Ischemia caused by time to freezing induces systematic microRNA and mRNA responses in cancer tissue.
Author(s): Borgan E, Navon R, Vollan HK, Schlichting E, Sauer T, Yakhini Z, Lingjærde OC, Sørlie T, Børresen-Dale AL
Publication: Mol Oncol, 2011, Vol. 5, Page 564-76
PubMed ID: 21917534 PubMed Review Paper? No
Purpose of Paper
The purpose of this paper was to assess the impact of cold ischemia time on miRNA and mRNA expression profiles of breast tumor specimens via microarray analysis.
Conclusion of Paper
While the greatest source of variation in mRNA and miRNA data was observed between patients, significant effects of cold ischemia time on mRNA and miRNA transcript levels were observed. Of the 507 miRNA and 24712 mRNA transcripts evaluated on the microarrays, levels of 56 miRNA and 1788 mRNA transcripts were significantly affected by a cold ischemia time of up to 6 h at room temperature (FDR<0.05), which included increases in miRNA levels and both increases and decreases in mRNA levels. While all miRNAs represented on the array remained stable after a cold ischemia time of ≤1 h, 119 and 494 mRNA transcripts differed significantly from 0 h controls after 0.5 and 1 h of cold ischemia, respectively; of those differentially expressed mRNA transcripts, 65% and 92% exhibited lower levels after a cold ischemia time of 0.5 and 1 h, respectively. Based on gene ontology, several of the miRNA and mRNA pairs that were consistently found to be significantly positively or negatively correlated were identified as being associated with a response to stimulus, signal transduction, or stress response (FDR<0.05).
Studies
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Study Purpose
The purpose of this study was to assess the impact of cold ischemia time on miRNA and mRNA expression profiles of breast tumor specimens. Ten primary breast tumor specimens were macrodissected by a pathologist to ensure the primary tumor was represented in all aliquots. Specimens were then placed in a cryotube and left at room temperature for 0.5, 1, 3, or 6 h. A core needle biopsy representing a cold ischemia time of 0 h was obtained immediately after surgical resection from five patients and served as a control. All specimens were then snap-frozen in liquid nitrogen. RNA was extracted using the Trizol method and RNA concentration and quality were determined by Nanodrop spectrophotometry and a bioanalyzer, respectively. Expression profiles of mRNA and miRNA were assessed using the Agilent Technologies 4 x 44 K Whole Human Genome Microarray and the Human miRNA Microarray Kit V2, respectively. Feature Extraction files containing mRNA and miRNA microarray data were processed and normalized using R (v2.10.1) and Bioconductor, respectively, and all data was log2 transformed.
Summary of Findings:
While the greatest source of variation in mRNA and miRNA data was observed between patients, significant effects of cold ischemia time on mRNA and miRNA transcript levels were observed. Of the 507 miRNA and 24712 mRNA transcripts evaluated on the microarrays, levels of 56 miRNA and 1788 mRNA transcripts were significantly affected by a cold ischemia time of up to 6 h at room temperature (FDR<0.05). Of these affected transcripts, all miRNA and the majority of mRNA transcripts displayed increased levels with cold ischemia, although several mRNAs exhibited lower levels or a nonlinear response. The five most affected miRNAs included hsa-miR-1224-5p, hsa-miR-1225-5p, Kshv-miR-K12-3, hsa-miR-125a-3p, and hsa-miR371-5p. The ten most affected mRNA transcripts included THC2271582, GADD45B, LIMS3, BG950086, BC032118, FOSB, JUND, ZFP36, FEM1B, and DFNB31. While all miRNAs represented on the array remained stable after a cold ischemia time of ≤1 h, 119 and 494 mRNA transcripts differed significantly from 0 h controls after 0.5 and 1 h of cold ischemia, respectively; of those differentially expressed mRNA transcripts, 65% and 92% exhibited decreased levels after a cold ischemia time of 0.5 and 1 h, respectively, although most were classified as “unknown” by Agilent Technologies. The mRNA transcripts displaying higher levels after a cold ischemia time of ≤1 h included JUN, ZFP36, and CD86. Based on gene ontology, several of the miRNA and mRNA pairs that were consistently found to be significantly positively or negatively correlated were identified as being associated with a response to stimulus, signal transduction, or stress response.
(FDR<0.05).
Biospecimens
Preservative Types
- Frozen
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform RNA DNA microarray RNA Spectrophotometry RNA Automated electrophoresis/Bioanalyzer Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Acquisition Cold ischemia time 0 h
0.5 h
1 h
3 h
6 h