ABSTRACT
Title
Tissue iron accumulation is strictly involved in pathological events occurring in SHRSP.
Authors
P. Gelosa 1, A. Pignieri 1, U. Guerrini 1, E. Tremoli 1,2, L. Sironi 1,2
1 Department of Pharmacological Sciences, University of Milan, Milan, Italy;
2 Centro Cardiologico Monzino, IRCCS, Milan, Italy.
1 Department of Pharmacological Sciences, University of Milan, Milan, Italy;
2 Centro Cardiologico Monzino, IRCCS, Milan, Italy.
Abstract
Background and aim The stroke-prone spontaneously hypertensive rat (SHRSP) is a widely used model of hypertensive cerebrovascular disease. The occurrence of brain damage is preceded by the development of proteinuria and systemic inflammation. Our recent data show renal iron accumulation before the appearance of brain damage, suggesting altered iron homeostasis. These results agree with previous data reporting on serum haptoglobin depletion (Sironi et al., 2001), decreased hematocrit, and elevated levels of haemoglobin in the urine of SHRSP (Smeda JS, 1997). The aim of this study was to characterize, by magnetic resonance imaging (MRI) gradient-echo (T2*), the iron accumulation in SHRSP and to investigate the role of iron in the pathological events occurring in SHRSP, by pharmacological approaches.
Methods and Results MaleSHRSP, fed a high-salt diet, were treated long-term i.p. with vehicle (n=12) or deferoxamine, an iron chelator, (n=12; 200 mg/kg/day); a control group of SHRSPs (n=4) fed a standard diet.
Once a week, whole blood was taken from each rat to determine hematocrit, the morphology of RBCs (red blood cells), and the number of reticulocytes. All rats were then housed individually in metabolic cages for 24 h to collect urine and determine 24-h urine protein.
The SHRSP underwent cerebral and renal MRI every week until 24-h proteinuria exceeded 40 mg/day; thereafter, MRI was performed every other day until brain damage was observed.
We observed a loss on T2* MRI signal in the renal cortex of vehicle-treated SHRSP at the appearance of proteinuria, as opposed to control SHR-SP and deferoxamine-treated rats. The T2* signal value decreased with the progression of renal damage and was related to iron oxide accumulation as localized in macrophages and in tubular epithelial cells.
The vehicle-treated rats developed brain abnormalities, as detected by MRI, after 32.8± 5.6 days; deferoxamine delayed the time of brain damage occurrence and enhancedthe survival time (p<0.01).
In addition, deferoxamine delayed the development of proteinuria and preserved renal structure, by preventing inflammatory cells infiltrations and superoxide production, accumulation of iron, extracellular matrix, vimentin, and heme oxygenase-1.
Vehicle-treated SHRSP also exhibited renal mitochondrial dysfunction, as evaluated by cytochromec oxidase activity, which was preserved by deferoxamine treatment.
Finally, as opposed to deferoxamine-treated rats, we observed a decrease of hematocrit in vehicle-treated SHRSP, together with a high number of schistocytes and reticolocyte in the blood and a depletion of haptoglobin in the serum.
ConclusionThis study shows for the first time an endogenous iron deposition in the kidney of SHRSP that is strictly involved in the end-organ damage. Iron deposition could have harmful consequences for the kidney by increasing oxidative stress and toxic free radical production, generated by the metabolism of hydrogen peroxide.
This excess of iron seems to originate from ruptured red blood cells that may be induced by systemic oxidative stress occurring in salt-loaded SHRSP. Enhanced intravascular hemolysis might lead to depletion of haptoglobin and accumulation of iron related compounds that are implicated in several toxic processes.
Treatment with deferoxamine delayed these pathological events and prevented loss of T2* MRI signals, iron deposition in the kidney, and renal damage.
These data suggest new directions for the pharmacological treatment of hypertensive renal diseases, in which a modulation of oxidative metabolism is warranted. Exploration of the relationship among iron deposition, oxidative stress, inflammation, and renal injury requires further studies.
- Sironi et al. Acute-phase proteins before cerebral ischemia in stroke-prone rats: identification by proteomics. Stroke. 2001;32(3):753-60.
- Smeda JS. Renal function in stroke-prone rats fed a high-K+ diet. Can J Physiol Pharmacol. 1997;75(7):796-806.
Methods and Results MaleSHRSP, fed a high-salt diet, were treated long-term i.p. with vehicle (n=12) or deferoxamine, an iron chelator, (n=12; 200 mg/kg/day); a control group of SHRSPs (n=4) fed a standard diet.
Once a week, whole blood was taken from each rat to determine hematocrit, the morphology of RBCs (red blood cells), and the number of reticulocytes. All rats were then housed individually in metabolic cages for 24 h to collect urine and determine 24-h urine protein.
The SHRSP underwent cerebral and renal MRI every week until 24-h proteinuria exceeded 40 mg/day; thereafter, MRI was performed every other day until brain damage was observed.
We observed a loss on T2* MRI signal in the renal cortex of vehicle-treated SHRSP at the appearance of proteinuria, as opposed to control SHR-SP and deferoxamine-treated rats. The T2* signal value decreased with the progression of renal damage and was related to iron oxide accumulation as localized in macrophages and in tubular epithelial cells.
The vehicle-treated rats developed brain abnormalities, as detected by MRI, after 32.8± 5.6 days; deferoxamine delayed the time of brain damage occurrence and enhancedthe survival time (p<0.01).
In addition, deferoxamine delayed the development of proteinuria and preserved renal structure, by preventing inflammatory cells infiltrations and superoxide production, accumulation of iron, extracellular matrix, vimentin, and heme oxygenase-1.
Vehicle-treated SHRSP also exhibited renal mitochondrial dysfunction, as evaluated by cytochromec oxidase activity, which was preserved by deferoxamine treatment.
Finally, as opposed to deferoxamine-treated rats, we observed a decrease of hematocrit in vehicle-treated SHRSP, together with a high number of schistocytes and reticolocyte in the blood and a depletion of haptoglobin in the serum.
ConclusionThis study shows for the first time an endogenous iron deposition in the kidney of SHRSP that is strictly involved in the end-organ damage. Iron deposition could have harmful consequences for the kidney by increasing oxidative stress and toxic free radical production, generated by the metabolism of hydrogen peroxide.
This excess of iron seems to originate from ruptured red blood cells that may be induced by systemic oxidative stress occurring in salt-loaded SHRSP. Enhanced intravascular hemolysis might lead to depletion of haptoglobin and accumulation of iron related compounds that are implicated in several toxic processes.
Treatment with deferoxamine delayed these pathological events and prevented loss of T2* MRI signals, iron deposition in the kidney, and renal damage.
These data suggest new directions for the pharmacological treatment of hypertensive renal diseases, in which a modulation of oxidative metabolism is warranted. Exploration of the relationship among iron deposition, oxidative stress, inflammation, and renal injury requires further studies.
- Sironi et al. Acute-phase proteins before cerebral ischemia in stroke-prone rats: identification by proteomics. Stroke. 2001;32(3):753-60.
- Smeda JS. Renal function in stroke-prone rats fed a high-K+ diet. Can J Physiol Pharmacol. 1997;75(7):796-806.