Article Text

PDF
327 FENOLDOPAM PROTECTS HUMAN RENAL TUBULAR CELLS AGAINST COLD-STORAGE INJURY THROUGH AN HO-1 MECHANISM: STUDIES UTILIZING HO-1 siRNA
  1. H. Huang1,
  2. D. E. Stec1,
  3. A. K. Salahudeen1
  1. 1Department of Medicine and Physiology, University of Mississippi Medical Center, Jackson

Abstract

We have demonstrated that infusion of donor rats with fenoldopam, a dopaminergic compound that maintains renal blood flow without increasing blood pressure, induces HO-1 expression and protects kidneys against cold-storage injury when transplanted. Since fenoldopam has vascular and other cellular effects, whether the protection is mediated through HO-1 remains uncertain. To test this, renal tubular (RPTE) cells were preincubated with fenoldopam (0-80 μg/mL) for 18 hours and subjected to 4°C cold storage for 24 hours. Fenoldopam significantly reduced cold-storage injury (e.g., % LDH release, mean ± SE, 53.4 ± 1.3 cold control vs. 16.2 ± 0.5 and 2.2 ± 0.2 for 10 and 80 μg/mL fenoldopam respectively (n = 3; p < .001). Similarly, inclusion of fenoldopam in UW solution during cold storage also protected the cells against cold-storage injury. The fenoldopam's protections of cells on both of these occasions were associated with a significant increase in HO-1 protein. To test whether HO-1 induction is directly responsible for fenoldopam's cytoprotection in RPTE cells, we employed siRNA oligonucleotides specifically directed against HO-1 mRNA to silence the HO-1 gene. Using real-time RT PCR, we first confirmed that fenoldopam caused a two-fold induction of HO-1 mRNA. In a separate experiment in which RPTE cells were stored in the cold, siRNA dose dependently knocked down the fenoldopam-induced HO-1 mRNA by 70%. The controls, oligofectamine, the vehicle for siRNA, or siRNA alone did not affect HO-1. To determine the effect of HO-1 on fenoldopam's protection, we repeated the experiment in the presence of HO-1 siRNA and measured LDH release. As before, fenoldopam significantly suppressed cold-induced LDH release in these cells. However, the addition of HO-1 siRNA completely abrogated the protective effect of fenoldopam (% LDH release: cold control 50.2 ± 0.4 vs. cold plus fenoldopam 10.1 ± 0.5, vs. cold plus fenoldopam 40 μg/mL plus HO-1 siRNA 52.2 ± 0.3; p < .001) demonstrating a central role for HO-1 in fenoldopam's cytoprotection. Use of fenoldopam may prove to be useful in reducing the cold-storage injury of transplanted organs.

Statistics from Altmetric.com

Request Permissions

If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.