Study On The Role And Mechanism Of Long Noncoding RNA AC 026166.2-001 In The Occurrence Of Laryngeal Carcinoma

ObjectiveLong noncoding RNA(lncRNA)is one of non-coding RNA and its length is generally greater than200 nt.Because they do not have the function of protein translation,they are largely present in the nucleus or cytoplasm after transcription.A large number of existing lncRNAs are considered to be useless for a period of time,but a large number of domestic and foreign scholars have found that lncRNAs are closely related to the occurrence and development of tumors.Previous studies have shown that the expression of lncRNA AC026166.2-001 in laryngeal cancer tissues and metastatic lymph nodes is different from that in paracancerous tissues,which can be used as a molecular marker for diagnosis and prognosis of laryngeal cancer patients.This study aimed to further explore the biological role of lncRNA AC026166.2-001 in laryngeal cancer cells and its related mechanisms.Methods1.Firstly,the lentiviral overexpression vector and 3 siRNAs were designed and synthesized according to the lncRNA AC026166.2-001 nucleic acid sequence.2.The expression of lncRNA AC026166.2-001 in laryngeal cancer cells was up-regulated and down-regulated by using lentivirus and small interference transfection.3.Quantitative reverse transcription-polymerase chain reaction(qRT-PCR)was used to verify the lncRNAAC026166.2-001 up-regulation and down-regulation effects in laryngeal cancer cells.4.Real-time cell analyzer(RTCA),colony formation analysis,flow cytometry,wound healing analysis and Transwell migration test were used to detect the role of lncRNA AC026166.2-001 in the growth,proliferation,apoptosis and migration of laryngeal carcinoma cells.5.Using miRcode and miRDataBase website databases,through sequence alignment,predict miRNAs that may interact with lncRNA AC026166.2-001.6.Dual-luciferase reporter assay used to confirm whether lncRNA AC026166.2-001 can interact with the predicted miR-24-3p,and detect the expression of miR-24-3p after upregulated lncRNA AC026166.2-001 by qRT-PCR.7.Western Blot was used to detect changes in the expression of the cell cycle-related proteins.8.A nude mouse laryngeal cancer xenograft model was established and the biological function of lncRNAAC026166.2-001 was verified in vivo.Results1.Real time cell analyzer(RTCA)and colony formation analysis showed that the proliferation and colony forming ability of laryngeal carcinoma cells were significantly inhibited after overexpression of AC026166.2-001.The down regulation of AC026166.2-001 can promote the proliferation and colony forming ability of laryngeal cancer cells.2.The results of flow cytometry showed that the cell cycle was blocked at G0/G1 and cell apoptosis was induced by overexpression of AC026166.2-001.While downregulation of AC026166.2-001 results inthe opposite result.3.Wound healing analysis and Transwell migration assay results showed that AC026166.2-001 can reduce cell migration.4.Dual-luciferase reporter assay and Western blotting indicated that AC026166.2-001 can act as a miR-24-3p sponge and regulates the expression of p27 and cyclin D1.5.In vivo results also showed that AC026166.2-001 significantly inhibited the growth of LSCC xenografts and promoted cell apoptosis.ConclusionsAC026166.2-001 acting as a tumor suppressor in LSCC.AC026166.2-001 inhibited proliferation and promoted apoptosis of LSCC cells in vitro and in vivo.The molecular mechanism of AC026166.2-001 may be associated with miR-24-3p/p27 pathway.AC026166.2-001 may be served as a potential therapeutic target for the treatment of LSCC.