학술논문
빌리루빈의 신경독성에 대한 MK-801의 신경학적 방어효과에 관한 연구
Neuroprotective Effects of MK-801 (Dizocilpine) on Bilirubin Neurotoxicity in Mouse Cortical Cell Culture
Neuroprotective Effects of MK-801 (Dizocilpine) on Bilirubin Neurotoxicity in Mouse Cortical Cell Culture
Document Type
Article
Author
Source
PERINATOLOGY (구 대한주산의학회잡지). Mar 25, 2004 15(1):34
Subject
Language
Korean
ISSN
2508-4887
Abstract
목적 : 일차배양을 통해 얻어진 마우스 태아의 대뇌피질세포를 이용하여 빌리루빈 세포독성에 대한 N-methyl-D-aspartate (NMDA) 수용체 길항제인 MK-801의 신경학적 방어효과에 대해 lactate dehydro genase (LDH) 활성도를 측정하여 정량적으로 평가하고자 하였다. 방법 : 14일 된 마우스 태아로부터 대뇌피질을 적출하여 일차배양을 통해 대뇌피질세포들을 얻었다. 100 μM 빌리루빈과 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohept-5,10-imine (MK-801, dizocilpine), 1,2,3,4-tetrahydroxy-6-nitro-2,3-dioxo-benzo [f] quinoxaline-7-sulfonamide disodium (NBQX), NG-nitro-L-arginine methyl ester hydrochloride (L-NAME)을 각각 혼합한 배지들을 만들어 여기에 대뇌피질세포들을 4시간 동안 노출시켰다. 그 후 각 배양액의 상층부를 얻어 대뇌피질세포로부터 배양액 내로 유리된 LDH의 활성도를 측정하여 빌리루빈에 의한 세포독성을 평가하였으며 형광염색 후 공초점 현미경으로 죽은 세포들을 관찰하였다. 결과 : 빌리루빈에만 노출된 세포와 MK-801, NBQX, L-NAME 등을 혼합한 세포에서 LDH 활성도로 평가한 세포독성은 각각 55±7%, 32±9%, 52±8%, 53±9%였으며 MK-801로 처리한 대뇌피질세포에서 세포독성이 가장 유의하게 감소하였다( p<0.05). 또한 공초점 현미경 상에서도 MK-801로 처리한 대뇌피질세포에서 죽은 세포가 가장 적게 관찰되었다. 결론 : 일차배양한 마우스 태아의 대뇌피질세포에서 빌리루빈의 흥분독성은 NMDA 수용체에 의해서 일어나며 비경쟁적 길항제인 MK-801은 신경세포의 손상을 감소시킬 수 있었다.
Objective : To evaluate whether MK-801, N-methyl-D-aspartate (NMDA) receptor antagonist, could protect against bilirubin neurotoxicity in mouse neuronal cells. Methods : Mouse cerebral cortical cells were obtained from 14 day-old mouse fetal cerebral cortex primary culture. Cerebral cortical cells were exposed to medium containing concentrations of bilirubin 100 μM and additionally treated with 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohept-5,10-imine (MK-801, dizocilpine), 1,2,3,4-tetrahydroxy-6-nitro-2,3-dioxo-benzo [f] quinoxaline-7-sulfonamide disodium (NBQX), NG-nitro-L-arginine methyl ester hydrochloride (L-NAME) for 4 hours. Then, the bilirubin cytotoxicity for cerebral cortical cells was quantitated by the activity of LDH released from cerebral cortical cells into the culture media. Cortical cells were stained with propidium iodide and visualized by confocal laser scanning microscopy for detection of apoptotic cells. Results : After exposure for 4 hours, the cytotoxicity for cortical cells was the least in bilirubin treated with MK-801 compared to bilirubin alone, bilirubin treated with NBQX, and L-NAME (32±9%, 55±7%, 52±8%, 53±9%, respectively, p<0.05). Cells treated with MK-801 had fewer apoptotic nuclei evident on confocal microscopy. Conclusion : Our study suggests bilirubin neurotoxicity is mediated NMDA receptors and MK-801 is capable to prevent bilirubin-induced neuronal damage.
Objective : To evaluate whether MK-801, N-methyl-D-aspartate (NMDA) receptor antagonist, could protect against bilirubin neurotoxicity in mouse neuronal cells. Methods : Mouse cerebral cortical cells were obtained from 14 day-old mouse fetal cerebral cortex primary culture. Cerebral cortical cells were exposed to medium containing concentrations of bilirubin 100 μM and additionally treated with 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohept-5,10-imine (MK-801, dizocilpine), 1,2,3,4-tetrahydroxy-6-nitro-2,3-dioxo-benzo [f] quinoxaline-7-sulfonamide disodium (NBQX), NG-nitro-L-arginine methyl ester hydrochloride (L-NAME) for 4 hours. Then, the bilirubin cytotoxicity for cerebral cortical cells was quantitated by the activity of LDH released from cerebral cortical cells into the culture media. Cortical cells were stained with propidium iodide and visualized by confocal laser scanning microscopy for detection of apoptotic cells. Results : After exposure for 4 hours, the cytotoxicity for cortical cells was the least in bilirubin treated with MK-801 compared to bilirubin alone, bilirubin treated with NBQX, and L-NAME (32±9%, 55±7%, 52±8%, 53±9%, respectively, p<0.05). Cells treated with MK-801 had fewer apoptotic nuclei evident on confocal microscopy. Conclusion : Our study suggests bilirubin neurotoxicity is mediated NMDA receptors and MK-801 is capable to prevent bilirubin-induced neuronal damage.