Treatment of calcific arterial disease via enhancement of autophagy using GSK343.

Vascular calcification is a hallmark of atherosclerotic disease and serves as a strong predictor and risk factor for cardiovascular events. Growing evidence suggests that autophagy may play a protective role in early atherosclerosis. The precise effects of autophagy on VSMC-mediated calcification remain unknown. In this study, we utilized multi-omic profiling to investigate impaired autophagy at the transcriptional level as a key driver of VSMC calcification. Our findings revealed that impaired autophagy is an essential determinant of VSMC calcification. We observed that an osteogenic environment affects the open chromatin status of VSMCs, compromising the transcriptional activation of autophagy initiation genes. In vivo experiments involve pharmacological and genetic activation of autophagy using mouse models of spontaneous large (Mgp-/-) and small (Abcc6-/-) artery calcification. Taken together, these data advance our mechanistic understanding of vascular calcification and provide important insights for a broad range of cardiovascular diseases involving VSMC phenotype switch.

Effects of hypertension on subcortical nucleus morphological alternations in patients with type 2 diabetes.

Objectives: Type 2 diabetes mellitus(T2DM) and hypertension(HTN) are common comorbidities, and known to affect the brain. However, little is known about the effects of the coexisting HTN on brain in T2DM patients. So we aim to investigate the impact of HTN on the subcortical nucleus morphological alternations in T2DM patients. Materials & methods: This work was registered by the clinicaltrials.gov (grant number NCT03564431). We recruited a total of 92 participants, comprising 36 only T2DM patients, 28 T2DM patients with HTN(T2DMH) and 28 healthy controls(HCs) in our study. All clinical indicators were assessed and brain image data was collected for each participant. Voxel-based morphometry(VBM), automatic volume and vertex-based shape analyses were used to determine the subcortical nucleus alternations from each participant's 3D-T1 brain images and evaluate the relationship between the alternations and clinical indicators. Results: T2DMH patients exhibited volumetric reduction and morphological alterations in thalamus compared to T2DM patients, whereas T2DM patients did not demonstrate any significant subcortical alterations compared to HCs. Furthermore, negative correlations have been found between thalamic alternations and the duration of HTN in T2DMH patients. Conclusion: Our results revealed that HTN may exacerbate subcortical nucleus alternations in T2DM patients, which highlighted the importance of HTN management in T2DM patients to prevent further damage to the brain health.

Stimulation of oxalate root exudate in arsenic speciation and fluctuation with phosphate and iron in anoxic mangrove sediment.

Mutual transformations of rhizospheric arsenic (As) in pollution-prone mangrove sediments affected by root exudate oxalate were simulated. This study focuses on the effect of oxalate on As release, mobilization, and phase speciation associated with P and Fe was examined under anoxic conditions in time-dependent changes. Results showed that oxalate addition significantly facilitated As-Fe-P release from As-contaminated mangrove sediments. Sediment As formed the adsorptive and the carbonate-binding fractionations, facilitating the re-adsorption processes. Solution As and As5+ correlated with NaOH-P positively but with NaHCO3-P and HCl-P negatively. Dominant Fe3+ (>84 %) from the amorphous Fe regulated suspension changes and then time-dependent co-precipitation with As and P. Sediment P formed strong complexes with Fe oxides and could be substituted for As via STEM analysis. Oxalate ligand exchange, competitive adsorption of oxalate, and Fe-reduced dissolution are confirmed to involve, allowing for an insight As/P/Fe mobilization and fate in mangrove wetland.

Whole-genome sequencing uncovers two loci for coronary artery calcification and identifies ARSE as a regulator of vascular calcification.

Coronary artery calcification (CAC) is a measure of atherosclerosis and a well-established predictor of coronary artery disease (CAD) events. Here we describe a genome-wide association study (GWAS) of CAC in 22,400 participants from multiple ancestral groups. We confirmed associations with four known loci and identified two additional loci associated with CAC (ARSE and MMP16), with evidence of significant associations in replication analyses for both novel loci. Functional assays of ARSE and MMP16 in human vascular smooth muscle cells (VSMCs) demonstrate that ARSE is a promoter of VSMC calcification and VSMC phenotype switching from a contractile to a calcifying or osteogenic phenotype. Furthermore, we show that the association of variants near ARSE with reduced CAC is likely explained by reduced ARSE expression with the G allele of enhancer variant rs5982944. Our study highlights ARSE as an important contributor to atherosclerotic vascular calcification, and a potential drug target for vascular calcific disease.

Short communication: TNF-α and IGF-1 regulates epigenetic mechanisms of HDAC2 and HDAC10.

Vascular restenosis often presents as a consequence of injury to the vessel wall, resulting from stenting and other interventional procedures. Such injury to the arteries induces proliferation of Vascular Smooth Muscle Cells (VSMCs), resulting in cellular hyperplasia and restenosis. We and others have previously reported de-novo production of different cytokines and growth factors such as Tumor Necrosis Factor Alpha (TNF-α) and Insulin like Growth Factor 1 (IGF-1), after vascular injury. As complex as it is, the profuse proliferation of VSMCs appears to be occurring due to several induced factors which initiate molecular mechanisms and exacerbate disease conditions. In many pathological events, the deleterious effects of TNF-α and IGF-1 in initiating disease mechanisms was reported. In the present work, we explored whether TNF-α and IGF-1 can regulate epigenetic mechanisms that promote proliferation of VSMCs. We investigated the mechanistic roles of proteins which can structurally interact with DNMT1 and initiate cellular pathways that promote proliferation of VSMCs. Our findings here, identify a novel molecular mechanism that is initiated by TNF-α and IGF-1. It was previously reported that DNMT1 expression is directly induced by TNF-α and IGF-1 treatment and increased/induced expression of DNMT1 causes silencing of genes that are essential to maintaining cellular homeostasis such as the tumor suppressor genes. We have earlier reported that TNF-α and IGF-1 treatment elevates DNMT1 expression in VSMCs and causes increased VSMC proliferation. However, the molecular mechanisms involved were not fully deciphered. Interestingly, in the present study we found that TNF-α and IGF-1 treatment failed to elevate DNMT1 expression levels in absence of HDAC2 and HDAC10. Also, while HDAC2 expression was not affected by HDAC10 knockdown, HDAC2 is essentially required for HDAC10 expression. Further, in TNF-α and IGF-1 induced epigenetic signaling mechanism, the expression of two important proteins EZH2 and PCNA seem to be regulated in an HDAC2-HDAC10 dependent manner. Our results show an inter-dependence of epigenetic mediators in inducing proliferation in VSMCs. To our knowledge, this is the first report that shows HDAC2 dependent expression of HDAC10, and suggests a novel mechanistic link between DNMT1, HDAC10 and HDAC2 that regulates EZH2 and PCNA to enhance cell proliferation of VSMCs which is the underlying cause for neointimal hyperplasia and restenosis.

Structural and Functional Analysis of Female Sex Hormones against SARS-CoV-2 Cell Entry.

Emerging evidence suggests that males are more susceptible to severe infection by the SARS-CoV-2 virus than females. A variety of mechanisms may underlie the observed gender-related disparities including differences in sex hormones. However, the precise mechanisms by which female sex hormones may provide protection against SARS-CoV-2 infectivity remains unknown. Here we report new insights into the molecular basis of the interactions between the SARS-CoV-2 spike (S) protein and the human ACE2 receptor. We further report that glycosylation of the ACE2 receptor enhances SARS-CoV-2 infectivity. Importantly, estrogens can disrupt glycan-glycan interactions and glycan-protein interactions between the human ACE2 and the SARS-CoV-2 thereby blocking its entry into cells. In a mouse model of COVID-19, estrogens reduced ACE2 glycosylation and thereby alveolar uptake of the SARS-CoV-2 spike protein. These results shed light on a putative mechanism whereby female sex hormones may provide protection from developing severe infection and could inform the development of future therapies against COVID-19.

Estimating risk of mechanical ventilation and in-hospital mortality among adult COVID-19 patients admitted to Mass General Brigham: The VICE and DICE scores.

BACKGROUND: Risk stratification of COVID-19 patients upon hospital admission is key for their successful treatment and efficient utilization of hospital resources. We sought to evaluate the risk factors on admission (including comorbidities, vital signs, and initial laboratory assessment) associated with ventilation need and in-hospital mortality in COVID-19. METHODS: We established a retrospective cohort of COVID-19 patients from Mass General Brigham hospitals. Demographic, clinical, and admission laboratory data were obtained from electronic medical records of patients admitted to the hospital with laboratory-confirmed COVID-19 before May 19, 2020. Multivariable logistic regression analyses were used to construct and validate the Ventilation in COVID Estimator (VICE) and Death in COVID Estimator (DICE) risk scores. FINDINGS: The entire cohort included 1042 patients (median age, 64 years; 56.8% male). The derivation and validation cohorts for the risk scores included 578 and 464 patients, respectively. We found four factors to be independently predictive for mechanical ventilation requirement (diabetes mellitus, SpO2:FiO2 ratio, C-reactive protein, and lactate dehydrogenase), and 10 factors to be predictors of in-hospital mortality (age, male sex, coronary artery disease, diabetes mellitus, chronic statin use, SpO2:FiO2 ratio, body mass index, neutrophil to lymphocyte ratio, platelet count, and procalcitonin). Using these factors, we constructed the VICE and DICE risk scores, which performed with C-statistics of 0.84 and 0.91, respectively. Importantly, the chronic use of a statin was associated with protection against death due to COVID-19. The VICE and DICE score calculators have been placed on an interactive website freely available to healthcare providers and researchers (https://covid-calculator.com/). INTERPRETATION: The risk scores developed in this study may help clinicians more appropriately determine which COVID-19 patients will need to be managed with greater intensity. FUNDING: COVID-19 Fast Grant (fastgrants.org).

Estimating Risk of Mechanical Ventilation and Mortality Among Adult COVID-19 patients Admitted to Mass General Brigham: The VICE and DICE Scores.

BACKGROUND: Risk stratification of COVID-19 patients upon hospital admission is key for their successful treatment and efficient utilization of hospital resources. OBJECTIVE: To evaluate the risk factors associated with ventilation need and mortality. DESIGN, SETTING AND PARTICIPANTS: We established a retrospective cohort of COVID-19 patients from Mass General Brigham hospitals. Demographic, clinical, and admission laboratory data were obtained from electronic medical records of patients admitted to hospital with laboratory-confirmed COVID-19 before May 19th, 2020. Using patients admitted to Massachusetts General Hospital (MGH, derivation cohort), multivariable logistic regression analyses were used to construct the Ventilation in COVID Estimator (VICE) and Death in COVID Estimator (DICE) risk scores. MEASUREMENTS: The primary outcomes were ventilation status and death. RESULTS: The entire cohort included 1042 patients (median age, 64 years; 56.8% male). The derivation and validation cohorts for the risk scores included 578 and 464 patients, respectively. We found seven factors to be independently predictive for ventilation requirement (diabetes mellitus, dyspnea, alanine aminotransferase, troponin, C-reactive protein, neutrophil-lymphocyte ratio, and lactate dehydrogenase), and 10 factors to be predictors of in-hospital mortality (age, sex, diabetes mellitus, chronic statin use, albumin, C-reactive protein, neutrophil-lymphocyte ratio, mean corpuscular volume, platelet count, and procalcitonin). Using these factors, we constructed the VICE and DICE risk scores, which performed with C-statistics of at least 0.8 in our cohorts. Importantly, the chronic use of a statin was associated with protection against death due to COVID-19. The VICE and DICE score calculators have been placed on an interactive website freely available to the public (https://covid-calculator.com/). LIMITATIONS: One potential limitation is the modest sample sizes in both our derivation and validation cohorts. CONCLUSION: The risk scores developed in this study may help clinicians more appropriately determine which COVID-19 patients will need to be managed with greater intensity.

Structural and functional analysis of female sex hormones against SARS-Cov2 cell entry.

Emerging evidence suggests that males are more susceptible to severe infection by the SARS-CoV-2 virus than females. A variety of mechanisms may underlie the observed gender-related disparities including differences in sex hormones. However, the precise mechanisms by which female sex hormones may provide protection against SARS-CoV-2 infectivity remains unknown. Here we report new insights into the molecular basis of the interactions between the SARS-CoV-2 spike (S) protein and the human ACE2 receptor. We further observed that glycosylation of the ACE2 receptor enhances SARS-CoV-2 infectivity. Importantly estrogens can disrupt glycan-glycan interactions and glycan-protein interactions between the human ACE2 and the SARS-CoV2 thereby blocking its entry into cells. In a mouse model, estrogens reduced ACE2 glycosylation and thereby alveolar uptake of the SARS-CoV-2 spike protein. These results shed light on a putative mechanism whereby female sex hormones may provide protection from developing severe infection and could inform the development of future therapies against COVID-19.

Predictors of mortality in hospitalized COVID-19 patients: A systematic review and meta-analysis.

Mortality rates of coronavirus disease-2019 (COVID-19) continue to rise across the world. Information regarding the predictors of mortality in patients with COVID-19 remains scarce. Herein, we performed a systematic review of published articles, from 1 January to 24 April 2020, to evaluate the risk factors associated with mortality in COVID-19. Two investigators independently searched the articles and collected the data, in accordance with PRISMA guidelines. We looked for associations between mortality and patient characteristics, comorbidities, and laboratory abnormalities. A total of 14 studies documenting the outcomes of 4659 patients were included. The presence of comorbidities such as hypertension (odds ratio [OR], 2.5; 95% confidence interval [CI], 2.1-3.1; P < .00001), coronary heart disease (OR, 3.8; 95% CI, 2.1-6.9; P < .00001), and diabetes (OR, 2.0; 95% CI, 1.7-2.3; P < .00001) were associated with significantly higher risk of death amongst patients with COVID-19. Those who died, compared with those who survived, differed on multiple biomarkers on admission including elevated levels of cardiac troponin (+44.2 ng/L, 95% CI, 19.0-69.4; P = .0006); C-reactive protein (+66.3 µg/mL, 95% CI, 46.7-85.9; P < .00001); interleukin-6 (+4.6 ng/mL, 95% CI, 3.6-5.6; P < .00001); D-dimer (+4.6 µg/mL, 95% CI, 2.8-6.4; P < .00001); creatinine (+15.3 µmol/L, 95% CI, 6.2-24.3; P = .001); and alanine transaminase (+5.7 U/L, 95% CI, 2.6-8.8; P = .0003); as well as decreased levels of albumin (-3.7 g/L, 95% CI, -5.3 to -2.1; P < .00001). Individuals with underlying cardiometabolic disease and that present with evidence for acute inflammation and end-organ damage are at higher risk of mortality due to COVID-19 infection and should be managed with greater intensity.

Non-coding RNAs as Epigenetic Gene Regulators in Cardiovascular Diseases.

Epigenetic gene regulations can be considered as de-novo initiation of abnormal molecular signaling events whose regulation is otherwise required during normal or specific developmental stages of the organisms. Primarily, three different mechanisms have been identified to participate in epigenetic gene regulations which include, DNA methylation, non-coding RNA species (microRNAs [miRNA], and long non-coding RNAs [LNC-RNA]) and histone modifications. These de-novo epigenetic mechanisms have been associated with altered normal cellular functions which eventually facilitate normal cells to transition into an abnormal phenotype. Among the three modes of regulation, RNA species which are usually considered to be less stable, can be speculated to initiate instant alterations in gene expression compared to DNA methylation or histone modifications. However, LNC-RNAs appear to be more stable in the cells than the other RNA species. Moreover, there is increasing literature which clearly suggests that a single specific LNC-RNA can regulate multiple mechanisms and disease phenotypes. With specific focus on cardiovascular diseases, here we attempt to provide UpToDate information on the functional role of miRNAs and LNC-RNAs. Here we discuss the role of these epigenetic mediators in different components of cardiovascular disease which include physiopathological heart development, athersclerosis, retenosis, diabetic hearts, myocardial infarction, ischemia-reperfusion, heart valve disease, aortic aneurysm, osteogenesis, angiogenesis and hypoxia in the heart. While there is abundant literature support that shows the involvement of many LNC-RNAs and miRNAs in cardiovascular diseases, very few RNA species have been identified which regulate epigenetic mechanisms which is the current focus in this article. Understanding the role of these RNA species in regulating epigenetic mechanisms in different cell types causing cardiovascular disease, would advance the field and promote disease prevention approaches that are aimed to target epigenetic mechanisms.

Modulation of Cathepsin L Expression in the Coronary Arteries of Atherosclerotic Swine.

INTRODUCTION: Neointimal hyperplasia (NIH) and restenosis after percutaneous transluminal coronary angioplasty (PTCA) and intravascular stenting remain a problem on a long-term basis by causing endothelial denudation and damage to the intima and media. Vascular sterile inflammation has been attributed to the formation of NIH. Cathepsin L (CTSL), a lysosome protease, is associated with diet-induced atherogenesis. Vitamin D regulates the actions and regulatory effects of proteases and protease inhibitors in different cell types. Objectives of this study are to evaluate the modulatory effect of vitamin D on CTSL activity in post-PTCA coronary arteries of atherosclerotic swine. METHODS: Yucatan microswine were fed with high-cholesterol atherosclerotic diets. The swine were stratified to receive three diets: (1) vitamin D-deficient diet, (2) vitamin D-sufficient diet, and (3) vitamin D-supplement diet. After 6 mo, PTCA was performed in the left circumflex coronary artery (LCx). After 1 y, angiography and optical coherence tomography imaging were performed, and swine was euthanized. Coronary arteries were embedded in paraffin. Tissue sections were stained with hematoxylin and eosin. Expression of Ki67 and CTSL were evaluated by immunofluorescence. RESULTS: Increased number of Ki67 + cells were observed in the postangioplasty LCx in vitamin D-deficient compared with vitamin D-sufficient or vitamin D-supplemented swine. Notably, the expression of CTSL was significantly increased in postangioplasty LCx of vitamin D-deficient swine compared with the vitamin D-sufficient or vitamin D-supplemented animal groups. CONCLUSIONS: Increased expression of CTSL correlates with the formation of NIH in the PTCA-injured coronary arteries. However, in the presence of sufficient or supplemented levels of vitamin D in the blood, CTSL expression was significantly reduced.

Inhibition of DNA methyltransferase-1 instigates the expression of DNA methyltransferase-3a in angioplasty-induced restenosis.

Increased expression of DNA methyltransferase-1 (DNMT1) associates with the progression of many human diseases. Because DNMT1 induces cell proliferation, drugs that inhibit DNMT1 have been used to treat proliferative diseases. Because these drugs are nonspecific inhibitors of DNMT1, subsidiary events or the compensatory mechanisms that are activated in the absence of DNMT1 limit their therapeutic application. Here, we studied the molecular mechanisms that occur during angioplasty-induced restenosis and found that DNMT1 inhibition in both in vitro and in vivo approaches resulted in the induction of DNA methyltransferase-3a (DNMT3a) expression. In vascular smooth muscle cells (VSMCs), the microRNA hsa-miR-1264 mimic, specifically inhibiting DNMT1, induced nuclear expression of DNMT3a. On the contrary, there was no induced expression of DNMT3a in VSMCs that were transfected with hsa-miR-1264 inhibitor. Further, ectopic expression of suppressor of cytokine signaling 3 (SOCS3) through adeno-associated virus (AAV)-mediated gene delivery in the coronary arteries of Yucatan microswine showed inhibition of both DNMT1 and DNMT3a in vivo. These findings show the existence of an inter-regulatory mechanism between DNMT1 and DNMT3a where, in the absence of DNMT1, induction of DNMT3a compensates for the loss of DNMT1 functions, suggesting that the inhibition of both DNMT1 and DNMT3a are required to prevent restenosis.

Cell­specific histone modifications in atherosclerosis (Review).

Histone modifications are the key epigenetic mechanisms that have been identified to regulate gene expression in many human diseases. However, in the early developmental stages, such as in utero and the postnatal stages, histone modifications are essential for gene regulation and cell growth. Atherosclerosis represents a classical example of the involvement of different cell types, and their cumulative effects in the development of atheroma and the progression of the disease. Post translational modifications on proteins either induces their functional activity or renders them inactive. Post translational modifications such as methylation or acetylation on histones have been well characterized, and their role in enhancing or inhibiting specific gene expression was clearly elucidated. In the present review article, the critical roles of different histone modifications that occur in atherosclerosis have been summarized. Different histone proteins have been identified to serve a critical role in the development of atherosclerosis. Specifically, histone methylation and histone acetylation in monocytes, macrophages, vascular smooth muscle cells and in endothelial cells during the progression of atherosclerosis, have been well reported. In recent years, different target molecules and genes that regulate histone modifications have been examined for their effects in the treatment of atherosclerosis in animal models and in clinical trials. An increasing body of evidence suggests that these epigenetic changes resulting from DNA methylation and non­coding RNA may also be associated with histone modifications, thereby indicating that novel therapeutic strategies can be developed by targeting these post translational modifications, which may in turn aid in the treatment of atherosclerosis.

Efficacy of and patient compliance with a ketogenic diet in adults with intractable epilepsy: a meta-analysis.

BACKGROUND AND PURPOSE: Despite the successful use of a ketogenic diet in pediatric epilepsy, its application in adults has been limited. The aim of this meta-analysis was to summarize the findings of relevant published studies in order to identify the efficacy of and compliance with a ketogenic diet and its main subtypes (i.e., classic ketogenic diet and modified Atkins diet) in adults with intractable epilepsy, and to provide useful information for clinical practice. METHODS: Electronic searches of PubMed, EMBASE, Google Scholar, and the ISI Web of Science were conducted to identify studies of the efficacy of and patient compliance with a ketogenic diet in adults with intractable epilepsy; the included studies were reviewed. Meta-analyses were performed using STATA to determine combined efficacy rates and combined rates of compliance with the ketogenic diet and its main subtypes. RESULTS: In total, 12 studies qualified for inclusion, and data from 270 patients were evaluated.The results of the meta-analysis revealed combined efficacy rates of all types of ketogenic diet, a classical ketogenic diet, and a modified Atkins diet were 42%, 52%, and 34%, respectively; the corresponding combined compliance rates were 45%, 38%, and 56%. CONCLUSIONS: The results indicate that a ketogenic diet is a promising complementary therapy in adult intractable epilepsy, and that while a classical ketogenic diet may be more effective, adult patients are likely to be less compliant with it than with a modified Atkins diet.