scholarly journals Silencing of miR-223 expression inhibits the apoptosis of H2O2-induced cardiomyocytes by increasing the activity of the IGF-1R/PI3K/AKT pathway

2020 ◽  
Vol 18 ◽  
pp. 205873922095087
Author(s):  
Qianqian Zhou ◽  
Yufen Li ◽  
Tianjin Gu
Keyword(s):  
Cell Apoptosis ◽  
Cell Activity ◽  
B Cell Lymphoma ◽  
Luciferase Reporter ◽  
Akt Pathway ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Luciferase Reporter Gene ◽  
Related Proteins ◽  
Low Expression

To study the: (1) function of micro (mi)R-223 on H2O2-induced H9C2 cells; (2) relationship between miR-223 and insulin-like growth factor 1 receptor (IGF-1R); and (3) role of miR-223 on the phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) pathway. H9C2 cells were selected to establish the H2O2-injury model. Overexpression/low expression of miR-223 in H9C2 cells was constructed, respectively. Flow cytometry and western blotting were applied to measure the apoptosis, cell activity, and expression of related proteins. Dual-luciferase reporter gene assays (DLRGAs) were applied to test if miR-223 targeted IGF-1R. Overexpression/low expression of IGF-1R was constructed to test if miR-223 regulated IGF-1R expression negatively. Increases in miR-223 expression were observed in H2O2-induced H9C2 cells. miR-223 absence improved H2O2-induced H9C2-cell apoptosis accompanied by an increase in B-cell lymphoma (Bcl)-2 expression and decrease in expression of Bax and cleaved caspase-3 ( P < 0.05). miR-223 silencing increased expression of IGF-1R, p-PI3K, and p-AKT in H2O2-induced H9C2 cells ( P < 0.05). miR-223 overexpression aggravated H2O2-induced H9C2-cell apoptosis and reduced expression of the proteins of IGF-1R, p-PI3K, and p-AKT. DLRGAs showed IGF-1R to be a downstream gene of miR-223. IGF-1R silencing significantly inhibited expression of p-PI3K and p-AKT proteins ( P < 0.05). miR-223 negatively regulated IGF-1R expression for H9C2-cell apoptosis and the PI3K/AKT pathway. miR-223 absence can ameliorate H2O2-induced cardiomyocyte apoptosis by targeting IGF-1R to regulate PI3K/AKT activity.

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 PROLIFERATION AND APOPTOSIS OF B-CELL LYMPHOMA CELLS UNDER TARGETED REGULATION OF FOXO3 BY miR-155

2020 ◽  
Vol 12 (1) ◽  
pp. e2020073
Author(s):  
Xiaoqiang Zheng ◽  
Hongbing Rui ◽  
Ying Liu ◽  
Jinfeng Dong
Keyword(s):  
B Cell ◽  
B Lymphocytes ◽  
Cell Apoptosis ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Luciferase Reporter ◽  
Lymphoma Cells ◽  
Apoptosis Rate ◽  
Proliferation And Apoptosis ◽  
Low Expression

This study aimed to explore the proliferation and apoptosis of B-cell lymphoma cells under targeted regulation of FOXO3 by miR-155. We analyzed the differences between B-cell lymphoma cells and B lymphocytes in expressions of miR-155 and FOXO3, explored the effects of miR-155 on proliferation and apoptosis of B-cell lymphoma cells, and relevant mechanisms, and also analyzed the relationship between expressions of miR-155 and FOXO3 in 42 patients with diffuse large B-cell lymphoma (DLBCL) and clinical characteristics of them. B-cell lymphoma cells showed a higher expression of miR-155 and a low expression of FOXO3 than B lymphocytes (both P<0.05). B-cell lymphoma cells transfected with miR-155-inhibitor showed significantly decreased expression of miR-155, significantly weakened cell proliferation ability and increased cell apoptosis rate (all P<0.05), and they also showed up-regulated expression of FOXO3 (P<0.05). Dual luciferase reporter assay revealed that there were targeted binding sites between miR-155 and FOXO3. Compared with B-cell lymphoma cells transfected with miR-155-inhibitor alone, those with co-transfection showed lower expression of FOXO3, higher proliferation and lower cell apoptosis rate (all P<0.05). The expression of miR-155 in DLBCL tissues was higher than that in tumor-adjacent tissues (P<0.05), and the expressions of miR-155 and FOXO3 were closely related to the international prognostic index (IPI) and the 5-year prognosis and survival of the patients (P<0.05). miR-155 can promote the proliferation of B-cell lymphoma cells and suppress apoptosis of them by targeted inhibiting FOXO3, and both over-expression of miR-155 and low expression of FOXO3 are related to poor prognosis of DLBCL patients.

miR-27b-3p Down-Regulation Prevents Hypoxia-Induced Cardiomyocyte Apoptosis Through Regulating Yes-Associated Protein 1 (YAP1) Expression

2021 ◽  
Vol 11 (5) ◽  
pp. 948-956
Author(s):  
Lilin Wang ◽  
Bo Feng ◽  
Shu Zhu
Keyword(s):  
Cell Viability ◽  
Target Genes ◽  
Luciferase Reporter ◽  
Pcr Analysis ◽  
H9c2 Cells ◽  
Luciferase Reporter Gene ◽  
Western Blot Assay ◽  
Heart Protection ◽  
Gene Assay ◽  
Related Proteins

Background: Congenital heart disease (CHD) is one of the most common birth defects. MicroR-NAs (miRNAs) are a group of endogenous, non-coding small RNAs and mediate the target genes expression. An increasing evidence showed that in recent years, miRNAs have given rise to more and more attention in heart protection and development. In our research, the main purpose was to determine the effect of miR-27b-3p in CHD and analyze related mechanisms. Methods: We performed qRT-PCR analysis to examine miR-27b-3p expression in myocardial tissue from 30 patients with CHD and hypoxia-induced H9C2 cells. Then, we performed biological software TargetScan to predict the relationship of miR-27b-3p and YAP1, and dual luciferase reporter gene assay was used to verify the results. H9C2 cells were transfected with inhibitor control, miR-27b-3p inhibitor, miR-27b-3p inhibitor + control-siRNA or miR-27b-3p inhibitor + YAP1-siRNA for 6 hours and then induced by hypoxia for 72 hours. Subsequently, we performed MTT and FCM analysis to detect cell viability and apoptosis. Finally, we used western blot assay to measure the expression of apoptosis-related proteins. Results: Our study indicated that miR-27b-3p expression in myocardial samples of cyanotic CHD patients was significantly higher than that of the acyanotic CHD patients. miR-27b-3p expression was gradually up-regulated with the increase of hypoxia induction time in H9C2 cells. Besides, we confirmed that YAP1 was a target gene of miR-27b-3p. Moreover, our results showed that miR-27b-3p inhibitor improved cell viability, decreased apoptosis, and affected apoptosis-related proteins expression in hypoxia induced H9C2 cells. These changes were reversed by YAP1-siRNA. All data demonstrated that miR-27b-3p/YAP1 might be new potential bio-marker and therapeutic target for CHD treatment.

scholarly journals Mir10a overexpression aggravates the renal ischemia-reperfusion injury associated with a decrease in PIK3CA

2020 ◽  
Author(s):  
Dongsheng Xu ◽  
Wenjun Li ◽  
Tao Zhang ◽  
Gang Wang
Keyword(s):  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Luciferase Reporter ◽  
Akt Pathway ◽  
Luciferase Reporter Gene ◽  
Gene Assay ◽  
Underlying Mechanisms ◽  
Related Proteins ◽  
Hypoxia Reoxygenation

Abstract Background: To investigate the effect of miR-10a on the renal tissues with ischemia-reperfusion (I/R) injury in rats and explore the underlying mechanisms of miR-10a in the HK-2 cells of hypoxia-reoxygenation. Methods: The miR-10a level was measured in renal tissues with I/R rats by RT-PCR. In order to research the role of miR-10a in the renal tissues, miR-10 agonist and miR-10a antagonist were used to treat I/R rats. The levels of serum creatinine (Scr) and blood urea nitrogen (BUN) in serum, renal histopathology, apoptosis of cells in renal tissues were analyzed, separately. The phosphatidylinositol 3-kinase (PI3K)/Akt pathway related proteins were measured by Western blot. The HK-2 cell was cultured to study the mechanism of miR-10a in the model of hypoxia-reoxygenation. The dual luciferase reporter gene assay was used to confirm the PI3K p100 catalytic subunit α (PIK3CA) was a target gene of miR-10a. Results: After renal I/R injury in rats, the miR-10a expression was significantly increased (p<0.05). Injection of miR-10a agonist significantly aggravated the injury of renal and raised the apoptosis of cells in renal in rats with renal I/R injury (p<0.05). However, administration of miR-10a antagonist obviously improved the injury of renal, decreased the renal cells apoptosis and inhibited the PI3K/Akt pathway activity (p<0.05). In vitro experiments, the negative relation between PIK3CA and miR-10a was confirmed. Further, overexpression of miR-10a significantly decreased the proliferation of HK-2 cells, and increased the cells apoptosis via up-regulating PI3K/Akt pathway (p<0.05). Conclusion: miR-10a could aggravate the renal I/R injury associated with a decrease in PIK3CA/PI3K/Akt 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 MiR-326 mediates malignant biological behaviors of lung adenocarcinoma by targeting ZEB1

2021 ◽  
Vol 104 (2) ◽  
pp. 003685042110093
Author(s):  
Mingxin Liu ◽  
Hong Wu ◽  
Yiqiang Liu ◽  
Yan Tan ◽  
Songtao Wang ◽  
...  
Keyword(s):  
Lung Adenocarcinoma ◽  
Cell Lines ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Invasion And Migration ◽  
Transwell Invasion Assay ◽  
Enhancer Binding Protein ◽  
Adenocarcinoma Cells ◽  
And Migration ◽  
Related Proteins

MiR-326 functions as an antioncogene in the several types of cancer. However, the underling mechanisms through which miRNA-326 regulates the anti-carcinogenesis of lung adenocarcinoma have remained elusive. The aim of this study was to explore the role and regulatory mechanism of miR-326 in cell proliferation, invasion, migration and apoptosis in lung adenocarcinoma. Quantitative real-time PCR (qRT-PCR) was used to detect the expression pattern of miR-326 in human bronchial epithelial cells (HBES-2B), 4 kinds of lung adenocarcinoma cell lines (H23, H1975, H2228, H2085) and 20 lung adenocarcinoma tissues. Then, H23 cells were infected with miR-326 mimics, miR-326 inhibitors and si-ZEB1 to build up-regulated miR-326 cell lines, down-regulated ZEB1(zinc-finger-enhancer binding protein 1)cell lines, simultaneous down-regulated ZEB1 and miR-326 cell lines. Moreover, CCK-8 assay, transwell invasion assay, wound healing assay and flow cytometry assay were employed to examine the effects of miR-326 and ZEB1 on the proliferation, invasion, migration and apoptosis abilities of H23 cells. Western blot was performed to explore the effects of miR-326 and ZEB1 on the expression of invasion and migration related proteins N-cadherin, E-cadherin, MMP7, MMP13, SLUG and apoptotic proteins PARP, BAX. On the mechanism, a dual-luciferase reporter gene was used to measure the target relationship between miR-326 and ZEB1. MiR-326 expression was significantly downregulated in lung adenocarcinoma tissues and cells. Overexpression of miR-326 significantly inhibited the malignant behaviors of H23 cells. Mechanically, luciferase reporter assay showed that ZEB1 was a direct target of miR-326. MiR-326 mimic downregulated the expression of ZEB1. Furthermore, knocking down ZEB1 strongly inhibited the proliferation, invasion and migration of H23 cells but promoted apoptosis. MiR-326 could target ZEB1 to inhibit the proliferation, invasion and migration of lung adenocarcinoma cells and promote apoptosis, which is a potential therapeutic target for lung adenocarcinoma.

scholarly journals MiR-206 regulates the progression of osteoporosis via targeting HDAC4

2021 ◽  
Vol 26 (1) ◽  
Author(s):  
Zhiyuan Lu ◽  
Dawei Wang ◽  
Xuming Wang ◽  
Jilong Zou ◽  
Jiabing Sun ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Apoptosis ◽  
Diagnostic Value ◽  
Expression Level ◽  
Luciferase Reporter ◽  
Control Group ◽  
Luciferase Reporter Gene ◽  
Mineral Density ◽  
Gene Assay

Abstract Background More and more studies have confirmed that miRNAs play an important role in maintaining bone remodeling and bone metabolism. This study investigated the expression level of miR-206 in the serum of osteoporosis (OP) patients and explored the effect and mechanism of miR-206 on the occurrence and development of osteoporosis. Methods 120 postmenopausal women were recruited, including 63 cases with OP and 57 women without OP. The levels of miR-206 were determined by qRT-PCR technology. Spearman correlation coefficient was used to evaluate the correlation of miR-206 with bone mineral density (BMD). An ROC curve was used to evaluate the diagnostic value of miR-206 in osteoporosis. The effects of miR-206 on cell proliferation and cell apoptosis of hFOBs were measured by CCK-8 assay and flow cytometry, respectively. Luciferase reporter gene assay was used to confirm the interaction of miR-206 and the 3′UTR of HDAC4. Results Serum miR-206 had low expression level in osteoporosis patient group compared with control group. The expression level of serum miR-206 had diagnostic value for osteoporosis, and the serum miR-206 levels were positively correlated with BMD. The down-regulated miR-206 could inhibit cell proliferation and promote cell apoptosis. Luciferase analysis indicated that HDAC4 was the target gene of miR-206. Conclusions MiR-206 could be used as a new potential diagnostic biomarker for osteoporosis, and in in vitro cell experiments, miR-206 may regulate osteoblast cell proliferation and apoptosis by targeting HDAC4.

scholarly journals Ganoderic Acid A Protects Rat H9c2 Cardiomyocytes from Hypoxia-Induced Injury via Up-Regulating miR-182-5p

2018 ◽  
Vol 50 (6) ◽  
pp. 2086-2096 ◽  
Author(s):  
Xiaohong  Zhang ◽  
Can Xiao ◽  
Hong Liu
Keyword(s):  
Cell Viability ◽  
Molecular Mechanisms ◽  
Akt Pathway ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Ganoderic Acid ◽  
Expression Levels ◽  
Protein Levels ◽  
Viability Loss ◽  
H9c2 Cardiomyocytes

Background/Aims: Ganoderic acid A (GAA) isolated from Ganoderma lucidum, shows various benefit activities, such as anti-tumor activity, anti-HIV activity and hepatoprotective activity. However, the potential effects of GAA on hypoxia-induced injury of cardiomyocytes are still unclear. In this study, we aimed to reveal the effects of GAA on hypoxic-induced H9c2 cell injury, as well as potential underlying molecular mechanisms. Methods: Rat H9c2 cardiomyocytes were cultured in hypoxia condition with different doses of GAA. Cell viability and apoptosis were detected by CCK-8 assay and flow cytometry, respectively. qRT-PCR was performed to assess the expression levels of microRNA-182-5p (miR-182-5p) and phosphatase and tensin homologue (PTEN). Cell transfection was conducted to change the expression levels of miR-182-5p and PTEN in H9c2 cells. Finally, protein levels of key factors involved in cell proliferation, cell apoptosis and PTEN/PI3K/AKT pathway were evaluated using western blotting. Results: Hypoxia treatment significantly induced H9c2 cell viability loss and apoptosis. GAA incubation remarkably protected H9c2 cells from hypoxia-induced viability loss, proliferation inhibition and apoptosis. In addition, GAA obviously enhanced the expression level of miR-182-5p in H9c2 cells. Suppression of miR-182-5p notably alleviated the protective effects of GAA on hypoxia-treated H9c2 cells. Furthermore, miR-182-5p negatively regulated the mRNA and protein levels of PTEN in H9c2 cells. GAA attenuated hypoxia-induced inactivation of PI3K/AKT pathway in H9c2 cells by up-regulating miR-182-5p and then down-regulating PTEN. Conclusion: GAA protected rat H9c2 cardiomyocytes from hypoxia-induced injury might via up-regulating miR-182-5p, down-regulating PTEN and then activating PI3K/AKT signaling pathway.

scholarly journals Mir-204 Regulates LPS-Induced A549 Cell Damage by Targeting FOXK2

2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Shufen Li ◽  
Lifen Zhao ◽  
Xujiong Li ◽  
Gaiping Shang ◽  
Lijing Gao ◽  
...  
Keyword(s):  
A549 Cell ◽  
Cell Apoptosis ◽  
Cell Injury ◽  
Cell Damage ◽  
A549 Cells ◽  
Inflammatory Factors ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Downstream Target ◽  
Apoptosis Related Protein

Objective. To assess whether miR-204 and HA affect A549 cell injury induced by lipopolysaccharide. Material and Methods. A549 cells were treated with hirsutanol A, and cell damage was induced by LPS followed by analysis of cell proliferation by CCK-8, cell apoptosis by flow cytometry, apoptosis-related protein expression by western blot, downstream target of miR-20 by dual-luciferase reporter gene, and inflammatory factors by ELISA and PCR. Results. LPS can significantly inhibit the viability of A549 cells, induce cell apoptosis, and promote the release of IL-6, IL-1β, and TNF-α, while HA pretreatment can target FOXK2 by upregulating miR-204 levels, thereby alleviating apoptosis and promoting cell viability and at the same time inhibiting the release of inflammatory factors by inhibiting the activation of NF-κB. Conclusions. miR-204 participates in the protection of HA acute lung injury by targeting FOXK2.

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.

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