ABSTRACT
Objective To explore the effect of expression of FHL1 on biological character of lung adenocarcinoma cell line A549.Methods Lung adenocarcinoma cell line A549 was transfected by FHL1 mRNA and shRNA lentivirus vector.The stable overexpression of FHL1 in lung adenocarcinoma cell line A549 model and low expression of cell line model were tested by Western blot and RT-PCR method.CCK-8 test,Transwell invasion,flow cytometry and Colony-forming Assay were used to test the proliferation,invasion,apoptosis and anchorage independence growth of the cell line.Results The difference between the stable overexpression of FHL1 group and its low expression group was obvious tested by Western blot and RT-PCR.CCK-8 test showed higher cell proliferation of the low expression group than that of the overexpression group and the blank group (P =0.002 2).Flow cytometry showed that the apoptosis rate of the overexpression group was 50.48 ％,significantly higher than that of the two low expression groups (16.41％,20.12％) and blank group (25.90％,P＜0.001).The invasiveness of the overexpression group was obviously lower than the low expression group and blank group in the Transwell invasion (P＜0.001).Colony-forming Assay showed that the colony number of the overexpression group was 28.7 ± 6.0,significantly less than the two low expression groups (157.0 ± 6.4,150.7 ± 9.5,P＜0.000 1).Conclusions The stable overexpression and low expression of FHL1 in lung cancer cell line A549 models were successfully constructed.The overexpression of FHL1 inhibited the proliferation,invasion and metastasis,and accelerated apoptosis in lung adenocarcinoma cell line A549.
ABSTRACT
<p><b>OBJECTIVE</b>To study the expression of heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) in non-small cell lung cancer (NSCLC), and the interaction between hnRNP A2/B1 protein and mRNA of DNA repair enzymes O(6)-methylguanine DNA-methyltransferase (MGMT), 8-oxoguanine DNA glycosylase (OGG1), redox factor 1(Ref-1), DNA-dependent protein kinase (including DNA-PKcs and ku).</p><p><b>METHODS</b>The expression and distribution of hnRNP A2/B1 were detected by immunohistochemistry and Western blot on 50 NSCLC samples from patients who underwent resection in Zhongshan Hospital. The hnRNP A2/B1 mRNA expression was tested by real-time PCR. Co-immunoprecipitation (co-IP) combined RT-PCR was used to investigate whether hnRNP A2/B1 could be bound with the mRNA of the above mentioned 5 DNA repair enzymes in human lung cancer cell line (HTB-182). Then immunohistochemistry and real-time PCR were used to detect the expression of MGMT in the same group of patients.</p><p><b>RESULTS</b>HnRNP A2/B1 protein and mRNA expressions were increased in the NSCLC tissues than that in the corresponding normal lung tissues. HnRNP A2/B1 was expressed predominantly in the nuclei of tumor cells. The positive rate and immunohistochemistry score of hnRNP A2/B1 in tumor tissue were significantly higher than that in normal tissue (P < 0.01). In stage III-IV NSCLC, hnRNP A2/B1 expression was higher than that in stage I-II. There was no significant differences of hnRNP A2/B1 expression among patients of different age, sex, histological type, and smoking history. The results of co-IP combined RT-PCR suggested that hnRNP A2/B1 is bound with MGMT mRNA, and MGMT expression is decreased in tumor tissue of NSCLC.</p><p><b>CONCLUSIONS</b>The results of this study show that hnRNP A2/B1 protein and mRNA are highly expressed in NSCLC, and hnRNP A2/B1 is bound with MGMT mRNA, which indicate that it might be one of the mechanisms of hnRNP A2/B1 participating in the pathogenesis of NSCLC.</p>