CAJ | 학술논문

目的利用神经元损伤模型和蛋白双向凝胶电泳技术研究亚低温脑保护机制.方法细胞裂解液浸融法建立神经元损伤模型,分别置于33℃和37℃温控培养箱中培养,测定培养上清液中乳酸脱氢酶含量,TUNEL细胞凋亡法比较两种培养温度下细胞凋亡情况.提取细胞总蛋白,蛋白双向凝胶电泳技术筛选差异蛋白质点,质谱分析法完成蛋白鉴定,Western blot方法进行蛋白质确认,统计学方法解析检测结果.结果处于亚低温培养条件下的受损神经细胞乳酸脱氢酶水平和发生原位凋亡的细胞数量均明显低于37℃培养组(p＜0.05).利用双向凝胶电泳共筛选出14个差异蛋白质点,其中12个得到有效认证,它们参与细胞能量代谢、再生、氧化应激等多种病理生理过程.结论亚低温通过干预调节细胞能量代谢、再生、氧化应激等多个分子事件发挥肯定的神经保护作用.
목적 이용신경원손상모형화단백쌍향응효전영기술연구아저온뇌보호궤제.방법 세포렬해액침융법건립신경원손상모형,분별치우33℃화37℃온공배양상중배양,측정배양상청액중유산탈경매함량,TUNEL세포조망법비교량충배양온도하세포조망정황.제취세포총단백,단백쌍향응효전영기술사선차이단백질점,질보분석법완성단백감정,Western blot방법진행단백질학인,통계학방법해석검측결과.결과 처우아저온배양조건하적수손신경세포유산탈경매수평화발생원위조망적세포수량균명현저우37℃배양조(p＜0.05).이용쌍향응효전영공사선출14개차이단백질점,기중12개득도유효인증,타문삼여세포능량대사、재생、양화응격등다충병리생리과정.결론 아저온통과간예조절세포능량대사、재생、양화응격등다개분자사건발휘긍정적신경보호작용.
Objective To study the neuroprotection mechanism of mild hypothermia using a neuronal injury model and protein two-dimensional gel electrophoresis.Methods A neuron damage model was induced by the cell lysates soak assay.Neurons were cultured in temperature-controlled incubator at 33 ℃ and 37 ℃ respectively.The lactic acid dehydrogenase level in the culture supernatant was tested.TUNEL apoptosis assay was use to compare the cell apoptosis under the 2 culture temperatures.The total cellular protein was extracted.The protein two-dimensional gel electrophoresis technique was used to screen the differential protein spots.Mass spectrometry was used to complete the protein identification.Western blot was used to conduct protein confirmation.Statistical methods were used to analyze the test results.Results The impaired neuronal lactate dehydrogenase level and the cell numbers of occurring in situ apoptosis were significantly lower than those of the 37 ℃ culture group under the low temperature condition (P ＜ 0.05).A total of 14 differential protein spots were screened using the two-dimensional gel electrophoresis,12 of them were identified effectively.They were involved in a variety of pathophysiological processes,including cellular energy metabolism,regeneration,and oxidative stress.Conclusion Mild hypothermia plays a certain neuroprotective effect by intervening and regulating multiple molecular events,including cellular energy metabolism,regeneration,and oxidative stress.