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Identification and Validation of a Potential Marker of Tissue Quality Using Gene Expression Analysis of Human Colorectal Tissue.

Lange N, Unger FT, Schöppler M, Pursche K, Juhl H, David KA - PLoS ONE (2015)

Bottom Line: Tissue quality can be measured by changes in levels of gene expression and can be influenced by many factors including pre-analytical conditions, ischemic effects and the surgical collection procedure itself.Both microarray analysis and qPCR revealed regulator of G-protein signaling 1 (RGS1) as a potential marker of CRC tissue quality and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) as a potential reference gene of post-operative tissue quality.Larger studies with additional time points and endpoints will be needed to confirm these results.

View Article: PubMed Central - PubMed

Affiliation: Indivumed GmbH, Hamburg, Germany.

ABSTRACT
Correlative studies have identified numerous biomarkers that are individualizing therapy across many medical specialties, including oncology. Accurate interpretation of these studies requires the collection of tissue samples of sufficient quality. Tissue quality can be measured by changes in levels of gene expression and can be influenced by many factors including pre-analytical conditions, ischemic effects and the surgical collection procedure itself. However, as yet there are no reliable biomarkers of tissue quality at researchers' disposal. The aim of the current study was to identify genes with expression patterns that fluctuated reproducibly in response to typical post-surgical stress (ischemia) in order to identify a specific marker of tissue quality. All tissue samples were obtained from patients with primary colorectal carcinoma (CRC) (N = 40) either via colonoscopy prior to surgery, or by surgical resection. Surgically resected tissue samples were divided into three groups and subjected to cold ischemia for 10, 20 or 45 minutes. Normal colorectal tissue and CRC tissue was analyzed using microarray and quantitative real-time PCR (qPCR). Comparing changes in gene expression between pre- and post-surgical tissue using microarray analysis identified a list of potential tissue quality biomarkers and this list was validated using qPCR. Results revealed that post-operative ischemia significantly alters gene expression in normal and CRC tissue samples. Both microarray analysis and qPCR revealed regulator of G-protein signaling 1 (RGS1) as a potential marker of CRC tissue quality and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) as a potential reference gene of post-operative tissue quality. Larger studies with additional time points and endpoints will be needed to confirm these results.

No MeSH data available.


Related in: MedlinePlus

Expression levels of RGS1 following normalization to UBC, GAPDH and EEF1A1 as well as to the combination of GAPDH and UBC.Comparison of RGS1 gene expression changes in normal (A) and tumor (B) ischemic tissue samples of 20 individual patients. Patients pre-surgery samples (0) were set to 1 and individual fold changes of ischemic samples were calculated based on normalization to UBC (white box), GAPDH (light grey box), EEF1A1 (grey box) or to a combination of GAPDH and UBC (dark grey box). Results were summarized by Box-Whisker Plots. Boxes represent first and third quartile, whiskers represent minima and maxima, solid lines within boxes indicate medians. Kruskal-Wallis test and Dunn’s multiple comparison test were used for statistical analysis to compare the four normalization approaches within each time point. n.s. = not significant; N = normal tissue; T = tumor tissue; 0 = before surgery; 10, 20, 45 = 10, 20, 45 minutes after resection.
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pone.0133987.g004: Expression levels of RGS1 following normalization to UBC, GAPDH and EEF1A1 as well as to the combination of GAPDH and UBC.Comparison of RGS1 gene expression changes in normal (A) and tumor (B) ischemic tissue samples of 20 individual patients. Patients pre-surgery samples (0) were set to 1 and individual fold changes of ischemic samples were calculated based on normalization to UBC (white box), GAPDH (light grey box), EEF1A1 (grey box) or to a combination of GAPDH and UBC (dark grey box). Results were summarized by Box-Whisker Plots. Boxes represent first and third quartile, whiskers represent minima and maxima, solid lines within boxes indicate medians. Kruskal-Wallis test and Dunn’s multiple comparison test were used for statistical analysis to compare the four normalization approaches within each time point. n.s. = not significant; N = normal tissue; T = tumor tissue; 0 = before surgery; 10, 20, 45 = 10, 20, 45 minutes after resection.

Mentions: Next, all RGS1 data were normalized to UBC, GAPDH or EEF1A1. RGS1 expression level changes were compared with RGS1 expression levels following normalization to the combination of GAPDH and UBC (Fig 4). Results revealed that the same RGS1 expression levels were generated regardless of the reference gene used. Furthermore, statistical analysis revealed no significant differences in RGS1 expression when comparing the four normalization approaches within each time point. These data support the assumption that EEF1A1 might act as a potential reference gene of post-resectional tissue quality.


Identification and Validation of a Potential Marker of Tissue Quality Using Gene Expression Analysis of Human Colorectal Tissue.

Lange N, Unger FT, Schöppler M, Pursche K, Juhl H, David KA - PLoS ONE (2015)

Expression levels of RGS1 following normalization to UBC, GAPDH and EEF1A1 as well as to the combination of GAPDH and UBC.Comparison of RGS1 gene expression changes in normal (A) and tumor (B) ischemic tissue samples of 20 individual patients. Patients pre-surgery samples (0) were set to 1 and individual fold changes of ischemic samples were calculated based on normalization to UBC (white box), GAPDH (light grey box), EEF1A1 (grey box) or to a combination of GAPDH and UBC (dark grey box). Results were summarized by Box-Whisker Plots. Boxes represent first and third quartile, whiskers represent minima and maxima, solid lines within boxes indicate medians. Kruskal-Wallis test and Dunn’s multiple comparison test were used for statistical analysis to compare the four normalization approaches within each time point. n.s. = not significant; N = normal tissue; T = tumor tissue; 0 = before surgery; 10, 20, 45 = 10, 20, 45 minutes after resection.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4519187&req=5

pone.0133987.g004: Expression levels of RGS1 following normalization to UBC, GAPDH and EEF1A1 as well as to the combination of GAPDH and UBC.Comparison of RGS1 gene expression changes in normal (A) and tumor (B) ischemic tissue samples of 20 individual patients. Patients pre-surgery samples (0) were set to 1 and individual fold changes of ischemic samples were calculated based on normalization to UBC (white box), GAPDH (light grey box), EEF1A1 (grey box) or to a combination of GAPDH and UBC (dark grey box). Results were summarized by Box-Whisker Plots. Boxes represent first and third quartile, whiskers represent minima and maxima, solid lines within boxes indicate medians. Kruskal-Wallis test and Dunn’s multiple comparison test were used for statistical analysis to compare the four normalization approaches within each time point. n.s. = not significant; N = normal tissue; T = tumor tissue; 0 = before surgery; 10, 20, 45 = 10, 20, 45 minutes after resection.
Mentions: Next, all RGS1 data were normalized to UBC, GAPDH or EEF1A1. RGS1 expression level changes were compared with RGS1 expression levels following normalization to the combination of GAPDH and UBC (Fig 4). Results revealed that the same RGS1 expression levels were generated regardless of the reference gene used. Furthermore, statistical analysis revealed no significant differences in RGS1 expression when comparing the four normalization approaches within each time point. These data support the assumption that EEF1A1 might act as a potential reference gene of post-resectional tissue quality.

Bottom Line: Tissue quality can be measured by changes in levels of gene expression and can be influenced by many factors including pre-analytical conditions, ischemic effects and the surgical collection procedure itself.Both microarray analysis and qPCR revealed regulator of G-protein signaling 1 (RGS1) as a potential marker of CRC tissue quality and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) as a potential reference gene of post-operative tissue quality.Larger studies with additional time points and endpoints will be needed to confirm these results.

View Article: PubMed Central - PubMed

Affiliation: Indivumed GmbH, Hamburg, Germany.

ABSTRACT
Correlative studies have identified numerous biomarkers that are individualizing therapy across many medical specialties, including oncology. Accurate interpretation of these studies requires the collection of tissue samples of sufficient quality. Tissue quality can be measured by changes in levels of gene expression and can be influenced by many factors including pre-analytical conditions, ischemic effects and the surgical collection procedure itself. However, as yet there are no reliable biomarkers of tissue quality at researchers' disposal. The aim of the current study was to identify genes with expression patterns that fluctuated reproducibly in response to typical post-surgical stress (ischemia) in order to identify a specific marker of tissue quality. All tissue samples were obtained from patients with primary colorectal carcinoma (CRC) (N = 40) either via colonoscopy prior to surgery, or by surgical resection. Surgically resected tissue samples were divided into three groups and subjected to cold ischemia for 10, 20 or 45 minutes. Normal colorectal tissue and CRC tissue was analyzed using microarray and quantitative real-time PCR (qPCR). Comparing changes in gene expression between pre- and post-surgical tissue using microarray analysis identified a list of potential tissue quality biomarkers and this list was validated using qPCR. Results revealed that post-operative ischemia significantly alters gene expression in normal and CRC tissue samples. Both microarray analysis and qPCR revealed regulator of G-protein signaling 1 (RGS1) as a potential marker of CRC tissue quality and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) as a potential reference gene of post-operative tissue quality. Larger studies with additional time points and endpoints will be needed to confirm these results.

No MeSH data available.


Related in: MedlinePlus