| Background: Lung cancer is one of the most common malignancies with higher morbidity and mortality worldwide.As the predominant type of lung cancer,non-small-cell lung cancer(NSCLC)accounts for approximately 85% of all lung cancer cases.Even though therapeutic strategies have made significant improvements over the past two decades,the 5-year survival rate of NSCLC patients is still just around 20%.That is related to the fact that some patients are in the middle and late stages when they are first diagnosed,the traditional treatment methods are not good,and the pathogenesis of lung cancer is complex and has not been fully elucidated.Therefore,elucidating the underlying mechanism is urgently required for exploration for NSCLC patients.Micro RNAs(mi RNAs or mi Rs)are a class of small noncoding RNAs.Studies have found that mi RNAs play key roles in many biological processes,including differentiation,cell proliferation,migration,invasion,and apoptosis.Accumulating literatures have indicated that mi RNAs have been widely proposed as tumor suppressors or oncogenes in different cancers.As one of mangy micro RNAs,MiR-924 has been reported to be a tumor suppressor in hepatocellular carcinoma.However,the functions and mechanisms of mi R-924 in NSCLC remain unclear.Rhomboid domain-containing protein 1(RHBDD1)is a new member of the Rhomboid family.Related studies showed that mi RNA interference mediated RHBDD1 silencing significantly suppressed cell proliferation and cell cycle progression in tumors such as hepatocellular carcinoma,glioblastoma and colorectal cancer.That is to say,RHBDD1 plays the role of an oncogene.Using bioinformatics software,we found that mi R-924 and RHBDD1 m RNA have complementary base pairing sequences in the 3’ untranslated region of RHBDD1 m RNA.It is speculated that the two may interact and affect tumor progression.The purpose of this study is to research the biological function and regulatory mechanism of mi R-924 in non-small cell lung cancer,to verify whether RHBDD1 was a direct target gene of mi R-924 and to explore the signaling pathways involved in targeting regulation in order to provide new targets for the treatment of non-small cell lung cancer.Methods:(1)The expression of mi R-924 was determined in NSCLC cell lines using quantitative real time PCR.(2)Cell proliferation was assessed by CCK-8 assay.(3)Cell migration and invasion were detected by transwell assay.(4)The expression level of protein was detected by western blot assay.(5)The combination of mi R-924 and RHBDD1 was analyzed via the luciferase reporter assay.(6)The expression level of RHBDD1 was evaluated in lung cancer tissues using public microarray datasets from Oncomine and its prognostic value was assessed by Kaplan–Meier Plotter databases.(7)A tumor xenograft mouse model was established to illustrate the effects of mi R-924 on the tumorigenesis of NSCLC in vivo.Results:(1)In this study,we found mi R-924 was strikingly decreased in NSCLC cell lines.(2)MiR-924 could directly complement the 3’UTR of RHBDD1 m RNA.(3)The expression level of RHBDD1 was significantly increased and inversely associated with prognosis using public microarray datasets from Oncomine and Kaplan–Meier Plotter databases.(4)MiR-924 overexpression suppressed cell proliferation,migration and invasion.(5)In vitro and in vivo experiments found that overexpression of mi R-924 could inhibit the expression of RHBDD1 and Wnt/β-catenin signaling pathway components Wnt1,β-catenin,p-GSK-3β,and up-regulate the expression of E-cadherin,a protein related to epithelial-mesenchymal transition,Vimentin expression was down-regulated.Conclusions:(1)MiR-924 function as a tumor suppressor in NSCLC cells.(2)RHBDD1 was predicted and confirmed as a direct target of mi R-924.(3)MiR-924 inhibited the proliferation,migration and invasion of non-small cell lung cancer cells by direct targeting of RHBDD1.(4)In vitro and in vivo studies further confirmed that mi R-924 exerts tumor suppressor effect through RHBDD1/Wnt/β-catenin signaling pathway. |
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