| Oral squamous cell carcinoma(OSCC)invasion and metastasis is the main factor affecting the prognosis of OSCC and the mechanism has not yet fully elucidated.In recent years,microRNA(miR)has been involved in the invasion and metastasis of cancer cells.Among them,miR-21-3p,miR-18a-3p,miR-210-3p,miR-155-5p,miR-181a-3p,miR-19a-3p are associated with oral cancer named oncology microRNA(Onco-miR),but few studies have been done on the editing and maturation of these Onco-miRs.Double-stranded RNA editing enzyme 1(Adenosine deaminases acting on RNA 1,ADAR1)can form allogeneic dimer with DICER to promote the maturation of miRNA.Onco-miR regulates the biological characteristics of cancer cells through post-transcriptional regulation.In this study,we discuss the expression of ADAR1 in oral cancer,and further explore whether ADAR1 can affect the migration and invasion of oral cancer cells by editing the maturation of oral cancer-related Onco-miR.Part1 Expression and significance of ADAR1 in oral cancerObjective:To investigate the expression of ADAR1 in oral cancer tissue and adjacent non-tumor tissue and its relationship with clinicopathological features.Methods:The expression of ADAR1 in oral cancer tissues and squamous cell carcinoma cells was detected by qRT-PCR and Western Blot and analyze the relationship between ADAR1 and TNM classification,clinical stage and pathological grade.Results:(1)ADAR1 expression in oral cancer tissue was significantly higher than that in adjacent tissues;(2)ADAR1 was significantly higher in patients with lymph node metastasis than in patients without lymph node metastasis;(3)it is same to patients with clinc stage ?-?;(4)The expression of ADAR1 in HN4,HN6,SCC9 and CAL27 cells was significantly higher than that in normal oral mucosa cells.Conclution:ADARl is highly expressed in oral cancer and is associated with lymph node metastasis.Part II the effect of ADAR1 on the invasion and migration of oral cancerObjective:To investigate the effect of ADAR1 on cell invasion and migration and its correlation with epithelial transformation.Methods:CAL27 and HN4 knocking down or over expression of ADAR1 were used to observe the ability of migration and invasion of ADAR1 by scratch assay and transwell invasion assay.The changes of EMT-related proteins such as Vim,and E-cad were detected by Western Blot afte knock-down or overexpression of ADAR1.Results:(1)After konck-down of ADAR1,for CAL27 and HN4 cells,the cell scratches assays showed that the healing percentage is more smaller for the experimental group being compared to the control group with statistical differences;(2)After overexpression of ADAR1,for CAL27 and HN4 cells,the cell scratches assays showed that the healing percentage is more bigger for the experimental group being compared to the control group with statistical differences;(3)After konck-down of ADAR1,for CAL27 and HN4 cells,the cell invasion assays showed that the nummer of the cells in the bottom of the chamber is more little for the experimental group being compared to the control group with statistical differences;(4)After overexpression of ADAR1,for CAL27 and HN4 cells,the the cell invasion assays showed that the the nummer of the cells in the bottom of the chamber is more for the experimental group being compared to the control group with statistical differences;(5)After konck-down of ADAR1,CAL27 and HN4 cells showed a decrease in Vim expression and an increase in E-cad expression;(6)After overexpression of ADAR1,CAL27 and HN4 cells showed a in crease in Vim expression and an decrease in E-cad expression.Conclution:(1)ADAR1 promoted the ability of cell migration and invasion.(2)ADAR1 promote epithelial-mesenchymal transition in oral cancer.Part? ADAR1 regulates the maturation of microRNAsObjective:To investigate the expression of Oncology-associated miRNA(Onco-miRNA)by qRT-PCR after knock-down and overexpresson of ADAR1 and the pre-miRNA and pri-miRNA after overexpresson of ADAR1.Methods:The specificity of microRNA and pre-microRNA primers was verified by Agarose gel electrophoresis assays.The Onco-miRNA(miR-21-3p,miR-18a-3p,miR-210-3p,miR-155-5p,miR-181a-3p,miR-19a-3p)expression was detected by qRT-PCR after knock-down and overexpression of ADAR1 in the cells of CAL27 and HN4,and the pre-miRNA and pri-miRNA was detected after overexpression of ADAR1 in the cells of CAL27 and HN4;Western blot and immunofluorescence staining were used to observe the distribution of ADAR1 cells;Immunoprecipitation was used to detect the interaction between AD AR1 and DICER.Results:(1)Agarose gel electrophoresis assays showed that the mature microRNA and precursor microRNA primers were specific.(2)The expression of oncology-associated microRNA(miR-21-3p,miR-18a-3p,miR-210-3p,miR-155-5p,miR-181a-3p,miR-19a-3p)was decreased in the experimental group being compared with control group after knockdown of ADAR1 in the cells CAL27 and HN4;(3)The expression of oncology-associated microRNA was increased in the experimental group being compared with control group after overexpression of ADAR1 in the cells CAL27 and HN4;(4)The expression of pre-microRNA relevant with oncology-associated microRNA was decreased in the experimental group being compared with control group after overexpression of ADAR1 in the cells CAL27 and HN4;(5)The expression of pri-microRNA relevant with oncology-associated microRNA was decreased in the experimental group being compared with control group after overexpression of ADAR1 in the cells CAL27 and HN4;(6)ADAR1 is mainly distributed in the cytoplasm;(7)ADAR1 interacts with DICER.Conclution:ADAR1 Forms a Complex with Dicer to Promote MicroRNA Processing. |
PDF Full Text Request