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High soy (HS) diets are neuroprotective and promote vascular dilatation in the periphery. We hypothesized that a HS diet would promote vascular dilatation in the cerebrovasculature by mimicking estradiol's actions on the endothelial nitric oxide synthase (eNOS) system including increasing eNOS expression and decreasing caveolin-1 expression to increase nitric oxide (NO) production. Ovariectomized rats were fed HS or a soy-free diet (SF) ± low physiological estradiol (E2) for 4 weeks. Neither E2 nor HS altered middle cerebral artery (MCA) structure or vascular responses to acetylcholine, serotonin, or phenylephrine. Estradiol enhanced bradykinin-induced relaxation in an eNOS-dependent manner. Although E2 and HS increased eNOS mRNA expression in the brain and cerebrovasculature, they had no effect on eNOS protein expression or phosphorylation in the MCA. However, E2 decreased caveolin-1 protein in the MCA. In MCAs neither E2 nor HS altered estrogen receptor (ER) alpha expression, but E2 did reduce ER beta levels. These data suggest that HS diets have no effect on vascular NO production, and that E2 may modulate basal NO production by reducing the expression of caveolin-1, an allosteric inhibitor of NOS activity. However, the effects of E2 and HS on the cerebrovasculature are small and may not underlie their protective actions in pathological states.

Conventional methods used for measuring regional renal blood flow, such as laser-Doppler flowmetry, are highly invasive, and each measurement is restricted to a discrete location. The aim of this study was to determine whether ultrasound imaging in conjunction with enhanced contrast agent (microbubbles; Vevo MicroMarker, VisualSonics) could provide a viable noninvasive alternative. This was achieved by determining changes in renal cortical and medullary rate of perfusion in response to a bolus injection of endothelin-1 (ET-1; 0.6, 1.0, or 2.0 nmol/kg) and comparing these responses to those observed in separate groups of mice with conventional laser-Doppler methods. Intravenous infusion of ET-1 in anesthetized male C57bl/6 mice resulted in a dose-dependent increase in mean arterial pressure and a dose-dependent decrease in total renal blood flow as measured by pulse-wave Doppler. ET-1 infusion resulted in a dose-dependent decrease in regional kidney perfusion as measured by both ultrasound with enhanced contrast agent and laser-Doppler measurements, verifying the use of ultrasound to measure regional kidney perfusion. Noted limitations of ultrasound imaging compared with laser-Doppler flowmetry included a lower degree of sensitivity to changes in tissue perfusion and the inability to assess rapid or transient changes in tissue perfusion. In conclusion, ultrasound represents an effective and noninvasive method for the measurement of relatively short-term, steady-state changes in regional blood flow in the mouse kidney.

A mircoarray analysis was performed to identify novel inflammatory genes that are differentially expressed in the mesenteric arteries of male and female spontaneously hypertensive rats (SHRs). Fractalkine was found to be the inflammatory gene with the greatest differential expression in mesenteric arteries, with the expression being greater in female SHRs compared with males. Greater inflammatory mediators in female SHRs were verified by measuring urinary monocyte chemoattractant protein-1, transforming growth factor-beta, and tumor necrosis factor-alpha (TNF-alpha) excretion, all of which were greater in female SHRs compared with males. Real-time PCR, Western blot analysis, and ELISA verified greater soluble fractalkine in mesenteric arteries of female SHRs. Consistent with increased fractalkine expression, TNF-alpha-converting enzyme and TNF-alpha levels in mesenteric arteries were also greater in female SHRs. We next tested the hypothesis that mesenteric arteries from female SHRs will have greater fractalkine-induced dysfunction. Acetylcholine, sodium nitroprusside, phenylephrine, and KCl concentration-response curves were performed in third-order mesenteric arteries from male and female SHRs pretreated with either vehicle or fractalkine (1 microg/ml). Fractalkine decreased sensitivity to 1) acetylcholine in arteries from male SHRs, 2) phenylephrine in arteries from both sexes, and 3) KCl in arteries from female SHRs. In conclusion, urinary and vascular markers of inflammation are greater in female SHRs compared with males, although blood pressure and cardiovascular risk are less in females.

Aim: Our laboratory and others have shown that endothelin (ET)-1 directly stimulates nitric oxide (NO) production in inner medullary collecting duct (IMCD) cells. The goal of this study was to determine which NO synthase (NOS) isoforms in IMCD are sensitive to ET-1, and the role of ETA and ETB receptor activation in vivo and in vitro. Methods: NOS enzymatic activity and NOS isoform protein expression were examined in cultured IMCD-3 cells and isolated renal inner medulla. ETB receptor-deficient homozygous rats (sl/sl) have elevated levels of circulating ET-1 and lack a functional ETB signalling pathway in kidneys, and furthermore provides a unique model to study ETA receptor signalling in the renal inner medulla in vivo. Results: Incubation of IMCD-3 cells with exogenous ET-1 (50 nm) resulted in ETA-dependent increased NOS1 protein expression in IMCD-3 cells with no effect on NOS2 or NOS3 expression. ETB receptor antagonism has no effect on NOS expression in IMCD-3 cells. Consistent with in vitro results, cytosolic NOS1 protein expression was significantly greater in the renal inner medulla of sl/sl rats compared with heterozygous (sl/+) controls, with no alteration in NOS3 expression. In contrast to protein expression data, NOS1- and NOS3-specific enzymatic activities decreased in the cytosolic fraction from the renal inner medulla of sl/sl compared with sl/+. Conclusion: These results provide evidence that both ETA and ETB receptors regulate NOS isoform activity in the renal inner medulla and specifically support the hypothesis that ETA receptor activation increases NOS1 expression.

There is a sex difference in hypertensive renal injury, with men experiencing greater severity and a more rapid progression of renal disease than women; however, the molecular mechanisms protecting against renal injury in women are unknown. The goal of this study was to determine whether sex hormones modulate blood pressure and the progression of albuminuria during the developmental phase of hypertension in male and female spontaneously hypertensive rats (SHR). Studies were also performed to examine how sex and sex hormones influence two major risk factors for albuminuria, overactivation of the renin-angiotensin system and oxidative stress. Blood pressure was measured by telemetry in gonad-intact and gonadectomized male and female SHR. Microalbumin excretion, measured over time, and macrophage infiltration were used to assess renal health. Male SHR had significantly higher blood pressures than female SHR, and gonadectomy decreased blood pressures in males with no effect in females. Male SHR displayed a gonad-sensitive increase in albuminuria over time, and female SHR had a gonad-sensitive suppression in macrophage infiltration. Female SHR had greater plasma ANG II levels and similar levels of renal cortical ANG II vs. levels shown in males but less AT1-receptor protein expression in the renal cortex. Female SHR also had a gonad-sensitive decrease in renal oxidative stress. Therefore, the renal protection afforded to female SHR is associated with lower blood pressure, decreased macrophage infiltration, and decreased levels of oxidative stress.

Endothelin-1 (ET-1) is a potent vasoconstrictor, and dysregulation of the endothelin (ET) system has been implicated in the development of hypertension. Sex differences in the ET system have been identified in ET receptor expression and activation, levels of ET-1, and downstream mediators of the ET system. More specifically, males have greater ET-1/ETA receptor activation, whereas females exhibit greater ETB receptor activation. These differences have been suggested to contribute to the sex differences observed in blood pressure control, with greater ETB receptor activation in females potentially acting as an important pathway contributing to the lower prevalence of hypertension in young females compared with age-matched males. This hypothesis is further supported by studies in pregnancy; the role of the ET system is enhanced during pregnancy, with dysregulation of the ET system resulting in preeclampsia. Further research is necessary to elucidate the relative roles of the ET system in blood pressure control in both sexes and to further explore the potential benefits of pharmacological ET blockade in women.

Background We previously reported that sexually mature female spontaneously hypertensive rats ( SHRs ) have greater nitric oxide ( NO ) synthase ( NOS ) enzymatic activity in the renal inner medulla ( IM ), compared to age‐matched males. However, the mechanisms responsible for this sexual dimorphism are unknown. The current study tested the hypothesis that sex differences in renal IM NOS activity and NOS 1 expression in adult SHRs develop with sexual maturation and increases in blood pressure (BP) in a female sex hormone‐dependent manner. Methods and Results Renal IM were isolated from sexually immature 5‐week‐old and sexually mature 13‐week‐old male and female SHRs . Whereas NOS activity and NOS 1 expression were comparable in 5‐ and 13‐week‐old male SHRs and 5‐week‐old female SHRs , 13‐week‐old females had greater NOS activity and NOS 1 expression, compared to 5‐week‐old female SHRs and age‐matched males. NOS 3 expression was greater in 5‐week‐old than 13‐week‐old SHRs regardless of sex. Treatment with antihypertensive therapy (hydrochlorothiazide and reserpine) from 6 to 12 weeks of age to attenuate age‐related increases in BP abolished the sex difference in NOS activity and NOS 1 expression between sexually mature SHR males and females. To assess the role of female sex hormones in age‐related increases in NOS , additional females were ovariectomized ( OVX ), and NOS activity was studied 8 weeks post‐ OVX . OVX decreased NOS activity and NOS 1 expression. Conclusions The sex difference in renal IM NOS in SHR is mediated by a sex hormone‐ and BP‐dependent increase in NOS 1 expression and NOS activity exclusively in females.

Th17 cells are pro-inflammatory cells that have been implicated in the pathogenesis of several autoimmune diseases and more recently linked to the progression of hypertension and related end-organ damage. Hv1 is a voltage-gated H+ channel highly expressed in immune cells, including T cells. Hv1-knockout mice have been reported to develop an autoimmune phenotype, including greater activation of circulating CD4+ T-cells and development of nephritis in aged mice, which is more pronounced in females. We have recently developed a Hv1-/- rat on the Dahl salt-sensitive rat (SS) genetic background. The goal of this study was to determine whether T-cells are activated in male and female rats lacking Hv1 and whether this is associated with alterations in blood pressure and/or renal injury. Flow cytometry analysis did not identify a difference in CD4+ T-cells or Th17 cells in whole blood from 12 week old female SS and Hv1-/- SS rats fed either a low (0.4%) or high (8%) NaCl diet for 14 days prior to sacrifice (TABLE...

Acute stress in both rodents and humans causes a transient rise in blood pressure associated with an increase in plasma endothelin-1 (ET-1). High salt (HS) intake also increases ET-1 production, and interestingly, blunts the pressor response to acute air jet stress in rats. We previously reported that female rats lacking functional ETB receptors everywhere except sympathetic nerves (ETB def) had a greater degree of hypertension in response to a HS diet compared to their male counterparts when measured by the tail cuff method. However, we now report that salt-induced hypertension is not different between sexes when measured by telemetry. Therefore, additional experiments were designed to test the hypothesis that female ETB def rats are more sensitive to acute stress when on a HS diet. The pressor response, measured by telemetry, to acute air jet stress was similar between male transgenic control (Tg control) and ETB def rats following chronic HS intake. In contrast, female ETB def rats had a significantly greater pressor response (about twofold higher) than female or male Tg control or male ETB def rats maintained on HS, a finding that cannot be explained by increased vascular reactivity to ET-1 in female rats as we observed that male ETB def rats had a greater pressor response to i.v. infusion of ET-1 compared to females. Furthermore, HS feeding exacerbated the pressor response to ET-1 in both male and female ETB def rats. Given our previous studies demonstrating that the ETA receptor functions to reduce the pressor response to acute stress, these findings further support a role for the ET receptor system in the pressor response to acute stress and that female rats have reduced ETA receptor activity when on a HS diet compared to males.

In experimental animal models of hypertension, angiotensin (1–7) [ANG-(1–7)] is higher in females compared with males; however, it is less clear whether the same applies to humans. Therefore, this study sought to compare circulating concentrations of ANG-(1–7) in apparently healthy men and women under normal physiological conditions. With the use of a cross-sectional experimental design, blood was collected in EDTA anticoagulant from 42 volunteers (21 men and 21 women; and age range, 19–48 yr) for analysis of plasma concentrations of ANG-(1–7) and ANG II. Blood pressure was measured and vascular endothelial function was determined ( n = 25) using the brachial artery flow-mediated dilation (FMD) test. As a result, women exhibited a higher circulating concentration of ANG-(1–7) ( P = 0.04) compared with men, whereas values of ANG II were similar between groups. Baseline arterial diameter, peak diameter, and shear rate were significantly greater ( P < 0.02) in men compared with women. No significant differences in FMD, FMD normalized for shear, or time to peak dilation were observed between men and women. In addition, a positive correlation between ANG-(1–7) and FMD ( P = 0.04) and negative association between ANG-(1–7) with ANG II ( P = 0.01) were only identified in men, whereas a positive relationship between ANG-(1–7) and diastolic blood pressure ( P = 0.03) was observed in women. In conclusion:, women exhibit significantly higher plasma concentrations of ANG-(1–7) compared with men. In addition, this study describes a relationship between ANG-(1–7), vascular function, and diastolic blood pressure that appears to be sex dependent.

The renin angiotensin aldosterone system (RAAS) is critical for controlling blood pressure and body fluid volume and inhibitors of the RAAS are among the most widely used and effective drugs for the treatment of hypertension in both men and women. However, there is an extensive literature detailing sex differences in the RAAS, both in the expression levels of RAAS components and in physiological/pathophysiological responses to RAAS activation. Angiotensin (Ang) II is the primary vasoconstrictor peptide of the RAAS and male experimental animals have greater increases in blood pressure following chronic Ang II infusion than females. This chapter details basic sciences studies that have examined how males vs. females differentially respond to Ang II infusion with the goal of identifying molecular mechanisms that are responsible for the observed sex differences in Ang II hypertension.

Epidemiological evidence suggests that advancing age affects the cardiovascular system of men and women differently. The purpose of this study was to determine whether the effects of aging on nitric oxide synthase (NOS), oxidative stress, and vascular function are different in males and females. Mesenteric arteries from young (3 mo) and old (24 mo) male and female Fischer 344/Brown Norway rats were studied. Western blot analysis and NOS activity were performed on the homogenized mesenteric arterial bed separated into cytosolic and membrane-associated fractions. Plasma 8-isoprostane measurements assessed oxidative stress. Vascular reactivity was determined by using a wire myograph in the absence and presence of a NOS inhibitor, Nω-nitro-l-arginine, to examine endothelial function and basal and stimulated nitric oxide release. In additional arteries, reactivity was performed in the presence of polyethylene glycol-SOD to assess the impact of superoxide on vascular function. Among females, aging was associated with a decline in membrane-associated NOS activity and membrane-associated NOS III protein expression. Advancing age in males was associated with increased cytosolic NOS III protein expression. Among both males and females, advancing age resulted in increased oxidative stress. Vascular function was maintained with age in arteries from both males and females, and there was no difference in either basal or stimulated nitric oxide release with age. Despite sex-specific effects of advancing age on the NOS system and increases in markers of oxidative stress, vascular function is maintained in mesenteric arteries from aged Fischer 344/Brown Norway rats. These data suggest that age-related alterations in the resistance vasculature are complex and likely involve multiple compensating vasoactive pathways.

Endothelium-derived nitric oxide (NO) is a key signalling molecule in the maintenance of cardiovascular health. Endothelial NO synthase (NOS 3), which catalyses the formation of NO, is targeted to the plasma membrane by dual acylation. In vitro studies suggest that membrane localization of NOS 3 is an important regulatory element of NO production. Dysfunction of the vascular endothelium and a decrease in NO bioavailability is associated with the development and progression of a number of cardiovascular diseases, including hypertension. Our laboratory has previously published that in salt-dependent hypertension there is an altered localization of NOS 3, with an increase in cytosolic expression. These data have led us to question whether the increased cytosolic NOS 3 expression is a form of compensation for endothelial dysfunction in hypertension, or an indicator and contributing factor to endothelial dysfunction. This review will outline the importance of subcellular localization in the regulation of NOS 3 in vitro, the role of NOS 3 in endothelial dysfunction associated with salt-dependent hypertension, and the potential physiological consequences of altered NOS 3 localization in vivo.

Recent studies suggest that sex of the animal and T cell impact ANG II hypertension in Rag−/− mice, with females being protected relative to males. This study tested the hypothesis that ANG II results in greater increases in proinflammatory T cells and cytokines in males than in females. Male and female Sprague-Dawley (SD) rats, aged 12 wk, were treated with vehicle or ANG II (200 ng·kg−1·min−1) for 2 wk. Renal CD4+ T cells and Tregs were comparable between vehicle-treated males and females, although males expressed more Th17 and IL-17+ T cells and fewer IL-10+ T cells than females. ANG II resulted in greater increases in CD4+ T cells, Th17 cells, and IL-17+ cells in males; Tregs increased only in females. We previously showed that ANG (1–7) antagonizes ANG II-induced increases in blood pressure in females and ANG (1–7) has been suggested to be anti-inflammatory. Renal ANG (1–7) levels were greater in female SD at baseline and following ANG II infusion. Additional rats were treated with ANG II plus the ANG (1–7)-mas receptor antagonist A-779 (48 μg·kg−1·h−1) to test the hypothesis that greater ANG (1–7) in females results in more Tregs relative to males. Inhibition of ANG (1–7) did not alter renal T cells in either sex. In conclusion, ANG II induces a sex-specific effect on the renal T cell profile. Males have greater increases in proinflammatory T cells, and females have greater increases in anti-inflammatory Tregs; however, sex differences in the renal T cell profile are not mediated by ANG (1–7).

The present study tested the hypothesis that spontaneously hypertensive rats (SHR) have impaired nitric oxide synthase (NOS)‐mediated regulation of vascular function versus Wistar‐Kyoto rats (WKY). Aorta and small mesenteric arteries were studied from male and female SHR (M SHR and F SHR) and WKY (M WKY and F WKY). Phenylephrine (PE)‐induced vasoconstriction was greater in aorta of M SHR versus all others (P < 0.05); there were neither sex nor strain differences in PE contraction in mesenteric arteries. The NOS inhibitor l‐Nitro‐Arginine Methyl Ester (l‐NAME) increased PE‐induced vasoconstriction in all rats, although the increase was the least in male SHR (P < 0.05), revealing a blunted vasoconstrictor buffering capacity of NOS. l‐NAME increased sensitivity to PE‐induced constriction only in mesenteric arteries of SHR, although, the maximal percent increase in contraction was comparable among groups. ACh‐induced relaxation was also less in aorta from M SHR versus all others (P < 0.05). ACh relaxation was comparable among groups in mesenteric arteries, although SHR exhibited a greater NOS component to ACh‐induced relaxation than WKY. To gain mechanistic insight into sex and strain differences in vascular function, NOS activity and NOS3 protein expression were measured. Aortic NOS activity was comparable between groups and M SHR had greater NOS3 expression than M WKY. In contrast, although vascular function was largely maintained in mesenteric arteries of SHR, NOS activity was less in SHR versus WKY. In conclusion, M SHR exhibit a decrease in NOS regulation of vascular function compared to F SHR and WKY, although this is not mediated by decreases in NOS activity and/or expression., The present study tested the hypothesis that spontaneously hypertensive rats (SHR) have impaired nitric oxide synthase (NOS)‐mediated regulation of vascular function versus Wistar‐Kyoto rats (WKY). Aorta and small mesenteric arteries were studied from male and female SHR and WKY. Male SHR showed a decreased NOS regulation of vascular function compared to F SHR and WKY, although this was not mediated by decreases in NOS activity and/or expression.

See related article, pp 384–390 There is an ever expanding literature base implicating T lymphocytes in the development and progression of numerous cardiovascular diseases, including hypertension. T lymphocytes contribute to the development of hypertension in genetic, angiotensin II (Ang-II), and salt-sensitive male experimental animals.1 Among the most definitive studies implicating T lymphocytes in hypertension are studies conducted in Rag-1−/− mice, which lack B and T lymphocytes. Guzik et al2 were the first to demonstrate that these mice have a blunted hypertensive response to Ang-II infusion. Adoptive transfer of T lymphocytes into male Rag−/− mice restored the hypertensive response to Ang-II; adoptive transfer of B lymphocytes did not alter the blood pressure (BP) response. Although low-grade inflammation, and T lymphocytes in particular, are now a recognized hallmark of hypertension, the majority of basic science literature in this field has been conducted exclusively in males, despite the fact that females account for ≈50% of all hypertensive cases in the United States. Therefore, it was with great interest that we read the study by Pollow et al3 in the current issue of Hypertension , which was designed to determine (1) whether there are sex differences in the ability of T lymphocytes to induce Ang-II–dependent hypertension and (2) whether sex affects central or renal T lymphocytes infiltration after Ang-II hypertension. Of particular interest, they found that male mice exhibited a significant increase in BP and renal damage to Ang-II after the adoptive transfer of CD3+ T lymphocytes from wild-type male mice. In contrast, BP responses and renal injury to Ang-II were not significantly altered in female Rag−/− mice after adoptive transfer of T lymphocytes from males. Male Rag−/− mice also had greater renal CD3+, CD4+, CD8+, and T-regulatory cells (Tregs) …

Female spontaneously hypertensive rats (SHR) have more regulatory T cells (Tregs) in their kidneys than males. The goal of this study was to determine the impact of blood pressure (BP) on the renal immune profile. We hypothesize that increases in BP promote a proinflammatory renal T cell and cytokine profile in SHR, although females will have greater hormone-dependent increases in Tregs and males will have greater increases in Th17 cells. Renal T cell and cytokine profiles were assessed in male and female Wistar-Kyoto rats and male and female SHR treated with vehicle or hydrochlorothiazide and reserpine (HCTZ) from 6 to 12 (6-HCTZ) or 11 to 13 weeks of age (2-HCTZ). Regardless of sex, SHR had a more proinflammatory renal immune profile than Wistar-Kyoto rats. 6-HCTZ attenuated age-related increases in BP and 2-HCTZ reversed hypertension compared with vehicle-treated SHR. Neither 6-HCTZ nor 2-HCTZ altered CD3 + , CD4 + , or CD8 + T cells in either sex. Both treatments decreased Tregs only in female SHR abolishing sex differences in Tregs. 6-HCTZ has no impact on Th17 cells in either sex and 2-HCTZ had a minimal impact on renal Th17 cells. To further assess mechanisms mediating sex differences in the renal immune profile, male and female SHR were gonadectomized to determine the impact of sex hormones. Gonadectomy increased proinflammatory markers in both sexes, suggesting that both male and female sex hormones are anti-inflammatory. In conclusion, BP contributes to sex differences in the renal T-cell profile of SHR; female SHR increase renal Tregs in response to increases in BP.

ER stress has recently been linked to the development of hypertension in male SHR. Little is known regarding ER stress in females, although blood pressure (BP) in female SHR is lower than in age-matched males. This study was designed to test the hypothesis that female SHR have less ER stress than males, and consequently will have a smaller decrease in BP in response to an ER stress inhibitor. 11-week old rats were treated with vehicle or the ER stress inhibitor phenylbutyric acid (PBA) for 2 weeks (100 mg/kg/day via drinking water; N=4) and BP was measured weekly via tail cuff. The ER stress markers glucose-regulated protein (GRP78) and C/EBP homologous protein (CHOP) were measured in the renal cortex via Western blot analysis. Basal expression of CHOP was higher in females when compared to males (female: 2.7±0.4 vs. male: 1.3±0.2, P=0.007) as well as with GRP (female: 1.8±0.2 vs. male: 1.3±0.1, P=0.04), however these differences were abolished after treatment. BP did not change in either sex following PB...

The kidney is critical in the long-term control of blood pressure, and renal inflammation has been linked to the progression of hypertension. Previous experiments have shown a sex difference in protein excretion and the renal immune cell profile: male SHRs have greater indices of renal injury and more pro-hypertensive renal T helper17 cells (Th17) while females had more BP-lowering T regulatory cells (Tregs). Male and female SHR were randomized to receive tap water (vehicle) or the BP-lowering drugs, hydrochlorothiazide and reserpine, in drinking water from 5 to 12 weeks of age. BP was measured by tail cuff, and kidneys were isolated for flow cytometric analysis of T cells. Treatment attenuated age-related increases in BP in males (174±4 vs. 137±4 mmHg, p

Angiotensin (Ang) II is pro-inflammatory, and T cells contribute to Ang II hypertension in male animal models; little is known in females. Initial studies treated 12 wk old male and female Sprague ...

Objective: The purpose of this study was to determine whether the effect of age on electrical field stimulation (EFS)-induced endothelium-dependent vasodilation was different in males and females. Methods: Young (3 month) and old (25 month) Fisher 344 rats were studied: young females (YF, n = 34), young males (YM, n = 28), old females (OF, n = 19), and old males (OM, n = 24). Isolated mesenteric resistance arteries (endothelium-intact and denuded) were pressurized, and outer diameter was monitored. EFS-response curves (0.1–8 Hz) were performed in preconstricted arteries in the presence of guanethidine. EFS responses were expressed as percent relaxation from preconstricted diameter. Area under the curve (AUC) was calculated and comparisons were made using ANOVA and t -tests. Results: Males became less responsive to EFS-induced vasodilation with age, while responses among females were unaffected (AUC: YM=344±23, OM=253±25, P = 0.008; YF=397±21, OF=365±25, P = 0.33). Endothelial denudation produced a significant decrease in EFS-induced dilation among all rat groups. The effect of denudation was greater in young animals compared to old. Incubation with the nitric oxide synthase inhibitor N ω-nitro-l-arginine (LNA) significantly decreased EFS responsiveness among all of the rat groups. Conclusions: EFS-induced vasodilation declines with age among males. In YF and YM endothelium-dependent EFS-induced vasodilation is mediated by the release of endothelium-derived NO. With age, endothelial function declines, which is likely due to a decrease in the production of endothelium-derived NO. A decline in endothelial-derived NO is likely responsible for the decrease in EFS-induced vasodilation with age among males.

This year represents the fifth annual Data Diuresis session of the Water and Electrolyte Homeostasis (WEH) section of the American Physiological Society (APS) at the 2015 Experimental Biology meeting. As opposed to taking a single organ approach to the study of physiology, the WEH section employs an integrative approach to encompass how the different organ systems interact to regulate numerous physiological and pathophysiological processes. The goal of this minireview is to highlight the broad spectrum of research themes that were presented over the first five years of Data Diuresis. Presentation topics include (but are not limited to) oxidative stress, inflammation, obesity, pregnancy, and hypertension spanning the brain, heart and vasculature, and kidney. WEH researchers continue to impact and help drive the direction of physiological research across multiple disciplines, leaving us excited to see what the next five years of Data Diuresis will bring.

See related article, pages 768–776 Endothelial dysfunction is seen early in the development of atherosclerosis, before overt vascular and structural changes. Nitric oxide (NO) is recognized as one of the major mediators of the maintenance of vascular homeostasis, and a decrease in NO bioavailability is associated with endothelial dysfunction. Endothelial NO synthase (eNOS; NOS3) catalyzes the formation of NO from L-arginine and O2 in a reaction requiring Ca2+–calmodulin, FAD, FMN, NADPH, and tetrahydrobiopterin (BH4). A decrease in NO bioavailability may be caused by: (1) a decrease in the expression or activity of NOS3, (2) uncoupling of NOS to produce superoxide (O2·−), or (3) degradation of NO by reacting with O2·− from other enzymatic sources resulting in the formation of peroxynitrite (ONOO−). Physiologically, NOS3-derived NO inhibits leukocyte–endothelial cell adhesion, vascular smooth muscle proliferation and migration, and platelet aggregation to maintain the health of the vascular endothelium. Under a number of pathological conditions, NOS3 enzymatic activity becomes uncoupled, resulting in the production of O2·−. NOS3-derived O2·− has been shown to contribute to the development and progression of atherosclerosis and hypertension.1,2 In this issue of Circulation Research , Gharavi et al report that treatment of endothelial cells with oxidized phospholipids results in increased interleukin-8 (IL-8) production through the activation and uncoupling of NOS3.3 When NOS is uncoupled, electrons flowing from the reductase domain to the heme are diverted to molecular oxygen instead of to L-arginine, resulting in the formation of O2·−. A number of potential mechanisms are responsible for uncoupling of NOS3, although the most consistent evidence exists for BH4 deficiency.4,5 Also, NOS3 uncoupling …

Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disorder with a high prevalence of hypertension and cardiovascular disease. Because SLE predominantly affects women, estrogen is commonly implicated as a contributor to SLE disease progression. Using an established mouse model of SLE (female NZBWF1), we tested whether estrogen has a causal role in the development of hypertension in adulthood. Thirty-week-old SLE and control mice (NZW/LacJ) underwent either a sham or ovariectomy (OVX) procedure. 17β-Estradiol (E2; 5 μg/mouse, twice/week, subcutaneously) was administered to a subset of OVX mice. Mean arterial pressure (in mm Hg) was increased in SLE mice (134±4 versus 119±3 in controls). Contrary to our hypothesis, OVX exacerbated the hypertension in female SLE mice (153±3; P0.05 versus SLE sham), and repletion of E2 prevented the OVX-induced increase in blood pressure (132±2). The prevalence of albuminuria was increased in SLE mice compared with controls (37% versus 0%). OVX increased the prevalence in SLE mice (70% versus 37% in SLE shams). Repletion of E2 completely prevented albuminuria in OVX SLE mice. Renal cortical tumor necrosis factor α was increased in SLE mice compared with controls and was further increased in OVX SLE. The OVX-induced increase in renal tumor necrosis factor α expression was prevented by repletion of E2. Treatment of OVX SLE mice with the tumor necrosis factor α inhibitor, etanercept, blunted the OVX-induced increase in blood pressure (140±2) and prevalence of albuminuria (22%). These data suggest that 17β-estradiol protects against the progression of hypertension during adulthood in SLE, in part, by reducing tumor necrosis factor α.

Inflammation plays an important role in the pathogenesis of hypertension, and we recently published that male spontaneously hypertensive rats (SHR) have more pro-hypertensive, pro-inflammatory Th17 cells in their kidneys than females. However, the mechanisms initiating the inflammatory response remain unclear. Damage-associated molecular patterns (DAMPs) are released from dying cells and induce immune system activation. High mobility group box 1 (HMGB1) is a DAMP actively secreted by apoptotic cells in an oxidized form, or passively released by necrotic cells in a reduced form. Reduced HMGB1 has prolonged activity and is more pro-inflammatory. We hypothesized that sex differences in cell death contribute to sex differences in the immune cell profile and hypertension. Renal necrosis and apoptosis were measured by flow cytometry in 12 wk old male and female SHR. Male SHR have greater necrotic cell death (% total renal cells: 5.3±0.8 vs.1.0±0.4; p=0.004; N=6), while female SHR have more apoptosis (% total renal cells: 1.6±0.4 vs.4.0±0.4; p=0.004; N=4-5). Male SHR also had greater reduced HMGB1 in renal cortex compared to females (HMGB1 oxidized/reduced ratio: 1.0±0.1 vs. 1.7±0.1; p=0.002; N=6). We next measured blood pressure by telemetry in male and female SHR treated with vehicle or necrosis inhibitor NecroX-5 (1 mg/kg per day; 2 wks). The renal T cell and macrophage levels were measured by flow cytometry. NecroX-5 treatment in male SHR prevented age-related increases in blood pressure compared to vehicle-treated rats (ΔmmHg/week: 0.8±1.1 vs. 4.2±0.3; p=0.05; N=3). Necrox5 also significantly decreased renal necrosis (% total renal cells: 5.3±0.8 vs. 0.8±0.1; p=0.005; N=3-6), macrophages (% total renal cells: 2.8±0.6 vs. 0.8± 0.1; p=0.04; N=3-4), Th17 cells (% T cells: 4.5±0.3 vs. 2.7±0.3; p=0.009; N=3-4) and the pro-inflammatory cytokine IL-17 (% total renal cells: 10.0±0.4 vs. 5.3±0.7; p=0.002; N=3-4) in male SHR. NecroX-5 had no effect on blood pressure or pro-inflammatory markers in females. We conclude that necrosis is an important contributor in progression of hypertension in male SHR. Sex differences in cell death may contribute to sex differences in blood pressure and the immune profile.

We have previously reported that there are sex differences in the renal T cell profile of spontaneously hypertensive rats (SHR), with females having more anti-inflammatory regulatory T cells (Tregs) and males expressing more pro-inflammatory Th17 T cells. High mobility group box 1 protein (HMGB1) is an endogenous danger molecule and when released by necrotic cells it activates pro-inflammatory immune cells. We hypothesized that males have higher HMGB1 mediated renal T cell activation compared to females. Serum and kidneys were isolated from 13 week old male and female SHR and spleens from normotensive male Wistar Kyoto rats (WKY). HMGB1 expression was measured in sera using an ELISA. To quantitate renal T cell activation, we performed a mixed lymphocyte reaction in the presence and absence of HMGB1 neutralizing antibody. For the reaction, T cells were isolated from kidneys using magnetic activated cell sorting and spleens were homogenized to obtain a single cell suspension of antigen presenting cells (APCs). T cells and APCs were plated together, incubated for 96 hours, proliferation was confirmed and T cell activation was assessed by the magnitude of CD71 using flow cytometry. Male SHR had greater serum HMGB1 levels than female SHR (ng/ml: 5.3±1.1 vs. 2.5±0.3; p=0.0225; N=4-6). In male SHR, 92% of T cells were activated by incubation with APCs in contrast to 65% of T cells in females (CD71 + T cells: 92±1% vs. 65±1%; p

Approximately 30% of American adults are diagnosed with hypertension; however, only ~50% achieve adequate blood pressure (BP) control with current therapeutic options. Preclinical data demonstrate that a sexual dimorphism exists for angiotensin 1-7 (Ang (1-7)), a vasodilatory peptide that has therapeutic promise in controlling BP. This study sought to determine if 1) sex and race differences in Ang (1-7) are present in normotensives adults, and 2) increases in Ang (1-7) are related to lower BP in hypertensive adults. Blood samples, BP, and flow-mediated dilation (FMD, n=29) were assessed in 45 adults (52% women, 44% African American (AA)). Systolic BP was lower (p=0.013) in women compared to men (111±4 mm Hg vs. 126±4 mm Hg) and higher (p=0.024) in AA compared to Caucasians (C; 125±4 mm Hg vs. 111±4 mm Hg). Diastolic BP was higher (p=0.021) in AA compared to C (77±3 mm Hg vs. 65±4 mm Hg). Overall, Ang (1-7) was highest in C women (44±5 pg/ml) followed by AA women (38±5 pg/ml), C men (33±4 pg/ml), and AA men (30±6 pg/ml). Only within C was a significant correlation between FMD and Ang (1-7) observed (r=0.573;p=0.041). Overall, the change in Ang (1-7) following treatment with candesartan (n=6) was positively associated with the change in FMD (r=0.485) and inversely associated with the change in systolic BP (r=-0.431). These data are the first to identify, in humans, the existence of a sexual and racial dimorphism in Ang (1-7). Our findings support the therapeutic potential of Ang (1-7) to reduce BP and improve endothelial function in hypertensive adults.

T cells contribute to angiotensin (Ang) II hypertension in male animal models yet less is known in females. We previously showed that hypertensive female rats have more immuno-suppressive T regulatory cells (Tregs) and males have more pro-inflammatory Th17 cells in their kidneys. This study tested the hypothesis that Ang II results in greater expression of renal Tregs in females and Th17 cells in males. 12 wk old male and female Sprague Dawley rats (SD) were treated with Ang II (200 ng/kg/min) or vehicle for 2 wks; blood pressure (BP) was measured via tail cuff and renal T cell profiles were measured via flow cytometry. Ang II infusion increased BP in both sexes (mmHg: males: 130±2 to 152±2; females: 123±1 to 146±1; p+ T cell counts at baseline vs. males (p+ T cells in both sexes (p+ T cell counts were comparable between male and female SD. Ang II infusion increased CD4 + T cells in both sexes (p

Nitric oxide is a critical regulator of blood pressure (BP) and inflammation, and female spontaneously hypertensive rats (SHR) have higher renal nitric oxide bioavailability than males. We hypothesize that female SHR will have a greater rise in BP and renal T cell infiltration in response to nitric oxide synthase (NOS) inhibition than males. Both male and female SHR displayed a dose-dependent increase in BP to the nonspecific NOS inhibitor NG-nitro-l-arginine methyl ester (l-NAME: 2, 5, and 7 mg·kg−1·day−1 for 4 days each); however, females exhibited a greater increase in BP than males. Treatment of male and female SHR with 7 mg·kg−1·day−1 l-NAME for 2 wk significantly increased BP in both sexes; however, prior exposure to l-NAME only increased BP sensitivity to chronic NOS inhibition in females. l-NAME-induced hypertension increased renal T cell infiltration and indices of renal injury in both sexes, yet female SHR exhibited greater increases in Th17 cells and greater decreases in regulatory T cells than males. Chronic l-NAME was also associated with larger increases in renal cortical adhesion molecule expression in female SHR. The use of triple therapy to block l-NAME-mediated increases in BP attenuated l-NAME-induced increases in renal T cell counts and normalized adhesion molecule expression in SHR, suggesting that l-NAME-induced increases in renal T cells were dependent on both increases in BP and NOS inhibition. Our data suggest that NOS is critical in the ability of SHR, females in particular, to maintain BP and limit a pro-inflammatory renal T cell profile.

Historically, tools to assess renal function have been developed to investigate the physiology of the kidney in an experimental setting, and certain of these techniques have utility in evaluating renal function in the clinical setting. The following work will survey a spectrum of these tools, their applications and limitations in four general sections. The first is clearance, including evaluation of exogenous and endogenous markers for determining glomerular filtration rate, the adaptation of estimated glomerular filtration rate in the clinical arena, and additional clearance techniques to assess various other parameters of renal function. The second section deals with in vivo and in vitro approaches to the study of the renal microvasculature. This section surveys a number of experimental techniques including corticotomy, the hydronephrotic kidney, vascular casting, intravital charge coupled device videomicroscopy, multiphoton fluorescent microscopy, synchrotron-based angiography, laser speckle contrast imaging, isolated renal microvessels, and the perfused juxtamedullary nephron microvasculature. The third section addresses in vivo and in vitro approaches to the study of renal blood flow. These include ultrasonic flowmetry, laser-Doppler flowmetry, magnetic resonance imaging (MRI), phase contrast MRI, cine phase contrast MRI, dynamic contrast-enhanced MRI, blood oxygen level dependent MRI, arterial spin labeling MRI, x-ray computed tomography, and positron emission tomography. The final section addresses the methodologies of metabolic balance studies. These are described for humans, large experimental animals as well as for rodents. Overall, the various in vitro and in vivo topics and applications to evaluate renal function should provide a guide for the investigator or physician to understand and to implement the techniques in the laboratory or clinic setting.

Tetrahydrobiopterin Supplementation Increases Renal Inner Medullary Nitric Oxide Synthase Enzymatic Activity in Male, But Not Female Spontaneously Hypertensive Rats Krystal N. Brinson and Jennifer C. Sullivan, Experimental Medicine, Georgia Health Sciences University, Augusta, GA 30912 We previously published that male spontaneously hypertensive rats (SHR) having less nitric oxide synthase (NOS) enzymatic activity in the renal inner medullary (IM) than female SHR, which cannot be explained by protein expression alone. Tetrahydrobiopterin (BH4) is an essential cofactor for NOS activity. BH4 is easily oxidized to BH2, which is not a NOS cofactor, and male SHR have greater levels of oxidative stress than females. Therefore, we hypothesized that 1) male SHR will have greater increases in NOS enzymatic activity following BH4 supplementation than females and 2) antioxidant treatment will result in greater increases in NOS enzymatic activity in male SHR regardless of BH4 supplementation. Studies measured renal IM NOS enzymatic activity in the presence and absence of BH4 from 12-13 week old control (N=5-6) or antioxidant treated (tempol, 30 mg/kg/day in drinking water) male and female SHR (N=4-5). Supplementation with BH4 increased NOS activity in male SHR (177±35 vs. 315±52 pmol/mg; p=0.04), with no effect in females (434±112 vs. 548±84 pmol/mg). Following 2 weeks of antioxidant treatment, NOS enzymatic activity in male SHR was significantly increased compared to control SHR (p=0.04), and supplementation with BH4 no longer altered NOS enzymatic activity in males (588±143 vs. 648±153 pmol/mg). Antioxidant treatment did not significantly alter renal IM NOS enzymatic activity in female SHR compared to untreated controls, and BH4 supplementation did not alter NOS activity in tempol-treated females (473±82 vs. 580±66 pmol/mg). Tempol treatment also abolished the sex difference in renal IM NOS enzymatic activity. In conclusion, our data suggest that greater levels of oxidative stress in male SHR contributes to lower levels of NOS enzymatic activity compared to females. We hypothesize that this results in greater BH4 deficiency in males contributing to decreases in NO and increases in blood pressure.

The prevalence of cardiovascular disease is greater in men than in women, and although the mechanism responsible is unknown, sex differences in Angiotensin (Ang) peptides have been suggested to contribute. In fact, evidence in rats indicates Ang (1-7), a vasodilatory peptide, is higher in females when compared to males. In humans, the flow-mediated dilation (FMD) test represents a bioassay of NO-dependent vasodilation and is a predictor of future cardiovascular disease. This study sought to test the hypotheses that 1) Ang (1-7) is higher in women compared to men, and 2) determine if relationships exist among concentrations of Ang (1-7), FMD, and blood pressure. A total of 29 subjects participated in this study; FMD was obtained on 14 (7 men; 7 women). Following an overnight fast, blood pressure and FMD were performed and blood samples were collected. Systolic blood pressure was greater in men (121±5 mm Hg) compared to women (101±3 mm Hg; p=0.01); however, no difference in diastolic blood pressure was observed (66±4 vs. 63±2 mm Hg, respectively). Plasma concentrations of Ang (1-7) were significantly higher in women compared to men (43±5 vs. 32±4 pg/mL; p=0.04). FMD was positively associated with Ang (1-7) independent of sex (r=0.592; p=0.02); however, the relationship between Ang (1-7) and diastolic blood pressure was evident only in women (women r=-0.717; p=0.07 and men r=0.186; p=0.69). These data support a sexual dimorphism in Ang (1-7) in humans, similar to what we have observed previously in our animal model. For the first time in humans, we have also observed a sex dependent relationship between Ang (1-7) and diastolic blood pressure supporting the hypotheses that Ang (1-7) may play an important role in BP regulation and maintenance of endothelial function particularly in women. These data may provide insight into the cardiovascular disease risk disparity between sexes.

Lymphocytes contribute to hypertension in male spontaneously hypertensive rats (SHR); data in females is lacking, yet young females have a lower blood pressure compared to age-matched males. We hypothesized that 12 week old female SHR have less T cell infiltration and cytokine expression in the kidney compared to males. Kidneys were isolated from male and female SHR and normotensive Wistar Kyoto rats (WKY) and the T cell profile was determined using analytical flow cytometry. In both sexes of SHR, all T cell subtypes and cytokines measured were significantly higher in the kidney compared to the same sex WKY (p+ T cell infiltration vs. female SHR (p+ T cell numbers were higher in female SHR (p=0.0114). Analysis of T cell subpopulations showed female SHR has greater expression of regulatory T cells (Foxp3 + CD4 + CD3 + ) than male SHR (p=0.0008), however, male SHR had more infiltrating pro-inflammatory Th17 cells (RORγ + CD4 + CD3 + ; p+ ; p=0.0009, n=3-6), CD4 + T cell (p=0.0010) and Th17 cell (p=0.0025) infiltration into the kidneys. Infiltration of CD8 + and Foxp3 + cells remained comparable between groups. Together these data suggest that hypertension in both sexes are associated with an increase in renal inflammatory markers; however female SHR have a more anti-inflammatory profile vs. males which may contribute to their lower blood pressure.

We recently demonstrated that female spontaneously hypertensive rats (SHR) have higher nitric oxide (NO) bioavailability in the kidney compared to males. Therefore, we hypothesize that female SHR are more dependent on the NO synthase (NOS) system to modulate their blood pressure, and as a result will have a greater rise in blood presure and renal injury in response to chronic NOS inhibition compared to males. Mean arterial pressure in SHR was very sensitive to NOS inhibition, with both sexes exhibiting a significant increase in blood pressure to the non-specific NOS inhibitor N G -nitro-L-arginine methyl ester (L-NAME) at a dose of 2 mg/kg/day. However, there was not a sex difference in the percent increase in mean arterial pressure. Treatment of female and male SHR with 7 mg/kg/day L-NAME for 2 weeks significantly increased mean arterial pressure and indices of renal injury in both females and males; the percent increases in mean arterial pressure and injury were greater in females during the last three days of L-NAME treatment (% increase in mean arterial pressure was 44± 1.3 in females vs. 35± 2.9 in males, Pvs. 4.6± 0.1 to 7.9± 0.3 ng/mg in females, Pvs. males following L-NAME treatment, the increase in cortical T cell infiltration and sICAM-1 were greater in males. These data suggest that the NO system plays an important role in modulating blood pressure, renal injury and inflammation in SHR, with females being more dependent on the NOS system than males.

T cells contribute to hypertension in male experimental models; data in females is lacking even though women are more likely to develop immune disorders. The goal of this study was to determine whether immune cells contribute to hypertension in female spontaneously hypertensive rats (SHR) and define the T cell profile in whole blood and kidneys of male and female SHR. We hypothesized that inflammatory cells contribute to hypertension in female SHR; however, male SHR have a higher blood pressure so we hypothesize they will have a heightened inflammatory profile. The lymphocyte inhibitor mycophenolate mofetil (MMF) was administered in a dose-dependent manner to SHR. At the highest dose (50 mg·kg−1·day−1), blood pressure was significantly decreased in both sexes, yet the percent decrease in blood pressure was greater in females (female: 12 ± 1%; males: 7 ± 1%, P = 0.01). Circulating and renal T cell profiles were defined using analytical flow cytometry. Female SHR had more circulating CD3+, CD4+, and pro-inflammatory CD3+CD4+RORγ+ Th17 cells, whereas males had more immune-suppressive CD3+CD4+Foxp3+ T regulatory cells. In the kidney, females had greater numbers of CD8+ and T regulatory cells than males, whereas males had greater CD4+ and Th17 cell infiltration. MMF decreased circulating and renal T cells in both sexes ( P < 0.0001), although the effect of MMF on T cell subtypes was sex specific with females having greater sensitivity to MMF-induced decreases in lymphocytes. In conclusion, there is a lymphocyte contribution to the maintenance of hypertension in the female SHR and sex of the animal impacts the T cell profile.

Induction of hemeoxygenase-1 (HO-1) lowers blood pressure and reduces organ damage in hypertensive animal models; however, a potential protective role for HO-1 induction against diabetic-induced glomerular injury remains unclear. We hypothesize that HO-1 induction will protect against diabetes-induced glomerular injury by maintaining glomerular integrity and inhibiting renal apoptosis, inflammation, and oxidative stress. Diabetes was induced with streptozotocin in spontaneously hypertensive rats (SHR) as a model where the coexistence of hypertension and diabetes aggravates the progression of diabetic renal injury. Control and diabetic SHR were randomized to receive vehicle or the HO-1 inducer cobalt protoporphyrin (CoPP). Glomerular albumin permeability was significantly greater in diabetic SHR compared with control, consistent with an increase in apoptosis and decreased glomerular nephrin and α3β1-integrin protein expression in diabetic SHR. CoPP significantly reduced albumin permeability and apoptosis and restored nephrin and α3β1-integrin protein expression levels in diabetic SHR. Glomerular injury in diabetic SHR was also associated with increases in NF-κB-induced inflammation and oxidative stress relative to vehicle-treated SHR, and CoPP significantly blunted diabetes-induced increases in glomerular inflammation and oxidative stress in diabetic SHR. These effects were specific to exogenous stimulation of HO-1, since incubation with the HO inhibitor stannous mesoporphyrin alone did not alter glomerular inflammatory markers or oxidative stress yet was able to prevent CoPP-mediated decreases in these parameters. These data suggest that induction of HO-1 reduces diabetic induced-glomerular injury and apoptosis and these effects are associated with decreased NF-κB-induced inflammation and oxidative stress.

NADPH oxidase has been implicated in ANG II-induced oxidative stress and hypertension in males; however, the contribution of oxidative stress to ANG II hypertension in females is unknown. In the present study, we tested the hypothesis that greater antioxidant capacity in female spontaneously hypertensive rats (SHR) blunts ANG II-induced oxidative stress and hypertension relative to males. Whole body and renal cortical oxidative stress levels were assessed in female and male SHR left untreated or following 2 wk of chronic ANG II infusion. Chronic ANG II infusion increased NADPH oxidase enzymatic activity in the renal cortex of both sexes; however, this increase only reached significance in female SHR. In contrast, male SHR demonstrated a greater increase in all measurements of reactive oxygen species production in response to chronic ANG II infusion. ANG II infusion increased plasma superoxide dismutase activity only in female SHR (76 ± 9 vs. 190 ± 7 Units·ml−1·mg−1, P < 0.05); however, cortical antioxidant capacity was unchanged by ANG II in either sex. To assess the functional implication of alterations in NADPH enzymatic activity and oxidative stress levels following ANG II infusion, additional experiments assessed the ability of the in vivo antioxidant apocynin to modulate ANG II hypertension. Apocynin significantly blunted ANG II hypertension in male SHR (174 ± 2 vs. 151 ± 1 mmHg, P < 0.05), with no effect in females (160 ± 11 vs. 163 ± 10 mmHg). These data suggest that ANG II hypertension in male SHR is more dependent on increases in oxidative stress than in female SHR.

To the Editor: Oxidative stress has been implicated in the pathogenesis of renal and cardiovascular diseases; however, there is still much to be learned. It is against this background that we read with great interest the article titled, “Superoxide Dismutase 1 Limits Renal Microvascular Remodeling and Attenuates Arteriole and Blood Pressure Responses to Angiotensin II via Modulation of Nitric Oxide Bioavailability,” by Carlstrom et al.1 The goal of this article was to determine the functional role of superoxide dismutase 1 (SOD1) in regulating renal microvascular function and blood pressure responses to angiotensin (Ang) II. The authors found that SOD1 …

We previously reported that small mesenteric arteries from hypertensive rats have increased NOS-derived H2O2 and reduced NO/cGMP signaling. We hypothesized that antihypertensive therapy lowers blood pressure through a tetrahydrobiopterin (BH4)-dependent mechanism restoring NO/cGMP signaling and endothelial NOS (NOS3; eNOS) phosphorylation in small arteries. To test this hypothesis, small mesenteric arteries from normotensive rats (NORM), angiotensin II-infused rats (ANG), ANG rats with triple therapy (reserperine, hydrochlorothiazide, and hydralazine), or ANG rats with oral BH4 therapy were studied. Both triple therapy and oral BH4 therapy attenuated the rise in systolic blood pressure in ANG rats and restored NO/cGMP signaling in small arteries similarly. Triple therapy significantly increased vascular BH4 levels and BH4-to-BH2 ratio similar to ANG rats with BH4 supplementation. Furthermore, triple therapy (but not oral BH4 therapy) significantly increased GTP cyclohydrolase I (GTPCH I) activity in small arteries without a change in expression. NOS3 phosphorylation at Ser1177 was reduced in small arteries from ANG compared with NORM, while NOS3 phosphorylation at Ser633 and Thr495 were similar in ANG and NORM. NOS3 phosphorylation at Ser1177 was restored with triple therapy or oral BH4 in ANG rats. In conclusion, antihypertensive therapy regulates NO/cGMP signaling in small arteries through increasing BH4 levels and NOS3 phosphorylation at Ser1177.

The prevalence of diabetes has dramatically increased worldwide due to the vast increase in the obesity rate. Diabetic nephropathy is one of the major complications of type 1 and type 2 diabetes and it is currently the leading cause of end-stage renal disease. Hyperglycemia is the driving force for the development of diabetic nephropathy. It is well known that hyperglycemia increases the production of free radicals resulting in oxidative stress. While increases in oxidative stress have been shown to contribute to the development and progression of diabetic nephropathy, the mechanisms by which this occurs are still being investigated. Historically, diabetes was not thought to be an immune disease; however, there is increasing evidence supporting a role for inflammation in type 1 and type 2 diabetes. Inflammatory cells, cytokines, and profibrotic growth factors including transforming growth factor-β (TGF-β), monocyte chemoattractant protein-1 (MCP-1), connective tissue growth factor (CTGF), tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1), interleukin-6 (IL-6), interleukin-18 (IL-18), and cell adhesion molecules (CAMs) have all been implicated in the pathogenesis of diabetic nephropathy via increased vascular inflammation and fibrosis. The stimulus for the increase in inflammation in diabetes is still under investigation; however, reactive oxygen species are a primary candidate. Thus, targeting oxidative stress-inflammatory cytokine signaling could improve therapeutic options for diabetic nephropathy. The current review will focus on understanding the relationship between oxidative stress and inflammatory cytokines in diabetic nephropathy to help elucidate the question of which comes first in the progression of diabetic nephropathy, oxidative stress, or inflammation.

High soy diets reduce injury in rat models of focal cerebral ischemia and are proposed as alternatives to hormone replacement therapy for postmenopausal women. The present study tests the hypothesis that the major soy isoflavone genistein and the daidzein metabolite equol are neuroprotective in transient focal cerebral ischemia in male and ovariectomized (OVX) female rats by inhibiting oxidative stress. Genistein is the primary circulating soy isoflavone in humans, whereas equol is the primary circulating isoflavone in rats. Male and OVX female Sprague-Dawley rats were fed an isoflavone-reduced diet alone or supplemented with genistein (500 ppm) or equol (250 ppm) for 2 wk prior to 90-min transient middle cerebral artery occlusion followed by reperfusion under isoflurane anesthesia. Indices of oxidative stress were determined 24 h after reperfusion, and cerebral injury was evaluated 3 days after reperfusion. Genistein and equol significantly reduced infarct size in both sexes. Further studies in OVX female rats revealed that this neuroprotection was accompanied by a decrease in NAD(P)H oxidase activity and superoxide levels in the brain. In addition, equol reduced plasma thiobarbituric acid reactive substances, and neurological deficits up to 7 days after injury. There were no significant differences in cerebral blood flow among treatment groups. In conclusion, dietary soy isoflavones are neuroprotective in transient focal cerebral ischemia in male and OVX female rats. These isoflavones may protect the brain via increases in endogenous antioxidant mechanisms and reduced oxidative stress.