Transesophageal approach to lung, adrenal biopsy and fiducial placement using endoscopic ultrasonography (EUS): An interventional pulmonology experience. Initial experience of the UCSF-FRETOC (fresno tracheobronchial & oesophageal center) study group.

BACKGROUND: Routine lung cancer surveillance has resulted in early detection of pulmonary nodules and masses. Combined endobronchial ultrasound (EBUS) and trans-esophageal endoscopic ultrasound (EUS) are approved methods for sampling lymph nodes or masses. Furthermore, EUS allows for adrenal sampling as part of staging, and can assist with fiducial placement for stereotactic body radiation therapy (SBRT). OBJECTIVES: Promote use of EUS by interventional pulmonologists in the United States when diagnosing and staging lung cancer or when placing fiducials. METHODS: All patients undergoing EUS and/or EBUS were serially entered into a prospectively maintained database. Only patients undergoing EUS guided lung and/or adrenal biopsy and/or fiducial placement were selected for analysis. All patients underwent a post-procedure chest radiograph and were followed outpatient. RESULTS: 20 of 39 patients underwent sampling of a suspicious lung mass. An adequate sample was obtained in 19 of 20 patients. In all 19 patients a definitive diagnosis was achieved (95%). In all 13 patients who underwent adrenal sampling, presence or absence of metastasis was conclusively established. 6 patients successfully underwent fiducial placement. In all 39 patients, no major procedure related complications were noted for a period of 30 days. One patient had a small pneumothorax that resolved spontaneously. CONCLUSIONS: EUS can be safely performed by a trained interventional pulmonologist for the diagnosis of lung, adrenal masses and placement of fiducials. We think that interventional pulmonologists in the United States involved in lung cancer staging should receive training in EUS techniques.

TPD52L1-ROS1, a new ROS1 fusion variant in lung adenosquamous cell carcinoma identified by comprehensive genomic profiling.

Crizotinib was approved for the treatment of ROS1-rearranged non-small cell lung cancer (NSCLC) patients in the US on 11 March, 2016. Interestingly no one companion diagnostic test (CDx) has been approved simultaneously with this approval of crizotinib. Hence, an ideal and adequate CDx will have to be able to identify ROS1 fusions without the knowledge of the fusion partners to ROS1, and as to date there are 13 fusion partners reported for ROS1 in NSCLC. Here we report a novel TPD52L1-ROS1 fusion variant in NSCLC. This novel TPD52L1-ROS1 fusion variant is generated by the fusion of exons 1-3 of TPD52L1 on chromosome 6q22-23 to the exons 33-43 of ROS1 on chromosome 6q22, likely from an intra-chromosomal deletion and subsequent fusion event similar to the generation of EML4-ALK. The predicted TPD52L1-ROS1 protein product contains 655 amino acids comprising of the N-terminal amino acids 1-95 of TPD52L1 and C-terminal amino acids of 1789-2348 of ROS1. In summary, TPD52L1-ROS1 is a novel ROS1 fusion variant in NSCLC identified by comprehensive genomic profiling and should be included in any ROS1 detecting assays that depend on identifying the corresponding fusion partners, such as reverse transcriptase-polymerase chain reaction (RT-PCR).

Involvement of EZH2, SUV39H1, G9a and associated molecules in pathogenesis of urethane induced mouse lung tumors: potential targets for cancer control.

In the present study, we showed the correlation of EZH2, SUV39H1 or G9a expression and histone modifications with the urethane induced mouse lung tumorigenesis in the presence or absence of antitumor agent, inositol hexaphosphate (IP6). Tumorigenesis and the molecular events involved therein were studied at 1, 4, 12 or 36 weeks after the exposure. There were no tumors at 1 or 4 weeks but tumors started appearing at 12 weeks and grew further till 36 weeks after urethane exposure. Among the molecular events, upregulation of EZH2 and SUV39H1 expressions appeared to be time dependent, but G9a expression was altered significantly only at later stages of 12 or 36 weeks. Alteration in miR-138 expression supports the upregulation of its target, EZH2. H3K9me2, H3K27me3 or H4K20me3 was found to be altered at 12 or 36 weeks. However, ChIP analysis of p16 and MLH1 promoters showed their binding with H3K9me2 and H3K27me3 which was maximum at 36 weeks. Thus, histone modification and their interactions with gene promoter resulted in the reduced expression of p16 and MLH1. IP6 prevented the incidence and the size of urethane induced lung tumors. IP6 also prevented the urethane induced alterations in EZH2, SUV39H1, G9a expressions and histone modifications. Our results suggest that the alterations in the histone modification pathways involving EZH2 and SUV39H1 expressions are among the early events in urethane induced mouse lung tumorigenesis and could be exploited for cancer control.

Sequestration of mitochondrial iron by silica particle initiates a biological effect.

Inhalation of particulate matter has presented a challenge to human health for thousands of years. The underlying mechanism for biological effect following particle exposure is incompletely understood. We tested the postulate that particle sequestration of cell and mitochondrial iron is a pivotal event mediating oxidant generation and biological effect. In vitro exposure of human bronchial epithelial cells to silica reduced intracellular iron, which resulted in increases in both the importer divalent metal transporter 1 expression and metal uptake. Diminished mitochondrial (57)Fe concentrations following silica exposure confirmed particle sequestration of cell iron. Preincubation of cells with excess ferric ammonium citrate increased cell, nuclear, and mitochondrial metal concentrations and prevented significant iron loss from mitochondria following silica exposure. Cell and mitochondrial oxidant generation increased after silica incubation, but pretreatment with iron diminished this generation of reactive oxygen species. Silica exposure activated MAP kinases (ERK and p38) and altered the expression of transcription factors (nF-κB and NF-E2-related factor 2), proinflammatory cytokines (interleukin-8 and -6), and apoptotic proteins. All of these changes in indexes of biological effect were either diminished or inhibited by cell pretreatment with iron. Finally, percentage of neutrophils and total protein concentrations in an animal model instilled with silica were decreased by concurrent exposure to iron. We conclude that an initiating event in the response to particulate matter is a sequestration of cell and mitochondrial iron by endocytosed particle. The resultant oxidative stress and biological response after particle exposure are either diminished or inhibited by increasing the cell iron concentration.

The role and regulation of microRNAs in asthma.

PURPOSE OF REVIEW: Asthma is a common chronic inflammatory airway disorder that is characterized by variable and recurring airflow obstruction, chronic airway inflammation and bronchial hyper-responsiveness. The etiopathogenesis of asthma remains a complex issue. The intricacy in developing a more effective therapeutic strategy may be due to a large diversity in causative agents and a lack of understanding of the precise molecular mechanism involved in asthma. However, recent identification of microRNAs (miRs) has enhanced technological abilities to understand the disease process. RECENT FINDINGS: miRs regulate gene expression by controlling the translation of a specific type of messenger RNA. miRs have been recently identified as key regulatory RNAs with immense significance in numerous biological processes including asthma. miRs have been implicated to have a fundamental role in acute and chronic asthma and in airway remodeling by the regulation of multiple signal transduction pathways that are involved in the pathogenesis of asthma. It is possible that miRs may bring a fundamental change to our understanding of the pathophysiology of asthma. This may then lead to the development of novel efficacious therapeutic strategies in asthma. SUMMARY: In this review, we highlight the current understanding of the role and regulation of miRs in asthma.

Inhibition of inflammatory mediators: role of statins in airway inflammation.

OBJECTIVE: To determine if statins induce anti-inflammatory effects in upper airway inflammation. Mediators of innate and adaptive immunity regulate airway inflammation. Release of these mediators involves enzymatic conversion of polyunsaturated fatty acids into biologically active mediators, which can be blocked by statins. Although upper airway inflammation and chronic sinusitis occur in millions of patients with asthma worldwide, the anti-inflammatory effects of statins in upper airway inflammation have not been previously studied. STUDY DESIGN: Laboratory research. SETTING: Tertiary referral center. SUBJECTS AND METHODS: Analysis of sinus tissues collected from patients with chronic rhinosinusitis revealed suppression of highly expressed inflammatory mediators in patients who were found to be on statins, suggesting that statins may induce anti-inflammatory effects. Therefore, the authors performed an in vitro study to determine if these anti-inflammatory effects were induced by statins. Cultured primary human airway epithelial cells were exposed to ambient air pollution particulates (PM) to trigger the inflammation, with and without statins, and the expression of inflammatory mediators was analyzed. RESULTS: The authors found that expression of CCL5, CCL11, and IL13RA was suppressed in patients on statins. In vitro exposure to PM enhanced the expression of these mediators, while pretreatment with statins completely blocked these effects. Furthermore, the effects of statins were blocked by inhibition of the statin pathway using isopentenyl-5-pyrophosphate. Statins did not have any significant effect on the viability of normal cells. CONCLUSION: Statins induce anti-inflammatory effects in human airway epithelial inflammation. Statins may play a role in the treatment and prevention of chronic rhinosinusitis and pulmonary exacerbation of obstructive airway diseases.

Novel effects of statins in enhancing efficacy of chemotherapy in vitro in nasopharyngeal carcinoma.

BACKGROUND: Nasopharyngeal cancer (NPC) is a relatively uncommon malignant epithelial cancer seen worldwide. The treatment of NPC has evolved toward combined modality treatment with radiation therapy and chemotherapy. However, chemotherapeutic agents currently have a secondary role, due to their lack of efficacy as curative agents. Recent identification of a novel property of statin drugs raises a promising hope that concurrent use of statins may enhance the efficacy of single-drug chemotherapy in NPC. However, the effects of statins have not been studied before in NPC. In this in vitro study, we demonstrate a unique property of statins that can enhance the efficacy of cisplatin in NPC. METHODS: Primary human NPC cells (CCL-30) were treated with statins and cisplatin concurrently, and the effects on cell proliferation, apoptosis, and cell colony formation were examined. RESULTS: Statins caused significant decrease in cell proliferation and viability in NPC. Statins also induced loss of cell attachment, change in cellular morphology, decrease in colony forming units, and loss of sphere formation in soft gel agar, which are the important properties of tumorigenicity in NPC. Furthermore, we found that the effects of statins occur by a mevalonate (MA)-mediated pathway in these cells. CONCLUSIONS: We demonstrate a unique property of statins that can enhance the antitumor effects of cisplatin in NPC. Statins may act as a relatively safe and cost effective chemoadjuvant agent in the treatment of NPC.

Nicotine induces resistance to chemotherapy in nasal epithelial cancer.

BACKGROUND: Epidemiological and clinical data implicate that in patients with cancer, continued smoking causes progression of cancer growth and resistance to therapy. The carcinogens possess the ability to block apoptosis, an important mechanism in the development of tumors and resistance to chemotherapy. We previously showed that nicotine enhances growth and proliferation in lung cancer. However, the effects of nicotine, a tobacco carcinogen that inhibits apoptosis, have not been studied before in nasal epithelial carcinoma (NC). In this study, we sought to determine the effects of nicotine on chemotherapy-induced apoptosis in human NC. METHODS: Primary human NC cells were grown per protocol, treated with combination chemotherapy, and the apoptosis was assessed by TUNEL (terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling) and DNA fragmentation assays. The regulation of mitogen-activated protein kinase (MAPK) and protein kinase B (AKT) signal transduction pathway was examined by real time quantitative polymerized chain reaction, and immunofluorescent staining assays. RESULTS: Combination chemotherapy with cisplatin (35 microM) plus etoposide (20 microM) caused a significant increase in NC apoptosis compared with single agent alone, and nicotine, in part, inhibited chemotherapy-induced apoptosis in NC. Furthermore, nicotine induced activation of AKT and MAPK pathways, while inhibition of MAPK using U0126 and AKT by phosphatidylinositol 3-kinase inhibitor, LY294002, in part, blocked the antiapoptotic effects of nicotine against cisplatin and etoposide-induced apoptosis in NC. CONCLUSION: Nicotine inhibits chemotherapy-induced apoptosis in NC via the AKT and MAPK-mediated signaling pathways. We speculate that nicotine may play a role in oncogenesis and resistance to cancer therapy in NC.

SPARC suppresses apoptosis of idiopathic pulmonary fibrosis fibroblasts through constitutive activation of beta-catenin.

Idiopathic pulmonary fibrosis (IPF) is a poorly understood progressive disease characterized by the accumulation of scar tissue in the lung interstitium. A hallmark of the disease is areas of injury to type II alveolar epithelial cells with attendant accumulation of fibroblasts in areas called fibroblastic foci. In an effort to better characterize the lung fibroblast phenotype in IPF patients, we isolated fibroblasts from patients with IPF and looked for activation of signaling proteins, which could help explain the exaggerated fibrogenic response in IPF. We found that IPF fibroblasts constitutively expressed increased basal levels of SPARC, plasminogen activator inhibitor-1 (PAI-1), and active beta-catenin compared with control cells. Control of basal PAI-1 expression in IPF fibroblasts was regulated by SPARC-mediated activation of Akt, leading to inhibition of glycogen synthase kinase-3beta and activation of beta-catenin. Additionally, IPF fibroblasts (but not control fibroblasts) were resistant to plasminogen-induced apoptosis and were sensitized to plasminogen-mediated apoptosis by inhibition of SPARC or beta-catenin. These findings uncover a newly discovered regulatory pathway in IPF fibroblasts that is characterized by elevated SPARC, giving rise to activated beta-catenin, which regulates expression of downstream genes, such as PAI-1, and confers an apoptosis-resistant phenotype. Disruption of this pathway may represent a novel therapeutic target in IPF.

Nicotine induces resistance to chemotherapy by modulating mitochondrial signaling in lung cancer.

Continued smoking causes tumor progression and resistance to therapy in lung cancer. Carcinogens possess the ability to block apoptosis, and thus may induce development of cancers and resistance to therapy. Tobacco carcinogens have been studied widely; however, little is known about the agents that inhibit apoptosis, such as nicotine. We determine whether mitochondrial signaling mediates antiapoptotic effects of nicotine in lung cancer. A549 cells were exposed to nicotine (1 muM) followed by cisplatin (35 muM) plus etoposide (20 muM) for 24 hours. We found that nicotine prevented chemotherapy-induced apoptosis, improved cell survival, and caused modest increases in DNA synthesis. Inhibition of mitogen-activated protein kinase (MAPK) and Akt prevented the antiapoptotic effects of nicotine and decreased chemotherapy-induced apoptosis. Small interfering RNA MAPK kinase-1 blocked antiapoptotic effects of nicotine, whereas small interfering RNA MAPK kinase-2 blocked chemotherapy-induced apoptosis. Nicotine prevented chemotherapy-induced reduction in mitochondrial membrane potential and caspase-9 activation. Antiapoptotic effects of nicotine were blocked by mitochondrial anion channel inhibitor, 4,4'diisothiocyanatostilbene-2,2'disulfonic acid. Chemotherapy enhanced translocation of proapoptotic Bax to the mitochondria, whereas nicotine blocked these effects. Nicotine up-regulated Akt-mediated antiapoptotic X-linked inhibitor of apoptosis protein and phosphorylated proapoptotic Bcl2-antagonist of cell death. The A549-rho0 cells, which lack mitochondrial DNA, demonstrated partial resistance to chemotherapy-induced apoptosis, but blocked the antiapoptotic effects of nicotine. Accordingly, we provide evidence that nicotine modulates mitochondrial signaling and inhibits chemotherapy-induced apoptosis in lung cancer. The mitochondrial regulation of nicotine imposes an important mechanism that can critically impair the treatment of lung cancer, because many cancer-therapeutic agents induce apoptosis via the mitochondrial death pathway. Strategies aimed at understanding nicotine-mediated signaling may facilitate the development of improved therapies in lung cancer.

Orbital emphysema in COPD with bronchopleural fistula.

Orbital emphysema is usually a benign, self-limited condition. Intraorbital extension of the entrapped air in the absence of an orbital fracture is extremely rare. Although benign, a careful periodic monitoring of intraocular pressure, optic nerve examinations, and prompt management are warranted in order to prevent tension pneumoobitus secondary to increasing intraorbital pressure by the orbital air.

Ambient particulate matter induces alveolar epithelial cell cycle arrest: role of G1 cyclins.

We hypothesized that the ambient air pollution particles (particulate matter; PM) induce cell cycle arrest in alveolar epithelial cells (AEC). Exposure of PM (25microg/cm(2)) to AEC induced cells cycle arrest in G1 phase, inhibited DNA synthesis, blocked cell proliferation and caused decrease in cyclin E, A, D1 and Cyclin E- cyclin-dependent kinase (CDK)-2 kinase activity after 4h. PM induced upregulation of CDK inhibitor, p21 protein and p21 activity in AEC. SiRNAp21 blocked PM-induced downregulation of cyclins and AEC G1 arrest. Accordingly, we provide the evidence that PM induces AEC G1 arrest by altered regulation of G1 cyclins and CDKs.

Cardiologists use pulmonary artery catheter information to make homogenous treatment decisions.

Medical intensivists make heterogenous decisions using pulmonary artery catheter (PAC) data in medical intensive care unit patients. The object was to determine if cardiologists given PAC data from critically ill cardiology patients make uniform management choices. A survey questionnaire containing 3 coronary care unit (CCU) clinical vignettes was designed and mailed to board-certified cardiologists who are members of the American College of Cardiology. Twenty board-certified medical intensivists were also asked to complete the vignettes. Each vignette contained PAC data and one-half of the surveys contained echocardiographic (ECHO) information. Every respondent was asked to select 1 of 6 interventions for each vignette. In 2 of 3 vignettes 1 intervention was selected by more than 70% of cardiologists. In the third vignette, 1 intervention was selected by more than 50% of cardiologists. For each vignette, 1 intervention was selected by at least 75% of medical intensivists. There was no significant difference in the distribution of management choices between the ECHO and the non-ECHO subgroups. There is relative homogeneity in selecting an intervention based on PAC data among cardiologists and medical intensivists in CCU patients and is probably due to patient-related factors. The presence of ECHO information did not change the intervention selected. Cardiology patients may represent an ideal group in which to evaluate PAC efficacy.

Recurrence of pulmonary intravascular bronchoalveolar tumor with mediastinal metastasis 20 years later.

Pulmonary intravascular bronchoalveolar tumor (IVBAT) also recognized as pulmonary epithelioid hemangioendothelioma, is a rare malignant vascular tumor of unknown etiology. IVBAT is a tumor of multicentric origin and the lungs are rarely involved, with only about 60 cases of pulmonary IVBAT described in the literature. The prognosis is unpredictable, with life expectancy ranging from 1 to 15 years. We report an unusual case of pulmonary IVBAT that recurred in the lung with metastasis to the mediastinum.

Lung surfactant gelation induced by epithelial cells exposed to air pollution or oxidative stress.

Lung surfactant lowers surface tension and adjusts interfacial rheology to facilitate breathing. A novel instrument, the interfacial stress rheometer (ISR), uses an oscillating magnetic needle to measure the shear viscosity and elasticity of a surfactant monolayer at the air-water interface. The ISR reveals that calf lung surfactant, Infasurf, exhibits remarkable fluidity, even when exposed to air pollution residual oil fly ash (ROFA), hydrogen peroxide (H2O2), or conditioned media from resting A549 alveolar epithelial cells (AEC). However, when Infasurf is exposed to a subphase of the soluble fraction of ROFA- or H2O2-treated AEC conditioned media, there is a prominent increase in surfactant elasticity and viscosity, representing two-dimensional gelation. Surfactant gelation is decreased when ROFA-AEC are pretreated with inhibitors of cellular reactive oxygen species (ROS), or with a mitochondrial anion channel inhibitor, as well as when A549-rho0 cells that lack mitochondrial DNA and functional electron transport are investigated. These results implicate both mitochondrial and nonmitochondrial ROS generation in ROFA-AEC-induced surfactant gelation. A549 cells treated with H2O2 demonstrate a dose-dependent increase in lung surfactant gelation. The ISR is a unique and sensitive instrument to characterize surfactant gelation induced by oxidatively stressed AEC.

Fibroblast growth factor-10 prevents asbestos-induced alveolar epithelial cell apoptosis by a mitogen-activated protein kinase-dependent mechanism.

Asbestos induces alveolar epithelial cell (AEC) DNA damage and apoptosis by the mitochondria-regulated death pathway and oxidative stress. Fibroblast growth factor-10 (FGF-10), an alveolar epithelial type II cell mitogen that is required for the lung development, prevents H(2)O(2)-induced AEC DNA damage by a mitogen activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK)-dependent mechanism. In this study, we show that FGF-10 attenuates asbestos-induced AEC DNA strand break formation and apoptosis. MAPK/ERK kinase (MEK) inhibitors, U0126 or PD98059, each blocked the protective effect of FGF-10 against asbestos-induced DNA damage and apoptosis, whereas a p38-MAPK inhibitor had a negligible effect, suggesting a crucial role for MEK/ERK activation in mediating the protective effects of FGF-10. Further, we show that FGF-10 attenuates asbestos-induced change in AEC mitochondrial membrane potential and caspase 9 activation, both of which are blocked by U0126. We conclude that FGF-10 decreases asbestos-induced AEC DNA damage and apoptosis in part by mechanisms involving MEK/ERK-dependent signaling that affects the mitochondria-regulated death pathway.

Fibroblast growth factor-10 attenuates H2O2-induced alveolar epithelial cell DNA damage: role of MAPK activation and DNA repair.

Fibroblast growth factor-10 (FGF-10), an alveolar epithelial cell (AEC) mitogen that is critical for lung development, may promote AEC repair. We determined whether FGF-10 attenuates H2O2-induced, A549 and rat alveolar type II cell DNA damage. We show that FGF-10 prevents H2O2-induced DNA damage assessed by an alkaline elution, ethidium bromide fluorescence as well as by a comet assay. Mitogen-activated protein kinase inhibitors abolished the protective effect of FGF-10 against H2O2-induced DNA damage yet had no effect on H2O2-induced DNA damage. A Grb2-SOS inhibitor (SH3 binding peptide), an Ras inhibitor (farnesyl transferase inhibitor 277), and an Raf-1 inhibitor (forskolin) each prevented FGF-10- and H2O2-induced A549 cell ERK1/2 phosphorylation. Also, FGF-10 and H2O2 each induced negligible ERK1/2 phosphorylation in Ras dominant-negative (N17) cells. Inhibitors of Ras and Raf-1 blocked the protective effect of FGF-10 against H2O2-induced DNA damage but had no effect on H2O2-induced DNA damage. Furthermore, cold conditions and aphidicolin, an inhibitor of DNA polymerase-alpha, -delta, and -epsilon, each blocked the protective effects of FGF-10, suggesting a role for DNA repair. We conclude that FGF-10 attenuates H2O2-induced AEC DNA damage by mechanisms that involve activation of Grb2-SOS/Ras/RAF-1/ERK1/2 pathway and DNA repair.

Variability in interventions with pulmonary artery catheter data.

OBJECTIVES: To determine if intensivists given PAC data from critically ill patients make uniform management choices. DESIGN: Cross-sectional survey of board-certified intensivists. SETTING: Medical intensive care unit. PARTICIPANTS: Board-certified intensivists who are members of the American College of Chest Physicians and Society of Critical Care Medicine. INTERVENTIONS: A survey questionnaire containing three medical intensive care clinical vignettes was mailed to critical care physicians. Each vignette contained PAC data and one-half of the surveys contained echocardiographic (Echo) information. Every respondent was asked to select one of six interventions for each vignette. MEASUREMENTS: There were 126 evaluable surveys returned. In vignette 1 an intervention (none of the above) was selected by more than 50% of respondents. In vignettes 2 and 3, the most frequent selection was chosen only 44 and 37% of the times, respectively. There was a significant difference in the distribution of management choices between te Echo and the non-Echo subgroups. CONCLUSIONS: There is a significant heterogeneity in selecting an intervention based on PAC data among intensivists. The presence of Echo information may change the intervention selected but does not reduce heterogeneity. Any randomized trial evaluating efficacy of PAC's will have to have strict treatment protocols.