Hypoxia and STAT3 signalling interactions regulate pro-inflammatory pathways in rheumatoid arthritis

2014 ◽  
Vol 74 (6) ◽  
pp. 1275-1283 ◽  
Author(s):  
Wei Gao ◽  
Jennifer McCormick ◽  
Mary Connolly ◽  
Emese Balogh ◽  
Douglas J Veale ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Target Genes ◽  
Ex Vivo ◽  
Notch 1 ◽  
Functional Link ◽  
Jak2 Inhibitor ◽  
Downstream Target ◽  
Inflammatory Pathways ◽  
And Migration ◽  
Spontaneous Secretion

ObjectiveTo examine the effect of hypoxia on Signal Transducer and Activator of Transcription 3 (STAT3)-induced pro-inflammatory pathways in rheumatoid arthritis (RA).MethodsDetection of phospho-STAT3 was assessed in RA synovial tissue and fibroblasts (RASFC) by immunohistology/immunofluorescence. Primary RASFCs and a normal synoviocyte cell line (K4IM) were cultured under hypoxic and normoxic conditions±Stat3-siRNA, HIF-siRNA or WP1066 (JAK2-inhibitor). HIF1α, p-STAT3, p-STAT1 and Notch-1IC protein expression were analysed by western blot. Functional mechanisms were quantified by invasion chamber, matrigel and migration assays. IL-6, IL-8, IL-10 and matrixmetalloproteinases (MMP)-3 were quantified by ELISA. Notch-1 receptor, its DLL-4 ligand and downstream target genes (hrt-1, hrt-2) were quantified by real-time PCR. The effect of WP1066 on spontaneous secretion of pro/anti-inflammatory cytokines and Notch signalling was examined in RA synovial explants ex vivo.Resultsp-STAT3 was increased in RA synovium compared with control (p<0.05). Hypoxia induced p-STAT3, p-STAT1 and HIF1α expression, an effect blocked by Stat3-siRNA and WP1066. Hypoxia-induced cell invasion, migration and cytokine production were inhibited by Stat3-siRNA (p<0.05) and WP1066 (p<0.05). While HIF1α siRNA inhibited hypoxia-induced p-STAT3 detection, Stat3-siRNA also inhibited hypoxia-induced HIF1α. Furthermore, hypoxia-induced Notch-1IC, DLL4, hrt-1 and -2 expression were significantly inhibited by WP1066 (p<0.05). Finally, in RA synovial explant cultures ex vivo, WP1066 decreased spontaneous secretion of IL-6, IL-8 and MMP3 (p<0.05), Notch-1 mRNA (p<0.05) and induced IL-10 (p<0.05).ConclusionsThis is the first study to provide evidence of a functional link between HIF1α, STAT3 and Notch-1 signalling in the regulation of pro-inflammatory mechanisms in RA, and further supports a role for STAT blockade in the treatment of RA.

scholarly journals si-SNHG5-FOXF2 inhibits TGF-β1-induced fibrosis in human primary endometrial stromal cells by the Wnt/β-catenin signalling pathway

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Limin Liu ◽  
Guobin Chen ◽  
Taoliang Chen ◽  
Wenjuan Shi ◽  
Haiyan Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Stromal Cells ◽  
Target Genes ◽  
Cell Model ◽  
Ex Vivo ◽  
Signalling Pathway ◽  
Endometrial Stromal Cells ◽  
Downstream Target ◽  
Related Proteins ◽  
Tgf Β1

Abstract Background Intrauterine adhesions (IUAs) are manifestations of endometrial fibrosis characterized by inflammation and fibrinogen aggregation in the extracellular matrix (ECM). The available therapeutic interventions for IUA are insufficiently effective in the clinical setting for postoperative adhesion recurrence and infertility problems. In this study, we investigated whether si-SNHG5-FOXF2 can serve as a molecular mechanism for the inhibition of IUA fibrosis ex vivo. Methods FOXF2, TGF-β1 and collagen expression levels were measured by microarray sequencing analysis in three normal endometrium groups and six IUA patients. We induced primary human endometrial stromal cells (HESCs) into myofibroblasts (MFs) to develop an IUA cell model with various concentrations of TGF-β1 at various times. Downstream target genes of FOXF2 were screened by chromatin immunoprecipitation combined with whole-genome high-throughput sequencing (ChIP-seq). We investigated ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs with FOXF2 downregulation by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blotting (WB), immunohistochemistry (IHC), flow cytometry, ethylenediurea (EdU) and CCK8 assays. We identified long noncoding RNAs (lncRNA) SNHG5 as the upstream regulatory gene of FOXF2 through RNA immunoprecipitation (RIP), RNA pulldown and fluorescence in situ hybridization (FISH). Finally, we examined FOXF2 expression, ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs upon FOXF2 downregulation. Results FOXF2 was highly expressed in the endometrium of patients with IUA. Treatment of primary HESCs with 10 ng/ml TGF-β1 for 72 h was found to be most effective for developing an IUA cell model. FOXF2 regulated multiple downstream target genes, including collagen, vimentin (VIM) and cyclin D2/DK4, by ChIP-seq and ChIP-PCR. FOXF2 downregulation inhibited TGF-β1-mediated primary HESC fibrosis, including ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related protein expression. We identified lncRNA SNHG5 as an upstream gene that directly regulates FOXF2 by RIP-seq, qRT-PCR, WB and FISH. SNHG5 downregulation suppressed FOXF2 expression in the IUA cell model, resulting in synergistic repression of the Wnt/β-catenin pathway, thereby altering TGF-β1-mediated ECM aggregation in endometrial stromal cells ex vivo. Conclusions Regulation of the Wnt/β-catenin signalling pathway and ECM formation by si-SNHG5-FOXF2 effectively inhibited the profibrotic effect of TGF-β1 on primary HESCs. This finding can provide a molecular basis for antagonizing TGF-β1-mediated fibrosis in primary HESCs.

Abstract 2178: Sonic Hedgehog In Adult Hypoxic Cerebellum

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Giuseppe Straface ◽  
Andrea Flex ◽  
Federico Biscetti ◽  
Eleonora Gaetani ◽  
Giovanni Pecorini ◽  
...  
Keyword(s):  
Sonic Hedgehog ◽  
Target Genes ◽  
Positive Staining ◽  
Lacz Gene ◽  
Downstream Target ◽  
Shh Pathway ◽  
And Migration ◽  
First Time ◽  
Proliferation And Migration

Background: Cerebellar hypoxia is responsible for important aspects of cognitive deterioration and motor disturbances in neurological disorders, such as stroke, vascular dementia, and neurodegeneration. In the cerebellum, VEGF is significantly upregulated after hypoxia and is able to induce angiogenesis, reduce neuronal apoptosis, and regulate neuronal differentiation, proliferation, and migration. But, VEGF is not sufficient to provide neuroprotection. A crucial role is played by growth associated protein-43 (GAP43), for which important activities have been described. The purpose of this study was to investigate the role of the developmental Sonic hedgehog (Shh) signaling pathway in postnatal hypoxic cerebellum and its relationship with VEGF and GAP43 expression. Methods: We used adult C57BL/6J mice, ptc1-lacZ mice, and GAP43−/− mice for these experiments. Ptc1-lacZ mice carry a non-disruptive insertion of the lacZ gene under the control of the ptc1 promoter. Ptc1 is a downstream-transcriptional target of Shh and its upregulation indicates activation of the Shh pathway. Mice were exposed to systemic normobaric hypoxia (6%O 2 ) for 6 hours and the expression of Shh, Ptc1, VEGF, and GAP43 were investigated. Results: After exposure to hypoxia, Shh-positive staining was detected in Purkinje cells (PCs). The same cells were also lacZ(ptc1)-positive, indicating that PCs are both Shh-producing and -responding elements. Also the cells of the internal granular layer (IGL) were lacZ(ptc1)-positive, indicating that these cells are Shh-responsive. LacZ(ptc1)-positive IGL cells were also immunopositive for VEGF and GAP-43. We also found that ptc1 expression is lost in PCs of GAP43−/− mice, indicating that Shh requires GAP43 to activate its downstream target genes in PCs. Finally, when cultures enriched in granular cells were stimulated with Shh recombinant protein, GAP43 phosphorylation was increased. This effect was inhibited by Shh-inhibitor cyclopamine. Conclusions: This is the first time that hypoxia is reported to activate the Shh pathway in the adult. Our data suggest that the Shh pathway might be important for the cerebellar response to hypoxia, through interactions with VEGF and GAP43.

scholarly journals si-SNHG5-FOXF2 inhibits TGF-β1-induced fibrosis in human primary endometrial stromal cells by the Wnt/β-catenin signalling pathway

2020 ◽  
Author(s):  
Limin Liu ◽  
Guobin Chen ◽  
Taoliang Chen ◽  
Wenjuan Shi ◽  
Haiyan Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Stromal Cells ◽  
Target Genes ◽  
Cell Model ◽  
Ex Vivo ◽  
Signalling Pathway ◽  
Endometrial Stromal Cells ◽  
Downstream Target ◽  
Related Proteins ◽  
Tgf Β1

Abstract Background: Intrauterine adhesions (IUAs) are manifestations of endometrial fibrosis characterized by inflammation and fibrinogen aggregation in the extracellular matrix (ECM). The available therapeutic interventions for IUA are insufficiently effective in the clinical setting for postoperative adhesion recurrence and infertility problems. In this study, we investigated whether si-SNHG5-FOXF2 can serve as a molecular mechanism for the inhibition of IUA fibrosis ex vivo.Methods: FOXF2, TGF-β1 and collagen expression levels were measured by microarray sequencing analysis in three normal endometrium groups and six IUA patients. We induced primary human endometrial stromal cells (HESCs) into myofibroblasts (MFs) to develop an IUA cell model with various concentrations of TGF-β1 at various times. Downstream target genes of FOXF2 were screened by chromatin immunoprecipitation combined with whole-genome high-throughput sequencing (ChIP-seq). We investigated ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs with FOXF2 downregulation by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blotting (WB), immunohistochemistry (IHC), flow cytometry, ethylenediurea (EdU) and CCK8 assays. We identified lncRNA SNHG5 as the upstream regulatory gene of FOXF2 through RNA immunoprecipitation (RIP), RNA pulldown and fluorescence in situ hybridization (FISH). Finally, we examined FOXF2 expression, ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related proteins in primary HESCs upon FOXF2 downregulation.Results: FOXF2 was highly expressed in the endometrium of patients with IUA. Treatment of primary HESCs with 10 ng/ml TGF-β1 for 72 h was found to be most effective for developing an IUA cell model. FOXF2 regulated multiple downstream target genes, including collagen, vimentin (VIM) and cyclin D2/DK4, by ChIP-seq and ChIP-PCR. FOXF2 downregulation inhibited TGF-β1-mediated primary HESC fibrosis, including ECM formation, cell proliferation and Wnt/β-catenin signalling pathway-related protein expression. We identified lncRNA SNHG5 as an upstream gene that directly regulates FOXF2 by RIP-seq, qRT-PCR, WB, and FISH. SNHG5 downregulation suppressed FOXF2 expression in the IUA cell model, resulting in synergistic repression of the Wnt/β-catenin pathway, thereby altering TGF-β1-mediated ECM aggregation in endometrial stromal cells ex vivo. Conclusions: Regulation of the Wnt/β-catenin signalling pathway and ECM formation by si-SNHG5-FOXF2 effectively inhibited the profibrotic effect of TGF-β1 on primary HESCs. This finding can provide a molecular basis for antagonizing TGF-β1-mediated fibrosis in primary HESCs.

Globular adiponectin and its downstream target genes are up-regulated locally in human colorectal tumors: ex vivo and in vitro studies

Metabolism ◽  
2014 ◽  
Vol 63 (5) ◽  
pp. 672-681 ◽  
Author(s):  
Katja Kannisto Vetvik ◽  
Tonje Sonerud ◽  
Mona Lindeberg ◽  
Torben Lüders ◽  
Ragnhild H. Størkson ◽  
...  
Keyword(s):  
Target Genes ◽  
Ex Vivo ◽  
In Vitro Studies ◽  
Colorectal Tumors ◽  
Downstream Target ◽  
Globular Adiponectin

scholarly journals RETRACTED ARTICLE: Circular RNA ABCB10 promotes hepatocellular carcinoma progression by increasing HMG20A expression by sponging miR-670-3p

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Yu Fu ◽  
Limin Cai ◽  
Xuexue Lei ◽  
Dunwei Wang
Keyword(s):  
Hepatocellular Carcinoma ◽  
Target Genes ◽  
Gene Prediction ◽  
Critical Role ◽  
Circular Rna ◽  
Luciferase Reporter ◽  
Brdu Incorporation ◽  
Downstream Target ◽  
Retracted Article ◽  
And Migration

Abstract Background/aims The dysregulation of circABCB10 may play an critical role in tumor progression. However, its function in liver cancer (HCC) is still unclear. Therefore, this experimental design is based on circABCB10 to explore the pathogenesis of HCC. Methods The expression of circABCB10 and miR-670-3p in HCC tissues was detected by RT-qPCR. CCK-8, Brdu incorporation, colony formation and transwell assays were used to determine the effect of circABCB10 on HCC cell proliferation and migration. Target gene prediction and screening, luciferase reporter assays were used to validate downstream target genes of circABCB10 and miR-670-3p. HMG20A expression was detected by RT-qPCR and Western blotting. The tumor changes in mice were detected by in nude mice. Results CircABCB10 was significantly increased in HCC tissues and cell lines, and high CircABCB10 expression was directly associated with low survival in HCC patients. Silencing of circABCB10 inhibited proliferation and invasion of hepatocellular carcinoma. In addition, circABCB10 acted as a sponge of miR-670-3p to upregulate HMG20A expression. In addition, overexpression of miR-670-3p or knockdown of HMG20A reversed the carcinogenic effects of circABCB10 in HCC. There was a negative correlation between the expression of circABCB10 and miR-670-3p, and a positive correlation between the expression of circABCB10 and HMG20A in HCC tissues. Conclusion circABCB10 promoted HCC progression by modulating the miR-670-3p/HMG20A axis, and circABCB10 may be a potential therapeutic target for HCC. Trail registration JL1H384739, registered at Sep 09, 2014.

Derlin-1 functions as a growth promoter in breast cancer

2020 ◽  
Vol 401 (3) ◽  
pp. 377-387 ◽  
Author(s):  
Yansong Liu ◽  
Ziming Wang ◽  
Handong Liu ◽  
Xin Wang ◽  
Zhonghua Zhang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Target Genes ◽  
Malignant Tumors ◽  
Expression Patterns ◽  
Growth Promoter ◽  
Downstream Target ◽  
Protein Levels ◽  
Kaplan Meier ◽  
And Migration ◽  
Predicting Factor

AbstractBreast cancer is one of the most common malignant tumors in women. Derlin-1 has been found to be overexpressed in several human cancers in addition to playing an important role in tumor processes; however, the expression patterns and functions of Derlin-1 in human breast cancer are not fully understood. In this study, we found that Derlin-1 overexpression was higher in breast cancer compared to normal samples through TCGA and GTEx database analyses. Kaplan-Meier plotter analysis showed that Derlin-1 was a predicting factor for patient prognosis. Derlin-1 expression was significantly up-regulated in breast cancer tissues (18/30, 60.00%) compared to corresponding paracancerous tissue (9/30, 30.00%, p < 0.05) as detected by immunohistochemistry, and the expression of Derlin-1 was correlated to pathological grading. siRNA interference of Derlin-1 inhibited cell proliferation, which is associated with the promotion of apoptosis and migration. Derlin-1 knockdown suppressed the protein levels of p-AKT and Cyclin D1 while up-regulating Caspase3 and Bax. GEPIA database analysis showed that MTDH and ATAD2 were downstream target genes, and the expression of MTDH and was suppressed in Derlin-1 knockdown cells. Taken together, our results demonstrated ATAD2 that Derlin-1 is overexpressed in breast cancer and promoted a malignant phenotype through the AKT signaling pathway.

scholarly journals Erratum

2021 ◽  
Vol 28 (6) ◽  
pp. 681-682
Author(s):  
Juan Gu ◽  
Chang-fu Cui ◽  
Li Yang ◽  
Ling Wang ◽  
Xue-hua Jiang
Keyword(s):  
Signaling Pathway ◽  
Target Genes ◽  
Mrna Level ◽  
Xenograft Model ◽  
Dependent Manner ◽  
Invasion And Migration ◽  
Downstream Target ◽  
And Migration

Colon cancer (CC) is the third most common cancer worldwide. Emodin is an anthraquinone-active substance that has the ability to affect tumor progression. Our study aims to explore the effects and the relevant mechanism of emodin on the invasion and migration of CC in vitro and in vivo. In our study, we found that emodin inhibited the invasion and migration abilities of RKO cells and decreased the expression of matrix metalloproteinase-7 (MMP-7), MMP-9, and vascular endothelial growth factor (VEGF) in a dose-dependent manner. Further research suggested that emodin inhibited EMT by increasing the mRNA level of E-cadherin and decreasing the expression of N-cadherin, Snail, and -catenin. Emodin also significantly inhibited the activation of the Wnt/-catenin signaling pathway by downregulating the expression of related downstream target genes, including TCF4, cyclin D1, and c-Myc. A Wnt/-catenin signaling pathway agonist abolished the effect of emodin on EMT and cell mobility, suggesting that emodin exerted its regulating role through the Wnt/-catenin pathway. The CC xenograft model was established to study the antitumor efficiency of emodin in vivo. The in vivo study further demonstrated that emodin (40 mg/kg) suppressed tumor growth by inhibiting EMT via the Wnt/-catenin signaling pathway in vivo. Taken together, we suggest that emodin inhibits the invasion and migration of CC cells in vitro and in vivo by blocking EMT, which is related with the inhibition of the Wnt/-catenin signaling pathway.

scholarly journals Baicalin alleviates hyperglycemia-induced endothelial impairment via Nrf2

2019 ◽  
Vol 240 (1) ◽  
pp. 81-98 ◽  
Author(s):  
Gen Chen ◽  
Xiangjuan Chen ◽  
Chao Niu ◽  
Xiaozhong Huang ◽  
Ning An ◽  
...  
Keyword(s):  
Oxidative Damage ◽  
Nuclear Export ◽  
Target Genes ◽  
Ex Vivo ◽  
Umbilical Vein ◽  
Protective Effects ◽  
Cellular Mechanisms ◽  
Downstream Target ◽  
Nrf2 Activation

Baicalin is the major component found in Scutellaria baicalensis root, a widely used herb in traditional Chinese medicine, which exhibits strong anti-inflammatory, anti-viral and anti-tumor activities. The present work was devoted to elucidate the molecular and cellular mechanisms underlying the protective effects of Baicalin against diabetes-induced oxidative damage, inflammation and endothelial dysfunction. Diabetic mice, induced by streptozotocin (STZ), were treated with intraperitoneal Baicalin injections. Human umbilical vein endothelial cells (HUVECs) were cultured either in normal glucose (NG, 5.5 mM) or high glucose (HG, 33 mM) medium in the presence or absence of Baicalin for 72 h. We observed an obvious inhibition of hyperglycemia-triggered oxidative damage and inflammation in HUVECs and diabetic aortal vasculature by Baicalin, along with restoration of hyperglycemia-impaired nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway activity. However, the protective effects of Baicalin almost completely abolished in HUVECs transduced with shRNA against Nrf2, but not with nonsense shRNA. Mechanistic studies demonstrated that HG decreased Akt and GSK3B phosphorylation, restrained nuclear export of Fyn and nuclear localization of Nrf2, blunted Nrf2 downstream target genes and subsequently induced oxidative stress in HUVECs. However, those destructive cascades were well prevented by Baicalin in HUVECs. Furthermore, LY294002 and ML385 (inhibitor of PI3K and Nrf2) attenuated Baicalin-mediated Nrf2 activation and the ability of facilitates angiogenesis in vivo and ex vivo. Taken together, the endothelial protective effect of Baicalin under hyperglycemia condition could be partly attributed to its role in downregulating reactive oxygen species (ROS) and inflammation via the Akt/GSK3B/Fyn-mediated Nrf2 activation.

scholarly journals Delivery of miR-26a Using an Exosomes-Based Nanosystem Inhibited Proliferation of Hepatocellular Carcinoma

2021 ◽  
Vol 8 ◽  
Author(s):  
Shaya Mahati ◽  
Xiangjun Fu ◽  
Xuexian Ma ◽  
Hua Zhang ◽  
Lei Xiao
Keyword(s):  
Hepatocellular Carcinoma ◽  
Target Genes ◽  
Great Promise ◽  
Antitumor Therapy ◽  
Human Cord Blood ◽  
Downstream Target ◽  
New Perspective ◽  
And Migration ◽  
Genetic Engineering Technology

Background: MicroRNA (abbreviated miRNA)-based treatment holds great promise for application as clinical antitumor therapy, but good carriers for delivery of the miRNA drug are lacking. Exosomes secreted by mesenchymal stem cells (MSCs) have proved to be safe, and exogenously modified exosomes may potentially represent an excellent drug delivery vehicle.Methods: In this study, we designed a delivery nano system using single-stranded variable fragment (scFv)-modified exosomes derived from human cord blood MSCs. Genetic engineering technology was used to obtain anti-Glypican 3 (GPC3) scFv-modified exosomes, which were then loaded with miR-26a mimics through electroporation.Results: Results of electron microscopy and dynamic light scattering indicated that the diameter of the drug-carrying exosomes was about 160 nm. Furthermore, anti-GPC3 scFv-modified exosomes effectively delivered miR-26a to GPC3-positive hepatocellular carcinoma cells, thereby inhibiting cell proliferation and migration by regulating the expression of downstream target genes of miR-26a. The exosomes-based nano system displayed favorable anti-tumor effect in vivo with no obvious side effects.Conclusion: Our data provided a new perspective for the use of exosome delivery systems for miRNA-based antitumor therapy.

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