In vivo mutagenesis induced by benzo[a]pyrene instilled into the lung of gpt delta transgenic mice

Benzo[a]pyrene (B[a]P) is a ubiquitous airborne pollutant whose mutagenicity has been evaluated previously by oral and intraperitoneal administration to experimental animals. In this study, mutagenesis in the lungs, the target organ of air pollutants, was examined after a single intratracheal instillation of 0-2 mg B[a]P into gpt delta transgenic mice. Intratracheal injection of B[a]P resulted in a statistically significant and dose-dependent increase in gpt mutant frequency as measured by 6-thioguanine selection. The mutant frequencies at B[a]P doses of 0.5, 1, and 2 mg were 2.8, 4.2, and 6.8 times higher than the frequency seen in nontreated mice (0.60 +/- 0.13 x 10(-5)). The most frequent mutations induced by B[a]P treatment were G:C-->T:A transversions, which are characteristic of B[a]P mutagenesis in other models, and single-base deletions of G:C base pairs. To characterize the hotspots of B[a]P-induced mutations in the gpt gene, we analyzed sequences adjacent to the mutated G:C base pairs. Guanine bases centered in the nucleotide sequences CGT, CGA, and CGG were the most frequent targets of B[a]P. Our results indicate that intratracheal instillation of B[a]P into gpt delta mice causes a dose-dependent increase in gpt mutant frequency in the lung, and that the predominant mutation induced is G:C-->T:A transversion.     

Pranlukast, a leukotriene receptor antagonist, inhibits interleukin-5 production via a mechanism distinct from leukotriene receptor antagonism

BACKGROUND: Pranlukast, a cysteinyl leukotriene receptor 1 (CysLTR1) antagonist, inhibits not only airway smooth muscle contraction, but also allergic inflammation. The aim of this study was to determine the mechanism of pranlukast-induced interleukin-5 (IL-5) inhibition in allergic inflammation. METHODS: Surgically resected human lung tissue was passively sensitized in vitro with mite-allergen-sensitized sera, followed by stimulation with mite allergen after pretreatment of the tissue with pranlukast, dexamethasone, or both. The IL-5 protein level in the culture medium was measured, and in situ hybridization of IL-5 and CysLTR1 mRNA was performed using lung tissues. RESULTS: Pretreatment of lung tissues with pranlukast alone significantly decreased the amount of IL-5 protein in the culture medium by 40%. The combination of pranlukast and dexamethasone synergistically enhanced this effect. Quantitative in situ hybridization with image analysis revealed abundant expression of IL-5 mRNA in eosinophils, lymphocytes, and mast cells in sensitized and allergen-stimulated lung tissues. CysLTR1 mRNA was detected in macrophages, smooth muscle cells, eosinophils, and mast cells, but was less expressed in lymphocytes. Pranlukast-induced inhibition of IL-5 mRNA expression was noted in various cells, irrespective of their CysLTR1 mRNA expression status. In addition, cysteinyl leukotrienes per se failed to upregulate the IL-5 production. CONCLUSION: Our results indicate that pranlukast inhibits IL-5 synthesis via a mechanism distinct from CysLTR1 antagonism.     

SAR by MS: discovery of a new class of RNA-binding small molecules for the hepatitis C virus: internal ribosome entry site IIA subdomain

A new class of small molecules that bind the HCV RNA IRES IIA subdomain with sub-micromolar affinity is reported. The benzimidazole 'hit' 1 with a KD approximately 100 microM to a 29-mer RNA model of Domain IIA was identified from a 180000-member library using mass spectrometry-based screening methods. Further MS-assisted SAR (structure-activity relationships) studies afforded benzimidazole derivatives with sub-micromolar binding affinity for the IIA RNA construct. The optimized benzimidazoles demonstrated activity in a cellular replicon assay at concentrations comparable to their KD for the RNA target.     

Smoking cessation increases gingival blood flow and gingival crevicular fluid

OBJECTIVES: The purpose of the present study was to determine the effect of smoking cessation on gingival blood flow (GBF) and gingival crevicular fluid (GCF). MATERIAL AND METHODS: Sixteen male smokers (aged 22-39 (25.3+/-4.0) years), with no clinical signs of periodontal and systemic diseases, were recruited. The experiment was performed before (baseline) and at 1, 3 and 5 days, and at 1, 2, 4 and 8 weeks after smoking cessation. The status of smoking and smoking cessation was verified by exhaled carbon monoxide (CO) concentration, and by serum nicotine and cotinine concentrations. A laser Doppler flowmeter was used to record relative blood flow continuously, on three gingival sites of the left maxillary central incisor (mid-labial aspect of the gingival margin and bilateral interdental papillae). The GCF was collected at the mesio- and disto-labial aspects of the left maxillary central incisor and the volume was calculated by the Periotron 6000(R) system. The same measurements except for the GBF were performed on 11 non-smoking controls (four females and seven males), aged 23-27 (24.4+/-1.2) years. RESULTS: Eleven of 16 smokers successfully completed smoking cessation for 8 weeks. At 1 day after smoking cessation, there was a significantly lower CO concentration than at baseline (p<0.01). Also, nicotine and cotinine concentrations markedly decreased at the second measurement. The GBF rate of smokers was significantly higher at 3 days after smoking cessation compared to the baseline (p<0.01). While the GCF volume was significantly increased at 5 days after smoking cessation compared to the baseline (p<0.01), it was significantly lower than that of non-smokers until 2 weeks after smoking cessation (p<0.01). CONCLUSION: The results show that the gingival microcirculation recovers to normal in the early stages of smoking cessation, which could activate the gingival tissues metabolism/remodeling, and contribute to periodontal health.     

Real-time endobronchial ultrasound-guided transbronchial needle aspiration of mediastinal and hilar lymph nodes

STUDY OBJECTIVES: Although various techniques are available for obtaining pathology specimens from the mediastinal lymph nodes, including conventional bronchoscopic transbronchial needle aspiration (TBNA), transesophageal ultrasonography-guided needle aspiration, and mediastinoscopy, there are limitations to these techniques, which include low yield, poor access, need for general anesthesia, or complications. To overcome these problems, we undertook the current study to evaluate the clinical utility of the newly developed ultrasound puncture bronchoscope to visualize and perform real-time TBNA of the mediastinal and hilar lymph nodes under direct endobronchial ultrasonography (EBUS) guidance. DESIGN: Prospective patient enrollment. SETTING: University teaching hospital. PATIENTS: From March 2002 to September 2003, 70 patients were included in the study. INTERVENTIONS: The new convex probe (CP) EBUS is integrated with a convex scanning probe on its tip with a separate working channel, thus permitting real-time EBUS-guided TBNA. The indications for CP-EBUS were the diagnosis of mediastinal and/or hilar lymphadenopathy for known or suspected malignancy. Lymph nodes and the surrounding vessels were first visualized with CP-EBUS using the Doppler mode. The dimensions of the lymph nodes were recorded, followed by real-time TBNA under direct EBUS guidance. Final diagnosis was based on cytology, surgical results, and/or clinical follow-up. RESULTS: All lymph nodes that were detected on the chest CT scan could be visualized using CP-EBUS. In 70 patients, CP-EBUS-guided TBNA was performed to obtain samples from mediastinal lymph nodes (58 nodes) and hilar lymph nodes (12 nodes). The sensitivity, specificity, and accuracy of CP-EBUS-guided TBNA in distinguishing benign from malignant lymph nodes were 95.7%, 100%, and 97.1%, respectively. The procedure was uneventful, and there were no complications. CONCLUSIONS: Real-time CP-EBUS-guided TBNA of mediastinal and hilar lymph nodes is a novel approach that is safe and has a good diagnostic yield. This new ultrasound puncture bronchoscope has an excellent potential for assisting in safe and accurate diagnostic interventional bronchoscopy.     

The effects of fluorescent lamp coated with a photocatalyst film on the number of floating microorganisms or smell molecules in a room installed with the lamp

Fluorescent lamp that is coated with a photocatalyst film, titanium dioxide (TiO2), can catalyze the microorganisms and smell producing molecules that touch the membrane. In this report, we examined the effect of this coated lamp by the number of floating microorganisms or smell molecules in a room installed with this lamp. The number of floating microorganisms was examined independently in different laboratories before and after installing the lamps in an animal facility and a vegetable processing room of a food company. We found that the number of floating microorganisms was significantly reduced after the installation of this coated lamp. To test for smell molecules, acetaldehyde was injected into a closed chamber installed with the coated lamp and the result was compared to a control where a regular fluorescent lamp was used. The acetaldehyde concentration did not change with the regular lamp but was halved after 1h with the photocatalyst coated lamp. To test the effect in real settings, questionnaires were filled out by restroom users in several different places. About 90% of those surveyed answered that there was a reduction of smell after the installation of the photocatalyst coated lamp. We concluded that the installation of this coated fluorescent lamp reduced not only the floating microorganisms but the smell in various practical situations.     

Verapamil prevents impairment in filterability of human erythrocytes exposed to oxidative stress

Effects of oxidative stress on intact human erythrocytes were investigated using tert-butyl hydroperoxide (tBHP). Exposure of erythrocytes to tBHP caused a marked decrease in filterability in a time-dependent manner. Erythrocytes exposed to tBHP also show an increase in mean corpuscular volume and a remarkable formation of methemoglobin (met-Hb) without any appearance of hemichromes that form Heinz bodies. High performance liquid chromatography demonstrated that the tBHP-treated erythrocytes exhibited an apparent decrease in the membrane phospholipid, phosphatidylethanolamine (PE). The decrease in PE was inhibited by pretreatment with ascorbate, but not with verapamil. SDS-polyacrylamide gel electrophoresis of the tBHP-treated erythrocyte membrane showed a degradation of spectrin, band 3, band 4.2, and band 4.5, accompanied by the appearance of low-molecular-weight products. The degradation of the membrane proteins was not prevented by pretreatment with verapamil or ascorbate. However, the pretreatment with verapamil but not with ascorbate revealed significant inhibition of the tBHP-induced impairment in filterability in the presence of extracellular Ca2+. Thus, the present study shows that verapamil, a potent drug in reperfusion therapy, plays an important role in protection against oxidative injury, based on a close linkage among decreased filterability, met-Hb formation, and impaired membrane integrity.