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Circadian disruption is a disturbance in biological timing, which can occur within or between different organizational levels, ranging from molecular rhythms within specific cells to the misalignment of behavioral and environmental cycles. Previous work from our group showed that less than 1 wk of food restriction to the light (inactive) period is sufficient to invert diurnal blood pressure rhythms in mice. However, kidney excretory rhythms and functions remained aligned with the light-dark cycle. Shift workers have an increased risk of cardiovascular disease that may different between sexes and often have irregular mealtimes, making the possibility of mistimed feeding as a potential contributor to the development of kidney disease. Thus, we hypothesized that chronic mistimed food intake would result in adverse cardiorenal effects, with sex differences in severity. Here, we show that chronic circadian disruption via mistimed feeding results in renal fibrosis and aortic stiffness in a sex-dependent manner. Our results indicate the importance of meal timing for the maintenance of blood pressure rhythms and kidney function, particularly in males. Our results also demonstrate that females are better able to acclimate to circadian-related behavioral change. NEW & NOTEWORTHY Circadian disruption through mistimed feeding resulted in nondipping blood pressure, renal fibrosis, and arterial stiffness that were less severe in females versus males. Mice fed exclusively during the daytime maintain their circadian rhythms of locomotor activity regardless of their loss of blood pressure rhythms. Although these mice ate less food, they maintained body weight, suggesting inefficiencies in overall metabolism. These findings demonstrate the importance of maintaining optimal food intake patterns to prevent cardiorenal pathophysiology.

Activation of G protein-coupled estrogen receptor 1 (GPER1) elicits antihypertensive actions in different animal models. The endothelin-1 signaling system plays a fundamental role in blood pressure regulation. Lack of functional endothelin receptor B (ET B ) evokes hypertension and salt sensitivity. GPER1 and ET B interact to promote urinary sodium excretion in female rats. We hypothesized that activation of GPER1 protects against hypertension and salt sensitivity induced by ET B antagonism in female rats. Female Sprague-Dawley rats were implanted with radiotelemetry. Animals were then subjected to ovariectomy and simultaneously implanted with minipumps to deliver either the GPER1 agonist G1 or its corresponding vehicle. Two weeks post surgery, we initiated treatment of rats with the ET B antagonist A-192621. Animals were maintained on a normal-salt diet and then challenged with a high-salt diet for an additional 5 days. Assessment of mean arterial blood pressure revealed an increase in vehicle-treated, but not G1-treated, rats in response to ovariectomy. A-192621 increased blood pressure in normal-salt diet-fed vehicle- and G1-treated rats. G1 improved the circadian blood pressure rhythms that were disrupted in A-192621-treated ovariectomized rats. Thus, although systemic GPER1 activation did not protect ovariectomized rats from hypertension and salt sensitivity induced by ET B antagonism, it maintained circadian blood pressure rhythms. Functional ET B is required to elicit the antihypertensive actions of GPER1. Additional studies are needed to improve our understanding of the interaction between G protein-coupled receptors in regulating circadian blood pressure rhythm. NEW & NOTEWORTHY Systemic G protein-coupled estrogen receptor 1 (GPER1) activation in rats prevents the increase in blood pressure evoked by ovariectomy. Blockade of endothelin receptor B negates the blood pressure-lowering impact of GPER1 in ovariectomized rats. Endothelin receptor B plays an important role in mediating the blood pressure-lowering action of GPER1 activation in female rats.

Persistent albuminuria (PA) is common in sickle cell anaemia (SCA). With the association of chronic kidney disease (CKD) with increased mortality, biomarkers that predict its development or progression are needed. We evaluated the association of select biomarkers with PA in adults with SCA using Kruskal-Wallis rank-sum test and logistic regression models, with adjustment for multiple testing. Of 280 subjects, 100 (35.7%) had PA. Median plasma levels of soluble vascular cell adhesion molecule-1 (VCAM-1) (1176.3 vs. 953.4 ng/mL, false discovery rate [FDR] q-value <0.003), thrombin-antithrombin complex (5.5 vs. 4.7 ng/mL, FDR q-value = 0.04), and urinary angiotensinogen (AGT) (12.2 vs. 5.3 ng/mg, FDR q-value <0.003), urinary nephrin (30.6 vs. 27.2 ng/mg, FDR q-value = 0.04), and urinary kidney injury molecule-1 (KIM-1) (0.8 vs. 0.5 ng/mg, FDR q-value <0.003), normalized to urine creatinine, were significantly higher in subjects with PA. In multivariable analysis, only urinary AGT (odds ratio = 1.058, FDR q-value <0.0001) remained a significant predictor of PA. In addition, soluble VCAM-1 (FDR q-value <0.0001), D-dimer (FDR q-value <0.0001), urinary AGT (FDR q-value <0.0001), KIM-1 (FDR q-value <0.0001), and nephrin (FDR q-value <0.0001) were significantly associated with urine albumin-creatinine ratio in multivariable analyses. Longitudinal studies to evaluate the predictive capacity of biomarkers for the development and progression of CKD in SCA are warranted., (© 2024 British Society for Haematology and John Wiley & Sons Ltd.)

Aim: We aimed to test the hypothesis that a high-salt diet (HS) impairs NO signaling in kidney microvascular endothelial cells through a histone deacetylase 1 (HDAC1)-dependent mechanism., Methods: Male Sprague Dawley rats were fed normal salt diet (NS; 0.49% NaCl) or HS (4% NaCl) for 2 weeks. NO signaling was assessed by measuring L-NAME induced vasoconstriction of the afferent arteriole using the blood perfused juxtamedullary nephron (JMN) preparation. In this preparation, kidneys were perfused with blood from a donor rat on a matching or different diet to that of the kidney donor. Kidney endothelial cells were isolated with magnetic activated cell sorting and HDAC1 activity was measured., Results: We found HS-induced impaired NO signaling in the afferent arteriole. This was restored by inhibition of HDAC1 with MS-275. Consistent with these findings, HDAC1 activity was increased in kidney endothelial cells. We further found the loss of NO to be dependent upon the diet of the blood donor rather than the diet of the kidney donor and the plasma from HS-fed rats to be sufficient to induce impaired NO signaling. This indicates the presence of a humoral factor we termed plasma-derived endothelial dysfunction mediator (PDEM). Pretreatment with the antioxidants, PEG-SOD and PEG-catalase, as well as the NOS cofactor, tetrahydrobiopterin, restored NO signaling., Conclusion: We conclude that HS activates endothelial HDAC1 through PDEM leading to decreased NO signaling. This study provides novel insights into the molecular mechanisms by which a HS decreases renal microvascular endothelial NO signaling., (© 2024 The Author(s). Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.)

Competing Interests: D.M.P. holds the position of Editor-in-Chief for Function and is blinded from reviewing or making decisions for the manuscript.

ET (endothelin) is a powerful vasoconstrictor 21-amino acid peptide present in many tissues, which exerts many physiological functions across the body and participates as a mediator in many pathological conditions. ETs exert their effects through ET A and ET B receptors, which can be blocked by selective receptor antagonists. ETs were shown to play important roles among others, in systemic hypertension, particularly when resistant or difficult to control, and in pulmonary hypertension, atherosclerosis, cardiac hypertrophy, subarachnoid hemorrhage, chronic kidney disease, diabetic cardiovascular disease, scleroderma, some cancers, etc. To date, ET antagonists are only approved for the treatment of primary pulmonary hypertension and recently for IgA nephropathy and used in the treatment of digital ulcers in scleroderma. However, they may soon be approved for the treatment of patients with resistant hypertension and different types of nephropathy. Here, the role of ETs is reviewed with a special emphasis on participation in and treatment of hypertension and chronic kidney disease., Competing Interests: E.L. Schiffrin has participated in advisory boards of Janssen Pharmaceuticals, USA. The other author reports no conflicts.

Behavior and function of living systems are synchronized by the 24-h rotation of the Earth that guides physiology according to time of day. However, when behavior becomes misaligned from the light-dark cycle, such as in rotating shift work, jet lag, and even unusual eating patterns, adverse health consequences such as cardiovascular or cardiometabolic disease can arise. The discovery of cell-autonomous molecular clocks expanded interest in regulatory systems that control circadian physiology including within the kidney, where function varies along a 24-h cycle. Our understanding of the mechanisms for circadian control of physiology is in the early stages, and so the present review provides an overview of what is known and the many gaps in our current understanding. We include a particular focus on the impact of eating behaviors, especially meal timing. A better understanding of the mechanisms guiding circadian function of the kidney is expected to reveal new insights into causes and consequences of a wide range of disorders involving the kidney, including hypertension, obesity, and chronic kidney disease.

Introduction: Current understanding of sodium (Na + ) handling is based on studies done primarily in males. Contrary to the gradual increase in high salt (HS) induced natriuresis over 3-5 days in males, female Sprague Dawley (SD) rats have a robust natriuresis after 1 day of HS. Renal endothelin-1 (ET-1) signaling, through ET receptor A and B, is an important natriuretic pathway and was implicated in our previous dietary salt acclimation studies, however, the contribution of ET receptors to sex-differences in acclimation to dietary Na + challenges has yet to be clarified. We hypothesized that ET receptors mediate the augmented natriuretic capacity of female rats in response to a HS diet., Methods: To test our hypothesis, male and female SD rats were implanted with telemeters and randomly assigned to treatment with A-182086, a dual ET A and ET B receptor antagonist, or control. 24-h urine samples were collected and assessed for electrolytes and ET-1. Studies were performed on a normal salt (NS, 0.3% NaCl) diet and after challenging rats with HS (4% NaCl) diet for 1 day., Results: We found that A-182086 increased blood pressure in male and female SD rats fed either diet. Importantly, A-182086 eliminated sex-differences in natriuresis on NS and HS. In particular, A-182086 promotes HS-induced natriuresis in male rats rather than attenuating the natriuretic capacity of females. Further, the sex-difference in urinary ET-1 excretion in NS-fed rats was eliminated by A-182086., Conclusion: In conclusion, ET receptors are crucial for mediating sex-difference in the natriuretic capacity primarily through their actions in male rats., (© 2023. Society for Women's Health Research and BioMed Central Ltd.)

Renal nerves have been an attractive target for interventions aimed at lowering blood pressure; however, the specific roles of renal afferent (sensory) versus efferent sympathetic nerves in mediating hypertension are poorly characterized. A number of studies have suggested that a sympathoexcitatory signal conveyed by renal afferents elicits increases in blood pressure, whereas other studies identified sympathoinhibitory afferent pathways. These sympathoinhibitory pathways have been identified as protective against salt-sensitive increases in blood pressure through endothelin B (ET B ) receptor activation. We hypothesized that ET B -deficient (ET B -def) rats, which are devoid of functional ET B receptors except in adrenergic tissues, lack appropriate sympathoinhibition and have lower renal afferent nerve activity following a high-salt diet compared with transgenic controls. We found that isolated renal pelvises from high salt-fed ET B -def animals lack a response to a physiological stimulus, prostaglandin E 2 , compared with transgenic controls but respond equally to a noxious stimulus, capsaicin. Surprisingly, we observed elevated renal afferent nerve activity in intact ET B -def rats compared with transgenic controls under both normal- and high-salt diets. ET B -def rats have been previously shown to have heightened global sympathetic tone, and we also observed higher total renal sympathetic nerve activity in ET B -def rats compared with transgenic controls under both normal- and high-salt diets. These data indicate that ET B receptors are integral mediators of the sympathoinhibitory renal afferent reflex (renorenal reflex), and, in a genetic rat model of ET B deficiency, the preponderance of sympathoexcitatory renal afferent nerve activity prevails and may contribute to hypertension. NEW & NOTEWORTHY Here, we found that endothelin B receptors are an important contributor to renal afferent nerve responsiveness to a high-salt diet. Rats lacking endothelin B receptors have increased afferent nerve activity that is not responsive to a high-salt diet.

Environmental cues such as light and timing of food intake influence molecular clocks that produce circadian rhythmicity of many biological functions. The master circadian clock is entrained by light input and synchronizes with peripheral clocks in every organ of the body. Careers that require rotating shift work schedules predispose workers to a constant desynchronization of these biological clocks and are associated with increased risk of cardiovascular disease. We used a stroke-prone spontaneously hypertensive rat model exposed to a known biological desynchronizer, chronic environmental circadian disruption (ECD), to test the hypothesis that it would accelerate the time to stroke onset. We then investigated whether time-restricted feeding could delay stroke onset and evaluated its usefulness as a countermeasure when combined with the constant disruption of the light cycle. We found that phase advancing of the light schedule accelerated stroke onset. Restricting food access time to 5 h/day regardless of lighting profoundly delayed stroke onset in both standard 12-h:12-h light/dark or ECD-lighting conditions compared with ad libitum feeding; however, acceleration by ECD versus control lighting conditions was still observed. Since hypertension is a precursor to stroke in this model, we assessed blood pressure in a small cohort longitudinally using telemetry. Mean daily systolic and diastolic blood pressure increased in a similar manner across rats in control and ECD conditions, thus hypertension was not grossly accelerated to cause earlier strokes. However, we observed intermittent dampening of rhythms after each shift of the light cycle reminiscent of a relapsing-remitting nondipping state. Our results suggest that constant disruption of environmental rhythms may be associated with an increased risk of cardiovascular complications in the presence of cardiovascular risk factors. NEW & NOTEWORTHY This stroke-prone spontaneously hypertensive rat model significantly delayed stroke onset with the timed food restriction intervention. Blood pressure recordings in this same model were continuous through the 3 mo and showed dampened systolic rhythms after each shift in the lighting schedule.

Despite the overwhelming evidence that the kidney is the principal regulator of chronic blood pressure though the ability to sense pressure and adjust blood volume accordingly, recent clinical and preclinical evidence suggests that skin clearance of Na+ through sweat significantly contributes to long-term blood pressure and risk of hypertension. Evidence indicates that changes in skin Na+ content negatively associate with renal function, and factors that influence the concentration of Na+ in sweat are affected by major regulators of Na+ excretion by the kidney such as angiotensin and aldosterone. In addition, known regulatory mechanisms that regulate the amount of sweat produced do not include changes in Na+ intake or blood volume. Because of these reasons, it will be hard to quantify the contribution of Na+ clearance through sweat to blood pressure regulation and hypertension. While Chen et al. demonstrate significant negative associations between sweat Na+ concentration and blood pressure, it is likely that Na+ clearance through the skin has a short-term influence on blood pressure and sweat Na+ concentration is most likely a biomarker of renal function and its key role in hypertension., (© 2023 The Author(s).)

Healthy individuals exhibit blood pressure variation over a 24-hour period with higher blood pressure during wakefulness and lower blood pressure during sleep. Loss or disruption of the blood pressure circadian rhythm has been linked to adverse health outcomes, for example, cardiovascular disease, dementia, and chronic kidney disease. However, the current diagnostic and therapeutic approaches lack sufficient attention to the circadian rhythmicity of blood pressure. Sleep patterns, hormone release, eating habits, digestion, body temperature, renal and cardiovascular function, and other important host functions as well as gut microbiota exhibit circadian rhythms, and influence circadian rhythms of blood pressure. Potential benefits of nonpharmacologic interventions such as meal timing, and pharmacologic chronotherapeutic interventions, such as the bedtime administration of antihypertensive medications, have recently been suggested in some studies. However, the mechanisms underlying circadian rhythm-mediated blood pressure regulation and the efficacy of chronotherapy in hypertension remain unclear. This review summarizes the results of the National Heart, Lung, and Blood Institute workshop convened on October 27 to 29, 2021 to assess knowledge gaps and research opportunities in the study of circadian rhythm of blood pressure and chronotherapy for hypertension.

Objective: Early life stress (ELS) occurring during childhood and adolescence is an established risk factor for later cardiovascular disease and dysregulated reactivity to acute social stress. This study examined whether ELS associations with baseline cardiovascular functioning, cardiovascular stress reactivity and recovery, and emotional stress reactivity vary across levels of emotion-oriented, task-oriented, and avoidant coping styles., Methods: The sample included 1027 adolescents and young adults (mean age = 19.29 years; 50% female; 64% Black, 34% non-Hispanic White) who reported on their ELS exposure and coping styles. Participants completed a standardized acute social stress test (the Trier Social Stress Test [TSST]), with heart rate (HR) and blood pressure (BP) measured before, during, and after the TSST. Self-reports of negative emotions during the TSST indexed emotional stress reactivity., Results: Multiple regression models adjusting for demographic factors and body mass index showed that ELS was associated with lower HR stress reactivity, avoidant coping was related to lower systolic BP and diastolic BP during stress and lower systolic BP during recovery, and higher emotion-oriented coping and lower task-oriented coping predicted greater emotional stress reactivity. A consistent pattern emerged where emotion-oriented coping amplified the associations between ELS and maladaptive stress responses (blunted cardiovascular stress reactivity and recovery; enhanced emotional stress reactivity), whereas lower levels of emotion-oriented coping were associated with resilient profiles among those who experienced ELS (lower resting HR, lower emotional stress reactivity, average HR and BP stress reactivity and recovery). However, low levels of emotion-oriented coping also conferred a risk of higher BP during recovery for those with high levels of ELS., Conclusions: These results suggest that low to moderate levels of emotion-oriented coping promote optimal cardiovascular and emotional reactivity to acute stress among individuals exposed to ELS., (Copyright © 2022 by the American Psychosomatic Society.)

Demographic studies reveal lower prevalence of hypertension among premenopausal females compared to age-matched males. The kidney plays a central role in the maintenance of sodium (Na + ) homeostasis and consequently blood pressure. Renal endothelin-1 (ET-1) is a pro-natriuretic peptide that contributes to sex differences in blood pressure regulation and Na + homeostasis. We recently showed that activation of renal medullary G protein-coupled estrogen receptor 1 (GPER1) promotes ET-1-dependent natriuresis in female, but not male, rats. We hypothesized that GPER1 upregulates the renal ET-1 signaling system in females, but not males. To test our hypothesis, we determined the effect of GPER1 deletion on ET-1 and its downstream effectors in the renal cortex, outer and inner medulla obtained from 12-16-week-old female and male mice. GPER1 knockout (KO) mice and wildtype (WT) littermates were implanted with telemetry transmitters for blood pressure assessment, and we used metabolic cages to determine urinary Na + excretion. GPER1 deletion did not significantly affect 24-h mean arterial pressure (MAP) nor urinary Na + excretion. However, GPER1 deletion decreased urinary ET-1 excretion in females but not males. Of note, female WT mice had greater urinary ET-1 excretion than male WT littermates, whereas no sex differences were observed in GPER1 KO mice. GPER1 deletion increased inner medullary ET-1 peptide content in both sexes but increased outer medullary ET-1 content in females only. Cortical ET-1 content increased in response to GPER1 deletion in both sexes. Furthermore, GPER1 deletion notably increased inner medullary ET receptor A (ET A ) and decreased outer medullary ET receptor B (ET B ) mRNA expression in male, but not female, mice. We conclude that GPER1 is required for greater ET-1 excretion in females. Our data suggest that GPER1 is an upstream regulator of renal medullary ET-1 production and ET receptor expression in a sex-specific manner. Overall, our study identifies the role of GPER1 as a sex-specific upstream regulator of the renal ET-1 system., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Guthrie, Almutlaq, Sugahara, Butt, Brooks, Pollock and Gohar.)

Podocyte foot processes are an important cellular layer of the glomerular barrier that regulates glomerular permeability. Insulin via the protein kinase G type Iα (PKGIα) signaling pathway regulates the balance between contractility and relaxation (permeability) of the podocyte barrier by regulation of the actin cytoskeleton. This mechanism was shown to be disrupted in diabetes. Rho family guanosine-5'-triphosphates (GTPases) are dynamic modulators of the actin cytoskeleton and expressed in cells that form the glomerular filtration barrier. Thus, changes in Rho GTPase activity may affect glomerular permeability to albumin. The present study showed that Rho family GTPases control podocyte migration and permeability. Moreover these processes are regulated by insulin in PKGIα-dependent manner. Modulation of the PKGI-dependent activity of Rac1 and RhoA GTPases with inhibitors or small-interfering RNA impair glomerular permeability to albumin. We also demonstrated this mechanism in obese, insulin-resistant Zucker rats. We propose that PKGIα-Rac1-RhoA crosstalk is necessary in proper organization of the podocyte cytoskeleton and consequently the stabilization of glomerular architecture and regulation of filtration barrier permeability., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Emerging evidence over the past several years suggests that diurnal control of sodium excretion is sex dependent and involves the renal endothelin system. Given recent awareness of disruptions of circadian function in obesity, we determined whether diet-induced obesity impairs renal handling of an acute salt load at different times of day and whether this varies by sex and is associated with renal endothelin dysfunction., Methods: Male and female Sprague-Dawley rats were placed on a high-fat diet for 8 weeks before assessing renal sodium handling and blood pressure., Results: Male, but not female, rats on high fat had a significantly reduced natriuretic response to acute NaCl injection at the beginning of their active period that was associated with lower endothelin 1 (ET-1) excretion, lower ET-1 mRNA expression in the cortex and outer medulla as well as lower ET B receptor expression in the outer medulla of the high-fat rats. Obese males also had significantly higher blood pressure (telemetry) that was exacerbated by adding high salt to the diet during the last 2 weeks. While female rats developed hypertension with a high-fat diet, they were not salt sensitive and ET-1 excretion was unchanged., Conclusions: These data identify diet-induced obesity as a sex-specific disruptive factor for maintaining proper sodium handling. Although high-fat diets induce hypertension in both sexes, these data reveal that males are at greater risk of salt-dependent hypertension and further suggest that females have more redundant systems that can be productive against salt-sensitive hypertension in at least some circumstances.

Progressive iron accumulation and renal impairment are prominent in both patients and mouse models of sickle cell disease (SCD). Endothelin A receptor (ET A ) antagonism prevents this iron accumulation phenotype and reduces renal iron deposition in the proximal tubules of SCD mice. To better understand the mechanisms of iron metabolism in the kidney and the role of the ET A receptor in iron chelation and transport, we studied renal iron handling in a nonsickle cell iron overload model, heme oxygenase-1 ( Hmox-1 -/- ) knockout mice. We found that Hmox-1 -/- mice had elevated plasma endothelin-1 (ET-1), cortical ET-1 mRNA expression, and renal iron content compared with Hmox-1 +/+ controls. The ET A receptor antagonist, ambrisentan, attenuated renal iron deposition, without any changes to anemia status in Hmox-1 -/- mice. This was accompanied by reduced urinary iron excretion. Finally, ambrisentan had an important iron recycling effect by increasing the expression of the cellular iron exporter, ferroportin-1 (FPN-1), and circulating total iron levels in Hmox-1 -/- mice. These findings suggest that the ET-1/ET A signaling pathway contributes to renal iron trafficking in a murine model of iron overload.

Increased adiposity is associated with dysregulation of the endothelin system, both of which increase the risk of cardiovascular disease (CVD). Preclinical data indicate that endothelin dysregulation also reduces resting energy expenditure (REE). The objective was to test the hypothesis that endothelin receptor antagonism will increase REE in people with obesity compared with healthy weight individuals. Using a double blind, placebo-controlled, crossover design, 32 participants [healthy weight (HW): n = 16, BMI: 21.3 ± 2.8 kg/m 2 , age: 26 ± 7 yr and overweight/obese (OB): n = 16, BMI: 33.5 ± 9.5 kg/m 2 , age: 31 ± 6 yr] were randomized to receive either 125 mg of bosentan (ET A/B antagonism) or placebo twice per day for 3 days. Breath-by-breath gas exchange data were collected and REE was assessed by indirect calorimetry. Venous blood samples were analyzed for concentrations of endothelin-1 (ET-1). Treatment with bosentan increased plasma ET-1 in both OB and HW groups. Within the OB group, the changes in absolute REE (PLA: -77.6 ± 127.6 vs. BOS: 72.2 ± 146.6 kcal/day; P = 0.046). The change in REE was not different following either treatment in the HW group. Overall, absolute plasma concentrations of ET-1 following treatment with bosentan were significantly associated with kcal/day of fat ( r = 0.488, P = 0.005), percentage of fat utilization ( r = 0.415, P = 0.020), and inversely associated with the percentage of carbohydrates ( r = -0.419, P = 0.019), and respiratory exchange ratio ( r = -0.407, P = 0.023). Taken together, these results suggest that modulation of the endothelin system may represent a novel therapeutic approach to increase both resting metabolism and caloric expenditure, and reduce CVD risk in people with increased adiposity. NEW & NOTEWORTHY Findings from our current translational investigation demonstrate that dual endothelin A/B receptor antagonism increases total REE in overweight/obese individuals. These results suggest that modulation of the endothelin system may represent a novel therapeutic target to increase both resting metabolism and caloric expenditure, enhance weight loss, and reduce CVD risk in seemingly healthy individuals with elevated adiposity.

Peroxiredoxin-2 (Prx-2) is a critical antioxidant protein in red blood cells (RBC). Prx-2 is oxidized to a disulfide covalently-bound dimer by H 2 O 2 , and then reduced back by the NADPH-dependent thioredoxin-thioredoxin reductase system. The reduction of oxidized Prx-2 is relatively slow in RBCs. Since Prx-2 is highly abundant, Prx-2s' peroxidase catalytic cycle is not considered to be limiting under normal conditions. However, whether Prx-2 recycling becomes limiting when RBCs are exposed to stress is not known. Using three different model systems characterized by increased oxidative damage to RBCs spanning the physiologic (endogenous RBCs of different ages), therapeutic (cold-stored RBCs in blood banks) and pathologic (RBCs from sickle cell disease (SCD) patients and humanized SCD mice) spectrum, basal levels of Prx-2 oxidation and Prx-2 recycling kinetics after addition of H 2 O 2 were determined. The reduction of oxidized Prx-2 was significantly slower in older versuin older versus younger RBCs, in RBCs stored for 4-5 weeks compared to 1 week, and in RBC from pediatric SCD patients compared to RBCs from control non-SCD patients. Similarly, the rate of Prx-2 recycling was slower in humanized SCD mice compared to WT mice. Treatment of RBC with carbon monoxide (CO) to limit heme-peroxidase activity had no effect on Prx-2 recycling kinetics. Treatment with glucose attenuated slowed Prx-2 recycling in older RBCs and SCD RBCs, but not stored RBCs. In conclusion, the reduction of oxidized Prx-2 can be further slowed in RBCs, which may limit the protection afforded by this antioxidant protein in settings associated with erythrocyte stress., (© 2022 Federation of American Societies for Experimental Biology.)

Night shift work increases risk of cardiovascular disease associated with an irregular eating schedule. Elevating this risk is the high level of salt intake observed in the typical Western diet. Renal Na + excretion has a distinct diurnal pattern, independent of time of intake, yet the interactions between the time of intake and the amount of salt ingested are not clear. The hypothesis of the present study was that limiting food intake to the typically inactive period in addition to high-salt (HS) feeding will disrupt the diurnal rhythm of renal Na + excretion. Male Sprague-Dawley rats were placed on either normal-salt (NS; 0.49% NaCl) or HS (4% NaCl) diets. Rats were housed in metabolic cages and allowed food ad libitum and then subjected to inactive period time-restricted feeding (iTRF) for 5 days. As expected, rats fed NS and allowed food ad libitum had a diurnal pattern of Na + excretion. The diurnal pattern of Na + excretion was not significantly different after 5 days of iTRF compared with ad libitum rats. In response to HS, the diurnal pattern of Na + excretion was similar to NS-fed rats. However, this pattern was attenuated after 5 days of HS iTRF. The diurnal excretion pattern of urinary aldosterone was abolished in both NS iTRF and HS iTRF rats. These data support the hypothesis that HS intake combined with iTRF impairs circadian mechanisms associated with renal Na + excretion. NEW & NOTEWORTHY Timing of food intake normally has little effect on the diurnal pattern of Na + and water excretion. However, rats on a high-salt diet were unable to maintain this pattern, yet K + excretion was more readily adjusted to match timing of intake. These data support the hypothesis that Na + and water homeostasis are impacted by timing of high-salt diets.

Background Premenopausal women are less likely to develop hypertension and salt-related complications than are men, yet the impact of sex on mechanisms regulating Na + homeostasis during dietary salt challenges is poorly defined. Here, we determined whether female rats have a more efficient capacity to acclimate to increased dietary salt intake challenge. Methods and Results Age-matched male and female Sprague Dawley rats maintained on a normal-salt (NS) diet (0.49% NaCl) were challenged with a 5-day high-salt diet (4.0% NaCl). We assessed serum, urinary, skin, and muscle electrolytes; total body water; and kidney Na + transporters during the NS and high-salt diet phases. During the 5-day high-salt challenge, natriuresis increased more rapidly in females, whereas serum Na + and body water concentration increased only in males. To determine if females are primed to handle changes in dietary salt, we asked the question whether the renal endothelin-1 natriuretic system is more active in female rats, compared with males. During the NS diet, female rats had a higher urinary endothelin-1 excretion rate than males. Moreover, Ingenuity Pathway Analysis of RNA sequencing data identified the enrichment of endothelin signaling pathway transcripts in the inner medulla of kidneys from NS-fed female rats compared with male counterparts. Notably, in human subjects who consumed an Na + -controlled diet (3314-3668 mg/day) for 3 days, women had a higher urinary endothelin-1 excretion rate than men, consistent with our findings in NS-fed rats. Conclusions These results suggest that female sex confers a greater ability to maintain Na + homeostasis during acclimation to dietary Na + challenges and indicate that the intrarenal endothelin-1 natriuretic pathway is enhanced in women.

Obesity is associated with dysregulation of the endothelin system. In individuals with obesity, an exaggerated pressor response to acute stress is accompanied by increased circulating endothelin-1 (ET-1). The impact of combined endothelin A/B receptor (ET A/B ) antagonism on the stress-induced pressor response in overweight/obese (OB) individuals is unknown. The objective of this study is to test the hypothesis that treatment with an ET A/B antagonist (bosentan) would reduce the stress-induced pressor response and arterial stiffness in overweight/obese compared with normal weight (NW) individuals. Forty participants [normal weight (NW): n = 20, body mass index (BMI): 21.7 ± 2.4 kg/m 2 and overweight/obese (OB): n = 20, BMI: 33.8 ± 8.2 kg/m 2 ] were randomized to placebo or 125 mg of bosentan twice a day (250 mg total) for 3 days. Hemodynamics were assessed before, during, and after a cold pressor test (CPT). Endothelin-1 was assessed at baseline and immediately after CPT. Following a washout period, the same protocol was repeated with the opposite treatment. The change from baseline in mean arterial pressure (MAP) during CPT following bosentan was significantly lower ( P = 0.039) in the OB group than in the NW group (OB: 28 ± 12 vs. NW: 34 ± 15 mmHg). These results suggest that ET A/B antagonism favorably blunts the pressor response to acute stress in overweight/obese individuals. NEW & NOTEWORTHY Findings from our current translational investigation demonstrate that dual endothelin A/B receptor antagonism blunts the pressor response to acute stress in overweight/obese individuals. These results suggest that modulation of the endothelin system may represent a novel therapeutic target to reduce cardiovascular disease (CVD) risk by blunting the stress response in overweight/obese individuals.

The attention for the control of dietary risk factors involved in the development of hypertension, includes a large effort on dietary salt restrictions. Ample studies show the beneficial role of limiting dietary sodium as a lifestyle modification in the prevention and management of essential hypertension. Not until the past decade or so have studies more specifically investigated diurnal variations in renal electrolyte excretion, which led us to the hypothesis that timing of salt intake may impact cardiovascular health and blood pressure regulation. Cell autonomous molecular clocks as the name implies, function independently to maintain optimum functional rhythmicity in the face of environmental stressors such that cellular homeostasis is maintained at all times. Our understanding of mechanisms influencing diurnal patterns of sodium excretion and blood pressure has expanded with the discovery of the circadian clock genes. In this review, we discuss what is known about circadian regulation of renal sodium handling machinery and its influence on blood pressure regulation, with timing of sodium intake as a potential modulator of the kidney clock., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Journal of Hypertension, Ltd. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The liver plays a central role that influences cardiovascular disease outcomes through regulation of glucose and lipid metabolism. It is recognized that the local liver molecular clock regulates some liver-derived metabolites. However, it is unknown whether the liver clock may impact cardiovascular function. Perivascular adipose tissue (PVAT) is a specialized type of adipose tissue surrounding blood vessels. Importantly, cross talk between the endothelium and PVAT via vasoactive factors is critical for vascular function. Therefore, we designed studies to test the hypothesis that cardiovascular function, including PVAT function, is impaired in mice with liver-specific circadian clock disruption. Bmal1 is a core circadian clock gene, thus studies were undertaken in male hepatocyte-specific Bmal1 knockout (HBK) mice and littermate controls (i.e., flox mice). HBK mice showed significantly elevated plasma levels of β-hydroxybutyrate, nonesterified fatty acids/free fatty acids, triglycerides, and insulin-like growth factor 1 compared with flox mice. Thoracic aorta PVAT in HBK mice had increased mRNA expression of several key regulatory and metabolic genes, Ppargc1a , Pparg , Adipoq , Lpl , and Ucp1 , suggesting altered PVAT energy metabolism and thermogenesis. Sensitivity to acetylcholine-induced vasorelaxation was significantly decreased in the aortae of HBK mice with PVAT attached compared with aortae of HBK mice with PVAT removed, however, aortic vasorelaxation in flox mice showed no differences with or without attached PVAT. HBK mice had a significantly lower systolic blood pressure during the inactive period of the day. These new findings establish a novel role of the liver circadian clock in regulating PVAT metabolic gene expression and PVAT-mediated aortic vascular function.

Feeding rodents a high-fat diet (HFD) disrupts normal behavioral rhythms, particularly meal timing. Within the brain, mistimed feeding shifts molecular rhythms in the hippocampus and impairs memory. We hypothesize that altered meal timing induced by an HFD leads to cognitive impairment and that restricting HFD access to the "active period" (i.e., night) rescues the normal hippocampal function. In male mice, ad-lib access to an HFD for 20 weeks increased body weight and fat mass, increased daytime meal consumption, reduced hippocampal long-term potentiation (LTP), and eliminated day/night differences in spatial working memory. Importantly, two weeks of time-restricted feeding (TRF) at the end of the chronic HFD protocol rescued spatial working memory and restored LTP magnitude, even though there was no change in body composition and total daily caloric intake. These findings suggest that short-term TRF is an effective mechanism for rescuing HFD-induced impaired cognition and hippocampal function., Competing Interests: No relevant disclosures., (© 2021 The Authors.)

Despite advancements in disease management, sickle cell nephropathy, a major contributor to mortality and morbidity in patients, has limited therapeutic options. Previous studies indicate hydroxyurea, a commonly prescribed therapy for sickle cell disease (SCD), can reduce renal injury in SCD but the mechanisms are uncertain. Because SCD is associated with reduced nitric oxide (NO) bioavailability, we hypothesized that hydroxyurea treatment would improve NO bioavailability in the humanized sickle cell mouse. Humanized male 12-wk-old sickle (HbSS) and genetic control (HbAA) mice were treated with hydroxyurea or regular tap water for 2 wk before renal and systemic NO bioavailability as well as renal injury were assessed. Untreated HbSS mice exhibited increased proteinuria, elevated plasma endothelin-1 (ET-1), and reduced urine concentrating ability compared with HbAA mice. Hydroxyurea reduced proteinuria and plasma ET-1 levels in HbSS mice. Untreated HbSS mice had reduced plasma nitrite and elevated plasma arginase concentrations compared with HbAA mice. Hydroxyurea treatment augmented plasma nitrite and attenuated plasma arginase in HbSS mice. Renal vessels isolated from HbSS mice also had elevated nitric oxide synthase 3 (NOS3) and arginase 2 expression compared with untreated HbAA mice. Hydroxyurea treatment did not alter renal vascular NOS3, however, renal vascular arginase 2 expression was significantly reduced. These data support the hypothesis that hydroxyurea treatment augments renal and systemic NO bioavailability by reducing arginase activity as a potential mechanism for the improvement on renal injury seen in SCD mice.

Baroreflex function is an integral component maintaining consistent blood pressure. Hypertension is often associated with baroreflex dysfunction, and environmental risk factors such as high salt diet exacerbate hypertension in subjects with baroreflex dysfunction. However, the interactions between high salt diet, baroreflex dysfunction, and hypertension are incompletely understood. The endothelin system is another potent mediator of blood pressure control especially in response to a high salt diet. We hypothesized that the endothelin B (ET B ) receptor activation on adrenergic nerves decreases baroreflex sensitivity. We utilized male ET B receptor deficient (ET B -def) rats that express functional ET B receptors only on adrenergic nerves and transgenic (TG) controls to evaluate baroreflex function during normal (0.49% NaCl) and high (4.0% NaCl) salt diets. In conscious rats equipped with telemetry, ET B -def rats had an increased lability of systolic blood pressure (SBP) compared to TG controls as indicated by higher standard deviation (SD) of SBP under both normal (10.2 ± 0.6 vs. 12.4 ± 0.9 mmHg, respectively, p = 0.0001) and high (11.7 ± 0.6 vs. 16.1 ± 1.0 mmHg, p = 0.0001) salt diets. In anesthetized preparations, ET B -def rats displayed reduced heart rate (p genotype = 0.0167) and renal sympathetic nerve (p genotype = 0.0022) baroreflex sensitivity. We then gave male Sprague-Dawley rats the selective ET B receptor antagonist, A-192621 (10 mg/kg/day), to block ET B receptors. Following ET B receptor antagonism, even though SBP increased (131 ± 7 before vs. 152 ± 8 mmHg after, p < 0.0001), the lability (standard deviation) of SBP decreased (9.3 ± 2.0 vs. 7.1 ± 1.1 mmHg, p = 0.0155). These data support our hypothesis that ET B receptors on adrenergic nerves contribute to baroreflex dysfunction., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Recent evidence indicates a crucial role for G protein-coupled estrogen receptor 1 (GPER1) in the maintenance of cardiovascular and kidney health in females. The current study tested whether GPER1 activation ameliorates hypertension and kidney damage in female Dahl salt-sensitive (SS) rats fed a high-salt (HS) diet. Adult female rats were implanted with telemetry transmitters for monitoring blood pressure and osmotic minipumps releasing G1 (selective GPER1 agonist, 400 μg/kg/day ip) or vehicle. Two weeks after pump implantation, rats were shifted from a normal-salt (NS) diet (0.4% NaCl) to a matched HS diet (4.0% NaCl) for 2 wk. Twenty-four hour urine samples were collected during both diet periods and urinary markers of kidney injury were assessed. Histological assessment of kidney injury was conducted after the 2-wk HS diet period. Compared with values during the NS diet, 24-h mean arterial pressure markedly increased in response to HS, reaching similar values in vehicle-treated and G1-treated rats. HS also significantly increased urinary excretion of protein, albumin, nephrin (podocyte damage marker), and KIM-1 (proximal tubule injury marker) in vehicle-treated rats. Importantly, G1 treatment prevented the HS-induced proteinuria, albuminuria, and increase in KIM-1 excretion but not nephrinuria. Histological analysis revealed that HS-induced glomerular damage did not differ between groups. However, G1 treatment preserved proximal tubule brush-border integrity in HS-fed rats. Collectively, our data suggest that GPER1 activation protects against HS-induced proteinuria and albuminuria in female Dahl SS rats by preserving proximal tubule brush-border integrity in a blood pressure-independent manner.

Timing of food intake has become a critical factor in determining overall cardiometabolic health. We hypothesized that timing of food intake entrains circadian rhythms of blood pressure (BP) and renal excretion in mice. Male C57BL/6J mice were fed ad libitum or reverse feeding (RF) where food was available at all times of day or only available during the 12-h lights-on period, respectively. Mice eating ad libitum had a significantly higher mean arterial pressure (MAP) during lights-off compared to lights-on (113 ± 2 mmHg vs 100 ± 2 mmHg, respectively; P  < 0.0001); however, RF for 6 days inverted the diurnal rhythm of MAP (99 ± 3 vs 110 ± 3 mmHg, respectively; P  < 0.0001). In contrast to MAP, diurnal rhythms of urine volume and sodium excretion remained intact after RF. Male Bmal1 knockout mice (Bmal1KO) underwent the same feeding protocol. As previously reported, Bmal1KO mice did not exhibit a diurnal MAP rhythm during ad libitum feeding (95 ± 1 mmHg vs 92 ± 3 mmHg, lights-off vs lights-on; P  > 0.05); however, RF induced a diurnal rhythm of MAP (79 ± 3 mmHg vs 95 ± 2 mmHg, lights-off vs lights-on phase; P  < 0.01). Transgenic PERIOD2::LUCIFERASE knock-in mice were used to assess the rhythm of the clock protein PERIOD2 in ex vivo tissue cultures. The timing of the PER2::LUC rhythm in the renal cortex and suprachiasmatic nucleus was not affected by RF; however, RF induced significant phase shifts in the liver, renal inner medulla, and adrenal gland. In conclusion, the timing of food intake controls BP rhythms in mice independent of Bmal1, urine volume, or sodium excretion., (© The Author(s) 2020. Published by Oxford University Press on behalf of American Physiological Society.)

The G protein-coupled estrogen receptor 1 (GPER1) mediates rapid estrogenic signaling. We recently reported that activation of GPER1 in the renal medulla evokes endothelin-1-dependent natriuresis in female, but not male, rats. However, the involvement of the ET receptors, ET A and ET B , underlying GPER1 natriuretic action remain unclear. In this study, we used genetic and pharmacologic methods to identify the contributions of ET A and ET B in mediating this female-specific natriuretic effect of renal medullary GPER1. Infusion of the GPER1-selective agonist G1 (5 pmol/kg per minute) into the renal medulla for 40 minutes increased Na + excretion and urine flow in anesthetized female ET B -deficient (ET B def) rats and littermate controls but did not affect blood pressure or urinary K + excretion in either group. Pretreatment with the selective ET A inhibitor ABT-627 (5 mg/kg, intravenous) abolished G1-induced natriuresis in ET B def rats. To further isolate the effects of inhibiting either receptor alone, we conducted the same experiments in anesthetized female Sprague-Dawley (SD) rats pretreated or not with ABT-627 and/or the selective ET B inhibitor A-192621 (10 mg/kg, intravenous). Neither antagonism of ET A nor antagonism of ET B receptor alone affected the G1-induced increase in Na + excretion and urine flow in SD rats. However, simultaneous antagonism of both receptors completely abolished these effects. These data suggest that ET A and ET B receptors can mediate the natriuretic and diuretic response to renal medullary GPER1 activation in female rats. SIGNIFICANCE STATEMENT: Activation of G protein-coupled estrogen receptor 1 (GPER1) in the renal medulla of female rats evokes natriuresis via endothelin receptors A and/or B, suggesting that GPER1 and endothelin signaling pathways help efficient sodium excretion in females. Thus, GPER1 activation could be potentially useful to mitigate salt sensitivity in females., (Copyright © 2020 by The Author(s).)

Background: Premenopausal women have a lower risk of hypertension compared to age-matched men and postmenopausal women. P2Y 2 and P2Y 4 purinoceptor can be considered potential contributors to hypertension due to their emerging roles in regulating renal tubular Na + transport. Activation of these receptors inhibits epithelial Na + channel activity (ENaC) via a phospholipase C (PLC)-dependent pathway resulting in natriuresis. We recently reported that activation of P2Y 2 and P2Y 4 receptors in the renal medulla by UTP promotes natriuresis in male and ovariectomized (OVX) rats, but not in ovary-intact females. This led us to hypothesize that ovary-intact females have greater basal renal medullary activity of P2 (P2Y 2 and P2Y 4 ) receptors regulating Na + excretion compared to male and OVX rats., Methods: To test our hypothesis, we determined (i) the effect of inhibiting medullary P2 receptors by suramin (750 μg/kg/min) on urinary Na + excretion in anesthetized male, ovary-intact female, and OVX Sprague Dawley rats, (ii) mRNA expression and protein abundance of P2Y 2 and P2Y 4 receptors, and (iii) mRNA expression of their downstream effectors (PLC-1δ and ENaCα) in renal inner medullary tissues obtained from these three groups. We also subjected cultured mouse inner medullary collecting duct cells (segment 3, mIMCD3) to different concentrations of 17ß-estradiol (E 2 , 0, 10, 100, and 1000 nM) to test whether E 2 increases mRNA expression of P2Y 2 and P2Y 4 receptors., Results: Acute P2 inhibition attenuated urinary Na + excretion in ovary-intact females, but not in male or OVX rats. We found that P2Y 2 and P2Y 4 mRNA expression was higher in the inner medulla from females compared to males or OVX. Inner medullary lysates showed that ovary-intact females have higher P2Y 2 receptor protein abundance, compared to males; however, OVX did not eliminate this sex difference. We also found that E 2 dose-dependently upregulated P2Y 2 and P2Y 4 mRNA expression in mIMCD3., Conclusion: These data suggest that ovary-intact females have enhanced P2Y 2 and P2Y 4 -dependent regulation of Na + handling in the renal medulla, compared to male and OVX rats. We speculate that the P2 pathway contributes to facilitated renal Na + handling in premenopausal females.

Background: Sickle cell disease (SCD) is characterized by chronic hemolytic anemia, vaso-occlusive crises, chronic inflammation, and activation of coagulation. The clinical complications such as painful crisis, stroke, pulmonary hypertension, nephropathy and venous thromboembolism lead to cumulative organ damage and premature death. High molecular weight kininogen (HK) is a central cofactor for the kallikrein-kinin and intrinsic coagulation pathways, which contributes to both coagulation and inflammation., Objective: We hypothesize that HK contributes to the hypercoagulable and pro-inflammatory state that causes end-organ damage and early mortality in sickle mice., Methods: We evaluated the role of HK in the Townes mouse model of SCD., Results/conclusions: We found elevated plasma levels of cleaved HK in sickle patients compared to healthy controls, suggesting ongoing HK activation in SCD. We used bone marrow transplantation to generate wild type and sickle cell mice on a HK-deficient background. We found that short-term HK deficiency attenuated thrombin generation and inflammation in sickle mice at steady state, which was independent of bradykinin signaling. Moreover, long-term HK deficiency attenuates kidney injury, reduces chronic inflammation, and ultimately improves survival of sickle mice., (© 2020 International Society on Thrombosis and Haemostasis.)

Histone deacetylase (HDAC) enzymes regulate transcription through epigenetic modification of chromatin structure, but their specific functions in the kidney remain elusive. We discovered that the human kidney expresses class I HDACs. Kidney medulla-specific inhibition of class I HDACs in the rat during high-salt feeding results in hypertension, polyuria, hypokalemia, and nitric oxide deficiency. Three new inducible murine models were used to determine that HDAC1 and HDAC2 in the kidney epithelium are necessary for maintaining epithelial integrity and maintaining fluid-electrolyte balance during increased dietary sodium intake. Moreover, single-nucleus RNA-sequencing determined that epithelial HDAC1 and HDAC2 are necessary for expression of many sodium or water transporters and channels. In performing a systematic review and meta-analysis of serious adverse events associated with clinical HDAC inhibitor use, we found that HDAC inhibitors increased the odds ratio of experiencing fluid-electrolyte disorders, such as hypokalemia. This study provides insight on the mechanisms of potential serious adverse events with HDAC inhibitors, which may be fatal to critically ill patients. In conclusion, kidney tubular HDACs provide a link between the environment, such as consumption of high-salt diets, and regulation of homeostatic mechanisms to remain in fluid-electrolyte balance.

The diurnal rhythms of sodium handling and blood pressure are thought to be regulated by clock genes, such as Bmal1. However, little is known about the regulation of these factors by Bmal1, especially in rats. Using a novel whole-body Bmal1 knockout rat model ( Bmal1 - /- ), we hypothesized that time of day regulation of sodium excretion is dependent on Bmal1. Using telemetry to continuously record mean arterial pressure, we observed that male and female Bmal1 -/- rats had significantly reduced mean arterial pressure over the course of 24 hours compared with littermate controls. The circadian mean arterial pressure pattern remained intact in both sexes of Bmal1 -/- rats, which is in contrast to the Bmal1 -/- mouse model. Male Bmal1 -/- rats had no significant difference in baseline sodium excretion between 12-hour active and inactive periods, indicating a lack of diurnal control independent of maintained mean arterial pressure rhythms. Female Bmal1 -/- rats, however, had significantly greater sodium excretion during the active versus inactive period similar to controls. Thus, we observed a clear dissociation between circadian blood pressure and control of sodium excretion that is sex dependent. These findings are consistent with a more robust ability of females to maintain control of sodium excretion, and furthermore, demonstrate a novel role for Bmal1 in control of diurnal blood pressure independent of sodium excretion.

Background The novel estrogen receptor, G-protein-coupled estrogen receptor (GPER), is responsible for rapid estrogen signaling. GPER activation elicits cardiovascular and nephroprotective effects against salt-induced complications, yet there is no direct evidence for GPER control of renal Na + handling. We hypothesized that GPER activation in the renal medulla facilitates Na + excretion. Methods and Results Herein, we show that infusion of the GPER agonist, G1, to the renal medulla increased Na + excretion in female Sprague Dawley rats, but not male rats. We found that GPER mRNA expression and protein abundance were markedly higher in outer medullary tissues from females relative to males. Blockade of GPER in the renal medulla attenuated Na + excretion in females. Given that medullary endothelin 1 is a well-established natriuretic factor that is regulated by sex and sex steroids, we hypothesized that GPER activation promotes natriuresis via an endothelin 1-dependent pathway. To test this mechanism, we determined the effect of medullary infusion of G1 after blockade of endothelin receptors. Dual endothelin receptor subtype A and endothelin receptor subtype B antagonism attenuated G1-induced natriuresis in females. Unlike males, female mice with genetic deletion of GPER had reduced endothelin 1, endothelin receptor subtype A, and endothelin receptor subtype B mRNA expression compared with wild-type controls. More important, we found that systemic GPER activation ameliorates the increase in mean arterial pressure induced by ovariectomy. Conclusions Our data uncover a novel role for renal medullary GPER in promoting Na + excretion via an endothelin 1-dependent pathway in female rats, but not in males. These results highlight GPER as a potential therapeutic target for salt-sensitive hypertension in postmenopausal women.

Kidney function follows a 24-h rhythm subject to regulation by circadian genes including the transcription factor Bmal1 . A high-salt diet induces a phase shift in Bmal1 expression in the renal inner medulla that is dependent on endothelin type B (ET B ) receptors. Furthermore, ET B receptor-mediated natriuresis is sex dependent. Therefore, experiments tested the hypothesis that collecting duct Bmal1 regulates blood pressure in a sex-dependent manner. We generated a mouse model that lacks Bmal1 expression in the collecting duct, where ET B receptor abundance is highest. Male, but not female, collecting duct Bmal1 knockout (CDBmal1KO) mice had significantly lower 24-h mean arterial pressure (MAP) than flox controls (105 ± 2 vs. 112 ± 3 mmHg for male mice and 106 ± 1 vs. 108 ± 1 mmHg for female mice, by telemetry). After 6 days on a high-salt (4% NaCl) diet, MAP remained significantly lower in male CDBmal1KO mice than in male flox control mice (107 ± 2 vs. 113 ± 1 mmHg), with no significant differences between genotypes in female mice (108 ± 2 vs. 109 ± 1 mmHg). ET B receptor blockade for another 6 days increased MAP similarly in both male and female CDBmal1KO and flox control mice. However, MAP remained lower in male CDBmal1KO mice than in male flox control mice (124 ± 2 vs. 130 ± 2 mmHg). No significant differences were observed between female CDBmal1KO and flox mice during ET B blockade (130 ± 2 vs. 127 ± 2 mmHg). There were no significant genotype differences in amplitude or phase of MAP in either sex. These data suggest that collecting duct Bmal1 has no role in circadian MAP but plays an important role in overall blood pressure in male, but not female, mice.

Many physiological processes, including most kidney-related functions, follow specific rhythms tied to a 24-h cycle. This is largely because circadian genes operate in virtually every cell type in the body. In addition, many noncanonical genes have intrinsic circadian rhythms, especially within the liver and kidney. This new level of complexity applies to the control of renal electrolyte excretion. Furthermore, there is growing evidence that paracrine and autocrine factors, especially the endothelin system, are regulated by clock genes. We have known for decades that excretion of electrolytes is dependent on time of day, which could play an important role in fluid volume balance and blood pressure control. Here, we review what is known about the interplay between paracrine and circadian control of electrolyte excretion. The hope is that recognition of paracrine and circadian factors can be considered more deeply in the future when integrating with well-established neuroendocrine control of excretion.

Genes for the epithelial sodium channel (ENaC) subunits are expressed in a circadian manner, but whether this results in time-of-day differences in activity is not known. Recent data show that protein expression of ENaC subunits is higher in kidneys from female rats, yet females are more efficient in excreting an acute salt load. Thus, our in vivo study determined whether there is a time-of-day difference as well as a sex difference in the response to ENaC inhibition by benzamil. Our results showed that the natriuretic and diuretic responses to a single dose of benzamil were significantly greater in male compared with female rats whether given at the beginning of the inactive period [Zeitgeber time 0 (ZT0), 7 AM] or active period (ZT12, 7 PM). However, the response to benzamil was not significantly different between ZT0 and ZT12 dosing in either male or female rats. There was no difference in renal cortical α-ENaC protein abundance between ZT0 and ZT12 or males and females. Given previous reports of flow-induced stimulation of endothelin-1 (ET-1) production and sex differences in the renal endothelin system, we measured urinary ET-1 excretion to assess the effects of increased urine flow on intrarenal ET-1. ET-1 excretion was significantly increased following benzamil administration in both sexes, but this increase was significantly greater in females. These results support the hypothesis that ENaC activity is less prominent in maintaining Na + balance in females independent of renal ET-1. Because ENaC subunit genes and protein expression vary by time of day and are greater in female rat kidneys, this suggests a clear disconnect between ENaC expression and channel activity.

Background: Ethnic differences in nighttime blood pressure (BP) have long been documented with African Americans (AAs) having higher BP than European Americans (EAs). At present, lower nighttime melatonin, a key regulator of circadian rhythms, has been associated with higher nighttime BP levels in EAs. This study sought to test the hypothesis that AAs have lower nighttime melatonin secretion compared with EAs. We also determined if this ethnic difference in melatonin could partially explain the ethnic difference in nighttime BP., Methods: A total of 150 young adults (71 AA; 46% females; mean age: 27.7 years) enrolled in the Georgia Stress and Heart study provided an overnight urine sample for the measurement of 6-sulfatoxymelatonin, a major metabolite of melatonin. Urine melatonin excretion (UME) was calculated as the ratio between 6-sulfatoxymelatonin concentration and creatinine concentration. Twenty-four-hour ambulatory BP was assessed and nighttime systolic BP (SBP) was used as a major index of BP regulation., Results: After adjustment of age, sex, body mass index, and smoking, AAs had significantly lower UME (P = 0.002) and higher nighttime SBP than EAs (P = 0.036). Lower UME was significantly associated with higher nighttime SBP and this relationship did not depend on ethnicity. The ethnicity difference in nighttime SBP was significantly attenuated after adding UME into the model (P = 0.163)., Conclusion: This study is the first to document the ethnic difference in nighttime melatonin excretion, demonstrating that AAs have lower melatonin secretion compared with EAs. Furthermore, the ethnic difference in nighttime melatonin can partially account for the established ethnic difference in nighttime SBP., (© American Journal of Hypertension, Ltd 2019. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Hyperfiltration, highly prevalent early in sickle cell disease (SCD), is in part driven by an increase in ultrafiltration coefficient (K f ). The increase in K f may be due to enlarged filtration surface area and/or increased glomerular permeability (P alb ). Previous studies have demonstrated that endothelin-1 (ET-1) contributes to P alb changes in models of diabetes and SCD. Thus, we performed longitudinal studies of renal function to determine the relationship between ET-1 and glomerular size and P alb that may contribute to hyperfiltration in humanized sickle cell (HbSS) and control (HbAA) mice at 8-32 weeks of age. HbSS mice were characterized by significant increases in plasma and glomerular ET-1 expression in both sexes although this increase was significantly greater in males. HbSS glomeruli of both males and females presented with a progressive and significant increase in glomerular size, volume, and K f During the onset of hyperfiltration, plasma and glomerular ET-1 expression were associated with a greater increase in glomerular size and K f in HbSS mice, regardless of sex. The pattern of P alb augmentation during the hyperfiltration was also associated with an increase in glomerular ET-1 expression, in both male and female HbSS mice. However, the increase in P alb was significantly greater in males and delayed in time in females. Additionally, selective endothelin A receptor (ET A ) antagonist prevented hyperfiltration in HbSS, regardless of sex. These results suggest that marked sex disparity in glomerular hyperfiltration may be driven, in part, by ET-1-dependent ultra-structural changes in filtration barrier components contributing to glomerular hyperfiltration in HbSS mice., (© 2019 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

We previously reported that humanized sickle cell (HbSS) mice develop spontaneous nephropathy, a major cause of morbidity and mortality in sickle cell disease (SCD). Because sex-dependent protective mechanisms in SCD have been reported, we examined the course of nephropathy in male and female HbSS mice to determine contributors and/or predictors of disease severity. In male HbSS mice, glomerular filtration rate was characterized by a rapid onset of hyperfiltration and subsequent progressive decline of renal function over 20 weeks. Early tubular injury presented with increased excretion of kidney injury marker 1 (KIM-1), progressive loss of tubular brush border, and interstitial fibrosis that preceded the onset of glomerular damage, suggesting a tubuloglomerular mechanism of kidney injury in these mice. Additionally, we observed a strong association between the magnitude of hyperfiltration and the degree of long-term kidney injury in male HbSS mice. Unlike males, female HbSS mice did not demonstrate a significant loss of renal function or severe kidney damage during the time course of the study. These results suggest that magnitude of hyperfiltration predicts the onset of chronic kidney damage in male HbSS mice, whereas protective mechanisms in female HbSS mice delay the onset of SCD nephropathy., (© 2019 by The American Society of Hematology.)

Aim: Chronic high salt intake exaggerates renal injury and inflammation, especially with the loss of functional ET B receptors. Tauroursodeoxycholic acid (TUDCA) is a chemical chaperone and bile salt that is approved for the treatment of hepatic diseases. Our aim was to determine whether TUDCA is reno-protective in a model of ET B receptor deficiency with chronic high salt-induced renal injury and inflammation., Methods: ET B -deficient and transgenic control rats were placed on normal (0.8% NaCl) or high salt (8% NaCl) diet for 3 weeks, receiving TUDCA (400 mg/kg/d; ip) or vehicle. Histological and biochemical markers of kidney injury, renal cell death and renal inflammation were assessed., Results: In ET B -deficient rats, high salt diet significantly increased glomerular and proximal tubular histological injury, proteinuria, albuminuria, excretion of tubular injury markers KIM-1 and NGAL, renal cortical cell death and renal CD4 + T cell numbers. TUDCA treatment increased proximal tubule megalin expression as well as prevented high salt diet-induced glomerular and tubular damage in ET B -deficient rats, as indicated by reduced kidney injury markers, decreased glomerular permeability and proximal tubule brush border restoration, as well as reduced renal inflammation. However, TUDCA had no significant effect on blood pressure., Conclusions: TUDCA protects against the development of glomerular and proximal tubular damage, decreases renal cell death and inflammation in the renal cortex in rats with ET B receptor dysfunction on a chronic high salt diet. These results highlight the potential use of TUDCA as a preventive tool against chronic high salt induced renal damage., (© 2018 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.)

Cardiovascular and renal physiology follow strong circadian rhythms. For instance, renal excretion of solutes and water is higher during the active period compared to the inactive period, and blood pressure peaks early in the beginning of the active period of both diurnal and nocturnal animals. The control of these rhythms is largely dependent on the expression of clock genes both in the central nervous system and within peripheral organs themselves. Although it is understood that the central and peripheral clocks interact and communicate, few studies have explored the specific mechanism by which various organ systems within the body are coordinated to control physiological processes. The renal sympathetic nervous innervation has long been known to have profound effects on renal function, and because the sympathetic nervous system follows strong circadian rhythms, it is likely that autonomic control of the kidney plays an integral role in modulating renal circadian function. This review highlights studies that provide insight into this interaction, discusses areas lacking clarity, and suggests the potential for future work to explore the role of renal autonomics in areas such as blood pressure control and chronic kidney disease., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Aim: The objective of this study is to determine if ambrisentan (ET A selective antagonist) and hydroxyurea (HU) treatment has a synergistic effect on renal injury in sickle cell nephropathy when compared to HU treatment alone. The premise of the study is based on recent studies showing that endothelin-1 (ET-1) contributes to the pathophysiology of nephropathy in sickle cell disease (SCD) and that ET A receptor blockade improves renal function and protects against renal injury. Hydroxyurea (HU) is commonly prescribed for the treatment of SCD and has been shown to reduce renal injury in patients with SCD., Methods: Male 12-week-old humanized sickle mice (HbSS) and their genetic controls (HbAA) were treated with vehicle, HU, ambrisentan, or HU with ambrisentan for 2 weeks and renal structure and function were assessed., Results: Vehicle treated HbSS mice exhibited significant proteinuria compared to vehicle treated HbAA mice. HbSS mice also displayed significantly elevated plasma ET-1 concentrations and decreased urine osmolality compared to HbAA controls. Proteinuria was significantly lower in both HU and ambrisentan treated animals compared to vehicle treated HbSS mice; however, there was no additional improvement in HbSS mice treated with combined ambrisentan and HU. The combination of HU and ambrisentan resulted in significantly lower KIM-1 excretion, glomerular injury, and interstitial inflammation than HU alone., Conclusion: These findings indicate that HU and ET A receptor blockade produce similar reductions in renal injury in the humanized sickle mouse suggesting that both treatments may converge on the same mechanistic pathway., (© 2018 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.)