Screening Of Differential Circular RNAs In Pancreatic Cancer And Bioinformatics Prediction And Experimental Validation Of The Pathogenic Mechanism Of Circ＿0007883

Objective : Pancreatic cancer is a kind of digestive tract malignant tumor with insidious onset,difficult diagnosis and poor prognosis.In recent years,the incidence of pancreatic cancer continues to rise,which has brought serious burden to the society.More importantly,pancreatic cancer is also the most deadly of all solid tumors,ranking seventh among all cancer-related causes of death.The early symptoms of pancreatic cancer patients are not obvious,and they are often in the advanced stage of the disease when diagnosed,losing the opportunity of surgical resection,which brings great challenges to the treatment.The 5-year survival rate for most patients is only10-20%,even if surgery is available.At the same time,the side effects and resistance of chemotherapy drugs are not conducive to the treatment of patients.At present,our understanding of pancreatic cancer is limited,so it is urgent to study the mechanism of its occurrence and development.Circular RNA（circRNA）is a kind of endogenous non-coding RNA.Unlike linear RNA,it has no free end,but a circular structure formed by covalent bonds.It can resist nuclease to a certain extent and is more stable in the body,so it is an ideal biomarker.A large number of studies have proved that compared with normal tissues,circ RNA is often expressed abnormally in tumors,which is closely related to tumor progression.Research on it may bring new hope for the diagnosis and treatment of pancreatic cancer.With the development of life science and computer science,bioinformatics has become an indispensable tool in medical research.Driven by high-throughput technology,a large number of circ RNAs have been discovered,and the study of circ RNAs playing a pathophysiological regulatory role as mi RNA sponges has aroused great interest of researchers.In this study,through bioinformatics analysis,circ₀007883,which is closely related to pancreatic cancer,was screened out,and the correlation between circ₀007883 and the clinicopathological data of patients with pancreatic cancer was studied from this point of view,and it was found that circ₀007883 was highly expressed in pancreatic cancer and promoted the occurrence of malignant behavior ofpancreatic cancer cells.Then,through bioinformatics prediction and experimental verification,it was proved that circ₀007883 / mi R-141-3p /HGF axis played an important role in the occurrence and development of pancreatic cancer.Methods: Firstly,the Gene Expression Omnibus（GEO）database was searched to screen the data set for the study of circ RNA Expression profile in pancreatic cancer.Filtering and quality control were carried out on the raw data of the sequencing data set,and difference analysis was carried out by using Deseq2 packet in R language.The difference analysis of chip data set after quality control was carried out by using limma packet in R language.The intersection of the difference analysis results of multiple data sets was taken to determine the research target as circ₀007883.Patients who underwent surgical resection for pancreatic cancer in our hospital were searched.A total of 33 patients with pancreatic cancer with detailed medical history were included and their cancer and paracancerous histopathological sections were obtained.Fluorescence in situ hybridization experiment proved that circ₀007883 expression was different between cancers and adjacent tissues,the relationship between circ₀007883 expression level in tumor tissues and clinicopathological data such as age,sex,related disease history,life history,tumor stage,tumor differentiation degree,serum CA19-9 level was analyzed by statistical method.Then,the expression of circ₀007883 in normal pancreatic cells HPDE6-C7 and pancreatic cancer cells Bx PC-3,Capan-2 and PANC-1 was detected by q RT-PCR.Then circ₀007883 was overexpressed and knocked down in pancreatic cancer cells,and cell phenotype tests were performed,including CCK-8 assay for proliferation,Transwell assay for migration and invasion,and flow cytometry for cell cycle.The circ Bank database was used to predict the mi RNA targeted by circ₀007883,and the survival analysis of mi RNA was performed in starbase database to screen out mi R-141-3p.Linked Omics and mi Rwalk databases were used to predict the target genes of mi R-141-3p,and the intersection of the above results was obtained.Protein-protein interaction analysis was performed on the target genes in the intersection,and the sub-network was extracted to screen out the hub genes.The circ₀007883 / mi R-141-3p /HGF axis was finally determined and constructed by analyzing the hub genes in GEPIA2 database.Subsequently,the expression levels ofmi R-141-3p and HGF in pancreatic cancer cells were verified by qRT-PCR.Finally,the binding relationships between circ₀007883 and mi R-141-3p and between mi R-141-3p and HGF were confirmed by dual luciferase reporter assay.Results: 1.Three data sets were obtained by searching GEO database,namely GSE136569,GSE69362 and GSE79634.The difference analysis found that circ₀007883 was highly expressed in all the data sets.2.The expression of circ₀007883 in pancreatic cancer cells was significantly higher than that in normal pancreatic cells（P<0.05）.3.The expression of circ₀007883 in pancreatic cancer tissues was significantly higher than that in paracancerous tissues（P<0.001）.The expression of circ₀007883in tumor tissues was significantly correlated with tumor T stage（P<0.001）,M stage（P=0.003）,TNM stage（P<0.001）,serum CA19-9（P=0.039）and tumor differentiation（P=0.016）.4.In Capan-2 and PANC-1 cell lines,overexpression of circ₀007883 promoted cell proliferation（P<0.05）,invasion（P<0.01）and migration（P<0.01）,and more cells were arrested in S phase.After knocking cric₀007883,the biological behaviors ofthe cells were reversed.5.Combined analysis of multiple bioinformatics methods showed that circ₀007883may be involved in the progression of pancreatic cancer through the circ₀007883 /mi R-141-3p /HGF axis.6.In Capan-2 and PANC-1 cell lines,the overexpression of circc₀007883 reduced the expression of mi R-141-3p and increased the expression of HGF,while the knockdown of circ₀007883 increased the expression of mi R-141-3p and decreased the expression of HGF.7.There were binding relationships between circ₀007883 and mi R-141-3p,and between HGF and mi R-141-3p in cells.Conclusion:1.circ₀007883 was highly expressed in pancreatic cancer tissues,and its expression level was closely related to clinicopathological data such as tumor stage,tumor differentiation and serum CA19-9.2.circ₀007883 was highly expressed in pancreatic cancer and played a carcinogenic role in pancreatic cancer,leading to the improvement of proliferation,migration andinvasion ability of pancreatic cancer cells.Knocking down circ₀007883 expression can inhibit the malignant behaviors.of pancreatic cancer cells.3.The existence of circ₀007883 / mi R-141-3p /HGF axis in pancreatic cancer was verified by bioinformatics and experiments.