缓释NT-3的PLGA纳米微球对大鼠脊髓损伤的修复作用

Repairing effects of PLGA nanospheres with sustained-release of NT-3 on spinal cord injury in the rat

目的:应用聚乳酸聚乙醇酸共聚物(PLGA)包载神经营养因子3(NT-3),形成缓释纳米微球,探究其对大鼠脊髓损伤(SCI)的修复作用。方法:制备包载NT-3的PLGA纳米微球,扫描电镜观察其形态,ELISA法检测其释放,检测其对PC12细胞的作用。将18只成年雌性SD大鼠随机分为3组:假手术组(sham)、脊髓损伤组(SCI)和PLGA微球注射组(PLGA/NT-3)。应用脊髓打击器制作脊髓T9撞击损伤模型,PLGA/NT-3组大鼠立即在损伤区注射PLGA/NT-3纳米微球。术后每周对大鼠进行旷场实验及BBB评分,尼氏染色观察损伤区尾侧脊髓前角运动神经元(MNs)的变化,生物素化葡聚糖胺(BDA)示踪检测损伤区尾侧下行脊髓固有束(dSPST)情况。结果:制备的PLGA/NT-3纳米微球能够持续释放NT-3;与sham组和PLGA组比较,PLGA/NT-3组PC12细胞平均突起长度增加(P<0.05)。PLGA/NT-3组大鼠术后第7周、8周BBB评分、旷场总移动距离均高于SCI组(P<0.05);尼氏染色结果显示,PLGA/NT-3组大鼠脊髓损伤区尾侧脊髓前角MNs多于SCI组(P<0.05);BDA示踪结果显示,PLGA/NT-3组大鼠脊髓损伤区尾侧BDA标记的dSPST多于SCI组(P<0.05)。结论:包载NT-3的PLGA纳米微球可持续释放NT-3;PLGA/NT-3纳米微球能够促进脊髓损伤区尾侧灰质前角MNs和dSPST存活,并可改善大鼠后肢运动功能。

Objective:To fabricate sustained-release nanospheres containing neurotrophin-3(NT-3)by poly lactic-co-glycolic acid(PLGA)and investigate its effects on the repair of spinal cord injury(SCI)in rat.Methods:PLGA nanospheres encapsulated with NT-3 were prepared.The morphology was observed by scanning electron microscopy,and the release was detected by ELISA.The effect of NT-3-encapsulated nanospheres on PC12 cells was examined.Eighteen female SD adult rats were randomized into three groups:sham group,SCI group,and PLGA/NT-3 group.T9 contusive SCI rat model was established.Rats in the PLGA/NT-3 group were immediately injected with PLGA/NT-3 nanospheres into the injury area.Open field test and BBB scoring were conducted weekly after surgery.Nissl staining was used to observe the changes in motor neurons(MNs)in the anterior horn of spinal cord gray matter on the caudal segment of the injury area.The biotinylated dextran amine(BDA)trace was used to detect the descending spinal propriospinal tract(dSPST)in the caudal segment of the injury area.Results:PLGA/NT-3 nanospheres were successfully prepared,capable of sustained-release of NT-3.Compared with the sham group and the PLGA group,the average axon length of PC12 cells in the PLGA/NT-3 group was increased(P<0.05).At the 7thand 8thweeks after surgery,the BBB scores and total open field distance increased in the PLGA/NT-3 group compared with the SCI group(P<0.05).The results of Nissl staining showed that the number of MNs in the anterior horn of spinal cord gray matter at the caudal segment of the injury site in the PLGA/NT-3 group was greater than that in the SCI group(P<0.05).BDA tracing results showed that the dSPST labeled with BDA at the caudal segment of the injury area in the PLGA/NT-3 group was greater than that in the SCI group(P<0.05).Conclusion:PLGA nanospheres containing NT-3 consistently release NT-3.The application of PLGA/NT-3 nanospheres could promote the survival of MNs and dSPST at the caudal segment of the SCI area and improve the locomotor recovery of the hind limbs in rat.