| Background:Lung cancer is the most common malignant tumor, seriously threatening human health, and the leading causes of death worldwide. At present, most cases of lung cancer are diagnosed at an advanced clinical stage. Although the improvements in diagnostics, surgery and chemotherapy, the 5-year survival rate of the patients is reported to be still less than 15%. As one of the main ways of epigenetics, DNA methylation plays an important role in the transcriptional silencing of tumor suppressor genes. More and more studies have indicated that the changes of DNA methylation in tumor suppressor gene promoter region are closely related to the occurrence of lung cancer. Therefore, identification of the change of DNA methylation in lung cancer will provide a new way for the early diagnosis and treatment of lung cancer.MPDZ gene is expressed in various tissues, and has important effects on the adaptive response of renal infiltration, normal development of retina, alcohol dependence and epileptic seizure, and the formation of electrical synapses in brain. Recent studies have found that MPDZ expression is down regulated in breast cancer, and may be associated with tumor metastasis. However, the expression pattern, specific function and molecular mechanism of MPDZ gene in tumor are still unknown. In our previous study, we have found that the DNA methylation of MPDZ gene in lung cancer and normal lung tissues were significantly different. Therefore, the DNA methylation, expression of MPDZ gene in lung cancer tissues, normal tissues and tumor cell lines, and their significance of prognosis in lung cancer patients were studied in the present study.Objective:1. To detect the methylation status of MPDZ gene in lung cancer cells, lung cancer tissues and para carcinoma tissues, and analyze the relationship between the methylation incidence and clinical data of the lung cancer patients.2. To analyze the relationship between the expression and DNA methylation of MPDZ gene in lung cancer cells and lung cancer tissues.3. The relationship between the expression of MPDZ protein and the survival time of the lung cancer patients, and the difference of the relationship between the two groups were analyzed by different clinical factors.Methods:1. Methylation status of MPDZ gene was detected by methylation specific PCR(MSP) and bisulfite sequencing PCR(BSP) in lung cancer cells, lung cancer tissues and adjacent tissues.2. The expression of MPDZ gene was detected by reverse transcription-polymerase chain reaction(RT-PCR) and immunohistochemistry in lung cancer cells, lung cancer tissues and adjacent tissues, and the lung cancer cells with and without 5-aza-2′-deoxycytidine(5-aza-dC) treatment.3. The correlation between MPDZ gene methylation and clinical pathological data of the lung cancer patients was analyzed by χ2 test, and the correlation between MPDZ gene expression and survival time was analyzed by Kaplan-Meier and Cox-regression survival analysis in SPSS 17.0.Results:1. MPDZ gene was methylated in 70%(7/10) of lung cancer cell lines, 61.2%(60/98) of lung cancer tissue, 55.6%(70/126) of lung cancer patient’s plasma samples and 48.1%(50/104) lung cancer patient’s sputum samples, while in 0%(0/1) of normal cell line and 0%(0/20) of normal tissues adjacent to cancer. The differences of methylation incidence between normal cell/normal lung tissues and lung cancer tissue have a statistical significance after statistical analysis.2. MPDZ gene was methylated in 43 cases of 62 cases with corresponding plasma and sputum specimens of lung cancer, and the methylation rate was 69.4%(43/62) in lung cancer tissues, 76.7%(33/43) in lung cancer tissues corresponding plasma samples, and 46.5%(20/43) in lung cancer tissue and corresponding sputum specimens. It has a significant correlation in lung cancer tissue, corresponding plasma and sputum specimens after statistical analysis.3. The methylation of MPDZ gene was significantly correlated with the clinical stages of lung cancer patients(P=0.047), but not correlated with the age(P=0.156), gender(P=0.950), smoking(P=0.455), tumor differentiation degree(P=0.445) and tumor pathological type(P=0.100). The incidence of MPDZ gene methylation was significantly higher in patients at III and IV(15/17, 88.2%) than the patients at I and II(28/45, 62.2%).4. MPDZ gene was not or down-regulated expressed in most of the methylated lung cancer cells and the lung cancer tissues, and the expression of MPDZ gene was obviously restored in lung cancer cells treated with 5-aza-d C. These results suggest that MPDZ is a gene that is silenced by DNA methylation in lung cancer.5. The expression of MPDZ protein is closely related to pathological grade(χ2=7.250, P=0.027) and clinical stage(χ2=5.773, P=0.016) of lung cancer patients, but not related to age, gender, tumor size, total lymph nodes, lymph node status and pathological type of lung cancer patients(P>0.05).6. The prognostic significance of MPDZ protein in patients at clinical stage I+II and stage III+IV was significantly different. The expression of MPDZ protein in patients with I+II was positively related to the survival time, and was an independent prognostic factor, however, the expression of MPDZ protein was not correlated with the survival time of patients at stage III+IV.Conclusion:MPDZ is a gene that is regulated by DNA methylation in lung cancer, and its expression is closely related to the clinical stage of lung cancer. The expression of MPDZ protein is closely related to the pathological grade and clinical stage of lung cancer. The MPDZ protein expression is positively related to the survival time of lung cancer patients, and its expression is an independent prognostic factor. From these results, we suggested that MPDZ gene methylation or expression can be used as a potential marker for diagnosis and prognosis of lung cancer. |
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