Analysis of p16 gene mutations and their expression using exhaled breath condensate in non-small-cell lung cancer.

The aim of the present study was to investigate the mutational status of exons 1 and 2 of the p16 gene in the exhaled breath condensate (EBC) of patients with non-small-cell lung cancer (NSCLC) and determine the feasibility and clinical significance of applying EBC in the diagnosis of NSCLC. Polymerase chain reaction and DNA sequencing were applied to detect exon 1 and 2 alterations of the p16 gene in EBC by comparing 58 samples from NSCLC patients and 30 from healthy controls. Of the 58 EBC samples from NSCLC patients, 54 were successfully tested and 8 cases of mutations were identified, of which 3 were in exon 1 and 5 in exon 2. The mutation rate was 14.81% (8/54). There were no p16 gene mutations in the 30 samples obtained from healthy controls. EBC p16 gene mutations exhibited no statistically significant differences according to gender, smoking history, pathological type, degree of differentiation and presence or absence of lymph node metastasis. The p16 gene mutation rate was proportional to the tumor stage (P<0.05). Therefore, the detection of the p16 gene mutation in EBC may be used as a novel molecular marker to assist in the diagnosis of NSCLC.

Methylation of P16 in exhaled breath condensate for diagnosis of non-small cell lung cancer.

BACKGROUND: Non-small cell lung cancer is the most frequently cause of cancer-related death in the world. To explore the technical feasibility, we detected aberrant promoter methylation of P16 in exhaled breath condensate which was a new, non-invasive tool for diagnosis and screening program of NSCLC. METHODS: We analyzed aberrant promoter methylation of P16 in 180 samples from 60 individuals, including 30 NSCLC patients (cancer tissues, adjacent normal lung tissues, blood plasma, and EBC), and 30 healthy controls (blood plasma and EBC) by fluorescent quantitative methylation-specific polymerase chain reaction (F-MSP). RESULTS: The positive rate of aberrant promoter methylation of P16 was 26 of 30 (86.66%) in tumor tissues, 15 of 30 (50%) in blood plasma, and 12 of 30 (40%) in EBC, we have not observed the positive methylation of P16 in the adjacent normal lung tissues, or in EBC or blood plasma from the healthy control group. CONCLUSION: We found that detected promoter methylation of P16 in EBC was feasibility, it should be an useful biomarker for diagnosis of NSCLC, it have potential prospect that detected the gene molecular in EBC because of noninvasive, specificity, convenient and repeatable.

[Determination and clinical implication of leukotriene B(4) and tumor necrosis factor-alpha in condensate of exhaled breath of chronic obstructive pulmonary disease patients].

OBJECTIVE: To study the changes and clinical implication of leukotriene B(4) (LTB(4)) and tumor necrosis factor-alpha (TNF-alpha) in exhaled breath condensate (EBC) of chronic obstructive pulmonary disease (COPD) patients. METHODS: EBC of 20 patients in acute episode of COPD (AECOPD), 20 patients in period of remission of COPD, and 20 persons who were having regular check-up (healthy control group) were enrolled. The concentrations of LTB(4) and TNF-alpha in EBC were assayed. Forced expiratory volume in one second (FEV1) and peak expiratory flow (PEF) of COPD patients were observed at the same time, pH, oxygenation index (PaO(2)/FiO(2)), partial pressure of oxygen in arterial blood (PaO(2)) and leukocyte count were also determined. RESULTS: (1)The concentrations of LTB(4) in EBC of AECOPD (35.43+/-14.19)ng/L and remission of COPD(24.39+/-13.75)ng/L, were significantly higher than that of the healthy control group (16.75+/-7.44)ng/L, and the concentration of LTB(4) in EBC during remission of COPD was significantly lower than that of AECOPD (all P<0.05). (2)The concentration of TNF-alpha in EBC of AECOPD (9.35+/-8.66) ng/L was significantly higher than that of remission of COPD (4.42+/-4.11)ng/L and healthy control group (4.45+/-3.92) ng/L, and the differences had statistical significance (both P<0.05). The concentration of TNF-alpha in EBC showed no significant difference between patients in remission of COPD and healthy control group. (3)The concentration of LTB(4) in EBC had negative correlation with FEV1 of AECOPD patients. Regression equation was y=or-0.51 x+0.22, r=-0.481 (P<0.05). CONCLUSION: The concentrations of LTB4 and TNF-alpha in EBC of AECOPD patients are raised when oxidation stress is reinforced, and its level reflects the severity and prognosis of COPD.

[Study of carvedilol on plaque stability in carotid atherosclerotic rabbits transfected by p53 gene].

OBJECTIVE: To investigate the effects of carvedilol on stabilizing atherosclerosis plaque. METHODS: Forty five male Japanese white rabbits were divided randomly into 5 groups with 9 for each. One group was fed up with normal diet as blank control. In other four groups, the common carotid artery of rabbits fed up with high cholesterol diet were injured by balloon. Three groups of them were transfected by wild-type p53 gene 8 weeks later, and then two groups of them were treated with carvedilol (3 mgxkg(-1)xd(-1)) and metoprolol (6 mgxkg(-1)xd(-1)) respectively, high cholesterol diet should be continued for other 4 weeks. Serum lipid, hypersensitive C-reaction protein (hsCRP), oxidized low density lipoprotein (oxLDL), superoxide dismutase (SOD), malondialdehyde (MDA) and glutathione peroxidase (GSH-PX) were measured in 0, 8, 12 weeks after experiment. The apoptosis rate of smooth muscle cell (SMC) in endomembrane and the local expression of p53, bcl-2, bax, alpha-actin were examined after experiment, and the carotid arteries were examined by light microscopy and transmission electron microscopy. RESULTS: The typical carotid atherosclerotic plaques were observed in balloon-injured groups. The local expression rates of p53 in groups transfected by wild type p53 gene were higher obviously than them in other two groups (P < 0.01). Compared with the rabbits received simple transfection, the thickness of the fibrous cap in rabbits received carvedilol and metoprolol were all increased, but the change could be observed significantly in carvedilol group (P < 0.05). Compared with metoprolol, carvedilol could reduce the level of serum hsCRP, oxLDL, MDA, and increase the concentration of SOD and GSH-PX significantly (P < 0.05 or 0.01), but two medicines had no obvious influence to serum lipid. The apoptosis rate of SMC in endomembrane, the local expression of bax gene and bax/bcl-2 ratio were decreased, the positive expression rates of alpha-actin and bcl-2 were enhanced in carvedilol group (P < 0.01). CONCLUSIONS: Both carvedilol and metoprolol can improve the stability of the plaque, but carvedilol is superior. Its mechanisms may lie in that carvedilol still has function of anti-inflammation, anti-oxidation, decreasing the apoptosis rate of SMC in addition to its function of blocking beta-receptor.