miRNA-103 promotes chondrocyte apoptosis by down-regulation of Sphingosine kinase-1 and ameliorates PI3K/AKT pathway in osteoarthritis

Abstract Objectives: The aim of the present study was to determine the effects of miRNA-103 on chondrocyte apoptosis and molecular mechanisms in osteoarthritis (OA) progression. Methods: The cell proliferation, apoptosis, and recovery ability were measured by cell counting kit-8 (CCK-8), flow cytometry, and wound healing assays. The interaction of miRNA-103 and Sphingosine kinase-1 (SPHK1) were determined by using luciferase reporter assay. The expression of mRNA and proteins were measured by qRT-PCR and Western blot. OA rat model was established by surgery stimulation. Results: miRNA-103 expression was significantly increased in the cartilage of OA patients and surgery-induced OA rat models. miRNA-103 transfection into primary rat chondrocytes reduced SPHK1 expression, induced apoptosis, inhibited cell proliferation, and impeded scratch assay wound closure. Moreover, expression of total AKT, and p-AKT were significantly reduced in miRNA-103-overexpressing chondrocytes while SPHK1 up-regulation increased the expression of phosphatidylinsitol-3-kinase (PI3K) and p-AKT, and reversed the proliferation suppression induced by the miRNA-103 mimic. Conclusions: Our studies suggest that miRNA-103 contributes to chondrocyte apoptosis, promoting OA progression by down-regulation of PI3K/AKT pathway through the reduction in SPHK1 activity.

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MiR-489-3p inhibits cell proliferation, migration, and invasion, and induces apoptosis, by targeting the BDNF-mediated PI3K/AKT pathway in glioblastoma

Open Life Sciences ◽

10.1515/biol-2020-0024 ◽

2020 ◽

Vol 15 (1) ◽

pp. 274-283

Author(s):

Bo Zheng ◽

Tao Chen

Keyword(s):

Cell Proliferation ◽

Luciferase Reporter ◽

Akt Pathway ◽

Migration And Invasion ◽

Bdnf Mrna ◽

Protein Levels ◽

Rna Immunoprecipitation ◽

Underlying Mechanisms ◽

Induced Apoptosis ◽

Protein Cell

AbstractAmong astrocyte tumors, glioblastoma (GBM) is the most malignant glioma, highly aggressive and invasive, with extremely poor prognosis. Previous research has reported that microRNAs (miRNAs) participate in the progression of many cancers. Thus, this study aimed to explore the role and the underlying mechanisms of microRNA (miR)-489-3p in GBM progression. The expression of miR-489-3p and brain-derived neurotrophic factor (BDNF) mRNA was measured by quantitative real-time polymerase chain reaction. Western blot analysis was used to detect BDNF protein and the PI3K/AKT pathway-related protein. Cell proliferation, apoptosis, migration, and invasion were analyzed using CKK-8 assay, flow cytometry, and transwell assay, respectively. The interaction between BDNF and miR-489-3p was explored by luciferase reporter assay and RNA immunoprecipitation (RIP) assay. MiR-489-3p was down-regulated and BDNF was up-regulated in GBM tissues and cells. MiR-489-3p re-expression or BDNF knockdown inhibited GBM cell proliferation, migration, and invasion, and promoted apoptosis. BDNF was a target of miR-489-3p, and BDNF up-regulation reversed the effects of miR-489-3p on GBM cells. The protein levels of p-AKT and p-PI3K were notably reduced in GBM cells by overexpression of miR-489-3p, but were rescued following BDNF up-regulation. Therefore, miR-489-3p inhibited proliferation, migration, and invasion, and induced apoptosis, by targeting the BDNF-mediated PI3K/AKT pathway in GBM, providing new strategies for clinical treatment of GBM.

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MiR-4310 Induced by SP1 targets PTEN to Promote Glioma Progression

10.21203/rs.3.rs-40999/v1 ◽

2020 ◽

Author(s):

Wu Zhiyong ◽

Luo Jie ◽

Huang Tengyue ◽

Yi Renhui ◽

Ding Shengfeng ◽

...

Keyword(s):

Cell Proliferation ◽

Molecular Mechanisms ◽

Luciferase Reporter ◽

Akt Pathway ◽

Migration And Invasion ◽

Boyden Chamber ◽

Luciferase Reporter Gene ◽

Mobility Shift ◽

Glioma Progression

Abstract Background: miRNAs have been reported to be involved in multiple biological processes of gliomas. Here, we aimed to analyze miR-4310 and its correlation genes involved in the tumor progression of human glioma.Methods: miR-4310 expression levels were examined in glioma and non-tumor brain (NB) tissues. The molecular mechanisms of miR-4310 expression and its effects on cell proliferation, migration, and invasion were explored by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) , Transwell chamber, Boyden chamber, and western blot analyses, as well as in vivo tumorigenesis in nude mice. The relationships among miR-4310, SP1, and phosphatase and tensin homolog (PTEN) were explored by chromatin immunoprecipitation (ChIP), agarose gel electrophoresis, electrophoresis mobility shift (EMSA), and dual luciferase reporter gene assays. Results: miR-4310 expression was upregulated in glioma tissues compared to NB. Overexpressed miR-4310 promoted glioma cell proliferation, migration, and invasion in vitro and tumorigenesis in vivo . Inhibition of miR-4310 was sufficient to reverse these results. Mechanistic analyses revealed that miR-4310 promoted glioma progression through the PI3K/AKT pathway by targeting PTEN. Additionally, SP1 induced the expression of miR-4310 by binding to its promoter region. Conclusion: miR-4310 promotes the progression of glioma by targeting PTEN and activating the PI3K/AKT pathway meanwhile the expression of miR-4310 is induced by SP1.

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LncRNA KCNQ1OT1 promotes the development of diabetic nephropathy by regulating miR-93-5p/ROCK2 axis

Diabetology & Metabolic Syndrome ◽

10.1186/s13098-021-00726-4 ◽

2021 ◽

Vol 13 (1) ◽

Author(s):

Li Zhao ◽

Huaqian Chen ◽

Lin Wu ◽

Zhengdong Li ◽

Ren Zhang ◽

...

Keyword(s):

Cell Proliferation ◽

Diabetic Nephropathy ◽

Mesangial Cells ◽

Coiled Coil ◽

Cell Counting ◽

Luciferase Reporter ◽

Glomerular Mesangial Cells ◽

Cell Models ◽

Protein Levels ◽

Induced Apoptosis

Abstract Background Long non-coding RNAs (lncRNAs) have been reported to play vital roles in diabetic nephropathy (DN). The aim of this study was to explore the function of mechanism of lncRNA KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1) in DN. Methods DN cell models were established using high glucose (HG) treatment in human glomerular mesangial cells (HGMC) and human renal glomerular endothelial cells (HRGEC). The expression levels of KCNQ1OT1, microRNA-93-5p (miR-93-5p), and Rho associated coiled-coil containing protein kinase 2 (ROCK2) mRNA was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Cell Counting Kit-8 (CCK-8) assay and flow cytometry were used to detect cell proliferation and apoptosis, respectively. ROCK2 and apoptosis/fibrosis-related protein levels were examined by western blot. The predicted interaction between miR-93-5p and KCNQ1OT1 or ROCK2 was verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. Results KCNQ1OT1 was upregulated in DN patients and DN cell models. KCNQ1OT1 knockdown inhibited cell proliferation and fibrosis and induced apoptosis in DN cell models. MiR-93-5p was a direct target of KCNQ1OT1, and miR-93-5p inhibition restored the KCNQ1OT1 knockdown-mediated effects on cell proliferation, fibrosis and apoptosis in DN cell models. In addition, ROCK2 was identified as a target of miR-93-5p, and miR-93-5p overexpression suppressed cell proliferation and fibrosis and accelerated apoptosis by targeting ROCK2 in DN cell models. Moreover, KCNQ1OT1 regulated ROCK2 expression by binding to miR-93-5p. Conclusion KCNQ1OT1 knockdown inhibited cell proliferation and fibrosis and induced apoptosis in DN by regulating miR-93-5p/ROCK2 axis, providing potential value for the treatment of DN.

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MiR-4310 induced by SP1 targets PTEN to promote glioma progression

10.21203/rs.3.rs-40999/v2 ◽

2020 ◽

Author(s):

Wu Zhiyong ◽

Luo Jie ◽

Huang Tengyue ◽

Yi Renhui ◽

Ding Shengfeng ◽

...

Keyword(s):

Cell Proliferation ◽

Molecular Mechanisms ◽

Luciferase Reporter ◽

Akt Pathway ◽

Migration And Invasion ◽

Boyden Chamber ◽

Luciferase Reporter Gene ◽

Mobility Shift ◽

Glioma Progression

Abstract Background: miRNAs have been reported to be involved in multiple biological processes of gliomas. Here, we aimed to analyze miR-4310 and its correlation genes involved in the tumor progression of human glioma. Methods: miR-4310 expression levels were examined in glioma and non-tumor brain (NB) tissues. The molecular mechanisms of miR-4310 expression and its effects on cell proliferation, migration, and invasion were explored by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) , Transwell chamber, Boyden chamber, and western blot analyses, as well as in vivo tumorigenesis in nude mice. The relationships among miR-4310, SP1, and phosphatase and tensin homolog (PTEN) were explored by chromatin immunoprecipitation (ChIP), agarose gel electrophoresis, electrophoresis mobility shift (EMSA), and dual luciferase reporter gene assays. Results: miR-4310 expression was upregulated in glioma tissues compared to NB. Overexpressed miR-4310 promoted glioma cell proliferation, migration, and invasion in vitro and tumorigenesis in vivo . Inhibition of miR-4310 was sufficient to reverse these results. Mechanistic analyses revealed that miR-4310 promoted glioma progression through the PI3K/AKT pathway by targeting PTEN. Additionally, SP1 induced the expression of miR-4310 by binding to its promoter region. Conclusion: miR-4310 promotes the progression of glioma by targeting PTEN and activating the PI3K/AKT pathway meanwhile the expression of miR-4310 is induced by SP1.

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circHUWE1 Exerts an Oncogenic Role in Inducing DDP-Resistant NSCLC Progression Depending on the Regulation of miR-34a-5p/TNFAIP8

International Journal of Genomics ◽

10.1155/2021/3997045 ◽

2021 ◽

Vol 2021 ◽

pp. 1-20

Author(s):

Xueliang Yang ◽

Quan Sun ◽

Yongming Song ◽

Wenli Li

Keyword(s):

Cell Cycle ◽

Molecular Mechanisms ◽

Cell Counting ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Circular Rnas ◽

Cell Cycle Distribution ◽

Necrosis Factor Alpha ◽

Induced Apoptosis

Background. Circular RNAs (circRNAs) are reported as competing endogenous RNAs (ceRNAs) and play key roles in non-small-cell lung cancer (NSCLC) progression. Thus, this study was aimed at clarifying underlying molecular mechanisms of circHUWE1 in NSCLC. Methods. The quantitative real-time polymerase chain reaction (RT-qPCR) and western blot analyses were used for examining circHUWE1, microRNA-34a-5p (miR-34a-5p), and tumor necrosis factor alpha-induced protein 8 (TNFAIP8). IC50 of cisplatin (DDP) in A549/DDP and H1299/DDP cells and cell viability were analyzed by the Cell Counting Kit 8 (CCK-8) assay. Colony forming assay was performed to assess colony forming ability. Cell apoptosis and cell cycle distribution were determined by flow cytometry. Migrated and invaded cell numbers were examined by transwell assay. The association among circHUWE1, miR-34a-5p, and TNFAIP8 was analyzed by dual-luciferase reporter and RNA immunoprecipitation assays. A xenograft experiment was applied to clarify the functional role of circHUWE1 in vivo. Results. circHUWE1 was upregulated in NSCLC tissues and cells, especially in DDP-resistant groups. circHUWE1 downregulation inhibited DDP resistance, proliferation, migration, and invasion while it induced apoptosis and cell cycle arrest of DDP-resistant NSCLC cells, which was overturned by silencing of miR-34a-5p. TNFAIP8 was a functional gene of miR-34a-5p, and the suppressive effects of miR-34a-5p overexpression on DDP-resistant NSCLC progression were dependent on the suppression of TNFAIP8. circHUWE1 inhibition also delayed tumor growth of DDP-resistant NSCLC cells. Conclusion. circHUWE1 functioned as a promoter in DDP-resistant NSCLC by interaction with miR-34a-5p-TNFAIP8 networks, providing novel insight into DDP-resistant NSCLC diagnosis and treatment.

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CircRNA RERE Promotes the Oxidative Stress-Induced Apoptosis and Autophagy of Nucleus Pulposus Cells through the miR-299-5p/Galectin-3 Axis

Journal of Healthcare Engineering ◽

10.1155/2021/2771712 ◽

2021 ◽

Vol 2021 ◽

pp. 1-12

Author(s):

Rong Wang ◽

Xingchao Zhou ◽

Guorui Luo ◽

Jin Zhang ◽

Min Yang ◽

...

Keyword(s):

Western Blot ◽

Cell Viability ◽

Nucleus Pulposus ◽

Molecular Mechanisms ◽

Cell Loss ◽

Cell Counting ◽

Luciferase Reporter ◽

Nucleus Pulposus Cells ◽

Galectin 3 ◽

Induced Apoptosis

Intervertebral disc degeneration (IDD) is widely accepted as a cause of low back pain and related degenerative musculoskeletal disorders. Nucleus pulposus (NP) cell loss is closely related to IDD progression. Thus, investigating the specifically targeted therapeutic agents against NP cell loss depends on understanding the molecular mechanisms. In this study, human NP cells were treated with hydrogen peroxide (H2O2). Cell viability was assessed by using the Cell Counting Kit-8 (CCK-8) kit. The expression of circRNA arginine-glutamic acid dipeptide repeats (hsa_circ_RERE) and miR-299-5p was analyzed by real-time quantitative PCR. Western blot analysis was used to assess the protein expression levels. The autophagy levels in NP cells were detected by using an electronic microscope, LC3B protein immunofluorescence, and western blot. The apoptosis levels of NP cells were detected by flow cytometry and western blot. Dual-luciferase reporter assay analyzed the miR-299-5p bound to circ_RERE and galectin-3. Our results revealed that H2O2 significantly inhibited the viability of NP cells, promoted apoptosis and autophagy, and upregulated galectin-3 expression. miR-299-5p was reduced in IDD and H2O2-induced NP cells. The overexpression of miR-299-5p promoted cell viability and attenuated apoptosis and autophagy under H2O2 treatment. Besides, circ_RERE was upregulated in IDD and H2O2-induced NP cells. However, knockdown of circ_RERE reversed the effects of miR-299-5p overexpression on cell viability, apoptosis, and autophagy in NP cells. We propose that circ_RERE promotes the H2O2-induced apoptosis and autophagy of NP cells through the miR-299-5p/galectin-3 axis and may provide a new target for the clinical treatment of IDD.

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miR-142-5p regulates the progression of diabetic retinopathy by targeting IGF1

International Journal of Immunopathology and Pharmacology ◽

10.1177/2058738420909041 ◽

2020 ◽

Vol 34 ◽

pp. 205873842090904 ◽

Cited By ~ 2

Author(s):

Xiuming Liu ◽

Jianchang Li ◽

Xiaofeng Li

Keyword(s):

Cell Proliferation ◽

Diabetic Retinopathy ◽

Growth Factor ◽

Signaling Pathway ◽

Molecular Mechanisms ◽

Cell Counting ◽

Luciferase Reporter ◽

Igf1r Signaling

As one of leading causes of blindness, diabetic retinopathy (DR) is a progressive microvascular complication of diabetes mellitus (DM). Despite significant efforts have been devoted to investigate DR over the years, the molecular mechanisms still remained unclear. Emerging evidences demonstrated that microRNAs (miRNAs) were tightly associated with pathophysiological development of DR. Hence, this study was aimed to illustrate the role and molecular mechanisms of miR-412-5p in progression of DR. Streptozotocin (STZ) treatment in rats and human retinal endothelial cell (HREC) models were used to simulate DR conditions in vivo and in vitro. Hematoxylin-eosin (HE) staining was used to demonstrate the morphology of retinal tissues of rats. Qualitative real-time polymerase chain reaction (qRT-PCR) detected miR-142-5p and vascular endothelial growth factor (VEGF) expression levels. Cell counting kit-8 (CCK8) assay and immunofluorescence (IF) measured the cell proliferation rates. Western blot tested the expression status of IGF1/IGF1R-mediated signaling pathway. Dual-luciferase reporter assays demonstrated the molecular mechanism of miR-142-5p. miR-142-5p level was down-regulated in retinal tissues of DR rats and high glucose (HG)-treated HRECs. Insulin-like growth factor 1 (IGF1) was identified as a direct target of miR-142-5p. The reduced miR-142-5p level enhanced HRECs proliferation via activating IGF/IGF1R-mediated signaling pathway including p-PI3K, p-ERK, p-AKT, and VEGF activation, ultimately giving rise to cell proliferation. Either miR-142-5p overexpression or IGF1 knockdown alleviated the pathological effects on retinal tissues in DR rats. Collectively, miR-142-5p participated in DR development by targeting IGF1/p-IGF1R signaling pathway and VEGF generation. This miR-142-5p/IGF1/VEGF axis provided a novel therapeutic target for DR clinical treatment.

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MiR-4310 induced by SP1 targets PTEN to promote glioma progression

Cancer Cell International ◽

10.1186/s12935-020-01650-9 ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Zhiyong Wu ◽

Jie Luo ◽

Tengyue Huang ◽

Renhui Yi ◽

Shengfeng Ding ◽

...

Keyword(s):

Cell Proliferation ◽

Molecular Mechanisms ◽

Luciferase Reporter ◽

Akt Pathway ◽

Migration And Invasion ◽

Boyden Chamber ◽

Luciferase Reporter Gene ◽

Mobility Shift ◽

Glioma Progression

Abstract Background miRNAs have been reported to be involved in multiple biological processes of gliomas. Here, we aimed to analyze miR-4310 and its correlation genes involved in the progression of human glioma. Methods miR-4310 expression levels were examined in glioma and non-tumor brain (NB) tissues. The molecular mechanisms of miR-4310 expression and its effects on cell proliferation, migration, and invasion were explored using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide, Transwell chamber, Boyden chamber, and western blot analyses, as well as its effect on tumorigenesis was explored in vivo in nude mice. The relationships between miR-4310, SP1, phosphatase, and tensin homolog (PTEN) were explored using chromatin immunoprecipitation, agarose gel electrophoresis, electrophoresis mobility shift, and dual-luciferase reporter gene assays. Results miR-4310 expression was upregulated in glioma tissues compared to that in NB tissues. Overexpressed miR-4310 promoted glioma cell proliferation, migration, and invasion in vitro, as well as tumorigenesis in vivo. The inhibition of miR-4310 expression was sufficient to reverse these results. Mechanistic analyses revealed that miR-4310 promoted glioma progression through the PI3K/AKT pathway by targeting PTEN. Additionally, SP1 induced the expression of miR-4310 by binding to its promoter region. Conclusion miR-4310 promotes the progression of glioma by targeting PTEN and activating the PI3K/AKT pathway; meanwhile, the expression of miR-4310 was induced by SP1.

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LncRNA MEG3 affects cell proliferation, apoptosis and migration by targeting miR-9-5p/MDK axis and activating PDK/AKT pathway in hepatocellular carcinoma

10.21203/rs.3.rs-21095/v1 ◽

2020 ◽

Author(s):

Dezhi Wu ◽

Zheng Ma ◽

Deyu Ma ◽

Qiquan Li

Keyword(s):

Hepatocellular Carcinoma ◽

Cell Proliferation ◽

Cell Lines ◽

Molecular Mechanisms ◽

Normal Liver ◽

Luciferase Reporter ◽

Akt Pathway ◽

Normal Liver Cell ◽

Non Coding Rna ◽

And Migration

Abstract Background Long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) was supposed to be a tumor suppressor in various cancers. However, the role of MEG3 in hepatocellular carcinoma (HCC) and the related molecular mechanisms are not well illustrated. This study aimed to determine the biological function of MEG3 in regulating HCC cell proliferation, apoptosis and migration. Moreover, the interaction among MEG3, microRNA (miR)-9-5p and Midkine (MDK), and the activation of phosphoinositide-dependent kinase (PDK)/AKT pathway in HCC cells were examined. Methods and Results Expression of MEG3 in a series of liver cancer cell lines was detected by RT-qPCR. Luciferase reporter assay, RT-qPCR and western blot were used to determine the interaction among MEG3, miR-9-5p and MDK, and the activation of PDK/AKT pathway. Cell proliferation and apoptosis were evaluated by CCK8, flow cytometry analysis for cell cycle and apoptosis, and Caspase 3/9 activity. Cell migration was determined by wound healing assay and MMP1 expression. We found MEG3 was decreased in HCC cell lines compared with the normal liver cell line. MEG3 suppressed HCC cell proliferation and migration, and induced cell apoptosis. Further, we found MEG3 targets miR-9-5p/MDK axis and modulates PDK/AKT pathway in HCC. Conclusion Our findings demonstrated that lncRNA MEG3 affects HCC cell proliferation, apoptosis and migration through its targeting of miR-9-5p/MDK and regulating of PDK/AKT pathway. This study suggested MEG3/miR-9-5p/MDK axis as the potential therapeutic target in HCC.

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NEAT1 Negatively Regulates Cell Proliferation and Migration of Neuroblastoma Cells by miR-183-5p/FOXP1 Via the ERK/AKT Pathway

Cell Transplantation ◽

10.1177/0963689720943608 ◽

2020 ◽

Vol 29 ◽

pp. 096368972094360

Author(s):

Weikang Pan ◽

Ali Wu ◽

Hui Yu ◽

Qiang Yu ◽

Baijun Zheng ◽

...

Keyword(s):

Cell Proliferation ◽

Neuroblastoma Cells ◽

Cell Counting ◽

Luciferase Reporter ◽

Akt Pathway ◽

Migration And Invasion ◽

Cell Proliferation And Migration ◽

Gene Assay ◽

And Migration ◽

Proliferation And Migration

Neuroblastoma, a malignant tumor of the sympathetic nervous system, is an aggressive extracranial tumor in childhood. Long noncoding RNAs (lncRNAs) have been discovered to play a key role in the eukaryotic regulatory gene network and be involved in a wide variety of biological processes. We observed that the expression of lncRNA nuclear-enriched abundant transcript-1 (NEAT1) was significantly decreased in human neuroblastoma tissues and cell lines, compared with the normal. We observed cell proliferation, migration, and invasion with Cell Counting Kit-8 assay, colony formation assay, and Transwell assay to investigate the effects of NEAT1, miR-183-5p, or FOXP1 on neuroblastoma cells. And we also used StarBase and luciferase reporter gene assay to predict and confirm the interaction of NEAT1, miR-183-5p, and FOXP1 in neuroblastoma cells. First, overexpression of NEAT1 suppressed cell proliferation and played a key role in cell migration and invasion. In addition, NEAT1 was demonstrated to directly interact with miR-183-5p and exerted its antioncogenic role in neuroblastoma by negatively regulating miR-183-5p expression. miR-183-5p suppressed the expression of FOXP1 and regulated cell proliferation and migration by directly targeting FOXP1 mRNA 3′-untranslated region. Moreover, FOXP1 antagonized the effect of miR-183-5p on the phosphorylation of extracellular-regulated kinase/protein kinase B (ERK/AKT), while FOXP1 siRNA increased the reduced phosphorylation of ERK/AKT caused by miR-183-5p inhibitor in neuroblastoma cells. Taken together, these data showed that NEAT1 negatively regulated cell proliferation and migration of neuroblastoma by the miR-183-5p/FOXP1 axis via suppression of the ERK/AKT pathway. Our findings may provide a new target for the study of pathogenesis and treatment of neuroblastoma.

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