Inorganic arsenic inhibits the nucleotide excision repair pathway and reduces the expression of XPC

Chronic exposure to arsenic, most often through contaminated drinking water, has been linked to several types of cancer in humans, including skin and lung cancer. However, the mechanisms underlying its role in causing cancer are not well understood. There is evidence that exposure to arsenic can enhance the carcinogenicity of UV light in inducing skin cancers and may enhance the carcinogenicity of tobacco smoke in inducing lung cancers. The nucleotide excision repair (NER) pathway removes different types of DNA damage including those produced by UV light and components of tobacco smoke. The aim of the present study was to investigate the effect of sodium arsenite on the NER pathway in human lung fibroblasts (IMR-90 cells) and primary mouse keratinocytes. To measure NER, we employed a slot-blot assay to quantify the introduction and removal of UV light-induced 6-4 photoproducts (6-4 PP) and cyclobutane pyrimidine dimers (CPDs). We find a concentration-dependent inhibition of the removal of 6-4 PPs and CPDs in both cell types treated with arsenite. Treatment of both cell types with arsenite resulted in a significant reduction in the abundance of XPC, a protein that is critical for DNA damage recognition in NER. The abundance of RNA expressed from several key NER genes was also significantly reduced by treatment of IMR-90 cells with arsenite. Finally, treatment of IMR-90 cells with MG-132 abrogated the reduction in XPC protein, suggesting an involvement of the proteasome in the reduction of XPC protein produced by treatment of cells with arsenic. The inhibition of NER by arsenic may reflect one mechanism underlying the role of arsenic exposure in enhancing cigarette smoke-induced lung carcinogenesis and UV light-induced skin cancer, and it may provide some insights into the emergence of arsenic trioxide as a chemotherapeutic agent.     

IGRT and motion management during lung SBRT delivery

Patient motion can cause misalignment of the tumour and toxicities to the healthy lung tissue during lung stereotactic body radiation therapy (SBRT). Any deviations from the reference setup can miss the target and have acute toxic effects on the patient with consequences onto its quality of life and survival outcomes. Correction for motion, either immediately prior to treatment or intra-treatment, can be realized with image-guided radiation therapy (IGRT) and motion management devices. The use of these techniques has demonstrated the feasibility of integrating complex technology with clinical linear accelerator to provide a higher standard of care for the patients and increase their quality of life.     

Dosimetric effect of uncorrected rotations in lung SBRT with stereotactic imaging guidance

PurposeTo evaluate the dosimetric impact of uncorrected rotations on the planning target volume (PTV) coverage for early stage non-small cell lung cancer patients treated with stereotactic body radiotherapy using Brainlab ExacTrac image guidance.MethodsTwenty-two patients were retrospectively selected. Two scenarios of uncorrected rotations were simulated with magnitude of 1°, 2°, 3° and 5°: (1) rotation around the treatment isocenter; and (2) roll and yaw rotations around a setup isocenter. The D₉₅ of PTV from recalculated dose on the rotated CT was compared to that from the clinical plan. A logistic regression model was used to predict the probability of dose differences between recalculated and original plans that are less than 2% based on the rotation angle, PTV volume, and distance between the treatment and setup isocenter.ResultsLogistic regression model showed the uncorrected isocentric rotations of up to 2.5° in all directions have negligible dosimetric impact. For non-isocentric rotations, a rotational error of 2° may cause significant under-dose of the PTV. Statistically significant (p < 0.05) parameters in the logistic regression model were angle for isocentric rotations, angle and distance for non-isocentric roll rotations, and angle, distance and the PTV volume for non-isocentric yaw rotations.ConclusionsThe severity of the dose deviations due to uncorrected rotations depends on the type and magnitude of the rotation, the volume of the PTV, and the distance between the treatment and setup isocenter, which should be taken into consideration when making clinical judgment of whether the rotational error could be ignored.     

uPARAP expression during murine lung development

Lung remodeling requires active collagen deposition and degradation. Urokinase plasminogen activator receptor-associated protein (uPARAP), or Endo 180, is a cell-surface receptor for collagens, which leads to collagen internalization and degradation. Thus, uPARAP-mediated collagen degradation is an additional pathway for matrix remodeling in addition to matrix remodeling mediated by matrix metalloproteinases and cathepsins. Using immunohistochemistry, we demonstrate extensive uPARAP expression in the mesenchyme throughout murine lung development. By immunofluorescence, we demonstrate significant overlap of uPARAP expression with collagen IV expression, but minimal overlap with collagen I expression in the developing murine lung. Finally, we compared lung development between wild-type and uPARAP(-/-) mice, and found no significant histologic differences, indicating the presence of alternative collagen degradation pathways during murine lung development.     

The role of nitric oxide in reducing deformability of Lewis lung tumor cell stimulated by inflammatory cytokines

Highly metastatic cells, especially in the lungs, are known to be resistant to nitric oxide (NO)-mediated cytotoxicity, compared with poorly or non-metastatic cells. However, the precise mechanisms connecting NO and metastasis remain to be determined. To clarify the role of NO in the characteristic changes in NO-resistant cells in response to inflammatory cytokines, we used Lewis lung tumor (LLT) cells, which are known to be highly metastatic NO-resistant cells, and determined the changes in cell deformability and the gene expression profile after the cells were stimulated using cytokine mixture or an NO donor. Both exogenous NO and endogenous NO via inducible NO synthase produced by cytokines decreased cell deformability by enhancing actin polymerization. The expression of several genes associated with actin polymerization was changed so as to increase actin filaments in the cells by enhancing actin polymerization and by suppressing actin depolymerization, actin filament severing, and barbed-end actin filament capping. In conclusion, inflammatory cytokine stimulation reduces deformability of LLT cells and enhances actin polymerization which is mainly controlled by the same genes induced by NO.     

Protein kinase A-mediated CREB phosphorylation is an oxidant-induced survival pathway in alveolar type II cells

Oxidant stress plays a role in the pathogenesis of pulmonary diseases, including fibrotic lung disease and cancer. We previously found that hydrogen peroxide (H₂O₂) initiates an increase in Ca²⁺/cAMP-response element binding protein (CREB) phosphorylation in C10 alveolar type II cells that requires activation of extracellular regulated kinases 1/2 (ERK1/2). Here, we investigated the role of crosstalk between protein kinase A (PKA) and epidermal growth factor receptor (EGFR) in oxidant-induced signaling to ERK1/2 and CREB in C10 cells. Application of H₂O₂ increased nuclear accumulation of PKA, and inhibition of PKA with H89 reduced oxidant-mediated phosphorylation of both CREB and ERK1/2. Single cell measurements of cAMP and redox status, using a FRET-based biosensor and a redox-sensitive GFP, respectively, indicated that H₂O₂ increases production of cAMP that correlates with redox state. Inhibition of EGFR activity decreased both H₂O₂-induced CREB phosphorylation and translocation of PKA to the nucleus, suggesting that crosstalk between PKA and EGFR underlies the oxidant-induced CREB response. Furthermore, knockdown of CREB expression using siRNA led to a decrease in bcl-2 and an increase in oxidant-induced apoptosis. Together these data reveal a novel role for crosstalk between PKA, ERK1/2 and CREB that mediates cell survival during oxidant stress.     

Autocrine and paracrine purinergic signaling in the most lethal types of cancer

Cancer comprises a collection of diseases that occur in almost any tissue and it is characterized by an abnormal and uncontrolled cell growth that results in tumor formation and propagation to other tissues, causing tissue and organ malfunction and death. Despite the undeniable improvement in cancer diagnostics and therapy, there is an urgent need for new therapeutic and preventive strategies with improved efficacy and fewer side effects. In this context, purinergic signaling emerges as an interesting candidate as a cancer biomarker or therapeutic target. There is abundant evidence that tumor cells have significant changes in the expression of purinergic receptors, which comprise the G-protein coupled P2Y and AdoR families of receptors and the ligand-gated ion channel P2X receptors. Tumor cells also exhibit changes in the expression of nucleotidases and other enzymes involved in nucleotide metabolism, and the concentrations of extracellular nucleotides are significantly higher than those observed in normal cells. In this review, we will focus on the potential role of purinergic signaling in the ten most lethal cancers (lung, breast, colorectal, liver, stomach, prostate, cervical, esophagus, pancreas, and ovary), which together are responsible for more than 5 million annual deaths.     

Changes in serum pneumoproteins caused by short-term exposures to nitrogen trichloride in indoor chlorinated swimming pools

Nitrogen trichloride (NCl₃) is an irritant gas released in the air of indoor pools sanitized with chlorine-based disinfectants. In the present study we investigated the effects of NCl 3 on the pulmonary epithelium of pool attendees by measuring the leakage into serum of three lung-specific proteins (pneumoproteins): the alveolar surfactant-associated proteins A and B (SP-A and SP-B) and the bronchiolar 16 kDa Clara cell protein (CC16). These pneumoproteins were measured in the serum of 29 recreational swimmers (16 children and 13 adults) before and after attending a chlorinated pool with a mean NCl₃ concentration of 490 µg m^-3. Pneumoprotein changes in serum were also studied in 14 trained swimmers performing an intensive 45 min standardized swimming session in a chlorinated pool (mean NCl₃ concentration of 355 µg m^-3) and for the purposes of comparison in a non-chlorinated pool sanitized by the copper/silver method. Serum CC16 was not increased in recreational swimmers, but in trained swimmers serum levels of this protein peaked immediately after strenuous exercise, both in the copper/silver pool and in the chlorinated pool. This acute increase in airway permeability is probably the consequence of the mechanical stress on the epithelial barrier caused by overinflation and/or hyperventilation during intense exercise. Serum levels of SP-A and SP-B were unaffected by strenuous exercise in the copper/silver pool. The two proteins were, however, significantly increased in a time-dependent manner in recreational and trained swimmers attending the chlorinated pool. The intravascular leakage of SP-A and SP-B was already statistically significant after only 1 h of exposure to pool air without exercising and remained elevated for 12 h after. These changes were not associated with decrements in lung function. The ability of NCl₃ to acutely disrupt the lung epithelium barrier was confirmed in mice using serum CC16 and plasma proteins in bronchoalveolar lavage fluid as permeability markers. The significance of these permeability changes induced by NCl₃ in the deep lung is presently unknown. In view of the increasing and widespread human exposure to this gas not only in indoor pools but also in a variety of other situations, these findings warrant further study.