Early microvascular coronary endothelial dysfunction precedes pembrolizumab-induced cardiotoxicity. Preventive role of high dose of atorvastatin.

Immune checkpoint inhibitors (ICIs) exhibit remarkable antitumor activity and immune-related cardiotoxicity of unknown pathomechanism. The aim of the study was to investigate the ICI class-dependent cardiotoxicity in vitro and pembrolizumab's (Pem's) cardiotoxicity in vivo, seeking for translational prevention means. Cytotoxicity was investigated in primary cardiomyocytes and splenocytes, incubated with ipilimumab, Pem and avelumab. Pem's cross-reactivity was assessed by circular dichroism (CD) on biotechnologically produced human and murine PD-1 and in silico. C57BL6/J male mice received IgG4 or Pem for 2 and 5 weeks. Echocardiography, histology, and molecular analyses were performed. Coronary blood flow velocity mapping and cardiac magnetic resonance imaging were conducted at 2 weeks. Human EA.hy926 endothelial cells were incubated with Pem-conditioned media from human mononuclear cells, in presence and absence of statins and viability and molecular signaling were assessed. Atorvastatin (20 mg/kg, daily) was administered in vivo, as prophylaxis. Only Pem exerted immune-related cytotoxicity in vitro. Pem's cross-reactivity with the murine PD-1 was confirmed by CD and docking. In vivo, Pem initiated coronary endothelial and diastolic dysfunction at 2 weeks and systolic dysfunction at 5 weeks. At 2 weeks, Pem induced ICAM-1 and iNOS expression and intracardiac leukocyte infiltration. At 5 weeks, Pem exacerbated endothelial activation and triggered cardiac inflammation. Pem led to immune-related cytotoxicity in EA.hy926 cells, which was prevented by atorvastatin. Atorvastatin mitigated functional deficits, by inhibiting endothelial dysfunction in vivo. We established for the first time an in vivo model of Pem-induced cardiotoxicity. Coronary endothelial dysfunction precedes Pem-induced cardiotoxicity, whereas atorvastatin emerges as a novel prophylactic therapy.

Tubulin-folding cofactor E deficiency promotes vascular dysfunction by increased endoplasmic reticulum stress.

AIMS: Assessment of endothelial function in humans by measuring flow-mediated dilation (FMD) risk-stratifies individuals with established cardiovascular disease, whereas its predictive value is limited in primary prevention. We therefore aimed to establish and evaluate novel markers of FMD at the population level. METHODS AND RESULTS: In order to identify novel targets that were negatively correlated with FMD and investigate their contribution to vascular function, we performed a genome-wide association study (GWAS) of 4175 participants of the population based Gutenberg Health Study. Subsequently, conditional knockout mouse models deleting the gene of interest were generated and characterized. GWAS analysis revealed that single-nucleotide polymorphisms (SNPs) in the tubulin-folding cofactor E (TBCE) gene were negatively correlated with endothelial function and TBCE expression. Vascular smooth muscle cell (VSMC)-targeted TBCE deficiency was associated with endothelial dysfunction, aortic wall hypertrophy, and endoplasmic reticulum (ER) stress-mediated VSMC hyperproliferation in mice, paralleled by calnexin up-regulation and exacerbated by the blood pressure hormone angiotensin II. Treating SMMHC-ERT2-Cre+/-TBCEfl/fl mice with the ER stress modulator tauroursodeoxycholic acid amplified Raptor/Beclin-1-dependent autophagy and reversed vascular dysfunction. CONCLUSION: TBCE and tubulin homeostasis seem to be novel predictors of vascular function and offer a new drug target to ameliorate ER stress-dependent vascular dysfunction.

Metabolomics Point out the Effects of Carfilzomib on Aromatic Amino Acid Biosynthesis and Degradation.

(1) Carfilzomib (Cfz) is an antineoplastic agent indicated for the treatment of multiple myeloma. However, its beneficial action is attenuated by the occurrence of cardiotoxicity and nephrotoxicity as the most common adverse effects. Presently, there is well-established knowledge on the pathomechanisms related to these side effects; however, the research on the metabolic alterations provoked by the drug is limited. (2) An in vivo simulation of Cfz-induced toxicity was developed in (i) Cfz-treated and (ii) control mice. An RP-HRMS-based protocol and an advanced statistical treatment were used to investigate the impact of Cfz on the non-polar metabolome. (3) The differential analysis classified the Cfz-treated and control mice and resulted in a significant number of identified biomarkers with AUC > 0.9. The drug impaired the biosynthesis and degradation of aromatic amino acids (AAA) and led to alterations of uremic toxins in the renal and urine levels. Furthermore, the renal degradation of tryptophan was affected, inducing its degradation via the kynurenine pathway. (4) The renal levels of metabolites showed impaired excretion and degradation of AAAs. Cfz was, finally, correlated with the biosynthesis of renal dopamine, explaining the biochemical causes of water and ion retention and the increase in systolic pressure.

Synergistic Pulmonoprotective Effect of Natural Prolyl Oligopeptidase Inhibitors in In Vitro and In Vivo Models of Acute Respiratory Distress Syndrome.

Acute respiratory distress syndrome (ARDS) is a highly morbid inflammatory lung disease with limited pharmacological interventions. The present study aims to evaluate and compare the potential pulmonoprotective effects of natural prolyl oligopeptidase (POP) inhibitors namely rosmarinic acid (RA), chicoric acid (CA), epigallocatechin-3-gallate (EGCG) and gallic acid (GA), against lipopolysaccharide (LPS)-induced ARDS. Cell viability and expression of pro-inflammatory mediators were measured in RAW264.7 cells and in primary murine lung epithelial and bone marrow cells. Nitric oxide (NO) production was also assessed in unstimulated and LPS-stimulated RAW264.7 cells. For subsequent in vivo experiments, the two natural products (NPs) with the most favorable effects, RA and GA, were selected. Protein, cell content and lipid peroxidation levels in bronchoalveolar lavage fluid (BALF), as well as histopathological changes and respiratory parameters were evaluated in LPS-challenged mice. Expression of key mediators involved in ARDS pathophysiology was detected by Western blotting. RA and GA favorably reduced gene expression of pro-inflammatory mediators in vitro, while GA decreased NO production in macrophages. In LPS-challenged mice, RA and GA co-administration improved respiratory parameters, reduced cell and protein content and malondialdehyde (MDA) levels in BALF, decreased vascular cell adhesion molecule-1 (VCAM-1) and the inducible nitric oxide synthase (iNOS) protein expression, activated anti-apoptotic mechanisms and down-regulated POP in the lung. Conclusively, these synergistic pulmonoprotective effects of RA and GA co-administration could render them a promising prophylactic/therapeutic pharmacological intervention against ARDS.

Cardioprotection by selective SGLT-2 inhibitors in a non-diabetic mouse model of myocardial ischemia/reperfusion injury: a class or a drug effect?

Major clinical trials with sodium glucose co-transporter-2 inhibitors (SGLT-2i) exhibit protective effects against heart failure events, whereas inconsistencies regarding the cardiovascular death outcomes are observed. Therefore, we aimed to compare the selective SGLT-2i empagliflozin (EMPA), dapagliflozin (DAPA) and ertugliflozin (ERTU) in terms of infarct size (IS) reduction and to reveal the cardioprotective mechanism in healthy non-diabetic mice. C57BL/6 mice randomly received vehicle, EMPA (10 mg/kg/day) and DAPA or ERTU orally at the stoichiometrically equivalent dose (SED) for 7 days. 24 h-glucose urinary excretion was determined to verify SGLT-2 inhibition. IS of the region at risk was measured after 30 min ischemia (I), and 120 min reperfusion (R). In a second series, the ischemic myocardium was collected (10th min of R) for shotgun proteomics and evaluation of the cardioprotective signaling. In a third series, we evaluated the oxidative phosphorylation capacity (OXPHOS) and the mitochondrial fatty acid oxidation capacity by measuring the respiratory rates. Finally, Stattic, the STAT-3 inhibitor and wortmannin were administered in both EMPA and DAPA groups to establish causal relationships in the mechanism of protection. EMPA, DAPA and ERTU at the SED led to similar SGLT-2 inhibition as inferred by the significant increase in glucose excretion. EMPA and DAPA but not ERTU reduced IS. EMPA preserved mitochondrial functionality in complex I&II linked oxidative phosphorylation. EMPA and DAPA treatment led to NF-kB, RISK, STAT-3 activation and the downstream apoptosis reduction coinciding with IS reduction. Stattic and wortmannin attenuated the cardioprotection afforded by EMPA and DAPA. Among several upstream mediators, fibroblast growth factor-2 (FGF-2) and caveolin-3 were increased by EMPA and DAPA treatment. ERTU reduced IS only when given at the double dose of the SED (20 mg/kg/day). Short-term EMPA and DAPA, but not ERTU administration at the SED reduce IS in healthy non-diabetic mice. Cardioprotection is not correlated to SGLT-2 inhibition, is STAT-3 and PI3K dependent and associated with increased FGF-2 and Cav-3 expression.

Myocardial Protection and Current Cancer Therapy: Two Opposite Targets with Inevitable Cost.

Myocardial protection against ischemia/reperfusion injury (IRI) is mediated by various ligands, activating different cellular signaling cascades. These include classical cytosolic mediators such as cyclic-GMP (c-GMP), various kinases such as Phosphatydilinositol-3- (PI3K), Protein Kinase B (Akt), Mitogen-Activated-Protein- (MAPK) and AMP-activated (AMPK) kinases, transcription factors such as signal transducer and activator of transcription 3 (STAT3) and bioactive molecules such as vascular endothelial growth factor (VEGF). Most of the aforementioned signaling molecules constitute targets of anticancer therapy; as they are also involved in carcinogenesis, most of the current anti-neoplastic drugs lead to concomitant weakening or even complete abrogation of myocardial cell tolerance to ischemic or oxidative stress. Furthermore, many anti-neoplastic drugs may directly induce cardiotoxicity via their pharmacological effects, or indirectly via their cardiovascular side effects. The combination of direct drug cardiotoxicity, indirect cardiovascular side effects and neutralization of the cardioprotective defense mechanisms of the heart by prolonged cancer treatment may induce long-term ventricular dysfunction, or even clinically manifested heart failure. We present a narrative review of three therapeutic interventions, namely VEGF, proteasome and Immune Checkpoint inhibitors, having opposing effects on the same intracellular signal cascades thereby affecting the heart. Moreover, we herein comment on the current guidelines for managing cardiotoxicity in the clinical setting and on the role of cardiovascular confounders in cardiotoxicity.

An Untargeted Metabolomics Approach on Carfilzomib-Induced Nephrotoxicity.

BACKGROUND: Carfilzomib (Cfz) is an anti-cancer drug related to cardiorenal adverse events, with cardiovascular and renal complications limiting its clinical use. Despite the important progress concerning the discovery of the underlying causes of Cfz-induced nephrotoxicity, the molecular/biochemical background is still not well clarified. Furthermore, the number of metabolomics-based studies concerning Cfz-induced nephrotoxicity is limited. METHODS: A metabolomics UPLC-HRMS-DIA methodology was applied to three bio-sample types i.e., plasma, kidney, and urine, obtained from two groups of mice, namely (i) Cfz (8 mg Cfz/ kg) and (ii) Control (0.9% NaCl) (n = 6 per group). Statistical analysis, involving univariate and multivariate tools, was applied for biomarker detection. Furthermore, a sub-study was developed, aiming to estimate metabolites' correlation among bio-samples, and to enlighten potential mechanisms. RESULTS: Cfz mostly affects the kidneys and urine metabolome. Fifty-four statistically important metabolites were discovered, and some of them have already been related to renal diseases. Furthermore, the correlations between bio-samples revealed patterns of metabolome alterations due to Cfz. CONCLUSIONS: Cfz causes metabolite retention in kidney and dysregulates (up and down) several metabolites associated with the occurrence of inflammation and oxidative stress.

Thiol-based redox-active proteins as cardioprotective therapeutic agents in cardiovascular diseases.

Thiol-based redox compounds, namely thioredoxins (Trxs), glutaredoxins (Grxs) and peroxiredoxins (Prxs), stand as a pivotal group of proteins involved in antioxidant processes and redox signaling. Glutaredoxins (Grxs) are considered as one of the major families of proteins involved in redox regulation by removal of S-glutathionylation and thereby reactivation of other enzymes with thiol-dependent activity. Grxs are also coupled to Trxs and Prxs recycling and thereby indirectly contribute to reactive oxygen species (ROS) detoxification. Peroxiredoxins (Prxs) are a ubiquitous family of peroxidases, which play an essential role in the detoxification of hydrogen peroxide, aliphatic and aromatic hydroperoxides, and peroxynitrite. The Trxs, Grxs and Prxs systems, which reversibly induce thiol modifications, regulate redox signaling involved in various biological events in the cardiovascular system. This review focuses on the current knowledge of the role of Trxs, Grxs and Prxs on cardiovascular pathologies and especially in cardiac hypertrophy, ischemia/reperfusion (I/R) injury and heart failure as well as in the presence of cardiovascular risk factors, such as hypertension, hyperlipidemia, hyperglycemia and metabolic syndrome. Further studies on the roles of thiol-dependent redox systems in the cardiovascular system will support the development of novel protective and therapeutic strategies against cardiovascular diseases.

Vascular conditioning prevents adverse left ventricular remodelling after acute myocardial infarction: a randomised remote conditioning study.

AIMS: Remote ischemic conditioning (RIC) alleviates ischemia-reperfusion injury via several pathways, including micro-RNAs (miRs) expression and oxidative stress modulation. We investigated the effects of RIC on endothelial glycocalyx, arterial stiffness, LV remodelling, and the underlying mediators within the vasculature as a target for protection. METHODS AND RESULTS: We block-randomised 270 patients within 48 h of STEMI post-PCI to either one or two cycles of bilateral brachial cuff inflation, and a control group without RIC. We measured: (a) the perfusion boundary region (PBR) of the sublingual arterial microvessels to assess glycocalyx integrity; (b) the carotid-femoral pulse wave velocity (PWV); (c) miR-144,-150,-21,-208, nitrate-nitrite (NOx) and malondialdehyde (MDA) plasma levels at baseline (T0) and 40 min after RIC onset (T3); and (d) LV volumes at baseline and after one year. Compared to baseline, there was a greater PBR and PWV decrease, miR-144 and NOx levels increase (p < 0.05) at T3 following single- than double-cycle inflation (PBR:ΔT0-T3 = 0.249 ± 0.033 vs 0.126 ± 0.034 µm, p = 0.03 and PWV:0.4 ± 0.21 vs -1.02 ± 0.24 m/s, p = 0.03). Increased miR-150,-21,-208 (p < 0.05) and reduced MDA was observed after both protocols. Increased miR-144 was related to PWV reduction (r = 0.763, p < 0.001) after the first-cycle inflation in both protocols. After one year, single-cycle RIC was associated with LV end-systolic volume reduction (LVESV) > 15% (odds-ratio of 3.75, p = 0.029). MiR-144 and PWV changes post-RIC were interrelated and associated with LVESV reduction at follow-up (r = 0.40 and 0.37, p < 0.05), in the single-cycle RIC. CONCLUSION: RIC evokes "vascular conditioning" likely by upregulation of cardio-protective microRNAs, NOx production, and oxidative stress reduction, facilitating reverse LV remodelling. CLINICAL TRIAL REGISTRATION: http://www.clinicaltrials.gov . Unique identifier: NCT03984123.

Molecular mechanisms of carfilzomib-induced cardiotoxicity in mice and the emerging cardioprotective role of metformin.

Carfilzomib (Cfz), an irreversible proteasome inhibitor licensed for relapsed/refractory myeloma, is associated with cardiotoxicity in humans. We sought to establish the optimal protocol of Cfz-induced cardiac dysfunction, to investigate the underlying molecular-signaling and, based on the findings, to evaluate the cardioprotective potency of metformin (Met). Mice were randomized into protocols 1 and 2 (control and Cfz for 1 and 2 consecutive days, respectively); protocols 3 and 4 (control and alternate doses of Cfz for 6 and 14 days, respectively); protocols 5A and 5B (control and Cfz, intermittent doses on days 0, 1 [5A] and 0, 1, 7, and 8 [5B] for 13 days); protocols 6A and 6B (pharmacological intervention; control, Cfz, Cfz+Met and Met for 2 and 6 days, respectively); and protocol 7 (bortezomib). Cfz was administered at 8 mg/kg (IP) and Met at 140 mg/kg (per os). Cfz resulted in significant reduction of proteasomal activity in heart and peripheral blood mononuclear cells in all protocols except protocols 5A and 5B. Echocardiography demonstrated that Cfz led to a significant fractional shortening (FS) depression in protocols 2 and 3, a borderline dysfunction in protocols 1 and 4, and had no detrimental effect on protocols 5A and 5B. Molecular analysis revealed that Cfz inhibited AMPKα/mTORC1 pathways derived from increased PP2A activity in protocol 2, whereas it additionally inhibited phosphatidylinositol 3-kinase/Akt/endothelial nitric oxide synthase pathway in protocol 3. Coadministration of Met prevented Cfz-induced FS reduction and restored AMPKα phosphorylation and autophagic signaling. Conclusively, Cfz decreased left ventricular function through increased PP2A activity and inhibition of AMPKα and its downstream autophagic targets, whereas Met represents a novel promising intervention against Cfz-induced cardiotoxicity.

Elucidating Carfilzomib's Induced Cardiotoxicity in an In Vivo Model of Aging: Prophylactic Potential of Metformin.

BACKGROUND: Carfilzomib is a first-line proteasome inhibitor indicated for relapsed/refractory multiple myeloma (MM), with its clinical use being hampered by cardiotoxic phenomena. We have previously established a translational model of carfilzomib cardiotoxicity in young adult mice, in which metformin emerged as a prophylactic therapy. Considering that MM is an elderly disease and that age is an independent risk factor for cardiotoxicity, herein, we sought to validate carfilzomib's cardiotoxicity in an in vivo model of aging. METHODS: Aged mice underwent the translational two- and four-dose protocols without and with metformin. Mice underwent echocardiography and were subsequently sacrificed for molecular analyses in the blood and cardiac tissue. RESULTS: Carfilzomib decreased proteasomal activity both in PBMCs and myocardium in both protocols. Carfilzomib induced mild cardiotoxicity after two doses and more pronounced cardiomyopathy in the four-dose protocol, while metformin maintained cardiac function. Carfilzomib led to an increased Bip expression and decreased AMPKα phosphorylation, while metformin coadministration partially decreased Bip expression and induced AMPKα phosphorylation, leading to enhanced myocardial LC3B-dependent autophagy. CONCLUSION: Carfilzomib induced cardiotoxicity in aged mice, an effect significantly reversed by metformin. The latter possesses translational importance as it further supports the clinical use of metformin as a potent prophylactic therapy.

Investigating the Vascular Toxicity Outcomes of the Irreversible Proteasome Inhibitor Carfilzomib.

BACKGROUND: Carfilzomib's (Cfz) adverse events in myeloma patients include cardiovascular toxicity. Since carfilzomib's vascular effects are elusive, we investigated the vascular outcomes of carfilzomib and metformin (Met) coadministration. METHODS: Mice received: (i) saline; (ii) Cfz; (iii) Met; (iv) Cfz+Met for two consecutive (acute) or six alternate days (subacute protocol). Leucocyte-derived reactive oxygen species (ROS) and serum NOx levels were determined and aortas underwent vascular and molecular analyses. Mechanistic experiments were recapitulated in aged mice who received similar treatment to young animals. Primary murine (prmVSMCs) and aged human aortic smooth muscle cells (HAoSMCs) underwent Cfz, Met and Cfz+Met treatment and viability, metabolic flux and p53-LC3-B expression were measured. Experiments were recapitulated in AngII, CoCl2 and high-glucose stimulated HAoSMCs. RESULTS: Acutely, carfilzomib alone led to vascular hypo-contraction and increased ROS release. Subacutely, carfilzomib increased ROS release without vascular manifestations. Cfz+Met increased PGF2α-vasoconstriction and LC3-B-dependent autophagy in both young and aged mice. In vitro, Cfz+Met led to cytotoxicity and autophagy, while Met and Cfz+Met shifted cellular metabolism. CONCLUSION: Carfilzomib induces a transient vascular impairment and oxidative burst. Cfz+Met increased vascular contractility and synergistically induced autophagy in all settings. Therefore, carfilzomib cannot be accredited for a permanent vascular dysfunction, while Cfz+Met exert vasoprotective potency.

Hydroxytyrosol ameliorates metabolic, cardiovascular and liver changes in a rat model of diet-induced metabolic syndrome: Pharmacological and metabolism-based investigation.

Metabolic syndrome is a clustering of interrelated risk factors for cardiovascular disease and diabetes. The Mediterranean diet has been proposed as an important dietary pattern to confer cardioprotection by attenuating risk factors of metabolic syndrome. Hydroxytyrosol (HT) is present in olive fruit and oil, which are basic constituents of the Mediterranean diet. In this study, we have shown that treatment with HT (20mg/kg/d for 8 weeks) decreased adiposity, improved impaired glucose and insulin tolerance, improved endothelial function with lower systolic blood pressure, decreased left ventricular fibrosis and resultant diastolic stiffness and reduced markers of liver damage in a diet-induced rat model of metabolic syndrome. These results were accompanied by reduced infiltration of monocytes/macrophages into the heart with reduced biomarkers of oxidative stress. Furthermore, in an HRMS-based metabolism study of HT, we have identified 24 HT phase I and II metabolites, six of them being over-produced in high-starch, low-fat diet fed rats treated with HT compared to obese rats on high-carbohydrate, high-fat diet. These results provide direct evidence for cardioprotective effects of hydroxytyrosol by attenuation of metabolic risk factors. The implications of altered metabolism of HT in high-carbohydrate, high-fat diet fed obese rats warrant further investigation.

Cardioprotection by H2S Donors: Nitric Oxide-Dependent and ­Independent Mechanisms.

Hydrogen sulfide (H2S) is a signaling molecule with protective effects in the cardiovascular system. To harness the therapeutic potential of H2S, a number of donors have been developed. The present study compares the cardioprotective actions of representative H2S donors from different classes and studies their mechanisms of action in myocardial injury in vitro and in vivo. Exposure of cardiomyocytes to H2O2 led to significant cytotoxicity, which was inhibited by sodium sulfide (Na2S), thiovaline (TV), GYY4137 [morpholin-4-ium 4 methoxyphenyl(morpholino) phosphinodithioate], and AP39 [(10-oxo-10-(4-(3-thioxo-3H-1,2-dithiol5yl)phenoxy)decyl) triphenylphospho-nium bromide]. Inhibition of nitric oxide (NO) synthesis prevented the cytoprotective effects of Na2S and TV, but not GYY4137 and AP39, against H2O2-induced cardiomyocyte injury. Mice subjected to left anterior descending coronary ligation were protected from ischemia-reperfusion injury by the H2S donors tested. Inhibition of nitric oxide synthase (NOS) in vivo blocked only the beneficial effect of Na2S. Moreover, Na2S, but not AP39, administration enhanced the phosphorylation of endothelial NOS and vasodilator-associated phosphoprotein. Both Na2S and AP39 reduced infarct size in mice lacking cyclophilin-D (CypD), a modulator of the mitochondrial permeability transition pore (PTP). Nevertheless, only AP39 displayed a direct effect on mitochondria by increasing the mitochondrial Ca(2+) retention capacity, which is evidence of decreased propensity to undergo permeability transition. We conclude that although all the H2S donors we tested limited infarct size, the pathways involved were not conserved. Na2S had no direct effects on PTP opening, and its action was nitric oxide dependent. In contrast, the cardioprotection exhibited by AP39 could result from a direct inhibitory effect on PTP acting at a site different than CypD.

Effects of the olive tree leaf constituents on myocardial oxidative damage and atherosclerosis.

The olive (Olea europaea) leaf is considered an important traditional herbal medicine utilized against infectious diseases, and for the treatment of diabetes and hypertension. Moreover, olive leaf constituents have been related to cardioprotection, probably due to their association with cellular redox modulating effects. The pathogenesis of certain common diseases, including those of the cardiovascular system, involves oxidative stress and tissue inflammation. Olive polyphenolic compounds, such as oleuropein, hydroxytyrosol, or tyrosol, possess antioxidant, anti-inflammatory, antiatherosclerotic, anti-ischemic, and hypolipidemic effects on the myocardium as demonstrated by various in vitro and in vivo studies. In this review article, we summarize the current knowledge on the role of the olive leaf constituents in the prevention of cardiac dysfunction and highlight future perspectives in their use as cardioprotective agents in therapeutics.

The natural olive constituent oleuropein induces nutritional cardioprotection in normal and cholesterol-fed rabbits: comparison with preconditioning.

Ischemic preconditioning, which is mediated by cell signaling molecules, protects the heart from ischemia-reperfusion injury by limiting the infarct size. Oleuropein, the main polyphenolic constituent of olives, reduced the infarct size in normal and cholesterol-fed rabbits when it was administered at a nutritional dose. The aim of the present study was to compare the effects of oleuropein and preconditioning in terms of the cell signaling and metabolism pathways underlying myocardial protection. Rabbits were randomly divided into six groups: the control group received 5 % dextrose for six weeks, the preconditioning group was subjected to two cycles of preconditioning with 5 min ischemia/10 min reperfusion, the O6 group was treated with oleuropein for six weeks, the Chol group was fed a cholesterol-enriched diet and 5 % dextrose for six weeks, and the CholO6 and CholO3 groups were treated with cholesterol and oleuropein for six and three weeks, respectively; oleuropein was dissolved in 5 % dextrose solution and was administered orally at a dose of 20 mg × kg(-1) × day(-1). All animals were subsequently subjected to 30 min myocardial ischemia followed by 10 min of reperfusion. At that time, myocardial biopsies were taken from the ischemic areas for the assessment of oxidative and nitrosative stress biomarkers (malondialdehyde and nitrotyrosine), and determination of phosphorylation of signaling molecules involved in the mechanism of preconditioning (PI3K, Akt, eNOS, AMPK, STAT3). The tissue extracts NMR metabolic profile was recorded and further analyzed by multivariate statistics. Oxidative biomarkers were significantly reduced in the O6, CholO6, and CholO3 groups compared to the control, preconditioning, and Chol groups. Considering the underlying signaling cascade, the phosphorylation of PI3K, Akt, eNOS, AMPK, and STAT-3 was significantly higher in the preconditioning and all oleuropein-treated groups compared to the control and Chol groups. The NMR-based metabonomic study, performed through the analysis of spectroscopic data, depicted differences in the metabolome of the various groups with significant alterations in purine metabolism. In conclusion, the addition of oleuropein to a normal or hypercholesterolemic diet results in a preconditioning-like intracellular effect, eliminating the deleterious consequences of ischemia and hypercholesterolemia, followed by a decrease of oxidative stress biomarkers. This effect is exerted through inducing preconditioning-involved signaling transduction. Nutritional preconditioning may support the low cardiovascular morbidity and mortality associated with the consumption of olive products.

Implications and hidden toxicity of cardiometabolic syndrome and early-stage heart failure in carfilzomib-induced cardiotoxicity.

BACKGROUND AND PURPOSE: Cancer therapy-related cardiovascular adverse events (CAEs) in presence of comorbidities, are in the spotlight of the cardio-oncology guidelines. Carfilzomib (Cfz), indicated for relapsed/refractory multiple myeloma (MM), presents with serious CAEs. MM is often accompanied with co-existing comorbidities. However, Cfz use in MM patients with cardiometabolic syndrome (CMS) or in heart failure with reduced ejection fraction (HFrEF), is questionable. EXPERIMENTAL APPROACH: ApoE-/- and C57BL6/J male mice received 14 weeks Western Diet (WD) (CMS models). C57BL6/J male mice underwent permanent LAD ligation for 14 days (early-stage HFrEF model). CMS- and HFrEF-burdened mice received Cfz for two consecutive or six alternate days. Daily metformin and atorvastatin administrations were performed additionally to Cfz, as prophylactic interventions. Mice underwent echocardiography, while proteasome activity, biochemical and molecular analyses were conducted. KEY RESULTS: CMS did not exacerbate Cfz left ventricular (LV) dysfunction, whereas Cfz led to metabolic complications in both CMS models. Cfz induced autophagy and Ca2+ homeostasis dysregulation, whereas metformin and atorvastatin prevented Cfz-mediated LV dysfunction and molecular deficits in the CMS-burdened myocardium. Early-stage HFrEF led to depressed LV function and increased protein phosphatase 2A (PP2A) activity. Cfz further increased myocardial PP2A activity, inflammation and Ca2+-cycling dysregulation. Metformin co-administration exerted an anti-inflammatory potential on the myocardium without improving LV function. CONCLUSION AND IMPLICATIONS: CMS and HFrEF seem to exacerbate Cfz-induced CAEs, by presenting metabolism-related hidden toxicity and PP2A-related cardiac inflammation, respectively. Metformin retains its prophylactic potential in the presence of CMS, while mitigating inflammation and Ca2+ signalling dysregulation in the HFrEF myocardium.

Antihypertensive Potential of Pistacia lentiscus var. Chia: Molecular Insights and Therapeutic Implications.

Background: Hypertension poses a significant global health burden and is associated with cardiovascular morbidity. Chios mastic gum (CMG), derived from Pistacia lentiscus var. Chia, shows potential as a phytotherapeutic agent, due to its multifaceted beneficial effects. However, its anti-hypertensive effects and vascular, circulatory, and renal-related dysfunction, have not been thoroughly investigated. Herein, we aimed to explore the antihypertensive potential of CMG, focusing on vascular and renal endothelium, in vivo. Methods: Two models of hypertension in male rats, induced by Angiotensin II and Deoxycorticosterone acetate (DOCA)-high-salt administration, were utilized. CMG was administered at 220 mg/kg daily for four weeks after hypertension onset and blood pressure was measured non-invasively. Whole blood RNA sequencing, metabolomics, real-time PCR, and Western blot analyses of kidney and aorta tissues were additionally performed. Results: CMG significantly lowered systolic, diastolic, and mean blood pressure in both models. RNA sequencing revealed that CMG modulated immunity in the Angiotensin II model and metabolism in the DOCA-HS model. CMG downregulated genes related to oxidative stress and endothelial dysfunction and upregulated endothelial markers such as Vegfa. Metabolomic analysis indicated improved endothelial homeostasis via lysophosphatidylinositol upregulation. Conclusions: CMG emerges as a potent natural antihypertensive therapy, demonstrating beneficial effects on blood pressure and renal endothelial function.

Myeloid cell-derived interleukin-6 induces vascular dysfunction and vascular and systemic inflammation.

Aims: The cytokine interleukin-6 (IL-6) plays a central role in the inflammation cascade as well as cardiovascular disease progression. Since myeloid cells are a primary source of IL-6 formation, we aimed to generate a mouse model to study the role of myeloid cell-derived IL-6 in vascular disease. Methods and results: Interleukin-6-overexpressing (IL-6OE) mice were generated and crossed with LysM-Cre mice, to generate mice (LysM-IL-6OE mice) overexpressing the cytokine in myeloid cells. Eight- to 12-week-old LysM-IL-6OE mice spontaneously developed inflammatory colitis and significantly impaired endothelium-dependent aortic relaxation, increased aortic reactive oxygen species (ROS) formation, and vascular dysfunction in resistance vessels. The latter phenotype was associated with decreased survival. Vascular dysfunction was accompanied by a significant accumulation of neutrophils, monocytes, and macrophages in the aorta, increased myeloid cell reactivity (elevated ROS production), and vascular fibrosis associated with phenotypic changes in vascular smooth muscle cells. In addition to elevated Mcp1 and Cxcl1 mRNA levels, aortae from LysM-IL-6OE mice expressed higher levels of inducible NO synthase and endothelin-1, thus partially accounting for vascular dysfunction, whereas systemic blood pressure alterations were not observed. Bone marrow (BM) transplantation experiments revealed that vascular dysfunction and ROS formation were driven by BM cell-derived IL-6 in a dose-dependent manner. Conclusion: Mice with conditional overexpression of IL-6 in myeloid cells show systemic and vascular inflammation as well as endothelial dysfunction. A decrease in circulating IL-6 levels by replacing IL-6-producing myeloid cells in the BM improved vascular dysfunction in this model, underpinning the relevant role of IL-6 in vascular disease.