• The Korean Journal of Physiology and Pharmacology
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• 제17권4호
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• pp.307-314
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• 2013

Connective tissue growth factor (CTGF) is a potent pro-fibrotic factor, which is implicated in fibrosis through extracellular matrix (ECM) induction in diabetic cardiovascular complications. It is an important downstream mediator in the fibrotic action of transforming growth factor ${\beta}$ ($TGF{\beta}$) and is potentially induced by hyperglycemia in human vascular smooth muscle cells (VSMCs). Therefore, the goal of this study is to identify the signaling pathways of CTGF effects on ECM accumulation and cell proliferation in VSMCs under hyperglycemia. We found that high glucose stimulated the levels of CTGF mRNA and protein and followed by VSMC proliferation and ECM components accumulation such as collagen type 1, collagen type 3 and fibronectin. By depleting endogenous CTGF we showed that CTGF is indispensable for the cell proliferation and ECM components accumulation in high glucose-stimulated VSMCs. In addition, pretreatment with the MEK1/2 specific inhibitors, PD98059 or U0126 potently inhibited the CTGF production and ECM components accumulation in high glucose-stimulated VSMCs. Furthermore, knockdown with ERK1/2 MAPK siRNA resulted in significantly down regulated of CTGF production, ECM components accumulation and cell proliferation in high glucose-stimulated VSMCs. Finally, ERK1/2 signaling regulated Egr-1 protein expression and treatment with recombinant CTGF reversed the Egr-1 expression in high glucose-induced VSMCs. It is conceivable that ERK1/2 MAPK signaling pathway plays an important role in regulating CTGF expression and suggests that blockade of CTGF through ERK1/2 MAPK signaling may be beneficial for therapeutic target of diabetic cardiovascular complication such as atherosclerosis.

• 한방재활의학과학회지
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• 제30권3호
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• pp.71-87
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• 2020

Objectives Even though the various alternative herbal medicine has applied for osteoarthritis (OA) treatment, its scientific proof remains uncertain. The aim of the present study evaluates the effects of Chulbu-tang on inflammatory responses in a monosodium iodoacetate (MIA)-induced osteoarthritis rat model. Methods OA rat model was established by MIA injection in intra-joint of rats. 7 days after, OA rats except OA control rats were administrated Chulbu-tang (100 or 200 mg/kg) or Indomathacin (5 mg/kg) once a day for 14 days. The weight-bearing ability of hind paws were measured when group isolation 0, 7, and 14 days. Western blotting was performed to examine the knockdown/overexpressing efficiency of Chulbu-tang. In addition, cartilage destruction was measured histologically. Results Chulbu-tang treatment significantly reduced the protein expressions of inflammatory mediators such as inducible nitric oxide synthase and cyclooxygenase 2, and inhibited inflammatory cytokines including tumor necrosis factor alpha, interleukin (IL)-1β, and IL-6 through nuclear factor-kappa B (NF-κB) inactivation. Moreover, anti-oxidant enzymes such as superoxide dismutase and glutathione peroxidase-1/2 through nuclear factor-erythroid 2-related factor 2 (Nrf2) pathway significantly increased. Our findings indicate that Chulbu-tang has the potential therapeutic effect on OA through inhibiting the inflammatory responses via inactivating NF-κB signaling pathway. In addition, upregulation of Nrf2 led to anti-oxidant effects. Conclusions Taken together, Chulbu-tang is believed to have antioxidant, anti-inflammatory effects, and cartilage protection for arthritis-causing rats.

• 대한수의학회지
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• 제56권1호
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• pp.15-21
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• 2016

NAD(P)H-quinone oxidoreductase-1 (NQO1) is a down-stream target gene of nuclear factor erythroid 2-related factor 2 (Nrf2), and performs diverse biological functions. Recently, NQO1 is recognized as an effective gene for the cytotoxic inserts with its diverse biological functions, which is focused on antioxidant properties. The aim of present study was to assess the impact of NQO1 knockdown on cytoprotection-related protein expression in cisplatin cytotoxicity by using small interfering (si) RNA targeted on NQO1 gene. Cytotoxicity of cisplatin on ACHN cells was assessed in a dose- and time-dependent manner after siScramble or siNQO1 treatment. After cisplatin treatment, cells were subjected to cell viability assay, western-blot analysis, and immunofluorescence study. The cell viability was decreased in the siNQO1 cells (50%) than the siScramble cells (70%) after 24 h of cisplatin ($20{\mu}M$) treatment. Moreover, cytoprotection-related protein expressions were markedly suppressed in the siNQO1 cells after cisplatin treatment. The expression of Nrf2 and Klotho were decreased by 20% and 40%, respectively, of that in siScramble cells. Nrf2 and Klotho activation were also decreased in cisplatin treated siNQO1 cells, confirmed by cytoplasm-tonuclear translocation. Our findings demonstrate that the increased cisplatin-induced cytotoxicity was accompanied by suppressed Nrf2 activation and Klotho expression in siNQO1 cells.

• Molecules and Cells
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• 제26권2호
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• pp.193-199
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• 2008

The pro-apoptotic Bcl-2 family member Bim acts as a sensor for apoptotic stimuli and initiates apoptosis through the mitochondrial pathway. To identify novel regulators of Bim, we employed the yeast two-hybrid system and isolated the human gene encoding macrophage migration inhibitory factor (MIF), a ubiquitously expressed proinflammatory mediator that has also been implicated in cell proliferation, the cell cycle and carcinogenesis. The interaction between MIF and Bim was confirmed by both in vitro and in vivo protein interaction assays. Intriguingly, protein complexes between MIF and the three major Bim isoforms (BimEL/BimL/BimS) could be detected in HEK293 and K562 cells, especially in cells undergoing apoptosis. Moreover, exogenous expression of MIF partially inhibited Bim-induced apoptosis in HEK293 cells. SiRNA-mediated knockdown of MIF increased apoptosis in K562 cells exposed to the chemical oxidant diamide. Endogenous MIF may regulate the pro-apoptotic activity of Bim and inhibit the release of cytochrome c from mitochondria.

• 대한골관절종양학회지
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• 제20권1호
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• pp.1-6
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• 2014

목적: 골육종에서 Connective Tissue Growth Factor (CTGF)의 발현 정도를 확인하고 발암기전에서의 역할을 살펴보고자 하였다. 대상 및 방법: 환자에서 수립한 23개의 골육종 세포주에서 CTGF의 발현 정도를 관찰하였으며, siRNA를 이용하여 CTGF의 발현을 억제한 후 세포침습과 세포 증식에서 CTGF의 역할을 분석하였다. 결과: 17명(74%)의 세포주에서 control 세포인 정상 골모세포보다 CTGF의 발현이 증가되어 있었다. CTGF의 발암기전에서의 역할을 관찰하기 위해 불멸화된 골육종 세포주 SaOS-2와 MG63에서 siRNA로 CTGF의 발현을 억제한 후 siRNA를 transfection한 세포에서 유의하게 세포침습이 억제되고 세포 증식이 억제되는 것을 관찰하였다. 결론: 골육종 세포주에서 CTGF의 발현이 높았고 세포침습, 세포 성장에 관여하는 바 CTGF가 골육종의 발암기전에서 중요한 역할을 하는 것으로 판단된다.

• BMB Reports
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• 제52권6호
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• pp.391-396
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• 2019

Receptor activator of nuclear factor kappa B ligand (RANKL) expression in osteoblasts is regulated by 1,25-dihydroxyvitamin D3 (1,25D3). CCAAT/enhancer-binding protein beta ($C/EBP{\beta}$) has been proposed to function as a transcription factor and upregulate RANKL expression, but it is still uncertain how $C/EBP{\beta}$ is involved in 1,25D3-induced RANKL expression of osteoblasts. 1,25D3 stimulation increased the expression of RANKL and $C/EBP{\beta}$ genes in osteoblasts and enhanced phosphorylation and stability of these proteins. Moreover, induction of RANKL expression by 1,25D3 in osteoblasts was downregulated upon knockdown of $C/EBP{\beta}$. In contrast, $C/EBP{\beta}$ overexpression directly upregulated RANKL promoter activity and exhibited a synergistic effect on 1,25D3-induced RANKL expression. In particular, 1,25D3 treatment of osteoblasts increased $C/EBP{\beta}$ protein binding to the RANKL promoter. In conclusion, $C/EBP{\beta}$ is required for induction of RANKL by 1,25D3.

• Yonsei Medical Journal
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• 제59권9호
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• pp.1041-1048
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• 2018

Purpose: Heat shock factor 1 (HSF1) is a key regulator of the heat shock response and plays an important role in various cancers. However, the role of HSF1 in gastric cancer is still unknown. The present study evaluated the function of HSF1 and related mechanisms in gastric cancer. Materials and Methods: The expression levels of HSF1 in normal and gastric cancer tissues were compared using cDNA microarray data from the NCBI Gene Expression Omnibus (GEO) dataset. The proliferation of gastric cancer cells was analyzed using the WST assay. Transwell migration and invasion assays were used to evaluate the migration and invasion abilities of gastric cancer cells. Protein levels of HSF1 were analyzed using immunohistochemical staining of tissue microarrays from patients with gastric cancer. Results: HSF1 expression was significantly higher in gastric cancer tissue than in normal tissue. Knockdown of HSF1 reduced the proliferation, migration, and invasion of gastric cancer cells, while HSF1 overexpression promoted proliferation, migration, and invasion of gastric cancer cells. Furthermore, HSF1 promoted the proliferation of gastric cancer cells in vivo. In Kaplan-Meier analysis, high levels of HSF1 were associated with poor prognosis for patients with gastric cancer (p=0.028). Conclusion: HSF1 may be closely associated with the proliferation and motility of gastric cancer cells and poor prognosis of patients with gastric cancer. Accordingly, HSF1 could serve as a prognostic marker for gastric cancer.

• BMB Reports
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• 제53권12호
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• pp.628-633
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• 2020

WNT11 is a member of the non-canonical Wnt family and plays a crucial role in tumor progression. However, the regulatory mechanisms underlying WNT11 expression are unclear. Tumor necrosis factor-alpha (TNFα) is a major inflammatory cytokine produced in the tumor microenvironment and contributes to processes associated with tumor progression, such as tumor invasion and metastasis. By using site-directed mutagenesis and introducing a serial deletion in the 5'-regulatory region of WNT11, we observed that TNFα activates the early growth response 1 (EGR1)-binding sequence (EBS) in the proximal region of WNT11 and that the transcription factor EGR1 is necessary for the TNFα-induced transcription of WNT11. EGR1 bound directly to the EBSs within the proximal 5'-regulatory region of WNT11 and ectopic expression of EGR1 stimulated WNT11 promoter activity, whereas the knockdown of EGR1 expression by RNA interference reduced TNFα-induced WNT11 expression in T47D breast cancer cells. We also observed that mitogen-activated protein kinases (MAPK), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 kinase mediated TNFα-induced transcription of WNT11 via EGR1. Our results suggest that EGR1 directly targets WNT11 in response to TNFα stimulation in breast cancer cells.

• Journal of Microbiology and Biotechnology
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• 제31권1호
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• pp.16-24
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• 2021

Hepatitis B virus (HBV) genome P-encoded protein HBV DNA polymerase (Pol) has long been known as a reverse transcriptase during HBV replication. In this study, we investigated the impact of HBV Pol on host cellular processes, mainly apoptosis, and the underlying mechanisms. We showed a marked reduction in apoptotic rates in the HBV Pol-expressed HepG2 cells compared to controls. Moreover, a series of assays, i.e., yeast two-hybrid, GST pull-down, co-immunoprecipitation, and confocal laser scanning microscopy, identified the host factor eEF1A2 to be associated with HBV Pol. Furthermore, knockdown of eEF1A2 gene by siRNA abrogated the HBV Pol-mediated anti-apoptotic effect with apoptosis induced by endoplasmatic reticulum (ER) stress-inducer thapsigargin (TG), thus suggesting that the host factor eEF1A2 is essential for HBV Pol's anti-apoptosis properties. Our findings have revealed a novel role for HBV Pol in its modulation of apoptosis through integrating with eEF1A2.

• Molecules and Cells
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• 제45권6호
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• pp.413-424
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• 2022

Suppressor of mothers against decapentaplegic homolog (SMAD) 4 is a pluripotent signaling mediator that regulates myriad cellular functions, including cell growth, cell division, angiogenesis, apoptosis, cell invasion, and metastasis, through transforming growth factor β (TGF-β)-dependent and -independent pathways. SMAD4 is a critical modulator in signal transduction and functions primarily as a transcription factor or cofactor. Apart from being a DNA-binding factor, the additional SMAD4 mechanisms in tumor suppression remain elusive. We previously identified methyl malonyl aciduria cobalamin deficiency B type (MMAB) as a critical SMAD4 binding protein using a proto array analysis. This study confirmed the interaction between SMAD4 and MMAB using bimolecular fluorescence complementation (BiFC) assay, proximity ligation assay (PLA), and conventional immunoprecipitation. We found that transient SMAD4 overexpression down-regulates MMAB expression via a proteasome-dependent pathway. SMAD4-MMAB interaction was independent of TGF-β signaling. Finally, we determined the effect of MMAB downregulation on cancer cells. siRNA-mediated knockdown of MMAB affected cancer cell metabolism in HeLa cells by decreasing ATP production and glucose consumption as well as inducing apoptosis. These findings suggest that SMAD4 controls cancer cell metabolism by regulating MMAB.