Analyzing miRNAs in ductal adenocarcinomas of the pancreas

TY - JOUR

T1 - Analyzing miRNAs in ductal adenocarcinomas of the pancreas

AU - Mees, Soeren Torge

AU - Schleicher, Christina

AU - Mardin, Wolf Arif

AU - Senninger, Norbert

AU - Colombo-Benkmann, Mario

AU - Haier, Joerg

N1 - Copyright © 2011 Elsevier Inc. All rights reserved.

PY - 2011/8

Y1 - 2011/8

N2 - BACKGROUND: MicroRNAs (miRNAs) have gained attention as an epigenetic component involved in the development of pancreatic ductal adenocarcinoma (PDAC). Several methods for miRNA profiling are in common use, but the validity of these methods is not defined. The aim of this study was to define the optimal method for miRNA detection in PDAC.METHODS: miRNA expression was determined using different and partially redundant methods (miRNA microarray, TaqMan low density array (TLDA), single tube quantitative RT-PCR). The data from different methods were statistically evaluated and tested for intermethodic consistency and reliability of the results. Finally, the miRNA expression status and the cell lines' ability to metastasize were correlated.RESULTS: Comparing low and high metastatic cells, miRNA-microarrays identified fewer differentially expressed and only upregulated miRNAs (n=27; 27 up-regulated) compared with TLDAs (n=54; 19 up- and 35 down-regulated). Evaluating miRNAs that target tumor suppressor genes, expression of all single tube quantitative real-time reverse transcriptase PCR (qRT-PCR) validated miRNAs was detected to be significantly altered in TLDA analysis (100%). MiRNA microarrays detected only 25% of qRT-PCR validated miRNAs. Furthermore, results from TLDA analysis correlated well with data from qRT-PCR and presented ΔΔCt values from 3.5±1.86 (range 0.8-5.62) compared with 3.74±1.86 (range 0.78-5.95) in qRT-PCR.CONCLUSION: Notable differences comparing data obtained from different screening methods were found. While TLDA and qRT-PCR correlated well in quantity and quality of the measured miRNAs, several tumor suppressor gene targeting and down-regulated miRNAs were not detected by miRNA-microarrays. This heterogeneity shows that care must be exercised when comparing results from different methods in PDAC.

AB - BACKGROUND: MicroRNAs (miRNAs) have gained attention as an epigenetic component involved in the development of pancreatic ductal adenocarcinoma (PDAC). Several methods for miRNA profiling are in common use, but the validity of these methods is not defined. The aim of this study was to define the optimal method for miRNA detection in PDAC.METHODS: miRNA expression was determined using different and partially redundant methods (miRNA microarray, TaqMan low density array (TLDA), single tube quantitative RT-PCR). The data from different methods were statistically evaluated and tested for intermethodic consistency and reliability of the results. Finally, the miRNA expression status and the cell lines' ability to metastasize were correlated.RESULTS: Comparing low and high metastatic cells, miRNA-microarrays identified fewer differentially expressed and only upregulated miRNAs (n=27; 27 up-regulated) compared with TLDAs (n=54; 19 up- and 35 down-regulated). Evaluating miRNAs that target tumor suppressor genes, expression of all single tube quantitative real-time reverse transcriptase PCR (qRT-PCR) validated miRNAs was detected to be significantly altered in TLDA analysis (100%). MiRNA microarrays detected only 25% of qRT-PCR validated miRNAs. Furthermore, results from TLDA analysis correlated well with data from qRT-PCR and presented ΔΔCt values from 3.5±1.86 (range 0.8-5.62) compared with 3.74±1.86 (range 0.78-5.95) in qRT-PCR.CONCLUSION: Notable differences comparing data obtained from different screening methods were found. While TLDA and qRT-PCR correlated well in quantity and quality of the measured miRNAs, several tumor suppressor gene targeting and down-regulated miRNAs were not detected by miRNA-microarrays. This heterogeneity shows that care must be exercised when comparing results from different methods in PDAC.

KW - Adenocarcinoma

KW - Carcinoma, Pancreatic Ductal

KW - Cell Line, Tumor

KW - Epigenesis, Genetic

KW - Gene Expression Profiling

KW - Humans

KW - MicroRNAs

KW - Microarray Analysis

KW - Pancreatic Neoplasms

KW - Reproducibility of Results

KW - Reverse Transcriptase Polymerase Chain Reaction

U2 - 10.1016/j.jss.2009.10.005

DO - 10.1016/j.jss.2009.10.005

M3 - SCORING: Journal article

C2 - 20080250

VL - 169

SP - 241

EP - 246

JO - J SURG RES

JF - J SURG RES

SN - 0022-4804

IS - 2

ER -