• Journal of Ginseng Research
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• v.41 no.1
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• pp.96-102
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• 2017

Background: Korean ginseng, Panax ginseng Meyer, has been used as a traditional oriental medicine to treat illness and promote health for several thousand years. Ginsenosides are the main constituents for the pharmacological effects of P. ginseng. Since several ginsenosides, including ginsenoside (G)-Rg3 and G-Rp1, have reported antiplatelet activity, here we investigate the ability of G-Rp4 to modulate adenosine diphosphate (ADP)-induced platelet aggregation. The ginsenoside Rp4, a similar chemical structure of G-Rp1, was prepared from G-Rg1 by chemical modification. Methods: To examine the effects of G-Rp4 on platelet activation, we performed several experiments, including antiplatelet ability, the modulation of intracellular calcium concentration, and P-selectin expression. In addition, we examined the activation of integrin ${\alpha}IIb{\beta}_3$ and the phosphorylation of signaling molecules using fibrinogen binding assay and immunoblotting in rat washed platelets. Results: G-Rp4 inhibited ADP-induced platelet aggregation in a dose-dependent manner. We found that G-Rp4 decreased calcium mobilization and P-selectin expression in ADP-activated platelets. Moreover, fibrinogen binding to integrin ${\alpha}IIb{\beta}_3$ by ADP was attenuated in G-Rp4-treated platelets. G-Rp4 significantly attenuated phosphorylation of extracellular signal-regulated protein kinases 1 and 2, p38, and c-Jun N-terminal kinase, as well as protein kinase B, phosphatidylinositol 3-kinase, and phospholipase C-${\gamma}$ phosphorylations. Conclusion: G-Rp4 significantly inhibited ADP-induced platelet aggregation and this is mediated via modulating the intracellular signaling molecules. These results indicate that G-Rp4 could be a potential candidate as a therapeutic agent against platelet-related cardiovascular diseases.

• Biomolecules & Therapeutics
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• v.7 no.3
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• pp.227-233
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• 1999

The effects of 2-bromo-3-(3,5-tert-butyl-4-hydroxylphenyl)-1,4-naphthalenedione(TPN2), a synthetic vitamin K derivative, on platelet aggregation and its action mechanisms were investigated in rat platelet. TPN2 inhibited the platelet aggregation induced by collagen($10\mu\textrm{g}$/ml), thrombin(0.1 U/ml), A23187($10\mu\textrm{M}$) and arachidonic acid($100\mu\textrm{M}$) in concentration-dependent manner with $IC_{50}$ values of 6.5$\pm$1.3, 59.3$\pm$4.5, 13.0$\pm$2.37 and 2.9$\pm$$1.0\mu\textrm{M}$, respectively. Collagen-induced serotonin release was significantly reduced by TPN2. The elevation of intracellular free $Ca^{2+}$ concentration ([$Ca^{2+}$]i) by collagen stimulation was greatly decreased by the pretreatment of TPN2, which was due to the inhibition of calcium release from intracellular store and influx from outside of the cell. TPN2 also significantly reduced the thromboxane $A_2$($TXA_2$) formation in a concentration-dependent manner. The collagen-induced arachidonic acid (AA) release in [$^3H$]-AA incorporated platelet, an indicative of the phospholipase $A_2$ activity, was decreased by TPN2 pretreatment. TPN2 significantly inhibited the activity of thromboxane synthase, but did not affect the cyclooxygenase activity. From these results. it is suggested that TPN2 exert its antiplatelet activity through the inhibition of the intra-cellular $Ca^{2+}$ mobilization and the decrease of the $TXA_2$ synthesis.

• Journal of Microbiology and Biotechnology
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• v.34 no.2
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• pp.379-386
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• 2024

Basophils and mast cells are specialized effector cells in allergic reactions. Haliotis discus hannai (abalone), is valuable seafood. Abalone male viscera, which has a brownish color and has not been previously reported to show anti-allergic activities, was extracted with acetone. Six different acetone/hexane fractions (0, 10, 20, 30, 40, and 100%) were obtained using a silica column via β-hexosaminidase release inhibitory activity-guided selection in phorbol myristate acetate and a calcium ionophore, A23187 (PMACI)-induced human basophils, KU812F cells. The 40% acetone/hexane fraction (A40) exhibited the strongest inhibition of PMACI-induced-β-hexosaminidase release. This fraction dose-dependently inhibited reactive oxygen species (ROS) production and calcium mobilization without cytotoxicity. Western blot analysis revealed that A40 down-regulated PMACI-induced MAPK (ERK 1/2, p-38, and JNK) phosphorylation, and the NF-κB translocation from the cytosol to membrane. Moreover, A40 inhibited PMACI-induced interleukin (IL)-1β, IL-6, and IL-8 production. Anti-allergic activities of A40 were confirmed based on inhibitory effects on IL-4 and tumor necrosis factor alpha (TNF-α) production in compound (com) 48/80-induced rat basophilic leukemia (RBL)-2H3 cells. A40 inhibited β-hexosaminidase release and cytokine production such as IL-4 and TNF-α produced by com 48/80-stimulated RBL-2H3 cells. Furthermore, it's fraction attenuated the IgE/DNP-induced passive cutaneous anaphylaxis (PCA) reaction in the ears of BALB/c mice. Our results suggest that abalone contains the active fraction, A40 is a potent therapeutic and functional material to treat allergic diseases.

• The Journal of Korean Medicine
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• v.30 no.6
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• pp.9-16
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• 2009

Objectives: To clarify the effects of Jungri-tang Gamibang on accelerating small intestinal movement induced by the stimulation of cholinergic neurotransmission. Methods: 500, 250 and 125mg Jungri-Tang Gamibang or 20mg domperidone were dissolved or suspended in distilled water and orally pretreated on the carbachol-accelerated small intestinal transit mice once a day for 7 days at a volume of 10ml/kg (of body weight) using a Zonde needle attached to 1 ml syringes containing test drugs. Result: Significantly (p<0.01) increase of % regions of activated charcoal transit in the small intestine was detected in carbachol control compared to that of intact control. However, significant (p<0.01) decreases of % regions of activated charcoal transit were dose-dependently observed in all Jungri-Tang Gamibang extracts or domperidone-pretreated groups. Conclusions: it was concluded that Jungri-tang Gamibang enhancement in the normal intestinal motility and normalization in the accelerated intestinal motility might interfere with a variety of muscarinic, adrenergic and histaminic receptor activities or with the mobilization of calcium ions required for smooth muscle contraction non-specifically.

• Biomolecules & Therapeutics
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• v.19 no.1
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• pp.1-8
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• 2011

G-protein coupled receptors (GPCRs) constitute an important class of drug targets and are involved in every aspect of human physiology including sleep regulation, blood pressure, mood, food intake, perception of pain, control of cancer growth, and immune response. Radiometric assays have been the classic method used during the search for potential therapeutics acting at various GPCRs for most GPCR-based drug discovery research programs. An increasing number of diverse small molecules, together with novel GPCR targets identified from genomics efforts, necessitates the use of high-throughput assays with a good sensitivity and specificity. Currently, a wide array of high-throughput tools for research on GPCRs is available and can be used to study receptor-ligand interaction, receptor driven functional response, receptor-receptor interaction,and receptor internalization. Many of the assay technologies are based on luminescence or fluorescence and can be easily applied in cell based models to reduce gaps between in vitro and in vivo studies for drug discovery processes. Especially, cell based models for GPCR can be efficiently employed to deconvolute the integrated information concerning the ligand-receptor-function axis obtained from label-free detection technology. This review covers various platform technologies used for the research of GPCRs, concentrating on the principal, non-radiometric homogeneous assay technologies. As current technology is rapidly advancing, the combination of probe chemistry, optical instruments, and GPCR biology will provide us with many new technologies to apply in the future.

• Journal of Applied Biological Chemistry
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• v.62 no.4
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• pp.425-431
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• 2019

Platelet activation is essential for hemostatic process on blood vessel damage. However, excessive platelet activation can cause some cardiovascular diseases including atherosclerosis, thrombosis, and myocardial infarction. Scoparone is commonly encountered in the roots of genus Artemisia or Scopolia, and has been studied for its potential pharmacological properties including immunosuppression and vasorelaxation, but antiplatelet effects of scoparone have not been reported yet. We investigated the effect of scoparone on human platelet activation prompted by an analogue of thromboxane A₂, U46619. As the results, scoparone dose-dependently increased cyclic adenosine monophosphate (cAMP) levels as well as cyclic guanosine monophosphate (cGMP) levels, both being aggregation-inhibiting molecules. In addition, scoparone strongly phosphorylated inositol 1, 4, 5-triphosphate receptor (IP3R) and vasodilator-stimulated phosphoprotein (VASP), substrates of cAMP dependent kinase and cGMP dependent kinase. Phosphorylation of IP₃R by scoparone resulted in inhibition of Ca²⁺ mobilization in calcium channels in a dense tubular system, and phosphorylation of VASP by scoparone led to an inability of fibrinogen being able to bind to αIIb/β3. Finally, scoparone inhibited thrombin-induced fibrin clotting, thereby reducing thrombus formation. Therefore, we suggest that scoparone has a strong antiplatelet effect and is highly probable to prevent platelet-derived vascular disease.

• The Korean Journal of Pharmacology
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• v.29 no.1
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• pp.107-120
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• 1993

Particulate or soluble stimuli appear to stimulate phagocytic cell's response by the change of $Ca^{2+}$ mobilization and by the activation of protein kinase C. In contrast, it is reported that activation of protein kinase C could attenuate agonist-stimulated elevation of $Ca^{2+}i$ in neutrophils. PAF elicited an increase of $Ca^{2+}i$ in peritoneal macrophages in a dose dependent fashion and $Ca^{2+}$ extrusion was accompanied. PAF-induced elevation of $Ca^{2+}i$ was not affected by TMB-8, verapamil and TTX. TEA stimulated PAF-induced mobilization of $Ca^{2+}i$ and delayed lowering of $Ca^{2+}i$. Five mM EGTA almost completely inhibited PAF-induced mobilization of $Ca^{2+}i$. After the addition of PAF, membrane permeability was markedly increased up to 5 min and then slowly increased. PAF-induced LDH release was slightly decreased by EGTA plus TMB-8. PAF-stimulated superoxide generation was inhibited by EGTA, TMB-8 and verapamil but not affected by TTX and TEA. PAF-induced elevation of $Ca^{2+}i$, increased membrane permeability and superoxide generation were inhibited by IQSP, chlorpromazine and propranolol. PAF-induced LDH release was significantly inhibited by chlorpromazine and minimally decreased by propranolol. After the pretreatment with PMA, the stimulatory effect of PAF on the elevation of $Ca^{2+}i$ and LDH release in macrophages was significantly decreased. These results suggest that PAF may exert the stimulatory action on peritoneal macrophages of mouse by the elevation of $Ca^{2+}i$ and by the activation of protein kinase C. Preactivation of protein kinase C appears to attenuate the stimulatory action of PAF on macrophage response.

• The Korean Journal of Pharmacology
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• v.26 no.1
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• pp.55-66
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• 1990

In both resting and opsonized zymosan activated neutrophils, ATP stimulated superoxide generation, whereas adenosine inhibited it slightly. The superoxide generation in activated neutrophils to ATP was greater than that of resting neutrophils. In $Ca^{++}$ free medium, inhibitory effect of adenosine on superoxide generation was detectable, whereas ATP did not have any effect. The stimulatory effect of ATP on superoxide generation was inhibited by adenosine in a dose dependent manner. Neither ATP nor adenosine had any effect on NADPH oxidase acitivity. Effects of ATP or adenosine on superoxide generation were more prominent than that by other triphosphate nucleotides or nucleosides. ATP and ADP further stimulated $Ca^{++}$ uptake and increased cytosolic free $Ca^{++}$ level in neutrophils activated by opsonized zymosan, but adenosine inhibited a $Ca^{++}$ mobilization. Verapamil effectively and tetrodotoxin slightly inhibited an increase of cytosolic free $Ca^{++}$ level induced by ATP. Inhibitory effect of either verapamil or tetrodotoxin on superoxide generation in the ATP plus opsonized zymosan-activated neutrophils was greater than in the cells activated by opsonized zymosan alone. Tetraethylammonium chloride had no apparent effect on superoxide generation. CCCP, 2,4-dinitrophenol, diphenylhydantoin and procaine all inhibited superoxide generation in neutrophils activated by opsonized zymosan. Among these, CCCP only inhibited a stimulatory effect of ATP. ATP further stimulated a loss of sulfhydryl groups in activated neutrophils, whereas adenosine had no effect on it. These results suggest that functional responses of neutrophils may be regulated at least partly by purines. ATP and adenosine may further after functional responses of activated neutrophils through their effect on $Ca^{++}$ uptake, membrane phosphorylation and oxidation of soluble sulfhydryl groups.

• Journal of Ginseng Research
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• v.41 no.4
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• pp.548-555
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• 2017

Background: Korean Red Ginseng has been used for several decades to treat many diseases, enhancing both immunity and physical strength. Previous studies have documented the therapeutic effects of ginseng, including its anticancer, antiaging, and anti-inflammatory activities. These activities are mediated by ginsenosides present in the ginseng plant. Ginsenoside Rg3, an effective compound from red ginseng, has been shown to have antiplatelet activity in addition to its anticancer and anti-inflammatory activities. Platelets are important for both primary hemostasis and the repair of the vessels after injury; however, they also play a crucial role in the development of acute coronary diseases. We prepared ginsenoside Rg3-enriched red ginseng extract (Rg3-RGE) to examine its role in platelet physiology. Methods: To examine the effect of Rg3-RGE on platelet activation in vitro, platelet aggregation, granule secretion, intracellular calcium ($[Ca^{2+}]_i$) mobilization, flow cytometry, and immunoblot analysis were carried out using rat platelets. To examine the effect of Rg3-RGE on platelet activation in vivo, a collagen plus epinephrine-induced acute pulmonary thromboembolism mouse model was used. Results: We found that Rg3-RGE significantly inhibited collagen-induced platelet aggregation and $[Ca^{2+}]_i$ mobilization in a dose-dependent manner in addition to reducing ATP release from collagen-stimulated platelets. Furthermore, using immunoblot analysis, we found that Rg3-RGE markedly suppressed mitogen-activated protein kinase phosphorylation (i.e., extracellular stimuli-responsive kinase, Jun N-terminal kinase, p38) as well as the PI3K (phosphatidylinositol 3 kinase)/Akt pathway. Moreover, Rg3-RGE effectively reduced collagen plus epinephrine-induced mortality in mice. Conclusion: These data suggest that ginsenoside Rg3-RGE could be potentially be used as an antiplatelet therapeutic agent against platelet-mediated cardiovascular disorders.

• The Korean Journal of Physiology and Pharmacology
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• v.14 no.2
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• pp.105-111
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• 2010

Inositol 1,4,5-trisphosphate receptors ($InsP_3Rs$) modulate $Ca^{2+}$ release from intracellular $Ca^{2+}$ store and are extensively expressed in the membrane of endoplasmic/sarcoplasmic reticulum and Golgi. Although caffeine and 2-aminoethoxydiphenyl borate (2-APB) have been widely used to block $InsP_3Rs$, the use of these is limited due to their multiple actions. In the present study, we examined and compared the ability of caffeine and 2-APB as a blocker of $Ca^{2+}$ release from intracellular $Ca^{2+}$ stores and $Ca^{2+}$ entry through store-operated $Ca^{2+}$ (SOC) channel in the mouse pancreatic acinar cell. Caffeine did not block the $Ca^{2+}$ entry, but significantly inhibited carbamylcholine (CCh)-induced $Ca^{2+}$ release. In contrast, 2-APB did not block CCh-induced $Ca^{2+}$ release, but remarkably blocked SOC-mediated $Ca^{2+}$ entry at lower concentrations. In permeabilized acinar cell, caffeine had an inhibitory effect on InsP3-induced $Ca^{2+}$ release, but 2-APB at lower concentration, which effectively blocked $Ca^{2+}$ entry, had no inhibitory action. At higher concentrations, 2-APB has multiple paradoxical effects including inhibition of Ins$P_3$-induced $Ca^{2+}$ release and direct stimulation of $Ca^{2+}$ release. Based on the results, we concluded that caffeine is useful as an inhibitor of $InsP_3R$, and 2-APB at lower concentration is considered a blocker of $Ca^{2+}$ entry through SOC channels in the pancreatic acinar cell.