Faculty Bibliography

Estimating the burden of exposure-related diffuse lung disease in terms of health effects and economic burden remains challenging. Labor statistics are inadequate to define the scope of the problem, and few studies have analyzed the prevalence of exposure-related illnesses and the subsequent health care cost. Well-defined exposures, such as those associated with coal mines, asbestos mines, and stonecutting, have led to more accurate assessment of prevalence and cost. As governmental regulation of workplace exposure has increased, the prevalence of diseases such as silicosis and coal workers' pneumoconiosis has diminished. However, the health and economic effects of diseases with long latency periods, such as asbestosis and mesothelioma, continue to increase in the short term. Newer exposures, such as those related to air pollution, nylon flock, and the World Trade Center collapse, have added to these costs. As a result, estimates of cost for occupational diseases, including respiratory illnesses, exceed $26 billion annually, and the true economic burden is likely much higher

BACKGROUND: Whole genome amplification (WGA) methods allow diagnostic laboratories to overcome the common problem of insufficient DNA in patient specimens. Further, body fluid samples useful for cancer early detection are often difficult to amplify with traditional PCR methods. In this first application of WGA on the entire human mitochondrial genome, we compared the accuracy of mitochondrial DNA (mtDNA) sequence analysis after WGA to that performed without genome amplification. We applied the method to a small group of cancer cases and controls and demonstrated that WGA is capable of increasing the yield of starting DNA material with identical genetic sequence. METHODS: DNA was isolated from clinical samples and sent to NIST. Samples were amplified by PCR and those with no visible amplification were re-amplified using the Multiple Displacement Amplificaiton technique of whole genome amplification. All samples were analyzed by mitochip for mitochondrial DNA sequence to compare sequence concordance of the WGA samples with respect to native DNA. Real-Time PCR analysis was conducted to determine the level of WGA amplification for both nuclear and mtDNA. RESULTS: In total, 19 samples were compared and the concordance rate between WGA and native mtDNA sequences was 99.995%. All of the cancer associated mutations in the native mtDNA were detected in the WGA amplified material and heteroplasmies in the native mtDNA were detected with high fidelity in the WGA material. In addition to the native mtDNA sequence present in the sample, 13 new heteroplasmies were detected in the WGA material. CONCLUSION: Genetic screening of mtDNA amplified by WGA is applicable for the detection of cancer associated mutations. Our results show the feasibility of this method for: 1) increasing the amount of DNA available for analysis, 2) recovering the identical mtDNA sequence, 3) accurately detecting mtDNA point mutations associated with cancer

We report an indigenous case of leprosy in New York City in an immunocompetent patient who was infected with a Mycobacterium leprae genotype that is consistent with an exogenous origin. Physicians in the eastern United States should be alerted that, although most patients who develop leprosy in the United States are foreign born, native-born Americans are also susceptible to the infection

We evaluated the mechanisms using immunohistochemistry whereby chrysotile asbestos and benzo(a)pyrene (BaP) instilled intratracheally into lung-specific dominant-negative p53 (dnp53) mice might interact in causing lung carcinomas and fibrosis. Chrysotile asbestos and benzo(a)pyrene (BaP) were instilled intratracheally into lung-specific dominant-negative p53 (dnp53) and control mice. The mice were sacrificed at 12 months and their lungs examined for lung carcinomas and fibrosis. Immunostains for proteins related to apoptosis, fibrogenesis, matrix remodeling and inflammation were performed. The dnp53 mice had increased numbers of lung adenocarcinomas with BaP alone and the combination of chrysotile and BaP (the latter was additive but not significant). Several atypical adenomatous hyperplasia lesions were found in the combined treatment group. dnp53 and FVBN control mice developed nodular buds of fibrotic lung tissue after chrysotile asbestos exposure that were localized in respiratory bronchioles; these lesions had significant increases in immunohistochemical staining for TGF-beta, MMP-7 and -9, MIG-1, and SDF-1. Fibrotic lesions in mice exposed to chrysotile had increased collagen demonstrated by picrosirius red staining. The dnp53 mice with adenocarcinomas had increased SDF-1, TGF-beta, MMP-9 and -7, Cyclin D, and MIG-1 immunostaining in the chrysotile and combined treatment groups. We conclude that BaP and the combination of BaP plus chrysotile asbestos are potent inducers of adenocarcinoma in dnp53 mice and that the inflammatory cytokines and proteases MMP-7 and -9, MIG-1, and SDF-1, and growth factors Cyclin D and TGF-beta are increased in the specific lesions

The interaction of Mycobacterium tuberculosis (MTB) with the immune system is mediated by cytokine and chemokine responses of macrophages and/or dendritic cells. Chemokine (C-C motif) ligand 18 (CCL18) and interleukin (IL)-10 are major factors secreted by phagocytes, postulated to recruit naive T lymphocytes and inhibit pro-inflammatory cells. Our study investigated the role of CCL18 and IL-10 in an in vitro model of infection by MTB in human macrophages. CD14(+) monocytes, obtained from the peripheral blood of eight healthy donors, differentiated in monocyte-derived macrophages (MDM) with monocyte-colony stimulating factor (100 ng/ml) for 6 days, were stimulated in vitro with lipopolysaccharide (LPS) (1 microg/ml) and with heat killed MTB Hv37Ra (multiplicity of infection 1:5) for 24 h. Alveolar macrophages from five healthy donors were infected with MTB Hv37RA. CCL18 protein and mRNA were detected by enzyme-linked immunosorbent assay (ELISA) and real-time PCR, IL-10 levels by ELISA. Stimulation of MDM with LPS or MTB led to a significant increase in CCL18 protein (control 2.67 +/- 0.46 ng/ml, LPS 4.05 +/- 0.56 ng/ml, with MTB 6.70 +/- 1.59 ng/ml, n = 5, P < 0.05) and specific mRNA levels (control 0.09 +/- 0.01, LPS 0.24 +/- 0.11, with MTB 0.34 +/- 0.08 CCL18/Glyceraldehyde 3-phosphate dehydrogenase (GAPDH), n = 3, P < 0.05). A significant increase of the production of CCL18 was observed in infected alveolar macrophages. IL-10 levels increased from 38.52 +/- 26.38 pg/ml in control cells to 1129.32 +/- 235.00 and 974.25 +/- 164.46 pg/ml in LPS and MTB treated cells, respectively (P < 0.05). Up-regulation of CCL18 and IL-10 in macrophages by MTB may be involved in the recruitment of naive T cells in association with local suppressive immunity against intracellular pathogens. This could represent a mechanism of tolerance during the early phases of infection

OBJECTIVE: To determine whether linezolid is safe and well tolerated in the treatment of extensively drug-resistant tuberculosis (XDR-TB). MATERIALS AND METHODS: The was conducted in a specialized tuberculosis ward for multidrug-resistant tuberculosis (MDR-TB) on the Chest Service of Bellevue Hospital Center, which is a 768-bed public hospital in New York City. Seven patients with confirmed MDR-TB or XDR-TB who were still culture positive despite appropriate directly observed therapy were treated with a regimen containing linezolid and at least one other active agent. RESULTS: The linezolid-containing regimen led to sustained negative conversion of sputum cultures and radiographic improvement in all patients. Long-term therapy (longest duration of therapy, 28 months) was well tolerated in most patients. Neutropenia developed in three patients, but was reversible, and peripheral neuropathy developed in two patients. CONCLUSIONS: Linezolid remains a promising possible addition to our therapeutic armamentarium against XDR-TB. Linezolid is associated with side effects that can be adequately managed. Further studies to define the mechanism of action and optimum dose should be performed

Exposure to ambient air pollution has been associated with adverse health effects including lung cancer. A recent epidemiology study has established that each 10mug/m(3) elevation in long-term exposure to average PM(2.5) ambient concentration was associated with approximately 8% of lung cancer mortality. The underlying mechanisms of how PM contributes to lung carcinogenesis, however, remain to be elucidated. We have recently found that transition metals such as nickel and chromium and oxidative stress induced lipid peroxidation metabolites such as aldehydes can greatly inhibit nucleotide excision repair (NER) and enhance carcinogen-induced mutations. Because PM is rich in metal and aldehyde content and can induce oxidative stress, we tested the effect of PM on DNA repair capacity in cultured human lung cells using in vitro DNA repair synthesis and host cell reactivation assays. We found that PM greatly inhibits NER for ultraviolet (UV) light and benzo(a)pyrene diol epoxide (BPDE) induced DNA damage in human lung cells. We further demonstrated that PM exposure can significantly increase both spontaneous and UV-induced mutagenesis. These results together suggest that the carcinogenicity of PM may act through its combined effect on suppression of DNA repair and enhancement of DNA replication errors

BACKGROUND: Mutations in the mitochondrial genome (mtgenome) have been associated with cancer and many other disorders. These mutations can be point mutations or deletions, or admixtures (heteroplasmy). The detection of mtDNA mutations in body fluids using resequencing microarrays, which are more sensitive than other sequencing methods, could provide a strategy to measure mutation loads in remote anatomical sites. METHODS: We determined the mtDNA mutation load in the entire mitochondrial genome of 26 individuals with different early stage cancers (lung, bladder, kidney) and 12 heavy smokers without cancer. MtDNA was sequenced from three matched specimens (blood, tumor and body fluid) from each cancer patient and two matched specimens (blood and sputum) from smokers without cancer. The inherited wildtype sequence in the blood was compared to the sequences present in the tumor and body fluid, detected using the Affymetrix Genechip Human Mitochondrial Resequencing Array 1.0 and supplemented by capillary sequencing for noncoding region. RESULTS: Using this high-throughput method, 75% of the tumors were found to contain mtDNA mutations, higher than in our previous studies, and 36% of the body fluids from these cancer patients contained mtDNA mutations. Most of the mutations detected were heteroplasmic. A statistically significantly higher heteroplasmy rate occurred in tumor specimens when compared to both body fluid of cancer patients and sputum of controls, and in patient blood compared to blood of controls. Only 2 of the 12 sputum specimens from heavy smokers without cancer (17%) contained mtDNA mutations. Although patient mutations were spread throughout the mtDNA genome in the lung, bladder and kidney series, a statistically significant elevation of tRNA and ND complex mutations was detected in tumors. CONCLUSION: Our findings indicate comprehensive mtDNA resequencing can be a high-throughput tool for detecting mutations in clinical samples with potential applications for cancer detection, but it is unclear the biological relevance of these detected mitochondrial mutations. Whether the detection of tumor-specific mtDNA mutations in body fluidsy this method will be useful for diagnosis and monitoring applications requires further investigation

BACKGROUND: A combination of biomarkers in a multivariate model may predict disease with greater accuracy than a single biomarker employed alone. We developed a non-linear method of multivariate analysis, weighted digital analysis (WDA), and evaluated its ability to predict lung cancer employing volatile biomarkers in the breath. METHODS: WDA generates a discriminant function to predict membership in disease vs no disease groups by determining weight, a cutoff value, and a sign for each predictor variable employed in the model. The weight of each predictor variable was the area under the curve (AUC) of the receiver operating characteristic (ROC) curve minus a fixed offset of 0.55, where the AUC was obtained by employing that predictor variable alone, as the sole marker of disease. The sign (+/-) was used to invert the predictor variable if a lower value indicated a higher probability of disease. When employed to predict the presence of a disease in a particular patient, the discriminant function was determined as the sum of the weights of all predictor variables that exceeded their cutoff values. The algorithm that generates the discriminant function is deterministic because parameters are calculated from each individual predictor variable without any optimization or adjustment. We employed WDA to re-evaluate data from a recent study of breath biomarkers of lung cancer, comprising the volatile organic compounds (VOCs) in the alveolar breath of 193 subjects with primary lung cancer and 211 controls with a negative chest CT. RESULTS: The WDA discriminant function accurately identified patients with lung cancer in a model employing 30 breath VOCs (ROC curve AUC=0.90; sensitivity=84.5%, specificity=81.0%). These results were superior to multilinear regression analysis of the same data set (AUC=0.74, sensitivity=68.4, specificity=73.5%). WDA test accuracy did not vary appreciably with TNM (tumor, node, metastasis) stage of disease, and results were not affected by tobacco smoking (ROC curve AUC=0.92 in current smokers, 0.90 in former smokers). WDA was a robust predictor of lung cancer: random removal of 1/3 of the VOCs did not reduce the AUC of the ROC curve by >10% (99.7% CI). CONCLUSIONS: A test employing WDA of breath VOCs predicted lung cancer with accuracy similar to chest computed tomography. The algorithm identified dependencies that were not apparent with traditional linear methods. WDA appears to provide a useful new technique for non-linear multivariate analysis of data