• Biomolecules & Therapeutics
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• v.21 no.4
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• pp.277-283
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• 2013

In this study, we investigated the effects of a selective urotensin II (UII) receptor antagonist, SB-657510, on the inflmmatory response induced by UII in human umbilical vein endothelial cells (EA.hy926) and human monocytes (U937). UII induced inflammatory activation of endothelial cells through expression of proinflammatory cytokines (IL-$1{\beta}$ and IL-6), adhesion molecules (VCAM-1), and tissue factor (TF), which facilitates the adhesion of monocytes to EA.hy926 cells. Treatment with SB-657510 significantly inhibited UII-induced expression of IL-$1{\beta}$, IL-6, and VCAM-1 in EA.hy926 cells. Further, SB-657510 dramatically blocked the UII-induced increase in adhesion between U937 and EA.hy926 cells. In addition, SB-657510 remarkably reduced UII-induced expression of TF in EA.hy926 cells. Taken together, our results demonstrate that the UII antagonist SB-657510 decreases the progression of inflammation induced by UII in endothelial cells.

• Biomolecules & Therapeutics
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• v.24 no.5
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• pp.523-528
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• 2016

Urotensin II (UII) is a potent vasoactive peptide and mitogenic agent to induce proliferation of various cells including vascular smooth muscle cells (VSMCs). In this study, we examined the effects of a novel UII receptor (UT) antagonist, KR-36676, on vasoconstriction of aorta and proliferation of aortic SMCs. In rat aorta, UII-induced vasoconstriction was significantly inhibited by KR-36676 in a concentration-dependent manner. In primary human aortic SMCs (hAoSMCs), UII-induced cell proliferation was significantly inhibited by KR-36676 in a concentration-dependent manner. In addition, KR-36676 decreased UII-induced phosphorylation of ERK, and UII-induced cell proliferation was also significantly inhibited by a known ERK inhibitor U0126. In mouse carotid ligation model, intimal thickening of carotid artery was dramatically suppressed by oral treatment with KR-36676 (30 mg/kg/day) for 4 weeks compared to vehicle-treated group. From these results, it is indicated that KR-36676 suppress UII-induced proliferation of VSMCs at least partially through inhibition of ERK activation, and that it also attenuates UII-induced vasoconstriction and vascular neointima formation. Our study suggest that KR-36676 may be an attractive candidate for the pharmacological management of vascular dysfunction.

• Biomolecules & Therapeutics
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• v.25 no.3
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• pp.308-314
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• 2017

Urotensin II (UII) is a mitogenic and hypertrophic agent that can induce the proliferation of vascular cells. UII inhibition has been considered as beneficial strategy for atherosclerosis and restenosis. However, currently there is no therapeutics clinically available for atherosclerosis or restenosis. In this study, we evaluated the effects of a newly synthesized UII receptor (UT) antagonist, KR-36996, on the proliferation of SMCs in vitro and neointima formation in vivo in comparison with GSK-1440115, a known potent UT antagonist. In primary human aortic SMCs (HASMCs), UII (50 nM) induced proliferation was significantly inhibited by KR-36996 at 1, 10, and 100 nM which showed greater potency ($IC_{50}$: 3.5 nM) than GSK-1440115 ($IC_{50}$: 82.3 nM). UII-induced proliferation of HASMC cells was inhibited by U0126, an ERK1/2 inhibitor, but not by SP600125 (inhibitor of JNK) or SB202190 (inhibitor of p38 MAPK). UII increased the phosphorylation level of ERK1/2. Such increase was significantly inhibited by KR-36996. UII-induced proliferation was also inhibited by trolox, a scavenger for reactive oxygen species (ROS). UII-induced ROS generation was also decreased by KR-36996 treatment. In a carotid artery ligation mouse model, intimal thickening was dramatically suppressed by oral treatment with KR-36996 (30 mg/kg) which showed better efficacy than GSK-1440115. These results suggest that KR-36996 is a better candidate than GSK-1440115 in preventing vascular proliferation in the pathogenesis of atherosclerosis and restenosis.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.16 no.2
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• pp.1200-1206
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• 2015

The major target for drug discovery, G-protein coupled receptor (GPCR) is involved in many physiological activities and related to various diseases and disorders. Among experimental techniques relating to the GPCR drug discovery process, various cell-based screening methods are influenced by cell conditions used in the overall process. Recently, the utilization of frozen cells is suggested in terms of reducing data variation and cost-effectiveness. The aim of this study is to evaluate various conditions in cell freezing such as temperature conditions and storage terms. The stable cell lines for calcium sensing receptor and urotensin receptor were established followed by storing cultured cells at $-80^{\circ}C$ up to 4 weeks. To compare with cell stored at liquid nitrogen, agonist and antagonist responses were recorded based on the luminescence detection by the calcium induced photoprotein activation. Cell signals were reduced as the storage period was increased without the changes in $EC_{50}$ and $IC_{50}$ values $EC_{50}:3.46{\pm}1.36mM$, $IC_{50}:0.49{\pm}0.15{\mu}M$). In case of cells stored in liquid nitrogen, cell responses were decreased comparing to those in live cells, however changes by storage periods and significant variations of $EC_{50}/IC_{50}$ values were not detected. The decrease of cell signals in various frozen cells may be due to the increase of cell damages. From these results, the best way for a long-term cryopreservation is the use of liquid nitrogen condition, and for the purpose of short-term storage within a month, $-80^{\circ}C$ storage condition can be possible to adopt. As a conclusion, the active implementation of frozen cells may contribute to decrease variations of experimental data during the initial cell-based screening process.