A comprehensive map of proteoglycan expression and deposition in the pulmonary arterial wall in health and pulmonary hypertension.

Changes in the extracellular matrix of pulmonary arteries (PAs) are a key aspect of vascular remodelling in pulmonary hypertension (PH). Yet, our understanding of the alterations affecting the proteoglycan (PG) family remains limited. We sought to investigate the expression and spatial distribution of major vascular PGs in PAs from healthy individuals and various PH groups (chronic obstructive pulmonary disease: PH-COPD, pulmonary fibrosis: PH-PF, idiopathic: IPAH). PG regulation, deposition, and synthesis were notably heightened in IPAH, followed by PH-PF, with minor alterations in PH-COPD. Single-cell analysis unveiled cell-type and disease-specific PG regulation. Agrin expression, a basement membrane PG, was increased in IPAH, with PA endothelial cells (PAECs) identified as a major source. PA smooth muscle cells (PASMCs) mainly produced large-PGs, aggrecan and versican, and small-leucine-like proteoglycan (SLRP) biglycan, while the major PGs produced by adventitial fibroblasts were SLRP decorin and lumican. In IPAH and PF-PH, the neointima-forming PASMC population increased the expression of all investigated large-PGs and SLRPs, except fibroblast-predominant DCN. Expression of lumican, versican, and biglycan also positively correlated with collagen 1α1/1α2 expression in PASMCs of IPAH and PH-PF patients. We demonstrated that TGF-ß regulates versican and biglycan expression, indicating their contribution to vessel fibrosis in IPAH and PF-PH. We furthermore show that certain circulating PG levels display a disease-dependent pattern, with increased decorin and lumican across all patient groups, while versican was elevated in PH-COPD and IPAH and biglycan reduced in IPAH. These findings suggest unique compartment-specific PG regulation in different forms of PH, indicating distinct pathological processes.

In-depth analysis of pre- and postoperative functional outcome parameters in patients receiving laryngotracheal surgery.

OBJECTIVES: Surgical treatment for airway stenosis necessitates personalized techniques based on the stenosis location and length, leading to favourable surgical outcomes. However, there is limited literature on functional outcomes following laryngotracheal surgery with an adequate number of patients. METHODS: We conducted a retrospective analysis of patients who underwent laryngotracheal surgery at the Department of Thoracic Surgery, Medical University of Vienna, from January 2017 to June 2021. The study included standardized functional assessments before and after surgery, encompassing spirometry, voice measurements, swallowing evaluation and subjective patient perception. RESULTS: The study comprised 45 patients with an average age of 51.9 ± 15.9 years, of whom 89% were female, with idiopathic being the most common aetiology (67%). Procedures included standard cricotracheal resection in 11%, cricotracheal resection with dorsal mucosal flap in 49%, cricotracheal resection with dorsal mucosal flap and lateral cricoplasty in 24% and single-stage laryngotracheal reconstruction in 16%. There were no in-hospital mortalities or restenosis cases during the mean follow-up period of 20.8 ± 13.2 months. Swallowing function remained intact in all patients. Voice evaluations showed a decrease in fundamental vocal pitch [203 (81-290) Hz vs 150 (73-364) Hz, P < 0.001] and dynamic voice range (23.5 ± 5.8 semitones vs 17.8 ± 6.7 semitones, P < 0.001). However, no differences in voice volume were observed (60.0 ± 4.1 dB vs 60.2 ± 4.8 dB, P = 0.788). The overall predicted voice profile changed from R0B0H0 to R1B0H1. CONCLUSIONS: Laryngotracheal surgery proves effective in fully restoring breathing capacity while preserving vocal function. Even in cases of high-grade and complex airway stenosis necessitating laryngotracheal reconstruction, favourable functional outcomes can be achieved.

Alternative venous access sites for dual-lumen extracorporeal membrane oxygenation cannulation.

OBJECTIVES: Dual-lumen cannulas for veno-venous (VV) extracorporeal membrane oxygenation (ECMO) support are typically inserted in the right internal jugular vein (RIJV); however, some scenarios can make this venous route inaccessible. This multicentre case series aims to evaluate if single-site cannulation using an alternative venous access is safe and feasible in patients with an inaccessible RIJV. METHODS: We performed a multi-institutional retrospective analysis including high-volume ECMO centres with substantial experience in dual-lumen cannulation (DLC) (defined as >10 DLC per year). Three centres [Freiburg (Germany), Toronto (Canada) and Vienna (Austria)] agreed to share their data, including baseline characteristics, technical ECMO and cannulation data as well as complications related to ECMO cannulation and outcome. RESULTS: A total of 20 patients received alternative DLC for respiratory failure. Cannula insertion sites included the left internal jugular vein (n = 5), the right (n = 7) or left (n = 3) subclavian vein and the right (n = 4) or left (n = 1) femoral vein. The median cannula size was 26 (19-28) French. The median initial target ECMO flow was 2.9 (1.8-3.1) l/min and corresponded with used cannula size and estimated cardiac output. No procedural complications were reported during cannulation and median ECMO runtime was 15 (9-22) days. Ten patients were successfully bridged to lung transplantation (n = 5) or lung recovery (n = 5). Ten patients died during or after ECMO support. CONCLUSIONS: Alternative venous access sites for single-site dual-lumen catheters are a safe and feasible option to provide veno-venous ECMO support to patients with inaccessible RIJV.

Computed Tomography-Based Body Composition is Related to Perioperative Morbidity in Older Lung Transplant Recipients.

BACKGROUND: In older patients, a limited physical reserve is considered a contraindication for lung transplantation (LTx). Herein, we aimed to establish a computed tomography (CT)-based quantification of physical reserve in older patients scheduled for transplantation. METHODS: This retrospective study included patients older than 60 years who received LTx. Semiautomatic measurements of the mediastinal fat area and the dorsal muscle group area in pretransplantation CT scans were performed, and normalized data were correlated with clinical parameters. RESULTS: Patients (n = 108) were assigned into three groups (Musclehighfatlow [n = 25], Musclelowfathigh [n = 24], and other combinations [n = 59]). The Musclelowfathigh group had a significantly increased risk of wound infections (p = 0.002) and tracheostomy (p = 0.001) compared with Musclehighfatlow patients. The median length of intensive care unit stay (25 vs. 3.5 days; p = 0.002) and the median length of hospital stay (44 vs. 22.5 days; p = 0.013) post-LTx were significantly prolonged in the Musclelowfathigh group. Significantly more patients in this group had a prolonged ventilation time (11 vs. 0; p < 0.001). CONCLUSION: Body composition parameters determined in pretransplant chest CT scans in older LTx candidates might aid in identifying high-risk patients with a worse perioperative outcome after LTx.

The ESTS-AIR database-initial results of a multi-institutional database on airway surgery.

OBJECTIVES: Compared to lung resections, airway procedures are relatively rare in thoracic surgery. Despite this, a growing number of dedicated airway centres have formed throughout Europe. These centres are characterized by a close interdisciplinary collaboration and they often act as supra-regional referring centres. To date, most evidence of airway surgery comes from retrospective, single-centre analysis as there is a lack of large-scale, multi-institutional databases. METHODS: In 2018, an initiative was formed, which aimed to create an airway database within the framework of the ESTS database (ESTS-AIR). Five dedicated airway centres were asked to test the database in a pilot phase. A 1st descriptive analysis of ESTS-AIR was performed. RESULTS: A total of 415 cases were included in the analysis. For adults, the most common indication for airway surgery was post-tracheostomy stenosis and idiopathic subglottic stenosis; in children, most resections/reconstructions had to be performed for post-intubation stenosis. Malignant indications required significantly longer resections [36.0 (21.4-50.6) mm] when compared to benign indications [26.6 (9.4-43.8) mm]. Length of hospital stay was 11.0 (4.1-17.3) days (adults) and 13.4 (7.6-19.6) days (children). Overall, the rates of complications were low with wound infections being reported as the most common morbidity. CONCLUSIONS: This evaluation of the 1st cases in the ESTS-AIR database allowed a large-scale analysis of the practice of airway surgery in dedicated European airway centres. It provides proof for the functionality of ESTS-AIR and sets the basis for rolling out the AIR subsection to all centres participating in the ESTS database.

Compartment-specific remodeling patterns in end-stage chronic obstructive pulmonary disease with and without severe pulmonary hypertension.

BACKGROUND: In patients with end-stage chronic obstructive pulmonary disease (COPD), severe pulmonary hypertension (PH) is frequently associated with less severe airway obstruction as compared to mild or no PH. However, the histologic correlate of this finding is not clear. We aimed to quantify remodeling of pulmonary arteries, airways, and parenchyma in random samples of explanted end-stage COPD lungs. METHODS: We quantified remodeling of small pulmonary arteries, small airways, and the degree of emphysema (mean interseptal distance [MID]) with dedicated software. As primary objective, we compared COPD patients with severe PH (SevPH-COPD) with age- and sex-matched MildPH-COPD. For comparison, we also investigated COPD lungs with no PH (NoPH-COPD), idiopathic PAH (IPAH), and healthy donors. RESULTS: We included n = 17 SevPH-COPD (mPAP = 43 [39-45]mm Hg), n = 17 MildPH-COPD (mPAP = 28 [24-31]mm Hg), n = 5 NoPH-COPD (mPAP = 18 [16-19]mm Hg), n = 10 IPAH (mPAP = 72 [65-91]mm Hg), and n = 10 healthy donor lungs. SevPH-COPD versus MildPH-COPD was characterized by better preserved forced vital capacity (51% vs 40% predicted, p < 0.05), less emphysema (MID 169 µm vs 279 µm, p < 0.001), and less PAS-positive and CD45-positive mucosa cells (15% vs 22%, p = 0.063% and 5% vs 7%, p = 0.058) suggesting less airway inflammation. In COPD patients, intimal and medial thickening were strongly correlated with mPAP (r = 0.676, p < 0.001 and r = 0.595, p < 0.001). MID was negatively correlated with mPAP (r = -0.556, p < 0.001) and was highest in NoPH-COPD (mean 281 µm), suggesting that emphysema per se is not associated with PH. CONCLUSIONS: End-stage COPD with severe PH is characterized by pronounced pulmonary vascular remodeling, less inflammation of small airways, and less emphysema as compared to COPD with mild PH or no PH, suggesting that COPD with severe PH may represent a unique phenotype of COPD.

A novel technique of voice-sparing cricotracheal resection.

Background: Cricotracheal resection (CTR) is considered the standard of care for patients suffering from idiopathic subglottic stenosis (iSGS). Although CTR results in permanent restoration of airway patency, it has a mild to moderate impact on voice quality. Here we propose modifications of the standard CTR technique to make it a voice-preserving procedure. Methods: Five women with iSGS underwent voice-sparing CTR between January 2022 and January 2023. In this procedure, through several technical adaptations, the function of the cricothyroid joint was preserved. Outcomes of these voice-sparing CTRs were compared to outcomes in patients who underwent standard CTR in our institution. All patients underwent full functional preoperative and postoperative workups, including spirometry, voice measurements, patient self-assessment, and fiberoptic endoscopic evaluation of swallowing. Results: All 5 patients in the study group suffered from iSGS with high-grade Myer-Cotton III° stenosis (100%); 1 patient had previously undergone endoscopic laser resection. Voice evaluation demonstrated a nearly unchanged fundamental pitch (mean preoperative, 191 ± 73.1 Hz; postoperative, 182 ± 64.2 Hz) and dynamic voice range (preoperative, 24.4 semitones; postoperative, 20.4 semitones). This was in contrast to the control group, in which significantly reduced voice quality was observed. Conclusions: In selected patients suffering from iSGS, excellent functional results can be obtained with voice-sparing CTR.

C-Myc protein expression indicates unfavorable clinical outcome in surgically resected small cell lung cancer.

BACKGROUND: By being highly involved in the tumor evolution and disease progression of small cell lung cancer (SCLC), Myc family members (C-Myc, L-Myc, and N-Myc) might represent promising targetable molecules. Our aim was to investigate the expression pattern and prognostic relevance of these oncogenic proteins in an international cohort of surgically resected SCLC tumors. METHODS: Clinicopathological data and surgically resected tissue specimens from 104 SCLC patients were collected from two collaborating European institutes. Tissue sections were stained by immunohistochemistry (IHC) for all three Myc family members and the recently introduced SCLC molecular subtype-markers (ASCL1, NEUROD1, POU2F3, and YAP1). RESULTS: IHC analysis showed C-Myc, L-Myc, and N-Myc positivity in 48%, 63%, and 9% of the specimens, respectively. N-Myc positivity significantly correlated with the POU2F3-defined molecular subtype (r = 0.6913, p = 0.0056). SCLC patients with C-Myc positive tumors exhibited significantly worse overall survival (OS) (20 vs. 44 months compared to those with C-Myc negative tumors, p = 0.0176). Ultimately, in a multivariate risk model adjusted for clinicopathological and treatment confounders, positive C-Myc expression was confirmed as an independent prognosticator of impaired OS (HR 1.811, CI 95% 1.054-3.113, p = 0.032). CONCLUSIONS: Our study provides insights into the clinical aspects of Myc family members in surgically resected SCLC tumors. Notably, besides showing that positivity of Myc family members varies across the patients, we also reveal that C-Myc protein expression independently correlates with worse survival outcomes. Further studies are warranted to investigate the role of Myc family members as potential prognostic and predictive markers in this hard-to-treat disease.

Coronavirus Receptor Expression Profiles in Human Mast Cells, Basophils, and Eosinophils.

A major problem in SARS-CoV-2-infected patients is the massive tissue inflammation in certain target organs, including the lungs. Mast cells (MC), basophils (BA), and eosinophils (EO) are key effector cells in inflammatory processes. These cells have recently been implicated in the pathogenesis of SARS-CoV-2 infections. We explored coronavirus receptor (CoV-R) expression profiles in primary human MC, BA, and EO, and in related cell lines (HMC-1, ROSA, MCPV-1, KU812, and EOL-1). As determined using flow cytometry, primary MC, BA, and EO, and their corresponding cell lines, displayed the CoV-R CD13 and CD147. Primary skin MC and BA, as well as EOL-1 cells, also displayed CD26, whereas primary EO and the MC and BA cell lines failed to express CD26. As assessed using qPCR, most cell lines expressed transcripts for CD13, CD147, and ABL2, whereas ACE2 mRNA was not detectable, and CD26 mRNA was only identified in EOL-1 cells. We also screened for drug effects on CoV-R expression. However, dexamethasone, vitamin D, and hydroxychloroquine did not exert substantial effects on the expression of CD13, CD26, or CD147 in the cells. Together, MC, BA, and EO express distinct CoV-R profiles. Whether these receptors mediate virus-cell interactions and thereby virus-induced inflammation remains unknown at present.

Red blood cell transfusion-related dynamics of extracellular vesicles in intensive care patients: a prospective subanalysis.

Extracellular vesicles (EVs) accumulate during packed red blood cell (PRBC) storage. To date, the involvement of EVs in transfusion-related immunomodulation (TRIM) has not been prospectively evaluated in intensive care unit (ICU) patients. This was a prospective subanalysis of a recent observational feasibility study in postoperative ICU patients after: (1) open aortic surgery (Aorta), (2) bilateral lung transplantation (LuTx), and (3) other types of surgery (Comparison). Patient plasma was collected three times each before and after leukoreduced PRBC transfusion at 30-min intervals. The total number of EVs and EVs derived from erythrocytes (EryEVs), total platelets (total PEVs), activated platelets, granulocytes (GEVs), monocytes, and myeloid cells in PRBC samples and patient plasma were analyzed by flow cytometry. Statistical analysis was performed by Spearman's correlation test, linear mixed models and pairwise comparisons by Wilcoxon matched-pairs test. Twenty-three patients (Aorta n = 5, LuTx n = 9, Comparison n = 9) were included in the final analysis. All EV subgroups analyzed were detectable in all PRBCs samples (n = 23), but concentrations did not correlate with storage time. Moreover, all EVs analyzed were detectable in all plasma samples (n = 138), and EV counts were consistent before transfusion. Concentrations of total EVs, EryEVs, total PEVs, and GEVs increased after transfusion compared with baseline in the entire cohort but not in specific study groups. Furthermore, the change in plasma EV counts (total EVs and EryEVs) after transfusion correlated with PRBC storage time in the entire cohort. Extracellular vesicles were detectable in all PRBC and plasma samples. Individual EV subtypes increased after transfusion in the entire cohort, and in part correlated with storage duration. Future clinical studies to investigate the role of EVs in TRIM are warranted and should anticipate a larger sample size.Trial registration: Clinicaltrials.gov: NCT03782623.

High Rate of Passenger Lymphocyte Syndrome after ABO Minor Incompatible Lung Transplantation.

Rationale: Passenger lymphocyte syndrome (PLS) may complicate minor ABO mismatched lung transplantation (LuTX) via donor-derived red cell antibody-induced hemolysis.Objectives: To ascertain the incidence and specificity of PLS-relevant antibodies among the study population as well as the dynamics of hemolysis parameters and the transfusion requirement of patients with or without PLS.Methods: In this cohort study, 1,011 patients who received LuTX between January 2010 and June 2019 were studied retrospectively. Prospectively, 87 LuTX (July 2019 to June 2021) were analyzed. Postoperative ABO antibody and hemolytic marker determinations, transfusion requirement, and duration of postoperative hospital care were analyzed. Retrospectively, blood group A recipients of O grafts with PLS were compared with those without.Measurements and Main Results: PLS affected 18.18% (retrospective) and 30.77% (prospective) of A recipients receiving O grafts, 5.13% of B recipients of O grafts, and 20% of AB patients receiving O transplants. Anti-A and anti-A1 were the predominant PLS-inducing antibodies, followed by anti-B and anti-A,B. Significantly lower hemoglobin values (median, 7.4 vs. 8.3 g/dl; P = 0.0063) and an approximately twice as high percentage of patients requiring blood transfusions were seen in PLS. No significant differences in other laboratory markers, duration of hospital stay, or other complications after LuTX were registered.Conclusions: Minor ABO incompatible LuTX recipients are at considerable risk of developing clinically significant PLS. Post-transplant monitoring combining red cell serology and hemolysis marker determination appears advisable so as not to overlook hemolytic episodes that necessitate antigen-negative transfusion therapy.

Pentastatin, a matrikine of the collagen IVα5, is a novel endogenous mediator of pulmonary endothelial dysfunction.

Deposition of basement membrane components, such as collagen IVα5, is associated with altered endothelial cell function in pulmonary hypertension. Collagen IVα5 harbors a functionally active fragment within its C-terminal noncollageneous (NC1) domain, called pentastatin, whose role in pulmonary endothelial cell behavior remains unknown. Here, we demonstrate that pentastatin serves as a mediator of pulmonary endothelial cell dysfunction, contributing to pulmonary hypertension. In vitro, treatment with pentastatin induced transcription of immediate early genes and proinflammatory cytokines and led to a functional loss of endothelial barrier integrity in pulmonary arterial endothelial cells. Mechanistically, pentastatin leads to ß1-integrin subunit clustering and Rho/ROCK activation. Blockage of the ß1-integrin subunit or the Rho/ROCK pathway partially attenuated the pentastatin-induced endothelial barrier disruption. Although pentastatin reduced the viability of endothelial cells, smooth muscle cell proliferation was induced. These effects on the pulmonary vascular cells were recapitulated ex vivo in the isolated-perfused lung model, where treatment with pentastatin-induced swelling of the endothelium accompanied by occasional endothelial cell apoptosis. This was reflected by increased vascular permeability and elevated pulmonary arterial pressure induced by pentastatin. This study identifies pentastatin as a mediator of endothelial cell dysfunction, which thus might contribute to the pathogenesis of pulmonary vascular disorders such as pulmonary hypertension.NEW & NOTEWORTHY This study is the first to show that pentastatin, the matrikine of the basement membrane (BM) collagen IVα5 polypeptide, triggers rapid pulmonary arterial endothelial cell barrier disruption, activation, and apoptosis in vitro and ex vivo. Mechanistically, pentastatin partially acts through binding to the ß1-integrin subunit and the Rho/ROCK pathway. These findings are the first to link pentastatin to pulmonary endothelial dysfunction and, thus, suggest a major role for BM-matrikines in pulmonary vascular diseases such as pulmonary hypertension.

Entinostat Enhances the Efficacy of Chemotherapy in Small Cell Lung Cancer Through S-phase Arrest and Decreased Base Excision Repair.

PURPOSE: Acquired chemoresistance is a frequent event in small cell lung cancer (SCLC), one of the deadliest human malignancies. Histone deacetylase inhibitors (HDACi) have been shown to synergize with different chemotherapeutic agents including cisplatin. Accordingly, we aimed to investigate the dual targeting of HDAC inhibition and chemotherapy in SCLC. EXPERIMENTAL DESIGN: The efficacy of HDACi and chemotherapy in SCLC was investigated both in vitro and in vivo. Synergistic drug interactions were calculated based on the HSA model (Combenefit software). Results from the proteomic analysis were confirmed via ICP-MS, cell-cycle analysis, and comet assays. RESULTS: Single entinostat- or chemotherapy significantly reduced cell viability in human neuroendocrine SCLC cells. The combination of entinostat with either cisplatin, carboplatin, irinotecan, epirubicin, or etoposide led to strong synergy in a subset of resistant SCLC cells. Combination treatment with entinostat and cisplatin significantly decreased tumor growth in vivo. Proteomic analysis comparing the groups of SCLC cell lines with synergistic and additive response patterns indicated alterations in cell-cycle regulation and DNA damage repair. Cell-cycle analysis revealed that cells exhibiting synergistic drug responses displayed a shift from G1 to S-phase compared with cells showing additive features upon dual treatment. Comet assays demonstrated more DNA damage and decreased base excision repair in SCLC cells more responsive to combination therapy. CONCLUSIONS: In this study, we decipher the molecular processes behind synergistic interactions between chemotherapy and HDAC inhibition. Moreover, we report novel mechanisms to overcome drug resistance in SCLC, which may be relevant to increasing therapeutic success.

Targeting YB-1 via entinostat enhances cisplatin sensitivity of pleural mesothelioma in vitro and in vivo.

Pleural mesothelioma (PM) is characterized by poor prognosis and limited therapeutic options. Y-box-binding protein 1 (YB-1) was shown to drive growth and migration of PM cells. Here, we evaluated the effect of genetic and pharmacological targeting of YB-1 on PM growth and response to cisplatin and radiation treatment. YB-1 knockdown via siRNA resulted in reduced PM cell growth, which significantly correlated with wt BAP1 and mutant NF2 and P53 status. Entinostat inhibited YB-1 deacetylation and its efficacy correlated with YB-1 knockdown-induced growth inhibition in 20 PM cell lines. Tumor growth inhibition by siRNA as well as entinostat was confirmed in mouse xenotransplant models. Furthermore, both YBX1-targeting siRNA and entinostat enhanced sensitivity to cisplatin and radiation. In particular, entinostat showed strong synergistic interactions with cisplatin which was linked to significantly increased cellular platinum uptake in all investigated cell models. Importantly, in a mouse model, the combination of cisplatin and entinostat also resulted in stronger growth inhibition than each treatment alone. Our study highlights YB-1 as an attractive target in PM and demonstrates that targeting YB-1 via entinostat is a promising approach to enhance cisplatin and radiation sensitivity.

How We Would Treat Our Own Lung Transplantation: A Multidisciplinary and International Perspective.

Lung transplantation is the ultimate treatment for end-stage lung disease (ESLD) arising from various etiologies. This manuscript will focus on selected aspects of a theoretical perioperative lung transplantation plan, highlighting data-driven and theoretical techniques the authors would employ if they were the index patient undergoing surgery. Beginning with looking for a transplant center using an Internet search, patient advocacy organizations, and artificial intelligence guidance, the authors lay out a course for multidisciplinary care throughout the entire patient journey, ending with ideal approaches to recovery 6 months posttransplantation. Although the index case will focus on a theoretical patient with pulmonary fibrosis, the general management principles will be applicable across the entire spectrum of patients with ESLD presenting for lung transplantation.

Microenvironmental regulation of T-cells in pulmonary hypertension.

Introduction: In pulmonary hypertension (PH), pulmonary arterial remodeling is often accompanied by perivascular inflammation. The inflammation is characterized by the accumulation of activated macrophages and lymphocytes within the adventitial stroma, which is comprised primarily of fibroblasts. The well-known ability of fibroblasts to secrete interleukins and chemokines has previously been implicated as contributing to this tissue-specific inflammation in PH vessels. We were interested if pulmonary fibroblasts from PH arteries contribute to microenvironmental changes that could activate and polarize T-cells in PH. Methods: We used single-cell RNA sequencing of intact bovine distal pulmonary arteries (dPAs) from PH and control animals and flow cytometry, mRNA expression analysis, and respirometry analysis of blood-derived bovine/human T-cells exposed to conditioned media obtained from pulmonary fibroblasts of PH/control animals and IPAH/control patients (CM-(h)PH Fibs vs CM-(h)CO Fibs). Results: Single-cell RNA sequencing of intact bovine dPAs from PH and control animals revealed a pro-inflammatory phenotype of CD4+ T-cells and simultaneous absence of regulatory T-cells (FoxP3+ Tregs). By exposing T-cells to CM-(h)PH Fibs we stimulated their proinflammatory differentiation documented by increased IFNγ and decreased IL4, IL10, and TGFß mRNA and protein expression. Interestingly, we demonstrated a reduction in the number of suppressive T-cell subsets, i.e., human/bovine Tregs and bovine γδ T-cells treated with CM-(h)PH-Fibs. We also noted inhibition of anti-inflammatory cytokine expression (IL10, TGFß, IL4). Pro-inflammatory polarization of bovine T-cells exposed to CM-PH Fibs correlated with metabolic shift to glycolysis and lactate production with increased prooxidant intracellular status as well as increased proliferation of T-cells. To determine whether metabolic reprogramming of PH-Fibs was directly contributing to the effects of PH-Fibs conditioned media on T-cell polarization, we treated PH-Fibs with the HDAC inhibitor SAHA, which was previously shown to normalize metabolic status and examined the effects of the conditioned media. We observed significant suppression of inflammatory polarization associated with decreased T-cell proliferation and recovery of mitochondrial energy metabolism. Conclusion: This study demonstrates how the pulmonary fibroblast-derived microenvironment can activate and differentiate T-cells to trigger local inflammation, which is part of the vascular wall remodeling process in PH.

Laser ablation-inductively coupled plasma-mass spectrometry analysis reveals differences in chemotherapeutic drug distribution in surgically resected pleural mesothelioma.

AIMS: Pleural mesothelioma (PM) is a highly aggressive thoracic tumour with poor prognosis. Although reduced tissue drug accumulation is one of the key features of platinum (Pt) resistance, little is known about Pt distribution in human PM. METHODS: We assessed Pt levels of blood samples and surgically resected specimens from 25 PM patients who had received neoadjuvant Pt-based chemotherapy (CHT). Pt levels and tissue distributions were measured by laser ablation-inductively coupled plasma-mass spectrometry and correlated with clinicopathological features. RESULTS: In surgically resected PM specimens, mean Pt levels of nontumourous (fibrotic) areas were significantly higher (vs tumourous regions, P = 0.0031). No major heterogeneity of Pt distribution was seen within the tumourous areas. Pt levels correlated neither with the microvessel area nor with apoptosis rate in the tumourous or nontumourous regions. A significant positive correlation was found between serum and both full tissue section and tumourous area mean Pt levels (r = 0.532, P = 0.006, 95% confidence interval [95% CI] 0.161-0.771 and r = 0.415, P = 0.039, 95% CI 0.011-0.702, respectively). Furthermore, a significant negative correlation was detected between serum Pt concentrations and elapsed time from the last cycle of CHT (r = -0.474, P = 0.017, 95% CI -0.738--0.084). Serum Pt levels correlated negatively with overall survival (OS) (P = 0.029). CONCLUSIONS: There are major differences in drug distribution between tumourous and nontumourous areas of PM specimens. Serum Pt levels significantly correlate with full section and tumourous area average Pt levels, elapsed time from the last CHT cycle, and OS. Further studies investigating clinicopathological factors that modulate tissue Pt concentration and distribution are warranted.

Comparative expression analysis of immune-related markers in surgically resected lung neuroendocrine neoplasms.

BACKGROUND: Although immunotherapy has led to a paradigm shift in the treatment of lung cancer, the therapeutic approaches for lung neuroendocrine neoplasms (LNENs) are still limited. Our aim was to explore the immunological landscape and the expression of immune checkpoint markers in LNENs. METHODS: Surgically removed tumor samples of 26 atypical carcinoid (AC), 30 large cell neuroendocrine carcinoma (LCNEC) and 29 small cell lung cancer (SCLC) patients were included. The immune phenotype of each tumor type was assessed by using a panel of 15 immune-related markers. As these markers are potentially expressed by immune cells and/or tumor cells, they might serve as putative targets for immunotherapy. Expression patterns were measured by immunohistochemistry and correlated with clinicopathological parameters and prognosis. RESULTS: Unsupervised hierarchical clustering revealed distinct immunologic profiles across tumor types. Specifically, AC tumors were characterized by high tumor cell CD40 expression and low levels of immune infiltrates whereas SCLC samples had a high CD47 and Inducible T Cell Costimulator (ICOS) expression in tumor cells and immune cells, respectively. High CD70 and CD137 expression by tumor cells as well as elevated expression of CD27, Lymphocyte Activation Gene 3 (LAG3), and CD40 by immune cells were characteristic for LCNEC samples. Overall, SCLC and LCNEC tumors had a more immunogenic phenotype than AC samples. High tumor cell CD47 and CD40 expressions were associated with impaired and improved survival outcomes, respectively. CONCLUSIONS: By providing insights into the widely divergent immunologic profiles of LNENs, our results might serve as a basis for the development of novel immunotherapy-related approaches in these devastating malignancies.

Clinical insights into small cell lung cancer: Tumor heterogeneity, diagnosis, therapy, and future directions.

Small cell lung cancer (SCLC) is characterized by rapid growth and high metastatic capacity. It has strong epidemiologic and biologic links to tobacco carcinogens. Although the majority of SCLCs exhibit neuroendocrine features, an important subset of tumors lacks these properties. Genomic profiling of SCLC reveals genetic instability, almost universal inactivation of the tumor suppressor genes TP53 and RB1, and a high mutation burden. Because of early metastasis, only a small fraction of patients are amenable to curative-intent lung resection, and these individuals require adjuvant platinum-etoposide chemotherapy. Therefore, the vast majority of patients are currently being treated with chemoradiation with or without immunotherapy. In patients with disease confined to the chest, standard therapy includes thoracic radiotherapy and concurrent platinum-etoposide chemotherapy. Patients with metastatic (extensive-stage) disease are treated with a combination of platinum-etoposide chemotherapy plus immunotherapy with an anti-programmed death-ligand 1 monoclonal antibody. Although SCLC is initially very responsive to platinum-based chemotherapy, these responses are transient because of the development of drug resistance. In recent years, the authors have witnessed an accelerating pace of biologic insights into the disease, leading to the redefinition of the SCLC classification scheme. This emerging knowledge of SCLC molecular subtypes has the potential to define unique therapeutic vulnerabilities. Synthesizing these new discoveries with the current knowledge of SCLC biology and clinical management may lead to unprecedented advances in SCLC patient care. Here, the authors present an overview of multimodal clinical approaches in SCLC, with a special focus on illuminating how recent advancements in SCLC research could accelerate clinical development.

Circulating FGF18 is decreased in pleural mesothelioma but not correlated with disease prognosis.

BACKGROUND: Pleural mesothelioma (PM) is a relatively rare malignancy with limited treatment options and dismal prognosis. We have previously found elevated FGF18 expression in PM tissue specimens compared with normal mesothelium. The objective of the current study was to further explore the role of FGF18 in PM and evaluate its suitability as a circulating biomarker. METHODS: FGF18 mRNA expression was analyzed by real-time PCR in cell lines and in silico in datasets from the Cancer Genome Atlas (TCGA). Cell lines overexpressing FGF18 were generated by retroviral transduction and cell behavior was investigated by clonogenic growth and transwell assays. Plasma was collected from 40 PM patients, six patients with pleural fibrosis, and 40 healthy controls. Circulating FGF18 was measured by ELISA and correlated to clinicopathological parameters. RESULTS: FGF18 showed high mRNA expression in PM and PM-derived cell lines. PM patients with high FGF18 mRNA expression showed a trend toward longer overall survival (OS) in the TCGA dataset. In PM cells with low endogenous FGF18 expression, forced overexpression of FGF18 resulted in reduced growth but increased migration. Surprisingly, despite the high FGF18 mRNA levels observed in PM, circulating FGF18 protein was significantly lower in PM patients and patients with pleural fibrosis than in healthy controls. No significant association of circulating FGF18 with OS or other disease parameters of PM patients was observed. CONCLUSIONS: FGF18 is not a prognostic biomarker in PM. Its role in PM tumor biology and the clinical significance of decreased plasma FGF18 in PM patients warrant further investigation.