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To Investigate the Expression of HnRNP A2/B1 in NSCLC and the Role of HnRNP A2/B1 in the Pathogenesis of NSCLC

Author: ShiYuan
Tutor: TanYunShan
School: Fudan University
Course: Pathology and Pathophysiology
Keywords: Heterogeneous nuclear ribonucleoprotein A2/B1 Lung cancer DNA repair enzymes O ~ 6 - methylguanine - DNA methyltransferase (MGMT ) Sputum Diagnosis Cell block Pleural effusion Immunohistochemistry
CLC: R734.2
Type: Master's thesis
Year: 2010
Downloads: 22
Quote: 0
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Abstract

Lung cancer incidence and mortality have been ranked in various malignancies, but its pathogenesis has not been elucidated, and the lack of early diagnosis. Nuclear ribonucleoprotein A2/B1 (heterogeneous nuclear ribonucleoprotein A2/B1, hnRNP A2/B1) is an important RNA-binding protein, is mainly responsible for gene transcription regulation. Known DNA repair enzyme O6-methylguanine-DNA methyl transferase (O6-methylguanine DNA-methyltransferase, MGMT) .8 - hydroxyguanine DNA glycosylase (8-oxoguanine DNA glycosylase OGGl). Oxidation / reduction factor -1 (redox factor ref-1) .2 DNA-dependent protein kinase (DNA-dependent protein kinase, DNA-PK)-DNA-PKcs ku expression in lung cancer tissue and normal lung tissue are associated with the occurrence of lung cancer. Based on the above background, the purpose of this study was to observe the hnRNP A2/B1 protein and mRNA in non-small cell lung cancer (non-smal l ce lll ung cancer, NSCLC) cancer tissues and corresponding normal lung tissue expression; detect hnRNP A2 / B1 protein and DNA repair enzymes of MGMT, OGG1, ref-1, DNA-PKcs and Ku-mRNA combined to explore its role of DNA repair enzymes existence of post-transcriptional regulation, trying to reveal its role in NSCLC pathogenesis ; and explore sputum detection the hnRNP A2/B1 significance in the early diagnosis of lung cancer and pleural effusion detection of hnRNP A2/B1 value for the identification of lung adenocarcinoma cells to mesothelial cells. Protein levels and mRNA levels detected hnRNP A2/B1 expression in NSCLC: Immunohistochemistry was used to detect the 50 cases, the Western Blot method detected 10 cases and fluorescence real-time quantitative PCR (real-t ime PCR) method to detect 22 patients with NSCLC cancer tissue and corresponding normal lung tissue hnRNPA2/B1 expression; using co-immunoprecipitation (Co-immunoprecipitation, co-IP) combined with reverse transcription polymerase chain reaction (RT-PCR) method in human lung cancer cell lines (HTB-182) whether in hnRNPA2/B1 protein with five kinds of DNA repair enzyme mRNA the mesh combined, after using immunohistochemical method detected 50 cases of specimens (specimens with detection of hnRNP A2/B1) and real-time PCR method detected 22 cases of NSCLC (specimens the same detection hnRNP A2/B1 specimens) cancer tissue and the corresponding expression of MGMT in normal lung tissue. Sputum samples collected another 63 cases of suspected lung cancer patients, sputum smear and cell block sections cytology (microscopic examination to find cancer that the diagnosis of lung cancer) and sputum cell block sections Immunohistochemical detection of hnRNP A2/B1 in (expression positive that the diagnosis of lung cancer) diagnosis, the results of the comparison of two diagnostic methods. The cell block technology combined with immunohistochemistry method to detect the expression of hnRNP A2/B1 100 cases of pleural effusion specimens observed expression differences in lung adenocarcinoma cells and mesothelial cells. All data processed using the SPSS 13.0 statistical package, p lt; 0.05 was considered statistically significant. Results HnRNP A2/B1 protein and mRNA expression in the the NSCLC cancer tissue in higher than corresponding normal lung tissue: Immunohistochemistry showed that hnRNP A2/B1 is located in the nucleus, the positive rate in NSCLC cancer tissue and score (100%, 5.3 ± 0.9) were significantly higher than in normal lung tissue (32%, 2.2 ± 0.7) (p lt; 0.01), slightly higher than in the Ⅲ - Ⅳ stage NSCLC cancer tissue expression Ⅰ - Ⅱ stage NSCLC cancer tissue expression with age, gender, tumor type and smoking history was no significant correlation (p gt; 0.05); the hnRNP A2/B1 Western Blot results show that the average gray NSCLC cancer tissue ratio (1.4 ± 0.5) was significantly higher than normal lung tissue (0.7 ± 0.2) (p lt; 0.01); real-time PCR test results show the average expression of hnRNP A2/B1 mRNA in NSCLC cancer tissue 37.4 (15.6 to 82.6) was significantly higher than in normal lung tissue 17.6 ( 7.8 ~ 27.8) (p lt; 0.01). Co-immunoprecipitation experiments first human lung squamous carcinoma cell line (HTB-182) hnRNP A2/B1 protein precipitate hnRNP A2/B1 monoclonal antigen-antibody complex, after expansion detected by RT-PCR product from the co-immunoprecipitation increase the MGMT mRNA, suggesting that hnRNP A2/B1 protein and the MGMT mRNA combined, after immunohistochemistry and real-time PCR detection reveals MGMT protein and mRNA expression in NSCLC cancer tissues compared to corresponding normal lung tissue to reduce: immunohistochemistry chemical display MGMT located in the nucleus, the positive rate in NSCLC cancer tissue and score (32%, 2.2 ± 0.8), were significantly lower than in normal lung tissue (78%, 4.1 ± 1.2) (p lt; 0.01); of hnRNP A2/B1 score of MGMT expression in the NSCLC cancer tissue in the correlation analysis, draw both a negative correlation (F = -0.49, p lt; 0.01); real-time PCR test results showed that MGMT in NSCLC cancer tissue the average expression level of 1.8 (0.6 to 3.1) was significantly lower than that in normal lung tissue 9.8 (6.8 to 18.3) (p lt; 0.01). Sputum smears and cell block sections cytological diagnosis of lung sensitivity 31.8%, specificity of 100%. The sputum cell the block detection sensitivity of the hnRNP A2/B1 diagnosis of lung cancer for 80%, specificity was 68.4%. The hnRNP A2/B1 sensitivity of sputum was significantly higher than the sputum cytology (p lt; 0.01). The HnRNP A2/B1 expression in pleural effusion lung adenocarcinoma cells positive rate was 100% (60/60), in the mesothelial cells positive rate was 30% (12/40) between the two groups significant differences (p lt; 0.01). Pleural effusion lung adenocarcinoma cells HnRNP A2/B1 mark a sensitivity of 100% and a specificity of 70%. Conclusions In this study, by immunohistochemistry, Western Blot and real-time fluorescence quantitative PCR revealed expression of hnRNP A2/B1 protein and mRNA in NSCLC cancer tissue compared with corresponding normal lung tissue increased, and immunohistochemistry results show higher the the lesions more late hnRNP A2/B1 expression, suggesting that it may be related to the occurrence and development of NSCLC, hnRNP A2/B1 expression in the NSCLC cancer tissue in age, gender, histological type and smoking history had no significant correlation. (2) In this study, by co-immunoprecipitation combined with RT-PCR technology found hnRNP A2/B1 protein combination MGMT mRNA, suggesting that hnRNP A2/B1 may exist MGMT mRNA transcription regulation. After immunohistochemistry and fluorescence real-time quantitative PCR showed that the expression of MGMT protein and mRNA in NSCLC cancer tissue than in normal lung tissue to reduce, by MGMT mRNA transcription regulation might speculate that hnRNP A2/B1 down the expression of MGMT protein weaken the cellular DNA damage repair capacity, and thus participate in the occurrence of non-small cell lung cancer. Sputum cell block combined with immunohistochemistry assay sensitivity of of hnRNP A2/B1 diagnosis of lung cancer was significantly higher than that sputum cytology detected more conducive to detect early lung cancer. Pleural effusion cell blocks combined immunohistochemical method to detect hnRNP A2/B1 helpful for the identification of lung adenocarcinoma cells and mesothelial cells.

To Investigate the Expression of HnRNP A2/B1 in NSCLC and the Role of HnRNP A2/B1 in the Pathogenesis of NSCLC

Abstract

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