Neuroprotective Effect of miR-483-5p Against Cardiac Arrest-Induced Mitochondrial Dysfunction Mediated Through the TNFSF8/AMPK/JNK Signaling Pathway.

Substantial morbidity and mortality are associated with postcardiac arrest brain injury (PCABI). MicroRNAs(miRNAs) are essential regulators of neuronal metabolism processes and have been shown to contribute to alleviated neurological injury after cardiac arrest. In this study, we identified miRNAs related to the prognosis of patients with neurological dysfunction after cardiopulmonary resuscitation based on data obtained from the Gene Expression Omnibus (GEO) database. Then, we explored the effects of miR-483-5p on mitochondrial biogenesis, mitochondrial-dependent apoptosis, and oxidative stress levels after ischemia‒reperfusion injury in vitro and in vivo. MiR-483-5p was downregulated in PC12 cells and hippocampal samples compared with that in normal group cells and hippocampi. Overexpression of miR-483-5p increased the viability of PC12 cells after ischemia‒reperfusion injury and reduced the proportion of dead cells. A western blot analysis showed that miR-483-5p increased the protein expression of PCG-1, NRF1, and TFAM and reduced the protein expression of Bax and cleaved caspase 3, inhibiting the release of cytochrome c from mitochondria and alleviating oxidative stress injury by inhibiting the production of ROS and reducing MDA activity. We confirmed that miR-483-5p targeted TNFSF8 to regulate the AMPK/JNK pathway, thereby playing a neuroprotective role after cardiopulmonary resuscitation. Hence, this study provides further insights into strategies for inhibiting neurological impairment after cardiopulmonary resuscitation and suggests a potential therapeutic target for PCABI.

Targeted Activation of HNF4α by AMPK Inhibits Apoptosis and Ameliorates Neurological Injury Caused by Cardiac Arrest in Rats.

Previous studies have shown that AMPK plays an important role in cerebral ischemia-reperfusion injury by participating in apoptosis, but the exact mechanism and target of action remains unclear. This study aimed to investigate the protective mechanism of AMPK activation on brain injury secondary to cardiac arrest. HE, Nills and TUNEL assays were used to evaluate neuronal damage and apoptosis. The relationships between AMPK, HNF4α and apoptotic genes were verified by ChIP-seq, dual-luciferase and WB assays. The results showed that AMPK improved the 7-day memory function of rats, and reduced neuronal cell injury and apoptosis in the hippocampal CA1 region after ROSC, while the use of HNF4α inhibitor weakened the protective effect of AMPK. Further research found that AMPK positively regulated the expression of HNF4α, and AMPK could promote the expression of Bcl-2 and inhibit the expression of Bax and Cleaved-Caspase 3. In vitro experiments showed that AMPK ameliorated neuronal injury by inhibiting apoptosis through the activation of HNF4α. Combined with ChIP-seq, JASPAR analysis and Dual-luciferase assay, the binding site of HNF4α to the upstream promoter of Bcl-2 was found. Taken together, AMPK attenuates brain injury after CA by activating HNF4α to target Bcl-2 to inhibit apoptosis.

IncRNA PLAC2 Upregulates CDK6 by Directly Targeting miR-29C to Promote Cell Proliferation in Lung Squamous Cell Carcinoma.

Long noncoding RN (IncRNA) placenta-specific 2 (PLAC2) plays a critical role in many cancer types. A previous study found that PLAC2 expression dysregulated in non-small cell lung cancer (NSCLC). However, its function in LSCC is not entirely known. The present study enrolled 68 lung squamous cell carcinoma (LSCC, a major subtype of NSCLC) patients (gender: 39 males and 29 females; age: 36 to 67 years old; mean age: 53.2 ± 5.5 years old). The expression levels of PLAC2 in two types of tissue (non-tumor and LSCC) were measured by quantitative PCR. LSCC cells H1581 and H1993 were transfected with PLAC2 and cyclin-dependent kinase 6 (CDK6) expression vectors and small interfering RNAs (siRNAs), as well as microRNA-29C (miR-29C) mimic and inhibitor to perform overexpression and knock-down experiment, respectively. PLAC2 was upregulated in LSCC and positively correlated with CDK6 but negatively correlated with miR-29C. During a 5-year follow-up, high expression levels of PLAC2 were found to be closely associated with poor survival. PLAC2 and CDK6 were significantly upregulated in LSCC cells, while miR-29C was remarkably downregulated. miR-29C was predicted to be a potential target of PLAC2, and RNA pull-down assay confirmed their direct interaction. Overexpression of PLAC2 led to upregulation of CDK6 and downregulation of miR-29C, while knock-down of PLAC2 showed opposite effects. Overexpression of miR-29C downregulated CDK6, while knock-down of miR-29C increased the expression levels of CDK6. However, the expression of PLAC2 was not affected by overexpression or knock-down of miR-29C. Overexpression of PLAC2 and CDK6 enhanced LSCC cell proliferation and cell cycle progression, while knock-down showed opposite effects. In addition, overexpression of miR-29C inhibited cell proliferation and cell cycle progression, while its knockdown display opposite effects. Moreover, overexpression of miR-29C suppressed the role of overexpression of PLAC2 in cell proliferation and cell cycle progression. In conclusion, PLAC2 upregulates CDK6 by downregulating miR-29C to promote LSCC cell proliferation.

Identification of Commonly Dysregulated Genes in Non-small-cell Lung Cancer by Integrated Analysis of Microarray Data and qRT-PCR Validation.

BACKGROUND: Non-small-cell lung cancer (NSCLC), the most common lung cancer, leads to the largest number of cancer-related deaths worldwide. There are many studies to identify the differentially expressed genes (DEGs) between NSCLC and normal control (NC) tissues by means of microarray technology. Because of the inconsistency of the microarray data sets, we performed an integrated analysis to identify DEGs and analyzed their biological function. METHODS AND RESULTS: We combined 15 microarray data sets and identified 1063 DEGs between NSCLC and NC tissues; in addition, we found that the DEGs were enriched in regulation of cell proliferation process and focal adhesion signaling pathway. The protein-protein interaction network analysis for the top 20 significantly DEGs revealed that CAV1, COL1A1, and ADRB2 were the significant hub proteins. Finally, we employed qRT-PCR to validate the meta-analysis approach by determining the expression of the top 10 most significantly DEGs and found that the expression of these genes were significantly different between tumor and NC tissues, in accordance with the results of meta-analysis. CONCLUSION: qRT-PCR results indicated that the meta-analysis approach in our study was acceptable. Our data suggested that some of the DEGs, including MMP12, COL11A1, THBS2, FAP, and CAV1, may participate in the pathology of NSCLC and could be applied as potential markers or therapeutic targets for NSCLC.

Inhibition of ASAP1 Modulates the Tumor Immune Microenvironment and Suppresses Lung Cancer Metastasis via the p-STAT3 Signaling Pathway.

ADP ribosylation factor guanylate kinase 1 (ASAP1), a key protein regulating cell migration and invasion, has attracted extensive attention in oncological research in recent years. This study aims to explore the effects of ASAP1 inhibition on lung cancer metastasis and its potential mechanisms, particularly how it modulates the tumor immune microenvironment through the p-STAT3 signaling pathway. In this study, shRNA technology was employed to specifically inhibit ASAP1 expression in lung cancer cell lines A549, NCI-H1299, and PC-9. The effects of ASAP1 inhibition on lung cancer cell viability, apoptosis, migration, and invasion were evaluated using CCK-8, TUNEL apoptosis detection, and cell migration and invasion assays. Furthermore, animal experiments were conducted to assess the in vivo effects of ASAP1 inhibition on lung cancer metastasis, and immunohistochemical analysis was performed to investigate changes in immune cells in lung metastasis models, further exploring its impact on the tumor immune microenvironment. The experimental results demonstrated that ASAP1 inhibition significantly reduced lung cancer cell viability, induced apoptosis in A549, NCI-H1299, and PC-9 cells, and suppressed the migration and invasion abilities of these cells. In vivo experiments revealed that ASAP1 inhibition effectively suppressed lung cancer metastasis and altered the tumor immune microenvironment by regulating immune cells. Moreover, we found that ASAP1 inhibition could decrease tumor cell proliferation and induce tumor apoptosis in lung metastasis models by inhibiting the p-STAT3 signaling pathway. This study confirms that ASAP1 inhibition can suppress lung cancer metastasis by modulating the tumor immune microenvironment through the inhibition of the p-STAT3 signaling pathway. These findings provide new targets for lung cancer treatment and a theoretical basis for developing novel strategies against lung cancer metastasis. Future research will further explore the mechanisms of ASAP1 in lung cancer metastasis and how to optimize treatment strategies for lung cancer patients by targeting ASAP1.

RNF6 activates TGF-ß1/c-Myb pathway to promote EMT in esophageal squamous cell carcinoma.

Objective: This study aimed to investigate RING-Finger Protein 6 (RNF6) expression in esophageal squamous cell carcinoma (ESCC) cells and whether it affects cell proliferation, invasion, and migration by regulating the TGF-ß1/c-Myb pathway. Methods: TCGA database was used to analyze RNF6 expression in normal tissues and esophageal cancer tissues. Kaplan-Meier method was used to examine the correlation between RNF6 expression and patient prognosis. SiRNA interference vector and RNF6 overexpression plasmid were constructed, and RNF6 was transfected into Eca-109 and KYSE-150 esophageal cancer cell line. In vitro scratch assay and Transwell assay were conducted to investigate the effects of RNF6 on the migration and invasion of Eca-109 and KYSE-150 cells. RT-PCR detected the expression of Snail, E-cadherin, and N-cadherin, and TUNEL detected the apoptosis of cells. Results: RNF6 up-regulation promoted the progression of esophageal cancer and predicted poor prognosis. RNF6 also enhanced the migration and invasion of ESCC cells in vitro. RNF6 silencing inhibited the migration and invasion of ESCC cells. TGF-ß inhibitors reversed the oncogenic effects of RNF6. RNF6 regulated the migration and invasion of ESCC cells by activating the TGF-ß pathway. RNF6/TGF-ß1 promoted esophageal cancer progression through c-Myb. Conclusion: RNF6 promotes the proliferation, invasion, and migration of ESCC cells possibly by activating the TGF-ß1/c-Myb pathway and affects the progression of ESCC.

Transcription factor EB (TFEB) improves ventricular remodeling after myocardial infarction by inhibiting Wnt/ß-catenin signaling pathway.

Background: Adverse left ventricular remodeling after myocardial infarction (MI) compromises cardiac function and increases heart failure risk. Until now, comprehension of the role transcription factor EB (TFEB) plays after MI is limited. Objectives: The purpose of this study was to describe the effects of TFEB on fibroblasts differentiation and extracellular matrix expression after MI. Methods: AAV9 (adeno-associated virus) mediated up- and down-regulated TFEB expressions were generated in C57BL/6 mice two weeks before the MI modeling. Echocardiography, Masson, Sirius red staining immunofluorescence, and wheat germ agglutinin staining were performed at 3 days, and 1, 2, and 4 weeks after MI modeling. Fibroblasts collected from SD neonatal rats were transfected by adenovirus and siRNA, and cell counting kit-8 (CCK8), immunofluorescence, wound healing and Transwell assay were conducted. Myocardial fibrosis-related proteins were identified by Western blot. PNU-74654 (100 ng/mL) was used for 12 hours to inhibit ß-catenin-TCF/LEF1 complex. Results: The up-regulation of TFEB resulted in reduced fibroblasts proliferation and its differentiation into myofibroblasts in vitro studies. A significant up-regulation of EF and down-regulation of myocyte area was shown in the AAV9-TFEB group. Meanwhile, decreased protein level of α-SMA and collagen I were observed in vitro study. TFEB didn't affect the concentration of ß-catenin. Inhibition of TFEB, which promoted cell migration, proliferation and collagen I expression, was counteracted by PNU-74654. Conclusions: TFEB demonstrated potential in restraining fibrosis after MI by inhibiting the Wnt/ß-catenin signaling pathway.

Identification and in vitro validation of diagnostic and prognostic biomarkers for lung squamous cell carcinoma.

Background: The aim of the present study was to find diagnostic and prognostic biomarkers for lung squamous cell carcinoma (LUSC) and to validate key biomarkers in vitro. Methods: RNA sequencing was used to identify differentially expressed mRNAs (DEmRNAs) and differentially expressed long non-coding RNAs (DElncRNAs) in LUSC tissues. RNA sequencing results were validated using a published dataset. Diagnostic and prognostic values of candidate genes were evaluated by receiver-operating characteristic (ROC) curve analysis and survival analysis, respectively. To determine the effect of MIR205HG in LUSC, MIR205HG expression was knocked down in NCI-H520 cells. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Transwell assay were used to respectively detect the effect of MIR205HG on cell proliferation and migration. Results: In total, 1,946 DEmRNAs and 428 DElncRNAs were identified in LUSC compared with normal tissues. A total of 851 DElncRNA-DEmRNA co-expression pairs were obtained. With the exception of NEAT1, MCM2, SERPINB5, ITGB8, CASC19, and MIR205HG were upregulated in LUSC. ROC curve analysis indicated that MCM2, SERPINB5, ITGB8, CASC19, and MIR205HG could predict LUSC. Survival analysis suggested that SERPINB5, NEAT1, and MIR205HG had potential prognostic value for LUSC. MIR205HG knockdown inhibited cell proliferation and migration, and significantly reduced the expression of ITGB8. Conclusions: The findings of the present study could help determine the pathogenesis of LUSC and provide new and accurate therapeutic targets for its treatment.

Laminin-bound integrin α6ß4 promotes non-small cell lung cancer progression via the activation of YAP/TAZ signaling pathway.

Laminin is an extracellular matrix multidomain trimeric glycoprotein, that has a potential role in tumor progression. Here, we studied the effects of non-small cell lung cancer (NSCLC) cells interaction on laminin and explored the underlying mechanism of laminin associated NSCLC progression. Culture of A549 and NCI-1299 cells on 2D collagen gels (containing laminin) significantly promoted the proliferative and tumorigenic characteristics, as well as cell invasion of tumor cells in vitro. Consistently, comparing the clinical NSCLC tumor tissues, a poor overall survival was observed in patients with high laminin expression. Mechanistically, the expression of integrin α6ß4 was required for the pro-tumor effects of laminin. Meanwhile, we showed that the downstream signaling of integrin α6ß4, involved the focal adhesion kinase (FAK)/Yes-Associated Protein (YAP)/TAZ signaling pathway. The activation of FAK/YAP/TAZ signaling pathway induced by laminin was validated in tumor tissues from NSCLC patients. Suppression of integrin α6ß4/FAK/YAP/TAZ signaling pathway efficiently suppressed the laminin-induced tumor growth, and strengthened the anticancer effects of chemotherapy, describing a novel target for NSCLC treatment.

Yin Yang 1-stimulated long noncoding RNA bladder cancer-associated transcript 1 upregulation facilitates esophageal carcinoma progression via the microRNA-5590-3p/programmed cell death-ligand 1 pathway.

Esophageal carcinoma (EC) is a common gastrointestinal malignancy that poses a threat to public health worldwide. Long noncoding RNA (lncRNA) bladder cancer-associated transcript 1 (BLACAT1) exerts a tumorigenic role in several malignant tumors; nevertheless, its function in EC remains largely unknown. Besides, programmed cell death-ligand 1 (PD-L1), an oncogene in numerous human cancers, has been identified as a therapeutic target for EC. Therefore, we intended to explore the potential regulatory network involving BLACAT1 and PD-L1 in EC. In this study, we observed increased BLACAT1 and PD-L1 levels in EC tissues and EC cell lines. Moreover, YY1 could activate BLACAT1 transcription in EC cells (TE-1 and EC9706). In addition, in vitro and in vivo experiments demonstrated that BLACAT1 facilitated EC cell proliferation and metastasis and EC tumor growth. Also, the effects of BLACAT1 silencing on EC cell functions were partially reversed by PD-L1 overexpression. Besides, it was identified that BLACAT1 competed with PD-L1 to bind to miR-5590-3p in EC cells. Furthermore, miR-5590-3p suppression could abrogate the functional effects of BLACAT1 knockdown on EC cells; while PD-L1 silencing partly abolished the promoting effects of miR-5590-3p suppression on the biological functions of EC cells. To sum up, YY1-induced BLACAT1 accelerated EC progression via regulating the miR-5590-3p/PD-L1 axis.

Identification and Validation of Novel Potential Pathogenesis and Biomarkers to Predict the Neurological Outcome after Cardiac Arrest.

Predicting neurological outcomes after cardiac arrest remains a major issue. This study aimed to identify novel biomarkers capable of predicting neurological prognosis after cardiac arrest. Expression profiles of GSE29540 and GSE92696 were downloaded from the Gene Expression Omnibus (GEO) database to obtain differentially expressed genes (DEGs) between high and low brain performance category (CPC) scoring subgroups. Weighted gene co-expression network analysis (WGCNA) was used to screen key gene modules and crossover genes in these datasets. The protein-protein interaction (PPI) network of crossover genes was constructed from the STRING database. Based on the PPI network, the most important hub genes were identified by the cytoHubba plugin of Cytoscape software. Eight hub genes (RPL27, EEF1B2, PFDN5, RBX1, PSMD14, HINT1, SNRPD2, and RPL26) were finally screened and validated, which were downregulated in the group with poor neurological prognosis. In addition, GSEA identified critical pathways associated with these genes. Finally, a Pearson correlation analysis showed that the mRNA expression of hub genes EEF1B2, PSMD14, RPFDN5, RBX1, and SNRPD2 were significantly and positively correlated with NDS scores in rats. Our work could provide comprehensive insights into understanding pathogenesis and potential new biomarkers for predicting neurological outcomes after cardiac arrest.

Comparison of Up-Front Minimally Invasive Esophagectomy versus Open Esophagectomy on Quality of Life for Esophageal Squamous Cell Cancer.

This study investigates whether minimally invasive esophagectomy (MIE) is a safe and effective way for patients with resectable esophageal cancer by comparing the short-term quality of life (QOL) after minimally invasive esophagectomy and open esophagectomy (OE). A total number of 104 patients who underwent esophagectomy from January 2013 to March 2014 were enrolled in this study. These patients were divided into two groups (MIE and OE group). Three scoring scales of quality of life were used to evaluate QOL before the operation and at the first, third, sixth and twelfth months after MIE or OE, which consist of Karnofshy performance scale (KPS), the European Organization for Research and Treatment questionnaire QLQC-30 (EORTC QLQC-30) and esophageal cancer supplement scale (OES-18). The MIE group was higher than the OE group in one-year survival rate (92.54% vs. 72.00%). Significant differences between the two groups were observed in intraoperative bleeding volume (158.53 ± 91.07 mL vs. 228.97 ± 109.33 mL, p = 0.001), and the incidence of postoperative pneumonia (33.33% vs. 58.62%, p = 0.018). The KPS of MIE group was significantly higher than the OE group at the first (80 vs. 70, p = 0.004 < 0.05), third (90 vs. 80, p = 0.006 < 0.05), sixth (90 vs. 80, p = 0.007 < 0.05) and twelfth months (90 vs. 80, p = 0.004 < 0.05) after surgery. The QLQC-30 score of MIE group was better than OE group at first and twelfth months after the operation. The OES-18 score of MIE group was significantly better than OE group at first, sixth and twelfth months after surgery. The short-term quality of life in MIE group was better than OE group.

[A retrospective comparison of thoracoscopic and laparoscopic esophagectomy between right neck anastomosis and left neck anastomosis].

OBJECTIVE: To evaluate the feasibility of right neck anastomosis in thoracoscopic and laparoscopic esophagectomy. METHODS: This study used a retrospective cohort study method. Clinical data of 169 patients with stage I-III esophageal squamous cell carcinoma undergoing neck anastomosis in thoracoscopic and laparoscopic esophagectomy at the Department 5 of Thoracic Surgery, the Fourth Hospital of Hebei Medical University from November 2013 to October 2016 were retrospectively analyzed. Eighty-two cases underwent right neck anastomosis (right neck anastomosis group) and 87 cases underwent left neck anastomosis(left neck anastomosis group). Both groups underwent routine thoracoscopic and laparoscopic radical resection of esophageal cancer. The entry of right and left neck anastomosis group was at the anterior edge of the right and left sternocleidomastoid muscle respectively. Anastomosis of the esophagogastric junction was performed and the drainage tube was placed in the neck incision. The operation time, intraoperative blood loss, lymph node dissection and morbidity of postoperative complications were compared between the two groups. RESULTS: There were 101 males and 68 females among 169 patients with esophageal cancer. There were no significant differences in age, gender, tumor location, clinical stage between two groups(all P>0.05). The total operation time of left and right neck anastomosis groups was (278.3±39.4) minutes and (287.8±39.4) minutes, respectively (t=1.563, P=0.120). The intraoperative blood loss was (134.9±71.5) ml and(147.9±85.5) ml, respectively (t=1.074, P=0.284). The number of lymph node dissections was (17.45±5.68) and (16.47±4.98), respectively (t=1.190, P=0.236). Seventeen cases(20.7%) in the right neck anastomosis group developed postoperative complications, while 31 cases (35.6%) in the left neck anastomosis group developed postoperative complications (χ²=4.609,P=0.032). Compared with left neck anastomosis group, right neck anastomosis group had lower rate of gastric emptying disorder (0% vs. 6.9%, P=0.029), anastomotic fistula (7.3% vs. 18.4%, χ²=4.572, P=0.033), pneumonia (18.3% vs. 32.2%, χ²=4.294, P=0.038) and ICU management (4.9% vs. 16.1%, χ²=4.726, P=0.030). CONCLUSION: Thoracoscopic and laparoscopic esophagectomy with right neck anastomosis is safe and effective, can completely remove the tumor, at the same time, has less complications than left neck anastomosis, and improve the quality of life.