新生小鼠炎症-缺氧缺血脑损伤模型的建立  

Establishment and evaluation of inflammation-sensitized hypoxic-ischemic injury in the immature brains

作　　者：朱将虎[1] 朱敏丽[1] 胡莹莹[1] 方明楚 盛汉松[2] 林振浪[1] Zhu Jianghu;Zhu Minli;Hu Yingying;Fang Mingchu;Sheng Hansong;Lin Zhenlang(The Second Affiliated Hospital of Wenzhou Medical University,Wenzhou 325027,China;Department of Neurosurgery,the Second Affiliated Hospital of Wenzhou Medical University,Wenzhou 325027,China)

机构地区：[1]温州医科大学附属第二医院,325027 [2]温州医科大学附属第二医院神经外科,325027

出　　处：《中华实验外科杂志》2019年第6期1136-1139,共4页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金(81701489、81771624);温州市科技局项目(Y20160020).

摘　　要：目的建立新生小鼠炎症-缺氧缺血脑损伤模型。方法本研究采用3d龄C57BL/6小鼠,每组有3个独立检测指标,每个实验指标各6只,共144只。3d龄新生小鼠腹腔内分别注射0.05mg/kg、0.50mg/kg的脂多糖(LPS),24h后检测小鼠体重增长情况,2,3,5-氯化三苯基四氮唑(TTC)染色观察脑梗死情况,苏木精-伊红(HE)染色观察脑组织病理情况,以及末端脱氧核苷酸转移酶介导的脱氧三磷酸尿苷缺口末端标记法(TUNEL)检测脑组织细胞凋亡指数,根据结果确定一个对小鼠损伤不明显的剂量(即亚损伤剂量)。3d龄新生小鼠结扎左侧颈总动脉后,置于8%O2缺氧舱分别缺氧(HI)20、40min,24h后检测小鼠体重、脑组织TTC、HE以及TUNEL染色,根据结果确定一个对小鼠脑损伤不明显的时间(即亚损伤时间)。最后,3d龄新生小鼠分为两组:实验组(LPS+HI组)及对照组(Sham组)。根据上述实验得出的结果,实验组小鼠予亚损伤剂量的LPS,2h后再给予亚损伤的缺氧缺血时间;对照组给予磷酸盐缓冲液(PBS)腹腔内注射,2h后假手术处理且不缺氧。24h后检测小鼠体重,脑组织TTC、HE以及TUNEL染色。应用SPSS 23.0统计软件分析,计量资料以均值±标准差(Mean±SD)表示,多组间比较采用单因素方差分析(ANOVA),方差齐性资料两两比较用SNK检验;方差不齐资料两两比较用Dunnett’s T3检验。两组间比较采用t检验。结果LPS 0.05mg/kg组体重[(2.81±0.14)g]与PBS组[(2.81±0.16)g]比较,差异无统计学意义(P>0.05),而LPS 0.50mg/kg组体重[(2.63±0.09)g]较PBS组轻,差异有统计学意义(P<0.05)。HI 20组新生小鼠未见明显脑梗死及少量细胞凋亡[(2.80±1.74)%],HI 40组新生小鼠大脑出现脑梗死灶及大量的细胞凋亡[(55.67±7.81)%]。最后,实验组(LPS+HI组)体重[(2.66±0.09)g]较Sham组[(2.83±0.12)g]轻,差异有统计学意义(t=2.565,P<0.05),同时LPS+HI组TTC染色显示明显脑梗死灶,HE染色显示神经细胞病理损伤形态,TUNEL法显示�Objective To establish a neonatal mouse model mimicking pathological changes of inflammation-sensitized hypoxic-ischemic brain injury in human preterm infants. Methods First, postnatal day 3 (P3) neonatal mice were randomly assigned into 3 different groups: phosphate buffer (PBS) group (injected intraperitoneally with PBS), lipopolysaccharide (LPS) 0.05 group (injected intraperitoneally with LPS 0.05 mg/kg), and LPS 0.50 mg/kg group (injected intraperitoneally with LPS 0.50 mg/kg). At 24 h after the LPS insult, the weight of mice was measured, and their brains were harvested: 2, 3, 5-triphenyltetrazoulium hydrochloride (TTC) staining was employed for detecting the brain infarct, hematoxylin and eosin (HE) staining for observing neural necrosis, and the terminal dexynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method for detecting cell apoptosis. We chose the dosage of LPS with little injury on the neonatal mice for further study. Second, P3 neonatal mice were randomly assigned into 3 different groups: Sham group [without hypox (HI) insult], HI 20 group (exposed to HI for 20 min), and HI 40 group (exposed to HI for 40 min). At 24 h after the HI insult, the weight of mice was measured, and their brains were harvested for TTC staining, HE staining, and TUNEL method. We chose the time of HI with little injury on the neonatal mice for further study. Last, P3 neonatal mice were randomly assigned into 2 different groups: Sham group (PBS injected without HI), and LPS+ HI group (LPS injected 2 hours before HI). We chose the dosage of LPS and the time of HI which had little injury on the neonatal mice for the study. At 24 h after the HI insult, the weight of mice was measured, and their brains were harvested for TTC staining, HE staining, and TUNEL method. Results First, LPS 0.5 group had lower body weight [(2.63±0.09) g] than both LPS 0.05 group [(2.81±0.14) g] and PBS group [(2.81±0.16) g], so we chose LPS 0.05 mg/kg for further study. Second, HI 40 group had significant brain infarct detected by TT

关 键 词：早产儿 炎症 缺氧缺血 脑损伤 

分 类 号：R722.1[医药卫生—儿科] R-332[医药卫生—临床医学]