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Xanthine oxidoreductase (XOR) is generally known as the final enzyme in purine metabolism and as a source of reactive oxygen species (ROS). In addition, this enzyme has been suggested to mediate nitric oxide (NO) formation via reduction of inorganic nitrate and nitrite. This NO synthase (NOS)-independent pathway for NO generation is of particular importance during certain conditions when NO bioavailability is diminished due to reduced activity of endothelial NOS (eNOS) or increased oxidative stress, including aging and cardiovascular disease. The exact interplay between NOS- and XOR-derived NO generation is not fully elucidated yet. The aim of the present study was to investigate if eNOS deficiency is associated with changes in XOR expression and activity and the possible impact on nitrite, NO and ROS homeostasis. Plasma levels of nitrate and nitrite were similar between eNOS deficient (eNOS -/- ) and wildtype (wt) mice. XOR activity was upregulated in eNOS -/- compared with wt, but not in nNOS -/- , iNOS -/- or wt mice treated with the non-selective NOS inhibitor L-NAME. Following an acute dose of nitrate, plasma nitrite increased more in eNOS -/- compared with wt, and this augmented response was abolished by the selective XOR inhibitor febuxostat. Livers from eNOS -/- displayed higher nitrite reducing capacity compared with wt, and this effect was attenuated by febuxostat. Dietary supplementation with nitrate increased XOR expression and activity, but concomitantly reduced superoxide generation. The latter effect was also seen in vitro after nitrite administration. Treatment with febuxostat elevated blood pressure in eNOS -/- , but not in wt mice. A high dose of dietary nitrate reduced blood pressure in naïve eNOS -/- mice, and again this effect was abolished by febuxostat. In conclusion, eNOS deficiency is associated with an upregulation of XOR facilitating the nitrate-nitrite-NO pathway and decreasing the generation of ROS. This interplay between XOR and eNOS is proposed to play a significant role in NO homeostasis and blood pressure regulation., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background and Purpose: NO deficiency and oxidative stress are crucially involved in the development or progression of cardiovascular disease, including hypertension and stroke. We have previously demonstrated that acute treatment with the newly discovered organic nitrate, 2-nitrate-1,3-dibuthoxypropan (NDBP), is associated with NO-like effects in the vasculature. This study aimed to further characterize the mechanism(s) and to elucidate the therapeutic potential in a model of hypertension and oxidative stress., Experimental Approach: A combination of ex vivo, in vitro and in vivo approaches was used to assess the effects of NDBP on vascular reactivity, NO release, NADPH oxidase activity and in a model of hypertension., Key Results: Ex vivo vascular studies demonstrated NDBP-mediated vasorelaxation in mesenteric resistance arteries, which was devoid of tolerance. In vitro studies using liver and kidney homogenates revealed dose-dependent and sustained NO generation by NDBP, which was attenuated by the xanthine oxidase inhibitor febuxostat. In addition, NDBP reduced NADPH oxidase activity in the liver and prevented angiotensin II-induced activation of NADPH oxidase in the kidney. In vivo studies showed that NDBP halted the progression of hypertension in mice with chronic angiotensin II infusion. This was associated with attenuated cardiac hypertrophy, and reduced NADPH oxidase-derived oxidative stress and fibrosis in the kidney and heart., Conclusion and Implications: The novel organic nitrate NDBP halts the progression of angiotensin II-mediated hypertension. Mechanistically, our findings suggest that NDBP treatment is associated with sustained NO release and attenuated activity of NADPH oxidase, which to some extent requires functional xanthine oxidase., (© 2016 The British Pharmacological Society.)

The mechanisms underlying the adaptive process in the remaining kidney after unilateral nephrectomy are not well known. In the present study adaptive changes in interstitial pressure conditions following uninephrectomy were investigated in male Sprague-Dawley rats. Direct subcapsular hydrostatic pressure recordings were made via microcatheters , renal lymph was collected for lymph flow rate estimations and determinations of lymph protein content and the urine flow rate and urinary excretion of sodium and potassium were measured, before and after contralateral nephrectomy. There was a significant rise in subcapsular hydrostatic pressure by about 50%, and in the lymph flow rate, by more than 100%, within 1 hr after nephrectomy. The protein concentration in collected renal lymph was significantly decreased (from 2.30 +/- 0.25 to 1.45 +/- 0.26 g/dl), compared with control collections in sham-operated animals (from 2.22 +/- 0.23 to 1.78 +/- 0.19 g/dl). These findings indicate an increased hydrostatic and decreased intersititial pressure in response to contralateral nephrectomy. Urine flow rate and electrolyte excretion were significantly elevated (urine flow rate by 92%, sodium excretion from 0.210 +/- 0.035 to 0.352 +/- 0.067 mumoles/min and potassium excretion from 0.306 +/- 0.074 to 1.617 +/- 0.228 mumoles/min). Inhibition of prostaglandin synthesis prior to nephrectomy abolished the lymph flow increase and oncotic pressure decrease and also the diuretic and natriuretic responses. The adaptively increased potassium excretion was, however, only blocked by diclofenac sodium and not by indomethacin. These results demonstrate the important role of the renal interstitium and the prostaglandin system in the renal adaptation to nephron loss.

Compared with the effects of a 6% protein diet, feeding rats a 40% protein diet for 10 days increases glomerular filtration rate and decreases the activity of the tubuloglomerular (TG) feedback control system. The decrease in TG feedback activity results from an increase in the threshold at which the loop of Henle flow rate initiates feedback responses. To determine whether this protein-dependent shift in the TG feedback response curve is caused by changes in either the signal or the sensing mechanism in the feedback pathway, we used micropuncture and microperfusion techniques to study the TG feedback system of rats fed high- or low-protein (40 or 6% casein) diets for approximately 7-10 days. Compared with the rats fed the low-protein diet, in the high-protein group distally measured single nephron glomerular filtration rate was 17% higher, and Na and Cl concentrations in early distal tubule fluid were 30-50% lower. Early distal osmolality was not different in the two groups. TG feedback responses assessed by changes in stop-flow pressure during perfusion of the distal nephron with NaCl solutions did not differ between diet groups. We conclude that the sensing mechanism in the TG feedback system is not altered by this manipulation of dietary protein, whereas the signal eliciting the TG feedback response is affected. Because rats fed a high-protein diet have higher rates of Na and Cl absorption between the late proximal and early distal tubules than do rats fed a low-protein diet, early distal Na and Cl concentrations are reduced, and the signal for TG feedback is diminished in rats fed the high-protein diet.