人脐带间充质干细胞在缺氧缺血性脑损伤幼鼠学习记忆中的潜在调节机制研究

Potential Regulatory Mechanism of Human Umbilical Cord Mesenchymal Stem Cells in Learning and Memory of Young Rats with Hypoxic Ischemic Brain Damage Injury

探讨人脐带间充质干细胞(Human umbilical cord mesenchymal stem cells,hUC-MSCs)在缺氧缺血性脑损伤(Hypoxic ischemic brain damage,HIBD)幼鼠学习记忆中的潜在调节机制。选取健康新生7日龄SD大鼠,将幼鼠分为3组(每组8只):对照组(Sham+0.9%NaCl),治疗组(HIBD+hUC-MSCs),损伤组(HIBD+0.9%NaCl)。通过缺血(右侧颈总动脉结扎离断)+缺氧(8%O_(2)+92%N_(2))法制备HIBD幼鼠模型,对照组仅行假手术(Sham)处理。通过Morris水迷宫实验检测幼鼠学习记忆力,HE和TUNEL染色观察大脑额叶皮层组织病变及神经元凋亡情况,Western-blot实验测定额叶皮层BDNF和p-CREB/CREB蛋白表达水平。hUC-MSCs治疗可减少HIBD幼鼠逃避潜伏时间,增加穿台次数,差异均具有统计学意义(P<0.05)。HE染色显示对照组幼鼠额叶皮层组织结构正常,细胞排列整齐,无明显变性坏死及炎症浸润,损伤组幼鼠皮层神经细胞排列杂乱,核固缩深染,大片组织液化坏死灶,空泡形成,神经细胞大量坏死,而经hUC-MSCs治疗后皮层损伤明显减轻。凋亡染色发现经hUC-MSCs治疗可抑制HIBD幼鼠额叶皮层神经元的凋亡,差异具有统计学意义(P<0.05)。进一步发现hUC-MSCs治疗促进了HIBD幼鼠额叶皮层中BDNF和CREB磷酸化水平,差异均具有显著统计学意义(P<0.05)。实验证实hUC-MSCs干预可减轻额叶皮层损伤、抑制神经细胞凋亡;hUC-MSCs干预可促进HIBD幼鼠学习,有助于记忆功能的改善,可能与额叶皮层BDNF、p-CREB的表达增强有关。

To investigate the potential regulatory mechanism of human umbilical cord mesenchymal stem cells(hUC-MSCs)in learning and memory of hypoxic-ischemic brain damage(HIBD)young rats,healthy newborn 7-day-old SD rats were selected and divided into three groups(8 rats in each group):control group(sham+0.9%NaCl),treatment group(HIBD+hUC-MSCs)and injury group(HIBD+0.9%NaCl).HIBD model was established by ischemia(ligation and disconnection of right common carotid artery)+hypoxia(8%O_(2)+92%N_(2)),and the control group was treated with sham operation only.Morris water maze test was used to detect the learning and memory of young rats,HE and TUNEL staining were used to observe the tissue lesions and neuronal apoptosis of frontal cortex,and Western blot test was used to measure the expression levels of BDNF and p-CREB/CREB protein in frontal cortex.hUC-MSCs treatment could reduce the escape latency of HIBD young rats and increase the number of passes through the platform,the difference was statistically significant(P<0.05).HE staining showed that the tissue structure of frontal cortex in the control group was normal,the cells were arranged orderly,without obvious degeneration,necrosis and inflammatory infiltration.The cortical nerve cells in the injury group were arranged disorderly,nuclear pyknosis and deep staining,large areas of tissue liquefaction and necrosis,vacuole formation,and a large number of nerve cells died.However,the cortical injury was significantly reduced after hUC-MSCs treatment.Apoptosis staining showed that hUC-MSCs treatment could inhibit the apoptosis of frontal cortical neurons in HIBD young rats,and the difference was statistically significant(P<0.05).It was further found that hUC-MSCs treatment promoted the phosphorylation levels of BDNF and CREB in the frontal cortex of HIBD young rats,and the difference was statistically significant(P<0.05).(1)hUC-MSCs intervention can reduce frontal cortex injury and inhibit neuronal apoptosis.(2)hUC-MSCs intervention promotes the improvement of learning and memory function in HIBD young rats,which may be related to the increased expression of BDNF and p-CREB in frontal cortex.