Forced activation of dystrophin transcription by CRISPR/dCas9 reduced arrhythmia susceptibility via restoring membrane Nav1.5 distribution.

Dystrophin deficiency due to genetic mutations causes cardiac abnormalities in Duchenne's muscular dystrophy. Dystrophin is also shown to be downregulated in conventional failing hearts. Whether restoration of dystrophin expression possesses any therapeutic potential for conventional heart failure (HF) remains to be examined. HF mouse model was generated by transverse aortic constriction (TAC). In vivo activation of dystrophin transcription was achieved by tail-vein injection of adeno-associated virus 9 carrying CRISPR/dCas system for dystrophin. We found that activation of dystrophin expression in TAC mice significantly reduced the susceptibility to arrhythmia of TAC mice and the mortality rate. We further demonstrated that over-expression of dystrophin increased cardiac conduction of hearts in TAC mice by optical mapping evaluation. Activation of dystrophin expression also increased peak sodium current in isolated ventricular myocytes from hearts of TAC mice as recorded by the patch-clamp technique. Immunoblotting and immunofluorescence showed that increased dystrophin transcription restored the membrane distribution of Nav1.5 in the hearts of TAC mice. In summary, correction of dystrophin downregulation by the CRISPR-dCas9 system reduced the susceptibility to arrhythmia of conventional HF mice through restoring Nav1.5 membrane distribution. This study paved the way to develop a new therapeutic strategy for HF-related ventricular arrhythmia.

The application of the spot the difference teaching method in clinical skills training for residents.

BACKGROUND: Clinical skill training (CST) is indispensable for first-year surgical residents. It can usually be carried out through video-based flipped learning (FL) within a web-based learning environment. However, we found that residents lack the process of reflection, blindly imitating results in losing interest and passion for learning in the traditional teaching pattern. The teaching method of "spot the difference" (SDTM), which is based on the fundamentals of the popular game of "spot the difference," is designed to improve students' participation and reflective learning during skill training. This study aimed to evaluate this novel educational model's short-term and long-term effectiveness for surgical residents in China. METHODS: First-year residents who required a three-month rotation in the head and neck surgery department were recruited to participate in a series of CSTs. They were randomized into SDTM and traditional FL (control) groups. Clinical skill performance was assessed with validated clinical skill scoring criteria. Evaluations were conducted by comparing the scores that contain departmental rotation skill examinations and the first China medical licensing examination (CMLE) performance on practical skills. In addition, two-way subjective evaluations were also implemented as a reference for the training results. Training effects were assessed using t tests, Mann-Whitney-Wilcoxon tests, chi-square tests, and Cohen' s effect size (d). The Cohen' s d value was considered to be small (<0.2), medium (0.2-0.8), or large (>0.8). RESULTS: The SDTM group was significantly superior to the control group in terms of after-department skill examination (t=2.179, p<0.05, d=0.5), taking medical history (t=2.665, p<0.05, d=0.59), and CMLE performance on practical skill (t=2.103, p<0.05, d=0.47). The SDTM members rated the curriculum more highly than the control on the items relating to interestingness and participation (p < 0.05) with large effect sizes (d >0.8). There were no significant differences between the two groups on clinical competence (t=0.819, p=0.415, d=0.18), the first-time pass rate for CMLE (χ2 =1.663, p=0.197, d=0.29), and short-term operational skills improvement (t=1.747, p=0.084, d=0.39). CONCLUSIONS: SDTM may be an effective method for enhancing residents' clinical skills, and the effect is significant both short- and long-term. The improvement effect seemed to be more significant in the peer-involved SDTM than training alone. However, despite positive objective results, SDTM still risks student learning burnout. TRIAL REGISTRATION: ISRCTN registry, ISRCTN10598469 , 02/04/2022，retrospectively registered.

Transcription factor Meis1 act as a new regulator of ischemic arrhythmias in mice.

INTRODUCTION: The principal voltage-gated Na+ channel, NaV1.5 governs heart excitability and conduction. NaV1.5 dysregulation is responsible for ventricular arrhythmias and subsequent sudden cardiac death (SCD) in post-infarct hearts. The transcription factor Meis1 performs a significant role in determining differentiation fate and regenerative capability of cardiomyocytes. However, the functions of Meis1 in ischemic arrhythmias following myocardial infarction (MI) are still largely undefined. OBJECTIVES: Here we aimed to study whether Meis1 could act as a key regulator to mediate cardiac Na+ channel and its underlying mechanisms. METHODS: Heart-specific Meis1 overexpression was established by AAV9 virus injection in C57BL/6 mice. The QRS duration, the incidence of ventricular arrhythmias and cardiac conduction velocity were evaluated by ECG, programmed electrical stimulation and optical mapping techniques respectively. The conventional patch clamp technique was performed to explore the INa characteristics of isolated mouse ventricular myocytes. In vitro, Meis1 was also overexpressed in hypoxic-treated neonatal cardiomyocytes. The analysis of immunoblotting and immunofluorescence were used to detect the changes in the expression of NaV1.5 in each group. RESULTS: We found that forced expression of Meis1 rescued the prolongation of QRS complex, produced anti-arrhythmic activity and improved epicardial conduction velocity in infarcted mouse hearts. In terms of mechanisms, cardiac electrophysiological changes of MI mice can be ameliorated by the recovery of Meis1, which is characterized by the restoration of INa current density and NaV1.5 expression level of cardiomyocytes in the marginal zone of MI mouse hearts. Furthermore, in vitro studies showed that Meis1 was also able to rescue hypoxia-induced decreased expression and dysfunction of NaV1.5 in ventricular myocytes. We further revealed that E3 ubiquitin ligase CDC20 led to the ubiquitination and degradation of Meis1, which blocked the transcriptional regulation of SCN5A by Meis1 and ultimately led to the electrophysiological remodeling in ischemic-hypoxic cardiomyocytes. CONCLUSION: CDC20 mediates ubiquitination of Meis1 to govern the transcription of SCN5A and cardiac electrical conduction in mouse cardiomyocytes. This finding uncovers a new mechanism of NaV1.5 dysregulation in infarcted heart, and provides new therapeutic strategies for malignant arrhythmias and sudden cardiac death following MI.

Cytoplasmic sequestration of p53 by lncRNA-CIRPILalleviates myocardial ischemia/reperfusion injury.

Myocardial ischemia/reperfusion (MI/R) injury is a pathological process that seriously affects the health of patients with coronary artery disease. Long non-coding RNAs (lncRNAs) represents a new class of regulators of diverse biological processes and disease conditions, the study aims to discover the pivotal lncRNA in MI/R injury. The microarray screening identifies a down-regulated heart-enriched lncRNA-CIRPIL (Cardiac ischemia reperfusion associated p53 interacting lncRNA, lncCIRPIL) from the hearts of I/R mice. LncCIRPIL inhibits apoptosis of cultured cardiomyocytes exposed to anoxia/reoxygenation (A/R). Cardiac-specific transgenic overexpression of lncCIRPIL alleviates I/R injury in mice, while knockout of lncCIRPIL exacerbates cardiac I/R injury. LncCIRPIL locates in the cytoplasm and physically interacts with p53, which leads to the cytoplasmic sequestration and the acceleration of ubiquitin-mediated degradation of p53 triggered by E3 ligases CHIP, COP1 and MDM2. p53 overexpression abrogates the protective effects of lncCIRPIL. Notably, the human fragment of conserved lncCIRPIL mimics the protective effects of the full-length lncCIRPIL on cultured human AC16 cells. Collectively, lncCIRPIL exerts its cardioprotective action via sequestering p53 in the cytoplasm and facilitating its ubiquitin-mediated degradation. The study highlights a unique mechanism in p53 signal pathway and broadens our understanding of the molecular mechanisms of MI/R injury.

A Special Report on 2019 International Planning Competition and a Comprehensive Analysis of Its Results.

PURPOSE: The aim of this work is to introduce the 2019 International Planning Competition and to analyze its results. METHODS AND MATERIALS: A locally advanced non-small cell lung cancer (LA-NSCLC) case using the simultaneous integrated boost approach was selected. The plan quality was evaluated by using a ranking system in accordance with practice guidelines. Planners used their clinical Treatment Planning System (TPS) to generate the best possible plan along with a survey, designed to obtain medical physics aspects information. We investigated the quality of the large population of plans designed by worldwide planners using different planning and delivery systems. The correlations of plan quality with relevant planner characteristics (work experience, department scale, and competition experience) and with technological parameters (TPS and modality) were examined. RESULTS: The number of the qualified plans was 287 with a wide range of scores (38.61-97.99). The scores showed statistically significant differences by the following factors: 1) department scale: the mean score (89.76 ± 8.36) for planners from the departments treating >2,000 patients annually was the highest of all; 2) competition experience: the mean score for the 107 planners with previous competition experience was 88.92 ± 9.59, statistically significantly from first-time participants (p = .001); 3) techniques: the mean scores for planners using VMAT (89.18 ± 6.43) and TOMO (90.62 ± 7.60) were higher than those using IMRT (82.28 ± 12.47), with statistical differences (p <.001). The plan scores were negligibly correlated with the planner's years of work experience or the type of TPS used. Regression analysis demonstrated that plan score was associated with dosimetric objectives that were difficult to achieve, which is generally consistent with a clinical practice evaluation. However, 51.2% of the planners abandoned the difficult component of total lung receiving a dose of 5 Gy in their plan design to achieve the optimal plan. CONCLUSION: The 2019 international planning competition was carried out successfully, and its results were analyzed. Plan quality was not correlated with work experiences or the TPS used, but it was correlated with department scale, modality, and competition experience. These findings differed from those reported in previous studies.

The association between soluble suppression of tumorigenicity-2 and long-term prognosis in patients with coronary artery disease: A meta-analysis.

OBJECTIVE: Findings regarding the prognostic value of soluble suppression of tumorigenecity-2 (sST2) in patients with coronary artery disease (CAD) remain inconsistent. Therefore, we conducted this meta-analysis to investigate the long-term prognostic value of sST2 in patients with CAD. METHODS: A comprehensive literature search was conducted across the PubMed, Embase, and Cochrane Library databases up to June 3, 2020. The primary outcome was major adverse cardiac events (MACEs). The secondary outcomes were all-cause mortality, cardiovascular (CV) death, heart failure (HF), and myocardial infarction (MI). Pooled estimations and 95% confidence intervals (CIs) were assessed using a random-effects model. RESULTS: Twenty-two articles that enrolled a total of 17,432 patients with CAD were included in the final analysis. CAD patients in the highest categories of baseline sST2 had a significantly higher risk of MACEs (HR: 1.42, 95% CI: 1.09-1.76), all-cause mortality (HR: 2.00, 95% CI: 1.54-2.46), and CV death (HR: 1.42, 95% CI: 1.15-1.68), HF (HR: 2.41, 95% CI: 1.87-2.94), but not that of MI (HR: 1.15, 95% CI: -0.73-3.04), than those in the lowest categories. These results were consistent when baseline sST2 was presented as continuous values in one unit increments. Moreover, subgroup analysis showed that elevated baseline sST2 levels increased the long-term risk of MACEs in the acute coronary syndrome (ACS) population (HR: 1.74, 95% CI: 1.39-2.09) but only showed a trend toward higher risk of MACEs in the non-ACS population (HR: 1.09, 95% CI: 0.87-1.30). CONCLUSIONS: The findings suggest that a higher concentration of baseline sST2 is associated with a higher risk of MACEs, all-cause mortality, CV death, and HF in patients with CAD. Elevated sST2 levels could significantly predict future MACEs in the ACS population but not in the non-ACS population.