Hyperbaric oxygen treatment improves GFR in rats with ischaemia/reperfusion renal injury: a possible role for the antioxidant/oxidant balance in the ischaemic kidney

Abstract

Background: Ischaemic kidney injury continues to play a dominant role in the pathogenesis of acute renal failure (ARF) in many surgical and medical settings. A major event in the induction of renal injury is related to the generation of oxygen-free radicals. Hyperbaric oxygen therapy (HBO) is indicated for treatment of many ischaemic events but not for ARF. Therefore, the present study examined the effects of HBO on kidney function and renal haemodynamics in rats with ischaemic ARF.

Methods: Renal ischaemia was induced by unilateral renal artery clamping (45 min) in rats. Within 24 h following ischaemia, rats were treated twice with HBO of 100% O(2) at 2.5 absolute atmospheres for 90 min each (+HBO). Untreated rats (-HBO) served as a control. Forty-eight hours later, GFR, RBF and endothelial-dependent vasorelaxation were measured. In addition, the immunoreactive staining of 4-hydroxy-2-noneal (4-HNE), a major product of endogenous lipid peroxidation, and superoxide dismutase (SOD) were assessed.

Results: In the -HBO group, GFR was reduced by 94% compared with the untouched normal kidney (ischaemic: 0.06 +/- 0.03 ml/min, normal: 1.02 +/- 0.13 ml). In contrast, in the +HBO group, GFR of the ischaemic kidney (0.36 +/- 0.07 ml/min) was reduced only by 68% compared with the contralateral normal kidney (1.12 +/- 0.12 ml/min). In line with these findings, HBO improved the vasodilatory response to ACh as expressed in enhancement of both total and regional renal blood flow. In addition, HBO reduced the formation of 4-HNE by 33% and 76% and increased SOD by 30% and 70% in the cortex and outer stripe region of the medulla of the ischaemic kidney, respectively.

Conclusion: HBO attenuates the decline in GFR following renal ischaemia, and improves endothelial-dependent vasorelaxation, suggesting that treatment with HBO may be beneficial in the setting of ischaemic ARF.

Figures

Fig. 1

(A) Effect of HBO treatment on glomerular filtration rate (GFR) of control and ischaemic kidneys. (B) GFR ratio of ischaemic/normal kidney in the +HBO group and the −HBO group. n = 6–9 in each group. (A) shows that in the −HBO group, GFR was reduced by 94% compared with the untouched normal kidney. In contrast, in the +HBO group GFR of the ischaemic kidney was reduced only by 68% compared with the contralateral normal kidney. (B) demonstrates that the ratio of GFR of ischaemic/normal kidney is significantly higher in the +HBO group compared with the −HBO group.

Fig. 2

Baseline values of (A) mean arterial pressure (MAP), (B) renal blood flow (RBF) and renal vascular resistance (RVR), in the sham-operated control group and in the ischaemic kidney of −HBO and +HBO treated groups, 48 h after the induction of ischaemia. n = 6–9 in each group. Baseline values of MAP, RBF and calculated RVR in the sham-operated control group and in the ischaemic kidney of −HBO and +HBO groups are summarized in Figure 3. It is clear that baseline RBF decreased by 26% in +HBO rats versus −HBO rats. However, both were significantly lower compared with normal control kidney. Since MAP does not differ significantly in the three experimental groups, the calculated baseline RVR is higher (by 52%) in the ischaemic kidney of the +HBO group compared with the −HBO group.

Fig. 3

Effects of acetylcholine (ACh) administration (1, 10 and 100 μg/kg/min) on (A) mean arterial pressure (MAP), (B) renal blood flow (RBF) and (C) renal vascular resistance (RVR), in the sham-operated control group and in the ischaemic kidney of −HBO and +HBO groups 48 h after the induction of ischaemia. n = 6–9 in each group. *P < 0.05 versus sham. As shown, infusion of incremental doses of ACh produced a similar decrease in MAP in association with rise in RBF in response to ACh in all the groups, apparently as a result of an endothelial-dependent renal vasodilatation.

Fig. 4

Effects of acetylcholine (ACh) administration (10 μg/kg/min) on (A) cortical blood flow (CBF) and (B) medullary blood flow (MBF) in the sham-operated control group and in the ischaemic kidney of −HBO and +HBO groups 48 h after the induction of ischaemia. n = 6–9 in each group. It is evident that administration of ACh to control rats caused a gradual increase in CBF, whereas MBF did not change significantly. In contrast, in the ischaemic kidney of the −HBO group, ACh elicited an attenuated increase in CBF. The cortical vasodilatory response to ACh significantly improved following HBO treatment. These findings may explain the observed stimulatory effects of HBO on total RBF, and suggest that treatment with HBO may improve endothelial-dependent vasorelaxation mediated by eNOS. P < 0.05 vs. −HBO.

Fig. 5

Representative pictures of 4-HNE staining in the medulla of −HBO (A) and +HBO (B) ischaemic kidneys (objective ×20). (C) Percentage staining area of 4HNE in the renal outer strip of the outer of −HBO and +HBO of control and ischaemic kidneys. n = 6–9 in each group. This figure indicates that 4-HNE is localized primarily in the renal tubular cells of the outer stripe of the medulla of the ischaemic kidneys, and is almost undetectable in the cortex. The area of the outer stripe of the medulla stained for 4-HNE is significantly less than in the ischaemic kidneys of rats treated with hyperbaric oxygen than in untreated rats.

Fig. 6

Representative pictures of CuZn-SOD staining in the medulla of −HBO (A) and +HBO (B) ischaemic kidneys (objective × 20). (C) Percentage staining area of CuZn-SOD in the renal medulla of −HBO and +HBO control and ischaemic kidney. n = 6–9 in each group. It is evident that immunostaining for Cu/Zn-SOD in the cortex and the outer stripe of the medulla of untouched kidneys is negligible. A marked increase in Cu/Zn-SOD immunoreactive staining in the outer stripe of the medulla of the ischaemic kidneys in both −HBO and +HBO groups, compared with the control non-ischaemic kidneys, is notable. However, in the +HBO group, the percentage of staining for Cu/Zn-SOD is significantly higher than that in the −HBO group. Thus, HBO treatment induces an increase in the immunohistochemical staining of the antioxidant Cu/Zn-SOD in the outer stripe of the medulla of the ischaemic kidneys.