| Objective Lung cancer has become one common malignant tumor in the world.The incurrence and death rate of lung cancer is also high,and it has the trend of increase with global environmental pollution and change of life style in recent years.In China,lung cancer is also a kind of malignant tumor that is a threat to the physical and mental health of Chines people.In Chinese urban and rural areas,lung cancer has become the leading tumor incurrence rate and mortality rate.Most of the lung cancer patients have insidious onset.The local invasion and distant metastasis have occurred at the time of initial diagnosis,and the chance of cure has been lost when lung cancer is diagnosed.Traditional lung cancer treatment mainly includes surgery radiotherapy and chemotherapy.Among them,surgical treatment is main radical operation for lung cancer patients,however,for lung cancer patients who spread and metastasize in the medium and late stage,the best treatment opportunity has lost.Although the treatment of lung cancer patients get greatly improvement with the further study of tumor pathogenesis,the survival rate has not been significantly improved,which makes the treatment of lung cancer still face great challenges.Therefore,it is of great significance for the early diagnosis and treatment of lung cancer and the prognosis of patients with medium and late stage lung cancer to deeply understand the molecular mechanism of lung cancer cell growth,migration and invasion.Recent study showed that non-coding RNAs play an important role in human body and other organisms.Long non-coding RNA is a kind of non-coding RNA with a length of more than 200 nucleotides.Recently,long non-coding RNA is found to conduct biological function in many ways,including regulating gene expression,m RNA alternative splicing,changing intracellular protein localization and mediating chromatin remodeling.Function of long non-coding RNA being the research hotspot of tumor field attracts more and more attention of researchers.A large number of long non-coding RNAs are investigated in many kinds of tumors.These long non-coding RNAs play an important role in tumorigenesis and development.miRNA was one kind of short chain non-coding RNA to be found in recent years,whose length is about 22 nucleotides.miRNA is also highly conservative in evolution and plays a crucial role in the process of biological development and evolution.miRNA can regulate biological growth and development,control gene expression,change cell cycle and regulate human metabolism.The occurrence and development of many tumors was closely related to misregulation of miRNA.miRNA regulates tumor cell proliferation,metastasis,apoptosis,angiogenesis of tumor tissue and energy metabolism.In recent years,more and more attention has been paid to the interaction between long non coding RNA and miRNA in tumorigenesis and development.In this study,the relationship between long non coding RNA UCA1 and miRNA138/193 was discussed.Methods1.20 patients diagnosed with lung cancer were collected in this study in the First Affiliated Hospital of Zhengzhou University.All patients provided written informed consent and this study was approved by the Ethics Committee at our local institution.2.Lung cancer cells A549 and H1650 were separately cultured in DMEM(Dulbecco’s modified Eagle’s medium)or 1640 medium(Gibco)containing 10%(v/v)fetal bovine serum(Gibco)with 100μg/ml penicillin and 100μg/ml streptomycin at 37°C in a humidity incubator with 5% CO2.The cells were subcultured every 2 to 3 days,and the cells in logarithmic growth phase were selected for the experiment.3.Real time PCR was used to detect the expression of UCA1,CDK6,p21,p27,HMGB1,Slug,MMP9,miR-193 and miR-138 in lung cancer tissues and cell lines.4.According to instruction of Lipofectamine 2000,the transfection of lung cells was performed.5.Cell Counting Kit-8 was used to analyze the proliferation ability of lung cancer cells.6.Construction of p GL3 dual luciferase reporter gene and mutant gene was performed by gene cloning technology.7.Targeting of micro RNA to UCA1 was detected by dual luciferase reporter gene detection kit.8.The protein expression level of CDK6,p21,p27、HMGB1,Slug and MMP9 was detected by Western blot。9.Wound healing assay was used to analyze lung cancer cell migration ability.10.Transwell assay was used to analyze lung cancer cell invasion ability.11.All data were analyzed by SPSS18.0 statistical software(SPSS Inc,Chicago,IL,USA).Results1.Real time PCR showed that the m RNA expression level of UCA1 was higher in lung cancer tissues compared with normal ones.2.The downregulation of UCA1 resulted in the decrease of cell proliferation ability in A549 and H1650 lung cancer cells.3.The downregulation of UCA1 resulted in the decrease of cell migration ability in A549 and H1650 lung cancer cells.4.Online prediction software showed that miR-138 and miR-193 were potential miRNAs,which bond to the 1395-1433 bp area in UCA1 RNA.5.Overexpression of miR-138 or miR-193 decreased the expression of UCA1.6.Inhibition of miR-138 or miR-193 enhanced the expression of UCA1.7.Dual luciferase reporter gene assay showed that miR-138 and miR-193 directly targeted long non-coding RNA UCA1.8.UCA1 reversed the suppression effect of miR-138/193 on cell growth.9.UCA1 reversed the suppression effect of miR-138/193 on cell migration.10.UCA1 reversed the suppression effect of miR-138/193 on cell invasion.11.miR-138 and miR-193 regulated the UCA1-downstream genes.12.UCA1 regulated the expression of CDK6 as a miR-138 and miR-193 common target.13.The high m RNA and protein level of CDK6 in lung cancer tissues were found.14.The RNA level of miR-138 and miR-193 in lung cancer tissues was lower than the normal ones.15.There was a negative relationship between miR-138/193 and UCA1 in lung cancer tissues.Conclusions There was a negative relationship between the low expression of miR-138/193 and the high expression of UCA1 in lung cancer tissues.UCA1 reversed the suppression effect of miR-138/193 on cell growth,migration and invasion. |
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