• The Korean Journal of Physiology and Pharmacology
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• v.13 no.5
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• pp.385-392
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• 2009

Angiotensin II (Ang II) plays an important role in vascular hypertension. The role of the chemokine CCL5 on Ang II-induced activities in vascular smooth muscle cells (VSMCs) has not been studied. In this study, we elucidated the effect of CCL5 on Ang II-induced 12-lipoxygenase (LO) expression and cell proliferation in spontaneously hypertensive rats (SHR) VSMCs. CCL5 decreased Ang II-induced 12-LO mRNA expression and protein production, and it increased Ang II type 2 ($AT_2$) receptor expression in SHR VSMCs. The inhibitory effect of CCL5 on Ang II-induced 12-LO mRNA expression was mediated through the $AT_2$ receptor. Although treatment of CCL5 alone induced SHR VSMCs proliferation, CCL5 inhibited Ang II-induced VSMCs proliferation and PD123,319, an $AT_2$ receptor antagonist, blocked the inhibitory effect of CCL5 on Ang II-induced VSMCs proliferation. Phosphorylation of p38 was detected in VSMCs treated with Ang II or CCL5 alone. But, decrease of p38 phosphorylation was detected in VSMCs treated with Ang II and CCL5 simultaneously (Ang II/CCL5) and PD123,319 increased p38 phosphorylation in VSMCs treated with Ang II/CCL5. Therefore, these results suggest that the inhibitory effect of CCL5 on Ang II-induced VSMCs proliferation is mediated by the $AT_2$ receptor via p38 inactivation, and CCL5 may play a beneficial role in Ang II-induced vascular hypertension.

• Archives of Pharmacal Research
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• v.19 no.5
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• pp.390-395
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• 1996

To investigate whether $AT_{1}$ receptor antagonists are acting by increasing endothelium-de-pendent and -independent relaxation of aortas in normotensive rats, $AT_{1}$ receptor antagonists, losartan and KR-30988, and angiotensin converting enzyme inhibitor, captopril, were orally administered for two weeks (50 mg/kg, b.i.d.). THe blood pressure, heart rate and body weight were not significantly changed by losartan, KR-30988 and captopril compared to the control group. In aortic preparations, the $pD_{2}$ of KR-30988 for ACh-induced relaxation was 8.33 $\pm$ 0.16, significantly (p <0.05) lower than that of control group $(7.71 \pm 0.15)$. ACh-induced relaxation was significantly increased on losartan-treated group (p<0.01) at $10^{-6}$ M of ACh, and in captopril-treated group (p<0.05) at the range of $10^{-7}$ -$10^{-5}$ M of ACh. The $pD_{2}$ values for histamine-induced relaxatio of losartan, KR-30988 and captopril were 5.57 $\pm$ 0.10, 5.85 $\pm$ 0.21 and 5.60 $\pm$ 0.01, respectively, with significant differences in all groups (p<0.01) compared to that of control group (5.13 $\pm$ 0.09). ACh-induced relaxations of aortic preparations were not changed by pretreatment of indomethacin ($10_{-5}$ M), and completely bolcked by pretreatment of L-NAME $(10_{-5}M)$ in all groups. Sodium nitroprusside-induced relaxations were not significantly changed by all drugs tested in this experiments. These results suggest that $AT_{1}$ receptor antagonists, losartan and KR-30988, enhance the endothelium-dependent relaxatio on aortic preparations through the release of, or increase sensitivity, to nitric oxide in nor-motensive rats.

• International Journal of Oral Biology
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• v.39 no.1
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• pp.49-56
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• 2014

We investigated the role of central P2X receptors in inflammatory pain transmission in the orofacial area in rats. Experiments were carried out using male Sprague-Dawley rats weighing 230-280g. Complete Freund's adjuvant (CFA, $40{\mu}L$) was applied subcutaneously to the vibrissa pad to produce inflammatory pain. The intracisternal administration of iso-PPADS tetrasodium salt, a non-selective P2X receptor antagonist, A317491 sodium salt hydrate, a $P2X_{2/3}$ receptor antagonist, 5-BDBD, a $P2X_4$ receptor antagonist, or A438079 hydrochloride, a $P2X_7$ receptor antagonist, was performed 5 days after CFA injection. Subcutaneous injections of CFA produced increases in thermal hypersensitivity. Intracisternal injections of iso-PPADS ($25{\mu}g$) or A438079 (25 or $50{\mu}g$) produced significant anti-hyperalgesic effects against thermal stimuli compared to the vehicle group. A317491 or 5-BDBD did not affect the head withdrawal latency times in rats showing an inflammatory response. Subcutaneous injections of CFA resulted in the up-regulation of OX-42, a microglia marker, and GFAP, an astrocyte marker, in the medullary dorsal horn. The intracisternal administration of A438079 reduced the numbers of activated microglia and astrocytes in the medullary dorsal horn. These results suggest that a blockade of the central $P2X_7$ receptor produces antinociceptive effects, mediated by inhibition of glial cell function in the medullary dorsal horn. These data also indicate that central $P2X_7$ receptors are potential targets for future therapeutic approaches to inflammatory pain in the orofacial area.

• Molecules and Cells
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• v.39 no.4
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• pp.322-329
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• 2016

Voltage-gated $Ca^{2+}$ ($Ca_V$) channels are dynamically modulated by Gprotein-coupled receptors (GPCR). The $M_1$ muscarinic receptor stimulation is known to enhance $Ca_V2.3$ channel gating through the activation of protein kinase C (PKC). Here, we found that $M_1$ receptors also inhibit $Ca_V2.3$ currents when the channels are fully activated by PKC. In whole-cell configuration, the application of phorbol 12-myristate 13-acetate (PMA), a PKC activator, potentiated $Ca_V2.3$ currents by ~two-fold. After the PMA-induced potentiation, stimulation of $M_1$ receptors decreased the $Ca_V2.3$ currents by $52{\pm}8%$. We examined whether the depletion of phosphatidylinositol 4,5-bisphosphate ($PI(4,5)P_2$) is responsible for the muscarinic suppression of $Ca_V2.3$ currents by using two methods: the Danio rerio voltage-sensing phosphatase (Dr-VSP) system and the rapamycin-induced translocatable pseudojanin (PJ) system. First, dephosphorylation of $PI(4,5)P_2$ to phosphatidylinositol 4-phosphate (PI(4)P) by Dr-VSP significantly suppressed $Ca_V2.3$ currents, by $53{\pm}3%$. Next, dephosphorylation of both PI(4)P and $PI(4,5)P_2$ to PI by PJ translocation further decreased the current by up to $66{\pm}3%$. The results suggest that $Ca_V2.3$ currents are modulated by the $M_1$ receptor in a dual mode-that is, potentiation through the activation of PKC and suppression by the depletion of membrane $PI(4,5)P_2$. Our results also suggest that there is rapid turnover between PI(4)P and $PI(4,5)P_2$ in the plasma membrane.

• Archives of Pharmacal Research
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• v.29 no.11
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• pp.1032-1041
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• 2006

Extracellular adenosine 5'-triphosphate (ATP) affects the function of many tissues and cells. To confirm the biological activity of ATP on human myeloid leukemic cells, F-36P and HL-60, cells were treated with a variety of concentrations of ATP. The stimulation with extracellular ATP induced the arrest of cell proliferation and cell death. from the analysis of Annexin-V staining and caspase activity by flow cytometry. The Annexin-V positive cells in both cell lines were dramatically increased following ATP stimulation. The expression of P2 purinergic receptor genes was confirmed, such as P2X1, P2X4, P2X5, P2X7 and P2Y1, P2Y2, P2Y4, P2Y5, P2Y6, P2Y11 in both leukemic cell lines. Interestingly, ATP induced intracellular calcium flux in HL-60 cells but not in F-36P cells, as determined by Fluo-3 AM staining. Cell cycle analysis revealed that ATP treatment arrested both F-36P and HL-60 cells at G1/G0. Taken together, these data showed that extracellular ATP via P2 receptor genes was involved in the cell proliferation and survival in human myeloid leukemic cells, HL-60 and F-36P cells by the induction of apoptosis and control of cell cycle. Our data suggest that treatment with extracellular nucleotides may be a novel and powerful therapeutic avenue for myeloid leukemic disease.

• Development and Reproduction
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• v.11 no.3
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• pp.187-193
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• 2007

Ethane 1,2-dimethane sulfonate(EDS), a Leydig cell specific toxicant, has been widely used to create the reversible testosterone withdrawal rat model. Though the maintenance of epididymal structure and function is highly dependent on the testosterone secreted from testis, its derivatives, dihydroxytestosterone(DHT) and estrogen, might have crucial roles. The aim of present study was to monitor the expression patterns of sex steroid receptors, cytochrome P450 aromatase(P450arom) and $5{\alpha}$-reductase in the rat epididymis up to 7 weeks after EDS injection. Adult male rats($350{\sim}400g$) were injected with a single does of EDS(75 mg/kg i.p.) and sacrificed on weeks 0, 1, 2, 3, 4, 5, 6 and 7. The transcriptional activities of the target genes were evaluated by semi-quantitative RT-PCRs. The transcript level of estrogen receptor alpha($ER{\alpha}$) in EDS group was significantly higher than control level on week 1(P<0.01). After week 2, there was no significant difference in $ER{\alpha}$ levels between EDS group and control. The transcript level of estrogen receptor beta($ER{\beta}$) in EDS group was significantly higher than control level on week 1(P<0.05), lowered on weeks 2 and 3(P<0.05 and P<0.01, respectively), fluctuated during weeks 4 and 6, and elevated on week 7(P<0.05). The androgen receptor (AR) message levels increased significantly week 2(P<0.01), then returned to control level on week 3. In contrast, expression of cytochrome P450 aromatase(P450arom) decreased sharply during weeks $1{\sim}3$(P<0.01 on weeks 1 and 2; P<0.05 on week 3), then went back to control level on week 4. The mRNA level of $5{\alpha}$-reductase type 2($5{\alpha}$-RT2) increased significantly on week 4(P<0.01), then returned to control level. The present study indicated that EDS administration could induce reversible alterations in the transcriptional activities of sex steroid hormone receptors and androgenconverting enzymes in rat epididymis. EDS injection model will be useful to clarify the regulation mechanism of mammalian epididymal physiology.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1993.04a
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• pp.61-61
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• 1993

사람의 혈액의 백혈구로부터 Polymerase Chain Reaction 방법으로 $\beta$$_2$-adrenergic receptor 유전자를 증폭하였다. 이 증폭된 유전자를 pbluescript KS(＋)에 cloning하였으며, 제한효소지도 작성과 부분적인 염기배열 확인으로 증폭된 유전자가 사람의 $\beta$$_2$-adrenergic receptor 유전자임을 확인하였다. $\beta$$_2$-adrenergic receptor 유전자를 N말단의 아미노산이 21개 제거되도록 결손시킨후 yeast의 pSec5의 Killer toxin signal sequence의 뒤에 in-frame으로 연결하였다. 현재 효모에서의 발현양상 및 발현된 단백질의 활성을 조사중이다.

• Archives of Pharmacal Research
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• v.27 no.10
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• pp.1065-1072
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• 2004

Depression is associated with a dysfunctional serotonin (5-hydroxytryptamine; 5-HT) system. More recently, several lines of evidence suggest that an important factor in the development of depression may be a deficit in the function and expression of $5-HT_{1A}$ receptors. The present study assessed if Nelumbinis Semen (N. s.) had an anti-depression effect through reversing a decrease in $5-HT_{1A}$receptor binding in rats with depression-like symptoms induced by chronic mild stress. Using a $5-HT_{1A}$ receptor binding assay, with a specific $5-HT_{1A}$receptor agonist, 8- OH-DPAT (8-hydroxy-2-(di-n-propylamino) tetralin), the mechanism of the anti-depression effect of N. s. on rats was investigated, and the effects compared with two well-known antidepressants, Hyperium Perforatum (St. Johns Wort) and fluoxetine (Prozac). Animals were divided into five groups: the normal (N) group without chronic mild stress (CMS), the control (C) group under CMS for 8 weeks, the Nelumbinis Semen (N. s.) treatment group under CMS for 8 weeks, the Hyperium Perforatum (H. p.) treatment group under CMS for 8 weeks and finally, the fluoxetine (F) treatment group under CMS for 8 weeks. Each treatment was administered to rats during the last 4 weeks of the 8-week CMS. A sucrose intake test was performed to test the anti-depression effect of N. s. The N. s. treatment significantly reversed the decreased sucrose intake under CMS (P<0.05 compared to control group under CMS). In the CA2 and CA3 regions of the hippocampus, both N. s. and H. p. reversed the CMS-induced decrease in $5-HT_{1A}$receptor binding. In the I to II regions of the frontal cortex, N. s. and H. p. also reversed the CMS-induced decrease in$5-HT_{1A}$receptor binding, and even showed a significant increase in $5-HT_{1A}$receptor binding compared to the F treatment group (N. s. vs. P, p<0.05, H. p. vs. P, p<0.05). However, in the hypothalamus, all treatments reversed the CMSinduced decrease in $5-HT_{1A}$receptor binding. This reversal effect of N. s. on the decrease in $5-HT_{1A}$receptor binding in the frontal cortex, hippocampus and hypothalamus of rat brains was very similar to that of H. p, but different from that of F. It is concluded that N. s. presents an anti-depression effect through enhancing $5-HT_{1A}$receptor binding.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.7
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• pp.3053-3059
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• 2012

Human epidermal growth factor receptor 2 (HER2) is a member of the epidermal growth factor receptor family of receptor tyrosine kinases that plays important roles in all processes of cell development. Their overexpression is related to many cancers, including examples in the breast, ovaries and stomach. Anticancer therapies targeting the HER2 receptor have shown promise, and monoclonal antibodies against subdomains II and IV of the HER2 extra-cellular domain (ECD), Pertuzumab and Herceptin, are currently used in treatments for some types of breast cancers. Since anti HER2 antibodies targeting distinct epitopes have different biological effects on cancer cells; in this research linear and conformational B cell epitopes of HER2 ECD, subdomain III, were identified by bioinformatics analyses using a combination of linear B cell epitope prediction web servers such as ABCpred, BCPREDs, Bepired, Bcepred and Elliprro. Then, Discotope, CBtope and SUPERFICIAL software tools were employed for conformational B cell epitope prediction. In contrast to previously reported epitopes of HER2 ECD we predicted conformational B cell epitopes $P1_C$: 378-393 (PESFDGDPASNTAPLQ) and $P2_C$: 500-510 (PEDECVGEGLA) by the integrated strategy and P4: PESFDGD-X-TAPLQ; P5: PESFDGDP X TAPLQ; P6: ESFDGDP X NTAPLQP; P7: PESFDGDP-X-NTAPLQ; P8: ESFDG-XX-TAPLQPEQL and P9: ESFDGDP-X-NTAPLQP by SUPERFICIAL software. These epitopes could be further used as peptide antigens to actively immune mice for development of new monoclonal antibodies and peptide cancer vaccines that target different epitopes or structural domains of HER2 ECD.

• Journal of Acupuncture Research
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• v.28 no.2
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• pp.57-67
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• 2011

목적 : Collagenase-induced osteoarthritis(OA) 동물 모델에서 전침의 adrenergic mechanism을 연구하고자 한다. 방법 : Collagenase-induced arthritis(CIA)를 유발하기 위하여 5주령의 male Sprague-Dawley rat의 뒷다리 좌측 무릎 관절에 0.05ml의 4mg/ml collagenase solution을 intra-articular 주입하고, 다시 4일 후에 같은 부위에 같은 농도의 collagenase solution을 intra-articular boosting injection 시행한 뒤, gross, histopathological features 및 biomarker activity 변화를 관찰하였다. 예비실험을 통하여 CIA rat model에서 진통효과를 발휘하는 것으로 확인한, 족삼리(足三里) ($ST_{36}$)에 대한 저빈도 train pulse EA stimulation (2Hz, 0.07 mA, 0.3ms)을 침치료 방법으로 적용하였다. 전침의 진통기전을 확인하기 위하여, ${\alpha}1$-adrenergic antagonist (prazosin, 1 mg/kg, i.p.), ${\alpha}2$-adrenergic receptor antagonist (yohimbine, 2mg/kg, i.p.), ${\alpha}1$-adrenergic receptor agonist(phenylephrine, 2mg/kg, i.p.), ${\alpha}2$-adrenergic receptor agonist(clonidine, $40{\mu}g$/kg, i.p.)을 전침시행 20분 전에 복강 내로 전처치하였다. Tail flick unit(Ugo Basile Model 7360)을 이용하여 열자극에 대한 통증역치를 측정하였다. 결과 : 퇴행성관절염 징후(gross, histopathological features)와 통증역치의 변화가 최대값을 나타내는 CIA 유발 4주차에 저빈도 전침자극(train pulse, 2Hz, 0.07mA, 0.3ms)을 족삼리($ST_{36}$)에 적용하였으며, 족삼리 전침의 진통효과는 ${\alpha}2$-adrenergic receptor antagonist(yohimbine, 2mg/kg, i.p.)전처치에 의해 억제되었으나, ${\alpha}1$-adrenergic antagonist(prazosin, 1 mg/kg, i.p.)전처치에는 억제되지 않았다. 또 ${\alpha}2$-adrenergic receptor agonist(clonidine, $40{\mu}g$/kg, i.p.)의 전처치를 통하여 유의한 synergistic analgesic effect가 관찰되었으나, ${\alpha}1$-adrenergic receptor agonist(phenylephrine, 2mg/kg, i.p.)의 전처치는 전침의 진통효과에 synergistic effect를 미치지 않는 것으로 나타났다. 결론 : 저빈도 족삼리 전침은 CIA로 유발된 염증성 통증에 대하여 진통효과를 발휘하며, 이는 ${\alpha}2$-adrenergic receptor에 의하여 매개되는 것으로 보이며 ${\alpha}1$-adrenergic receptor는 영향을 미치지 않는 것으로 사료된다.