Dimethylarginine dimethylaminohydrolase 1 protects PM2.5 exposure-induced lung injury in mice by repressing inflammation and oxidative stress.

BACKGROUND: Airborne fine particulate matter with aerodynamic diameter ≤ 2.5 µm (PM2.5) pollution is associated with the prevalence of respiratory diseases, including asthma, bronchitis and chronic obstructive pulmonary disease. In patients with those diseases, circulating asymmetric dimethylarginine (ADMA) levels are increased, which contributes to airway nitric oxide deficiency, oxidative stress and inflammation. Overexpression of dimethylarginine dimethylaminohydrolase 1 (DDAH1), an enzyme degrading ADMA, exerts protective effects in animal models. However, the impact of DDAH1/ADMA on PM2.5-induced lung injury has not been investigated. METHODS: Ddah1-/- and DDAH1-transgenic mice, as well as their respective wild-type (WT) littermates, were exposed to either filtered air or airborne PM2.5 (mean daily concentration ~ 50 µg/m3) for 6 months through a whole-body exposure system. Mice were also acutely exposed to 10 mg/kg PM2.5 and/or exogenous ADMA (2 mg/kg) via intratracheal instillation every other day for 2 weeks. Inflammatory response, oxidative stress and related gene expressions in the lungs were examined. In addition, RAW264.7 cells were exposed to PM2.5 and/or ADMA and the changes in intracellular oxidative stress and inflammatory response were determined. RESULTS: Ddah1-/- mice developed more severe lung injury than WT mice after long-term PM2.5 exposure, which was associated with greater induction of pulmonary oxidative stress and inflammation. In the lungs of PM2.5-exposed mice, Ddah1 deficiency increased protein expression of p-p65, iNOS and Bax, and decreased protein expression of Bcl-2, SOD1 and peroxiredoxin 4. Conversely, DDAH1 overexpression significantly alleviated lung injury, attenuated pulmonary oxidative stress and inflammation, and exerted opposite effects on those proteins in PM2.5-exposed mice. In addition, exogenous ADMA administration could mimic the effect of Ddah1 deficiency on PM2.5-induced lung injury, oxidative stress and inflammation. In PM2.5-exposed macrophages, ADMA aggravated the inflammatory response and oxidative stress in an iNOS-dependent manner. CONCLUSION: Our data revealed that DDAH1 has a marked protective effect on long-term PM2.5 exposure-induced lung injury.

New Genetic Biomarkers of the Overlap Syndrome Tension-Type Headache and Arterial Hypertension.

BACKGROUND: Nitric oxide (NO) is an important autocrine and paracrine signaling molecule that plays a crucial role in cardiovascular physiology and pathology regulation. NO is an important molecule involved in regulation of cerebral and extra cerebral cranial blood flow and arterial diameters. Reduced bioavailability of NO in the endothelium is an important precursor for impaired vasodilation and arterial hypertension (AH). Furthermore, NO is involved in nociceptive processing. A NO-induced biphasic response with immediate and a delayed headache is typical for chronic tension-type headaches (TTH) in humans. The aim was to study the association of allelic variants and genotypes of the single nucleotide variant (SNV) rs3782218 of the NOS1 gene with the TTH and AH overlap syndrome development in middle age adults. MATERIALS AND METHODS: We observed 91 Caucasian participants who resided in Krasnoyarsk city: group 1 (TTH and AH overlap syndrome)-30 patients; group 2 (AH without headache)-30 patients; group 3 (control)-31 healthy volunteers. The diagnosis of AH was based on criteria of the European Society of Cardiology and the European Society of Hypertension (2018) и criteria of the Russian Society of Cardiology (2020). Diagnosis of TTH was based on criteria of the International Classification of Headache Disorders (2018). Real-time polymerase chain reaction was used for the determination of allelic variants and genotypes of the SNV rs3782218 of the NOS1 gene in all groups of participants. RESULTS: The frequency of the minor allele T of rs3782218 was statistically significantly higher by 16.7 times in group 1 (TTH and AH) compared to group 3 (control): 26.7% versus 1.6%, respectively (p-value = 0.000065) and 3.2 times higher in group 1 (TTH and AH) compared to group 2 (AH without headache): 26.7% versus 8.3%, respectively (p-value = 0.008). The frequency of the heterozygous (CT) genotype was statistically significantly higher in group 1 (TTH and AH) compared to group 3 (control): 40.0% versus 3.2% (p-value = 0.000454) and in group 1 (TTH and AH) compared to group 2 (AH without headache): 40.0% versus 16.7% (p-value = 0.045). The minor allele T was statistically significantly associated with a high risk of developing the TTH and AH overlap syndrome compared with the controls (odds ratio (OR) = 22.2 (95% confidential interval (CI): 2.8-173.5)) and compared with AH without headache (OR = 4.0 (95% CI: 1.4-11.8)). Although the frequency of the minor allele T was 5.2 times higher in group 2 (AH without headache) compared with group 3 (control), there were not statistically significantly differences (p-value = 0.086). CONCLUSION: Thus, the minor allele T of rs3782218 of the NOS1 gene is an important genetic biomarker for a high risk of developing the TTH and AH overlap syndrome in hypertensive patients.

Nitric Oxide Signaling and Its Association with Ubiquitin-Mediated Proteasomal Degradation in Plants.

Nitric oxide (NO) is a versatile signaling molecule with diverse roles in plant biology. The NO-mediated signaling mechanism includes post-translational modifications (PTMs) of target proteins. There exists a close link between NO-mediated PTMs and the proteasomal degradation of proteins via ubiquitylation. In some cases, ubiquitin-mediated proteasomal degradation of target proteins is followed by an NO-mediated post-translational modification on them, while in other cases NO-mediated PTMs can regulate the ubiquitylation of the components of ubiquitin-mediated proteasomal machinery for promoting their activity. Another pathway that links NO signaling with the ubiquitin-mediated degradation of proteins is the N-degron pathway. Overall, these mechanisms reflect an important mechanism of NO signal perception and transduction that reflect a close association of NO signaling with proteasomal degradation via ubiquitylation. Therefore, this review provides insight into those pathways that link NO-PTMs with ubiquitylation.

A New Player in Jasmonate-Mediated Stomatal Closure: The Arabidopsis thaliana Copper Amine Oxidase ß.

Plant defence responses to adverse environmental conditions include different stress signalling, allowing plant acclimation and survival. Among these responses one of the most common, immediate, and effective is the modulation of the stomatal aperture, which integrates different transduction pathways involving hydrogen peroxide (H2O2), calcium (Ca2+), nitric oxide (NO), phytohormones and other signalling components. The Arabidopsis thaliana copper amine oxidases ß (AtCuAOß) encodes an apoplastic CuAO expressed in guard cells and root protoxylem tissues which oxidizes polyamines to aminoaldehydes with the production of H2O2 and ammonia. Here, its role in stomatal closure, signalled by the wound-associated phytohormone methyl-jasmonate (MeJA) was explored by pharmacological and genetic approaches. Obtained data show that AtCuAOß tissue-specific expression is induced by MeJA, especially in stomata guard cells. Interestingly, two Atcuaoß T-DNA insertional mutants are unresponsive to this hormone, showing a compromised MeJA-mediated stomatal closure compared to the wild-type (WT) plants. Coherently, Atcuaoß mutants also show compromised H2O2-production in guard cells upon MeJA treatment. Furthermore, the H2O2 scavenger N,N1-dimethylthiourea (DMTU) and the CuAO-specific inhibitor 2-bromoethylamine (2-BrEtA) both reversed the MeJA-induced stomatal closure and the H2O2 production in WT plants. Our data suggest that AtCuAOß is involved in the H2O2 production implicated in MeJA-induced stomatal closure.

Molecular Cloning, Expression and Macrophage Activation of an Immunoregulatory Protein from Cordyceps militaris.

Protein components of C. militaris have been reported to possess various biological activities. In our previous research, a Cordyceps militaris-derived immunoregulatory protein (CMIP) was naturally isolated and showed the activity of inhibiting the metastasis of breast cancer cells. This study aimed to obtain recombinant CMIP (rCMIP) using recombinant expression and elucidate its ability to activate macrophages. Recombinant CMIP showed one band at approximately 15 kDa or 30 kDa, or two bands at 15 kDa and 30 kDa, under different denaturation conditions of electrophoresis. The cell binding assay showed that rCMIP selectively binds to the surface of macrophages. After adhesion, it did not induce the apoptosis of RAW 264.7 cells, but promoted their proliferation. Moreover, rCMIP significantly induced the expression of M1 macrophage polarization-related molecules. The mean fluorescence intensity (MFI) of CD 86 was enhanced by 2.1-fold and 3.2-fold under 0.64 µM and 1.6 µM of rCMIP treatment, respectively. Cytokines typically expressed in M1 macrophages, such as TNF-α, iNOS, IL-6, CCL 4, CCL 5 and CXCL 10, were also considerably induced by rCMIP, while the expression of cytokines in typical M2 macrophages, like Arg-1, CCL17 and CCL22, were not changed or slightly decreased. Under rCMIP treatment, the release of NO was also appreciably induced. In the present study, we reported cloning, expression and functional characterization of rCMIP, which was naturally isolated from the fruiting body of C. militaris in our previous study. The data imply that rCMIP possesses immunomodulatory activity in macrophages.

Study of the Role of the Tyrosine Kinase Receptor MerTK in the Development of Kidney Ischemia-Reperfusion Injury in RCS Rats.

Renal ischaemia reperfusion (I/R) triggers a cascade of events including oxidative stress, apoptotic body and microparticle (MP) formation as well as an acute inflammatory process that may contribute to organ failure. Macrophages are recruited to phagocytose cell debris and MPs. The tyrosine kinase receptor MerTK is a major player in the phagocytosis process. Experimental models of renal I/R events are of major importance for identifying I/R key players and for elaborating novel therapeutical approaches. A major aim of our study was to investigate possible involvement of MerTK in renal I/R. We performed our study on both natural mutant rats for MerTK (referred to as RCS) and on wild type rats referred to as WT. I/R was established by of bilateral clamping of the renal pedicles for 30' followed by three days of reperfusion. Plasma samples were analysed for creatinine, aspartate aminotransferase (ASAT), lactate dehydrogenase (LDH), kidney injury molecule -1 (KIM-1), and neutrophil gelatinase-associated lipocalin (NGAL) levels and for MPs. Kidney tissue damage and CD68-positive cell requirement were analysed by histochemistry. monocyte chemoattractant protein-1 (MCP-1), myeloperoxidase (MPO), inducible nitric oxide synthase (iNOS), and histone 3A (H3A) levels in kidney tissue lysates were analysed by western blotting. The phagocytic activity of blood-isolated monocytes collected from RCS or WT towards annexin-V positive bodies derived from cultured renal cell was assessed by fluorescence-activated single cell sorting (FACS) and confocal microscopy analyses. The renal I/R model for RCS rat described for the first time here paves the way for further investigations of MerTK-dependent events in renal tissue injury and repair mechanisms.

Repurposing Riociguat to Target a Novel Paracrine Nitric Oxide-TRPC6 Pathway to Prevent Podocyte Injury.

Increased expression and activity of the Ca2+ channel transient receptor potential channel 6 (TRPC6) is associated with focal segmental glomerulosclerosis, but therapeutic strategies to target TRPC6 are currently lacking. Nitric oxide (NO) is crucial for normal glomerular function and plays a protective role in preventing glomerular diseases. We investigated if NO prevents podocyte injury by inhibiting injurious TRPC6-mediated signaling in a soluble guanylate cyclase (sGC)-dependent manner and studied the therapeutic potential of the sGC stimulator Riociguat. Experiments were performed using human glomerular endothelial cells and podocytes. Podocyte injury was induced by Adriamycin incubation for 24 h, with or without the NO-donor S-Nitroso-N-acetyl-DL-penicillamine (SNAP), the sGC stimulator Riociguat or the TRPC6 inhibitor Larixyl Acetate (LA). NO and Riociguat stimulated cGMP synthesis in podocytes, decreased Adriamycin-induced TRPC6 expression, inhibited the Adriamycin-induced TRPC6-mediated Ca2+ influx and reduced podocyte injury. The protective effects of Riociguat and NO were blocked when sGC activity was inhibited with 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) or when TRPC6 activity was inhibited by LA. Our data demonstrate a glomerular (e)NOS-NO-sGC-cGMP-TRPC6 pathway that prevents podocyte injury, which can be translated to future clinical use by, e.g., repurposing the market-approved drug Riociguat.

The evolutionary origins of peroxynitrite signalling.

Peroxynitrite is a reactive intermediate formed in vivo through uncatalysed reaction of superoxide and nitric oxide radicals. Despite significant interest in detecting peroxynitrite in vivo and understanding its production, little attention has been given to the evolutionary origins of peroxynitrite signalling. Herein we focus on two enzymes that are key to the biosynthesis of superoxide and nitric oxide, NADPH oxidase 5 (NOX5) and endothelial nitric oxide synthase (eNOS), respectively. Multiple sequence alignments of both enzymes including homologues from all domains of life, coupled with a phylogenetic analysis of NOX5, suggest eNOS and NOX5 are present in animals as the result of horizontal gene transfer from ancestral cyanobacteria to ancestral eukaryotes. Therefore, biochemical studies from other laboratories on a NOX5 homologue in Cylindrospermum stagnale and an eNOS homologue in Synechococcus sp. PCC 7335 are likely to be of relevance to human NOX5 and eNOS and to the production of superoxide, nitric oxide and peroxynitrite in humans.

Hybrids as NO Donors.

Cinnamic acid and its derivatives have been studied for a variety of biological properties, including anti-inflammatory, antioxidant, anticancer, antihypertensive, and antibacterial. Many hybrids of cinnamic derivatives with other bioactive molecules have been synthesized and evaluated as nitric oxide (NO) donors. Since NO plays a significant role in various biological processes, including vasodilation, inflammation, and neurotransmission, NO donor groups are incorporated into the structures of already-known bioactive molecules to enhance their biological properties. In this review, we present cinnamic hybrids with NO-donating ability useful in the treatment of several diseases.

Nitrosative Stress and Human Disease: Therapeutic Potential of Denitrosylation.

Proteins dynamically contribute towards maintaining cellular homeostasis. Posttranslational modification regulates the function of target proteins through their immediate activation, sudden inhibition, or permanent degradation. Among numerous protein modifications, protein nitrosation and its functional relevance have emerged. Nitrosation generally initiates nitric oxide (NO) production in association with NO synthase. NO is conjugated to free thiol in the cysteine side chain (S-nitrosylation) and is propagated via the transnitrosylation mechanism. S-nitrosylation is a signaling pathway frequently involved in physiologic regulation. NO forms peroxynitrite in excessive oxidation conditions and induces tyrosine nitration, which is quite stable and is considered irreversible. Two main reducing systems are attributed to denitrosylation: glutathione and thioredoxin (TRX). Glutathione captures NO from S-nitrosylated protein and forms S-nitrosoglutathione (GSNO). The intracellular reducing system catalyzes GSNO into GSH again. TRX can remove NO-like glutathione and break down the disulfide bridge. Although NO is usually beneficial in the basal context, cumulative stress from chronic inflammation or oxidative insult produces a large amount of NO, which induces atypical protein nitrosation. Herein, we (1) provide a brief introduction to the nitrosation and denitrosylation processes, (2) discuss nitrosation-associated human diseases, and (3) discuss a possible denitrosylation strategy and its therapeutic applications.

Polygalaxanthone III downregulates inflammation in the lipopolysaccharide-stimulated RAW264.7 macrophages: A quantibody array analysis.

Polygala japonica Houtt. (PJ), a member of the Polygala L. family that is suggested to exhibit detoxification properties in traditional Chinese medicine, is often used to treat upper respiratory tract infections. The anti-inflammatory effects of four main components of PJ (POL, PS-XLIX, PS-E, and PS-F) were examined using the LPS(0.3 µg·mL-1)-stimulated RAW264.7 macrophage model. The levels of NO, ROS, and iNOS were examined to analyze the anti-inflammatory activity of POL. Additionally, the levels of extracellular inflammation-related cytokines and chemokines were measured using quantibody array. The KEGG pathway analysis was performed to examine the anti-inflammatory mechanism of POL. The levels of NO in the POL-pretreated group were significantly downregulated when compared with those in the PS-E-pretreated, PS-F-pretreated, and PS-XLIX-pretreated groups. POL significantly inhibited the changes of iNOS, ROS, and inflammatory factors caused by LPS stimulation (p < 0.001). The expression levels of IL21 and GM-CSF were examined using qPCR, while those of JAK-STAT signaling pathway-related proteins in the LPS-stimulated RAW264.7 macrophages were analyzed using western blotting. POL significantly downregulated the expression of IL-21 and GM-CSF. The anti-inflammatory mechanism of POL is mediated through the JAK-STAT pathway. Thus, this study demonstrated that POL is an anti-inflammatory component of PJ and elucidated its mechanism.

Carbon Monoxide and Nitric Oxide as Examples of the Youngest Class of Transmitters.

The year 2021 is the 100th anniversary of the confirmation of the neurotransmission phenomenon by Otto Loewi. Over the course of the hundred years, about 100 neurotransmitters belonging to many chemical groups have been discovered. In order to celebrate the 100th anniversary of the confirmation of neurotransmitters, we present an overview of the first two endogenous gaseous transmitters i.e., nitric oxide, and carbon monoxide, which are often termed as gasotransmitters.

High total cholesterol and triglycerides levels increase arginases metabolism, impairing nitric oxide signaling and worsening fetoplacental endothelial dysfunction in gestational diabetes mellitus pregnancies.

Maternal physiological dyslipidemia (MPD) supports fetal development in human pregnancy. However, some women develop maternal supraphysiological dyslipidemia (MSPD: increased total cholesterol (TC) and triglycerides (TG) levels). MSPH is present in normal and also in gestational diabetes mellitus (GDM) pregnancies. MSPD and GDM associate with fetoplacental endothelial dysfunction, producing alterations in nitric oxide (NO)-L-arginine/arginase metabolism. Nevertheless, the effect of MSPD on GDM, and how this synergy alters fetoplacental endothelial function is unknown. Therefore, the aim of this study was to evaluate in human umbilical vein endothelial cells, the effects of MSPD in GDM and how these pathologies together affect the fetoplacental endothelial function. 123 women at term of pregnancy were classified as MPD (n = 40), MSPD (n = 35), GDM with normal lipids (GDM-MPD, n = 23) and with increased lipids (GDM-MSPD, n = 25). TC ≥291 mg/dL and TG ≥275 mg/dL were considered as MSPD. Endothelial NO synthase (eNOS), human cationic amino acid transporter 1 (hCat1), and arginase II protein abundance and activity, were assayed in umbilical vein endothelial cells. In MSPD and GDM-MSPD, TC and TG increased respect to MPD and GDM-MPD. eNOS activity was reduced in MSPD and GDM-MSPD, but increased in GDM-MPD compared with MPD. However, decreased tetrahydrobiopterin levels were observed in all groups compared with MPD. Increased hCat1 protein and L-arginine transport were observed in both GDM groups compared with MPD. However, the transport was higher in GDM-MSPD compared to GDM-MPD. Higher Arginase II protein and activity were observed in GDM-MSPD compared with MPD. Thus, MSPD in GDM pregnancies alters fetal endothelial function associated with NO metabolism.

NAMPT single-nucleotide polymorphism rs1319501 and visfatin/NAMPT affect nitric oxide formation, sFlt-1 and antihypertensive therapy response in preeclampsia.

Aim: We examined the relationships between visfatin/NAMPT and nitrite concentrations (a marker of nitric oxide [NO] formation) or sFlt-1 levels in 205 patients with preeclampsia (PE) responsive or nonresponsive to antihypertensive therapy, and whether NAMPT SNPs rs1319501 and rs3801266 affect nitrite concentrations in PE and 206 healthy pregnant women. Patients & methods: Circulating visfatin/NAMPT and sFlt-1 levels were measured by ELISA, and nitrite concentrations by using an ozone-based chemiluminescence assay. Results: In nonresponsive PE patients, visfatin/NAMPT levels were inversely related to nitrite concentrations and positively related to sFlt-1 levels. NAMPT SNP rs1319501 affected nitrite concentrations in nonresponsive PE patients and was tightly linked with NAMPT functional SNPs in Europeans. Conclusion:NAMPT SNP rs1319501 and visfatin/NAMPT affect NO formation, sFlt-1 levels and antihypertensive therapy response in PE.

Ligustilide inhibited Angiotensin II induced A7r5 cell autophagy via Akt/mTOR signaling pathway.

Autophagy is essential to vessel homeostasis and function in the cardiovascular system. Ligustilide (LIG) is one of the main active ingredients extracted from traditional Chinese medicines, such as Ligusticum chuanxiong, Angelica, and other umbelliferous plants, and reported to have cardiovascular protective effects. In this study, we explore the effects and the potential mechanism of ligustilide on the Ang II-induced autophagy in A7r5 cells. Our results showed that ligustilide inhibited the Ang II-induced autophagy in A7r5 cells and down regulated the expression of autophagy-related proteins LC3, ULK1, and Beclin-1. Ligustilide exerted a protective effect on the reduction of the concentrations of reactive oxygen species and Ca2+ and upregulated the nitric oxide concentration in A7r5 cells with Ang II-induced autophagy. Additionally, the analyses of network pharmacological targets and potential signal pathways indicated that the target of ligustilide to regulate autophagy was related to the Akt/mTOR signaling pathway. Furthermore, ligustilide could upregulate the expression of p-Akt and p-mTOR and inhibit the expression of LC3II in A7r5 cells with Ang II-induced autophagy. These findings showed that ligustilide inhibited the autophagic flux in A7r5 cells induced by Ang II via the activation of the Akt/mTOR signaling pathway.

Neurodegeneration: Impact of S-nitrosylated Parkin, DJ-1 and PINK1 on the pathogenesis of Parkinson's disease.

Parkinson's disease (PD) is one of the fastest-growing neurodegenerative disorders of increasing global prevalence. It represents the second most common movement disorder after tremor and the second most common neurodegenerative disorder after Alzheimer's disease. The incidence rate of idiopathic PD increases steadily with age, however, some variants of autosomal recessive inheritance are present with an early age-at-onset (ARPD). Approximately 50 percent of ARPD cases have been linked to bi-allelic mutations in genes encoding Parkin, DJ-1, and PINK1. Each protein has been implicated in maintaining proper mitochondrial function, which is particularly important for neuronal health. Aberrant post-translational modifications of these proteins may disrupt their cellular functions and thus contributing to the development of idiopathic PD. Some post-translational modifictions can be attributed to the dysregulation of potentially harmful reactive oxygen and nitrogen species inside the cell, which promote oxidative and nitrosative stress, respectively. Unlike oxidative modifications, the covalent modification by Nitric Oxide under nitrosative stress, leading to S-nitrosylation of Parkin, DJ-1; and PINK1, is less studied. Here, we review the available literature on S-nitrosylation of these three proteins, their implications in the pathogenesis of PD, and provide an overview of currently known, denitrosylating systems in eukaryotic cells.

Activation of RAW264.7 cells by PCp-I, a polysaccharide from Psoralea corylifolia L, through NF-κB/MAPK signalling pathway.

PCp-I is a polysaccharide isolated and identified from the Psoralea corylifolia L. by our research group. In this study, the immunomodulatory effects of PCp-I on RAW264.7 cells was evaluated. PCp-I could enhance the level of NO along with up-regulation of iNOS mRNA in RAW264.7 cells. The PCp-I could significantly up-regulate the mRNA expression of TNF-α and IL-6 in RAW264.7 cells, and then the expression of TNF-α, IL-6, ROS and the phagocytic activity were increased. Additionally, PCp-I could significantly up-regulate the phosphorylation level of p65, p38, ERK and JNK proteins, which proved that PCp-I could activate the macrophages by MAPKs and NF-κB signalling pathway and the TLR4 may be one of the receptors of PCp-I regulate the RAW264.7 cells.

Metabolomics in Severe Aortic Stenosis Reveals Intermediates of Nitric Oxide Synthesis as Most Distinctive Markers.

BACKGROUND: Calcific aortic valve disease (CAVD) is a rapidly growing global health problem with an estimated 12.6 million cases globally in 2017 and a 112% increase of deaths since 1990 due to aging and population growth. CAVD may develop into aortic stenosis (AS) by progressive narrowing of the aortic valve. AS is underdiagnosed, and if treatment by aortic valve replacement (AVR) is delayed, this leads to poor recovery of cardiac function, absence of symptomatic improvement and marked increase of mortality. Considering the current limitations to define the stage of AS-induced cardiac remodeling, there is need for a novel method to aid in the diagnosis of AS and timing of intervention, which may be found in metabolomics profiling of patients. METHODS: Serum samples of nine healthy controls and 10 AS patients before and after AVR were analyzed by untargeted mass spectrometry. Multivariate modeling was performed to determine a metabolic profile of 30 serum metabolites which distinguishes AS patients from controls. Human cardiac microvascular endothelial cells (CMECs) were incubated with serum of the AS patients and then stained for ICAM-1 with Western Blot to analyze the effect of AS patient serum on endothelial cell activation. RESULTS: The top 30 metabolic profile strongly distinguishes AS patients from healthy controls and includes 17 metabolites related to nitric oxide metabolism and 12 metabolites related to inflammation, in line with the known pathomechanism for calcific aortic valve disease. Nine metabolites correlate strongly with left ventricular mass, of which three show reversal back to control values after AVR. Western blot analysis of CMECs incubated with AS patient sera shows a significant reduction (14%) in ICAM-1 in AS samples taken after AVR compared to AS patient sera before AVR. CONCLUSION: Our study defined a top 30 metabolic profile with biological and clinical relevance, which may be used as blood biomarker to identify AS patients in need of cardiac surgery. Future studies are warranted in patients with mild-to-moderate AS to determine if these metabolites reflect disease severity and can be used to identify AS patients in need of cardiac surgery.

Role of Rho in Salt-Sensitive Hypertension.

A high amount of salt in the diet increases blood pressure (BP) and leads to salt-sensitive hypertension in individuals with impaired renal sodium excretion. Small guanosine triphosphatase (GTP)ase Rho and Rac, activated by salt intake, play important roles in the pathogenesis of salt-sensitive hypertension as key switches of intracellular signaling. Focusing on Rho, high salt intake in the central nervous system increases sodium concentrations of cerebrospinal fluid in salt-sensitive subjects via Rho/Rho kinase and renin-angiotensin system activation and causes increased brain salt sensitivity and sympathetic nerve outflow in BP control centers. In vascular smooth muscle cells, Rho-guanine nucleotide exchange factors and Rho determine sensitivity to vasoconstrictors such as angiotensin II (Ang II), and facilitate vasoconstriction via G-protein and Wnt pathways, leading to increased vascular resistance, including in the renal arteries, in salt-sensitive subjects with high salt intake. In the vascular endothelium, Rho/Rho kinase inhibits nitric oxide (NO) production and function, and high salt amounts further augment Rho activity via asymmetric dimethylarginine, an endogenous inhibitor of NO synthetase, causing aberrant relaxation and increased vascular tone. Rho-associated mechanisms are deeply involved in the development of salt-sensitive hypertension, and their further elucidation can help in developing effective protection and new therapies.