• Journal of Periodontal and Implant Science
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• v.34 no.4
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• pp.807-818
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• 2004

산화질소는 생리적 농도에서 세포내 신호전달자로 작용하지만 높은 농도에서는 세포독성을 일으킨다. 최근 치은 섬유아세포와 치주인대 섬유아세포는 산화질소 합성효소를 가지고 있고 세균의 lipopolysaccharide나 cytokine에 의해 대량의 높은 농도의 산화질소가 합성된다는 보고가 있음에도 지금까지 치은 조직에서 산화질소의 세포독성에 대한 연구는 아직 이루어 지지않고 있다. 본 연구는 사람의 치은 섬유아세포에서, 산화질소유도세포 고사기전을 밝히는데 목적이 있다. 세포 생장력은 MTT 방법으로 측정하였고, 세포의 형태적 변화는 Diff-Quick 염색법으로 조사하였다. Bcl-2 famly와 Fas 발현 정도는 RT-PCR 방법에 의해 확인하였으며, caspase-3, -8 와 -9의 활성은 spectrophotometer로 reactive oxygen species (ROS)는 형광분광계에 의해 측정되었다. 미토콘드리아에서 세포질로 분비된 cytochrome c는 western blot으로 조사하였다. 산화질소 유리제인 sodium nitroprusside (SNP) 처리는 사람 섬유아세포의 생존률을 시간과 농도 의존적으로 감소시켰고, 세포용적축소, 염색사 용축, DNA 절편화를 일으켰다. 또한, SNP 처리로 미토콘드리아에서 세포질로 유리되는 cytochrome c 양이 증가되었고, caspase-9 과 caspase-3 의 활성이 증가되었다. 한편, SNP 처리에 의해 death receptor 구성요소인 Fas 발현이 증가되었고, caspase-8의 활성이 증가되었다. Bcl-2 family 에 대한 RT-PCR 분석결과, 세포고사를 억제하는 Bcl-2 발현은 감소되었으나 세포고사를 자극하는 Bax와 Bid의 발현은 증가되었다. Soluble guanylate cyclase 억제제인 ODQ는 SNP에 의한 세포 생존율 감소를 차단하지 못했다. 따라서, 본 실험의 결과들은 사람 섬유아세포에서 산화질소유도 세포고사에 Bcl-2 family나 ROS가 매개하는 미토콘드리아 의존 및 death receptor 의존 세포고사기전이 관여함을 시사하였다.

• Korean Journal of Veterinary Research
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• v.44 no.3
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• pp.357-366
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• 2004

We studied the effect of propofol (PPF) on the endothelium-dependent vascular responses in isolated rat thoracic aorta. In aortic rings with endothelium, PPF inhibited the phenylephrine (PE)-induced contraction in a concentration-dependent manner. In PE-precontracted preparations, PPF attenuated the endothelium-dependent relaxation by acetylcholine but not by A23187. And PPF did not attenuate the endothelium-independent relaxation by sodium nitroprusside (SNP). The relaxation induced by acetylcholine in PE-precontracted aortic rings was significantly augmented by zaprinast, a cGMP-specific phosphodiesterase inhibitor, and this augmentation was inhibited by PPF. Although SNP-induced relaxation was significantly augmented by zaprinast, this augmentation was not inhibited by PPF. In preparations preconstricted with PE, the PPF-induced relaxation was inhibited by atropine. In addition, PPF attenuated the vasorelaxation by phosphodiesterase inhibitors (IBMX, Ro20-1724 or zaprinast except milrinone). In vivo, the infusion of acetylcholine and SNP showed decreased arterial blood pressure in rats. The pre-injection of PPF inhibited the acetylcholine-induced blood pressure lowering, but not the SNP-induced blood pressure lowering. These results suggest that PPF can attenuate in part the acetylcholine-induced vasorelaxation and blood pressure lowering through the inhibition of the acetylcholine receptor-mediated endothelium-derived relaxing factor by acting on endothelium. It is considered that the inhibitory effect of PPF on the vasorelaxation is due to the decreased level of cGMP which can be attributed to the inhibition of the muscarinic receptor and/or receptor-G-protein interaction.

• The Korean Journal of Physiology and Pharmacology
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• v.18 no.4
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• pp.353-358
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• 2014

Control of blood pressure is maintained by the interaction between the arterial baroreflex and vestibulosympathetic reflex during postural changes. In this study, the contributions of vestibular receptors and baroreceptors to the maintenance of blood pressure following acute hypotension were compared in terms of phosphorylated extracellular regulated protein kinase (pERK) expression in the nucleus tractus solitaries (NTS). Expression of pERK in the NTS was measured in conscious rats that had undergone bilateral labyrinthectomy (BL) and/or sinoaortic denervation (SAD) 5, 10, 20, and 40 min following acute hypotension induced by sodium nitroprusside (SNP) infusion. Expression of pERK increased significantly in the NTS in the control group following SNP infusion, and the expression peaked at 10 min after SNP infusion. The number of pERK positive neurons increased following SNP infusion in BL, SAD, and BL+SAD groups, although the increase was smaller than in control group. The BL group showed a relatively higher reduction in pERK expression than the SAD group, and the pERK expression in the NTS was localized to the caudal portion of the nuclei in the BL and SAD groups. These results suggest that the vestibular receptors may play a key role in maintaining blood pressure following acute hypotension; thus, the vestibular system may contribute to compensate for orthostatic hypotension.

• Toxicological Research
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• v.16 no.2
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• pp.133-139
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• 2000

This study was designed to evaluate the mechanism by which reactive nitrogen intermediates (RNI) induced the cytotoxicity of human doparminergic neuroblastoma SH-SY5Y cells. 3-Morpholino-sydnonimine (SIN-l), a donor of peroxynitrite (ONOO) and sodium nitroprusside (SNP), a donor of nitric oxide (NO) induced cell detachment and apoptotic death, as characterized by chromatin condensation, the ladder pattern fragmentation of genomic DNA and morphological nuclear changes. SIN-l also induced the activation of caspase 3-like protease in a time-dependent manner. Exogenous antioxidants, such as reduced glutathione (GSH), N-acetylcysteine (NAC), and selenium protected the cells from apoptotic death and reduced the activation of caspase 3-like protease by SIN-1. Furthermore, SIN-l directly reduced the intracellular levels of glutathione. Taken together, these data suggested that RNI including NO and peroxynitrite decrease the concentration of intracellular antioxidant such as GSH, which lead to the apoptotic death of human neuroblastoma SH-SY5Y cells.

• IMMUNE NETWORK
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• v.5 no.4
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• pp.199-204
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• 2005

Background: ADP-ribosyl pyrophosphatases (ADPRase) has been known to catalyze the hydrolysis of ADP-ribose to ribose-5-phosphate and AMP. The role of ADPRase has been suggested to sanitize the cell by removing potentially toxic ADP-ribose. In this study, we examined the effect of nitric oxide on ADPRase activity in macrophages. Methods: ADPRase activity was measured in NO-inducing J774 cells. For in vitro experiments, recombinant human ADPRase was prepared in bacteria. Results: ADPRase activity was increased by the treatment of exogenous NO generating reagent, sodium nitroprusside (SNP), in J774 cells. The increased ADPRase activity was mediated by the post-translational modification, likely to cause cADP-ribosylation via nitrosylation of cysteine residue on the enzyme. The stimulation with endogeneous NO inducers, $TNF-{\alpha}/IFN-{\gamma}$, also increased ADPRase activity through NO synthesis. Futhermore, ADPRase activity may be mediated by the post-translational modification of ADPRase, ADP-ribosylation. Conclusion: These results indicate that NO synthesized by macrophage activation plays a critical role in the increase in ADPRase activity following ADP-ribose metabolism.

• Proceedings of the PSK Conference
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• 2003.10b
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• pp.98.1-98.1
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• 2003

Bee venom (BY) has been utilized to relieve pain and to treat inflammatory diseases such as rheumatoid arthritis (RA). However, the molecular mechanism by which BV-induced anti-arthritis effect has been not reported yet. Therefore, in the present study we investigated anti-inflammatory effect of BV in a murine marcrophage cell line Raw 264.7 cell and synoviocyte obtained from RA patients. The present data showed that BV has a preventive effect on lipopolysaccharide (LPS) and sodium nitroprusside (SNP) induced induction of COX-2, cPLA2 and iNOS. (omitted)

• Journal of Acupuncture Research
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• v.21 no.3
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• pp.107-119
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• 2004

Objective : The purpose of this study was to investigate the effect of Bee Venom and melittin on the lipopolysaccharide(LPS) and sodium nitroprusside(SNP)-induced expressions of Cell viability, nitric oxide(NO), inducible nitric oxide synthase(iNOS), extra-signal response kinase(ERK), jun N-terminal Kinase(JNK) and p38 kinase(p38)- mitogen activated protein kinase(MAPK) Family- in RAW 264.7 cells, a murine macrophage cell line. Methods : The expressions of cell viability by MTT assay, NO by Nitrite assay and iNOS, ERK, JNK and p38 were determined by Western blotting. Results : 1. Compared with the control group, 0.5, 1, $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin increased cell viability of RAW 264.7 induced by LPS and SNP significantly respectively. 2. Compared with the control group, 0.5, 1, $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin inhibited expression of NO induced by LPS and SNP significantly respectively. 3. Compared with the control group, 1, $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin inhibited expression of iNOS induced by LPS significantly and 0.5, 1, $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin inhibited expression of iNOS induced by SNP significantly. 4. Compared with the control group, the expression of ERK induced by LPS and SNP decreased significantly in the treatment groups of $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin, which of p-ERK by LPS also did in 1, $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin, but which of p-ERK by SNP did not decrease. 5. Compared with the control group, the. expression of JNK induced by LPS and SNP decreased significantly in the treatment groups of 5, $10{\mu}g/m{\ell}$ melittin, which of p-JNK by LPS in 5, $10{\mu}g/m{\ell}$ melittin and by SNP in $1{\mu}g/m{\ell}$ bee venom and $10{\mu}g/m{\ell}$ melittin decreased significantly. 6. Compared with the control group, the expression of p38 induced by LPS did not have significant difference, which induced by SNP decreased significantly in the treatment groups of 1, $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin. p-p38 induced by LPS decreased significantly in the treatment group of $10{\mu}g/m{\ell}$ of melittin, which induced by SNP also decreased significantly in 0.5, 1, $5{\mu}g/m{\ell}$ bee venom and 5, $10{\mu}g/m{\ell}$ melittin.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.2
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• pp.190-197
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• 2011

The objective of the present study was to elucidate the fundamental mechanism of bovine oviduct epithelial cell (BOEC) co-culture on developmental capacity of bovine IVM/IVF embryos and to determine whether or not melatonin acts as an antioxidant in BOEC culture and subsequent embryo development. These studies examined the effects of melatonin against NO-induced oxidative stress on cell viability, lipid peroxidation (LPO) and the expression of antioxidant genes (CuZnSOD, MnSOD and Catalase) or apoptosis genes (Bcl-2, Caspase-3 and Bax) during BOECs culture. We also evaluated the developmental rates of bovine IVM/IVF embryos with BOEC co-culture, which were pre-treated with melatonin ($1,000\;{\mu}M$) in the presence or absence of sodium nitroprusside (SNP, $1,000\;{\mu}M$) for 24 h. Cell viability in BOECs treated with SNP (50-$2,000\;{\mu}M$) decreased while melatonin addition (1-$1,000\;{\mu}M$) increased viability in a dose-dependent manner. Cell viability in melatonin plus SNP ($1,000\;{\mu}M$) gradually recovered according to increasing melatonin addition (1-$1,000\;{\mu}M$). The LPO products were measured by thiobarbituric acid (TBA) reaction for malondialdehyde (MDA). Addition of melatonin in BOEC culture indicated a dose-dependent decrease of MDA, and in the SNP group among BOECs treated with SNP or melatonin plus SNP groups MDA was significantly increased compared with SNP plus melatonin groups (p<0.05). In expression of apoptosis or antioxidant genes detected by RT-PCR, Bcl-2 and antioxidant genes were detected in melatonin or melatonin plus SNP groups, while Caspase-3 and Bax genes were only found in the SNP group. When bovine IVM/IVF embryos were cultured for 6-7 days under the BOEC co-culture system pre-treated with melatonin in the presence or absence of SNP, the highest developmental ability to blastocysts was obtained in the $1,000\;{\mu}M$ melatonin group. These results suggest that melatonin has an anti-oxidative effect against NO-induced oxidative stress on cell viability of BOECs and on the developmental competence of bovine IVM/IVF embryo co-culture with BOEC.

• The Korean Journal of Physiology and Pharmacology
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• v.13 no.6
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• pp.503-510
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• 2009

To elucidate the mechanism of cyclic nucleotides, such as adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP), in the regulation of human gastric motility, we examined the effects of forskolin (FSK), isoproterenol (ISO) and sodium nitroprusside (SNP) on the spontaneous, high $K^+$ and acetylcholine (ACh)-induced contractions of corporal circular smooth muscle in human stomach. Gastric circular smooth muscle showed regular spontaneous contraction, and FSK, ISO and SNP inhibited its phasic contraction and basal tone in a concentration-dependent manner. High $K^+$ (50 mM) produced sustained tonic contraction, and ACh $(10\;{\mu}M)$ produced initial transient contraction followed by later sustained tonic contraction with superimposed phasic contractions. FSK, ISO and SNP inhibited high $K^+$-induced tonic contraction and also ACh-induced phasic and tonic contraction in a reversible manner. Nifedipine $(1\;{\mu}M)$, inhibitor of voltage-dependent L-type calcium current $(VDCC_L)$, almost abolished ACh-induced phasic contractions. These findings suggest that FSK, ISO and SNP, which are known cyclic nucleotide stimulators, inhibit smooth muscle contraction in human stomach partly via inhibition of $VDCC_L$.

• The Korean Journal of Physiology and Pharmacology
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• v.15 no.5
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• pp.273-277
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• 2011

Nitric oxide (NO) and atrial natriuretic peptide (ANP) may induce vascular relaxation by increasing the production of cyclic guanosine monophosphate (cGMP), an important mediator of vascular tone during sepsis. This study aimed to determine whether regulation of NO and the ANP system is altered in lipopolysaccharide (LPS)-induced kidney injury. LPS (10 $mg{\cdot}kg^{-1}$) was injected in the tail veins of male Sprague-Dawley rats; 12 hours later, the kidneys were removed. Protein expression of NO synthase (NOS) and neutral endopeptidase (NEP) was determined by semiquantitative immuno-blotting. As an index of synthesis of NO, its stable metabolites (nitrite/nitrate, NOx) were measured using colorimetric assays. mRNA expression of the ANP system was determined by real-time polymerase chain reaction. To determine the activity of guanylyl cyclase (GC), the amount of cGMP generated in response to sodium nitroprusside (SNP) and ANP was calculated. Creatinine clearance decreased and fractional excretion of sodium increased in LPS-treated rats compared with the controls. Inducible NOS protein expression increased in LPS-treated rats, while that of endothelial NOS, neuronal NOS, and NEP remained unchanged. Additionally, urinary and plasma NOx levels increased in LPS-treated rats. SNP-stimulated GC activity remained unchanged in the glomerulus and papilla in the LPS-treated rats. mRNA expression of natriuretic peptide receptor (NPR)-C decreased in LPS-treated rats, while that of ANP and NPR-A did not change. ANP-stimulated GC activity reduced in the glomerulus and papilla. In conclusion, enhancement of the NO/cGMP pathway and decrease in ANP clearance were found play a role in the pathogenesis of LPS-induced kidney injury.