Effect of the calcium channel blocker nitrendipine in normotensive and spontaneously hypertensive, diabetic rats on kidney morphology and urinary albumin excretion.

OBJECTIVE: To investigate the effect of nitrendipine on the development of renal changes in experimental diabetes. DESIGN: Streptozotocin (STZ)-induced diabetic normotensive Wistar rats (WIS) and spontaneously hypertensive rats (SHR) were randomly allocated to nitrendipine treatment (250 mg/kg fodder) or placebo treatment for 6 months. METHODS: Blood pressure was assessed by the tail-cuff method, urinary albumin excretion (UAE) was determined, and glomerular basement membrane (GBM) thickness, mesangial volume, and mean glomerular volume (MGV) were estimated by morphometric measurements. RESULTS: In diabetic WIS, nitrendipine significantly reduced UAE after 2 months of treatment (P< 0.05), while no effect was was seen after 4-6 months. In diabetic SHR, no effect on UAE was seen at any time. Nitrendipine was unable to inhibit the renal and glomerular enlargement in diabetic WIS and SHR. Diabetes plus hypertension was associated with significant increase in GBM thickness, while diabetes or hypertension alone showed no significant increase in GBM. Nitrendipine treatment was unable to prevent increased GBM in diabetic SHR. CONCLUSION: Nitrendipine inhibits an early increase in UAE in normotensive, diabetic rats, but fails to sustain this effect in long-term diabetes. No effect of nitrendipine was observed in SHR.     

Genetic hypertension accelerates nephropathy in the streptozotocin diabetic rat

To evaluate whether hypertension is a cause or just an association with diabetic renal disease, diabetes was induced in both normotensive Wistar-Kyoto and spontaneously hypertensive rats (WKY and SHR). Animals were assessed monthly for 8 months before sacrifice. When compared to normotensive diabetic rats (WKY-STZ), hypertensive diabetic rats (SHR-STZ) had an earlier and more rapid rise in urinary albumin excretion. In addition, SHR-STZ had increased glomerular basement membrane thickness when compared to WKY-STZ or SHR. In a separate experiment, Enalapril therapy (35 mg/L) was administered in drinking water to WKY-STZ and SHR-STZ. Enalapril significantly reduced blood pressure in both animal groups, and this was associated with a decrease in urinary albumin excretion. The SHR-STZ model has accelerated nephropathy as determined by both functional and structural parameters. Angiotensin-converting enzyme inhibition is associated with a reduction in albuminuria in both hypertensive and normotensive models of diabetic nephropathy.     

Preventive effect of cerivastatin on diabetic nephropathy through suppression of glomerular macrophage recruitment in a rat model

AIMS/HYPOTHESIS: We investigated the effect of cerivastatin, a statin, on the development of diabetic nephropathy in spontaneously hypertensive rats (SHR) with streptozotocin-induced diabetes. METHODS: Diabetic SHR were given standard chow or chow containing cerivastatin at a dose of 0.1 mg/kg or 1.0 mg/kg for 12 weeks. Effects of cerivastatin on urinary albumin excretion, mesangial expansion, glomerular macrophage infiltration, and the number of anionic sites on the glomerular basement membrane (GBM) were assessed. RESULTS: Cerivastatin did not affect the blood glucose concentration, blood pressure or serum cholesterol concentration in diabetic SHR. However, cerivastatin treatment caused a dose-dependent decrease of albuminuria and hyperfiltration. At 1.0 mg/kg, cerivastatin inhibited the diabetes-induced expansion of mesangial and tuft areas on histological examination of the kidneys, as well as the loss of anionic sites from the GBM evaluated with polyetyleneimine and the intraglomerular infiltration of ED1-positive macrophages evaluated by immunohistochemistry. Whole-kidney expression of mRNA for MCP-1 and TGF-beta, estimated by the real-time quantitative RT-PCR, was increased (both 2.6-fold) in untreated diabetic SHR at 12 weeks. Cerivastatin treatment (1.0 mg/kg) inhibited the up-regulated expression of MCP-1 and TGF-beta mRNA (decreased to 48% and 34%, respectively) in diabetic SHR. CONCLUSION/INTERPRETATION: In this hypertensive model of diabetic nephropathy, cerivastatin decreased albuminuria through suppression of glomerular hyperfiltration, mesangial expansion, and the loss of charge barrier independently of a cholesterol-lowering effect. These preventive effects could be at least partly due to inhibition of macrophage recruitment and activation, and inhibition of TGF-beta overexpression.     

Structural involvement in type 1 and type 2 diabetic nephropathy

Structural changes underlying diabetic nephropathy in Type 1 diabetes are prodominant in the glomerulus [thickening of glomerular basement membrane (GBM) and mesangial expansion], but also include arteriolar, tubular and interstitial lesions. The structural measure that correlates best with all renal functional parameters in Type 1 diabetes is mesangial fractional volume [Vv(mes/glom)], an estimate of mesangial expansion. Structural-functional relationships in Type 2 diabetes are much less known. These studies investigated renal structure in the early stages of nephropathy [microalbuminuria (MA)] in patients with Type 1 and Type 2 diabetes. Diabetic glomerulopathy was quite advanced in Type 1 diabetic patients with MA, and both Vv (mes/glom) and GBM width were increased as compared to normoalbuminuric (NA) patients when the albumin excretion rate (AER) was > 30 microgram/min. Serial renal biopsies were performed 5 years apart in 11 Type 1 diabetic patients to evaluate whether glomerular and interstitial lesions progress jointly. AER increased significantly in 5 years, while the glomerular filtration rate remained unchanged. All structural parameters were initially abnormal. Vv(mes/glom) and mean glomerular volume increased significantly, whereas GBM width and the interstitial volume fraction were unchanged. Moreover, the change in Vv (mes/glom) was correlated with the change in AER (r =0.64, p <0.05). Thus, at the disease stage during which some patients progress to MA or proteinuria, continuing mesangial expansion is the main variable, whereas further interstitial expansion does not occur. A large number of Type 2 patients were also studied. Early diabetic glomerulopathy was detected by electron microscopy in NA patients and found to be more advanced in those with MA and proteinuria. However, lesions were milder than in Type 1 diabetic patients, and there was considerable overlap between groups. Morphometric results by electron microscopy were similar to those by light microscopy, demonstrating the heterogeneity of renal structure in Type 2 diabetic patients. In fact, only 30% of MA patients had typical diabetic glomerulopathy, while 40% had more advanced tubulo-interstitial and/or vascular lesions and 30% had normal renal structure.     

Effect of octreotide and insulin on manifest renal and glomerular hypertrophy and urinary albumin excretion in long-term experimental diabetes in rats

Summary Treatment of diabetic rats with octreotide can inhibit early diabetic renal hypertrophy. Octreotide administration for 6 months from the day of diabetes induction inhibits renal hypertrophy and diminishes increase in urinary albumin excretion. To investigate the effect of octreotide on manifest diabetic renal changes, octreotide treatment was given for 3 weeks after an untreated diabetic period of 3 or 6 months. In addition, following 6 months of diabetes, a group of diabetic rats was treated with insulin for 3 weeks. Renal and glomerular hypertrophy, and increased urinary albumin excretion were observed in diabetic rats compared to non-diabetic control rats from 3 months and throughout the study period. Octreotide treatment did not affect body weight, food intake, blood glucose or serum fructosamine levels. We observed no effect of octreotide treatment on renal and glomerular hypertrophy or urinary albumin excretion compared to placebotreated diabetic rats. Insulin treatment for 3 weeks after 6 months of untreated diabetes normalized blood glucose and serum fructosamine levels, and furthermore renal hypertrophy was significantly diminished compared to the placebo-treated diabetic rats. However, insulin treatment had no effect on glomerular hypertrophy or urinary albumin excretion. In conclusion, octreotide treatment for 3 weeks following an untreated diabetic period of 3 or 6 months is unable to reduce the increased renal and glomerular volume or urinary albumin excretion. However, insulin treatment for 3 weeks with induction of euglycaemia diminishes the renal hypertrophy but has no effect on glomerular volume or urinary albumin excretion.Abbreviations UAE Urinary albumin excretion - IGF-I insulin-like growth factor I - IGFBP insulin-like growth factor binding protein - STZ streptozotocin - TGV total glomerular volume - BW body weight - GH growth hormone - RPF renal plasma flow     

Differential effects of angiotensin II type-1 receptor antisense oligonucleotides on renal function in spontaneously hypertensive rats

The effect of selectively decreasing renal angiotensin II type 1 (AT1) receptor expression on renal function and blood pressure has not been determined. Therefore, we studied the consequences of selective renal inhibition of AT1 receptor expression in normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) in vivo. Vehicle, AT1 receptor antisense oligodeoxynucleotides (AS-ODN), or scrambled oligodeoxynucleotides were infused chronically into the cortex of the remaining kidney of conscious, uninephrectomized WKY and SHR on a 4% NaCl intake. Basal renal cortical membrane AT1 receptor protein was greater in SHR than in WKY. In WKY and SHR, AS-ODN decreased renal but not cardiac AT1 receptors. AT1 receptor AS-ODN treatment increased plasma renin activity to a greater extent in WKY than in SHR. However, plasma angiotensin II and aldosterone were increased by AS-ODN to a similar degree in both rat strains. In SHR, sodium excretion was increased and sodium balance was decreased by AS-ODN but had only a transient ameliorating effect on blood pressure. Urinary protein and glomerular sclerosis were markedly reduced by AS-ODN-treated SHR. In WKY, AS-ODN had no effect on sodium excretion, blood pressure, or renal histology but also modestly decreased proteinuria. The major consequence of decreasing renal AT1 receptor protein in the SHR is a decrease in proteinuria, probably as a result of the amelioration in glomerular pathology but independent of systemic blood pressure and circulating angiotensin II levels.     

Diabetic glomerulopathy in the SHR/N-corpulent rat: role of dietary carbohydrate in a model of NIDDM

Summary We evaluated the course of diabetes and nephropathy in the SHR/N-cp (corpulent) rat characterized by genetic obesity, non-insulin-dependent diabetes (NIDDM), and hypertension, and examined whether the nephropathy in this model is influenced by the type of carbohydrate intake. Two groups of obese and lean SHR/N-cp rats were fed diets containing 54 % carbohydrate, as either sucrose or starch for 3 months (group I) and 9 months (group II). After 3 months on either diet, group I obese rats had higher 2-h response serum glucose levels and urinary glucose excretion than lean rats. Sucrose feeding was associated with greater proteinuria and a higher percentage of abnormal glomeruli in obese rats. Morphometric evaluation of glomeruli (by computerized image analysis) showed greater mean renal corpuscular volume and mesangial fraction in obese than in lean rats fed similar diets. Mean renal corpuscular volume and mesangial fraction were also greater in sucrose-fed obese rats than in starch-fed obese rats. After 9 months, group II obese rats had substantial reductions in serum and urine glucose levels but they were still hyperinsulinaemic and showed more proteinuria than lean rats and a higher percentage of sclerotic glomeruli compared with group I obese rats. At this time, mean mesangial fraction but not renal corpuscular volume was still higher in obese than in lean rats. In group I obese rats, a significant correlation was found between mesangial fraction and urinary protein excretion (r = 0.67, p < 0.05). In group II obese rats, renal corpuscular volume was correlated with percentage of glomerular sclerosis (r = 0.60, p < 0.05). Thus, obese SHR/N-cp rats develop persistent proteinuria and glomerulopathy marked by glomerular enlargement, increased mesangial matrix, and progressive glomerular sclerosis. Sucrose feeding accentuates mesangial expansion and glomerulosclerosis in obese rats. [Diabetologia (1995) 38: 31–38] Received: 14 February 1994 and in revised form: 5 July 1994     

Functional Evidence of Inhibitory Reno-renal Reflexes in Spontaneously Hypertensive Rats

The experiments were performed to study the role of the renal nerves and the reno-renal reflexes in the control of water and sodium excretion in spontaneously hypertensive rats (SHR) compared to their normotensive controls, Wistar Kyoto (WKY) rats. Unilateral renal denervation in anaesthetized animals produced a slight, progressive decrease in arterial pressure in both WKY and SHR rats. The glomerular filtration rate temporarily increased in the kidney that underwent the denervation in the SHR group only. After unilateral renal denervation a sharp increase in water and sodium excretion from the ipsilateral kidney was observed in both WKY and SHR. One hour after the denervation, the percent changes in water and sodium excretion were smaller in WKY (+32 ± 19% and +24 ± 17%) than in SHR rats (+84 ± 15% and +93 ± 20%). In the kidney contralateral to the denervation a reduction in water and sodium excretion was observed and this reduction was prompter in SHR than in WKY rats. One hour after the denervation, the percent changes in water and sodium excretion were similar in WKY (-21 ± 8% and -18 ± 7%) and SHR (-19 ± 6% and -19 ± 7%). In control groups, sham denervation did not cause significant changes in glomerular filtration rate, and urinary water and sodium excretion. Arterial pressure slightly and progressively decreased in both control groups. Electrical stimulation of the efferent renal nerves performed in WKY and SHR produced similar decreases in renal blood flow, glomerular filtration rate, and water and sodium excretion in the two groups for the same frequencies of stimulation. As this finding indicates that renal targets in hypertensive rats are normally responsive to the neural drive, our data demonstrate that renal responses to unilateral renal denervation in hypertensive rats are equal to the responses observed in normotensive rats. Our results indicate that tonically active inhibitory reno-renal reflexes normally operate in spontaneously hypertensive rats.     

Glomerular basement membrane polyanion distribution and nitric oxide in spontaneous hypertensive rats: effects of salt loading and antihypertensive therapy with propranolol

Cationic colloidal gold (CCG), a polycationic histochemical probe, was used to analyze the distribution of glomerular basement membrane (GBM) polyanions, mainly heparan sulfate proteoglycan in spontaneous hypertensive rats (SHR) with or without salt loading and antihypertensive treatment with propranolol. The changes of mean GBM width and anionic sites distribution were assessed by electron microscopy. Plasma and urinary nitrates (NO(x)) were measured by nitrite (NO2) + nitrate (NO3), stable metabolites of NO. SHR had decreased NO production and increased GBM width (27%) compared with the control Wistar-Kyoto (WKY) rats. The chronic high dietary salt intake resulted in a significant increase in blood pressure, proteinuria, and renal function in the SHR rats. The chronic high salt dietary intake resulted in a decrease in NO in the WKY and a further reduction in NO production in the SHR. The GBM anionic sites count was similar in the SHR and WKY nonsalt-loaded groups, 13.5 +/- 0.5 and 12.8 +/- 0.4 CCG counts/microm GBM, respectively, but significantly lower in both salt-loaded SHR and WKY, 9.9 +/- 0.55 (P < .01) and 9.6 +/- 0.55 (P < .01) CCG counts/microm GBM, respectively. Antihypertensive treatment with propranolol in the salt-loaded SHR group resulted in lower blood pressure, a further decrease in NO production, but no significant changes in GBM width and anionic sites count. It is concluded that chronic high salt intake may be deleterious to the permselectivity of the GBM. A low NO production state that results from chronic salt loading in already hypertensive rats will result in more severe organ (renal) damage, most probably by the addition of the loss of GBM permselectivity to the existing pathomorphologic changes.     

Renal function of conscious spontaneously hypertensive rats

Renal clearance studies were performed in conscious 13-week-old spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY) before and during volume expansion by intravenous infusion of isotonic saline. Mean arterial pressure and filtration fraction were greater in SHR, whereas fractional and absolute excretion of sodium and water, glomerular filtration rate, and renal plasma flow in SHR and WKY were not statistically different. This was the case during hydropenia and volume expansion. We did not observe as exaggerated natriuresis after intravenous loading when unanesthetized SHR were compared with the response of WKY. These observations suggest that the kidneys of genetically hypertensive rats of the Okamoto-Aoki strain have adapted to an elevated renal perfusion pressure or that hypertension is required to normalize renal function so that excretion is appropriately matched with intake.     

Increased urinary type IV collagen marks the development of glomerular pathology in diabetic d/db mice.

The diabetic db/db mouse exhibits increased albumin excretion soon after the onset of obesity and hyperglycemia, and later manifests glomerular mesangial matrix expansion resembling that found in human diabetic nephropathy. Since the glomerular lesion in this rodent model of type 2 diabetes is associated with renal overexpression of mRNA encoding type IV collagen, we postulated that changes in the urinary excretion of collagen IV may reflect developing glomerular pathology. To explore this hypothesis, we monitored urinary collagen IV (measured by immunoassay) in db/db mice during the course of evolution of nephropathy. At age 8 weeks, collagen IV excretion was not different in diabetic compared to nondiabetic animals despite marked albuminuria, but was significantly increased in db/db compared to db/m mice at age 12 and 16 weeks. Serum levels of collagen IV did not significantly differ between normal versus diabetic mice at any age. Glomerular morphometry revealed mesangial matrix expansion at age 12 weeks, coincident with the rise in collagen IV excretion, which became more marked at age 16 weeks in association with reduced creatinine clearance and elevated serum creatinine. The findings suggest that increased urinary type IV collagen is a better indicator than albuminuria of developing glomerular matrix accumulation that results in compromised renal filtration function.     

Cerebrovascular and brain microanatomy in spontaneously hypertensive rats with streptozotocin-induced diabetes

The influence of hypertension associated with diabetes on cerebrovascular and frontal cortex or hippocampus microanatomy was investigated in 20-week-old spontaneously hypertensive rats (SHR) in which diabetes was induced by treatment with streptozotocin (STZ) and in control or STZ-diabetic age-matched normotensive Wistar Kyoto (WKY) rats. At the beginning of experiment, systolic pressure values were similar in WKY rats either control, or exposed to STZ and remarkably higher in control or STZ-treated SHR. Systolic pressure values increased in the different animal groups examined along the course of experiment. Blood glucose levels were increased in either STZ-WKY rats or -SHR compared to WKY rats and SHR respectively. The main changes occurring in pial and intracerebral arteries of SHR and STZ-SHR were thickening of the arterial wall accompanied by luminal narrowing. In medium sized pial arteries of STZ-WKY rats luminal narrowing and a decreased thickness of arterial wall were noticeable. Intracerebral arteries of STZ-WKY diabetic rats showed a not homogeneous sensitivity of different sized branches. The volume of zones III and IV of frontal cortex was decreased in SHR and STZ-SHR compared to control WKY rats. The number of nerve cells in these cerebrocortical layers was decreased to a similar extent in SHR. STZ-WKY rats or STZ-SHR compared to control WKY rats. In dentate gyrus, followed by the CA1 subfield of hippocampus, decreased volume and number of neurons were found in SHR and STZ-SHR compared to control WKY rats. The occurrence of astrogliosis was observed in hypertensive, diabetic or hypertensive plus diabetic rats. The above findings indicate the occurrence of cerebrovascular and brain microanatomical changes in SHR and to a lesser extent in STZ-diabetic rats compared to control normotensive and normoglicemic WKY rats. Association of hypertension and diabetes caused more pronounced changes than in the single disease models. These results support the view that hypertension and diabetes affect the structure of cerebrovascular tree and of brain and that association of the two diseases results in an increased risk of target-organ damage, involving brain.     

The characteristics of renal hemodynamics in diabetic spontaneously hypertensive rats in comparison with diabetic Wistar-Kyoto rats

Diabetic Sprague-Dawley (SD) rats are known to exhibit renal hyperfiltration and hyperperfusion accompanied by renal hypertrophy. We examined whether such characteristics of renal hemodynamics in diabetic SD rats are also observed in diabetic spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats. SHR and WKY rats were divided into four groups: D-S, diabetic SHR; N-S, nondiabetic SHR; D-W, diabetic WKY rats; and N-W, nondiabetic WKY rats. Streptozotocin (STZ), 90 mg, was intraperitoneally injected to induce diabetes. Renal blood flow (RBF) and glomerular filtration rate (GFR) were measured by a clearance method with paraaminohypurate and insulin, respectively, 7–12 days after diabetes induction. In D-S and D-W, there was no increase in the kidney weight and RBF, in spite of significant increases in GFR and fasting blood sugar levels. These results indicate that, in both WKY and SHR, diabetes does not always produce renal hypertrophy and does not result in an increase in RBF.     

Effects of aminoguanidine on glomerular basement membrane thickness and anionic charge in a diabetic rat model

We investigated the effect of aminoguanidine (AG) administration on GBM thickness, glomerular heparan sulfate (HS) content, and urinary albumin and HS excretion in diabetic rats. After induction of diabetes, female Wistar rats were divided into 2 groups: Group AGDM (n = 11) received 1 g/L aminoguanidine bicarbonate in drinking water, group DC (n = 12) was given only tap water. Control rats received AG (group AGH, n = 8) or tap water (group HC, n = 8). At the end of a period of 8 weeks, urinary albumin and glycosaminoglycan (GAG) excretion was detected. GBM heparan sulfate distribution and count was determined under the electron microscope. The AGDM group had lower urinary albumin and GAG excretion than diabetic controls. GBM thickness was increased in diabetic rats compared to groups of AGDM and HC. In AGDM group alcian blue stained particle distribution and count in the GBM was similar to healthy controls. In conclusion AG prevents the decrease of anionic charged molecules in the GBM and GBM thickening. This can be one of the mechanisms by which AG decreases albuminuria in diabetic rats.     

Longitudinal evaluation of the renal clearance of glycated albumin in the diabetic rat

This study has examined glycation of serum albumin and its role in evolving diabetic proteinuria. Renal clearances of endogenous glycated and nonglycated albumin were studied in groups of normal and streptozotocin-induced diabetic Wistar-Kyoto rats over a 32 week period. Concentrations of glycated and nonglycated albumin in serum and urine were measured by rat albumin radioimmunoassay following separation on m-aminophenylboronate affinity columns. Levels of glycated serum albumin in diabetic rats were significantly higher than in normal rats (5.9 +/- 0.7% vs 4.4 +/- 0.3%, P less than 0.05). Median total urinary albumin excretion increased from 120 micrograms/24 h at baseline to 879 micrograms/24 h (P less than 0.05) 28-32 weeks after induction of diabetes. The renal clearance of glycated albumin was approximately twice as great as that of nonglycated albumin in both normal (P less than 0.01) and diabetic (P less than 0.01) rats. However, the glycated albumin/nonglycated albumin clearance ratio in diabetic rats did not correlate with duration of diabetes or with the level of albuminuria. These results indicate that glycation of albumin does not contribute disproportionately to the development of proteinuria in the diabetic rat, during which median renal albumin clearance increased 7-fold. Other factors, such as glycation of the glomerular filtration surface, may have a more important role in the pathogenesis of proteinuria in experimental diabetes.     

Thiazolidinedione derivatives ameliorate albuminuria in streptozotocin-induced diabetic spontaneous hypertensive rat

Recent reports have shown that thiazolidinediones have preventive effects on urinary albumin excretion in diabetes. However, the mechanism leading to these effects has not yet been elucidated. We studied here the effects of thiazolidinediones on albuminuria and hemodynamic and morphological changes in the kidneys of streptozotocin (STZ)-induced diabetic spontaneous hypertensive rats (SHRs). Diabetes was induced in SHRs by intravenous injection of STZ (50 mg/kg). The diabetic SHRs were divided into the following 3 groups: (1) STZ-SHRs given normal chow (STZ), (2) STZ-SHRs given chow mixed with 0.1% troglitazone (STZ + tro), and (3) STZ-SHRs given chow mixed with 0.001% pioglitazone (STZ + pio). Three groups of nondiabetic SHRs were also investigated: (4) SHR, (5) tro, and (6) pio. We evaluated the urinary albumin excretion rate (AER) every 4 weeks. After 12 weeks of treatment, the animals were killed and renal morphological examinations were performed. Thiazolidinediones did not affect blood pressure or blood glucose levels. Urinary AER were markedly increased in STZ-SHRs. After 12 weeks of treatment with thiazolidinediones, the urinary AER was significantly decreased while creatinine (Cr) clearance was left unchanged. Histologically, the loss of anionic sites of glomerular basement membranes (GBM) evaluated with polyetyleneimine was suppressed significantly in the diabetic SHRs treated with thiazolidinediones. In conclusion, administration of thiazolidinediones in diabetic SHRs decreased the urinary AER and suppressed the loss of anionic sites of GBM without affecting blood pressure, blood glucose levels, or Cr clearance. These results clarify the novel therapeutic action of thiazolidinediones on diabetic nephropathy.     

Increased urinary type IV collagen marks the development of glomerular pathology in diabetic d/db mice

The diabetic db/db mouse exhibits increased albumin excretion soon after the onset of obesity and hyperglycemia, and later manifests glomerular mesangial matrix expansion resembling that found in human diabetic nephropathy. Since the glomerular lesion in this rodent model of type 2 diabetes is associated with renal overexpression of mRNA encoding type IV collagen, we postulated that changes in the urinary excretion of collagen IV may reflect developing glomerular pathology. To explore this hypothesis, we monitored urinary collagen IV (measured by immunoassay) in db/db mice during the course of evolution of nephropathy. At age 8 weeks, collagen IV excretion was not different in diabetic compared to nondiabetic animals despite marked albuminuria, but was significantly increased in db/db compared to db/m mice at age 12 and 16 weeks. Serum levels of collagen IV did not significantly differ between normal versus diabetic mice at any age. Glomerular morphometry revealed mesangial matrix expansion at age 12 weeks, coincident with the rise in collagen IV excretion, which became more marked at age 16 weeks in association with reduced creatinine clearance and elevated serum creatinine. The findings suggest that increased urinary type IV collagen is a better indicator than albuminuria of developing glomerular matrix accumulation that results in compromised renal filtration function.     

Perinatal L-arginine and antioxidant supplements reduce adult blood pressure in spontaneously hypertensive rats

Embryo cross-transplantation and cross-fostering between spontaneously hypertensive rats (SHR) and normotensive rats (WKY) suggest that perinatal environment modulates the genetically determined phenotype. In SHR the balance between NO and reactive oxygen species (ROS) is disturbed. We hypothesized that increasing NO and diminishing ROS in perinatal life would ameliorate hypertension in adult SHR. Pregnant SHR and WKY and their offspring received l-arginine plus antioxidants (vitamin C, vitamin E, and taurine) during the last 2 weeks of pregnancy and then until either 4 or 8 weeks after birth. Systolic blood pressure (SBP) and urinary excretion of protein, nitrates (NO(x)), and thiobarbituric acid reactive substances (TBARS) were measured. At 48 weeks of age rats were euthanized for glomerular counts. Perinatal supplements reduced SBP persistently in SHR and prevented the SBP increase observed in aging WKY. Initially NO(x) excretion was lower and TBARS excretion higher in SHR than WKY. There was a direct effect on NO(x) excretion in supplemented pregnant SHR and their offspring, but no increase was observed after stopping the supplements. TBARS excretion was only depressed up to 14 weeks by the supplements despite persistent differences in SBP. Consistent effects on nephron number were absent. Mild proteinuria, present in control SHR at 48 weeks, was prevented in all supplemented rats. Perinatal supplementation of NO substrate and antioxidants results in persistent reduction of SBP and renal protection in SHR, although effects on NO(x) and TBARS were only transient. This suggests a critical role for perinatal pro- and antioxidant balance in programming BP later in life.     

Interactions between hypertension and diabetes on vascular function and structure in rats.

Regional 125I-albumin permeation and glomerular structural changes were assessed in male Sprague-Dawley rats with diabetes and/or hypertension. All rats underwent unilateral nephrectomy 2 weeks after induction of diabetes with streptozotocin. At the same time, one-half of the nondiabetic and diabetic animals were placed on 1% saline drinking water and given weekly intramuscular injections of deoxycorticosterone acetate to induce hypertension (systolic blood pressure greater than 150 mm Hg). Vascular permeability studies were performed after 1 and 3 months of hypertension. Hypertension, alone or in combination with diabetes, had no effect on weight gain, plasma glucose, or food consumption, but did increase 24-h urine volume in nondiabetics. In normotensive diabetics and in nondiabetic hypertensive rats, vascular 125I-albumin permeation was increased in eyes, aorta, and new granulation tissue (formed in a subcutaneous fabric implant), and glomerular basement membranes were thickened without any change in the fractional volume of the glomerulus occupied by mesangium. Urinary albumin and IgG excretion in nondiabetic hypertensive rats was increased much more than in normotensive diabetics. Hypertension and diabetes were additive in their effects on 125I-albumin permeation in eyes, aorta, and granulation tissue, and on glomerular basement membrane thickening, but were synergistic in their effects on urinary albumin excretion and mesangial fractional volume. The magnitude of the increase in vascular albumin permeation and urinary albumin and IgG excretion between and 1 and 3 months was much larger in diabetic hypertensive rats than in rats with hypertension or diabetes alone. Neither diabetes nor hypertension, alone or in combination, had any effect on albumin permeation in skeletal muscle, skin, heart, or brain. These findings demonstrate that hypertension and diabetes increase vascular albumin permeation in rats preferentially in tissues that correspond to sites of clinically significant vascular disease in human diabetics. They also attest to an important interaction between blood pressure-induced and diabetes-induced increases in vascular permeability in these tissues and in structural changes in the glomerular vasculature.