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Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses

BACKGROUND: We aimed to identify prognostic biomarkers of upper tract urothelial carcinomas (UTUCs), including microRNAs (miRNAs) and genes which account for only 5% to 10% of all urothelial carcinomas (UCs). In Taiwan, this figure is markedly higher, where it can reach up to 30% of UC cases. MATERI...

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Autores principales: Lee, Hsiang-Ying, Li, Ching-Chia, Li, Wei-Ming, Hsu, Ya-Ling, Yeh, Hsin-Chih, Ke, Hung-Lung, Yeh, Bi Wen, Huang, Chun-Nung, Li, Chien-Feng, Kuo, Po-Lin, Wu, Wen-Jeng
Formato: Online Artículo Texto
Lenguaje:English
Publicado: PeerJ Inc. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8086570/
https://www.ncbi.nlm.nih.gov/pubmed/33987019
http://dx.doi.org/10.7717/peerj.11343
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author Lee, Hsiang-Ying
Li, Ching-Chia
Li, Wei-Ming
Hsu, Ya-Ling
Yeh, Hsin-Chih
Ke, Hung-Lung
Yeh, Bi Wen
Huang, Chun-Nung
Li, Chien-Feng
Kuo, Po-Lin
Wu, Wen-Jeng
author_facet Lee, Hsiang-Ying
Li, Ching-Chia
Li, Wei-Ming
Hsu, Ya-Ling
Yeh, Hsin-Chih
Ke, Hung-Lung
Yeh, Bi Wen
Huang, Chun-Nung
Li, Chien-Feng
Kuo, Po-Lin
Wu, Wen-Jeng
author_sort Lee, Hsiang-Ying
collection PubMed
description BACKGROUND: We aimed to identify prognostic biomarkers of upper tract urothelial carcinomas (UTUCs), including microRNAs (miRNAs) and genes which account for only 5% to 10% of all urothelial carcinomas (UCs). In Taiwan, this figure is markedly higher, where it can reach up to 30% of UC cases. MATERIALS AND METHODS: Using next-generation sequencing (NGS), we analyzed two pairs of renal pelvis tumors and adjacent normal urothelial tissues to screen miRNAs and messenger RNAs. By combining bioinformatics analysis from miRmap, Gene Expression Omnibus (GEO), and Oncomine and Ingenuity(®) Pathway Analysis databases, we identified candidate genes. To search for upstream miRNAs with exact target binding sites, we used miRmap, TargetScan, and miRDB to enforce evidence. Then, we clarified gene and protein expression through an in vitro study using western blot analysis and quantitative real-time reverse transcriptase-PCR. RESULTS: Interactions between selected target genes obtained using the NGS and miRmap methods were assessed through a Venn diagram analysis. Six potential genes, namely, PDE5A, RECK, ZEB2, NCALD, PLCXD3 and CYBRD1 showed significant differences. Further analysis of gene expression from the GEO dataset indicated lower expression of PDE5A, RECK, ZEB2, and CYBRD1 in bladder cancer tissue than in normal bladder mucosa, which indicated that PDE5A, RECK, ZEB2, and CYBRD1 may act as tumor suppressors in UTUC. In addition, we compared the expression of these genes in various UC cell lines (RT4, BFTC905, J82, T24, UMUC3, 5637, BFTC 909, UMUC14) and found decreased expression of PDE5A in muscle-invasive UC cells compared with the RT4 cell line. Furthermore, by using paired UTUC and normal tissues from 20 patients, lower PDE5A expression was also demonstrated in tumor specimens. CONCLUSIONS: Our findings suggest these candidate genes may play some roles in UTUC progression. We propose that these markers may be potential targets clarified by in vitro and in vivo experiments. PDE5A also potentially presents tumor suppressor genes, as identified by comparing the expression between normal and tumor specimens.
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spelling pubmed-80865702021-05-12 Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses Lee, Hsiang-Ying Li, Ching-Chia Li, Wei-Ming Hsu, Ya-Ling Yeh, Hsin-Chih Ke, Hung-Lung Yeh, Bi Wen Huang, Chun-Nung Li, Chien-Feng Kuo, Po-Lin Wu, Wen-Jeng PeerJ Bioinformatics BACKGROUND: We aimed to identify prognostic biomarkers of upper tract urothelial carcinomas (UTUCs), including microRNAs (miRNAs) and genes which account for only 5% to 10% of all urothelial carcinomas (UCs). In Taiwan, this figure is markedly higher, where it can reach up to 30% of UC cases. MATERIALS AND METHODS: Using next-generation sequencing (NGS), we analyzed two pairs of renal pelvis tumors and adjacent normal urothelial tissues to screen miRNAs and messenger RNAs. By combining bioinformatics analysis from miRmap, Gene Expression Omnibus (GEO), and Oncomine and Ingenuity(®) Pathway Analysis databases, we identified candidate genes. To search for upstream miRNAs with exact target binding sites, we used miRmap, TargetScan, and miRDB to enforce evidence. Then, we clarified gene and protein expression through an in vitro study using western blot analysis and quantitative real-time reverse transcriptase-PCR. RESULTS: Interactions between selected target genes obtained using the NGS and miRmap methods were assessed through a Venn diagram analysis. Six potential genes, namely, PDE5A, RECK, ZEB2, NCALD, PLCXD3 and CYBRD1 showed significant differences. Further analysis of gene expression from the GEO dataset indicated lower expression of PDE5A, RECK, ZEB2, and CYBRD1 in bladder cancer tissue than in normal bladder mucosa, which indicated that PDE5A, RECK, ZEB2, and CYBRD1 may act as tumor suppressors in UTUC. In addition, we compared the expression of these genes in various UC cell lines (RT4, BFTC905, J82, T24, UMUC3, 5637, BFTC 909, UMUC14) and found decreased expression of PDE5A in muscle-invasive UC cells compared with the RT4 cell line. Furthermore, by using paired UTUC and normal tissues from 20 patients, lower PDE5A expression was also demonstrated in tumor specimens. CONCLUSIONS: Our findings suggest these candidate genes may play some roles in UTUC progression. We propose that these markers may be potential targets clarified by in vitro and in vivo experiments. PDE5A also potentially presents tumor suppressor genes, as identified by comparing the expression between normal and tumor specimens. PeerJ Inc. 2021-04-27 /pmc/articles/PMC8086570/ /pubmed/33987019 http://dx.doi.org/10.7717/peerj.11343 Text en ©2021 Lee et al. https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.
spellingShingle Bioinformatics
Lee, Hsiang-Ying
Li, Ching-Chia
Li, Wei-Ming
Hsu, Ya-Ling
Yeh, Hsin-Chih
Ke, Hung-Lung
Yeh, Bi Wen
Huang, Chun-Nung
Li, Chien-Feng
Kuo, Po-Lin
Wu, Wen-Jeng
Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses
title Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses
title_full Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses
title_fullStr Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses
title_full_unstemmed Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses
title_short Identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses
title_sort identification of potential genes in upper tract urothelial carcinoma using next-generation sequencing with bioinformatics and in vitro analyses
topic Bioinformatics
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8086570/
https://www.ncbi.nlm.nih.gov/pubmed/33987019
http://dx.doi.org/10.7717/peerj.11343
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