Regulation and clinical significance of circ_0003926 for esophageal squamous cell carcinoma
Objective To investigate the role and mechanism of circ_0003926 in esophageal squamous cell carcinoma(ESCC).Methods Plasma samples from 60 ESCC patients and 60 age-and gender-matiched healthy controls were collected at Af-filiated Hospital of North China University of Science and Technology from May 1,2022,to January 30,2023.The ex-pression levels of circ_0003926 and miR-139-3p were detected by fluorescence in situ hybridization in a ESCC tissue mi-croarray and their clinical relevance was analyzed.The loop structure and stability of circ_0003926 were verified by Sanger sequencing,back-to-back primer validation and ribonuclease R(Rnase R)tolerance assay.Real-time quantitative poly-merase chain reaction(qRT-PCR)was used to detected the expression levels of circ-0003926 and miR-139-3p in ESCC cell lines and plasma.Counting Kit-8(CCK-8),colony formation,trans well invasion and migration assays,wound healing as-says,and flow cytometry assay were used to measure the effects of circ_0003926 and miR-139-3p on cell proliferation,migration,invasion and apoptosis,respectively.The interaction between circ_0003926 and miR-139-3p was further identi-fied by dual luciferase reporter gene assay and qRT-PCR.Nude mouse subcutaneous xenograft tumor model was used to investigate the impact of circ_0003926 on tumor growth.Results The circular structure of circ_0003926 was identified by Sanger sequencing,back-to-back primer and Rnase R assay.The results of fluorescence in situ hybridization showed that the mean fluorescence intensity of circ_0003926 expressed in ESCC tissues and adjacent tissues were 18.00±5.10 and 11.80±4.22(t=8.792,P＜0.001),respectively,and the mean fluorescence intensity of miR-139-3p were 18.26±2.81 and 26.23±2.97(t=17.713,P＜0.001),respectively.The expression level of circ_0003926 was positively correlated with the clinical stage(x2=5.312,P=0.021),but negatively related to overall survival time,the differences were statis-tically significant,P=0.002;while the expression level of miR-139-3p was positively correlated with lymph node metas-tasis(x2=4.114,P=0.043)and pathological grade(x2=4.267,P=0.039).qRT-PCR results demonstrated that the ex-pression of circ_0003926 in the ESCC and normal plasma samples was 3.54±2.15 and 1.03±0.05(t=6.993,P＜0.001),respectively,and the expression level of miR-139-3p was 0.36±0.17,and 1.03±0.03(t=27.001,P＜0.001),respectively.Compared with HEEC,circ_0003926 was highly expressed in ESCC cell lines(F=281.577,P＜0.001),and the expression of miR-139-3p was low(F=87.034,P＜0.001).After knock-down of circ_0003926,the proliferation,migration and invasion abilities of KYSE-150 and KYSE-410 cells were significantly decreased,and the level of apoptosis was significantly increased(all P＜0.05).The miR-139-3p silencing rescued the proliferation,apoptosis,migration and invasion abilities in circ_0006168-silenced ESCC cells(P＜0.05).The results of subcutaneous transplanted assay in nude mice demonstrated that the tumor volume and tumor weight of si-circ_0003926 group were lower than those in si-NC group,and the differences were statistically significant from day 10 to day 28(all P＜0.001);and the tumor volume and body weight in the si-circ_0003926+antagomiR-NC group were lower than those in the si-circ_0003926+antagomiR-139-3p group,and the differences were statistically significant from day 10 to day 28(P＜0.001).Conclusion circ_0003926 was high-ly expressed in ESCC tissue,plasma,and cells,and and regulates the progression of ESCC through miR-139-3p,which could become a new biomarker of diagnosis,treatment,and prognosis for ESCC.
esophageal squamous cell carcinomacirc_0003926miR-139-3pproliferation and apoptosismigration and invasion
NETL
NSTL
万方数据
