Alternative pathways for angiotensin II production as an important determinant of kidney damage in endotoxemia.

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.

Overexpression of urinary N-domain ACE in chronic kidney dysfunction in Wistar rats.

Local activation of the renin-angiotensin system (RAS) has been implicated in the pathogenesis of several renal disorders. In this study we investigated how chronic kidney disease (CKD) modulates RAS components in an experimental model. Male Wistar rats were divided into three groups: sham, nephrectomized, and nephrectomized receiving losartan. Chronic kidney disease animals presented decreased renal N-domain angiotensin-converting enzyme (ACE) activity but overexpression of N-domain ACE in urine. Remnant kidneys presented high angiotensin II levels. Losartan treatment increased urine and tissue ACE activity and tissue levels of angiotensins, mainly angiotensin (1-7), and improved renal and histopathologic parameters. Taken together, the authors' results indicate that pathophysiological changes due to CKD could lead to an increased expression of somatic and N-domain ACE, mainly the 65 kDa isoform, suggesting that this enzyme could be used as a biological urinary marker in CKD.

Proteomic approaches in understanding a detected relationship between chemotherapy-induced nephrotoxicity and cell respiration in HK-2 cells.

BACKGROUND/AIMS: Nephrotoxicity is a prominent component of the profile of chemotherapeutic agents and to date proteomics has represented the main technique to identify protein profiles in response to xenobiotic exposure. METHODS: We made use of two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight analysis to evaluate chemotoxicity effects of cisplatin (CPT) and carboplatin (CB) on proteins from human renal proximal tubule epithelial cells (HK-2). RESULTS: Tandem mass spectrometry analysis showed that ATP synthase subunit α and serine hydroxymethyltransferase were only expressed in HK-2 cells exposed to CPT. Since CPT causes damage in cellular respiration, we suggest that this might be a protective adaptation to CPT-induced nephrotoxicity. Thioredoxin-dependent peroxide reductase disappeared in the CPT group and was upregulated in the CB group, suggesting that CB exposure stimulates preventive apoptotic mechanisms. We suggest a relationship between chemotherapeutic agent-induced nephrotoxicity and cell respiration. The identification of proteins differentially expressed in HK-2 cells, when exposed to CPT and CB, not only supplies important information to understand the molecular action mechanisms, which are triggered by metal-based drugs in cell nephrotoxicity, but also can lead to the design of more effective anticancer drugs. CONCLUSION: These results provide important insights into the investigation of possible biomarker(s) of toxicity that could eventually reduce the side effects of chemotherapeutic agents.

Catecholamines production by kidney tissue and mesangial cell culture is differentially modulated by diabetes.

INTRODUCTION: According to the International Diabetes Federation, the number of people with diabetes mellitus may reach 700 million in 2045. Catecholamines are involved in the regulation of several kidney functions. This study investigates the effects of hyperglycemia on catecholamines' metabolism in kidney tissue from control, diabetic, and insulin-treated diabetic rats, both in vivo and in vitro. METHODS: Male Wistar-Hannover rats were randomized into: control, diabetic, and insulin-treated diabetic groups. Diabetes was induced by a single injection of streptozotocin, and diabetic treated group also received insulin. After 60 days, blood and kidney tissue from all groups were collected for catecholamines' quantification and mesangial cells culture. RESULTS: diabetic rats had lower body weight, hyperglycemia, and increase water intake and diuresis. Additionally, diabetes promoted a sharp decrease in creatinine clearance compared to control group. Regarding the whole kidney extracts, both diabetic groups (treated and non-treated) had significant reduction in norepinephrine concentration. In mesangial cell culture, catecholamines' concentration were lower in the culture medium than in the intracellular compartment for all groups. Norepinephrine, epinephrine, and dopamine medium levels were increased in the diabetic group. CONCLUSION: The major finding of the present study was that 8 weeks of diabetes induction altered the kidney catecholaminergic system in a very specific manner, once the production of catecholamines in the excised kidney tissue from diabetic rats was differentially modulated as compared with the production and secretion by cultured mesangial cells.

Diabetes induces changes of catecholamines in primary mesangial cells.

Diabetes mellitus is a frequent cause of kidney function damage with diabetic nephropathy being predominantly related to glomerular dysfunction. Diabetes is capable of interfering with distinct hormonal systems, as well as catecholamine metabolism. Since mesangial cells, the major constituent of renal glomerulus, constitute a potential site for catecholamine production, the present study was carried out to investigate alterations in catecholamine metabolism in cultured mesangial cells from the nonobese diabetic mouse, a well-established model for type I diabetes. We evaluated mesangial cells from normoglycemic and hyperglycemic nonobese diabetic mice, as well as cells from normoglycemic Swiss mice as control. Mesangial cells from normoglycemic mice presented similar profiles concerning all determinations. However, cells isolated from hyperglycemic animals presented increased dopamine and norepinephrine production/secretion. Among the studied mechanisms, we observed an upregulation of tyrosine hydroxylase expression accompanied by increased tetrahydrobiopterin consumption, the tyrosine hydroxylase enzymatic cofactor. However, this increase in synthetic pathways was followed by decreased monoamine oxidase activity, which corresponds to the major metabolic pathway of catecholamines in mesangial cells. In addition, whole kidney homogenates from diabetic animals also presented increased dopamine and norepinephrine levels when compared to normoglycemic animals. Thus, our results suggest that diabetes alters catecholamine production by interfering with both synthesizing and degrading enzymes, suggesting a possible role of catecholamine in the pathogenesis of acute and chronic renal complications of diabetes mellitus.

Catecholamines production by kidney tissue and mesangial cell culture is differentially modulated by diabetes / A produção de catecolaminas pelo tecido renal e a cultura de células mesangiais é modulada diferencialmente pelo diabetes

Abstract Introduction: According to the International Diabetes Federation, the number of people with diabetes mellitus may reach 700 million in 2045. Catecholamines are involved in the regulation of several kidney functions. This study investigates the effects of hyperglycemia on catecholamines' metabolism in kidney tissue from control, diabetic, and insulin-treated diabetic rats, both in vivo and in vitro. Methods: Male Wistar-Hannover rats were randomized into: control, diabetic, and insulin-treated diabetic groups. Diabetes was induced by a single injection of streptozotocin, and diabetic treated group also received insulin. After 60 days, blood and kidney tissue from all groups were collected for catecholamines' quantification and mesangial cells culture. Results: diabetic rats had lower body weight, hyperglycemia, and increase water intake and diuresis. Additionally, diabetes promoted a sharp decrease in creatinine clearance compared to control group. Regarding the whole kidney extracts, both diabetic groups (treated and non-treated) had significant reduction in norepinephrine concentration. In mesangial cell culture, catecholamines' concentration were lower in the culture medium than in the intracellular compartment for all groups. Norepinephrine, epinephrine, and dopamine medium levels were increased in the diabetic group. Conclusion: The major finding of the present study was that 8 weeks of diabetes induction altered the kidney catecholaminergic system in a very specific manner, once the production of catecholamines in the excised kidney tissue from diabetic rats was differentially modulated as compared with the production and secretion by cultured mesangial cells.

Resumo Introdução: Segundo a Federação Internacional de Diabetes, o número de pessoas com diabetes mellitus pode chegar a 700 milhões em 2045. As catecolaminas estão envolvidas na regulação de várias funções renais. Este estudo investiga os efeitos da hiperglicemia no metabolismo das catecolaminas no tecido renal de ratos controle, diabéticos e diabéticos tratados com insulina, tanto in vivo como in vitro. Métodos: Os ratos Wistar-Hannover machos foram randomizados em: grupos controle, diabéticos e diabéticos tratados com insulina. O diabetes foi induzido por uma única injeção de estreptozotocina, e o grupo diabético tratado também recebeu insulina. Após 60 dias, sangue e tecido renal dos grupos foram coletados para quantificação de catecolaminas e cultura de células mesangiais. Resultados: ratos diabéticos apresentaram peso corporal mais baixo, hiperglicemia, e aumento da ingestão de água e diurese. Ademais, o diabetes promoveu uma redução acentuada na depuração de creatinina comparado com o grupo controle. Quanto aos extratos de rim total, ambos os grupos diabéticos (tratados/não tratados) tiveram redução significativa na concentração de noradrenalina. Na cultura de células mesangiais, a concentração de catecolaminas foi menor no meio de cultura do que no compartimento intracelular para todos os grupos. Níveis médios de noradrenalina, adrenalina e dopamina estavam aumentados no grupo diabético. Conclusão: O principal achado deste estudo foi que 8 semanas de indução de diabetes alteraram o sistema catecolaminérgico renal de maneira muito específica, já que a produção de catecolaminas no tecido renal excisado de ratos diabéticos foi modulada diferencialmente comparada com produção e secreção por células mesangiais cultivadas.

Tissue-specific expression of transgenic secreted ACE in vasculature can restore normal kidney functions, but not blood pressure, of Ace-/- mice.

Angiotensin-converting enzyme (ACE) regulates normal blood pressure and fluid homeostasis through its action in the renin-angiotensin-system (RAS). Ace-/- mice are smaller in size, have low blood pressure and defective kidney structure and functions. All of these defects are cured by transgenic expression of somatic ACE (sACE) in vascular endothelial cells of Ace-/- mice. sACE is expressed on the surface of vascular endothelial cells and undergoes a natural cleavage secretion process to generate a soluble form in the body fluids. Both the tissue-bound and the soluble forms of ACE are enzymatically active, and generate the vasoactive octapeptide Angiotensin II (Ang II) with equal efficiency. To assess the relative physiological roles of the secreted and the cell-bound forms of ACE, we expressed, in the vascular endothelial cells of Ace-/- mice, the ectodomain of sACE, which corresponded to only the secreted form of ACE. Our results demonstrated that the secreted form of ACE could normalize kidney functions and RAS integrity, growth and development of Ace-/- mice, but not their blood pressure. This study clearly demonstrates that the secreted form of ACE cannot replace the tissue-bound ACE for maintaining normal blood pressure; a suitable balance between the tissue-bound and the soluble forms of ACE is essential for maintaining all physiological functions of ACE.

Renin-angiotensin system may trigger kidney damage in NOD mice.

Diabetic nephropathy is a complication of diabetes and one of the main causes of end-stage renal disease. A possible causal link between renin-angiotensin aldosterone system (RAAS) and diabetes is widely recognized but the mechanisms by which the RAAS may lead to this complication remains unclear. The aim of this study was to evaluate angiotensin-I converting enzyme (ACE) activity and expression in numerous tissues, especially kidney, of non-obese diabetic mouse. Kidney, lung, pancreas, heart, liver and adrenal tissues from diabetic and control female NOD mice were homogenized for measurement of ACE activity, SDS-PAGE and Western blotting for ACE and ACE2, immunohistochemistry for ACE and angiotensins I, II and 1-7 and bradykinin quantification. ACE activity was higher in kidney, lung and adrenal tissue of diabetic mice compared with control mice. In pancreas, activity was decreased in the diabetic group. Western blotting analysis indicated that both groups presented ACE isoforms with molecular weights of 142 and 69 kDa and a decrease in ACE2 protein expression. Angiotensin concentrations were not altered within groups, although bradykinin levels were higher in diabetic mice. The immunohistochemical study in kidney showed an increase in tubular ACE expression. Our results show that the RAAS is affected by diabetes and the elevated ACE/ACE2 ratio may contribute to renal damage.