Zymosan A Improved Doxorubicin-Induced Ventricular Remodeling by Evoking Heightened Cardiac Inflammatory Responses and Healing in Mice.

Background Doxorubicin-induced myocardial injury is reflected by the presence of vacuolization in both clinical and animal models. The lack of scar tissue to replace the vacuolizated cardiomyocytes indicates that insufficient cardiac inflammation and healing occurred following doxorubicin injection. Whether improved macrophage activity by zymosan A (zymosan) ameliorates doxorubicin-induced ventricular remodeling in mice is unknown. Methods and Results Mice were intravenously injected with vehicle or doxorubicin (5 mg/kg per week, 4 weeks), and cardiac structure and function were assessed by echocardiography. Two distinct macrophage subsets in hearts following doxorubicin injection were measured at different time points by flow cytometry. Moreover, cardiomyocyte vacuolization, capillary density, collagen content, and ventricular tensile strength were assessed. The therapeutic effect of zymosan (3 mg/kg, single injection) on doxorubicin-induced changes in the aforementioned parameters was determined. At the cellular level, the polarization of monocytes to proinflammatory or reparative macrophages were measured, with or without doxorubicin (0.25 and 0.5 µmol/L). Doxorubicin led to less proinflammatory and reparative macrophage infiltration in the heart in the early phase, with decreased cardiac capillary density and collagen III in the chronic phase. In cell culture, doxorubicin (0.5 µmol/L) repressed macrophage transition toward both proinflammatory and reparative subset. Zymosan enhanced both proinflammatory and reparative macrophage infiltration in doxorubicin-injected hearts, evoking a heightened acute inflammatory response. Zymosan alleviated doxorubicin-induced cardiomyocyte vacuolization in the chronic phase, in parallel with enhanced collagen content, capillary density, and ventricular tensile strength. Conclusions Zymosan improved cardiac healing and ameliorated doxorubicin-induced ventricular remodeling and dysfunction by activating macrophages at an optimal time.

The pathogenesis of aging-induced left atrial appendage thrombus formation and cardioembolic stroke in mice is influenced by inflammation-derived matrix metalloproteinases.

Elderly people without atrial fibrillation (AF) still have a high incidence of cardioembolic stroke, suggesting that thrombus formation within the left atrial appendage (LAA) may also occur in an AF-independent manner. In the present study, we explored the potential mechanisms for aging-induced LAA thrombus formation and stroke in mice. We monitored stroke events in 180 aging male mice (14-24 months) and assessed left atrium (LA) remodeling by echocardiography at different ages. Mice that had stroke were implanted with telemeters to confirm AF. Histological features of LA and LAA thrombi were examined, as well as collagen content, expression of matrix metalloproteinases (MMPs), and leukocyte density in the atria at different ages, in mice with or without stroke. Also, the effects of MMP inhibition on stroke incidence and atrial inflammation were tested. We detected 20 mice (11 %) with stroke, 60 % of which were within 18-19 months of age. Although we did not detect AF in mice with stroke, we detected the presence of LAA thrombi, suggesting that stroke originated from the hearts of these mice. Compared with 18-month-old mice without stroke, 18-month-old stroke mice had enlarged LA with a very thin endocardium, that was associated with less collagen and heightened MMP expression in the atria. During aging, we found that the expression of mRNAs for atrial MMP7, MMP8, and MMP9 peaked at 18 months, which closely correlated with reductions in collagen content and the time-window for cardioembolic stroke in these mice. Treatment of mice with an MMP inhibitor at 17-18 months of age reduced atrial inflammation and remodeling, and stroke incidence. Taken together, our study demonstrates that aging-induced LAA thrombus formation occurs through a mechanism involving upregulation of MMPs and breakdown of collagen, and that treatment with an MMP inhibitor may be effective as a treatment strategy for this heart condition.

The predictive value of changes in left atrial volume index for rehospitalization in heart failure with preserved ejection fraction.

AIMS: Left atrial volume index (LAVI) is an adequate analysis to predicate the left ventricle (LV) filling pressures, providing a powerful predictive marker of LV diastolic dysfunction. LAVI is a dynamic morphophysiological marker, and whether LAVI changes can predicate clinical outcomes in HF with preserved ejection fraction (HFpEF) is unknown. METHODS: HFpEF patients were retrospectively studied from the First Affiliated Hospital of Dalian Medical University. Patients were classified into deteriorated, stable and improved groups according to the change in LAVI. Rehospitalization was defined as the main endpoint, the composite outcome of rehospitalization or all-cause death was defined as the secondary endpoint. RESULTS: A total of 409 patients were included. In this cohort, the percentage of deteriorated, stable, and improved LAVI were 99 (24.2%), 235 (57.4%), and 75 (18.4%), respectively. During the 22 months follow-up period, 168 patients (41.1%) were rehospitalized, 31 patients (7.5%) died and 182 patients (44.5%) experienced a composite outcome. Multivariate Cox regression showed that compared to improved LAVI, those with deteriorated and stable LAVI experienced higher risk of rehospitalization. Logistic regression showed atrial fibrillation (AF) and higher creatinine were independent predictors of deteriorated LAVI, whereas the use of loop diuretics, calcium channel blockers (CCB), and high level of high-density lipoprotein cholesterol (HDL-C) were significantly associated with improved LAVI. CONCLUSIONS: Change in LAVI provides a powerful and dynamic morphophysiological marker of LV filling status and can be used to evaluate the rehospitalization in HFpEF patients.

Leucine Supplementation in Middle-Aged Male Mice Improved Aging-Induced Vascular Remodeling and Dysfunction via Activating the Sirt1-Foxo1 Axis.

Vascular aging is associated with metabolic remodeling, and most studies focused on fatty acid and glucose metabolism. Based on our metabolomic data, leucine was significantly reduced in the aortas of aged mice. Whether leucine supplementation can reverse aging-induced vascular remodeling remains unknown. To investigate the effectiveness of leucine, male mice at 15 or 18 months were supplemented with leucine (1.5%) for 3 months. All the aged mice, with or without leucine, were sacrificed at 21 months. Blood pressure and vascular relaxation were measured. H&E, Masson's trichrome, and Elastica van Gieson staining were used to assess aortic morphology. Vascular inflammation, reactive oxidative stress (ROS), and vascular smooth muscle cell (VSMC) phenotype were also measured in mouse aortas. Compared with the 21-month-old mice without leucine, leucine supplementation from 15 months significantly improved vascular relaxation, maintained the contractile phenotype of VSMCs, and repressed vascular inflammation and ROS levels. These benefits were not observed in the mice supplemented with leucine starting from 18 months, which was likely due to the reduction in leucine transporters Slc3a2 or Slc7a5 at 18 months. Furthermore, we found benefits from leucine via activating the Sirt1-induced Foxo1 deacetylation. Our findings indicated that leucine supplementation in middle-aged mice improved aging-induced vascular remodeling and dysfunction.

Anthracycline-Induced Atrial Structural and Electrical Remodeling Characterizes Early Cardiotoxicity and Contributes to Atrial Conductive Instability and Dysfunction.

Aims: Cancer patients treated with anthracyclines are susceptible to atrial fibrillation (AF), while the mechanisms remain unclear. Due to sudden and unpredictable features, prediction of anthracycline-induced AF at early phase is difficult. Clinically, we tested whether anthracycline-induced early atrial remodeling in patients could be detected by echocardiography. Experimentally, we investigated the mechanisms of doxorubicin-induced atrial remodeling and AF in mice, and the protective effects of dexrazoxane and antioxidants. Methods and Results: Postsurgery breast cancer patients with an anthracycline-containing or anthracycline exclusion regimen were recruited for echocardiography before chemotherapy, and 3 and 6 months after chemotherapy. Mice were injected with doxorubicin or vehicle (5 mg/kg/week, 4 weeks), and left atrial diameter, electrical transmission, and AF inducibility were measured. Meanwhile, the level of reactive oxygen species (ROS), activity of antioxidant enzymes, cardiomyocyte size, vacuolization, inflammation, and fibrosis were also measured in mouse atria. The therapeutic effects of dexrazoxane and antioxidants on doxorubicin-induced changes in the aforementioned parameters were also determined. While ventricular parameters and functions were unchanged in cancer patients receiving anthracyclines before and after chemotherapy, left atrial reservoir and conduit function were decreased at 3 months postchemotherapy versus prechemotherapy. Doxorubicin-induced susceptibility to AF occurred in mice before onset of ventricular dysfunction. Doxorubicin-induced AF was via inducing structural remodeling (cardiomyocyte death, hypotrophy, and vacuolization) and electrical remodeling (reduction and redistribution of connexin 43) in atria, which was effectively prevented by dexrazoxane or antioxidants through inhibiting ROS generation or enhancing ROS elimination. Innovation and Conclusion: AF inducibility was induced after doxorubicin injection, which can be inhibited by repressing the ROS level. Antioxid. Redox Signal. 37, 19-39. The Clinical Trial Registration number is PJ-KS-KY-2019-73.

Circulating Soluble Suppression of Tumorigenicity 2 Predicts Recurrence After Radiofrequency Ablation of Persistent Atrial Fibrillation.

Objective: A more extensively fibrotic left atrium contributes to atrial fibrillation (AF) occurrence, persistence, and recurrence. The soluble suppression of tumorigenicity 2 (sST2) has emerged as a ventricular fibrotic biomarker for patients with heart failure. The present study is to investigate associations between circulating sST2 and risk of recurrence after ablation in AF patients. Methods: We measured the baseline plasma level of sST2 from patients with persistent AF (n = 117) and paroxysmal AF (n = 93) patients. Patients were followed up for 15 months after ablation. The relationship between circulating sST2 and recurrence was assessed by multivariable Cox regression. The cutoff value of sST2 was determined by receiver operating characteristic curve. The relationship between baseline sST2 level and left atrial volume index (LAVI) was assessed by multivariate linear regression analysis. Serial sST2 measurements were also conducted after 24 h, 6 months, and 15 months of ablation. ST2 localization was examined in left atrial appendages of persistent AF patients by immunohistochemistry and Western blot. Results: Baseline sST2 positively associated with LAVI in the persistent AF group, and elevated sST2 (≥39.25 ng/ml) independently increased the risk of recurrence after ablation (area under the curve = 0.748), with hazard ratio of 1.038 (95% confidence interval 1.017-1.060, P < 0.001) when adjusted for co-variables. In contrast, elevated sST2 cannot predict recurrence in paroxysmal AF. Conclusions: In persistent AF patients, increased sST2 serves as a marker of recurrence after radiofrequency ablation. Patients with sST2 ≥ 39.25 ng/ml are more likely to develop recurrence within a year.

A Three-Dimensional Microfluidic Device for Monitoring Cancer and Chemotherapy-Associated Platelet Activation.

Platelet activation and the risk of thrombosis are increased in cancer patients, especially after chemotherapy. Our previous studies indicated that chemotherapy-induced platelet activation is largely due to endothelial cell damage. Thus, simple in vitro tests, such as aggregometry, are not desirable tests to predict platelet responsiveness to different chemotherapeutic agents because other contributory factors, such as tumor cells, endothelial cells, and the flow rate of platelets, also contribute to the formation of cancer-associated thrombosis. Therefore, developing a platelet detection system, which includes all possible risk parameters, is necessary. In the present study, we described a microengineered microfluidic system that contained a drug concentration generator, cancer cell culture chip, and three-dimensional (3D) circular microvascular model covered with a confluent endothelial layer and perfused with human platelets at a stable flow rate. Doxorubicin was injected through two injection sites. Endothelial cell injury was evaluated by counting, cell cytoskeleton observation, and the level of IACM1 and ET-1 in endothelial cells or a culture medium. Prestained platelets were perfused into the artificial blood vessel, and platelet-endothelial cell adhesion was measured. We found that (i) MCF7 cell-released factors had a cytotoxicity effect on both endothelial cells and platelets. (ii) We confirmed that doxorubicin-induced platelet activation was endothelial cell-dependent. (iii) A lower dosage of doxorubicin (0-2.0 µM) induced platelet activation, while a higher dosage of doxorubicin (2.0-4.0 µM) led to platelet death. Our findings indicated that platelet-endothelial cell adhesion could be used as a diagnostic marker of platelet activation, providing a simple and rapid detective way to predict platelet responsiveness before or during chemotherapy.

Platelets Promote Ang II (Angiotensin II)-Induced Atrial Fibrillation by Releasing TGF-ß1 (Transforming Growth Factor-ß1) and Interacting With Fibroblasts.

Hypertension is a risk factor of atrial fibrillation (AF), and a certain number of patients with hypertension were found with an enlarged left atrium. Platelet activation is found in patients with hypertension or pressure overload/Ang II (angiotensin II)-induced hypertensive animal models and contribute to ventricular fibrosis. Whether hypertension-induced atrial fibrosis is mediated by platelets remains unknown. Our previous experimental data showed that platelet-derived TGF-ß1 (transforming growth factor-ß1) was reduced in patients with hypertensive AF. The present study is to investigate whether platelet-derived TGF-ß1 promotes Ang II-induced atrial fibrosis and AF. Platelet activation and atrial platelet accumulation were measured in sinus rhythm controls, normotensive AF, and patients with hypertensive AF. Ang II (1500 ng/kg per minute, 3 weeks) infused mice with pharmacological (clopidogrel) and genetic platelet inhibition (TGF-ß1 deletion in platelets) were used. Platelet activation, atrial structural remodeling, atrial electrical transmission, AF inducibility, inflammation, and fibrosis were measured in mice. We found that circulating platelets were activated in patients with hypertensive AF. A large amount of platelet was accumulated in the atriums of patients with hypertensive AF. Both clopidogrel treatment and platelet-specific deletion of TGF-ß1 attenuated Ang II-induced structural remodeling, atrial electrical transmission, AF inducibility, as well as atrial inflammation and fibrosis than mice without interventions. Furthermore, clopidogrel blocked atrial platelet accumulation and platelet-fibroblast conjugation. Platelets promoted atrial fibroblast differentiation in cell culture. Profibrotic actions of platelets are largely via activation of atrial fibroblasts by releasing TGF-ß1 and inducing platelet-fibroblast conjugation, and platelet inhibition is sufficient to inhibit atrial fibrosis and AF inducibility.

Doxorubicin contributes to thrombus formation and vascular injury by interfering with platelet function.

In clinical studies, platelet aggregation and risk of thrombosis are increased in patients after doxorubicin treatment. However, the exact role of doxorubicin in platelet functions and thrombus formation in vivo remain unclear. The present study is to investigate the role of doxorubicin in platelet function in relation to thrombus formation and vascular toxicity, as well as the efficacy of antiplatelet therapy. Mice were treated with doxorubicin or vehicle (5 mg/kg iv, 4 wk), and the following parameters were determined: platelet count and size, platelet surface adhesive receptors by flow cytometry, density of granules by electron microscopy, platelet aggregation and degranulation at resting or agonist-stimulated state, platelet adhesion on fibrinogen or endothelial cells, and thrombus formation on collagen matrix. The efficacy of clopidogrel (15 mg·kg-1·day-1, followed by 5 mg·kg-1·day-1) on doxorubicin-induced changes in the aforementioned parameters as well as vascular injury were also determined. Whereas platelet count and size were similar between doxorubicin-treated and vehicle-treated mice, doxorubicin promoted thrombus formation evidenced by greater platelet aggregation, degranulation, and adhesion to endothelial cells evoked by agonists. Clopidogrel treatment attenuated the enhanced platelet activity and thrombus formation by doxorubicin, as well as vascular platelet infiltration and reactive oxygen species generation. Collectively, this study demonstrates that platelet functions are enhanced after long-term doxorubicin administration, which leads to thrombus formation and vascular toxicity, and that doxorubicin-induced changes in the functionality of platelets can be effectively inhibited by antiplatelet drugs.NEW & NOTEWORTHY Doxorubicin therapy in mice (antitumor dosage) markedly enhanced platelet functions measured as agonist-induced platelet aggregation, degranulation, and adhesion to endothelial cells, actions leading to thrombus formation and thrombosis-independent vascular injury. Clopidogrel treatment ameliorated thrombus formation and vascular toxicity induced by doxorubicin via inhibiting platelet activity.

The ECG Characteristics of Patients With Isolated Hypomagnesemia.

BACKGROUND: Electrocardiographic (ECG) characteristics of patients with isolated hypomagnesemia are not well defined. We aimed to investigate these ECG characteristics in order to define clearly the features of isolated hypomagnesemia. HYPOTHESIS: Lower serum magnesium could affect ECG parameters after excluding potential confounders. METHODS: This retrospective study was of patients with low serum magnesium <0.65 mmol/L compared with the same patients after restoration to normal serum magnesium. Patients with hypokalemia, hypocalcemia and other electrolyte disturbances were excluded. ECG parameters manually determined and analyzed were P wave dispersion, PR interval, QRS duration, ST-T changes, T wave amplitude, T peak-to-end interval (Tpe), corrected Tpe (Tpec), QT, corrected QT (QTc), QT peak corrected (QTpc) and Tpe dispersion, Tpe/QT ratio. RESULTS: Two-hundred-and-fourteen patients with isolated hypomagnesemia were identified with 50 of them (56.9 ± 13.6 years; 25 males) being eligible for final analysis from 270,997 patients presenting April 2011-October 2017. In the period of isolated hypomagnesemia, P wave duration was found prolonged (p ≤ 0.02); as was QTc (439 ± 27 vs. 433 ± 22, p = 0.01). Tpec (122 ± 24vs. 111 ± 22, p = 0.000) and Tpe/QT ratio (0.29 ± 0.05 vs. 0.27 ± 0.05, p = 0.000) were increased. QTpc decreased during hypomagnesemia (334 ± 28 vs. 342 ± 21, p = 0.02). However, no significant differences were found in PR interval, QRS duration (85 ± 12 ms vs. 86 ± 12 ms, p = 0.122) and ST-T segments between the patients and their own controls. CONCLUSIONS: In patients with isolated hypomagnesemia, P wave duration, QTc, Tpec, and Tpe/QT ratio suggesting atrial depolarization and ventricular repolarization dispersion were significantly increased compared with normal magnesium levels in the same patients after restoration to normal levels.