| Lung cancer is one of the most threatening malignant tumor to human health, which now is the leading cause of cancer death in the Urben China, and the mortality rate of lung cancer is still rising. However, if the patients with stage I lung cancer could be early diagnosed and provided with timely effective surgical treatment, their 5-year survival rate could be reach to 88%. Breath test might be used as a new mode, non-invasive,and convinient diagnostic method, which can make the assessment of disease through detecting the range of the VOCs exhaled or consumed by the patients, and reflecting the metabolism of the corresponding tissue cell. Considering the location, lung cancer which are occurred in the bronchial tree and alveolar epithelium is closely related to VOCs, therefore, to find out the lung cancer-associated specific VOCs might be provided us a new idea and target for early diagnosis of lung cancer.This reseach compared advantages of different VOC detecting systems of breath which lung cancer patients exhaled, and selected the best analytical method. Lung cancer patients were enrolled as study objects, while health person and lung benign disease patients were as the control group. Analyzing the expression of VOCs of exhaled breath, and selecting the special VOCs markers associated with lung cancer, which can be uesd as potential target of early detection to lung cancer. Meanwhile, the study further established a lung cancer diagnostic model by exhaled breath, searched its value of early diagnosis to lung cancer, and verified it at the level of tissue cell.The study confirmed that there are stable conclusion and good repeatability to analyze special VOCs from exhaled breath by SPME-GCMS, which is superior to SPME-GS, Tenxa-TA-GS. We analyzed exhaled breath VOCs of 243 samples by SPME-GCMS system, including 88 samples of lung cancer,70 samples of lung benign disease,85 samples of health person, and further researched the sensitivity and specificity of the diagnostic models based on different combinations of special VOCs by applying statistical approaches, such as cluster analysis or stepwise discriminant analysis. In the end, we selected age, as well as 4 to 6 VOCs, as the diagnostic models for lung cancer, including 2,6-dimethylnaphthalene, palmitic aldehyde, nonadecyl alcohol, etc, basically with high sensitivity and specificity more than 85%. Take one model for example, which is consisted of palmitic aldehyde, nonadecyl alcohol, 2,6-dimethylnaphthalene, age, and so on, its specificity is 94.9%, and 78.7% of early stage lung cancer patients were correctly identificated, however,31.9% of them were misclassificated to advanced stage. Therefore, this kind of diagnostic model could be applied to early diagnosis of lung cancer, but with poor effect to differentiate stages, which indicates that breath test diagnostic model for lung cancer might be one of the screening and early warning methods, although is not identical well to clinical staging of lung cancer. Furthermore, the study found that several special VOCs expression are associated with pathological classification, lesion locations, surgery and smoking situation.Although the result of exhaled breath test in lung cancer patients was satisfied and several kind of specific VOCs were found, we still known less about the VOCs exhaled by the lung cancer tissue and the cancer cell lines. Huge variations existed between results of different studies of exhaled breath test in the cancer cell lines, and the analyzing of breath test for specific VOCs in the lung cancer tissue has not been reported yet.The aim of this study was to confirm the existence of specific VOCs released or consumed by the lung cancer tissue and the cancer cell lines, while chose the normal lung tissue and the primary human bronchial epithelial cell as contrast.The lung cancer tissues were sampled from eighteen surgeried patients. These tissues were cultured with the four kinds of cell lines(A549, NCI-H446, SK-MES-1, BEAS-2B) respectively. We used the solid-phase micro-extraction and gas chromatography-mass spectroscopy technique (SPME-GCMS) to analyze the VOCs exhaled by the lung cancer cells which confined in the headspace of culture flasks and screen for the specific VOCs. As the result it has been found,2-pentadecanone and nonadecane were detected in all the three lung cancer cell lines A549, NCI-H446 and SK-MES-1 (the concentration of 2-pentadecanone were (1.382±0.171)×10-5 mg/L, (1.681±0.190)×10-4 mg/L,(2.835±0.401)×10-6 mg/L, the nonadecane's were (8.382±0.606)×10-6 mg/L, (1.845±0.130)×10-5 mg/L, (6.220±0.362)×10-6 mg/L), while eicosane were detected in A549 and NCI-H446 (the concentration were (8.313±1.130)×10-6 mg/L, (1.020±0.141)×10-5 mg/L), all the three VOCs were not detected in the BEAS-2B. Twelve VOCs such as decane,2-pentadecanone, nonadecane and eicosane had greater concentration in the eighteen lung cancer tissues (the concentration of 2-pentadecanone was 5.421×10-6～3.621×10-5 mg/L, the nonadecane's was 5.805×10-6～1.830×10-5 mg/L, the eicosane's was 2.730×10-6～2.343×10-5 mg/L), but there were no difference between patients with different cancer differentiation. Eicosane in the non-squamous carcinoma was more notable than that in the squamous carcinoma, which was confirmed in the cell lines'results. Only the 2-pentadecanone was stably detected in all of the lung cancer patients, the lung cancer tissues and the cell lines.The reasons for differences in VOC release or consumption among the investigated cell lines or tissues in vitro or patients'breath in viro are currently unknown, to solve this issue will require an understanding of the underlying molecular mechanisms for VOC production. This study firstly identified the specific VOCs produced by the lung cancer tissue and the lung cancer cell lines, which supported the breath test as a complementary noninvasive diagnostic method for lung cancer in the tissue level.This research constructed the exhaled breath test for specific VOCs as the diagnostic model for lung cancer with high sensitivity and specificity. Further study is required to understand the molecular mechanisms for VOC production, and its biochemical pathways and exhalation kinetics. |
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