莱菔硫烷抑制急性一氧化碳中毒大鼠脑组织神经细胞凋亡的分子机制研究  

The molecular mechanism study of Sulforaphane inhibiting neurocellular apoptosis in brain tissue of rats with acute carbon monoxide poisoning

作　　者：岳傲春 宋慧平[3] 周诩栋 姬忠良 韩伟 李琴 Yue Aochun;Song Huiping;Zhou Xudong;Ji Zhongliang;Han Wei;Li Qin(Department of Emergency,Shenzhen University General Hospital,Shenzhen 518055,Guangdong,China;Integrative Medicine Center,School of Medicine,Qingdao University,Qingdao 266071,Shandong,China;Department of Integrated Traditional Chinese and Western Medicine,the First Clinical Hospital of Shandong University of Traditional Chinese Medicine,Jinan 250399,Shandong,China)

机构地区：[1]深圳大学总医院急诊科,广东深圳518055 [2]青岛大学医学部中西医结合中心,山东青岛266071 [3]山东中医药大学第一临床医院,山东济南250399

出　　处：《中国中西医结合急救杂志》2024年第6期714-719,共6页Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care

基　　金：国家自然科学基金(82071465);广东省深圳市医疗卫生三名工程项目(SZSM201911007);山东省自然科学基金(ZR2020MH154);医学救援关键技术装备应急管理部重点实验室项目(YJBKFKT202416)。

摘　　要：目的观察莱菔硫烷(SFN)对急性一氧化碳中毒(ACOP)大鼠脑组织凋亡相关蛋白天冬氨酸特异性半胱氨酸蛋白酶(caspase-3,caspase-9)表达的影响,探讨其干预治疗ACOP脑损伤的分子机制。方法按随机数字表法将健康雄性SD大鼠分为正常对照组(NC组)、ACOP组、SFN组,每组36只。使用高压氧舱暴露大鼠于一氧化碳(CO)以建立ACOP动物模型;NC组则自由呼吸新鲜空气。SFN组于中毒后2 h内腹腔注射SFN 20 mg/kg进行干预,每日1次,直到处死;NC组和ACOP组注射等量生理盐水。各组干预1、3、7 d处死大鼠取脑组织,苏木素-伊红(HE)染色检测脑组织病理学损伤情况;尼氏染色检测脑组织神经元病理学改变;采用免疫组化法检测脑组织皮质区caspase-3、caspase-9的阳性表达;采用蛋白质免疫印迹试验(Western blotting)和荧光定量反转录-聚合酶链反应(qRT-PCR)分别检测脑组织中caspase-3、caspase-9的蛋白及mRNA表达。结果CO染毒后,ACOP组脑组织损伤逐渐加重,尼氏小体数量逐渐减少,脑组织皮质区caspase-3、caspase-9阳性细胞数量逐渐增多,脑组织中caspase-3、caspase-9的蛋白及mRNA表达逐渐升高,与同一时间点NC组比较,差异均有统计学意义〔尼氏小体(个):69.33±0.94比91.33±1.25,caspase-3阳性表达(A值):0.149±0.003比0.113±0.004,caspase-9阳性表达(A值):0.178±0.002比0.111±0.010,caspase-3蛋白表达(caspase-3/GAPDH):1.634±0.045比0.844±0.021,caspase-9蛋白表达(caspase-9/GAPDH):1.754±0.024比0.811±0.053,caspase-3 mRNA(2-ΔΔCt):1.718±0.052比1,caspase-3 mRNA(2-ΔΔCt):1.722±0.066比1,均P<0.05〕。与同时间点ACOP组相比,SFN组大鼠脑组织损伤程度得到改善,尼氏小体数量明显增加(个:84.67±1.53比69.33±0.94,P<0.05);脑组织皮质区caspase-3、caspase-9阳性细胞数量明显减少(A值:0.126±0.002比0.149±0.003,0.127±0.002比0.178±0.002,均P<0.05),脑组织中caspase-3、caspase-9的蛋白及mRNA表达明显降低〔caspase-3蛋白表达(caspasObjective To investigate the effect of sulforaphane(SFN)on the expression of apoptosis-related proteins(caspase-3 and caspase-9),in brain tissue of rats with acute carbon monoxide poisoning(ACOP),and to explore the molecular mechanism underlying its intervention in ACOP-induced brain injury.Methods The healthy male Sprague-Dawley(SD)rats were randomly assigned to three groups:normal control(NC)group,ACOP group,and SFN group,with 36 rats in each group.An ACOP animal model was established by exposing the rats to carbon monoxide(CO)in a hyperbaric oxygen chamber,while the rats in the NC group were allowed to breathe fresh air.The SFN group received an intraperitoneal injection of SFN 20 mg/kg within 2 hours after poisoning,once daily,until euthanasia.The NC and ACOP groups were injected with an equivalent volume of saline.Rats from each group were sacrificed on days 1,3,and 7 of the intervention to collect brain tissue,hematoxylin-eosin(HE)staining was performed to assess pathological damage in the brain tissue;Nissl staining was used to examine neuronal pathological changes;Immunohistochemistry was employed to detect the positive expression of caspase-3 and caspase-9 in the cortical region of the brain.Western blotting and quantitative reverse transcription-polymerase chain reaction(qRT-PCR)were conducted to measure the protein and mRNA expression of caspase-3 and caspase-9 in the brain tissue.Results After CO poisoning,brain tissue damage in the ACOP group progressively worsened,with a gradual decrease in the number of Nissl bodies and a gradual increase in the number of positive cells for caspase-3 and caspase-9 in the cortical region of the brain.The protein and mRNA expression of caspase-3 and caspase-9 in the brain tissue also gradually increased.Compared with the NC group at the same time point,the differences were statistically significant[Nissl bodies:69.33±0.94 vs.91.33±1.25;caspase-3 positive expression(A value):0.149±0.003 vs.0.113±0.004;caspase-9 positive expression(A value):0.178±0.002 vs.0.111±0

关 键 词：莱菔硫烷 急性一氧化碳中毒 细胞凋亡 凋亡相关蛋白 

分 类 号：R59[医药卫生—内科学]