High glucose concentration stimulates intracellular renin activity and angiotensin II generation in rat mesangial cells

Increased intrarenal renin-angiotensin system activity contributes to diabetic nephropathy. ANG II generation in mesangial cells (MC) is increased by high-glucose (HG) exposure. This study assessed the mechanisms involved in the glucose-induced ANG II generation in rat MC. Under basal conditions, MC mainly secreted prorenin. HG decreased prorenin secretion and induced a striking 30-fold increase in intracellular renin activity. After 72 h of HG exposure, only the mRNA levels for angiotensinogen and angiotensin-converting enzyme (ACE) were significantly elevated. However, after shorter periods of 24 h of HG stimulation the mRNA levels of the enzymes prorenin and cathepsin B, besides that for ACE, were significantly increased. the results suggest that the HG-induced increase in ANG II generation in MC results from an increase in intracellular renin activity mediated by at least three factors: a time-dependent stimulation of ( pro) renin gene transcription, a reduction in prorenin enzyme secretion, and an increased rate of conversion of prorenin to active renin, probably mediated by cathepsin B. the increase in angiotensinogen mRNA in parallel to increased renin activity indicates that HG also increased the availability of the renin substrate. the consistent upregulation of ACE mRNA suggests that, besides renin, ACE is directly involved in the increased mesangial ANG II generation induced by HG.UNIFESP, Div Renal, BR-04023900 São Paulo, BrazilUNIFESP, Div Renal, BR-04023900 São Paulo, BrazilWeb of Scienc

Receptor-Induced Dilatation in the Systemic and Intrarenal Adaptation to Pregnancy in Rats

Normal pregnancy is associated with systemic and intrarenal vasodilatation resulting in an increased glomerular filtration rate. This adaptive response occurs in spite of elevated circulating levels of angiotensin II (Ang II). In the present study, we evaluated the potential mechanisms responsible for this adaptation. The reactivity of the mesangial cells (MCs) cultured from 14-day-pregnant rats to Ang II was measured through changes in the intracellular calcium concentration ([Cai]). The expression levels of inducible nitric oxide synthase (iNOS), the Ang II-induced vasodilatation receptor AT2, and the relaxin (LGR7) receptor were evaluated in cultured MCs and in the aorta, renal artery and kidney cortex by real time-PCR. The intrarenal distribution of LGR7 was further analyzed by immunohistochemistry. The MCs displayed a relative insensitivity to Ang II, which was paralleled by an impressive increase in the expression level of iNOS, AT2 and LGR7. These results suggest that the MCs also adapt to the pregnancy, thereby contributing to the maintenance of the glomerular surface area even in the presence of high levels of Ang II. The mRNA expression levels of AT2 and LGR7 also increased in the aorta, renal artery and kidney of the pregnant animals, whereas the expression of the AT1 did not significantly change. This further suggests a role of these vasodilatation-induced receptors in the systemic and intrarenal adaptation during pregnancy. LGR7 was localized in the glomeruli and on the apical membrane of the tubular cells, with stronger labeling in the kidneys of pregnant rats. These results suggest a role of iNOS, AT2, and LGR7 in the systemic vasodilatation and intrarenal adaptation to pregnancy and also suggest a pivotal role for relaxin in the tubular function during gestation

Mesenchymal stem cells (MSC) prevented the progression of renovascular hypertension, improved renal function and architecture

Renovascular hypertension induced by 2 Kidney-1 Clip (2K-1C) is a renin-angiotensin-system (RAS)-dependent model, leading to renal vascular rarefaction and renal failure. RAS inhibitors are not able to reduce arterial pressure (AP) and/or preserve the renal function, and thus, alternative therapies are needed. Three weeks after left renal artery occlusion, fluorescently tagged mesenchymal stem cells (MSC) (2×10(5) cells/animal) were injected weekly into the tail vein in 2K-1C hypertensive rats. Flow cytometry showed labeled MSC in the cortex and medulla of the clipped kidney. MSC prevented a further increase in the AP, significantly reduced proteinuria and decreased sympathetic hyperactivity in 2K-1C rats. Renal function parameters were unchanged, except for an increase in urinary volume observed in 2K-1C rats, which was not corrected by MSC. The treatment improved the morphology and decreased the fibrotic areas in the clipped kidney and also significantly reduced renal vascular rarefaction typical of 2K-1C model. Expression levels of IL-1β, TNF-α angiotensinogen, ACE, and Ang II receptor AT1 were elevated, whereas AT2 levels were decreased in the medulla of the clipped kidney. MSC normalized these expression levels. In conclusion, MSC therapy in the 2K-1C model (i) prevented the progressive increase of AP, (ii) improved renal morphology and microvascular rarefaction, (iii) reduced fibrosis, proteinuria and inflammatory cytokines, (iv) suppressed the intrarenal RAS, iv) decreased sympathetic hyperactivity in anesthetized animals and v) MSC were detected at the CNS suggesting that the cells crossed the blood-brain barrier. This therapy may be a promising strategy to treat renovascular hypertension and its renal consequences in the near future.Coordenação de Aperfeiçoamento de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Effect of renal ischemia/reperfusion on gene expression of a pH-sensitive K+ channel

Background: Sodium reabsorption depends on the Na/K/ATPase activity coupled to basolateral K+ recycling through K+ channels. ATP depletion reduces pump activity and increases K+ leak resulting in transport dysfunction. Kir4.1 is a pH-sensitive K+ channel expressed in the basolateral membrane of distal tubules. in this study, we evaluated whether Kir4.1 is also expressed in proximal tubules (PTs) and whether renal ischemia alters Kir4.1 mRNA expression levels. Methods: the presence of Kir4.1 mRNA was evaluated in PTs microdissected from collagenase-treated rat kidneys. Kir4.1 expression levels were estimated in the renal cortex by multiplex RT-PCR after 30 or 60 min of renal ischemia followed by 1, 24, 48 or 72 h of reperfusion. Results: the PCR product obtained from isolated tubules was sequenced and showed similar to 98% homology with rat Kir4.1 cDNA. Ischemia/reperfusion for 30 min induced a time-dependent reduction in Kir4.1 mRNA expression in parallel with plasma creatinine, however recovery was delayed after 60 min of ischemia, remaining reduced after 72 h of reperfusion when plasma creatinine was already normalized. Conclusion: Kir4.1 mRNA expression was decreased by renal ischemia. the ischemia-induced cellular K+ loss may be minimized by Kir4.1 down-regulation and may contribute to the mechanism by which cellular acidification induces cell protection against ATP depletion. Copyright (c) 2007 S. Karger AG, BaselUniversidade Federal de São Paulo, Disciplina Nefrol, Rua Botucatu 740, BR-04023900 São Paulo, SP, BrazilUniversidade Federal de São Paulo, Disciplina Nefrol, Rua Botucatu 740, BR-04023900 São Paulo, SP, BrazilWeb of Scienc

Effect of renal ischemia/reperfusion on gene expression of a pH-sensitive K+ channel

Background: Sodium reabsorption depends on the Na/K/ATPase activity coupled to basolateral K+ recycling through K+ channels. ATP depletion reduces pump activity and increases K+ leak resulting in transport dysfunction. Kir4.1 is a pH-sensitive K+ channel expressed in the basolateral membrane of distal tubules. in this study, we evaluated whether Kir4.1 is also expressed in proximal tubules (PTs) and whether renal ischemia alters Kir4.1 mRNA expression levels. Methods: the presence of Kir4.1 mRNA was evaluated in PTs microdissected from collagenase-treated rat kidneys. Kir4.1 expression levels were estimated in the renal cortex by multiplex RT-PCR after 30 or 60 min of renal ischemia followed by 1, 24, 48 or 72 h of reperfusion. Results: the PCR product obtained from isolated tubules was sequenced and showed similar to 98% homology with rat Kir4.1 cDNA. Ischemia/reperfusion for 30 min induced a time-dependent reduction in Kir4.1 mRNA expression in parallel with plasma creatinine, however recovery was delayed after 60 min of ischemia, remaining reduced after 72 h of reperfusion when plasma creatinine was already normalized. Conclusion: Kir4.1 mRNA expression was decreased by renal ischemia. the ischemia-induced cellular K+ loss may be minimized by Kir4.1 down-regulation and may contribute to the mechanism by which cellular acidification induces cell protection against ATP depletion. Copyright (c) 2007 S. Karger AG, BaselUniversidade Federal de São Paulo, Disciplina Nefrol, Rua Botucatu 740, BR-04023900 São Paulo, SP, BrazilUniversidade Federal de São Paulo, Disciplina Nefrol, Rua Botucatu 740, BR-04023900 São Paulo, SP, BrazilWeb of Scienc

The effect of Phyllanthus niruri on urinary inhibitors of calcium oxalate crystallization and other factors associated with renal stone formation

Objective To evaluate the effect of an aqueous extract of Phyllanthus niruri (Pn), a plant used in folk medicine to treat lithiasis. on the urinary excretion of endogenous inhibitors of lithogenesis, citrate, magnesium and glycosaminoglycans (GAGs).Materials and methods the effect of chronic (42 days) administration of Pn (1.25 mg/mL/day, orally) was evaluated in a rat model of urolithiasis induced by the introduction of a calcium oxalate (CaOx) seed into the bladder of adult male Wistar rats. the animals were divided into four groups: a sham control (16 rats): a control+Pri (six): CaOx+water instead of Pn (14): and CaOx+Pn (22). Plasma and urine were collected after 42 days of treatment for biochemical analysis and the determination of urinary excretion of citrate, magnesium and GAGs. the animals were then killed and the calculi analysed.Results the creatinine clearance or urinary and plasma concentrations of Na+, K+, Ca2+, oxalate, phosphate and uric acid were unaffected by Pn or the induction of lithiasis. Treatment with Pn strongly inhibited the growth of the matrix calculus and reduced the number of stone satellites compared with the group receiving water. the calculi were eliminated or dissolved in some treated animals (three of 22). the urinary excretion of citrate and magnesium was unaffected by Pn treatment. However, the mean (si)) urinary concentration of GAGs was significantly lower in rats treated with CaOx + Pn, at 5.64 (0.8 6) mg/g crcatinine. than when treated with CaOx+water, at 11.78(2.21)mg/g creatinine. in contrast, the content of GAGs in the calculi was higher in the CaOx + Pn rats. at 48.0 (10.4) g/g calculus. than in the CaOx+water group, at 16.6 (9.6) g/g calculus.Conclusion These results show that Pn has an inhibitory effect on crystal growth. which is independent of changes in the urinary excretion of citrate and Mg. but might be related to the higher incorporation of GAGs into the calculi.Universidade Federal de São Paulo, Escola Paulista Med, Div Nephrol, São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Div Nephrol, São Paulo, BrazilWeb of Scienc

ACUTE, SUBACUTE, and CHRONIC X-RAY-EFFECTS ON GLOMERULAR HEMODYNAMICS in RATS

In order to evaluate the effects of x-rays on glomerular hemodynamics, surgically exposed left kidneys of Munich-Wistar rats were irradiated with 15 Gy in a single dose. the animals were studied 45 min (acute group, n = 8), 14 days (subacute group, n = 7), and 60 days (chronic group, n = 7) after irradiation and compared with their respective controls. A decrease in total glomerular filtration (55%) and renal plasma flow (40%) rates with marked elevation of total renal vascular resistance (180%), p < 0.05, occurred within 45 min. Significant changes also occurred in the microcirculation; i.e., single-nephron glomerular filtration (SNGFR), glomerular plasma flow (Q(A)), and glomerular capillary hydraulic pressure (P-GC) declined by 35%, 40%, and 12%, respectively, due to an increase in total arteriolar resistance (90%), p < 0.05. Within 14 days, SNGFR was similar to control in spite of a moderate elevation of afferent arteriolar resistance (26%) and reduction in P-GC (11%), p < 0.05, and Q(A) (20%). K-f was significantly elevated (46%), p < 0.05. the chronic group presented a response pattern similar to that of the acute group, although less severe. Histopathological changes were not relevant and were restricted to tubules. the present results suggest that: (a) Acutely, there was a marked reduction in filtration, flow, and P-GC with significant elevation of resistances. (b) Within 14 days, the maintenance of SNGFR was probably the result of an offsetting effect between Q(A) and P-GC decreases and K-f elevation. (c) After 60 days, the homeostatic mechanism was not sufficient to maintain normal renal function. (d) A functional effect is probably the most important pathogenetic mechanism, at least during the initial phase, for the development of radiation nephropathy since no morphological alterations were observed.ESCOLA PAULISTA MED,DEPT MED,DIV NEPHROL,BR-04023062 São Paulo,BRAZILESCOLA PAULISTA MED,DEPT MED,DIV NEPHROL,BR-04023062 São Paulo,BRAZILWeb of Scienc