Mokine (C-X-C motif) ligand209774_x_atCXCLchemokine (C-X-C motif) ligand225664_atCOL12Acollagen, type XII, alpha209395_atCHI3Lchitinase 3-like201195_s_atSLC7Asolute carrier family 7, member205828_atMMPmatrix metallopeptidaseBiomarkers for Dysplasia-Carcinoma TransitionFC = fold change. doi:10.1371/journal.pone.0048547.tBiomarkers for Dysplasia-Carcinoma TransitionFigure 1. Discriminatory power of the Itacitinib site classifier set with 11 transcripts ?Principal component analysis. A. Original sample set (53 samples, microarray) B. Independent sample set (94 samples, microarray) C. Independent sample set (68 samples, RT-PCR) D. GSE8671 (64 samples, microarray) E. GSE18105 (111 samples, microarray) a = adenoma, n = normal, crc = colorectal cancer. doi:10.1371/journal.pone.0048547.ghighlight refers to 6 and 11 adenoma samples which were above or near to the threshold. Green highlight refers to adenoma 10 samples which were clustered with CRC samples but ROC statistic shows clear separation from that group (Figure 3B, D). After patient follow the aforementioned samples transferred into CRC group and new multiple logistic regression was applied. Comparison of 9 high-grade dyslpalstic adenoma 12 and early cancer resulted 100 sensitivity and 100 specificity (Figure 3D), thereby optimize sensitivity (100 ) and specificity (90.9 ) of original sample classification (Figure 3B).DiscussionIn this study a characteristic transcript set was determined which is specific for the colorectal dysplasia-carcinoma transition using whole genomic microarray in 53 biopsy samples. In order to test the differentiation power of the discriminatory gene panel, an additional 94 microarrays with independent colonic biopsy specimen and microarray datasets downloaded from the Gene Expression Omnibus were also analyzed. With further validation conducted by array real-time PCR cards that contained the characteristic transcript panel. The identified set of 11 transcripts can be used for separation of CRC, adenoma and normal biopsy samples, moreover it is suitable for discrimination between highgrade dysplastic adenoma and early stage CRC cases by high specificity and sensitivity.The use of whole genomic microarray analyses represents an important tool for high-throughput gene expression screening, but equipment and reagent costs do not qualify it as for a cost effective diagnostic tool. Therefore quantitative array real-time PCR cards with assays for selected set of classifiers offer a more viable alternative for diagnostic application with lower costs and automation possibility for the whole process from RNA isolation to the RT-PCR analysis [22]. The current method of determining colorectal cancers and adenomas is histological analysis. Colon biopsy specimens are evaluated from 4? pieces of small sections of 3? mm thick taken from different areas of the colon. However critical areas may remain Naringin supplier hidden in the uncut specimen block or due to inadequate orientation including aberrant crypt foci in hyperplastic polyps, in situ carcinoma in adenomas, dysplastic areas and carcinomas in long-time IBD specimens [23?4]. In this study, whole biopsy specimens containing mixed cell populations were applied for mRNA expression microarray and real-time PCR analysis in order to overcome the potential sampling errors of conventional histological analysis. Though histological laser microdissection can provide accurate cell type specific information, its major limitation is the need of a very skilled oper.Mokine (C-X-C motif) ligand209774_x_atCXCLchemokine (C-X-C motif) ligand225664_atCOL12Acollagen, type XII, alpha209395_atCHI3Lchitinase 3-like201195_s_atSLC7Asolute carrier family 7, member205828_atMMPmatrix metallopeptidaseBiomarkers for Dysplasia-Carcinoma TransitionFC = fold change. doi:10.1371/journal.pone.0048547.tBiomarkers for Dysplasia-Carcinoma TransitionFigure 1. Discriminatory power of the classifier set with 11 transcripts ?Principal component analysis. A. Original sample set (53 samples, microarray) B. Independent sample set (94 samples, microarray) C. Independent sample set (68 samples, RT-PCR) D. GSE8671 (64 samples, microarray) E. GSE18105 (111 samples, microarray) a = adenoma, n = normal, crc = colorectal cancer. doi:10.1371/journal.pone.0048547.ghighlight refers to 6 and 11 adenoma samples which were above or near to the threshold. Green highlight refers to adenoma 10 samples which were clustered with CRC samples but ROC statistic shows clear separation from that group (Figure 3B, D). After patient follow the aforementioned samples transferred into CRC group and new multiple logistic regression was applied. Comparison of 9 high-grade dyslpalstic adenoma 12 and early cancer resulted 100 sensitivity and 100 specificity (Figure 3D), thereby optimize sensitivity (100 ) and specificity (90.9 ) of original sample classification (Figure 3B).DiscussionIn this study a characteristic transcript set was determined which is specific for the colorectal dysplasia-carcinoma transition using whole genomic microarray in 53 biopsy samples. In order to test the differentiation power of the discriminatory gene panel, an additional 94 microarrays with independent colonic biopsy specimen and microarray datasets downloaded from the Gene Expression Omnibus were also analyzed. With further validation conducted by array real-time PCR cards that contained the characteristic transcript panel. The identified set of 11 transcripts can be used for separation of CRC, adenoma and normal biopsy samples, moreover it is suitable for discrimination between highgrade dysplastic adenoma and early stage CRC cases by high specificity and sensitivity.The use of whole genomic microarray analyses represents an important tool for high-throughput gene expression screening, but equipment and reagent costs do not qualify it as for a cost effective diagnostic tool. Therefore quantitative array real-time PCR cards with assays for selected set of classifiers offer a more viable alternative for diagnostic application with lower costs and automation possibility for the whole process from RNA isolation to the RT-PCR analysis [22]. The current method of determining colorectal cancers and adenomas is histological analysis. Colon biopsy specimens are evaluated from 4? pieces of small sections of 3? mm thick taken from different areas of the colon. However critical areas may remain hidden in the uncut specimen block or due to inadequate orientation including aberrant crypt foci in hyperplastic polyps, in situ carcinoma in adenomas, dysplastic areas and carcinomas in long-time IBD specimens [23?4]. In this study, whole biopsy specimens containing mixed cell populations were applied for mRNA expression microarray and real-time PCR analysis in order to overcome the potential sampling errors of conventional histological analysis. Though histological laser microdissection can provide accurate cell type specific information, its major limitation is the need of a very skilled oper.
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