Differential Expressions of Aquaporin Subtypes in the Adult Mouse Testis

배혜란,임지우,김동환,Elsayed A. Mohamed 한국발생생물학회 2022 발생과 생식 Vol.26 No.2

Many efforts have been made to study the expression of aquaporins (AQP) in the mammalian reproductive system, but there are not enough data available regarding their localized expression to fully understand their specific roles in male reproduction. The present study investigated the expression and localization patterns of different AQP subtypes in the adult mouse testes and testicular spermatozoa using an immunofluorescence assay. All the studied AQPs were expressed in the testes and revealed subtype-specific patterns in the intensity and localization depending on the cell types of the testes. AQP7 was the most abundant and intensive AQP subtype in the seminiferous tubules, expressing in Leydig cells and Sertoli cells as well as all stages of germ cells, especially the spermatids and testicular spermatozoa. The expression pattern of AQP3 was similar to that of AQP7, but with higher expression in the basal and lower adluminal compartments rather than the upper adluminalcompartment. AQP8 expression was limited to the spermatogonia and Leydig cells whereas AQP9 expression was exclusive to tails of the testicular spermatozoa and elongated spermatids. Taken together, the abundance and distribution of the AQPs across the different cell types in the testes indicating to their relavance in spermatogenesis, as well as in sperm maturation, transition, and function.

HGF prevents H202-induced cell death via Erk-dependent pathway in MDCK cells

배혜란 ( Bae Hye Lan ),나서희 ( Na Seo Hui ),안원석 ( An Won Seog ),김성은 ( Kim Seong Eun ),김기현 ( Kim Gi Hyeon ) 대한신장학회 2003 춘계학술대회 초록집 Vol.22 No.1

미성숙 희쥐 해마에서 열성경련에 의한 신경흥분성의 증가

최봉근,배혜란*,황규근 대한소아청소년과학회 2004 Clinical and Experimental Pediatrics (CEP) Vol.47 No.9

Purpose:To determine whether febrile seizure enhances neuroexcitability by altering synaptic transmission and whether febrile seizure-induced hyperexcitability leads to long-lasting neuronal death. Methods:We investigated the expression of synaptic and postsynaptic proteins and the apoptosis of neuronal cells in rat pup hippocampus after hyperthermic seizure using immunoblotting and confocal microscopy. Results:Hyperthermic seizure enhanced the long-term expressions of presynaptic proteins such as syntaxin, VAMP, SNAP-25 and nSec1, whereas that of NSF was decreased. The expressions of postsynaptic NMDA receptors 1, 2a and 2b were up-regulated. The expression of postsynaptic AMPA glutamate receptors 1 month after hyperthermic seizures altered by way of increasing the ratio of GluR1 to GluR2 and decreasing NSF-GluR2 interaction, which leads to the formation of Ca2+ permeable AMPA receptors and enhanced toxicity. However, in spite of enhanced neuroexcitability, there was a transient increase of neuronal death in hipocampus one week after hyperthermic seizure, but returned to baseline one month later. Conclusion:These results demonstrate both presynaptic and postsynaptic forms of long-term enhancement of glutamate synaptic transmission after hyperthermic seizure and support the idea that early-life febrile seizure might have persistent effects on neuronal excitability in the hippocampus. 목 적:열성경련은 영유아에서 가장 흔한 경련이다. 열성경련이 지속적인 신경 흥분성을 유발하는지와 열성경련이 측두엽 간질과 연관성이 있는지가 중요한 관심사이다. 이에 미성숙 쥐 모델을 이용하여 열성경련이 해마의 시냅스 단백질 및 glutamate 수용체 발현에 미치는 효과를 관찰하여 신경 흥분성과 신경세포 손상을 유발하는지 알아보고자 이 연구를 시도하였다. 방 법:실험동물로는 생후 10일된 Sprague-Dawley계의 rat pups 흰쥐를 사용하였다. 열성경련은 더운물 (47℃)로 유발시켰다. 경련 유발 후 생후 1일째, 1주째, 1개월째 뇌를 적출하여 western blot를 이용하여 해마에서 시냅스 단백의 발현을 조사하였으며 tunel study로 해마 신경세포의 세포사를 관찰하였다. 결 과:열성경련을 유발시킨 쥐의 해마에서 고체온만 유발시킨 대조군에 비해 신경 연접부 세포막과 연접소포의 융합에 관여하는 시냅스 단백질인 VAMP, SNAP-25, syntaxin 및 nSec1의 발현이 경련 한달 후까지 증가되었으나, NSF의 발현은 감소되었다. 열성경련 후 시냅스 후 신경막에 있는 NMDA 수용체 1, 2a 및 2b의 발현은 증가하였다. AMPA 수용체 중 GluR1의 발현은 열성경련 후 증가된 반면 GluR2의 발현은 감소되었다. 시냅스 후 신경막의 GluR2의 수를 증가시키는 것으로 알려진 NSF와 GluR2의 수를 증가시키는 것으로 알려진 NSF와 GluR2 복합체의 형성이 열성경련 한 달 후까지 현저히 감소되어 있는 것을 관찰하였다. TUNEL 염색으로 관찰한 신경세포의 손상은 열성경련 일주일 후에는 유의한 증가를 보였으나 한 달 후에는 유의한 차이가 없었다 결 론:열성경련은 시냅스 전 시냅스단백질의 발현을 증가시키고, 시냅스 후 신경막에서 GluR1/GluR2 발현비를 증가시킬 뿐만 아니라, GluR2와 NSF의 결합을 감소시킴으로써 Ca2+ permeable AMPA 수용체 형성을 증가시킴으로써 신경흥분성이 유발되는 것으로 보인다.

간세포성장인자가 메산지움 세포의 TGF-β, Fibronectin, Collagen Ia 생성에 미치는 영향

안원석,배혜란,나서희,김성은,김기현 대한신장학회 2004 Kidney Research and Clinical Practice Vol.23 No.6

장시간 탈진운동 시 골격근 수축 및 에너지대사에서 AQP3의 역할

김미자,배혜란,김동환,윤미연,박성철,소용석,김영준 한국운동생리학회(구 한국운동과학회) 2013 운동과학 Vol.22 No.4

본 연구는 AQP3 -/-(AQP3 유전자결핍생쥐)와 AQP3 +/+(야생형생쥐)에서 장시간 중강도 탈진운동을 실시하여 운동 수행능력의 차이를 조사하고 글리세롤과 관련된 운동에너지원의 혈장 및 조직 내 농도 변화를 분석하였다. AQP3-/-와 AQP3+/+ 각각 16마리를 대상으로 트레드밀을 이용하여 10° 경사에서 최대 12 m/min 속도까지 증가시키면서 탈진할 때까지 장시간 운동을 실시하였다. AQP3 -/-의 조직학적 차이와 AQP3 단백의 세포내 위치를 알아보기 위해 넙다리뒤근, 긴발가락폄근 및 가자미근을 적출하여 면역조직화학 및 형광염색을 실시하였고, 장시간 탈진운동 후 운동에너지원의 변화를 분석하기 위해 혈장 및 조직 내 포도당, 당원, 중성지방 및 글리세롤 농도를 측정하였다. AQP3 -/-은 AQP3 +/+에 비해 넙다리뒤근의 직경과 무게가 감소되어 있었으나, 조직학적 차이는 관찰되지 않았다. 운동지속시간은 AQP3 -/-가 현저히 낮았다. 운동수행 전 간, 근육 및 혈장 내 포도당, 당원, 중성지방 및 글리세롤 농도는 모두 AQP3 -/-가 유의하게 감소되어 있었으나, 근육 내 중성지방 농도는 오히려 증가되어 있었다. 장시간 탈진운동 후 간의 당원함량과 근육 내 글리세롤 농도가 AQP3 +/+에서는 감소된 반면, AQP3 -/-는 오히려 증가되었다. 이상의 결과를 종합하면 골격근에서 AQP3은 지구성 운동 시 골격근 내 지방의 분해로 생성되는 글리세롤의 유출을 촉진시킴으로써 간에서 글리세롤을 이용한 당신생과정을 통해 지속적인 운동에너지원의 공급에 기여하여 운동 수행능력에 영향을 주는 것으로 사료된다. The purpose of this study was to investigate the difference in the capacity to perform a single bout of exhausting exercise between AQP3 -/-(AQP3 knock-out mice) and AQP3 +/+(wild-type mice), and to compare the concentrations of glycerol and related metabolic parameters in plasma, liver and skeletal muscle after exhaustion. AQP3 -/- and AQP3 +/+ , 16 per each group, were exposed to a single bout of treadmill running at the speed up to 12 m/min with 10° incline until exhaustion. AQP3 -/- exhibited earlier fatigue and decrease in endurance exercise performance with shorter average time to exhaustion than the wild-type control. Both immunohistochemistry and double immunofluorescence staining revealed that AQP3 expressed at the cell surface. Although the longitudinal length and weight of hamstring muscle were reduced in AQP3 -/-, no significant histological difference was observed. At rest, the levels of glucose, glycogen, triglyceride, and glycerol in plasma, liver and skeletal muscle, were lower in AQP3 -/- than AQP3 +/+, intramuscular triglyceride level was higher in AQP3 -/-. After exercise, plasma and muscle glucose, muscle glycogen, and plasma and liver triglyceride levels decreased, whereas plasma and liver glycerol levels increased in both AQP3 -/- and AQP3 +/+ compared to those at rest. However, muscle glycerol as well as liver glycogen concentrations decreased after exercise in wild-type mice, but rather increased in AQP3 -/-. These findings suggest that decreased glycerol efflux from skeletal muscle in AQP3 knock-out mice might result in low exercise capacity due to the limitation in constant energy supply through hepatic gluconeogenesis from glycerol during the prolonged endurance exercise. Therefore, AQP3 may play a role in skeletal muscle fat metabolism and influence the endurance exercise capacity.

Succinate Transport in Rabbit Renal Basolateral Membrane Vesicles

김용근,배혜란,임병용,Kim, Yong-Keun,Bae, Hae-Rahn,Rhim, Byung-Yong The Korean Physiological Society 1988 대한생리학회지 Vol.22 No.2

가토 신장 피질에서 Percoll density gradient방법으로 분리한 basolateral membrane vesicle (BLMV)에서 rapid filtration technique을 이용하여 succinate의 이동 특성을 관찰하였다. $Na^+$은 succinate의 이동을 증가시켜 "overshoot"현상을 보였으며 이러한 효과는 $K^+,{\;}Li^+,{\;}Rb^+,{\;}choline$과 같은 다른 양이온들에 의해 나타나지 않았다. $Na^+$농도변화에 따른 succinate의 이동율은 sigmoid모양을 보였고, $Na^+$에 대한 Hill coefficient는 2.0이었다. soccinate의 이동은 vesicle 내부가 음전압일 때 더욱 증가되었다. BLMV에서 succinate이동은 용액내 pH변화에 따라 영향을 받았으나 brush border membrane vesicle (BBMV)에서는 영향을 받지 않았다. 동력학적 분석결과 succinate의 Km값은 $15.5{\pm}0.94{\;}{\mu}M$이었고 Vmax는 $16.22{\pm}0.25{\;}n{\;}mole/mg{\;}protein/min$이었다. succinate의 이동은 $4{\sim}5$탄소를 가진 dicarboxylate들에 의해 강력하게 억제되었으나 monocarboxylate나 다른 유기음이온들에 의해 영향을 적게 받거나 받지 않았다. succinate의 이동은 DIDS, SITS, furosemide와 같은 음이온 이동 억제제와 harmaline과 같은 $Na^+$ 이동 억제제에 의해 억제되었다. 이들 결과들은 BLMV에서 succinate는 $Na^+$에 의존하여 이동하며 다른 Krebs cycle중간 산물들과 동일한 운반기전을 이용함을 가르킨다. 또한 BLMV에서 succinate의 이동은 그 기질특이성에 있어서 다른 연구자에 의해 보고된 BBMV에서 이동특성과 유사함을 보였다. Properties of succinate transport were examined in basolaterat membrane vesicles (BLMV) isolated from rabbit renal cortex. An inwardly directed $Na^+$ gradient stimulated succinate uptake and led to a transient overshoot. $K^+,{\;}Li^+,{\;}Rb^+$ and choline could not substitute for $Na^+$ in the uptake process. The dependence of the initial uptake rate of succinate on $Na^+$ concentration exhibited sigmoidal kinetics, indicating interaction of more than one $Na^+$ with transporter Hill coefficient for $Na^+$ was calculated to be 2.0. The $Na^+-dependent$ succinate uptake was electrogenic, resulting in the transfer of positive charge across the membrane. The succinate uptake into BLMV showed a pH optimum at external pH $7.5{\sim}8.0$, whereas succinate uptake into brush border membrane vesicles (BBMV) did not depend on external pH. Kinetic analysis showed that a Na-dependent succinate uptake in BLMV occurred via a single transport system, with an apparent Km of $15.5{\pm}0.94{\;}{\mu}M$ and Vmax of $16.22{\pm}0.25{\;}nmole/mg{\;}protein/min$. Succinate uptake was strongly inhibited by $4{\sim}5$ carbon dicarboxylates, whereas monocarboxylates and other organic anions showed a little or no effect. The succinate transport system preferred dicarboxylates in trans-configuration (furmarate) over cis-dicarboxylates (maleate). Succinate uptake was inhibited by the anion transport inhibitors DIDS, SITS and furosemide, and $Na^+-coupled$ transport inhibitor harmaline. These results indicate the existence of a $Na^+-dependent$ succinate transport system in BLMV that may be shared by the other Krebs cycle intemediates. This transport system seems to be very similar to the luminal transport system for dicarboxylates.

쥐 대동맥 평활근 장력조절에 대한 Ehanol의 효과

서덕준,배혜란,정동근,이중희 대한순환기학회 2001 Korean Circulation Journal Vol.31 No.2

트레드밀 운동이 심근경색증 생쥐모델의 심기능 회복에 미치는 효과

김동환,배혜란,김미자,소용석,윤미연,장성동,김영준 한국운동생리학회(구 한국운동과학회) 2013 운동과학 Vol.22 No.2

연구는 생쥐 심근경색모델을 이용하여 다양한 강도의 유산소 운동이 심장기능 회복에 미치는 효과를 분석하여, 심근경색 후 심장기능 회복에 적절한 운동 강도 및 최소한의 운동 기간을 조사하고자 하였다. 생후 10∼12주된 CD1 수컷 생쥐의 좌전하행지를 차단하여 심근경색증을 유발시킨 후 운동을 시키지 않은 심근경색 대조 군과 심근경색 후 트레드밀을 이용하여 10도 경사에서 8 m/min(저강도), 12 m/min(중강도) 및 16 m/min(고강도)의 다양한 강도로 매일 30분씩 8주까지 운동을 부하시킨 운동군 간의 심장기능 회복 정도를 심장초음파 검사와 조직검사로 분석하였다. 운동군은 심근경색 대조군에 비해 8주 운동 훈련 후 심실벽 비후와 모세혈관 수의 증가가 관찰되었으며, 심장조직 헤마톡실린-에오신 염색 결과 10도 경사 12 m/min의 속도로 매일 30분씩 8주간 운동시킨 운동군에서 4주부터 심근경색 흉터의 크기가 유의하게 감소되었고, 심장초음파검사에서 좌심실 수축기말용적과 좌심실 내경이 감소되고 심박출량과 좌심실 구획단축 율은 증가되어 심실 수축계수가 증가되었다. 하지만 고강도 운동군은 심근경색 후 운동을 하지 않은 대조군에 비해 오히려 심근경색 흉터가 더 커지고 심기능 지수들이 악화되었으며, 저강도 운동군은 그 효과가 운동을 하지 않은 대조군과 차이가 없었다. 이상의 결과를 종합하면 심근경색 발생 후 중강도의 트레드밀 운동을 8주 이상 지속하는 것이 심장기능 회복에 가장 효과적일 것으로 사료된다. The p urpose o f t his s tud y w as t o i nvestigate t he a d equate i ntensity a nd the m inimal d uration o f a erobic e xercise t o o btain t he cardioprotective effect after myocardial infarction. Myocardial infarction model was established by occluding the left descending anterior artery in heart of CD 1 mice. Three days after myocardial infarction, mice were exercised with treadmill running with a speed of 8 m/min(low intensity), 12 m/min(moderate intensity) or 16 m/min(high intensity) at 10 degree incline 30 minutes per day for 8 weeks. Hematoxylin-eosin s tain showed significant decrease in the s ize of infarct scar and increase in the thickness of normal myocardial tissue after 8 weeks of moderate intensity exercise. However, infarct scar was increased and further thinned after 8 weeks of high intensity exercise. Echocardiography revealed that both stroke volume and fractional shortening increased while both left ventricular internal diameter and endsystolic volume decreased after 8 weeks of moderate intensity exercise. Fractional ejection, a representative of cardiac function, was also increased. However, the echocardiographic parameters either became worse after 8 weeks of high intensity exercise or were not significant changed after 8 weeks of low intensity exercise. These results suggest that treadsmill exercise of moderate intensity at least for 8 weeks after myocardial infarction has beneficial effect in recovery of cardiac function in post-infarcted patients.

Elevated Levels of α-Synuclein Oligomer in the Cerebrospinal Fluid of Drug-Naïve Patients with Parkinson’s Disease

박민정,천상명,배혜란,김상호,김재우 대한신경과학회 2011 Journal of Clinical Neurology Vol.7 No.4

Background and Purpose The detection of α-synuclein in the body fluids of patients with synucleinopathy has yielded promising but inconclusive results, in part because of conformational changes of α-synuclein in response to environmental conditions. The aim of this study was to determine the feasibility of using α-synuclein as a biological marker for Parkinson’s disease (PD). Methods Twenty-three drug-naïve patients with PD (age 62.4±12.7 years, mean±SD; 11 males)and 29 age- and sex-matched neurologic control subjects (age 60.1±16.2 years; 16 males) were recruited. The levels of oligomeric and total α-synuclein in the cerebrospinal fluid (CSF) and plasma were measured using two simultaneous enzyme-linked immunosorbent assays. Results The level of α-synuclein oligomer in the CSF of PD patients was significantly higher in PD patients than in neurological controls, but other findings (plasma α-synuclein oligomer and total α-synuclein in CSF and plasma) did not differ significantly between the two groups. When the control subjects were divided into a symptomatic control group (11 patients who complained of parkinsonian symptoms and were diagnosed with hydrocephalus and drug-induced or vascular parkinsonism) and a neurologic control group (10 normal subjects and 8 patients with diabetic ophthalmoplegia), the level of α-synuclein oligomer in the CSF was still significantly higher in PD patients than in both of the control subgroups. Conclusions These findings provide further evidence for a pathogenic role of the α-synuclein oligomer and suggest that CSF levels of α-synuclein oligomer can be a reliable marker for PD.

Selective Susceptibility of Human Dopaminergic Neural Stem Cells to Dopamine-Induced Apoptosis

전성만,천상명,배혜란,김재우,Seung U Kim 한국뇌신경과학회 2010 Experimental Neurobiology Vol.19 No.3

Dysfunctions of ubiquitin-proteasome system and toxicity of dopamine have been known as the key mechanisms in the pathogenesis of Parkinson's disease (PD) and proteasome inhibitors are widely used in experimental models of PD to reproduce cell death of dopaminergic neurons. In the present study, immortalized human neural stem cells (HB1.F3, F3) and those transfected with human aromatic acid decarboxylase gene (F3.AADC), were used to investigate the mechanism of selective dopaminergic neuronal cell death mediated by dopamine or proteasome inhibitors. Flow cytometric analysis revealed that F3.AADC was more susceptible to dopamine than parental F3 cell which does not carry dopaminergic phenotype. The dopamine-induced apoptosis was mediated by activation of caspases 3 and 9 and cleavage of PARP. Proteasome inhibitors also induced apoptosis in dose-dependent manner but there was no difference between cell types. Prolonged exposure to subtoxic dose of proteasome inhibitors further enhanced dopamine-induced apoptosis in the F3.AADC, and increased presence of alpha-synuclein and ubiquitin-positive inclusions was noted in the cytoplasm of apoptotic cells by immunocytochemistry. These findings indicate that dopaminergic cells are selectively susceptible to dopamine toxicity and prolonged suppression of proteasome system further enhances selective sensitivity to dopamine toxicity. Chronic subtoxic proteasomal dysfunction of dopaminergic cells might contribute to selective cell death of dopaminergic neurons during the pathogenesis of Parkinson's disease.