DNA methylation-based machine learning classification distinguishes pleural mesothelioma from chronic pleuritis, pleural carcinosis, and pleomorphic lung carcinomas.

OBJECTIVES: Our goal was to evaluate the diagnostic value of DNA methylation analysis in combination with machine learning to differentiate pleural mesothelioma (PM) from important histopathological mimics. MATERIAL AND METHODS: DNA methylation data of PM, lung adenocarcinomas, lung squamous cell carcinomas and chronic pleuritis was used to train a random forest as well as a support vector machine. These classifiers were validated using an independent validation cohort including pleural carcinosis and pleomorphic variants of lung adeno- and squamous cell carcinomas. Furthermore, we performed differential methylation analysis and used a deconvolution method to estimate the composition of the tumor microenvironment. RESULTS: T-distributed stochastic neighbor embedding clearly separated PM from lung adenocarcinomas and squamous cell carcinomas, but there was a considerable overlap between chronic pleuritis specimens and PM with low tumor cell content. In a nested cross validation on the training cohort, both machine learning algorithms achieved the same accuracies (94.8%). On the validation cohort, we observed high accuracies for the support vector machine (97.8%) while the random forest performed considerably worse (89.5%), especially in distinguishing PM from chronic pleuritis. Differential methylation analysis revealed promoter hypermethylation in PM specimens, including the tumor suppressor genes BCL11B, EBF1, FOXA1, and WNK2. Deconvolution of the stromal and immune cell composition revealed higher rates of regulatory T-cells and endothelial cells in tumor specimens and a heterogenous inflammation including macrophages, B-cells and natural killer cells in chronic pleuritis. CONCLUSION: DNA methylation in combination with machine learning classifiers is a promising tool to reliably differentiate PM from chronic pleuritis and lung cancer, including pleomorphic carcinomas. Furthermore, our study highlights new candidate genes for PM carcinogenesis and shows that deconvolution of DNA methylation data can provide reasonable insights into the composition of the tumor microenvironment.

Genome-wide association study of lung lesions and pleurisy in New Zealand lambs.

Pneumonia is an important issue for sheep production, leading to reduced growth rate and a predisposition to pleurisy. The objective of this study was to identify loci associated with pneumonic lesions and pleurisy in New Zealand progeny test lambs. The lungs from 3,572 progeny-test lambs were scored for presence and severity of pneumonic lesions and pleurisy at slaughter. Animals were genotyped using the Illumina Ovine Infinium HD SNP BeadChip (606,006 markers). The heritability of lung lesion score and pleurisy were calculated using the genomic relationship matrix, and genome-wide association analyses were conducted using EMMAX and haplotype trend regression. At slaughter, 35% of lambs had pneumonic lesions, with 9% showing lesions on more than half of any individual lobe. The number of lambs recorded as having pleurisy by the processing plants was 9%. Heritability estimates for pneumonic lesions and pleurisy scores adjusted for heteroscedasticity (CPSa and PLEURa) were 0.16 (± 0.03) and 0.05 (± 0.02), respectively. Five single-nucleotide polymorphisms (SNPs) were significantly associated with pneumonic lesions at the genome-wide level, and additional 37 SNPs were suggestively significant. Four SNPs were significantly associated with pleurisy, with an additional 11 SNPs reaching the suggestive level of significance. There were no regions that overlapped between the 2 traits. Multiple SNPs were in regions that contained genes involved in either the DNA damage response or the innate immune response, including several that had previously been reported to have associations with respiratory disease. Both EMMAX and HTR analyses of pleurisy data showed a significant peak on chromosome 2, located downstream from the transcription factor SP3. SP3 activates or suppresses the expression of numerous genes, including several genes with known functions in the immune system. This study identified several SNPs associated with genes involved in both the innate immune response and the response to DNA damage that are associated with pneumonic lesions and pleurisy in lambs at slaughter. Additionally, the identification in sheep of several SNPs within genes that have previously been associated with the respiratory system in cattle, pigs, rats, and mice indicates that there may be common pathways that underlie the response to invasion by respiratory pathogens in multiple species.

Pulmonary and pleural inflammation after intratracheal instillation of short single-walled and multi-walled carbon nanotubes.

Relationships between the physical properties of carbon nanotubes (CNTs) and their toxicities have been studied. However, little research has been conducted to investigate the pulmonary and pleural inflammation caused by short-fiber single-walled CNTs (SWCNTs) and multi-walled CNTs (MWCNTs). This study was performed to characterize differences in rat pulmonary and pleural inflammation caused by intratracheal instillation with doses of 0.15 or 1.5mg/kg of either short-sized SWCNTs or MWCNTs. Data from bronchoalveolar lavage fluid analysis, histopathological findings, and transcriptional profiling of rat lungs obtained over a 90-day period indicated that short SWCNTs caused persistent pulmonary inflammation. In addition, the short MWCNTs markedly impacted alveoli immediately after instillation, with the levels of pulmonary inflammation following MWCNT instillation being reduced in a time-dependent manner. MWCNT instillation induced greater levels of pleural inflammation than did short SWCNTs. SWCNTs and MWCNTs translocated in mediastinal lymph nodes were observed, suggesting that SWCNTs and MWCNTs underwent lymphatic drainage to the mediastinal lymph nodes after pleural penetration. Our results suggest that short SWCNTs and MWCNTs induced pulmonary and pleural inflammation and that they might be transported throughout the body after intratracheal instillation. The extent of changes in inflammation differed following SWCNT and MWCNT instillation in a time-dependent manner.

Grape seed proanthocyanidin extract protects against carrageenan-induced lung inflammation in mice through reduction of pro-inflammatory markers and chemokine expressions.

Proanthocyanidins are the most abundant phenolic compounds and have been reported to exert anti-inflammatory actions. The aim of this study was to investigate the effects of grape seed proanthocyanidin extract (GSPE) in a mouse model of carrageenan-induced pleurisy. Following the induction of pleurisy using λ-carrageenan (Cg, 1 %), GSPE (25, 50 and 100 mg/kg) was administered per-oral (p.o.), and the glucocorticoid-induced tumour necrosis factor receptor (GITR), IL-17A expressing cells and other markers, such as cytokines (Th1/Th2 and Th17), were studied. We evaluate the effects of GSPE on the mRNA expression of pro-inflammatory and anti-inflammatory mediators. The results illustrated that the cell numbers of IL-17A and GITR expressing cells and the cytokine levels in Th1/Th17 cells were markedly increased in the Cg-group, whereas the cytokines produced by Th2 cells were significantly decreased in the same group. Treatment with GSPE reversed these effects. Histological examinations revealed anti-inflammatory effects of GSPE.

Systemic administration of Rosmarinus officinalis attenuates the inflammatory response induced by carrageenan in the mouse model of pleurisy.

Rosmarinus officinalis, also named rosemary, is a native plant from the Mediterranean region that is useful for the treatment of inflammatory diseases. Studies using experimental models and/or in vitro tests have shown the important biological effects of rosemary. In this context, the mechanism of the anti-inflammatory activity of rosemary must be investigated to support the discovery of new substances with anti-inflammatory effects. The aim of the present study was to investigate the anti-inflammatory effects of crude extract oil free obtained from the leaves of rosemary in an animal model of inflammation, thus evaluating its medicinal use for the treatment of inflammatory conditions. Also its ethanol, hexane, and ethyl acetate fractions, as well as its isolated compounds carnosol and rosmarinic acid were analyzed. Swiss mice were used for the in vivo experiments. The effect of this herb on the inhibition of the leukocytes, exudation, myeloperoxidase, and adenosine-deaminase activities, nitrite/nitrate, interleukin 17A, and interleukin 10 levels and mRNA expression was determined. The crude extract and its derived fractions, in addition to its isolated compounds, inhibited leukocytes and decreased exudation and myeloperoxidase and adenosine-deaminase activities, as well as nitrite/nitrate and interleukin 17A levels and mRNA expression, besides increasing interleukin 10 levels and mRNA expression. Rosemary showed important anti-inflammatory activity by inhibiting leukocytes and decreasing exudation. These effects were associated with a decrease in the proinflammatory parameters (myeloperoxidase, adenosine-deaminase, nitrite/nitrate, and interleukin 17A) and an increase in the anti-inflammatory cytokine (interleukin 10). This study confirms the anti-inflammatory properties of rosemary and validates its use in folk medicine to treat inflammatory diseases such as rheumatism and asthma.

Diagnostic usefulness of p16/CDKN2A FISH in distinguishing between sarcomatoid mesothelioma and fibrous pleuritis.

The distinction between sarcomatoid mesothelioma and fibrous pleuritis is difficult based on histology, especially when the amount of tumor tissue examined via biopsy is small and immunohistochemical examination is inconclusive. We studied the usefulness of deletion of p16 with fluorescence in situ hybridization (FISH) and p16 hypermethylation with polymerase chain reaction for the diagnosis and prognosis of malignant pleural mesothelioma (MPM). We analyzed 50 MPMs, including 22 sarcomatoid mesothelioma cases and 10 fibrous pleuritis cases. We set the cutoff value of homozygous deletion pattern as 14.4% based on FISH signaling patterns using samples of fibrous pleuritis. The percentage of homozygous deletion pattern was higher than 14.4% in 55.6% of the epithelioid mesotheliomas (10/18) and in all of the sarcomatoid mesotheliomas (22/22). Methylation of p16 was observed in 7 (20.6%) of 34 informative cases. p16 FISH analysis can be a reliable test for distinguishing between sarcomatoid mesothelioma and fibrous pleuritis and a prognostic factor for MPM.

Genomic regions associated with ventro-cranial chronic pleuritis in pig.

Ventro-cranial chronic pleuritis can be a result of pleuropneumonia and enzootic pneumonia. These diseases cause severe losses in intensive pig production worldwide, but host resistance is difficult to breed for. It could be beneficial to use marker-assisted selection, and a step towards this is to identify genomic regions associated with the trait. For this purpose, 7304 pigs from 11 boar families were analysed for associations between single nucleotide polymorphisms and ventro-cranial chronic pleuritis. The pigs were genotyped by the use of the iSelect Custom 7 K porcine SNP Chip. Quantitative trait loci (QTL), significant at the chromosome-wide level, were identified on Sus scrofa chromosomes (SSC) 2, 4, 11, 12 and 13 in four different boar families. The QTL on SSC 4 in family G was also significant at the genome-wide threshold according to Bonferroni correction. We have identified a number of candidate genes, but the causative mutations still need to be identified. Markers closely associated with the resistance traits have a strong potential for use in breeding towards animals with improved characteristics concerning ventro-cranial chronic pleuritis.

[Periodic fever with increased IgD]. / Fièvre périodique avec élévation des immunoglobulines D.

Periodic fever or hereditary inflammatory fevers are characterized by intermittent inflammatory attacks. Many entities are well recognized today such as familial mediterranean fever (FMF) and hyperimmunoglobulinemia D syndrome (HIDS). We report on the case of a 6-year-old boy referred for evaluation of a recurrent fever associated with chest pain, pneumonitis, or pleuritis since the age of 5 years. Laboratory data showed leukocytosis, a high erythrocyte sedimentation rate, and C-reactive protein; however, a permanent high serum level IgD was noted. Stereotypical episodes of fever appeared every 4-6 weeks, while infectious, malignant, and auto-immune causes were eliminated. A search for the most common mutations of the FMF gene in Tunisian patients (M694V, M680I, V726A, E148Q, M694I, and A744S) were negative. Likewise, urinary leukotriene E(4), which may be increased in HIDS, was normal in this patient. Mevalonate kinase activity in lymphocytes was not assayed. Ethnic origin and clinical presentation suggest FMF with an increased IgD rather than authentic HIDS, in spite of the lack of improvement under colchicine treatment and the negativity of the main mutations involved in FMF.

[Thoracic impairment from familial Mediterranean fever: review of the literature and a case study]. / Atteintes thoraciques de la fièvre méditerranéenne familiale : revue de la littérature - à propos d'un cas.

Familial Mediterranean fever (FMF) is a recessive autosomal disease, predominantly affecting the population around the Mediterranean. The main clinical signs consist of attacks of fever associated with abdominal, articular and thoracic pain. Based on a case report, the authors describe the main thoracic forms of this illness comprising pleural pain, pleural effusion and pulmonary amyloidosis. The authors also discuss the association of mesothelioma and FMF. Colchicine is successfully used in the treatment of FMF.

Proinflammatory role of glucocorticoid-induced TNF receptor-related gene in acute lung inflammation.

Glucocorticoid-induced TNFR-related gene (GITR) participates in the immune/inflammatory response. Because GITR expression has been described in cells other than T lymphocytes, we investigated whether it also modulates acute inflammatory response. Using GITR-deficient (GITR(-/-)) mice, we analyzed the role of GITR in the development of carrageenan-induced lung inflammation (pleurisy) by studying several proinflammatory markers 2-8 h after carrageenan injection. When compared with GITR(+/+), GITR(-/-) mice exhibited decreased production of turbid exudate containing a lower number of leukocytes. This was correlated with the reduction of inflammatory markers (including TNF-alpha, IL-1beta, myeloperoxidase, inducible NO synthase, and cyclooxygenase 2) in the pleural exudate and/or in the lung. Moreover, endothelial cells expressed lower levels of adhesion molecules. In lungs of GITR(+/+) mice, GITR ligand expression was not modulated during pleurisy, while that of GITR increased, as a consequence of increased infiltration by GITR-expressing cells and of GITR up-regulation in macrophages and endothelial cells. Finally, cotreatment of GITR(+/+) mice with carrageenan and Fc-GITR fusion protein decreased the number of inflammatory cells (pleural macrophages and lung neutrophils) as compared with carrageenan treatment alone, confirming that GITR plays a role in the modulation of pleurisy.

Role of monocyte chemotactic protein-1/CC chemokine ligand 2 on gamma delta T lymphocyte trafficking during inflammation induced by lipopolysaccharide or Mycobacterium bovis bacille Calmette-Guérin.

Gammadelta T lymphocytes are involved in a great variety of inflammatory and infectious responses. However, the mechanisms by which gammadelta T lymphocytes migrate to inflamed sites are poorly understood. In this study we investigate the role of monocyte chemotactic protein (MCP)-1 in regulating gammadelta T cell migration after LPS or Mycobacterium bovis bacille Calmette-Guérin (BCG) challenge. LPS-induced gammadelta T cell influx was significantly inhibited by either pretreatment with dexamethasone or vaccinia virus Lister 35-kDa chemokine binding protein, vCKBP, a CC chemokine neutralizing protein, suggesting a role for CC chemokines in this phenomenon. LPS stimulation increased the expression of MCP-1 mRNA and protein at the inflammation site within 6 h. It is noteworthy that LPS was unable to increase MCP-1 production or gammadelta T cell recruitment in C3H/HeJ, indicative of the involvement of Toll-like receptor 4. Gammadelta T cells express MCP-1 receptor CCR2. Pretreatment with anti-MCP-1 mAb drastically inhibited LPS-induced in vivo gammadelta T cell mobilization. Indeed, MCP-1 knockout mice were unable to recruit gammadelta T cells to the pleural cavity after LPS stimulation, effect that could be restored by coadministration of MCP-1. In addition, BCG-induced gammadelta lymphocyte accumulation was significantly reduced in MCP-1 knockout mice when compared with wild-type mice. In conclusion, our results indicate that LPS-induced gammadelta T lymphocyte migration is dependent on Toll-like receptor 4 and sensitive to both dexamethasone and CC chemokine-binding protein inhibition. Moreover, by using MCP-1 neutralizing Abs and genetically deficient mice we show that LPS- and BCG-induced gammadelta T lymphocyte influx to the pleural cavity of mice is mainly orchestrated by the CC chemokine MCP-1.

Role of IL-6 in the pleurisy and lung injury caused by carrageenan.

In the present study we used IL-6 knockout mice (IL-6KO) to evaluate the role of IL-6 in the inflammatory response caused by injection of carrageenan into the pleural space. Compared with carrageenan-treated IL-6 wild-type (IL-6WT) mice, carrageenan-treated IL-6KO mice exhibited a reduced degree of pleural exudation and polymorphonuclear cell migration. Lung myeloperoxidase activity and lipid peroxidation were significantly reduced in IL-6KO mice compared with those in IL-6WT mice treated with carrageenan. Immunohistochemical analysis for nitrotyrosine and poly(A)DP-ribose polymerase revealed a positive staining in lungs from carrageenan-treated IL-6WT mice. No positive staining for nitrotyrosine or PARS was found in the lungs of the carrageenan-treated IL-6KO mice. Staining of lung tissue sections obtained from carrageenan-treated IL-6WT mice with an anti-cyclo-oxygenase-2 Ab showed a diffuse staining of the inflamed tissue. Furthermore, expression of inducible nitric oxide synthase was found mainly in the macrophages of the inflamed lungs from carrageenan-treated IL-6WT mice. The intensity and degree of the staining for cyclo-oxygenase-2 and inducible nitric oxide synthase were markedly reduced in tissue sections obtained from carrageenan-treated IL-6KO mice. Most notably, the degree of lung injury caused by carrageenan was also reduced in IL-6KO mice. Treatment of IL-6WT mice with anti-IL-6 (5 microg/day/mouse at 24 and 1 h before carrageenan treatment) also significantly attenuated all the above indicators of lung inflammation. Taken together, our results clearly demonstrate that IL-6KO mice are more resistant to the acute inflammation of the lung caused by carrageenan injection into the pleural space than the corresponding WT mice.

Peroxynitrite-mediated DNA strand breakage activates poly (ADP-ribose) synthetase and causes cellular energy depletion in carrageenan-induced pleurisy.

The aim of the present study was to investigate the role of poly (ADP-ribose) synthetase in acute local inflammation (carrageenan-induced pleurisy), where oxyradicals, nitric oxide and peroxynitrite are known to play a crucial role in the inflammatory process. DNA single-strand breakage and activation of the nuclear enzyme poly (ADP-ribose) synthetase (PARS) triggers an energy-consuming, inefficient repair cycle, which contributes to peroxynitrite-induced cellular injury. Here we investigated whether peroxynitrite production and PARS activation are involved in cytotoxicity in macrophages collected from rats subjected to carrageenan-induced pleurisy. Macrophages harvested from the pleural cavity exhibited a significant production of peroxynitrite, as measured by the oxidation of the fluorescent dye dihydrorhodamine 123, and by nitrotyrosine Western blotting at 4 hr after carrageenan injection. Furthermore, carrageenan-induced pleurisy caused a suppression of macrophage mitochondrial respiration, DNA strand breakage, activation of PARS and reduction of NAD+ cellular levels. In vivo treatment with 3-aminobenzamide (10 mg/kg intraperitoneally, 1 hr after carrageenin injection) significantly inhibited the decrease in mitochondrial respiration and the activation of PARS and partially restored the cellular level of NAD+. In a separate group of experiments, in vivo pretreatment with NG-nitro-L-arginine methyl ester, a non-selective inhibitor of nitric oxide (NO) synthesis (10 mg/kg intraperitoneally, 15 min before carrageenan administration), reduced peroxynitrite formation and prevented the appearance of DNA damage, the decrease in mitochondrial respiration and the loss of cellular levels of NAD+. Our study suggests that formation of peroxynitrite and subsequent activation of PARS may alter macrophage function in inflammatory processes and inhibition of NO and PARS may be a novel pharmacological approach to prevent cell injury in inflammation.

Urokinase-type plasminogen activator-deficient mice are predisposed to staphylococcal botryomycosis, pleuritis, and effacement of lymphoid follicles.

Urokinase-type plasminogen activator (uPA) is thought to be an important mediator in the proteolytic degradation of extracellular matrix components observed in a wide variety of normal physiological and pathological conditions. However, the phenotype of a recently developed strain of urokinase-deficient (uPA-/-) mice appears to be normal when maintained under ideal nonstressful conditions. We report an outbreak of botryomycosis, an unusual staphylococcal infection, in a colony of uPA-deficient mice. A detailed histological examination of these uPA-deficient animals also revealed a variety of previously unreported phenotypic abnormalities such as pleuritis and the effacement of lymphoid follicles in the regional lymph nodes and spleen. Additional phenotypic abnormalities such as dystrophic calcifications and rectal prolapse were also observed in the uPA-deficient population. These abnormalities were also noted in ostensibly healthy uPA-deficient animals. Botryomycosis did not affect a colony of wild-type (uPA+/+) animals maintained concurrently under identical conditions in the same room. The peculiar predisposition of the uPA-deficient animals to this rare bacterial infection and the development of phenotypic abnormalities associated with the targeted disruption the uPA gene suggests that uPA contributes significantly to the cutaneous microenvironment and is additional evidence of the extensive involvement of the plasminogen activators in mammalian physiology.

Familial cryptogenic fibrosing pleuritis with Fanconi's syndrome (renal tubular acidosis). A new syndrome.

We describe two siblings with a progressive unrelenting and unique syndrome of bilateral fibrosing pleuritis of unknown cause occurring in association with Fanconi's syndrome (renal tubular acidosis). The parents of the siblings were second cousins. Both siblings had identical pleural histologic characteristics and identical urinary metabolic defects. This condition resulted in the development of severe respiratory failure in both patients and ultimately the death of the older sibling at the age of 21 years.

Pathogenetic and functional characterization of lymphocytes in influenza patients.

The kinetics of acrocentric chromosome associations and chromosome aberrations in peripheral blood and pleural exudate lymphocytes has been studied in 25 influenza patients and 7 exudative pleurisy patients. Lymphocytes without associations and with 2 associated acrocentric chromosomes were activated in the body, since their frequency appeared to be positively correlated with the immunoresponsiveness indices and with clinical symptoms. The number of these lymphocytes in pleural exudate was 2.5 times higher than in the peripheral blood. When comparing the frequency of chromosome aberrations in the patients' lymphocytes to the level of immunity, cytogenetic changes corresponded to the indices of cellular rather than humoral immunity.

[Chromosome analysis performed in 176 cases of pleural effusions of various types]. / Studio sull'analisi cromosomica effettuata in 176 casi di versamenti pleurici di varia natura.

Reference is made to the results of a chromosome analysis conducted on pleural liquid sediments in 71 cases of neoplastic effusion and 105 transudative and inflammatory cases. Numeric and morphological karyotype alterations conclusively indicative of the neoplastic origin of the effusion were observed in 79% cases where a tumour was actually present, and in none of the non-neoplastic cases. The diagnostic sensitivity of the examination was compared with that of the Pap-test, which gave positive results in 63% of cases. When both methods were used, diagnosis was possible in 90% of cases. Some suggestions are made for checking with other studies on the possibilities offered by chromosome as a means of prognosis as well as diagnosis.