摘要
目的:探讨神经干细胞促进大鼠局部神经元中降钙素基因相关肽(CGRP)生成和脊柱骨折愈合的机制。方法:将30只成年SD雄性大鼠分为对照组、脊柱骨折组和神经干细胞组,每组10只。采用酶联免疫吸附试验(ELISA)检测受损脊髓组织蛋白样品中的神经营养素-3(NT-3)和CGRP水平。通过蛋白印迹分析巢蛋白(Nestin)、CGRP和神经元核心抗原(NeuN)的蛋白表达。通过网格显微镜对切片组织中的神经元进行计数分析。使用ELISA试剂盒检测血清骨钙素(OC)、Ⅰ型前胶原氨基端延长肽(P1NP)、抗酒石酸酸性磷酸酶(TRACP-5b)和Ⅰ型胶原交联C-末端肽(CTX-1)。使用高分辨率显微CT扫描检测大鼠骨骼微结构。对大鼠骨组织形态学进行分析。结果:脊柱骨折组NT-3和CGRP水平较对照组降低,神经干细胞组NT-3和CGRP水平较脊柱骨折组升高(均P<0.05)。脊柱骨折组Nestin蛋白表达较对照组升高,神经干细胞组Nestin蛋白表达较脊柱骨折组升高(均P<0.05)。脊柱骨折组CGRP和NeuN蛋白表达较对照组降低,神经干细胞组CGRP和NeuN蛋白表达较脊柱骨折组升高(均P<0.05)。脊柱骨折组锥体外系、内锥体层和L_(1)脊髓节段处神经元数量较对照组减少,神经干细胞组锥体外系、内锥体层和L_(1)脊髓节段处神经元数量较脊柱骨折组增多(均P<0.05)。脊柱骨折组骨钙素和P1NP水平较对照组降低,TRAcP-5b和CTX-1水平较对照组升高(均P<0.05)。神经干细胞组骨钙素和P1NP水平较脊柱骨折组升高,TRAcP5-b和CTX-1水平较脊柱骨折组降低(均P<0.05)。脊柱骨折组骨矿物质密度(BMD)、每总体积的骨体积(BV/TV)、小梁数(Tb.N)、小梁厚度(Tb.Th)和皮质厚度(Ct.Th)较对照组减小,神经干细胞组BMD、BV/TV、Tb.N、Tb.Th和Ct.Th较脊柱骨折组增加(均P<0.05)。脊柱骨折组矿物沉积率(MAR)、骨形成率/骨表面(BFR/BS)和成骨细胞数量/骨表面(N.Ob/BS)较对照组降低,神经干细胞组MAR、BFR/BS和N.Ob/BS较脊柱骨折组升高(均P<0.05)。结论:神经干细胞能促进局部神经元中CGRP生成和神经元存活,从而促进大鼠骨折愈合和功能恢复。
Objective:To investigate the mechanism of neural stem cell promoting the generation of calcitonin gene-related peptide(CGRP)in local neurons and the healing of spinal fractures in rats.Methods:A total of 30 adult SD male rats were divided into control group,spinal fracture group and neural stem cell group.Enzyme-linked immunosorbent assay(ELISA)was used to detect the levels of NT-3 and CGRP in protein samples of injured spinal cord tissue.The protein expression of Nestin,CGRP and NeuN was analyzed by Western blot.Neurons in sliced tissues were counted and analyzed by grid microscopy.Osteocalcin,P1NP,TRAcP-5b and CTX-1 were measured by using ELISA kits.Bone microstructure was detected in rats by using high-resolution micro-CT scans.Bone histomorphology of rats was analyzed.Results:The levels of NT-3 and CGRP in the spinal fracture group were lower than those in the control group,and the levels of NT-3 and CGRP in the neural stem cell group were higher than those in the spinal fracture group(all P<0.05).The expression of Nestin protein in the spinal fracture group was higher than that in the control group,and the expression of Nestin protein in the neural stem cell group was higher than that in the spinal fracture group(all P<0.05).The protein expressions of CGRP and NeuN in the spinal fracture group were lower than those in the control group,and the protein expressions of CGRP and NeuN in the neural stem cell group were higher than those in the spinal fracture group(all P<0.05).The number of neurons in the extrapyramidal system,inner pyramidal layer and L_(1)spinal cord segment in the spinal fracture group were decreased in comparison with the control group,and the number of neurons in the extrapyramidal system,inner pyramidal layer and L_(1)spinal cord segment in the neural stem cell group were increased in comparison with the spinal fracture group(all P<0.05).The levels of osteocalcin and P1NP in the spinal fracture group were lower than those in the control group,and the levels of TRAcP-5b and CTX-1 in the spinal fracture group were higher than those in the control group(all P<0.05).The levels of osteocalcin and P1NP in the neural stem cell group were higher than those in the spinal fracture group,and the levels of TRAcP-5b and CTX-1 in the neural stem cell group were higher than those in the spinal fracture group(all P<0.05).BMD,BV/TV,Tb.N,Tb.Th and Ct.Th were decreased in the spinal fracture group in comparison with the control group,while BMD,BV/TV,Tb.N,Tb.Th and Ct.Th were increased in the neural stem cell group in comparison with the spinal fracture group(all P<0.05).MAR,BFR/BS and N.Ob/BS in the spinal fracture group were lower than those in control group,and MAR,BFR/BS and N.Ob/BS in the neural stem cell group were higher than those in the spinal fracture group(all P<0.05).Conclusion:Neural stem cell can promote the generation of CGRP in local neurons and the survival of neurons,thus promoting fracture healing and functional recovery in rats.
作者
李鹏
李晓博
王林钦
LI Peng;LI Xiaobo;WANG Linqin(Department of Spine Surgery,Ankang People’s Hospital,Ankang 725000,China)
出处
《陕西医学杂志》
CAS
2022年第9期1066-1070,1075,共6页
Shaanxi Medical Journal
基金
陕西省安康市科学技术研究发展计划项目(AK2019SF-05)。
关键词
脊柱骨折
神经干细胞
降钙素基因相关肽
神经元
骨折愈合
机制
Spinal fracture
Neural stem cell
Calcitonin gene-related peptide
Neuron
Fracture healing
Mechanism
