CAJ | 학술논문

目的评价海马神经细胞线粒体通透性转换孔(mPTP)在富氢液减轻大鼠全脑缺血再灌注损伤中的作用.方法雄性SD大鼠72只,体重250 ～ 300 g,采用随机数字表法,将其随机分为6组(n=12):假手术组(S组)、缺血再灌注组(IR组)、生理盐水组(NS组)、富氢液组(H组)、苍术苷组(A组)和富氢液+苍术苷组(HA组).采用四血管阻塞法建立大鼠全脑缺血再灌注模型,缺血15 min后恢复灌注.H组和HA组于再灌注即刻腹腔注射富氢液5 ml/kg,其余组腹腔注射等容量生理盐水；A组和HA组于再灌注前10 min行侧脑室注射苍术苷15 μl,NS组和H组侧脑室注射等容量生理盐水.再灌注24h时行神经行为学损伤评分后各组随机处死8只大鼠,迅速断头,分离海马神经细胞线粒体,采用分光光度计法测定mPTP的开放程度,Rhodamine123法测定线粒体膜电位.再灌注72 h时各组处死4只大鼠,取海马组织,光镜下观察CA1区病理学结果,计数该区神经细胞存活数.结果与S组比较,其余组再灌注24h时行为学损伤加重,mPTP活性升高,线粒体膜电位降低(P＜0.05)；与IR组比较,H组和HA组再灌注24h时行为学损伤减轻,mPTP活性降低,线粒体膜电位升高(P＜0.05)；与H组比较,HA组行为学损伤加重,mPTP活性升高,线粒体膜电位降低(P＜0.05).再灌注72 h时HA组较IR组神经细胞存活数增加(P＜0.05),H组海马CA1区神经元损伤较IR组、NS组、A组和HA组减轻.结论富氢液可减轻大鼠全脑缺血再灌注损伤,其机制与抑制海马神经细胞mPTP开放,减少线粒体膜电位降低,从而维持线粒体功能有关.
목적 평개해마신경세포선립체통투성전환공(mPTP)재부경액감경대서전뇌결혈재관주손상중적작용.방법 웅성SD대서72지,체중250 ～ 300 g,채용수궤수자표법,장기수궤분위6조(n=12):가수술조(S조)、결혈재관주조(IR조)、생리염수조(NS조)、부경액조(H조)、창술감조(A조)화부경액+창술감조(HA조).채용사혈관조새법건립대서전뇌결혈재관주모형,결혈15 min후회복관주.H조화HA조우재관주즉각복강주사부경액5 ml/kg,기여조복강주사등용량생리염수；A조화HA조우재관주전10 min행측뇌실주사창술감15 μl,NS조화H조측뇌실주사등용량생리염수.재관주24h시행신경행위학손상평분후각조수궤처사8지대서,신속단두,분리해마신경세포선립체,채용분광광도계법측정mPTP적개방정도,Rhodamine123법측정선립체막전위.재관주72 h시각조처사4지대서,취해마조직,광경하관찰CA1구병이학결과,계수해구신경세포존활수.결과 여S조비교,기여조재관주24h시행위학손상가중,mPTP활성승고,선립체막전위강저(P＜0.05)；여IR조비교,H조화HA조재관주24h시행위학손상감경,mPTP활성강저,선립체막전위승고(P＜0.05)；여H조비교,HA조행위학손상가중,mPTP활성승고,선립체막전위강저(P＜0.05).재관주72 h시HA조교IR조신경세포존활수증가(P＜0.05),H조해마CA1구신경원손상교IR조、NS조、A조화HA조감경.결론 부경액가감경대서전뇌결혈재관주손상,기궤제여억제해마신경세포mPTP개방,감소선립체막전위강저,종이유지선립체공능유관.
Objective To investigate the role of mitochondrial permeability transition pore (mPTP) of hippocampal neurons in process of hydrogen-rich saline attenuating global cerebral ischemia-reperfusion (I/R) injury in rats.Methods Seventy-two male Sprague Dawley rats,weighing 250-300 g,were randomly divided into six groups ( n =12 each):sham operation group (group S),cerebral ischemia-reperfusion group (group IR),normal saline group (group NS),hydrogen-rich saline group (group H),atractyloside group (group A) and hydrogen-rich saline + atractyloside group (group HA).Global cerebral I/R injury was produced by four-vessel occlusion method.Bilateral vertebral arteries were cauterized.Then bilateral common carotid arteries were occluded for 15min and followed by reperfusion.In groups H and HA,hydrogen-rich saline 5 ml/kg was injected intraperitoneally immediately after reperfusion,while equal volume of normal saline was injected in the other four groups.The rats in groups A and HA received intracerebroventricular injection of atractyloside 15 μl 10 min before reperfusion,while groups NS and H received intracerebroventricular injection of equal volume of normal saline.After the neurological behavior was evaluated at 24 h of reperfusion,8 rats in each group were sacrificed and the hippocampi were immediately isolated and homogenized followed by density gradient centrifugation.The opening degree of mPTP was assayed with spectrophotometry and the mitochondrial membrane potential (MMP) was detected with Rhodamine 123 method.Four rats in each group were killed at 72 h of reperfusion and the brains were removed for microscopic examination of the area CA1 of the hippocampus and determination of the number of normal pyramidal neurons.Results Compared with group S,the neurological behavior was compromised,MMP was decreased and mPTP opening degree was enhanced in the other five groups ( P ＜ 0.05).The neurological behavior was better,MMP was increased and mPTP opening degree was decreased in groups H and HA as compared with group IR ( P ＜ 0.05).Compared with group H,the neurological behavior was compromised,MMP was decreased and mPTP opening degree was enhanced in group HA ( P ＜ 0.05).Compared with group IR,the number of normal pyramidal neurons at 72 h of reperfusion in the CA1 region of the hippocampus was higher in group HA ( P ＜0.05).The injury of the CA1 region of the hippocampus at 72 h of reperfusion was attenuated in group H as compared with groups IR,NS,A and HA.Conclusion Hydrogen-rich saline can attenuate global cerebral I/R injury throngh inhibiting the mPTP opening and reducing the dissipation of MMP,thus maintaining the mitochondrial function.