| Objective: Acute B lymphocytic leukemia(B-ALL)is a childhood malignant tumor characterized by abnormal proliferation,aggregation and tissue infiltration of B lymphocytes,which is relatively rare in adults.At present,most patients can be treated with multiple doses of combination therapy,but the prognosis is poor for patients with drug resistance and treatment effect is not significant.Therefore,further research is needed on the pathogenesis and therapeutic targets of B-ALL.Circular RNA(circRNA)is a class of endogenous non-coding RNAs with a single-stranded closed loop structure.CircRNA molecules are mainly produced by reverse splicing of exons and/or introns derived from precursor RNA,and their formation is regulated positively and negatively by a variety of proteins.CircRNA is widely distributed in various tissue cells in the body,and has certain characteristics of organization,timing and disease specificity,high conservation and structural stability.circRNA can be used as a miRNA sponge to cis-regulate the expression of parental genes and to act as a biomarker of disease.circRNA plays an important role in gene expression in a variety of physiological and pathological processes,particularly during tumor formation and progression.The study of circRNA in leukemia is still in its infancy,and differential expression and molecular function of only a few circRNAs have been initially revealed in acute myeloid leukemia.However,it is unclear whether circRNA is differentially expressed in acute B lymphocytic leukemia and its expression mechanism.This study aims to provide new diagnostic indicators and therapeutic targets for the diagnosis and treatment of acute B lymphocytic leukemia by exploring the expression mechanism of circRNA in BALL-1cells.Methods:1.Total RNA of human B lymphoblastic cell Hmy2-cir cell line and human acute B lymphocyte leukemia BALL-1 cell line were extracted to construct a sequencing library.They were subjected to circRNA sequencing using an Illumina HiSeq sequencer to obtain circRNA expression profiles of the two cell lines.2.Bioinformatics analysis of circRNA expression profiles of the two cell lines.Using Hmy2-cir cells as a normal control,it was found that 12561 circRNAs weredown-regulated in BALL-1 leukemia cells compared with normal cells.We selected 7circRNAs with the most significant difference in expression down-regulation,and 7kinds of circRNAs were detected by RT-qPCR.The relative expression levels of circRNAs in the two cell lines were verified to be consistent with the circRNA sequencing results.3.Based on the RT-qPCR results and the ease of construction of the overexpression vector(determined by the length of the circRNA sequence),chr12: 130884224-130907106 was selected for further study and named circRIMBP2 according to its source gene.The circRIMBP2 overexpression vector was transfected into BALL-1 cells using adenovirus,and its overexpression effect was verified by RT-qPCR.4.Using blank(Control group)and no-load(NC group)as control,the biological behavior of acute B lymphocytic leukemia cells after overexpression of circRIMBP2 was studied by CCK-8 cell proliferation assay,cell cycle assay and apoptosis assay influences.5.The control group and NC group were used as controls.The expression of ERK1/2and p-ERK1/2 in BALL-1 cells was detected by Western-blot after over-expression of circRIMBP2.Results:1.CircRNA expression profiles of human B lymphoblastic Hmy2-cir cell line and acute B lymphocyte leukemia BALL-1 cell line were detected.There were 14720 circRNAs expressed in Hmy2-cir cells and 7516 circRNAs expressed in Ball-1 cells.2.The results of the selected 7 circRNAs in the RT-qPCR experiment were consistent with the trend of circRNA sequencing results.CircRIMBP2 is underexpressed in Ball-1cells compared to Hmy2-cir cells.3.Using Control group and NC group,after over-expressing circRIMBP2 in Ball-1cells,the apoptosis rate of Ball-1 cells increased,the cell proliferation ability decreased,and the number of cells in G1 phase increased.4.Compared with Control group and NC group,the expression of ERK1/2 was not significantly changed and the expression of p-ERK1/2 was decreased after overexpression of circRIMBP2 in Ball-1 cells.Conclusion:1.Human B lymphoblastic cell Hmy2-cir was used as a normal control,and circRIMBP2 was lowly expressed in acute B lymphocyte leukemia cell line BALL-1.2.Overexpression of circRIMBP2 significantly inhibited the proliferation of BALL-1cells and promoted apoptosis of BALL-1 cells.3.Overexpression of circRIMBP2 significantly inhibited the expression of p-ERK1/2in the MAPK signaling pathway,thereby affecting the proliferation and apoptosis of BALL-1 cells. |
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