CRISPR-targeted mutagenesis of mitogen-activated protein kinase phosphatase 1 improves both immunity and yield in wheat.

Plants have evolved a sophisticated immunity system for specific detection of pathogens and rapid induction of measured defences. Over- or constitutive activation of defences would negatively affect plant growth and development. Hence, the plant immune system is under tight positive and negative regulation. MAP kinase phosphatase1 (MKP1) has been identified as a negative regulator of plant immunity in model plant Arabidopsis. However, the molecular mechanisms by which MKP1 regulates immune signalling in wheat (Triticum aestivum) are poorly understood. In this study, we investigated the role of TaMKP1 in wheat defence against two devastating fungal pathogens and determined its subcellular localization. We demonstrated that knock-down of TaMKP1 by CRISPR/Cas9 in wheat resulted in enhanced resistance to rust caused by Puccinia striiformis f. sp. tritici (Pst) and powdery mildew caused by Blumeria graminis f. sp. tritici (Bgt), indicating that TaMKP1 negatively regulates disease resistance in wheat. Unexpectedly, while Tamkp1 mutant plants showed increased resistance to the two tested fungal pathogens they also had higher yield compared with wild-type control plants without infection. Our results suggested that TaMKP1 interacts directly with dephosphorylated and activated TaMPK3/4/6, and TaMPK4 interacts directly with TaPAL. Taken together, we demonstrated TaMKP1 exert negative modulating roles in the activation of TaMPK3/4/6, which are required for MAPK-mediated defence signalling. This facilitates our understanding of the important roles of MAP kinase phosphatases and MAPK cascades in plant immunity and production, and provides germplasm resources for breeding for high resistance and high yield.

Takotsubo Syndrome: Finally Emerging From the Shadows?

It is now 30 years since Japanese investigators first described Takotsubo Syndrome (TTS) as a disorder occurring mainly in ageing women, ascribing it to the impact of multivessel coronary artery spasm. During the intervening period, it has become clear that TTS involves relatively transient vascular injury, followed by prolonged myocardial inflammatory and eventually fibrotic changes. Hence symptomatic recovery is generally slow, currently an under-recognised issue. It appears that TTS is induced by aberrant post-ß2-adrenoceptor signalling in the setting of "surge" release of catecholamines. Resultant activation of nitric oxide synthases and increased inflammatory vascular permeation lead to prolonged myocardial infiltration with macrophages and associated oedema formation. Initially, the diagnosis of TTS was made via exclusion of relevant coronary artery stenoses, plus the presence of regional left ventricular hypokinesis. However, detection of extensive myocardial oedema on cardiac MRI imaging offers a specific basis for diagnosis. No adequate methods are yet available for definitive diagnosis of TTS at hospital presentation. Other major challenges remaining in this area include understanding of the recently demonstrated association between TTS and antecedent cancer, the development of effective treatments to reduce risk of short-term (generally due to shock) and long-term mortality, and also to accelerate symptomatic recovery.

Neutrophil-Initiated Myocardial Inflammation and Its Modulation by B-Type Natriuretic Peptide: A Potential Therapeutic Target.

Activation of neutrophils is a critically important component of the innate immune response to bacterial and chemical stimuli, and culminates in the "neutrophil burst", which facilitates neutrophil phagocytosis via the release of superoxide anion radical (O2-) from NADPH oxidase. Excessive and/or prolonged neutrophil activation results in substantial tissue injury and increases in vascular permeability-resulting in sustained tissue infiltration with neutrophils and monocytes, and persistent vasomotor dysfunction. Cardiovascular examples of such changes include acute and chronic systolic and diastolic heart failure ("heart failure with preserved ejection fraction"), and the catecholamine-induced inflammatory disorder takotsubo syndrome. We have recently demonstrated that B-type natriuretic peptide (BNP), acting via inhibition of activation of neutrophil NADPH oxidase, is an important negative modulator of the "neutrophil burst", though its effectiveness in limiting tissue injury is partially lost in acute heart failure. The potential therapeutic implications of these findings, regarding the development of new means of treating both acute and chronic cardiac injury states, are discussed.

Hypoxic potentiation of nitrite effects in human vessels and platelets.

Previous studies in non-human blood vessels and in platelets have demonstrated that under hypoxic conditions release of NO from nitrite (NO2(-)) is potentiated by deoxyhaemoglobin. In the current study, we characterized hypoxic potentiation of NO2(-) effects in human vasculature and platelets in vitro, addressing underlying mechanisms. The vasodilator efficacy of NO2(-), in comparison with glyceryl trinitrate (GTN), was evaluated in vitro, using segments of human saphenous vein. Under hypoxic conditions, there was a leftward shift of the NO2(-) concentration-response curve (EC50: 22 µM in hyperoxia vs 3.5 µM in hypoxia; p<0.01), but no significant potentiation of GTN effect. In the presence of red blood cells, hypoxic potentiation of NO2(-) vasodilator effect was accentuated. In whole blood samples and platelet-rich plasma (PRP) we assessed inhibition of platelet aggregation by NO2(-) (1mM), in comparison with that of sodium nitroprusside (SNP, 10 µM). In individual subjects (n=37), there was a strong correlation (r=0.75, p<0.0001) between anti-aggregatory effects of NO2(-) and SNP in whole blood, signifying that resultant sGC activation underlies biological effect and responses to NO2(-) are diminished in the presence of NO resistance. In PRP, the effects of NO2(-) were less pronounced than in whole blood (p=0.0001), suggesting an important role of Hb (within RBCs) in the bioconversion of NO2(-) to NO. Inhibition of platelet aggregation by NO2(-) was almost 3-fold greater in venous than in arterial blood (p<0.0001), and deoxyHb concentration directly correlated (r=0.69, p=0.013) with anti-aggregatory response. Incremental hypoxia applied to venous blood samples (in hypoxic chamber) caused a progressive increase in both deoxyHb level and anti-aggregatory effect of NO2(-). When subjects inhaled a 12% O2 mixture for 20 min, there was a 3-fold rise in blood deoxyHb fraction (p<0.01). In PRP, response to NO2(-) also increased under hypoxia, and was further enhanced (p<0.01) by deoxyHb. Furthermore, deoxyHb exerted significant anti-aggregatory effects even in the absence of added NO2(-), suggesting a role for endogenous NO2(-). The results of this work provide further mechanistic insights into hypoxic potentiation of vasodilator and anti-aggregatory actions of NO2(-). In human saphenous veins and blood, the balance of evidence suggests differential rates of NO release from NO2(-) (largely modulated by deoxyHb) as the fundamental mechanism.

Thioredoxin-interacting protein: pathophysiology and emerging pharmacotherapeutics in cardiovascular disease and diabetes.

The thioredoxin system, which consists of thioredoxin (Trx), nicotinamide adenine dinucleotide phosphate (NADPH) and thioredoxin reductase (TrxR), has emerged as a major anti-oxidant involved in the maintenance of cellular physiology and survival. Dysregulation in this system has been associated with metabolic, cardiovascular, and malignant disorders. Thioredoxin-interacting protein (TXNIP), also known as vitamin D-upregulated protein or thioredoxin-binding-protein-2, functions as a physiological inhibitor of Trx, and pathological suppression of Trx by TXNIP has been demonstrated in diabetes and cardiovascular diseases. Furthermore, TXNIP effects are partially Trx-independent; these include direct activation of inflammation and inhibition of glucose uptake. Many of the effects of TXNIP are initiated by its dissociation from intra-nuclear binding with Trx or other SH-containing proteins: these effects include its migration to cytoplasm, modulating stress responses in mitochondria and endoplasmic reticulum, and also potentially activating apoptotic pathways. TXNIP also interacts with the nitric oxide (NO) signaling system, with apparent suppression of NO effect. TXNIP production is modulated by redox stress, glucose levels, hypoxia and several inflammatory activators. In recent studies, it has been shown that therapeutic agents including insulin, metformin, angiotensin converting enzyme inhibitors and calcium channel blockers reduce TXNIP expression, although it is uncertain to what extent TXNIP suppression contributes to their clinical efficacy. This review addresses the role of TXNIP in health and in cardiovascular and metabolic disorders. Finally, the potential advantages (and disadvantages) of pharmacological suppression of TXNIP in cardiovascular disease and diabetes are summarized.

B-Type natriuretic peptide suppression of neutrophil superoxide generation: mechanistic studies in normal subjects.

Many acute cardiovascular disease states are associated with neutrophil infiltration of myocardium and subsequent release of superoxide (O2 (-) ) and myeloperoxidase (MPO), which contribute to inflammatory reactions. B-Type natriuretic peptide (BNP) is known to exert anti-inflammatory and antifibrotic effects, but it is not known whether these may include interactions with neutrophils. In neutrophils isolated from 20 healthy subjects, we assessed the effect of BNP on the 'neutrophil burst' (O2 (-) production and MPO release) stimulated by phorbol myristate acetate (PMA) and N-formyl-methionyl-leucyl-phenylalanine (fMLP), respectively. Effects of BNP on cGMP accumulation, and the effects of the cell-permeable cGMP analogue 8-(4-chlorophenylthio) guanosine-cGMP (8-p-CPT-cGMP) and protein kinase G (PKG) inhibition with KT5823 on the neutrophil-BNP interaction were also evaluated. B-Type natriuretic peptide suppressed O2 (-) release from neutrophils by 23 ± 6% (P < 0.001) and 24 ± 8% (P < 0.05) following PMA and fMLP stimulation, respectively. Although BNP did not significantly increase cGMP formation, 8-p-CPT-cGMP suppressed both PMA- and fMLP-induced neutrophil O2 (-) release by 16% and 28%, respectively (P < 0.05). The PKG inhibitor KT5823 attenuated the effects of BNP on both fMLP- and PMA-associated O2 (-) production. Neither BNP nor 8-p-CPT-cGMP significantly affected MPO release from neutrophils. Suppression of O2 (-) release from neutrophils by BNP may contribute to its anti-inflammatory and antifibrotic actions.

Functional Evolution of Pseudofabraea citricarpa as an Adaptation to Temperature Change.

Citrus target spot, caused by Pseudofabraea citricarpa, was formerly considered a cold-tolerant fungal disease. However, it has now spread from high-latitude regions to warmer low-latitude regions. Here, we conducted physiological observations on two different strains of the fungus collected from distinct regions, and evaluated their pathogenicity. Interestingly, the CQWZ collected from a low-latitude orchard, exhibited higher temperature tolerance and pathogenicity when compared to the SXCG collected from a high-latitude orchard. To further understand the evolution of temperature tolerance and virulence in these pathogens during the spread process, as well as the mechanisms underlying these differences, we performed genomic comparative analysis. The genome size of CQWZ was determined to be 44,004,669 bp, while the genome size of SXCG was determined to be 45,377,339 bp. Through genomic collinearity analysis, we identified two breakpoints and rearrangements during the evolutionary process of these two strains. Moreover, gene annotation results revealed that the CQWZ possessed 376 annotated genes in the "Xenobiotics biodegradation and metabolism" pathway, which is 79 genes more than the SXCG. The main factor contributing to this difference was the presence of salicylate hydroxylase. We also observed variations in the oxidative stress pathways and core pathogenic genes. The CQWZ exhibited the presence of a heat shock protein (HSP SSB), a catalase (CAT2), and 13 core pathogenic genes, including a LysM effector, in comparison to the SXCG. Furthermore, there were significant disparities in the gene clusters responsible for the production of seven metabolites, such as Fumonisin and Brefeldin. Finally, we identified the regulatory relationship, with the HOG pathway at its core, that potentially contributes to the differences in thermotolerance and virulence. As the global climate continues to warm, crop pathogens are increasingly expanding to new territories. Our findings will enhance understanding of the evolution mechanisms of pathogens under climate change.

Diagnostic and prognostic significance of circulating secreted frizzled-related protein 5 in colorectal cancer.

BACKGROUND: Secreted Frizzled-Related Protein 5 (SFRP5) modulates Wnt signalling pathways, affecting diverse biological processes. We assessed the diagnostic and prognostic value of circulating SFRP5 (cSFRP5) in colorectal cancer (CRC) METHODS: Plasma cSFRP5 concentrations were measured using enzyme-linked immunosorbent assay (ELISA) in healthy donors (n = 133), individuals diagnosed with CRC (n = 449), colorectal polyps (n = 85), and medical conditions in other organs including cancer, inflammation, and benign states (n = 64). RESULTS: Patients with CRC, polyps, and other conditions showed higher cSFRP5 levels than healthy individuals (p < 0.0001). Receiver operating characteristic curves comparing healthy donors with medical conditions, polyps and CRC were 0.814 (p < 0.0001), 0.763 (p < 0.0001) and 0.762 (p < 0.0001), respectively. In CRC, cSFRP5 correlated with patient age (p < 0.0001), tumour stage (p < 0.0001), and histological differentiation (p = 0.0273). Levels, adjusted for patient age, sex, plasma age and collection institution, peaked in stage II versus I (p < 0.0001), III (p = 0.0002) and IV (p < 0.0001), were lowest in stage I versus III (p = 0.0002) and IV (p = 0.0413), with no difference between stage III and IV. Elevated cSFRP5 levels predicted longer overall survival in stages II-III CRC (univariate: HR 1.82, 95% CI: 1.02-3.26, p = 0.024; multivariable: HR 2.34, 95% CI: 1.12-4.88, p = 0.015). CONCLUSION: This study confirms cSFRP5 levels are elevated in CRC compared to healthy control and reveals a correlation between elevated cSFRP5 and overall survival in stages II-III disease.

LysM Proteins TaCEBiP and TaLYK5 are Involved in Immune Responses Mediated by Chitin Coreceptor TaCERK1 in Wheat.

Plant lysin motif (LysM) ectodomain receptors interact with pathogen-associated molecular patterns (PAMPs) and have critical functions in plant-microbe interactions. In this study, 65 LysM family genes were identified using the recent version of the reference sequence of bread wheat (Triticum aestivum), in which 23, 16, 20, and 6 members belonged to LysM-containing receptor-like kinases (LYKs), LysM-containing receptor-like proteins (LYPs), extracellular LysM proteins (LysMes), and intracellular nonsecretory LysM proteins (LysMns), respectively. The study found that TaCEBiP, TaLYK5, and TaCERK1 were highly responsive to PAMP elicitors and phytopathogens, with TaCEBiP and TaLYK5 binding directly to chitin. TaCERK1 acted as a coreceptor with TaCEBiP and TaLYK5 at the plasma membrane. Overexpression of TaCEBiP, TaLYK5, and TaCERK1 in Nicotiana benthamiana leaves exhibited enhanced resistance to Sclerotinia sclerotiorum. Subsequently, knocking down TaCEBiP, TaLYK5, and TaCERK1 genes with barley stripe mosaic virus-VIGS compromised the wheat defense response to an avirulent strain of Puccinia striiformis. The study concluded that wheat has two synergistic chitin perception systems for detecting pathogen elicitors, with the activated CERK1 intracellular kinase domain leading to signaling transduction. This research provides valuable insights into the functional roles and regulatory mechanisms of wheat LysM members under biotic stress.

A chitin deacetylase PsCDA2 from Puccinia striiformis f. sp. tritici confers disease pathogenicity by suppressing chitin-triggered immunity in wheat.

Plants have the ability to recognize the essential chitin molecule present in the fungal cell wall, which stimulates the immune response. Phytopathogenic fungi have developed various strategies to inhibit the chitin-triggered immune response. Here, we identified a chitin deacetylase of Puccinia striiformis f. sp. tritici (Pst), known as PsCDA2, that was induced during the initial invasion of wheat and acted as an inhibitor of plant cell death. Knockdown of PsCDA2 in wheat enhanced its resistance against Pst, highlighting the significance of PsCDA2 in the host-pathogen interaction. Moreover, PsCDA2 can protect Pst urediniospores from being damaged by host chitinase in vitro. PsCDA2 also suppressed the basal chitin-induced plant immune response, including the accumulation of callose and the expression of defence genes. Overall, our results demonstrate that Pst secretes PsCDA2 as a chitin deacetylase involved in establishing infection and modifying the acetyl group to prevent the breakdown of chitin in the cell wall by host endogenous chitinases. Our research unveils a mechanism by which the fungus suppresses plant immunity, further contributing to the understanding of wheat stripe rust control. This information could have significant implications for the development of suitable strategies for protecting crops against the devastating effects of this disease.

Fibulin-3 is necessary to prevent cardiac rupture following myocardial infarction.

Despite the high prevalence of heart failure in the western world, there are few effective treatments. Fibulin-3 is a protein involved in extracellular matrix (ECM) structural integrity, however its role in the heart is unknown. We have demonstrated, using single cell RNA-seq, that fibulin-3 was highly expressed in quiescent murine cardiac fibroblasts, with expression highest prior to injury and late post-infarct (from ~ day-28 to week-8). In humans, fibulin-3 was upregulated in left ventricular tissue and plasma of heart failure patients. Fibulin-3 knockout (Efemp1-/-) and wildtype mice were subjected to experimental myocardial infarction. Fibulin-3 deletion resulted in significantly higher rate of cardiac rupture days 3-6 post-infarct, indicating a weak and poorly formed scar, with severe ventricular remodelling in surviving mice at day-28 post-infarct. Fibulin-3 knockout mice demonstrated less collagen deposition at day-3 post-infarct, with abnormal collagen fibre-alignment. RNA-seq on day-3 infarct tissue revealed upregulation of ECM degradation and inflammatory genes, but downregulation of ECM assembly/structure/organisation genes in fibulin-3 knockout mice. GSEA pathway analysis showed enrichment of inflammatory pathways and a depletion of ECM organisation pathways. Fibulin-3 originates from cardiac fibroblasts, is upregulated in human heart failure, and is necessary for correct ECM organisation/structural integrity of fibrotic tissue to prevent cardiac rupture post-infarct.

Integrated Metabolo-transcriptomics Reveals the Defense Response of Homogentisic Acid in Wheat against Puccinia striiformis f. sp. tritici.

Stripe rust is a widespread and harmful wheat disease caused by Puccinia striiformis f. sp. tritici (Pst) worldwide. Targeted metabolome and transcriptomics analyses of CYR23 infected leaves were performed to identify the differential metabolites and differentially expressed genes related to wheat disease resistance. We observed upregulation of 33 metabolites involved in the primary and secondary metabolism, especially for homogentisic acid (HGA), p-coumaroylagmatine, and saccharopine. These three metabolites were mainly involved in the phenylpropanoid metabolic pathway, hydroxycinnamic acid amides pathway, and saccharopine pathway. Combined with transcriptome data on non-compatible interaction, the synthesis-related genes of these three differential metabolites were all upregulated significantly. The gene regulatory network involved in response to Pst infection was constructed, which revealed that several transcription factor families including WRKYs, MYBs, and bZIPs were identified as potentially hubs in wheat resistance response against Pst. An in vitro test showed that HGA effectively inhibited the germination of stripe rust fungus urediniospores and reduced the occurrence of wheat stripe rust. The results of gene silencing and overexpression of HGA synthesis-related gene 4-hydroxyphenylpyruvate dioxygenase proved that HGA was involved in wheat disease resistance. These results provided a further understanding of the disease resistance of wheat and indicated that HGA can be developed as a potential agent against Pst.

The Role of Hydroxycinnamic Acid Amide Pathway in Plant Immunity.

The compounds involved in the hydroxycinnamic acid amide (HCAA) pathway are an important class of metabolites in plants. Extensive studies have reported that a variety of plant hydroxycinnamamides exhibit pivotal roles in plant-pathogen interactions, such as p-coumaroylagmatine and ferulic acid. The aim of this review is to discuss the emerging findings on the functions of hydroxycinnamic acid amides (HCAAs) accumulation associated with plant defenses against plant pathologies, antimicrobial activity of HCAAs, and the mechanism of HCAAs involved in plant immune responses (such as reactive oxygen species (ROS), cell wall response, plant defense hormones, and stomatal immunity). However, these advances have also revealed the complexity of HCAAs participation in plant defense reactions, and many mysteries remain to be revealed. This review provides an overview of the mechanistic and conceptual insights obtained so far and highlights areas for future exploration of phytochemical defense metabolites.

Perhexiline Therapy in Patients with Type 2 Diabetes: Incremental Insulin Resistance despite Potentiation of Nitric Oxide Signaling.

Perhexiline (Px) inhibits carnitine palmitoyltransferase 1 (CPT1), which controls uptake of long chain fatty acids into mitochondria. However, occasional cases of hypoglycaemia have been reported in Px-treated patients, raising the possibility that Px may also increase sensitivity to insulin. Furthermore, Px increases anti-aggregatory responses to nitric oxide (NO), an effect which may theoretically parallel insulin sensitization. We therefore sought to examine these relationships in patients with stable Type 2 diabetes (T2D) and cardiovascular disease (n = 30). Px was initiated, and dosage was titrated, to reach the therapeutic range and thus prevent toxicity. Investigations were performed before and after 2 weeks, to examine changes in insulin sensitivity and, utilizing aggregometry in whole blood, platelet responsiveness to the anti-aggregatory effects of the NO donor sodium nitroprusside (SNP). Other parameters that affect may affect NO signalling were also evaluated. Px substantially potentiated inhibition of platelet aggregation by SNP (from 16.7 ± 3.0 to 27.3 ± 3.7%; p = 0.005). Px did not change fasting blood glucose concentrations but reduced insulin sensitivity (HOMA-IR score increased from median of 4.47 to 6.08; p = 0.028), and increased fasting plasma insulin concentrations (median 16.5 to 19.0 mU/L; p = 0.014). Increases in SNP responses tended (r = -0.30; p = 0.11) to be reciprocally related to increases in HOMA-IR, and increases in HOMA-IR were greater (p = 0.002) in patients without NO-sensitizing effects. No patient developed symptomatic hypoglycaemia, nor was there any other short-term toxicity of Px. Thus, in patients with stable T2D and cardiovascular disease, Px increases anti-aggregatory responsiveness to NO, but is not an insulin sensitizer, and does not induce hypoglycaemia. Absence of NO-sensitizing effect occurs in approximately 30% of Px-treated patients with T2D, and is associated with induction of insulin resistance in these patients.

In vitro evaluation of gaseous microemboli handling of cardiopulmonary bypass circuits with and without integrated arterial line filters.

The delivery of gaseous microemboli (GME) by the cardiopulmonary bypass circuit should be minimized whenever possible. Innovations in components, such as the integration of arterial line filter (ALF) and ALFs with reduced priming volumes, have provided clinicians with circuit design options. However, before adopting these components clinically, their GME handling ability should be assessed. This study aims to compare the GME handling ability of different oxygenator/ALF combinations with our currently utilized combination. Five commercially available oxygenator/ALF combinations were evaluated in vitro: Terumo Capiox SX25RX and Dideco D734 (SX/D734),Terumo Capiox RX25R and AF125 (RX/AF125), Terumo FX25R (FX), Sorin Synthesis with 102 microm reservoir filter (SYN102), and Sorin Synthesis with 40 microm reservoir filter (SYN40). GME handling was studied by introducing air into the venous return at 100 mL/min for 60 seconds under two flow/ pressure combinations: 3.5 L/min, 150 mmHg and 5 L/min, 200 mmHg. Emboli were measured at three positions in the circuit using the Emboli Detection and Classification (EDAC) Quantifier and analyzed with the General Linear Model. All circuits significantly reduced GME. The SX/D734 and SYN40 circuits were most efficient in GME removal whilst the SYN102 handled embolic load (count and volume) least efficiently (p < .001). A greater number of emboli <70 microm were observed for the SYN102, FX and RX/AF125 circuits (p < .001). An increase in embolic load occurred with higher flow/pressure in all circuits (p < .001). The venous reservoir significantly influences embolic load delivered to the oxygenator (p < .001). The majority of introduced venous air was removed; however, significant variation existed in the ability of the different circuits to handle GME. Venous reservoir design influenced the overall GME handling ability. GME removal was less efficient at higher flow and pressure, and for smaller sized emboli. The clinical significance of reducing GME requires further investigation.

Impairment of platelet NO signalling in coronary artery spasm: role of hydrogen sulphide.

BACKGROUND AND PURPOSE: The pathophysiology of coronary artery spasm (CAS), with its associated ischaemic crises, is currently poorly understood and treatment is frequently ineffective. In view of increasing evidence that platelet-based defects may occur in CAS patients, we investigated platelet reactivity in CAS patients and whether symptomatic crises reflect activation of platelet-endothelial interactions. EXPERIMENTAL APPROACH: CAS patients were evaluated during acute and/or chronic symptomatic phases and compared with healthy control subjects. Inhibition of ADP-induced platelet aggregation by the NO donor sodium nitroprusside (SNP) and plasma concentrations of syndecan 1 (glycocalyx shedding marker), tryptase (mast cell activation marker) and platelet microparticles were measured. KEY RESULTS: Inhibition of platelet aggregation by SNP was diminished in chronic CAS, with further (non-significant) deterioration during symptomatic crises, whereas plasma concentrations of syndecan 1, tryptase and platelet microparticles increased. Treatment of patients with high-dose N-acetylcysteine (NAC) plus glyceryl trinitrate rapidly increased platelet responsiveness to SNP and decreased plasma syndecan 1 concentrations. The effect of NAC on platelet responsiveness to SNP was confirmed in vitro and mimicked by the H2 S donor NaHS. Conversely, inhibition of enzymatic production of H2 S attenuated NAC effect. CONCLUSION AND IMPLICATIONS: CAS is associated with substantial impairment of platelet NO signalling. During acute symptomatic exacerbations, platelet resistance to NO is aggravated, together with mast cell activation and damage to both vasculature and platelets. NAC, via release of H2 S, reverses platelet resistance to NO and terminates glycocalyx shedding during symptomatic crises: This suggests that H2 S donors may correct the pathophysiological anomalies underlying CAS.

Prolonged suppression of the anti-oxidant/anti-inflammatory effects of BNP post-Takotsubo syndrome.

AIMS: Takotsubo syndrome (TTS) episodes are primarily initiated by 'pulse' release of catecholamines inducing neutrophil infiltration and myocardial inflammation in susceptible individuals (largely ageing women). Evidence of myocardial inflammation and associated energetic impairment persists for ≥ 3 months post-acute TTS episodes, suggesting the existence of additional 'perpetuating' mechanisms. The effects of B-type natriuretic peptide (BNP) in suppressing superoxide (O2- ) release from neutrophils are transiently impaired in acute heart failure. We also evaluated the extent and duration of BNP-induced suppression of O2- release post-TTS. METHODS AND RESULTS: TTS patients were studied acutely (n = 34) and 3 months thereafter (n = 13) and compared with control subjects (n = 25). O2- generation from neutrophils, triggered by N-formyl-methionyl-leucyl-phenylalanine and phorbol myristate acetate, and its suppression by BNP, were measured in vitro. Determinants of variability in BNP effect were sought via univariate and multivariate analyses. Relative to control subjects, in TTS patients, BNP suppression of both phorbol myristate acetate and N-formyl-methionyl-leucyl-phenylalanine-induced O2- release was impaired acutely (P < 0.05 for both); this did not improve over the 3-month recovery period, despite treatment with conventional anti-failure medication in 85% of patients. No significant correlates of BNP effect (other than TTS) were identified. CONCLUSIONS: (1) While TTS is associated with marked and prolonged release of BNP, there is virtually total loss of the ability of BNP to suppress neutrophil O2- release and its impact on tissue inflammation. (2) BNP responses do not recover for at least 3 months post-attacks, suggesting that this might contribute to perpetuation of myocardial inflammation in TTS patients.

Does cardiac resynchronization therapy restore peripheral circulatory homeostasis?

AIMS: To evaluate whether peripheral circulatory 'remodelling' as measured by changes in vascular compliance and in markers of nitric oxide signalling contributes to patient response to cardiac resynchronization therapy (CRT). METHODS AND RESULTS: Effects of CRT were evaluated in 33 patients pre-procedure and 6 months post-procedure. Peak oxygen consumption, 6 min walk distance, New York Heart Association class, and quality of life score were evaluated. Augmentation index and its interactions with nitric oxide (NO) were evaluated by applanation tonometry. Platelet NO responsiveness and content of thioredoxin-interacting protein were assessed. Plasma concentrations of N-terminal proBNP, asymmetric and symmetric dimethylarginine (SDMA), high sensitivity C-reactive protein, catecholamines, and matrix metalloproteinases-2 and -9 were assessed. Despite significant improvement in 6 min walk distance (P = 0.005), New York Heart Association class (P < 0.001), quality of life (P = 0.001), and all echocardiographic parameters post-CRT, there were no significant changes in augmentation index measurements, thioredoxin-interacting protein content, and platelet NO response. Significant falls in N-terminal proBNP (P = 0.008) and SDMA (P = 0.013; independent of renal function) occurred. Falls in SDMA predicted reduction in high-sensitivity C-reactive protein (P = 0.04) and increases in peak oxygen consumption (P = 0.04). There were no correlations between changes in echocardiographic parameters and those in vascular function. CONCLUSIONS: These data suggest that the beneficial effects of CRT over 6 months are independent of any change in peripheral NO-related signalling. However, there is evidence that suppression of inflammation occurs, and its magnitude predicts extent of clinical improvement.

Glycocalyx shedding is markedly increased during the acute phase of Takotsubo cardiomyopathy.

BACKGROUND: Acute myocardial infarction (AMI) and other forms of myocardial acute oxidative stress are associated with variable "shedding" of the endothelial glycocalyx (GCS) which can be quantitated ex vivo by release into plasma of glycocalyx components such as Syndecan-1 (SD-1). Previous studies have implicated release of both catecholamines and BNP as potential accentuating factors in GCS: since these are prominent aspects of the pathogenesis of Takotsubo cardiomyopathy (TTC), we hypothesised that TTC is associated with increased GCS and the extent of GCS is predictable on the basis of NT-proBNP and catecholamine releases. METHODS: SD-1 concentrations were measured in 48 TTC patients acutely and after 3months, and compared with those in 12 healthy controls, and 17 patients with AMI. Correlations were sought between SD-1 levels markers of severity of TTC episodes in individual patients. RESULTS: Acute SD-1 concentrations in TTC patients were elevated significantly (p<0.0001, 1-way ANOVA) compared to control values. There were no significant correlations between SD-1 concentrations and any markers of severity of acute TTC episodes, such as NT-proBNP or catecholamine release. Over 3months, SD-1 concentrations fell significantly (p=0.0002) to approximately the same values as in control subjects. CONCLUSIONS: TTC is associated acutely with a marked increase in GCS. Potentially, GCS might contribute to increased coronary vascular permeability in TTC, thus dissociating development of myocardial oedema from severity of associated inflammation. Prevention of GCS represents a potential therapeutic option in TTC.