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In type 1 diabetes mellitus (T1DM) there is increased prorenin secretion by the principal cells of the collecting duct. Binding of prorenin to prorenin receptor (PRR) on intercalated cells increases its catalytic activity, increases local angiotensin (Ang) II formation, and stimulates intracellular MAPK signaling responsible for inflammation and tissue fibrosis. Thus, changes in the amount of membrane bound PRR may be a key factor in stimulating these pathways. However, it has not been established that activation of PRR in the collecting duct contributes to increased intrarenal Ang II and tubulointerstitial inflammation via stimulation of inflammatory pathways including transforming growth factor-beta (TGF-β). This study tested the hypothesis that hyperglycemia increases the PRR abundance at the plasma membrane (PM) in the collecting duct cells, thus allowing greater capability to be activated by locally produced prorenin. Streptozotocin (STZ; 60 mg/kg; ip single dose) was used to induce T1DM in Sprague-Dawley rats (N=10) and compared to control rats (N=8). After 7-days induction, STZ-rats showed plasma glucose levels of 428±13 vs. 138±9 mg/dL and insulin of 0.05±0.02 vs. 2.4±0.6 ng/mL, compared to control. Although PRR transcript in the renal medulla were not different between groups; PRR localized predominantly on the apical aspects of collecting duct cells in STZ-induced rats; while in controls it was primarily found intracellularlly. These changes were accompanied by greater levels of active renin and Ang II in the urine and increased TGF-β mRNA levels in the renal medulla of STZ-rats (Renin: 186± 34 vs. 6± 3 ng Ang I/mL/h; P<0.01; Ang II: 884± 147 vs. 42± 14 fmol/h; P<0.05; TGF-β: 1.22 ± 0.06 vs. 0.97 ± 0.03 mRNA ratio; P<0.01). To further assess if hyperglycemia induced in vitro PRR trafficking alterations, collecting duct M-1 cells were treated with normal glucose (NG; 1mM glucose + 1 mM mannitol) and high glucose (HG; 4mM) for 5, 60, and 360 min. PRR protein levels were higher in the PM fractions in cells treated with HG, compared to cells treated with NG. Thus, hyperglycemia increases PRR abundance in the PM of the collecting duct and stimulates TGF-β synthesis in the renal medulla which may underlie the development of tubulointerstitial inflammation. [ABSTRACT FROM AUTHOR]

Prior study of our group shown that previous aerobic exercise training improved the damage caused by diabetes mellitus on renal and cardiovascular system. Resistance exercise training, also known as strength training, is traditionally performed to gain muscle mass; however, it is not clear whether this type of exercise modulates renal system. Additionally, it is also unknown whether previous resistance exercise training can potentially influence the kidney and skeletal muscle. Wistar rats were submitted to resistance exercise training in an apparatus developed especially to this type of exercise (8 - 12 climbs/day, 5 days/week, 12 weeks). Previous resistance exercise trained group (PTD) performed for 4 weeks before the establishment of the disease and after this period they were followed by 8 weeks of resistance exercise training. Additional trained groups such as trained diabetic (TD) and trained control (TC) groups were followed by 8 weeks of resistance exercise training. Control groups were also followed (control - C, diabetes - D). We have found that PTD suppressed abnormalities linked to renal system such as, water consumption and amount of urine PTD=71mL vs. DT=127mL vs. D=138mL (measured during metabolic cage period), as well as attenuated proteinuria and kidney weight. Regarding to skeletal muscle, PTD group had increased muscle weight (extensor digitorium longus - EDL; C=192mg, D=116mg, TD=106mg and PTD=126mg; Tibialis anterior, C=780mg, D=496mg, DT=450mg and PTD=535mg); we also found a great muscle force level in the PTD group (C=573g, CT=1037, D=414g, TD=737g and PTD=825g), suggesting a protective effect of previous exercise in this group. PTEN was suppressed in PTD group and Akt and 4EBP1 (upstream and downstream of mTOR) were activated in PTD group, measured by western blot. These data suggest that, resistance exercise performed prior the establishment of the diabetes mellitus can protect kidney from diabetic nephropathy and skeletal muscle from atrophy. The mechanisms by which kidney and skeletal muscle have been improved are linked to mTOR signaling pathway. Further studies will be performed to confirm the potential involvement of this signaling pathway.Support: FAPESP, CAPES, CNPq. [ABSTRACT FROM AUTHOR]

Overactivity of the intrarenal renin-angiotensin system (RAS) has been implicated in the pathogenesis of diabetic nephropathy. Our group has already demonstrated that aerobic exercise reduces kidney angiotensin II levels and attenuates renal dysfunction under concurrence of diabetes and hypertension. Resistance training (RT) has recently been recognized as a useful therapeutic tool for the treatment chronic diseases and similar to aerobic exercise, has been reported to improve glycemic control. Therefore, the aim of this study was to evaluate the effect of RT on renal function and RAS in diabetic animals, to understand whether this type of exercise is also associated with renoprotection. Wistar rats (3 months old) were randomized into: sedentary control (SC); trained control (TC); sedentary diabetic (SD) and trained diabetic (TD). Animals were made diabetic with a single tail injection of streptozotocin (STZ, 50 mg/Kg). RT was performed on an 110-cm ladder (8 ladder climbs, once/day, 5 days/week, 8 weeks), carrying a load of 50-80% body weight (BW) appended to the tail. At week 8, 24 hr urine volume and albuminuria were evaluated. Kidney was excised and ACE and ACE2 activities were determined (ZPhe-HL and 7-Mca-APK(Dnp), respectively) (Two way ANOVA + Tukey test; P<0.05). RT significantly reduced blood glucose (TD = 449 ± 17 vs. SD = 572 ± 18 mg/dL) and attenuated BW loss of diabetic animals. DM reduced renal ACE activity in sedentary and trained groups (SD = 3.72 ± 0.48, TD = 3.85 ± 0.40 vs. SC = 9.2 ± 0.59 nmol/min/mg), while RT reduced enzyme activity only in control group (TC = 5.14 ± 0.26 vs. SC = 9.2 ± 0.59 nmol/min/mg). RT reduced renal ACE2 in the control group compared to the others (TC = 0.05 ± 0.0001 vs. SC = 0.09 ± 0.004, SD = 0.09 ± 0.003, TD = 0.10 ± 0.002 μM/min/mg), with no effect of diabetes on enzyme activity. RT improved renal function, decreasing urinary volume and albuminuria (DT = 4.13 ± 0.84 vs. SD = 11 ± 2.11 mg/24h) in DT group. The results from the present study show that RT is strongly associated with renoprotection in an experimental model of diabetic nephropathy. Moreover, results show that this improvement on renal function is modulated by other pathways apart from ACE and ACE2 converting enzymes. Financial Support: FAPESP, CAPES, CNPq. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

High prorenin levels in the circulation are associated with the progression of diabetic nephropathy and microvascular changes in diabetes. In spite of high plasma prorenin, the urine of diabetes mellitus patients exhibits very low pH and contains high levels of “active” renin, but low levels of prorenin. Prorenin binding to the prorenin receptor (PRR) results in non-proteolytic activation of prorenin and stimulation of intracellular synthesis of transforming growth factor-beta (TGF-β). Furthermore, the PRR cytosolic domain is associated with the vacuolar H+-ATPase, which is involved in acid-base homeostasis by renal intercalated cells. In diabetes, the collecting duct is the major source of prorenin and PRR is upregulated in the kidneys of diabetic rats. We examined the relationships among urine levels of renin (uRen), prorenin (uPro), soluble PRR (sPRR) and TGF-β (uTGF-β) in 93 subjects, including non-diabetes mellitus (Non-DM: n=64); and type-2 diabetes mellitus (DM; n=29) patients. Urine albumin (uAlb), creatinine (uCr) and pH were also assessed. DM patients showed higher uRen levels (DM: 81±21 vs Non-DM: 29±4.5 ng AngI/mL/hr; p<0.001), which were positively associated with uAlb/uCr (p<0.05, r=0.21) and inversely correlated with urine pH (p<0.01, r=-0.38). These values contrasted with uPro levels, which were no different between groups. DM patients showed increased uTGF-β compared to Non-DM (DM: 22±5.3 vs Non-DM: 5.6±1.5 pg/mg uCr; p<0.0001). Urine sPRR levels were positively correlated with urine pH (p<0.0001, r=0.56). No differences were observed in sPRR plasma and urine levels between non-DM and DM patients. However, the plasma levels of sPRR were 12.5 and 18.5-fold higher than in urine of Non-DM and DM, respectively. Treatment with RAS blockers increased sPRR in plasma (DM with RAS blockade: 24,455±2,151 vs DM without RAS blockade: 17,726±1,255 pg/mL, p<0.001) but not in urine. These data suggest that the cell membrane-bound PRR, not the soluble form, might be responsible for the increases in renin activity and TGF-β, as well as in the low pH in the urine of type-2 DM patients. It is likely that prorenin-mediated activation of membrane bound PRR promotes diabetic nephropathy by increasing urinary formation of TGF-β. [ABSTRACT FROM AUTHOR]

To investigate whether thyroid hormone (TH) interacts with the sympathetic nervous system (SNS) to modulate bone mass and structure, we studied the effects of daily T3 treatment in a supraphysiological dose for 12 wk on the bone of young adult mice with chronic sympathetic hyperactivity owing to double-gene disruption of adrenoceptors that negatively regulate norepinephrine release, α(2A)-AR, and α(2C)-AR (α(2A/2C)-AR(-/-) mice). As expected, T3 treatment caused a generalized decrease in the areal bone mineral density (aBMD) of WT mice (determined by DEXA), followed by deleterious effects on the trabecular and cortical bone microstructural parameters (determined by μCT) of the femur and vertebra and on the biomechanical properties (maximum load, ultimate load, and stiffness) of the femur. Surprisingly, α(2A/2C)-AR(-/-) mice were resistant to most of these T3-induced negative effects. Interestingly, the mRNA expression of osteoprotegerin, a protein that limits osteoclast activity, was upregulated and downregulated by T3 in the bone of α(2A/2C)-AR(-/-) and WT mice, respectively. β1-AR mRNA expression and IGF-I serum levels, which exert bone anabolic effects, were increased by T3 treatment only in α(2A/2C)-AR(-/-) mice. As expected, T3 inhibited the cell growth of calvaria-derived osteoblasts isolated from WT mice, but this effect was abolished or reverted in cells isolated from KO mice. Collectively, these findings support the hypothesis of a TH-SNS interaction to control bone mass and structure of young adult mice and suggests that this interaction may involve α2-AR signaling. Finally, the present findings offer new insights into the mechanisms through which TH regulates bone mass, structure, and physiology., (Copyright © 2014 the American Physiological Society.)