Generation of a TBX20 homozygous knockout stem cell line (WAe009-A-1E) by episomal vector-based CRISPR/Cas9 system.

The T-box family transcription factor gene TBX20 plays a crucial role in cardiac development and function. TBX20 mutations are associated with congenital heart disease, dilated cardiomyopathy, arrhythmias, and heart failure. To further study the role of TBX20 in human heart, here we generated a homozygous TBX20 knockout (TBX20-KO) human embryonic stem cell line using the CRISPR/Cas9 system. This TBX20-KO cell line maintains normal morphology, pluripotency, and karyotype, making it a valuable tool for investigating TBX20's role in cardiac biology.

Explainable Federated Medical Image Analysis Through Causal Learning and Blockchain.

Federated learning (FL) enables collaborative training of machine learning models across distributed medical data sources without compromising privacy. However, applying FL to medical image analysis presents challenges like high communication overhead and data heterogeneity. This paper proposes novel FL techniques using explainable artificial intelligence (XAI) for efficient, accurate, and trustworthy analysis. A heterogeneity-aware causal learning approach selectively sparsifies model weights based on their causal contributions, significantly reducing communication requirements while retaining performance and improving interpretability. Furthermore, blockchain provides decentralized quality assessment of client datasets. The assessment scores adjust aggregation weights so higher-quality data has more influence during training, improving model generalization. Comprehensive experiments show our XAI-integrated FL framework enhances efficiency, accuracy and interpretability. The causal learning method decreases communication overhead while maintaining segmentation accuracy. The blockchain-based data valuation mitigates issues from low-quality local datasets. Our framework provides essential model explanations and trust mechanisms, making FL viable for clinical adoption in medical image analysis.

Effect of CDM3 on co-culture of human-induced pluripotent stem cells with Matrigel-covered polycaprolactone to prepare cardiac patches.

To promote the differentiation of human-induced pluripotent stem cells (hiPSCs) into myocardium through a standard chemically defined and small-molecule-based induction protocol (CDM3), and preliminarily prepare myocardial patches that provide experimental data and theoretical support for further maturation through other in vitro experiments and safety studies in vivo. After resuscitation, culture, and identification of hiPSCs, they were inoculated onto Matrigel-coated polycaprolactone (PCL). After 24 h, cell growth was observed by DAPI under a fluorescence microscope and the stemness of hiPSCs was identified by OCT4 fluorescence. After fixation, scanning electron microscopy was performed to observe the morphology of cells on the patch surface. On days 1, 3, 5, and 7 of culture, cell viability was determined by Cell Counting Kit-8 (CCK-8) assay and a curve was drawn to observe cell growth and proliferation. After co-culture with Matrigel-covered PCL for 24 h, hiPSCs were divided into control and CDM3 groups, and cultured for an additional 6 d. On the eighth day, cell growth was observed by DAPI under a fluorescence microscope, hiPSC stemness was identified by OCT4 fluorescence, and cardiomyocytes were identified by cardiac troponin T (cTnT) and α-actin expression. hiPSCs co-cultured with Matrigel-covered PCL for 24 h emitted green fluorescence indicating OCT4, showing that hiPSCs maintained their stemness on Matrigel-covered PCL scaffolds. DAPI emitted blue fluorescence, indicating that cells grew clonally with uniform cell morphology. Scanning electron microscopy showed that hiPSCs adhered and grew on PCL covered with Matrigel, with clearly visible cell outlines indicating normal morphology. Assessment of cell viability by the CCK-8 method showed that hiPSCs proliferated and grew on PCL scaffolds covered with Matrigel. After 6 d of culture, immunofluorescence showed that control group hiPSCs highly expressed the stem cell marker OCT4 but not myocardial markers cTnT or α-actin. In contrast, notable expression of myocardial markers cTnT and α-actin but not OCT4 occurred in the CDM3 group. hiPSCs can proliferate and grow on PCL scaffolds covered with Matrigel. Under the influence of CDM3, hiPSCs differentiated into cardiomyocyte-like cells, allowing the preliminary preparation of myocardial patches that can provide a better method for clinical treatment of myocardial infarction.

Experimental study on co-culture of DiI-labeled rat bone marrow mesenchymal stem cells and neonatal rat cardiomyocytes to induce differentiation into cardiomyocyte-like cells.

BACKGROUND: Myocardial infarction is a serious clinical disease with high mortality and poor prognosis. Cardiomyocytes (CMs) have limited regeneration abilities after ischemic injury. Their growth and differentiation can be enhanced by contact co-culture with stem cells. OBJECTIVE: The aim was to study the contact co-culture of Dil-labeled bone marrow mesenchymal stem cells (BMSCs) and CMs for inducing differentiation of CMs from stem cells for treating myocardial infarction. METHODS: After contact co-culture, the differentiation of BMSCs into CMs was analyzed qualitatively by detecting myocardial markers (cardiac troponin T and α-smooth muscle actin) using immunofluorescence and quantitatively using flow cytometry. To examine the mechanism, possible gap junctions between BMSCs and CMs were analyzed by detecting gap junction protein connexin 43 (C×43) expression in BMSCs using immunofluorescence. The functionality of gap junctions was analyzed using dye transfer experiments. RESULTS: The results revealed that BMSCs in contact with CMs exhibited myocardial markers and a significant increase in differentiation rate (P < 0.05); they also proved the existence and function of gap junctions between BMSCs and CMs. CONCLUSIONS: It was shown that contact co-culture can induce Dil-labeled BMSCs to differentiate into CM-like cells and examined the principle of gap junction-mediated signaling pathways involved in inducing stem cells to differentiate into cardiomyocytes.

Retrospective, observational analysis of cardiac function associated with global preoperative myocardial scar in patients with ischemic cardiomyopathy after coronary artery bypass grafting.

Background: Drawing on accumulated patient data from a hospital database, the goal of this retrospective study was to analyze cardiac function associated with global preoperative myocardial scarring assessed by cardiac magnetic resonance with late gadolinium enhancement (CMR-LGE) in patients with ischemic cardiomyopathy (ICM) after coronary artery bypass grafting (CABG). Methods: A total of 57 patients diagnosed with ICM who underwent isolated CABG at Beijing Anzhen Hospital between September 2017 and September 2019 were enrolled in this retrospective study. All these patients underwent a preoperative CMR-LGE examination. Based on postoperative echocardiography results at 6 months, cases were divided into the following 2 groups: improved cardiac function [a difference of left ventricular ejection fraction (LVEF) greater than or equal to 5%] and unimproved cardiac function. The factors contributing to these patients' unimproved cardiac function were investigated. Results: At 6 months after surgery, 64.9% (37/57) of cases had improved cardiac function, and 35.1% (20/57) had no improvement. There was no statistical difference between the 2 groups in the Synergy Between Percutaneous Coronary Intervention With Taxus and Cardiac Surgery (SYNTAX) score (41.7±7.6 vs. 42.8±8.3; P=0.603), but compared to the improved group, preoperative myocardial scarring was significantly enlarged in the unimproved group (41.9%±6.4% vs. 27.8%±8.5%; P<0.001). In regression analysis, only preoperative myocardial scarring [odds ratio (OR) =1.44; 95% confidence interval (CI): 1.13-1.83; P=0.003] was associated with no change in cardiac function evaluated by echocardiography after CABG. The median follow-up of 1.6 years (range, 0.6-4.1 years) found that the unimproved group had a higher incidence of major adverse cardiovascular and cerebrovascular events (MACCEs) (8.1% vs. 25.0%; P=0.044), and that the New York Heart Association (NYHA) classification of the unimproved group was higher than that of the improved group (P=0.018). Conclusions: In ICM patients, a greater amount of preoperative myocardial scarring is associated with unimproved cardiac function after CABG. The measurement of preoperative myocardial scarring may aid clinicians in identifying patients who would benefit from CABG.

Preparation of myocardial patches from DiI-labeled rat bone marrow mesenchymal stem cells and neonatal rat cardiomyocytes contact co-cultured on polycaprolactone film.

To provide better treatment of myocardial infarction, DiI-labeled bone marrow mesenchymal stem cells (BMSCs) were contact co-cultured with cardiomyocytes (CMs) on polycaprolactone (PCL) film to prepare myocardial patches. BMSCs from Sprague Dawley rats were isolated, cultured, and characterized for expression of surface markers by flow cytometry. CMs were isolated from suckling rats. After BMSCs were cultured for three generations, they were labeled with DiI dye. DiI-labeled BMSCs were co-cultured with CMs on PCL film in the experimental group, while CMs were replaced with the same amount of unlabeled BMSCs in the control group. After 24 h, cell growth was observed by light microscopy and cells were fixed for scanning electron microscopy (SEM). After 7 d of co-culture, cells were stained for immunofluorescence detection of myocardial markers cardiac troponin T (cTnT) andα-actin. Differentiation of BMSCs on PCL was observed by fluorescence microscopy. The efficiency of BMSC differentiation into CMs was analyzed by flow cytometry on the first and seventh days of co-culture. CMs were stained with calcein alone and contact co-cultured with DiI-labeled BMSCs on PCL film to observe intercellular dye transfer. Finally, cells were stained for immunofluorescence detection of connexin 43 (Cx43) expression and to observe the relationship between gap junctions and contact co-culture. BMSCs were identified by flow cytometry as strongly positive for CD90 and CD44H, and negative for CD11b/c and CD45. After co-culture for 24 h, cells were observed to have attached to PCL by light microscopy. Upon appropriate excitation, DiI-labeled BMSCs exhibited red fluorescence, while unlabeled CMs did not. SEM revealed a large number of cells on the PCL membrane and their cell state appeared normal. On the seventh day, some DiI-labeled BMSCs expressed cTnT andα-actin. Flow cytometry showed that the rate of stem cell differentiation in the experimental group was significantly higher than the control group on the seventh day (20.12% > 3.49%,P< 0.05). From the second day of co-culture, immunofluorescence staining for Cx43 revealed green fluorescent puncta in some BMSCs; from the third day of co-culture, a portion of BMSCs exhibited green fluorescence in dye transfer tests. Contact co-culture of DiI-labeled BMSCs and CMs on PCL film generated primary myocardial patches. The mechanism by which contact co-culture promoted differentiation of the myocardial patch may be related to gap junctions and gap junction-mediated intercellular signaling pathways.

Outcomes of surgical repair of anterior or posterior ventricular septal rupture after myocardial infarction.

Background: Ventricular septal rupture (VSR) is a rare and fatal complication of myocardial infarction. Surgery is the main treatment for the condition. It is currently believed that surgery is less effective for posterior VSR than for anterior VSR. The objective of this study was to investigate the clinical outcomes of surgical treatment for myocardial infarction combined with an anterior or posterior VSR. Methods: This was a single-center, retrospective, observational, cohort study. Clinical data of 68 patients with myocardial infarction combined with VSR were retrospectively analyzed. According to the site of the VSR, patients were divided into the anterior (43 cases) and posterior (25 cases) VSR groups, and the general clinical data, preoperative examination results, surgery, and follow-up results were compared between the two groups. Results: Compared with the anterior VSR group, the operative time in the posterior VSR group was longer {300 [240, 360] vs. 360 [300, 400] min; P=0.003}, and the cardiopulmonary bypass time was longer (142.0±52.2 vs. 180.2±52.3 min; P=0.005), and the aortic clamp time was longer (84.0±32.5 vs. 115.9±39.8 min; P=0.001). There were no significant differences in the incidence of perioperative complications, including bleeding, low cardiac output, pulmonary, and cerebrovascular complications, and the incidence of perioperative death between the two groups (P>0.05). The patients were followed up for 1.0-10.5 (median, 4.2) years. There were no significant differences in the survival rate and the incidence of major adverse cardiovascular and cerebrovascular events (MACCEs) including myocardial infarction, heart failure, revascularization, and cerebrovascular events between the two groups (P>0.05). Conclusions: The perioperative risks and medium- and long-term outcomes of the surgical repair of anterior or posterior VSR after myocardial infarction were similar.

The number of myocardial infarction segments connected to papillary muscle is associated with the improvement in moderate ischemic mitral regurgitation.

Background: We evaluated whether the number of myocardial infarction (MI) segments connected to the papillary muscle (PM), as assessed using cardiac magnetic resonance (CMR) with late gadolinium enhancement (LGE), predicts whether moderate ischemic mitral regurgitation (IMR) improves after isolated coronary artery bypass grafting (CABG) to guide the choice of surgical strategy. Methods: A total of 54 patients diagnosed with coronary heart disease (CHD) complicated with moderate IMR who underwent isolated CABG were selected continuously in this retrospective study at Beijing Anzhen Hospital. All patients underwent preoperative LGE. The patients were divided into the IMR improved group (37 patients) and the unimproved group (17 patients) according to 1-year postoperative echocardiography. The factors associated with no IMR improvement after isolated CABG were analyzed. There was no trial registration and no publication of the study protocol. Results: The number of MI segments connected to PM was an independent risk factor for no IMR improvement after isolated CABG [odds ratio 4.39; 95% confidence interval (CI): 1.93-9.98; P<0.001]. The optimal receiver operating characteristic (ROC) curve cut-off value for no IMR improvement was ≥2 (sensitivity: 82.4%; specificity: 83.8%). Follow-up at 1-5 years (median, 2.8 years) showed that the incidences of major adverse cardiovascular and cerebrovascular events (5.4% vs. 23.5%; P=0.041) and New York Heart Association (NYHA) grade (P=0.026) were higher in the unimproved group. Conclusions: In patients with CHD complicated with moderate IMR, the number of MI segments connected to PM is an independent risk factor for no IMR improvement after isolated CABG. Mitral valve surgery should be performed simultaneously with CABG in patients with ≥2 MI segments connected to the PM.

The use of electrical stimulation to induce cardiac differentiation of stem cells for the treatment of myocardial infarction.

Ischemic heart disease resulting from a myocardial infarction (MI), is a major health issue. Stem cell therapies may play an important role in this field. However, cardiomyocytes induced from stem cells are characterized by low rates of differentiation and immaturity. After transplantation into the damaged heart, they may even increase the risk of arrhythmias. Studies have demonstrated that electrical stimulation (ES) can promote the cardiac differentiation of stem cells. This review summarizes the latest research on the effects of applying different electrical stimulation (ES) parameters to different types of stem cells and the related mechanisms that may be involved.

Recent Advances in Maturation of Pluripotent Stem Cell-Derived Cardiomyocytes Promoted by Mechanical Stretch.

Stem cells have significant potential use in tissue regeneration, especially for treating cardiac diseases because of their multi-directional differentiation capability. By mimicking the in vivo physiological environment of native cardiomyocytes during their development and maturation, researchers have been able to induce pluripotent stem cell-derived cardiomyocytes (PSC-CMs) at high purity. However, the phenotype of these PSC-CMs is immature compared with that of adult cardiomyocytes. Various strategies have been explored to improve the maturity of PSC-CMs, such as long-term culturing, mechanical stimuli, chemical stimuli, and combinations of these strategies. Among these strategies, mechanical stretch as a key mechanical stimulus plays an important role in PSC-CM maturation. In this review, the optimal parameters of mechanical stretch, the effects of mechanical stretch on maturation of PSC-CMs, underlying molecular mechanisms as well as existing problems are discussed. Mechanical stretch is a powerful approach to promote the maturation of SC-CMs in terms of morphology, structure, and functionality. Nonetheless, further research efforts are needed to reach a satisfactory standard for clinical applications of PSC-CMs in treating cardiac diseases.

A Further Exploration of Multi-Slot Based Spectrum Sensing.

Spectrum sensing (SS) exhibits its advantages in the era of Internet of Things (IoT) due to limited spectrum resource and a lower utilization rate of authorized spectrum. In consequence, the performance improvement of SS seems a matter of great significance for the development of wireless communication and IoT. Motivated by this, this paper is devoted to multi-slot based SS in specialty and several important conclusions are drawn. Firstly, SS with one slot outperforms those with multiple slots if decision fusion rule is considered for multi-slot based SS. Secondly, multi-slot based SS is conducive to the performance improvement of SS when instantaneous strong noise occurs in the radio environment. Thirdly, for multi-slot based cooperative spectrum sensing (CSS), majority voting rule among multiple nodes obtains the optimal sensing performance. Both theoretical analysis and simulation experiment validate the conclusions drawn in this paper.

Hypoxic three-dimensional culture microenvironment promotes proliferation of bone marrow mesenchymal stem cells through HIF-1α signaling pathway / 中国胸心血管外科临床杂志

@#Objective To investigate the effects of hypoxic three-dimensional culture microenvironment on the proliferation of bone marrow mesenchymal stem cells and its mechanism. Methods P5 generation mouse bone marrow mesenchymal stem cells and P (3HB-co-4HB) were co-cultured under normoxic three-dimensional (20%) and hypoxic three-dimensional microenvironment (4%) respectively. After 24 hours, the proliferation of the two groups was determined by CCK-8 method. The expression of HIF-1α gene was detected by real-time quantitative PCR after 12 hours. Western blotting was used to detect the expression of HIF-1α protein after 24 hours. Results After 24 hours, the CCK-8 method showed that the OD value of the hypoxia group was significantly higher than that of the normoxia group (0.455±0.027 vs. 0.352±0.090, n=12, P<0.05). After 12 hours of hypoxic culture, the expression level of HIF-1α mRNA in the hypoxia group was significantly higher than that in the normoxia group (P<0.05). Compared with the normoxia group (0.47± 0.05), the relative expression level of HIF-1α protein in the hypoxia group (0.63±0.06) significantly increased in the Western blotting after 24 hours (n=3, P<0.05). Conclusion The hypoxic three-dimensional microenvironment can promote the proliferation of bone marrow mesenchymal stem cells, which may be related to the activation of HIF-1α signaling pathway.

Treatment of Myocardial Infarction and Mitral Regurgitation in a Patient with Congenitally Corrected Transposition of the Great Arteries.

Congenitally corrected transposition of the great arteries (CTGA) is a rare congenital heart disease. In patients with functional CTGA with circumflex artery occlusion and mitral regurgitation (MR), the right ventricle functions as the left ventricle. Coronary artery bypass grafting with mitral valve replacement is an effective treatment for CTGA with concomitant myocardial infarction (MI) and MR.

[Culture effects of mouse induced pluripotent stem cells by three-dimensional films in vitro].

OBJECTIVE: To observe the proliferation and differentiation capacities of mouse induced pluripotent stem cells (miPSCs) cultured in 3-hydroxybutyrate-co-3-hydroxyhexanoate(PHBHHx) three-dimensional films three-dimension films for the purpose of finding a suitable polymeric biomaterials for forming myocardial patches. METHODS: miPSCs were recovered, passaged, cultured and identified, then miPSCs divided into the experimental and control groups. MiPSCs in the experimental group were cultured with PHBHHx three-dimension films. MiPSCs in the control group were cultured with traditional culture dish. Stem cell culture medium or differential medium were added to miPSCs to detecte cell vitality by CCK-8 after 72 hours or to measure the cTnT expression of miPSCs through immunofluorescence or the cTnT expression quantity through flow cytometry after 15 days. RESULTS: Cell activity assay showed that the absorbance values were 0.836 ± 0.038 in the experimental group, 0.312 ± 0.004 in the control group (P<0.05). Scanning electron microscope (SEM) observation showed that miPSCs grew well on the PHBHHx dimensional films with normal shape. Immunofluorescence results demonstrated positive cTnT expression in both groups and flow cytometry measured cTnT expression was (60.32 ± 1.76)% in the experimental group and (47.54 ± 1.46)% in the control group (P<0.05). CONCLUSIONS: miPSCs can survive, proliferate and differentiate on PHBHHx dimensional films. miPSCs proliferation and differentiation capacities are significantly higher in PHBHHx three-dimensional films culture compared with the traditional cell culture.

Comparative study of directional differentiation of human and mouse embryonic stem cells into cardiomyocytes.

This comparative study investigates the method, efficiency, and anti-hypoxic ability of cardiomyocytes, directionally induced from human (h) and mouse (m) embryonic stem cells (ESCs). hESCs were induced into cardiomyocytes by suspension culture, without inducers, or adherent culture using the inducers activin A and BMP4. mESCs were induced into cardiomyocytes by hanging-drop method, without inducers or induced with vitamin C. All four methods successfully induced ESCs to differentiate into cardiomyocytes. There was a significant difference between groups with and without inducers. A significant difference was found between mESC and hESC groups with inducers. The average beating frequency of cardiomyocytes differentiated from hESC was lower than cardiomyocytes differentiated from mESC, while the average beating frequency of cardiomyocytes differentiated from the same cell line, despite different culture methods, did not differ. Beating cardiomyocytes of each group were positive for cTnT staining. Spontaneous action potentials of beating cardiomyocytes were detected by patch-clamp experiments in each group. Different apoptotic ratios were detected in beating cardiomyocytes in each group and the difference between cardiomyocytes induced from mESCs and hESCs was statistically significant. The differentiation efficiencies in the groups without inducers were significantly higher than those without inducers. The induction of mESCs was more simple and efficient compared with hESCs. Without the presence of other protective factors, the anti-hypoxic ability of cardiomyocytes induced from hESCs was stronger and the beating times were longer in vitro compared with mESCs.