人脐带间充质干细胞对新生大鼠脑白质损伤后水通道蛋白4的影响

doi:10.3969/j.issn.1006-5725.2026.08.002

摘要/Abstract

摘要：

目的观察人脐带间充质干细胞（human umbilical cord mesenchymal stem cells，HUC-MSCs）移植对新生大鼠脑白质损伤（white matter injury，WMI）后水通道蛋白4（aquaporin-4，AQP4）的影响，评估对血脑屏障的修复作用及其可能机制。方法将2日龄Sprague-Dawley（SD）大鼠90只随机分为假手术组、WMI组及HUC-MSCs组（各组n = 30）。依次采用低剂量脂多糖腹腔注射（0.05 mg/kg）、单侧颈总动脉结扎联合缺氧（8%氧气＋92%氮气）构建新生大鼠WMI模型。HUC-MSCs组在建模后2 h内通过脑立体定位仪移植2×10⁵个HUC-MSCs，移植剂量为2 μL。建模后不同时间点进行样本的采集和检测。采用干湿重测量法分别检测各组大鼠脑含水量；苏木素-伊红染色观察各组大鼠脑组织病理形态变化；免疫组织化学染色观察各组大鼠脑组织中胶质纤维酸性蛋白（glial fibrillary acidic protein，GFAP）、AQP4及髓鞘碱性蛋白（myelin basic protein，MBP）表达部位及水平；免疫印迹实验检测各组大鼠AQP4、GFAP及MBP蛋白表达水平；采用Morris水迷宫实验观察各组大鼠的空间记忆能力；采用伊文思蓝实验检测各组大鼠脑组织中伊文思蓝含量。结果与假手术组相比，WMI组可见神经纤维排列疏松、走形紊乱，可见核固缩及核溶解表现；HUC-MSCs组较WMI组情况有所改善。与假手术组相比，WMI组脑含水量增多（均P < 0.05）；HUC-MSCs组较WMI组脑含水量下降（均P < 0.05）。与假手术组相比，WMI组和HUC-MSCs组脑组织中伊文思蓝含量增加，HUC-MSCs组较WMI组脑组织中伊文思蓝含量有所减少（均P < 0.05）。与假手术组相比，WMI组及HUC-MSCs组MBP蛋白表达降低，HUC-MSCs组较WMI组MBP蛋白表达升高（均P < 0.05）。与假手术组相比，WMI组及HUC-MSCs组GFAP蛋白表达升高，HUC-MSCs组较WMI组GFAP蛋白表达降低（均P < 0.05）。AQP4与GFAP表达趋势相同（均P < 0.05）。与假手术组相比，WMI组和HUC-MSCs组的逃逸潜伏期延长、穿越平台次数及在目标象限停留的时间减少；与WMI组相比，HUC-MSCs组的逃逸潜伏期缩短、穿越平台的次数及在目标象限停留的时间增多（均P<0.05）。结论 HUC-MSCs对新生大鼠WMI血脑屏障具有一定的修复作用，其机制可能与抑制星形胶质细胞活化并下调AQP4蛋白表达相关。

关键词: 脑白质损伤, 人脐带间充质干细胞, 血脑屏障, 星形胶质细胞, 水通道蛋白4

Abstract:

Objective To observe the effect of human umbilical cord mesenchymal stem cells （HUC-MSCs） transplantation on aquaporin-4（AQP4）after white matter injury（WMI）in neonatal rats，and to evaluate the repair effect on blood-brain barrier and its possible mechanism. Methods A total of 90 2-day-old Sprague-Dawley（SD）rats were randomly divided into three groups：the sham operation group，the WMI group，and the HUC-MSCs group，with 30 rats in each group.The WMI model of neonatal rats was constructed by intraperitoneal injection of low dose lipopolysaccharide（0.05 mg/kg），unilateral common carotid artery ligation combined with hypoxia（8% oxygen + 92% nitrogen）.The HUC-MSCs group was transplanted with 2×10⁵ HUC-MSCs at a dose of 2 μL by a brain stereotaxic apparatus within 2 hours after modeling.Samples were collected and tested at different time points after modeling.The brain water content of rats in each group was detected by wet and dry weight measurement；The pathological changes of brain tissue in each group were observed by hematoxylin-eosin staining； Immunohistochemical staining was used to observe the expression sites and levels of glial fibrillary acidic protein （GFAP）， AQP4 and myelin basic protein （MBP） in brain tissues，the expression levels of AQP4，GFAP and MBP protein in each group were detected by Weston blot，the memory and spatial exploration ability of rats in each group were observed by Morris water maze test，and the concentration of Evans blue in each group was detected by Evans blue test. Results Compared with the sham operation group，the WMI group showed loose nerve fiber arrangement，disordered shape，nuclear pyknosis and nuclear lysis. The situation in the HUC-MSCs group was improved compared with the WMI group. Compared with the sham group， the brain water content in the WMI group increased （all P < 0.05）； Brain water content decreased in the HUC-MSCs group compared with the WMI group （all P < 0.05）. Compared with the sham group， the content of Evans blue in the brain tissue of the WMI group and the HUC-MSCs group increased， and decreased in the HUC-MSCs group compared with the WMI group （all P < 0.05）. The expression of MBP in the WMI group and HUC-MSCs group decreased compared with the sham group， which increased in the HUC-MSCs group compared with the WMI group （all P < 0.05）. And the expression of GFAP in the WMI group and HUC-MSCs group increased compared with the sham group， and it decreased in the HUC-MSCs group compared with the WMI group （all P < 0.05）. AQP4 had the same trend as GFAP （all P < 0.05）. The escape incubation period of WMI group and HUC-MSCs group was prolonged， the number of platforms crossed and the time stayed in the target quadrant were shortened compared with the sham group； and the escape incubation period was shortened， the number of platforms crossed and the time spent in the target quadrant increased in the HUC-MSCs group compared with the WMI group（all P < 0.05）. Conclusion HUC-MSCs have a certain repair effect on WMI blood-brain barrier in neonatal rats， and the mechanism may be related to the inhibition of astrocyte activation and down-regulation AQP4 protein expression.

Key words: white matter injury, human umbilical cord mesenchymal stem cells, blood-brain barrier, astrocytes, aquaporin-4

中图分类号:

R722

柳雅蒙,王超,王梦欣,甘姗宁,朱艳萍. 人脐带间充质干细胞对新生大鼠脑白质损伤后水通道蛋白4的影响[J]. 实用医学杂志, 2026, 42(8): 1312-1321.

Yameng LIU,Chao WANG,Mengxin WANG,Shanning GAN,Yanping ZHU. Effects of human umbilical cord mesenchymal stem cells on aquaporin-4 after white matter injury in neonatal rats[J]. The Journal of Practical Medicine, 2026, 42(8): 1312-1321.

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组别	脑含水量
组别	7 d	14 d	t值	P值
假手术组	85.74 ± 0.18	82.81 ± 0.18	28.339	< 0.001
WMI组	86.40 ± 0.25^*	83.94 ± 0.27^*	16.172	< 0.001
HUC-MSCs组	85.76 ± 0.50^#	82.91 ± 0.13^#	13.492	< 0.001
F值	7.235	56.961
P值	0.006	< 0.001

组别	MBP		GFAP		AQP4
组别	平均光密度值	相对蛋白表达量	平均光密度值	相对蛋白表达量	平均光密度值	相对蛋白表达量
假手术组	0.22 ± 0.01	1.27 ± 0.11	0.17 ± 0.01	0.82 ± 0.08	0.16 ± 0.02	0.82 ± 0.09
WMI组	0.16 ± 0.00^*	0.76 ± 0.07^*	0.26 ± 0.02^*	1.19 ± 0.09^*	0.24 ± 0.17^*	1.18 ± 0.10^*
HUC-MSCs组	0.20 ± 0.01^*#	1.01 ± 0.08^*#	0.23 ± 0.02^*#	1.01 ± 0.04^*#	0.20 ± 0.01^*#	0.99 ± 0.04^*#
F值	106.690	46.550	42.125	37.641	48.215	29.828
P值	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001

组别	逃逸潜伏期/s					穿越平台次数	目标象限停留时间/s
组别	DAY1	DAY2	DAY3	DAY4	DAY5	穿越平台次数	目标象限停留时间/s
假手术组	53.82 ± 3.34	43.11 ± 2.06	36.87 ± 3.10	23.53 ± 2.11	9.06 ± 2.36	8.33 ± 1.75	48.60 ± 3.63
WMI组	51.39 ± 6.47	52.44 ± 4.54^*	46.13 ± 5.05^*	35.89 ± 1.82^*	21.47 ± 5.80^*	4.83 ± 0.75^*	35.32 ± 3.03^*
HUC-MSCs组	54.58 ± 3.98	47.12 ± 2.18^*#	41.46 ± 2.07^*#	28.96 ± 1.26^*#	14.85 ± 2.13^*#	6.67 ± 0.82^*#	42.08 ± 5.90^*#
F值	0.723	13.292	9.789	85.813	15.876	12.830	13.880
P值	0.501	< 0.001	0.002	< 0.001	< 0.001	0.001	< 0.001