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Myocardial ischemia-reperfusion (I/R) injury is a prevalent cause of myocardial injury, involving a series of interconnected pathophysiological processes. However, there is currently no clinical therapy for effectively mitigating myocardial I/R injury. Here, we show that p85α protein levels increase in response to I/R injury through a comprehensive analysis of cardiac proteomics, and confirm this in the I/R-injured murine heart and failing human myocardium. Genetic inhibition of p85α in mice activates the Akt-GSK3β/Bcl-x(L) signaling pathway and ameliorates I/R-induced cardiac dysfunction, apoptosis, inflammation, and mitochondrial dysfunction. p85α silencing in cardiomyocytes alleviates hypoxia-reoxygenation (H/R) injury through activating the Akt-GSK3β/Bcl-x(L) signaling pathway, while its overexpression exacerbates the damage. Mechanistically, the interaction between MG53 and p85α triggers the ubiquitination and degradation of p85α, consequently enhancing Akt phosphorylation and ultimately having cardioprotective effects. Collectively, our findings reveal that substantial reduction of p85α and subsequently activated Akt signaling have a protective effect against cardiac I/R injury, representing an important therapeutic strategy for mitigating myocardial damage., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Aims: Regulated necrosis (necroptosis) and apoptosis are important biological features of myocardial infarction, ischaemia-reperfusion (I/R) injury, and heart failure. However, the molecular mechanisms underlying myocardial necroptosis remain elusive. Ischaemic preconditioning (IPC) is the most powerful intrinsic cardioprotection against myocardial I/R injury. In this study, we aimed to determine whether IPC suppresses I/R-induced necroptosis and the underlying molecular mechanisms., Methods and Results: We generated p55γ transgenic and knockout mice and used ligation of left anterior descending coronary artery to produce an in vivo I/R model. The effects of p55γ and its downstream molecules were subsequently identified using mass spectroscopy and co-immunoprecipitation and pulldown assays. We found that p55γ expression was down-regulated in failing human myocardium caused by coronary heart disease as well as in I/R mouse hearts. Cardiac-specific p55γ overexpression ameliorated the I/R-induced necroptosis. In striking contrast, p55γ deficiency (p55γ-/-) and cardiac-specific deletion of p55γ (p55γc-KO) worsened I/R-induced injury. IPC up-regulated p55γ expression in vitro and in vivo. Using reporter and chromatin immunoprecipitation assays, we found that Hif1α transcriptionally regulated p55γ expression and mediated the cardioprotection of IPC. IPC-mediated suppression of necroptosis was attenuated in p55γ-/- and p55γc-KO hearts. Mechanistically, p55γ overexpression decreased the protein levels of RIP3 rather than the mRNA levels, while p55γ deficiency increased the protein abundance of RIP3. IPC attenuated the I/R-induced up-regulation of RIP3, which was abolished in p55γ-deficient mice. Up-regulation of RIP3 attenuated the p55γ- or IPC-induced inhibition of necroptosis in vivo. Importantly, p55γ directly bound and degraded RIP3 in a ubiquitin-dependent manner. We identified MG53 as the E3 ligase that mediated the p55γ-induced degradation of RIP3. In addition, we also found that p55γ activated the RISK pathway during IPC., Conclusions: Our findings reveal that activation of the MG53-RIP3 signal pathway by p55γ protects the heart against I/R-induced necroptosis and underlies IPC-induced cardioprotection., Competing Interests: Conflict of interest: The authors declare no conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: A previously published meta-analysis found that about one-third of the general population experienced some mental health problem during the early phase of the COVID-19 pandemic, potentially leading to a late mental health crisis. We aimed to describe the acute, short-term, and long-term effects of the COVID-19 pandemic on mental health., Methods: A one-year online survey (S) was conducted in Spain (April 2020 - March 2021). We recruited 18 180 subjects using a virtual respondent-driven snowball sampling method (S1 April 2020, n = 6108; S2 October-November 2020, n = 6418; S3 March 2021, n = 5654). Participants completed the Spanish Depression, Anxiety, and Stress Scale (DASS-21)., Results: Overall, our results suggest a progressive increase in the prevalence of anxiety and stress throughout the pandemic waves and relative stability of depression. Women had a greater probability of having depression, anxiety, or stress than men in each survey ( P  < 0.001). The youngest group (aged 18-24) reported a higher probability ( P  < 0.05) of having depression, anxiety, or stress than the older groups in S1 and S2. Middle-aged people (25-59) had a greater probability of being a case in the DASS-21 scales than the oldest group (60+), except for depression in men ( P  = 0.179). In S3, the trend changed: the youngest group showed a decrease in depression and stress while the oldest group showed a dramatic increase (anxiety: men = 664.5%, women = 273.52%; stress: men = 786%, women = 431.37%)., Conclusions: It is plausible to conclude that COVID-19 psychological fatigue exists, especially in middle-aged and older adults. Strategies to assist people who have fewer coping skills should be implemented in the near future., Competing Interests: Competing interests: The authors completed the ICMJE Unified Competing Interest form (available upon request from the corresponding author), and declare no conflicts of interest., (Copyright © 2022 by the Journal of Global Health. All rights reserved.)

Background: Epidemic outbreaks have significant impact on psychological well-being, increasing psychiatric morbidity among the population. We aimed to describe the early psychological impact of COVID-19 and its contributing factors in a large Spanish sample, globally and according to mental status (never mental disorder NMD, past mental disorder PMD, current mental disorder CMD)., Methods: An online questionnaire was conducted between 19 and 26 March, five days after the official declaration of alarm and the lockdown order. Data included sociodemographic and clinical information and the DASS-21 and IES questionnaires. We analysed 21 207 responses using the appropriate descriptive and univariate tests as well as binary logistic regression to identify psychological risk and protective factors., Results: We found a statistically significant gradient in the psychological impact experienced in five domains according to mental status, with the NMD group being the least affected and the CMD group being the most affected. In the three groups, the depressive response was the most prevalent (NMD = 40.9%, PMD = 51.9%, CMD = 74.4%, F  = 1011.459, P  < 0.001). Risk factors were female sex and classification as a case in any psychological domain. Protective factors were younger age and ability to enjoy free time. Variables related to COVID-19 had almost no impact except for having COVID-19 symptoms, which was a risk factor for anxiety in all three groups., Conclusions: Our results can help develop coping strategies addressing modifiable risk and protective factors for each mental status for early implementation in future outbreaks., Competing Interests: Competing interests: The authors completed the ICMJE Unified Competing Interest form (available upon request from the corresponding author), and declare no conflicts of interest., (Copyright © 2020 by the Journal of Global Health. All rights reserved.)

Background: Ischemic heart disease is the leading cause of morbidity and mortality worldwide. Ischemic preconditioning (IPC) is the most powerful intrinsic protection against cardiac ischemia/reperfusion injury. Previous studies have shown that a multifunctional TRIM family protein, MG53 (mitsugumin 53; also called TRIM72), not only plays an essential role in IPC-mediated cardioprotection against ischemia/reperfusion injury but also ameliorates mechanical damage. In addition to its intracellular actions, as a myokine/cardiokine, MG53 can be secreted from the heart and skeletal muscle in response to metabolic stress. However, it is unknown whether IPC-mediated cardioprotection is causally related to MG53 secretion and, if so, what the underlying mechanism is., Methods: Using proteomic analysis in conjunction with genetic and pharmacological approaches, we examined MG53 secretion in response to IPC and explored the underlying mechanism using rodents in in vivo, isolated perfused hearts, and cultured neonatal rat ventricular cardiomyocytes. Moreover, using recombinant MG53 proteins, we investigated the potential biological function of secreted MG53 in the context of IPC and ischemia/reperfusion injury., Results: We found that IPC triggered robust MG53 secretion in rodents in vivo, perfused hearts, and cultured cardiac myocytes without causing cell membrane leakage. Mechanistically, IPC promoted MG53 secretion through H 2 O 2 -evoked activation of protein kinase-C-δ. Specifically, IPC-induced myocardial MG53 secretion was mediated by H 2 O 2 -triggered phosphorylation of protein kinase-C-δ at Y311, which is necessary and sufficient to facilitate MG53 secretion. Functionally, systemic delivery of recombinant MG53 proteins to mimic elevated circulating MG53 not only restored IPC function in MG53-deficient mice but also protected rodent hearts from ischemia/reperfusion injury even in the absence of IPC. Moreover, oxidative stress by H 2 O 2 augmented MG53 secretion, and MG53 knockdown exacerbated H 2 O 2 -induced cell injury in human embryonic stem cell-derived cardiomyocytes, despite relatively low basal expression of MG53 in human heart., Conclusions: We conclude that IPC and oxidative stress can trigger MG53 secretion from the heart via an H 2 O 2 -protein kinase-C-δ-dependent mechanism and that extracellular MG53 can participate in IPC protection against cardiac ischemia/reperfusion injury.

Background: Mitsugumin 53 (MG53 or TRIM72), a striated muscle-specific E3 ligase, promotes ubiquitin-dependent degradation of the insulin receptor and insulin receptor substrate-1 and subsequently induces insulin resistance, resulting in metabolic syndrome and type 2 diabetes mellitus (T2DM). However, it is unknown how MG53 from muscle regulates systemic insulin response and energy metabolism. Increasing evidence demonstrates that muscle secretes proteins as myokines or cardiokines that regulate systemic metabolic processes. We hypothesize that MG53 may act as a myokine/cardiokine, contributing to interorgan regulation of insulin sensitivity and metabolic homeostasis., Methods: Using perfused rodent hearts or skeletal muscle, we investigated whether high glucose, high insulin, or their combination (conditions mimicking metabolic syndrome or T2DM) alters MG53 protein concentration in the perfusate. We also measured serum MG53 levels in rodents and humans in the presence or absence of metabolic diseases, particularly T2DM. The effects of circulating MG53 on multiorgan insulin response were evaluated by systemic delivery of recombinant MG53 protein to mice. Furthermore, the potential involvement of circulating MG53 in the pathogenesis of T2DM was assessed by neutralizing blood MG53 with monoclonal antibodies in diabetic db/db mice. Finally, to delineate the mechanism underlying the action of extracellular MG53 on insulin signaling, we analyzed the potential interaction of MG53 with extracellular domain of insulin receptor using coimmunoprecipitation and surface plasmon resonance assays., Results: Here, we demonstrate that MG53 is a glucose-sensitive myokine/cardiokine that governs the interorgan regulation of insulin sensitivity. First, high glucose or high insulin induces MG53 secretion from isolated rodent hearts and skeletal muscle. Second, hyperglycemia is accompanied by increased circulating MG53 in humans and rodents with diabetes mellitus. Third, systemic delivery of recombinant MG53 or cardiac-specific overexpression of MG53 causes systemic insulin resistance and metabolic syndrome in mice, whereas neutralizing circulating MG53 with monoclonal antibodies has therapeutic effects in T2DM db/db mice. Mechanistically, MG53 binds to the extracellular domain of the insulin receptor and acts as an allosteric blocker., Conclusions: Thus, MG53 has dual actions as a myokine/cardiokine and an E3 ligase, synergistically inhibiting the insulin signaling pathway. Targeting circulating MG53 opens a new therapeutic avenue for T2DM and its complications.

Background: Lipids, a group of primary metabolites, could be used as quality markers of Traditional Chinese medicine., Purpose: The present study was designed to develop a research method to explore lipid markers of the quality of coix seeds with different geographical origins., Study Design: The geographical origins of coix seeds were divided into three regions based on the latitude. A central composite design (CCD test) was used to optimize the chromatographic parameters of supercritical fluid chromatography to obtain optimal lipid profile of coix seed., Methods: An untargeted method based on ultra-performance convergence chromatography - quadrupole/time-of-flight hybrid mass spectrometry (UPC 2 -QTOF) was developed. Four chromatographic parameters were optimized using CCD test, and a fusion index established by Derringer function was used to evaluate. The lipid profile of 27 batches of coix seeds were acquired and processed by Progenesis QI software, and the MS/MS spectrums were obtained to identify, simultaneously. The difference lipids were explored by orthogonal partial least squares discriminant analysis (OPLS-DA). The lipids that showed differences depending on their seeds' geographical origin were selected as markers of the quality of coix seeds from the three regions., Results: A Torus 2-PIC (1.7 µm, 100 mm × 3.0 mm) was selected as the optimal column of the untargeted method which the run time was only 8 minutes. From the CCD test, the interaction of chromatographic parameters between column temperature and backpressure was founded which the optimal parameters were 55 °C and 2600 psi, respectively. Thirty-two peaks in the lipid profile of coix seed were tentatively identified, of which 20 were triglyceride, and 12 were diglyceride. Nine features that could potentially be used to distinguish the coix seeds by their geographical origin were identified, most of which were diglycerides, such as OP., Conclusions: Our findings confirm that UPC 2 -QTOF combined with chemometrics could be used as an efficient method for exploring potential lipid markers of the quality of herbal medicine., (Copyright © 2018. Published by Elsevier GmbH.)

Triglycerides are the primary constituents of some seed kernels used in traditional Chinese medicine. Quality control of seed kernels containing multiple components with an environmentally friendly method is indispensable for establishing their quality standards (called monographs) in pharmacopeia. Using coix seeds (Semen Coicis) as an example, a green quantification strategy was proposed by combining C 8 core-shell particles with single standard to determine multicomponent technologies to quantify seven triglycerides simultaneously. A core-shell column, namely, Halo C 8 (3.0 × 100 mm, 2.7 µm), was used. Methanol was used as the mobile phase at a flow rate of 0.3 mL/min, enabling UV detection of the elutes. Seven triglycerides were well separated in 20 min, and simultaneously quantified using triolein as a single standard. The conversion factor for each standard was set as 1.0 on ELSD, while for the conversion factors at 203 nm, the values increased with the reduction of linoleate. The recovery values were all in the range of 97 - 107% (RSD < 3.0%). The RSD values of precision, including intraday and intermediate precision, were < 3.0% when the total content of triglycerides was calculated. The linearity reached r ≥ 0.9990, and the limit of quantitation reached 40 - 70 ng. Forty-nine batches of coix seeds from four different places of origins and eight batches of adulterants were evaluated and differentiated using principal component analysis. In addition, the validated method was used successfully to quantity seven triglycerides in Semen Persicae, Semen Armeniacae Amarum, and Semen Pruni., Competing Interests: The authors declare no conflict of interest., (Georg Thieme Verlag KG Stuttgart · New York.)

Though vascular smooth muscle cell (VSMC) proliferation underlies all cardiovascular hyperplastic disorders, our understanding of the molecular mechanisms responsible for this cellular process is still incomplete. Here we report that SRSF1 (serine/arginine-rich splicing factor 1), an essential splicing factor, promotes VSMC proliferation and injury-induced neointima formation. Vascular injury in vivo and proliferative stimuli in vitro stimulate SRSF1 expression. Mice lacking SRSF1 specifically in SMCs develop less intimal thickening after wire injury. Expression of SRSF1 in rat arteries enhances neointima formation. SRSF1 overexpression increases, while SRSF1 knockdown suppresses the proliferation and migration of cultured human aortic and coronary arterial SMCs. Mechanistically, SRSF1 favours the induction of a truncated p53 isoform, Δ133p53, which has an equal proliferative effect and in turn transcriptionally activates Krüppel-like factor 5 (KLF5) via the Δ133p53-EGR1 complex, resulting in an accelerated cell-cycle progression and increased VSMC proliferation. Our study provides a potential therapeutic target for vascular hyperplastic disease.

Regulated necrosis (necroptosis) and apoptosis are crucially involved in severe cardiac pathological conditions, including myocardial infarction, ischemia-reperfusion injury and heart failure. Whereas apoptotic signaling is well defined, the mechanisms that underlie cardiomyocyte necroptosis remain elusive. Here we show that receptor-interacting protein 3 (RIP3) triggers myocardial necroptosis, in addition to apoptosis and inflammation, through activation of Ca(2+)-calmodulin-dependent protein kinase (CaMKII) rather than through the well-established RIP3 partners RIP1 and MLKL. In mice, RIP3 deficiency or CaMKII inhibition ameliorates myocardial necroptosis and heart failure induced by ischemia-reperfusion or by doxorubicin treatment. RIP3-induced activation of CaMKII, via phosphorylation or oxidation or both, triggers opening of the mitochondrial permeability transition pore and myocardial necroptosis. These findings identify CaMKII as a new RIP3 substrate and delineate a RIP3-CaMKII-mPTP myocardial necroptosis pathway, a promising target for the treatment of ischemia- and oxidative stress-induced myocardial damage and heart failure.

The aims of this study were to evaluate the characteristics of hypercoagulable states in patients with membranous nephropathy (MN) via thromboelastography (TEG) and to identify risk factors. 235 MN patients who had undergone TEG examinations from 2011 to 2014 were included. An abnormality in at least two TEG parameters is considered a hypercoagulable state. Patient data was compared between the hypercoagulable and non-hypercoagulable groups. Potential risk factors for hypercoagulability were analyzed by logistic regression models. Subgroup analysis was performed in hypercoagulable patients. Compared to the non-hypercoagulable MN patients, the hypercoagulable patients showed a significantly higher proportion of female patients, urinary protein, platelet count, triglyceride and fibrinogen level, along with more severe hypoproteinemia and a reduction of serum antithrombin III. Correlation analysis showed that hypoproteinemia was the primary risk factor for hypercoagulability in MN patients. Among the hypercoagulable MN patients, a subgroup TEG parameter analysis showed that glucocorticoids-used subgroup and smoker subgroup had shortened time to initial fibrin formation (R value) and increased coagulation index respectively (P < 0.05), indicating a more serious hypercoagulable state. Meanwhile, the time to initial fibrin formation (R value) and time to clot formation (K value) of the statin-used patients were remarkably higher than those of the non-statin patients. TEG examinations facilitated the detection of hypercoagulable states in MN patients, and hypoproteinemia was the most important risk factor for hypercoagulability in these patients. The use of glucocorticoids and smoking may help to aggravate hypercoagulable states, while statin drugs may alleviate hypercoagulability.

Four withanolides (1-4) and two sucrose esters (5, 6) were isolated from the aerial parts of Physalis neomexicana. The structures of 1-6 were elucidated through a variety of spectroscopic techniques. Cytotoxicity studies of the isolates revealed that 2 inhibited human breast cancer cell lines (MDA-MB-231 and MCF-7) with IC50 values of 1.7 and 6.3 μM, respectively.

Unlabelled: Proliferation and migration disorders of vascular smooth muscle cells (VSMCs) contribute to the pathogenesis of proliferative cardiovascular diseases. Although, over the past two decades, a large panel of drugs has been developed for targeting VSMC proliferation, cardiovascular disease remains the leading cause of death worldwide. Thus, there is a compelling need to identify novel signaling pathways and molecules controlling VSMC proliferation and migration, to provide not only mechanistic insights but also safe and effective therapies for the treatment of cardiovascular diseases. Our recent studies have demonstrated that p55γ, a regulatory subunit of phosphoinositide 3-kinase, functions as an endogenous brake on VSMC proliferation. Here, we demonstrate that the small peptide N24, the first 24 amino acids of the NH2 terminus of p55γ, is a functional mimetic which negatively regulates VSMC proliferation and migration. Specifically, luminal delivery of adenovirus expressing N24 or local administration of Tat transactivator protein (TAT)-tagged N24 by pluronic gel alleviates neointimal formation following balloon injury in rat carotid arteries. Enforced expression of N24 suppresses the proliferation and migration of VSMCs induced by serum- or platelet-derived growth factor-BB. Mechanistically, N24 induces cell cycle arrest via activating the p53-p21 signal pathway, without triggering cell death. N24 interacts with and stabilizes p53 by blocking its ubiquitin-dependent degradation, subsequently promotes p21 transcription, and arrests cell cycle progression. Indeed, knockdown of p21 or p53 abrogates the N24-mediated cell growth arrest. Thus, N24 is a p55γ mimetic inhibiting VSMC proliferation as well as migration, thereby conferring important therapeutic implications for anti-proliferative treatment., Key Message: • N24 attenuates balloon injury-induced neointimal formation. • Overexpression of N24 inhibits cultured VSMC proliferation and migration. • Overexpression of N24 arrests the cell cycle at S phase. • N24 interacts with and stabilizes p53 resulting in growth suppression.

Background: Insulin resistance (IR) is present at all stages of chronic kidney disease (CKD) and is associated with CKD progression. Probucol can improve the prognosis of IR in diabetes mellitus (DM) patients. This study aimed to observe the effect of probucol on IR and kidney protection in non-diabetic CKD patients., Methods: This was an open-label, non-placebo-controlled, randomized study. A total of 59 patients were randomized to the probucol group (0.5 g, twice daily) or the control group using a 1: 1 treatment ratio. IR was determined using a homeostatic model assessment-IR (HOMA-IR) index. An Excel database was established to analyze follow-up data at weeks 0, 12, and 24. The primary outcome of interest was changes in the HOMA-IR, and the secondary outcomes of interest were changes in the estimated glomerular filtration rate (eGFR), body mass index (BMI), cholesterol, triglycerides, high-density lipoprotein (HDL), low-density lipoprotein (LDL), and 24-h urinary protein., Results: The HOMA-IR index of the probucol group after 24 weeks was significantly decreased (P < 0.001) compared to the value before treatment (average decrease: 1.45; range: -2.90 to -0.43). The HOMA-IR index in the control group increased (average increase: 0.54; range: -0.38 to 1.87). For the secondary outcomes of interest, the changes between these two groups also exhibited significant differences in eGFR (P = 0.041), cholesterol (P = 0.001), fasting insulin (P < 0.001), and fasting C-peptide (P = 0.001)., Conclusions: Compared to angiotensin receptor blockers alone, the combination with probucol ameliorates IR in non-diabetic CKD patients and delays disease progression.

An analytical method based on high-performance liquid chromatography-photodiode array detection was developed for the simultaneous determination of three anti-proliferative withanolides [withalongolide A ( 1: ), withaferin A ( 2: ) and withalongolide B ( 3: )] present in the aboveground biomass of the long-leaf groundcherry, Physalis longifolia. This method was achieved by biomass extraction followed by chromatographic separation on C18 column eluted with a gradient acetonitrile-water mobile phase. Calibration curves produced satisfactory linear regression (r(2) > 0.9995) for each examined sample. The method was also validated for accuracy, precision and limits of detection and quantification. Such an approach is applicable for the rapid detection and quantitative assessment of withanolides in various P. longifolia accessions., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Aims: Phosphatidylinositol 3 kinases (PI3Ks) play a pivotal role in vascular physiology and pathophysiology. We aimed to investigate the role of p55γ, a regulatory subunit of PI3Ks, in vascular smooth muscle cell (VSMC) proliferation and neointimal formation., Methods and Results: We identified p55γ as an important factor that suppresses VSMC proliferation and injury-evoked neointimal formation. Western blot and mRNA analyses showed that p55γ expression declined in balloon-injured rat carotid arteries and in response to PDGF-BB and serum treatment in cultured VSMCs. Overexpression of p55γ inhibited, whereas short hairpin RNA knockdown of p55γ promoted PDGF-BB- and serum-induced VSMC proliferation. Importantly, in vivo adenoviral gene transfer of p55γ into carotid arteries attenuated, while knockdown of p55γ enhanced balloon injury-induced neointimal formation. Furthermore, p55γ sequentially up-regulated p53 and p21, resulting in cell-cycle arrest in S phase; small-interfering RNA knockdown of either p53 or p21 blocked p55γ-induced VSMC growth arrest. Mechanistically, p55γ interacted with and stabilized p53 protein by blocking mouse double minute 2 homologue-mediated p53 ubiquitination and degradation, subsequently activating its target gene p21. Concurrently, p55γ up-regulated Bcl-xl expression, resulting in non-apoptotic growth arrest effect., Conclusion: These findings mark p55γ as a novel upstream regulator of the p53-p21 signalling pathway that negatively regulates VSMC proliferation, suggesting that malfunction of p55γ may trigger vascular proliferative disorders., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2014. For permissions please email: journals.permissions@oup.com.)

Nine new withanolides (1-9), withahisolides A-I, were isolated along with nine known compounds (10-18) from the aerial parts of Physalis hispida. The structures of 1-9 were elucidated through a variety of spectroscopic techniques, while the structures of 1 and 2 were confirmed by X-ray crystallographic analysis. Compounds 1-3 are the first withanolides with nonaromatic six-membered ring D moieties. In addition, withanolide 8 represents a novel withanolide skeleton due to the absence of a C-13-C-17 bond within the steroidal nucleus.

Withanolides 2,3-dihydrotrechonolide A (1) and 2,3-dihydro-21-hydroxytrechonolide A (2) were isolated along with two known withanolides trechonolide A (3) and jaborosalactone 39 (4) from Jaborosa caulescens var. bipinnatifida (Solanaceae). The structures of 1-2 were elucidated through 2D NMR and other spectroscopic techniques. In addition, the structure of withanolide 1 was confirmed by X-ray crystallographic analysis., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

To date, our work on solanaceous species (Datura wrightii, Jaborosa caulescens, Physalis hispida, Physalis longifolia, Vassobia breviflora and Withania somnifera) has resulted in the isolation of 65 withanolides, 31 of which were new, as well as the semi-synthesis of a further 30 withanolides. Structure identification and MTS assay-based antiproliferative evaluation of these 95 compounds revealed that a Δ(2)-1-oxo functionality in ring A, in conjunction with either a 5β,6β-epoxy or 5α-chloro-6β-hydroxy moiety in ring B, is the minimum structural requirement for withanolides to produce potent cytotoxic activity. Such structure-activity relationship analysis also revealed that oxygenation (the -OH or -OR groups) at C-4, 7, 11 and 12, as well as C-14 to C-28, did not contribute towards the observed antiproliferative activity. Herein, we present a complete overview of our work as it relates to the withanolides reported from 1965 to 2013.

Objective: Osteosarcoma is the most common primary malignancy, mainly arising from the metaphysis of the long bones of adolescents and young adults. Its poor prognosis is strongly associated with invasion and distant metastasis. The calcium-binding protein S100A4 promotes metastasis in several experimental animal models, including osteosarcoma (OS), and S100A4 protein expression is associated with patient outcome in a number of tumor types. In the present study, we investigated the expression of S100A4 and its clinicopathologic significance in OSs., Patients and Methods: S100A4 were examined immunohistochemically in resected OSs from 120 patients with OS to clarify their clinicopathologic significance. Multivariate survival analyses were carried out on all investigated parameters., Results: The immunohistochemical assays revealed that S1004A expression in osteosarcoma tissues was significantly higher than that in corresponding noncancerous bone tissues (p < 0.001). In addition, positive S100A4 expression more frequently occurred in osteosarcoma tissues with advanced clinical stage (p = 0.003), positive distant metastasis (p = 0.001) and poor response to chemotherapy (p = 0.04). In Kaplan-Meier analysis, only S100A4 positively stained cases showed a significantly decreased overall survival time and disease-free survival compared with negatively stained cases (both p < 0.001). On Cox multivariate analysis, positive S100A4 expression was an independent and significant prognostic factor to predict poor overall survival and disease-free survival (both p = 0.001)., Conclusions: Expression of S100A4 protein in OS may be related to the prediction of metastasis potency, response to chemotherapy and poor prognosis for osteosarcoma patients, suggesting that S100A4 may serve as a prognostic marker for the optimization of clinical treatments.

Methanol solutions of the main withanolides (6-8) naturally present in Physalis longifolia yielded five artificial withanolides (1-5), including three new compounds (1-3). Withanolides 1 and 2 were identified as intramolecular Michael addition derivatives, while withanolides 3-5 were the result of intermolecular Michael addition. A comprehensive literature investigation was conducted to identify potential withanolide Michael addition artifacts isolated from Solanaceous species to date.

Low-temperature conditioning of garlic "seed" cloves accelerated the development of the crop cycle, decreased plant growth, and increased the synthesis of phenolic compounds and anthocyanins in the outer scale leaves of the bulbs at harvest time, leading to 3-fold content increase compared with those conditioned at room temperature. Cold conditioning of "seed" cloves also altered the anthocyanin profile during bulb development and at harvest. Two new anthocyanins are reported for the first time in garlic. The high phenolics and anthocyanin contents in bulbs of plants generated from "seed" cloves conditioned at 5 °C for 5 weeks were preceded by overexpression of some putative genes of the phenolic metabolism [6-fold for phenylalanine ammonia lyase (PAL)] and anthocyanin synthesis [1-fold for UDP-sugar:flavonoid 3-O-glycosyltransferase (UFGT)] compared with those conditioned at room temperature.

Insulin resistance is a fundamental pathogenic factor present in various metabolic disorders including obesity and type 2 diabetes. Although skeletal muscle accounts for 70-90% of insulin-stimulated glucose disposal, the mechanism underlying muscle insulin resistance is poorly understood. Here we show in mice that muscle-specific mitsugumin 53 (MG53; also called TRIM72) mediates the degradation of the insulin receptor and insulin receptor substrate 1 (IRS1), and when upregulated, causes metabolic syndrome featuring insulin resistance, obesity, hypertension and dyslipidaemia. MG53 expression is markedly elevated in models of insulin resistance, and MG53 overexpression suffices to trigger muscle insulin resistance and metabolic syndrome sequentially. Conversely, ablation of MG53 prevents diet-induced metabolic syndrome by preserving the insulin receptor, IRS1 and insulin signalling integrity. Mechanistically, MG53 acts as an E3 ligase targeting the insulin receptor and IRS1 for ubiquitin-dependent degradation, comprising a central mechanism controlling insulin signal strength in skeletal muscle. These findings define MG53 as a novel therapeutic target for treating metabolic disorders and associated cardiovascular complications.

Aims: Recent studies show that ischaemic postconditioning (PostC), similar to the well-established ischaemic preconditioning (IPC), confers cardioprotection against ischaemia/reperfusion (IR) injury, and both IPC and PostC can activate the reperfusion injury salvage kinase (RISK) pathway and the survivor activating factor enhancement (SAFE) pathway. PostC is clinically more attractive because of its therapeutic application at the predictable onset of reperfusion. Our previous studies have demonstrated that MG53 is a primary component of the IPC machinery. Here, we investigated the potential role of MG53 in PostC-mediated myocardial protection and explored the underlying mechanism., Methods and Results: Using Langendorff perfusion, we investigated IR injury in wild-type (wt) and MG53-deficient (mg53(-/-)) mouse hearts with or without PostC. IR-induced myocardial damage was markedly exacerbated in mg53(-/-) hearts compared with wt controls. PostC protected wt hearts against IR-induced myocardial infarction, myocyte necrosis, and apoptosis, but failed to protect mg53(-/-) hearts. The loss of PostC protection in mg53(-/-) hearts was attributed to selectively impaired PostC-activated RISK signalling. Mechanistically, MG53 is required for the interaction between caveolin 3 (CaV3) and the p85 subunit of phosphoinositide 3-kinase (p85-PI3K) and PostC-mediated activation of the RISK pathway. Importantly, a structure-function study revealed that the MG53 tripartite motif (TRIM) domain (aa1-284) physically interacted with CaV3 but not p85-PI3K, whereas the MG53 SPRY domain (aa285-477) interacted with p85-PI3K but not CaV3, indicating that MG53 binds to CaV3 and p85 at its N- and C-terminus, respectively., Conclusions: We conclude that MG53 participates in PostC-mediated cardioprotection largely through tethering CaV3 and PI3K and subsequent activation of the RISK pathway.

Plants subjected to postharvest abiotic stresses synthesize secondary metabolites with health-promoting properties. Here, we report the potential use of carrots (Daucus carota) as biofactories of caffeoylquinic acids when subjected to wounding and hyperoxia stresses. Wounding stress induced an increase of ∼287% in total phenolic content (PC) in carrots stored for 48 h at 20 °C. This increase was higher (∼349%) in the wounded tissue treated with hyperoxia stress. To further understand the physiological role of reactive oxygen species (ROS) as a signaling molecule for the stress-induced accumulation of phenolics in carrots, the respiration rate as well as the enzymatic activities of NADPH oxidase, superoxide dismutase, ascorbate peroxidase, and catalase were evaluated. Likewise, shredded carrots were treated with diphenyleneiodonium chloride solution to block NADPH oxidase ROS productions, and the phenylalanine ammonia lyase activity and total PC were evaluated. Results demonstrated that ROS play a key role as a signaling molecule for the stress-induced accumulation of PC in carrots.

Inducible gene expression is a powerful tool to study and engineer genes whose overexpression could be detrimental for the host organisms. However, only limited systems have been adopted in plant biotechnology. We have developed an osmotically inducible system using three components of plant origin, RD29a (Responsive to Dehydration 29A) promoter, CBF3 (C-repeat Binding Factor 3) transcription factor and cpl1-2 (CTD phosphatase-like 1) mutation. The osmotic stress responsible RD29a promoter contains the CBF3 binding sites and thus RD29A-CBF3 feedforward cassette enhances induction of RD29a promoter under stress. The cpl1-2 mutation in a host repressor CPL1 promotes stress responsible RD29a promoter expression. The efficacy of this system was tested using PAP1 (Production of Anthocyanin Pigment 1) transgene, a model transcription factor that regulates the anthocyanin pathway in Arabidopsis. While transgenic plants with only one or two of three components did not reproducibly accumulate anthocyanin pigments above the control level, transgenic cpl1 plants containing homozygous RD29a-PAP1 and RD29a-CBF3 transgenes produced 30-fold higher level of total anthocyanins than control plants upon cold treatment. Growth retardation and phytochemical production of transgenic plants were minimum under normal conditions. The flavonoid profile in cold-induced transgenic plants was determined by LC/MS/MS, which resembled that of previously reported pap1-D plants but enriched for kaempferol derivatives. These results establish the functionality of the inducible three-component gene expression system in plant metabolic engineering. Furthermore, we show that PAP1 and environmental signals synergistically regulate the flavonoid pathway to produce a unique flavonoid blend that has not been produced by PAP1 overexpression or cold treatment alone.

Chemical study of the whole plants of Sarcandra hainanensis yielded two new biflavonoids with a flavan-flavanone skeleton, sarcandrone C (1), D (2) and 6 known compounds (3-8). Structures were elucidated on the basis of NMR spectroscopic methods.

Background: Ischemic heart disease is the greatest cause of death in Western countries. The deleterious effects of cardiac ischemia are ameliorated by ischemic preconditioning (IPC), in which transient ischemia protects against subsequent severe ischemia/reperfusion injury. IPC activates multiple signaling pathways, including the reperfusion injury salvage kinase pathway (mainly PI3K-Akt-glycogen synthase kinase-3beta [GSK3beta] and ERK1/2) and the survivor activating factor enhancement pathway involving activation of the JAK-STAT3 axis. Nevertheless, the fundamental mechanism underlying IPC is poorly understood., Methods and Results: In the present study, we define MG53, a muscle-specific TRIM-family protein, as a crucial component of cardiac IPC machinery. Ischemia/reperfusion or hypoxia/oxidative stress applied to perfused mouse hearts or neonatal rat cardiomyocytes, respectively, causes downregulation of MG53, and IPC can prevent ischemia/reperfusion-induced decrease in MG53 expression. MG53 deficiency increases myocardial vulnerability to ischemia/reperfusion injury and abolishes IPC protection. Overexpression of MG53 attenuates whereas knockdown of MG53 enhances hypoxia- and H(2)O(2)-induced cardiomyocyte death. The cardiac protective effects of MG53 are attributable to MG53-dependent interaction of caveolin-3 with phosphatidylinositol 3 kinase and subsequent activation of the reperfusion injury salvage kinase pathway without altering the survivor activating factor enhancement pathway., Conclusions: These results establish MG53 as a primary component of the cardiac IPC response, thus identifying a potentially important novel therapeutic target for the treatment of ischemic heart disease.

Proliferation of vascular smooth muscle cells (VSMCs) is a primary mechanism underlying cardiovascular proliferative disorders. Phosphoinositide 3-kinase (PI3K)-Akt (or protein kinase B) axis has been assigned at the center of pathways that regulate cell proliferation. Here we demonstrate that enhanced PI3K-Akt signaling by mitogenic stimulation or arterial injury profoundly elevates expression of receptor interacting protein 3 (RIP3) in primary cultured rat VSMCs and in vivo and that the up-regulation of RIP3 leads to VSMC growth arrest and apoptosis via inhibiting the PI3K-Akt signaling pathway, thereby alleviating balloon injury-induced neointimal formation. Specifically, mitogenic stimulation with platelet-derived growth factor-BB or angiotensin II leads to a profound increase in RIP3 expression, which is abolished by inhibition of PI3K or Akt, and increased PI3K-Akt signaling by expression of a constitutively active PI3K mutant also elevates RIP3 expression. Importantly, adenoviral overexpression of RIP3 not only triggers apoptosis but also causes cell cycle arrest at G(1)/G(0) phases that is associated with suppressed Akt activation. In sharp contrast, RIP3 gene silencing enhances serum- and platelet-derived growth factor-induced cell proliferation and Akt activation. In vivo adenoviral gene delivery of rat RIP3 (rRIP3) increased apoptosis and reduced VSMC proliferation, thus, effectively alleviating balloon injury-induced neointimal formation. The growth-suppressive and pro-apoptotic effects are independent of rRIP3 Ser/Thr kinase activity, because overexpression of a kinase-inactive mutant of rRIP3, similar to its wild type, is sufficient to induce growth arrest and apoptosis. These findings reveal a novel growth-suppressive action of RIP3, marking RIP3 as an important factor to prevent excessive mitogenic stimulation- or injury-induced vascular smooth muscle cells hyperplasia.

Rationale: Ca2+/calmodulin-dependent protein kinase (CaMK)II is a multifunctional kinase involved in vital cellular processes such as Ca(2+) handling and cell fate regulation. In mammalian heart, 2 primary CaMKII isoforms, deltaB and deltaC, localize in nuclear and cytosolic compartments, respectively. Although previous studies have established an essential role of CaMKII-deltaC in cardiomyocyte apoptosis, the functional role of the more abundant isoform, CaMKII-deltaB, remains elusive., Objective: Here, we determined the potential role of CaMKII-deltaB in regulating cardiomyocyte viability and explored the underlying mechanism., Methods and Results: In cultured neonatal rat cardiomyocytes, the expression of CaMKII-deltaB and CaMKII-deltaC was inversely regulated in response to H2O2-induced oxidative stress with a profound reduction of the former and an increase of the later. Similarly, in vivo ischemia/reperfusion (IR) led to an opposite regulation of these CaMKII isoforms in a rat myocardial IR model. Notably, overexpression of CaMKII-deltaB protected cardiomyocytes against oxidative stress-, hypoxia-, and angiotensin II-induced apoptosis, whereas overexpression of its cytosolic counterpart promoted apoptosis. Using cDNA microarray, real-time PCR and Western blotting, we demonstrated that overexpression of CaMKII-deltaB but not CaMKII-deltaC elevated expression of heat shock protein (HSP)70 family members, including inducible (i)HSP70 and its homolog (Hst70). Moreover, overexpression of CaMKII-deltaB led to phosphorylation and activation of heat shock factor (HSF)1, the primary transcription factor responsible for HSP70 gene regulation. Importantly, gene silencing of iHSP70, but not Hst70, abolished CaMKII-deltaB-mediated protective effect, indicating that only iHSP70 was required for CaMKII-deltaB elicited antiapoptotic signaling., Conclusions: We conclude that cardiac CaMKII-deltaB and CaMKII-deltaC were inversely regulated in response to oxidative stress and IR injury, and that in contrast to CaMKII-deltaC, CaMKII-deltaB serves as a potent suppressor of cardiomyocyte apoptosis triggered by multiple death-inducing stimuli via phosphorylation of HSF1 and subsequent induction of iHSP70, marking both CaMKII-delta isoforms as promising therapeutic targets for the treatment of ischemic heart disease.

Two new flavonoid dimers (1, 2) consisting of flavan-chalcone together with three known compounds (3-5) were isolated from the ethanol extract of the whole plants of Sarcandra hainanensis. Their structures were determined by extensive spectroscopic analysis. Human immunodeficiency virus-1 integrase inhibition activities of compounds 1 and 2 were evaluated and they showed weak activities with IC(50) at 18.05 and 25.27 muM, respectively.

Progresses in the studies on chemical constituents and bioactivities of Chloranthus S. were systemically reviewed in this article. The whole plants of most species in Chloranthus have long been used as folk medicine in China for detumescence, treating on snake bite and wound. 68 compounds obtained from the plants in Chloranthus fall into the following compound classes: terpenoids, coumarins, amides, organic acids and sterols. Fifty one sesquiterpenes were got. The results of studies on their bioactivities show that most of the plants in this genus have antibacterial and antitumor activities. Further phytochemical and biological studies on the species in Chloranthus are needed for better medicinal utilization.

Two new taxanes were isolated from the MeOH extract of the needles and branches bark of the Japanese yew, Taxus cuspidata. The structures were established as (2alpha,5alpha,7beta,9alpha,10beta,13alpha)-5,10,13,20-tetraacetoxytax-11-ene-2,7,9-triol (1) and (2alpha,5alpha,9alpha,10beta)-2,9,10-triacetoxy-5-[(beta-D-glucopyranosyl)oxy]-3,11-cyclotax-11-en-13-one (2) on the basis of in-depth 1D- and 2D-NMR analyses. Compound 2 is the first example of a transannular taxane glycoside isolated from a natural source.

Objective: To study the chemical constituents in the heartwood of Taxus cuspidata., Method: Silica gel column chromatography, preparative HPLC and preparative TLC were used to isolate and purify the chemical constituents; 1H- and 13C-NMR spectroscopic methods were used for structural identification., Result: Ten compounds, taxinine (1), taxusin (2), beta-sitosterol (3), 1 beta-hydroxybaccatin I (4), 2alpha, 5alpha, 10beta-triacetoxy-14beta-(2'-methyl) butanoyloxy-4 (20), 11-taxadiene (5), 2alpha, 5alpha, 10beta-triacetoxy-14beta-(2'-methyl-3'-hydroxy-butanoyloxyl-4 (20), 11-taxadiene (yunnanxane) (6), 9alpha, 10beta, 13alpha-triacetoxy-5alpha-cinnamoyltaxa-4 (20), 11-diene (7), 2-deacetoxytaxinine J (8), taxezopidine G (9), 2alpha, 7beta, 9alpha, 10beta, 13alpha-pentaacetoxyl-taxa-4 (20), 11-dien-5-ol (5-decinnamoyltaxinine J) (10), were isolated and identified from the heartwood of T. cuspidata., Conclusion: Three taxanes, 1 beta-hydroxybaccatin I (4), 2alpha, 5alpha, 10beta-triacetoxy-14beta-(2'-methyl-3'-hydroxy-butanoyloxy)-4 (20), 11-taxadiene (yunnanxane) (6), and 2alpha, 7beta, 9alpha, 10beta, 13alpha-pentaacetoxyltaxa-4 (20) , 11-dien-5-ol (10), were obtained from this plant for the first time.

Aim: To investigate whether mitochondrial calcium uniporter participates in the cardioprotection of tumor necrosis factor alpha (TNFalpha) pretreatment in isolated rat hearts subjected to ischemia/reperfusion., Methods: Isolated perfused rat hearts were subjected to 30 min regional ischemia (occlusion of left anterior descending artery) and 120 min reperfusion. The infarct size, coronary flow (CF) and lactate dehydrogenase (LDH) release during reperfusion were measured. The mitochondria of the heart were isolated and suspended in the swelling buffer for measurement of absorbance at 520 nm., Results: Pretreatment with TNFa at 10 U/ml for 7 min followed by 10 min washout reduced the infarct size and LDH release, and improved the recovery of CF during reperfusion. Administration of spermine (20 micromol/L), an opener of mitochondrial calcium uniporter, for 10 min during early reperfusion attenuated the reduction of infarct size and LDH release, and improvement of CF induced by TNFalpha. In isolated mitochondria of the heart pretreated with TNFalpha, the absorbance at 520 nm decreased less than that of mitochondria without TNFalpha pretreatment. Administration of spermine (50 micromol/L) attenuated the change of the absorbance induced by TNFalpha., Conclusion: The findings indicate that TNFalpha protects myocardium against ischemia/reperfusion injury via inhibiting mitochondrial calcium uniporter opening as well as mitochondrial permeability transition pore opening.

Aim: Attenuation of mitochondrial Ca2+ ([Ca2+]m), but not cytosolic Ca2+ ([Ca2+]c), overload improves contractile recovery. We hypothesized that attenuation of [Ca2+]m, but not [Ca2+]c, overload confers cardioprotection against ischemia/reperfusion-induced injury., Methods: Infarct size from isolated perfused rat heart, cell viability, and electrically-induced Ca2+ transient in isolated rat ventricular myocytes were measured. We determined the effects of BAPTA-AM, a Ca2+ chelator, at concentrations that abolish the overload of both [Ca2+]c and [Ca2+]m, and ruthenium red, an inhibitor of mitochondrial uniporter of Ca2+ transport, at concentrations that abolish the overload of [Ca2+]m, but not [Ca2+]c, on cardiac injury induced by ischemia/reperfusion., Results: Attenuation of both [Ca2+]m and [Ca2+]c by BAPTA-AM, and attenuation of [Ca2+]m, but not [Ca2+]c, overload by ruthenium red, reduced the cardiac injury observations, indicating the importance of [Ca2+]m in cardioprotection and contractile recovery in response to ischemia/reperfusion., Conclusion: The study has provided unequivocal evidence using a cause-effect approach that attenuation of [Ca2+]m, but not [Ca2+]c, overload is responsible for cardioprotection against ischemia/reperfusion-induced injury. We also confirmed the previous observation that attenuation of [Ca2+]m, but not [Ca2+]c, by ruthenium red improves contractile recovery following ischemia/reperfusion.

Four new epoxy taxoids were isolated from the needles of Taxus cuspidata. Their structures were established as 2a,9a-diacetoxy-5a-cinnamoyloxy-11,12-epoxy-10ss-hydroxytax-4(20)-en-13-one (1), 2a,10ss-diacetoxy-5a-cinnamoyloxy-11,12-epoxy-9a-hydroxytax-4(20)-en-13-one (2), 2a,9a-diacetoxy-11,12-epoxy-10ss,20-dihydroxytax-4-en-13-one (3) and 2a,10ss-diacetoxy-11,12-epoxy-9a,20-dihydroxytax-4-en-13-one (4) on the basis of spectral analysis including 1H-NMR, 13C-NMR, 1H-1H-COSY, HSQC, HMBC and HRFABMS. Compounds 3 and 4 are the first example of 11,12-epoxy taxoids with C-4 double bond found in T. cuspidata.

The objective of the present study was to determine whether the mitochondrial calcium uniporter plays a role in the cardioprotection induced by ischemic preconditioning (IPC). Isolated rat hearts were subjected to 30 min of regional ischemia by ligation of the left anterior descending artery followed by 120 min of reperfusion. IPC was achieved by two 5-min periods of global ischemia separated by 5 min of reperfusion. IPC reduced the infarct size and lactate dehydrogenase release in coronary effluent, which was associated with improved recovery of left ventricular contractility. Treatment with ruthenium red (RR, 5 microM), an inhibitor of the uniporter, or with Ru360 (10 microM), a highly specific uniporter inhibitor, provided cardioprotective effects like those of IPC. The cardioprotection induced by IPC was abolished by spermine (20 microM), an activator of the uniporter. Cyclosporin A (CsA, 0.2 microM), an inhibitor of the mitochondrial permeability transition pore, reversed the effects caused by spermine. In mitochondria isolated from untreated hearts, both Ru360 (10 microM) and RR (1 microM) decreased pore opening, while spermine (20 microM) increased pore opening which was blocked by CsA (0.2 microM). In mitochondria from preconditioned hearts, the opening of the pore was inhibited, but this inhibition did not occur in the mitochondria from hearts treated with IPC plus spermine. These results indicate that the mitochondrial calcium uniporter is involved in the cardioprotection conferred by ischemic preconditioning.

Objective: To determine the possible difference in vasodialtation effect of quercetin and rutin., Methods: The isolated rat thoracic aorta was treated with phenylephrine (PE), and the effects of quercetin and rutin on the preconstricted aorta rings with or without endothelium were determined by organ bath technique. Nitric oxide synthase inhibitor L-N(G)-nitroarginine methyl-ester (L-NAME), guanylyl cyclase inhibitor methylene blue, cyclooxygenase inhibitor indomethacin were used to explore the mechanism., Results: Quercetin (10-160 micromol/L) caused vasorelaxation of aorta rings preconstricted with PE in endothelium-intact and denuded aorta rings in a dose-dependent manner. Rutin(10-160 micromol/L) caused dose-dependent vasorelaxation in endothelium-intact rings preconstricted with phenylephrine, but not in denuded aorta rings. The maximal response (Rmax) values calculated from vasorelaxation curves of quercetin and rutin were (77.20+/-6.11)% and (44.28+/-7.48)%, respectively. There was no difference between median effective concentration (EC(50)) values of quercetin and rutin. Pretreatment with L-NAME (0.1 mmol/L) abolished the vasorelaxation by rutin,but did not influence the vasodilating effect of quercetin in endothelium-intact rings. Pretreatment with methylene blue (10 mmol/L) canceled the vasorelaxation both by quercetin and rutin. Pretreatment with indomethacin (10 micromol/L) attenuated the vasodilatation of quercetin, but did not affect the vascular effect of rutin., Conclusion: The vasodilatation effect of quercetin is more potent than rutin. The vasodilatation effect of quercetin might be mediated by guanylyl cyclase and cyclooxygenase-dependent pathway, while the vasodilatation by rutin might be via nitric oxide-guanylyl cyclase pathway.