M.F. Cavanal

Long-term effects of maternal diabetes on vascular reactivity and renal function in rat male offspring.

Fetal growth impairment can occur in pregnancy complicated by diabetes. Although several studies have focused the effects of nutritional status on intrauterine development, the long-term impact of maternal diabetes on vascular and renal function in the offspring is poorly investigated. In the present study, blood pressure profiles and renal function parameters were investigated in the offspring of diabetic rats (DO). Female rats were made diabetic throughout gestation with a single dose of streptozotocyn (STZ) 10 d before mating. After weaning, the offspring had free access to food and water. Arterial pressure was evaluated every 15 d. Functional and morphometric kidney studies were performed in newborn, 3, 6 and 12-mo-old male rats in DO and in controls, C. Although maternal diabetes did not affect nephron number in the young adult rat, glomerular hypertrophy developed from 3 mo on. Glomerular Filtration Rate and Renal Plasma Flow were observed to be significantly decreased in DO when compared with C, from 3 mo on. In DO, hypertension was observed from 8 wk on and persisted elevated throughout the experimental period (12 mo). Vascular reactivity, evaluated in mesenteric arterial bed showed a decreased endothelium-dependent vasodilatation in 12-mo-old DO animals, while preserved response to sodium nitroprusside was demonstrated. Our data show that exposure to intrauterine diabetes induced by STZ does not affect nephron number in the young offspring but can cause permanent changes in Nitric Oxide (NO)-related vascular response, which, in turn may accelerate the natural age-related nephron loss.

The Influence of L-Arginine on Blood Pressure, Vascular Nitric Oxide and Renal Morphometry in the Offspring from Diabetic Mothers

The present study was designed to evaluate the effects of l-arginine (l-arg) supplementation on blood pressure, vascular nitric oxide content, and renal morphometry in the adult offspring from diabetic mothers. Diabetes mellitus was induced in female rats with a single dose of streptozotocin (50 mg/kg), before mating. The offspring was divided into four groups: group C (controls); group DO (diabetic offspring); group CA (controls receiving 2% l-arg solution dissolved in 2% sucrose in the drinking water) and group DA (DO receiving the l-arg solution). Oral supplementation began after weaning and continued until the end of the experiments. In DO, hypertension was observed, from 3 mo on. In DA, pressure levels were not different from C and CA. In 6-mo-old animals, basal NO production (assessed by DAF-2) was significantly depressed in DO in comparison to controls. The NO production was significantly increased after stimulation with Ach or BK in all groups, the increase being greater in control than in DO rats. l-Arg was able to improve the NO production and to prevent the glomerular hypertrophy in the DO. Our data suggest that the bioavailability of NO is reduced in the DO, because l-arg corrected both the hypertension and glomerular hypertrophy.

Effects of intrauterine food restriction and long-term dietary supplementation with L-arginine on age-related changes in renal function and structure of rats.

We have previously demonstrated that restricting intrauterine food by 50% in 3-mo-old rats produced lower nephron numbers and early-onset hypertension, the latter being normalized by l-arginine administration. In 18-mo-old rats, such restriction increased glomerulosclerosis. In this study, we expanded our investigation, evaluating functional, morphologic, and immunohistochemical parameters in intrauterine-food-restricted 18-mo-old rats, either receiving l-arginine (RA18) or not (R18). Age-matched, non-food-restricted controls were assigned to similar groups with l-arginine (CA18) and without (C18). After weaning, l-arginine was given daily for 17 mo. No functional or morphologic changes were observed in C18 rats. The R18 rats developed early-onset hypertension, which persisted throughout the observation period, as well as significant proteinuria from 12 mo on. In RA18 rats, l-arginine decreased both blood pressure levels and proteinuria, and glomerular diameter was significantly smaller than in R18 rats (115.63 ± 2.2 versus 134.8 ± 1.0 μm, p < 0.05). However, in RA18 rats, glomerular filtration rate remained depressed. Although l-arginine prevented glomerulosclerosis (R18 = 14%, RA18 = 4%; p < 0.05), glomerular expression of fibronectin and desmin was still greater in RA18 rats than in controls. Our data show that, although l-arginine prevented hypertension and proteinuria, glomerular injury still occurred, suggesting that intrauterine food restriction may be one of the leading causes of impaired renal function in adult life.

Effect of D-alpha-tocopherol on tubular nephron acidification by rats with induced diabetes mellitus

The objective of the present study was to determine if treatment of diabetic rats with D-alpha-tocopherol could prevent the changes in glomerular and tubular function commonly observed in this disease. Sixty male Wistar rats divided into four groups were studied: control (C), control treated with D-alpha-tocopherol (C + T), diabetic (D), and diabetic treated with D-alpha-tocopherol (D + T). Treatment with D-alpha-tocopherol (40 mg/kg every other day, ip) was started three days after diabetes induction with streptozotocin (60 mg/kg, ip). Renal function studies and microperfusion measurements were performed 30 days after diabetes induction and the kidneys were removed for morphometric analyses. Data are reported as means +/- SEM. Glomerular filtration rate increased in D rats but decreased in D + T rats (C: 6.43 +/- 0.21; D: 7.74 +/- 0.45; D + T: 3.86 +/- 0.18 ml min-1 kg-1). Alterations of tubular acidification observed in bicarbonate absorption flux (JHCO3) and in acidification half-time (t/2) in group D were reversed in group D + T (JHCO3, C: 2.30 +/- 0.10; D: 3.28 +/- 0.22; D + T: 1.87 +/- 0.08 nmol cm-2 s-1; t/2, C: 4.75 +/- 0.20; D: 3.52 +/- 0.15; D + T: 5.92 +/- 0.19 s). Glomerular area was significantly increased in D, while D + T rats exhibited values similar to C, suggesting that the vitamin prevented the hypertrophic effect of hyperglycemia (C: 8334.21 +/- 112.05; D: 10,217.55 +/- 100.66; D + T: 8478.21 +/- 119.81 microm(2)). These results suggest that D-alpha-tocopherol is able to protect rats, at least in part, from the harmful effects of diabetes on renal function.

Influence of Parathyroidectomy and Calcium on Rat Renal Function

Parathyroid hormone (PTH) has multiple effects on water and electrolyte transport along the nephron. However, the influences of PTH and calcium on the urinary concentration ability are not fully understood. In this study, clearance and microperfusion studies were performed in thyroparathyroidectomized (TPTX) rats either supplemented (TPTX+Ca2+) or not with calcium added to the ingested food as CaCl2 (1.6 g/100 g). Acid-base data and renal functional parameters were measured in TPTX and TPTX+Ca2+ rats. Additional studies were performed in the isolated inner medullary collecting tubules of intact and TPTX rats to evaluate the osmotic permeability of this segment in the presence of 10–6M PTH added to the bath. In these experiments the possible influence of PTH on antidiuretic hormone induced changes of the osmotic permeability in TPTX and TPTX+Ca2+ rats was also investigated. In the TPTX+Ca+ group, the glomerular filtration rate increased significantly when compared to the TPTX group (6.04 ± 0.42 vs. 4.88 ± 0.20 ml·min–1·kg–1; p < 0.05), but the U/P inulin ratio remained lower than control values (30.8 ± 1.48 vs. 54.0 ± 3.5; p < 0.05), which suggests that normal levels of PTH are necessary to maintain the concentrating ability. In a group of TPTX rats, an acute infusion of PTH (0.5 µg·min–1·kg–1) significantly decreased the urinary flow and increased the renal plasma flow, results that agree with the vasomodulator action of this hormone on the renal vasculature. A significant increase in the fractional K+ excretion observed in the TPTX+Ca2+ group as compared with both control and TPTX, groups suggests that the excreted load of Ca2+ may interfere with tubular K+ handling in the absence of PTH. PTH (10–6M) added to the bath of the isolated inner medullary collecting tubules did not change the osmotic permeability, of intact, TPTX, and TPTX+Ca2+ rats. Furthermore, it did not modify the antidiuretic hormone induced changes in the osmotic permeability. These results suggest that this segment of the nephron is PTH insensitive as far as water and ion transport are concerned.

Inflammatory milieu as an early marker of kidney injury in offspring rats from diabetic mothers.

The present study investigated the early presence of inflammatory response in renal tissue of young offspring from diabetic mothers. The effect of L-arginine (L-arg) supplementation was also investigated. The offspring was divided into four groups: group CO (controls); group DO (diabetic offspring); group CA (CO receiving 2% L-arg solution) and group DA (DO receiving the 2% L-arg solution). Glycemia, arterial pressure and renal function were evaluated; gene and protein expression of pro-inflammatory cytokines were also measured. Blood pressure levels were significantly increased in 2 and 6 month-old DO rats, whereas L-arg administration caused a significant decrease in the DA group, at both ages. DO rats showed a significantly blunted glycemic response to exogenous insulin. In 2 month-old DO animals, renal protein expression of pro-inflammatory molecules was significantly increased. At six months of age, we also observed an increase in gene expression of pro-inflammatory molecules, whereas L-arg supplementation prevented this increase at both ages. Our data suggest that activation of inflammatory pathways is present early in the kidney of DO rats, and that L-arg can attenuate the expression of these markers of tissue inflammation. Our results also reinforce the concept that intrauterine environmental factors are a fundamental determinant in the development of metabolic and vascular diseases later in life.

L-Arginine supplementation improves insulin sensitivity and beta cell function in the offspring of diabetic rats through AKT and PDX-1 activation

Maternal hyperglycemia can result in defects in glucose metabolism and pancreatic β-cell function in offspring. The purpose of this study was to evaluate the impact of maternal diabetes mellitus on pancreatic islets, muscle and adipose tissue of the offspring, with or without oral l-Arginine supplementation. The induction of diabetes was performed using streptozotocin (60mg/kg). Animals were studied at 3 months of age and treatment (sucrose or l-Arginine) was administered from weaning. We observed that l-Arg improved insulin sensitivity in the offspring of diabetic mothers (DA), reflected by higher insulin-induced phosphorylation of Akt in muscle and adipose tissue. Insulin resistance is associated with increased oxidative stress and the NADPH oxidase enzyme plays an important role. Our results showed that the augmented interaction of p47PHOX with gp91PHOX subunits of the enzyme in skeletal muscle tissue in the offspring of diabetic rats (DV) was abolished after l-Arg treatment in DA rats. Maternal diabetes caused alterations in the islet functionality of the offspring leading to increased insulin secretion at both low (2.8mM) and high (16.7mM) concentrations of glucose. l-Arg reverses this effect, suggesting that it may be an important modulator in the insulin secretory process. In addition it is possible that l-Arg exerts its effects directly onto essential molecules for the maintenance and survival of pancreatic islets, decreasing protein expression of p47PHOX while increasing Akt phosphorylation and PDX-1 expression. The mechanism by which l-Arg exerts its beneficial effects may involve nitric oxide bioavailability since treatment restored NO levels in the pancreas.

Role of thyroid hormone in protecting against gentamicin-induced nephrotoxicity

Abstract This study investigates the protective effect of thyroid hormone (T 4 ) in an in vivo model of gentamicin nephorotoxicity. Rats were divided into a control group and groups treated with either T 4 alone (C + T 4 ), gentamicin 40 mg/kg/d alone (G 40 ), or T 4 for 14 days with G 40 in the last 10 days (T 4 + G 40 ). The glomerular variables assessed showed similar results in groups G 40 and T 4 + G 40 (glomerular filtration rate, 5.31 ± 0.27 mL · min −1 · kg −1 with G 40 and 5.40 ± 0.23 mL · min −1 · kg −1 with T 4 + G 40 ; renal plasma flow, 14.87 ± 0.97 mL · min −1 · kg −1 with G 40 and 14.71 ± 0.97 mL · min −1 · kg −1 with T 4 + G 40 . Some values of tubular functions that were impaired in the group that received gentamicin alone were similar to the values in the group treated with T 4 + G 40 . Urine HCO 3 − concentration was 1.98 ± 0.44 mEq/mL/kg with G 40 , 0.85 ± 0.26 mEq/mL/kg with T 4 + G 40 , 0.879 ± 0.117 mEq/mL/kg with C + T 4 , and 0.882 ± 0.06 mEq/mL/kg in the control group; net acid excretion was 4.01 ± 0.11 μEq · min −1 · kg −1 with G 40 , 5.24 ± 0.34 μEq · min −1 · kg −1 with T 4 + G 40 , 5.47 ± 0.24 μEq · min −1 · kg −1 with C + T 4 , and 5.86 ± 0.29 μEq · min −1 · kg −1 in the control group; and sodium excretion was 10.32 ± 1.09 μEq · min −1 · kg −1 with G 40 , 4.72 ± 0.43 μEq · min −1 · kg −1 with T 4 + G 40 , 4.84 ± 0.20 μEq · min −1 · kg −1 for C + T 4 , and 7.12 ± 0.80 μEq · min −1 · kg −1 in the control group. These results suggest that T 4 protects the tubular function against epithelial injury but does not prevent the glomerular changes induced by gentamicin.

Aminoglycoside and nephrotoxicity.

The effect of three aminoglycosides--gentamicin, netilmicin and amikacin--on renal acid excretion was studied in male rats treated with doses equivalent to those clinically used. The amikacin and netilmicin groups showed no important changes in the values of glomerular filtration rate (GFR), effective renal plasma flow (ERPF) and U/P inulin ratio during normal and acidotic conditions. The gentamicin group, however, showed a clear tendency to decreases in these functional parameters even in normal conditions, a finding that reinforces the concept that gentamicin is more nephrotoxic than other aminoglycosides. During normal conditions net acid excretion (BH) did not change with any of the three tested drugs. However, after an acute acid load BH markedly fell regardless of the antibiotic used. The capacity to elevate the urine-blood pCO2 was preserved after an alcaline overload, suggesting that the distal tubule was not significantly affected by aminoglycoside treatment. These data suggest that the clinical use of aminoglycosides during metabolic acidosis deserves close attention due to the possible deleterious effect that can emerge as the result of an inappropriate retention of acid loads.

Parathyroidectomy enhances the nephrotoxicity of gentamicin

Abstract The objective of this study was to evaluate the effect of parathyroidectomy on the renal function in rats treated with saline (control), low-dose gentamicin (GM, 4 mg/kg per day), or high-dose GM (40 mg/kg per day) for 10 consecutive days. Gentamicin sulfate was dissolved in saline. A group of rats were subjected to thyroparathyroidectomy (PTX) 2 to 3 days before the beginning of the GM treatment. Thyroxine (1 μg/100 g body weight) was subcutaneously injected daily into PTX rats. In rats with saline infusion, a significant reduction in renal functional variables was obtained after PTX. When GM treatment and PTX were both present, a further impairment in the glomerular filtration rate and the urine/plasma inulin ratio was observed, suggesting additive effects. In rats infused with ammonium chloride to cause metabolic acidosis, a decline in glomerular filtration rate was obtained, a condition that was aggravated when GM treatment and PTX were both present. Acid excretion was significantly reduced in PTX groups, mainly during acidosis. In acidotic PTX rats treated with both low and high GM doses, the excreted amount of acid was strongly impaired, compared with the acidotic non-PTX GM group. These results show that in the PTX rats nephrotoxicity of GM is increased, which suggests that in this experimental model, parathyroid hormone may exert a protective effect during aminoglycoside treatment.