• The Korean Journal of Physiology and Pharmacology
• /
• v.2 no.3
• /
• pp.353-359
• /
• 1998

In gastrointestinal smooth muscle, muscarinic stimulation by carbachol (CCh) activates nonselective cation channel current ($I_{CCh}$) which is facilitated by intracellular [$Ca^{2+}$] increase. Caffeine is widely used in experiments to mobilize $Ca^{2+}$ from intracellular stores. This study shows a strong inhibitory effect of caffeine on $I_{CCh}$ in guinea-pig gastric myocyte. In this study, the underlying mechanism of the inhibitory effect of caffeine was investigated. $I_{CCh}$ was completely suppressed by the addition of caffeine (10 mM) to the superfusing solution. Inhibition of $I_{CCh}$ by caffeine was not related to the intracellular cAMP accumulation which was expected from the phosphodiesterase-inhibiting effect of caffeine. The blockade of $InsP_3-induced$ $Ca^{2+}$ release by heparin had no significant effects on the activation of $I_{CCh}$. When the same cationic current had been induced by intracellular dialysis of $GTP[{\gamma}S]$ in order to bypass the muscarinic receptor, the inhibitory effect of caffeine was significantly attenuated. The results of this study indicate that both intracellular signalling pathways for $I_{CCh}$, proximal and distal to G-protein activation, are suppressed by caffeine. A major inhibition was observed at the proximal level.

• Journal of Life Science
• /
• v.32 no.10
• /
• pp.762-770
• /
• 2022

Hyperglycemia in type 2 diabetes can be alleviated by promoting cellular glucose uptake. Betulinic acid (3β,-3-hydroxy-lup-20(29)-en-28-oic acid) is a pentacyclic lupane-type triterpenoid compound. Although there have been studies on the antidiabetic activity of betulinic acid, studies on cellular glucose uptake are lacking. We investigated the effects of betulinic acid on glucose uptake and its mechanism of action in 3T3-L1 adipocytes. Betulinic acid significantly stimulated glucose uptake in 3T3-L1 adipocytes by increasing the phosphorylation of the insulin receptor substrate 1-tyrosine (IRS-1tyr) in the insulin signaling pathway, which in turn stimulated the activation of phosphoinositide 3-kinase (PI3K) and the phosphorylation of protein kinase B (Akt). The activation of PI3K and Akt by betulinic acid translocated glucose transporter 4 to the plasma membrane (PM-GLUT4), thereby increasing the expression of PM-GLUT4 and thus stimulating cellular glucose uptake. Betulinic acid also significantly increased the phosphorylation/activation of AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase. The activation of PI3K and AMPK by betulinic acid was confirmed using the PI3K inhibitor wortmannin and the AMPK inhibitor compound C. The increase in glucose uptake induced by betulinic acid was significantly decreased by wortmannin and compound C in the 3T3-L1 adipocytes. These results suggest that betulinic acid stimulates glucose uptake by activating PI3K and AMPK in 3T3-L1 adipocytes.

• The Korea Journal of Herbology
• /
• v.28 no.1
• /
• pp.23-31
• /
• 2013

Objectives : The purpose of this study was to determine the anti-obesity effect and molecular mechanism of YY312, a herbal extract composed of Imperatae Rhizoma, Citri Unshius Pericarpium Immaturus, and Evodiae Fructus, on a high-fat diet-induced animal model and on 3T3-L1 cells. Methods : C57BL/6 mice were fed for 6 weeks with a normal diet or a high-fat diet (HFD). Then they orally administered daily with 300 mg/kg YY312 for next 10 weeks. Body weight and food consumption were recorded weekly and daily, respectively. Tissue weights, serum lipid, and glucose levels were analyzed at the end of the study. Additionally, the effects of YY312 on adipocyte differentiation in 3T3-L1 cells were examined. After differentiating 3T3-L1 cells were treated with YY312, Oil-red O staining, RT-PCR, and Western blotting were performed for lipid accumulation, mRNA expression of adipogenesis gene, and AMP-activated protein kinase (AMPK) phosphorylation, respectively. Results : YY312-administered mice showed a significant reduction of body weights and abdominal adipose tissue weights. YY312 also reduced the serum levels of triglycerides and total cholesterol, compared with the HFD group. Treatment with YY312 inhibited lipid accumulation and blocked expression of adipogenic transcription factors and lipogenesis genes, including peroxisome proliferator-activated receptor ${\gamma}$, CCAT/enhancer binding protein ${\alpha}$ and fatty acid synthase. YY312 increased AMPK phosphorylation in 3T3-L1 adipocytes. Conclusions : This study showed that herbal extract YY312 has an anti-obesity effect in vitro and in vivo. Thus, YY312 could be developed as a supplement for reduction of body weight gain induced by an HFD.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.24 no.2
• /
• pp.228-235
• /
• 2010

This study was performed to investigate the memory enhancing effect of Poria cocos Wolf (Hoelen cum radix) against scopolamine-induced amnesia in Sprague-Dawley (SD) rats. To induce amnesia, scopolamine (0.75 mg/kg) was intraperitonically injected into SD rats 30 min before starting behavior tests. We have conducted Morris water-maze and Y-maze tests to monitor learning and memory functions. Poria cocos effectively reversed scopolamine-induced memory impairment in SD rats which was represented by an improvement of mean escape latency in water-maze test and spontaneous alterations in Y-maze test. To elucidate possible molecule mechanism, we have measured mRNA as well as protein expression of acetylcholine esterase (AchE), choline acetyltransferase (ChAT), muscarinic acetylcholine receptor (mAchR), and brain-derived neurotrophic factor (BDNF) using RT-PCR and Western blot analysis, respectively. Poria cocos increased mRNA levels of ChAT and mAchR in rat hippocampus compared with those in the scopolamine-injected amnesic group. In addition, protein expression of ChAT and BDNF was also elevated by Poria cocos intake. Furthermore, as an upstrem regulator, the activation of cAMP response element-binding protein (CREB) was assessed by immunohistochemistry. In this immunohistochemical analysis, the phosphorylation of CREB (p-CREB) was reduced by scopolamine injection, which was restored back to control levels by administration of Poria cocos. These results suggest that Poria cocos may improve memory and cognitive deficit in amnesia and have therapeutic potentials through up-regulation of ChAT, mAchR, and BDNF, which seemed to be mediated by activation of CREB.

• Journal of Life Science
• /
• v.31 no.8
• /
• pp.729-735
• /
• 2021

Type 2 diabetes occurs when there is an abnormality in the tissue's ability to absorb glucose. Glucose uptake and metabolism by insulin are the basic mechanisms that maintain blood sugar. Glucose uptake goes through various signaling steps initiated by the binding of insulin to receptors on the cell surface. In line with the foregoing, the purpose of this study was to investigate the effect of 2,7-phloroglucinol-6,6-bieckol (PHB), an active compound isolated from Ecklonia cava, on glucose uptake in 3T3-L1 adipocytes. Notably, PHB increased glucose uptake in a dose-dependent manner owing to the enhanced glucose transporter type 4 (GLUT4) expression in the plasma membrane of 3T3-L1 adipocytes. These effects of PHB were attributed to the phosphorylation of insulin receptor substrate-1 and protein kinase B (PKB or AKT), as well as to the phosphoinositide 3-kinase (PI3K) activation in the insulin signaling pathway. PHB also stimulated 5' AMP-activated protein kinase (AMPK) phosphorylation and activation. The phosphorylation and activation of the PI3K/AKT and AMPK pathways by PHB were identified using wortmannin (a PI3K inhibitor) and compound C (an AMPK inhibitor). In this study, we showed that PHB can increase glucose uptake in 3T3-L1 adipocytes by promoting GLUT4 translocation to the plasma membrane via the PI3K and AMPK pathways. The results indicate that PHB may help improve insulin sensitivity.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.27 no.6
• /
• pp.802-808
• /
• 2013

Here we tried to uncover the potential anti-lipogenic effect and the underlying mechanism of Phaseolus angularis seed in a cellular model of nonalcoholic fatty liver disease (NAFLD) induced in HepG2 cells. Ethanol extract of Phaseolus angularis seed (JSD) was prepared. HepG2 cells were incubated in palmitate containing media to induce intracellular lipid accumulation, and co-treated with JSD for 16 hrs before examine intracellular lipid content. In control group, the cells were not co-treated with JSD. We measured the effects of JSD on liver X receptor ${\alpha}$ ($LXR{\alpha}$) and sterol regulatory element-binding transcription factor-1c (SREBP-1c) expression, transcription level of lipogenic genes, including acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), stearoyl-CoA desaturase-1 (SCD-1), and AMP-activated protein kinase (AMPK) activation in HepG2 cells. JSD markedly reduced palmitate-induced intracellular lipid accumulation in HepG2 cells. JSD suppressed $LXR{\alpha}$/SREBP-1c expression, and SREBP-1c mediated induction of ACC, FAS, and SCD-1. Furthermore, JSD activated AMPK, which plays a major role in the control of hepatic lipid metabolism. Taken together, it is suggested that JSD has a potential to alleviate hepatic steatosis, at least in part, by suppressing $LXR{\alpha}$/SREBP-1c mediated induction of lipogenic genes. In addtion, the anti-lipogenic potential may be associated with activation of AMPK. Therefore, the Phaseolus angularis seed could be applied as a potential therapeutics for NAFLD with additional clinical studies.

• Proceedings of the Plant Resources Society of Korea Conference
• /
• 2023.04a
• /
• pp.50-50
• /
• 2023

Prostaglandin E2(PGE2), a major product of cyclooxygenase-2 (COX-2), plays an important role in the carcinogenesis of many solid tumors, including colorectal cancer. Because PGE2 functions by signaling through PGE2 receptors (Eps), which regulate tumor cell growth, invasion, and migration, there has been a growing amount of interest in the therapeutic potential of targeting Eps. In the present study, we investigated the role of EP4 on the effectiveness of cordycepin in inhibititing the migration and invasion of HCT116 human colorectal carcinoma cells. Our data indicate that cordycepin suppressed lipopolysaccharide (LPS)-enhanced cell migration and invasion through the inactivation of matrix metalloproteinases (MMP)-9 as well as the down-regulation of COX-2 expression and PGE2 production. These events were shown to be associated with the inactivation of EP4 and activation of AMP-activated protein kinase (AMPK). Moreover, the AMPK inhibitor, compound C, as well as AMPK knockdown via siRNA, attenuated the cordycepin-induced inhibition of EP4 expression. Cordycepin treatment also reduced the activation of CREB. These findings indicate that cordycepin suppresses the migration and invasion of HCT116 cells. Through modulating EP4 expression and the AMPK-CREB signaling pathway. Therefore, cordycepin has the potential to serve as a potent anti-cancer agent in therapeutic strategies against colorectal cancer metastasis.

• Journal of Life Science
• /
• v.27 no.2
• /
• pp.155-163
• /
• 2017

Mori Folium, the leaf of Morus alba, is a traditional medicinal herb that shows various pharmacological activities such as antiinflammatory, antidiabetic, antimelanogenesis, antioxidant, antibacterial, antiallergic, and immunomodulatory activities. However, the mechanisms of their inhibitory effects on adipocyte differentiation and adipogenesis remain poorly understood. In the present study, we investigated the inhibition of adipocyte differentiation and adipogenesis by ethanol extracts of Mori Folium (EEMF) in 3T3-L1 preadipocytes. Treatment with EEMF suppressed the terminal differentiation of 3T3-L1 preadipocytes in a dose-dependent manner, as confirmed by a decrease in the lipid droplet number and lipid content through Oil Red O staining. EEMF significantly reduced the accumulation of cellular triglyceride, which is associated with a significant inhibition of pro-adipogenic transcription factors, including sterol regulatory element-binding protein-1c (SREBP-1c), peroxisome proliferator-activated receptor-${\gamma}$ ($PPAR{\gamma}$), and CCAAT/enhancer-binding proteins ${\alpha}$ ($C/EBP{\alpha}$) and ${\beta}$ ($C/EBP{\beta}$). In addition, EEMF potentially downregulated the expression of adipocyte-specific genes, including adipocyte fatty acid binding protein (aP2) and leptin. Furthermore, EEMF treatment effectively increased the phosphorylation of the AMP-activated protein kinase (AMPK) and acetyl CoA carboxylase (ACC); however, treatment with a potent inhibitor of AMPK, compound C, significantly restored the EEMF-induced inhibition of pro-adipogenic transcription factors and adipocyte-specific genes. These results together indicate that EEMF has preeminent effects on the inhibition of adipogenesis through the AMPK signaling pathway, and further studies will be needed to identify the active compounds in Mori Folium.

• Biomolecules & Therapeutics
• /
• v.21 no.4
• /
• pp.299-306
• /
• 2013

In the present study, we investigated the effect of ethanolic extract of the seed of Zizyphus jujuba var. spinosa (EEZS) on cholinergic blockade-induced memory impairment in mice. Male ICR mice were treated with EEZS. The behavioral tests were conducted using the passive avoidance, the Y-maze, and the Morris water maze tasks. EEZS (100 or 200 mg/kg, p.o.) significantly ameliorated the scopolamine-induced cognitive impairment in our present behavioral tasks without changes of locomotor activity. The ameliorating effect of EEZS on scopolamine-induced memory impairment was significantly reversed by a sub-effective dose of MK-801 (0.0125 mg/kg, s.c.). In addition, single administration of EEZS in normal naive mouse enhanced latency time in the passive avoidance task. Western blot analysis was employed to confirm the mechanism of memory-ameliorating effect of EEZS. Administration of EEZS (200 mg/kg) increased the level of memory-related signaling molecules, including phosphorylation of extracellular signal-regulated kinase or cAMP response element-binding protein in the hippocampal region. Also, the time-dependent expression level of brain-derived neurotrophic factor by the administration of EEZS was markedly increased from 3 to 9 h. These results suggest that EEZS has memory-ameliorating effect on scopolamine-induced cognitive impairment, which is mediated by the enhancement of the cholinergic neurotransmitter system, in part, via NMDA receptor signaling, and that EEZS would be useful agent against cognitive dysfunction such as Alzheimer's disease.

• Reproductive and Developmental Biology
• /
• v.34 no.1
• /
• pp.33-40
• /
• 2010

Equine chorionic gonadotropin (eCG) is a heavily glycosylated glycoprotein composed of non-covalently linked $\alpha$- and $\beta$-subunits. To study the function and signal transduction of tethered recombinant-eCG (rec-eCG), a single chain eCG molecule was constructed, and the rec-eCG protein was prepared. In this study, we constructed 5 mutants (${\Delta}1$, ${\Delta}2$, ${\Delta}3$, ${\Delta}4$, and ${\Delta}5$) of rec-eCG using data about known glycoprotein hormones to analyze the role of specific follicle stimulating homone (FSH)-like activity. Three amino acids of certain specific sites were replaced with alanine. The expression vectors were transfected into CHO cells and subjected to G418 selection for 2~3 weeks. The media were collected and the quantity of secreted tethered rec-eCGs was quantified by ELISA. The LH- and FSH-like activities were assayed in terms of cAMP production by rat LH/CG and rat FSH receptors. Then, the metabolic clearance rate analyzed by the injection of rec-eCG (5 IU) into the tail vein was analyzed. The mutant eCGs (${\Delta}l$, ${\Delta}4$, and ${\Delta}5$) were transcripted, but not translated into proteins. Rec-eCG A2 was secreted in much lower amounts than the wild type. Only the rec-eCG ${\Delta}3$ ($\beta$-subunit: $Gln^{94}-Ile^{95}-Lys^{96}{\rightarrow}Ala^{94}-Ala^{95}-Ala^{96}$) was efficiently secreted. Although activity is low, its LH-like activity was similar to that of tethered $eCG{\beta\alpha}$. However, the FSH-like activity of rec-$eCG{\beta\alpha\Delta}3$ was completely flat. The result of the analysis of the metabolic clearance rate shoed the persistence of the mutant in the blood until 4 hours after the injection. After then, it almost disappeared at 8 hours. Taken together, these data suggest that 94~96 amino acid sequences in eCG $\beta$-subunit appear to be of utmost importance for signal transduction of the FSH receptor.