• Biomolecules & Therapeutics
• /
• v.20 no.3
• /
• pp.299-305
• /
• 2012

Endochondral bone formation is the process by which mesenchymal cells condense to become chondrocytes, which ultimately form new bone. The process of chondrogenic differentiation and hypertrophy is critical for bone formation and as such is regulated by many factors. In this study, we aimed to indentify novel factors that regulate chondrogenesis. We investigated the possible role of isopsoralen in induction of chondrogenic differentiation in clonal mouse chondrogenic ATDC5 cells. Isopsoralen treatment stimulated the accumulation of cartilage nodules in a dose-dependent manner. Further, ATDC5 cells treated with isopsoralen were stained more intensely with Alcian blue than control cells, suggesting that isopsoralen increases the synthesis of matrix proteoglycans. Similarly, isopsoralen markedly induced the activation of alkaline phosphatase activity compared with control cells. Isopsoralen enhanced the expressions of chondrogenic marker genes such as collagen II, collagen X, OCN, Smad4 and Sox9 in a time-dependent manner. Furthermore, isopsoralen induced the activation of extracellular signal-regulated kinase (ERK) and p38 MAP kinase, but not that of c-jun N-terminal kinase (JNK). Isopsoralen significantly enhanced the protein expression of BMP-2 in a time-dependent manner. PD98059 and SB 203580, inhibitors of ERK and p38 MAPK, respectively, decreased the number of stained cells treated with isopsoralen. Taken together, these results suggest that isopsoralen mediates a chondromodulating effect by BMP-2 or MAPK signaling pathways, and is therefore a possible therapeutic agent for bone growth disorders.

• Journal of the Korean Society of Food Culture
• /
• v.38 no.3
• /
• pp.176-183
• /
• 2023

In this study, we investigated the effect of the extracts of Cyrtomium fortunei J.Sm. (CFJ) on lipopolysaccharide (LPS) induced inflammation in mouse BV-2 microglial cells. Nitric oxide (NO) production and cell viability were measured using the Griess reagent and the (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) (MTT) assay. Inflammatory cytokines were detected by quantitative polymerase chain reaction (qPCR) in BV-2 microglial cells with and without CFJ extracts. Subsequently, mitogen-activated protein kinases (MAPKs) and antioxidant markers were assessed by western blot analysis. It was found that the CFJ extract significantly decreased the production of pro-inflammatory cytokines (interleukin [IL]-6, tumor necrosis factor [TNF]-α, and IL-1β) and NO in BV-2 microglial cells that were stimulated with LPS. In addition, the expression levels of the phosphorylation of the MAPK family (p38, c-Jun N-terminal kinases [JNK], and extracellular-signal regulated kinase [ERK]) were reduced by CFJ. Also, treatment with CFJ significantly increased the activities of superoxide dismutase type 1(SOD1) and Catalase in BV-2 microglial cells. Our results indicate that CFJ has a potent suppressive effect on the pro-inflammatory responses of activated BV-2 microglia. Therefore, CFJ has the potential to be an effective treatment for neurodegenerative diseases, as it can inhibit the production of inflammatory mediators in activated BV-2 microglial cells.

• Journal of Life Science
• /
• v.33 no.11
• /
• pp.859-867
• /
• 2023

Lupeol is a type of pentacyclic triterpene that has been reported to have therapeutic effects for treating many diseases; however, its effect on insulin resistance is unclear clear. This study examined the inhibitory effect of lupeol on the serine phosphorylation of insulin receptor substrate-1 in insulin resistance-induced 3T3-L1 adipocytes. 3T3-L1 cells were cultured and treated with tumor necrosis factor-α (TNF-α) for 24 hours to induce insulin resistance. Cells treated with different concentrations of lupeol (15 μM or 30 μM) or 100 nM of rosiglitazone were incubated. Then, lysed cells underwent western blotting. Lupeol exhibited a positive effect on the negative regulator of insulin signaling and inflammation-activated protein kinase caused by TNF-α in adipocytes. Lupeol inhibited the activation of protein tyrosine phosphatase-1B (PTP-1B)-a negative regulator of insulin signaling-and c-Jun N-terminal kinase (JNK); it was also an inhibitor of nuclear factor kappa-B kinase (IKK) and inflammation-activated protein kinases. In addition, Lupeol downregulated serine phosphorylation and upregulated tyrosine phosphorylation in insulin receptor substrate-1. Then, the downregulated phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway was activated, the translocation of glucose transporter type 4 was stimulated to the cell membrane, and intracellular glucose uptake increased in the insulin resistance-induced 3T3-L1 adipocytes. Lupeol may improve TNF-α-induced insulin resistance by downregulating the serine phosphorylation of insulin receptor substrate 1 by inhibiting negative regulators of insulin signaling and inflammation-activated protein kinases in 3T3-L1 adipocytes.

• Journal of Ginseng Research
• /
• v.40 no.4
• /
• pp.423-430
• /
• 2016

Background: The induction of cellular defensive genes such as phase II detoxifying and antioxidant enzymes is a highly effective strategy for protection against carcinogenesis as well as slowing cancer development. Transcription factor Nrf2 (nuclear factor E2-related factor 2) is responsible for activation of phase II enzymes induced by natural chemopreventive compounds. Methods: Red ginseng oil (RGO) was extracted using a supercritical $CO_2$ extraction system and chemical profile of RGO was investigated by GC/MS. Effects of RGO on regulation of the Nrf2/antioxidant response element (ARE) pathway were determined by ARE-luciferase assay, western blotting, and confocal microscopy. Results: The predominant components of RGO were 9,12-octadecadienoic acid (31.48%), bicyclo[10.1.0] tridec-1-ene (22.54%), and 22,23-dihydrostigmasterol (16.90%). RGO treatment significantly increased nuclear translocation of Nrf2 as well as ARE reporter gene activity, leading to upregulation of heme oxygenase-1 and NAD(P)H:quinone oxidoreductase 1. Phosphorylation of the upstream kinases such as apoptosis signal-regulating kinase (ASK)1, mitogen-activated protein kinase (MAPK) kinase (MKK)4/7, c-Jun N-terminal kinase (JNK), and p38 MAPK were enhanced by treatment with RGO. In addition, RGO-mediated Nrf2 expression and nuclear translocation was attenuated by JNK inhibitor SP600125 and p38 MAPK inhibitor SB202190. Conclusion: RGO could be used as a potential chemopreventive agent, possibly by induction of Nrf2/ARE-mediated phase II enzymes via ASK1-MKK4/7-JNK and p38 MAPK signaling pathways.

• The Korea Journal of Herbology
• /
• v.28 no.5
• /
• pp.7-11
• /
• 2013

Objectives : Seungmagalgeun-tang (SMGGT) is traditional medicine widely used for inflammatory disease and flu. But SMGGT exhibits potent anti-inflammatory activity with an unknown mechanism. To elucidate the molecular mechanisms of SMGGT water extract on pharmacological and biochemical actions in inflammation, we examined the effect of SMGGT on pro-inflammatory mediators in Phorbol-12-myristate-13-acetate (PMA)+A23187-stimulated mast cells. Methods : In the present study, pro-inflammatory cytokine production was determined by performing enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), and western blot analysis to measure the activation of MAPKs. Cells were treated with SMGGT 1 h prior to the addition of 50 nM of PMA and $1{\mu}M$ of A23187. Cell viability was measured by MTS assay. The investigation focused on whether SMGGT inhibited the expressions of interleukin-6 (IL-6), interleukin-8 (IL-8) and mitogen-activated protein kinases (MAPKs) in PMA+A23187-stimulated mast cells. Results : SMGGT has no cytotoxicity at examined concentration (100, 250, and $500{\mu}g/ml$). Also, gene expression of IL-6 and IL-8 in HMC-1 cells stimulated by PMA+A23187 was down regulated by SMGGT. Furthermore, SMGGT suppressed the PMA+A23187-induced phosphorylation of extracellular signal-regulated kinase (ERK) and c-jun N-terminal Kinase(JNK). But, SMGGT could not regulate phosphorylation of p38 MAPK. Conclusions : These results suggest that SMGGT has inhibitory effects on PMA+A23187-induced IL-6 and IL-8 production. These inhibitory effects occur through blockades on the phosphorylation of ERK and JNK.

• Journal of Life Science
• /
• v.31 no.11
• /
• pp.987-994
• /
• 2021

Our previous studies have reported that antimicrobial peptides (AMPs) derived from the larvae of white-spotted flower chafer (Protaetia brevitarsis seulensis) exert anti-inflammatory and neuroprotective activities. This study explored the anti-inflammatory effects of protaetiamycine 9 (CVLKKAYFLTNLKLRG-NH₂), a novel AMP, derived from P. b. seulensis against lipopolysaccharide (LPS)-mediated inflammatory response in RAW264.7 macrophage cells. Protaetiamycine 9 (25, 50, 75, and 100 ㎍/ml) did not cause cytotoxic effects against RAW264.7 cells. The RAW264.7 cells were pre-treated with various concentrations of protaetiamycine 9 (25-100 ㎍/ml) for 1 hr and then exposed to LPS (100 ng/ml) for 24 hr. Protaetiamycine 9 treatments decreased the LPS-induced secretion of inflammatory mediators, such as nitric oxide (NO), in a dose-dependent manner. Protaetiamycine 9 (25-100 ㎍/ml) effectively downregulated the LPS-induced increase in mRNA and the protein expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2), which are involved in the production of inflammatory mediators. Protaetiamycine 9 also suppressed the production and gene expression of pro-inflammatory cytokines, including interleukin (IL)-6 and IL-1β, compared to the presence of LPS alone. Furthermore, protaetiamycine 9 inhibited the degradation of inhibitory kappa B alpha (IκB-α) and the phosphorylation of mitogen-activated protein kinases (MAPKs), such as extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38. In conclusion, these results suggest that protaetiamycine 9 exhibits LPS-mediated inflammatory responses by blocking IκB-α degradation and MAPK phosphorylation.

• Journal of Life Science
• /
• v.25 no.9
• /
• pp.984-992
• /
• 2015

Sanguinarine is a benzophenanthridine alkaloid originally isolated from the roots of Sanguinaria canadensis. It has multiple biological activities (e.g., antioxidant and antiproliferative) and immune-enhancing potential. In this study, we explored the proapoptotic properties and modes of action of sanguinarine in human hepatocellular carcinoma HepG2 cells. Our results revealed that sanguinarine inhibited HepG2 cell growth and induced apoptosis in a dose-dependent manner. The induction of apoptosis by sanguinarine was associated with the up-regulation of Fas and Bax, the release of cytochrome c from the mitochondria to the cytosol, and the loss of the mitochondrial membrane potential. In addition, sanguinarine activated caspase-9 and -8, initiator caspases of the intrinsic and death extrinsic pathways, respectively, and caspase-3, accompanied by proteolytic degradation of poly (ADP-ribose) polymerase. Sanguinarine also triggered the generation of reactive oxygen species (ROS). The elimination of ROS by N-acetylcysteine reversed sanguinarine-induced apoptosis. Furthermore, sanguinarine induced the dephosphorylation of Akt and the phosphorylation of mitogen-activated protein kinases, including extracellular signal-regulated kinase (ERK), c-jun N-terminal kinase (JNK), and p38. The growth inhibition was enhanced by the combined treatment of sanguinarine with a phosphatidylinositol 3'-kinase (PI3K) inhibitor and an ERK inhibitor but not JNK and p38 inhibitors. Overall, our data indicate that the proapoptotic effects of sanguinarine in HepG2 cells depend on ROS production and the activation of both intrinsic and extrinsic signaling pathways, which is mediated by blocking PI3K/Akt and activating the ERK pathway. Thus, our data suggest that sanguinarine may be a natural compound with potential for use as an antitumor agent in liver cancer.

• Journal of the Korean Applied Science and Technology
• /
• v.30 no.1
• /
• pp.173-182
• /
• 2013

Atopic dermatitis is a chronic, relapsing inflammatory skin disease associated with dysfunction of skin barrier and cutaneous hyper-reactivity to environmental triggers. In the previous study, cytotoxicity, antioxidant, anti-inflammatory and anti-allergic activities were investigated for various herbal extracts such as Aloe vera L. (AV), Viola mandshurica W. Becker (VM), Punica granatum L. (PG), and Dendrobium nobile L. (DN) in order to develop effective therapeutic herbal extracts for atopic dermatitis, In this study, anti-inflammatory activities of these herb extracts in lipopolysaccharide (LPS)-induced macrophage RAW264.7 cells were further examined to find the underlying molecular mechanisms. The RT-PCR (reverse transcription polymerase chain reaction) analysis showed that PG, DN and AV inhibited effectively the gene expression of pro-inflammatory cytokines IL-6 and IL-$1{\beta}$ in LPS-stimulated macrophages, while VM did not. The transfection and luciferase analysis exhibited that all herbal extracts hindered the activation of transcription nuclear factor kappa B (NF-${\kappa}B$). The western blot analysis indicated that AV blocked the activation of only JNK MAP (c-Jun N-terminal kinase mitogen-activated protein) kinase not p38 MAP kinase, while VM, PG and DN did not show the activation of both JNK and p38 MAP kinases. These results suggest that AV, VM, PG, and DN have anti-inflammatory activities and thus have the potential to reduce and alleviate the symptoms of atopic dermatitis.

• Biomolecules & Therapeutics
• /
• v.25 no.5
• /
• pp.519-527
• /
• 2017

Excessive activation of microglia causes the continuous production of neurotoxic mediators, which further causes neuron degeneration. Therefore, inhibition of microglial activation is a possible target for the treatment of neurodegenerative disorders. Balanophonin, a natural neolignoid from Firmiana simplex, has been reported to have anti-inflammatory and anti-cancer effects. In this study, we aimed to evaluate the anti-neuroinflammatory effects and mechanism of balanophonin in lipopolysaccharide (LPS)-stimulated BV2 microglia cells. BV2 microglia cells were stimulated with LPS in the presence or absence of balanophonin. The results indicated that balanophonin reduced not only the LPS-mediated TLR4 activation but also the production of inflammatory mediators, such as nitric oxide (NO), prostaglandin E2 (PGE2), $Interleukin-1{\beta}$ ($IL-1{\beta}$), and tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$), in BV2 cells. Balanophonin also inhibited LPS-induced inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX2) protein expression and mitogen activated protein kinases (MAPKs), including extracellular signal-regulated kinase (ERK1/2), c-Jun N-terminal kinase (JNK), and p38 MAPK. Interestingly, it also inhibited neuronal cell death resulting from LPS-activated microglia by regulating cleaved caspase-3 and poly ADP ribose polymerase (PARP) cleavage in N2a cells. In conclusion, our data indicated that balanophonin may delay the progression of neuronal cell death by inhibiting microglial activation.

• Journal of Animal Science and Technology
• /
• v.59 no.7
• /
• pp.16.1-16.6
• /
• 2017

Background: This study was performed to investigate the impact of exogenous ghrelin on the pancreatic ${\alpha}$-amylase outputs and responses of pancreatic proteins to ghrelin that may relate to pancreatic exocrine. Methods: Sprague-Dawley male rats (9 weeks old, $300{\pm}10g$) were injected with ghrelin via intraperitoneal (i.p.) infusion at dosage of 0, 0.1, 1.0 and $10.0{\mu}g/kg$ body weight (BW), respectively. The plasma ghrelin and cholecystokinin (CCK) level were determined using enzyme immunoassay kit; the mRNA expression of ghrelin receptor ($GHSR-1{\alpha}$) and growth hormone (GH) receptor were assessed by reverse transcription PCR; the expressions of pancreatic ${\alpha}$-amylase activity, extracellular-signal-regulated kinases (ERK), phosphorylated extracellular-signal-regulated kinases (pERK) and c-Jun N-terminal kinase (JNK) were evaluated by western blotting; moreover the responses of pancreatic proteins to ghrelin were analyzed using the two-dimensional gel electrophoresis system. Results: The exogenous ghrelin (1.0 and $10.0{\mu}g/kg\;BW$) elevated the level of plasma ghrelin (p < 0.05), and suppressed the expression of pancreatic ${\alpha}$-amylase at a dose of $10.0{\mu}g/kg\;BW$ (p < 0.05). No difference in the level of plasma CCK was observed, even though rats were exposed to any dose of exogenous ghrelin. In addition, a combination of western blot and proteomic analysis revealed exogenous ghrelin ($10.0{\mu}g/kg\;BW$) induced increasing the JNK and ERK expressions (p < 0.05) and four proteins such as Destrin, Anionic trypsin-1, Trypsinogen, and especially eukaryotic translation initiation factor 3 in rat pancreas. Conclusions: Taken together, exogenous ghrelin by i.p. infusion plays a role in the pancreatic exocrine secretion via mitogen-activated protein kinase signaling pathway.