Activation of epidermal growth factor receptor gene is involved in transforming growth factor-β-mediated fibronectin expression in a chondrocyte progenitor cell line

Takeda, Hiroyuki; Inoue, Hirofumi; Kutsuna, Tastuhiko; Matsushita, Natsuki; Takahashi, Toshiaki; Watanabe, Shohei; Higashiyama, Shigeki; Yamamoto, Haruyasu

doi:10.3892/ijmm_00000381

Int J Mol Med

2010

DOI: 10.3892/ijmm_00000381

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Activation of epidermal growth factor receptor gene is involved in transforming growth factor-β-mediated fibronectin expression in a chondrocyte progenitor cell line

Hiroyuki Takeda

Hirofumi Inoue²,

Tastuhiko Kutsuna³

et al.

Abstract: Abstract. Chondrocytes produce many types of ECM to maintain elasticity and plasticity in articular cartilage of revolute joints. Both transforming growth factor ß (TGF-ß) and bone morphogenetic proteins (BMPs) induce extracellular matrix proteins such as type II·1 collagen and aggrecan during chondrogenic differentiation in vitro. However, differences in the matrix gene expression pattern by the stimulation of TGF-ßs and BMPs remains unclear. In the present study, we created a customized PCR-based ECM array t… Show more

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Cited by 3 publications

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Periodic Mechanical Stress Activates PKCδ-Dependent EGFR Mitogenic Signals in Rat Chondrocytes via PI3K-Akt and ERK1/2

Shen

Gao

et al. 2016

Cell Physiol Biochem

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Background/Aims: The present study aimed to analyze the mechanisms by which periodic mechanical stress is translated into biochemical signals, and to verify the important role of signaling molecules including phosphatidylinositol-3-kinase (PI3K)-Akt, protein kinase C (PKC), and epidermal growth factor receptor (EGFR) in chondrocyte proliferation. The effects of periodic mechanical stress on the mitogenesis of chondrocytes have been studied extensively in recent years. However, the mechanisms underlying the ability of chondrocytes to sense and respond to periodic mechanical stress need further investigation. Methods: Two steps were undertaken in the experiment. In the first step, the cells were pretreated with shRNA targeted to Akt or EGFR or PKCδ or control scrambled shRNA. Moreover, they were pretreated with LY294002, GF109203X, Gö6976, rottlerin, and AG1478. They were maintained under static conditions or periodic mechanical stress for 3 days, 8 h per day, prior to direct cell counting and CCK-8 assay, respectively. In the second step, the cells were pretreated with shRNA targeted to Akt or EGFR or PKCδ or control scrambled shRNA. Moreover, they were pretreated with LY294002, AG1478, and rottlerin. They were maintained under static conditions or periodic mechanical stress for 1 h prior to Western blot analysis. Results: Proliferation was inhibited by pretreatment with PKC or PKCδ inhibitor GF109203X or rottlerin and by short hairpin RNA (shRNA) targeted to PKCδ, but not by PKCα inhibitor Gö6976 in chondrocytes in response to periodic mechanical stress. Meantime, rottlerin and shRNA targeted to PKCδ also attenuated EGFR, Akt, and ERK1/2 activation. Furthermore, inhibiting EGFR activity by AG1478 and shRNA targeted to EGFR abrogated chondrocyte proliferation and phosphorylation levels of Akt and extracellular signal-regulated kinase (ERK)1/2 subjected to periodic mechanical stress, while the phosphorylation site of PKCδ was not affected. In addition, pretreatment with the PI3K-Akt-selective inhibitor LY294002 and shRNA targeted to Akt reduced periodic mechanical stress-induced chondrocyte proliferation and phosphorylation of ERK1/2, while the phosphorylation levels of EGFR and PKCδ were not inhibited. Conclusion: These findings suggested that periodic mechanical stress promoted chondrocyte proliferation through PKCδ-EGFR-PI3K-Akt-ERK1/2. They provide a stronger viewpoint for further investigations into chondrocyte mechanobiology under periodic mechanical stress and the ways to improve the quality of tissue-engineered cartilage.

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Periodic Mechanical Stress Activates PKCδ-Dependent EGFR Mitogenic Signals in Rat Chondrocytes via PI3K-Akt and ERK1/2

Shen

Gao

et al. 2016

Cell Physiol Biochem

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Therapeutic Potential of Human Nasal Inferior Turbinate-Derived Stem Cells: Microarray Analysis of Multilineage Differentiation

Park

Kim

Lim

et al. 2021

ORL

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Introduction: Human nasal inferior turbinate-derived stem cells (hNTSCs) are attractive sources of adult stem cells for medical application because they can be easily obtained and cultivated with a highly proliferative capacity. The ability of hNTSCs to differentiate into chondrocytes, osteocytes, and neural cells makes them potential replacement therapeutic candidates in intractable disease. Nevertheless, detailed expression pattern of genes associated with trilineage differentiation (osteogenesis, chondrogenesis, and neurogenesis) in hNTSCs has not been revealed yet. Methods: In this study, we aimed to evaluate gene expression patterns of various transcription factors and marker genes associated with a particular lineage (osteogenesis, chondrogenesis, and neurogenesis) of differentiation of hNTSCs by DNA microarrays. Results: In microarrays, 36 transcripts such as E2F transcription factor 1, activating transcription factor 5, and AKR1B10 were upregulated and 36 transcripts such as CA9, PPFIA4, HAS2, and COL4A4 were downregulated in osteogenically differentiated hNTSCs. In chondrogenically differentiated hNTSCs, 3 transcripts (NUDT14, CPA4, and heparin-binding epidermal growth factor-like growth factor) were upregulated and 82 transcripts such as PTGS1, CLEC2D, and TET1 were downregulated. In neurally differentiated hNTSCs, 61 transcripts such as insulin-like growth factor-binding protein-1, nerve growth factor receptor, FGF1, OLFML1, and EPGN were upregulated and 98 transcripts such as ACAN, RUNX2, and C21orf96 were downregulated. In gene ontology (GO) analysis, cell signal-related GO terms were highly expressed. By contrast, catalysis GO terms and GO terms related to oxidoreductase were overrepresented in chondrogenically differentiated hNTSCs and osteogenically differentiated hNTSCs, respectively. Conclusion: Considering overall results, hNTSCs-specific genetic information may promote further studies on intracellular mechanisms defining key features of these cells.

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Blocking ERK1/2 signaling impairs TGF-β1 tumor promoting function but enhances its tumor suppressing role in intrahepatic cholangiocarcinoma cells

et al. 2017

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BackgroundTransforming growth factor-β (TGF-β) plays a paradoxical role in cancer: it suppresses proliferation at early stages but promotes metastasis at late stages. This cytokine is upregulated in cholangiocarcinoma and is implicated in cholangiocarcinoma invasion and metastasis. Here we investigated the roles of non-Smad pathway (ERK1/2) and Smad in TGF-β tumor promoting and suppressing activities in intrahepatic cholangiocarcinoma (ICC) cells.MethodsTGF-β1 effects on proliferation, invasion and migration of ICC cells, KKU-M213 and/or HuCCA-1, were investigated using MTT, colony formation, in vitro Transwell and wound healing assays. Levels of mRNAs and proteins/phospho-proteins were measured by quantitative (q)RT-PCR and Western blotting respectively. E-cadherin localization was examined by immunofluorescence and secreted MMP-9 activity was assayed by gelatin zymography. The role of ERK1/2 signaling was evaluated by treating cells with TGF-β1 in combination with MEK1/2 inhibitor U0126, and that of Smad2/3 and Slug using siSmad2/3- and siSlug-transfected cells.Resultsh-TGF-β1 enhanced KKU-M213 cell invasion and migration and induced epithelial-mesenchymal transition as shown by an increase in vimentin, Slug and secreted MMP-9 levels and by a change in E-cadherin localization from membrane to cytosol, while retaining the cytokine’s ability to attenuate cell proliferation. h-TGF-β1 stimulated Smad2/3 and ERK1/2 phosphorylation, and the MEK1/2 inhibitor U0126 attenuated TGF-β1-induced KKU-M213 cell invasion and MMP-9 production but moderately enhanced the cytokine growth inhibitory activity. The latter effect was more noticeable in HuCCA-1 cells, which resisted TGF-β-anti-proliferative activity. Smad2/3 knock-down suppressed TGF-β1 ability to induce ERK1/2 phosphorylation, Slug expression and cell invasion, whereas Slug knock-down suppressed cell invasion and vimentin expression but marginally affected ERK1/2 activation and MMP-9 secretion. These results indicate that TGF-β1 activated ERK1/2 through Smad2/3 but not Slug pathway, and that ERK1/2 enhanced TGF-β1 tumor promoting but repressed its tumor suppressing functions.ConclusionsInhibiting ERK1/2 activation attenuates TGF-β1 tumor promoting effect (invasion) but retains its tumor suppressing role, thereby highlighting the importance of ERK1/2 in resolving the TGF-β paradox switch.Electronic supplementary materialThe online version of this article (doi:10.1186/s12935-017-0454-2) contains supplementary material, which is available to authorized users.

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Activation of epidermal growth factor receptor gene is involved in transforming growth factor-β-mediated fibronectin expression in a chondrocyte progenitor cell line

Cited by 3 publications

References 29 publications

Periodic Mechanical Stress Activates PKCδ-Dependent EGFR Mitogenic Signals in Rat Chondrocytes via PI3K-Akt and ERK1/2

Periodic Mechanical Stress Activates PKCδ-Dependent EGFR Mitogenic Signals in Rat Chondrocytes via PI3K-Akt and ERK1/2

Therapeutic Potential of Human Nasal Inferior Turbinate-Derived Stem Cells: Microarray Analysis of Multilineage Differentiation

Blocking ERK1/2 signaling impairs TGF-β1 tumor promoting function but enhances its tumor suppressing role in intrahepatic cholangiocarcinoma cells

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