• Journal of Microbiology and Biotechnology
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• v.30 no.5
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• pp.649-661
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• 2020

This study examined the laxative effects of hot-water extracts of Hovenia dulcis Thunb. (HD), Phyllostachys pubescens Mazel (PM), and a 2:8 mixture of both (HP) in two chronic constipation models. For the loperamide-induced constipation model, animals were divided into an untreated group, negative control group (loperamide 4 mg/kg), positive control group (bisacodyl 4 mg/kg) group, and six treatment groups (HP 100 or 400, HD 50 or 100, and PM 100 or 400 mg/kg). For the low-fiber diet-induced constipation model, animals were divided into an untreated group (normal diet), negative control group (low-fiber diet), positive control group (Agio granule, 620 mg/kg), and the same treatment groups. Fecal number, weight, fecal water content, and intestinal transit ratio were higher in the groups treated with HP, HD, and PM than in the groups treated with loperamide or low-fiber diet. Thickness of colon mucosa and muscle layers were increased in the treated groups. Colon tension increased in the HP groups, and [Ca²⁺]i measurements using fura-2 as an indicator showed that HP inhibits ATP-mediated Ca²⁺ influx in IEC-18 cells. These results showed that the HP mixture has laxative activity by increased mucin secretion and inducing contractile activity and relaxation. It may be a useful therapeutic strategy for ameliorating in chronic constipation.

• The Korean Journal of Pain
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• v.29 no.4
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• pp.229-238
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• 2016

Background: The goal of this in vitro study was to investigate the effect of lipid emulsion on vasodilation caused by toxic doses of bupivacaine and mepivacaine during contraction induced by a protein kinase C (PKC) activator, phorbol 12,13-dibutyrate (PDBu), in an isolated endothelium-denuded rat aorta. Methods: The effects of lipid emulsion on the dose-response curves induced by bupivacaine or mepivacaine in an isolated aorta precontracted with PDBu were assessed. In addition, the effects of bupivacaine on the increased intracellular calcium concentration ($[Ca^{2+}]_i$) and contraction induced by PDBu were investigated using fura-2 loaded aortic strips. Further, the effects of bupivacaine, the PKC inhibitor GF109203X and lipid emulsion, alone or in combination, on PDBu-induced PKC and phosphorylation-dependent inhibitory protein of myosin phosphatase (CPI-17) phosphorylation in rat aortic vascular smooth muscle cells (VSMCs) was examined by western blotting. Results: Lipid emulsion attenuated the vasodilation induced by bupivacaine, whereas it had no effect on that induced by mepivacaine. Lipid emulsion had no effect on PDBu-induced contraction. The magnitude of bupivacaine-induced vasodilation was higher than that of the bupivacaine-induced decrease in $[Ca^{2+}]_i$. PDBu promoted PKC and CPI-17 phosphorylation in aortic VSMCs. Bupivacaine and GF109203X attenuated PDBu-induced PKC and CPI-17 phosphorylation, whereas lipid emulsion attenuated bupivacaine-mediated inhibition of PDBu-induced PKC and CPI-17 phosphorylation. Conclusions: These results suggest that lipid emulsion attenuates the vasodilation induced by a toxic dose of bupivacaine via inhibition of bupivacaine-induced PKC and CPI-17 dephosphorylation. This lipid emulsion-mediated inhibition of vasodilation may be partly associated with the lipid solubility of local anesthetics.

• The Korean Journal of Pain
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• v.27 no.3
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• pp.229-238
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• 2014

Background: A toxic dose of bupivacaine produces vasodilation in isolated aortas. The goal of this in vitro study was to investigate the cellular mechanism associated with bupivacaine-induced vasodilation in isolated endothelium-denuded rat aortas precontracted with phenylephrine. Methods: Isolated endothelium-denuded rat aortas were suspended for isometric tension recordings. The effects of nifedipine, verapamil, iberiotoxin, 4-aminopyridine, barium chloride, and glibenclamide on bupivacaine concentration-response curves were assessed in endothelium-denuded aortas precontracted with phenylephrine. The effect of phenylephrine and KCl used for precontraction on bupivacaine-induced concentration-response curves was assessed. The effects of verapamil on phenylephrine concentration-response curves were assessed. The effects of bupivacaine on the intracellular calcium concentration ($[Ca^{2+}]_i$) and tension in aortas precontracted with phenylephrine were measured simultaneously with the acetoxymethyl ester of a fura-2-loaded aortic strip. Results: Pretreatment with potassium channel inhibitors had no effect on bupivacaine-induced relaxation in the endothelium-denuded aortas precontracted with phenylephrine, whereas verapamil or nifedipine attenuated bupivacaine-induced relaxation. The magnitude of the bupivacaine-induced relaxation was enhanced in the 100mM KCl-induced precontracted aortas compared with the phenylephrine-induced precontracted aortas. Verapamil attenuated the phenylephrine-induced contraction. The magnitude of the bupivacaine-induced relaxation was higher than that of the bupivacaine-induced $[Ca^{2+}]_i$ decrease in the aortas precontracted with phenylephrine. Conclusions: Taken together, these results suggest that toxic-dose bupivacaine-induced vasodilation appears to be mediated by decreased calcium sensitization in endothelium-denuded aortas precontracted with phenylephrine. In addition, potassium channel inhibitors had no effect on bupivacaine-induced relaxation. Toxic-dose bupivacaine-induced vasodilation may be partially associated with the inhibitory effect of voltage-operated calcium channels.

• Journal of Ginseng Research
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• v.32 no.3
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• pp.244-249
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• 2008

Vascular inflammation is an important step in the development of cardiovascular disorder. Since it has not been known whether Korean red ginseng has a role to play on the vascular inflammation, we investigated the effects of Korean red ginseng extract (KRGE) on monocyte adhesion and its underlying signaling mechanism. Monocyte adhesion assay and Western blot were conducted on the human umbilical vein endothelial cells to study monocyte adhesion and the expression of adhesion molecules. Intracellular calcium was measured with Fura-2 fluorescent staining, and superoxide production was measured with lucigenin chemiluminescence in the endothelial cells. KRGE inhibits tumor necrosis factor (TNF)-alpha-induced monocyte adhesion on the endothelial cells at the range of $0.03{\sim}1$ mg/ml. TNF-alpha-induced vascular cell adhesion molecule-1 and intercellular cell adhesion molecule-1 expression were inhibited by the pretreatment of KRGE in the endothelial cells. KRGE also inhibits TNF-alpha-induced intracellular calcium and the superoxide production in the endothelial cells. This study first demonstrated that KRGE inhibits TNF-alpha-induced monocyte adhesion by inhibiting the adhesion molecule expression, intracellular calcium and superoxide production in the endothelial cells. Therefore, the anti-inflammatory function of KRGE may be contributed to protecting the endothelial dysfunction in the vascular inflammatory disorders.

• The Korean Journal of Pharmacology
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• v.32 no.1
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• pp.39-49
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• 1996

In the present study, we observed change in intracellular $Ca^{2+}$$([Ca^{2+}]_i)$ as measured with the fluorescent $Ca^{2+}-indicator$ fura-2 in association with force development of the rat basilar arteries during activation by$K^+$ depolarizing solution and U46619, a thromboxane analogue, in the absence and the presence of calcitonin-gent related peptide (CGRP). CGRP (30 and 100 nM) caused a concentration-dependent inhibition of U46619-induced contraction with decrease in $[Ca^{2+}]_i$, whereas it did not exert any effect on the $K^+$ (90 mM)-induced contraction and increase in $[Ca^{2+}]_i$, Further, $[Ca^{2+}]_i-force$ relationships were determined by plotting the ratio of $F_{340}/F_{380}$ $([Ca^{2+}]_i)$ as a function of the force induced by U46619, and the results were compared with those obtained in the presence of CGRP. The curves obtained in the presence of CGRP (30 and 100 nM) were significantly moved to downward without right shift of the curves suggesting that CGRP inhibited the U46619-induced contraction only by mediation of reduction in $[Ca^{2+}]_i$ with out any change in the sensitivity of contractile apparatus to $Ca^{2+}$. The CGRP-induced attenuation of $[Ca^{2+}]_i$ and force development was significantly inhibited under pretreatment with CGRP $(8{\sim}37)$ fragment (100 nM), a CGRP1 receptor antagonist. Both the reduced contraction and reduction in $[Ca^{2+}]_i$ caused by CGRP were fully reversed by pretreatment with charybdotoxin (100 nM) and iberiotoxin (100 nM), large conductance $Ca^{2+}-activated$ $K^+$ channel blockers, but not by apamin (300 nM), a small conductance $Ca^{2+}-activated$ $K^+$ channel blocker, and glibenclamide ( 1 ${\mu}M$), an ATP-sensitive $K^+$ channel blocker. In conclusion, it is suggested that the CGRP1 receptor, upon activation by CGRP, are coupled to opening of $Ca^{2+}-activated$ $K^+$ channel and cause to decrease in $[Ca^{2+}]_i$, thereby leading to vasodilation of the rat basilar artery. However, it is not defined that the mechanism underlying vasodilation whether the $K^+$ channel blockers, charybdotoxin and iberiotoxin directly block the CGRP receptors and that CGRP-evoked relaxation is dependent on the cyclic AMP or $K^+$ channel opening or both actions.

• Journal of Chest Surgery
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• v.43 no.4
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• pp.345-355
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• 2010

Background: Extracellular and intracellular pH ($pH_o$ and $pH_i$), which can be changed in various pathological conditions such as hypoxia, affects vascular contractility. To elucidate the mechanism to alter vascular contractility by pH, the effects of pH on reactivity to vasocontracting agents, intracellular $Ca^{2+}$ influx, and $Ca^{2+}$ sensitivity in vascular smooth muscle were examined. Material and Method: Isometric contractions in rat superior mesenteric arteries (SMA) were observed. Intracellular $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) was recorded by microfluorometer using Fura-2/acetoxylmethyl ester in muscle cells. $pH_o$ was increased from 7.4 to 7.8 or decreased to 6.9 or 6.4. $pH_i$ was decreased by applying $NH_4^+$ or propionic acid or modulated by changing $pH_o$ after increasing membrane permeability using $\beta$-escin. Result: Decreases in $pH_o$ from 7.4 to 6.9 or 6.4 shifted concentration-response curve by norepinephrine (NE) or serotonin (SE) to the right and significantly increased half maximal effective concentration (EC50) to NE or SE. Increase in $pH_o$ from 7.4 to 7.8 shifted concentration-response curve by norepinephrine (NE) or serotonin (SE) to the left and significantly reduced EC50 to NE or SE. NE increased $[Ca^{2+}]_i$ in cultured smooth muscle cells from SMA and the increased $[Ca^{2+}]_i$ was reduced by decreases in $pH_o$. NE-induced contraction was inhibited by $NH_4^+$, whereas the resting tension was increased by $NH_4^+$ or propionic acid. When the cell membrane of SMA was permeabilized using ${\beta}$-escin, SMA was contracted by increasing extracellular $Ca^{2+}$ concentration from 0 to $10{\mu}M$ and the magnitude of contraction was decreased by a decrease in $pH_o$ and vice versa. Conclusion: From these results, it can be concluded that a decrease in $pH_o$ might inhibit vascular contraction by reducing the reactivity of vascular smooth muscle to vasoactive agents, $Ca^{2+}$ influx and the sensitivity of vascular smooth muscle to $Ca^{2+}$.

• Journal of Chest Surgery
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• v.42 no.5
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• pp.588-596
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• 2009

Background: To clarify the effect of hypoxia on vascular contractility, we tried to show whether hypoxia induced the release of endothelium-derived relaxing factor (EDRF) and the nature of the underlying mechanism for this release. Material and Method: Isometric contractions were observed in rabbit aorta, and the released EDRF from the rabbit aorta was bioassayed by using rabbit denuded carotid artery. The intracellular $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) in the cultured rabbit aortic endothelial cells was recorded by a microfluorimeter with using Fura-2/AM. Hypoxia was evoked to the blood vessels or endothelial cells by eliminating the $O_2$ in the aerating gases in the external solution. Chemical hypoxia was evoked by applying deoxyglucose or $CN^-$. Result: Hypoxia relaxed the precontracted rabbit thoracic aorta that had its endothelium, and the magnitude of the relaxation was gradually increased by repetitive bouts of hypoxia. In contrast, hypoxia-induced relaxation was not evoked in the aorta that was denuded of endothelium. In a bioassay experiment, hypoxia released endothelium-derived relaxing factor (EDRF) and the release was inhibited by L-NAME or the $K^+$ channel blocker tetraethylammonium (TEA). In the cultured endothelial cells, hypoxia augmented the ATP-induced increase of the intracellular $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) and this increase was inhibited by TEA. Furthermore, chemical hypoxia also increased the $Ca^{2+}$ influx. Conclusion: From these results, it can be concluded that hypoxia might induce the release of NO from rabbit aortic endothelial cells by increasing $[[Ca^{2+}]_i$.

• Journal of Chest Surgery
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• v.37 no.3
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• pp.210-219
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• 2004

Extracellular $K^{+}$ concentration ([ $K^{+}$]$_{0}$ ) can be increased within several mM by the efflux of intracellular $K^{+}$. To investigate the effect of an increase in [ $K^{+}$]$_{0}$ on vascular contractility, we attempted to examine whether extracellular $K^{+}$ might modulate vascular contractility, endothelium-dependent relaxation (EDR) and intracellular $Ca^2$$^{+}$ concentration ([C $a^2$$^{+}$]$_{i}$ ) in endothelial cells (EC). We observed isometric contractions in rabbit carotid, superior mesenteric, basilar arteries and movse aorta. [C $a^2$$^{+}$]$_{i}$ was recorded by microfluorimeter using Fura-2/AM in EC. No change in contractility was recorded by the increase in [ $K^{+}$]$_{0}$ from 6 to 12 mM in conduit artery such as rabbit carotid artery. whereas resistant vessels, such as basilar and branches of superior mesenteric arteries (SMA), were relaxed by the increase. In basilar artery, the relaxation by the increase in [ $K^{+}$]$_{0}$ to from 1 to 3 mM was bigger than that by the increase from 6 to 12 mM. In contrast, in branches of SMA, the relaxation by the increase in [ $K^{+}$]$_{0}$ to from 6 to 12 mM is bigger than that by the increase from 1 to 3 mM. $Ba^2$$^{+}$ (30 $\mu$M) did not inhibit the relaxation by the increase in [ $K^{+}$]$_{0}$ from 1 to 3 mM but did inhibit the relaxation by the increase from 6 to 12 mM. In the mouse aorta without the endothelium or treated with $N^{G}$_nitro-L-arginine (30 $\mu$M), nitric oxide synthesis blocker, the increase in [ $K^{+}$]$_{0}$ from 6 to 12 mM did not change the magnitude of contraction induced either norepinephrine or prostaglandin $F_2$$_{\alpha}$. The increase in [ $K^{+}$]$_{0}$ up to 12 mM did not induce contraction of mouse aorta but the increase more than 12 mM induced contraction. In the mouse aorta, EDR was completely inhibited on increasing [ $K^{+}$]$_{0}$ from 6 to 12 mM. In cultured mouse aorta EC, [C $a^2$$^{+}$]$_{i}$ , was increased by acetylcholine or ATP application and the increased [C $a^2$$^{+}$]$_{i}$ , was reduced by the increase in [ $K^{+}$]$_{0}$ reversibly and concentration-dependently. In human umbilical vein EC, similar effect of extracellular $K^{+}$ was observed. Ouabain, a N $a^{+}$ - $K^{+}$ pump blocker, and N $i^2$$^{+}$, a N $a^{+}$ - $Ca^2$$^{+}$ exchanger blocker, reversed the inhibitory effect of extracellular $K^{+}$. In resistant arteries, the increase in [ $K^{+}$]$_{0}$ relaxes vascular smooth muscle and the underlying mechanisms differ according to the kinds of the arteries; $Ba^2$$^{+}$-insensitive mechanism in basilar artery and $Ba^2$$^{+}$ -sensitive one in branches of SMA. It also inhibits [C $a^2$$^{+}$]$_{i}$ , increase in EC and thereby EDR. The initial mechanism of the inhibition may be due to the activation of N $a^{+}$ - $K^{+}$pump. activation of N $a^{+}$ - $K^{+}$pump.p.p.p.