Nitric oxide synthase isoforms and glomerular hyperfiltration in early diabetic nephropathy

This study tested the hypothesis that nitric oxide (NO)-mediated renal vasodilation due to the activity of the inducible nitric oxide synthase (iNOS) contributes to glomerular hyperfiltration in diabetic rats. Two weeks after induction of diabetes mellitus by streptozotocin, mean arterial BP (MAP), GFR (inulin clearance), and renal plasma flow (RPF) (para-aminohippurate clearance) were measured in conscious instrumented rats. Diabetic rats had elevated GFR (3129 +/- 309 microl/min versus 2297 +/- 264 microl/min in untreated control rats, P < 0.05) and RPF (10526 +/- 679 microl/min versus 8005 +/- 534 microl/min), which was prevented by chronic insulin treatment. Intravenous administration of 0.1 and 1 mg of L-imino-ethyl-lysine (L-NIL), an inhibitor of iNOS, did not affect MAP, GFR, or RPF, either in diabetic or control rats. A higher L-NIL dose (10 mg) increased MAP and decreased RPF in diabetic rats significantly (n = 6, P < 0.05), but not in controls (n = 6). In addition, 0.1 mg of NG-nitro-L-arginine methyl ester (L-NAME), a nonselective blocker of NOS isoforms, decreased GFR (2389 +/- 478 microl/min) and RPF (7691 +/- 402 microl/min) in diabetic animals to control levels, while renal hemodynamics in normoglycemic rats were not altered. Higher L-NAME doses (1 and 10 mg) reduced GFR and RPF in diabetic and control rats to identical levels. In glomeruli isolated from diabetic and control rats, neither iNOS mRNA nor iNOS protein expression was detected. In contrast, increased protein levels of endothelial constitutive NOS (ecNOS) were found in glomeruli of diabetic rats compared with controls. By immunohistochemistry, ecNOS but not iNOS staining was observed in the endothelium of preglomerular vessels and in diabetic glomeruli. These results support the notion that increased NO availability due to greater abundance of ecNOS contributes to the pathogenesis of glomerular hyperfiltration in early experimental diabetic nephropathy. In contrast, we found no functional or molecular evidence for increased glomerular expression and activity of iNOS in diabetic rats.     

Endothelial-derived vasoactive mediators in polycystic kidney disease

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is characterized by hypertension and renal vasoconstriction. Mediators of these hemodynamic changes are not well understood, but evidence suggests that endothelial-derived mediators may participate. METHODS: Baseline measurements of blood pressure, proteinuria, and urinary nitrite/nitrate excretion were performed in control and cystic male Han:SPRD rats (6 weeks of age). They were then treated with the nitric oxide (NO), nitric oxide synthase (NOS) inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), or vehicle, for 6 weeks. After repeat systemic measurements, renal function was determined using inulin and para-aminohippurate (PAH) clearances. Levels of renal endothelin-1 (ET-1) and renal endothelial NOS (eNOS) proteins were determined, and immunohistochemistry localized renal eNOS and neuronal NOS (nNOS). RESULTS: Administration of L-NAME aggravated systemic hypertension and renal vasoconstriction in the cystic rats, but did not affect the progression of proteinuria or cystic expansion. Cystic rats demonstrated marked increases in renal ET-1 and eNOS levels. L-NAME reduced eNOS expression in the membrane compartment, but increased eNOS in the cytosol. Localization studies indicated that renal eNOS was abundant in nonvascular compartments, but not in renal vascular and glomerular structures, whereas renal nNOS was diffusely diminished. CONCLUSION: These alterations of endothelial-derived mediators (up-regulation of ET-1, and dysfunction of the NO system) contribute to vasoconstriction, and thereby are likely to contribute to the progressive loss of renal function in polycystic kidney disease (PKD).     

Visualization of renal autoregulation in the split hydronephrotic kidney of rats

The autoregulatory ability of the renal vascular system in the split hydronephrotic kidney was quantitated with intravital microscopy. The luminal diameters of the arcuate and interlobular arteries as well as the afferent and efferent arterioles were measured. Glomerular blood flow was determined by the dual slit technique. In the first series of experiments, the renal perfusion pressure was reduced by graded clamping of the abdominal aorta. Pressure reduction from 118 mm Hg to 95 mm Hg induced dilation of all preglomerular vessels except for the distal afferent arteriole; there was no change in the efferent arteriole and the blood flow was maintained. Further pressure reductions to 71 and 43 mm Hg caused additional dilations of the preglomerular vessels, a marginal reduction in diameter of proximal efferent arterioles and flow reductions by 15% and 41%, respectively. In the second series, systemic blood pressure was increased by continuous i.v. infusions of norepinephrine (NE). NE constricted pre- and postglomerular vessels except for the distal afferent arteriole; glomerular flow was decreased. Reduction of renal perfusion pressure during NE infusion to the preinfusion value did not diminish glomerular blood flow, but reduced the constrictor response to NE in preglomerular vessels. In a third series of experiments we examined the effect of atrial natriuretic factor (ANF) on renal autoregulation. Addition of ANF (10(-9) to 10(-7) M) to the renal bath induced a dose-dependent dilation of all preglomerular vessels and a constriction of the efferent arteriole. Pressure reduction from 120 to 95 mm Hg resulted in a further preglomerular vasodilation. These experiments demonstrate that autoregulation is mediated primarily by diameter changes in all preglomerular vessels excluding the distal segment of the afferent arteriole. Further, these data suggest that ANF induced dilation of preglomerular vessels does not impair the myogenic response of these vessels.     

Renal sodium handling in experimental diabetes: role of NO

Recent studies have suggested that diabetes is a state of increasedrenal nitric oxide (NO) activity as assessed by urinary excretionof nitrites and nitrates (NO_x), and that NO synthase inhibitorsreverse the increased glomerular filtration rate (GFR) observedin experimental diabetes. In addition to being a potent vasodilatorin the renal vasculature, NO also plays a role in modulationof renal sodium excretion. To explore the role of NO in diabetes-associatedalterations in renal excretory function, renal haemodynamicand sodium handling parameters were evaluated in conscious control(C) and streptozotocin diabetic rats (D) and correlated to therenal activity of NO, as assessed by urinary excretion of itsmetabolites NO_x. To further explore this issue, the changesin renal haemodynamics and sodium handling were also assessedafter NO synthase inhibition with a non-pressor dose of L-nitro-arginine-methyl-ester(L-NAME) and after administration of the NO donor, glyceryltrinitrate (GTN). Systolic blood pressure was not differentbetween C and D rats. D rats exhibited marked hyperglycaemia(P<0.001), and increases in GFR (P<0.001), renal plasmaflow, filtration fraction, urinary sodium excretion (U_NaV, P<0.001),filtered load of sodium (FL_Na, P<0.01), and a decrease infractional reabsorption of sodium (FR_Na, P<0.0001). In contrast,total reabsorption of sodium (TR_Na) was increased in D ratscompared to C rats (P<0.0001). The urinary excretion of NOwas markedly increased in D rats (P<0.01). Regression analysesperformed in D rats revealed a close relationship between U_NaVand GFR and a weaker correlation with urinary NO_x. AlthoughFR_Na correlated only with urinary excretion of NO_x, there wasa strong relationship between TR_Na and GFR. In contrast to Drats, control rats demonstrated only a relationship betweenTR_Na and GFR and no other correlations were found, in D rats,NO inhibition with L-NAME (1 mg/kg body weight) resulted ina marked decrease in GFR and urinary NO_x associated with decreasesin FL_Na and TR_Na but did not influence FR_Na. In contrast, inC rats the post-L-NAME decrease in NO_x was not associated withsignificant changes in GFR and renal sodium handling. GTN-treatedC rats exhibited a renal vasodilatory response and an increasein natriuresis and urinary NO_x whereas no renal changes wereobserved in D rats during GTN administration. The present dataindicate that changes in renal sodium handling before and afterNO modulation in experimental diabetes are related to changesin GFR rather than to the renal activity of NO. Therefore, incontrast to the effects on renal haemodynamics, NO does notplay an important role in the altered renal sodium handlingobserved in experimental diabetes.     

Oral sorbent AST-120 increases renal NO synthesis in uremic rats.

OBJECTIVE: The urine level of nitric oxide (NO) metabolites, i.e., nitrates/nitrites (NOx), in chronic renal failure (CRF) is decreased because of reduced renal synthesis of NO. We determined whether the administration of an oral sorbent, AST-120, increases the urine level of NOx and the renal expression of nitric oxide synthase (NOS) isoforms in CRF rats. METHODS: Chronic renal failure rats were produced by 4/5 nephrectomy. Rats were randomized into two groups: CRF control rats, and AST-120-treated CRF rats. The AST-120 was administered to the rats at a dose of 4 g/kg with powder chow for 16 weeks, whereas powder chow alone was administered to control rats. The urine levels of NOx were measured by using a NOx colorimetric assay kit. The expression of endothelial NOS (eNOS), inducible NOS (iNOS), and neuronal NOS (nNOS) in the kidney was determined by immunohistochemistry. Serum and urine levels of indoxyl sulfate were determined by high-performance liquid chromatography. RESULTS: Urine levels of NOx and the expression of glomerular eNOS and tubulointerstitial nNOS were significantly decreased in CRF rats compared with normal rats. The administration of AST-120 to CRF rats significantly increased urine levels of NOx and the expression of glomerular eNOS and tubulointerstitial nNOS. The administration of AST-120 to CRF rats significantly decreased urine and serum levels of indoxyl sulfate. CONCLUSIONS: The oral sorbent AST-120 increases NO synthesis in the kidneys of uremic rats by increasing the renal expression of eNOS and nNOS, through alleviation of indoxyl sulfate overload on the kidney.     

Acute renal hemodynamic effects of dimanganese decacarbonyl and cobalt protoporphyrin

BACKGROUND: Heme oxygenase (HO) products have a protective role in acute renal failure (ARF) that may be hemodynamically mediated because the HO-derived carbon monoxide (CO) is an important control system of arteriolar tone. The vascular effects of HO may be caused directly through changes in CO synthesis, and indirectly by alterations in nitric oxide (NO) release. The present study evaluated in vivo the renal effects of a heme oxygenase inhibitor, Co(III)Protoporphyrin (CoPP) alone or in combination with the CO donor dimanganese decacarbonyl (Mn2(CO)10). METHODS: All drugs were administered into the renal artery of anesthetized rats. Changes in renal cortical nitric oxide concentration were measured in vivo electrochemically. RESULTS: The intrarenal administration of the CO donor Mn2(CO)10 increased blood carboxyhemoglobin levels (+74%), renal blood flow (+54%), glomerular filtration (+38%), and urinary cGMP excretion (+128%). On the other hand, the inhibition of renal HO with CoPP progressively induced an ARF characterized by a drop in renal blood flow (-77%), glomerular filtration (-93%), and urinary cGMP excretion (-93%). These deleterious effects of HO inhibition on renal function were nearly abolished by supplementing CO with the coadministration of Mn2(CO)10+ CoPP, indicating that they may be caused by inhibition of CO synthesis and the resulting hemodynamic changes. In addition, CoPP lowered the renal cortical NO concentration (-21%) and also decreased the urinary excretion of nitrates/nitrites, while Mn2(CO)10 increased renal NO levels (+20%) and raised the excretion of nitrates/nitrites, suggesting that changes in NO release may contribute to the renal effects of the HO-CO system. CONCLUSION: These results indicate that heme oxygenase-derived CO plays a cardinal role in the control of renal hemodynamics and glomerular filtration.     

Antibodies against vascular endothelial growth factor improve early renal dysfunction in experimental diabetes

Vascular endothelial growth factor (VEGF) is a cytokine that potently stimulates angiogenesis, microvascular hyperpermeability, and endothelium-dependent vasodilation, effects that are largely mediated by endothelial nitric oxide synthase (eNOS). The expression of VEGF is pronounced in glomerular visceral epithelial cells, but its function in renal physiology and pathophysiology is unknown. VEGF expression is upregulated by high ambient glucose concentrations in several cell types in vitro and in glomeruli of diabetic rats. To assess the role of VEGF in the pathophysiology of early renal dysfunction in diabetes, monoclonal anti-VEGF antibodies (Ab) were administered to control and streptozotocin-induced diabetic rats for 6 wk after induction of diabetes. Based on in vitro binding studies, an adequate serum VEGF inhibitory activity was achieved during the entire course of anti-VEGF Ab administration. Anti-VEGF Ab treatment but not administration of isotype-matched control Ab decreased hyperfiltration, albuminuria, and glomerular hypertrophy in diabetic rats. VEGF blockade also prevented the upregulation of eNOS expression in glomerular capillary endothelial cells of diabetic rats. Antagonism of VEGF had no effect on GFR and glomerular volume in control rats. These results identify VEGF as a pathogenetic link between hyperglycemia and early renal dysfunction in diabetes. Targeting VEGF may prove useful as a therapeutic strategy for the treatment of early diabetic nephropathy.     

Altered endothelial nitric oxide synthase targeting and conformation and caveolin-1 expression in the diabetic kidney

Experimental diabetes is associated with complex changes in renal nitric oxide (NO) bioavailability. We explored the effect of diabetes on renal cortical protein expression of endothelial NO synthase (eNOS) with respect to several determinants of its enzymatic function, such as eNOS expression, membrane localization, phosphorylation, and dimerization, in moderately hyperglycemic streptozotocin-induced diabetic rats compared with nondiabetic control rats and diabetic rats with intensive insulin treatment to achieve near-normal metabolic control. We studied renal cortical expression and localization of caveolin-1 (CAV-1), an endogenous modulator of eNOS function. Despite similar whole-cell eNOS expression in all groups, eNOS monomer and dimer in membrane fractions were reduced in moderately hyperglycemic diabetic rats compared with control rats; the opposite trend was apparent in the cytosol. Stimulatory phosphorylation of eNOS (Ser1177) was also reduced in moderately hyperglycemic diabetic rats. eNOS colocalized and interacted with CAV-1 in endothelial cells throughout the renal vascular tree both in control and moderately hyperglycemic diabetic rats. However, the abundance of membrane-localized CAV-1 was decreased in diabetic kidneys. Intensive insulin treatment reversed the effects of diabetes on each of these parameters. In summary, we observed diabetes-mediated alterations in eNOS and CAV-1 expression that are consistent with the view of decreased bioavailability of renal eNOS-derived NO.     

Analysis of NO-synthase expression and clinical risk factors in human diabetic nephropathy.

BACKGROUND: Changes of renal nitric oxide (NO) production have been associated with glomerular hyperfiltration, vascular permeability, albuminuria, glomerulosclerosis and tubulointerstitial fibrosis. Several studies demonstrated an up- as well as downregulated expression of NO-synthases (NOS) in experimental diabetic nephropathy. It is still not yet specified whether the regulation and activity of NOS is changed in human diabetic nephropathy. METHODS: Renal biopsies and clinical data of 45 patients with diabetic nephropathy and of 10 control subjects were investigated. Glomerular and cortical endothelial NOS (eNOS) and inducible NOS (iNOS) expression were assessed by immunohistochemical staining and related to clinical data such as the duration of diabetes, insulin therapy and arterial hypertension, albuminuria/proteinuria, eGFR according to the formula modification of diet in renal disease (MDRD), presence of vascular complications or diabetic retinopathy. RESULTS: The mean age of patients at biopsy was 60.3 years and the mean duration of diabetes 12.9 years. Expression of cortical and glomerular eNOS was increased in type 2 diabetes (P < 0.05). Increased expression of glomerular and cortical eNOS correlated with more severe vascular complications (r = 0.44; P < 0.05). Glomerular eNOS was strongly increased among different degrees of proteinuria (P < 0.01). In contrast to expression levels of eNOS, the glomerular expression pattern of iNOS changed from an endothelial pattern in glomeruli with preserved morphology towards expression predominantly by inflammatory cells. CONCLUSIONS: Thus, increased eNOS expression by the renal endothelium could be demonstrated in type 2 diabetic nephropathy, whereas iNOS was unchanged but spatially differentially expressed. The eNOS expression was related to vascular lesions and the degree of proteinuria.     

Role of postglomerular microvessels in pathophysiology of diabetic nephropathy. Assessment and hypothesis

Although glomerular damage plays a well-established and important role in the pathomechanism of diabetic nephropathy, it alone does not fully explain the progression of renal complications in long-term diabetes mellitus. We discuss experimental evidence showing involvement of the postglomerular microvessels (peritubular capillaries and venules) in diabetic microangiopathy. This involvement is manifest in increased permeability of these vessels to plasma proteins and in highly augmented lymphatic drainage of the extravasated proteins from the renal interstitium. We suggest that in the advanced phase of diabetic nephropathy, proteinuria (corresponding to excess leakage of proteins through the glomerular capillary wall) indicates the probability that postglomerular microvessels have also allowed leakage of plasma proteins. As long as lymphatic drainage is capable of removing the increased quantity of extravasated plasma proteins from the interstitium, renal function should not be deleteriously affected. However, if the excess amount of extravasated proteins exceeds the capacity of lymphatic drainage, increases in interstitial volume and pressure are unavoidable with detrimental consequences for glomerular filtration and tubular reabsorption. Under these conditions, a potential positive-feedback loop can be visualized that involves increased extravasation of plasma proteins leading to increased interstitial pressure that through dilation of the afferent and efferent arterioles results in a further increase in protein extravasation. These conditions combined with glomerular damage should lead to the eventual collapse of renal function.     

Possible role of nitric oxide in the protective effect of resveratrol in 5/6th nephrectomized rats

BACKGROUND: Nitric oxide (NO) plays an important role in the modulation of glomerular disease. The renal protective effect of resveratrol (RVT), a polyphenolic phytoalexin, was investigated in the 5/6th nephrectomized rats. MATERIALS AND METHODS: Resveratrol (5 mg/kg, PO) was administered for 12 weeks to 5/6th nephrectomized (NX) rats together with and without nitro L-arginine methyl ester (L-NAME) (10 mg/kg, IP). We evaluated the effect of these agents on proteinuria, hypertension, renal function, glomerulosclerosis, and urinary excretion of nitric oxide metabolites. RESULTS: 5/6th NX resulted in elevation in systolic blood pressure (SBP), reduced the urinary excretion of NO metabolites, increased urinary protein excretion, and deranged renal function and glomerulosclerosis. Treatment of animals with resveratrol significantly attenuated the increase in SBP, preserved the normal renal function, reduced the urinary protein excretion, increased the urinary excretion of NO metabolites, and prevented the glomerulosclerosis. Co-administration of animals with L-NAME along with resveratrol prevented the protection observed with resveratrol. CONCLUSION: These findings indicate that resveratrol exerts its protective effect in 5/6 NX rats through a nitric oxide pathway.     

Preglomerular and postglomerular resistance responses to different levels of sympathetic activation by hypoxia.

This study investigated the effects of graded reflex increases in renal sympathetic nerve activity (RSNA) on renal preglomerular and postglomerular vascular resistances. With the use of hypoxia to reflexly elicit increases in RSNA without affecting mean arterial pressure, renal function and stop-flow pressures were measured in three groups of rabbits before and after exposure to room air and moderate (14% O2) or severe (10% O2) hypoxia. Moderate and severe hypoxia increased RSNA, primarily by increasing the amplitude of the sympathetic bursts rather than their frequency. RSNA amplitude increased by 20 +/- 6% (P < 0.05) and 60 +/- 16% (P < 0.05), respectively. Moderate hypoxia decreased estimated renal blood flow (ERBF; 26 +/- 7%; P = 0.07), whereas estimated glomerular capillary pressure (32 +/- 1 versus 34 +/- 1 mmHg; P < 0.05) and filtration fraction (FF; P < 0.01) increased. In response to moderate hypoxia, calculated preglomerular (approximately 20%) and postglomerular (approximately 70%) resistance both increased, but only the increase in postglomerular resistance was significant (P < 0.05). In contrast, severe hypoxia decreased ERBF (56 +/- 8%; P < 0.01), GFR (55 +/- 9%; P < 0.001), and glomerular capillary pressure (32 +/- 1 versus 29 +/- 1 mmHg; P < 0.001), with no change in FF, reflecting similar preglomerular (approximately 240%; P < 0.05) and postglomerular ( approximately 250%; P < 0.05) contributions to the vasoconstriction and a decrease in calculated K(f) (P < 0.05). These results provide evidence that reflexly induced increases in RSNA amplitude may differentially control preglomerular and postglomerular vascular resistances.     

Adenosine regulates renal nitric oxide production in hypothyroid rats.

In the hypothyroid kidney, exogenous adenosine (ADO) produces vasodilation and restores renal function to near-normal values. This study evaluates whether this response is mediated by nitric oxide synthesis stimulated by adenosine. GFR and urinary excretion of NO2-/NO3- (UNO2-/NO3-) were measured in normal (NL) and hypothyroid (HTX) rats under basal conditions and during infusion of: intra-aortic ADO, intravenously, 1,3-dipropyl-8p-sulfophenylxanthine (DPSPX), 8-cyclopentyl-1,3-dipropyl xanthine (DPCPX), N(omega)-nitro-L-arginine methylester (L-NAME) + ADO, L-NAME + PSPX, L-NAME + DPCPX, and intrarenal (IR) ADO or DPCPX + IR ADO. Intra-aortic ADO induced a fall in GFR and increased UNO2-/NO3- slightly in NL rats; in HTX rats, both GFR and UNO2-/NO3- increased significantly. DPSPX and DPCPX increased UNO2-/NO3- excretion in NL animals with minor changes in GFR; the blockers increased both GFR and UNO2-/ NO3- in HTX rats. L-NAME completely blocked the increase in NO2-/NO3- induced by ADO, DPSPX, and DPCPX. The intrarenal infusion of ADO at 1, 10, and 35 nmol/kg per min progressively decreased GFR with a slight increase in UNO2-/ NO3- in NL rats; in the HTX, GFR increased with the highest dose and UNO2-/NO3- progressively increased. DPCPX prevented the fall in GFR induced by intrarenal ADO in NL rats, with no further changes in UNO2-/NO3-; in HTX rats, intrarenal ADO under A1 blockade further increased GFR and UNO2-/NO3-. Arterial and venous ADO concentrations were lower in the HTX rats. In the HTX kidney, NO production was stimulated by ADO, most likely through activation of A2 or A3 receptors, whereas A1 receptors had an inhibitory effect. Thus, ADO receptors are involved in the regulation of kidney function in pathophysiologic conditions.     

Radical scavenging effect of gliclazide in diabetic rats fed with a high cholesterol diet

BACKGROUND: Gliclazide is a sulphonylurea antidiabetic drug with anti-oxidant effect due to its azabicyclo-octyl ring. We hypothesized that gliclazide may have a beneficial effect on diabetic nephropathy via radical scavenging. METHODS: Streptozotocin-induced diabetic rats fed a 4% cholesterol diet [high cholesterol-diabetes mellitus (HC-DM)] (N= 12) were treated with gliclazide (HC-DM + gliclazide) (N= 12) or glibenclamide (HC-DM + glibenclamide) (N= 12) after 2 weeks of the diabetes induction, and normal rat fed with 4% cholesterol served as control [high cholesterol-control (HC-control)] (N= 12). Renal expression of endothelial nitric oxide synthase (eNOS) and intracellular adhesion molecule-1 (ICAM-1), oxidative stress production via nicotinamide adenine dinucleotide phosphate (NAD(P)H) oxidase and antioxidant enzyme manganese superoxide dismutase (MnSOD) were evaluated at 4 weeks and renal damage was examined at 8 weeks. RESULTS: HC-DM showed significant increase in renal NAD(P)H oxidase and reduction in MnSOD with a significant increase in urinary lipid peroxidation products and H2O2 excretion compared to HC-control. Gliclazide treatment, but not glibenclamide, significantly reduced the oxidative products and NAD(P)H oxidase. There was no difference in renal eNOS expression between HC-DM and HC-control rats, and only gliclazide treatment enhanced eNOS expression. Renal damage evaluated by increased glomerular macrophage migration via enhanced ICAM-1 expression, mesangial matrix expansion, and albuminuria was significantly increased in HC-DM, and they were ameliorated by gliclazide but not by glibenclamide. CONCLUSION: Gliclazide reduced oxidative stress in diabetic rats fed a high cholesterol diet with reduction of renal NAD(P)H oxidase expression, enhanced MnSOD and eNOS expression, and had a beneficial effect on glomerular macrophage infiltration and mesangial expansion.     

Development of diabetic nephropathy in the Milan normotensive strain, but not in the Milan hypertensive strain: possible permissive role of hemodynamics

BACKGROUND: Rats of the Milan normotensive strain develop spontaneous glomerulosclerosis, whereas those of the Milan hypertensive strain are resistant to renal disease, possibly due to intrarenal artery hypertrophy protecting from systemic hypertension. To assess the role of hemodynamic versus metabolic factors in diabetic nephropathy, we investigated whether streptozotocin-induced diabetes accelerates glomerulosclerosis in Milan normotensive and/or removes (the hemodynamic) protection in Milan hypertensive rats by reducing preglomerular vascular resistance. METHODS: Diabetic and nondiabetic Milan normotensive, hypertensive, and progenitor Wistar rats were followed for 6 months for the assessment of renal function and structure. RESULTS: Proteinuria increased in nondiabetic and diabetic normotensive and, to a lesser extent, in diabetic Wistar, but not hypertensive rats. Serum creatinine increased and creatinine clearance decreased in nondiabetic and diabetic normotensive rats at 6 months. At 1.5 months, diabetic normotensive, but not hypertensive rats showed increased glomerular filtration rate and filtration fraction, suggesting glomerular hypertension. Diabetic nephropathy was detected in diabetic normotensive and Wistar, but not hypertensive rats. Glomerular extracellular matrix and TGF-beta mRNA levels increased with diabetes (and age) in normotensive, but not hypertensive rats. Arterioles and interlobular arteries showed increased media thickness in hypertensive versus normotensive rats, with diabetes reducing it only in the normotensive. CONCLUSION: These data show that Milan hypertensive rats are not susceptible to diabetic nephropathy, at variance with glomerulosclerosis-prone Milan normotensive rats, thus indicating the importance of genetic background. Our study suggests that the nature of this (genetic) protection might be hemodynamic, with intrarenal artery hypertrophy preventing diabetes-induced loss of autoregulation.     

Decompensated benign nephrosclerosis and secondary malignant nephrosclerosis

Using semiquantitative morphometric methods, the clinical picture of decompensated benign nephrosclerosis is distinguished from that of secondary malignant nephrosclerosis, designated as the consequence of high pressure. It is shown that hypertensive glomerulopathy triggered by high pressure and postglomerular interstitial fibrosis with tubular atrophy are in the foreground of pathologic changes in decompensated benign nephrosclerosis, whereas the preglomerular vessel network is most often affected in secondary malignant nephrosclerosis. The preglomerular vascular lesions in secondary malignant nephrosclerosis lead to such heavy stenosis of the afferent vessels that the clinical picture of hypertensive glomerulopathy is rarely observed, while that of ischemic glomerular capillary collapse is frequent. The preferred affliction of the glomeruli and the postglomerular vessel network leads in decompensated benign nephrosclerosis to severe interstitial fibrosis, which has a pyramidal form, decreasing from the base of the pyramid at the corticomedullary boundary to the outer renal cortex. In secondary malignant nephrosclerosis fibrosis of the renal cortical interstitium is homogeneous in all layers of the renal cortex. Clinically, decompensated benign nephrosclerosis and secondary malignant nephrosclerosis, which occur predominantly in young to middle-aged males, manifest malignant hypertension. They are also accompanied by progressive renal insufficiency.     

Shor- and long-term effect of a calcium channel blocker, barnidipine, on renal hemodynamics in spontaneously hypertensive, diabetic rats

Background The purpose of this study was to examine the short- and long-term effects of the calcium channel blocker, barnidipine, on renal hemodynamics and urinary albumin excretion in spontaneously hypertensive rats with streptozotocin-induced diabetes. Methods Diabetic and nondiabetic spontaneously hypertensive rats and nonhypertensive rats were treated with barnidipine or placebo (vehicle). In the short-term experiment, barnidipine was given as a single bolus injection (3 μg/kg); in the long-term experiment, barnidipine was administered orally (15 mg/kg per day) for 16 to 20 weeks. Results Renal hyperfiltration was observed in both hypertensive and nonhypertensive rats at 1 to 2 weeks after induction of diabetes, without changes in renal blood flow. Although short-term administration of barnidipine significantly decreased mean arterial pressure and renal vascular resistance, barnidipine did not affect renal blood flow or glomerular filtration rate in hypertensive, diabetic rats. At 16 to 20 weeks after induction of diabetes, renal hyperfiltration and increased urinary albumin excretion were still observed in hypertensive rats given placebo, compared to values for hypertensive nondiabetic rats given placebo. Long-term administration of barnidipine to hypertensive, diabetic rats suppressed the increase in both glomerular filtration rate and urinary albumin excretion, and reduced systolic blood pressure without any change in renal blood flow, renal vascular resistance, or filtration fraction. Conclusions These results indicate that in hypertensive, diabetic rats short-term administration of barnidipine, despite reducing renal vascular resistance, is less effective than long-term administration in restoring normal renal filtration, although long-term administration may normalize renal filtration and reduce urinary albumin excretion.     

Effects of NADPH oxidase inhibitor in diabetic nephropathy

BACKGROUND: We used apocynin to test the hypothesis that superoxide anion (O(-) (2)) from nicotinamide adenine dinucleotide phosphate (NADPH) oxidase underlies the development of diabetic nephropathy in the rat. METHODS: Rats received apocynin (16 mg/kg/day) from 2 to 8 weeks after inducing diabetes mellitus (DM) with streptozotocin. RESULTS: DM increased excretion of hydrogen peroxide (H(2)O(2)), lipid peroxidation products (LPO), nitric oxide products (NOx), and protein. The kidneys of rats with DM had increased expression of p47phox and gp91phox and endothelial nitric oxide synthase (eNOS), and increased mesangial matrix with expression of fibronectin and collagen I. Apocynin prevented the increase in excretion of H(2)O(2), LPO, and protein in diabetic rats, increased renal NOx generation, and prevented the increased renal expression of gp91phox and the membrane fraction of p47phox, and reverted the mesangial matrix expansion. CONCLUSION: Activation of NADPH oxidase with translocation of p47phox to the membrane underlies the oxidative stress and limited NO generation, despite enhanced eNOS expression in a model of diabetic nephropathy. Apocynin prevents these changes and the associated proteinuria.     

Blockade of serotonin 2A receptor improves glomerular endothelial function in rats with streptozotocin-induced diabetic nephropathy

Background Serotonin (5-HT) is involved in vascular inflammation and atherosclerogenesis. Serum 5-HT concentrations are elevated in diabetes, and 5-HT is involved in diabetic vasculopathies. Sarpogrelate hydrochloride, a 5-HT2A receptor antagonist, has renoprotective effects, but its effect in diabetic nephropathy is not elucidated. The aim of this study was to examine the effects of sarpogrelate on endothelial dysfunction in rats with streptozotocin (STZ)-induced diabetes. Methods Rats with STZ-induced diabetes were either untreated or treated with sarpogrelate (30 mg/kg P.O.) for 8 weeks. At the end of the experiment, we measured urinary albumin excretion, serum adiponectin concentration and platelet-derived microparticles. Intraglomerular coagulation was detected by immunostaining for platelets. Production of renal reactive oxygen species (ROS) and nitric oxide (NO) was investigated by confocal laser microscopy and used as an index of glomerular endothelial dysfunction. Results Diabetic nephropathy was associated with enhanced production of ROS and diminished bioavailable NO in the glomeruli. Treatment with sarpogrelate improved ROS/NO imbalance in glomeruli, suppressed platelet aggregation in glomeruli, reduced platelet-derived microparticles, increased serum adiponectin level and reduced the level of albuminuria, compared with non-treated diabetic rats. Conclusions Our results indicate that sarpogrelate improves endothelial function in rats with STZ-induced diabetes through a reduction of glomerular platelet activation and an increase in serum adiponectin concentrations and suggest that sarpogrelate is potentially useful for the treatment of diabetic nephropathy.     

The determinants of renal hemodynamics in pregnancy

Studies in the rat have permitted a complete characterization of the glomerular hemodynamic changes that occur during a normal pregnancy. The rise in glomerular filtration rate (GFR) is the result of an evenly distributed increase in plasma flow to all nephrons due to vasodilation of both pre- and postglomerular resistance vessels. Pregnancy is not associated with any change in blood pressure in the glomerular capillaries; neither is there any detectable alteration in the glomerular water permeability or filtration surface area. Despite the chronically maintained renal vasodilation of pregnancy, gravid rats exhibit substantial renal reserve when challenged with an amino acid load, indicating that a residual renal vasodilatory capacity exists in the kidney of the normal pregnant animal. Despite the concomitant plasma volume expansion of pregnancy, the tubuloglomerular feedback system (a volume regulatory system that modulates GFR) remains fully active in the pregnant rat, suggesting that the plasma volume in pregnancy is not sensed as expanded by this system. The factor(s) that initiates the gestational rise in GFR is currently unknown, although since similar renal hemodynamic changes occur in the pseudopregnant rat, the stimulus is maternal rather than fetoplacental in origin.