백서에서 방광 출구 단기부분폐색 초기의 방광 내 Nitric Oxide Synthase의 발현 변화

임동현,정현철,송재만 대한비뇨의학회 2008 Investigative and Clinical Urology Vol.49 No.7

Purpose: Nitric oxide synthase(NOS) is an important enzyme in the production of nitric oxide(NO). The constitutive type(cNOS) is expressed in the normal physiologic state, and the inducible type(iNOS) in expressed in the active immune state. cNOS is divided into an endothelial type (eNOS) and a neuronal type(nNOS). eNOS affects blood vessels, while nNOS affects nerve fibers. In the present study, we evaluated the expression of eNOS and nNOS in rat bladders with short-term partial outlet obstructions. We presupposed that NO is responsible for prolonged micturition problems after partial outlet obstruction. Materials and Methods: Specific pathogen-free Sprague-Dawley rats weighing 250-300g were used for the study. Individual bladders were obtained from sham-operated control rats(n=5) and from experimental rats at 12 hours and 1, 2, 3, and 7 days after partial urethral obstruction(n=25). eNOS and nNOS were detected using immunochemical staining and analyzed with confocal microscopy and an image analyzer. Results: eNOS and nNOS expression were detected in both the control group and in the group with partial outlet obstruction. The expression of eNOS showed a sharp increase at 3 days after obstruction and returned to normal at 7 days. The expression of nNOS was not significantly different between the two groups. Conclusions: In this study, we showed that eNOS increases in the rat bladder after partial outlet obstruction. This finding suggests that overproduction of NO may be the result of ischemic injury sustained during partial bladder outlet obstruction. Purpose: Nitric oxide synthase(NOS) is an important enzyme in the production of nitric oxide(NO). The constitutive type(cNOS) is expressed in the normal physiologic state, and the inducible type(iNOS) in expressed in the active immune state. cNOS is divided into an endothelial type (eNOS) and a neuronal type(nNOS). eNOS affects blood vessels, while nNOS affects nerve fibers. In the present study, we evaluated the expression of eNOS and nNOS in rat bladders with short-term partial outlet obstructions. We presupposed that NO is responsible for prolonged micturition problems after partial outlet obstruction. Materials and Methods: Specific pathogen-free Sprague-Dawley rats weighing 250-300g were used for the study. Individual bladders were obtained from sham-operated control rats(n=5) and from experimental rats at 12 hours and 1, 2, 3, and 7 days after partial urethral obstruction(n=25). eNOS and nNOS were detected using immunochemical staining and analyzed with confocal microscopy and an image analyzer. Results: eNOS and nNOS expression were detected in both the control group and in the group with partial outlet obstruction. The expression of eNOS showed a sharp increase at 3 days after obstruction and returned to normal at 7 days. The expression of nNOS was not significantly different between the two groups. Conclusions: In this study, we showed that eNOS increases in the rat bladder after partial outlet obstruction. This finding suggests that overproduction of NO may be the result of ischemic injury sustained during partial bladder outlet obstruction.

대뇌 기저핵 신경세포에서 Nitric Oxide를 매개로 한 망간의 세포독성

정용욱,배재훈,송대규,박원균,고복현,김두희,신동훈,Jung, Yong-Wook,Bae, Jae-Hoon,Song, Dae-Kyu,Park, Won-Kyun,Ko, Bok-Hyun,Kim, Doo-Hie,Shin, Dong-Hoon 대한예방의학회 1999 예방의학회지 Vol.32 No.4

Objectives:eurotoxicity is mediated by nitric oxide(NO) in the rat primary neuronal cultures and assess the effect of $Mn^{2+}$ on the N-methyl-D aspartate(NMDA) receptors. Methods: We have used 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)assay to examine the effect of cytotoxicity of $MnCl_2$ in neuronal cells , NO production was determined by measuring nirites, a stable oxidation product of NO. The neurons in the rat that contains neuronal nitric oxide synthase(nNOS) were examined by immunofluorescence and confocal microscopy. The effects of $Mn^{2+}$ on the NMDA receptors was assesed by the whole cell voltage clamp technique. Results: We showed that the NO release and NOS expression was increased with 500uM $MnCl_2$ treatment and an NOS inhibitors, $N^G-nitro-L-arginine$, prevented neurotoxicity elicited by manganese. In the electrophysiological study, $Mn^{2+}$ does not block or activate the NMDA receptors and not pass through the NMDA receptors in a neurons of basal ganglia. Conclusions: It is concluded that manganese neurotoxicity in basal ganglia was partially mediated by nitric oxide in the cell culture model.

Agmatine Attenuates Nitric Oxide Synthesis and Protects ER-structure from Global Cerebral Ischemia in Rats

문진희,김재환,박경아,이원택,백자현,이종은 대한해부학회 2009 Anatomy & Cell Biology Vol.42 No.3

In ischemic strokes, apoptosis is caused by excitotoxicity, ionic imbalance, oxidative/nitrosative stress, and apoptotic-like pathways. Nitric oxide (NO), a free radical, is elevated after ischemic insult. NO, which is generated primarily by neuronal nitric oxide synthase (nNOS) and inducible nitric oxide synthase (iNOS), promotes neuronal damage following ischemia. Evidence obtained in recent years has demonstrated that endoplasmic reticulum (ER)-mediated cell death plays an important role in cerebral ischemia. Agmatine is an endogenous substance synthesized from L-arginine by arginine decarboxylase (ADC) and is present in mammalian brain. We had previously reported that agmatine contributes to neuroprotection against ischemic injury. In continuation of our earlier work, we intended to investigate whether agmatine protects brain from transient global ischemia, and also tried to determine the neuroprotective mechanism of agmatine. Twenty minutes of transient global ischemia was induced by 4 vessel occlusion (4-VO). Agmatine (100 mg/kg, IP) was administered simultaneously with reperfusion. Samplings of brain were done at 6, 24, 48, and 72 h after reperfusion to determine the effect of agmatine on ischemic injured hippocampus. ER-damage was also investigated using electron microscope. Results showed that agmatine treatment prevented delayed neuronal cell death in hippocampal CA1 neurons after global cerebral ischemia. It also blocked NOS expression in the rat brain. Agmatine induced the increased expression of glucose-regulated protein 78 (Grp78). These results suggest that agmatine inhibits the production of NO by decreasing the expression of nNOS and iNOS on global forebrain ischemia and the neuroprotective effect of agmatine were concerned with the ER stress-mediated condition.

Kainic acid에 의해 유발된 경련이 흰쥐 해마의 Nitric Oxide Synthase의 미치는 영향

박종권(Jong Kwon Park),박 찬(Chan Park),강민정(Min Jeong Kang),강경란(Kyoung Lan Kang),이재룡(Jae Ryong Lee),김정혜(Jung Hye Kim),유진화(Jin Hwa Yoo),허영범(Young Buhm Huh),안희경(Hee Kyoung Ahn) 대한해부학회 2000 Anatomy & Cell Biology Vol.33 No.5

지금까지의 연구에 의하면 kainic acid (KA)-유도 경련은 해마에 존재하는 신경세포를 손상시키고 nitric oxide synthase(NOS)신경세포의 소실을 유발시키는 것으로 알려졌다. 또한 nitric oxide (NO)는 신경세포의 손상을 유발하는 것으로 알려졌다. 따라서 본 연구에서 KA를 흰쥐에게 투여하여 경련을 유발한 다음 6시간, 1일, 3일, 6일 경과 후에 해마에 일어나는 신경세포의 손상과 NOS신경세포의 활성과 발현의 변화를 은 침착법 (silver impregnation), nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) 조직화학 그리고 역전사효소 연쇄반응 (reverse transcriptase-polymerase chain reaction)을 이용하여 분석하였다. 그 결과, 신경세포의 손상은 1일군에서 부터 관찰되었으며, 3일군에서 가장 많은 세포 손상이 관찰되었다. NADPH-d의 염색성은 대조군에 비해 KA를 투여한 실험군에서 통계적으로 유의성 있게 증가하였다. 특히 1일과 3일군에서 강한 염색 성을 보였고, 6일군에서는 염색성이 약해지는 경향을 보였다. 해마의 부위에 따른 NADPH-d 신경세포 수의 변화를 측정한 결과 KA 투여 후 3일군에서 CA1/CA2과 CA3/CA4 부위에서 NADPH-d 신경세포가 통계적으로 유의성 있게 감소하였으 나, 치아이랑 (dentate gyrus)에서는 유의성 있는 변화를 보이지 않았다. 또한 nNOS, iNOS 그리고 eNOS의 mRNA의 발현은 모두 대조군에 비해 KA 투여 후 6시간과 1일군에서 증가하며, 3일군과 6일군에서는 감소하는 경향을 보였다. 이상의 결과에서 경련 발생후 유발되는 신경세포의 손상은 NOS 신경세포의 활성의 증가와 시기적으로 일치하는 것으로 보아 경련에 의한 해마 신경세포의 손상에 NOS 관여하는 것으로 생각되며, NOS의 활성은 mRNA 수준에서 조절되는 것으로 생각된다. We have investigated the neural cell damage and the change in the expression of NOS in the rat hippocampus, one of the brain structures most vulnerable to seizures. Rats were injected with kainic acid (KA) and sacrificed 6 h, 1 d, 3 d and 6 d after KA administration. The neural cell damage and the expression pattern of NOS was studied using silver impregnation, NADPH-diaphorase (NADPH-d) histochemistry and reverse transcription-polymerase chain reaction (RT-PCR) analysis. Silver impregnation revealed that kainic acid caused pyramical cell damage which was most severe in the CA1/CA2 subfield and hilus and to a lesser degree in the CA3 region. The optical densities of NADPH-dpositive neurons in the CA1, CA3 and dentate gyrus (DG) regions of the hippocampus were shown to have increased in samples obtained 1 d and 3 d after injection of KA. The number of NADPH-d-positive neurons in the CA1 and CA3 regions of the hippocampus was shown to have decreased in samples obtained 3 d and 6 d after injection of KA. However, the number of NADPH-d-positive neurons in the DG region did not change significantly. The increase in the levels of nNOS, iNOS and eNOS mRNA reached maximal values in samples obtained 1 d after KA treatment. Our findings indicate that the KA-induced seizures induce neural cell damage, increase NOS activity and upregulate the expression of NOS mRNA, which suggests the possibility of a functional role of NOS in bringing about changes in the cells in the hippocampus following seizures.

Nitric Oxide Is an Essential Mediator for Neuronal Differentiation of Rat Primary Cortical Neuron Cells

Oh, Soo-Jin,Heo, Jee-In,Kho, Yoon-Jung,Kim, Jeong-Hyeon,Kang, Hong-Joon,Park, Seong-Hoon,Kim, Hyun-Seok,Shin, Jong-Yeon,Kim, Min-Ju,Kim, Sung Chan,Park, Jae-Bong,Kim, Jaebong,Lee, Jae-Yong The Korean Society for Brain and Neural Science 2010 Experimental Neurobiology Vol.19 No.2

<P>Nitric oxide (NO) regulates proliferation, differentiation and survival of neurons. Although NO is reported to involve in NGF-induced differentiation of PC12 cells, the role of NO has not been characterized in primary neuron cells. Therefore, we investigated the role of NO in neuronal differentiation of primary cortical neuron cells. Primary cortical neuron cells were prepared from rat embryos of embryonic day 18 and treated with NMMA (NOS inhibitor) or PTIO (NO scavenger). Neurite outgrowth of neuron cells was counted and the mRNA levels of p21, p27, c-jun and c-myc were measured by RT-PCR. Neurite outgrowth of primary cortical neuron cells was inhibited a little by NOS inhibitor and completely by NO scavenger. The mRNA levels of p21 and p27, differentiation-induced growth arrest genes were increased during differentiation, but they were decreased by NOS inhibitor or NO scavenger. On the other hand, the level of c-jun mRNA was not changed and the level of c-myc mRNA was increased during differentiation differently from previously reported. The levels of these mRNA were reversed in NOS inhibitor- or NO scavenger-treated cells. The level of nNOS protein was not changed but NOS activity was inhibited largely by NOS inhibitor or NO scavenger. These results suggest that NO is an essential mediator for neuronal differentiation of primary cortical neuron cells.</P>

Neuronal Nitric Oxide-mediated Cytotoxicity in Trophoblast Cells Induced by Increase of Intracellular Calcium

Shin, Mi-Kyung,Kwon, Yong-Hyun,Shin, Jong-Chul,Yang, Dong-Eun,Lee, Sung-Keun,Kang, Ju-Hee,Park, Chang-Shin The Korean Society of Toxicogenomics and Toxicopro 2008 Molecular & cellular toxicology Vol.4 No.1

Cell death of trophoblast, particularly by abnormal release of physiological nitric oxide (NO) has been known to be a causative factor of pre-eclampsia. In the present study, effects of intracellular calcium increase enhancing the activity of NO synthases (neuronal NO synthase, nNOS in this trophoblast cells) on the cell death were examined in a human placental full-term cell line (HT-1). Furthermore, we analyzed the possible mechanisms underlying the augmentation of $Ca^{++}$-mediated NOS activity mediated by protein kinases like PKC, PKA, or CaM-KII. In experiments for cell toxicity, a calcium ionophore (ionomycin $10{\mu}M$) enhanced cell death confirmed by MTT assay, and increased significantly nNOS activity determined with a hemoglobin oxidation assay. This cell death was partially protected by pre-treatment of 7-nitroindazole (7-NI, $10{\mu}M$ and $100{\mu}M$), a nNOS-specific inhibitor. Additionally, $Ca^{++}$-ionophore -induced increase of nNOS activity also was partially normalized by pre-treatment of specific inhibitors of protein kinases, PKC, PKA or CaM-KII. Therefore, we suggest that an increase of calcium influx, leading to the activation of nNOS activity, which in turn may result in the death of trophoblast cells by involvement of signaling mechanisms of protein kinases.

Reconsideration of the Autonomic Cranial Ganglia: An Immunohistochemical Study of Mid‐Term Human Fetuses

Kiyokawa, Hiromichi,Katori, Yukio,Cho, Kwang Ho,Murakami, Gen,Kawase, Tetsuaki,Cho, Baik Hwan Wiley Subscription Services, Inc., A Wiley Company 2012 The anatomical record Vol.295 No.1

<P><B>Abstract</B></P><P>The cranial parasympathetic ganglia have been reported to paradoxically contain the sympathetic nerve marker, tyrosine hydroxylase (TH), in addition to neurons expressing parasympathetic markers such as vasoactive intestinal peptide (VIP) and neuronal nitric oxide synthase (nNOS). However, the distribution of these molecules in the cranial ganglia of human fetuses has not yet been examined. Using paraffin sections from 10 mid‐term human fetuses (12–15 weeks), we performed immunohistochemistry for TH, VIP, and nNOS in the parasympathetic ciliary, pterygopalatine, otic, and submandibular ganglia, and for comparison, the sensory inferior vagal ganglion. The ciliary and submandibular ganglia contained abundant TH‐positive neurons. In the former, TH‐positive neurons were much more numerous than nNOS‐positive neurons, whereas in the latter, nNOS immunoreactivity was extremely strong. No or a few cells in the pterygopalatine, otic, and inferior vagal ganglia expressed TH. Ciliary TH neurons appeared to compensate for classically described sympathetic fibers arising from the superior cervical ganglion, whereas in the submandibular ganglion, nNOS‐positive neurons as well as TH neurons might innervate the lingual artery in addition to the salivary glands. Significant individual variations in the density of all these markers suggested differences in sensitivity to medicine affecting autonomic nerve function. Consequently, in the human cranial autonomic ganglia, it appears that there is no simple dichotomy between sympathetic and parasympathetic function. Anat Rec, 2012. © 2011 Wiley Periodicals, Inc.</P>

Thrombin-activated microglia contribute to death of dopaminergic neurons in rat mesencephalic cultures: Dual roles of mitogen-activated protein kinase signaling pathways

Lee, Da Yong,Oh, Young J.,Jin, Byung Kwan Wiley Subscription Services, Inc., A Wiley Company 2005 GLIA Vol.51 No.2

<P>This study evaluated the role of thrombin-activated microglia in the neurodegeneration of mesencephalic cultures. Immunocytochemical and biochemical evidence indicated that in co-cultures consisting of rat cortical microglia and mesencephalic neurons, thrombin led to nonselective loss of mesencephalic neurons. Accompanying neurodegeneration, microglial activation was obvious, evidenced by expression of tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) and by increasing production of TNF-α and nitric oxide (NO). In mesencephalic neurons treated with conditioned media (CM) taken from thrombin-activated microglia, the number of dopaminergic neurons was significantly attenuated. The neurotoxicity of the CM was diminished when it was derived from microglia co-treated with thrombin and either an extracellular signal-regulated kinase 1/2 (ERK1/2) pathway inhibitor (PD98059) or a p38-mitogen-activated protein kinase (p38-MAPK) inhibitor (SB203580). Moreover, jun N-terminal kinase (JNK) and p38-MAPK were activated in mesencephalic neurons treated with CM of thrombin-activated microglia. Inhibition of JNK and p38-MAPK rescued the dopaminergic neurons. Collectively, these results indicate that thrombin-activated microglia induce neurodegeneration in cultured mesencephalic neurons and that the MAPKs actively participate in both microglial activation and neurodegeneration. The present data carefully suggest that microglial activation triggered by thrombin may be involved in the neuropathological processes of dopaminergic neuronal cell death that occur in Parkinson's disease. © 2005 Wiley-Liss, Inc.</P>

영아 비후성 유문 협착증에서 신경성 Nitric Oxide Synthase의 mRNA 발현감소

허민희(Min-Hee Hur),서정민(Jeong-Meen Seo),조응호(Eung-Ho Cho),이현규(Hyeon-Gyu Yi),박창신(Chang-Shin Park),차영남(Young-Nam Cha),홍기천(Kee-Chun Hong),우제홍(Ze-Hong Woo) 대한외과학회 2000 Annals of Surgical Treatment and Research(ASRT) Vol.59 No.3

Ganglion cardiacum or juxtaductal body of human fetuses

김지현,조광호,Zhe Wu Jin,Gen Murakami,Hiroshi Abe,채옥희 대한해부학회 2018 Anatomy & Cell Biology Vol.51 No.4

The ganglion cardiacum or juxtaductal body is situated along the left recurrent laryngeal nerve in the aortic window and is an extremely large component of the cardiac nerve plexus. This study was performed to describe the morphologies of the ganglion cardiacum or juxtaductal body in human fetuses and to compare characteristics with intracardiac ganglion. Ganglia were immunostained in specimens from five fetuses of gestational age 12–16 weeks and seven fetuses of gestational age 28–34 weeks. Many ganglion cells in the ganglia were positive for tyrosine hydroxylase (TH; sympathetic nerve marker) and chromogranin A, while a few neurons were positive for neuronal nitric oxide synthase (NOS; parasympathetic nerve marker) or calretinin. Another ganglion at the base of the ascending aorta carried almost the same neuronal populations, whereas a ganglion along the left common cardinal vein contained neurons positive for chromogranin A and NOS but no or few TH-positive neurons, suggesting a site-dependent difference in composite neurons. Mixtures of sympathetic and parasympathetic neurons within a single ganglion are consistent with the morphology of the cranial base and pelvic ganglia. Most of the intracardiac neurons are likely to have a non-adrenergic non-cholinergic phenotype, whereas fewer neurons have a dual cholinergic/noradrenergic phenotype. However, there was no evidence showing that chromogranin A‒ and/or calretinin-positive cardiac neurons corresponded to these specific phenotypes. The present study suggested that the ganglion cardiacum was composed of a mixture of sympathetic and parasympathetic neurons, which were characterized the site-dependent differences in and near the heart.