LDOC1 silenced by cigarette exposure and involved in oral neoplastic transformation.

Previously, we identified global epigenetic aberrations in smoking-associated oral squamous cell carcinoma (OSCC). We hypothesized that cigarette exposure triggers OSCC through alteration of the methylome of oral cells. Here we report that cigarette smoke condensate (CSC) significantly changes the genomic 5-methyldeoxycytidine content and nuclear accumulation of DNA methyltransferase 1 (DNMT1) and DNMT3A in human untransformed oral cells. By using integrated analysis of cDNA and methylation arrays of the smoking-associated dysplastic oral cell line and OSCC tumors, respectively, we identified four epigenetic targets--UCHL1, GPX3, LXN, and LDOC1--which may be silenced by cigarette. Results of quantitative methylation-specific PCR showed that among these four genes, LDOC1 promoter was the most sensitive to CSC. LDOC1 promoter hypermethylation and gene silencing followed 3 weeks of CSC treatment. LDOC1 knockdown led to a proliferative response and acquired clonogenicity of untransformed oral cells. Immunohistochemistry showed that LDOC1 was downregulated in 53.3% (8/15) and 57.1% (20/35) of premalignant oral tissues and early stage OSCCs, respectively, whereas 76.5% (13/17) of normal oral tissues showed high LDOC1 expression. Furthermore, the microarray data showed that LDOC1 expression had decreased in the lung tissues of current smokers compared with that in those of never smokers and had significantly decreased in the lung tumors of smokers compared with that in normal lung tissues. Our data suggest that CSC-induced promoter methylation may contribute to LDOC1 downregulation, thereby conferring oncogenic features to oral cells. These findings also imply a tumor suppressor role of LDOC1 in smoking-related malignancies such as OSCC and lung cancer.

Identification of trans,trans-2,4-decadienal metabolites in mouse and human cells using liquid chromatography-mass spectrometry.

trans,trans-2,4-Decadienal (tt-DDE), a lipid peroxidation product of linolieic acid, is the most abundant aldehyde identified in cooking oil fumes and is readily detectable in food products as well as in restaurant emissions. Previously, we have reported the toxicological effects of tt-DDE in vitro and in vivo. However, the metabolic pathways of tt-DDE in vivo remain unclear. In our present study, we combined liquid chromatography-mass spectrometry with triple quadrupole and time-of-flight to identify tt-DDE metabolites in the urine of mice orally administered tt-DDE. We identified two tt-DDE metabolites, 2,4-decadienoic acid and cysteine-conjugated 2,4-decadien-1-ol, in the urine of mice gavaged with tt-DDE and in human hepatoma cell cultures. The structure of 2,4-decadienoic acid was confirmed upon comparison of its tandem mass spectrometry (MS/MS) spectrum and retention time with those of synthetic standards. The moieties of cysteine and alcohol on cysteine-conjugated 2,4-decadien-1-ol were validated by treating cell cultures with stable-isotope-labeled cysteine and 4-methylpyrazole, an alcohol dehydrogenase inhibitor. The MS/MS spectra of a cysteine standard and ionized cysteine detached from cysteine-conjugated 2,4-decadien-1-ol were identical. Two metabolic pathways for the biotransformation of tt-DDE in vivo are proposed: (i) the oxidation of tt-DDE to the corresponding carboxylic acid, 2,4-decadienoic acid, in liver cells and (ii) glutathione (GHS) conjugation, GSH breakdown, and aldehyde reduction, which generate cysteine-conjugated 2,4-decadien-1-ol in both liver and lung cells. In conclusion, this platform can be used to identify tt-DDE metabolites, and cysteine-conjugated 2,4-decadien-1-ol can serve as a biomarker for assessing exposure to tt-DDE.

Aryl hydrocarbon receptor is a target of 17-Allylamino-17-demethoxygeldanamycin and enhances its anticancer activity in lung adenocarcinoma cells.

We have demonstrated that aryl hydrocarbon receptor (AhR) is overexpressed in lung adenocarcinoma (AD). AhR is usually associated with heat shock protein 90 (Hsp90) in the cytoplasm. 17-Allylamino-17-demethoxygeldanamycin (17-AAG), an Hsp90 inhibitor, is currently under evaluation for its anticancer activity in clinical trials. Here we investigated whether AhR plays a role in 17-AAG-mediated anticancer activity by functioning as a downstream target or by modulating its anticancer efficacy. AhR expression in lung AD cells was modulated by siRNA interference or overexpression. Tumor growth was determined with colony formation in vitro or in vivo. Anticancer activity of 17-AAG was determined by measuring cell viability, cell cycle distribution, and expression of cell cycle regulators. Proteins and mRNA levels were examined by immunoblotting and the real-time reverse transcription-polymerase chain reaction, respectively. In this study, AhR overexpression positively modulated growth of lung AD cells, at least partially, via RelA-dependent mechanisms. Although treatment with 17-AAG reduced AhR levels and AhR-regulated gene expression in lung AD cells, AhR expression increased anticancer activity of 17-AAG. In addition, 17-AAG treatment reduced cell viability, CDK2, CDK4, cyclin E, cyclin D1, and phosphorylated Rb levels in AhR-expressing lung AD cells. NAD(P)H:quinone oxidoreductase (NQO1), which is regulated by AhR, was shown to increase anticancer activity of 17-AAG in cells. Knockdown of NQO1 expression attenuated the reduction of cell cycle regulators by 17-AAG treatment in AhR overexpressed cells. We demonstrated that AhR protein not only functions as a downstream target of 17-AAG, but also enhances anticancer activity of 17-AAG in lung AD cells.

Hypersensitivity of lung vagal C fibers induced by acute intermittent hypoxia in rats: role of reactive oxygen species and TRPA1.

Obstructive sleep apnea, manifested by intermittent hypoxia and excess production of reactive oxygen species (ROS) in airways, is associated with hyperreactive airway diseases, but the mechanism remains unclear. Sensitization of lung vagal C fibers (LVCFs) contributes to the airway hypersensitivity. We investigated the mechanisms underlying the sensitization of LVCFs with acute intermittent hypoxia (AIH), by 10 episodes of exposure to 30 s of hypoxic air (0%, 5%, or 10% O(2)) followed by 30 s of room air in anesthetized, open-chest, and artificially ventilated rats. Reflex apneic response to intravenous capsaicin (an LVCF stimulant), as measured by phrenic nerve activity, was concentration dependently augmented by AIH. Similarly, reflex apneic response to intravenous α,ß-methylene-ATP (another LVCF stimulant) was augmented by AIH (0% O(2)). The reflex apnea evoked by these two stimulants was abolished by bilateral vagotomy, which suggests the involvement of lung vagal afferents. The AIH-augmented apneic response to these two stimulants was prevented by pretreatment with dimethylthiourea (a hydroxyl radical scavenger), N-acetyl-l-cysteine (an antioxidant) and HC-030031 [a transient receptor potential ankyrin 1 (TRPA1) receptor antagonist]. Consistently, electrophysiological study revealed the afferent responses of LVCFs to capsaicin or α,ß-methylene-ATP were augmented by AIH, and this sensitization of LVCFs was prevented by dimethylthiourea, N-acetyl-l-cysteine, and HC-030031. In contrast, AIH did not alter the afferent response of LVCFs to mechanical stimulation by lung hyperinflation. We concluded that AIH sensitizes LVCFs in rats, thus resulting in exaggerated airway reflexogenic responses to chemical stimulants, possibly by ROS action and activation of TRPA1 receptors.

Changes of dipalmitoyl phosphatidyl choline after mechanical ventilation in patients with acute cerebral injury.

OBJECTIVE: To detect the levels of dipalmitoyl phosphatidyl choline (DPPC) in the sputum of the patients with acute cerebral injury without primary pulmonary injury after mechanical ventilation treatment. METHODS: DPPC levels in sputum of 35 patients with acute cerebral injury but without pulmonary injury were detected with high performance liquid chromatography at the beginning of ventilation and 16-20 days, 21-40 days, and 41-60 days after ventilation, respectively. RESULTS: There was no significant difference of the DPPC levels between 16-20 days after ventilation (3.36+/-0.49) and at the beginning of ventilation (3.37+/-0.58) (P>0.05). The mean levels of DPPC decreased significantly at 21-40 days (2.87 mg/ml+/-0.26 mg/ml, P<0.05) and 41-60 days (1.93 mg/ml+/-0.21 mg/ml, P<0.01) after ventilation compared with that at the beginning of ventilation. At the same period, the peak inspiratory pressure and the mean pressure of airway increased significantly, whereas the static compliance and the partial pressure of oxygen in artery decreased significantly. Among the 25 patients who received ventilation for more than 20 days, 8 (32%) had slightly-decreased partial pressure of oxygen in artery compared with that at the beginning of ventilation. CONCLUSIONS: Mechanical ventilation can decrease the DPPC levels, decrease the lung compliance and increase the airway pressure, even impair the oxygenation function in patients with acute cerebral injury. Abnormal DPPC is one of the major causes of ventilator-associated lung injury.

Comparative study of pressure-control ventilation and volume-control ventilation in treating traumatic acute respiratory distress syndrome.

OBJECTIVE: To observe the clinical therapeutic effect and side effect of pressure-control ventilation (PCV) on traumatic acute respiratory distress syndrome (ARDS) compared with volume-control ventilation (VCV). METHODS: Forty patients with traumatic ARDS were hospitalized in our department from June 1996 to December 2002. Twenty were treated with PCV (PCV group) and 20 with VCV (VCV group). The changes of the peak inflating pressure and the mean pressure of the airway were observed at the very beginning of the mechanical ventilation and the following 12 and 24 hours, respectively. The transcutaneous saturation of oxygen pressure, the pressure of oxygen in artery, the mean blood pressure, the central venous pressure, the heart rate and the incidence of the pressure injury were also monitored before ventilation and 12 hours after ventilation. RESULTS: The pressure of oxygen in artery, the transcutaneous saturation of oxygen pressure, the heart rate and the respiratory rate in the PCV group were obviously improved after ventilation treatment. The peak inflating pressure, the mean pressure of the airway and the central venous pressure in the PCV group were lower than in the VCV group. The incidence of pressure injury was 0 in the PCV group while 10% in the VCV group. CONCLUSIONS: The clinical effect of PCV on traumatic ARDS is better and the incidence rate of pressure injury is lower than that of VCV. PCV has minimal effects on the hemodynamics.

[Effect of thymosin alpha 1 on cellular immune function in elderly patients with malignant tumor].

OBJECTIVE: To investigate the effect of thymosin alpha 1 on cellular immune function in the elderly patients with malignant tumor. METHODS: Thirty patients with malignant tumor were injected with thymosin alpha 1 subcutaneously at the dose of 1.6 mg q.d. for the first month and q.o.d. for the following month. The number of T cell subgroups and the activity of NK cell in peripheral blood were detected and the quality of life of the patients were evaluated before treatment and at the end of treatment. RESULT: Treatment of thymosin alpha 1 increased the number of CD4 cells and improved the NK activity, and also improved the quality of life of the elderly patients with malignant tumor. There were no side effects found. CONCLUSION: Thymosin alpha 1 can enhance the cellular immune function of the elderly patients with malignant tumor.

[Study on anxiety and depressive disorder of inpatients in general hospital].

OBJECTIVE: To assess the co-morbidity of anxiety and depression in hospitalized patients and to analyze the degree of symptoms and the influential factors. METHODS: Thirty patients with malignant tumor were injected with thymosin alpha 1 subcutaneously at the dose of 1.6 mg q.d. for the first month and q.o.d. for the following month. The number of T cell subgroups and the activity of NK cell in peripheral blood were detected and the quality of life of the patients were evaluated before treatment and at the end of treatment. RESULT: (1) About 39.9 % of the patients presented the symptoms of anxiety and depression and out of them 6.7 % were taken psychotropic medication. (2) Stepwise Logistic Regression showed that the factors related to anxiety and depression could be classified into four categories: economic factor, sleeping status, cognition of disease, the other factors such as course of disease and the environment of hospital. (3) Data analysis of oncology patients showed that the cognition of disease would be an important factor, which would affect anxiety and depression status. CONCLUSION: Multiple factors can affect anxiety and depression symptoms of inpatients. The results suggest that psychological intervention such as cognitive therapy should be considered in clinical practice.