Impact of in Vivo Ischemic Time on RNA Quality—Experiences from a Colon Cancer Biobank

Abstract

Considerable effort has been made to improve differentiated diagnostics as well as personalized treatment for colorectal cancer patients. High-quality fresh frozen tissue is often required to investigate relevant molecular signatures in these patients. In RNA expression studies, the "RNA integrity number" is widely accepted as a reliable marker of RNA quality. Here, we investigate the feasibility of obtaining high-quality tissue from a colon cancer biobank and the impact of in vivo ischemic time and various technical and clinicopathological factors on RNA quality. Biopsies were obtained immediately following the tumor removal. The time from clamping the main arterial supply to resection and removal of the tumor was used to estimate the in vivo ischemic time. We did not observe a significant difference in RNA quality between normal tissue and tumor tissue. We observed a significant correlation between in vivo ischemic time and RNA quality in normal tissue (r = -0.24, p<0.001) but not in tumor tissue. Male gender and laparoscopic procedure were also significantly associated with lower RNA quality in normal tissue only. In tumor tissue, poor differentiation was associated with low RNA quality. In conclusion, in vivo ischemic time, surgical procedure, and gender have minor but significant effects on the quality of RNA from normal colon tissue but not tumor tissue. Poorly differentiated tumors are associated with lower RNA quality. Although its impact is low, it can still be considered to note in vivo ischemic time in colon cancer specimen procurement.

Impact of different stabilization methods on RT-qPCR results using human lung tissue samples

“…All pre-analytical parameters included in this range could alter the integrity of biomolecules contained in tissues and thus also introduce a bias in the results obtained. For example, it has been demonstrated that ischemia time (IT), both warm and cold, as one of the most influencing factors on RNA integrity [9][10][11] . However, no consensus has yet been reached to determine the exact moment in which RNA quality and integrity begin to decrease.…”

Colorectal cancer-associated SNP rs17042479 is involved in the regulation of NAF1 promoter activity

“…The patient material in the clinical data was compiled from the cancer Biobank at the University of Copenhagen [ 37 ]. The clinical data were collected between September 2006 and May 2012 from Roskilde University Hospital [ 17 ].…”

Effects of ex�vivo ischemia time and delayed processing on quality of specimens in tissue biobank

“…an agilent 2100 Bioanalyzer and an agilent rna 6000 nano kit (agilent Technologies, inc.) were used to determine rna quality, according to the manufacturers' protocol. rna integrity numbers (rins) were calculated using agilent Technologies 2100 Bioanalyzer expert software (version B.02.0x; agilent Technologies, inc.), and were presented as a value (1)(2)(3)(4)(5)(6)(7)(8)(9)(10) to indicate the degree of degradation.…”

“…Pre-analytical variables are divided into the pre-acquisition and post-acquisition phases. In the pre-acquisition phase, factors such as anesthesia ( 7 ), surgical trauma ( 8 , 9 ) and in vivo ischemia ( 10 ) might affect the cell integrity and gene expression profiles of the biospecimen, but they cannot be controlled very well because of the uncertainty of clinical treat requirements ( 11 ). Rather, factors such as ex vivo ischemia time, delayed processing time and processing temperature, which are crucial for bio-specimen quality ( 12 ), can be controlled and standardized effectively.…”