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.

Expression patterns of novel immunotherapy targets in intermediate- and high-grade lung neuroendocrine neoplasms.

BACKGROUND: Advancements in immunotherapeutic approaches only had a modest impact on the therapy of lung neuroendocrine neoplasms (LNENs). Our multicenter study aimed to investigate the expression patterns of novel immunotherapy targets in intermediate- and high-grade LNENs. METHODS: The expressions of V-domain Ig suppressor of T cell activation (VISTA), OX40L, Glucocorticoid-induced TNF receptor (GITR), and T cell immunoglobulin and mucin domain 3 (TIM3) proteins were measured by immunohistochemistry in surgically resected tumor samples of 26 atypical carcinoid (AC), 49 large cell neuroendocrine lung cancer (LCNEC), and 66 small cell lung cancer (SCLC) patients. Tumor and immune cells were separately scored. RESULTS: Tumor cell TIM3 expression was the highest in ACs (p < 0.001), whereas elevated tumor cell GITR levels were characteristic for both ACs and SCLCs (p < 0.001 and p = 0.011, respectively). OX40L expression of tumor cells was considerably lower in ACs (vs. SCLCs; p < 0.001). Tumor cell VISTA expression was consistently low in LNENs, with no significant differences across histological subtypes. ACs were the least immunogenic tumors concerning immune cell abundance (p < 0.001). Immune cell VISTA and GITR expressions were also significantly lower in these intermediate-grade malignancies than in SCLCs or in LCNECs. Immune cell TIM3 and GITR expressions were associated with borderline prognostic significance in our multivariate model (p = 0.057 and p = 0.071, respectively). CONCLUSIONS: LNEN subtypes have characteristic and widely divergent VISTA, OX40L, GITR, and TIM3 protein expressions. By shedding light on the different expression patterns of these immunotherapy targets, the current multicenter study provides support for the future implementation of novel immunotherapeutic approaches.

Standardized Classification of Lung Adenocarcinoma Subtypes and Improvement of Grading Assessment Through Deep Learning.

The histopathologic distinction of lung adenocarcinoma (LADC) subtypes is subject to high interobserver variability, which can compromise the optimal assessment of patient prognosis. Therefore, this study developed convolutional neural networks capable of distinguishing LADC subtypes and predicting disease-specific survival, according to the recently established LADC tumor grades. Consensus LADC histopathologic images were obtained from 17 expert pulmonary pathologists and one pathologist in training. Two deep learning models (AI-1 and AI-2) were trained to predict eight different LADC classes. Furthermore, the trained models were tested on an independent cohort of 133 patients. The models achieved high precision, recall, and F1 scores exceeding 0.90 for most of the LADC classes. Clear stratification of the three LADC grades was reached in predicting the disease-specific survival by the two models, with both Kaplan-Meier curves showing significance (P = 0.0017 and 0.0003). Moreover, both trained models showed high stability in the segmentation of each pair of predicted grades with low variation in the hazard ratio across 200 bootstrapped samples. These findings indicate that the trained convolutional neural networks improve the diagnostic accuracy of the pathologist and refine LADC grade assessment. Thus, the trained models are promising tools that may assist in the routine evaluation of LADC subtypes and grades in clinical practice.

Mesothelioma in the pleura, pericardium and peritoneum: Recommendations from the International Collaboration on Cancer Reporting (ICCR).

AIMS: Mesothelioma is a rare malignancy of the serosal membranes that is commonly related to exposure to asbestos. Despite extensive research and clinical trials, prognosis to date remains poor. Consistent, comprehensive and reproducible pathology reporting form the basis of all future interventions for an individual patient, but also ensures that meaningful data are collected to identify predictive and prognostic markers. METHODS AND RESULTS: This article details the International Collaboration on Cancer Reporting (ICCR) process and the development of the international consensus mesothelioma reporting data set. It describes the 'core' and 'non-core' elements to be included in pathology reports for mesothelioma of all sites, inclusive of clinical, macroscopic, microscopic and ancillary testing considerations. An international expert panel consisting of pathologists and a medical oncologist produced a set of data items for biopsy and resection specimens based on a critical review and discussion of current evidence, and in light of the changes in the 2021 WHO Classification of Tumours. The commentary focuses particularly upon new entities such as mesothelioma in situ and provides background on relevant and essential ancillary testing as well as implementation of the new requirement for tumour grading. CONCLUSION: We recommend widespread and consistent implementation of this data set, which will facilitate accurate reporting and enhance the consistency of data collection, improve the comparison of epidemiological data, support retrospective research and ultimately help to improve clinical outcomes. To this end, all data sets are freely available worldwide on the ICCR website (www.iccr-cancer.org/data-sets).

From Morphology to Molecules: Advances in the Distinction of Multiple Primary Lung Cancers From Intrapulmonary Metastases in Non-Small Cell Lung Cancer.

The increasing incidence of multiple lung nodules underscores the need for precise differentiation between multiple primary lung cancers (MPLCs) and intrapulmonary metastases (IPMs). This distinction impacts patient prognosis and treatment strategies. The prevalence of multiple lung nodules, ranging from 19.7% to 55.5%, highlights the clinical significance of this challenge. Historically, the role of histopathology, particularly comprehensive histology assessment (CHA), has been pivotal in differentiating MPLCs and IPMs. However, CHA has significant limitations, resulting in a constant search for a better way to distinguish those lesions. The best strategy for delineating MPLCs from IPMs is a multidisciplinary approach combining clinical data, radiology, histology, and molecular methods. Histology provides architectural and cellular characteristics, radiology contributes anatomic context and lesion characterization, and molecular methods reveal molecular features critical for accurate differentiation. Incorporating clinical data further enhances diagnostic precision. This review presents current knowledge and current approaches to multiple lung tumors. It is clear that even with a combination of pathology, radiology, and molecular data, definitive classification of multifocal lung tumors is not always possible.

Detection of antibody subclasses IgA, IgM and IgG against HPV L1 in HPV-positive oropharyngeal squamous cell carcinoma patients: a pilot study.

PURPOSE: Despite prognostic superiority of HPV-positive oropharyngeal squamous cell carcinoma (OPSCC), up to 25% of patients will suffer from recurrence within the first 5 years. Therefore, it is of great scientific interest to find relevant biomarkers to identify patients at risk. In this prospective observational study, we aimed to investigate the dynamics of HPV-L1 capsid protein specific antibody (AB) subclasses IgA, IgM, and IgG in HPV-positive OPSCC patients under therapy. METHODS: Serum samples from HPV-positive OPSCC patients, identified by positive p16-immunohistochemistry, were collected before and during tumor-specific therapy and 3-6 months during follow-up. They were analyzed for the presence of HPV-L1 AB subclasses IgA, IgM, and IgG using an HPV-L1-specific immuno-assay. Additionally, a PCR-based HPV-DNA detection from the tumor tissue was performed. RESULTS: Altogether, 33 patients with a mean follow-up of 55 months were included. Analysis of a total of 226 serum samples revealed that the most common L1-AB-subclass pattern was characterized by IgG > > IgA > IgM without significant fluctuation during the course of disease. Patients with excessive IgG levels tended to higher tumor stages and three out of three patients with disease recurrence showed increasing IgG AB titers beforehand. Seven patients showed an IgA dominance at diagnosis, which was associated with a better disease-free survival. CONCLUSION: Despite limited cases, our prospective pilot study revealed promising trends in HPV L1 AB subclasses and may contribute useful information for future risk stratification and post-treatment monitoring in HPV-positive OPSCC patients.

NR4A3 fusions characterize a distinctive peritoneal mesothelial neoplasm of uncertain biological potential with pure adenomatoid/microcystic morphology.

A focal adenomatoid-microcystic pattern is not uncommon in peritoneal mesothelioma, but tumors composed almost exclusively of this pattern are distinctly rare and have not been well characterized. A small subset of mesotheliomas (mostly in children and young adults) are characterized by gene fusions including EWSR1/FUS::ATF1, EWSR1::YY1, and NTRK and ALK rearrangements, and often have epithelioid morphology. Herein, we describe five peritoneal mesothelial neoplasms (identified via molecular screening of seven histologically similar tumors) that are pure adenomatoid/microcystic in morphology and unified by the presence of an NR4A3 fusion. Patients were three males and two females aged 31-70 years (median, 40 years). Three presented with multifocal/diffuse and two with a localized disease. The size of the individual lesions ranged from 1.5 to 8 cm (median, 4.7). The unifocal lesions originated in the small bowel mesentery and the mesosigmoid. Treatment included surgery, either alone (three) or combined with hyperthermic intraperitoneal chemotherapy (two), and neoadjuvant or adjuvant chemotherapy (one case each). At the last follow-up (6-13 months), all five patients were alive and disease-free. All tumors were morphologically similar, characterized by extensive sieve-like microcystic growth with bland-looking flattened cells lining variably sized microcystic spaces and lacked a conventional epithelioid or sarcomatoid component. Immunohistochemistry confirmed mesothelial differentiation, but most cases showed limited expression of D2-40 and calretinin. Targeted RNA sequencing revealed an NR4A3 fusion (fusion partners were EWSR1 in three cases and CITED2 and NIPBL in one case each). The nosology and behavior of this morphomolecularly defined novel peritoneal mesothelial neoplasm of uncertain biological potential and its distinction from adenomatoid variants of conventional mesothelioma merit further delineation as more cases become recognized.

Salivary carcinosarcoma: insight into multistep pathogenesis indicates uniform origin as sarcomatoid variant of carcinoma ex pleomorphic adenoma with frequent heterologous elements.

AIMS: The formal pathogenesis of salivary carcinosarcoma (SCS) remained unclear, both with respect to the hypothetical development from either preexisting pleomorphic adenoma (PA) or de novo and the clonal relationship between highly heterogeneous carcinomatous and sarcomatous components. METHODS AND RESULTS: We performed clinicopathological and molecular (targeted RNA sequencing) analyses on a large series of 16 cases and combined this with a comprehensive literature search (111 cases). Extensive sampling (average 11.6 blocks), combined with immunohistochemistry and molecular studies (PA-specific translocations including PLAG1 or HMGA2 proven in 6/16 cases), enabled the morphogenetic identification of PA in 15/16 cases (93.8%), by far surpassing a reported rate of 49.6%. Furthermore, we demonstrated a multistep (intraductal/intracapsular/extracapsular) adenoma-carcinoma-sarcoma-progression, based on two alternative histogenetic pathways (intraductal, 56.3%, versus myoepithelial pathway, 37.5%). Thereby, early intracapsular stages are identical to conventional carcinoma ex PA, while later extracapsular stages are dominated by secondary, frequently heterologous sarcomatous transformation with often large tumour size (>60 mm). CONCLUSION: Our findings strongly indicate that SCS (almost) always develops from PA, with a complex multistep adenoma-carcinoma-sarcoma-sequence, based on two alternative histogenetic pathways. The findings from this novel approach strongly suggest that SCS pathogenetically is a rare (3-6%), unique, and aggressive variant of carcinoma ex PA with secondary sarcomatous overgrowth. In analogy to changes of terminology in other organs, the term "sarcomatoid carcinoma ex PA with/without heterologous elements" might be more appropriate.

Expression patterns and prognostic relevance of subtype-specific transcription factors in surgically resected small-cell lung cancer: an international multicenter study.

The tissue distribution and prognostic relevance of subtype-specific proteins (ASCL1, NEUROD1, POU2F3, YAP1) present an evolving area of research in small-cell lung cancer (SCLC). The expression of subtype-specific transcription factors and P53 and RB1 proteins were measured by immunohistochemistry (IHC) in 386 surgically resected SCLC samples. Correlations between subtype-specific proteins and in vitro efficacy of various therapeutic agents were investigated by proteomics and cell viability assays in 26 human SCLC cell lines. Besides SCLC-A (ASCL1-dominant), SCLC-AN (combined ASCL1/NEUROD1), SCLC-N (NEUROD1-dominant), and SCLC-P (POU2F3-dominant), IHC and cluster analyses identified a quadruple-negative SCLC subtype (SCLC-QN). No unique YAP1-subtype was found. The highest overall survival rates were associated with non-neuroendocrine subtypes (SCLC-P and SCLC-QN) and the lowest with neuroendocrine subtypes (SCLC-A, SCLC-N, SCLC-AN). In univariate analyses, high ASCL1 expression was associated with poor prognosis and high POU2F3 expression with good prognosis. Notably, high ASCL1 expression influenced survival outcomes independently of other variables in a multivariate model. High POU2F3 and YAP1 protein abundances correlated with sensitivity and resistance to standard-of-care chemotherapeutics, respectively. Specific correlation patterns were also found between the efficacy of targeted agents and subtype-specific protein abundances. In conclusion, we investigated the clinicopathological relevance of SCLC molecular subtypes in a large cohort of surgically resected specimens. Differential IHC expression of ASCL1, NEUROD1, and POU2F3 defines SCLC subtypes. No YAP1-subtype can be distinguished by IHC. High POU2F3 expression is associated with improved survival in a univariate analysis, whereas elevated ASCL1 expression is an independent negative prognosticator. Proteomic and cell viability assays of human SCLC cell lines revealed distinct vulnerability profiles defined by transcription regulators. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Impairment of the NKT-STAT1-CXCL9 Axis Contributes to Vessel Fibrosis in Pulmonary Hypertension Caused by Lung Fibrosis.

Rationale: Pulmonary hypertension (PH) is a common, severe comorbidity in interstitial lung diseases such as pulmonary fibrosis (PF), and it has limited treatment options. Excessive vascular fibrosis and inflammation are often present in PH, but the underlying mechanisms are still not well understood. Objectives: To identify a novel functional link between natural killer T (NKT) cell activation and vascular fibrosis in PF-PH. Methods: Multicolor flow cytometry, secretome, and immunohistological analyses were complemented by pharmacological NKT cell activation in vivo, in vitro, and ex vivo. Measurements and Main Results: In pulmonary vessels of patients with PF-PH, increased collagen deposition was linked to a local NKT cell deficiency and decreased IL-15 concentrations. In a mouse model of PH caused by lung fibrosis, pharmacological NKT cell activation using a synthetic α-galactosylceramide analog (KRN7000) restored local NKT cell numbers and ameliorated vascular remodeling and right ventricular systolic pressure. Supplementation with activated NKT cells reduced collagen deposition in isolated human pulmonary arterial smooth muscle cells (hPASMCs) and in ex vivo precision-cut lung slices of patients with end-stage PF-PH. Coculture with activated NKT cells induced STAT1 signaling in hPASMCs. Secretome analysis of peripheral blood mononuclear cells identified CXCL9 and CXCL10 as indicators of NKT cell activation. Pharmacologically, CXCL9, but not CXCL10, potently inhibited collagen deposition in hPASMCs via the chemokine receptor CXCR3. Conclusions: Our results indicate that the absence of NKT cells impairs the STAT1-CXCL9-CXCR3 axis in PF-PH and that restoration of this axis by NKT cell activation may unravel a novel therapeutic strategy to target vascular fibrosis in interstitial lung disease.

Cancer-Associated Fibroblasts Influence Survival in Pleural Mesothelioma: Digital Gene Expression Analysis and Supervised Machine Learning Model.

The exact mechanism of desmoplastic stromal reaction (DSR) formation is still unclear. The interaction between cancer cells and cancer-associated fibroblasts (CAFs) has an important role in tumor progression, while stromal changes are a poor prognostic factor in pleural mesothelioma (PM). We aimed to assess the impact of CAFs paracrine signaling within the tumor microenvironment and the DSR presence on survival, in a cohort of 77 PM patients. DSR formation was evaluated morphologically and by immunohistochemistry for Fibroblast activation protein alpha (FAP). Digital gene expression was analyzed using a custom-designed CodeSet (NanoString). Decision-tree-based analysis using the "conditional inference tree" (CIT) machine learning algorithm was performed on the obtained results. A significant association between FAP gene expression levels and the appearance of DSR was found (p = 0.025). DSR-high samples demonstrated a statistically significant prolonged median survival time. The elevated expression of MYT1, KDR, PIK3R1, PIK3R4, and SOS1 was associated with shortened OS, whereas the upregulation of VEGFC, FAP, and CDK4 was associated with prolonged OS. CIT revealed a three-tier system based on FAP, NF1, and RPTOR expressions. We could outline the prognostic value of CAFs-induced PI3K signaling pathway activation together with FAP-dependent CDK4 mediated cell cycle progression in PM, where prognostic and predictive biomarkers are urgently needed to introduce new therapeutic strategies.

Expression of phosphorylated ribosomal protein S6 in mesothelioma patients - correlation with clinico-pathological characteristics and outcome: results from the European Thoracic Oncology Platform (ETOP) Mesoscape project.

Pleural mesothelioma (PM) is an aggressive malignancy with poor prognosis. Although histology and pathologic stage are important prognostic factors, better prognostic biomarkers are needed. The ribosomal protein S6 is a downstream target of the phosphatidylinositol 3-kinase (PI3K) pathway involved in protein synthesis and cell proliferation. In previous studies, low phosphorylated S6 (pS6) immunoreactivity was significantly correlated with longer progression-free survival (PFS) and overall survival (OS) in PM patients. We aimed to correlate pS6 expression to clinical data in a large multi-centre PM cohort as part of the European Thoracic Oncology Platform (ETOP) Mesoscape project. Tissue Micro Arrays (TMAs) of PM were constructed and expression of pS6 was evaluated by a semi-quantitatively aggregate H-score. Expression results were correlated to patient characteristics as well as OS/PFS. pS6 IHC results of 364 patients from 9 centres, diagnosed between 1999 and 2017 were available. The primary histology of included tumours was epithelioid (70.3%), followed by biphasic (24.2%) and sarcomatoid (5.5%). TMAs included both treatment-naïve and tumour tissue taken after induction chemotherapy. High pS6 expression (181 patients with H-score>1.41) was significantly associated with less complete resection. In the overall cohort, OS/PFS were not significantly different between pS6-low and pS6-high patients. In a subgroup analysis non-epithelioid (biphasic and sarcomatoid) patients with high pS6 expression showed a significantly shorter OS (p < 0.001, 10.7 versus 16.9 months) and PFS (p < 0.001, 6.2 versus 10.8 months). In subgroup analysis, in non-epithelioid PM patients high pS6 expression was associated with significantly shorter OS and PFS. These exploratory findings suggest a clinically relevant PI3K pathway activation in non-epithelioid PM which might lay the foundation for future targeted treatment strategies.

Abnormal pulmonary hemodynamics during exercise is associated with exercise capacity in COPD.

BACKGROUND: Pulmonary hypertension (PH) is a frequent complication in COPD and it is associated with decreased exercise capacity and poor prognosis. We hypothesized that even in COPD patients without significant PH at rest, abnormal pulmonary hemodynamics during exercise affect exercise capacity. METHODS: Consecutive COPD patients with clinically indicated right heart catheterization and resting mean pulmonary arterial pressure (mPAP) < 25 mmHg and age- and sex-matched controls with the same limits of pulmonary hemodynamics but no chronic lung disease who underwent clinical work-up including invasive hemodynamic assessment during exercise, were retrospectively analyzed. Chi-square tests were used to evaluate differences between groups for categorical data and Fisher's exact test or Mann-Whitney-U-tests for continuous variables. Associations were analyzed with Spearman rank correlation tests. RESULTS: We included n = 26 COPD patients (female/male: 16/10, 66 ± 11 yr, FEV1: 56 ± 25%predicted) and n = 26 matched controls (FEV1: 96 ± 22%predicted). At rest, COPD patients presented with slightly increased mPAP (21 (18-23) vs. 17 (14-20) mmHg, p = 0.022), and pulmonary vascular resistance (PVR) [2.5 (1.9-3.0) vs. 1.9 (1.5-2.4) WU, p = 0.020] as compared to controls. During exercise, COPD patients reached significantly higher mPAP [47 (40-52) vs. 38 (32-44) mmHg, p = 0.015] and PVR [3.1 (2.2-3.7) vs. 1.7 (1.1-2.9) WU, p = 0.028] values despite lower peak exercise level [50 (50-75) vs. 100 (75-125) Watt, p = 0.002]. The mPAP/cardiac output slope was increased in COPD vs. controls [6.9 (5.5-10.9) vs. 3.7 (2.4-7.4) mmHg/L/min, p = 0.007] and negatively correlated with both peak oxygen uptake (r = - 0.46, p = 0.007) and 6-min walk distance (r = - 0.46, p = 0.001). CONCLUSION: Even in the absence of significant PH at rest, COPD patients reveal characteristic abnormalities in pulmonary hemodynamics during exercise, which may represent an important exercise-limiting factor.

The impact of COVID-19 on oral squamous cell carcinoma's diagnostic stage-A retrospective study.

OBJECTIVE: The COVID-19 pandemic has had an impact on patients' access to primary care services. A timely diagnosis of oral squamosa cell carcinoma is paramount. This study aims to quantify the pandemic's effect on tumor volume at the time of initial diagnosis. MATERIALS AND METHODS: In a retrospective study, all primarily diagnosed cancer patients between March 2018 and March 2022 were compiled; the TNM stage and the tumor volume were evaluated. Tumor volumes were calculated using pathology or radiology reports. RESULTS: In total, 162 newly diagnosed tumor cases were included in this study. Of these, 76 (46.9%) cases were allocated in the "pre-COVID-19" group and 86 (53.1%) in the "COVID-19" group. Patients diagnosed during the "COVID-19" period showed a significantly advanced T stage (p < 0.001) and larger mean tumor volumes (53.16 ± 73.55 cm3 vs. 39.89 ± 102.42 cm3 ; p = 0.002) when compared to the control group. CONCLUSION: Tumor volume and T stage were significantly advanced in the "COVID-19" group when compared to prepandemic data. We hypothesize that the postponement of routine dental check-ups may explain this finding. Hence, dentists and primary care providers are encouraged to place particular emphasis on screening during routine check-ups.

Lung fibrosis in autoimmune diseases and hypersensitivity: how to separate these from idiopathic pulmonary fibrosis.

Lung involvement in autoimmune diseases (AID) is uncommon, but may precede other organ manifestations. A diagnostic problem is chronicity presenting with lung fibrosis. A new category of interstitial pneumonia with autoimmune features for patients with clinical symptoms of AID and presenting with usual interstitial pneumonia (UIP) enables antifibrotic treatment for these patients. Hypersensitivity pneumonia (HP) and other forms of lung fibrosis were not included into this category. As these diseases based on adverse immune reactions often present with unspecific clinical symptoms, a specified pathological diagnosis will assist the clinical evaluation. We aimed to establish etiology-relevant differences of patterns associated with AID or HP combined with lung fibrosis. We retrospectively evaluated 51 cases of AID, and 29 cases of HP with lung fibrosis, and compared these to 24 cases of idiopathic pulmonary fibrosis (UIP/IPF). Subacute AID and HP most often presented with organizing pneumonia (OP), whereas chronicity was associated with UIP. Unspecified fibrosis was seen in a few cases, whereas NSIP pattern was rare. In 9 cases, the underlying etiology could not be defined. Statistically significant features differentiating chronic AID or HP from UIP/IPF are lymphocytic infiltrations into myofibroblastic/fibroblastic foci. Other features significantly associated with AID and HP were granulomas, isolated Langhans giant cells, and protein deposits, but seen in only a minority of cases. A combination of UIP with one of these features enabled a specific etiology-based diagnosis. Besides the antifibrotic drug regimen, additional therapies might be considered.

State-of-the-art cytology of pleural fluid, focusing on the diagnosis of mesothelioma.

Mesothelioma has always been a challenging diagnosis to render in body cavity cytology samples. This review is a timely update on pleural fluid cytology and ancillary studies that should be considered in the diagnosis of mesothelial proliferations, specifically mesotheliomas. Information about new diagnostic approaches and ancillary studies in mesothelioma was obtained from the peer-reviewed literature and the authors' experiences. Although the morphological diagnosis of mesothelioma is fraught with numerous challenges given the overlap with other diagnostic entities, there are a variety of immunohistochemical and fluorescence in situ hybridization studies available to help in determining mesothelial origin and in distinguishing malignant proliferations from the more common benign or reactive mesothelial proliferations. Although ancillary studies can be helpful, there are important pitfalls to be aware of when interpreting these cases, and this review highlights some of the challenges that require caution.

Cancer-Associated Fibroblasts Regulate Kinase Activity in Mesothelioma Cell Lines via Paracrine Signaling and Thereby Dictate Cell Faith and Behavior.

BACKGROUND: Malignant pleural mesothelioma (MPM) has an infaust prognosis due to resistance to systemic treatment with platin-analoga. MPM cells modulate the immune response to their benefit. They release proinflammatory cytokines, such as TGF-ß, awakening resting fibrocytes that switch their phenotype into activated fibroblasts. Signaling interactions between cancer cells and cancer-associated fibroblasts (CAFs) play an integral part in tumor progression. This study aimed to investigate the role CAFs play in MPM progression, analyzing the impact this complex, symbiotic interaction has on kinase-related cell signaling in vitro. METHODS: We simulated paracrine signaling in vitro by treating MPM cell lines with conditioned medium (CM) from fibroblasts (FB) and vice versa. NCI-H2052, MSTO-211H, and NCI-H2452 cell lines representing the three mayor MPM subtypes, while embryonal myofibroblast cell lines, IMR-90 and MRC-5, provide a CAFs-like phenotype. Subsequently, differences in proliferation rates, migratory behavior, apoptosis, necrosis, and viability were used as covariates for data analysis. Kinase activity of treated samples and corresponding controls were then analyzed using the PamStation12 platform (PamGene); Results: Treatment with myofibroblast-derived CM revealed significant changes in phosphorylation patterns in MPM cell lines. The observed effect differs strongly between the analyzed MPM cell lines and depends on the origin of CM. Overall, a much stronger effect was observed using CM derived from IMR-90 than MRC-5. The phosphorylation changes mainly affected the MAPK signaling pathway.; Conclusions: The factors secreted by myofibroblasts in fibroblasts CM significantly influence the phosphorylation of kinases, mainly affecting the MAPK signaling cascade in tested MPM cell lines. Our in vitro results indicate promising therapeutic effects by the use of MEK or ERK inhibitors and might have synergistic effects in combination with cisplatin-based treatment, improving clinical outcomes for MPM patients.

[Predictive immunocytochemistry in non-small cell lung carcinoma]. / Prädiktive Immunzytochemie beim nicht-kleinzelligen Lungenkarzinom.

Predictive immunochemistry is a time-, tumor sample- and cost-efficient method for testing the increasing number of predictive biomarkers in advanced non-small cell lung cancer (NSCLC). Immunohistochemistry (IHC) on formalin-fixed, paraffin-embedded (FFPE) tumor tissue has an established role in detecting PD-L1 expression and in ALK, ROS1, and more recently NTRK testing. Cytology specimens as a source for predictive biomarker testing in NSCLC is very important as up to 40% of all NSCLC are diagnosed by cytology alone.Despite the established role of cytology in lung cancer diagnosis, no commercial IHC assays have been validated for cytology specimens.FFPE cell blocks (CB) are the most straightforward cytology preparation for predictive immunocytochemistry (ICC) as the results are valid using protocols standardized for FFPE histology. But CB are not always available.With non-CB cytology specimens being less standardized than FFPE histology and with considerable preanalytical variability, rigorous cytology-specific ICC protocol optimization, validation, and quality control are required. With this prerequisite, predictive ICC, most commonly performed on Papanicolaou-stained cytology specimens, is robust and reliable on non-CB preparations. This valuable material should not be underutilized for predictive biomarker testing, as this would put patients at risk of unnecessary repeat sampling. This review highlights preanalytical, analytical, and postanalytical aspects that may influence ICC results and summarizes the published data on predictive ICC for PD-L1, ALK, and ROS1 in NSCLC.

Malignant peritoneal mesothelioma: prognostic significance of clinical and pathologic parameters and validation of a nuclear-grading system in a multi-institutional series of 225 cases.

Malignant peritoneal mesothelioma historically carried a grim prognosis, but outcomes have improved substantially in recent decades. The prognostic significance of clinical, morphologic, and immunophenotypic features remains ill-defined. This multi-institutional cohort comprises 225 malignant peritoneal mesotheliomas, which were assessed for 21 clinical, morphologic, and immunohistochemical parameters. For epithelioid mesotheliomas, combining nuclear pleomorphism and mitotic index yielded a composite nuclear grade, using a previously standardized grading system. Correlation of clinical, morphologic, and immunohistochemical parameters with overall and disease-free survival was examined by univariate and multivariate analyses. On univariate analysis, longer overall survival was significantly associated with diagnosis after 2000 (P = 0.0001), age <60 years (P = 0.0001), ECOG performance status 0 or 1 (P = 0.01), absence of radiographic lymph-node metastasis (P = 0.04), cytoreduction surgery (P < 0.0001), hyperthermic intraperitoneal chemotherapy (P = 0.0001), peritoneal carcinomatosis index <27 (P = 0.01), absence of necrosis (P = 0.007), and epithelioid histotype (P < 0.0001). Among epithelioid malignant mesotheliomas only, longer overall survival was further associated with female sex (P = 0.03), tubulopapillary architecture (P = 0.005), low nuclear pleomorphism (P < 0.0001), low mitotic index (P = 0.0007), and low composite nuclear grade (P < 0.0001). On multivariate analyses, the low composite nuclear grade was independently associated with longer overall and disease-free survival (P < 0.0001). Our data further clarify the interactions of clinical and pathologic features in peritoneal mesothelioma prognosis and validate the prognostic significance of a standardized nuclear-grading system in epithelioid malignant mesothelioma of the peritoneum.

Proteomic Analysis of Vocal Fold Fibroblasts Exposed to Cigarette Smoke Extract: Exploring the Pathophysiology of Reinke's Edema.

Reinke's edema is a smoking-associated, benign, mostly bilateral lesion of the vocal folds leading to difficulties in breathing and voice problems. Pronounced histological changes such as damaged microvessels or immune cell infiltration have been described in the vocal fold connective tissue, the lamina propria Thus, vocal fold fibroblasts, the main cell type of the lamina propria, have been postulated to play a critical role in disease mediation. Yet information about the pathophysiology is still scarce and treatment is only surgical, i.e. symptomatic. To explore the pathophysiology of Reinke's edema, we exposed near-primary human vocal fold fibroblasts to medium conditioned with cigarette smoke extract for 24 h as well as 4 days followed by quantitative mass spectrometry.Proteomic analyses after 24 h revealed that cigarette smoke increased proteins previously described to be involved in oxidative stress responses in other contexts. Correspondingly, gene sets linked to metabolism of xenobiotics and reactive oxygen species were significantly enriched among cigarette smoke-induced proteins. Among the proteins most downregulated by cigarette smoke, we identified fibrillar collagens COL1A1 and COL1A2; this reduction was validated by complementary methods. Further, we found a significant increase of UDP-glucose 6-dehydrogenase, generating a building block for biosynthesis of hyaluronan, another crucial component of the vocal fold lamina propria In line with this result, hyaluronan levels were significantly increased because of cigarette smoke exposure. Long term treatment of 4 days did not lead to significant changes.The current findings corroborate previous studies but also reveal new insights in possible disease mechanisms of Reinke's edema. We postulate that changes in the composition of the vocal folds' extracellular matrix -reduction of collagen fibrils, increase of hyaluronan- may lead to the clinical findings. This might ease the identification of better, disease-specific treatment options.