Follicle-stimulating hormone receptor expression in advanced atherosclerotic plaques.

Experimental evidence indicates that follicle-stimulating hormone (FSH), an essential hormone for reproduction, can act directly on endothelial cells inducing atherosclerosis activation and development. However, it remains unknown whether the FSH-receptor (FSHR) is expressed in human atherosclerosis plaques. To demonstrate the FSHR presence, we used immunohistochemical and immunoelectron microscopy involving a specific monoclonal antibody FSHR1A02 that recognizes an epitope present in the FSHR-ectodomain. In all 55 patients with atherosclerotic plaques located in carotid, coronary, femoral arteries, and iliac aneurysm, FSHR was selectively expressed in arterial endothelium covering atherosclerotic plaques and endothelia lining intraplaque neovessels. Lymphatic neovessels were negative for FSHR. M1-macrophages, foam cells, and giant multinucleated cells were also FSHR-positive. FSHR was not detected in normal internal thoracic artery. Immunoelectron microscopy performed in ApoEKO/hFSHRKI mice with atherosclerotic plaques, after injection in vivo with mouse anti-hFSHR monoclonal antibody FSHR1A02 coupled to colloidal gold, showed FSHR presence on the luminal surface of arterial endothelial cells covering atherosclerotic plaques. Therefore, FSHR can bind, internalize, and deliver into the plaque circulating ligands to FSHR-positive cells. In conclusion, we report FSHR expression in endothelial cells, M1-macrophages, M1-derived foam cells, giant multinucleated macrophages, and osteoclasts associated with human atherosclerotic plaques.

CCL18 aggravates atherosclerosis by inducing CCR6-dependent T-cell influx and polarization.

Introduction: The CC chemokine ligand 18 (CCL18) is a chemokine highly expressed in chronic inflammation in humans. Recent observations of elevated CCL18 plasma levels in patients with acute cardiovascular syndromes prompted an investigation into the role of CCL18 in the pathogenesis of human and mouse atherosclerosis. Methods and results: CCL18 was profoundly upregulated in ruptured human atherosclerotic plaque, particularly within macrophages. Repeated administration of CCL18 in Western-type diet-fed ApoE -/- mice or PCSK9mut-overexpressing wild type (WT) mice led to increased plaque burden, enriched in CD3+ T cells. In subsequent experimental and molecular modeling studies, we identified CCR6 as a functional receptor mediating CCL18 chemotaxis, intracellular Ca2+ flux, and downstream signaling in human Jurkat and mouse T cells. CCL18 failed to induce these effects in vitro in murine spleen T cells with CCR6 deficiency. The ability of CCR6 to act as CCL18 receptor was confirmed in vivo in an inflammation model, where subcutaneous CCL18 injection induced profound focal skin inflammation in WT but not in CCR6-/- mice. This inflammation featured edema and marked infiltration of various leukocyte subsets, including T cells with a Th17 signature, supporting CCR6's role as a Th17 chemotactic receptor. Notably, focal overexpression of CCL18 in plaques was associated with an increased presence of CCR6+ (T) cells. Discussion: Our studies are the first to identify the CCL18/CCR6 axis as a regulator of immune responses in advanced murine and human atherosclerosis.

Vitamin C as Scavenger of Reactive Oxygen Species during Healing after Myocardial Infarction.

Currently, coronary artery bypass and reperfusion therapies are considered the gold standard in long-term treatments to restore heart function after acute myocardial infarction. As a drawback of these restoring strategies, reperfusion after an ischemic insult and sudden oxygen exposure lead to the exacerbated synthesis of additional reactive oxidative species and the persistence of increased oxidation levels. Attempts based on antioxidant treatment have failed to achieve an effective therapy for cardiovascular disease patients. The controversial use of vitamin C as an antioxidant in clinical practice is comprehensively systematized and discussed in this review. The dose-dependent adsorption and release kinetics mechanism of vitamin C is complex; however, this review may provide a holistic perspective on its potential as a preventive supplement and/or for combined precise and targeted therapeutics in cardiovascular management therapy.

Metabolic reprogramming of immune cells by mitochondrial division inhibitor-1 to prevent post-vascular injury neointimal hyperplasia.

BACKGROUND AND AIMS: New treatments are needed to prevent neointimal hyperplasia that contributes to post-angioplasty and stent restenosis in patients with coronary artery disease (CAD) and peripheral arterial disease (PAD). We investigated whether modulating mitochondrial function using mitochondrial division inhibitor-1 (Mdivi-1) could reduce post-vascular injury neointimal hyperplasia by metabolic reprogramming of macrophages from a pro-inflammatory to anti-inflammatory phenotype. METHODS AND RESULTS: In vivo Mdivi-1 treatment of Apoe-/- mice fed a high-fat diet and subjected to carotid-wire injury decreased neointimal hyperplasia by 68%, reduced numbers of plaque vascular smooth muscle cells and pro-inflammatory M1-like macrophages, and decreased plaque inflammation, endothelial activation, and apoptosis, when compared to control. Mdivi-1 treatment of human THP-1 macrophages shifted polarization from a pro-inflammatory M1-like to an anti-inflammatory M2-like phenotype, reduced monocyte chemotaxis and migration to CCL2 and macrophage colony stimulating factor (M-CSF) and decreased secretion of pro-inflammatory mediators. Finally, treatment of pro-inflammatory M1-type-macrophages with Mdivi-1 metabolically reprogrammed them to an anti-inflammatory M2-like phenotype by inhibiting oxidative phosphorylation and attenuating the increase in succinate levels and correcting the decreased levels of arginine and citrulline. CONCLUSIONS: We report that treatment with Mdivi-1 inhibits post-vascular injury neointimal hyperplasia by metabolic reprogramming macrophages towards an anti-inflammatory phenotype thereby highlighting the therapeutic potential of Mdivi-1 for preventing neointimal hyperplasia and restenosis following angioplasty and stenting in CAD and PAD patients.

Multiomics analyses reveal dynamic bioenergetic pathways and functional remodeling of the heart during intermittent fasting.

Intermittent fasting (IF) has been shown to reduce cardiovascular risk factors in both animals and humans, and can protect the heart against ischemic injury in models of myocardial infarction. However, the underlying molecular mechanisms behind these effects remain unclear. To shed light on the molecular and cellular adaptations of the heart to IF, we conducted comprehensive system-wide analyses of the proteome, phosphoproteome, and transcriptome, followed by functional analysis. Using advanced mass spectrometry, we profiled the proteome and phosphoproteome of heart tissues obtained from mice that were maintained on daily 12- or 16 hr fasting, every-other-day fasting, or ad libitum control feeding regimens for 6 months. We also performed RNA sequencing to evaluate whether the observed molecular responses to IF occur at the transcriptional or post-transcriptional levels. Our analyses revealed that IF significantly affected pathways that regulate cyclic GMP signaling, lipid and amino acid metabolism, cell adhesion, cell death, and inflammation. Furthermore, we found that the impact of IF on different metabolic processes varied depending on the length of the fasting regimen. Short IF regimens showed a higher correlation of pathway alteration, while longer IF regimens had an inverse correlation of metabolic processes such as fatty acid oxidation and immune processes. Additionally, functional echocardiographic analyses demonstrated that IF enhances stress-induced cardiac performance. Our systematic multi-omics study provides a molecular framework for understanding how IF impacts the heart's function and its vulnerability to injury and disease.

In-vivo assessment of vascular injury for the prediction of in-stent restenosis.

BACKGROUND: Despite optimizations of coronary stenting technology, a residual risk of in-stent restenosis (ISR) remains. Vessel wall injury has important impact on the development of ISR. While injury can be assessed in histology, there is no injury score available to be used in clinical practice. METHODS: Seven rats underwent abdominal aorta stent implantation. At 4 weeks after implantation, animals were euthanized, and strut indentation, defined as the impression of the strut into the vessel wall, as well as neointimal growth were assessed. Established histological injury scores were assessed to confirm associations between indentation and vessel wall injury. In addition, stent strut indentation was assessed by optical coherence tomography (OCT) in an exemplary clinical case. RESULTS: Stent strut indentation was associated with vessel wall injury in histology. Furthermore, indentation was positively correlated with neointimal thickness, both in the per-strut analysis (r = 0.5579) and in the per-section analysis (r = 0.8620; both p ≤ 0.001). In a clinical case, indentation quantification in OCT was feasible, enabling assessment of injury in vivo. CONCLUSION: Assessing stent strut indentation enables periprocedural assessment of stent-induced damage in vivo and therefore allows for optimization of stent implantation. The assessment of stent strut indentation might become a valuable tool in clinical practice.

Phospholipids, the Masters in the Shadows during Healing after Acute Myocardial Infarction.

Phospholipids are major components of cell membranes with complex structures, high heterogeneity and critical biological functions and have been used since ancient times to treat cardiovascular disease. Their importance and role were shadowed by the difficulty or incomplete available research methodology to study their biological presence and functionality. This review focuses on the current knowledge about the roles of phospholipids in the pathophysiology and therapy of cardiovascular diseases, which have been increasingly recognized. Used in singular formulation or in inclusive combinations with current drugs, phospholipids proved their positive and valuable effects not only in the protection of myocardial tissue, inflammation and fibrosis but also in angiogenesis, coagulation or cardiac regeneration more frequently in animal models as well as in human pathology. Thus, while mainly neglected by the scientific community, phospholipids present negligible side effects and could represent an ideal target for future therapeutic strategies in healing myocardial infarction. Acknowledging and understanding their mechanisms of action could offer a new perspective into novel therapeutic strategies for patients suffering an acute myocardial infarction, reducing the burden and improving the general social and economic outcome.

Vitamin C Regulates the Profibrotic Activity of Fibroblasts in In Vitro Replica Settings of Myocardial Infarction.

Extracellular collagen remodeling is one of the central mechanisms responsible for the structural and compositional coherence of myocardium in patients undergoing myocardial infarction (MI). Activated primary cardiac fibroblasts following myocardial infarction are extensively investigated to establish anti-fibrotic therapies to improve left ventricular remodeling. To systematically assess vitamin C functions as a potential modulator involved in collagen fibrillogenesis in an in vitro model mimicking heart tissue healing after MI. Mouse primary cardiac fibroblasts were isolated from wild-type C57BL/6 mice and cultured under normal and profibrotic (hypoxic + transforming growth factor beta 1) conditions on freshly prepared coatings mimicking extracellular matrix (ECM) remodeling during healing after an MI. At 10 µg/mL, vitamin C reprogramed the respiratory mitochondrial metabolism, which is effectively associated with a more increased accumulation of intracellular reactive oxygen species (iROS) than the number of those generated by mitochondrial reactive oxygen species (mROS). The mRNA/protein expression of subtypes I, III collagen, and fibroblasts differentiations markers were upregulated over time, particularly in the presence of vitamin C. The collagen substrate potentiated the modulator role of vitamin C in reinforcing the structure of types I and III collagen synthesis by reducing collagen V expression in a timely manner, which is important in the initiation of fibrillogenesis. Altogether, our study evidenced the synergistic function of vitamin C at an optimum dose on maintaining the equilibrium functionality of radical scavenger and gene transcription, which are important in the initial phases after healing after an MI, while modulating the synthesis of de novo collagen fibrils, which is important in the final stage of tissue healing.

Magnesium Improves Cardiac Function in Experimental Uremia by Altering Cardiac Elastin Protein Content.

Cardiovascular complications are accompanied by life-threatening complications and represent the major cause of death in patients with chronic kidney disease (CKD). Magnesium is important for the physiology of cardiac function, and its deficiency is common in CKD. In the present study, we investigated the impact of oral magnesium carbonate supplementation on cardiac function in an experimental model of CKD induced in Wistar rats by an adenine diet. Echocardiographic analyses revealed restoration of impaired left ventricular cardiac function in animals with CKD. Cardiac histology and real-time PCR confirmed a high amount of elastin protein and increased collagen III expression in CKD rats supplemented with dietary magnesium as compared with CKD controls. Both structural proteins are crucial in maintaining cardiac health and physiology. Aortic calcium content increased in CKD as compared with tissue from control animals. Magnesium supplementation numerically lowered the increases in aortic calcium content as it remained statistically unchanged, compared with controls. In summary, the present study provides evidence for an improvement in cardiovascular function and aortic wall integrity in a rat model of CKD by magnesium, as evidenced by echocardiography and histology.

Platelet-instructed SPP1+ macrophages drive myofibroblast activation in fibrosis in a CXCL4-dependent manner.

Fibrosis represents the common end stage of chronic organ injury independent of the initial insult, destroying tissue architecture and driving organ failure. Here we discover a population of profibrotic macrophages marked by expression of Spp1, Fn1, and Arg1 (termed Spp1 macrophages), which expands after organ injury. Using an unbiased approach, we identify the chemokine (C-X-C motif) ligand 4 (CXCL4) to be among the top upregulated genes during profibrotic Spp1 macrophage differentiation. In vitro and in vivo studies show that loss of Cxcl4 abrogates profibrotic Spp1 macrophage differentiation and ameliorates fibrosis after both heart and kidney injury. Moreover, we find that platelets, the most abundant source of CXCL4 in vivo, drive profibrotic Spp1 macrophage differentiation. Single nuclear RNA sequencing with ligand-receptor interaction analysis reveals that macrophages orchestrate fibroblast activation via Spp1, Fn1, and Sema3 crosstalk. Finally, we confirm that Spp1 macrophages expand in both human chronic kidney disease and heart failure.

Endogenous Modulation of Extracellular Matrix Collagen during Scar Formation after Myocardial Infarction.

Myocardial infarction is remains the leading cause of death in developed countries. Recent data show that the composition of the extracellular matrix might differ despite similar heart function and infarction sizes. Because collagen is the main component of the extracellular matrix, we hypothesized that changes in inflammatory cell recruitment influence the synthesis of different collagen subtypes in myofibroblasts, thus changing the composition of the scar. We found that neutrophils sustain the proliferation of fibroblasts, remodeling, differentiation, migration and inflammation, predominantly by IL-1 and PPARγ pathways (n = 3). They also significantly inhibit the mRNA expression of fibrillar collagen, maintaining a reduced stiffness in isolated myofibroblasts (n = 4-5). Reducing the neutrophil infiltration in CCR1-/- resulted in increased mRNA expression of collagen 11, moderate expression of collagen 19 and low expression of collagen 13 and 26 in the scar 4 weeks post infarction compared with other groups (n = 3). Mononuclear cells increased the synthesis of all collagen subtypes and upregulated the NF-kB, angiotensin II and PPARδ pathways (n = 3). They increased the synthesis of collagen subtypes 1, 3, 5, 16 and 23 but reduced the expression of collagens 5 and 16 (n = 3). CCR2-/- scar tissue showed higher levels of collagen 13 (n = 3), in association with a significant reduction in stiffness (n = 4-5). Upregulation of the inflammation-related genes in myofibroblasts mostly modulated the fibrillar collagen subtypes, with less effect on the FACIT, network-forming and globular subtypes (n = 3). The upregulation of proliferation and differentiation genes in myofibroblasts seemed to be associated only with the fibrillar collagen subtype, whereas angiogenesis-related genes are associated with fibrillar, network-forming and multiplexin subtypes. In conclusion, although we intend for our findings to deepen the understanding of the mechanism of healing after myocardial infarction and scar formation, the process of collagen synthesis is highly complex, and further intensive investigation is needed to put together all the missing puzzle pieces in this still incipient knowledge process.

Intraoperative Tumor Detection Using Pafolacianine.

Cancer is a leading cause of death worldwide, with increasing numbers of new cases each year. For the vast majority of cancer patients, surgery is the most effective procedure for the complete removal of the malignant tissue. However, relapse due to the incomplete resection of the tumor occurs very often, as the surgeon must rely primarily on visual and tactile feedback. Intraoperative near-infrared imaging with pafolacianine is a newly developed technology designed for cancer detection during surgery, which has been proven to show excellent results in terms of safety and efficacy. Therefore, pafolacianine was approved by the U.S. Food and Drug Administration (FDA) on 29 November 2021, as an additional approach that can be used to identify malignant lesions and to ensure the total resection of the tumors in ovarian cancer patients. Currently, various studies have demonstrated the positive effects of pafolacianine's use in a wide variety of other malignancies, with promising results expected in further research. This review focuses on the applications of the FDA-approved pafolacianine for the accurate intraoperative detection of malignant tissues. The cancer-targeting fluorescent ligands can shift the paradigm of surgical oncology by enabling the visualization of cancer lesions that are difficult to detect by inspection or palpation. The enhanced detection and removal of hard-to-detect cancer tissues during surgery will lead to remarkable outcomes for cancer patients and society, specifically by decreasing the cancer relapse rate, increasing the life expectancy and quality of life, and decreasing future rates of hospitalization, interventions, and costs.

Promising Therapies for Atrial Fibrillation and Ventricular Tachycardia.

Sudden cardiac death due to arrhythmias, such as atrial fibrillation or ventricular tachycardia, account for 15-20% of all deaths. Myocardial infarction increases the burden of atrial fibrillation and ventricular tachycardia by structural and electrical remodeling of the heart. The current management of new-onset atrial fibrillation includes electric cardioversion with very high conversion rates and pharmacologic cardioversion, with less a than 50% conversion rate. If atrial fibrillation cannot be converted, the focus becomes the control of the symptoms ensuring a constant rhythm and rate control, without considering other contributory factors such as autonomic imbalance. Recently, a huge success was obtained by developing ablation techniques or addressing the vagal nerve stimulation. On the other hand, ventricular tachycardia is more sensitive to drug therapies. However, in cases of non-responsiveness to drugs, the usual therapeutic choice is represented by stereotactic ablative therapy or catheter ablation. This review focuses on these newly developed strategies for treatment of arrhythmias in clinical practice, specifically on vernakalant and low-level tragus stimulation for atrial fibrillation and stereotactic ablative therapy for drug-refractory ventricular tachycardia. These therapies are important for the significant improvement of the management of atrial fibrillation and ventricular tachycardia, providing: (1) a safer profile than current therapies, (2) higher success rate than current solutions, (3) low cost of delivery.

Pro-oxidative priming but maintained cardiac function in a broad spectrum of murine models of chronic kidney disease.

AIMS: Patients with chronic kidney disease (CKD) have an increased risk of cardiovascular events and exhibit myocardial changes including left ventricular (LV) hypertrophy and fibrosis, overall referred to as 'uremic cardiomyopathy'. Although different CKD animal models have been studied for cardiac effects, lack of consistent reporting on cardiac function and pathology complicates clear comparison of these models. Therefore, this study aimed at a systematic and comprehensive comparison of cardiac function and cardiac pathophysiological characteristics in eight different CKD models and mouse strains, with a main focus on adenine-induced CKD. METHODS AND RESULTS: CKD of different severity and duration was induced by subtotal nephrectomy or adenine-rich diet in various strains (C57BL/6J, C57BL/6 N, hyperlipidemic C57BL/6J ApoE-/-, 129/Sv), followed by the analysis of kidney function and morphology, blood pressure, cardiac function, cardiac hypertrophy, fibrosis, myocardial calcification and inflammation using functional, histological and molecular techniques, including cardiac gene expression profiling supplemented by oxidative stress analysis. Intriguingly, despite uremia of variable degree, neither cardiac dysfunction, hypertrophy nor interstitial fibrosis were observed. However, already moderate CKD altered cardiac oxidative stress responses and enhanced oxidative stress markers in each mouse strain, with cardiac RNA sequencing revealing activation of oxidative stress signaling as well as anti-inflammatory feedback responses. CONCLUSION: This study considerably expands the knowledge on strain- and protocol-specific differences in the field of cardiorenal research and reveals that several weeks of at least moderate experimental CKD increase oxidative stress responses in the heart in a broad spectrum of mouse models. However, this was insufficient to induce relevant systolic or diastolic dysfunction, suggesting that additional "hits" are required to induce uremic cardiomyopathy. TRANSLATIONAL PERSPECTIVE: Patients with chronic kidney disease (CKD) have an increased risk of cardiovascular adverse events and exhibit myocardial changes, overall referred to as 'uremic cardiomyopathy'. We revealed that CKD increases cardiac oxidative stress responses in the heart. Nonetheless, several weeks of at least moderate experimental CKD do not necessarily trigger cardiac dysfunction and remodeling, suggesting that additional "hits" are required to induce uremic cardiomyopathy in the clinical setting. Whether the altered cardiac oxidative stress balance in CKD may increase the risk and extent of cardiovascular damage upon additional cardiovascular risk factors and/or events will be addressed in future studies.

Dose-dependent impact of statin therapy intensity on circulating progenitor cells in patients undergoing percutaneous coronary intervention for the treatment of acute versus chronic coronary syndrome.

BACKGROUND: By low-density lipoprotein (LDL) reduction, statins play an important role in cardiovascular risk modification. Incompletely understood pleiotropic statin effects include vasoprotection that might originate from mobilisation and differentiation of vascular progenitor cells. Data on the potentially differential impact of statin treatment intensity on circulating progenitor cells in patients undergoing percutaneous coronary intervention (PCI) are scarce. This study examines the potential association of different permanent statin treatment regimens on circulating progenitor cells in patients with coronary syndrome. METHODS AND RESULTS: In a monocentric prospective all-comers study, 105 consecutive cases scheduled for coronary angiography due to either (A) non-invasive proof of ischemia and chronic coronary syndrome (CCS) or (B) troponin-positive acute coronary syndrome (ACS) were included. According to the 2018 American College of Cardiology Guidelines on Blood Cholesterol, patients were clustered depending on their respective permanent statin treatment regimen in either a high- to moderate-intensity statin treatment (HIST) or a low-intensity statin treatment (LIST) group. Baseline characteristics including LDL levels were comparable. From blood drawn at the time of PCI, peripheral blood mononuclear cells were isolated, cultivated and counted and, by density gradient centrifugation, levels of circulating progenitor cells were determined using fluorescence-activated cell sorting (FACS) analysis. In ACS patients both absolute and relative numbers of circulating early-outgrowth endothelial progenitor cells (EPCs) concurrently were significantly lower in the HIST group as compared to the LIST group. This effect was more pronounced in ACS patients than in CCS patients. Both in ACS and CCS patients, HIST caused a significant reduction of the number of circulating SMPCs. CONCLUSIONS: In patients undergoing PCI, a dose intensity-dependent and LDL level-independent pro-differentiating vasoprotective pleiotropic capacity of statins for EPC and SMPC is demonstrated.

Fibroblast Growth Factor 23 and Outcome Prediction in Patients with Acute Myocardial Infarction.

(1) Background: Fibroblast growth factor 23 (FGF23) is associated with mortality in patients with heart failure (HF); however, less is known about mortality associations in patients with myocardial infarction (MI). (2) Methods: FGF23 was assessed in 180 patients with acute MI, 99 of whom presented with concomitant acute HF. Patients were followed up for one year, and outcome estimates by FGF23 were compared to GRACE score estimates. (3) Results: Log-transformed serum levels of intact FGF23 (logFGF23) did not differ between MI patients with and without HF, and no difference in logFGF23 was observed between 14 MI patients who died and those who survived. However, when only MI patients with concomitant HF were considered, logFGF23 was significantly higher among non-survivors compared to that in survivors. While logFGF23 was not associated with the outcome in the entire cohort, logFGF23 was fairly predictive for one-year mortality in patients with concomitant HF (AUC 0.78; 95%CI 0.61-0.95), where it outperformed GRACE score estimates (AUC 0.70; 95%CI 0.46-0.94). (4) Conclusions: FGF23 was associated with one-year mortality only in MI patients who concomitantly presented with HF, surpassing the predictive ability of GRACE score estimates. No associations were observed in patients without HF despite similar FGF23 levels at admission. Further studies are warranted to investigate whether FGF23 is causal for dismal outcome of HF.

Engagement of the CXCL12-CXCR4 Axis in the Interaction of Endothelial Progenitor Cell and Smooth Muscle Cell to Promote Phenotype Control and Guard Vascular Homeostasis.

Endothelial progenitor cells (EPCs) are involved in vascular repair and modulate properties of smooth muscle cells (SMCs) relevant for their contribution to neointima formation following injury. Considering the relevant role of the CXCL12-CXCR4 axis in vascular homeostasis and the potential of EPCs and SMCs to release CXCL12 and express CXCR4, we analyzed the engagement of the CXCL12-CXCR4 axis in various modes of EPC-SMC interaction relevant for injury- and lipid-induced atherosclerosis. We now demonstrate that the expression and release of CXCL12 is synergistically increased in a CXCR4-dependent mechanism following EPC-SMC interaction during co-cultivation or in response to recombinant CXCL12, thus establishing an amplifying feedback loop Additionally, mechanical injury of SMCs induces increased release of CXCL12, resulting in enhanced CXCR4-dependent recruitment of EPCs to SMCs. The CXCL12-CXCR4 axis is crucially engaged in the EPC-triggered augmentation of SMC migration and the attenuation of SMC apoptosis but not in the EPC-mediated increase in SMC proliferation. Compared to EPCs alone, the alliance of EPC-SMC is superior in promoting the CXCR4-dependent proliferation and migration of endothelial cells. When direct cell-cell contact is established, EPCs protect the contractile phenotype of SMCs via CXCL12-CXCR4 and reverse cholesterol-induced transdifferentiation toward a synthetic, macrophage-like phenotype. In conclusion we show that the interaction of EPCs and SMCs unleashes a CXCL12-CXCR4-based autoregulatory feedback loop promoting regenerative processes and mediating SMC phenotype control to potentially guard vascular homeostasis.

Caffeine prevents restenosis and inhibits vascular smooth muscle cell proliferation through the induction of autophagy.

Caffeine is among the most highly consumed substances worldwide, and it has been associated with decreased cardiovascular risk. Although caffeine has been shown to inhibit the proliferation of vascular smooth muscle cells (VSMCs), the mechanism underlying this effect is unknown. Here, we demonstrated that caffeine decreased VSMC proliferation and induced macroautophagy/autophagy in an in vivo vascular injury model of restenosis. Furthermore, we studied the effects of caffeine in primary human and mouse aortic VSMCs and immortalized mouse aortic VSMCs. Caffeine decreased cell proliferation, and induced autophagy flux via inhibition of MTOR signaling in these cells. Genetic deletion of the key autophagy gene Atg5, and the Sqstm1/p62 gene encoding a receptor protein, showed that the anti-proliferative effect by caffeine was dependent upon autophagy. Interestingly, caffeine also decreased WNT-signaling and the expression of two WNT target genes, Axin2 and Ccnd1 (cyclin D1). This effect was mediated by autophagic degradation of a key member of the WNT signaling cascade, DVL2, by caffeine to decrease WNT signaling and cell proliferation. SQSTM1/p62, MAP1LC3B-II and DVL2 were also shown to interact with each other, and the overexpression of DVL2 counteracted the inhibition of cell proliferation by caffeine. Taken together, our in vivo and in vitro findings demonstrated that caffeine reduced VSMC proliferation by inhibiting WNT signaling via stimulation of autophagy, thus reducing the vascular restenosis. Our findings suggest that caffeine and other autophagy-inducing drugs may represent novel cardiovascular therapeutic tools to protect against restenosis after angioplasty and/or stent placement.

Inhibiting cardiac myeloperoxidase alleviates the relaxation defect in hypertrophic cardiomyocytes.

AIMS: Hypertrophic cardiomyopathy (HCM) is characterized by cardiomyocyte hypertrophy and disarray, and myocardial stiffness due to interstitial fibrosis, which result in impaired left ventricular filling and diastolic dysfunction. The latter manifests as exercise intolerance, angina, and dyspnoea. There is currently no specific treatment for improving diastolic function in HCM. Here, we investigated whether myeloperoxidase (MPO) is expressed in cardiomyocytes and provides a novel therapeutic target for alleviating diastolic dysfunction in HCM. METHODS AND RESULTS: Human cardiomyocytes derived from control-induced pluripotent stem cells (iPSC-CMs) were shown to express MPO, with MPO levels being increased in iPSC-CMs generated from two HCM patients harbouring sarcomeric mutations in the MYBPC3 and MYH7 genes. The presence of cardiomyocyte MPO was associated with higher chlorination and peroxidation activity, increased levels of 3-chlorotyrosine-modified cardiac myosin binding protein-C (MYBPC3), attenuated phosphorylation of MYBPC3 at Ser-282, perturbed calcium signalling, and impaired cardiomyocyte relaxation. Interestingly, treatment with the MPO inhibitor, AZD5904, reduced 3-chlorotyrosine-modified MYBPC3 levels, restored MYBPC3 phosphorylation, and alleviated the calcium signalling and relaxation defects. Finally, we found that MPO protein was expressed in healthy adult murine and human cardiomyocytes, and MPO levels were increased in diseased hearts with left ventricular hypertrophy. CONCLUSION: This study demonstrates that MPO inhibition alleviates the relaxation defect in hypertrophic iPSC-CMs through MYBPC3 phosphorylation. These findings highlight cardiomyocyte MPO as a novel therapeutic target for improving myocardial relaxation associated with HCM, a treatment strategy which can be readily investigated in the clinical setting, given that MPO inhibitors are already available for clinical testing.

Endothelial Progenitor Cells Modulate the Phenotype of Smooth Muscle Cells and Increase Their Neointimal Accumulation Following Vascular Injury.

BACKGROUND: Smooth muscle cells (SMCs) are the main driver of neointima formation and restenosis following vascular injury. In animal models, endothelial progenitor cells (EPCs) accelerate endothelial regeneration and reduce neointima formation after arterial injury; however, EPC-capture stents do not reduce target vessel failure compared with conventional stents. Here we examined the influence of EPCs on features of SMCs pivotal for their impact on injury-induced neointima formation including proliferation, migration, and phenotype switch. METHODS AND RESULTS: EPCs, their conditioned medium, and EPC-derived microparticles induced proliferation of SMCs while limiting their apoptosis. In transwell membrane experiments and scratch assays, EPCs stimulated migration of SMCs and accelerated their recovery from scratch-induced injury. Treatment of SMCs with an EPC-derived conditioned medium or microparticles triggered transformation of SMCs toward a synthetic phenotype. However, co-cultivation of EPCs and SMCs enabling direct cell-cell contacts preserved their original phenotype and protected from the transformative effect of SMC cholesterol loading. Adhesion of EPCs to SMCs was stimulated by SMC injury and reduced by blocking CXCR2 and CCR5. Interaction of EPCs with SMCs modulated their secretory products and synergistically increased the release of selected chemokines. Following carotid wire injury in athymic mice, injection of EPCs resulted not only in reduced neointima formation but also in altered cellular composition of the neointima with augmented accumulation of SMCs. CONCLUSION: EPCs stimulate proliferation and migration of SMCs and increase their neointimal accumulation following vascular injury. Furthermore, EPCs context-dependently modify the SMC phenotype with protection from the transformative effect of cholesterol when a direct cell-cell contact is established.