To Explore The Expression Characteristics And Regulatory Network Of CircRNAs In Tuberculosis Patients Based On Transcriptome

Objective:To understand the expression pattern of circRNAs in pulmonary tuberculosis patients and healthy individuals and screen out the biomarker for early diagnosis of pulmonary tuberculosis,the peripheral blood leucocyte extracted from the pulmonary tuberculosis patients and healthy individuals were used to analyze with transcriptome sequencing.Metheds:According to the clinical detection,the peripheral blood leucocyte extracted from the 3 pulmonary tuberculosis patients and 3 healthy individuals were used as samples for circRNAs identification with Solexa transcriptome sequencing.Depend on the clinical classification,the sequencing data were analyzed by clustering.Differential expressed circRNAs from the compared with two groups were screened out the the threshold value false discovery rate(FDR)<0.05 and Foldchange(FC)>2.The consistent of the sequencing data was further validated with real-time PCR,Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway analysis were applied to explore the potential circRNA functions of the significantly deregulated genes.Competing endogenous RNAs(ceRNA)were constructed and inferred that aberrant expression of circRNA-associated ceRNA resulted in extensive variation in gene expression by miRNA-mediated circRNA-mRNA crosstalk interactions.It will help us to predict the potential roles of these dysregulated circRNAs in the peripheral blood leucocyte of pulmonary tuberculosis patients.Results:Transcriptome sequencing was used to identify circRNAs from 3 PTB patients and 3 healthy individuals to determine the expression pattern of circRNAs in blood and the circRNA molecular regulatory networks in pulmonary tuberculosis pathogenesis.Total 16288 circRNAs were identified from the samples.After compared with circbase,7775 circRNAs were considered as the known circRNAs,and 8513 circRNs were regarded as the novel circRNAs.Compared with two group,one hundred and seventy differentially expressed(≥2-fold change,FDR<0.05)circRNAs were dysregulated in pulmonary tuberculosis,compared with in healthy individuals.Quantitative real-time polymerase chain reaction was used to validate the RNA sequencing analysis from 20 pulmonary tuberculosis patients,and the results were consistent with the sequencing data.These circRNs can be used as the potential biomarker for diagonosis of pulmonary tuberculosis.To known the regulatory mechanism of these dysregualted circRNAs in pulmonary tuberculosis patients,we constructed a circRNA-associated ceRNA network by integrating the expression profiles and regulatory relationships of the circRNAs,miRNAs and mRNAs from the sequencing data of our 6 samples.A total of 4350 ceRNA relationships was found from the interaction of 41 differentially expressed miRNAs,2565 differentially expressed mRNAs and 170 differentially expressed circRNAs.According to the expression tendency of the differentially expressed genes in the ceRNAs,two networks were obtained.The first network of 1721 circRNA-miRNA-mRNAs contained 18 upregulated circRNAs and 472 mRNAs and 14 downregulated miRNAs.The second network of 666 circRNA-miRNA-mRNAs contained both 6 downregulated circRNAs and 307 mRNAs and 8 upregulated miRNAs.Conclusions:The screen out differential expressed circRNA may be used as the potential biomarker for diagonosis of pulmonary tuberculosis patients and these eeRNA regulatory networks included circRNAs,miRNAs and mRNAs that might play a pivotal role in the pathogenesis of pulmonary tuberculosis and provided important theoretical basis for circRNAs in the pathogenesis of pulmonary tuberculosis.