• Journal of Life Science
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• v.17 no.6 s.86
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• pp.783-790
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• 2007

G protein coupled receptors (GPCRs) transmit various extracellular signals into the cells. Upon binding of the ligands, conformational changes in the extracellular and/or transmembrane (TM) domains of CPCRs were propagated into the cytoplasmic (CP) domain of the molecule leading to the activation of their cognate heterotrimeric C proteins and kinases. Constitutively active GPCR mutants causing the activation of C Protein signaling even in the absence of ligand binding are of interest for the study of activation mechanism of GPCRs. Two classes of constitutively active mutations, categorized by their effects on the salt bridge between Ell3 and K296, were found in the TM domain of rhodopsin. Opsin mutants containing combinations of the mutations were constructed to study the conformational changes required for the activation of rhodopsin. Rhodopsin chromophore regenerated with 11-cis-retinal showed a thermal stability inversely correlated with its constitutive activity. In contrast, rhodopsin mutants exhibited a binding affinity to an agonist, all-trans-retinal, in a constitutive activity-dependent manner. In order to test whether the conformational changes responsible for the activation of trans-ducin (Gt) are the same as the conformation required for the recognition of rhodopsin kinase, analysis of the mutants were carried out with phosphorylation by rhodopsin kinase. Rhodopsin mutants containing combinations of different classes of the mutations showed a strong synergistic effect on the phosphorylation of the mutants in the dark as similar to that of Gt activation. The results suggest that at least two or three kinds of segmental and independent conformational changes are required for the activation of rhodopsin and the conformational changes responsible for activating rhodopsin kinase and Gt are similar to each other.

• Molecules and Cells
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• v.39 no.7
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• pp.557-565
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• 2016

The paired immunoglobulin-like type 2 receptor (PILR) family consists of two functionally opposite members, inhibitory $PILR{\alpha}$ and activating $PILR{\beta}$ receptors. PILRs are widely expressed in various immune cells and interact with their ligands, especially CD99 expressed on activated T cells, to participate in immune responses. Here we investigated whether PILR-derived agonists inhibit ${\beta}1$ integrin activity as ligands for CD99. PILR-derived peptides as well as PILR-Fc fusion proteins prevented cell adhesion to fibronectin through the regulation of ${\beta}1$ integrin activity. Especially, PILRpep3, a representative 3-mer peptide covering the conserved motifs of the PILR extracellular domain, prevented the clustering and activation of ${\beta}1$ integrin by dephosphorylating FAK and vinculin, which are major components of focal adhesion. In addition, PILRpep3 inhibited transendothelial migration of monocytes as well as endothelial cell tube formation. Furthermore, upon intraperitoneal injection of PILRpep3 into mice with collagen-induced arthritis, the inflammatory response of rheumatoid arthritis was strongly suppressed. Taken together, these results suggest that PILR-derived agonist ligands may prevent the inflammatory reactions of rheumatoid arthritis by activating CD99.

• Journal of Korean Neurosurgical Society
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• v.29 no.7
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• pp.868-876
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• 2000

Objective : Parkinsonian rat models have generally been characterized by unilateral destruction of both the nigrostriatal pathway and the mesolimbic pathway using the neurotoxin 6-hydroxydopamine. The induction of contraversive turning by apomorphine in these models is thought to reflect the stimulation of supersensitive dopamine D2 receptor or receptor-mediated mechanisms in denervated neostriatum. The present study was undertaken to investigate the expression of dopamine D2 receptor in denervated striatum according to modalities of apomorphine(dopamine agonist) treatment after creating a hemiparkinsonian rat model in which there is 6-hydroxydopamine induced destruction of the unilateral dopaminergic nigrostriatal pathway. Methods : After making complete lesion in left side substantia nigra pars compacta(SNpc) by stereotactic injection of 6-hydroxydopamine into medial and lateral areas of SNpc, and confirming successful animal model by apomorphine induced contraversive turning behavior without recovery and complete destruction of ipsilateral SNpc with tyrosine hydroxylase immunostaining in 7th day after operation, 15 rats of parkinsonian model were studied with or without administration of apomorphine at varying doses and durations. According to the modalities of apomorphine treatment for 4 days, these rats were divided into 3 groups, as not-treated group, intermittently treated group and constantly treated group. For investigating the extent of the expression of dopamine D2 receptor in denervated striatum, immunohistochemical staining by dopamine D2 receptor antibody and Western blot were performed. Results : In the D2 receptor antibody immunohistochemical staining, the mean number of positive stained neurons was highest in not-treated group($20.5{\pm}1.14$) of 3 groups. In constantly treated group, the mean number of positive stained neurons was less($3.9{\pm}1.79$) than intermittently treated group(p<0.05). The Western blotting with the D2 receptor antibody revealed that expression of receptors was also highest in not-treated group and less in constantiy treated group than intermittently treated group. Conclusion : Dopamine D2 receptors in denervated striatum of parkinsonian rat models, which were not treated with apomorphine, revealed to be most highly expressed. And, according to doses and durations of apomorphine administration, desensitization of the receptor was more apt to develop with constant treatment than intermittent treatment. In clinical setting, the authors believe that, in long-term treated parkinsonian patients, desensitization of dopamine receptors due to chronic dopaminergic stimulation seems to be partially related to mechanisms of drug tolerance.

• Biomedical Science Letters
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• v.23 no.4
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• pp.327-332
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• 2017

A2B adenosine receptor (A2BAR) is known to be a regulator of bone homeostasis, but the regulatory mechanism of A2BAR on the osteoclast proliferation are poorly explored. Recently, we have shown that stimulation with BAY 60-6583, a specific agonist of A2BAR, significantly reduced macrophage-colony stimulating factor (M-CSF)-induced osteoclast proliferation by inducing cell cycle arrest at G1 phase and increasing the apoptosis of osteoclasts. The objective of this study was to investigate the regulatory mechanisms of cell cycle and apoptosis by A2BAR stimulation. The expression of A2BAR and M-CSF receptor, c-Fms, was not changed by A2BAR stimulation whereas M-CSF effectively induced c-Fms expression during osteoclast proliferation. Interestingly, A2BAR stimulation remarkably increased the expression of $p27^{Kip-1}$, a cell cycle inhibitor, but the expression of Cyclin D1 and cdk4 was not affected. In addition, while BAY 60-6583 treatment reduced the expression of Bcl2, an anti-apoptotic oncogene, it failed to regulate the expression of Bax, a pro-apoptotic marker. Taken together, these results imply that the increase of $p27^{Kip-1}$ inducing cell cycle arrest at G1 phase and the decrease of Bcl2 inducing anti-apoptotic response by A2BAR stimulation contribute to the down-regulation of osteoclast proliferation.

• Toxicological Research
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• v.23 no.2
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• pp.135-142
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• 2007

Formononetin is an isoflavonoid phytoestrogen found in certain foodstuffs such as soy and red clover. In this study, we examined the action of formononetin with the carcinogen activation pathway mediated through the aryl hydrocarbon receptor (AhR) in MCF-7 breast carcinoma cells. Treating the cells with formononetin alone caused the accumulation of CYP1A1 mRNA as well as elevation in CYP1A1-specific 7-ethoxyresorufin O-deethylase (EROD) activity in a dose dependent manner. However, a concomitant treatment with 7,12-dimethylbenz[a]anthracene (DMBA) and formononetin markedly reduced both the DMBA-inducible EROD activity and CYP1A1 mRNA level. Under the same conditions, formononetin inhibited the DMBA-induced AhR transactivation, as shown by reporter gene analysis using a xenobiotic responsive element (XRE). Additionally, formononetin inhibited both DMBA-inducible nuclear localization of the aryl hydrocarbon receptor (AhR) and metabolic activation of DMBA, as measured by the formation of the DMBA-DNA adducts. Furthermore, formononetin competed with the prototypical AhR ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), for binding to the AhR in an isolated rat cytosol. These results suggest that formononetin might be considered as a natural ligand to bind on AhR and consequently produces a potent protective effect against DMBA-induced genotoxicity. Therefore, that's the potential to act as a chemopreventive agent that is related to its effect on AhR pathway as antagonist/agonist.

• The Korean Journal of Physiology and Pharmacology
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• v.1 no.1
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• pp.13-17
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• 1997

Carbamazepine (CBZ), an anticonvulsant, has beeen reported to displace ligands at adenosine receptors. Several studies have demonstrated that as far as $A_2$adenosine receptors is concerned, CBZ acts as an antagonist. However, the situation with regard to Al receptors is less straightforward. In this study, we describe the effects of one-week CBZ treatment (25 mg/kg/day) on cerebrocortical $A_1$ adenosine receptors. $A_1$ adenosine receptor bindings as determined by using $[^3CH]DPCPX$ was not significantly altered in membranes prepared from CBZ-treated rats. However, there was a significant decrease in the $A_1$ adenosine receptor-mediated stimulation of $[^{35}S]GTP_{\gamma}S$ binding to cerebrocortical membranes prepared from CBZ-treated rats (20.0% decrease in basal activity; 17.8% decrease in maximal activity). The basal and $10^{-4}$ M forskolin-stimulated adenylyl cyclase activities were relatively unaffected by CBZ treatment, but 10 mM NaF-stimulated adenylyl cyclase activity was significantly reduced in CBZ-treated rats. It appears that one-week CBZ treatment caused an uncoupling of adenosine receptors from G proteins without alteration of $A_1$ adenosine receptor molecules, suggesting that CBZ acts as an agonist at $A_1$ adenosine receptors in rat brain.

• YAKHAK HOEJI
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• v.44 no.6
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• pp.558-565
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• 2000

SK-1080 is one of the newly developed orally active nonpeptide angiotensinII $AT_1-receptor$ antagonist that selectively acts at $AT_1$ receptor with high affinity. The cardiac effect on ischemia/reperfusion injury of SK-1080 was compared with those of losartan, a prototype of this class, in isolated rat hearts. Isolated perfused rat heart was pretreated with drug for 10 min and then subjected to global ischemia for 30 min followed by reperfusion with- or without drug for 30 min. The possible additive effect of SK-1080 on the platelet aggregation and coagulation in human blood was also studied. We investigated whether SK-1080 effects the platelet aggregation induced by ADP, a platelet agonist partially dependent on $thromboxaneA_2$. The clotting times in the prothrombin time (PT) and activated partial thromboplastin time (APTT) were also examined in human plasma in vitro as coagulation screening test. SK-1080 improved reperfusion function (LVDP, left ventricular developed pressure; PRP, rate-pressure product) in a dose-dependent manner. SK-1080 reduced ADP-induced platelet aggregation compared with vehicle but less than losartan, and did not affect clotting times.

• Molecules and Cells
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• v.41 no.3
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• pp.234-243
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• 2018

In recent years, the interest towards the relationship between incretins and bone has been increasing. Previous studies have suggested that glucagon-like peptide-1 (GLP-1) and its receptor agonists exert beneficial anabolic influence on skeletal metabolism, such as promoting proliferation and differentiation of osteoblasts via entero-osseous-axis. However, little is known regarding the effects of GLP-1 on osteoblast apoptosis and the underlying mechanisms involved. Thus, in the present study, we investigated the effects of liraglutide, a glucagon-like peptide-1 receptor agonist, on apoptosis of murine MC3T3-E1 osteoblastic cells. We confirmed the presence of GLP-1 receptor (GLP-1R) in MC3T3-E1 cells. Our data demonstrated that liraglutide inhibited the apoptosis of osteoblastic MC3T3-E1 cells induced by serum deprivation, as detected by Annexin V/PI and Hoechst 33258 staining and ELISA assays. Moreover, liraglutide upregulated Bcl-2 expression and downregulated Bax expression and caspase-3 activity at intermediate concentration (100 nM) for maximum effect. Further study suggested that liraglutide stimulated the phosphorylation of AKT and enhanced cAMP level, along with decreased phosphorylation of $GSK3{\beta}$, increased ${\beta}-catenin$ phosphorylation at Ser675 site and upregulated nuclear ${\beta}-catenin$ content and transcriptional activity. Pretreatment of cells with the PI3K inhibitor LY294002, PKA inhibitor H89, and siRNAs GLP-1R, ${\beta}-catenin$ abrogated the liraglutide-induced activation of cAMP, AKT, ${\beta}-catenin$, respectively. In conclusion, these findings illustrate that activation of GLP-1 receptor by liraglutide inhibits the apoptosis of osteoblastic MC3T3-E1 cells induced by serum deprivation through $cAMP/PKA/{\beta}-catenin$ and $PI3K/Akt/GSK3{\beta}$ signaling pathways.

• YAKHAK HOEJI
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• v.49 no.5
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• pp.370-374
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• 2005

Lactoferrin is a multifunctional protein that is found in milk, neutrophils, and other biological fluids, and its receptors have also been identified in the central nervous system. Recently, it was reported that bovine milk-derived lacto­ferrin (BLF) produced analgesia via a $\mu$-opioid receptor-mediated response in the spinal cord. However the precise mech­anism of this analgesic effect is remains unclear. In Randall-Selitto paw pressure study, each single administration of morphine (10 mg/kg) and BLF (50, 100 and 200 mg/kg) induced analgesia, however, NMDA receptor antagonist MK-801 (0.1, 0.2 and 0.3 mg/kg), inhibited analgesia induced by BLF (100 mg/kg). Intracerebroventricular infusion (I.C.V.) of N­methyl-D-aspartic acid (NMDA) ($0.3\;{\mu}g/8.0\;{\mu}l/hr/day$), as a NMDA receptor agonist, reversed inhibition of MK-801 (0.3 mg/kg) on analgesia induced by BLF (100 mg/kg). These results suggest that BLF have analgesic effect, through NMDA recep­tor activation.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.6
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• pp.675-686
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• 2017

Orthostatic hypotension (OH) is associated with symptoms including headache, dizziness, and syncope. The incidence of OH increases with age. Attenuation of the vestibulosympathetic reflex (VSR) is also associated with an increased incidence of OH. In order to understand the pathophysiology of OH, we investigated the physiological characteristics of the VSR in the disorder. We applied sodium nitroprusside (SNP) to conscious rats with sinoaortic denervation in order to induce hypotension. Expression of pERK in the intermediolateral cell column (IMC) of the T4~7 thoracic spinal regions, blood epinephrine levels, and blood pressure were evaluated following the administration of glutamate and/or SNP. SNP-induced hypotension led to increased pERK expression in the medial vestibular nucleus (MVN), rostral ventrolateral medullary nucleus (RVLM) and the IMC, as well as increased blood epinephrine levels. We co-administered either a glutamate receptor agonist or a glutamate receptor antagonist to the MVN or the RVLM. The administration of the glutamate receptor agonists, AMPA or NMDA, to the MVN or RVLM led to elevated blood pressure, increased pERK expression in the IMC, and increased blood epinephrine levels. Administration of the glutamate receptor antagonists, CNQX or MK801, to the MVN or RVLM attenuated the increased pERK expression and blood epinephrine levels caused by SNP-induced hypotension. These results suggest that two components of the pathway which maintains blood pressure are involved in the VSR induced by SNP. These are the neurogenic control of blood pressure via the RVLM and the humoral control of blood pressure via epinephrine release from the adrenal medulla.