Increasing evidence points towards an inflammatory component underlying pulmonary hypertension. However, the conclusive characterisation of multiple inflammatory cell populations in the lung is challenging due to the complexity of marker specificity and tissue inaccessibility. We used an unbiased computational flow cytometry approach to delineate the inflammatory landscape of idiopathic pulmonary arterial hypertension (IPAH) and healthy donor lungs.Donor and IPAH samples were discriminated clearly using principal component analysis to reduce the multidimensional data obtained from single-cell flow cytometry analysis. In IPAH lungs, the predominant CD45⁺cell type switched from neutrophils to CD3⁺T-cells, with increases in CD4⁺, CD8⁺and γδT-cell subsets. Additionally, diversely activated classical myeloid-derived dendritic cells (CD14⁻HLA-DR⁺CD11c⁺CD1a^+/−) and nonclassical plasmacytoid dendritic cells (pDCs; CD14⁻CD11c⁻CD123⁺HLA-DR⁺), together with mast cells and basophils, were more abundant in IPAH samples. We describe, for the first time, the presence and regulation of two cell types in IPAH, γδT-cells and pDCs, which link innate and adaptive immunity.With our high-throughput flow cytometry with multidimensional dataset analysis, we have revealed the interactive interplay between multiple inflammatory cells is a crucial part of their integrative network. The identification of γδT-cells and pDCs in this disease potentially provides a missing link between IPAH, autoimmunity and inflammation.

Expression of the telomerase reverse transcriptase (TERT) might be altered by activating mutations of the rs2853669 polymorphism within the promoter region. Here we investigate the impact of these genomic alterations on telomerase activation and dissect their prognostic potential in glioblastoma (GBM). The respective TERT promoter region was sequenced in 126 GBM tissues and compared with clinical parameters and glioma biomarkers MGMT promoter methylation and IDH1 mutation. TERT mRNA expression, telomerase activity, and telomere lengths were determined by reverse transcriptase PCR, TRAP assay, and real-time PCR, respectively. Seventy-three percent of GBM patients harbored TERT promoter mutations associated with enhanced telomerase activity and TERT mRNA expression but reduced telomere lengths (P < .001 for all). Patients with mutated tumors exhibited significantly shorter overall survival in the entire cohort (11.5 vs 23.1 months; P < .0001) and in the primary GBM patient subgroup lacking IDH1 mutations (n = 120; P = .0084). This prognostic impact was confined to younger patients (aged <65 years), while the negative prognostic power of enhanced age at diagnosis was limited to those patients lacking TERT promoter mutations. Presence of the common single nucleotide polymorphism rs2853669, disrupting an endogenous Ets2 transcription factor-binding site, was associated with improved survival exclusively in patients with a wild-type TERT promoter. On the contrary, the shortest mean overall survival was detected in those patients harboring both an activating TERT promoter mutation and homozygous rs2853669 alleles. In summary, TERT promoter mutations are powerful prognosticators for worse course of disease in human GBM patients but their prognostic value is influenced by the rs2853669 polymorphism and age at diagnosis.

Background. The introduction of the lung allocation score has brought lung transplantation (LTX) of patients on extracorporeal membrane oxygenation (ECMO) bridge into the focus of interest. We reviewed our institutional experience with ECMO as a bridge to LTX. Methods. Between 1998 and 2011, 38 patients (median age 30.1 years, range 13–66 years) underwent ECMO support with intention to bridge to primary LTX. The underlying diagnosis was cystic fibrosis (n=17), pulmonary hypertension (n=4), idiopathic pulmonary fibrosis (n=9), adult respiratory distress syndrome (n=4), hemosiderosis (n=1), bronchiolitis obliterans (n=1), sarcoidosis (n=1), and bronchiectasis (n=1). The type of extracorporeal bridge was venovenous (n=18), venoarterial (n=15), interventional lung assist (n=1), or a stepwise combination of them (n=4). The median bridging time was 5.5 days (range 1–63) days. The type of transplantation was double LTX (n=7), size-reduced double LTX (n=8), lobar LTX (n=16), split LTX (n=2), and lobar LTX after ex vivo lung perfusion (n=1). Results. Four patients died before transplantation. Thirty-four patients underwent LTX, of them eight patients died in the hospital after a median stay of 24.5 days (range 1–180 days). Twenty-six patients left the hospital and returned to normal life (median hospital stay=47.5 days; range 21–90 days). The 1-, 3-, and 5-year survival for all transplanted patients was 60%, 60%, and 48%, respectively. The 1-, 3-, and 5-year survival conditional on 3-month survival for patients bridged with ECMO to LTX (78%, 78%, and 63%) was not worse than for other LTX patients within the same period of time (90%, 80%, and 72%, respectively, P=0.09, 0.505, and 0.344). Conclusion. Transplantation of patients bridged on ECMO to LTX is feasible and results in acceptable outcome.

Rationale: The development of pulmonary hypertension (PH) in patients with idiopathic pulmonary fibrosis (IPF) or chronic obstructive pulmonary disease (COPD) is associated with increased morbidity. Objectives: To elucidate whether vascular remodeling in a well-characterized PH-COPD and PH-IPF patient cohort results from similar or divergent molecular changes. Methods: Vascular remodeling of donor, PH-COPD, and PH-IPF pulmonary arteries was assessed. Laser capture microdissected pulmonary artery profiles in combination with whole genome microarrays were performed. Measurements and Main Results: Pulmonary arteries from patients with COPD and IPF with PH exhibited remodeling of vascular layers and reduction of lumen area. Pathway analyses comparing normalized gene expression profiles obtained from patients with PH-IPF or PH-COPD revealed the retinol and extracellular matrix (ECM) receptor interaction to be the most perturbed processes. Within the ECM-receptor pathway, differential regulation of 5 out of the top 10 results (collagen, type III, α-1; tenascin C; collagen, type VI, α-3; thrombospondin 2; and von Willebrand factor) were verified by real-time polymerase chain reaction and immunohistochemical staining. Conclusions: Despite clinical and histologic vascular remodeling in all patients with PH-COPD and PH-IPF, differential gene expression pattern was present in pulmonary artery profiles. Several genes involved in retinol metabolism and ECM receptor interaction enable discrimination of vascular remodeling in PH-IPF or PH-COPD. This suggests that pulmonary arterial remodeling in PH-COPD and PH-IPF is caused by different molecular mechanisms and may require specific therapeutic options.

The published experience with advanced broncho-plastic procedures performed with extracorporeal membrane oxygenation (ECMO) support is very limited. We examined our results to assess the risks and benefits of this approach. We retrospectively analysed all patients with thoracic malignancies who underwent complex tracheo-bronchial reconstruction under ECMO support in our department between 2001 and 2013. Ten patients (age range 21–81 years, mean 54 ± 11 years) underwent complex tracheo-bronchial reconstructions under veno-arterial ECMO support. In 7 patients, the underlying pathology was non-small-cell lung cancer, in 2 cases carcinoid tumour and in 1 case adenoid cystic carcinoma. ECMO cannulation was central (n = 7) or peripheral (n = 3). Mean time on bypass was 113 ± 17 min (range 70–135 min). A complete resection (R0) was achieved in 8 patients (80%). There was no perioperative mortality. Patients were discharged from the hospital after 7–52 days (median 11 days). Median time on ICU was 1 day (range 1–36 days). There was no complication related to the use of ECMO in this series. Mean follow-up time was 1694 ± 1385 days (range 12–4338). The 1-, 3- and 5-year Kaplan–Meier survival was 100, 74 and 56%, respectively. Based on this experience, we consider veno-arterial ECMO support as a safe and valuable approach for complex airway surgery.

Alterations in extracellular matrix (ECM) have been implicated in the pathophysiology of pulmonary hypertension. Here, we have undertaken a compartment-specific study to elucidate the expression profile of collagens and their processing enzymes in donor and idiopathic pulmonary arterial hypertension (IPAH) pulmonary arteries. Predominant intimal, but also medial and perivascular, remodeling and reduced lumen diameter were detected in IPAH pulmonary arteries. Two-photon microscopy demonstrated accumulation of collagen fibers. Quantification of collagen in pulmonary arteries revealed collagen accumulation mainly in the intima of IPAH pulmonary arteries compared with donors. Laser capture-microdissected pulmonary artery profiles (intima+media and perivascular tissue) were analyzed by real-time PCR for ECM gene expression. In the intima+media of IPAH vessels, collagens ( COL4A5, COL14A1, and COL18A1), matrix metalloproteinase (MMP) 19, and a disintegrin and metalloprotease (ADAM) 33 were higher expressed, whereas MMP10, ADAM17, TIMP1, and TIMP3 were less abundant. Localization of COLXVIII, its cleavage product endostatin, and MMP10, ADAM33, and TIMP1 was confirmed in pulmonary arteries by immunohistochemistry. ELISA for collagen XVIII/endostatin demonstrated significantly elevated plasma levels in IPAH patients compared with donors, whereas circulating MMP10, ADAM33, and TIMP1 levels were similar between the two groups. Endostatin levels were correlated with pulmonary arterial wedge pressure, and established prognostic markers of IPAH, right atrial pressure, cardiac index, 6-min walking distance, NH₂-terminal pro-brain natriuretic peptide, and uric acid. Expression of unstudied collagens, MMPs, ADAMs, and TIMPs were found to be significantly altered in IPAH intima+media. Elevated levels of circulating collagen XVIII/endostatin are associated with markers of a poor prognosis.