title: Impact of ischemia and procurement conditions on gene expression in renal cell carcinoma.
abstract: OBJECTIVE
  Previous studies have shown that ischemia alters gene expression in normal and malignant tissues. There are no studies that evaluated effects of ischemia in renal tumors. This study examines the impact of ischemia and tissue procurement conditions on RNA integrity and gene expression in renal cell carcinoma.
  
  
  METHODS
  Ten renal tumors were resected without renal hilar clamping from 10 patients with renal clear cell carcinoma. Immediately after tumor resection, a piece of tumor was snap frozen. Remaining tumor samples were stored at 4°C, 22°C, and 37°C and frozen at 5, 30, 60, 120, and 240 minutes. Histopathologic evaluation was conducted on all tissue samples, and only those with greater than 80% tumor were selected for further analysis. RNA integrity was confirmed by electropherograms and quantitated using RNA integrity number index. Altered gene expression was assessed by paired, two-sample t test between the zero time point and aliquots from various conditions obtained from the same tumor.
  
  
  RESULTS
  One hundred and forty microarrays were conducted. Some RNA degradation was observed 240 minutes after resection at 37°C. The expression of more than 4,000 genes was significantly altered by ischemia times or storage conditions. The greatest gene expression changes were observed with longer ischemia time and warmer tissue procurement conditions.
  
  
  CONCLUSIONS
  RNA from kidney cancer remains intact for up to 4 hours post surgical resection regardless of storage conditions. Despite excellent RNA preservation, time after resection and procurement conditions significantly influence gene expression profiles. Meticulous attention to preacquisition variables is of paramount importance for accurate tumor profiling.
authors.affiliation: Urologic Oncology Branch, National Cancer Institute, NIH, Bethesda, MD, USA.
authors.forename: Nick W
authors.initials: NW
authors.lastname: Liu
authors.forename: Thomas
authors.initials: T
authors.lastname: Sanford
authors.forename: Ramaprasad
authors.initials: R
authors.lastname: Srinivasan
authors.forename: Jack L
authors.initials: JL
authors.lastname: Liu
authors.forename: Kiranpreet
authors.initials: K
authors.lastname: Khurana
authors.forename: Olga
authors.initials: O
authors.lastname: Aprelikova
authors.forename: Vladimir
authors.initials: V
authors.lastname: Valero
authors.forename: Charles
authors.initials: C
authors.lastname: Bechert
authors.forename: Robert
authors.initials: R
authors.lastname: Worrell
authors.forename: Peter A
authors.initials: PA
authors.lastname: Pinto
authors.forename: Youfeng
authors.initials: Y
authors.lastname: Yang
authors.forename: Maria
authors.initials: M
authors.lastname: Merino
authors.forename: W Marston
authors.initials: WM
authors.lastname: Linehan
authors.forename: Gennady
authors.initials: G
authors.lastname: Bratslavsky
mesh_terms: D000328:Adult; D000368:Aged; D002292:Carcinoma, Renal Cell / Q000098:blood supply / Q000235:genetics* / Q000473:pathology; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007511:Ischemia; D007680:Kidney Neoplasms / Q000098:blood supply / Q000235:genetics* / Q000473:pathology; D008875:Middle Aged; D009367:Neoplasm Staging; D015203:Reproducibility of Results; D013048:Specimen Handling; D013696:Temperature; D013997:Time Factors