Angiotensin II type 2 receptor agonist, compound 21, prevents tubular epithelial cell damage caused by renal ischemia.

During ischemic acute kidney injury (AKI), loss of cytoskeletal integrity and disruption of intercellular junctions are rapid events in response to ATP depletion. Angiotensin II type 2 receptor (AT2R) is overexpressed in injury situations and its stimulation by angiotensin II (AngII) is related to beneficial renal effects. Its role on ischemic AKI has not been deeply studied. The aim of the present study was to investigate whether pretreatment with the AT2R agonist, C21, prevents ischemic renal epithelial cell injury. Studies in a model of 40 min of renal ischemia followed by 24 h of reperfusion (IR) in rats demonstrated that C21 pretreatment attenuated renal dysfunction and induced better preservation of tubular architecture. In addition, we studied the expression of Rho GTPases, RhoA and Cdc42, since they are key proteins in the regulation of the actin cytoskeleton and the stability of epithelial intercellular junctions. IR downregulated RhoA and Cdc42 abundance in rat kidneys. C21 pretreatment prevented RhoA reduction and increased Cdc42 abundance compared to controls. We also used an in vitro model of ATP depletion in MDCK cells grown on filter support. Using immunofluorescence we observed that in MDCK cells, C21 pretreatment prevented the ATP depletion-induced reduction of actin in brush border microvilli and in stress fibers. Moreover, C21 prevented membrane E-cadherin reduction, and RhoA and Cdc42 downregulation. The present study describes for the first time a renoprotective effect of the AT2R agonist, C21, against AKI, and provides evidence supporting that stimulation of AT2R triggers cytoprotective mechanisms against an ischemic event.

Effects of "in vivo" administration of baclofen on rat renal tubular function.

The effects of the in vivo administration of baclofen on renal tubular transport and aquaporin-2 (AQP2) expression were evaluated. In conscious animals kept in metabolic cages, baclofen (0.01-1mg/kg, s.c.) induced a dose-dependent increment in the urine flow rate (UFR) and in sodium and potassium excretion, associated with an increased osmolal clearance (Closm), a diminished urine to plasma osmolality ratio (Uosm/Posm) and a decrease in AQP2 expression. The above mentioned baclofen effects on functional parameters were corroborated by using conventional renal clearance techniques. Additionally, this model allowed the detection of a diminution in glucose reabsorption. Some experiments were performed with water-deprived or desmopressin-treated rats kept in metabolic cages. Either water deprivation or desmopressin treatment decreased the UFR and increased the Uosm/Posm. Baclofen did not change the Uosm/Posm or AQP2 expression in desmopressin-treated rats; but it increased the UFR and diminished the Uosm/Posm and AQP2 expression in water-deprived animals. These results indicate that in vivo administration of baclofen promotes alterations in proximal tubular transport, since glucose reabsorption was decreased. The distal tubular function was also affected. The increased Closm indicates an alteration in solute reabsorption at the ascending limb of the Henle's loop. The decreased Uosm/Posm and AQP2 expression in controls and in water-deprived, but not in desmopressin-treated rats, lead us to speculate that some effect of baclofen on endogenous vasopressin availability could be responsible for the impaired urine concentrating ability, more than any disturbance in the responsiveness of the renal cells to the hormone.

Localization and functional activity of cytochrome P450 side chain cleavage enzyme (CYP11A1) in the adult rat kidney.

Cumulative evidence demonstrated effective downstream metabolism of pregnenolone in renal tissue. The aim of this study was to evaluate the expression and functional activity of cytochrome P450 side chain cleavage enzyme (CYP11A1), which converts cholesterol into pregnenolone, in adult rat kidney. Immunohistochemical labeling for CYP11A1 was observed in renal cortex and medulla, on structures identified as distal convoluted tubule and thick ascending limb of Henle's loop, respectively. Immunoblotting analysis corroborated the renal expression of the protein in inner mitochondrial membrane fractions. The incubation of isolated mitochondria with the membrane-permeant cholesterol analogue 22R-hydroxycholesterol resulted in efficient formation of pregnenolone, the immediate precursor for the synthesis of all the steroid hormones. The low progesterone production rate observed in these experiments suggested a poor activity of 3ß-hydroxysteroid dehydrogenase enzyme in renal mitochondria. The steroidogenic acute regulatory protein (StAR), involved in the mitochondrial import of cholesterol, was detected in renal tissue at both mRNA and protein level. Immunostaining for StAR showed similar distribution to that observed for CYP11A1. The expression of StAR and CYP11A1 was found to be higher in medulla than in cortex. This enhanced expression of steroidogenesis-related proteins correlated with a greater pregnenolone synthesis rate and higher steroid hormones tissular content measured in medulla. In conclusion, we have established the expression and localization of StAR and CYP11A1 protein, the ability of synthesizing pregnenolone and a region-specific content of sex hormones in the adult rat kidney. These data clearly show that the kidney is a steroid hormones synthesizing organ. It is proposed that the existence in the kidney of complete steroidogenic machinery would respond to a physiological significance.