Abstract
BackgroundClinical diagnostic research relies upon the collection of tissue samples, and for those samples to be representative of the in vivo situation. Tissue collection procedures, including post-operative ischemia, can impact the molecular profile of the tissue at the genetic and proteomic level. Understanding the influence of factors such as ischemia on tissue samples is imperative in order to develop both markers of tissue quality and ultimately accurate diagnostic tests.MethodsUsing NanoPro1000 technology, a rapid and highly sensitive immunoassay platform, the phosphorylation status of clinically relevant cancer-related biomarkers in response to ischemia was quantified in tissue samples from 20 patients with primary colorectal cancer. Tumor tissue and adjacent normal tissue samples were collected and subjected to cold ischemia prior to nanoproteomic analysis of AKT, ERK1/2, MEK1/2, and c-MET. Ischemia-induced relative changes in overall phosphorylation and phosphorylation of individual isoforms were calculated and statistical significance determined. Any differences in baseline levels of phosphorylation between tumor tissue and normal tissue were also analyzed.ResultsChanges in overall phosphorylation of the selected proteins in response to ischemia revealed minor variations in both normal and tumor tissue; however, significant changes were identified in the phosphorylation of individual isoforms. In normal tissue post-operative ischemia, phosphorylation was increased in two AKT isoforms, two ERK1/2 isoforms, and one MEK1/2 isoform and decreased in one MEK1/2 isoform and one c-MET isoform. Following ischemia in tumor tissue, one AKT isoform showed decreased phosphorylation and there was an overall increase in unphosphorylated ERK1/2, whereas an increase in the phosphorylation of two MEK1/2 isoforms was observed. There were no changes in c-MET phosphorylation in tumor tissue.ConclusionThis study provides insight into the influence of post-operative ischemia on tissue sample biology, which may inform the future development of markers of tissue quality and assist in the development of diagnostic tests.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-015-0752-1) contains supplementary material, which is available to authorized users.
Highlights
Clinical diagnostic research relies upon the collection of tissue samples, and for those samples to be representative of the in vivo situation
It is well known that the tissue collection procedure itself can have a significant impact on the analysis of data [3, 4]; understanding and controlling exogenous factors is crucial to the accurate interpretation of results [2, 3, 5]
Results revealed decreased phosphorylation of ERK1/2 in response to ischemia in normal as well as tumor tissue after 10 min of ischemia, similar to MEK1/2 in tumor tissue
Summary
Clinical diagnostic research relies upon the collection of tissue samples, and for those samples to be representative of the in vivo situation. Reliable development of diagnostic tools, identification of therapeutic targets, and especially accurate molecular stratification of patients all rely on the availability and appropriate collection of human tissue samples [1, 2]. Studies have described how post-surgical ischemia influences the molecular composition of tissues at the mRNA level [6,7,8], as well as at the proteome level [9,10,11]. These observations indicate that exogenous parameters can influence tissue samples in a way that may not reflect the real-life molecular profile of the tissue in vivo. An improved description of ischemiainduced changes in tissues may provide a better understanding of how clinically relevant markers have to be interpreted and how this will affect patient stratification, staging, prognosis, and selection of appropriate targeted treatment [14, 15]
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