Susan E. Mulroney

Omega-3 fatty acid rich diet prevents diabetic renal disease

Omega-3 polyunsaturated fatty acids (n-3 PUFA) show beneficial effects in cardiovascular disease, IgA, and diabetic nephropathy; however, the mechanisms underlying these benefits are unknown. The study was performed in male Sprague-Dawley rats randomly divided into four treatment groups: nondiabetic (ND), streptozotocin-induced diabetic (D), diabetic and fed a high n-3 PUFA diet (D+canola), and diabetic and fed a high n-6 (omega-6) PUFA diet (D+corn). Study treatments were carried out for 30 wk. D+canola significantly decreased diabetes-associated increases in urine albumin excretion (ND 17.8 +/- 6.4; D 97.3 +/- 9.4; D+canola 8.3 +/- 2.2 mg/day); systolic blood pressure (ND 153 +/- 9; D 198 +/- 7; D+canola 162 +/- 9 mmHg); glomerulosclerosis (ND 0.6 +/- 0.2; D 1.8 +/- 0.2; D+canola 0.8 +/- 0.1 AU); and tubulointerstitial fibrosis in the renal cortex (ND 1.2 +/- 0.2; D 2.0 +/- 0.2; D+canola 1.1 +/- 0.1) and the inner stripe of the outer medulla (ND 1.0 +/- 0.2; D 2.1 +/- 0.2; D+canola 1.1 +/- 0.2 AU). D+corn also exerted renoprotection, but not to the same degree as D+canola (urine albumin excretion, 33.8 +/- 6.1 mg/day; systolic blood pressure, D+corn 177 +/- 6 mmHg; glomerulosclerosis, D+corn 1.2 +/- 0.3 AU; cortical tubulointerstitial fibrosis, D+corn 1.6 +/- 0.1 AU; medullary tubulointerstitial fibrosis, D+corn 1.5 +/- 0.1 AU). In addition, D+canola attenuated D-associated increase in collagen type I and type IV, IL-6, MCP-1, transforming growth factor-beta, and CD68 expression. These observations indicate a beneficial effect of high dietary intake of n-3 PUFA in reducing diabetic renal disease.

Impaired Nitric Oxide–Mediated Renal Vasodilation in Rats With Experimental Heart Failure Role of Angiotensin II

BACKGROUNDnCongestive heart failure (CHF) is associated with a decrease in renal perfusion. Because endothelium-derived NO is important in the regulation of renal blood flow (RBF), we tested the hypothesis that an impairment in the NO system may contribute to the decrease in RBF in rats with experimental CHF.nnnMETHODS AND RESULTSnStudies were performed in rats with experimental high-output CHF induced by aortocaval (AV) fistula and sham-operated controls. In controls, incremental doses of acetylcholine (ACh, 1 to 100 microg x kg(-1) x min(-1)) increased RBF and caused a dose-related decrease in renal vascular resistance (RVR). However, the increase in RBF and decrease in RVR were markedly attenuated in rats with CHF. Likewise, the effects of ACh on urinary sodium and cGMP excretion were also diminished in CHF rats, as was the renal vasodilatory effect of the NO donor S-nitroso-N-acetylpenicillamine (SNAP). These attenuated responses to endothelium-dependent and -independent renal vasodilators in CHF rats occurred despite a normal baseline and stimulated NO2+NO3 excretion and normal expression of renal endothelial NO synthase (eNOS), as determined by eNOS mRNA levels and immunoreactive protein. Infusion of the NO precursor L-arginine did not affect baseline RBF or the response to ACh in rats with CHF. However, administration of the nonpeptide angiotensin II receptor antagonist A81988 before ACh completely restored the renal vasodilatory response to ACh in CHF rats.nnnCONCLUSIONSnThis study demonstrates that despite a significant attenuation in the NO-related renal vasodilatory responses, the integrity of the renal NO system is preserved in rats with chronic AV fistula. This impairment in NO-mediated renal vasodilation in experimental CHF appears to be related to increased activity of the renin-angiotensin system and may contribute further to the decrease in renal perfusion seen in CHF.

Female Protection in Progressive Renal Disease Is Associated with Estradiol Attenuation of Superoxide Production

BACKGROUNDnSeveral types of renal disease progress at a faster rate in men compared with women, but the reasons for this sex difference are not well understood. Chronic renal disease is associated with elevated levels of toxic reactive oxygen species (ROS). Superoxide, the major ROS in the kidney, is generated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase.nnnOBJECTIVEnTo determine if female protection from renal disease progression is consistent with 17beta-estradiol (E2) attenuation of superoxide production, this study was conducted to assess superoxide production in the renal cortex of male and female control and renal wrap (RW) rats, as well as in ovariectomized rats treated with vehicle or E2.nnnMETHODSnSprague-Dawley rats were divided into 2 sham operation male (Sham-M) and female (Sham-F) control groups, and 4 RW hypertensive groups: RW-M; RW-F; RW ovariectomized females treated with vehicle (RW-OVX); and RW ovariectomized females treated with E2, supplied as a 0.24 mg/60-day release pellet (RW-OVX+E2). All groups were maintained on a high-sodium (4% NaCl) diet for 6 weeks.nnnRESULTSnMean (SEM) markers of renal injury and oxidative stress, including urinary protein (mg/24 h: RW-M, 298 [31] vs RW-F, 169 [22]; P < 0.001), microalbuminuria (RW/Sham arbitrary units [AU]/24 h: M, 8.78 [0.58] vs F, 4.31 [1.0]; P < 0.005), and malondialdehyde (nmol/24 h: RW-M, 167 [23] vs RW-F, 117 [8.5]; P < 0.05) levels, as well as mean glomerular volume (microm3 x 10(6): RW-M, 2.25 [0.16] vs RW-F, 1.25 [0.04]; P < 0.001) and the glomerulosclerotic index (AU: RW-M, 2.64 [0.19] vs RW-F, 1.10 [0.09]; P < 0.001) were greater in both control and RW males compared with females in the same treatment groups. Though RW surgery increased mean arterial pressure in both male and female rats, no sex difference was observed. Under these conditions, mean (SEM) renal cortical NADPH oxidase activity was 1.3-fold higher in RW males compared with RW females (relative light units [RLU]/180 sec: RW-M, 4080 [240] vs RW-F, 3200 [260]; P < 0.05). Ovariectomy increased NADPH oxidase activity by 1.4-fold (RLU/180 sec: RW-OVX, 4520 [184]; P < 0.01) under conditions in which the mean glomerular volume and glomerulosclerotic index were both increased by 1.5-fold, whereas E2 replacement (RLU/180 sec: RW-OVX+E2, 2745 [440]) prevented these effects. Furthermore, the effects on NADPH oxidase activity were mirrored by changes in the protein abundance of NADPH oxidase subunit p22P(phox).nnnCONCLUSIONnThese results suggest that E2 protects the female kidney in part by attenuating injury-induced increases in renal superoxide production.

The Insulin-Like Growth Factor Family of Peptides, Binding Proteins and Receptors: Their Potential Role in Tissue Regeneration

The insulin-like growth factors (IGF-I and IGF-II) are mitogenic pep tides that are structurally related to insulin (Figure 1). Until recently, these growth factors were thought to be produced exclusively by the liver and to act solely in an endocrine manner.1-3 According to the “somatomedin hypothesis,” the synthesis of IGF-I by the liver and its secretion are regulated by growth hormone (GH) and, following its release into the circulation, IGF-I reaches its target tissues where it induces growth and development, thereby mediating the effect of GH during the growth period.

Regulation of renal phosphate reabsorption during development: implications from a new model of growth hormone deficiency

It has been hypothesized that the high rate of renal phosphate (Pi) reabsorption in the immature animal is a consequence of the increased demand for Pi associated with the rapid rate of growth. Although growth hormone (GH) has been proposed to play a role in this process, investigations of the relationship between GH, growth and the renal Pi transport have been hampered by the lack of methods available to specifically alter circulating GH levels. This review summarizes the findings from recent studies using a newly developed peptidic antagonist to GH-releasing factor (GRF-AN) as a method of specifically inhibiting GH release. Systemic injection of GRF-AN was effective in suppressing the pulsatile release of GH, and in significantly attenuating the rate of growth, in both immature and adult rats. However, the inhibition of growth was associated with a reduction in net Pi retention only in immature rats, resulting in a doubling in the urinary excretion of Pi. GRF-AN treatment of immature rats lead to a decrease in the maximum tubular capacity to transport Pi-down to the level seen in adult rats. However, GRF-AN treatment did not alter renal Pi reabsorption in adult rats. We conclude that chronic administration of an antagonist to GRF in rats provides a new model of GH deficiency with which to study the interrelationships between growth, GH and other physiological systems. Furthermore, the findings suggest that the pulsatile release of GH, directly or indirectly, contributes to the high rate of renal Pi reabsorption in young, growing animals and may play a critical role in regulating Pi homeostasis during development.

Evidence for induction of a phosphate appetite in juvenile rats

This study examined whether dietary phosphate (Pi) restriction stimulates an appetite for Pi in the juvenile rat, which normally has a high metabolic Pi demand for growth. Juvenile Wistar rats were placed in individual cages with unrestricted access to tap water and a low (LPD, 0.02% Pi) or normal Pi diet (NPD, 0.6% Pi) for 7 days. On day 8, both groups of rats were given unlimited access to a solution of 0.3 M potassium phosphate water (PiH2O) for 8 additional days. Rats fed LPD consumed 70-100% more PiH2O then those rats fed NPD (P < 0.001). The increase in PiH2O intake resulted in a marked rise in the growth rate of rats fed LPD during days 8-15. A similar Pi intake was inducible after only 2 days of LPD and was associated with significant reductions in both plasma and cerebrospinal fluid (CSF) Pi levels; these levels remained low throughout Pi restriction, despite a significant PiH2O intake. Furthermore, the renal adaptation to enhance Pi reabsorption (TmPi) during Pi deprivation remained elevated despite enhanced PiH2O intake. Replenishment with a high-Pi diet rapidly quenched the PiH2O appetite and was associated with restoration of both plasma and CSF Pi levels. These findings suggest that an appetite for Pi can be induced in juvenile rats, perhaps through lowered plasma and CSF Pi levels. This behavioral response may serve as an additional mechanism to maintain an adequate supply of Pi necessary for growth and development of the animal.

The accumulation of IGF-I in kidneys of streptozotocin-diabetic adult rats is not associated with elevated plasma GH or IGF-I levels.

Nephropathy is a major complication of diabetes mellitus and is associated with expansion of the mesangium and an increase in kidney size in both humans and rats. Interestingly, early kidney enlargement occurs only in postpubertal animals, and is preceded by a significant increase in the levels of extractable renal IGF-I. This study examined the possibility that this difference is GH dependent, and that very early changes in plasma GH and/or IGF-I in the adult animal are associated with an early accumulation of renal IGF-I. Silastic jugular catheters were placed in adult (13–14 week) male Sprague-Dawley (S-D) rats for blood collection and drug injection. Serial blood samples were taken every 30 min in groups of saline control and streptozotocin (STZ) (50 μg/kg, IV) rats from 1–6 h, 9–15 h, and 24–30 h post-injection, and plasma GH profiles were determined by RIA. Renal IGF-I content was assessed following acid extraction. Following STZ, there was an immediate, step-wise reduction in peak GH levels (saline controls, 54±7 ng/mlvs 30±5 (1–6 h); 23±10 (9–15 h); and 13±3 ng/ml (24–30 h post-STZ);P<0.05 for all STZ groupsvs control). The same significant step-wise reduction was observed in the integrated area under the curve for GH. A separate group of rats were treated with a GH-releasing factor antagonist (GRF-AN) for 5 days prior to STZ, to suppress pulsatile GH release, and reduce plasma IGF-I. Chronic GRF-AN administration reduced plasma IGF-I levels significantly to 63% of control values (P<0.01). However, despite the reduction in plasma IGF-I, renal IGF-I remained significantly elevated 24 h post-STZ compared with controls and not significantly different from animals treated with STZ alone (467±49 ng IGF-I/g KW in control salinevs 778±100 in saline/STZ and 705±87 ng IGF-I/g KW in chronic GRF-AN/STZ rats (P<0.05)). In conclusion, following STZ administration in the adult rat, there is an immediate reduction in GH levels, indicating the renal IGF-I accumulation occurs without initial increases in plasma GH levels. Furthermore, when plasma IGF-I levels in the adult are significantly reduced renal IGF-I content remains elevated. These data suggest that early diabetic renal growth is not associated with elevated circulating GH levels, and that high basal plasma IGF-I levels are not necessary for IGF-I accumulation.

Induction of a Phosphate Appetite in Adult Male and Female Rats

We have reported that dietary inorganic phosphate (Pi) deprivation induces a Pi-seeking behavior in juvenile male rats. The purpose of the present study was to determine whether the Pi appetite is present in adult animals, and if so, whether it is altered during times of increased demand for Pi, such as pregnancy and lactation. Both male and female animals fed a low-phosphate diet (LPD) ingested significantly greater amounts of PiH20 daily than their normal phosphate diet (NPD) controls, and per 100 g of body weight (BW), the female animals fed LPD tended to ingest greater amounts of PiH20 than male rats fed LPD. Pregnant and lactating rats fed LPD ingested significantly more PiH20 than those fed NPD, however, neither group displayed a Pi appetite different than virgin females. However, lactation further reduced Pi levels in plasma and cerebral spinal fluid compared with control values. Despite the additional Pi from the PiH20 in the mothers fed LPD, pup birth weight was significantly lower than in NPD litters, and this was exacerbated 9 days after birth. This attenuated BW gain was associated with lower plasma Pi levels in the pups. In conclusion, a mild but consistent Pi-seeking behavior is induced in adult male and female rats after only 2 days of dietary Pi restriction. On a relative basis, the amount of PiH20 ingested is greater in female than in male animals, but does not increase further during pregnancy and lactation.

Changes in renal phosphate reabsorption in the aged rat.

Abstract Depletion of inorganic phosphate (Pi) reserves occurs frequently in aged animals and can result in diminished bone mineralization and osteoporosis. This altered Pi balance results from a reduction in intestinal Pi absorption and an elevation in renal Pi excretion. Since the kidney plays a central role in maintaining Pi homeostasis, we tested whether the increased phosphaturia seen with aging is a consequence of changes in the intrinsic tubular capacity to reabsorb Pi (TmPi). Male Wistar rats (12-, 18-, and 24-months-old) were acutely thyroparathyroidectomized (TPTX) and prepared for renal clearance studies in the presence and absence of fixed levels of parathyroid hormone (synthetic PTH-(1–34), 1U/kg/min). The maximum capacity for Pi transport (TmPi) was assessed by infusion of Pi at progressively higher rates (0–6 μmol/min) to increase the filtered load of Pi and facilitate the determination of the TmPi. TmPi declined significantly with age (3.51 ± 0.12 vs 3.04 ± 0.19 vs 2.30 ± 0.18 μmol/ml, for 12-, 18-, and 24-month-old rats, respectively, P < 0.05) in TPTX rats. Administration of PTH markedly reduced the TmP, in all age groups. Although the TmP, attained was similar among the age groups (1.15 ± 0.13 vs 1.15 ± 0.06 vs 1.03 ± 0.09 μmol/ml, for 12-, 18-, and 24-month-old rats, respectively), the magnitude of the reduction in the presence of PTH declined from 67% in 12-month-old rats to 62% and 55% in 18- and 24-month-old rats, respectively. These results demonstrate that aging is associated with a PTH-independent decrease in the intrinsic capacity of the kidney to reabsorb phosphate. Further, the kidney of the aged rat can respond to a pharmacological dose of PTH with appropriate reductions in the TmP, although the magnitude of the response declines with age.

Restoration of a phosphaturic response to parathyroid hormone in the immature rat.

ABSTRACT: Recent studies have shown that immature rats display a diminished sensitivity to the phosphaturic effects of parathyroid hormone (PTH), and that the responsiveness to PTH increases with age. The attenuated phosphaturia may reflect an inability of the neonate to respond to the hormone because of functional immaturity of the developing kidney. Alternatively, PTH may actually inhibit tubular phosphate reabsorption in the neonate but, due to other phosphate conservation mechanisms, no phosphaturia occurs. Our objective was to determine whether a phosphaturic response to PTH would be elicited in immature rats during infusion of moderate amounts of phosphate (Pi). Clearance experiments were performed on 26 acutely thyroparathyroidectomized immature Wistar rats (3-5 wk of age) fed a normal Pi diet (0.63%). In response to infusion of either Pi (1 µmol/min · 100 g) (group I) or PTH (8.3 ng/min · 100 g) (group II) alone, the fractional excretion of phosphate rose minimally (from 0.01 ± 0.01% to 4.9 ± 1.9% and from 0.12 ± 0.12% to 2.9 ± 1.4% for groups I and II, respectively). However, when Pi and PTH were combined either Pi first followed by PTH (group III) or PTH first followed by Pi (group IV), the fractional excretion of Pi rose dramatically (from 0.01 ± 0.01 to 21.8 ± 3.5% and from 0.04 ± 0.04 to 27.7 ± 3.3% for groups III and IV, respectively). A significant increase in urinary cAMP excretion occurred during infusion of PTH even when Pi excretion was minimal, but there was no further increase in urinary cAMP during the combined infusion of Pi and PTH. These results indicate that a phosphaturic response to PTH can be elicited in immature rats during infusion of moderate amounts of phosphate, and that the phosphaturia is dissociated from the increase in urinary excretion of cAMP. Thus, the attenuated phosphaturic response to PTH seen in developing rats is not due to functional immaturity of the kidney, but rather could be attributed to other factors that promote Pi reabsorption and prevent the phosphaturic effect from being expressed.