Bark Constituents from Mushroom-detoxified Rhus verniciflua Suppress Kainic Acid-induced Neuronal Cell Death in Mouse Hippocampus

변종선,한윤희,홍성준,황성미,권용수,전완주,이희제,김성수,김명조 대한약리학회 2010 The Korean Journal of Physiology & Pharmacology Vol.14 No.5

Urushinol, a plant allergen, has significantly restricted the medical application of Rhus verniciflua,although it has been reported to possess a wide variety of biological activities such as anti-inflammatory,antioxidant, and anti-cancer actions. To reduce the urushinol content while maintaining the beneficial biological activities, mushroom-mediated fermentation of Rhus verniciflua was carried out and this method resulted in significantly attenuated allergenicity [1]. In the present study, to examine the neuroprotective properties of mushroom-fermented stem bark of Rhus verniciflua, two constituents were isolated from mushroom-fermented bark and their neuroprotective properties were examined in a mouse model of kainic acid (KA)-induced excitotoxicity. KA resulted in significant apoptotic neuronal cell death in the CA3 region of mouse hippocampus. However, seven daily administrations of RVH-1or RVH-2 prior to KA injection significantly attenuated KA-induced pyramidal neuronal cell death in the CA3 region. Furthermore, pretreatment with RVH-1 and RVH-2 also suppressed KA-induced microglial activation in the mouse hippocampus. The present study demonstrates that RVH-1 and RVH-2 isolated from Rhus verniciflua and detoxified using mushroom species possess neuroprotective properties against KA-induced excitotoxicity. This leads to the possibility that detoxified Rhus verniciflua can be a valuable asset in herbal medicine.

Kainic Acid-induced Neuronal Death is Attenuated by Aminoguanidine but Aggravated by L-NAME in Mouse Hippocampus

변종선,이상현,전성호,권용수,이희재,김성수,김영명,김명조,전완주 대한약리학회 2009 The Korean Journal of Physiology & Pharmacology Vol.13 No.4

Nitric oxide (NO) has both neuroprotective and neurotoxic effects depending on its concentration and the experimental model. We tested the effects of NG-nitro-L-arginine methyl ester (L-NAME), a nonselective nitric oxide synthase (NOS) inhibitor, and aminoguanidine, a selective inducible NOS (iNOS) inhibitor, on kainic acid (KA)-induced seizures and hippocampal CA3 neuronal death. L-NAME (50 mg/kg, i.p.) and/or aminoguanidine (200 mg/kg, i.p.) were administered 1 h prior to the intracerebroventricular (i.c.v.) injection of KA. Pretreatment with L-NAME significantly increased KA-induced CA3 neuronal death, iNOS expression, and activation of microglia. However, pretreatment with aminoguanidine significantly suppressed both the KA-induced and L-NAME-aggravated hippocampal CA3 neuronal death with concomitant decreases in iNOS expression and microglial activation. The protective effect of aminoguanidine was maintained for up to 2 weeks. Furthermore, iNOS knockout mice (iNOS−/−) were resistant to KA-induced neuronal death. The present study demonstrates that aminoguanidine attenuates KA-induced neuronal death, whereas L-NAME aggravates neuronal death, in the CA3 region of the hippocampus, suggesting that NOS isoforms play different roles in KA-induced excitotoxicity.

ICR 생쥐의 해마에서 스타니오칼신 2가 세포생성에 미치는 영향

이재원,변종선,이균영,오달균,전완주,김성수,이희제 대한정신약물학회 2010 대한정신약물학회지 Vol.21 No.3

본 연구에서는 STC2가 신경생성에 미치는 영향을 알아보기 위하여 100 nM STC2와 BrdU를 ICR 생쥐(23~25 g)의뇌실 내와 복강에 각각 차례대로 투여하였다. STC2와 BrdU 처치 후 24시간과 3주 후에 각 군의 실험동물을 희생하고그 뇌 절편으로 BrdU와 인산화 CREB의 면역조직화학법을시행하였다. 24시간 경과한 STC2 투여군의 해마 치상회에서 선조세포들의 증식이 과립구하 지역에서 대조군과 비교하여 유의하게 증가하였다. 하지만 STC2 처치 후 3주 후의뇌 조직에서는 신경생성의 차이는 없었다. 한편 해마 치상회에서의 CREB 인산화 변화는 24시간 경과한 STC2 투여군에서 증가하였다. STC2 투여군에서 과립구하 지역에서의선조세포들이 증가했다는 점과 이 영역에서 CREB 인산화가증가했다는 점은 STC2가 CREB 신호전달 체계를 통해 선조세포들의 증식을 증가시킬 가능성을 시사한다. Objective Stanniocalcin 2 (STC2) is a glycoprotein hormone that is widely expressed in mammalian kidney, heart, and thymus. However, the potential function of STC2 in the brain is less understood. In this study, we investigated whether treatment with STC2 influenced cell proliferation and neurogenesis in imprinting control region (ICR) mice. Methods 100 nM STC2 and 5’-bromo-2’-deoxyuridine (BrdU) (50 mg/kg) were administered intracerebroventricularly and intraperitoneally, respectively. On days 1 and 21 after the BrdU injection, sections of STC2-treated group and controls were carried out immunohistochemistry using anti-BrdU and anti-phosphor-cAMP-response element-binding protein (CREB) antibody. Results We found that the number of BrdU-positive cells was significantly increased in the hippocampal dentate gyrus (DG),compared with controls. Next, we observed that CREB phosphorylation was decreased in the hippocampal DG of STC2-treated group versus the controls. Conclusion These results suggest that STC2 treatment may increase cell proliferation by increasing CREB phosphorylation in the subgranular zone of the DG.

Melatonin Induces Akt Phosphorylation through Melatonin Receptor- and PI3K-Dependent Pathways in Primary Astrocytes

공필재,변종선,임소영,이재준,홍성준,권광준,김성수 대한약리학회 2008 The Korean Journal of Physiology & Pharmacology Vol.12 No.2

Melatonin has been reported to protect neurons from a variety of neurotoxicity. However, the underlying mechanism by which melatonin exerts its neuroprotective property has not yet been clearly understood. We previously demonstrated that melatonin protected kainic acid-induced neuronal cell death in mouse hippocampus, accompanied by sustained activation of Akt, a critical mediator of neuronal survival. To further elucidate the neuroprotective action of melatonin, we examined in the present study the causal mechanism how Akt signaling pathway is regulated by melatonin in a rat primary astrocyte culture model. Melatonin resulted in increased astrocytic Akt phosphorylation, which was significantly decreased with wortmannin, a specific inhibitor of PI3K, suggesting that activation of Akt by melatonin is mediated through the PI3K-Akt signaling pathway. Furthermore, increased Akt activation was also significantly decreased with luzindole, a non-selective melatonin receptor antagonist. As downstream signaling pathway of Akt activation, increased levels of CREB phoshorylation and GDNF expression were observed, which were also attenuated with wortmannin and luzindole. These results strongly suggest that melatonin exerts its neuroprotective property in astrocytes through the activation of plasma membrane receptors and then PI3K-Akt signaling pathway. Melatonin has been reported to protect neurons from a variety of neurotoxicity. However, the underlying mechanism by which melatonin exerts its neuroprotective property has not yet been clearly understood. We previously demonstrated that melatonin protected kainic acid-induced neuronal cell death in mouse hippocampus, accompanied by sustained activation of Akt, a critical mediator of neuronal survival. To further elucidate the neuroprotective action of melatonin, we examined in the present study the causal mechanism how Akt signaling pathway is regulated by melatonin in a rat primary astrocyte culture model. Melatonin resulted in increased astrocytic Akt phosphorylation, which was significantly decreased with wortmannin, a specific inhibitor of PI3K, suggesting that activation of Akt by melatonin is mediated through the PI3K-Akt signaling pathway. Furthermore, increased Akt activation was also significantly decreased with luzindole, a non-selective melatonin receptor antagonist. As downstream signaling pathway of Akt activation, increased levels of CREB phoshorylation and GDNF expression were observed, which were also attenuated with wortmannin and luzindole. These results strongly suggest that melatonin exerts its neuroprotective property in astrocytes through the activation of plasma membrane receptors and then PI3K-Akt signaling pathway.

韓國人胎兒 肝臟의 形態學的硏究

羅鳳鎭,卞鍾善,姜在福,李洸鎬 中央醫學社 1969 中央醫學 Vol.17 No.2

Inhibition of eNOS/sGC/PKG Pathway Decreases Akt Phosphorylation Induced by Kainic Acid in Mouse Hippocampus

이상현,변종선,공필재,이희제,김덕경,손종희,이재준,임소영,김성수,전완주,김해성 대한약리학회 2010 The Korean Journal of Physiology & Pharmacology Vol.14 No.1

The serine/threonine kinase Akt has been shown to play a role of multiple cellular signaling pathways and act as a transducer of many functions initiated by growth factor receptors that activate phosphatidylinositol 3-kinase (PI3K). It has been reported that phosphorylated Akt activates eNOS resulting in the production of NO and that NO stimulates soluble guanylate cyclase (sGC), which results in accumulation of cGMP and subsequent activation of the protein kinase G (PKG). It has been also reported that PKG activates PI3K/Akt signaling. Therefore, it is possible that PI3K, Akt, eNOS, sGC, and PKG form a loop to exert enhanced and sustained activation of Akt. However, the existence of this loop in eNOS-expressing cells, such as endothelial cells or astrocytes, has not been reported. Thus, we examined a possibility that Akt phosphorylation might be enhanced via eNOS/ sGC/PKG/PI3K pathway in astrocytes in vivo and in vitro. Phosphorylation of Akt was detected in astrocytes after KA treatment and was maintained up to 72 h in mouse hippocampus. 2 weeks after KA treatment, astrocytic Akt phosphorylation was normalized to control. The inhibition of eNOS, sGC, and PKG significantly decreased Akt and eNOS phosphorylation induced by KA in astrocytes. In contrast, the decreased phosphorylation of Akt and eNOS by eNOS inhibition was significantly reversed with PKG activation. The above findings in mouse hippocampus were also observed in primary astrocytes. These data suggest that Akt/eNOS/sGC/PKG/PI3K pathway may constitute a loop, resulting in enhanced and sustained Akt activation in astrocytes.

청소년기 ICR 생쥐에서 비선택적 산화질소 생성 효소 억제제인 L-NAME이 구속 스트레스 후의 행동양식과 해마에서의 GDNF 발현에 미치는 영향

권광준,변종선,이재원,전완주,김성수,이희제 대한정신약물학회 2012 대한정신약물학회지 Vol.23 No.1

Objective Depending on genetic or environmental effects over adolescent development, typical behavioral responses come out in adolescence. Also, alteration of nitric oxide (NO) levels in the brain has been associated with modifications of stress related behavior. Present study was designed to investigate the possible influence of chronic stress from restraint on the generation of depression in adolescent mice, and also to evaluate whether NO has modulatory roles in the behavioral and biological reactions. Methods ICR mice exposed to stressful restraint, 2 h per day, was treated with NG-nitro L-arginine methyl ester (L-NAME) (10 mg/kg), a non-selective NO synthase (NOS) inhibitor. To evaluate depression-like behavior in the mice, forced swim test and open field test were performed after the last restraint. To investigate stress-induced changes in the expression level of glial cell-derived neurotrophic factor (GDNF), free-floating immunohistochemistry was performed. Results The results showed that stressed group has longer immobility time and less crossing number in forced swimming and open field test, and that these stress responses were significantly prevented by L-NAME. Furthermore, decreased GDNF expression in the hippocampus by stress was prevented to that of controls within the L-NAME treated group. Conclusion The results suggest that stress and NO signaling could be involved in generation of depression in adolescence. It also suggested that GDNF might contribute to prevent stress-related behaviors. 산화질소는 신경전달물질 가운데 하나로 알려져 있으며 환경에 따라 의존적으로 유리 농도가 결정된다. 또한 유리 농도에 따라 다양한 반응들을 나타내는 것으로 알려져 있다. 그 중 스트레스는 산화질소의 유리를 증가시키는 것으로 알려져 있으며 스트레스 관련 행동양식과도 밀접한 관계가 있다. 산화질소를 생성하는 효소로는 NOS가 있으며, 아형으로는 eNOS, nNOS, iNOS가 있다. L-NAME은 비특이적으로 이들 NOS의 기능을 억제하는 것으로 알려져 있다. 본 연구에서는 구속 스트레스 동물 모델에서 스트레스 관련 동물행동 양식과 해마에서의 생물학적 변화 등을 통해 NO의 영향을 살펴보고자 하였다. L-NAME 투여군에서 스트레스 군보다 강제 수영 시험에서 부동시간이 유의하게 감소하였으며 이동 칸 수도 증가하였다. 이 결과들을 통해 L-NAME이 스트레스에 의해 유발된 행동들을 억제하는 것으로 보여진다. 뇌 해마에서 아교세포 유래 신경생장인자(GDNF)의 발현은 스트레스 군에서는 감소하나 L-NAME 투여군에서는 대조군과 유사하였다. 위 결과들을 통해 청소년기의 ICR 생쥐에서 NO 신호전달체계가 중요하게 작용하는 것으로 사료된다.

急性腹症에 對한 外科的 考察

兪鎭一,朴麟浩,金彩圭,卞鍾善,鄭埈 中央醫學社 1941 中央醫學 Vol.10 No.4

We have attempted to review our experience with 483 cases of acute abdomen, which were treated surgically at the Nam-Boo City Hospital from January 1962 to December 1965. Recently many surgeons emphasized important items utilized in the differential diagnosis of the acute abdomen. There is no substitute for good history taking and careful physical examination in most incidences. The decision for the urgent operation will be led in this way laboratory aids including the X-ray examination. The series of 483 cases analysis as follow; 302 cases were men and 181 cases were women, sex incidences was 2 : 1 Among the 483 cases of acute abdomen, acute appendicitis were 337 cases (69,7%), ileus were 59 cases (12.1%)peptic ulcer perforation 24 cases (5.0%) biliary disease 42 cases(8. 6%) and accidental trauma 12 cases(2. 5%) The mortality rate mainly dependent on the length of the time from the onsest to laparotomy. The mortality rate of intestinal obstruction are 6.8% and peptic ulcer 4,1% and our series showed high incidence of mortality in the intestinal obstruction. We think above high mortality caused by delayed Laparotomy.

감초추출물의 지방세포와 조골세포에 대한 분화효과

서초롱(Cho-Rong Seo),변종선(Jong Seon Byun),안재진(Jae Jin An),이재환(JaeHwan Lee),홍정우(Joung-Woo Hong),장상호(Sang Ho Jang),박계원(Kye Won Park) 한국식품영양과학회 2013 한국식품영양과학회지 Vol.42 No.7

창과감초(Glycyrrhiza inflata Batal)는 한약재의 조화를 돕고 해독, 항염증, 항궤양 등의 약리작용으로 한방에서 널리 이용되는 약용식물이다. 본 연구에서는 지방세포와 조골세포에서 감초의 생리활성을 확인하고자 감초에탄올추출물(GBE)을 이용하여 세포분화 촉진여부를 조사하였다. GBE의 세포독성여부를 통해 안전하다고 확인된 1~30 μg/mL의 농도 내에서 실험이 진행되었고 지방세포 분화조건에서 다분화능 세포 C3H10T1/2과 지방전구세포 3T3-L1의 Oil Red O 염색을 통해 GBE의 지방세포분화 촉진효과를 확인할 수 있었다. 또한 지방세포의 핵심전사조절인자인 PPARγ와 그의 표지유전자 aP2, AdipoQ, C/EBPα의 발현량 증가를 통해 GBE의 지방세포분화 촉진효과를 재확인하였다. 이와 일관된 결과로서 조골세포 분화조건에서 다분화능 세포 C3H10T1/2과 조골전구세포 MC3T3-E1의 ALP 염색을 통해 GBE의 조골세포분화 촉진효과를 확인하였고 조골세포 표지유전자인 ALP, RUNX2, osterix, collagen의 발현량 증가를 통해 GBE의 조골세포분화 촉진효과를 재확인하였다. 이에 따라 본 연구에서는 GBE의 지방세포와 조골세포 분화효과를 매개하는 감초의 구성성분을 조사하기 위해 GA(glycyrrhizic acid)와 LA(licochalcone A)의 분화촉진 여부를 확인한 결과, GA는 영향을 주지 않으나 LA가 GBE의 세포분화효과를 매개한다고 사료된다. 따라서 본 연구에서는 제2형 당뇨와 그에 수반되는 골질환과 골다공증에 대한 치료 소재로서 GBE와 그의 생리활성을 매개할 수 있는 LA의 가능성을 보았으며 GBE에서 분리되는 다양한 화합물의 동정 및 생리활성 효과, LA와의 상승효과 등의 추후 연구가 기대된다. Glycyrrhiza inflata Batal, an important species of licorice, is one of the most widely used medicinal plants for over 4000 years. Glycyrrhiza plant species has been well known for its various therapeutic activities such as anti-inflammatory, anti-allergic, and anti-ulcer. The purpose of this study was to determine the effects of Glycyrrhiza inflata Batal ethanol extracts (GBE) on adipocyte and osteoblast differentiation. Mesenchymal C3H10T1/2 cells were treated with sub-cytotoxic doses of GBE, and its effects on adipocyte differentiation were assessed. We found that GBE dose-dependently increased lipid accumulation and also induced the expression of adipocyte markers, such as PPARγ and its target genes, aP2, and adiponectin, in C3H10T1/2 cells. Consistently, similar effects of GBE on lipid accumulation were also observed in preadipocyte 3T3-L1 cells that further supports the pro-adipogenic activities of GBE. We also investigated the effects of GBE on osteoblast differentiation of mesenchymal C3H10T1/2 cells. As a results, we found that GBE increased the activity of alkaline phosphatase in a dose-dependent manner and also promoted the expression of osteoblast markers, such as ALP and RUNX2, during osteoblast differentiation of C3H10T1/2 cells. Similar pro-osteogenic effects of GBE were also observed in preosteoblast MC3T3-E1 cells. Finally, our data show that a major bioactive compound found in Glycyrrhiza inflata Batal, licochalcone A (LA) but not glycyrrhizic acid (GA), can mediate the pro-adipogenic and pro-osteogenic effects of GBE. Taken together, this study provides data to show the possibility of GBE and its bioactive component LA as putative strategies for type 2 diabetes and bone diseases.