ACE2 deficiency increases NADPH-mediated oxidative stress in the kidney.

Abstract Angiotensin-converting enzyme 2 (ACE2) is highly expressed in the kidney and hydrolyzes angiotensin II (Ang II) to Ang(1-7). Since Ang II is a strong activator of oxidative stress, we reasoned that ACE2 could be involved in the regulation of renal oxidative stress by governing the levels of Ang II. We, therefore, assessed levels of oxidative stress in kidney cortex of ACE2 knockout and wild-type littermate mice under baseline conditions. We found multiple markers of increased oxidative stress in ACE2KO mice. NADPH oxidase activity was increased in kidney cortex from ACE2KO mice as compared to WT (227 ± 24% vs.100 ± 19%, P < 0.001). However, kidney catalase and superoxide dismutase activities were not different between groups. Exogenous Ang II was degraded less efficiently by kidneys from ACE2KO mice than WT mice, and administration of an AT1R blocker (losartan 30 mg/kg/day) resulted in normalization of NADPH oxidase activity in the ACE2KO. These findings suggest that an AT1R-dependent mechanism contributes to increased ROS observed in the ACE2KO. This study demonstrates that genetic deficiency of ACE2 activity in mice fosters oxidative stress in the kidney in the absence of overt hypertension and is associated with reduced kidney capacity to hydrolyze Ang II. ACE2KO mice serve as a novel in vivo model to examine the role of overactivity of NADPH oxidase in kidney function.

Angiotensin-converting enzyme 2-independent action of presumed angiotensin-converting enzyme 2 activators: studies in vivo, ex vivo, and in vitro.

Angiotensin (Ang)-converting enzyme 2 (ACE2) is a key enzyme in the metabolism of Ang II. XNT (1-[(2-dimethylamino)ethylamino]-4-(hydroxymethyl)-7-[(4-methylphenyl) sulfonyl oxy]-9H-xanthene-9-one) and diminazene have been reported to exert various organ-protective effects, which are attributed to the activation of ACE2. To test the effect of these compounds, we studied Ang II degradation in vivo and in vitro as well as their effect on ACE2 activity in vivo and in vitro. In a model of Ang II-induced acute hypertension, blood pressure (BP) recovery was markedly enhanced by XNT (slope with XNT, -3.26±0.2 versus -1.6±0.2 mm Hg/min without XNT; P<0.01). After Ang II infusion, neither plasma nor kidney ACE2 activity was affected by XNT. Plasma Ang II and Ang (1-7) levels also were not significantly affected by XNT. The BP-lowering effect of XNT seen in wild-type animals was also observed in ACE2 knockout mice (slope with XNT, -3.09±0.30 versus -1.28±0.22 mm Hg/min without XNT; P<0.001). These findings show that the BP-lowering effect of XNT in Ang II-induced hypertension cannot be because of the activation of ACE2. In vitro and ex vivo experiments in both mice and rat kidney confirmed a lack of enhancement of ACE2 enzymatic activity by XNT and diminazene. Moreover, Ang II degradation in vitro and ex vivo was unaffected by XNT and diminazene. We conclude that the biological effects of these compounds are ACE2-independent and should not be attributed to the activation of this enzyme.

Proximal renal tubular acidosis: a not so rare disorder of multiple etiologies.

Proximal renal tubular acidosis (RTA) (Type II RTA) is characterized by a defect in the ability to reabsorb HCO(3) in the proximal tubule. This is usually manifested as bicarbonate wastage in the urine reflecting that the defect in proximal tubular transport is severe enough that the capacity for bicarbonate reabsorption in the thick ascending limb of Henle's loop and more distal nephron segments is overwhelmed. More subtle defects in proximal bicarbonate transport likely go clinically unrecognized owing to compensatory reabsorption of bicarbonate distally. Inherited proximal RTA is more commonly autosomal recessive and has been associated with mutations in the basolateral sodium-bicarbonate cotransporter (NBCe1). Mutations in this transporter lead to reduced activity and/or trafficking, thus disrupting the normal bicarbonate reabsorption process of the proximal tubules. As an isolated defect for bicarbonate transport, proximal RTA is rare and is more often associated with the Fanconi syndrome characterized by urinary wastage of solutes like phosphate, uric acid, glucose, amino acids, low-molecular-weight proteins as well as bicarbonate. A vast array of rare tubular disorders may cause proximal RTA but most commonly it is induced by drugs. With the exception of carbonic anhydrase inhibitors which cause isolated proximal RTA, drug-induced proximal RTA is associated with Fanconi syndrome. Drugs that have been recently recognized to cause severe proximal RTA with Fanconi syndrome include ifosfamide, valproic acid and various antiretrovirals such as Tenofovir particularly when given to human immunodeficiency virus patients receiving concomitantly protease inhibitors such as ritonavir or reverse transcriptase inhibitors such as didanosine.

Genetic causes and mechanisms of distal renal tubular acidosis.

The primary or hereditary forms of distal renal tubular acidosis (dRTA) have received increased attention because of advances in the understanding of the molecular mechanism, whereby mutations in the main proteins involved in acid-base transport result in impaired acid excretion. Dysfunction of intercalated cells in the collecting tubules accounts for all the known genetic causes of dRTA. These cells secrete protons into the tubular lumen through H(+)-ATPases functionally coupled to the basolateral anion exchanger 1 (AE1). The substrate for both transporters is provided by the catalytic activity of the cytosolic carbonic anhydrase II (CA II), an enzyme which is also present in the proximal tubular cells and osteoclasts. Mutations in ATP6V1B1, encoding the B-subtype unit of the apical H(+) ATPase, and ATP6V0A4, encoding the a-subtype unit, lead to the loss of function of the apical H(+) ATPase and are usually responsible for patients with autosomal recessive dRTA often associated with early or late sensorineural deafness. Mutations in the gene encoding the cytosolic CA II are associated with the autosomal recessive syndrome of osteopetrosis, mixed distal and proximal RTA and cerebral calcification. Mutations in the AE1, the gene that encodes the Cl(-)/HCO(3)(-) exchanger, usually present as dominant dRTA, but a recessive pattern has been recently described. Several studies have shown trafficking defects in the mutant protein rather than the lack of function as the major mechanism underlying the pathogenesis of dRTA from AE1 mutations.

Erythrocyte markers of oxidative stress in higher age-group preeclamptic and normal pregnant mothers.

Preeclampsia can have significant impact on health of both mother and fetus. It had been proposed that maternal endothelial cell dysfunction is the key event resulting in the diverse clinical manifestations of preeclampsia and evidence has since accumulated. Research in recent times is indicative of the role of oxidative stress in the endothelial cell dysfunction. Preeclampsia is more common in first pregnancy and studies have further shown an increase in risk of preeclampsia with maternal age. The aim of this study was to explore the status of oxidative stress in higher age-group preeclamptic and normal pregnant mother. The study included 20 normal pregnant women and 60 preeclamptic women. They were allocated into 4 subgroups between ages 20-25, 26-30, 31-35, and 36-40. Erythrocytes were analyzed for the following antioxidant enzymes, i.e., glutathione peroxidase, superoxide dismutase and catalase.Lipid peroxidation product, malondialdehyde, was analyzed to determine oxidative stress. The results showed an increase in oxidative stress, and high magnitude suppression/decrease in antioxidant enzymes activities in erythrocytes with increase in age groups in both preeclamptic and normal pregnant women. This indicates that an increase in the risk of preeclampsia with maternal age could be due to an increase in oxidative stress with age. This further attests to the role of oxidative stress in preeclampsia.