scholarly journals miR-20b-5p attenuates hypoxia-induced apoptosis in cardiomyocytes via the HIF-1α/NF-κB pathway

2020 ◽  
Vol 52 (9) ◽  
pp. 927-934 ◽  
Author(s):  
Zhongquan Zhou ◽  
Songwen Chen ◽  
Zhiming Tian ◽  
Shibing Deng ◽  
Xuying Yi ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Target Genes ◽  
Annexin V ◽  
Pcr Analysis ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Flow Cytometry Analysis ◽  
Low Oxygen

Abstract Chronic hypoxia is a common inducer of end-stage cardiovascular disease. In cells under hypoxia, the hypoxia-inducible factor-1 (HIF-1) plays a vital role in regulating downstream target genes. However, the mechanism of hypoxia in cardiomyocytes is still unclear. In this study, we aimed to identify novel downstream epigenetic targets of HIF-1α in cardiomyocytes under hypoxia. H9c2 cells were exposed to hypoxia condition, and quantitative real-time PCR analysis was performed to evaluate the expression of miR-20b-5p. The results indicated that the expression of miR-20b-5p was down-regulated in H9c2 cells under low oxygen condition. Meanwhile, HIF-1α overexpression further down-regulated the miR-20b-5p expression in H9c2 cells transfected with HIF-1α plasmids. In addition, Annexin-V-FITC/PI flow cytometry analysis suggested that overexpression of miR-20b-5p attenuated cell apoptosis under hypoxia condition in H9c2 cells. Western blot analysis showed that the hypoxia apparently increased Bax and cleaved-caspase-3, but decreased Bcl-2 expression in H9c2 cells, indicating that hypoxia-induced NF-κB signaling pathway activation is mediated by miR-20b-5p. Hypoxia-induced H9c2 cell apoptosis was reduced after HIF-1α knockdown as shown by the flow cytometry analysis. In conclusion, we identified that miR-20b-5p plays an important role in mediating cardiomyocytes apoptosis under hypoxia, which is mediated by the HIF-1/NF-κB signaling pathway.

scholarly journals Propofol Alleviates Apoptosis Induced by Chronic High Glucose Exposure via Regulation of HIF-1α in H9c2 Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-13 ◽  
Author(s):  
Jinjun Pu ◽  
Shun Zhu ◽  
Dandan Zhou ◽  
Lidong Zhao ◽  
Ming Yin ◽  
...  
Keyword(s):  
Cell Viability ◽  
Proinflammatory Cytokines ◽  
High Glucose ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Mrna Level ◽  
Pcr Analysis ◽  
H9c2 Cell ◽  
Protective Mechanism ◽  
H9c2 Cells

Background. The sedative anesthetic, propofol, is a cardioprotective agent for hyperglycemia-induced myocardial hypertrophy and dysfunction in rats. However, the specific protective mechanism has not been clarified. Methods and Results. In this experiment, we used H9c2 cells subjected to 22 mM glucose lasting for 72 hours as an in vitro model of cardiomyocyte injury by hyperglycemia and investigated the potential mechanism of propofol against hyperglycemic stress in cells. Propofol (5, 10, or 20 μM) was added to the cell cultures before and during the high glucose culture phases. Cell viability and levels of ROS were measured. The levels of proinflammatory cytokines were tested by ELISA. The levels of SIRT3, SOD2, PHD2, HIF-1α, Bcl-2, P53, and cleaved caspase-3 proteins were detected by western blotting. Our data showed that propofol attenuated high glucose-induced cell apoptosis accompanied by a decrease in the level of reactive oxygen species (ROS) and proinflammatory cytokines. Meanwhile, propofol decreased the apoptosis of H9c2 cells via increasing the expression of Bcl-2, SIRT3, SOD2, and PHD2 proteins and decreasing the expression of cleaved caspase-3, P53, and HIF-1α. Real-time PCR analysis showed that propofol did not significantly change the HIF-1α but increase PHD2 at mRNA level. HIF-1α silence significantly decreased apoptosis and inflammation in H9c2 cell during high glucose stress. Pretreatment of IOX2 (the inhibitor of PHD2) inhibited cell viability until the concentration reached 200 μM during high glucose stress. However, 50 μM TYP (the inhibitor of SIRT3) significantly inhibited cell viability during high glucose stress. Delayed IOX2 treatment for 6 hours significantly inhibited cell viability during high glucose stress. Conclusions. Propofol might alleviate cell apoptosis via SIRT3-HIF-1α axis during high glucose stress.

Propranolol induced apoptosis and autophagy via the ROS/JNK signaling pathway in human ovarian cancer

2020 ◽  
Author(s):  
Shujun Zhao ◽  
Suzhen Fan ◽  
Yanyu Shi ◽  
Hongyan Ren ◽  
Hanqing Hong ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Flow Cytometry ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Human Ovarian Cancer ◽  
Dependent Manner ◽  
Flow Cytometry Analysis ◽  
Jnk Signaling ◽  
Jnk Signaling Pathway ◽  
Induced Apoptosis

Abstract Background: Propranolol has a significant anti-cancer effect on various cancers. The present study aimed to investigate the underlying mechanism behind the therapeutic effect of Propranolol on the ovarian cancer.Materials and methods: The effect of Propranolol on cell viability was examined by MTT analysis. Cellular apoptosis was evaluated by flow cytometry analysis. Autophagy was defined by autophagosome observed by confocal microscopy after infected with GFP-LC3 adenovirus. In addition, the expression of marker proteins involved in cell apoptosis, autophagy, and ROS/JNK signaling pathway were estimated by Western Blotting assay.Results: Propranolol significantly reduced the viability of human ovarian cancer cell lines SKOV-3 and A2780 in a dose- and time-dependent manner. Flow cytometry analysis revealed that Propranolol induced the cell cycle arrest at G2/M phase and resulted in apoptosis. Moreover, autophagy inhibitor 3-MA markedly enhanced the Propranolol-induced apoptosis. In addition, reactive oxygen species (ROS) was demonstrated dramatically increased after Propranolol treatment and Propranolol activated the phosphorylation of JNK. What is more, p38 inhibitor SB203580 and JNK inhibitor SP600125 attenuated the upregulated expression of LC3-II and cleaved-caspase-3 by the effect of Propranolol. ROS exclusive inhibitor antioxidant N-acetyl cysteine (NAC) weaken the phosphorylation of JNK proteins induced by Propranolol.Conclusions：In summary, our results suggested that Propranolol induced cell apoptosis and protective autophagy through the ROS/JNK signaling pathway in human ovarian cancer cells.

scholarly journals Naoxintong/PPARαSignaling Inhibits H9c2 Cell Apoptosis and Autophagy in Response to Oxidative Stress

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
Huimin Xu ◽  
Jianhua Jin ◽  
Lu Chen ◽  
Chunxiao Li ◽  
Qinggang Xu ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Flow Cytometry ◽  
Chinese Medicine ◽  
Cardiovascular Diseases ◽  
Protective Effect ◽  
Cell Apoptosis ◽  
H9c2 Cell ◽  
Protective Mechanism ◽  
H9c2 Cells ◽  
Protein Levels

Naoxintong (NXT) is an empirical formula based on the principle of traditional Chinese medicine, which has been approved by China Food and Drug Administration (CFDA) and is widely used for treatment of patients with cerebrovascular and cardiovascular diseases in China. The aim of this study is to investigate the protective mechanism of NXT on H9c2 cells (cardiogenic cell line) in response to H2O2. MTT, Western blot, and flow cytometry (FCM) methods were used to identify the protective effect of NXT extract on H2O2-induced H9c2 cells. Here we found that NXT extract significantly increased H9c2 cell viability and reduced H2O2-induced cell apoptosis and autophagy. More importantly, NXT inhibited H2O2-induced H9c2 cell apoptosis and autophagy by increasing PPARαprotein levels. In contrast, silenced PPARαterminated NXT protective effect on H2O2-induced H9c2 cells. These findings suggest that NXT/PPARαsignaling suppressed H2O2-induced H9c2 cell apoptosis and autophagy.

scholarly journals MiR-218 Promotes Adriamycin-Induced H9C2 Apoptosis by Inhibiting Stress-Associated Endoplasmic Reticulum Protein 1

2022 ◽  
Vol 2022 ◽  
pp. 1-10
Author(s):  
Qinghua Chen ◽  
Gang Chen ◽  
Shuofang Zhao
Keyword(s):  
Signaling Pathway ◽  
P38 Mapk ◽  
Cell Apoptosis ◽  
Cell Activity ◽  
Mapk Signaling ◽  
Luciferase Reporter ◽  
Tunel Staining ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Gene Assay

Objective. Adriamycin is a clinically important chemotherapeutic drug, but its use is restricted due to its myocardial toxicity. Therefore, it is especially important to explore the toxicity mechanism of Adriamycin (ADR) to cardiomyocytes. Methods. The myocardial toxicity model of ADR was constructed in vitro, and the effect of miR-218 inhibitor and sh-Serp1 on the activity of H9C2 cells induced by ADR was detected by MTT method. Also, flow cytometry, real-time polymerase chain reaction (RT-PCR), and TUNEL staining were used to detect the cell apoptosis. The activity of LDH was detected by colorimetry, and the interaction of miR-218 with Serp1 was detected by double-luciferase reporter gene assay. Western blotting technique was used to detect the expression level of caspase3 and p38 MAPK signal pathway. Results. miR-218 inhibitor can obviously inhibit ADR-induced decrease in cell activity of H9C2 cells, inhibit cell apoptosis, and inhibit p38 MAPK signaling pathway activation. Conversely, sh-Serp1 aggravated the decrease in H9C2 cell activity and promoted cell apoptosis. Conclusion. Upregulation of miR-218 expression will promote ADR-induced apoptosis of H9C2 cells. At the same time, we confirmed that the mechanism by which miR-218 promotes myocardial apoptosis was through the Serp1/p38 MAPK/caspase-3 signaling pathway.

scholarly journals miR-146a-5p Mediates Intermittent Hypoxia-Induced Injury in H9c2 Cells by Targeting XIAP

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Guofu Lin ◽  
Jiefeng Huang ◽  
Qingshi Chen ◽  
Lida Chen ◽  
Dehuai Feng ◽  
...  
Keyword(s):  
Cell Apoptosis ◽  
Intermittent Hypoxia ◽  
Target Genes ◽  
Cardiac Injury ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Obstructive Sleep ◽  
Apoptosis Protein

MicroRNAs (miRNAs) have emerged as key modulators in the pathophysiologic processes of cardiovascular diseases. However, its function in cardiac injury induced by obstructive sleep apnea (OSA) remains unknown. The aim of the current study was to identify the effect and potential molecular mechanism of miR-146a-5p in intermittent hypoxia(IH)- induced myocardial damage. We exposed H9c2 cells to IH condition; the expression levels of miR-146a-5p were detected by RT-qPCR. Cell viability, cell apoptosis, and the expressions of apoptosis-associated proteins were assessed via Cell Counting Kit-8 (CCK-8), flow cytometry, and western blotting, respectively. Target genes of miR-146a-5p were confirmed by dual-luciferase reporter assay. IH remarkably lowered viability but enhanced cell apoptosis. Concomitantly, the miR-146a-5p expression level was increased in H9c2 cells after IH. Subsequent experiments showed that IH-induced injury was alleviated through miR-146a-5p silence. X-linked inhibitor of apoptosis protein (XIAP) was predicted by bioinformatics analysis and further confirmed as a direct target gene of miR-146a-5p. Surprisingly, the effect of miR-146a-5p inhibition under IH may be reversed by downregulating XIAP expression. In conclusion, our results demonstrated that miR-146a-5p could attenuate viability and promote the apoptosis of H9c2 by targeting XIAP, thus aggravating the H9c2 cell injury induced by IH, which could enhance our understanding of the mechanisms for OSA-associated cardiac injury.

scholarly journals LncRNA PVT1 Promotes Hypoxia-Induced Cardiomyocyte Injury by Inhibiting miR-214-3p

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Chuanliang Liu ◽  
Jieqiong Zhang ◽  
Xuejie Lun ◽  
Lei Li
Keyword(s):  
Cell Viability ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Flow Cytometry Analysis ◽  
Chain Reaction ◽  
Cell Vitality ◽  
Gene Assay ◽  
Polymerase Chain

Objective. To explore the effect and related mechanism of LncRNA PVT1 on hypoxia-induced cardiomyocyte injury. Methods. PVT1RNA and miR-214-3p levels were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Cell vitality and apoptosis were, respectively, evaluated by Cell Counting Kit-8 (CCK-8) and flow cytometry analysis. Starbase and Dual luciferase reporter (DLR) gene assay was employed to validate the interaction between miR-214-3p and PVT1. Results. PVT1 was statistically upregulated, and miR-214-3p was statistically downregulated in hypoxia-induced H9c2 cells. The survival rate of H9c2 cells induced by hypoxia decreased statistically, while the apoptosis rate increased statistically ( P < 0.05 ). PVT1 knockdown upregulated the hypoxia-induced H9c2 cell viability and inhibited apoptosis. DLR assay verified the targeting relationship between PVT1 and miR-214-3p. In addition, miR-214-3p inhibitors reversed the viability of H9c2 cells with PVT1 knockout and promoted apoptosis. Conclusion. Silencing PVT1 can enhance the hypoxia-induced H9c2 cell viability and inhibit apoptosis, providing a potential target for the treatment of cardiovascular diseases.

Dysregulation of tRNA-derived small RNAs and their potential roles in lupus nephritis

Lupus ◽  
2021 ◽  
pp. 096120332110614
Author(s):  
Yan Liang ◽  
Ji Zhang ◽  
Wenxian Qiu ◽  
Bo Chen ◽  
Ying Zhou ◽  
...  
Keyword(s):  
Lupus Nephritis ◽  
Signaling Pathway ◽  
Small Rnas ◽  
High Throughput Sequencing ◽  
Target Genes ◽  
Expression Profiles ◽  
Mapk Signaling ◽  
Pcr Analysis ◽  
Clinical Indicators ◽  
Qrt Pcr

Objective Lupus nephritis (LN) is a major end-organ complication of systemic lupus erythematosus (SLE), and the molecular mechanism of LN is not completely clear. Accumulating pieces of evidence indicate the potential vital role of tRNA-derived small RNAs (tsRNAs) in human diseases. Current study aimed to investigate the potential roles of tsRNAs in LN. Methods We herein employed high‐throughput sequencing to screen the expression profiles of tsRNAs in renal tissues of the LN and control groups. To validate the sequencing data, we performed quantitative real-time PCR (qRT-PCR) analysis. Correlational analysis of verified tsRNAs expression and clinical indicators was conducted using linear regression. The potential target genes were also predicted. The biological functions of tsRNAs were annotated by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Results Our findings revealed that the expression profiles of tsRNAs were significantly altered in the kidney tissues from LN patients compared with control. Overall, 160 tsRNAs were significantly dysregulated in the LN group, of which 79 were upregulated, whereas 81 were downregulated. Subsequent qRT-PCR results confirmed the different expression of candidate tsRNAs. Correlation analysis results found that expression of verified tsRNAs were correlated to clinical indicators. The target prediction results revealed that verified tsRNAs might act on 712 target genes. Further bioinformatics analysis uncovered tsRNAs might participate in the pathogenesis of LN through several associated pathways, including cell adhesion molecules, MAPK signaling pathway, PI3K-Akt signaling pathway and B cell receptor signaling pathway. Conclusion This study provides a novel insight for studying the mechanism of LN.

scholarly journals MACC1 Suppresses Cell Apoptosis in Hepatocellular Carcinoma by Targeting the HGF/c-MET/AKT Pathway

2015 ◽  
Vol 35 (3) ◽  
pp. 983-996 ◽  
Author(s):  
Yingmin Yao ◽  
Chanwei Dou ◽  
Zhongtang Lu ◽  
Xin Zheng ◽  
Qingguang Liu
Keyword(s):  
Cell Apoptosis ◽  
Target Genes ◽  
Nuclear Translocation ◽  
Annexin V ◽  
Akt Pathway ◽  
Caspase 9 ◽  
Dapi Staining ◽  
In Vivo Experiments

Background & Aims: To investigate the expression and prognostic value of MACC1 in patients with HCC and identify the mechanism by which MACC1 inhibits HCC cell apoptosis. Methods: MACC1 and p-AKT expression was studied using immunohistochemistry of both HCC tissues and adjacent liver tissues. qRT-PCR and western immunoblotting were used to examine the expression of target genes at the mRNA and protein levels, respectively. The MTT assay was used to assess cell viability, and cell apoptosis was determined by DAPI staining, Annexin V/PI staining and Caspase 3/7 assay. Nude mice were used to perform in vivo experiments. Results: The overexpression of MACC1 was found in HCC tissues and was correlated with poor postsurgical prognosis. There was a positive relationship between MACC1 and p-AKT expression in HCC tissues. In vitro experiments showed that MACC1 repressed HCC cell apoptosis and promoted cell growth. Knockdown of c-MET abolished the anti-apoptotic function of MACC1. Next, MACC1 was verified to activate PI3K/AKT signaling by sensitizing HGF/c-MET signaling in HCC. MACC1 overexpression enhanced the HGF-driven phosphorylation of BAD, Caspase 9 and FKHRL1 and inhibited their pro-apoptotic functions in HCC cells. Finally, MACC1 was shown to inhibit cell apoptosis and promote HCC growth in vivo. Conclusions: This investigation revealed that MACC1 overexpression predicted worse prognosis after liver resection, which was attributed to the repression of HCC cell apoptosis via a molecular mechanism in which MACC1 accelerated the activation of the HGF/c-MET/PI3K/AKT pathway and phosphorylated BAD, Caspase 9 and FKHRL1, ultimately preventing their nuclear translocation and their pro-apoptotic function.

scholarly journals Long non-coding RNA RP11-468E2.5 curtails colorectal cancer cell proliferation and stimulates apoptosis via the JAK/STAT signaling pathway by targeting STAT5 and STAT6

2019 ◽  
Vol 38 (1) ◽  
Author(s):  
Li Jiang ◽  
Xu-Hai Zhao ◽  
Yin-Ling Mao ◽  
Jun-Feng Wang ◽  
Hui-Jun Zheng ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Cycle ◽  
Cell Proliferation ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Target Genes ◽  
Biological Activities ◽  
Janus Kinase ◽  
Normal Tissues ◽  
Stat Signaling

Abstract Background Long non-coding RNAs (lncRNAs) are tumor-associated biological molecules and have been found to be implicated in the progression of colorectal cancer (CRC). This study aims to examine the effects of lncRNA RP11-468E2.5 and its target genes (STAT5 and STAT6) on the biological activities of CRC cells via the Janus kinase-signal transducer and activator of transcription (JAK/STAT) signaling pathway. Methods We initially screened the GEO database for differentially expressed lncRNAs related to CRC and then made a prediction of the implicated target genes. Then we collected CRC tissues and adjacent normal tissues from 169 CRC patients. Human CRC HCT116 and SW480 cells were treated with small interference RNA (siRNA) against RP11-468E2.5, AG490 (an inhibitor of the JAK/STAT signaling pathway), or both in combination. Next, we measured the effects of RP11-468E2.5 treatment on cellular activities such as cell viability, cycle distribution and cell apoptosis, and studied interactions among RP11-468E2.5, STAT5/STAT6, and the JAK/STAT signaling pathway. Finally, an in vivo tumor formation assay was performed to observe the effect of RP11-468E2.5 on tumor growth. Results The CRC-related gene microarray data showed low expression of RP11-468E2.5 in CRC surgical specimens. However, RP11-468E2.5 was confirmed to target STAT5 and STAT6, which participate in the JAK/STAT signaling pathway. CRC tissues showed lower expression of RP11-468E2.5, higher expression of STAT5, STAT6 and of the cell cycle marker Cyclin D1 (CCND1), compared to the findings in adjacent normal tissues. The treatment of siRNA against RP11-468E2.5 increased expression of JAK2, STAT3, STAT5, STAT6, CCND1 and Bcl-2 along with the extent of STAT3, STAT5 and STAT6 phosphorylation, while lowering expression of P21 and P27. Treatment with AG490 exhibited approximately opposite effects, whereas siRNA against RP11-468E2.5 treatment stimulated CRC cell proliferation and reduced cell apoptosis, while promoting cell cycle entry; AG490 treatment reversed these results. Conclusions Altogether, we conclude that up-regulation of RP11-468E2.5 inhibits the JAK/STAT signaling pathway by targeting STAT5 and STAT6, thereby suppressing cell proliferation and promoting cell apoptosis in CRC.

scholarly journals Oxymatrine Induces Apoptosis in Human Myeloma Cells By Cell Cycle Arrest and Activation of Caspase-Dependent Apoptosis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5727-5727
Author(s):  
Wenjun Wu ◽  
Cai Wu ◽  
Fuming Zi ◽  
Yi Li ◽  
Li Yang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Flow Cytometry ◽  
Growth Inhibition ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Western Blotting ◽  
Cell Apoptosis ◽  
Conflicts Of Interest ◽  
Dependent Manner ◽  
Rt Pcr

Abstract Background : Multiple myeloma (MM) is a B cell malignant hematologic cancer. Despite the introduction of new drugs and improvement of chemotherapy, MM is still an incurable disease. Oxymatrine (OMT), the active ingredients of traditional Chinese herbal medicine sophora, has been reported to have antitumor activity. This study was to estimate the therapeutic efficacy of OMT in MM. Methods: The growth inhibition of myeloma cell lines (RPMI8226, U266, ARP-1) or primary cells by OMT was assessed by MTT assay. Apoptosis of MM cells was examined by annexin V-FITC using flow cytometry analysis. DNA content was analyzed by flow cytometry. RT-PCR and western-blot analysis were used to assess the expression of Bcl-2 family proteins and the IAP family proteins. Western blotting was also used to elucidate the signaling pathway that may mediate OMT-induced apoptosis of MM cells. Results: OMT treatment resulted in cell growth inhibition and apoptosis in primary MM cells and all tested MM cell lines in a dose-dependent manner (P <0.05). To elucidate OMT -induced MM cell apoptosis, MM cell lines were treated with or without OMT for 24h and assessed for caspase activation and signaling pathway by Western blotting. The results showed the cleavage of PARP, caspase-3, and caspase-9, and p-AKT were down-regulated after OMT treatment. The mRNA expression of survivin and HIAP by RT-PCR was down-regulated. OMT treatment at 5mM for 48h resulted in increased G-phase cells and decreased S-phase cells in MM cell lines (P <0.05). Cell cycle repressor P21 protein was up-regulated while CDK4, CDK6 and CyclinD1 expression was down-regulated. Our finding also showed a synergistic anti-MM activity of OMT and dexamethasone or adriamycin at a low does (CI<1). In addition, LC3-II expression was significantly increased both in RPMI8226 and U266 cells after treatment with OMT. However, treatment with different doses of OMT and 5 mM autophagy inhibitor 3-MA, significant increased cell apoptosis (P <0.05). Conclusion: Our findings demonstrate the anti-MM activity of OMT and indicate that OMT alone or together with other MM chemotherapeutics may be a prospective treatment for MM. Disclosures No relevant conflicts of interest to declare.

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