TUG-891对缺血缺氧诱导的缺血性脑卒中的保护作用及其机制

doi:10.13481/j.1671-587X.20250413

摘要/Abstract

摘要：

目的探讨TUG-891对缺血缺氧诱导的缺血性脑卒中（IS）的保护作用，并阐明其可能的作用机制。方法将60只健康雄性C57BL/6小鼠随机分为假手术组、模型组［远端大脑中动脉闭塞（dMCAO）组］和dMCAO+TUG-891组（dMCAO+TUG-891组），每组20只，造模后连续3 d给予小鼠腹腔注射TUG-891溶液（35 mg·kg^-1·d^-1）。采用改良神经功能缺损评分（mNSS）和转棒实验评估各组小鼠神经功能情况，2，3，5-氯化三苯基四氮唑（TTC）染色法观察各组小鼠脑梗死体积，生化法检测各组小鼠脑组织上清液中丙二醛（MDA）水平和超氧化物歧化酶（SOD）活性，苏木精-伊红（HE）和尼氏（NISSL）染色观察各组小鼠脑组织病理形态表现，末端脱氧核苷酸转移酶dUTP缺口末端标记法（TUNEL）染色检测各组小鼠脑组织中神经细胞凋亡指数，Western blotting法检测各组小鼠脑组织中G蛋白偶联受体78（GPR78）、PKR样内质网激酶（PERK）和磷酸化PERK（p-PERK）和CHOP蛋白表达水平。结果 mNSS和转棒实验，与假手术组比较，dMCAO组小鼠mNSS明显升高（P<0.01），在棒时间明显减少（P<0.01）；与dMCAO组比较，dMCAO+TUG-891组小鼠mNSS降低（P<0.05），在棒时间增加（P<0.05）。TTC染色法，与假手术组比较，dMCAO组小鼠脑梗死体积增大（P<0.01）；与dMCAO组比较，dMCAO+TUG-891组小鼠脑梗死体积明显减小（P<0.01）。HE染色，与假手术组比较，dMCAO组小鼠大脑皮层严重受损，表现为梗死区神经细胞排列紊乱及细胞核明显固缩。dMCAO+TUG-891组小鼠脑梗死区神经细胞形态损伤得到明显改善。NISSL染色，dMCAO组小鼠大脑皮层梗死区尼氏小体变细拉长，脱失增多；dMCAO+TUG-891组小鼠脑组织病理损伤均得到明显改善。与假手术组比较，dMCAO组大鼠脑组织中MDA水平明显升高（P<0.01），SOD活性降低（P<0.01）；与dMCAO组比较，dMCAO+TUG-891组小鼠脑组织中MDA水平明显降低（P<0.01），SOD活性明显升高（P<0.01）。TUNEL染色，与假手术组比较，dMCAO组小鼠脑组织中神经细胞凋亡指数升高（P<0.01）；与dMCAO组比较，dMCAO+TUG-891组小鼠脑组织中神经细胞凋亡指数降低（P<0.01）。与假手术组比较，dMCAO组小鼠脑组织中GRP78、p-PERK和CHOP蛋白表达水平升高（P<0.05）；与dMCAO组比较，dMCAO+TUG-891组小鼠脑组织中GRP78、p-PERK和CHOP蛋白表达水平降低（P<0.05）。结论 TUG-891可以减轻IS引起的神经损伤，其作用机制可能与TUG-891抑制内质网应激和细胞凋亡有关。

关键词: 缺血性脑卒中, G蛋白偶联受体78, TUG-891, 内质网应激, 细胞凋亡

Abstract:

Objective To discuss the protective effect of TUG-891 on ischemic stoke（IS） induced by ischemia-hypoxia， and to clarify its potential mechanism. Methods A total of 60 healthy male C57BL/6 mice were randomly divided into sham operation group（ n=20）， model group ［distal middle cerebral artery occlusion（dMCAO） group， n=20］， and model+TUG-891 group （dMCAO+TUG-891 group， n=20）. After modeling， the mice were intraperitoneally injected with TUG-891 solution （35 mg·kg?1·d?1） for 3 consecutive days. Modified neurological severity score （mNSS） and rotarod test were used to evaluate the neurological function of the mice in various groups； 2，3，5-triphenyltetrazolium chloride （TTC） staining was used to observe the cerebral infarction volumes of the mice in various groups； biochemical method was used to detect the malondialdehyde （MDA） level and superoxide dismutase （SOD） activity in the supernatant of brain tissue of the mice in various groups； Hematoxylin-Eosin （HE） and NISSL staining were used to observe the pathomerphology of brain tissue of the mice in various groups； terminal deoxynucleotidyl transferase dUTP nick-end labeling （TUNEL） staining was used to detect the apoptotic indexes of neuronal cells in brain tissue of the mice in various groups； Western blotting method was used to detect the expression levels of glucose-regulated protein 78 （GRP78）， protein kinase R-like endoplasmic reticulum kinase （PERK）， phosphorylated PERK （p-PERK）， and C/EBP homologous protein （CHOP） proteins in brain tissue of the mice in various groups. Results The mNSS and rotarod test results shoued that compared with sham operation group， the mNSS of the mice in dMCAO group was significantly increased （P<0.01）， and the time on the rod was significantly decreased （P<0.01）； compared with dMCAO group， the mNSS of the mice in dMCAO+TUG-891 group was decreased （P<0.05）， and the time on the rod was increased （P<0.05）. The TTC staining results shoued that compared with sham operation group， the volume of white infarct foci in the cerebral cortex of the mice in dMCAO group was increased （P<0.01）； compared with dMCAO group， the cerebral infarction volume of the mice in dMCAO+TUG-891 group was significantly decreased （P<0.01）. The HE staining results showed that compared with sham operation group， the cortex of the mice in dMCAO group was severely damaged， manifested by disordered arrangement of neuronal cells and obvious nuclear pyknosis in the infarct area， and the morphology of cortical infarct area of the mice in dMCAO+TUG-891 group was improved； the NISSL staining results showed that the Nissl bodies in the cortical infarct area of the mice in dMCAO group became thinner， elongated， and lost more. The pathological damage of brain tissue of the mice in dMCAO+TUG-891 group was significantly improved. Compared with sham operation group， the MDA level in brain tissue of the mice in model group was significantly increased （P<0.01）， and the SOD activity was decreased （P<0.01）； compared with model group， the MDA level in brain tissue of the mice in TUG-891 group was significantly decreased （P<0.01）， and the SOD activity was significantly increased （P<0.01）. The TUNEL staining results showed that compared with sham operation group， the apoptotic index of neuronal cells in brain tissue of the mice in dMCAO group was increased （P<0.01）； compared with dMCAO group， the apoptotic index of neuronal cells in brain tissue of the mice in dMCAO+TUG-891 group was decreased （P<0.01）. Compared with sham operation group， the expression levels of GRP78， p-PERK， and CHOP proteins in brain tissue of the mice in dMCAO group were increased （P<0.05）； compared with dMCAO group， the expression levels of GRP78， p-PERK， and CHOP proteins in brain tissue of the mice in dMCAO+TUG-891 group were decreased （P<0.05）. Conclusion TUG-891 can alleviate neurological injury caused by ischemic stroke， and its mechanism may be related to the inhibition of endoplasmic reticulum stress and apoptosis.

Key words: Ischemic stroke, G protein-coupled receptor 78, TUG-891, Endoplasmic reticulum stress, Apoptosis

中图分类号:

R285.5

孙攀喜,秦雪,张重阳,罗佳,陈勇,魏丽丽. TUG-891对缺血缺氧诱导的缺血性脑卒中的保护作用及其机制[J]. 吉林大学学报(医学版), 2025, 51(4): 968-975.

Panxi SUN,Xue QIN,Chongyang ZHANG,Jia LUO,Yong CHEN,Lili WEI. Protective effect of TUG-891 on ischemic stroke induced by ischemia and hypoxia and its mechanism[J]. Journal of Jilin University(Medicine Edition), 2025, 51(4): 968-975.

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Group	mNSS	Beam walking time (t/s)
Sham operation	0.24±0.10	289.00±23.93
dMCAO	13.37±0.82^*	163.00±30.05^*
dMCAO+TUG-891	9.88±0.53^△	208.00±19.27^△

Group	MDA [m_B/(μmol·g^-1)]	SOD[λ_B/（U·mg^-1）]
Sham operation	5.20±0.20	76.47±3.80
dMCAO	7.31±0.13^*	48.40±4.84^*
dMCAO+TUG-891	5.99±0.14^△	63.73±4.08^△