involvement, 4) pulmonary and intrathoracic lymph node involvement, and 5) extrapulmonary involvement.These five new clinical phenotypes will be useful to recruit homogenous cohorts in future biomedical studies.

Identification of Immune-Relevant Factors Conferring Sarcoidosis Genetic Risk.

RATIONALE: Genetic variation plays a significant role in the etiology of sarcoidosis. However, only a small fraction of its heritability has been explained so far. OBJECTIVES: To define further genetic risk loci for sarcoidosis, we used the Immunochip for a candidate gene association study of immune-associated loci. METHODS: Altogether the study population comprised over 19,000 individuals. In a two-stage design, 1,726 German sarcoidosis cases and 5,482 control subjects were genotyped for 128,705 single-nucleotide polymorphisms using the Illumina Immunochip for the screening step. The remaining 3,955 cases, 7,514 control subjects, and 684 parents of affected offspring were used for validation and replication of 44 candidate and two established risk single-nucleotide polymorphisms. MEASUREMENTS AND MAIN RESULTS: Four novel susceptibility loci were identified with genome-wide significance in the European case-control populations, located on chromosomes 12q24.12 (rs653178; ATXN2/SH2B3), 5q33.3 (rs4921492; IL12B), 4q24 (rs223498; MANBA/NFKB1), and 2q33.2 (rs6748088; FAM117B). We further defined three independent association signals in the HLA region with genome-wide significance, peaking in the BTNL2 promoter region (rs5007259), at HLA-B (rs4143332/HLA-B*0801) and at HLA-DPB1 (rs9277542), and found another novel independent signal near IL23R (rs12069782) on chromosome 1p31.3. CONCLUSIONS: Functional predictions and protein network analyses suggest a prominent role of the drug-targetable IL23/Th17 signaling pathway in the genetic etiology of sarcoidosis. Our findings reveal a substantial genetic overlap of sarcoidosis with diverse immune-mediated inflammatory disorders, which could be of relevance for the clinical application of modern therapeutics.

Genome-wide association analysis reveals 12q13.3-q14.1 as new risk locus for sarcoidosis.

Sarcoidosis is a systemic inflammatory disease of unknown aetiology, influenced by genetic and environmental factors. However, the loci so far identified for sarcoidosis explain only a part of its assumed heritability. To identify further susceptibility loci, we performed a genome-wide association analysis using the Affymetrix 6.0 Human GeneChip followed by validation and replication stages. After quality control, 637 cases, 1233 controls and 677 619 single-nucleotide polymorphisms (SNPs) were available for an initial screening. 99 SNPs were selected for validation in an independent study panel (1664 patients, 2932 controls). SNP rs1050045 was significantly associated with sarcoidosis (corrected p=0.0215) in the validation panel and yielded a p-value of 9.22 × 10(-8) (OR 1.24) in the meta-analysis of the screening and validation stage. A meta-analysis of three populations from Germany, the Czech Republic and Sweden confirmed this finding (p = 0.024; OR 1.14). Fine-mapping and mRNA expression studies pointed to osteosarcoma amplified 9 (OS9) as the most likely candidate for the underlying risk factor. The OS9 protein plays an important role in endoplasmic reticulum-associated protein degradation and acts during Toll-like receptor induced activation of myeloid cells. Expression analyses of OS9 mRNA provide evidence for a functional mechanism underlying the detected association signal.

Strong-field many-body physics and the giant enhancement in the high-harmonic spectrum of xenon.

We resolve an open question about the origin of the giant enhancement in the high-harmonic generation spectrum of atomic xenon around 100 eV. By solving the many-body time-dependent Schrödinger equation with all 4d, 5s, and 5p orbitals active, we truly demonstrate the enhancement results from the collective many-body excitation induced by the returning photoelectron via two-body interchannel interactions. Without the many-body interactions, which promote a 4d electron into the 5p vacancy created by strong-field ionization, no collective excitation and no enhancement in the high-harmonic generation spectrum exist.

A novel sarcoidosis risk locus for Europeans on chromosome 11q13.1.

RATIONALE: Sarcoidosis is a complex inflammatory disease with a heterogeneous clinical picture. Among others, an acute and chronic clinical course can be distinguished, for which specific genetic risk factors are known. OBJECTIVES: To identify additional risk loci for sarcoidosis and its acute and chronic subforms, we analyzed imputed data from a genome-wide association scan for these phenotypes. METHODS: After quality control, the genome-wide association scan comprised nearly 1.3 million imputed single-nucleotide polymorphisms based on an Affymetrix 6.0 Gene Chip dataset of 564 German sarcoidosis cases, including 176 acute and 354 chronic cases and 1,575 control subjects. MEASUREMENTS AND MAIN RESULTS: We identified chromosome 11q13.1 (rs479777) as a novel locus influencing susceptibility to sarcoidosis with genome-wide significance. The marker was significantly associated in three distinct German case-control populations and in an additional German family sample with odds ratios ranging from 0.67 to 0.77. This finding was further replicated in two independent European case-control populations from the Czech Republic (odds ratio, 0.75) and from Sweden (odds ratio, 0.79). In a meta-analysis of the included European case-control samples the marker yielded a P value of 2.68 × 10(-18). The locus was previously reported to be associated with Crohn disease, psoriasis, alopecia areata, and leprosy. For sarcoidosis, fine-mapping and expression analysis suggest KCNK4, PRDX5, PCLB3, and most promising CCDC88B as candidates for the underlying risk gene in the associated region. CONCLUSIONS: This study provides striking evidence for association of chromosome 11q13.1 with sarcoidosis in Europeans, and thus identified a further genetic risk locus shared by sarcoidosis, Crohn disease and psoriasis.

Pulmonary aspergillosis: therapeutic management and prognostic factors from 16 years of monocenter experience.

Twenty three patients of the University Hospital Bonn were reviewed following surgical procedures for pulmonary aspergilloma, including the choice of antifungal therapy, diagnostic findings, decision-making in treatment, and treatment outcomes of the past 16 years. We used pathological records to identify aspergilloma patients. A review of patients' records and follow-up phone calls to patients, families, or general practitioners were done. Data collected from 1995 to 2011 included patients with aspergilloma (n = 15), multiple aspergillomas (n = 2) and chronic necrotizing pulmonary aspergillosis (n = 6). Classification and diagnosis were based on pathological records. The decision to use systemic antimycotic therapy was based on perioperative findings suspecting parenchymal involvement of the fungal infection. Seventeen patients received systemic antimycotic chemotherapy. Compared with the use of Amphotericin B, newer drugs such as voriconazol, caspofungin, or posaconazol showed no better result in the morbidity and mortality of the patients. Postoperative complications requiring extended therapy and/or prolonged ICU stay (>48 h) were seen in 12 (52.2%) patients. During follow-up there were ten deaths; one death (4.4%) from aspergillus-associated sepsis, nine deaths from patients' underlying diseases (n = 4 within <3 months, n = 6 within >3 months of follow-up). In conclusion, in our cohort, immunocompromised patients with no documented preexisting lung-cavities were most likely to develop pulmonary aspergilloma. Postoperative morbidity (52.2%) was high, but related mainly to patient co-morbidity; postoperative mortality was reasonably low. Patients showing classical symptoms or immunocompromised patients should be considered for surgery. Encapsulated Aspergilloma without invasion of surrounding parenchyma requires no antifungal chemotherapy.

Toll-like receptor-9 polymorphisms in sarcoidosis and chronic obstructive pulmonary disease.

The etiology of inflammatory diseases of the lung like sarcoidosis and chronic obstructive pulmonary disease (COPD) is multifactorial. The main trigger for developing a COPD is tobacco smoking while exogenous factors causing sarcoidosis are unclear. In both diseases there is an underlying genetic susceptibility determining both the onset and the course of the diseases. Toll-like receptor (TLR)-9 plays an important role in innate immunity by recognizing bacterial CpG-DNA motifs. It is unclear whether single nucleotide polymorphisms (SNPs) in TLR-9 are able to alter the course of sarcoidosis or COPD, or raise the susceptibility for developing one of the disorders. We examined two SNPs in the promoter region of the TLR-9 gene (T1486C and T1237C) in 175 COPD patients (59% with a stable course of the disease, 41% with an instable course with more than 3 exacerbations over the last 3 years) and 166 sarcoidosis patients (19% with an acute and 81% with a chronic course of the disease lasting >2 years) comparing each group to 233 healthy controls. Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis was used for genotyping. The C-allele frequency of T1486C was significantly elevated in COPD patients (p = 0.008). For T1237C there were no significant associations comparing the COPD cohort with the controls. In the sarcoidosis cohort, we could observe a significantly higher prevalence of the C-allele for T1237C in the chronic sarcoidosis cohort in comparison to the control group (p = 0.026). For T1486 there no statistical association was observed. This is the first study showing an association between a SNP (T1486C) in the TLR-9 gene and the onset of COPD. Moreover, we could demonstrate that T1237C is able to alter the course of sarcoidosis as a disease-modifying gene. This study underlines that SNPs in TLR-9 might be involved in acquiring and maintaining lung diseases such as sarcoidosis and COPD.

Pulmonary arterial hypertension in patients with sarcoidosis: the Pulsar single center experience.

Sarcoidosis is a systemic granulomatous disease with unknown etiology. Lungs and lymph nodes are commonly affected. Also, cases of pulmonary hypertension (PH) and pulmonary arterial hypertension (PAH) are described. However, the exact prevalence of PAH in patients with sarcoidosis is unclear. A 111 patients with proven sarcoidosis were recruited from January 2010 to October 2010. All patients were studied prospectively by transthoracic echocardiography (TTE) for the presence of PH. In assumed PH, a right heart catheterization (RHC) followed if there were no other reasons for PH. In 23 of the 111 patients (21%) PH was assumed in TTE. Three patients presented with severe mitral insufficiency III° and IV°, in eight patients PH was supposed to be caused by chronic heart failure or relevant diastolic dysfunction > II°, two patients declined undergoing RHC. Of the ten patients investigated with RHC, four showed a precapillary pulmonary arterial hypertension and in one patient a postcapillary hypertension was diagnosed. All four patients with precapillary PH had a radiologic stage III and IV. In three of the four patients a significantly reduced transfer factor for carbon monoxide (TLCO) <50% was found. All patients with precapillary PH had a chronic course of sarcoidosis lasting ≥13 years. This is the first study which prospectively investigated a large cohort of patients with sarcoidosis for the prevalence of PH and PAH. The prevalence of precapillary PH was found to be at least 3.6% (4/111) and therefore exceeds the prevalence of PAH in the normal population by far. A chronic and progressive lung involvement due to sarcoidosis seems to be the most evident risk factor for developing a sarcoidosis PH.

Genetic and geographic influence on phenotypic variation in European sarcoidosis patients.

Introduction: Sarcoidosis is a highly variable disease in terms of organ involvement, type of onset and course. Associations of genetic polymorphisms with sarcoidosis phenotypes have been observed and suggest genetic signatures. Methods: After obtaining a positive vote of the competent ethics committee we genotyped 1909 patients of the deeply phenotyped Genetic-Phenotype Relationship in Sarcoidosis (GenPhenReSa) cohort of 31 European centers in 12 countries with 116 potentially disease-relevant single-nucleotide polymorphisms (SNPs). Using a meta-analysis, we investigated the association of relevant phenotypes (acute vs. sub-acute onset, phenotypes of organ involvement, specific organ involvements, and specific symptoms) with genetic markers. Subgroups were built on the basis of geographical, clinical and hospital provision considerations. Results: In the meta-analysis of the full cohort, there was no significant genetic association with any considered phenotype after correcting for multiple testing. In the largest sub-cohort (Serbia), we confirmed the known association of acute onset with TNF and reported a new association of acute onset an HLA polymorphism. Multi-locus models with sets of three SNPs in different genes showed strong associations with the acute onset phenotype in Serbia and Lublin (Poland) demonstrating potential region-specific genetic links with clinical features, including recently described phenotypes of organ involvement. Discussion: The observed associations between genetic variants and sarcoidosis phenotypes in subgroups suggest that gene-environment-interactions may influence the clinical phenotype. In addition, we show that two different sets of genetic variants are permissive for the same phenotype of acute disease only in two geographic subcohorts pointing to interactions of genetic signatures with different local environmental factors. Our results represent an important step towards understanding the genetic architecture of sarcoidosis.

Regulatory T cells with reduced repressor capacities are extensively amplified in pulmonary sarcoid lesions and sustain granuloma formation.

Sarcoidosis can evolve into a chronic disease with persistent granulomas accompanied by progressive fibrosis. While an unlimited inflammatory response suggests an impaired immune control in sarcoid lesions, it stands in contrast to the massive infiltration with CD4(+)CD25(high)FoxP3(+) regulatory T cells. We here revealed that those Treg cells in affected lung lesions were mainly derived from activated natural Treg cells with GARP (LRRC32)-positive phenotype but exhibited reduced repressor capacities despite high IL-10 and TGF-beta 1 levels. The repressive capacity of blood Treg cells, in contrast, was not impaired compared to age-matched healthy donors. Treg derived cells in granuloma lesions have undergone extensive rounds of amplifications indicated by shortened telomeres compared to blood Treg cells of the same patient. Lesional Treg derived cells moreover secreted pro-inflammatory cytokines including IL-4 which sustains granuloma formation through fibroblast amplification and the activation of mast cells, the latter indicated by the expression of membrane-bound oncostatin M.

Decoherence in attosecond photoionization.

The creation of superpositions of hole states via single-photon ionization using attosecond extreme-ultraviolet pulses is studied with the time-dependent configuration-interaction singles (TDCIS) method. Specifically, the degree of coherence between hole states in atomic xenon is investigated. We find that interchannel coupling not only affects the hole populations, but it also enhances the entanglement between the photoelectron and the remaining ion, thereby reducing the coherence within the ion. As a consequence, even if the spectral bandwidth of the ionizing pulse exceeds the energy splittings among the hole states involved, perfectly coherent hole wave packets cannot be formed. For sufficiently large spectral bandwidth, the coherence can only be increased by increasing the mean photon energy.

Health and functional status of tiotropium/olodaterol-treated patients with COPD: results from the AERIAL® non-interventional study.

Patients with COPD often have reduced physical activity, which can impair health status. Real-world data can provide valuable information on the health and functional status of patients with COPD treated with tiotropium/olodaterol. AERIAL® (ClinicalTrials.gov NCT03165045) was a German, non-interventional study of patients with COPD receiving treatment with tiotropium/olodaterol under real-world conditions for ∼6 weeks. The primary end-point was the proportion of patients achieving a decrease of ≥0.4 points in Clinical COPD Questionnaire (CCQ) score. The CCQ-4 subdomain was used to assess functional status, and the Physician's Global Evaluation (PGE) scale was used to assess the patients' general condition. Safety was assessed, as well as patient satisfaction and willingness to continue treatment. Out of 1351 screened patients, 1322 were treated and 1140 comprised the full analysis set. The primary end-point was met: 66.3% of patients achieved a ≥0.4-point decrease in overall CCQ score (mean±sd decrease 0.78±0.95). Mean±sd decreases in CCQ symptoms and functional state subdomains were 0.84±1.06 and 0.75±1.05 points, respectively. PGE scores improved. One fatality (not treatment-related) and 23 drug-related adverse events were recorded, most commonly nausea and vertigo. >85% of patients were satisfied/very satisfied with tiotropium/olodaterol overall and with the Respimat® device, both in terms of inhalation and handling. Most patients (95.2%) expressed willingness to continue treatment. Patients with COPD treated with tiotropium/olodaterol via Respimat® in routine clinical practice had clinically relevant improvements in health and functional status compared with baseline.

Pathogen burden in degenerative aortic valves is associated with inflammatory and immune reactions.

BACKGROUND AND AIM OF THE STUDY: The presence of five pathogens was assessed, together with a possible correlation of the total pathogen burden on inflammation and (auto)immunity in aortic stenosis (AS) and degenerative aortic valve bioprosthesis (BP). METHODS: Diseased valve specimens from a total of 68 patients (52 with AS, 16 with BP) were studied. The presence and localization was assessed of Chlamydia pneumoniae (cHSP60), Helicobacter pylori (HP), cytomegalovirus (CMV), Epstein-Barr virus (EBV) and herpes simplex virus (HSV), as well as of macrophages (CD68), C-reactive protein (CRP) and human heat shock protein 60 (hHSP60), by using immunohistochemical and morphometric analyses. RESULTS: In the majority of degenerative aortic valves, specific pathogens, inflammation and immunity were localized predominantly in the fibrosa of AS patients, and in superficial regions of the BP. The categorization of valves as having four or more pathogens (n = 37) or fewer pathogens (n = 31) demonstrated an increased signaling of CD68 (p = 0.03) and CRP (p = 0.02). Specifically, cHSP60, HP and hHSP60 levels were increased in valves where one or two bacteria were identified (n = 59) compared to those without bacterial presence (n = 9) (p = 0.04). CONCLUSION: The pathogen burden may contribute to valvular degeneration by promoting further deleterious inflammatory and (auto)immune processes at the level of the valvular fibrosa.

Toll-like receptor 2 gene polymorphisms Arg677Trp and Arg753Gln in chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a leading cause of death worldwide, with a continually rising mortality rate. As COPD is driven by abnormal pulmonary and systemic inflammation, Toll-like receptors (TLRs) seem to be important. TLRs play a key role in innate response, and in particular TLR2 gene polymorphisms Arg677Trp and Arg753Gln have been linked to an increased risk of infection. The purpose of this study was to investigate whether there is a link between polymorphisms in TLR2 and the onset or course of COPD. We analyzed 149 Caucasian COPD patients and 150 healthy individuals by using polymerase chain reaction (PCR) followed by restriction fragment length polymorphism (RFLP) analysis. To further characterize the disease, patients were classified according to GOLD and divided into two subgroups comprising a stable (60/149) course and an unstable (89/149) course. The TLR2 Arg677Trp mutant allele was not found in any of the subjects. With a prevalence of 8.72% (13/149) for TLR2 Arg753Gln, the patients did not differ from the controls, with a prevalence of 10.67% (16/150). No significant difference was apparent (P = 0.571). None of the individuals showed homozygosity for TLR2 Arg753Gln. With regard to the course of COPD, the prevalence of TLR2 Arg753Gln in the control group did not differ significantly either from the stable subgroup (P = 0.196) or from the unstable subgroup (P = 0.891). Our results suggest that there is no association of the TLR2 polymorphisms Arg677Trp and Arg753Gln with either the onset or the course of COPD.

Methods to study SUMO dynamics in yeast.

Covalent modification of proteins with the small ubiquitin-related modifier (SUMO) is found in all eukaryotes and is involved in many important processes. SUMO attachment may change interaction properties, subcellular localization, or stability of a modified protein. Usually, only a small fraction of a protein is modified at a given time because sumoylation is a highly dynamic process. The sumoylated state of a protein is controlled by the activity of the sumoylation enzymes that promote either their mono- or poly-sumoylation (SUMO chain formation), by SUMO proteases that reverse these modifications, and by SUMO-targeted ubiquitin ligases (STUbL, ULS) that mediate their degradation by the proteasome. While some organisms, such as humans, express multiple isoforms, budding yeast SUMO is encoded by a single and essential gene termed SMT3. The analysis of the simpler SUMO system in budding yeast has been instrumental in the identification of enzymes acting on this modification and controlling its dynamics. Sumoylation of proteins changes dramatically during the cell division cycle and under various stress conditions. Here we summarize various approaches that employ Saccharomyces cerevisiae as a model system to study the dynamics of sumoylation and how it is controlled.

Assessment of physical functioning and handling of tiotropium/olodaterol Respimat® in patients with COPD in a real-world clinical setting.

Background: Patients with chronic obstructive pulmonary disease (COPD) show signs of reduced physical activity from the early stages of the disease, impacting morbidity and mortality. Data suggest treatment with tiotropium, a long-acting muscarinic antagonist, and olodaterol, a long-acting ß2-agonist (LABA), as monotherapies and in combination, increases exercise capacity. This study assessed the effects of fixed-dose tiotropium/olodaterol (delivered via Respimat®) on physical function in Global Initiative for Chronic Obstructive Lung Disease A-D patients requiring long-acting dual bronchodilation treatment in a real-world setting. Methods: This open-label, single arm, noninterventional study measured changes in physical function in COPD patients treated with tiotropium/olodaterol 5/5 µg for approximately 6 weeks (between Visit 1 [baseline] and Visit 2). Primary end point was therapeutic success, defined as a minimum 10-point increase in Physical Functioning Questionnaire (PF-10) score. Secondary end points included change in PF-10 from Visit 1 to Visit 2, the patient's general condition (measured by Physician's Global Evaluation score) at Visit 1 and Visit 2, and patient satisfaction with treatment delivered via the Respimat® device (assessed by Patient Satisfaction Questionnaire) at study end. Results: Therapeutic success was observed in 51.5% of 1578 patients (95% confidence interval [CI] 49.0, 54.0) after approximately 6 weeks of treatment with tiotropium/olodaterol. Mean change in PF-10 score between Visit 1 and Visit 2 was 11.6 points (95% CI 10.7, 12.6). Patient general condition improved as indicated by a general improvement in scores between visits. Most patients were very satisfied or satisfied with tiotropium/olodaterol treatment (82.5%), inhalation (87.5%), and handling of Respimat® (85.2%). One percent of patients reported an investigator-defined drug-related adverse events (AE). Conclusion: Tiotropium/olodaterol treatment improved physical functioning in COPD patients. An associated increase in patient general condition was observed. Most patients were very satisfied or satisfied with tiotropium/olodaterol treatment, inhaling, and handling of the Respimat® device. No unexpected drug-related AE occurred.

Arkadia/RNF111 is a SUMO-targeted ubiquitin ligase with preference for substrates marked with SUMO1-capped SUMO2/3 chain.

Modification with SUMO regulates many eukaryotic proteins. Down-regulation of sumoylated forms of proteins involves either their desumoylation, and hence recycling of the unmodified form, or their proteolytic targeting by ubiquitin ligases that recognize their SUMO modification (termed STUbL or ULS). STUbL enzymes such as Uls1 and Slx5-Slx8 in budding yeast or RNF4 and Arkadia/RNF111 in humans bear multiple SUMO interaction motifs to recognize substrates carrying poly-SUMO chains. Using yeast as experimental system and isothermal titration calorimetry, we here show that Arkadia specifically selects substrates carrying SUMO1-capped SUMO2/3 hybrid conjugates and targets them for proteasomal degradation. Our data suggest that a SUMO1-specific binding site in Arkadia with sequence similarity to a SUMO1-binding site in DPP9 is required for targeting endogenous hybrid SUMO conjugates and PML nuclear bodies in human cells. We thus characterize Arkadia as a STUbL with a preference for substrate proteins marked with distinct hybrid SUMO chains.

Definition, clinical classification and initial diagnosis of pulmonary hypertension: Updated recommendations from the Cologne Consensus Conference 2018.

In the summer of 2016, delegates from the German Society of Cardiology (DGK), the German Respiratory Society (DGP), and the German Society of Pediatric Cardiology (DGPK) met in Cologne, Germany, to define consensus-based practice recommendations for the management of patients with pulmonary hypertension (PH). These recommendations were built on the 2015 European Pulmonary Hypertension guidelines, aiming at their practical implementation, considering country-specific issues, and including new evidence, where available. To this end, a number of working groups was initiated, one of which was specifically dedicated to the definition, clinical classification and initial diagnosis of PH. While the European guidelines provide a detailed clinical classification and a structured approach for diagnostic testing, their application in routine care may be challenging, particularly given the changing phenotype of PH patients who are nowadays often elderly and may present with multiple potential causes of PH, as well as comorbid conditions. Specifically, the working group addresses the thoroughness of diagnostic testing, and the roles of echocardiography, exercise testing, and genetic testing in diagnosing PH. Furthermore, challenges in the diagnostic work-up of patients with various causes of PH including "PAH with comorbidities", CTEPH and coexisting conditions are highlighted, and a modified diagnostic algorithm is provided. The detailed results and recommendations of the working group on definition, clinical classification and initial diagnosis of PH, which were last updated in the spring of 2018, are summarized in this article.