• Korean Journal of Plant Resources
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• v.33 no.4
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• pp.271-278
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• 2020

In this study, we evaluated the anti-cancer activity and potential molecular mechanism of 70% ethanol extracts of the Berchemia floribunda (BF) which belongs to Rhamnaceae against human colorectal cancer cells. The treatment of BF decreased the cell proliferation in HCT116 cell and suppressed cellular accumulation of Cyclin D1 protein. Inhibition of proteasomal activity by MG132 attenuated BF-mediated Cyclin D1 downregulation and Cyclin D1 was decreased in the cell treated with BF. These findings indicates that BF-mediated Cyclin D1 downregulation may be result from Cyclin D1 proteasomal degradation. Additionally, BF-mediated Cyclin D1 degradation was blocked in the presence of LiCl, a GSK3β inhibitor, but not PD98059, SP600125, SB203580, Bay11-7082, LY294002 an ERK1/2 inhibitor, JNK inhibitor, p38 inhibitor, IκK inhibitor and PI3K inhibitor. Furthermore, BF phosphorylated Cyclin D1 at threonine-286 (Thr286), and LiCl-induced GSK3β inhibition reduced the BF mediated phosphorylation of Cyclin D1 at Thr286. These results suggested that BF may downregulate Cyclin D1 expression as a potential anti-cancer target through GSK3β dependent Cyclin D1 degradation. Therefore, this study provides that the extract of BF has anticancer activity against human colorectal cancer cells.

• Journal of Physiology & Pathology in Korean Medicine
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• v.29 no.1
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• pp.18-26
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• 2015

Akebiae Caulis is a galenical originated from Akebia quinata Decaisne species. It is commonly used in the treatment of oposiuria, inflammation, nociceptive and fever. Here, we investigated the effect of Akebiae Caulis extract (ACE) to protect hepatocyte against the malfunction of mitochondria and apoptosis. Arachidonic acid (AA)+iron promoted excessive reactive oxygen species (ROS) production and exerted a deleterious effect on mitochondria. Treatment with ACE protected hepatocytes from AA+iron-induced cytotoxicity, as shown by alterations in the protein levels related with apoptosis such as poly(ADP-ribose) polymerase, pro-caspase 3, Bcl-XL and Bcl-2. Moreover, AA+iron-induced $H_2O_2$ production, GSH depletion and mitochondrial dysfunction were alleviated by ACE pretreatment. As a potential molecular mechanism for the ACE-mediated cytoprotection, phosphorylation of AMP-activated protein kinase (AMPK), a key regulator in determining cell survival or death, was increased by ACE. Moreover, ACE treatment enhanced inactive phosphorylation of glycogen synthase kinase-$3{\beta}$ ($GSK3{\beta}$), downstream substrate kinase of AMPK. More importantly, ACE prevented a decrease in the $GSK3{\beta}$ phosphorylation derived by AA+iron, which might contribute to mitohondiral protection and cell survival. To further identify essential compounds in Akebiae Caulis for the protection of AA+iron-mediated cytotoxicity, we found that betulin in combination with hederagenin protected from AA+iron-induced mitochondrial dysfunction. Betulin+hederagenin treatment also increased inactive phosphorylation of $GSK3{\beta}$ in common with ACE. These results suggest that ACE protected hepatocytes against oxidative stress and mitochondrial dysfunction, which is mediated with inactive $GSK3{\beta}$ phosphorylation downstream of AMPK.

• Molecules and Cells
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• v.41 no.3
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• pp.234-243
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• 2018

In recent years, the interest towards the relationship between incretins and bone has been increasing. Previous studies have suggested that glucagon-like peptide-1 (GLP-1) and its receptor agonists exert beneficial anabolic influence on skeletal metabolism, such as promoting proliferation and differentiation of osteoblasts via entero-osseous-axis. However, little is known regarding the effects of GLP-1 on osteoblast apoptosis and the underlying mechanisms involved. Thus, in the present study, we investigated the effects of liraglutide, a glucagon-like peptide-1 receptor agonist, on apoptosis of murine MC3T3-E1 osteoblastic cells. We confirmed the presence of GLP-1 receptor (GLP-1R) in MC3T3-E1 cells. Our data demonstrated that liraglutide inhibited the apoptosis of osteoblastic MC3T3-E1 cells induced by serum deprivation, as detected by Annexin V/PI and Hoechst 33258 staining and ELISA assays. Moreover, liraglutide upregulated Bcl-2 expression and downregulated Bax expression and caspase-3 activity at intermediate concentration (100 nM) for maximum effect. Further study suggested that liraglutide stimulated the phosphorylation of AKT and enhanced cAMP level, along with decreased phosphorylation of $GSK3{\beta}$, increased ${\beta}-catenin$ phosphorylation at Ser675 site and upregulated nuclear ${\beta}-catenin$ content and transcriptional activity. Pretreatment of cells with the PI3K inhibitor LY294002, PKA inhibitor H89, and siRNAs GLP-1R, ${\beta}-catenin$ abrogated the liraglutide-induced activation of cAMP, AKT, ${\beta}-catenin$, respectively. In conclusion, these findings illustrate that activation of GLP-1 receptor by liraglutide inhibits the apoptosis of osteoblastic MC3T3-E1 cells induced by serum deprivation through $cAMP/PKA/{\beta}-catenin$ and $PI3K/Akt/GSK3{\beta}$ signaling pathways.

• International Journal of Stem Cells
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• v.16 no.1
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• pp.52-65
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• 2023

Background and Objectives: Epithelial-Mesenchymal transition (EMT) is one of the origins of myofibroblasts in renal interstitial fibrosis. Mesenchymal stem cells (MSCs) alleviating EMT has been proved, but the concrete mechanism is unclear. To explore the mechanism, serum-free MSCs conditioned medium (SF-MSCs-CM) was used to treat rat renal tubular epithelial cells (NRK-52E) fibrosis induced by transforming growth factor-β1 (TGF-β1) which ameliorated EMT. Methods and Results: Galectin-3 knockdown (Gal-3 KD) and overexpression (Gal-3 OE) lentiviral vectors were established and transfected into NRK-52E. NRK-52E fibrosis model was induced by TGF-β1 and treated with the SF-MSCs-CM for 24 h after modelling. Fibrosis and autophagy related indexes were detected by western blot and immunocytochemistry. In model group, the expressions of α-smooth muscle actin (α-SMA), fibronectin (FN), Galectin-3, Snail, Kim-1, and the ratios of P-Akt/Akt, P-GSK3β/GSK3β, P-PI3K/PI3K, P-mTOR/mTOR, TIMP1/MMP9, and LC3B-II/I were obviously increased, and E-Cadherin (E-cad) and P62 decreased significantly compared with control group. SF-MSCs-CM showed an opposite trend after treatment compared with model group. Whether in Gal-3 KD or Gal-3 OE NRK-52E cells, SF-MSCs-CM also showed similar trends. However, the effects of anti-fibrosis and enhanced autophagy in Gal-3 KD cells were more obvious than those in Gal-3 OE cells. Conclusions: SF-MSCs-CM probably alleviated the EMT via inhibiting Galectin-3/Akt/GSK3β/Snail pathway. Meanwhile, Gal-3 KD possibly enhanced autophagy via inhibiting Galectin-3/Akt/mTOR pathway, which synergistically ameliorated renal fibrosis. Targeting galectin-3 may be a potential target for the treatment of renal fibrosis.

• Journal of Microbiology and Biotechnology
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• v.20 no.11
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• pp.1513-1520
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• 2010

A bacterium capable of producing melanin pigment in the presence of L-tyrosine was isolated from a crop field soil sample and identified as Klebsiella sp. GSK based on morphological, biochemical, and 16S rDNA sequencing. The polymerization of this pigment occurs outside the cell wall, which has a granular structure as melanin ghosts. Chemical characterization of the pigment particles showed then to be acid resistant, alkali soluble, and insoluble in most of the organic solvents and water. The pigment got bleached when subjected to the action of oxidants as well as reductants. This pigment was precipitated with $FeCl_3$, ammoniacal silver nitrate, and potassium ferricynide. The pigment showed high absorbance in the UV region and decreased absorbance when shifted towards the visible region. The melanin pigment was further charecterized by FT-IR and EPR spectroscopies. A key enzyme, 4-hydroxyphenylacetic acid hydroxylase, that catalyzes the formation of melanin pigment by hydroxylation of L-tyrosine was detected in this bacterium. Inhibition studies with specific inhibitors, kojic acid and KCN, proved that melanin is synthesized by the DOPA-melanin pathway.

• BMB Reports
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• v.42 no.2
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• pp.119-124
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• 2009

PI(3,4,5)$P_3$ produced by the activated PI3-kinase is a key lipid second messenger in cell signaling downstream of insulin. Skeletal muscle and kidney-enriched inositol phosphatase (SKIP) identified as a 5'-inositol phosphatase that hydrolyzes PI(3,4,5) $P_3$ to PI(3,4)$P_2$, negatively regulates the insulin-induced glycogen synthesis in skeletal muscle. However the mechanism by which this occurs remains unclear. To elucidate the function of SKIP in glycogen synthesis, we employed RNAi techniques to knockdown the SKIP gene in differentiating C2C12 myoblasts. Insulininduced phosphorylation of Akt (protein kinase B) and GSK-3$\beta$ (Glycogen synthase kinase), subsequent dephosphorylation of glycogen synthase and glycogen synthesis were increased by inhibiting the expression of SKIP, whereas the insulin-induced glycogen synthesis was decreased by overexpression of WT-SKIP. Our results suggest that SKIP plays a negative regulatory role in Akt/ GSK-3$\beta$/GS (glycogen synthase) pathway leading to glycogen synthesis in myocytes.

• The Korea Journal of Herbology
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• v.39 no.1
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• pp.23-29
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• 2024

Objectives : Polyporus umbellatus is a medicinal mushroom that has been used for over thousands years in Chinese medicine as a powerful diuretic to relieve fluid retention and edema. Dermal papilla is located at the bottom of the hair follicle and connected to the blood vessels where it gets the nutrients and oxygen to nurture hair follicle. This study examined the mechanism through which the ethanol extract of Polyporus umbellatus (EPU) promoted the proliferation of human dermal papilla cells (HHDPCs). Methods : To estimate the proliferative effects of EPU on HHDPCs, cell viability was estimated by thiazolyl blue tetrazolium bromide (MTT) assay. Western blotting was used to investgate the activation of ERK, phosphoinositide 3-kinase (PI3K)/Akt, β-catenin, GSK-3β and heme oxygenase-1 (HO-1). Cells were treated with inhibitors of ERK and Akt prior to EPU treatment. Results : EPU promoted the proliferation of HHDPCs and the phosphorylation of ERK and Akt in dose dependent manner. However, the proliferative effect of EPU on HHDPCs was inhibited by pre-treatment of ERK inhibitor (PD98059) and Akt inhibitor (LY294002). Furthermore, EPU respectively stimulated the protein expression of β-catenin and phosphorylated GSK-3β. EPU significantly increased the protein expression levels of proliferation and cytoprotection related genes such as Bcl-2, SIRT-1, and HO-1 in cells. Conclusion : This results suggest that EPU promoted the proliferation of HHDPCs via activating PI3K/Akt and Wnt/β-catenin signaling pathway in HHDPCs.

• Nutrition Research and Practice
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• v.12 no.1
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• pp.3-12
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• 2018

BACKGROUND/OBJECTIVES: Sageretia thea is traditionally used as a medicinal herb to treat various diseases, including skin disorders, in China and Korea. This study evaluated the inhibitory effect of Sageretia thea fruit on melanogenesis and its underlying mechanisms in B16F10 mouse melanoma cells. The active chemical compounds in anti-melanogenesis were determined in Sageretia thea. MATERIALS/METHODS: Solvent fractions from the crude extract were investigated for anti-melanogenic activities. These activities and the mechanism of anti-melanogenesis in B16F10 cells were examined by determining melanin content and tyrosinase activity, and by performing western blotting. RESULTS: The n-hexane fraction of Sageretia thea fruit (HFSF) exhibited significant anti-melanogenic activity among the various solvent fractions without reducing viability of B16F10 cells. The HFSF suppressed the expression of tyrosinase and tyrosinase-related protein 1 (TRP1). The reduction of microphthalmia-associated transcription factor (MITF) expression by the HFSF was mediated by the Akt/glycogen synthase kinase 3 beta ($GSK3{\beta}$) signaling pathway, which promotes the reduction of ${\beta}-catenin$. Treatment with the $GSK3{\beta}$ inhibitor 6-bromoindirubin-3'-oxime (BIO) restored HFSF-induced inhibition of MITF expression. The HFSF bioactive constituents responsible for anti-melanogenic activity were identified by bioassay-guided fractionation and gas chromatography-mass spectrometry analysis as methyl linoleate and methyl linolenate. CONCLUSIONS: These results indicate that HFSF and its constituents, methyl linoleate and methyl linolenate, could be used as whitening agents in cosmetics and have potential for treating hyperpigmentation disorders in the clinic.

• Korean Journal of Pharmacognosy
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• v.35 no.3 s.138
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• pp.244-249
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• 2004

The effects of hesperetin and naringenin on $NF-{\kappa}B$ activation were investigated in normal rat kidney cells transformed by temperature sensitive Rous Sarcoma Virus (tsNRK). The flavonoids, naringenin and hesperetin, significantly reduced v-Src-induced $NF-{\kappa}B$ activation as well as phosphorylation of Akt and GSK-3 in tsNRK cells, whereas these compounds did not effect on platelet-derived growth factor (PDGF)-induced $NF-{\kappa}B$ activation in $NIH3T3{\gamma}l$ cells. In addition, the DNA binding activity of SP-I was also reduced but that of AP-1 was not affected by the compounds. Our study suggests that Src-induced $NF-{\kappa}B$ activation could occur via Akt-GSK-3 pathway without $IkB{\alpha}$ degradation and that naringenin and hesperetin could be used in the treatment of cancer through the inhibition of $NF-{\kappa}B$ activation.

• Tuberculosis and Respiratory Diseases
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• v.72 no.4
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• pp.343-351
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• 2012

Background: The phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling axis has emerged as a novel target for cancer therapy. Agents that inhibit this pathway are currently under development for lung cancer treatment. In the present study, we have tested whether dual inhibition of PI3K/Akt/mTOR signaling can lead to enahnced antitumor effects. We have also examined the role of autophagy during this process. Methods: We analyzed the combination effect of the mTOR inhibitor, temsirolimus, and the Akt inhibitor, GSK690693, on the survival of NCI-H460 and A549 non-small cell lung cancer cells. Cell proliferation was determined by MTT assay and apoptosis induction was evaluated by flow cytometry and terminal deoxynucleotidyl transferase dUTP nick end labeling assay. Autophagy induction was also evaluated by acridine orange staining. Changes of apoptosis or autophagy-related proteins were evaluated by western blot analysis. Results: Combination treatment with temsirolimus and GSK690693 caused synergistically increased cell death in NCI-H460 and A549 cells. This was attributable to increased induction of apoptosis. Caspase 3 activation and poly(ADP-ribose) polymerase cleavage accompanied these findings. Autophagy also increased and inhibition of autophagy resulted in increased cell death, suggesting its cytoprotective role during this process. Conclusion: Taken together, our results suggest that the combination of temsirolimus and GSK690693 could be a novel strategy for lung cancer therapy. Inhibition of autophagy could also be a promising method of enhancing the combination effect of these drugs.