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Assessing wetland vulnerability to chronic and episodic physical drivers is fundamental for establishing restoration priorities. We synthesized multiple data sets from E.B. Forsythe National Wildlife Refuge, New Jersey, to establish a wetland vulnerability metric that integrates a range of physical processes, anthropogenic impact and physical/biophysical features. The geospatial data are based on aerial imagery, remote sensing, regulatory information, and hydrodynamic modeling; and include elevation, tidal range, unvegetated to vegetated marsh ratio (UVVR), shoreline erosion, potential exposure to contaminants, residence time, marsh condition change, change in salinity, salinity exposure and sediment concentration. First, we delineated the wetland complex into individual marsh units based on surface contours, and then defined a wetland vulnerability index that combined contributions from all parameters. We applied principal component and cluster analyses to explore the interrelations between the data layers, and separate regions that exhibited common characteristics. Our analysis shows that the spatial variation of vulnerability in this domain cannot be explained satisfactorily by a smaller subset of the variables. The most influential factor on the vulnerability index was the combined effect of elevation, tide range, residence time, and UVVR. Tide range and residence time had the highest correlation, and similar bay-wide spatial variation. Some variables (e.g., shoreline erosion) had no significant correlation with the rest of the variables. The aggregated index based on the complete dataset allows us to assess the overall state of a given marsh unit and quickly locate the most vulnerable units in a larger marsh complex. The application of geospatially complete datasets and consideration of chronic and episodic physical drivers represents an advance over traditional point-based methods for wetland assessment.

High energy laser, particularly 532 nm, is widely used in defense and medical applications and there is need to address its occupational safety. Thermal and non-thermal effects of 532 nm high energy laser on skin are cause of concern. This study indicates impact of 532 nm laser on rat skin and first of its kind of attempt to understand transcriptional activation of genes as an early response following laser exposure. Skin of experimental rats were exposed to 532 nm radiance at 0.1, 0.25 and 0.50 W/cm2 for 10 sec. Thermographic changes of skin exposed to 532 nm laser exhibited increased Tmax temperature in radiance dependent manner. After thermal imaging, skin of experimental rats was collected 1 h post laser exposure for studying differential gene expression. The skin exposed to lower power density (0.1 W/cm2) did not show significant changes in expression of gene pathways studied. At moderate radiance (0.25 W/cm2), predominantly canonical wnt/B-catenin pathway genes notch1, axin2, ccdn1, wnt5a and redox homeostasis genes; txn1, nqo1 and txnrd1 were expressed. At higher radiance (0.5 W/cm2), significant repression of genes related to wound healing process particularly notch/wnt pathway viz. hes5, wnt1, wn3b with higher expression of dab2 was recorded. The data obtained from these studies would help in drawing safety limits for skin exposure to 532 nm laser. Further, genes expressed at moderate and high level of radiance exposure to skin were distinct and differential and provide new avenue to configure pathway to counteract laser induced delay in tissue injury and hair follicular damage.

BACKGROUND:High altitude associated hypobaric hypoxia is one of the cellular and environmental perturbation that alters proteostasis network and push the healthy cell towards loss of muscle mass. The present study has elucidated the robust proteostasis network and signaling mechanism for skeletal muscle atrophy under chronic hypobaric hypoxia (CHH). METHODS:Male Sprague Dawley rats were exposed to simulated hypoxia equivalent to a pressure of 282 torr for different durations (1, 3, 7 and 14 days). After CHH exposure, skeletal muscle tissue was excised from the hind limb of rats for biochemical analysis. RESULTS:Chronic hypobaric hypoxia caused a substantial increase in protein oxidation and exhibited a greater activation of ER chaperones, glucose-regulated protein-78 (GRP-78) and protein disulphide isomerase (PDI) till 14d of CHH. Presence of oxidized proteins triggered the proteolytic systems, 20S proteasome and calpain pathway which were accompanied by a marked increase in [Ca2+]. Upregulated Akt pathway was observed upto 07d of CHH which was also linked with enhanced glycogen synthase kinase-3β (GSk-3β) expression, a negative regulator of Akt. Muscle-derived cytokines, tumor necrosis factor-α (TNF-α), interferon-ϒ (IFN-©) and interleukin-1β (IL-1β) levels significantly increased from 07d onwards. CHH exposure also upregulated the expression of nuclear factor kappa-B (NF-κB) and E3 ligase, muscle atrophy F-box-1 (Mafbx-1/Atrogin-1) and MuRF-1 (muscle ring finger-1) on 07d and 14d. Further, severe hypoxia also lead to increase expression of ER-associated degradation (ERAD) CHOP/ GADD153, Ub-proteasome and apoptosis pathway. CONCLUSIONS:The disrupted proteostasis network was tightly coupled to degradative pathways, altered anabolic signaling, inflammation, and apoptosis under chronic hypoxia. Severe and prolonged hypoxia exposure affected the protein homeostasis which overwhelms the muscular system and tends towards skeletal muscle atrophy.

Hypoxia induced oxidative stress incurs pathophysiological changes in hypertrophied cardiomyocytes by promoting translocation of p53 to mitochondria. Here, we investigate the cardio-protective efficacy of nanocurcumin in protecting primary human ventricular cardiomyocytes (HVCM) from hypoxia induced damages. Hypoxia induced hypertrophy was confirmed by FITC-phenylalanine uptake assay, atrial natriuretic factor (ANF) levels and cell size measurements. Hypoxia induced translocation of p53 was investigated by using mitochondrial membrane permeability transition pore blocker cyclosporin A (blocks entry of p53 to mitochondria) and confirmed by western blot and immunofluorescence. Mitochondrial damage in hypertrophied HVCM cells was evaluated by analysing bio-energetic, anti-oxidant and metabolic function and substrate switching form lipids to glucose. Nanocurcumin prevented translocation of p53 to mitochondria by stabilizing mitochondrial membrane potential and de-stressed hypertrophied HVCM cells by significant restoration in lactate, acetyl-coenzyme A, pyruvate and glucose content along with lactate dehydrogenase (LDH) and 5' adenosine monophosphate-activated protein kinase (AMPKα) activity. Significant restoration in glucose and modulation of GLUT-1 and GLUT-4 levels confirmed that nanocurcumin mediated prevention of substrate switching. Nanocurcumin prevented of mitochondrial stress as confirmed by c-fos/c-jun/p53 signalling. The data indicates decrease in p-300 histone acetyl transferase (HAT) mediated histone acetylation and GATA-4 activation as pharmacological targets of nanocurcumin in preventing hypoxia induced hypertrophy. The study provides an insight into propitious therapeutic effects of nanocurcumin in cardio-protection and usability in clinical applications.

Metastatic melanoma is the most aggressive form of this cancer. It is important to understand factors that increase or decrease metastatic activity in order to more effectively research and implement treatments for melanoma. Increased cell invasion through the extracellular matrix is required for metastasis and is enhanced by matrix metalloproteinases (MMPs). Tissue inhibitor of metalloproteinases 3 (TIMP3) inhibits MMP activity. It was previously shown by our group that miR-21, a potential regulator of TIMP3, is over-expressed in cutaneous melanoma. It was therefore hypothesized that increased levels of miR-21 expression would lead to decreased expression of TIMP3 and thereby enhance the invasiveness of melanoma cells. miR-21 over-expression in the melanoma cell lines WM1552c, WM793b, A375 and MEL 39 was accomplished via transfection with pre-miR-21. Immunoblot analysis of miR-21-overexpressing cell lines revealed reduced expression of TIMP3 as compared to controls. This in turn led to a significant increase in the invasiveness of the radial growth phase cell line WM1552c and the vertical growth phase cell line WM793b (p < 0.05), but not in the metastatic cell lines A375 or MEL 39. The proliferation and migration of miR-21 over-expressing cell lines was not affected. Reduced expression of TIMP3 was achieved by siRNA knockdown and significantly enhanced invasion of melanoma cell lines, mimicking the effects of miR-21 over-expression. Treatment of tumor cells with a linked nucleic acid antagomir to miR-21 inhibited tumor growth and increased tumor expression of TIMP3 in vivo in 01B74 Athymic NCr-nu/nu mice. Intra-tumoral injections of anti-miR-21 produced similar effects. This data shows that increased expression of miR-21 enhanced the invasive potential of melanoma cell lines through TIMP3 inhibition. Therefore, inhibition of miR-21 in melanoma may reduce melanoma invasiveness.

Patient-derived xenograft (PDX) models generated from surgical specimens are gaining popularity as preclinical models of cancer. However, establishment of PDX lines from small cell lung cancer (SCLC) patients is difficult due to very limited amount of available biopsy material. We asked whether SCLC cells obtained from endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) could generate PDX lines that maintained the phenotypic and genetic characteristics of the primary tumor. Following successful EBUS-TBNA sampling for diagnostic purposes, we obtained an extra sample for cytologic analysis and implantation into the flanks of immunodeficient mice. Animals were monitored for engraftment for up to 6 months. Histopathologic and immunohistochemical analysis, and targeted next-generation re-sequencing, were then performed in both the primary sample and the derivative PDX line. A total of 12 patients were enrolled in the study. EBUS-TBNA aspirates yielded large numbers of viable tumor cells sufficient to inject between 18,750 and 1,487,000 cells per flank, and to yield microgram quantities of high-quality DNA. Of these, samples from 10 patients generated xenografts (engraftment rate 83%) with a mean latency of 104 days (range 63-188). All but one maintained a typical SCLC phenotype that closely matched the original sample. Identical mutations that are characteristic of SCLC were identified in both the primary sample and xenograft line. EBUS-TBNA has the potential to be a powerful tool in the development of new targeting strategies for SCLC patients by providing large numbers of viable tumor cells suitable for both xenografting and complex genomic analysis.

BackgroundGuatemala is currently undergoing an epidemiologic transition. Preventive services are key to reducing the burden of non-communicable diseases, and smoking counseling and cessation are among the most cost-effective and wide-reaching strategies. Internal medicine physicians are fundamental to providing such services, and their knowledge is a cornerstone of non-communicable disease control.MethodsA national cross-sectional survey was conducted in 2011 to evaluate knowledge of clinical preventive services for non-communicable diseases. Interns, residents, and attending physicians of the internal medicine departments of all teaching hospitals in Guatemala completed a self-administered questionnaire. Participants' responses were contrasted with the Guatemalan Ministry of Health (MoH) prevention guidelines and the US Preventive Services Task Force (USPSTF) recommendations. Analysis compared knowledge of recommendations within and between hospitals.ResultsIn response to simulated patient scenarios, all services were recommended by more than half of physicians regardless of MoH or USPSTF recommendations. Prioritization was adequate according to the MoH guidelines but not including other potentially effective services (e.g. colorectal cancer and lipid disorder screenings). With the exception of colorectal and prostate cancer screening, less frequently recommended by interns, there was no difference in recommendation rates by level.ConclusionGuatemalan internal medicine physicians' knowledge on preventive services recommendations for non-communicable diseases is limited, and prioritization did not reflect cost-effectiveness. Based on these data we recommend that preventive medicine training be strengthened and development of evidence-based guidelines for low-middle income countries be a priority.

CD200 is a cell surface glycoprotein that functions through engaging CD200 receptor on cells of the myeloid lineage and inhibits their functions. Expression of CD200 has been implicated in a variety of human cancer cells including melanoma cells and has been thought to play a protumor role. To investigate the role of cancer cell expression of CD200 in tumor formation and metastasis, we generated CD200-positive and CD200-negative B16 melanoma cells. Subcutaneous injection of CD200-positive B16 melanoma cells inhibited tumor formation and growth in C57BL/6 mice but not in Rag1⁻/⁻C57BL/6 mice. However, i.v. injection of CD200-positive B16 melanoma cells dramatically inhibited tumor foci formation in the lungs of both C57BL/6 and Rag1⁻/⁻C57BL6 mice. Flow cytometry analysis revealed higher expression of CD200R in Gr1⁺ myeloid cells in the lung than in peripheral myeloid cells. Depletion of Gr1⁺ cells or stimulation of CD200R with an agonistic antibody in vivo dramatically inhibited tumor foci formation in the lungs. In addition, treatment with tumor antigen specific CD4 or CD8 T cells or their combination yielded a survival advantage for CD200 positive tumor bearing mice over mice bearing CD200-negative tumors. Taken together, we have revealed a novel role for CD200-CD200R interaction in inhibiting tumor formation and metastasis. Targeting CD200R may represent a novel approach for cancer immunotherapy.

BACKGROUND:The pro-fibrogenic cytokine connective tissue growth factor (CTGF) plays an important role in the development and progression of fibrosis in many organ systems, including liver. However, its role in the pathogenesis of hepatitis C virus (HCV)-induced liver fibrosis remains unclear. METHODS:In the present study, we assessed CTGF expression in HCV-infected hepatocytes using replicon cells containing full-length HCV genotype 1 and the infectious HCV clone JFH1 (HCV genotype 2) by real-time PCR, Western blot analysis and confocal microscopy. We evaluated transforming growth factor β1 (TGF-β1) as a key upstream mediator of CTGF production using neutralizing antibodies and shRNAs. We also determined the signaling molecules involved in CTGF production using various immunological techniques. RESULTS:We demonstrated an enhanced expression of CTGF in two independent models of HCV infection. We also demonstrated that HCV induced CTGF expression in a TGF-β1-dependent manner. Further dissection of the molecular mechanisms revealed that CTGF production was mediated through sequential activation of MAPkinase and Smad-dependent pathways. Finally, to determine whether CTGF regulates fibrosis, we showed that shRNA-mediated knock-down of CTGF resulted in reduced expression of fibrotic markers in HCV replicon cells. CONCLUSION:Our studies demonstrate a central role for CTGF expression in HCV-induced liver fibrosis and highlight the potential value of developing CTGF-based anti-fibrotic therapies to counter HCV-induced liver damage.

Cannabinoids bind to cannabinoid receptors CB(1) and CB(2) and have been reported to possess anti-tumorigenic activity in various cancers. However, the mechanisms through which cannabinoids modulate tumor growth are not well known. In this study, we report that a synthetic non-psychoactive cannabinoid that specifically binds to cannabinoid receptor CB(2) may modulate breast tumor growth and metastasis by inhibiting signaling of the chemokine receptor CXCR4 and its ligand CXCL12. This signaling pathway has been shown to play an important role in regulating breast cancer progression and metastasis.We observed high expression of both CB(2) and CXCR4 receptors in breast cancer patient tissues by immunohistochemical analysis. We further found that CB(2)-specific agonist JWH-015 inhibits the CXCL12-induced chemotaxis and wound healing of MCF7 overexpressing CXCR4 (MCF7/CXCR4), highly metastatic clone of MDA-MB-231 (SCP2) and NT 2.5 cells (derived from MMTV-neu) by using chemotactic and wound healing assays. Elucidation of the molecular mechanisms using various biochemical techniques and confocal microscopy revealed that JWH-015 treatment inhibited CXCL12-induced P44/P42 ERK activation, cytoskeletal focal adhesion and stress fiber formation, which play a critical role in breast cancer invasion and metastasis. In addition, we have shown that JWH-015 significantly inhibits orthotopic tumor growth in syngenic mice in vivo using NT 2.5 cells. Furthermore, our studies have revealed that JWH-015 significantly inhibits phosphorylation of CXCR4 and its downstream signaling in vivo in orthotopic and spontaneous breast cancer MMTV-PyMT mouse model systems.This study provides novel insights into the crosstalk between CB(2) and CXCR4/CXCL12-signaling pathways in the modulation of breast tumor growth and metastasis. Furthermore, these studies indicate that CB(2) receptors could be used for developing innovative therapeutic strategies against breast cancer.

S100A7 is a small calcium binding protein, which has been shown to be differentially expressed in psoriatic skin lesions, as well as in squamous cell tumors of the skin, lung and breast. Although its expression has been correlated to HER+ high-grade tumors and to a high risk of progression, the molecular mechanisms of these S100A7-mediated tumorigenic effects are not well known. Here, we showed for the first time that epidermal growth factor (EGF) induces S100A7 expression in both MCF-7 and MDA-MB-468 cell lines. We also observed a decrease in EGF-directed migration in shRNA-downregulated MDA-MB-468 cell lines. Furthermore, our signaling studies revealed that EGF induced simultaneous EGF receptor phosphorylation at Tyr1173 and HER2 phosphorylation at Tyr1248 in S100A7-downregulated cell lines as compared to the vector-transfected controls. In addition, reduced phosphorylation of Src at tyrosine 416 and p-SHP2 at tyrosine 542 was observed in these downregulated cell lines. Further studies revealed that S100A7-downregulated cells had reduced angiogenesis in vivo based on matrigel plug assays. Our results also showed decreased tumor-induced osteoclastic resorption in an intra-tibial bone injection model involving SCID mice. S100A7-downregulated cells had decreased osteoclast number and size as compared to the vector controls, and this decrease was associated with variations in IL-8 expression in in vitro cell cultures. This is a novel report on the role of S100A7 in EGF-induced signaling in breast cancer cells and in osteoclast formation.