Neuritin Up-regulates Kv4.2 α-Subunit of Potassium Channel Expression and Affects Neuronal Excitability by Regulating the Calcium-Calcineurin-NFATc4 Signaling Pathway

Neuritin is an important neurotrophin that regulates neural development, synaptic plasticity, and neuronal survival. Elucidating the downstream molecular signaling is important for potential therapeutic applications of neuritin in neuronal dysfunctions. We previously showed that neuritin up-regulates transient potassium outward current (IA) subunit Kv4.2 expression and increases IA densities, in part by activating the insulin receptor signaling pathway. Molecular mechanisms of neuritin-induced Kv4.2 expression remain elusive. Here, we report that the Ca2+/calcineurin (CaN)/nuclear factor of activated T-cells (NFAT) c4 axis is required for neuritin-induced Kv4.2 transcriptional expression and potentiation of IA densities in cerebellum granule neurons. We found that neuritin elevates intracellular Ca2+ and increases Kv4.2 expression and IA densities; this effect was sensitive to CaN inhibition and was eliminated in Nfatc4−/− mice but not in Nfatc2−/− mice. Stimulation with neuritin significantly increased nuclear accumulation of NFATc4 in cerebellum granule cells and HeLa cells, which expressed IR. Furthermore, NFATc4 was recruited to the Kv4.2 gene promoter loci detected by luciferase reporter and chromatin immunoprecipitation assays. More importantly, data obtained from cortical neurons following adeno-associated virus-mediated overexpression of neuritin indicated that reduced neuronal excitability and increased formation of dendritic spines were abrogated in the Nfatc4−/− mice. Together, these data demonstrate an indispensable role for the CaN/NFATc4 signaling pathway in neuritin-regulated neuronal functions.

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The TLR-2/TonEBP signaling pathway regulates 29-kDa fibronectin fragment-dependent expression of matrix metalloproteinases

Scientific Reports ◽

10.1038/s41598-021-87813-8 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Hyun Sook Hwang ◽

Mi Hyun Lee ◽

Hyun Ah Kim

Keyword(s):

Matrix Metalloproteinases ◽

Signaling Pathway ◽

Mapk Signaling ◽

Nuclear Accumulation ◽

Activated T Cells ◽

Inflammatory Conditions ◽

Enhancer Binding Protein ◽

Fibronectin Fragments ◽

Voltage Dependent ◽

Voltage Dependent Calcium Channels

AbstractTonicity-responsive enhancer-binding protein (TonEBP; nuclear factor of activated T cells 5) is a transcription factor that responds to changes in osmolality. However, recent studies have shown that it also modulates immune responses under inflammatory conditions independently of hyperosmolality. Fibronectin fragments (FN-fs), which are abundant in the synovial fluid of patients with osteoarthritis (OA), induce expression of matrix metalloproteinases (MMPs) via the toll-like receptor-2 (TLR-2) signaling pathway. In this study we examined whether TonEBP is involved in 29-kDa FN-f-induced expression of MMPs. The expression of TonEBP was significantly higher in human osteoarthritis compared with normal cartilage samples. 29-kDa FN-f affected the expression of MMPs 1, 3, and 13 via TonEBP, and expression and nuclear accumulation of TonEBP were induced by activation of the phospholipase C/NF-κB/MAPK signaling pathway and, in particular, modulated by TLR-2. In addition, 29-kDa FN-f induced the expression of osmoregulatory genes, including Tau-T, SMIT, and AR, as well as voltage-dependent calcium channels via the TonEBP/TLR-2 signaling pathway. These results show that 29-kDa FN-f upregulates MMPs in chondrocytes via the TLR-2/TonEBP signaling pathway.

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Neuron-specific activation of necroptosis signaling in multiple sclerosis cortical grey matter

Acta Neuropathologica ◽

10.1007/s00401-021-02274-7 ◽

2021 ◽

Vol 141 (4) ◽

pp. 585-604 ◽

Cited By ~ 1

Author(s):

Carmen Picon ◽

Anusha Jayaraman ◽

Rachel James ◽

Catriona Beck ◽

Patricia Gallego ◽

...

Keyword(s):

Multiple Sclerosis ◽

Signaling Pathway ◽

Cortical Neurons ◽

Grey Matter ◽

Molecular Mechanisms ◽

Fold Increase ◽

Cell Types ◽

Meningeal Inflammation ◽

Cortical Grey Matter

AbstractSustained exposure to pro-inflammatory cytokines in the leptomeninges is thought to play a major role in the pathogenetic mechanisms leading to cortical pathology in multiple sclerosis (MS). Although the molecular mechanisms underlying neurodegeneration in the grey matter remain unclear, several lines of evidence suggest a prominent role for tumour necrosis factor (TNF). Using cortical grey matter tissue blocks from post-mortem brains from 28 secondary progressive MS subjects and ten non-neurological controls, we describe an increase in expression of multiple steps in the TNF/TNF receptor 1 signaling pathway leading to necroptosis, including the key proteins TNFR1, FADD, RIPK1, RIPK3 and MLKL. Activation of this pathway was indicated by the phosphorylation of RIPK3 and MLKL and the formation of protein oligomers characteristic of necrosomes. In contrast, caspase-8 dependent apoptotic signaling was decreased. Upregulation of necroptotic signaling occurred predominantly in macroneurons in cortical layers II–III, with little expression in other cell types. The presence of activated necroptotic proteins in neurons was increased in MS cases with prominent meningeal inflammation, with a 30-fold increase in phosphoMLKL+ neurons in layers I–III. The density of phosphoMLKL+ neurons correlated inversely with age at death, age at progression and disease duration. In vivo induction of chronically elevated TNF and INFγ levels in the CSF in a rat model via lentiviral transduction in the meninges, triggered inflammation and neurodegeneration in the underlying cortical grey matter that was associated with increased neuronal expression of TNFR1 and activated necroptotic signaling proteins. Exposure of cultured primary rat cortical neurons to TNF induced necroptosis when apoptosis was inhibited. Our data suggest that neurons in the MS cortex are dying via TNF/TNFR1 stimulated necroptosis rather than apoptosis, possibly initiated in part by chronic meningeal inflammation. Neuronal necroptosis represents a pathogenetic mechanism that is amenable to therapeutic intervention at several points in the signaling pathway.

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The TLR-2/TonEBP Signaling Pathway Regulates 29-kDa Fibronectin Fragment-dependent Expression of Matrix Metalloproteinases

10.21203/rs.3.rs-148101/v1 ◽

2021 ◽

Author(s):

Hyun Sook Hwang ◽

Mi Hyun Lee ◽

Hyun Ah Kim

Keyword(s):

Matrix Metalloproteinases ◽

Signaling Pathway ◽

Mapk Signaling ◽

Nuclear Accumulation ◽

Activated T Cells ◽

Inflammatory Conditions ◽

Enhancer Binding Protein ◽

Fibronectin Fragments ◽

Voltage Dependent ◽

Voltage Dependent Calcium Channels

Abstract Tonicity-responsive enhancer-binding protein (TonEBP; nuclear factor of activated T cells 5) is a transcription factor that responds to changes in osmolality. However, recent studies have shown that it also modulates immune responses under inflammatory conditions independently of hyperosmolality. Fibronectin fragments (FN-fs), which are abundant in the synovial fluid of patients with osteoarthritis (OA), induce expression of matrix metalloproteinases (MMPs) via the toll-like receptor-2 (TLR-2) signaling pathway. In this study we examined whether TonEBP is involved in 29-kDa FN-f-induced expression of MMPs. The expression of TonEBP was significantly higher in human osteoarthritis compared with normal cartilage samples. 29-kDa FN-f affected the expression of MMPs 1, 3, and 13 via TonEBP, and expression and nuclear accumulation of TonEBP were induced by activation of the phospholipase C/NF-κB/MAPK signaling pathway and, in particular, modulated by TLR-2. In addition, 29-kDa FN-f induced the expression of osmoregulatory genes, including Tau-T, SMIT, and AR, as well as voltage-dependent calcium channels via the TonEBP/TLR-2 signaling pathway. These results show that 29-kDa FN-f upregulates MMPs in chondrocytes via the TLR-2/TonEBP signaling pathway.

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MicroRNA-138-1-3p sensitizes sorafenib to hepatocellular carcinoma by targeting PAK5 mediated β-catenin/ABCB1 signaling pathway

10.21203/rs.3.rs-97867/v1 ◽

2020 ◽

Author(s):

Tong-tong Li ◽

Jie Mou ◽

Yao-jie Pan ◽

Fu-chun Huo ◽

Wen-qi Du ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Signaling Pathway ◽

Molecular Mechanisms ◽

Nuclear Translocation ◽

Kinase Inhibitor ◽

Flow Cytometric Analysis ◽

Luciferase Reporter ◽

Advanced Hepatocellular Carcinoma ◽

Cell Viability Assay ◽

Resistant Cells

Abstract Background: Kinase inhibitor sorafenib is the first-line targeted drug for advanced hepatocellular carcinoma (HCC) patients. However, the appearance of anti-cancer agents’ resistance has limited its therapeutic effect. Methods: In this study, quantitative real-time PCR (qPCR) and Western Blot were utilized to detect the levels of PAK5 in HCC sorafenib-resistant cells and their parental cells. The biological functions of miR-138-1-3p and PAK5 in sorafenib-resistant cells and their parental cells were explored by cell viability assay, plate colony formation assay and flow cytometric analysis. The potential mechanisms of PAK5 were evaluated via co-immunoprecipitation (co-IP), immunofluorescence, dual luciferase reporter assay and chromatin immunoprecipitation (ChIP). The effects of miR-138-1-3p and PAK5 on HCC sorafenib chemoresistant characteristics were investigated by a xenotransplantation model. Results: We detected significant down-regulation of miR-138-1-3p and up-regulation of PAK5 in HCC sorafenib resistance cell lines. Mechanical studies revealed that miR-138-1-3p reduced the protein expression of PAK5 by directly targeting the 3′-UTR of PAK5 mRNA. In addition, we verified that PAK5 elevated the phosphorylation and nuclear translocation of β-catenin that enhanced the transcriptional activity of multidrug resistance protein ABCB1. Conclusions: PAK5 contributed to the sorafenib chemoresistant characteristics of HCC by activity β-catenin/ABCB1 signaling pathway. Our findings identified the correlation between miR-138-1-3p and PAK5 and the molecular mechanisms of PAK5-mediated HCC sorafenib resistance, which provided a potential therapeutic target for advanced HCC patients.

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SNHG 6 promotes the progression of Colon and Rectal adenocarcinoma via miR-101-3p and Wnt/β-catenin signaling pathway

BMC Gastroenterology ◽

10.1186/s12876-019-1080-3 ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 10

Author(s):

Qianwen Shao ◽

Jing Xu ◽

Rong Deng ◽

Wei Wei ◽

Bing Zhou ◽

...

Keyword(s):

Signaling Pathway ◽

Molecular Mechanisms ◽

Rectal Adenocarcinoma ◽

Human Colon ◽

Luciferase Reporter ◽

Survival Times ◽

Reporter Assays ◽

Polymerase Chain ◽

Nucleolar Rna ◽

Mtt Assays

Abstract Background Small nucleolar RNA host gene 6 (SNHG6) regulates diverse biological processes in cancers. Potential function of SNHG6 in human colon and rectal adenocarcinoma (CRC) was evaluated. Methods Quantitative real-time polymerase chain reaction, MTT assays, Colony formation assays, Transwell assay, Western Blotting and Luciferase reporter assays were performed to measure the biological functions and potential molecular mechanisms of SNHG6 in CRC. Results SNHG6 was over-expressed in CRC, and high expression of s SNHG6 were associated with short survival times. We then identified miR-101-3p as an inhibitory target of SNHG6. Knockdown of SNHG6 significantly decreased miR-101-3p expression. Moreover, silenced SNHG6 obviously inhibited CRC cell growth, weakened cell invasion capacity and blocked the Wnt/β-catenin signaling pathway. Conclusion SNHG6 could regulate the progression of CRC via modulating the expression levels of miR-101-3p and the activity of Wnt/β-catenin signaling.

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miR-3188 Regulates Cell Proliferation, Apoptosis, and Migration in Breast Cancer by Targeting TUSC5 and Regulating the p38 MAPK Signaling Pathway

Oncology Research Featuring Preclinical and Clinical Cancer Therapeutics ◽

10.3727/096504017x14953948675421 ◽

2018 ◽

Vol 26 (3) ◽

pp. 363-372 ◽

Cited By ~ 11

Author(s):

Xiaowen Chen ◽

Jianli Chen

Keyword(s):

Breast Cancer ◽

Cell Proliferation ◽

Signaling Pathway ◽

P38 Mapk ◽

Molecular Mechanisms ◽

Mapk Signaling ◽

Mapk Signaling Pathway ◽

Luciferase Reporter ◽

And Migration ◽

Mcf 7

This study intended to investigate the effects of miR-3188 on breast cancer and to reveal the possible molecular mechanisms. miR-3188 was upregulated and TUSC5 was downregulated in breast cancer tissues and MCF-7 cells compared to normal tissue and MCF-10 cells. After MCF-7 cells were transfected with miR-3188 inhibitor, cell proliferation and migration were inhibited, whereas apoptosis was promoted. Luciferase reporter assay suggested that TUSC5 was a target gene of miR-3188. In addition, miR-3188 overexpression increased the p-p38 expression, while miR-3188 suppression decreased the p-p38 expression significantly. miR-3188 regulated breast cancer progression via the p38 MAPK signaling pathway. In conclusion, miR-3188 affects breast cancer cell proliferation, apoptosis, and migration by targeting TUSC5 and activating the p38 MAPK signaling pathway. miR-3188 may serve as a potential therapeutic agent for the treatment of breast cancer.

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TIPE Regulates DcR3 Expression and Function by Activating the PI3K/AKT Signaling Pathway in CRC

Frontiers in Oncology ◽

10.3389/fonc.2020.623048 ◽

2021 ◽

Vol 10 ◽

Author(s):

Mengya Zhong ◽

Xingfeng Qiu ◽

Yu Liu ◽

Yan Yang ◽

Lei Gu ◽

...

Keyword(s):

Cell Proliferation ◽

Signaling Pathway ◽

Molecular Mechanisms ◽

Akt Signaling ◽

Luciferase Reporter ◽

Akt Signaling Pathway ◽

Cell Proliferation And Migration ◽

And Migration ◽

Hct116 Cells ◽

Proliferation And Migration

Tumor necrosis factor-induced protein-8 (TIPE) is highly expressed in colorectal cancer (CRC). Decoy receptor 3 (DcR3) is a soluble secreted protein that can antagonize Fas ligand (FasL)-induced apoptosis and promote tumorigenesis. It remains unclear whether TIPE can regulate DcR3 expression. In this study, we examined this question by analyzing the relationship between these factors in CRC. Bioinformatics and tissue microarrays were used to determine the expression of TIPE and DcR3 and their correlation in CRC. The expression of TIPE and DcR3 in colon cancer cells was detected. Plasma samples were collected from CRC patients, and DcR3 secretion was measured. Then, dual-luciferase reporter gene analysis was performed to assess the interaction between TIPE and DcR3. We exogenously altered TIPE expression and analyzed its function and influence on DcR3 secretion. Lipopolysaccharide (LPS) was used to stimulate TIPE-overexpressing HCT116 cells, and alterations in signaling pathways were detected. Additionally, inhibitors were used to confirm molecular mechanisms. We found that TIPE and DcR3 were highly expressed in CRC patients and that their expression levels were positively correlated. DcR3 was highly expressed in the plasma of cancer patients. We confirmed that TIPE and DcR3 were highly expressed in HCT116 cells. TIPE overexpression enhanced the transcriptional activity of the DcR3 promoter. TIPE activated the PI3K/AKT signaling pathway to regulate the expression of DcR3, thereby promoting cell proliferation and migration and inhibiting apoptosis. In summary, TIPE and DcR3 are highly expressed in CRC, and both proteins are associated with poor prognosis. TIPE regulates DcR3 expression by activating the PI3K/AKT signaling pathway in CRC, thus promoting cell proliferation and migration and inhibiting apoptosis. These findings may have clinical significance and promise for applications in the treatment or prognostication of CRC.

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MiR-128-3p Post-Transcriptionally Inhibits WISP1 to Suppress Apoptosis and Inflammation in Human Articular Chondrocytes via the PI3K/AKT/NF-κB Signaling Pathway

Cell Transplantation ◽

10.1177/0963689720939131 ◽

2020 ◽

Vol 29 ◽

pp. 096368972093913

Author(s):

Shujun Chen ◽

Bo Li

Keyword(s):

Signaling Pathway ◽

Proinflammatory Cytokines ◽

Molecular Mechanisms ◽

Articular Chondrocytes ◽

Negative Regulator ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Gene Assay ◽

Induced Apoptosis

In osteoarthritis (OA), the synthesis and decomposition of the extracellular matrix (ECM) are imbalanced. High expression levels of Wnt1-inducible signaling pathway protein 1 (WISP1) promote the synthesis of matrix metalloproteinases and induce the degradation of cartilage, which aggravates the OA. The aim of this study was to explore the role of miR-128-3p in the development of OA. In the present study, the expression of WISP1 and miR-128-3p in osteoarthritic tissues and chondrocytes was detected using quantitative reverse transcription PCR (RT-qPCR) and Western blotting. Then we predicted that WISP1 might be a potential target gene of miR-128-3p by TargetScan and verified using luciferase reporter gene assay. The effect of miR-128-3p or WISP1 on chondrocytes was evaluated by cell proliferation assay, apoptosis, and caspase-3 activity assay. To further reveal the molecular mechanisms of miR-128-3p in osteoarthritic development, the degradation of chondrocyte matrix and production of proinflammatory cytokines in osteoarthritic chondrocyte model were detected by ELISA. To mimic the osteoarthritic microenvironment in vitro studies, chondrocytes were stimulated with interleukin (IL)-1β, and then we found that the expression of miR-128-3p was downregulated. Overexpression of WISP1 inhibited the proliferation of chondrocytes, which induced apoptosis, degradation of chondrocyte matrix, production of proinflammatory cytokines, and activated the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt)/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway. Then, we identified that miR-128-3p was a negative regulator of WISP1 by directly targeting its 3′-untranslated region (UTR). Moreover, the PI3K allosteric activator 740 Y-P abolished the inhibition of miR-128-3p in apoptosis, degradation of chondrocyte matrix, and inflammation. Our results showed that miR-128-3p targets WISP1 to regulate chondrocyte proliferation, apoptosis, degradation of chondrocyte matrix, and production of proinflammatory cytokines via the PI3K/Akt/NF-κB pathway, which plays a suppressed role in OA.

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FOXH1 promotes lung cancer progression by activating the Wnt/β-catenin signaling pathway

Cancer Cell International ◽

10.1186/s12935-021-01995-9 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Jun Zhang ◽

Xian Zhang ◽

Shasha Yang ◽

Yanqiu Bao ◽

Dongyuan Xu ◽

...

Keyword(s):

Lung Cancer ◽

Cancer Cells ◽

Signaling Pathway ◽

Cancer Progression ◽

Molecular Mechanisms ◽

Lung Cancer Cells ◽

Cell Counting ◽

Luciferase Reporter ◽

Mesenchymal Transition

Abstract Background The expression of forkhead box protein H1 (FOXH1) is frequently upregulated in various cancers. However, the molecular mechanisms underlying the association between FOXH1 expression and lung cancer progression still remain poorly understood. Thus, the main objective of this study is to explore the role of FOXH1 in lung cancer. Methods The Cancer Genome Atlas dataset was used to investigate FOXH1 expression in lung cancer tissues, and the Kaplan–Meier plotter dataset was used to determine the role of FOXH1 in patient prognosis. A549 and PC9 cells were transfected with short hairpin RNA targeting FOXH1 mRNA. The Cell Counting Kit-8, colony formation, soft agar, wound healing, transwell invasion and flow cytometry assays were performed to evaluate proliferation, migration and invasion of lung cancer cells. Tumorigenicity was examined in a BALB/c nude mice model. Western blot analysis was performed to assess the molecular mechanisms, and β-catenin activity was measured by a luciferase reporter system assay. Results Higher expression level of FOXH1 was observed in tumor tissue than in normal tissue, and this was associated with poor overall survival. Knockdown of FOXH1 significantly inhibited lung cancer cell proliferation, migration, invasion, and cycle. In addition, the mouse xenograft model showed that knockdown of FOXH1 suppressed tumor growth in vivo. Further experiments revealed that FOXH1 depletion inhibited the epithelial-mesenchymal transition of lung cancer cells by downregulating the expression of mesenchymal markers (Snail, Slug, matrix metalloproteinase-2, N-cadherin, and Vimentin) and upregulating the expression of an epithelial marker (E-cadherin). Moreover, knockdown of FOXH1 significantly downregulated the activity of β-catenin and its downstream targets, p-GSK-3β and cyclin D1. Conclusion FOXH1 exerts oncogenic functions in lung cancer through regulation of the Wnt/β-catenin signaling pathway. FOXH1 might be a potential therapeutic target for patients with certain types of lung cancer.

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

Cellular Physiology and Biochemistry ◽

10.1159/000490932 ◽

2018 ◽

Vol 47 (4) ◽

pp. 1589-1603 ◽

Cited By ~ 18

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.

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