【作者】 刘磊；

【导师】 黄强；

【作者基本信息】 山东大学 ， 外科学（普外）（专业学位）， 2019， 博士

【摘要】 胰腺癌(Pancreatic ductal adenocarcinoma,PDAC)是一种恶性程度较高、预后较差的消化系肿瘤,在我国由于生活水平的提高和不良饮食生活习惯等因素,导致胰腺癌近20年来发病率增长约6倍,死亡率位居恶性肿瘤第5位。手术切除仍然是胰腺癌首选治疗方法,所以,如何提高胰腺癌的术前诊断率,严格掌握手术指征,提高手术切除率是保证胰腺癌患者获得良好疗效的关键。胰腺癌近几年流行病学研究表明其5年生存率仍然较低,约为8.5%,其中最重要的原因为其生物学行为恶性程度高,侵袭性强,病程较早时就可能伴有淋巴结转移和神经脉管的浸润,故很多患者确诊时已是疾病的进展期,较快出现局部和远处转移。由于胰腺癌缺少早期发现的标志物,因此,如何寻找胰腺癌早期发生发展的标志物,是提高患者预后的关键因素。近年来,越来越多的学者发现既往没有被受到重视的非编码基因如微小 RNA(microRNA,miRNA)、环状 RNA(circular RNA,circRNA)、长链非编码RNA(Long non-coding RNA,lncRNA)等与肿瘤发生、发展密切相关,部分非编码基因甚至可以作为肿瘤早期发现的标志物用于临床诊断。miRNA是真核生物中存在的一类约22～24个核苷酸大小,参与转录后基因调节的非编码单链小分子RNA,可通过特异性识别靶基因的Messenger ribonucleic acid(mRNA)的3’-UTR非翻译区(3’-UTR)并与之结合,引起靶基因mRNA翻译的抑制,从而对靶基因进行转录后表达调节,最终通过影响靶基因表达而参与肿瘤的恶性进程。通过Pubmed、万方数据库和Google学术搜索发现miRNA-7作为细胞中广泛存在的非编码抑癌基因,在众多肿瘤(包括肝癌、宫颈癌、肺癌等)的增殖及转移中起重要作用,并且是患者早期诊断的标志物及影响患者预后的关键因素,然而,在胰腺癌的增殖和转移过程中miRNA-7研究目前罕见报道。circRNA是具有基因表达调控作用的非编码RNA,其主要通过套索内含子或反向拼接的方式产生,具有结构稳定和组织表达特异性等一系列特征,这些特征赋予了 circRNA不同的功能,例如,作为miRNA海绵,能与miRNA竞争性结合,干扰miRNA对靶mRNA的抑制和调控作用,从而影响下游靶基因的表达及其蛋白的合成,此外,有研究显示circRNA不但可以抑制miRNA活性,而且,还可抑制miRNA的表达。其中具有代表性的研究即发现小脑变性相关蛋白1反义转录物(CDRlas),也称做ciRS-7,其包含了 70多个miRNA的别位点,最具特征性的就是可以抑制miRNA-7活性和表达,从而提高miRNA-7的靶基因表达影响生物学功能。最近在喉鳞状细胞癌和非小细胞肺癌的研究中,证实ciRS-7可以通过抑制miRNA-7调节信号转导和转录活化因子 3(Signal transducers and activators of transcription 3,STAT3)、细胞周期蛋白E1(Cyclin E1,CCNE1)、表皮生长因子受体(Epidermal growth factor receptor,EGFR)、磷脂酰肌醇-3激酶催化亚基 δ(Phosphoinositide-3 kinase,catalytic subunit delta,PIK3CD)等靶基因信号通路关键蛋白的表达影响肿瘤的增殖和侵袭特性。研究已证实EGFR/STAT3信号转导通路在乳腺癌、胶质瘤、肺癌等许多肿瘤中异常表达,并影响肿瘤的增殖和转移。基于此,我们拟探索在胰腺癌中ciRS-7和miRNA-7表达,及其对胰腺癌生物性特性的影响。进一步明确ciRS-7通过与miRNA-7竞争性结合而调节miRNA-7靶基因EGFR等表达,进而影响EGFR/STAT3信号通路活性的作用和机制,探索ciRS-7影响胰腺癌生物学特性的机制。查阅文献后发现迄今为止相关文献报道较少。本课题主要分为三个部分:第一部分ciRS-7与miRNA-7在胰腺癌组织中的表达及临床意义研究目的:研究ciRS-7与miRNA-7等在胰腺癌组织中的表达情况,并探索ciRS-7在组织中的表达与胰腺癌生物学特性的关系,如与肿瘤的发生发展,侵袭和转移的关系,并分析ciRS-7对胰腺癌临床预后预测的意义。研究方法:本研究获得安徽省立医院伦理委员会批准,在安徽省立医院普外科共获取胰腺癌及癌旁组织标本41例,所有胰腺癌患者均签署知情同意书,进行后续研究。1.通过实时定量 PCR(Quantitative Real-time PCR,QRT-PCR)方法检测 41 例胰腺癌及其癌旁新鲜冰冻组织标本中ciRS-7和miRNA-7表达情况,并将ciRS-7和miRNA-7的表达情况进行相关性分析,探索ciRS-7和miRNA-7的表达与肿瘤发生、发展的关系。2.将胰腺癌组织中ciRS-7的表达与临床病例资料进行统计分析,探索其表达与肿瘤位置、大小、淋巴结转移以及局部神经浸润等因素是否相关。3.通过免疫组化(Immunohistochemistry,IHC)检测分析 EGFR、STAT3 在胰腺癌及癌旁等组织中的表达情况,并与临床病例资料进行统计分析。4.术后进行患者随访,分析ciRS-7的高表达对胰腺癌临床预后预测的意义。结果:1.ciRS-7在胰腺癌组织中表达高于癌旁组织,差异具有统计学意义(P=0.002),而miRNA-7在胰腺癌组织中表达低于癌旁组织,差异具有统计学意义(P=0.048)。Spearman相关性分析研究发现在胰腺癌中ciRS-7和miRNA-7表达具有负相关性(P=0.023。2.对于临床资料的统计发现,ciRS-7在胰腺癌中表达水平与肿瘤是否有淋巴结转移(P=0.016)及是否有神经浸润(P=0.028)相关,而与患者的性别、年龄、有无糖尿病、肿瘤位置、肿瘤大小等无明显相关,差异无统计学意义(民P>0.05)。3.免疫组化检测发现EGFR、STAT3在胰腺癌中表达高于癌旁组织,EGFR、STAT3在胰腺癌中表达水平均与肿瘤是否有淋巴结转移(P<0.05)相关,而与患者的性别、年龄、肿瘤位置等无明显相关,差异无统计学意义(P>0.05)。4.通过随访1年后,发现在胰腺癌患者中ciRS-7高表达者的无瘤生存率低于ciRS-7低表达者,差异具有统计学意义(P<0.05)。结论:ciRS-7在胰腺癌中表达高于癌旁组织,miRNA-7在胰腺癌中表达低于癌旁组织,并且两者表达具有负相关性。ciRS-7的表达与肿瘤的有无淋巴结转移和有无神经浸润相关。在胰腺癌组织中EGFR、STAT3高表达并与肿瘤增殖、侵袭生物学特性有关。术后随访显示ciRS-7高表达可能是胰腺癌预后的一个评估指标。故组织学研究表明ciRS-7和miRNA-7等表达与肿瘤发生发展可能相关,并为胰腺癌的诊断和预后分析提供依据。第二部分 体外探讨ciRS-7通过miRNA-7调节EGFR/STAT3信号通路促进胰腺癌增殖和转移的作用与机制研究目的:通过体外研究探讨ciRS-7和miRNA-7在胰腺癌增殖和转移中的作用及其机制研究方法:1.通过QRT-PCR法检测胰腺癌细胞株(BXPC-3、PANC-1)以及正常胰腺细胞HPC-Y5 中 ciRS-7、miRNA-7 表达情况,通过小干扰 RNA(Small interfering RNA,siRNA)siciRS-7、simiRNA-7 实现胰腺癌 BXPC-3、PANC-1 细胞中 ciRS-7、miRNA-7的沉默。2.QRT-PCR检测ciRS-7沉默组、ciRS-7与miRNA-7同时沉默组、对照(Control)组以及阴性对照(Negative control,NC)组等各组中miRNA-7表达情况。3.通过MTT细胞增殖实验检测BXPC-3、PANC-1各组细胞增殖能力变化情况。4.Transwell小室检测BXPC-3、PANC-1各组细胞侵袭能力变化情况。5.通过QRT-PCR法及Western blotting法检测各组中EGFR/STAT3的表达情况。6.各实验组细胞中加入EGFR/STAT3信号通路的特异性阻断剂吉非替尼后,再通过MTT细胞增殖试验和Transwell小室法检测各组细胞的增殖和侵袭能力变化的情况。结果:1.胰腺癌细胞BXPC-3及PANC-1中ciRS-7表达均高于正常胰腺细胞HPC-Y5(P<0.05)。siciRS-7、simiRNA-7 可有效沉默 ciRS-7、miRNA-7 在 BXPC-3 和 PANC-1中的表达。并分别在BXPC-3及PANC-1细胞系中筛选出ciRS-7低表达最明显的siciRS-7-1,和 ciRS-7 与 miRNA-7 同时低表达的 siciRS-7-1+simiRNA-7 用于后续研究。2.QRT-PCR检测发现siciRS-7-1组中miRNA-7表达情况高于Control组和NC组(P<0.05);siciRS-7-1+simiRNA-7 组中 miRNA-7 表达与 Control 组相比差异无统计学意义(P>0.05)。3.通过MTT法发现siciRS-7-1组癌细胞的增殖能力明显低于Control组和NC组(P<0.05),而siciRS-7-1+simiRNA-7组细胞的增殖能力与Control组相比差异无统计学意义(P>0.05)。Transwell小室法发现siciRS-7-1组癌细胞的侵袭能力明显低于Control 组和 NC 组(P<0.05),siciRS-7-l+simiRNA-7 组细胞的侵袭力与 Control 组相比差异无统计学意义(P>0.05)。4.siciRS-7-1组胰腺癌细胞中EGFR、STAT3的表达低于Control组(P<0.05),siciRS-7-1+simiRNA-7组中EGFR、STAT3的表达与Control组相比差异无统计学意义(P>0.05)。5.加入EGFR/STAT3阻断剂吉非替尼后,siciRS-7-1组中对于细胞的增殖和侵袭能力无明显影响(P>0.05),而Control组加入吉非替尼后的增殖和侵袭能力变化明显(P<0.05)。结论:体外研究阐明了在胰腺癌细胞中ciRS-7高表达,下调ciRS-7可促进miRNA-7上调表达,从而抑制miRNA-7下游的靶基因信号通路EGFR/STAT3表达,最终抑制肿瘤的增殖和转移过程,表明了 ciRS-7可促进胰腺癌细胞的增殖和转移,这些为肿瘤靶向治疗提供研究基础。第三部分 体内探讨c iRS-7通过miRNA-7调节EGFR/STAT3信号通路促进胰腺癌增殖和转移的作用与机制研究目的:通过体内实验研究验证ciRS-7可通过miRNA-7调控EGFR/STAT3信号通路影响胰腺癌的增殖和转移。研究方法:1.将siciRS-7-1、siciRS-7-1+simiRNA-7等胰腺癌PANC-1细胞分别注射在裸鼠皮下和腹腔,成功建立胰腺癌的裸鼠皮下移植瘤和腹腔转移瘤模型。2.对于皮下移植瘤模型,饲养4周后,处死裸鼠剥取肿瘤后称重和测量体积。对于腹腔转移瘤模型,观察裸鼠腹部情况,饲养4周后处死各组裸鼠,观察各组裸鼠的肝肺转移情况。3.应用QRT-PCR法检测各种模型肿瘤组织中miRNA-7表达情况。4.应用免疫组化检测各种模型肿瘤组织中EGFR、STAT3蛋白的表达情况。结果:1.成功建立胰腺癌裸鼠皮下移植瘤和腹腔转移瘤模型。2.siciRS-7-1组皮下移植瘤的体积和瘤重明显小于Control组(P<0.05)。腹腔转移瘤模型中siciRS-7-1组的肝肺转移的发生情况明显少于Control组(P<0.05)。3.QRT-PCR发现miRNA-7在两种模型的siciRS-7-1组中表达均高于Control组(P<0.05)。4.免疫组化结果显示两种模型中siciRS-7-1组的EGFR、STAT3蛋白阳性表达均低于Control组(P<0.05)。结论:体内实验进一步证实在胰腺癌中下调ciRS-7可促进miRNA-7的表达,并可通过抑制其下游的靶基因信号通路EGFR/STAT3表达抑制肿瘤的增殖和转移过程。全文结论:ciRS-7在胰腺癌组织中的表达明显高于癌旁组织,其表达与肿瘤的淋巴结转移和神经浸润进程有一定的相关性,是评估胰腺癌预后的重要指标。在胰腺癌细胞中ciRS-7高表达,下调ciRS-7表达可促进miRNA-7上调表达从而抑制EGFR/STAT3信号通路抑制肿瘤的增殖和侵袭。综合研究表明ciRS-7通过miRNA-7调节EGFR/STAT3信号通路促进胰腺癌增殖与转移。ciRS-7有望为胰腺癌的早期诊断和治疗提供新的潜在靶点,为后续肿瘤基因靶向治疗提供研究基础。更多还原

【Abstract】 Pancreatic cancer(Pancreatic ductal adenocarcinoma PDAC)is a kind of digestive system tumor with high degree of malignancy.Due to the development of living standard and poor diet and living habits,the incidence of Pancreatic cancer has increased about sixfold in the past 20 years.Surgical resection is still the preferred treatment for pancreatic cancer.Therefore,how to improve the preoperative diagnosis rate of pancreatic cancer,master the surgical indications,and improve the surgical resection rate is the key to ensure good efficacy of pancreatic cancer patients.The epidemiological studies of pancreatic cancer in recent years have found that the 5-year survival rate is still low,about 8.5%,the most important reason is the high malignant degree,strong invasion and may associate with lymph node metastasis along with nerve vascular infiltration in an early stage,therefore many patients have been in an advanced stage when diagnosed,and can soon appear local and distant metastasis.For lack of the early detection markers of pancreatic cancer,finding markers for early occurrence and development of pancreatic cancer will be a key factor in improving the prognosis of patients,In recent years,more and more scholars have noticed that the non-coding genes including micro RNA,circular RNA,Long non-coding RNA which were neglected before were closely correlated with occurrence and development of cancer,and part of them can even be applied to tumor diagnosis as early detection markers.MiRNA is a kind of non-coding single strand small RNA with the size of about 22-24 nucleotides,existing in eukaryotic cells and is involved in gene regulation after transcription.It can specifically identify and bind to the 3’-untranslated area(3’UTR)in Messenger ribonucleic acid(mRNA)of target gene,causing inhibition of target gene mRNA translation,thus implementing regulation on post transcriptional expression of target gene,and finally taking part in development of malignant tumor by affecting target gene expression.It is via schorlarly searching engines like Pubmed,Wan Fang Data and Google scholar can we find that the miRNA-7,a kind of non-coding anti-cancer gene widely exists in cells,palys an important role in the proliferation and metastasis of numerous tumors(including liver cancer,cervical cancer,lung cancer,etc.),and it is also the marker for early diagnosis and key factor affecting patients’ prognosis,however the research of miRNA-7 on proliferation and metastasis of pancreatic cancer have been rarely reported until now.CicrRNA is a kind of non-coding RNA,with the function of gene expression regulation,can be produced by means of reverse stitching or lasso introns,having a series of characteristics including evalutionary conservative,structure stability and tissue expression specificity,which give different functions to circRNA,for instance,as the miRNA sponges,can compete with mRNA for binding to miRNA,can interfere the inhibition and regulation function of miRNA to target mRNA,thus affecting the expression and relevant protein synthesis of downstream gene.One of the representative studies found that CDR1as,also known as ciRS-7,contains more than 70 miRNA loci,which can competitively inhibit miRNA-7 activity,thereby improving the expression of miRNA-7 target genes and affecting biological functions.In recent studies of squamous cell carcinoma of the larynx and non-small cell lung cancer.It was confirmed that ciRS could regulate the expression of PIK3CD,EGFR,CCNE1,STAT3 by inhibiting miRNA-7,and affect the proliferation and invasion of tumors.Previous studies have confirmed that EGFR/STAT3 signal transduction pathway is abnormally expressed in many tumors,such as breast cancer,glioma and lung cancer,and affects the proliferation and metastasis of tumors.Based on this,we aim to explore the expression of ciRS-7 and miRNA in pancreatic cancer and the effect of which on biological characteristics of pancreatic cancer,to further identify the effect and mechanism of ciRS-7 affecting the activity of EGFR/STAT3 signaling pathway by regulating the expression of EGFR etc.,the target genes of miRNA-7 via competitively binds to miRNA-7,and explore the mechanism of ciRS-7 influencing the biological characteristics of pancreatic cancer.It was found that few relevant literatures were reported so far after reviewing literatures.This topic is mainly divided into three parts:Part I Expression of CiRS-7 and miRNA-7 expression in pancreatic cancer tissues and their clinical significanceResearch objectives:To study the expression of ciRS-7 and miRNA-7 in pancreatic cancer tissues,and to explore the relationship between ciRS-7 expression in tissues and the biological characteristics of pancreatic cancer,such as the relationship with tumor development,invasion and metastasis,and analyze the significance of ciRS-7 in the prediction of clinical prognosis of pancreatic cancer.Research methods:This study was approved by the Ethics Committee of Anhui Provincial Hospital.A total of 41 PDAC tumors and their paracancerous tissues were obtained from General Surgery Department of Anhui Provincial Hospitall.All patients had signed the informed consent before the follow-up study.1.Quantitative real-time PCR(QRT-PCR)was used to detect the expression of ciRS-7 and miRNA-7 in 41 cases of PDAC tumors and paracancerous fresh frozen tissues.Correlation analysis was conducted on the expression of ciRS-7 and miRNA-7 to explore the relationship between the expression of ciRS-7 and miRNA-7 along with the occurrence and development of tumor.2.The expression of ciRS-7 in pancreatic cancer tissues and the clinical data were statistically analyzed to explore whether the expression of ciRS-7 was correlated with location,size of tumor,lymph node metastasis,local nerve infiltration and other factors.3.Expression of EGFR and STAT3 in pancreatic cancer and paracancerous tissues detected and analyzed by Immunohistochemistry(MHI)along with the clinical data were statistically analyzed.4.Postoperative follow-up was conducted,analyzing the significance of high ciRS-7 expression on clinical prognostic prediction of pancreatic cancer.Results:1.CiRS-7 expression in pancreatic cancer tissues was higher than that in adjacent tissues,and the difference was statistically significant(P<0.01),while miRNA-7 expression in pancreatic cancer tissues was lower than that in adjacent tissues,and the difference was statistically significant(P<0.05).Spearman correlation analysis revealed a negative correlation between ciRS-7 and miRNA-7 expression in pancreatic cancer(r=-0.035,P<0.05).2.According to statistical analysis of clinical data,expression level of ciRS-7 in pancreatic cancer was associated with lymph node metastasis(P=0.016)and nerve invasion(P=0.028)of tumor,but not gander and age of patients,with diabetes or not,location,size of tumor(P>0.05).3.After detected by IHC,the expression level of EGFR,STAT3 in pancreatic cancer tissue was higher than adjacent tissue,and was correlated to lymph node metastasis of tumor(P<0.05),but not gender and age of patients,location of tumor(P>0.05).4.After 1 year of follow-up,it was found that the tumor-free survival rate of pancreatic cancer patients with up-regulated ciRS-7 expression was lower than those with down-regulated ciRS-7 expression,and the difference was statistically significant(P<0.05).Conclusion:CiRS-7 expression in pancreatic cancer is higher than that in adjacent tissues,while miRNA-7 expression in pancreatic cancer is lower than that in adjacent tissues,and the expressions of the two are negatively correlated.CiRS-7 expression is correlated with lymph node metastasis and nerve invasion.Expression of EGFR and STAT3 could be found in pancreatic cancer tissue and was correlated with biological characteristics like proliferation and invasion etc.of tumor.Follow-up survey showed that higher ciRS-7 expression may be an indicator of prognosis of pancreatic cancer.Histological studies have shown that ciRS-7 and miRNA-7 expression may be correlated with tumor development,and provide a basis for the diagnosis and prognosis of pancreatic cancer.Part II To investigate the effect and mechanism of circular RNA ciRS-7 promoting proliferation and metastasis of pancreatic cancer by regulating miRNA-7 mediated EGFR/STAT3 signaling pathway in vitroResearch objectives:The role and mechanism of ciRS-7 and miRNA-7 in the proliferation and metastasis of pancreatic cancer were investigated in vitroResearch methods:1.The expression of ciRS-7,miRNA-7 in pancreatic cancer cell lines(BXPC-3,PANC-1)and normal pancreatic cells HPC-Y5 was examined by method of QRT-PCR.Silenced the expression of ciRS-7 and miRNA-7 in pancreatic cancer cells BXPC-3,PANC-1 by small interfering RNA(siRNA),and screened the ciRS-7-1 that stably low expressing ciRS-7 along with siciRS-7-1+simiRNA-7 that silenced expression of miRNA-7.2.Detecting expression of miRNA-7 in siciRS-7 group,siciRS-7&simiRNA-7 group,control group and negative control group(NC)by QRT-PCR.3.Testing the proliferation of cells in every group through MTT cell proliferation test.4.Cell invasiveness in every group was tested by Transwel Small chamber.5.Expression of EGFR/STAT3 in every group were examined by QRT-PCR and Western blotting.6.Change of cell proliferation and invasiveness ability in every group was examined by MTT cell proliferation test and Transwell Small chamber after Gefitnib,the specific blocker of EGFR/STAT3 signaling pathway was added.Results:1.CiRS-7 expression in pancreatic cancer cells was higher than that in normal pancreatic cells(P<0.05),and low expression ciRS-7 group was selected for further study.2.QRT-PCR detection revealed that the expression of miRNA-7 in the siciRS-7-1 group was higher than that in the control group and the blank control group(P<0.05).3.MTT assay showed that the proliferation of cancer cells in the siciRS-7-1 group was significantly lower than that in the control group and the blank control group(P<0.05).while no significant difference of cells proliferation ability was found between siciRS-7-1+simiRNA-7 group and control group(P>0.05).Transwell chamber method also found that the invasion ability of cancer cells in the siciRS-7-1 group was significantly lower than that in the control group and the blank control group(P<0.05),while no significant difference of cells invasion ability was found between siciRS-7-1+simiRNA-7 group and control group(P>0.05).4.The expression of EGFR and STAT3 in pancreatic cancer cells of siciRS-7-1 group was lower than that in control group(P<0.05),while the expression of EGFR and STAT3 in siciRS-7-1+simiRNA-7 group was similar to that in control group(P>0.05).5.After Gefitnid,the blocker of EGFR/STAT3 signaling pathway was added,no influence was found on proliferation and invasion ability of cells in siciRS-7-1 group(P>0.05),while proliferation and invasion ability changed significantly in control group(P<0.05).Conclusion:It has been clarified by the vitro study that down regulating ciRS-7 expression could promote miRNA-7 expressing,and regulated the expression of miRNA-7 downstream signaling pathway,the EGFR/STAT3 signaling pathway,thus inhibiting the proliferation and metastasis of tumor.This study show that ciRS-7 promotes the proliferation and metastasis of pancreatic cancer.It provided a research basis for target treatment of tumor.Part III To investigate the effect and mechanism of circular RNA ciRS-7 promoting proliferation and metastasis of pancreatic cancer by regulating miRNA-7 mediated EGFR/STAT3 signaling pathway in vivoResearch objectives:To verify ciRS-7 can affect proliferation and metastasis of pancreatic cancer by regulating miRNA-7 mediated EGFR/STAT3 signaling pathway by vivo study.Research methods:1.Injected recombinant siciRS-7-1 and siciRS-7-1+simiRNA-7 pancreatic cancer cells PANC-1 in subcutis and abdominal cavity of nude mice respectively,and established the nude mice model of subcutaneous transplantation tumor and peritoneal metastatic tumor.2.All nude mice were executed after being raised for 4 weeks.Measured the weight and volume of tumors stripped from mice along with observed the liver metastasis.3.Measured the expression of miRNA-7 in tumor model tissue by QRT-PCR method4.Tested the expression of EGFR and STAT3 in tumor model tissue by IHC.Results:1.Subcutaneous transplantation and peritoneal metastasis model of pancreatic cancer in nude mice were established2.Volume and weight of subcutaneous transplantation tumor in siciRS-7-1 group was significantly lower than control group(P<0.05).Hepatopulmonary metastasis in siciRS-7-1 group was significantly less than that in control group in abdominal metastasis group(P<0.05).3.By QRT-PCR method,we could find that the expression of miRNA-7 in siciRS-7-1 group of both models were than that in control group(P<0.05).4.The IHC showed that protein expression of EGFR and STAT3 of siciRS-7-1 group was lower than control group in both models.Conclusion:The in vivo study has confirmed the results of the in vitro study that the down-regulation of ciRS-7 can promote the expression of miRNA-7,then up-regulated miRNA-7 can inhibit the proliferation and metastasis of tumor by down-regulating EGFR/STAT3 signaling pathway.Conclusion of the article:CiRS-7 expression in pancreatic cancer tissues is obviouse higher than in adjacent tissues,and its expression is correlated with lymph node metastasis and nerve infiltration,which is an important indicator for the prognosis of pancreatic cancer.The low expression of ciRS-7 can inhibit the proliferation and invasion of pancreatic cancer,by up regulating the expression of miRNA-7 and obstructing EGFR/STAT3 signaling pathway.Comprehensive studies show that ciRS-7 promotes the proliferation and metastasis of pancreatic cancer by regulating miRNA-7 mediated EGFR/STAT3 signaling pathway.ciRS-7 is expected to be a new potential target for the early diagnosis and treatment of pancreatic cancer,and provides a research basis for the follow-up targeted therapy of tumor genes.更多还原