Genome-wide analysis of RNA-binding proteins co-expression with alternative splicing events in mitral valve prolapse.

Objectives: We investigated the role and molecular mechanisms of RNA-binding proteins (RBPs) and their regulated alternative splicing events (RASEs) in the pathogenesis of mitral valve prolapse (MVP). Methods: For RNA extraction, we obtained peripheral blood mononuclear cells (PBMCs) from five patients with MVP, with or without chordae tendineae rupture, and five healthy individuals. High-throughput sequencing was used for RNA sequencing (RNA-seq). Differentially expressed genes (DEGs) analysis, alternative splicing (AS) analysis, functional enrichment analysis, co-expression of RBPs, and alternative splicing events (ASEs) analysis were conducted. Results: The MVP patients exhibited 306 up-regulated genes and 198 down-regulated genes. All down- and up-regulated genes were enriched in both Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. Furthermore, MVP was closely associated with the top 10 enriched terms and pathways. In MVP patients, 2,288 RASEs were found to be significantly different, and four suitable RASEs (CARD11 A3ss, RBM5 ES, NCF1 A5SS, and DAXX A3ss) were tested. We identified 13 RNA-binding proteins (RBPs) from the DEGs and screened out four RBPs (ZFP36, HSPA1A, TRIM21, and P2RX7). We selected four RASEs based on the co-expression analyses of RBPs and RASEs, including exon skipping (ES) of DEDD2, alternative 3' splice site (A3SS) of ETV6, mutually exclusive 3'UTRs (3pMXE) of TNFAIP8L2, and A3SS of HLA-B. Furthermore, the selected four RBPs and four RASEs were validated by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and showed high consistency with RNA sequencing (RNA-seq). Conclusion: Dysregulated RBPs and their associated RASEs may play regulatory roles in MVP development and may therefore be used as therapeutic targets in the future.

Myocardial work and energy loss of left ventricle obtained by pressure-strain loop and vector flow mapping: a new perspective on idiopathic left bundle branch block.

Background: To date, no research has been conducted on the electrical activity and mechanical dyssynchrony of idiopathic left bundle branch block (iLBBB) with normal left ventricular ejection fraction (LVEF). This study sought to assess the left ventricular summation of energy loss (EL-SUM) and average energy loss (EL-AVE) using vector flow mapping as well as myocardial work using pressure-strain loop (PSL) in patients with iLBBB and normal LVEF. Methods: We prospectively recruited 35 patients with iLBBB and 35 control participants with normal LVEF. Echocardiography was performed. Conventional echocardiographic parameters, myocardial work, and energy loss (i.e., the EL-SUM and EL-AVE) were calculated. Results: In relation to global myocardial work, compared to the control participants, the iLBBB patients showed decreased global longitudinal strain (GLS; -15.32%±2.58% vs. -18.27%±2.12%; P=0.001), a decreased global work index (GWI; 1,428.24±338.18 vs. 1,964.87±264.16 mmHg%; P<0.001), decreased global work efficiency (GWE) (84.48±5.19 vs. 91.73±5.31 mmHg%; P<0.001), and significantly increased global waste work (GWW; 341.60±132.62 vs. 161.80±106.81 mmHg%; P<0.001). In relation to the regional index, the iLBBB patients had a significantly reduced basal anteroseptal segment (879.15±370.50 vs. 1,746.38±154.44 mmHg%; P<0.001), basal inferoseptal segment (1,111.42±389.04 vs. 1,677.25±223.10 mmHg%; P<0.001), mid-anteroseptal segment (1,097.54±394.83 vs. 1,815.06±291.22 mmHg%; P<0.001), mid-inferoseptal segment (1,012.54±353.33 vs. 1,880.88±254.39 mmHg%; P<0.001), apical anterior segment (1,592.42±366.64 vs. 1,910.00±170.27 mmHg%; P=0.001), apical lateral segment (1,481.62±342.95 vs. 1,817.19±227.55 mmHg%; P=0.001), apical septal segment (1,437.65±428.22 vs. 1,852.25±275.19 mmHg%; P=0.001), and apex (1,542.62±342.89 vs. 1,907.06±197.94 mmHg%; P<0.001). The iLBBB patients had increased EL-AVE and EL-SUM during the late-diastole, isovolumic-systole, and rapid-ejection periods [EL-AVE in T2: 28.3 (8.7, 49.0) vs. 6.8 (5.4, 9.4) J/(s·m3); P=0.029]; [EL-AVE in T3: 24.7 (13.0, 46.8) vs. 7.2 (5.4, 10.8) J/(s·m3), P<0.001]; [EL-AVE in T4: 18.3 (12.0, 27.6) vs. 7.7 (4.1, 11.6) J/(s·m3), P=0.002]; [EL-SUM in T2: 8.3 (2.2, 14.5) vs. 2.1 (1.6, 3.2) J/(s·m), P=0.049]; [EL-SUM in T3: 7.6 (4.0, 14.5) vs. 2.2 (1.7, 3.3) J/(s·m), P<0.001]; [EL-SUM in T4: 5.3 (3.6, 9.7) vs. 2.2 (1.4, 3.0) J/(s·m), P=0.004]. Conclusions: The GWI and GWE were reduced in patients with iLBBB, especially in the septum and apex. The EL-SUM and EL-AVE were higher in patients with iLBBB during the late-diastole, isovolumic-systole, and rapid-ejection periods. EL and PSL reflect the LV hemodynamics of patients with iLBBB.

Functional analysis of differently expressed ferroptosis-related genes in patients with mitral valve prolapse.

Background: The prevalence of mitral valve prolapse (MVP) in heart valvular diseases is globally increasing. However, the understanding of its etiology and pathogenesis is limited. So far, the relationship between ferroptosis-related genes and long non-coding RNAs (lncRNAs) in MVP remains unexplored. This study investigates the potential pathogenesis of ferroptosis-related genes in MVP and provides a therapeutic target for the disease. Methods: Blood samples from patients with MVP and healthy volunteers were collected for transcriptomic sequencing to analyze the expression of ferroptosis-related differentially expressed genes (DEGs) and differentially expressed long non-coding RNAs (DElncRNAs Co-expression network of ferroptosis-related DEGs and DElncRNAs. Furthermore, this work conducted GO and KEGG enrichment analyses. Results: CDKN2A, SLC1A4, ATF3, and other core genes related to the mitral valve prolapse were screened out. CDKN2A, SLC1A4, and ATF3 genes were at the core position of the network, regulated by numerous lncRNAs. Notably, these genes are primarily involved in the extracellular region and p53 signaling pathway. Conclusion: In summary, CDKN2A, SLC1A4, and ATF3 regulate the pathophysiological process of MVP and are potential therapeutic targets.

Aortic valve repair for the treatment of rheumatic aortic valve disease: a systematic review and meta-analysis.

Valvuloplasty for rheumatic aortic valve disease remains controversial. We conducted this study to explore whether aortic valvuloplasty is appropriate for the rheumatic population. A comprehensive search was conducted, and 7 eligible retrospective studies were identified from PubMed, Embase, Medline and Cochrane (up to April 7, 2020) according to the inclusion and exclusion criteria. The data for hospital mortality, 5-year survival, 5-year reoperation, aortic insufficiency grade (AIG) and aortic valve gradient (AVG) were extracted by 2 independent reviewers and were analysed to evaluate the safety and availability of aortic valvuloplasty for rheumatic patients. The heterogeneity of the results was estimated using the Q test and I2 statistics. The fixed pooling model was used when I2 ≤ 50%; otherwise, the random pooling model was selected. 7 articles with 418 patients were included. The pooled hospital mortality, 5-year survival and 5-year reoperation rates were 3.2%, 94.5% and 9.9%, respectively. The heterogeneities of the weighted mean differences (WMD) values of the AIG and AVG between preoperation and postoperation were extremely high (I2 = 81.5%, p < 0.001 in AIG, I2 = 97.6%, p = 0.003 in AVG). Subgroup analysis suggested that the AIG and AVG were improved by 3.03 grades (I2 = 0%, p < 0.001) and 3.16 mmHg (I2 = 0%, p < 0.001) in the European group, respectively. In the Asian group, the AIG and AVG were improved by 2.57 grades (I2 = 0%, p < 0.001) and 34.39 mmHg (I2 = 0%, p < 0.001), respectively. Compared with the values at discharge, the AIG was increased by 0.15 grades (I2 = 0%, p = 0.031) and the AVG was still decreased by 2.07 mmHg (I2 = 0%, p = 0.031) at the time of follow up. Valvuloplasty is safe and effective to treat rheumatic aortic insufficiency and stenosis, and the duration of maintenance required to improve stenosis was longer than that of insufficiency.

Telocytes-derived extracellular vesicles alleviate aortic valve calcification by carrying miR-30b.

AIMS: Calcific aortic valve disease (CAVD) is frequent in the elderly. Telocytes (TCs) are implicated in intercellular communication by releasing extracellular vesicles (EVs). This study investigated the role of TC-EVs in aortic valve calcification. METHODS AND RESULTS: TCs were obtained and identified using enzymolysis method and flow cytometry. EVs were isolated from TCs using differential high-speed centrifugation method and identified using transmission electron microscope, western blot, and qNano analysis. The mouse model of CAVD was established. The changes of aortic valve activity-related indicators were analysed by ultrasound, and the expressions of TC markers CD34 and vimentin in mouse valve tissues were detected using RT-qPCR and western blot. The model mice were injected with TC-derived EVs. The expressions of Runx2, osteocalcin, and caspase-3 were detected using RT-qPCR and western blot. The calcification model of valvular interstitial cells (VICs) was established. TC-EVs were co-cultured with calcified VICs, and calcium deposition was detected using alizarin red S staining. miR-30b expression in calcified valvular tissues and cells was detected after EV treatment. miR-30b expression in TCs was knocked down and then EVs were extracted and co-cultured with calcified VICs. The target of miR-30b was predicted through bioinformatics website and verified using dual-luciferase assay. The levels of Wnt/ß-catenin pathway-related proteins were detected. ApoE-/- mice fed with a high-fat diet showed decreased aortic valve orifice area, increased aortic transvalvular pressure difference and velocity, reduced left ventricular ejection fraction, decreased CD34 and vimentin, and increased caspase-3, Runx2, and osteocalcin. The levels of apoptosis- and osteogenesis- related proteins were inhibited after EV treatment. TC-EVs reduced calcium deposition and osteogenic proteins in calcified VICs. EVs could be absorbed by VICs. miR-30b expression was promoted in calcified valvular tissues and cells after EV treatment. Knockdown of miR-30b weakened the inhibitory effects of TC-EVs on calcium deposition and osteogenic proteins. miR-30b targeted Runx2. EV treatment inhibited the Wnt/ß-catenin pathway, and knockdown of miR-30b in TCs attenuated the inhibitory effect of TC-EVs on the Wnt/ß-catenin pathway. CONCLUSION: TC-EVs played a protective role in aortic valve calcification via the miR-30b/Runx2/Wnt/ß-catenin axis.

Tocilizumab for treating COVID-19: a systemic review and meta-analysis of retrospective studies.

OBJECTIVES: COVID-19 has become a global epidemic, and effective therapies have not been discovered up to now. We conducted this study to explore the effectiveness and safety of tocilizumab recently used for treating COVID-19. METHOD: A comprehensive search was conducted (up to September 27, 2020), and 19 eligible records were identified according to the inclusion and exclusion criteria. The data of the studies were extracted by 2 independent reviewers and were analyzed to evaluate the safety and availability of tocilizumab for treating COVID-19. RESULTS: Thirteen retrospective case-control studies (n = 2285 patients) and 6 retrospective single-armed studies (n = 208) were retrieved in this study. In the comparison of tocilizumab treatment group (TCZ) and standard treatment group (ST), significant associations with a lower risk of admission to ICU, use of ventilation, and mortality (OR, 95% CI: 0.53, 0.26~1.09; 0.66, 0.46~0.94; 0.44, 0.36~0.55) were found in the tocilizumab treatment group. What is more, patients treated with tocilizumab had better clinical improvement compared with the patients treated with ST (OR, 1.24; 95% CI, 0.96~1.62). After taking tocilizumab, the patients had lower C-reactive protein (CRP), white blood cell count (WBC), aspartate aminotransferase (AST) (WMD, 95% CI: - 99.66, - 156.24~- 43.09; - 0.95, - 1.8~- 0.11; - 12.58, - 18.88~-6.29) but higher troponin (WMD, 7.61; 95% CI, 3.06~12.15) than before. In addition, tocilizumab did not have significant influence on patients' neutrophil count (Neut), lymphocyte count (Lymp), platelet count (Plt), alanine aminotransferase (ALT), and creatine (WMD, 95% CI: - 0.29, - 2.91~2.33; 0.42, - 0.23~1.07; 5.2, - 2.85~13.25; 22.49, - 2.73~47.7; - 44.78, - 93.37~3.81). CONCLUSION: Tocilizumab may have potential effectiveness to treat COVID-19 according to the results of this study. However, more large-scale studies are needed for more accurate conclusions.

MiR-889 promotes cell growth in human non-small cell lung cancer by regulating KLF9.

Currently, non-small cell lung cancer (NSCLC) is still the most common malignancy worldwide. Although miR-889 has been reported to play an important role in various malignancies, the physiological function of miR-889 in NSCLC remains unknown. This paper places emphasis on the influence of miR-889 on the development and progression of non-small cell lung cancer. To detect the expression level of miR-889 in NSCLC tissues and cell lines, quantitative real-time polymerase chain reaction (qRT-PCR) assay and In Situ Hybridization (ISH) were adopted in this study. Cell proliferation and colony forming ability were examined by Cell Counting Kit-8 (CCK-8) and colony formation assays. Furthermore, transwell experiments were conducted to determine the influence of miR-889 on migration. KLF9 expression was evaluated by qRT-PCR and Western blotting. First, miR-889 expression was increased in the cancer tissues of non-small cell lung cancer patients (n = 40) compared with adjacent tissues. Subsequently, knockdown of miR-889 significantly inhibited cell proliferation and migration, while overexpression of miR-889 had the opposite effect. KLF9 may be a potential target of miR-889. In addition, upregulation of miR-889 promotes tumorigenesis in vitro, and KLF9 protein levels are also reduced. The current study suggests that miR-889 may play a potential therapeutic role for NSCLC by targeting KLF9 to control NSCLC proliferation and migration.

Total endoscopic repair of atrial septal defect under on-pump beating heart.

BACKGROUND: We previously reported the techniques for total endoscopic atrial septal defect (ASD) repair on hearts arrested with cardioplegia through three small incisions in the chest wall without aid of a surgical robotic system. The optimal results motivated us to use total thoracoscopic technology for ASD on perfused beating hearts. METHODS: From 2010 to 2017, 161 patients with a mean age of 28.31±12.34 years who underwent non-robotically assisted total thoracoscopic closure for ASD were included in this study, and those patients were also divided into two groups, including group A and group B. In group A, 115 patients underwent the procedure on beating hearts without aorta cross-clamped; in group B, 46 patients underwent the procedure on hearts arrested with cardioplegia with aorta cross-clamped. Cardiopulmonary bypass (CPB) was peripherally achieved as well. RESULTS: Total thoracoscopic ASD closures were successfully performed without in-hospital mortality or other serious complications in all patients of both groups. Dacron or bovine patches were used in 81 and 32 patients in the two groups, respectively. Duration of operation, duration of CPB, aorta cross-clamped time, duration of mechanical ventilation, the length of intensive care unit (ICU) and post-operative hospital stay in group A, were all shorter than those in group B (P<0.05). There was no statistically significant difference in blood transfusion during operation or post-operation thoracic drainage. During follow-up, echocardiograms at 3, 30, 90 and 365, showed no residual shunt or tricuspid regurgitation. CONCLUSIONS: Total thoracoscopic closure of ASD without assistance of a surgical robotic system on beating heart is safe and feasible and can be used as a therapeutic option for ASD, and by using the mentioned technique, less injuries are applied to patients.

MiR-155 inhibits proliferation and invasion by directly targeting PDCD4 in non-small cell lung cancer.

BACKGROUND: MicroRNAs are often abnormally expressed in human non-small cell lung cancer (NSCLC) and are thought to play a critical role in the emergence or maintenance of NSCLC by binding to its target messenger RNA. We assessed the effects of miR-155 on cell proliferation and invasion to elucidate the role played by miR-155/PDCD4 in NSCLC. METHODS: Quantitative reverse transcription-PCR, Western blotting, and cell counting kit-8, luciferase, and transwell invasion assays were conducted on a normal human bronchial epithelial cell line (BEAS-2B) and three NSCLC cell lines (SPC-A-1, A549, and H2170). RESULTS: We confirmed that miR-155 was upregulated, while PDCD4 messenger RNA and protein levels were downregulated in NSCLC cell lines. miR-155 negatively regulated PDCD4 at both transcriptional and post-transcriptional levels. Moreover, PDCD4 was forecast as an assumed target of miR-155 using bioinformatic methods and we demonstrated that PDCD4 was a direct target of miR-155 using luciferase reporter assays. Furthermore, PDCD4 overexpression could restrain NSCLC proliferation and invasion induced by miR-155. CONCLUSION: Our results collectively demonstrate that miR-155 exerts an oncogenic role in NSCLC by directly targeting PDCD4.

Essential role of STX6 in esophageal squamous cell carcinoma growth and migration.

Abnormalities in endosomes, or dysregulation in their trafficking, play an important role directly in many diseases including oncogenesis. Syntaxin-6 (STX6) is involved in diverse cellular functions in a variety of cell types and has been shown to regulate many intracellular membrane trafficking events such as endocytosis, recycling and anterograde and retrograde trafficking. However, its expression pattern and biological functions in esophageal squamous cell carcinoma (ESCC) remained unknown. Here, we have found that the expression of STX6 was up-regulated in ESCC samples, its expression was significantly correlated with tumor size, histological differentiation, lymph node metastasis and depth. On one hand, STX6 silencing inhibited ESCC cells viability and proliferation in a p53-dependent manner. On the other hand, STX6 effect integrin trafficking and regulate ESCC cells migration. Taken together, our study revealed the oncogenic roles of STX6 in the progression of ESCC, and it might be a valuable target for ESCC therapy.

The development of early-onset ventilator-associated pneumonia after cardiac surgery with cardiopulmonary bypass is associated with toll-like receptor 4 signal transduction pathways.

We evaluated the hypothesis that early-onset ventilator-associated pneumonia (VAP) after cardiac surgery with cardiopulmonary bypass (CPB) requires Toll-like receptor 4 (TLR4)-dependent signaling pathways. We enrolled 50 patients undergoing mitral and aortic double valve replacement, collecting left atrial blood samples from all patients preoperatively (T1), when opening the atrial septum (T2), closing the atrial septum (T3), and at the end of CPB (T4). TLR4 expression on monocytes and lymphocytes was detected immediately with flow cytometry. Serum levels of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-10 (IL-10) were measured using enzyme-linked immunosorbent assays (ELISA). Data from flow cytometry, ELISA, and the clinical outcomes in the two groups were evaluated and compared. Seventeen patients were included for analysis, and assigned to group A (without early-onset VAP, n = 10) and group B (with early-onset VAP, n = 7). TLR4 expression on lymphocytes in group B at T1, T3, and T4 were significantly higher than those in group A. Serum levels of IL-10 in group A at T4 were significantly higher than for group B. We found characteristic patterns in TLR4 expression on lymphocytes in left atrial blood of patients undergoing cardiac surgery with CPB. Our findings clarify the role of TLR4 and its association with the development of postoperative pneumonia.

Totally thoracoscopic repair of atrial septal defect reduces systemic inflammatory reaction and myocardial damage in initial patients.

BACKGROUND: To compare the effect of totally thoracoscopic with conventional, open repair of atrial septal defect. METHODS: Forty atrial septal defect cases were divided into two groups by surgical approach: totally thoracoscopic approach (group A, n = 20) and conventional open approach (group B, n = 20). In group A, surgical procedures were performed through three portal incisions in the right lateral chest wall under thoracoscopic vision without the aid of a computerized robotic surgical system. Notably, all operations were completed by one surgeon who had just begun using this technique. In group B, the atrial septal defects were repaired in conventional open fashion. Clinical outcomes and serum levels of tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), interleukin-10 (IL-10), intercellular adhesion molecule 1 (ICAM-1), and creatine kinase isoenzyme-myocardial band (CK-MB) for the two groups were evaluated and compared. RESULTS: All operations were performed successfully without serious complications. Durations of cardiopulmonary bypass (CPB), CPB setup, aortic cross-clamping, and operative procedure were significantly longer in group A than in group B (P < 0.05). The recovery times for body temperature and laboratory values of leukocytes were significantly shorter for group A than for group B (P < 0.05). There were no differences in durations of postoperative assisted ventilation or intensive care unit and hospital stays, volumes of blood transfused intraoperatively or thoracic drainage, or medical costs between the two groups. Serum levels of inflammatory factors (TNF-α, IL-6, IL-10, and ICAM-1) and CK-MB increased significantly in both groups after surgery. However, 6 h and 12 h after surgery, levels of these inflammatory factors and CK-MB were significantly lower in group A than in group B (P < 0.05). CONCLUSIONS: Thoracoscopic cardiac surgery is technically feasible and safe, with less trauma and quicker recovery even when done by a surgeon newly introduced to the technique.