MYELOID-SPECIFIC FERRITIN LIGHT CHAIN DELETION DOES NOT EXACERBATE SEPSIS-ASSOCIATED AKI.

Sepsis-associated acute kidney injury (SA-AKI) is a key contributor to the life threatening sequalae of sepsis. Mechanistically, SA-AKI is a consequence of unabated myeloid cell activation and oxidative stress that induces tubular injury. Iron mediates inflammatory pathways directly and through regulating the expression of ferritin, an iron storage protein comprised of ferritin light (FtL) and heavy chain (FtH). Previous work revealed myeloid FtH deletion leads to a compensatory increase in intracellular and circulating FtL and is associated with amelioration of SA-AKI. We designed this study to test the hypothesis that loss of myeloid FtL will exacerbate the sepsis-induced inflammatory response and worsen SA-AKI. We generated a novel myeloid-specific FtL knockout mouse and induced sepsis via cecal ligation and puncture or lipopolysaccharide endotoxemia. As expected, myeloid FtL and serum ferritin levels were significantly lower in the knockout mice. Interestingly, while sepsis led to production of pro- and anti-inflammatory cytokines, there was no statistical difference between the genotypes. There was a similar loss of kidney function and injury, identified by expression of kidney injury molecule-1 and neutrophil gelatinase associated lipocalin. RNA sequencing revealed upregulation of pathways for cell cycle arrest and autophagy post-sepsis, but no significant differences were observed between genotypes, including in key genes associated with ferroptosis, an iron-mediated form of cell death. FtL deletion did not impact activation of NFkB or HIF-1a signaling, key inflammatory pathways associated with dysregulated host response. Taken together, while FtL overexpression was shown to be protective, loss of FtL did not influence sepsis pathogenesis.

Physiologic homeostasis after pig-to-human kidney xenotransplantation.

Demand for kidney grafts outpaces supply, limiting kidney transplantation as a treatment for kidney failure. Xenotransplantation has the potential to make kidney transplantation available to many more patients with kidney failure, but the ability of xenografts to support human physiologic homeostasis has not been established. A brain-dead adult decedent underwent bilateral native nephrectomies followed by 10 gene-edited (four gene knockouts, six human transgenes) pig-to-human xenotransplantation. Physiologic parameters and laboratory values were measured for seven days in a critical care setting. Data collection aimed to assess homeostasis by measuring components of the renin-angiotensin-aldosterone system, parathyroid hormone signaling, glomerular filtration rate, and markers of salt and water balance. Mean arterial blood pressure was maintained above 60 mmHg throughout. Pig kidneys secreted renin (post-operative day three to seven mean and standard deviation: 47.3 ± 9 pg/mL). Aldosterone and angiotensin II levels were present (post-operative day three to seven, 57.0 ± 8 pg/mL and 5.4 ± 4.3 pg/mL, respectively) despite plasma renin activity under 0.6 ng/mL/hr. Parathyroid hormone levels followed ionized calcium. Urine output down trended from 37 L to 6 L per day with 4.5 L of electrolyte free water loss on post-operative day six. Aquaporin 2 channels were detected in the apical surface of principal cells, supporting pig kidney response to human vasopressin. Serum creatinine down trended to 0.9 mg/dL by day seven. Glomerular filtration rate ranged 90-240 mL/min by creatinine clearance and single-dose inulin clearance. Thus, in a human decedent model, xenotransplantation of 10 gene-edited pig kidneys provided physiologic balance for seven days. Hence, our in-human study paves the way for future clinical study of pig-to-human kidney xenotransplantation in living persons.

Mild dehydration effects on the murine kidney single-nucleus transcriptome and chromatin accessibility.

Daily, we may experience mild dehydration with a rise in plasma osmolality that triggers the release of vasopressin. Although the effect of dehydration is well characterized in collecting duct principal cells (CDPCs), we hypothesized that mild dehydration (<12 h) results in many kidney cell-specific changes in transcriptomes and chromatin accessibility. Single-nucleus (sn) multiome (RNA-assay for transposase-accessible chromatin) sequencing and bulk RNA sequencing of kidneys from male and female mice that were mildly water deprived or not were compared. Water-deprived mice had a significant increase in plasma osmolality. sn-multiome-seq resulted in 19,837 nuclei that were annotated into 33 clusters. In CDPCs, aquaporin 2 (Aqp2) and aquaporin 3 (Apq3) were greater in dehydrated mice, but there were novel genes like gremlin 2 (Grem2; a cytokine) that were increased compared with ad libitum mice. The transcription factor cAMP-responsive element modulator (Crem) was greater in CDPCs of dehydrated mice, and the Crem DNA motif was more accessible. There were hundreds of sex- and dehydration-specific differentially expressed genes (DEGs) throughout the kidney, especially in the proximal tubules and thin limbs. In male mice, DEGs were enriched in pathways related to lipid metabolism, whereas female DEGs were enriched in organic acid metabolism. Many highly expressed genes had a positive correlation with increased chromatin accessibility, and mild dehydration exerted many transcriptional changes that we detected at the chromatin level. Even with a rise in plasma osmolality, male and female kidneys have distinct transcriptomes suggesting that there may be diverse mechanisms used to remain in fluid balance.NEW & NOTEWORTHY The kidney consists of >30 cell types that work collectively to maintain fluid-electrolyte balance. Kidney single-nucleus transcriptomes and chromatin accessibility profiles from male and female control (ad libitum water and food) or mildly dehydrated mice (ad libitum food, water deprivation) were determined. Mild dehydration caused hundreds of cell- and sex-specific transcriptomic changes, even though the kidney function to conserve water was the same.

Lipopolysaccharide Pretreatment Prevents Medullary Vascular Congestion following Renal Ischemia by Limiting Early Reperfusion of the Medullary Circulation.

BACKGROUND: Vascular congestion of the renal medulla-trapped red blood cells in the medullary microvasculature-is a hallmark finding at autopsy in patients with ischemic acute tubular necrosis. Despite this, the pathogenesis of vascular congestion is not well defined. METHODS: In this study, to investigate the pathogenesis of vascular congestion and its role in promoting renal injury, we assessed renal vascular congestion and tubular injury after ischemia reperfusion in rats pretreated with low-dose LPS or saline (control). We used laser Doppler flowmetry to determine whether pretreatment with low-dose LPS prevented vascular congestion by altering renal hemodynamics during reperfusion. RESULTS: We found that vascular congestion originated during the ischemic period in the renal venous circulation. In control animals, the return of blood flow was followed by the development of congestion in the capillary plexus of the outer medulla and severe tubular injury early in reperfusion. Laser Doppler flowmetry indicated that blood flow returned rapidly to the medulla, several minutes before recovery of full cortical perfusion. In contrast, LPS pretreatment prevented both the formation of medullary congestion and its associated tubular injury. Laser Doppler flowmetry in LPS-pretreated rats suggested that limiting early reperfusion of the medulla facilitated this protective effect, because it allowed cortical perfusion to recover and clear congestion from the large cortical veins, which also drain the medulla. CONCLUSIONS: Blockage of the renal venous vessels and a mismatch in the timing of cortical and medullary reperfusion results in congestion of the outer medulla's capillary plexus and promotes early tubular injury after renal ischemia. These findings indicate that hemodynamics during reperfusion contribute to the renal medulla's susceptibility to ischemic injury.

Kidney cell type-specific changes in the chromatin and transcriptome landscapes following epithelial Hdac1 and Hdac2 knockdown.

Recent studies have identified at least 20 different kidney cell types based upon chromatin structure and gene expression. Histone deacetylases (HDACs) are epigenetic transcriptional repressors via deacetylation of histone lysines resulting in inaccessible chromatin. We reported that kidney epithelial HDAC1 and HDAC2 activity is critical for maintaining a healthy kidney and preventing fluid-electrolyte abnormalities. However, to what extent does Hdac1/Hdac2 knockdown affect chromatin structure and subsequent transcript expression in the kidney? To answer this question, we used single nucleus assay for transposase-accessible chromatin-sequencing (snATAC-seq) and snRNA-seq to profile kidney nuclei from male and female, control, and littermate kidney epithelial Hdac1/Hdac2 knockdown mice. Hdac1/Hdac2 knockdown resulted in significant changes in the chromatin structure predominantly within the promoter region of gene loci involved in fluid-electrolyte balance such as the aquaporins, with both increased and decreased accessibility captured. Moreover, Hdac1/Hdac2 knockdown resulted different gene loci being accessible with a corresponding increased transcript number in the kidney, but among all mice only 24%-30% of chromatin accessibility agreed with transcript expression (e.g., open chromatin and increased transcript). To conclude, although chromatin structure does affect transcription, ∼70% of the differentially expressed genes cannot be explained by changes in chromatin accessibility and HDAC1/HDAC2 had a minimal effect on these global patterns. Yet, the genes that are targets of HDAC1 and HDAC2 are critically important for maintaining kidney function.

Endothelin system expression in the kidney following cisplatin-induced acute kidney injury in male and female mice.

The chemotherapeutic agent cisplatin accumulates in the kidney and induces acute kidney injury (AKI). Preclinical and clinical studies suggest that young female mice and women show greater recovery from cisplatin-AKI compared to young male mice and men. The endothelin (ET) and ET receptors are enriched in the kidney and may be dysfunctional in cisplatin-AKI; however, there is a gap in our knowledge about the putative effects of sex and cisplatin on the renal ET system. We hypothesized that cisplatin-AKI male and female mice will have increased expression of the renal ET system. As expected, all cisplatin-AKI mice had kidney damage and body weight loss greater than control mice. Cisplatin-AKI mice had greater cortical Edn1, Edn3, Ednra, and Ednrb, while outer medullary Ednra was significantly suppressed in both sexes. Of the ∼25 000 genes sequenced from the inner medulla, only 91 genes (comparing saline mice) and 134 genes (comparing cisplatin-AKI mice) were differentially expressed and they were unrelated to the ET system. However, Edn1 was significantly greater in the inner medulla of male and female cisplatin-AKI mice. Thus, RNA profiles of the ET system were significantly affected by cisplatin-AKI throughout the kidney regardless of sex and this may help determine the therapeutic potential of targeting the ET receptors in cisplatin-AKI.

The Seventeenth International Conference on Endothelin (ET-17).

The Seventeenth International Conference on Endothelin (ET-17) was held during 4-7 October 2021 and because of the SARS-CoV-2 pandemic it was held virtually. Sponsored by the American Physiological Society, ET-17 was held over 4 half-days, with exciting studies related to all organ systems presented. Since the Lancet article reporting the successful SONAR clinical trial with endothelin receptor A blockade in diabetic nephropathy, there has been renewed interest in the use of endothelin receptor antagonists in the treatment of a variety of diseases. From the rigorous preclinical studies to the latest clinical trials, ET-17 was full of exciting science, some of which is reported in this special issue. We welcomed new labs to the meeting and everyone left with the impression that ET-related research is a vibrant field with very significant discoveries being made.

Sex-Dependent Effects of Nephron Ift88 Disruption on BP, Renal Function, and Cystogenesis.

BACKGROUND: Primary cilia regulation of renal function and BP in health and disease is incompletely understood. This study investigated the effect of nephron ciliary loss on renal physiology, BP, and ensuing cystogenesis. METHODS: Mice underwent doxycycline (DOX)-inducible nephron-specific knockout (KO) of the Ift88 gene at 2 months of age using a Cre-LoxP strategy. BP, kidney function, and renal pathology were studied 2 and 9 months after DOX (Ift88 KO) or vehicle (control). RESULTS: At 2 months post-DOX, male, but not female, Ift88 KO, compared with sex-matched control, mice had reduced BP, enhanced salt-induced natriuresis, increased urinary nitrite and nitrate (NOx) excretion, and increased kidney NOS3 levels, which localized to the outer medulla; the reductions in BP in male mice were prevented by L-NAME. At 9 months post-DOX, male, but not female, Ift88 KO mice had polycystic kidneys, elevated BP, and reduced urinary NOx excretion. No differences were observed in plasma renin concentration, plasma aldosterone, urine vasopressin, or urine PGE2 between Ift88 KO and control mice at 2 or 9 months post-DOX. CONCLUSIONS: Nephron cilia disruption in male, but not female, mice (1) reduces BP prior to cyst formation, (2) increases NOx production that may account for the lower BP prior to cyst formation, and (3) induces polycystic kidneys that are associated with hypertension and reduced renal NO production.

High salt intake induces collecting duct HDAC1-dependent NO signaling.

We reported that high salt (HS) intake stimulates renal collecting duct (CD) endothelin (ET) type B receptor (ETBR)/nitric oxide (NO) synthase 1ß (NOS1ß)-dependent NO production inhibiting the epithelial sodium channel (ENaC) promoting natriuresis. However, the mechanism underlying the HS-induced increase of NO production is unclear. Histone deacetylase 1 (HDAC1) responds to increased fluid flow, as can occur in the CD during HS intake. The renal inner medulla (IM), in particular the IMCD, has the highest NOS1 activity within the kidney. Hence, we hypothesized that HS intake provokes HDAC1 activation of NO production in the IM. HS intake for 1 wk significantly increased HDAC1 abundance in the IM. Ex vivo treatment of dissociated IM from HS-fed mice with a selective HDAC1 inhibitor (MS-275) decreased NO production with no change in ET-1 peptide or mRNA levels. We further investigated the role of the ET-1/ETBR/NOS1ß signaling pathway with chronic ETBR blockade (A-192621). Although NO was decreased and ET-1 levels were elevated in the dissociated IM from HS-fed mice treated with A-192621, ex vivo MS-275 did not further change NO or ET-1 levels suggesting that HDAC1-mediated NO production is regulated at the level or downstream of ETBR activation. In split-open CDs from HS-fed mice, patch clamp analysis revealed significantly higher ENaC activity after MS-275 pretreatment, which was abrogated by an exogenous NO donor. Moreover, flow-induced increases in mIMCD-3 cell NO production were blunted by HDAC1 or calcium inhibition. Taken together, these findings indicate that HS intake induces HDAC1-dependent activation of the ETBR/NO pathway contributing to the natriuretic response.

Aquaporin-3 in the epidermis: more than skin deep.

The skin is essential for terrestrial life. It is responsible for regulating water permeability and functions as a mechanical barrier that protects against environmental insults such as microbial infection, ultraviolet light, injury, and heat and cold, which could damage the cells of the body and compromise survival of the organism. This barrier is provided by the outer layer, the epidermis, which is composed predominantly of keratinocytes; keratinocytes undergo a program of differentiation to form the stratum corneum comprising the cornified squame "bricks" and lipid "mortar." Dysregulation of this differentiation program can result in skin diseases, including psoriasis and nonmelanoma skin cancers, among others. Accumulating evidence in the literature indicates that the water-, glycerol-, and hydrogen peroxide-transporting channel aquaporin-3 (AQP3) plays a key role in various processes involved in keratinocyte function, and abnormalities in this channel have been observed in several human skin diseases. Here, we discuss the data linking AQP3 to keratinocyte proliferation, migration, differentiation, and survival as well as its role in skin properties and functions like hydration, water retention, wound healing, and barrier repair. We also discuss the mechanisms regulating AQP3 levels, localization, and function and the anomalies in AQP3 that are associated with various skin diseases.

Loss of endothelin type B receptor function improves insulin sensitivity in rats.

High salt intake (HS) is associated with obesity and insulin resistance. ET-1, a peptide released in response to HS, inhibits the actions of insulin on cultured adipocytes through ET-1 type B (ETB) receptors; however, the in vivo implications of ETB receptor activation on lipid metabolism and insulin resistance is unknown. We hypothesized that activation of ETB receptors in response to HS intake promotes dyslipidemia and insulin resistance. In normal salt (NS) fed rats, no significant difference in body mass or epididymal fat mass was observed between control and ETB deficient rats. After 2 weeks of HS, ETB-deficient rats had significantly lower body mass and epididymal fat mass compared to controls. Nonfasting plasma glucose was not different between genotypes; however, plasma insulin concentration was significantly lower in ETB-deficient rats compared to controls, suggesting improved insulin sensitivity. In addition, ETB-deficient rats had higher circulating free fatty acids in both NS and HS groups, with no difference in plasma triglycerides between genotypes. In a separate experiment, ETB-deficient rats had significantly lower fasting blood glucose and improved glucose and insulin tolerance compared to controls. These data suggest that ET-1 promotes adipose deposition and insulin resistance via the ETB receptor.

The contribution of collecting duct NOS1 to the concentrating mechanisms in male and female mice.

The collecting duct (CD) concentrates the urine, thereby maintaining body water volume and plasma osmolality within a normal range. The endocrine hormone arginine vasopressin acts in the CD to increase water permeability via the vasopressin 2 receptor (V2R)-aquaporin (AQP) axis. Recent studies have suggested that autocrine factors may also contribute to the regulation of CD water permeability. Nitric oxide is produced predominantly by nitric oxide synthase 1 (NOS1) in the CD and acts as a diuretic during salt loading. The present study sought to determine whether CD NOS1 regulates diuresis during changes in hydration status. Male and female control and CD NOS1 knockout (CDNOS1KO) mice were hydrated (5% sucrose water), water deprived, or acutely challenged with the V2R agonist desmopressin. In male mice, water deprivation resulted in decreased urine flow and increased plasma osmolality, copeptin concentration, and kidney AQP2 abundance independent of CD NOS1. In female control mice, water deprivation reduced urine flow, increased plasma osmolality and copeptin, but did not significantly change total AQP2; however, there was increased basolateral AQP3 localization. Surprisingly, female CDNOS1KO mice while on the sucrose water presented with symptoms of dehydration. Fibroblast growth factor 21, an endocrine regulator of sweetness preference, was significantly higher in female CDNOS1KO mice, suggesting that this was reducing their drive to drink the sucrose water. With acute desmopressin challenge, female CDNOS1KO mice failed to appropriately concentrate their urine, resulting in higher plasma osmolality than controls. In conclusion, CD NOS1 plays only a minor role in urine-concentrating mechanisms.

Dynamic changes in histone deacetylases following kidney ischemia-reperfusion injury are critical for promoting proximal tubule proliferation.

Deranged histone deacetylase (HDAC) activity causes uncontrolled proliferation, inflammation, fibrosis, and organ damage. It is unclear whether deranged HDAC activity results in acute kidney injury in the renal hypoperfusion model of bilateral ischemia-reperfusion injury (IRI) and whether in vivo inhibition is an appropriate therapeutic approach to limit injury. Male mice were implanted with intraperitoneal osmotic minipumps containing vehicle, the class I HDAC inhibitor, MS275, or the pan-HDAC inhibitor, trichostatin A (TSA), 3 days before sham/bilateral IRI surgery. Kidney cortical samples were analyzed using histological, immunohistochemical, and Western blotting techniques. HDAC-dependent proliferation rate was measured in immortalized rat epithelial cells and primary mouse or human proximal tubule (PT) cells. There were dynamic changes in cortical HDAC localization and abundance following IRI including a fourfold increase in HDAC4 in the PT. HDAC inhibition resulted in a significantly higher plasma creatinine, increased kidney damage, but reduced interstitial fibrosis compared with vehicle-treated IRI mice. HDAC-inhibited mice had reduced interstitial α-smooth muscle actin, fibronectin expression, and Sirius red-positive area, suggesting that IRI activates HDAC-mediated fibrotic pathways. In vivo proliferation of the kidney epithelium was significantly reduced in TSA-treated, but not MS275-treated, IRI mice, suggesting class II HDACs mediate proliferation. Furthermore, HDAC4 activation increased proliferation of human and mouse PTs. Kidney HDACs are activated during IRI with isoform-specific expression patterns. Our data point to mechanisms whereby IRI activates HDACs resulting in fibrotic pathways but also activation of PT proliferation and repair pathways. This study demonstrates the need to develop isoform-selective HDAC inhibitors for the treatment of renal hypoperfusion-induced injury.

Endothelin.

The endothelins comprise three structurally similar 21-amino acid peptides. Endothelin-1 and -2 activate two G-protein coupled receptors, ETA and ETB, with equal affinity, whereas endothelin-3 has a lower affinity for the ETA subtype. Genes encoding the peptides are present only among vertebrates. The ligand-receptor signaling pathway is a vertebrate innovation and may reflect the evolution of endothelin-1 as the most potent vasoconstrictor in the human cardiovascular system with remarkably long lasting action. Highly selective peptide ETA and ETB antagonists and ETB agonists together with radiolabeled analogs have accurately delineated endothelin pharmacology in humans and animal models, although surprisingly no ETA agonist has been discovered. ET antagonists (bosentan, ambrisentan) have revolutionized the treatment of pulmonary arterial hypertension, with the next generation of antagonists exhibiting improved efficacy (macitentan). Clinical trials continue to explore new applications, particularly in renal failure and for reducing proteinuria in diabetic nephropathy. Translational studies suggest a potential benefit of ETB agonists in chemotherapy and neuroprotection. However, demonstrating clinical efficacy of combined inhibitors of the endothelin converting enzyme and neutral endopeptidase has proved elusive. Over 28 genetic modifications have been made to the ET system in mice through global or cell-specific knockouts, knock ins, or alterations in gene expression of endothelin ligands or their target receptors. These studies have identified key roles for the endothelin isoforms and new therapeutic targets in development, fluid-electrolyte homeostasis, and cardiovascular and neuronal function. For the future, novel pharmacological strategies are emerging via small molecule epigenetic modulators, biologicals such as ETB monoclonal antibodies and the potential of signaling pathway biased agonists and antagonists.

Proteomic determination of the lysine acetylome and phosphoproteome in the rat native inner medullary collecting duct.

Phosphorylation and lysine (K)-acetylation are dynamic posttranslational modifications of proteins. Previous proteomic studies have identified over 170,000 phosphorylation sites and 15,000 K-acetylation sites in mammals. We recently reported that the inner medullary collecting duct (IMCD), which functions in the regulation of water-reabsorption, via the actions of vasopressin, expresses many of the enzymes that can modulated K-acetylation. The purpose of this study was to determine the K-acetylated or phosphorylated proteins expressed in IMCD cells. Second we questioned whether vasopressin V2 receptor activation significantly affects the IMCD acetylome or phosphoproteome? K-acetylated or serine-, threonine-, or tyrosine-phosphorylated peptides were identified from native rat IMCDs by proteomic analysis with four different enzymes (trypsin, chymotrypsin, ASP-N, or Glu-C) to generate a high-resolution proteome. K-acetylation was identified in 431 unique proteins, and 64% of the K-acetylated sites were novel. The acetylated proteins were expressed in all compartments of the cell and were enriched in pathways including glycolysis and vasopressin-regulated water reabsorption. In the vasopressin-regulated water reabsorption pathway, eight proteins were acetylated, including the novel identification of the basolateral water channel, AQP3, acetylated at K282; 215 proteins were phosphorylated in this IMCD cohort, including AQP2 peptides that were phosphorylated at four serines: 256, 261, 264, and 269. Acute dDAVP did not significantly affect the IMCD acetylome; however, it did significantly affect previously known vasopressin-regulated phosphorylation sites. In conclusion, presence of K-acetylated proteins involved in metabolism, ion, and water transport in the IMCD points to multiple roles of K-acetylation beyond its canonical role in transcriptional regulation.

High dietary sodium causes dyssynchrony of the renal molecular clock in rats.

Speed JS, Hyndman KA, Roth K, Heimlich JB, Kasztan M, Fox BM, Johnston JG, Becker BK, Jin C, Gamble KL, Young ME, Pollock JS, Pollock DM. High dietary sodium causes dyssynchrony of the renal molecular clock in rats. Am J Physiol Renal Physiol 314: F89-F98, 2018. First published September 27, 2017; doi:10.1152/ajprenal.00028.2017.-Dyssynchrony of circadian rhythms is associated with various disorders, including cardiovascular and metabolic diseases. The cell autonomous molecular clock maintains circadian control; however, environmental factors that may cause circadian dyssynchrony either within or between organ systems are poorly understood. Our laboratory recently reported that the endothelin (ET-1) B (ETB) receptor functions to facilitate Na+ excretion in a time of day-dependent manner. Therefore, the present study was designed to determine whether high salt (HS) intake leads to circadian dyssynchrony within the kidney and whether the renal endothelin system contributes to control of the renal molecular clock. We observed that HS feeding led to region-specific alterations in circadian clock components within the kidney. For instance, HS caused a significant 5.5-h phase delay in the peak expression of Bmal1 and suppressed Cry1 and Per2 expression in the renal inner medulla, but not the renal cortex, of control rats. The phase delay in Bmal1 expression appears to be mediated by ET-1 because this phenomenon was not observed in the ETB-deficient rat. In cultured inner medullary collecting duct cells, ET-1 suppressed Bmal1 mRNA expression. Furthermore, Bmal1 knockdown in these cells reduced epithelial Na+ channel expression. These data reveal that HS feeding leads to intrarenal circadian dyssynchrony mediated, in part, through activation of ETB receptors within the renal inner medulla.

Diurnal pattern in skin Na+ and water content is associated with salt-sensitive hypertension in ETB receptor-deficient rats.

Impairment in the ability of the skin to properly store Na+ nonosmotically (without water) has recently been hypothesized as contributing to salt-sensitive hypertension. Our laboratory has shown that endothelial production of endothelin-1 (ET-1) is crucial to skin Na+ handling. Furthermore, it is well established that loss of endothelin type B receptor (ETB) receptor function impairs Na+ excretion by the kidney. Thus we hypothesized that rats lacking functional ETB receptors (ETB-def) will have a reduced capacity of the skin to store Na+ during chronic high-salt (HS) intake. We observed that ETB-def rats exhibited salt-sensitive hypertension with an approximate doubling in the diurnal amplitude of mean arterial pressure compared with genetic control rats on a HS diet. Two weeks of HS diet significantly increased skin Na+ content relative to water; however, there was no significant difference between control and ETB-def rats. Interestingly, HS intake led to a 19% increase in skin Na+ and 16% increase in water content (relative to dry wt.) during the active phase (zeitgeber time 16) versus inactive phase (zeitgeber time 4, P < 0.05) in ETB-def rats. There was no significant circadian variation in total skin Na+ or water content of control rats fed normal or HS. These data indicate that ETB receptors have little influence on the ability to store Na+ nonosmotically in the skin during long-term HS intake but, rather, appear to regulate diurnal rhythms in skin Na+ content and circadian blood pressure rhythms associated with a HS diet.

Dynamic regulation of lysine acetylation: the balance between acetyltransferase and deacetylase activities.

Reversible posttranslational modification of proteins is a critically important process in physiological regulation in all tissues, including the kidney. Lysine acetylation occurs in all organisms, including prokaryotes, and is regulated by a balance between the lysine acetyltransferases (adding an acetyl group to the ε-amino group of a lysine) and deacetylases (removing it). The kidney is an organ rich with acetylated lysines, which map to >2,000 unique histone and nonhistone proteins. However, the functional significance of these modifications remains to be discovered. Here, we have compiled gene lists of the acetyltransferases and deacetylases in the mammalian genomes and mapped their mRNA expression along the renal tubule. These lists will be useful for generating targeted approaches to test the physiological or pathophysiological significance of lysine acetylation changes in the kidney.

Early life stress in male mice induces superoxide production and endothelial dysfunction in adulthood.

Early life stress (ELS) is a risk for cardiovascular disease in adulthood although very little mechanistic insight is available. Because oxidative stress and endothelial dysfunction are major contributors to cardiovascular risk, we hypothesized that ELS induces endothelial dysfunction in adult male mice via increased superoxide production. Studies employed a mouse model of ELS, maternal separation with early weaning (MSEW), in which litters were separated from the dam for 4 h/day [postnatal days (PD) 2-5] and 8 h/day (PD6-16), and weaned at PD17. Control litters remained undisturbed until weaning at PD21. When compared with control mice, thoracic aortic rings from adult male MSEW mice displayed significant endothelial dysfunction that was reversed by the superoxide scavenger, polyethylene glycol-superoxide dismutase (PEG-SOD). PEG-SOD-inhibitable superoxide production by aortae from MSEW mice was significantly greater than observed in control aortae, although unaffected by nitric oxide synthase inhibition, suggesting that uncoupled nitric oxide synthase was not responsible for the accelerated superoxide production. Aortic SOD expression, plasma SOD activity, and total antioxidant activity were similar in MSEW and control mice, indicating unaltered antioxidant capacity in MSEW mice. Increased expression of the NADPH oxidase subunits, NOX2 and NOX4, was evident in the aortae of MSEW mice. Moreover, endothelial dysfunction and superoxide production in MSEW mice was reversed with the NADPH oxidase inhibitor, apocynin, indicating increased NADPH oxidase-dependent superoxide production and endothelial dysfunction. The finding that MSEW induces superoxide production and endothelial dysfunction in adult mice may provide a mechanistic link between ELS and adult cardiovascular disease risk.