• Molecules and Cells
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• v.37 no.9
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• pp.685-690
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• 2014

Osteoclasts are large polykaryons that have the unique capacity to degrade bone and are generated by the differentiation of myeloid lineage progenitors. To identify the genes involved in osteoclast development, we performed microarray analysis, and we found that carboxypeptidase E (CPE), a prohormone processing enzyme, was highly upregulated in osteoclasts compared with their precursors, bone marrow-derived macrophages (BMMs). Here, we demonstrate a novel role for CPE in receptor activator of NF-${\kappa}B$ ligand (RANKL)-induced osteoclast differentiation. The overexpression of CPE in BMMs increases the formation of tartrate-resistant acid phosphatase (TRAP)-positive multinuclear osteoclasts and the expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which are key regulators in osteoclastogenesis. Furthermore, employing CPE knockout mice, we show that CPE deficiency attenuates osteoclast formation. Together, our data suggest that CPE might be an important modulator of RANKL-induced osteoclast differentiation.

• Journal of Microbiology and Biotechnology
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• v.26 no.6
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• pp.1063-1066
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• 2016

Longan (Dimocarpus longan Lour.) has been used as a traditional oriental medicine and possesses a number of physiological activities. In this study, we used cell-based herbal extract screening to identify longan fruit extract (LFE) as an activator of osteoblast differentiation. LFE up-regulated alkaline phosphatase (ALP) activity, induced mineralization, and activated Runx2 gene expression in MC3T3-E1 cells. Furthermore, treatment of MC3T3-E1 cells with LFE promoted the phosphorylation of extracellular signal-regulated kinase1/2 (Erk1/2); however, abrogation of Erk1/2 activation with PD98059 resulted in down-regulation of the phospho-SMAD1/5/8 and Runx2 levels, which in turn reduced the ALP activity. Our findings suggest that LFE exerts its osteogenic activity through activation of the ERK signaling pathway and may have potential as an herbal therapeutic or a preventive agent for the treatment of osteoporosis.

• The Journal of Korean Obstetrics and Gynecology
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• v.15 no.4
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• pp.61-75
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• 2002

Recombinant human $interleukin-1{\beta}$ $(rhIL-1{\beta})$ regulates several activities of the osteoblast cells derived from mouse calvarial bone explants in vitro. $rhIL-1{\beta}$ stimulated cellular proliferation and the synthesis of prostaglandin $E_2(PGE_2)$ and plasminogen activator activity in the cultured cells in a dose-dependent manner. However, the induction of osteocalcin synthesis and alkaine phosphatase activity in response to vitamine D, two characteristics of the osteoblast phenotype, were antagonized by $rhIL-1{\beta}$ over a similar dose range. This study supports the role of $IL-1{\beta}$ in the pathological modulation of bone cell metabolism, with regard to implication in the pathogenesis of osteoporosis by $IL-1{\beta}$. When the mouse calvarial bone cells were used, the bone resorption induced by $IL-1{\beta}$ was strongly inhibited by calcitonin treatment, indicating osteoclast-mediated bone resorption. On the other hand, the medicinal extracts of Taeyoungjon-Jahage (T.Y.J-J.H.G extracts) was tested for whether they could inhibit $IL-1{\beta}-induced$ $PGE_2$ production. Cell viability was not significantly affected by treatment with the indicated concentration of the extracts. The T.Y.J.-J.H.G. extracts were shown to have the inhibitory effects against the synthesis of $PGE_2$. We also examined the effect of the pretreatment with a various concentrations of the T.Y.J.-J.H.G. extracts then treated the $PGE_2-induction$ agents. Pretreatment of the T.Y.J.-J.H.G. extracts for 1 h, which by itself had little effect on cell survival, did not enhance the synthesis of $PGE_2$. Furthermore, the T.Y.J-J.H.G. extracts were shown to have the protective effects against plasminogen dependent fibrinolysis induced by the bone resorption agents of $IL-1{\beta}$. Pretreatment of the T.Y.J.-J.H.G. extracts for 1 h did not enhance the plasminogen dependent fibrinolysis. Finally, calcitonin showed the inhibitory activity the $IL-1{\beta}-stimulated$ bone resorption in the mouse calvarial bone cells having both of the osteoblast and osteoclast cells. Seemingly, pretreatment of the T.Y.J.-J.H.G. extracts for 1 h reduced the bone resorption. These results clearly indicated that calcitonin and T.Y.J.-J.H.G. extracts play key roles in inhibition of the osteoclast-mediated bone resorption.

• Reproductive and Developmental Biology
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• v.33 no.2
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• pp.113-117
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• 2009

Animals produced by somatic cell nuclear transfer (SCNT) using genetically modified cells are almost always transgenic, implying that this method is more efficient than the traditional pronuclear microinjection method. Most somatic cells for SCNT in animals are fetus-derived primary cells and successful gene integration in somatic cells will depend on transfection condition. The objective of this study is to evaluate the efficiency of electroporation (Microporator) and liposome reagents (F-6, F-HD, W-EX, W-Q, W-M) for tissue-type plasminogen activator (tPA) gene transfection and to estimate the overall efficiency of transfection of Korean native pig fetal fibroblast cells (KNPFF). Electroporation showed significantly higher transfection efficiency than liposome reagents with regard to the transfection of in vitro cultures in the early stages of development (41.7% with Microporator vs. 18.3% with F-6, 20.0% with F-HD 18.5% with W-EX, 5.0% with W-M and 6.3% W-Q,). Colonies identified as tPA-positives were treated once more with G418 for 10 to 14 days and growing colonies were selected again. When the cells of newly selected colonies were subjected to single-cell PCR, reselection of colonies following second round of G418 selection increased the rate of transgene integration per each colony. These results suggest that transfection with electroporation is the most efficient and the second rounds of G418 selection may be an effective method for transfection of porcine fetal fibroblast cells.

• Journal of Physiology & Pathology in Korean Medicine
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• v.38 no.1
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• pp.32-37
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• 2024

The aim of this study was to investigate the role of Daesiho-tang (Da Chai Hu-Tang) water extract (DSTE) in regulating chronic stress-induced cancer progression, focusing on its activity in modulating tumor-associated macrophages (TAMs). Different stimuli can polarize TAMs into immune-stimulating M1 macrophages or immunosuppressive M2 macrophages. During cancer progression, M2 phenotype increases and supports tumor growth, angiogenesis and metastasis. Notably, chronic stress-induced catecholamines promote M2 macrophage polarization. In this study, we investigated whether DSTE regulates norepinephrine (NE)-induced M2 macrophage polarization in RAW 264.7 mouse macrophage cells. Even though NE itself did not increase the expression of M2 markers, the conditioned media of NE-treated 4T1 mouse breast cancer cells (NE CM) significantly up-regulated M2 markers in RAW 264.7 cells, suggesting that NE-regulated cancer cell secretome stimulated M2 polarization. However, such increase was abrogated by DSTE. NE CM also induced phosphorylation of signal transducer and activator of transcription 6 (STAT6) in RAW 264.7 cells, which was clearly reversed by pretreatment with DSTE, demonstrating that DSTE inhibited M2 polarization by inactivating STAT6. Finally, M2-polarized RAW264.7 cells by NE CM markedly increased the migration of 4T1 cells. However, such increase was completely reversed by co-treating RAW264.7 cells with NE CM and DSTE, indicating that DSTE attenuated cancer cell migration by blocking M2 polarization. Taken together, our results suggest a probable use of DSTE for cancer patients under chronic stress by regulating M2 macrophage polarization.

• Biomolecules & Therapeutics
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• v.16 no.2
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• pp.100-105
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• 2008

The sphingolipid metabolites act as lipid mediator for cell proliferation and apoptosis in mammalian cells. In bacteria, sphingolipid metabolism remains unknown. The purpose of this study was to investigate whether sphingolipid metabolism is potential target for fumonisin $B_1$($FB_1$) and desipramine in Sphingomonas chungbukensis, Gram-negative bacteria, by comparing the intracellular contents of bacterial sphingolipids with ones of HIT-T15 ${\beta}$-cells, hamster pancreatic cells. The concentrations of ceramide and dihydroceramide were 18.0 ${\pm}$ 12.0 and 0.025 ${\pm}$ 0.018 nmol/mg protein, respectively, in HIT-T15 cells. However, the concentrations of ceramide and dihydroceramide in the bacterial culture were 2.0 ${\pm}$ 1.2 and 10.6 ${\pm}$ 5.5 nmol/mg protein, respectively. $FB_1$ decreased the level of ceramide from 18.0 to 3.8 nmol/mg protein in HIT-T15 ${\beta}$-cells. However, dihydroceramide content in $FB_1$-treated HIT-T15 cells was slightly decreased compared with the control culture. When S. chungbukensis was treated with either $FB_1$ or desipramine, dihydroceramide level was increased by 5- and 4-fold, respectively, compared with the control bacteria. These results indicate that $FB_1$ and desipramine may act as an activator in bacterial sphingolipid biosynthetic pathway, and bacterial sphingolipid metabolism pathway appears to be different from the pathway of mammalian cells.

• Molecules and Cells
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• v.37 no.10
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• pp.734-741
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• 2014

Histone modifications on major transcription factor target genes are one of the major regulatory mechanisms controlling adipogenesis. Plant homeodomain finger 2 (PHF2) is a Jumonji domain-containing protein and is known to demethylate the histone H3K9, a repressive gene marker. To better understand the function of PHF2 in adipocyte differentiation, we constructed stable PHF2 knock-down cells by using the mouse pre-adipocyte cell line 3T3-L1. When induced with adipogenic media, PHF2 knock-down cells showed reduced lipid accumulation compared to control cells. Differential expression using a cDNA microarray revealed significant reduction of metabolic pathway genes in the PHF2 knock-down cell line after differentiation. The reduced expression of major transcription factors and adipokines was confirmed with reverse transcription- quantitative polymerase chain reaction and Western blotting. We further performed co-immunoprecipitation analysis of PHF2 with four major adipogenic transcription factors, and we found that CCATT/enhancer binding protein (C/EBP)${\alpha}$ and C/EBP${\delta}$ physically interact with PHF2. In addition, PHF2 binding to target gene promoters was confirmed with a chromatin immunoprecipitation experiment. Finally, histone H3K9 methylation markers on the PHF2-binding sequences were increased in PHF2 knock-down cells after differentiation. Together, these results demonstrate that PHF2 histone demethylase controls adipogenic gene expression during differentiation.

• BMB Reports
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• v.54 no.5
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• pp.266-271
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• 2021

Estrogen-related receptor γ (ERRγ), a member of the orphan nuclear receptor family, is a key mediator in cellular metabolic processes and energy homeostasis. Therefore, ERRγ has become an attractive target for treating diverse metabolic disorders. We recently reported that ERRγ acts as a negative regulator of osteoclastogenesis induced by receptor activator of nuclear factor-κB ligand (RANKL). In the present study, we explored the effects of an ERRγ-specific modulator, GSK5182, on ERRγ-regulated osteoclast differentiation and survival. Interestingly, GSK5182 increased ERRγ protein levels much as does GSK4716, which is an ERRγ agonist. GSK5182 inhibited osteoclast generation from bone-marrow-derived macrophages without affecting cytotoxicity. GSK5182 also attenuated RANKL-mediated expression of cFos and nuclear factor of activated T-cells cytoplasmic 1 (NFATc1), pivotal transcription factors for osteoclastogenesis. Arrested osteoclast differentiation was associated with reduced RANK expression, but not with the M-CSF receptor, c-Fms. GSK5182 strongly blocked the phosphorylation of IκBα, c-Jun N-terminal kinase, and extracellular signal-regulated kinase in response to RANKL. GSK5182 also suppressed NF-κB promoter activity in a dose-dependent manner. In addition to osteoclastogenesis, GSK5182 accelerated osteoclast apoptosis by caspase-3 activation. Together, these results suggest that GSK5182, a synthetic ERRγ modulator, may have potential in treating disorders related to bone resorption.

• Toxicological Research
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• v.24 no.1
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• pp.17-22
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• 2008

Sopungsan (SS) is a traditional Korean decoction used for the treatment of dermatitis. The aim of this study is to confirm whether or not SS has a preventive effect on the development of atopic dermatitis in dinitrochlorobenzene-applied Nc/Nga mice. SS was administered orally to Nc/Nga mice, which led to the remarkable suppression of the development of dermatitis, as determined by a histological examination and the serum IgE levels. Moreover, SS inhibited the production of thymus- and activation-regulated chemokine (TARC) and its mRNA expression in a keratinocyte cell line, HaCaT, which had been stimulated with tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) and interferon-${\gamma}$ (IFN-${\gamma}$). Activation of the nuclear factor-${\kappa}B$ (NF-${\kappa}B$) or activator protein-1 (AP-1) is one of key steps in the signaling pathways mediating induction of TARC. In this study, SS selectively suppressed NF-${\kappa}B$ activation which may be essential for TARC expression in $TNF-{\alpha}/IFN-{\gamma}$ treated keratinocytes. The inhibitory effect of SS on NF-${\kappa}B$ activation and TARC production might be associated with the anti-dermatitic effects of SS.

• Reproductive and Developmental Biology
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• v.38 no.2
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• pp.63-70
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• 2014

Plasminogen activators (PAs) are serine proteases that convert plasminogen to plasmin. Two type of PAs are urokinase-type PA (uPA) and tissue-type PA (tPA). Plasminogen is present in most extracellular fluids. PAs play in various reproductive processes including implantation, ovulation and fertilization. In the spermatozoa, PAs and PAIs play a role in sperm motility and fertilization. PAs in the sertoli cell are stimulated spermatozoa maturation and sperm activation through the phospholipase A2. The oocyte maturation is the process for fertilization and implantation. PAs in cumulus-oocyte complexes (COCs) are related to oocyte maturation by protein kinase A and C. In the ovulatory process, PAs activity are changed and it are related to reducing the tensile strength of ovarian follicle wall. The uterine environment is important for reproduction and the uterus undergo tissue remodeling. In the uterus and oviduct of mammals, expression and activity of PAs are changed during estrous cycle. Thus, expression and activity of PAs are concerned to many reproductive functions. Therefore, PAs seem to important factor of regulator in reproductive events.