scholarly journals MicroRNA-142-3p Induces Atherosclerosis-Associated Endothelial Cell Apoptosis by Directly Targeting Rictor

2018 ◽  
Vol 47 (4) ◽  
pp. 1589-1603 ◽  
Author(s):  
Bing Qin ◽  
Yaqing Shu ◽  
Ling Long ◽  
Haiyan Li ◽  
Xuejiao Men ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Apoptotic Cell ◽  
Luciferase Reporter ◽  
In Vivo Model ◽  
Bioinformatics Analyses ◽  
Endothelial Apoptosis

Background/Aims: Atherosclerosis, a multifactorial chronic disease, is the main cause of death and impairment in the world. Endothelial cells (ECs) apoptosis plays a crucial role in the onset and development of atherosclerosis, whereas the underlying molecular mechanisms are unclear. MicroRNA-142-3p (miR-142-3p) is a well-defined tumor suppressor in several types of cancer, while the role of miR-142-3p in ECs apoptosis and the development of atherosclerosis has yet to be elucidated. Therefore, the present study aimed to investigate the role of miR-142-3p in ECs apoptosis during atherosclerosis and the underlying mechanism. Methods: Human aortic endothelial cells (HAECs) were treated with oxidized low-density lipoprotein (ox-LDL). The expression level of miR-142-3p was detected using qRT-PCR. Apoptosis was determined via flow cytometry and Caspase-3 activity assay. Prediction of the binding between miR-142-3p and 3’-UTR of Rictor mRNA was performed by bioinformatics analyses and confirmed by a dual luciferase reporter assay. The effects of miR-142-3p on endothelial apoptosis and atherosclerosis were further analyzed in an in vivo model using ApoE-/- mice fed with high-fat diet (HFD). Results: MiR-142-3p expression was substantially up-regulated during the ox-LDL-elicited apoptosis in HAECs. Forced expression of miR-142-3p exacerbated apoptosis in ECs whereas inhibition of miR-142-3p could partly alleviate apoptotic cell death mediated by ox-LDL. Further analysis identified Rictor as a direct target of miR-142-3p, and Rictor knockdown abolished the anti-apoptotic effect of miR-142-3p inhibitor. Moreover, the Akt/endothelial nitric oxide synthase (eNOS) signaling pathway was found to mediate the beneficial effect of miR-142-3p inhibitor on endothelial apoptosis. Finally, systemic treatment with miR-142-3p antagomir attenuated endothelial apoptosis and retarded the progression of atherosclerosis in the aorta of ApoE-/- mice. Conclusions: Down-regulation of miR-142-3p inhibited ECs apoptosis and atherosclerotic development by up-regulating the expression of Rictor and activating the Akt/eNOS signaling pathway. This indicates that miR-142-3p may be a potential target for the prevention and treatment of atherosclerosis.

scholarly journals MAZ-LINC00645-GP73 Axis Promotes Hepatocellular Carcinoma Proliferation and Metastasis

2020 ◽  
Author(s):  
Yang Chen ◽  
Huiyan Li ◽  
Chunxun Liu ◽  
Yongmei Han ◽  
Yubao Zhang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Molecular Mechanisms ◽  
Tumor Promoter ◽  
Transcriptional Analysis ◽  
Luciferase Reporter ◽  
Chip Assay ◽  
Bioinformatics Analyses

Abstract BACKGROUND: Long non-coding RNAs (lncRNA) have been shown to play important roles in the development and progression of hepatocellular carcinoma (HCC). In this report, we examined the role of lncRNA LINC00645 in HCC. MATERIAL AND METHODS: Based on public databases and integrating bioinformatics analyses, the over-expression of LINC00645 in HCC tissues was detected and further validated in a cohort of liver tissues. A series of in vitro and in vivo functional experiments were executed to investigate the role of LINC00645 in the carcinogenesis and development of HCC. Comprehensive transcriptional analysis, chromatin immunoprecipitation (ChIP) assay, dual-luciferase reporter assay and western blot etc. were performed to explore the molecular mechanisms underlying the functions of LINC00645. RESULTS: LINC00645 was significantly upregulated in HCC cell lines and HCC tissues, which was correlated with poor prognosis in HCC patients. LINC00645 knockdown remarkably suppressed tumor growth in vitro and in vivo. Mechanistically, LINC00645 could competitively bind with miR-141-3p to prevent the degradation of its target gene GP73, which acts as a tumor-promoter in HCC. Furthermore, the ChIP assay showed that the transcription factor MAZ could bind to the LINC00645 promoter and increase its transcription. CONCLUSIONS: Collectively, this study demonstrated that LINC00645 plays a critical regulatory role in hepatocellular carcinoma cells and LINC00645 may serve as a potential diagnostic biomarker and therapeutic target of HCC. Thus, targeting MAZ/LINC00645/miR-141-3p/GP73 signaling axis may prevent the progression of HCC.

scholarly journals Has-miR-875-5p Inhibits Tumorigenesis by Targeting USF2 via TGF-β Signaling Pathway in Gastric Cancer

2021 ◽  
Author(s):  
Shenshuo Gao ◽  
Zhikai Zhang ◽  
Xubin Wang ◽  
Yan Ma ◽  
Chensheng Li ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Signaling Pathway ◽  
Research Direction ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Gastric Cancer Cells ◽  
New Research

Abstract Background: Gastric cancer (GC) is one of the most common malignancies, and more and more evdiences show that the pathogenesis is regulated by various miRNAs.In this study, we investigated the role of miR-875 in GC. Methods:The expression of miR-875-5p was detected in human GC specimens and cell lines by miRNA RT-PCR. The effect of miR-875-5p on GC proliferation was determined by CCK-8 proliferation assay and EDU assay. Migration and invasion were examined by transwell migration and invasion assay and wound healing assay. The interaction between miR-875-5p and its target gene USF2 was verified by a dual luciferase reporter assay. The effects of miR-875-5p in vivo were studied in xenograft nude mice models.Related proteins were detected by Western blot.Results:The results showed that miR-875-5p inhibited the proliferation, migration and invasion of gastric cancer cells in vitro, and inhibited tumorigenesis in vivo. USF2 proved to be a direct target of miR-875-5p. Knockdown of USF2 partially counteracts the effects of miR-875-5p inhibitors.Overexpression of miR-875-5p can inhibit proliferation, migration, and invasion through the TGF-β signaling pathway by down-regulation of USF2 in GC, providing a new research direction for the diagnosis and targeted therapy of GC.Conclusions: MiR-875-5pcan inhibited the progression of GC by directly targeting USF2 and negatively regulating TGF-β signaling pathway.In the future, miR-875-5p is expected to be used as a potential therapeutic target for GC therapy.

Matrix metalloproteinase-9 : surrogate marker and therapeutic target against neurovascular impairment after cerebral ischemia

2015 ◽  
Author(s):  
◽  
Shanyan Chen
Keyword(s):  
Endothelial Cells ◽  
Ischemic Stroke ◽  
Cerebral Ischemia ◽  
Molecular Mechanisms ◽  
Combined Treatment ◽  
Cell Penetrating Peptides ◽  
Therapeutic Window ◽  
Gelatinase Activity

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] Stroke ranks fourth among all causes of death, and acute ischemic stroke is the most common form. The neurovascular unit (NVU) describes a basic functional structure in the brain and is primarily composed of endothelial cells, pericytes, astrocytes, microglia and neurons. The dynamic structure of the NVU is highly regulated due to interactions between different cells and extracellular matrix (ECM) components. Proteolysis of the ECM by matrix metalloproteinases (MMPs), especially MMP-9, plays an important role in the pathophysiology of cerebral ischemia and administration of tissue plasminogen activator (tPA). The activation of gelatinases (MMP-2/9) is considered a key mechanism involved in the impairment of NVU. The overall goal of this research project is to examine the role of MMP-9 in the neurovascular impairment after ischemic stroke in mice. In this project, we implemented a new strategy using gelatinase-activatable cell-penetrating peptides (ACPPs) tagged with fluorescence and/or gadolinium-based contrast agents to investigate proteolysis of gelatinases as surrogate markers of neurovascular integrity. We presented evidence that the combination of a sensitive fluorescent chromatophore and MRI contrast enhancement agent can be used to monitor gelatinase activity and its distribution in cultured neurons as well as in mice after focal cerebral ischemia. Detection of the activity of gelatinases in vivo using ACPPs could provide insights into the underlying mechanism for gelatinase proteolysis that mediate ischemia-related neurovascular impairment. We also applied a two-dimensional (2D) gelatin zymography technique that combines isoelectric focusing (IEF) with zymographic electrophoresis. We demonstrated that the 2D zymography approach can improve separation of different isoforms of gelatinases in both in vitro and in vivo conditions. 2D zymography is an effective method to separate posttranslational modification isoforms of gelatinases and to identify modifications that regulate their enzymatic activity in acute brain injuries. In work that follows, we used a fibrin-rich blood clot to occlude the middle cerebral artery (MCA) in mice as a model to represent the critical thromboembolic features of ischemic stroke in humans. In this study, we evaluated effects of SB-3CT, a mechanism-based inhibitor selective for gelatinases. We demonstrated MMP-9 activation and neurovasculature impairment in this stroke model, and showed the ability of SB-3CT to inhibit MMP-9 activity in vivo, which in turn resulted in maintenance of laminin, antagonism of pericyte contraction and loss, preservation of laminin-positive pericytes and endothelial cells, and thus rescuing neurons from apoptosis and preventing intracerebral hemorrhage. We further demonstrated that SB-3CT/tPA combined treatment could attenuate MMP-9 -- mediated degradation of endothelial laminin, impairment of endothelial cells, and decrease of caveolae -- mediated transcytosis. Early inhibition of MMP-9 proteolysis by SB-3CT decreased brain damage, reduced BBB disruption, and prevented hemorrhagic transformation after delayed tPA treatment. Therefore usage of SB-3CT will be helpful in accessing combination therapy with tPA in ischemic stroke. Results from these studies indicate the important role of MMP-9 in cerebral ischemia and thus the need for further studies to explore the molecular mechanisms underlying its activation and regulation. Results further demonstrated that the combined use of MMP-9 inhibitor with tPA may extend tPA therapeutic window for mitigating stroke damage.

scholarly journals DEPTOR Deficiency-Mediated mTORc1 Hyperactivation in Vascular Endothelial Cells Promotes Angiogenesis

2018 ◽  
Vol 46 (2) ◽  
pp. 520-531 ◽  
Author(s):  
Yan Ding ◽  
Lanlan Shan ◽  
Wenqing Nai ◽  
Xiaojun Lin ◽  
Ling Zhou ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Gene Knockout ◽  
Umbilical Vein ◽  
Vascular Endothelial Cell ◽  
Tube Formation ◽  
Vascular Endothelial

Background/Aims: The mechanistic target of rapamycin (mTOR) signaling pathway is essential for angiogenesis and embryonic development. DEP domain-containing mTOR-interacting protein (DEPTOR) is an mTOR binding protein that functions to inhibit the mTOR pathway In vitro experiments suggest that DEPTOR is crucial for vascular endothelial cell (EC) activation and angiogenic responses. However, knowledge of the effects of DEPTOR on angiogenesis in vivo is limited. This study aimed to determine the role of DEPTOR in tissue angiogenesis and to elucidate the molecular mechanisms. Methods: Cre/loxP conditional gene knockout strategy was used to delete the Deptor gene in mouse vascular ECs. The expression or distribution of cluster of differentiation 31 (CD31), vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1 alpha (HIF-1α) were detected by immunohistochemical staining or western blot. Tube formation assay was used to measure angiogenesis in vitro. Results: Deptor knockdown led to increased expression of CD31, VEGF and HIF-1α in heart, liver, kidney and aorta. After treatment with rapamycin, their expression was significantly down regulated. In vitro, human umbilical vein endothelial cells (HUVECs) were transfected with DEPTOR-specific small interfering RNA (siRNA), which resulted in a significant increase in endothelial tube formation and migration rates. In contrast, DEPTOR overexpression markedly reduced the expression of CD31, VEGF and HIF-1α. Conclusions: Our findings demonstrated that deletion of the Deptor gene in vascular ECs resulted in upregulated expression of CD31 and HIF-1α, and further stimulated the expression of VEGF which promoted angiogenesis, indicating that disruption of normal angiogenic pathways may occur through hyperactivation of the mTORC1/HIF-1α/VEGF signaling pathway.

Platelet-Derived Microparticles Induce Angiogenesis and Stimulate Post-Ischemic Revascularization.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3916-3916
Author(s):  
Olga Dashevsky ◽  
Alexander Brill ◽  
Julia Rivo ◽  
David Varon
Keyword(s):  
Endothelial Cells ◽  
Aortic Ring ◽  
In Vivo Model ◽  
Preliminary Evaluation ◽  
Vegf Receptor ◽  
In Vivo Models ◽  
Ischemic Region

Abstract Platelet attachment to the subcellular matrix at injured sites of the vasculature is followed by their activation and release of microparticles. Platelet-derived microparticles (PMP) have been shown to be involved in the regulation of hemostasis. However, little is known about the role of PMP in the regulation of angiogenesis and related clinical conditions. We have recently demonstrated that platelets as a cellular system induce angiogenic responses both in vitro and in vivo. In the present study, we investigated the potential role of PMP in angiogenesis. A strong dose-dependent pro-angiogenic effect of PMP in the rat aortic ring model (5.3±2.1 mm2 surface covered with sprouting vessels versus 0.24±0.2 mm2 in the control, p<0.001) was observed. This effect was reversed by selective inhibition of VEGF, bFGF and PDGF (surface covered with vessels 0.7±0.5 mm2, 1.7±1.5 mm2, and 2.4±1.2 mm2, respectively, p<0.02 versus control), but not by inhibition of heparanase (5.1±0.8 mm2, p>0.5 versus control). PMP exert their stimulatory effect via PI3-kinase, Src kinase and ERK, whereas protein kinase C seems not to be involved, as judged by the aortic ring sprouting model. Using confocal and electron microscopy, we also demonstrate that PMP bind to non-activated endothelial cells. In addition, PMP markedly increased invasion of human endothelial cells through a layer of matrigel. This effect was abolished by an inhibitor of VEGF receptor tyrosine phosphorylation or laminaran sulfate (heparanase inhibitor). It was also partially reduced by PDGF blocking mAb, whereas blocking of bFGF had no effect. Furthermore, we have demonstrated that PMP induce angiogenesis in an in vivo model, in which beads (30 μl) of 4% agarose gel containing the substances under study were transplanted subcutaneously into mice. Image analysis of the capillary area revealed the following: control beads − 0.2±0.05 mm2, VEGF + bFGF containing beads − 4.8±1.1 mm2, PMP (100 μg/ml) containing beads − 5.1±1.3 mm2, p<0.001 versus control. The latter finding was further supported by immunohistochemical staining of the skin in the vicinity of the beads for von Willebrand factor, a marker of endothelial cells (control − 4.0±3.2, VEGF+bFGF − 12±4.4, PMP − 17±6.5 capillaries per view field, p<0.05 versus control). Finally, we explored the potential effect of PMP in a rat myocardial infarction model. Ischemia was induced by LAD ligation followed by injection of either PMP or PBS into the ischemic region. Preliminary evaluation of the LAD myocardial territory in sham-operated animals revealed 157±42.0 capillaries per view field. In contrast, number of capillaries observed 3 weeks after induction of ischemia was reduced to 34±21.5. When PMP were injected into the ischemic region, there was an increase in capillary number up to 97±27.3. In conclusion, PMP induce angiogenesis in both in vitro and in vivo models. Local injection of PMP into the ischemic myocardium may improve revascularization.

scholarly journals Exosomal miR-1290 promotes angiogenesis in hepatocellular carcinoma via targeting SMEK1

2020 ◽  
Author(s):  
Qiong Wang ◽  
Guanwen Wang ◽  
Lianjie Niu ◽  
Shaorong Zhao ◽  
Jianjun Li ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Endothelial Cells ◽  
Tumor Angiogenesis ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Primary Liver Cancer ◽  
Tumor Model ◽  
Tube Formation ◽  
Luciferase Reporter

Abstract Background: Hepatocellular carcinoma (HCC), the most common primary liver cancer, rely on the formation of new blood vessel for growth and frequent intrahepatic and extrahepatic metastasis. Therefore, it is important to explore the underlying molecular mechanisms of tumor angiogenesis of HCC. Recently, microRNAs have been shown to modulate angiogenic processes by modulating the expression of critical angiogenic factors. However, the potential roles of tumor-derived exosomal microRNAs in regulating tumor angiogenesis remain to be elucidated. Methods: MiRNome sequencing was performed to uncover the miRNAs that are dysregulated in HCC patient serum-derived exosomes. Expression levels of miR-1290 in tissues and cells were determined by quantitative real-time PCR. The effect of mir-1290 on proliferation was evaluated by CCK-8 assay. The angiogenic ability of cells were determined by transwell, wound-healing, tube formation and matrigel plug assays. SMMC-7721 xenograft tumor model was established in NOD-SCID nude mice using miR-1290 and NC antagomirs to determin the angiogenic effect of mir-1290 in vivo. Target protein expression was determined by western blotting. Dual luciferase reporter assay was performed to confirm the action of miR-1290 on downstream target genes including SMEK1. Results are reported as means ± S.D. and differences were tested for significance using 2-sided Student’s t-test.Results: In this study, our miRNome sequencing demonstrated that miR-1290 was overexpressed in HCC patient serum-derived exosomes, and we found that delivery of miR-1290 into human endothelial cells enhanced their angiogenic ability. Our results further revealed that SMEK1 is a direct target of miR-1290 in endothelial cells. MiR-1290 exerted its pro-angiogenic function, at least in part, by alleviating the inhibition of VEGFR2 phosphorylation done by SMEK1. Conclusions: Collectively, our findings provide evidence that miR-1290 is overexpressed in HCC and promotes tumor angiogenesis via exosomal secretion, implicating its potential role as a therapeutic target for HCC.

scholarly journals LINC00467 Promotes Tumor Progression via Regulation of the NF-kb Signal Axis in Bladder Cancer

2021 ◽  
Vol 11 ◽  
Author(s):  
Jiawei Xiao ◽  
Lian Gong ◽  
Mengqing Xiao ◽  
Dong He ◽  
Liang Xiang ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Animal Experiments ◽  
Bioinformatic Analysis ◽  
New Strategy ◽  
Patient Prognosis

PurposeLong non-coding RNAs (lncRNAs) play an important role in the occurrence and development of bladder cancer, but the underlying molecular mechanisms remain largely unknown. In this study, we found that LINC00467 was significantly highly expressed in bladder cancer through bioinformatic analysis. The present study aimed to explore the role of LINC00467 in bladder cancer and its possible underlying molecular mechanisms.MethodsThe expression of LINC00467 was obtained from GEO (GSE31189), the TCGA database, and qRT-PCR. The role of LINC00467 in bladder cancer was assessed both in vitro and in vivo. RIP, RNA pulldown, and CO-IP were used to demonstrate the potential mechanism by which LINC00467 regulates the progression of bladder cancer.ResultsThrough the analysis of GEO (GSE133624) and the TCGA database, it was found that LINC00467 was highly expressed in bladder cancer tissues and that the expression of LINC00467 was significantly negatively correlated with patient prognosis. Cell and animal experiments suggest that LINC00467 promotes the proliferation and invasion of bladder cancer cells. On the one hand, LINC00467 can directly bind to NF-kb-p65 mRNA to stabilize its expression. On the other hand, LINC00467 can directly bind to NF-kb-p65 to promote its translocation into the nucleus to activate the NF-κB signaling pathway, which promotes the progression of bladder cancer.ConclusionsLINC00467 is highly expressed in bladder cancer and can promote the progression of bladder cancer by regulating the NF-κB signaling pathway. Therefore, targeting LINC00467 is very likely to provide a new strategy for the treatment of bladder cancer and for improving patient prognosis.

scholarly journals EPC-Derived Exosomal miR-1246 and miR-1290 Regulate Phenotypic Changes of Fibroblasts to Endothelial Cells to Exert Protective Effects on Myocardial Infarction by Targeting ELF5 and SP1

2021 ◽  
Vol 9 ◽  
Author(s):  
Yulang Huang ◽  
Lifang Chen ◽  
Zongming Feng ◽  
Weixin Chen ◽  
Shaodi Yan ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Endothelial Cells ◽  
Molecular Mechanisms ◽  
Cardiac Injury ◽  
Immunofluorescence Staining ◽  
Luciferase Reporter ◽  
Protective Effects ◽  
Expression Levels ◽  
Phenotypic Changes

Myocardial infarction (MI) remains a leading cause of morbidity and mortality worldwide. Endothelial progenitor cell (EPC)-derived exosomes have been found to be effective in alleviating MI, while the detailed mechanisms remain unclear. The present study aimed to determine the protective effects of EPC-derived exosomal miR-1246 and miR-1290 on MI-induced injury and to explore the underlying molecular mechanisms. The exosomes were extracted from EPCs; gene expression levels were determined by quantitative real-time PCR, and protein expression levels were determined by western blot and immunofluorescence staining, respectively. The angiogenesis and proliferation of human cardiac fibroblasts (HCFs) were determined by tube formation assay and immunofluorescence staining of PKH67, respectively. Luciferase reporter, CHIP, and EMSA assays determined the interaction between miR-1246/1290 and the targeted genes (EFL5 and SP1). The protective effects of miR-1246/1290 on MI were evaluated in a rat model of MI. EPC-derived exosomes significantly upregulated miR-1246 and miR-1290 expression and promoted phenotypic changes of fibroblasts to endothelial cells, angiogenesis, and proliferation in HCFs. Exosomes from EPCs with miR-1246 or miR-1290 mimics transfection promoted phenotypic changes of fibroblasts to endothelial cells and angiogenesis in HCFs, while exosomes from EPCs with miR-1246 or miR-1290 knockdown showed opposite effects in HCFs. Mechanistically, miR-1246 and miR-1290 from EPC-derived exosomes induced upregulation of ELF5 and SP1, respectively, by targeting the promoter regions of corresponding genes. Overexpression of both ELF5 and SP1 enhanced phenotypic changes of fibroblasts to endothelial cells and angiogenesis in HCFs pretreated with exosomes from EPCs with miR-1246 or miR-1290 mimics transfection, while knockdown of both EFL5 and SP1 exerted the opposite effects in HCFs. Both ELF5 and SP1 can bind to the promoter of CD31, leading to the upregulation of CD31 in HCFs. Furthermore, in vivo animal studies showed that exosomes from EPCs with miR-1246 or miR-1290 overexpression attenuated the MI-induced cardiac injury in the rats and caused an increase in ELF5, SP1, and CD31 expression, respectively, but suppressed α-SMA expression in the cardiac tissues. In conclusion, our study revealed that miR-1246 and miR-1290 in EPC-derived exosomes enhanced in vitro and in vivo angiogenesis in MI, and these improvements may be associated with amelioration of cardiac injury and cardiac fibrosis after MI.

scholarly journals Exosomal miR-1290 promotes angiogenesis in hepatocellular carcinoma via targeting SMEK1

2019 ◽  
Author(s):  
Qiong Wang ◽  
Guanwen Wang ◽  
Lianjie Niu ◽  
Shaorong Zhao ◽  
Jianjun Li ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Endothelial Cells ◽  
Tumor Angiogenesis ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Primary Liver Cancer ◽  
Tumor Model ◽  
Luciferase Reporter ◽  
Dependent Manner

Abstract Abstract Background: Hepatocellular carcinoma (HCC), the most common primary liver cancer, rely on the formation of new blood vessel for growth and frequent intrahepatic and extrahepatic metastasis. Therefore, it is important to explore the underlying molecular mechanisms of tumor angiogenesis of HCC. Recently, microRNAs have been shown to modulate angiogenic processes by modulating the expression of critical angiogenic factors. However, the potential roles of tumor-derived exosomal microRNAs in regulating tumor angiogenesis remain to be elucidated. Methods: MiRNome sequencing was performed to uncover the miRNAs that are dysregulated in HCC patient serum-derived exosomes. Expression levels of miR-1290 in tissues and cells were determined by quantitative real-time PCR. The effect of mir-1290 on proliferation was evaluated by CCK-8 assay. The angiogenic ability of cells were determined by transwell, wound-healing, tube formation and matrigel plug assays. SMMC-7721 xenograft tumor model was established in NOD-SCID nude mice using miR-1290 and NC antagomirs to determin the angiogenic effect of mir-1290 in vivo. Target protein expression was determined by western blotting. Dual luciferase reporter assay was performed to confirm the action of miR-1290 on downstream target genes including SMEK1. Results are reported as means ± S.D. and differences were tested for significance using 2-sided Student’s t-test. Results: In this study, our miRNome sequencing demonstrated that miR-1290 was overexpressed in HCC patient serum-derived exosomes, and we found that delivery of miR-1290 into human endothelial cells enhanced their angiogenic ability. Our results further revealed that SMEK1 is a direct target of miR-1290 in endothelial cells. MiR-1290 exerted its pro-angiogenic function, at least in part, by inhibiting the VEGFR2 signaling pathway in a SMEK1-dependent manner. Conclusions: Collectively, our findings provide evidence that miR-1290 is overexpressed in HCC and promotes tumor angiogenesis via exosomal secretion, implicating its potential role as a therapeutic target for HCC.

scholarly journals Epigenetic regulation of ferroptosis via ETS1/miR-23a-3p/ACSL4 axis mediates sorafenib resistance in human hepatocellular carcinoma

2022 ◽  
Vol 41 (1) ◽  
Author(s):  
Yuanjun Lu ◽  
Yau-Tuen Chan ◽  
Hor-Yue Tan ◽  
Cheng Zhang ◽  
Wei Guo ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Drug Resistance ◽  
Molecular Mechanisms ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Bioinformatics Analyses ◽  
Sorafenib Treatment ◽  
Hcc Cells

Abstract Background Drug resistance to sorafenib greatly limited the benefits of treatment in patients with hepatocellular carcinoma (HCC). MicroRNAs (miRNAs) participate in the development of drug resistance. The key miRNA regulators related to the clinical outcome of sorafenib treatment and their molecular mechanisms remain to be identified. Methods The clinical significance of miRNA-related epigenetic changes in sorafenib-resistant HCC was evaluated by analyzing publicly available databases and in-house human HCC tissues. The biological functions of miR-23a-3p were investigated both in vitro and in vivo. Proteomics and bioinformatics analyses were conducted to identify the mechanisms that regulating miR-23a-3p. Luciferase reporter assay and chromatin immunoprecipitation (ChIP) assay were used to validate the binding relationship of miR-23a-3p and its targets. Results We found that miR-23a-3p was the most prominent miRNA in HCC, which was overexpressed in sorafenib non-responders and indicated poor survival and HCC relapse. Sorafenib-resistant cells exhibited increased miR-23a-3p transcription in an ETS Proto-Oncogene 1 (ETS1)-dependent manner. CRISPR-Cas9 knockout of miR-23a-3p improved sorafenib response in HCC cells as well as orthotopic HCC tumours. Proteomics analysis suggested that sorafenib-induced ferroptosis was the key pathway suppressed by miR-23a-3p with reduced cellular iron accumulation and lipid peroxidation. MiR-23a-3p directly targeted the 3′-untranslated regions (UTR) of ACSL4, the key positive regulator of ferroptosis. The miR-23a-3p inhibitor rescued ACSL4 expression and induced ferrotoptic cell death in sorafenib-treated HCC cells. The co-delivery of ACSL4 siRNA and miR-23a-3p inhibitor abolished sorafenib response. Conclusion Our study demonstrates that ETS1/miR-23a-3p/ACSL4 axis contributes to sorafenib resistance in HCC through regulating ferroptosis. Our findings suggest that miR-23a-3p could be a potential target to improve sorafenib responsiveness in HCC patients.

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