| BackgroundBladder cancer(BC)is one of the most common malignancies of the urinary system.Its diagnosis mainly depends on pathology,and the pathogenesis remains to be elucidated,and there are no reliable tumor markers.We combined the application of transcriptomics and quantitative proteomics to further analyze the differentially expressed gene of tumor tissues obtained by BC radical surgery,and to screen for key biological signal molecules specific to BC tissues.Our research provides a theoretical basis for an in-depth understanding of the pathogenesis of BC,as well as new information for possible and molecular diagnostic marker discovery for the preventive diagnosis of BC.MethodsAll the clinical data of patients who went to the hospital for diagnosis and treatment of bladder tumor between January 2016 and December 2017 were collected.Transcriptomic analysis and quantitative proteomic analysis were performed on 9 patients with radical cystectomywere resected tumor tissue samples and adjacent cancer control samples that met the diagnosis of T2 stage muscle invasive bladder cancer(MIBC).The transcriptometrics analysis:total RNA of tissue samples was extracted and reversed transcription into cDNA following qualified quality control,then the sequencing library was prepared.Next,the library was sequenced by the next generation sequencing technology,and genes differentially expressed between BC tissues and control tissues and their possible biological functions were analyzed.Ten differentially expressed genes were randomly selected for verification.Expression of these DEGs was assayed using real-time quantitative PCR The quantitative proteomics analysis:proteins were extracted from tissue samples,and digested into peptides after qualified quality control.Then the peptides were labeled with TMT reagent and detected by liquid chromatography-mass spectrometry(LC-MS).Proteins differentially expressed in BC tissues and control tissues and their possible biological functions were analyzed.Results(1)81 cases of BC patients were collected,including 63 cases of non-muscle invasive bladder cancer(NMIBC),and all tumors were removed by urethral 2-micron laser monolithic resection,and then,pathology confirmed 59 cases of NMIBC of and 4 cases MIBC.Combined with the optimized specimen fixation methods,the muscle layer could be found on all the specimens.(2)Transcriptomics analysis revealed that there were a total of 7,094 significantly differently expressed genes in T2 stage BC tumor tissues(up-regulated expression genes 4040,down-regulated expression genes 3054),including CYP1A1,KRT5,MYH11,and ACTC1 known closely related genes.It was also show that nine out of ten DEGs analyzed by real-time quantitative PCR were consistent with the expression pattern of transcriptional data.(3)A total of 5,911 proteins were identified by quantitative proteomics,and 596 proteins were significantly differentially expressed(210 proteins were up-regulated and 386 proteins were down-regulated),including urea transporter 1(SLC14A1),calcium-binding protein S100-A12 and receptors.Subunit α-1(KPNA2)and other known closely related proteins of BC.(4)Analysis of integrated transcriptomics and proteomics data revealed 367 genes with the same differential expression trend.Further analysis indicated that these genes were significantly enriched in signal pathways such as cell adhesion,extracellular matrix receptor interaction,and complement and coagulation cascades,and could play an important role in BC cell differentiation,cell cycle regulation and cell proliferation.(5)Draw a differential molecular PPI network and find four hub genes,namely COL14A1,FBN1,HSPG2,and LAMC1.It was verified in the TCGA database that the expression of hub genes in tumor tissues was significantly lower than that in normal tissues.Survival prognosis is closely related.ConclusionsTransurethral 2 micron laser monolithectomy is an effective method for NMIBC,and can improve the accuracy of pathological staging by combined with optimized specimen fixation methods.There are a large number of differentially expressed genes in tumor tissue from patients with BC.These abnormally expressed products can be found at both the transcription and protein levels.Integrating analysis of genes with consistent expression trends in BC tissues in transcriptomics and proteomics,especially the 4 hub genes that were screened,could explain the pathogenesis of BC and provide theoretical basis for the development of diagnostic molecular markers. |
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