A Comparative Study Of Serum Exosomes MiRNA And CT Findings Based On Pancreatic Cancer

PART ? Screening and verification of miRNA markers in serum exosomes of pancreatic cancerObjective: The mic RNA in the serum exosomes of pancreatic cancer was screened as a diagnostic biomarker by Illumina second-generation sequencing,and the sample size was expanded to verify the selected mic RNA.Materials and Methods:(1)Case selection: According to the inclusion and exclusion criteria,from June 2017 to January 2019,Jiangsu Provincial Hospital of Traditional Chinese Medicine,Nanjing Drum Tower and the PLA General Hospital of the Eastern Military Region totaled 56 patients,including 28 patients with pancreatic cancer.There were 23 patients with benign pancreatic lesions and 5 normal controls.Among them,10 patients with pancreatic cancer,5 patients with benign pancreatic lesions,5 normal controls were used for Illumina second generation sequencing;18 patients with pancreatic cancer and 18 patients with benign pancreatic lesions were used to validate the biomarkers.(2)Exosome extraction and identification: exosomes were extracted using exo Easy Maxi Kit manufactured by Qiagen,Germany,and extracted exosomes were identified by transmission electron microscopy,nanoparticle tracking analysis and Western blot.(3)Total RNA extraction from exosomes: RNA from serum exosomes was extracted using the miRNeasy Serum/Plasma Kit from Qiagen,Germany.(4)Quality analysis of the extracted Total RNA: The obtained Total RNA was subjected to quality inspection using an Agilent RNA 6000 Nano Reagents Part I kit manufactured by Agilent.(5)Sequencing of RNA: Deep sequencing of the library to be passed through the assay using Ilbisa's Hiseq Xten.(6)Screening miRNA as a biomarker: analyzing the data obtained by sequencing,and screening miRNA as a biomarker according to the miRBase database using GO and KEGG enrichment analysis.(7)Biomarker verification: The screened miRNA was verified by RT-PCR technique.For the difference between the two,the difference multiple is greater than 1.5,which means that it is meaningful.Result: The results of sequencing of 20 samples were analyzed by two groups.The first group was pancreatic cancer group and normal control group(5 vs 5),and the second group was pancreatic cancer and benign pancreatic lesion group(5 vs 5).In the sequencing data of the group,there were 11 miRNAs with the same expression trend,and 6 of them were up-regulated(hsa-miR-203b-5p,hsa-miR-342-5p,hsa-miR-337-5p,hsa-miR-149-5p,hsa-miR-877-5p,hsa-miR-203a-3p),5 expressions down-regulated(hsa-miR-1226-3p,hsa-miR-3182,hsa-miR-625-3p,hsa-miR-624-5p,hsa-miR-664a-5p).By comparing the miRBase database and target gene GO analysis,KEGG pathway analysis finally selected hsa-miR-1226-3p,which is down-regulated in pancreatic cancer serum exosomes,as a potential diagnostic marker for pancreatic cancer for further verification.For the biomarker hsa-miR-1226-3p,RT-PCR was used to verify the serum of patients with expanded samples.It was found that hsa-miR-1226-3p was down-regulated in pancreatic cancer patients,compared with serum in patients with benign pancreatic lesions.The down-regulation factor is 3.1,and the difference is clinically significant.Conclusion: The content of serum exosomes hsa-miR-1226-3p in patients with pancreatic cancer is different from that in patients with benign pancreatic lesions and normal controls,and the difference is clinically significant.hsa-miR-1226-3p in serum exosomes can be used as a specific biomarker for the diagnosis of pancreatic cancer.part ? A comparative study of miRNA-1226 in serum exosomes and CT imagings of pancreatic cancerObjective: The expression level of miRNA-1226 in serum exosomes of pancreatic cancer was compared with the CT imagings to explore the relationship between the expression of miRNA-1226 and imaging features.Materials and Methods:(1)Case selection: Select the images of 18 patients with pancreatic cancer who were used in the first part of the trial.(2)Analysis of the patient's age,gender,lesion size,location,morphology,border,density,enhanced CT scan and special signs(such as calcification,pancreaticobiliary dilatation,etc.)and adjacent tissue invasion or distant metastasis,etc.Imaging findings were compared with the expression of mic RNA-1226 in serum exosomes of pancreatic cancer,statistical software was used for SPSS 16.0 data analysis,and Mann-Whitney U test and one-way ANOVA were used to compare the measurement data.Spearman correlation analysis was used to calculate the correlation between measurement data and count data,Pearson correlation analysis was used to analyze the two groups of measurement data.RESULTS: When 18 patients were divided into two groups with or without invasion and metastasis,the2-??Ct value of miRNA-1226 in the serum exosomes of the invasion or metastasis group was smaller than that of the non-invasive metastasis group.The P value was 0.018.(less than 0.05).Correlation analysis using Spearman correlation analysis revealed that the 2-??Ct value of miRNA-1226 in exosomes was correlated with invasion and metastasis.The less 2-??Ct value of miRNA-1226 in exosomes was invaded or The greater the possibility of transfer.The expression levels of miRNA-1226 in serum exosomes were compared with the tumor size,CT value of each stage and pancreatic duct dilatation.No significant correlation was found between them.When the cut off value was 1.0122,the sensitivity and specificity for the diagnosis of invasion or metastasis were 81.8%,85.2%,and the area under the ROC curve was 0.805.Conclusion: There is a correlation between the expression of miRNA-1226 in serum exosomes of patients with pancreatic cancer and local invasion or metastasis in patients with pancreatic cancer.The lower the expression,the greater the possibility of local invasion or metastasis.