Recent advances in interstitial lung disease research

The interstitial lung diseases are a diverse collection of disorders characterized by impaired gas exchange, restricted physiology on lung function testing, and diffuse parenchymal lung infiltrates on radiography. Although the interstitial lung diseases are many, in routine clinical practice, the most commonly encountered in general internal medicine practice are sarcoidosis, idiopathic pulmonary fibrosis, and connective tissue disease-associated interstitial lung diseases. In immunocompromised patients, infection is the most common cause of diffuse lung infiltrates and must be ruled out before any attempt to treat with immune altering agents like corticosteroids. This review will focus on the more clinically significant recent advances in the broad field of interstitial lung disease research, with emphasis on the more common interstitial lung diseases occurring in immunocompetent hosts.peer-reviewe

Pulmonary langerhans cell histiocytosis

Pulmonary Langerhans Cell Histiocytosis (PLCH) is a relatively uncommon lung disease that generally, but not invariably, occurs in cigarette smokers. The pathologic hallmark of PLCH is the accumulation of Langerhans and other inflammatory cells in small airways, resulting in the formation of nodular inflammatory lesions. While the overwhelming majority of patients are smokers, mechanisms by which smoking induces this disease are not known, but likely involve a combination of events resulting in enhanced recruitment and activation of Langerhans cells in small airways. Bronchiolar inflammation may be accompanied by variable lung interstitial and vascular involvement. While cellular inflammation is prominent in early disease, more advanced stages are characterized by cystic lung destruction, cicatricial scarring of airways, and pulmonary vascular remodeling. Pulmonary function is frequently abnormal at presentation. Imaging of the chest with high resolution chest CT scanning may show characteristic nodular and cystic abnormalities. Lung biopsy is necessary for a definitive diagnosis, although may not be required in instances were imaging findings are highly characteristic. There is no general consensus regarding the role of immunosuppressive therapy in smokers with PLCH. All smokers must be counseled on the importance of smoking cessation, which may result in regression of disease and obviate the need for systemic immunosuppressive therapy. The prognosis for most patients is relatively good, particularly if longitudinal lung function testing shows stability. Complications like pneumothoraces and secondary pulmonary hypertension may shorten life expectancy. Patients with progressive disease may require lung transplantation

Aryl hydrocarbon receptor (AHR) is a potential tumour suppressor in pituitary adenomas

Pituitary adenomas (PA) represent the largest group of intracranial neoplasms and yet the molecular mechanisms driving this disease remain largely unknown. The aim of this study was to use a high-throughput screening method to identify molecular pathways that may be playing a significant and consistent role in PA. RNA profiling using microarrays on eight local PAs identified the aryl hydrocarbon receptor (AHR) signalling pathway as a key canonical pathway downregulated in all PA types. This was confirmed by real-time PCR in 31 tumours. The AHR has been shown to regulate cell cycle progression in various cell types; however, its role in pituitary tissue has never been investigated. In order to validate the role of AHR in PA behaviour, further functional studies were undertaken. Over-expression of AHR in GH3 cells revealed a tumour suppressor potential independent of exogenous ligand activation by benzo α-pyrene (BαP). Cell cycle analysis and quantitative PCR of cell cycle regulator genes revealed that both unstimulated and BαP-stimulated AHR reduced E2F-driven transcription and altered expression of cell cycle regulator genes, thus increasing the percentage of cells in G0/G1 phase and slowing the proliferation rate of GH3 cells. Co-immunoprecipitation confirmed the interaction between AHR and retinoblastoma (Rb1) protein supporting this as a functional mechanism for the observed reduction. Endogenous Ahr reduction using silencing RNA confirmed the tumour suppressive function of the Ahr. These data support a mechanistic pathway for the putative tumour suppressive role of AHR specifically in PA, possibly through its role as a cell cycle co-regulator, even in the absence of exogenous ligands.peer-reviewe

Pulmonary mantle cell lymphoma: a rare manifestation of an uncommon condition

Herein we describe the case of a 64-year old man with a history of mantle cell lymphoma found to have evidence of pulmonary parenchymal involvement by recurrence of his lymphoma. While lung involvement is not necessarily uncommon with Non-Hodgkin's lymphomas as a group, it is very rare for mantle cell lymphoma to involve the lung parenchyma. In addition, the radiographic manifestation of his pulmonary lymphoma as a discrete FDG-avid ground-glass lesion on chest imaging was also distinctly uncommon for pulmonary lymphoma which classically appears in one of three patterns: scattered ill-defined nodules, a bronchovascular/lymphangitic process, or pneumonic/alveolar consolidation effectively indistinguishable from bacterial pneumonia. Due to significant underlying lung disease our patient was not a candidate for high-dose conditioning and autologous stem cell transplantation. He was ultimately treated with rituximab and cladribine therapy and had early signs of clinical response at last correspondence

Management of threatened Aphanius Fasciatus at Il-Maghluq, Malta

Over the last decade the distribution of Aphanius fasciatus Nardo has regressed sharply across the Maltese Islands despite numerous legal conservation instruments. In this study we present the results of a one-year phenological study at the protected wetland site known as Il-Maghluq. The A. fasciatus population structure and a number of water physical characteristics were monitored. The biotic data collected was found to be consistent with that of a highly vulnerable population. The authors make a number of management recommendations to improve the conservation status of this population.peer-reviewe

RNASeq of pituitary adenomas reveals dysfunctional metabolic, secretory and differentiation molecular pathways

Pituitary adenomas consist of a group of highly heterogeneous intracranial tumours with variable presentation, clinical prognosis and management. High throughput sequencing was used in order to try and identify common de-regulated pathways and characterize tumours according to specific molecular behaviour. RNA sequencing (RNAseq) was chosen since it provides not only information regarding the expression profile but also the mutational load of each specific tumour analysed. 58 locally resected tumours (36 non-functional tumours; 17 growth hormone-secreting; 3 prolactin-secreting; 2 Cushing’s) were stored in RNAlater (Qiagen, US) and RNA was extracted to purified. RNAseq was performed on all samples plus a control on the BGI-Seq500 platform (Beijing Genomics Ind., China). Bioinformatic analyses was also performed by BGI with additional analyses still being carried out. Preliminary data reveals a number of known and novel de-regulated pathways which characteristically differentiate between different tumour types such as hormone signalling and production pathways. However, novel metabolic pathways also appear to differ significantly, not only between controls and tumours but also between different tumour types with changes in lipid transport and glucose metabolism being observed. Additionally various hormone receptor signalling pathways were also found to be altered. Verification and additional bioinformatic analyses will be required to further delve into the vast data that is generated by this technique which has provided a wealth of information.peer-reviewe

Obsidian from the Site of Piano dei Cardoni, Ustica (Palermo, Italy): Preliminary Results on the First Occupation of the Island

Abstract New investigations on Ustica (Palermo, Sicily) originated from the need to improve our knowledge of the island's archaeological and environmental heritage. Through field surveys, particular attention was paid to human occupation in the Neolithic phases and focused on the less investigated southern side of the island. The systematic survey of the area of Piano dei Cardoni in 2018 brought to light a new Middle/Late Neolithic site, already partially documented in the literature. The island was settled for the first time during these phases, as also testified from the area of Punta Spalmatore. The presence of Serra d'Alto, incised dark burnished, and Diana styles suggests that the site and the archaeological assemblage dates from the mid to late 5th millennium BC, as confirmed by AMS dating. In addition to pottery, obsidian artifacts were also recovered, and a preliminary study of these materials is presented here. Portable XRF analyses on a sample of 41 obsidian artifacts, representing a high percentage of the lithic assemblage compared to chert tools, show that the provenance of the raw material is Gabellotto Gorge (Lipari) and Balata dei Turchi (Pantelleria). These results provide new insight into broader regional debates about obsidian technology and its exchange during the Neolithic and open an important consideration for sites that are far from the raw material sources

The down-regulated tumour suppressor Wnt inhibitory factor 1 (WIF1) regulates non-canonical Wnt signalling in pituitary adenomas (PA)

The Wnt developmental pathway has been implicated in tumour growth and development in a number of tissues. Most frequently, the canonical Wnt pathway acting through the nuclear translocation of B-catenin has been the key player in driving tumour growth. However, non-canonical Wnt pathways, namely the Wnt-Calcium signalling and the Wnt-planar polarity pathways have also been found de-regulated in a number of cancers. In this study, microarray analysis on locally resected PA revealed strong down-regulation Wnt pathway antagonists, namely the Wnt inhibitory factor 1 (WIF1) and the secreted frizzled-related proteins 2 and 4 (SFRP2 and 4). These results have been confirmed by qPCR and shown that WIF1 is under – expressed in all tumour types while SFRP’s tend to be repressed in functional PAs. The aim of this study was to functionally assess the role of WIF1 in PA in relation to the different Wnt signalling pathways using two established cell lines, the rat sommatotroph/lactrotroph GH3 and prolactinoma MMQ cell lines in the presence of known canonical and non-canonical Wnt ligands, Wnt3, Wnt4 and Wnt5a. WIF1 over-expression reduced significantly GH3 and MMQ cell proliferation using a fluorescent-based Alamar Blue assay, both in the absence and presence of Wnt ligands. However, both Wnt ligands and lithium chloride, an established canonical Wnt pathway activator, failed to activate β-catenin driven transcription using the TOP/FOP flash luciferase system. In fact, canonical Wnt signalling appears to be completely absent in GH3 and MMQ cells. In order to study the influence of Wnt ligands and their inhibitor, WIF1, on other non-canonical Wnt pathways, the Wnt-Calcium signalling pathway was chosen, owing to the important role that calcium signalling plays in regulating hormone release from these cells. Using the FluoForte Calcium assay (Enzo Biologicals, US) to assess free calcium in real-time in the chosen cell lines, we studied the effect of the Wnt ligands in the absence and presence of the WIF1 inhibitor. Wnt ligands activated calcium release with variable potentials with WIF1 displaying an inhibitory but selective role on this effect. Real-time PCR of targets of the canonical and non-canonical Wnt pathways is also being undertaken together with analysis of growth hormone and prolactin secretion from both cell lines. Preliminary data reveals that the Wnt agonists may activate the Wnt-Calcium signalling pathway and WIF1 could play a role in PA by inhibiting specific aspects of this pathway.peer-reviewe

Novel and classical molecular pathways identified in pituitary tumorigenesis using mRNA profiling

Pituitary tumorigenesis has been analysed from multiple perspectives, yet mRNA expression profiling studies are limited. In this study, microarray analysis was used to identify pathways and networks related to pituitary tumour physiology using key de-regulated genes and bioinformatics analysis. Eight pituitary adenomas (five non-functional tumours, two GH-secreting tumours and a TSH/prolactin-secreting tumour) and a pool of random normal control RNA were profiled for RNA expression using the 29 kb Affymetrix HuGene 1.0 ST chip. Microarray data was analysed using GeneSpring GX 11.0 and network analysis was done using the Ingenuity Pathway Analysis (IPA) software. Data obtained from the microarray was verified on 30 tumours (20 non-functional tumours, six GH-secreting, two prolactinomas and two Cushing’s) using quantitative PCR of key genes involved in the networks identified by the IPA. Different analyses between controls and tumour types were carried out. Among the classical networks discovered known to be involved in pituitary tumorigenesis were the cAMP signalling pathway and the Wnt developmental pathway although both networks were driven by genes not previously described in any other study. Different tumour types were also found to be characterised by variable novel de-regulated molecular pathways, such as the GABA signalling and aryl hydrocarbon receptor-signalling pathways in GH-secreting tumours, and the p53 signalling de-regulation in the non-functional tumours. Cluster analysis from microarray data was also able to distinguish between tumour types, identifying one non-functional tumour as belonging to the functional tumours by its mRNA expression profile. This study validates the use for gene expression profiling for the correct characterisation of pituitary adenoma sub-types and identification of key pathways involved in pituitary tumorigenesis thereby proposing possible therapeutic targets.peer-reviewe

Functional analysis of aryl hydrocarbon receptor (AHR) polymorphisms in pituitary adenomas (PAs) in the presence of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)

BACKGROUND: PAs are the most frequent pituitary neoplasms, however molecular pathogenesis is largely unknown. The AHR is a ligand-activated transcription factor that regulates expression of various genes that mediate cellular response to xenobiotics. The exact functional role of two AHR single nucleotide polymorphisms (SNPs); Arginine554Lysine (Arg554Lys) and Valine570Isoleucine (Val570Ile) has not yet been established, however studies suggest that these mutations might increase risk of developing PAs. To date, functional analysis of regarding the significance of these AHR SNPs in pituitary pathophysiology has never been analysed.AIMS: • Elucidate the effect of wildtype and polymorphic AHR on GH3 cell proliferation and on AHR-transcriptional response in the presence and absence of TCDD. • Determine the allele frequency of the most common AHR SNP; the Arg554Lys in PA patients and in a small cohort of the Maltese population.METHOD: The two missense mutations were introduced within the AHR-expressing vector and transfected in GH3 cells by magnetofaction, followed by the exposure to TCDD. Cell viability of GH3 transfected cells was measured using the MTT assay. Functional analysis of GH3 transfected cells treated with TCDD was carried out using luciferase assay and real-time PCR to detect and quantify the AHR-transcriptional activity. Genotyping of the Arg554Lys was performed on PA patients and neonatal controls using allele specific PCR. The Mann-Whitney test was used to compare two groups and Kruskall-Wallis test was used to compare three groups or more.RESULTS: In the absence and presence of low TCDD concentrations (1 and 10 nM), over-expression of wildtype AHR (wtAHR) did not affect GH3 cell proliferation. GH3 cells transfected with the AHR mutants did not exhibit any significant differences in their proliferative ability when compared with the wtAHR, both in the presence and absence of TCDD. Luciferase reporter analysis showed that there was a significant difference between the treated and untreated wtAHR (P=0.016), however this difference was not observed between the treated and untreated AHR mutants. Statistically significant difference in Cyp1a1 gene expression analysis was detected between the treated and untreated wtAHR (P=0.021), Arg554Lys (P=0.005) and Val570Ile (P=0.054). Genotyping of the Arg554Lys in patients with PA gave a minor allele frequency (MAF) of 3% vs 0% in neonatal controls.CONCLUSION: Gene expression and quantification analyses of AHR-target genes suggests that these AHR mutants might interfere with AHR target gene expression. Genotyping results suggested that this mutation is quite rare and may be similar to the frequencies of other European populations.peer-reviewe