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Wei Cao,* Shuai Song,* Guojian Fang,* Yingze Li, Yuepeng Wang, Qun-Shan Wang Department of Cardiology, Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai, 200092, People’s Republic of China*These authors contributed equally to this workCorrespondence: Qun-Shan Wang; Yuepeng Wang Email wangqunshan@xinhuamed.com.cn; wangyuepeng@xinhuamed.com.cnBackground: Atrial fibrosis serves as a disease initiating mechanism in the development of atrial fibrillation. Angiotensin II (Ang-II), a key mediator for atrial fibrosis, aberrantly activates atrial fibroblasts (AFs) into myofibroblasts, resulting in subsequent excessive synthesis and deposition of extracellular matrix (ECM). Cadherin-11 (CDH11) is essential in the development of non-cardiac fibrotic diseases. In this study, we investigated its role in the pathogenesis and underlying mechanism of atrial fibrillation.Methods: We obtained left atrial tissues from either patients with atrial fibrillation or Ang-II-induced atrial fibrosis mice. We utilized a global CDH11 knockout mouse (CDH11−/-) model to determine the effect of CDH11 on AF cell proliferation, migration, ECM synthesis/deposition. RNA-Seq of isolated AFs from CDH11−/- or normal mice was performed and differential expressed genes were analyzed. The mouse susceptibility to atrial fibrillation was examined by cardiac electrophysiology.Results: We found that cadherin-11 was significantly up-regulated in fibrotic atrial tissue from patients with atrial fibrillation and Ang-II-induced mice. Both normal and CDH11−/- mice did not develop atrial fibrosis at resting state. However, after Ang-II infusion, unlike severe atrial fibrosis occurred in normal mice, CDH11−/- mice displayed a reduced atrial fibrosis. Atrial fibroblasts with CDH11 deletion from CDH11−/- mice showed reduction in Ang-II-induced cell proliferation, migration and ECM synthesis/deposition, indicating the involvement of CDH11 in atrial fibrosis. Consistently, RNA-Seq of CDH11-null AFs uncovered significant decrease in pro-fibrotic gene expression. In addition, we identified reduction of transcripts associated with Smad2/3, ERK1/2 and JNK pathways. Further, CDH11−/- mice showed a significantly attenuated Ang-II-induced susceptibility to atrial fibrillation.Conclusion: Our results indicate that CDH11 potentiates Ang-II-induced activation of AFs. The pathogenesis of atrial fibrosis is through CDH11 mediated stimulation of Smad2/3, ERK1/2 and JNK pathways. Thus, CDH11 might serve as a novel therapeutic target for ameliorating the development of atrial fibrillation.Keywords: atrial fibrillation, atrial fibrosis, atrial fibroblast, cadherin-11, angiotensin-II

Fibrosis serves a critical role in driving atrial remodelling‐mediated atrial fibrillation (AF). Abnormal levels of the transcription factor PU.1, a key regulator of fibrosis, are associated with cardiac injury and dysfunction following acute viral myocarditis. However, the role of PU.1 in atrial fibrosis and vulnerability to AF remain unclear. Here, an in vivo atrial fibrosis model was developed by the continuous infusion of C57 mice with subcutaneous Ang‐II, while the in vitro model comprised atrial fibroblasts that were isolated and cultured. The expression of PU.1 was significantly up‐regulated in the Ang‐II‐induced group compared with the sham/control group in vivo and in vitro. Moreover, protein expression along the TGF‐β1/Smads pathway and the proliferation and differentiation of atrial fibroblasts induced by Ang‐II were significantly higher in the Ang‐II‐induced group than in the sham/control group. These effects were attenuated by exposure to DB1976, a PU.1 inhibitor, both in vivo and in vitro. Importantly, in vitro treatment with small interfering RNA against Smad3 (key protein of TGF‐β1/Smads signalling pathway) diminished these Ang‐II‐mediated effects, and the si‐Smad3‐mediated effects were, in turn, antagonized by the addition of a PU.1‐overexpression adenoviral vector. Finally, PU.1 inhibition reduced the atrial fibrosis induced by Ang‐II and attenuated vulnerability to AF, at least in part through the TGF‐β1/Smads pathway. Overall, the study implicates PU.1 as a potential therapeutic target to inhibit Ang‐II‐induced atrial fibrosis and vulnerability to AF.

Aims Activation of the renin-angiotensin-aldosterone system plays an important role in the pathophysiology of heart failure (HF) and has been associated with poor prognosis. There are limited data on the associations of renin and aldosterone levels with clinical profiles, treatment response, and study outcomes in patients with HF.Methods and results We analysed 2,039 patients with available baseline renin and aldosterone levels in BIOSTAT-CHF (a systems BIOlogy study to Tailored Treatment in Chronic Heart Failure). The primary outcome was the composite of all-cause mortality or HF hospitalization. We also investigated changes in renin and aldosterone levels after administration of mineralocorticoid receptor antagonists (MRAs) in a subset of the EPHESUS trial and in an acute HF cohort (PORTO). In BIOSTAT-CHF study, median renin and aldosterone levels were 85.3 (percentile(25-75) = 28-247) mu IU/mL and 9.4 (percentile(25-75) = 4.4-19.8) ng/dL, respectively. Prior HF admission, lower blood pressure, sodium, poorer renal function, and MRA treatment were associated with higher renin and aldosterone. Higher renin was associated with an increased rate of the primary outcome [highest vs. lowest renin tertile: adjusted-HR (95% CI) = 1.47 (1.16-1.86), P = 0.002], whereas higher aldosterone was not [highest vs. lowest aldosterone tertile: adjusted-HR (95% CI) = 1.16 (0.93-1.44), P = 0.19]. Renin and/or aldosterone did not improve the BIOSTAT-CHF prognostic models. The rise in aldosterone with the use of MRAs was observed in EPHESUS and PORTO studies.Conclusions Circulating levels of renin and aldosterone were associated with both the disease severity and use of MRAs. By reflecting both the disease and its treatments, the prognostic discrimination of these biomarkers was poor. Our data suggest that the "point" measurement of renin and aldosterone in HF is of limited clinical utility.

Radiation-induced heart disease (RIHD) is a potential late side-effect of thoracic radiotherapy resulting in left ventricular hypertrophy (LVH) and fibrosis due to a complex pathomechanism leading to heart failure. Angiotensin-II receptor blockers (ARBs), including losartan, are frequently used to control heart failure of various etiologies. Preclinical evidence is lacking on the anti-remodeling effects of ARBs in RIHD, while the results of clinical studies are controversial. We aimed at investigating the effects of losartan in a rat model of RIHD. Male Sprague-Dawley rats were studied in three groups: (1) control, (2) radiotherapy (RT) only, (3) RT treated with losartan (per os 10 mg/kg/day), and were followed for 1, 3, or 15 weeks. At 15 weeks post-irradiation, losartan alleviated the echocardiographic and histological signs of LVH and fibrosis and reduced the overexpression of chymase, connective tissue growth factor, and transforming growth factor-beta in the myocardium measured by qPCR; likewise, the level of the SMAD2/3 protein determined by Western blot decreased. In both RT groups, the pro-survival phospho-AKT/AKT and the phospho-ERK1,2/ERK1,2 ratios were increased at week 15. The antiremodeling effects of losartan seem to be associated with the repression of chymase and several elements of the TGF-β/SMAD signaling pathway in our RIHD model.

Targeting the collecting duct water channel aquaporin 2 (AQP2) to the plasma membrane is essential for the maintenance of mammalian water homeostasis. The vasopressin V2 receptor (V2R), which is a GS protein-coupled receptor that increases intracellular cAMP levels, has a major role in this targeting process. Although a rise in cAMP levels and activation of protein kinase A are involved in facilitating the actions of V2R, studies in knockout mice and cell models have suggested that cAMP signalling pathways are not an absolute requirement for V2R-mediated AQP2 trafficking to the plasma membrane. In addition, although AQP2 phosphorylation is a known prerequisite for V2R-mediated plasma membrane targeting, none of the known AQP2 phosphorylation events appears to be rate-limiting in this process, which suggests the involvement of other factors; cytoskeletal remodelling has also been implicated. Notably, several regulatory processes and signalling pathways involved in AQP2 trafficking also have a role in the pathophysiology of autosomal dominant polycystic kidney disease, although the role of AQP2 in cyst progression is unknown. Here, we highlight advances in the field of AQP2 regulation that might be exploited for the treatment of water balance disorders and provide a rationale for targeting these pathways in autosomal dominant polycystic kidney disease.

Background: In healthy pregnancies, components of the Renin-Angiotensin system (RAS) are present in the placental villi and contribute to invasion, migration, and angiogenesis. At the same time, soluble fms-like tyrosine kinase 1 (sFlt-1) production is induced after binding of ANG-II to its receptor (AT-1R) in response to hypoxia. As RAS plays an essential role in the pathogenesis of COVID-19, we hypothesized that angiogenic marker (sFlt-1) and RAS components (ANG-II and ACE-2) may be related to adverse outcomes in pregnant women with COVID-19; Methods: Prospective cohort study. Primary outcome was severe pneumonia. Secondary outcomes were ICU admission, intubation, sepsis, and death. Spearman’s Rho test was used to analyze the correlation between sFlt-1 and ANG-II levels. The sFlt-1/ANG-II ratio was determined and the association with each adverse outcome was explored by logistic regression analysis and the prediction was assessed using receiver-operating-curve (ROC); Results: Among 80 pregnant women with COVID-19, the sFlt-1/ANG-II ratio was associated with an increased probability of severe pneumonia (odds ratio [OR]: 1.31; p = 0.003), ICU admission (OR: 1.05; p = 0.007); intubation (OR: 1.09; p = 0.008); sepsis (OR: 1.04; p = 0.008); and death (OR: 1.04; p = 0.018); Conclusion: sFlt-1/ANG-II ratio is a good predictor of adverse events such as pneumonia, ICU admission, intubation, sepsis, and death in pregnant women with COVID-19.

[EN] Hypertension induces significant aortic remodelling, often adaptive but sometimes not. To identify immuno-mechanical mechanisms responsible for differential remodelling, we studied thoracic aortas from 129S6/SvEvTac and C57BL/6 J mice before and after continuous 14-day angiotensin II infusion, which elevated blood pressure similarly in both strains. Histological and biomechanical assessments of excised vessels were similar at baseline, suggesting a common homeostatic set-point for mean wall stress. Histology further revealed near mechano-adaptive remodelling of the hypertensive 129S6/SvEvTac aortas, but a grossly maladaptive remodelling of C57BL/6 J aortas. Bulk RNA sequencing suggested that increased smooth muscle contractile processes promoted mechano-adaptation of 129S6/SvEvTac aortas while immune processes prevented adaptation of C57BL/6 J aortas. Functional studies confirmed an increased vasoconstrictive capacity of the former while immunohistochemistry demonstrated marked increases in inflammatory cells in the latter. We then used multiple computational biomechanical models to test the hypothesis that excessive adventitial wall stress correlates with inflammatory cell infiltration. These models consistently predicted that increased vasoconstriction against an increased pressure coupled with modest deposition of new matrix thickens the wall appropriately, restoring wall stress towards homeostatic consistent with adaptive remodelling. By contrast, insufficient vasoconstriction permits high wall stresses and exuberant inflammation-driven matrix deposition, especially in the adventitia, reflecting compromised homeostasis and gross maladaptation., This work was supported by grants from NIH (grant nos. R01HL105297, P01 HL134605, U01 HL142518, R01 HL146723), Netherlands Organisation for Scientific Research (grant no. Rubicon 452172006) and the European Union¿s Horizon 2020 research and innovation program (grant no. 793805)

Pathological left ventricular hypertrophy (LVH) occurs in response to pressure overload and remains the single most important clinical predictor of cardiac mortality. The molecular pathways in the induction of pressure overload LVH are potential targets for therapeutic intervention. Current treatments aim to remove the pressure overload stimulus for LVH, but do not completely reverse adverse cardiac remodelling. Although numerous molecular signalling steps in the induction of LVH have been identified, the initial step by which mechanical stretch associated with cardiac pressure overload is converted into a chemical signal that initiates hypertrophic signalling remains unresolved. In this study, we show that selective deletion of transient receptor potential melastatin 4 (TRPM4) channels in mouse cardiomyocytes results in an approximately 50% reduction in the LVH induced by transverse aortic constriction. Our results suggest that TRPM4 channel is an important component of the mechanosensory signalling pathway that induces LVH in response to pressure overload and represents a potential novel therapeutic target for the prevention of pathological LVH. ispartof: ELIFE vol:10 ispartof: location:England status: published

Introduction: Myocardial remodeling due to large atrial septum defect (ASD) is macroscopically characterized by dilation of the right-sided cardiac cavities secondary to volume overload, the cellular mechanisms of which are not yet understood. We postulated that inflammation, fibrosis, and cell death are actors of right atrial remodeling secondary to ASD. Patients and Methods: In 12 children with large ASD (median age: 63 months), expression of genes coding for proteins involved in the response to cell stress and -protection, inflammation, growth and angiogenesis, fibrosis, and apoptosis was assessed by RT-PCR in right atrial myocardial biopsies taken during cardiac surgery. The presence of cytokines in myocardial cells was confirmed by immunohistochemistry and effective apoptosis by TUNEL assay. Results: In all patients investigated, a cellular response to early mechanical stress with the initiation of early protective mechanisms, of inflammation (and its control), -growth, and -angiogenesis, of fibrosis and apoptosis was present. The apoptotic index assessed by TUNEL assay averaged 0.3%. Conclusions: In children with large ASD, macroscopic right atrial remodeling relates to cellular mechanisms involving the expression of numerous genes that either still act to protect cells and tissues but that also harm as they initiate and/or sustain inflammation, fibrosis, and cell death by apoptosis. This may contribute to long term morbidity in patients with ASD. ispartof: Frontiers In Pediatrics vol:8 ispartof: location:Switzerland status: published

International journal of molecular sciences 21(18), 6547 (2020). doi:10.3390/ijms21186547 special issue: "Special Issue "The Role of Non-coding RNAs in Kidney Diseases" / Special Issue Editors: Prof. Dr. Laurent Metzinger, Guest Editor; Dr. Juan Antonio Moreno, Guest Editor; Dr. Valérie Metzinger-Le Meuth, Guest Editor", Published by Molecular Diversity Preservation International, Basel

Context No consensus exists for optimization of mineralocorticoid therapy in patients with primary adrenal insufficiency. Objective To explore the relationship between mineralocorticoid (MC) replacement dose, plasma renin concentration (PRC), and clinically important variables to determine which are most helpful in guiding MC dose titration in primary adrenal insufficiency. Design Observational, retrospective, longitudinal analysis. Patients A total of 280 patients (with 984 clinical visits and plasma renin measurements) with primary adrenal insufficiency were recruited from local databases and the international congenital adrenal hyperplasia (CAH) registry (www.i-cah.org). Thirty-seven patients were excluded from the final analysis due to incomplete assessment. Data from 204 patients with salt-wasting CAH (149 adults and 55 children) and 39 adult patients with Addison disease (AD) were analysed. Main outcome measures PRC, electrolytes, blood pressure (BP), and anthropometric parameters were used to predict their utility in optimizing MC replacement dose. Results PRC was low, normal, or high in 19%, 36%, and 44% of patients, respectively, with wide variability in MC dose and PRC. Univariate analysis demonstrated a direct positive relationship between MC dose and PRC in adults and children. There was no relationship between MC dose and BP in adults, while BP increased with increasing MC dose in children. Using multiple regression modeling, sodium was the only measurement that predicted PRC in adults. Longitudinally, the change in MC dose was able to predict potassium, but not BP or PRC. Conclusions The relationship between MC dose and PRC is complex and this may reflect variability in sampling with respect to posture, timing of last MC dose, adherence, and concomitant medications. Our data suggest that MC titration should not primarily be based only on PRC normalization, but also on clinical parameters such as BP and electrolyte concentration.

Background/Aims: Chronic heart failure (HF) disrupts normal kidney function and leads to cardiorenal syndrome that further promotes HF progression. To identify potential participants in HF-related injury, we analyzed kidney proteome in an established HF model. Methods: HF was induced by chronic volume overload in male HanSD rats using aorto-caval fistula. After 21 weeks, cardiac and renal functions (in-situ kidney study) and renal proteomics were studied in sham-operated (controls) and HF rats, using iTRAQ labeling and LC-MS with Orbitrap Fusion, leading to identification and quantification of almost 4000 proteins. Results: Compared to controls, HF rats had cardiac hypertrophy, systemic and pulmonary congestion. Kidneys of HF rats had reduced renal blood flow, sodium excretion and urine production. While glomerular filtration rate, serum cystatin C and creatinine were still normal compared to controls, HF kidneys showed albuminuria and markedly increased tissue angiotensin-II levels (5-fold). HF kidneys (versus controls) displayed differential expression (˃1.5-fold) of 67 proteins. The most upregulated were angiotensin-converting enzyme (ACE, ˃20-fold), advanced glycosylation product-specific receptor (RAGE, 14-fold), periostin (6.8-fold), caveolin-1 (4.5-fold) and other proteins implicated in endothelial function (vWF, cavins 1-3, T-kininogen 2), proinflammatory ECM activation (MFAP4, collagen-VI, galectin-3, FHL-1, calponin) and proteins involved in glomerular filtration membrane integrity (CLIC5, ZO-1). Carboxylesterase-1D (CES1D), an enzyme that converts ACE inhibitors or sacubitril into active drugs, was also upregulated in HF kidneys. Conclusion: Chronic HF leads to latent kidney injury, associated with deep changes in kidney protein composition. These alterations may act in concert with intrarenal renin-angiotensin system activation and may serve as markers and/or targets to tackle cardiorenal syndrome.

Kidney damage is a common consequence of arterial hypertension, but is also a cause of atherogenesis. Dysfunction and/or harm of the endothelium in glomeruli and tubular interstitium damage the function of these structures and translates into dynamic changes of filtration fraction, with progressive reduction in glomerular filtration rate, expansion of extracellular fluid volume, abnormal ion balance, and hypoxia, ultimately leading to chronic kidney disease. Considering the key role played by endothelial dysfunction in chronic kidney disease, the Working Group on Endothelin and Endothelial Factors of the European Society of Hypertension and the Japanese Society of Hypertension have critically reviewed available knowledge on the mechanisms underlying endothelial cell injury. This resulted into two articles: in the first, we herein examine the mechanisms by which endothelial factors induce vascular remodeling and the role of different players, including endothelin-1, the renin-angiotensin-aldosterone system and their interactions, and of oxidative stress; in the second, we discuss the role of endothelial dysfunction in the major disease conditions that affect the kidney.

Pressure overload causes cardiac fibroblast activation and transdifferentiation, leading to increased interstitial fibrosis formation and subsequently myocardial stiffness, diastolic and systolic dysfunction, and eventually heart failure. A better understanding of the molecular mechanisms underlying pressure overload-induced cardiac remodeling and fibrosis will have implications for heart failure treatment strategies. The microRNA (miRNA)-221/222 family, consisting of miR-221-3p and miR-222-3p, is differentially regulated in mouse and human cardiac pathology and inversely associated with kidney and liver fibrosis. We investigated the role of this miRNA family during pressure overload-induced cardiac remodeling. In myocardial biopsies of patients with severe fibrosis and dilated cardiomyopathy or aortic stenosis, we found significantly lower miRNA-221/222 levels as compared to matched patients with nonsevere fibrosis. In addition, miRNA-221/222 levels in aortic stenosis patients correlated negatively with the extent of myocardial fibrosis and with left ventricular stiffness. Inhibition of both miRNAs during AngII (angiotensin II)-mediated pressure overload in mice led to increased fibrosis and aggravated left ventricular dilation and dysfunction. In rat cardiac fibroblasts, inhibition of miRNA-221/222 derepressed TGF-beta (transforming growth factor-beta)-mediated profibrotic SMAD2 (mothers against decapentaplegic homolog 2) signaling and downstream gene expression, whereas overexpression of both miRNAs blunted TGF-beta-induced profibrotic signaling. We found that the miRNA-221/222 family may target several genes involved in TGF-beta signaling, including JNK1 (c-Jun N-terminal kinase 1), TGF-beta receptor 1 and TGF-beta receptor 2, and ETS-1 (ETS proto-oncogene 1). Our findings show that heart failure-associated downregulation of the miRNA-221/222 family enables profibrotic signaling in the pressure-overloaded heart.

Aims: Currently, no acute heart failure (AHF) therapy definitively improves outcomes. Reducing morbidity and mortality from acute heart failure (AHF) remains an unmet need. TRV027 is a novel ` biased' ligand of the angiotensin II type 1 receptor (AT1R), selectively antagonizing the negative effects of angiotensin II, while preserving the potential pro-contractility effects of AT1R stimulation. BLAST-AHF was designed to determine the safety, efficacy, and optimal dose of TRV027 to advance into future studies.Methods and results: BLAST-AHF was a multi-centre, international, randomized, double-blind, placebo-controlled, parallel group, phase IIb dose-ranging study, enrolling patients with AHF into 4 groups: placebo, 1, 5, or 25mg/h of TRV027. Treatment was by IV infusion for 48-96 h. The primary composite endpoint was comprised of the following: (i) time from baseline to death through day 30, (ii) time from baseline to heart failure re-hospitalization through day 30, (iii) the first assessment time point following worsening heart failure through day 5, (iv) change in dyspnea visual analogue scale (VAS) score calculated as the area under the curve (AUC) representing the change from baseline over time from baseline through day 5, and (v) length of initial hospital stay (in days) from baseline. Analyses were by modified intention-to-treat. Overall, 621 patients were enrolled. After 254 patients, a pre-specified interim analysis resulted in several protocol changes, including a lower blood pressure inclusion criterion as well as a new allocation scheme of 2: 1: 2: 1, overweighting both placebo, and the 5mg/h dose. TRV027 did not confer any benefit over placebo at any dose with regards to the primary composite endpoint or any of the individual components. There were no significant safety issues with TRV027.Conclusion: In this phase IIb dose-ranging AHF study, TRV027 did not improve clinical status through 30-day follow-up compared with placebo.

Hyperactivity of the renin-angiotensin system (RAS) is associated with the pathogenesis of Alzheimer’s disease (AD) believed to be mediated by angiotensin-II (Ang-II) activation of the angiotensin type 1 receptor (AT1R). We previously showed that angiotensin-converting enzyme-1 (ACE-1) activity, the rate-limiting enzyme in the production of Ang-II, is increased in human postmortem brain tissue in AD. Angiotensin-III (Ang-III) activates the AT1R and angiotensin type-2 receptor (AT2R), but its potential role in the pathophysiology of AD remains unexplored. We measured Ang-II and Ang-III levels by ELISA, and the levels and activities of aminopeptidase-A (AP-A) and aminopeptidase-N (AP-N) (responsible for the production and metabolism of Ang-III, respectively) in human postmortem brain tissue in the mid-frontal cortex (Brodmann area 9) in a cohort of AD (n = 90) and age-matched non-demented controls (n = 59), for which we had previous measurements of ACE-1 activity, Aβ level, and tau pathology (also in the mid-frontal cortex). We found that both Ang-II and Ang-III levels were significantly higher in AD compared to age-matched controls and that Ang-III, rather than Ang-II, was strongly associated with Aβ load and tau load. Levels of AP-A were significantly reduced in AD but AP-A enzyme activity was unchanged whereas AP-N activity was reduced in AD but AP-N protein level was unchanged. Together, these data indicate that the APA/Ang-III/APN/Ang-IV/AT4R pathway is dysregulated and that elevated Ang-III could contribute to the pathogenesis of AD.

Mesenchymal stem cells (MSCs) pretreatment is an effective route for improving cell-based therapy of endothelial cell survival, vascular stabilization, and angiogenesis. We hypothesized that the application of human umbilical cord-MSCs (hUC-MSCs) pretreated with angiotensin-II (Ang-II) might be a potential therapeutic approach for severe acute pancreatitis (SAP). Therefore, the effect of Ang-II pretreated hUC-MSCs on SAP was investigated in vitro and in vivo. Methods In the present study, human umbilical cord-derived MSCs pretreated with or without Ang-II were delivered through the tail vein of rats 12 h after induction of SAP. Pancreatitis severity scores and serum lipase levels, as well as the levels of VEGF and VEGFR2 were evaluated. Results We found that the administration of Ang-II-MSCs significantly inhibited pancreatic injury, as reflected by reductions of pancreatitis severity scores, serum amylase and serum lipase levels. Furthermore, the reduced apoptotic rate and increased tube formation in human umbilical vein endothelial Cells (HUVEC) were found resulting from the administration of Ang-II-MSC-CM. Moreover, knockdown of VEGFR2 can block the effect of Ang-II-MSC-CM on preventing HUVEC from apoptosis, as well as the capacity of tube formation was also suppressed. In addition, the expression of increased Bcl-2 and alleviated caspase-3 were observed in HUVEC and HUVEC transfectants exposure to Ang-II-MSC-CM. Conclusion Collectively, these results elucidated that the pretreatment of hUC-MSCs with Ang-II improved the outcome of MSC-based therapy for SAP via enhancing angiogenesis and ameliorating endothelial cell dysfunction in a VEGFR2 dependent manner.

The epidermal growth factor receptor (EGFR) pathway has a critical role in renal development, tissue repair and electrolyte handling. Numerous studies have reported an association between dysregulation of this pathway and the initiation and progression of various chronic kidney diseases such as diabetic nephropathy, chronic allograft nephropathy and polycystic kidney disease through the promotion of renal cell proliferation, fibrosis and inflammation. In the oncological setting, compounds that target the EGFR pathway are already in clinical use or have been evaluated in clinical trials; in the renal setting, therapeutic interventions targeting this pathway by decreasing ligand availability with disintegrin and metalloproteinase inhibitors or with ligand-neutralizing antibodies, or by inhibiting receptor activation with tyrosine kinase inhibitors or monoclonal antibodies are only just starting to be explored in animal models of chronic kidney disease and in patients with autosomal dominant polycystic kidney disease. In this Review we focus on the role of the EGFR signalling pathway in the kidney under physiological conditions and during the pathophysiology of chronic kidney diseases and explore the clinical potential of interventions in this pathway to treat chronic renal diseases.

Distal tubular sodium retention is a potent driver of hypertension, and the thiazide–sensitive sodium-chloride cotransporter (NCC) has a key role in this process. In humans, factors regulating NCC are unclear, but in animal models, aldosterone is a potent regulator, possibly via effects on plasma potassium. We studied the effects of the mineralocorticoid fludrocortisone on the abundance of NCC and its phosphorylated form (pNCC) as well as WNK lysine deficient protein kinase 4 (WNK4) and STE20/SPS1–related, proline alanine–rich kinase (SPAK) in human urinary exosomes. We isolated exosomes from daily urine samples in 25 patients undergoing fludrocortisone suppression testing (100 μg every 6 hours for 4 days) to diagnose or exclude primary aldosteronism. Over the course of the test, NCC levels increased 3.68-fold (P

Summary Cardiac ischemia leads to the loss of myocardial tissue and the activation of a repair process that culminates in the formation of a scar whose structural characteristics dictate propensity to favorable healing or detrimental cardiac wall rupture. To elucidate the cellular processes underlying scar formation, here we perform unbiased single-cell mRNA sequencing of interstitial cells isolated from infarcted mouse hearts carrying a genetic tracer that labels epicardial-derived cells. Sixteen interstitial cell clusters are revealed, five of which were of epicardial origin. Focusing on stromal cells, we define 11 sub-clusters, including diverse cell states of epicardial- and endocardial-derived fibroblasts. Comparing transcript profiles from post-infarction hearts in C57BL/6J and 129S1/SvImJ inbred mice, which displays a marked divergence in the frequency of cardiac rupture, uncovers an early increase in activated myofibroblasts, enhanced collagen deposition, and persistent acute phase response in 129S1/SvImJ mouse hearts, defining a crucial time window of pathological remodeling that predicts disease outcome., Graphical Abstract, Highlights • Longitudinal transcriptional profiling of cardiac interstitial cells post-infarct • Identification of epicardial versus endocardial origin of cardiac stromal cells • A distinct early injury-response signature precedes appearance of myofibroblasts • Modulation of early fibrosis predicts cardiac rupture and pathological remodeling, Using single-cell transcriptional profiling of mouse hearts carrying a reporter for epicardial-derived cells, Forte et al. provide a dynamic view of cardiac interstitial responses across acute and chronic phases of remodeling post-infarction. Comparing responses on diverse genetic backgrounds reveals novel cellular and transcriptional features of cardiac rupture propensity.

Targeted HFpEF therapy based on matchmaking of human and animal models. Am J Physiol Heart Circ Physiol 315: H1670-H1683, 2018. First published September 21, 2018; doi:10.1152/ajpheart.00024.2018. The diversity in clinical phenotypes and poor understanding of the underlying pathophysiology of heart failure with preserved ejection fraction (HFpEF) is the main reason why no effective treatments have been found yet. Targeted, instead of one size fits all, treatment seems the only promising approach for treating 1114pE14. To be able to design a targeted, phenotype-specific HFpEF treatment, the matrix relating clinical phenotypes and underlying pathophysiological mechanisms has to be clarified. This review discusses the opportunities for additional evaluation of the underlying pathophysiological processes, e.g., to evaluate biological phenotypes on top of clinical routine, to guide us toward a phenotype specific HFpEF treatment. Moreover, a translational approach with matchmaking of animal models to biological HFpEF phenotypes will be a valuable step to test the effectiveness of novel, targeted interventions in IIFpEF.

Systemic sclerosis (SSc) is a rare connective tissue disease characterized by autoimmunity, vasculopathy, and progressive fibrosis typically affecting multiple organs including the skin. SSc often is a lethal disorder, because effective disease-modifying treatment still remains unavailable. Vasculopathy with endothelial dysfunction, perivascular infiltration of mononuclear cells, vascular wall remodeling and rarefaction of capillaries is the hallmark of the disease. Most patients present with vasospastic attacks of the digital arteries referred to as ‘Raynaud’s phenomenon,’ which is often an indication of an underlying widespread vasculopathy. Although autoimmune responses and inflammation are both found to play an important role in the pathogenesis of this vasculopathy, no definite initiating factors have been identified. Recently, several studies have underlined the potential role of oxidative stress in the pathogenesis of SSc vasculopathy thereby proposing a new aspect in the pathogenesis of this disease. For instance, circulating levels of reactive oxygen species (ROS) related markers have been found to correlate with SSc vasculopathy, the formation of fibrosis and the production of autoantibodies. Excess ROS formation is well-known to lead to endothelial cell (EC) injury and vascular complications. Collectively, these findings suggest a potential role of ROS in the initiation and progression of SSc vasculopathy. In this review, we present the background of oxidative stress related processes (e.g., EC injury, autoimmunity, inflammation, and vascular wall remodeling) that may contribute to SSc vasculopathy. Finally, we describe the use of oxidative stress related read-outs as clinical biomarkers of disease activity and evaluate potential anti-oxidative strategies in SSc.

Background Renal hypoxia, implicated as crucial factor in onset and progression of chronic kidney disease, may be attributed to reduced nitric oxide because nitric oxide dilates vasculature and inhibits mitochondrial oxygen consumption. We hypothesized that chronic nitric oxide synthase inhibition would induce renal hypoxia. Methods and Results Oxygen‐sensitive electrodes, attached to telemeters, were implanted in either renal cortex (n=6) or medulla (n=7) in rats. After recovery and stabilization, baseline oxygenation ( pO 2 ) was recorded for 1 week. To inhibit nitric oxide synthase, N‐ω‐nitro‐ l ‐arginine (L‐NNA; 40 mg/kg/day) was administered via drinking water for 2 weeks. A separate group (n=8), instrumented with blood pressure telemeters, followed the same protocol. L‐NNA rapidly induced hypertension (165±6 versus 108±3 mm Hg; P P pO 2 , after initially dipping, returned to baseline and then increased. Medullary pO 2 decreased progressively (up to −19±6% versus baseline; P pO 2 was decreased (3.7 [2.2–5.3] versus 7.9 [7.5–8.4]; P pO 2 were unaltered. Terminal glomerular filtration rate (1374±74 versus 2098±122 μL/min), renal blood flow (5014±336 versus 9966±905 μL/min), and sodium reabsorption efficiency (13.0±0.8 versus 22.8±1.7 μmol/μmol) decreased (all P Conclusions For the first time, we show temporal development of renal cortical and medullary oxygenation during chronic nitric oxide synthase inhibition in unrestrained conscious rats. Whereas cortical pO 2 shows transient changes, medullary pO 2 decreased progressively. Chronic L‐NNA leads to decreased renal perfusion and sodium reabsorption efficiency, resulting in progressive medullary hypoxia, suggesting that juxtamedullary nephrons are potentially vulnerable to prolonged nitric oxide depletion.

Bicuspid valve disease is associated with the development of thoracic aortic aneurysm. The molecular mechanisms underlying this association still need to be clarified. Here, we evaluated the circulating levels of T and B lymphocyte subsets associated with the development of vascular diseases in patients with bicuspid aortic valve or tricuspid aortic valve with and without thoracic aortic aneurysm. We unveiled that the circulating levels of the MAIT, CD4+IL−17A+, and NKT T cell subsets were significantly reduced in bicuspid valve disease cases, when compared to tricuspid aortic valve cases in either the presence or the absence of thoracic aortic aneurysm. Among patients with tricuspid aortic valve, these cells were higher in those also affected by thoracic aortic aneurysm. Similar data were obtained by examining CD19+ B cells, naïve B cells (IgD+CD27−), memory unswitched B cells (IgD+CD27+), memory switched B cells (IgD−CD27+), and double-negative B cells (DN) (IgD−CD27−). These cells resulted to be lower in subjects with bicuspid valve disease with respect to patients with tricuspid aortic valve. In whole, our data indicate that patients with bicuspid valve disease show a quantitative reduction of T and B lymphocyte cell subsets. Future studies are encouraged to understand the molecular mechanisms underlying this observation and its pathophysiological significance.

Standard therapies for heart failure with preserved ejection fraction (HFpEF) have been unsuccessful, demonstrating that the contribution of the underlying diastolic dysfunction pathophysiology differs from that of systolic dysfunction in heart failure and currently is far from being understood. Complicating the investigation of HFpEF is the contribution of several comorbidities. Here, we selected three established rat models of diastolic dysfunction defined by three major risk factors associated with HFpEF and researched their commonalities and differences. The top differentially expressed genes in the left ventricle of Dahl salt sensitive (Dahl/SS), spontaneous hypertensive heart failure (SHHF), and diabetes 1 induced HFpEF models were derived from published data in Gene Expression Omnibus and used for a comprehensive interpretation of the underlying pathophysiological context of each model. The diversity of the underlying transcriptomic of the heart of each model is clearly observed by the different panel of top regulated genes: the diabetic model has 20 genes in common with the Dahl/SS and 15 with the SHHF models. Advanced analytics performed in Ingenuity Pathway Analysis (IPA (R)) revealed that Dahl/SS heart tissue transcripts triggered by upstream regulators lead to dilated cardiomyopathy, hypertrophy of heart, arrhythmia, and failure of heart. In the heart of SHHF, a total of 26 genes were closely linked to cardiovascular disease including cardiotoxicity, pericarditis, ST-elevated myocardial infarction, and dilated cardiomyopathy. IPA Upstream Regulator analyses revealed that protection of cardiomyocytes is hampered by inhibition of the ERBB2 plasma membrane-bound receptor tyrosine kinases. Cardioprotective markers such as natriuretic peptide A (NPPA), heat shock 27 kDa protein 1 (HSPB1), and angiogenin (ANG) were upregulated in the diabetes 1 induced model; however, the model showed a different underlying mechanism with a majority of the regulated genes involved in metabolic disorders. In conclusion, our findings suggest that multiple mechanisms may contribute to diastolic dysfunction and HFpEF, and thus drug therapies may need to be guided more by phenotypic characteristics of the cardiac remodeling events than by the underlying molecular processes.

Within the lungs, fibrosis can affect both the parenchyma and the airways. Fibrosis is a hallmark pathological change in the parenchyma in patients with idiopathic pulmonary fibrosis (IPF), whilst in asthma or chronic obstructive pulmonary disease (COPD) fibrosis is a component of the remodelling of the airways. In the past decade, significant advances have been made in understanding the disease behaviour and pathogenesis of parenchymal and airway fibrosis and as a result a variety of novel therapeutic targets for slowing or preventing progression of these fibrotic changes have been identified. This review highlights a number of these targets and discusses the potential for treating parenchymal or airway fibrosis through these mediators/pathways in the future.

Background Besides hypertension, obesity and the metabolic syndrome have recently emerged as risk factors for atrial fibrillation. This study sought to delineate the development of an arrhythmogenic substrate for atrial fibrillation in hypertension with and without concomitant obesity and metabolic syndrome. Methods and Results We compared obese spontaneously hypertensive rats ( SHR ‐obese, n=7–10) with lean hypertensive controls ( SHR ‐lean, n=7–10) and normotensive rats (n=7–10). Left atrial emptying function (MRI) and electrophysiological parameters were characterized before the hearts were harvested for histological and biochemical analyses. At the age of 38 weeks, SHR ‐obese, but not SHR ‐lean, showed increased body weight and impaired glucose tolerance together with dyslipidemia compared with normotensive rats. Mean blood pressure was similarly increased in SHR ‐lean and SHR ‐obese when compared with normotensive rats (178±9 and 180±8 mm Hg [not significant] versus 118±5 mm Hg, P SHR ‐obese than in SHR ‐lean. Impairment of left atrial emptying function, increase in total atrial activation time, and conduction heterogeneity, as well as prolongation of inducible atrial fibrillation durations, were more pronounced in SHR ‐obese as compared with SHR ‐lean. Histological and biochemical examinations revealed enhanced triglycerides and more pronounced fibrosis in the left atrium of SHR ‐obese. Besides increased expression of profibrotic markers in SHR ‐lean and SHR ‐obese, the profibrotic extracellular matrix protein osteopontin was highly upregulated only in SHR ‐obese. Conclusions In addition to hypertension alone, concomitant obesity and metabolic syndrome add to the atrial arrhythmogenic phenotype by impaired left atrial emptying function, local conduction abnormalities, interstitial atrial fibrosis formation, and increased propensity for atrial fibrillation.

Binding of renin and prorenin to the (pro)renin receptor (PRR) increases their enzymatic activity and upregulates the expression of pro-fibrotic genes in vitro. Expression of PRR is increased in the heart and kidney of hypertensive and diabetic animals, but its causative role in organ damage is still unclear. To determine whether increased expression of PRR is sufficient to induce cardiac or renal injury, we generated a mouse that constitutively overexpresses PRR by knocking-in the Atp6ap2/PRR gene in the hprt locus under the control of a CMV immediate early enhancer/chicken beta-actin promoter. Mice were backcrossed in the C57BI/6 and FVB/N strain and studied at the age of 12 months. In spite of a 25- to 80-fold renal and up to 400-fold cardiac increase in Atp6ap2/PRR expression, we found no differences in systolic blood pressure or albuminuria between wild-type and PRR overexpressing littermates. Histological examination did not show any renal or cardiac fibrosis in mutant mice. This was supported by real-time PCR analysis of inflammatory markers as well as of pro-fibrotic genes in the kidney and collagen in cardiac tissue. To determine whether the concomitant increase of renin would trigger fibrosis, we treated PRR overexpressing mice with the angiotensin receptor-1 blocker losartan over a period of 6 weeks. Renin expression increased eightfold in the kidney but no renal injury could be detected. In conclusion, our results suggest no major role for PRR in organ damage per se or related to its function as a receptor of renin.

Preeclampsia is a multisystem pregnancy complication, which affects 2% to 8% of all pregnancies.1 It is characterized by hypertension and proteinuria in the second half of pregnancy.2 Although the complete pathophysiology is still unknown, it is thought to consist of 2 phases. The first phase is poor placentation, which may result in hypoxia of the placenta.3,4 The second phase is characterized by the release of proinflammatory factors from the hypoxic placenta, resulting in systemic inflammation and endothelial cell dysfunction. As a result, hypertension and proteinuria associated with potential damage to multiple organs may develop.3,4 Delivery of the placenta and the fetus is the only effective treatment option for the maternal symptoms. High levels of ATP, which is now recognized as a danger signal, are found in preeclampsia.5,6 ATP is released by hypoxic and necrotic tissue, for instance by the hypoxic placenta.7,8 Release into the circulation causes activation of immune and endothelial cells,5,9 which in turn can also produce ATP, resulting in a cascade of activation.5,10 As a protective mechanism, ATP can be hydrolyzed into adenosine by various extracellular enzymes present in multiple cells including endothelial cells and placental trophoblast cells.11,12 Adenosine is also increased in preeclampsia and has opposite effects from ATP.13 Hence, the final effect of ATP and adenosine in preeclampsia depends on the balance between the 2 molecules. In the current review, we will discuss the role of ATP and adenosine in the pathogenesis of preeclampsia. We will first discuss the current knowledge on the biology of the 2 molecules in vascular function and the immune system, followed by an overview of how ATP and adenosine can play a role in pregnancy and preeclampsia. …

Reduction of dietary sodium intake or diuretic treatment increases renin-angiotensin-aldosterone system (RAAS) blockade efficacy in non-diabetic nephropathy. We aimed to investigate the effect of sodium restriction and the diuretic hydrochlorothiazide, separately and in combination, added to RAAS blockade on residual albuminuria in patients with type 2 diabetic nephropathy.In this multicentre, double-blind, placebo-controlled, crossover randomised trial, we included patients with type 2 diabetic nephropathy. Main entry criteria were microalbuminaria or macroalbuminuria, and creatinine clearance of 30 mL/min or higher with less than 6 mL/min decline in the previous year. We tested the separate and combined effects of sodium restriction (dietary counselling in the outpatient setting) and hydrochlorothiazide (50 mg daily), added to standardised maximal angiotensin-converting enzyme (ACE) inhibition (lisinopril 40 mg daily), on albuminuria (primary endpoint). Patients were given hydrochlorothiazide (50 mg per day) or placebo during four treatment periods of 6 weeks. Both treatments were combined with regular sodium diet or sodium restriction (target sodium intake 50 mmol Na(+) per day). The 6-week treatment periods were done consecutively in a random order. Patients were randomised in blocks of two patients. The trial was analysed by intention to treat. The trial is registered with TrialRegister.nl, number 2366.Of 89 eligible patients, 45 were included in the study. Both sodium restriction and hydrochlorothiazide significantly reduced albuminuria, irrespective of treatment sequence. Residual geometric mean albuminuria with baseline treatment was 711 mg per day (95% CI 485-1043); it was significantly reduced by sodium restriction (393 mg per day [258-599], p=0·0002), by hydrochlorothiazide (434 mg per day [306-618], p=0·0003), and to the greatest extent by their combination (306 mg per day [203-461], p0·0001). Orthostatic complaints were present in two patients (4%) during baseline treatment, five (11%) during addition of sodium restriction, five (11%) during hydrochlorothiazide treatment, and 12 (27%) during combination treatment. No serious adverse events occurred.We conclude that sodium restriction is an effective non-pharmacological intervention to increase RAAS blockade efficacy in type 2 diabetic nephropathy.None.

Combined cardiac and renal dysfunction has gained considerable attention. Hypotheses about its pathogenesis have been formulated, albeit based on a relatively small body of experimental studies, and a clinical classification system has been proposed. Cardiorenal syndrome, as presently defined, comprises a heterogeneous group of acute and chronic clinical conditions, in which the failure of one organ ( heart or kidney) initiates or aggravates failure of the other. This conceptual framework, however, has two major drawbacks: the first is that, despite worldwide interest, universally accepted definitions of cardiorenal syndrome are lacking and characterization of heart and kidney failure is not uniform. This lack of consistency hampers experimental studies on mechanisms of the disease. The second is that, although progress has been made in developing hypotheses for the pathogenesis of cardiorenal syndrome, these initiatives are at an impasse. No hierarchy has been identified in the myriad of haemodynamic and non-haemodynamic factors mediating cardiorenal syndrome. This Review discusses current understanding of cardiorenal syndrome and provides a roadmap for further studies in this field. Ultimately, discussion of the definition and characterization issues and of the lack of organization among pathogenetic factors is hoped to contribute to further advancement of this complex field.

Women with a history of preeclampsia have a 5- to 12-fold increased risk to develop end-stage kidney disease. Previous observations in small cohorts suggest that former preeclamptic (fPE) women have subtle abnormalities in renal hemodynamics and renal function, which might predispose them to renal failure in later life. In this study, we analyzed renal function in a cross-sectional cohort consisting of former early-onset preeclamptic (fPE, n=339) and former healthy pregnant women (fHP, n=332), overall with a mean age of 39 years at 10 years postpartum. Estimated glomerular filtration rate (eGFR), assessed by the modification of diet in renal disease (MDRD) and chronic kidney disease–epidemiology (CKD-epi) equations, and urinary protein:creatinine ratios were assessed 10 years postpartum. Median MDRD and CKD-epi eGFR did not significantly differ between fHP and fPE groups, whereas a comparison of distribution of eGFR revealed a shift toward a high-normal MDRD eGFR in the fPE group (χ 2 , P =0.02) with the same trend for CKD-epi eGFR (χ 2 , P =0.18). The odds ratio for fPE women having MDRD eGFR >110 mL/min per 1.73 m 2 was 1.6 (1.1–2.4). In addition, the median urinary protein:creatinine ratio was slightly higher in fPE (8.5 versus 7.1 mg/mmol; P Clinical Trial Registration— URL: http://www.trialregister.nl . Unique identifier: NTR2668.

Weight excess and/or central body fat distribution are associated with increased long-term renal risk, not only in subjects with renal disease or renal transplant recipients, but also in the general population. As the prevalence of weight excess is rising worldwide, this may become a main renal risk factor on a population basis, even more so because the risk extends to the overweight range. Understanding the mechanisms of this detrimental effect of weight excess on the kidneys is needed in order to design preventive treatment strategies. The increased risk associated with weight excess is partly attributed to associated comorbid conditions, such as hypertension, dyslipidaemia, insulin resistance and diabetes; however, current evidence supports a direct pathogenetic role for renal haemodynamics as well. Weight excess is associated with an altered renal haemodynamic profile, i.e. an increased glomerular filtration rate relative to effective renal plasma flow, resulting in an increased filtration fraction (FF). This renal haemodynamic profile is considered to reflect glomerular hyperfiltration and glomerular hypertension, resulting from a dysbalance between afferent and efferent arterial vasomotor balance. This unfavorable renal haemodynamic profile was found to be associated with renal outcome in experimental models and in human renal transplant recipients, and is associated with a blunted sodium excretion, and reversible by weight loss, renin-angiotensin-aldosterone system blockade or by dietary sodium restriction. More recent evidence showed that a central body fat distribution is also associated with an increased FF, even independent of overall weight excess. In this review, we provide an overview on current literature on the impact of weight excess and central body fat distribution on the renal haemodynamic profile in humans, and its possible role in progressive renal damage.

Metabolic syndrome is associated with an increased risk of colorectal cancer. This study investigated the impact of hypertension, a component of metabolic syndrome, on azoxymethane (AOM)-induced colorectal carcinogenesis using SHRSP/Izm (SHRSP) non-diabetic/hypertensive rats and SHRSP.Z-Leprfa/IzmDmcr (SHRSP-ZF) diabetic/hypertensive rats. Male 6-week-old SHRSP, SHRSP-ZF, and control non-diabetic/normotensive Wister Kyoto/Izm (WKY) rats were given 2 weekly intraperitoneal injections of AOM (20 mg/kg body weight). Two weeks after the last injection of AOM, the SHRSP and SHRSP-ZF rats became hypertensive compared to the control WKY rats. Serum levels of angiotensin-II, the active product of the renin-angiotensin system, were elevated in both SHRSP and SHRSP-ZF rats, but only the SHRSP-ZF rats developed insulin resistance, dyslipidemia, and hyperleptinemia and exhibited an increase in adipose tissue. The development of AOM-induced colonic preneoplastic lesions and aberrant crypts foci, was significantly accelerated in both SHRSP and SHRSP-ZF hypertensive rats, compared to WKY normotensive rats. Furthermore, induction of oxidative stress and exacerbation of inflammation were observed in the colonic mucosa and systemically in SHRSP and SHRSP-ZF rats. Our findings suggest that hypertension plays a role in the early stage of colorectal carcinogenesis by inducing oxidative stress and chronic inflammation, which might be associated with activation of the renin-angiotensin system.

Transactivation of epidermal growth factor receptor (EGFR) signaling by G protein-coupled receptors has been implicated in several cardiovascular (CV) conditions, including hypertension, heart failure, and cardiac and vascular hypertrophy. However, the therapeutic potential of EGFR inhibition in these conditions is currently unknown. The main objective of the present study was to investigate cardiac, vascular, and renal effects of EGFR inhibition by 4-[4-[[(1R)-1-phenylethyl]amino]-7H-pyrrolo[2,3-d]pyrimidin-6-yl]phenol (PKI-166) in the hypertensive chronic kidney disease model. Rats underwent 5/6 nephrectomy (5/6Nx) and were treated with PKI-166, lisinopril or vehicle from week 6 after disease induction until week 12. Sham animals received either PKI-166 or vehicle. Treatment with PKI-166 did not affect the development of the characteristic renal features in 5/6Nx, including proteinuria, diminished creatinine clearance, and increased glomerulosclerosis, whereas these were attenuated by lisinopril. Despite absence of effects on progressive renal damage, PKI-166 attenuated the progression of hypertension and maintained cardiac function (left ventricle end-diastolic pressure) to a similar extent as lisinopril. Also, PKI-166 attenuated the increase in phosphorylated EGFR in the heart as induced by 5/6Nx. Moreover, PKI-166 and lisinopril restored the impaired contraction of isolated thoracic aortic rings to phenylephrine and angiotensin II and impaired myogenic constriction of small mesenteric arteries in 5/6Nx rats. Blockade of the EGFR displays a CV benefit independent of limiting the progression of renal injury. Our findings extend the evidence on EGFR signaling as a target in CV disorders.

Objective To review the inter-relationships between calcium, phosphorus, parathyroid hormone (PTH), parent and activated vitamin D metabolites (vitamin D, 25(OH)-vitamin D, 1,25(OH)2-vitamin D, 24,25(OH)2-vitamin D), and fibroblast growth factor-23 (FGF-23) during chronic kidney disease (CKD) in dogs and cats. Data Sources Human and veterinary literature. Human Data Synthesis Beneficial effects of calcitriol treatment during CKD have traditionally been attributed to regulation of PTH but new perspectives emphasize direct renoprotective actions independent of PTH and calcium. It is now apparent that calcitriol exerts an important effect on renal tubular reclamation of filtered 25(OH)-vitamin D, which may be important in maintaining adequate circulating 25(OH)-vitamin D. This in turn may be vital for important pleiotropic actions in peripheral tissues through autocrine/paracrine mechanisms that impact the health of those local tissues. Veterinary Data Synthesis Limited information is available reporting the benefit of calcitriol treatment in dogs and cats with CKD. Conclusions A survival benefit has been shown for dogs with CKD treated with calcitriol compared to placebo. The concentrations of circulating 25(OH)-vitamin D have recently been shown to be low in people and dogs with CKD and are related to survival in people with CKD. Combination therapy for people with CKD using both parental and activated vitamin D compounds is common in human nephrology and there is a developing emphasis using combination treatment with activated vitamin D and renin-angiotensin-aldosterone-system (RAAS) inhibitors.

Background: Preeclampsia is a common and heterogeneous vascular syndrome of pregnancy. Its genetic risk profile is yet unknown and may vary between individuals and populations. The rs4606 3'UTR polymorphism of the Regulator of G-protein signaling 2 gene (RGS2) in the mother has been implicated in preeclampsia as well as in the development of chronic hypertension after preeclampsia. The RGS2 protein acts as an inhibitor of physiological vasoconstrictive pathways, and a low RGS2 level is associated with hypertension and obesity, two conditions that predispose to preeclampsia. We genotyped the rs4606 polymorphism in 1339 preeclamptic patients and in 697 controls from the Finnish Genetics of Preeclampsia Consortium (FINNPEC) cohort to study the association of the variant with preeclampsia. Results: No association between rs4606 and preeclampsia was detected in the analysis including all women. However, the polymorphism was associated with preeclampsia in a subgroup of overweight women (body mass index >= 25 kg/m(2), and

Angiotensin‐II (Ang‐II) infusion is associated with the development of interstitial fibrosis in both heart and kidney as a result of chemokine‐dependent uptake of monocytes and subsequent development of myeloid fibroblasts. This study emphasizes on the synergistic role of tumor necrosis factor (TNF) on the time course of Ang‐II‐induced fibrosis and inflammation in heart and kidney. In wild‐type (WT) hearts, Ang‐II‐induced fibrosis peaked within 1 week of infusion and remained stable over a 6‐week period, while the myeloid fibroblasts disappeared; TNF receptor‐1‐knockout (TNFR1‐KO) hearts did not develop a myeloid response or cardiac fibrosis during this time. WT hearts developed more accelerated cardiac hypertrophy and remodeling than TNFR1‐KO. In the kidney, 1‐week Ang‐II infusion did not evoke a fibrotic response; however, after 6 weeks, WT kidneys displayed modest but significant tubulointerstitial collagen deposition associated with the appearance of myeloid cells and profibrotic gene activation. Renal fibrosis was not seen in Ang‐II‐infused TNFR1‐KO. By contrast, while hypertension increased and cardiac function decreased more slowly in TNFR1‐KO than WT, they were equivalently abnormal at 6 weeks. Similarly, serum markers for renal dysfunction were not different after 6 weeks. In conclusion, Ang‐II infusion initiated fibroinflammatory responses with different time courses in heart and kidney, both requiring TNFR1 signaling, and both associated with monocyte‐derived myeloid fibroblasts. TNFR1 deletion obviated the fibroinflammatory effects of Ang‐II, but did not alter changes in blood pressure and cardiorenal function after 6 weeks. Thus, the synergy of TNF with Ang‐II targets the fibroinflammatory component of Ang‐II signaling.

Renal artery stenosis is frequently associated with resistant hypertension, which is defined as failure to normalize blood pressure (BP) even when combined drugs are used. Inhibition of PDE5 by sildenafil has been shown to increase endothelial function and decrease blood pressure in experimental models. However, no available study evaluated the baroreflex sensitivity nor autonomic balance in renovascular hypertensive rats treated with sildenafil. In a translational medicine perspective, our hypothesis is that sildenafil could improve autonomic imbalance and baroreflex sensitivity, contributing to lower blood pressure. Renovascular hypertensive 2-kidney-1-clip (2K1C) and sham rats were treated with sildenafil (45 mg/Kg/day) during 7 days. At the end of treatment, BP and heart rate (HR) were recorded in conscious rats after a 24-hour-recovery period. Spontaneous and drug-induced baroreflex sensitivity and autonomic tone were evaluated; in addition, lipid peroxidation was measured in plasma samples. Treatment was efficient in increasing both spontaneous and induced baroreflex sensitivity in treated hypertensive animals. Inhibition of PDE5 was also capable of ameliorating autonomic imbalance in 2K1C rats and decreasing systemic oxidative stress. Taken together, these beneficial effects resulted in significant reductions in BP without affecting HR. We suggest that sildenafil could be considered as a promising alternative to treat resistant hypertension.

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) OBJECTIVES: Considering that changes in the maternal environment may result in changes in progeny, the aim of this study was to investigate the influence of sleep restriction during the last week of pregnancy on renal function and autonomic responses in male descendants at an adult age. METHODS: After confirmation of pregnancy, female Wistar rats were randomly assigned to either a control or a sleep restriction group. The sleep-restricted rats were subjected to sleep restriction using the multiple platforms method for over 20 hours per day between the 14 th and 20 th day of pregnancy. After delivery, the litters were limited to 6 offspring that were designated as offspring from control and offspring from sleep-restricted mothers. Indirect measurements of systolic blood pressure (BPi), renal plasma flow, glomerular filtration rate, glomerular area and number of glomeruli per field were evaluated at three months of age. Direct measurements of cardiovascular function (heart rate and mean arterial pressure), cardiac sympathetic tone, cardiac parasympathetic tone, and baroreflex sensitivity were evaluated at four months of age. RESULTS: The sleep-restricted offspring presented increases in BPi, glomerular filtration rate and glomerular area compared with the control offspring. The sleep-restricted offspring also showed higher basal heart rate, increased mean arterial pressure, increased sympathetic cardiac tone, decreased parasympathetic cardiac tone and reduced baroreflex sensitivity. CONCLUSIONS: Our data suggest that reductions in sleep during the last week of pregnancy lead to alterations in cardiovascular autonomic regulation and renal morpho-functional changes in offspring, triggering increases in blood pressure. Escola Paulista de Medicina - UNIFESP, Departamento de Fisiologia, São Paulo/SP, Brazil Escola Paulista de Medicina - UNIFESP, Departamento de Psicobiologia, São Paulo/SP, Brazil Centro Universitário São Camilo, São Paulo/SP, Brazil Escola Paulista de Medicina - UNIFESP, Departamento de Fisiologia, São Paulo/SP, Brazil Escola Paulista de Medicina - UNIFESP, Departamento de Psicobiologia, São Paulo/SP, Brazil FAPESP: 2013-23622-7 Web of Science

Vavrinec P, Henning RH, Goris M, Vavrincova-Yaghi D, Buikema H, van Dokkum RP. Vascular smooth muscle function of renal glomerular and interlobar arteries predicts renal damage in rats. Am J Physiol Renal Physiol 303: F1187-F1195, 2012. First published July 11, 2012; doi:10.1152/ajprenal.00653.2011.-Previously, it was shown that individuals with good baseline (a priori) endothelial function in isolated (in vitro) renal arteries developed less renal damage after 5/6 nephrectomy (5/6Nx; Gschwend S, Buikema H, Navis G, Henning RH, de Zeeuw D, van Dokkum RP. J Am Soc Nephrol 13: 2909-2915, 2002). In this study, we investigated whether preexisting glomerular vascular integrity predicts subsequent renal damage after 5/6Nx, using in vivo intravital microscopy and in vitro myogenic constriction of small renal arteries. Moreover, we aimed to elucidate the role of renal ANG II type 1 receptor (AT1R) expression in this model. Anesthetized rats underwent intravital microscopy to visualize constriction to ANG II of glomerular afferent and efferent arterioles, with continuous measurement of blood pressure, heart rate, and renal blood flow. Thereafter, 5/6Nx was performed, interlobar arteries were isolated from the extirpated kidney, and myogenic constriction was assessed in a perfused vessel setup. Blood pressure and proteinuria were assessed weekly for 12 wk, and focal glomerulosclerosis (FGS) was determined at the end of study. Relative expression AT1R in the kidney cortex obtained at 5/6Nx was determined by PCR. Infusion of ANG II induced significant constriction of both afferent and efferent glomerular arterioles, which strongly positively correlated with proteinuria and FGS at 12 wk after 5/6Nx. Furthermore, in vitro measured myogenic constriction of small renal arteries negatively correlated with proteinuria 12 wk after 5/6Nx. Moreover, in vivo vascular reactivity negatively correlated with in vitro reactivity. Additionally, relative expression of AT1R positively correlated with responses of glomerular arterioles and with markers of renal damage. Both in vivo afferent and efferent responses to ANG II and in vitro myogenic constriction of small renal arteries in the healthy rat predict the severity of renal damage induced by 5/6Nx. This vascular responsiveness is highly dependent on AT1R expression. Intraorgan vascular integrity may provide a useful tool to guide the prevention and treatment of renal end-organ damage.

Aims Hypertensive patients develop cardiac hypertrophy and fibrosis with increased stiffness, contractile deficit and altered perfusion. Angiotensin II (AngII) is an important factor in the promotion of this pathology. The effects of AngII are partly mediated by endothelin-1 (ET-1) and transforming growth factor-β. The exact feature of these pathways and the intercellular communications involved remain unclear. In this study, we explored the role of endothelial cell-derived ET-1 in the development of AngII-induced cardiac fibrosis and hypertrophy. Main methods We used mice with vascular endothelial cell specific ET-1 deficiency (VEETKO) and their wild type littermates (WT). Mice were infused for one week with AngII (3.2 mg/kg/day, n = 12) or vehicle (0.15 mol/L NaCl and 1 mmol/L acetic acid, n = 5), using subcutaneous mini-pumps. Hearts were stained with hematoxylin–eosin and masson's trichrome for histology. Cardiac gene expression and protein abundance were measured by Northern Blot, real time PCR and Western Blot. Key findings AngII-induced cardiac hypertrophy, interstitial and perivascular fibrosis were less pronounced in VEETKO mice compared to WT. Blood pressure increased similarly in both genotypes. Expression of connective tissue growth factor, tumor growth factor-β, collagen I and III in response to AngII required endothelial ET-1. Endothelial ET-1 was also necessary to the elevation in protein kinase C δ abundance and ERK1/2 activation. AngII-induced elevation in PKCe abundance was however ET-1 independent. Significance This study underscores the significance of ET-1 from the vasculature in the process of AngII-induced cardiac hypertrophy and fibrosis, independently from blood pressure. Endothelial ET-1 represents therefore a possible pharmacological target.

BACKGROUND: A simulated upper gastrointestinal (UGI) bleed in cirrhotic patients has been shown to induce hyperammonaemia. The kidney was the site of this exaggerated ammoniagenesis with alanine as substrate. Administration of alanine to decompensated cirrhotic patients did not change hepatic gluconeogenesis, but resulted in increased ammoniagenesis. We hypothesise that reduced hepatic glycogen stores result in hyperglucagonaemia which may drive increased renal gluconeogenesis and therefore alanine uptake and renal ammoniagenesis. AIM: To determine whether an overnight glucose infusion lowers renal ammoniagenesis by reducing hyperglucagonaemia and renal ammoniagenesis. METHODS: Patients with decompensated cirrhosis were studied in a cross-over design. An UGI bleed was simulated via intragastric administration of an amino acids mixture mimicking the haemoglobin molecule after a 12-h overnight fast (F-group) or after a 12-h treatment with 20% glucose solution (G-group). RESULTS: Before the simulated bleed the glucagon levels were 21 (15-31) pmol/L in the F-group and 15 (9-21) pmol/L in the G-group (P < 0.01). After the simulated bleed, arterial ammonia levels increased in both groups [F-group: 73-118 mumol/L (P = 0.01); G-group 64-87 mumol/L (P = 0.01)]. The enhancement of hyperammonaemia was significantly higher in the F-group (45 [19-71] mumol/L) compared with the G-group (23 [13-39] mumol/L) (P = 0.01). The difference in renal ammoniagenesis during the simulated bleed in the F-group was 399 (260-655) nmol/kg/bwt/min and was significantly higher than in the G-group 313 (1-498) nmol/kg/bwt/min (P = 0.05). CONCLUSIONS: Overnight glucose infusion results in reduced renal ammoniagenesis and attenuates ammonia levels. These observations have implications for the development of nutritional strategies in hyperammonaemic patients.

The incidence of heart failure and renal failure is increasing and is associated with poor prognosis. Moreover, these conditions do often coexist and this coexistence results in worsened outcome. Various mechanisms have been proposed as an explanation of this interrelation, including changes in hemodynamics, endothelial dysfunction, inflammation, activation of renin-angiotensin-aldosterone system, and/or sympathetic nervous system. However, the exact mechanisms initializing and maintaining this interaction are still unknown. In many experimental studies on cardiac or renal dysfunction, the function of the other organ was either not addressed or the authors failed to show any decline in its function despite histological changes. There are few studies in which the dysfunction of both heart and kidney function has been described. In this review, we discuss animal models of combined cardiorenal dysfunction. We show that translation of the results from animal studies is limited, and there is a need for new and better models of the cardiorenal interaction to improve our understanding of this syndrome. Finally, we propose several requirements that a new animal model should meet to serve as a tool for studies on the cardiorenal syndrome.

Background The renin-angiotensin-aldosterone system (RAAS) plays a key role in the progression of heart failure (HF) and concomitant kidney dysfunction. Despite the use of RAAS blockade, sustained activation of RAAS has been suggested to link with adverse outcome. We aimed to investigate the prognostic value of active plasma renin concentration (APRC) and prorenin in patients with HF treated with RAAS-blocking agents and its relationship with kidney function parameters.Methods One hundred clinically stable patients with HF, treated with RAAS-blocking agents, were studied. Renal function parameters including effective renal plasma flow and glomerular filtration rate were measured invasively. The combined end point consisted of all-cause mortality, heart transplantation, and admission to hospital for HF.Results Mean age was 58 +/- 12 years, and 76% were men. Mean left ventricular ejection fraction was 28 +/- 9, and median APRC levels were 24.3 ng/mL per hour. Active plasma renin concentration was most strongly associated with mean arterial pressure (r = 0.60, P Conclusions Our data indicate that APRC is a strong prognostic factor in patients with HF in the presence of RAAS inhibition, especially in patients with kidney dysfunction. (Am Heart J 2011;162:487-93.)

Background and objectives Connective Tissue Growth Factor (CTGF/CCN-2) is a key player in fibrosis. Plasma CTGF levels predict end-stage renal disease and mortality in diabetic chronic kidney disease (CKD), supporting roles in intra- and extrarenal fibrosis. Few data are available on CTGF in nondiabetic CKD. We investigated CTGF levels and effects of antiproteinuric interventions in nondiabetic proteinuric CKD.Design, setting, participants, & measurements In a crossover randomized controlled trial, 33 nondiabetic CKD patients (3.2 [2.5 to 4.0] g/24 h proteinuria) were treated during 6-week periods with placebo, ARB (100 mg/d losartan), and ARB plus diuretics (100 mg/d losartan plus 25 mg/d hydrochlorothiazide) combined with consecutively regular and low sodium diets (193 +/- 62 versus 93 +/- 52 mmol Na+/d).Results CTGF was elevated in plasma (464 [387 to 556] pmol/L) and urine (205 [135 to 311] pmol/24 h) of patients compared with healthy controls (n = 21; 96 [86 to 108] pmol/L and 73 [55 to 98] pmol/24 h). Urinary CTGF was lowered by antiproteinuric intervention, in proportion to the reduction of proteinuria, with normalization during triple therapy (CTGF 99 [67 to 146] in CKD versus 73 [55 to 98] pmol/24 h in controls). In contrast, plasma CTGF was not affected.Conclusions Urinary and plasma CTGF are elevated in nondiabetic CKD. Only urinary CTGF is normalized by antiproteinuric intervention, consistent with amelioration of tubular dysfunction. The lack of effect on plasma CTGF suggests that its driving force might be independent of proteinuria and that short-term antiproteinuric interventions are not sufficient to correct the systemic profibrotic state in CKD. Clin J Am Soc Nephrol 6: 1845-1850, 2011. doi: 10.2215/CJN.08190910

Background— Adipose tissue (AT) dysfunction in obesity contributes to chronic, low-grade inflammation that predisposes to type 2 diabetes mellitus and cardiovascular disease. Recent in vitro studies suggest that AT hypoxia may induce inflammation. We hypothesized that adipose tissue blood flow (ATBF) regulates AT oxygen partial pressure (AT P o 2 ), thereby affecting AT inflammation and insulin sensitivity. Methods and Results— We developed an optochemical measurement system for continuous monitoring of AT P o 2 using microdialysis. The effect of alterations in ATBF on AT P o 2 was investigated in lean and obese subjects with both pharmacological and physiological approaches to manipulate ATBF. Local administration of angiotensin II (vasoconstrictor) in abdominal subcutaneous AT decreased ATBF and AT P o 2 , whereas infusion of isoprenaline (vasodilator) evoked opposite effects. Ingestion of a glucose drink increased ATBF and AT P o 2 in lean subjects, but these responses were blunted in obese individuals. However, AT P o 2 was higher (hyperoxia) in obese subjects despite lower ATBF, which appears to be explained by lower AT oxygen consumption. This was accompanied by insulin resistance, lower AT capillarization, lower AT expression of genes encoding proteins involved in mitochondrial biogenesis and function, and higher AT gene expression of macrophage infiltration and inflammatory markers. Conclusions— Our findings establish ATBF as an important regulator of AT P o 2 . Nevertheless, obese individuals exhibit AT hyperoxia despite lower ATBF, which seems to be explained by lower AT oxygen consumption. This is accompanied by insulin resistance, impaired AT capillarization, and higher AT gene expression of inflammatory cell markers. Clinical Trial Registration— URL: http://www.trialregister.nl . Unique identifier: NTR2451.

Introduction: Transforming growth factor-β (TGF-β) and connective tissue growth factor (CTGF) are often described as the initial pro-fibrotic mediators upregulated early in fibrosis models dependent on angiotensin II (Ang-II). In the present study, we explore the mechanistic link between TGF-β and CTGF expression by using a novel TGF-β trap. Materials and methods: NIH/3T3 fibroblasts were subjected to TGF-β with or without TGF-β trap or 1D11 antibody, CTGF or CTGF plus TGF-β for six or 24 hours, and then used for quantitative real-time polymerase chain reaction (qRT-PCR) or immunocytochemistry. Male C57BL/6 mice were infused with Ang-II and randomly assigned TGF-β trap for six or 24 hours. Hearts were harvested for histological analyses, qRT-PCR and western blotting. Results: Exogenous TGF-β-induced fibroblasts resulted in significant upregulation of CTGF, TGF-β and type I collagen transcript levels in vitro. Additionally, TGF-β promoted the differentiation of fibroblasts into α-SMA+ myofibroblasts. CTGF expression was reduced by the addition of TGF-β trap or neutralizing antibody, confirming that its expression is dependent on TGF-β signaling. In contrast, exogenous CTGF did not appear to have an effect on fibroblast production of pro-fibrotic transcripts or fibroblast differentiation. Ang-II infusion in vivo led to a significant increase in TGF-β and CTGF mRNA expression at six and 24 hours with corresponding changes in Smad2 phosphorylation (pSmad2), indicative of increased TGF-β signaling. Ang-II animals that received the TGF-β trap demonstrated reduced CTGF mRNA levels and pSmad2 at six hours, suggesting that early CTGF expression is dependent on TGF-β signaling. Conclusions: We demonstrated that CTGF expression is dependent on TGF-β signaling both in vitro and in vivo in a model of myocardial fibrosis. This also suggests that early myocardial CTGF mRNA expression (six hours) after Ang-II exposure is likely dependent on latent TGF-β activation via the canonical Smad-dependent pathway in resident cardiac cells.

Recognition of the link between non-alcoholic fatty liver disease (NAFLD) and cardiovascular disease (CVD) has boosted research in this area. The main objective of this paper is to review the literature on NAFLD in the context of CVD, focussing on underlying mechanisms and treatment. Besides excessive fatty acid influx, etiologic factors may include components of the metabolic syndrome, cytokines and mitochondrial dysfunction. NAFLD is associated with both hepatic and systemic insulin resistance. In the case of NAFLD, the liver overproduces several atherogenic factors, notably inflammatory cytokines, glucose, lipoproteins and coagulation factors, and factors increasing blood pressure. Intervention studies on diet and laparoscopic surgery revealed improvements of hepatic fat content and CVD risk profile. Pharmacological approaches with potential benefit have been developed as well, but effects are often confounded by weight change. NAFLD is associated with an increased CVD risk profile (and hepatic risk). In order to improve CVD risk profile, prevention and treatment of NAFLD seem advisable. However, well-designed intervention studies, randomized clinical trials and long-term follow-up studies are scarce.

Background In patients with chronic heart failure, erythropoietin (Epo) levels are increased and related to a poor prognosis. Furthermore, Epo levels in these patients show a weak correlation with hemoglobin levels.Methods This is a retrospective analysis of a subgroup of the OPTIMAAL (Optimal Trial in Myocardial Infarction with the Angiotensin II Antagonist Losartan) trial in which serum Epo levels were measured at baseline, at 1 month, and at 1 and 2 years in 224 patients with an acute myocardial infarction complicated by signs or symptoms of heart failure. We investigated the determinants and the prognostic role of elevated Epo levels in these patients, and we studied the change in Epo levels by either captopril or losartan.Results The correlation between Epo and hemoglobin at baseline (r* = 0.348, P Conclusion In this substudy of the OPTIMAAL trial, the correlation between Epo and hemoglobin disappeared in early post-acute myocardial infarction heart failure patients. Furthermore, elevated Epo levels at baseline predicted increased mortality. (Am Heart J 2009;157:91-6.)