Select Different Expression Long Noncoding RNA Genes In Urothelial Carcinoma Of Bladder By Microarray

Objective:Select different expression Long noncoding RNA genes in urothelial carcinoma of bladder by microarray,and verify screening results.Methods:After obtaining all the subjects themselves or their immediate relatives’ consent and the hospital Medical Ethics Committee agreement. We got specimens from March2010to March2011in our hospital from the patients who had bladder tumor transurethral resection, partial cystectomy or radical cystectomy.35cases pathologically diagnosed as urothelial carcinoma of the bladder and10cases are normal bladder mucosa. Then we choosed12cases specimens of urothelial carcinoma of the bladder and3cases specimens of normal bladder mucosa(2cases of them were women,13cases of them were men,their age were from35to78years old(average61years old)), we extracted RNA,transcribe it into cDNA and labeled it. Then we used the probe which have already be labeled hybridize with microarray.After washing, microarrays were scanned with the Agilent DNA Microarray Scanner. Data was extracted using Agilent Feature Extraction software. To identify differentially expressed LncRNAs with statistical significance, we performed a Volcano Plot filtering between two groups of samples (Fold Change>=2.0, P-value<=0.05). Then we used RT-PCR assay to amplify three of the differentially expressed genes. By using the house-keeping gene GAPDH as internal control,the quantity of genes devided by the quantity of GAPDH was the relative content of genes.After that we got the average value of the relative content of genes.Results:Compared with nomarl bladder tissues and urothelial carcinoma of the bladder tissues,we found25differentially expressed genes by Rinn lincRNAs profiling, lincRNA-TSPN8、AC079305.6、lincRNA-BBOX1-2were3genes which were downregulated in tumor tissues. lincRNA-SLC39A10-1、lincRNA-LMO1-2、BC039493、AL355916.1、 lincRNA-NFE2L3-3、AX746599、lincRNA-SNAI2-1、lincRNA-LOC285733-1、 lincRNA-DACT2-3、lincRNA-RPUSD4-2、AC004987.7、lincRNA-MEF2A、lincRNA-ALX1-1、 lincRNA-LOC100500938-1、lincRNA-KLRB1、BDNFOS、lincRNA-STK35、RP11-572M11.2. lincRNA-PRSS16、RP11-315H15.1、lincRNA-LOXL1-2were22genes which were upregulateed in tumor tisses.And we found7differentially expressed genes by Enhnacer lincRNAs profiling, BC039493、RP11-102N11.1、BC041451、AF127936.3、RP11-552E20.1、AC068610.3、 RP11-315H15.1were7genes which were upregulated in tumor tisses. Then we used RT-PCR assay to amplify lincRNA-PRSS16, RP11-315H15.1and BC039493.The lincRNA-PRSS16had maximum difference of chip signal, RP11-315H15.1and BC039493were two genes which we can be found by both Enhnacer lincRNAs profiling and Rinn lincRNAs profiling.The final results showed that the expression of three kinds of genes in tumor tissue does exist high degrees of expression:the lincRNA the-RPSS16tumor/normal=11.5887; BC039493tumor/normal=2.4924; RP11-315H15.1tumor/normal=5.5211.In all the genes which have different expression between human urothelial carcinoma and normal bladder tissues,we found that AC079305.6is downregulateed in non-muscle invasive bladder cancer,normal in muscle invasive bladder cancer and lincRNA-LOC285733-1,AC068610.3are upregulateed in non-muscle invasive bladder cancer,normal in muscle invasive bladder cancer.Conclusion:Long noncoding RNA genes have expression differences between human urothelial carcinoma and normal bladder tissues.We found25different expression genes by Rinn lincRNA profiling and7different expression genes by Enhancer LncRNA profiling. Some of these genes may become new targets for treatment of bladder urothelial carcinoma or new tumor markers.We choosed3genes which was the most likely to become tumor markers to do the RT-PCR assay,the PCR results met the results of microarrays.Compared with NMIBC and MIBC,we found3genes have different expression,which may have relationship with canceration.