The immunoproteasome subunit ß2i ameliorates myocardial ischemia/reperfusion injury by regulating Parkin-Mfn1/2-mediated mitochondrial fusion.

Mitochondrial dynamics are critical for maintaining mitochondrial morphology and function during cardiac ischemia and reperfusion (I/R). The immunoproteasome complex is an inducible isoform of the proteasome that plays a key role in modulating inflammation and some cardiovascular diseases, but the importance of immunoproteasome catalytic subunit ß2i (also known as LMP10 or MECL1) in regulating mitochondrial dynamics and cardiac I/R injury is largely unknown. Here, using ß2i-knockout (KO) mice and rAAV9-ß2i-injected mice, we discovered that ß2i expression and its trypsin-like activity were significantly attenuated in the mouse I/R myocardium and in patients with myocardial infarction (MI). Moreover, ß2i-KO mice exhibited greatly enhanced I/R-mediated cardiac dysfunction, infarct size, myocyte apoptosis and oxidative stress accompanied by excessive mitochondrial fission due to Mfn1/2 and Drp1 imbalance. Conversely, cardiac overexpression of ß2i in mice injected with recombinant adeno-associated virus 9 (rAAV9)-ß2i ameliorated cardiac I/R injury. Mechanistically, I/R injury reduced ß2i expression and activity, which increased the expression of the E3 ligase Parkin protein and promoted the degradation of mitofusin 1/2 (Mfn1/2), leading to excessive mitochondrial fission. In conclusion, our data suggest for the first time that ß2i exerts a protective role against cardiac I/R injury and that increasing ß2i expression may be a new therapeutic option for cardiac ischemic disease in clinical practice. Graphical abstract showing how the immunoproteasome subunit ß2i ameliorates myocardial I/R injury by regulating Parkin-Mfn1/2-mediated mitochondrial fusion.

Selective blocking of CXCR2 prevents and reverses atrial fibrillation in spontaneously hypertensive rats.

Atrial fibrillation (AF) is associated with inflammation and oxidative stress. Recently, we demonstrated that the chemokine-receptor CXCR2 plays a critical role in the recruitment of monocytes/macrophages and the development of hypertension and cardiac remodelling. However, the role of CXCR2 in the pathogenesis of hypertensive AF remains unclear. AF was induced in Wistar-Kyoto rats (WKYs) and spontaneously hypertensive rats (SHRs) administered with the CXCR2 inhibitor SB225002. Atrial remodelling, pathological changes and electrophysiology were examined. Our results showed that the chemokine CXCL1 and its receptor CXCR2 were markedly increased in atrial tissue of SHRs compared with WKYs. The administration of SB225002 to SHRs significantly reduced the elevation of blood pressure, AF inducibility and duration, atrial remodelling, recruitment of macrophages, superoxide production and conduction abnormalities compared with vehicle treatment. The administration of SB225002 to SHRs also reversed pre-existing AF development, atrial remodelling, inflammation and oxidative stress. These effects were associated with the inhibition of multiple signalling pathways, including TGF-ß1/Smad2/3, NF-κB-P65, NOX1, NOX2, Kir2.1, Kv1.5 and Cx43. In conclusion, this study provides new evidence that blocking CXCR2 prevents and reverses the development of AF in SHRs, and suggests that CXCR2 may be a potential therapeutic target for hypertensive AF.

Genetic ablation and pharmacological inhibition of immunosubunit ß5i attenuates cardiac remodeling in deoxycorticosterone-acetate (DOCA)-salt hypertensive mice.

Hypertensive cardiac remodeling is a major cause of heart failure. The immunoproteasome is an inducible form of the proteasome and its catalytic subunit ß5i (also named LMP7) is involved in angiotensin II-induced atrial fibrillation; however, its role in deoxycorticosterone-acetate (DOCA)-salt-induced cardiac remodeling remains unclear. C57BL/6 J wild-type (WT) and ß5i knockout (ß5i KO) mice were subjected to uninephrectomy (sham) and DOCA-salt treatment for three weeks. Cardiac function, fibrosis, and inflammation were evaluated by echocardiography and histological analysis. Protein and gene expression levels were analyzed by quantitative real-time PCR and immunoblotting. Our results showed that after 21 days of DOCA-salt treatment, ß5i expression and chymotrypsin-like activity were the most significantly increased factors in the heart compared with the sham control. Moreover, DOCA-salt-induced elevation of blood pressure, adverse cardiac function, chamber and myocyte hypertrophy, interstitial fibrosis, oxidative stress, and inflammation were markedly attenuated in ß5i KO mice. These findings were verified in ß5i inhibitor PR-957-treated mice. Moreover, blocking of PTEN (the gene of phosphate and tensin homolog deleted on chromosome ten) markedly attenuated the inhibitory effect of ß5i knockout on DOCA-salt-induced cardiac remodeling. Mechanistically, DOCA-salt stress upregulated the expression of ß5i, which promoted the degradation of PTEN and the activation of downstream signals (AKT/mTOR, TGF-ß1/Smad2/3, NOX, and NF-κB), which ultimately led to cardiac hypertrophic remodeling. This study provides new evidence of the critical role of ß5i in DOCA-salt-induced cardiac remodeling through the regulation of PTEN stability, and indicates that the inhibition of ß5i may be a promising therapeutic target for the treatment of hypertensive heart diseases.

N-arachidonoylphenolamine alleviates ischaemia/reperfusion-induced cardiomyocyte necroptosis by restoring proteasomal activity.

Necroptosis and apoptosis contribute to the pathogenesis of myocardial ischaemia/reperfusion (I/R) injury and subsequent heart failure. N-arachidonoylphenolamine (AM404) is a paracetamol lipid metabolite that has pleiotropic activity to modulate the endocannabinoid system. However, the protective role of AM404 in modulating I/R-mediated myocardial damage and the underlying mechanism remain largely unknown. A murine I/R model was generated by occlusion of the left anterior descending artery. AM404 (20 mg/kg) was injected intraperitoneally into mice at 2 and 24 h before the I/R operation. Our data revealed that AM404 administration to mice greatly ameliorated I/R-triggered impairment of myocardial performance and reduced infarct area, myocyte apoptosis, oxidative stress and inflammatory response accompanied by the reduction of receptor interacting protein kinase (RIPK)1/3- mixed lineage kinase domain-like (MLKL)-mediated necroptosis and upregulation of the immunosubunits (ß2i and ß5i). In contrast, administration of epoxomicin (a proteasome inhibitor) dramatically abolished AM404-dependent protection against myocardial I/R damage. Mechanistically, AM404 treatment increases ß5i expression, which interacts with Pellino-1 (Peli1), an E3 ligase, to form a complex with RIPK1/3, thereby promoting their degradation, which leads to inhibition of cardiomyocyte necroptosis in the I/R heart. In conclusion, these findings demonstrate that AM404 could prevent cardiac I/R damage and may be a promising drug for the treatment of ischaemic heart disease.

The mechanosensitive Piezo1 channel exacerbates myocardial ischaemia/reperfusion injury by activating caspase-8-mediated PANoptosis.

PANoptosis is a newly discovered type of cell death characterized by pyroptosis, apoptosis and/or necroptosis and has been implicated in the inflammatory response. Piezo1 is a mechanosensitive ion channel that plays important roles in physiological development and various diseases. However, whether cardiomyocytes undergo PANoptosis during myocardial ischaemia/reperfusion (I/R) injury and the role of Piezo1 in this process remain largely unexplored. In this study, our results revealed that the expression levels of the main components of the PANoptosome, including caspase-8, caspase-3, NLRP3, caspase-1, GSDMD, RIPK1, RIPK3 and MLKL, were significantly upregulated in I/R heart tissues over time, indicating the occurrence of PANoptosis in I/R hearts. Accordingly, Piezo1 expression was significantly upregulated in I/R-injured hearts and hypoxia/reoxygenation (H/R)-treated cardiomyocytes. In contrast, pharmacological inhibition of Piezo1 by the inhibitor GsMTx4 in mice markedly attenuated the I/R-mediated decline in cardiac contractile function and increases in infarct size, apoptosis, oxidative stress and inflammation accompanied by the inhibition of PANoptosis-related mediators in I/R hearts. Consistently, the effects of Piezo1 on calcium influx and PANoptosis were further verified by GsMTx4 and Piezo1 activator Yoda1 in H/R-treated cardiomyocytes in vitro. Moreover, caspase-8 rather than calcium influx was required for H/R-induced PANoptosis in vitro. Mechanistically, Piezo1 interacts with caspase-8, a key initial activator of the PANoptosome complex, which subsequently activates cardiomyocyte PANoptosis, leading to cardiac dysfunction. In summary, these data suggest that Piezo1 is a new cardiac mechanosensor that promotes cardiac I/R injury possibly through the caspase-8-mediated activation of cardiomyocyte PANoptosis and highlight that Piezo1 may represent a new target for treating ischaemic heart disease.

Ursolic Acid Ameliorates Myocardial Ischaemia/Reperfusion Injury by Improving Mitochondrial Function via Immunoproteasome-PP2A-AMPK Signalling.

Cardiac ischaemia/reperfusion (I/R) injury causes cardiomyocyte apoptosis and mitochondrial dysfunction. Ursolic acid (UA), as a pentacyclic triterpenoid carboxylic acid, exerts several bioactivities in animal models of different diseases, but the preventive role of UA in I/R-induced myocardial dysfunction remains largely unknown. Male wild-type mice were pre-administered with UA at a dosage of 80 mg/kg i.p. and then subjected to cardiac I/R injury for 24 h. Cardiac function and pathological changes were examined by echocardiography and histological staining. The protein and mRNA levels of the genes were determined using qPCR and immunoblotting analysis. Our results revealed that UA administration in mice significantly attenuated the I/R-induced decline in cardiac function, infarct size, myocyte apoptosis, and oxidative stress. Mechanistically, UA increased three immunoproteasome catalytic subunit expressions and activities, which promoted ubiquitinated PP2A degradation and activated AMPK-PGC1α signalling, leading to improved mitochondrial biosynthesis and dynamic balance. In vitro experiments confirmed that UA treatment prevented hypoxia/reperfusion (H/R)-induced cardiomyocyte apoptosis and mitochondrial dysfunction through activation of AMPK signalling. In summary, our findings identify UA as a new activator of the immunoproteasome that exerts a protective role in I/R-induced myocardial dysfunction and suggest that UA supplementation could be beneficial for the prevention of cardiac ischaemic disease.

TCH-165 attenuates cardiac ischaemia/reperfusion injury by balancing mitochondrial dynamics via increasing proteasome activity.

The proteasome is the main complex responsible for maintaining intracellular protein homeostasis, impairment of which is associated with cardiac ischaemia/reperfusion (I/R) injury. The small molecule TCH-165 has been found to activate the 20S proteasome to remove disordered proteins in multiple myeloma and glioblastoma. However, the preventive effect of TCH-165 against I/R-mediated cardiac impairment in mice remains largely unknown. Here, a cardiac I/R model was established in mice. Heart function was assessed with echocardiography. Cardiac infarction, myocyte death, and superoxide level were evaluated by 2,3,5-triphenyltetrazolium chloride (TTC)-Evans blue staining, terminal deoxynucleotidyl transferase-mediated dUTP nick and labelling (TUNEL) assay and immunostaining, respectively. Our results showed that TCH-165 treatment markedly ameliorated I/R-mediated cardiac dysfunction and decreased the infarct size, apoptosis, and superoxide levels. Mechanistically, TCH-165 increased immunoproteasome subunit expression/activity, increasing pro-fission protein dynamin-1-like protein (DNM1L, also known as DRP1) degradation and the expression of the pro-fusion proteins mitofusin 1/2 (Mfn1/2) and thereby leading to mitochondrial fission/fusion balance. In vitro experiments confirmed that inhibition of proteasome activity by epoxomicin abolished the protective effect of TCH-165 against hypoxia/reoxygenation (H/R)-induced increases in cardiomyocyte apoptosis, superoxide production and mitochondrial fission. In summary, TCH-165 is a newly discovered inducer of immunoproteasome activity that exerts a preventive effect against cardiac I/R damage by targeting Drp1 degradation, indicating that it may be as a potential therapeutic candidate for ischaemic heart disease.

MK-886 protects against cardiac ischaemia/reperfusion injury by activating proteasome-Keap1-NRF2 signalling.

Oxidative stress is considered a key factor contributing to the initiation and development of cardiac injury following ischaemia‒reperfusion (I/R). Arachidonate 5-lipoxygenase (ALOX5) is a rate-limiting enzyme for leukotriene biosynthesis. MK-886 is an inhibitor of ALOX5 that exhibits anti-inflammatory and antioxidant activities. However, the significance of MK-886 in preventing I/R-mediated cardiac injury and the underlying mechanism remain unclear. Cardiac I/R model was produced by ligation/release of the left anterior descending artery. MK-886 (20 mg/kg) was administered intraperitoneally into mice at 1 and 24 h before I/R. Our results indicated that MK-886 treatment significantly attenuated I/R-mediated cardiac contractile dysfunction and decreased the infarct area, myocyte apoptosis, and oxidative stress accompanied with reduction of Kelch-like ECH-associated protein 1 (keap1) and upregulation of nuclear factor erythroid 2-related factor 2 (NRF2). Conversely, administration of the proteasome inhibitor epoxomicin and NRF2 inhibitor ML385 greatly abrogated MK-886-mediated cardioprotection after I/R injury. Mechanistically, MK-886 enhanced the expression of the immunoproteasome subunit ß5i, which interacted with keap1 and enhanced its degradation, leading to activation of the NRF2-dependent antioxidant response and improvement of mitochondrial fusion-fission balance in the I/R-treated heart. In summary, our present findings indicated that MK-886 could protect the heart against I/R injury and highlight that MK-886 may represent a promising therapeutic candidate for preventing ischaemic disease.

Blockage of Fibronectin 1 Ameliorates Myocardial Ischemia/Reperfusion Injury in Association with Activation of AMP-LKB1-AMPK Signaling Pathway.

Myocardial ischemia/reperfusion injury (I/RI) is closely associated with energy substrate metabolism. Fibronectin 1 (Fn1) was markedly elevated in the heart of I/R pigs and ischemic patients, but its role in myocardial I/RI is controversial and the precise mechanism involved remains elusive. Herein, we tested whether blockage of Fn1 with its inhibitor (fibronectin tetrapeptide, RGDS) would alleviate myocardial I/RI. Wild-type (WT) mice were administered with RGDS once 3 h before I/R operation and once at 24 or 48 h postreperfusion, and sacrificed at 24 or 72 h post-I/R, respectively. Cardiac function was evaluated by echocardiography. Myocardial infarction size, apoptosis, fibrosis, and inflammation were examined via histological staining. Uptake of glucose and fatty acids were detected by positron emission tomography (PET) and computer tomography (CT) with [18F]-2-fluoro-2-deoxy-D-glucose (FDG) and [18F]-fluoro-6-thia-heptadecanoic acid (FTHA), respectively. Our results showed that administration of RGDS to mice remarkably limited the I/R-induced myocardial infarct size, myocyte apoptosis, inflammation, oxidative stress, and fibrosis and improved cardiac contractile dysfunction. These protective effects were associated with upregulation of the AMP/ATP ratio and the activation of LKB1-AMPK signaling, which subsequently increased AS160-GLUT4-mediated glucose and fatty acid uptake, improved mitochondrial dynamic imbalance, and inactivated TGF-ß and NF-κB signals in the I/R heart. In conclusion, the current study identified that blocking Fn1 protects against myocardial I/RI likely through activating the LKB1-AMPK-dependent signals and highlights that inhibition of Fn1 may be a novel therapeutic option for treating ischemic heart diseases.

Time Series Transcriptomic Analysis by RNA Sequencing Reveals a Key Role of PI3K in Sepsis-Induced Myocardial Injury in Mice.

Septic cardiomyopathy is the main complication and cause of death of severe sepsis with limited therapeutic strategy. However, the molecular mechanism of sepsis-induced cardiac injury remains unclear. The present study was designed to investigate differentially expressed genes (DEGs) involved in the pathogenesis of septic cardiomyopathy induced by cecal ligation and puncture (CLP) in mice. Male C57BL/6J mice (8-10 weeks old) were subjected to CLP with 21-gauge needles for 24, 48, and 72 h. Myocardial function was assessed by echocardiography. The pathological changes of the heart were evaluated by hematoxylin and eosin as well as immunohistochemical staining. Time series RNA sequencing was utilized to investigate the gene expression profiles. CLP surgery resulted in a significant decrease of animal survival rate and left ventricle contractile function, and an increase in cardiac dilation and infiltration of proinflammatory cells including Mac-2+ macrophages in a time-dependent manner. RNA sequencing identified 5,607 DEGs in septic myocardium at 24, 48, and 72 h after CLP operation. Moreover, gene ontology analysis revealed that these DEGs were mainly associated with the biological processes, including cell adhesion, immune system process, inflammatory response, and positive regulation of cell migration. KEGG pathway enrichment analysis indicated that Staphylococcus aureus infection, osteoclast differentiation, leishmaniasis, and ECM-receptor interaction were significantly altered in septic hearts. Notably, Pik3r1 and Pik3r5 were localized in the center of the gene co-expression network, and were markedly upregulated in CLP-induced septic myocardium. Further, blocking PI3Kγ by the specific inhibitor CZC24832 significantly protected against sepsis-induced cardiac impairment. The present study uncovers the gene expression signatures of CLP-induced myocardial injury and sheds light on the role of Pik3r5 in septic cardiomyopathy.

Time series proteome profile analysis reveals a protective role of citrate synthase in angiotensin II-induced atrial fibrillation.

BACKGROUND: Angiotensin (Ang) II and elevated blood pressure are considered to be the main risk factors for atrial fibrillation. However, the proteome profiles and key mediators/signaling pathways involved in the development of Ang II-induced atrial fibrillation remain unclear. METHODS: Male wild-type C57BL/6 mice (10-week old) were infused with Ang II (2000 ng/kg per min) for 1, 2, or 3 weeks, respectively. Time series proteome profiling of atrial tissues was performed using isobaric tags for relative and absolute quantitation and liquid chromatography coupled with tandem mass spectrometry. RESULTS: We identified a total of 1566 differentially expressed proteins (DEPs) in the atrial tissues at weeks 1, 2, and 3 after Ang II infusion. These DEPs were predominantly involved in mitochondrial oxidation-reduction and tricarboxylic acid cycle in Ang II-infused atria. Moreover, coexpression network analysis revealed that citrate synthase, a rate-limiting enzyme in the tricarboxylic acid cycle, was localized at the center of the mitochondrial oxidation-reduction process, and its expression was significantly downreguated in Ang II-infused atria at different time points. Cardiomyocyte-specific overexpresion of citrate synthase markedly reduced atrial fibrillation susceptibility and atrial remodeling in mice. These beneficial effects were associated with increased ATP production and mitochondrial oxidative phosphorylation system complexes I-V expression and inhibition of oxidative stress. CONCLUSION: The current study defines the dynamic changes of the DEPs involved in Ang II-induced atrial fibrillation, and identifies that citrate synthase plays a protective role in regulating atrial fibrillation development, and increased citrate synthase expression may represent a potential therapeutic option for atrial fibrillation treatment.

Blockage of UCHL1 activity attenuates cardiac remodeling in spontaneously hypertensive rats.

Cardiac remodeling is an important pathological process ultimately leading to heart failure. Ubiquitin carboxy-terminal hydrolase 1 (UCHL1) is a deubiquitinase that plays a critical role in neurodegenerative diseases and cancer. However, its role in cardiac remodeling in spontaneously hypertensive rats remains unclear. Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHRs) were administered the UCHL1 inhibitor LDN-57444 (20 µg/kg/day) from 2 months of age for 4 months. Blood pressure, cardiac hypertrophy, fibrosis, inflammation, and oxidative stress were evaluated by the tail-cuff system, echocardiography, and histological analysis. Gene and protein expression levels were examined by real-time PCR and immunoblotting analysis. At 6 months of age, the expression of UCHL at the mRNA and protein levels was significantly upregulated in SHRs compared with WKYs. Moreover, systolic blood pressure, cardiac performance, left ventricular (LV) hypertrophy, fibrosis, inflammation, and superoxide production were significantly increased in SHRs compared with WKYs, and these effects were markedly attenuated by LDN-57444 after 4 months of administration. These beneficial actions were possibly associated with a reduction in blood pressure and inactivation of multiple signaling pathways, including AKT, ERK1/2, STAT3, calcineurin A, TGF-ß/Smad2/3, and NF-κB. In conclusion, the results indicate that UCHL1 is involved in hypertensive cardiac remodeling in SHRs, and targeting UCHL1 activity may be a novel potential therapeutic approach for the treatment of hypertensive heart diseases.

Chemokine Receptor CXCR-2 Initiates Atrial Fibrillation by Triggering Monocyte Mobilization in Mice.

Atrial fibrillation (AF) is frequently associated with increased inflammatory response characterized by infiltration of monocytes/macrophages. The chemokine receptor CXCR-2 is a critical regulator of monocyte mobilization in hypertension and cardiac remodeling, but it is not known whether CXCR-2 is involved in the development of hypertensive AF. AF was induced by infusion of Ang II (angiotensin II; 2000 ng/kg per minute) for 3 weeks in male C57BL/6 wild-type mice, CXCR-2 knockout mice, bone marrow-reconstituted chimeric mice, and mice treated with the CXCR-2 inhibitor SB225002. Microarray analysis revealed that 4 chemokine ligands of CXCR-2 were significantly upregulated in the atria during 3 weeks of Ang II infusion. CXCR-2 expression and the number of CXCR2+ immune cells markedly increased in Ang II-infused atria in a time-dependent manner. Moreover, Ang II-infused wild-type mice had increased blood pressure, AF inducibility, atrial diameter, fibrosis, infiltration of macrophages, and superoxide production compared with saline-treated wild-type mice, whereas these effects were significantly attenuated in CXCR-2 knockout mice and wild-type mice transplanted with CXCR-2-deficient bone marrow cells or treated with SB225002. Moreover, circulating blood CXCL-1 levels and CXCR2+ monocyte counts were higher and associated with AF in human patients (n=31) compared with sinus rhythm controls (n=31). In summary, this study identified a novel role for CXCR-2 in driving monocyte infiltration of the atria, which accelerates atrial remodeling and AF after hypertension. Blocking CXCR-2 activation may serve as a new therapeutic strategy for AF.

Resveratrol Attenuates Pressure Overload-Induced Cardiac Fibrosis and Diastolic Dysfunction via PTEN/AKT/Smad2/3 and NF-κB Signaling Pathways.

SCOPE: Cardiac fibrosis is a key feature of cardiac remodeling. Recently, a protective role for resveratrol (RES) in pressure-overload-induced cardiac hypertrophy and contractile dysfunction has been demonstrated. However, the effect of RES on cardiac fibrosis and diastolic function in this model remains unclear. METHODS AND RESULTS: Cardiac remodeling is induced in mice by transverse aortic constriction (TAC) for 2-4 weeks. RES is administered at dose of 5 or 50 mg kg-1  d-1 for 2 weeks. It is found that RES administration at 50 mg kg-1  d-1 significantly attenuates TAC-induced adverse cardiac systolic and diastolic function, fibrosis, inflammation, and oxidative stress via inhibiting PTEN degradation and the downstream mediators. However, RES at 5 mg kg-1  d-1 has no significant effects. RES at 50 mg kg-1  d-1 also ameliorates pre-established adverse cardiac function and remodeling induced by TAC. Treatment with PTEN inhibitor VO-OHpic (10 mg kg-1  d-1 ) for 2 weeks abolishes RES-mediated protective effects. Additionally, the effect of RES (100 µm) on inhibition of Ang II-induced fibroblast proliferation and activation in vitro is verified. CONCLUSIONS: The findings provide new evidence that RES plays a critical role in the progression of cardiac fibrosis and diastolic dysfunction, and suggest that RES may be a promising therapeutic agent for cardiac fibrosis.

Chronic inhibition of chemokine receptor CXCR2 attenuates cardiac remodeling and dysfunction in spontaneously hypertensive rats.

System hypertension is a major risk factor for cardiac hypertrophy and heart failure. Our recent findings reveal that the ablation or inhibition of C-X-C chemokine receptor (CXCR) 2 blocks this process in mice; however, it is not clear whether the pharmacological inhibition of CXCR2 attenuates hypertension and subsequent cardiac remodeling in spontaneously hypertensive rats (SHRs). In the present study, we showed that chemokines (CXCL1 and CXCL2) and CXCR2 were significantly upregulated in SHR hearts compared with Wistar-Kyoto rat (WKY) hearts. Moreover, the administration of CXCR2-specific inhibitor N-(2-hydroxy-4-nitrophenyl)-N'-(2-bromophenyl)-urea (SB225002) in SHRs (at 2 months of age) for an additional 4 months significantly suppressed the elevation of blood pressure, cardiac myocyte hypertrophy, fibrosis, inflammation, and superoxide production and improved heart dysfunction in SHRs compared with vehicle-treated SHRs. SB225002 treatment also reduced established hypertension, cardiac remodeling and contractile dysfunction. Moreover, CXCR2-mediated increases in the recruitment of Mac-2-positive macrophages, proinflammatory cytokines, vascular permeability and ROS production in SHR hearts were markedly attenuated by SB225002. Accordingly, the inhibition of CXCR2 by SB225002 deactivates multiple signaling pathways (AKT/mTOR, ERK1/2, STAT3, calcineurin A, TGF-ß/Smad2/3, NF-κB-p65, and NOX). Our results provide new evidence that the chronic blocking of CXCR2 activation attenuates progression of cardiac hypertrophic remodeling and dysfunction in SHRs. These findings may be of value in understanding the benefits of CXCR2 inhibition for hypertensive cardiac hypertrophy and provide further support for the clinical application of CXCR2 inhibitors for the prevention and treatment of heart failure.