摘要
背景:非侵入式脊髓损伤治疗方法亟待开发。纳米材料能够递送药物,提高治疗效果,具有明显优势。目的:制备缓释神经营养因子3和神经节苷脂GD1a的聚乳酸-羟基乙酸共聚物纳米微球,探究其对大鼠脊髓损伤的修复作用。方法:采用油水乳液挥发有机溶剂法,制备缓释神经营养因子3的聚乳酸-羟基乙酸共聚物纳米微球和缓释神经节苷脂GD1a的聚乳酸-羟基乙酸共聚物纳米微球,检测微球的缓释性能。采用随机数字表法将60只雌性SD成年大鼠分为5组:假手术组打开椎板后直接缝合,脊髓损伤组建立脊髓T9撞击损伤模型,神经营养因子组脊髓T9损伤区域注射缓释神经营养因子3的纳米微球悬液,神经节苷脂组脊髓T9损伤区域注射缓释经节苷脂GD1a的纳米微球悬液,实验组脊髓T9损伤区域注射缓释神经营养因子3的纳米微球和缓释神经节苷脂GD1a的纳米微球混合悬液,每组12只。术后每周进行旷场实验及BBB评分,术后4,8周进行脊髓组织形态学观察。结果与结论:(1)体外浸泡于PBS 14 d内,缓释纳米微球可持续释放神经营养因子3和神经节苷脂GD1a。(2)术后7,8周,与脊髓损伤组比较,神经营养因子组、实验组大鼠的BBB评分、旷场总移动距离增加(P<0.05)。尼氏染色显示,实验组术后4,8周的脊髓灰质前角运动神经元多于脊髓损伤组(P<0.05),神经营养因子组术后8周的脊髓灰质前角运动神经元多于脊髓损伤组(P<0.05)。免疫荧光染色显示,与脊髓损伤组比较,神经营养因子组术后8周、实验组术后4,8周的脊髓白质腹侧神经纤维增多(P<0.05),神经节苷脂组、实验组术后4,8周的脊髓白质腹侧髓鞘碱性蛋白表达增加(P<0.05),神经营养因子组术后8周的髓鞘碱性蛋白表达增加(P<0.05),神经营养因子组、实验组术后8周的下行脊髓固有束增加(P<0.05)。(3)结果表明,神经营养因子3微球单独应用,或是与神经节苷脂GD1a微球联合应用,能够促进脊髓损伤区周边运动神经元及神经纤维存活,并可改善大鼠后肢运动功能。
BACKGROUND:Non-invasive treatment of spinal cord injury needs to be developed urgently.Nanomaterials have obvious advantages,which can deliver drugs and improve the therapeutic effect.OBJECTIVE:To fabricate sustained-release nanospheres containing neurotrophin-3 and ganglioside GD1 a by poly(lactic-co-glycolic acid)and investigate their effects on the repair of spinal cord injury in rats.METHODS:Poly(lactic-co-glycolic acid)nanospheres encaps ulated with neurotrophin-3 and ganglioside GD1 a were prepared using oil-water emulsion volatile organic solvent method.The sustained release prope rties of microspheres were tested.Sixty female adult SD rats were randomly divided into five groups with12 rats in each group.In the sham ope ration group,the lamina was opened and sutured directly.In the spinal cord injury group,a spinal cord T9 impingement injury model was established.In the neurotrophin group,the nanosphere suspension of slow-release neurotrophin-3 was injected into the injured area of spinal cord T9.In the ganglioside GD1 a group,the sustained-release ganglioside GD1 a nanosphere suspension was injected into the spinal cord T9 injury area.The expe rimental group was injected with nanosphere suspension of sustained-release neurotrophin-3 and ganglioside GD1 a in the injured area of spinal co rd T9.Open field test and BBB sco ring were conducted weekly after operation.The sections were to ken 4 and 8 weeks after surgery for morphological observation.RESULTS AND CONCLUSION:(1)After soaking in PBS for 14 days in vitro,the sustained-release nanospheres could continuously release neurotrophin-3 and ganglioside GD1 a.(2)At 7 and 8 wee ks after surgery,compared with the spinal cord injury group,the BBB score and the total open field distance of the rats increased in the neurotrophin group and the expe rimental group(P<0.05).The results of Nissl staining showed that at 4 and 8 wee ks after surgery,the number of motor neurons increased in the anterior horn of caudal gray matter in the experimental group compared with the spinal cord inju ry group(P<0.05).At 8 weeks after surgery,the number of motor neurons increased in the anterior horn of caudal gray matter in the neurotrophin group compared with the spinal co rd injury group(P<0.05).The res ults of immunofluorescence staining showed that compared with the spinal co rd injury group,the neurotrophin group at 8 weeks and the experimental group at 4 and 8 weeks had increased spinal white matter ventral nerve fibers(P<0.05);the expression of myelin basic protein on the ventral side of the spinal co rd white matter increased at 4 and 8 weeks in the ganglioside GDla and experimental groups(P<0.05);the expression of myelin basic protein in the neurotrophin group increased at 8 wee ks after operation(P<0.05);the descending proprios pinal tra ct increased in the neurotrophin group and the experimental group at 8 weeks(P<0.05).(3)The results showed that neurotrophin-3 microspheres alone or in combination with ganglioside GD1 a microspheres could promote the survival of motor neurons and nerve fibers around the spinal co rd injury site and improve the motor function of the hind limbs of rats.
作者
夏宇
孙佳
齐争艳
马琳
马文倩
牛建国
文玉军
Xia Yu;Sun Jia;Qi Zhengyan;Ma Lin;Ma Wenqian;Niu Jianguo;Wen Yujun(Department of Human Anatomy and Histology Embryology,School of Basic Medicine,Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Ningxia Key Laboratory of Craniocerebral Diseases,Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China)
出处
《中国组织工程研究》
CAS
北大核心
2023年第16期2518-2524,共7页
Chinese Journal of Tissue Engineering Research
基金
宁夏自然科学基金项目(2020AAC03182),项目负责人:文玉军
国家自然科学基金项目(81660216),项目负责人:文玉军
宁夏重点研发计划项目(2021BEG03096),项目负责人:文玉军。
作者简介
第一作者:夏宇,男,1994年生,湖南省邵阳市人,汉族,宁夏医科大学在读硕士,主要从事脊髓损伤修复研究。https://orcid.org/0000-0002-2120-7786;通讯作者:文玉军,教授,硕士生导师,宁夏医科大学基础医学院人体解剖与组织胚胎学系,颅脑疾病重点实验室,宁夏回族自治区银川市750004。
