摘要
目的探究雷公藤甲素(Triptolide,T10)对深低温保存动物模型神经活性和功能的影响。方法收集30只SD大鼠双侧15 mm长的坐骨神经组织,随机分为3组:冻存组、低浓度T10组(L-T10)和高浓度T10(H-T10)组,分别在含有0、1×10^(-9)、1×10^(-8)mol/L的T10的冻存液中、在-4℃的条件下低温保存4周。另取10条新鲜的长度为15 mm坐骨神经组织作为对照组。30只Wistar大鼠随机分为3组:冻存组、L-T10组和H-T10组,均构建坐骨神经10 mm损伤模型,将低温保存的坐骨神经修剪成10 mm进行异体移植。另取10只Wistar大鼠做坐骨神经自体移植模型。用TEM观察坐骨神经结构;利用Calcein-AM/DAPI荧光染色检测坐骨神经活性;神经电生理检测肌肉复合动作电位(CMAP)和运动神经传导速度(MNCV);Western blot检测NGF和GDNF蛋白。结果在冻存4周后,各组均出现不同程度的脱髓鞘样改变,并且出现空泡和轴浆萎缩,其中冻存组坐骨神经损伤最严重,H-T10组损伤情况较冻存组和L-T10组轻。对照组的细线坐骨神经组织中活细胞最多。冻存组活细胞减少。L-T10组和H-T10组的存活细胞较冻存组多,并且H-T10组的存活细胞多于L-T10组。冻存组的NGF和GDNF水平显著低于对照组(P<0.05),L-T10组和H-T10组的NGF和GDNF显著高于冻存组(P<0.05),并且H-T10组的NGF和GDNF水平高于L-T10组(P<0.05)。在移植20周后,冻存组的CMAP和MNCV显著低于对照组(P<0.05),L-T10组和H-T10组的CMAP和MNCV显著高于冻存组(P<0.05),并且H-T10组的CMAP和MNCV水平高于L-T10组(P<0.05);冻存组的神经再生的情况显著低于对照组(P<0.05),L-T10组和H-T10组的有髓神经纤维数目和髓鞘厚度/纤维直径显著高于冻存组(P<0.05),并且H-T10组的有髓神经纤维数目和髓鞘厚度/纤维直径水平高于L-T10组(P<0.05)。结论T10可提高深低温保存坐骨神经的活性,并且促进异体移植后坐骨神经的神经再生和功能恢复。
Objective To explore the effects of Triptolide(PG490 or T10)on the biological activity of sciatic nerve and the neural regeneration and functional recovery after transplantation in cryopreserved animal models.Methods Thirty SD rat bilateral 15 mm long sciatic nerve tissues were collected and randomly divided into 3 groups:cryopreservation group,low concentration T10 group(L-T10)and high concentration T10(H-T10)group,each containing 0 mol/L,1×10^(9)mol/L and 1×10^(8)mol/L T10 cryopreserved at-4℃for 4 weeks at low temperature.10 fresh 15 mm long sciatic nerve tissues were collected as a control group.Thirty Wistar rats were randomly divided into three groups:the cryopreserved group,the L-T10 group,and the H-T10 group.All were constructed with a sciatic nerve 10 mm injury model,and the cryopreserved sciatic nerve was trimmed to 10 mm for allogeneic transplantation.Another 10 Wistar rats were used as autologous transplantation models of fresh sciatic nerve.Transmission electron microscope(TEM)observation of sciatic nerve structure.Calcein-AM/DAPI fluorescence staining was used to detect sciatic nerve activity.Neuroelectrophysiology measures muscle complex action potential(CMAP)and motor nerve conduction velocity(MNCV).Nerve growth factor(NGF)and glialcellline-derivedneurotrophic factor(GDNF)proteins were detected by Western blot.Results After 4 weeks of cryopreservation,all groups showed varying degrees of demyelinating changes,and vacuoles and axonal atrophy occurred.The sciatic nerve injury was the most severe in the cryopreservation group.H-T10 group was lighter than the frozen group and L-T10 group.The control group had the most living cells in the thin-line sciatic nerve tissue.Viable cells were reduced in the cryopreservation group.There were more viable cells in the L-T10 and H-T10 groups than in the cryopreserved group.There were more surviving cells in the H-T10 group than in the L-T10 group.The levels of NGF and GDNF in the cryopreservation group were significantly lower than those in the control group(
作者
王维
李楠
田宇
WANG Wei;LI Nan;TIAN Yu(Department of Hand and Foot Surgery,Qinhuang-dao First Hospital,Qinhuangdao,Hebei Province 066000,China;Department of Ophthalmology,Qinhuangdao maternal and child health hospital,Qinhuangdao,Hebei Province 066000,China)
出处
《解剖学研究》
CAS
2022年第1期33-38,共6页
Anatomy Research
关键词
雷公藤甲素
低温保存
坐骨神经
神经再生
Triptolide
Cryopreservation
Sciatic nerve
Nerve regeneration
