Ruth Rasch

Maternal Protein Restriction Suppresses the Newborn Renin-Angiotensin System and Programs Adult Hypertension in Rats

Restriction of maternal protein intake during rat pregnancy produces offspring that are hypertensive in adulthood, but the mechanisms are not well understood. Our purpose was to determine whether this adult hypertension could be programmed during development by suppression of the fetal/newborn renin-angiotensin system (RAS) and a consequent reduction in nephron number. Pregnant rats were fed a normal protein (19%, NP) or low-protein (8.5%, LP) diet throughout gestation. Birth weight was reduced by 13% (p < 0.0005), and the kidney/body weight ratio was reduced in LP pups. Renal renin mRNA levels were significantly reduced in newborn LP pups; renal renin concentration and renin immunostaining were suppressed. Renal tissue angiotensin II levels were also suppressed in newborn LP (0.079 ± 0.002 ng/mg, LP versus 0.146 ± 0.016 ng/mg, NP, p < 0.01). Mean arterial pressure in conscious, chronically instrumented adult offspring (21 wk) was higher in LP (135 ± 1 mm Hg, LP versus 126 ± 1 mm Hg, NP, p < 0.00007), and GFR normalized to kidney weight was reduced in LP (p < 0.04). The number of glomeruli per kidney was lower in adult LP offspring (21,567 ± 1,694, LP versus 28,917 ± 2,342, NP, p < 0.03), and individual glomerular volume was higher (1.81 ± 0.16 106 μm3, LP versus 1.11 ± 0.10 106 μm3, NP, p < 0.005); the total volume of all glomeruli per kidney was not significantly different. Thus, perinatal protein restriction in the rat suppresses the newborn intrarenal RAS and leads to a reduced number of glomeruli, glomerular enlargement, and hypertension in the adult.

Renal enlargement: comparative autoradiographic studies of 3H-thymidine uptake in diabetic and uninephrectomized rats

SummaryIncorporation of 3H-thymidine into renal cortical tissue has been studied by light microscopic autoradiography in streptozotocin-diabetic rats, uninephrectomized rats, uninephrectomized diabetic rats, insulin-treated diabetic rats and control rats. The percentage of labelled cortical nuclei (the labelling index) was determined separately in glomeruli, proximal tubules and distal tubules after 2, 4 and 6 days on autoradiographs from 1 μm thick plastic embedded sections. The incorporation of thymidine in glomerular nuclei was consistantly low (< 1%) and no differences were found between the control and experimental groups. In both proximal and distal tubules an increase in thymidine incorporation was seen on day 2 followed by a decline on days 4 and 6. The maximal labelling on day 2 in proximal tubules was 9.1% in the uninephrectomized diabetic group, 3.7% in the diabetic group and 1.4% in the uninephrectomized group. In distal tubules the corresponding values were 5.2, 3.5 and 1.1%. The increase in kidney weight after 6 days was 83, 62 and 37%, respectively. Estimates of the net increase in the number of cortical tubular cells in the different experimental groups showed that the kidney enlargement followed different patterns with respect to the extent of cellular hyperplasia and hypertrophy. The kidney growth in uninephrectomized diabetic rats was dominated by tubular cellular hyperplasia, in the diabetic group hyperplasia and hypertrophy participated to approximately the same extent, whereas cellular hypertrophy was most pronounced in the uninephrectomized animals. Nuclear labelling in the insulin-treated diabetic rats was not different from that of control rats and consequently a hyperplastic effect of streptozotocin can be ruled out.It is concluded that combined tubular cellular hyperplasia and hypertrophy is involved in the kidney cortical enlargement seen in the present experimental groups and that each group follows different cellular reactions. In glomerular enlargement, however, no cellular hyperplasia is obserbed.

Perinatal ANG II programs adult blood pressure, glomerular number, and renal function in rats

ANG II is known to be important in normal renal development, but the long-term consequences of a suppressed renin-angiotensin system (RAS) during the developmental period are not completely understood. This study tested the hypothesis that the RAS in the developing animal is important in long-term regulation of renal function and arterial pressure. Newborn Sprague-Dawley rat pups were given the ANG II AT1 receptor antagonist losartan (25 mg . kg-1 . day-1 sc) for the first 12 days of postnatal life (Los). Body weights at weaning (22 days) were significantly reduced in Los (53.4 +/- 3.2 vs. 64.5 +/- 3.6 g in controls); however, at the time of study (approximately 22 wk), body weights and the kidney-to-body weight ratios were not different. In chronically instrumented conscious animals, glomerular filtration rate and effective renal plasma flow were reduced by 27 and 20%, respectively, in Los; the filtration fraction was not different. Maximal urine concentrating ability was also reduced in Los (1,351 +/- 45 vs. 2,393 +/- 52 mosmol/kg in controls). Mean arterial pressure was significantly higher in Los (134 +/- 3 vs. 120 +/- 1 mmHg). The number of glomeruli per kidney was reduced by 42% in Los, but the total glomerular volume was unchanged. Thus perinatal blockade of ANG II AT1 receptors results in fewer but enlarged glomeruli, reduced renal function, and an increased arterial pressure in adulthood. These data indicate that perinatal ANG II, acting via AT1 receptors, plays an important role in renal development and long-term control of renal function and arterial pressure. Physiological conditions that cause suppression of the RAS in the developing animal may have long-term consequences for renal function and blood pressure.ANG II is known to be important in normal renal development, but the long-term consequences of a suppressed renin-angiotensin system (RAS) during the developmental period are not completely understood. This study tested the hypothesis that the RAS in the developing animal is important in long-term regulation of renal function and arterial pressure. Newborn Sprague-Dawley rat pups were given the ANG II AT1 receptor antagonist losartan (25 mg ⋅ kg-1 ⋅ day-1sc) for the first 12 days of postnatal life (Los). Body weights at weaning (22 days) were significantly reduced in Los (53.4 ± 3.2 vs. 64.5 ± 3.6 g in controls); however, at the time of study (∼22 wk), body weights and the kidney-to-body weight ratios were not different. In chronically instrumented conscious animals, glomerular filtration rate and effective renal plasma flow were reduced by 27 and 20%, respectively, in Los; the filtration fraction was not different. Maximal urine concentrating ability was also reduced in Los (1,351 ± 45 vs. 2,393 ± 52 mosmol/kg in controls). Mean arterial pressure was significantly higher in Los (134 ± 3 vs. 120 ± 1 mmHg). The number of glomeruli per kidney was reduced by 42% in Los, but the total glomerular volume was unchanged. Thus perinatal blockade of ANG II AT1 receptors results in fewer but enlarged glomeruli, reduced renal function, and an increased arterial pressure in adulthood. These data indicate that perinatal ANG II, acting via AT1 receptors, plays an important role in renal development and long-term control of renal function and arterial pressure. Physiological conditions that cause suppression of the RAS in the developing animal may have long-term consequences for renal function and blood pressure.

The impact of experimental diabetes mellitus in rats on glomerular capillary number and sizes

SummaryThe structural counterpart of the increased glomerular filtration found in acute diabetes mellitus and experimental diabetes has been ascribed to the increased glomerular filtration surface. Using modern design-based stereological methods and light microscopy on perfusionfixed rat kidneys the average total surface area of capillaries per glomerulus in control rats was 291±42 10−3 mm2 (±SD) increasing to 383±55 10−3 mm2 in 10-day-diabetic rats. There was a further increase to 469±70 10−3 mm2 in 50-day-diabetic rats. The average total length of capillaries per glomerulus increased from 12.5±2.2 mm in the control rats to 16.9±2.4 mm in the 10-day-diabetic rats whereas the further increase to 19.4±3.0 mm in the 50-day-diabetic rats failed to reach statistical significance. The average number of topologically defined capillaries per glomerulus increased from 215±29 in the control rats to 260±45 and 316±29 in the 10-day-diabetic and 50-day-diabetic rats, respectively. The results showed just a slight increase in the mean length of a capillary from 58.1±6.2 μm in the control rats to 65.6±2.6 μm in the 10-day-diabetic rats after which it normalized in the 50-day-diabetic rats to 61.3±3.6 μm. The geometric factor or “resistance” in Poiseuilles law did not differ between the three groups, neither did the diameter of the capillaries, nor the number of glomeruli. The implications of the formation of capillaries in renal glomeruli are discussed and it is concluded that glomerular capillaries in experimental diabetes grow mainly by the formation of new capillaries.

Tubular lesions in streptozotocin-diabetic rats.

SummaryRenal tubular lesions have been studied in streptozotocin-diabetic rats after 50 days of diabetes and compared with age-matched controls. The kidney weight increased by 67% in the diabetic animals and the length of the proximal tubules increased by 22%, but no abnormalities were found. The length of the distal tubules increased by 20% and the total increase was due to abnormal distal tubules. These abnormalities were confined to the cortex and the outer stripe of the outer medulla, but they were not seen in the inner stripe of the outer medulla. Abnormal cells were found also in the distal tubular cells of the macula densa. The total length of the collecting ducts was the same in the two groups and the cells appeared normal. The cells of the abnormal distal tubules appeared either empty or full of a PAS-positive material, digestable with α-amylase. At the electron microscope level, the cytoplasm of the cells contained glycogen-like granules, strikingly few organelles and the basal infoldings were greatly reduced. It is suggested that these tubular lesions might play a role in the development of renal functional changes in diabetes.

Quantitative morphology of the rat kidney during diabetes mellitus and insulin treatment

Summary A morphometric study was performed on moderately hyperglycaemic streptozotocin diabetic rats after 10 and 50 days of diabetes, and on groups of rats that, after initial hyperglycaemia for 50 days, were insulin treated for 2 h or for 5, 15 or 38 days. A group of hyperglycaemic diabetic animals were fasted for 18 h. Another group of rats had acute hyperglycaemia induced by intravenous glucose injection. After 10 and 50 days of diabetes, kidney weight was increased by 55 and 93 %. Glomerular volume, tubule length, and tubular and interstitial volume increased in diabetic animals compared with controls. After 4 h insulin treatment, the kidney weight was 20 % decreased; after 5 days it was 31 % decreased. After 38 days the kidney weight was still 26 % greater than in controls. In diabetic animals, 18 h fasting induced a 30 % decrease in kidney weight. In normal animals, acute hyperglycaemia induced a 22 % increase in kidney weight. Volume fractions of most kidney structures remained similar in all groups. However, the glomerular volume fraction was smaller during kidney enlargement, and the tubular volume fraction was larger after induced hyperglycaemia compared with controls. In conclusion, high blood glucose levels in diabetic and normal animals are associated with increased kidney weight. In hyperglycaemic diabetic animals, normalization of blood glucose after insulin treatment or fasting was followed by a decrease in kidney weight. [Diabetologia (1997) 40: 802–809]

Mannose-binding lectin deficiency attenuates renal changes in a streptozotocin-induced model of type 1 diabetes in mice

Aims/hypothesisAn increasing amount of evidence indicates that mannose-binding lectin (MBL) plays a role in the development of diabetic nephropathy. The main objective of the study was to analyse whether MBL influences the effects of diabetes on the kidneys.Materials and methodsIn one group of wild-type mice and in one group of MBL double knockout mice we induced diabetes by the use of streptozotocin as a model of type 1 diabetes. Two groups of non-diabetic mice, wild-type and MBL knockout, were also included. By two-way ANOVA we evaluated if MBL modulated the effects of diabetes by testing the interaction between diabetes and MBL.ResultsMBL interacted with the effects of diabetes on three outcome measures: kidney weight (p < 0.001), urinary albumin excretion (p = 0.001) and the expression of collagen IVα1 (Col4a1) mRNA (p = 0.002). This means that the effects that diabetes normally has on these parameters were significantly modified by MBL. MBL showed a tendency to interact with the effects of diabetes on glomerular basement membrane thickness and total mesangial volume (p = 0.065 and p = 0.063, respectively). Glomerular volume and total mesangial volume were significantly smaller in animals lacking MBL than in wild-type animals (p = 0.006 and p = 0.047, respectively).Conclusions/interpretationThese findings, for the first time, show that the degree of kidney alteration as a consequence of diabetes is modified by MBL. These findings support a pivotal role of MBL in the development of diabetic kidney disease.

Pathophysiological role of vascular endothelial growth factor in the remnant kidney.

Background: Subtotal renal ablation is characterized by initial glomerular hypertrophy, followed by progressive development of glomerulosclerosis and interstitial fibrosis. Vascular endothelial growth factor (VEGF) is involved in glomerular hypertrophy and dysfunction in several pathophysiological conditions. On the other hand, progressive glomerulosclerosis and tubulo-interstitial fibrosis in the remnant kidney have been associated with loss of VEGF expression. Methods: To explore the pathophysiological role of VEGF in the development of glomerular hypertrophy and renal damage in the remnant kidney model, we examined the effect of a neutralizing VEGF antibody on glomerular volume and kidney function in rats after subtotal nephrectomy or sham operation. Erythropoietin was administered to exclude a confounding effect of anaemia. Results: Six weeks after subtotal nephrectomy, plasma urea and creatinine concentrations, urinary albumin excretion, and mean glomerular volume were elevated in the placebo-treated uraemic rats as compared with the sham-operated rats. Inhibition of VEGF partially prevented the glomerular hypertrophy and largely prevented the rise in urinary albumin excretion, but did not affect creatinine clearance in uraemic rats. Conclusions: VEGF is a mediator of glomerular hypertrophy after subtotal renal ablation. In view of glomerular hypertrophy as the initial deleterious event ultimately leading to progressive glomerulosclerosis, agents that block this glomerular growth could be useful in preventing scarring in progressive renal disease.

Quantitative changes in rat renin secretory granules after acute and chronic stimulation of the renin system

Abstract In order to study the cellular mode of renin secretion, stereological methods were used to estimate number and volume of rat renin secretory granules during stimulation of the renin system. An acute decrease in renal perfusion pressure to 40 mmHg for 5 min increased plasma renin concentration (PRC) twofold, but did not significantly change the number of renin granules per arteriole or the renin-containing volume of the arteriole. Chronic stimulation was achieved by a combination of low-salt diet and inhibition of angiotensin-converting enzyme (ACE) for 14 days, and resulted in a 36-fold increase in PRC, a 20-fold increase in the number of granules per arteriole, and a 17-fold increase in the arteriolar volume that contained renin. An acute decrease in renal perfusion pressure to 40 mmHg for 5 min in the chronically stimulated rats increased PRC further (1.6-fold), and significantly reduced the number of granules per arteriole by 4000 (45% reduction), but did not change the renin-containing arteriolar volume significantly. The average renin granule size was 0.35 μm3 with no significant differences among the groups. We conclude that recruited granular cells contribute significantly to renin release, and that all granular cells along the arteriole participate in secretory responses. The reduced number of renin granules after acute stimulation is compatible with exocytosis as the dominating mechanism of renin release.

The Involvement of Growth Hormone (GH), Insulin-Like Growth Factors (IGFs) and Vascular Endothelial Growth Factor (VEGF) in Diabetic Kidney Disease

At present, diabetic kidney disease affects about 15-25 % of patients with type 1 diabetes (T1D) and 30-40 % of patients with type 2 diabetes (T2D). Several decades of extensive research have elucidated various pathways to be implicated in the development of diabetic kidney disease. These include metabolic factors beyond blood glucose (e.g. advanced glycation endproducts (AGEs)), haemodynamic factors (e.g. the renin angiotensin system (RAS)), intracellular signaling molecule proteins (e.g. protein kinase C (PKC)) and growth factors/cytokines (e.g. growth hormone (GH), insulin-like growth factors (IGFs), transforming growth factor beta (TGF-beta) and vascular endothelial growth factor (VEGF)). This review focuses on the role of three of these growth factors, i.e. GH, IGFs and VEGF. A brief discussion of each system is followed by description of its expression in the normal kidney. Then, for each system, in vitro, experimental and clinical evidence addressing the role of the system in diabetic kidney disease is presented. The interplay of each system to other potential pathways will also be addressed. Finally, well-known and potential therapeutic strategies targeting the GH/IGF and VEGF systems in a specific or indirect way will discussed.