磁共振成像2024,Vol.15Issue(2) :122-128.DOI:10.12015/issn.1674-8034.2024.02.018

磁共振髓鞘探针Gd-DTDAS在多发性硬化大鼠髓鞘损伤模型中的实验研究

Experimental study of magnetic resonance targeted myelin probe Gd-DTDAS in multiple sclerotic rat myelin injury model

刘彩云 邵翠杰 翁娜 李国栋 黄丹琪 刘珈 宾莉 王旭
磁共振成像2024,Vol.15Issue(2) :122-128.DOI:10.12015/issn.1674-8034.2024.02.018

磁共振髓鞘探针Gd-DTDAS在多发性硬化大鼠髓鞘损伤模型中的实验研究

Experimental study of magnetic resonance targeted myelin probe Gd-DTDAS in multiple sclerotic rat myelin injury model

刘彩云 1邵翠杰 2翁娜 1李国栋 1黄丹琪 1刘珈 1宾莉 1王旭1
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作者信息

  • 1. 滨州医学院附属医院核医学科,滨州 256600
  • 2. 滨州医学院附属医院医学研究中心,滨州 256600
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摘要

目的 探讨MRI对比剂Gd-DTDAS在多发性硬化(multiple sclerosis,MS)大鼠髓鞘损伤模型中的应用价值.材料与方法细胞实验中,将少突胶质细胞前体细胞(oligodendrocyte precursor cells,OLN-93)随机分为对照组2(n=3)和溶血磷脂酰胆碱(lysophosphatidylcholine,LPC)组(n=3),LPC组细胞置于无菌共聚焦培养皿中与1 mL 800 μM LPC溶液共孵育30 min.通过噻唑蓝比色法(methyl thiazolyl tetrazolium,MTT)评价细胞毒性,计算OLN-93与Gd-DTDAS共孵育24 h后的吸光度和存活率;细胞摄取实验中,对照组2和LPC组对比,定量两组细胞对Gd-DTDAS的摄取值以及相应荧光强度的变化.动物实验中,将6~8周龄SD大鼠随机分为对照组(n=12)与实验组(n=18),实验组大鼠左侧胼胝体注射1%LPC溶液(1%LPC溶于PBS).造模后(1、3、7 d)进行行为学观察,并在注射后7天进行T1WI及T2WI序列扫描.根据MRI异常信号部位进行大鼠脑组织Gd-DTDAS染色(n=6)以及浸泡(n=6),评估Gd-DTDAS与髓鞘部位的结合情况,其中,染色实验分组命名为对照组3与实验组3,浸泡实验分组命名为对照组4与实验组4;通过尾静脉注射Gd-DTDAS,MR评估实验组(n=6)注射Gd-DTDAS前后大脑髓鞘变化.结果 细胞毒性实验中,当Gd-DTDAS浓度增加到400 μM时,OLN-93细胞的存活率约为95%,细胞存活率差异无统计学意义(t=4.20,P>0.05).细胞摄取实验中,两组细胞均能摄取Gd-DTDAS,LPC组摄取量显著低于对照组2,差异具有统计学意义(t=31.75,P<0.01).动物体外实验中,与对照组3比较,Gd-DTDAS染色的实验组3脑组织切片荧光强度显著下降,差异有统计学意义(U=9,P<0.01);Gd-DTDAS浸泡中,对照组4(n=6)与实验组4(n=6)脑组织切片浸泡后MRI分辨率显著升高,差异有统计学意义(对照组4,t=8.76,P<0.01)(实验组4,t=2.89,P<0.01).体内实验中,与尾静脉注射前比较,注射后胼胝体区域MRI T1maps弛豫性显著降低(t=14.46,P<0.01).结论 髓鞘探针Gd-DTDAS能够更好地结合髓磷脂丰富的区域,髓鞘靶向MRI显像更佳,能特异性显示多发性硬化髓鞘损伤部位.

Abstract

Objective:To investigate the application value of MRI contrast agent Gd-DTDAS in multiple sclerosis(MS)rat myelin injury model.Materials and Methods:In cell experiments,oligodendrocyte precursor cells(OLN-93)were randomly divided into control group 2(n=3)and lysophosphatidylcholine(LPC)group(n=3),and the cells of LPC group were incubated with 1 mL of 800 μM LPC in a sterile confocal dish for 30 min.Cytotoxicity was evaluated by methyl thiazolyl tetrazolium(MTT),and the absorbance and survival rate of OLN-93 after incubation with Gd-DTDAS for 24 h were calculated.In the uptake experiment,the control group 2 and the LPC group were compared to quantify the uptake value of Gd-DTDAS and the corresponding fluorescence intensity of the two groups.In animal experiments,6-8 week-old SD rats were randomly divided into control group(n=12)and experimental group(n=18),and the left corpus callosum of rats in the experimental group was injected with 1%LPC solution(1%LPC dissolved in PBS).After molding,behavioral observation was performed(1,3,7 d),and T1WI and T2WI sequence scanning were performed 7 d after injecting.Gd-DTDAS staining(n=6)and soaking(n=6)of rat brain tissue were performed according to the MRI abnormal signal site to evaluate the binding of Gd-DTDAS to the myelin site.Among them,the staining experiment was named as control group 3 and experimental group 3,while the soaking experiment group was named as control group 4 and experimental group 4.Gd-DTDAS was injected by tail vein,MRI assessed cerebral myelin sheath changes before and after Gd-DTDAS injection in the experiment group(n=6).Results:In the cytotoxicity experiment,when the concentration of Gd-DTDAS increased to 400 μM,the survival rate of OLN-93 cells was about 95%,and there was no significant difference in cell survival between concentrations(t=4.20,P>0.05).In the cell uptake experiment,both groups of cells could uptake Gd-DTDAS,and the uptake of LPC group was significantly lower than that of the control group 2,and the difference was statistically significant(t=31.75,P<0.01).In vitro experiments,compared with the control group 3,the fluorescence intensity of brain tissue sections in the experiment 3 group stained with Gd-DTDAS decreased significantly,and the difference was statistically significant(U=9,P<0.01).After immersion of brain tissue slices in Gd-DTDAS,the MRI resolution significantly increased in both the control group 4(n=3)and the experiment group 4(n=6),with statistically significant differences(control group 4,t =8.76,P<0.01;experiment group 4,t =2.89,P<0.01).In vivo experiments,MRI T1maps relaxation in the medullary region was significantly reduced after injection compared with before tail vein injection(t =14.46,P<0.01).Conclusions:The myelin probe Gd-DTDAS can better bind to myelin-rich regions,and the myelin sheath can be better targeted for MRI,and can specifically show the damage site of myelin sheath in multiple sclerosis.

关键词

自身免疫性疾病/多发性硬化/髓鞘探针/分子成像/磁共振成像

Key words

autoimmune diseases/multiple sclerosis/myelin probe/molecular imaging/magnetic resonance imaging

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基金项目

国家自然科学基金(81771828)

出版年

2024
磁共振成像
中国医院协会 首都医科大学附属北京天坛医院

磁共振成像

CSTPCD北大核心
影响因子:1.38
ISSN:1674-8034
参考文献量42
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