The Effect of Nitric Oxide on Remote Ischemic Preconditioning in Renal Ischemia Reperfusion Injury in Rats

Abstract

Although remote ischemic preconditioning (RIPC) is an organ-protective maneuver from subsequent ischemia reperfusion injury (IRI) by application of brief ischemia and reperfusion to other organs, its mechanism remains unclear. However, it is known that RIPC reduces the heart, brain, and liver IRI, and that nitric oxide (NO) is involved in the mechanism of this effect. To identify the role of NO in the protective effect of RIPC in renal IRI, this study examined renal function, oxidative status, and histopathological changes using N-nitro-L-arginine methyl ester (L-NAME), an NO synthase inhibitor. Remote ischemic preconditioning was produced by 3 cycles of 5 minutes ischemia and 5 minutes reperfusion. Blood urea nitrogen, creatinine (Cr), and renal tissue malondialdehyde levels were lower, histopathological damage was less severe, and superoxide dismutase level was higher in the RIPC + IRI group than in the IRI group. The renoprotective effect was reversed by L-NAME. Obtained results suggest that RIPC before renal IRI contributes to improvement of renal function, increases antioxidative marker levels, and decreases oxidative stress marker levels and histopathological damage. Moreover, NO is likely to play an important role in this protective effect of RIPC on renal IRI.

Keywords: ischemia reperfusion injury; nitric oxide; remote ischemic preconditioning.

Figure 1.

Schematic overview of the study design and experimental groups. The number of rats per group is 6. IRI indicates ischemia reperfusion injury; L-NAME, N-nitro-L-arginine methyl ester; RIPC, remote ischemic preconditioning.

Figure 2.

Plasma BUN (A) and Cr (B) levels in the experimental groups. Plasma BUN and Cr levels were significantly lower in the RIPC + IRI group than in the IRI group. Compared with the RIPC + IRI group, the L-NAME + RIPC + IRI group showed significantly high plasma BUN and Cr levels. *P < .05 versus the sham group; ^† P < .05 versus the IRI group; ^§ P < .05 versus the RIPC + IRI group; ^‡ P < .05 versus the L-NAME + RIPC + IRI group. Data are expressed as mean ± SEM (n = 6 rats/group). BUN indicates blood urea nitrogen; Cr, creatinine; IRI, ischemia reperfusion injury; L-Name, N-nitro-l-arginine methyl ester; RIPC, remote ischemic preconditioning; SEM, standard error of the mean.

Figure 3.

Renal tissue MDA (A) and SOD (B) levels in the experimental groups. (A) the RIPC + IRI group showed lower MDA levels than the IRI group. The MDA levels in the L-NAME + RIPC + IRI group were higher than in the RIPC + IRI group. (B) Compared with the IRI group, the RIPC + IRI group showed significantly high SOD levels. *P < .05 and .001 for MDA and SOD, respectively, versus the sham group; ^† P < .05 for MDA and SOD, versus the IRI group; ^§ P < .05 for MDA, versus the RIPC + IRI group; P < .05 for MDA, versus the L-NAME + RIPC + IRI group. Data are expressed as mean ± SEM (n = 6 rats/group). MDA, malondialdehyde; SOD, superoxide dismutase; IRI, ischemia reperfusion injury; RIPC, remote ischemic preconditioning; l-name, N-nitro-l-arginine methyl ester; SEM, standard error of the mean.

Figure 4.

Histopathological photographs of renal tissue in the experimental groups. A, Periodic acid-Schiff (PAS) stain, original magnification × 100. (1) Sham group: no damage; (2) IRI group: destroyed tubules with flat epithelial cells lacking nuclear staining and lumen congestion; (3) RIPC + IRI group: mild damage with rounding of epithelial cells and dilated tubular lumen; and (4) L-NAME + RIPC + IRI group: severe damage with flattened epithelial cells, loss of nuclear staining, dilated lumen, and lumen congestion; (5) RIPC group: no damage. B, Damage score on the basis of the PAS staining. Renal tissue damage was lower in the RIPC + IRI group than in the IRI group. In the L-NAME + RIPC + IRI group, renal tissue damage was significantly higher than in the RIPC + IRI group. *P < .05 versus the sham group; ^† P <.05 versus the IRI group; ^§ P <.05 versus the RIPC + IRI group; ^‡ P < .05 versus the RIPC + IRI group. Data are expressed as mean ± SEM (n = 6 rats/group). IRI indicates ischemia reperfusion injury; L-NAME, N-nitro-L-arginine methyl ester; PAS, periodic acid-Schiff; RIPC, remote ischemic preconditioning; SEM, standard error of the mean.

Figure 5.

Hypothesis of RIPC-NO-mediated organ protection. eNOS indicates endothelial nitric oxide synthase; NO, nitric oxide; NO₂ ⁻, nitrite; O₂ ⁻, superoxide; RIPC, remote ischemic preconditioning; XO, xanthine oxidase.