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Population studies have shown an association between diabetic nephropathy (DN) and insertion/deletion (I/D) polymorphism of the angiotensin-converting enzyme (ACE) gene (ACE in humans, Ace in mice). The aim was to evaluate the modulation of Ace copies number and diabetes mellitus (DM) on renal RAS and correlate it with indicators of kidney function. Increased number of copies of the Ace gene, associated with DM, induces renal dysfunction. The susceptibility to the development of DN in 3 copies of animals is associated with an imbalance in activity of RAS enzymes leading to increased synthesis of Ang II and Ang-(1–7). Increased concentration of renal Ang-(1–7) appears to potentiate the deleterious effects triggered by Ang II on kidney structure and function. Results also show increased bradykinin concentration in 3 copies diabetic group. Taken together, results indicate that the deleterious effects described in 3 copies diabetic group are, at least in part, due to a combination of factors not usually described in the literature. Thus, the data presented here show up innovative and contribute to understanding the complex mechanisms involved in the development of DN, in order to optimize the treatment of patients with this complication.

Sepsis is an uncontrolled systemic inflammatory response against an infection and a major public health issue worldwide. This condition affects several organs, and, when caused by Gram-negative bacteria, kidneys are particularly damaged. Due to the importance of renin-angiotensin system (RAS) in regulating renal function, in the present study, we aimed to investigate the effects of endotoxemia over the renal RAS. Wistar rats were injected with Escherichia coli lipopolysaccharide (LPS) (4 mg/kg), mimicking the endotoxemia induced by Gram-negative bacteria. Three days after treatment, body mass, blood pressure, and plasma nitric oxide (NO) were reduced, indicating that endotoxemia triggered cardiovascular and metabolic consequences and that hypotension was maintained by NO-independent mechanisms. Regarding the effects in renal tissue, inducible NO synthase (iNOS) was diminished, but no changes in the renal level of NO were detected. RAS was also highly affected by endotoxemia, since renin, angiotensin-converting enzyme (ACE), and ACE2 activities were altered in renal tissue. Although these enzymes were modulated, only angiotensin (ANG) II was augmented in kidneys; ANG I and ANG 1-7 levels were not influenced by LPS. Cathepsin G and chymase activities were increased in the endotoxemia group, suggesting alternative pathways for ANG II formation. Taken together, our data suggest the activation of noncanonical pathways for ANG II production and the presence of renal vasoconstriction and tissue damage in our animal model. In summary, the systemic administration of LPS affects renal RAS, what may contribute for several deleterious effects of endotoxemia over kidneys.

The present study investigated the angiotensin II (Ang II) responses in rat femoral veins taken from 2-kidney-1clip (2K1C) hypertensive rats at 4 weeks after clipping, as well as the effects of exercise on these responses. In this manner, femoral veins taken from 2K1C rats kept at rest or exposed to acute exercise or to exercise training were challenged with Ang II or endothelin-1 (ET-1) in organ bath. Simultaneously, the presence of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) were determined in these preparations by western blotting. In these experiments, femoral veins exhibited subdued Ang II responses. However, after nitric oxide (NO) synthesis blockade, the responses were higher in the femoral veins taken from animals kept at rest [0.137(0.049–0.245); n = 10] than those obtained in trained animals kept at rest [0.008(0.001–0.041); n = 10] or studied after a single bout of exercise [0.001(0.001–0.054); n = 11]. In preparations in which, in addition to NO synthesis, both the local production of prostanoids and the action of ET-1 on type A (ETA) or B (ETB) receptors were inhibited, the differences induced by exercise were no longer observed. In addition, neither ET-1 responses nor the presence of COX-1 and COX-2 in these preparations were modified by the employed exercise protocols. In conclusion, NO maintains Ang II responses reduced in femoral veins of 2K1C animals at rest. However, vasodilator prostanoids as well as other relaxing mechanisms, activated by ETB stimulation, are mobilized by exercise to cooperate with NO in order to maintain controlled Ang II responses in femoral veins.

The mechanisms underlying the link between high constitutive levels of ACE and diabetic nephropathy has not been completely understood, but an imbalance between angiotensin I (ACE) and II (ACE2) converting enzymes homeostasis has been described in diabetic kidney disease. The aim of this study was to evaluate ACE/ACE2 homeostasis in kidney from diabetic mice presenting increased dosage of ACE gene. Male mice (3 months old) genetically engineered to harbor one or three copies of the ACE gene were made diabetic (streptozotocin - STZ, 50 mg/Kg) and randomly assigned into: 1-copy control (1CC), 1-copy diabetic (1CD), 3-copy control (3CC) and 3-copy diabetic. At the end of experimental period body weight was evaluated and kidney was excised. Kidney-to-body weight ratio and ACE and ACE 2 activities were determined using specific substrates (ZPhe-HL and 7-Mca-APK(Dnp), respectively) (Two way ANOVA + Tukey test; P<0.05). Diabetes increased blood glucose (1CD : 436 ± 25 vs. 1CC: 90 ± 2; 3CD: 556 ± 6 vs. 3CC: 112 ± 4 mg/dL) and kidney-to-body weight ratio (1CD: 7.5 ± 0.2 vs. 1CC: 5.8 ± 0.2; 3CD: 7.8 ± 0.1 vs. 3CC: 5.8 ± 0.1 mg/g) with no influence of ACE genotype. As expected, renal ACE activity was directly related to ACE gene copy number in control group (3CC: 9.4 ± 2.11 vs. 1CC:5.6 ± 0.9 mU/mg protein). Renal ACE activity was decreased in diabetic groups (1CD: 3.6 ± 0.2 vs. 1CC: 5.6 ± 0.9; 3CD: 2.3 ± 0.4 vs. 3CC: 9.4 ± 2.1 mU/mg protein) with no influence of ACE genotype. Under physiological condition, renal ACE2 activity remained unchanged regardless of the ACE genotype (1CC: 1.9 ± 0.2 = 3CC: 1.4 ± 0.1 μM/min/mg). However upon a pathological stimulus, renal ACE2 activity was efficiently increased only in 1CD group, but not in 3CD, as compared with the others (1CD: 5.1 ± 0.9 vs. 1CC: 1.9 ± 0.2 = 3CC: 1.4 ± 0.1 = 3CD: 2.2 ± 0.2 μM/min/mg). Taken together, our results show for the first time, that susceptibility for the development of diabetic nephropathy associated with increased ACE gene dosage may be, at least in part, caused by a decrease on renal ACE2 activity. This may result in increased local levels of angiotensin II and decreased angiotensin (1-7), leading to altered glomerular permeability and albuminuria, functional alterations presented by 3CD animals. Financial Support: FAPESP, CAPES, CNPq.

The aim of this study is to analyze the catecholamines (CA) profile in mesangial cells (MC) from normal and diabetic animals. The diabetes mellitus has been induced in Wistar‐Hannover by injection of streptozotocin (50mg/kg). Three groups of study ‐ Control (CT), Hyperglycemic (HG) and treated Diabetic (TD), were evaluated during 60 days. MCs were obtained through sieve fractionation and collagenase‐treatment, and collected in the third subculture. CA concentrations were determined by high performance liquid chromatography. CA concentrations in the MC extracellular medium (EM) and intracellular compartment (IC) of the CT group expressed in pg/mg of cellular protein were respectively: Norepinephrine (NE), 18.1±1.1; Epinephrine (EP), 10.0±0.6; Dopamine (DE), and NE, 117.3±10.1; AD, 62.6±9.4; DE, 39.1±7.7. The levels of CA detected for HG in the EM were NE, 43.5±5.8; EP, 149.0±33.6; DE, 57.9±11.2 and in the IC were NE, 109.4±41.3; EP, 269.6±22,3; DE, 189.1±48.9. The comparison between groups demonstrated statistically significant differences in the CA levels of TD‐HG and for CT‐HG (TD and CT had a similar behavior). Thus, we can suggest that besides the renal damage induced directly by high blood glucose levels, DM could disturb CA metabolism and contribute to the development of diabetic nephropathy. Supported by FAPESP 02/13290‐2.

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