马索罗酚（NDGA）对实验性自身免疫性脑脊髓炎小鼠炎症因子表达的影响
发布时间：2019-06-14 06:58
【摘要】：目的:多发性硬化(MS)是一种以炎性脱髓鞘和轴索损伤为特征的免疫介导的中枢神经系统疾病。多发性硬化的病因和机制尚不明确,但炎症是这个疾病早期最重要的环节。各种细胞因子以自分泌或旁分泌方式,组成了一个复杂的动态网络结构,参与了免疫反应的所有过程及阶段,在调节免疫反应和组织修复中起着关键性作用,决定着多发性硬化的病情严重程度及发展趋势。我们课题组前期对Nrf2通路激活剂莱菔硫烷的研究中发现其增强小鼠抗氧化应激能力和调节免疫机制,在实验性自身免疫性脑脊髓炎(EAE)小鼠中起到神经保护作用。然而莱菔硫烷尚未应用于临床,通过文献查阅,我们发现了可应用于人体的Nrf2通路激活剂——马索罗酚。马索罗酚在MS的免疫调节机制中的作用尚未明确,需要进一步的研究。本实验以马索罗酚作为干预药物,观察EAE小鼠及马索罗酚治疗小鼠发病严重程度及病程的影响。研究马索罗酚对EAE小鼠IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β表达的调节作用。进而探索相关细胞因子在EAE病程中的作用,并研究马索罗酚在EAE免疫学机制中是否发挥神经保护作用。方法:将54只8至10周雌性C57BL/6小鼠,体重18g至20g,随机分成control组、EAE组和NDGA组,各组分别为18只。每组分为2个亚组,10天组及20天组,各亚组为9只。建立EAE动物模型。NDGA组小鼠自发病(weaver评分≥1分)当天(记为第0天),每只每日腹腔注射NDGA溶液10mg/kg。EAE组及control组小鼠自第0天,每只每日腹腔注射5%DMSO 10ml/kg。每日一次,直至处死。每天2次对其进行称重和神经功能的评分。利用实时定量PCR方法检测脊髓及脾中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β表达水平,用ELISA方法检测脑组织中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β表达水平。结果:1在发病第10天及第20天,NDGA组平均神经功能评分均较EAE组显著降低(P0.05)。2各组小鼠脊髓及脾组织中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β的m RNA水平:在发病10天时,与Control组对比,EAE组IL-6、IL-12、IL-17、IFN-γm RNA水平均增高,NDGA组IL-6、IL-12、IL-17、IFN-γm RNA水平较EAE组低,并有统计差异(P0.05)。EAE组TGF-β、IL-4 m RNA水平降低,NDGA组TGF-β、IL-4 m RNA水平较EAE组高,并有统计差异(P0.05)。在发病20天时,与Control组对比,EAE组IL-6、IL-12、IL-17、IFN-γm RNA水平均增高,NDGA组IL-12、IL-17、IFN-γm RNA表达较EAE组低,并有统计差异(P0.05)。EAE组TGF-β、IL-4 m RNA水平降低,NDGA组TGF-βm RNA水平较EAE组高,并有统计差异(P0.05)。3各组小鼠脑组织中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β含量变化情况:在发病10天时,与Control组对比,EAE组IL-6、IL-12、IL-17、IFN-γ的表达均增高,NDGA组IL-6、IL-12、IL-17、IFN-γ的表达较EAE组低,并有统计差异(P0.05)。EAE组TGF-β、IL-4的表达降低,NDGA组TGF-β、IL-4的表达较EAE组高,并有统计差异(P0.05)。在发病20天时,与Control组对比,EAE组IL-17、IFN-γ的表达均增高,NDGA组IL-17、IFN-γ表达较EAE组低,并有统计差异(P0.05)。NDGA组TGF-β的表达较EAE组高,并有统计差异(P0.05)。结论:1 EAE的发病和缓解与炎症因子的动态变化相关。2 NDGA可以减轻EAE小鼠神经功能损害严重程度,促进神经功能恢复,对EAE小鼠具有保护作用。3 NDGA可以降低EAE脑、脊髓及脾组织中促炎症细胞因子IL-6、IL-12、IL-17、IFN-γ水平,增加抗炎症细胞因子IL-4和TGF-β水平,具有免疫抗炎作用。4调节免疫炎症平衡可能是NDGA对EAE发挥保护作用的机制之一。
[Abstract]:Objective: Multiple sclerosis (MS) is an immune-mediated central nervous system disease characterized by inflammatory defibrination and axonal injury. The cause and mechanism of multiple sclerosis is not clear, but inflammation is the most important step in the early stage of this disease. The various cytokines form a complex dynamic network structure in the form of autocrine or paracrine, and participate in all the processes and stages of the immune response, play a key role in regulating immune response and tissue repair, and determine the severity and development trend of multiple sclerosis. In the early stage of our research group, the anti-oxidative stress ability and the regulation of the immune mechanism of the mice were found in the study of the Nrf2 pathway activator, Lepithionane, and played a role in neuroprotection in the experimental autoimmune encephalomyelitis (EAE) mice. However, that levonthiane has not been applied to clinical use, and it has been found by the literature that the Nrf2 pathway activator of the human body, the assorophenol, can be found. The role of Massorophenol in the immunoregulation mechanism of MS is not clear, and further research is needed. The effect of EAE on the severity of the disease and the course of the course of the treatment of mice with EAE and Moxinol was observed in this experiment as an intervention drug. The regulation of the expression of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in EAE mice was studied. In order to explore the role of relevant cytokines in the course of EAE, and to study whether the neuroprotective effect of the Moxinol in the EAE immunological mechanism was used. Methods:54 female C57BL/6 mice from 8 to 10 weeks were divided into control group, EAE group and NDGA group, and 18 rats were randomly divided into control group, EAE group and NDGA group. Each component was 2 subgroups,10 days and 20 days, and 9 in each subgroup. An EAE animal model was established. In the NDGA group, the mice were intraperitoneally injected with an NDGA solution of 10 mg/ kg (day 0) on the same day (as day 0), each of which was injected with 5% DMSO at 10 ml/ kg per day on the day 0 of each daily intraperitoneal injection of the NDGA solution. Once daily, until termination. It was scored twice a day for weighing and neurological function. The expression levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in the spinal cord and the spleen were detected by real-time quantitative PCR. The levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in the brain tissue were detected by ELISA. Results:1 In the 10 and 20 days of the attack, the mean neurological score of the NDGA group was significantly lower than that of the EAE group (P0.05). The levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in the spinal and spleen tissues of the mice were significantly lower than that in the EAE group (P0.05). The levels of IL-6, IL-12, IL-17 and IFN-were lower in the NDGA group than in the EAE group, and there was a statistical difference (P0.05). The levels of TGF-1 and IL-4 mRNA in the EAE group were lower than that of the EAE group, and there was statistical difference (P0.05). Compared with the control group, the levels of IL-6, IL-12, IL-17 and IFN-mRNA in EAE group were higher in EAE group than in control group, and the level of IL-12, IL-17 and IFN-mRNA in NDGA group was lower than that of EAE group, and there was statistical difference (P0.05). The level of TGF-1 and IL-4 mRNA in the EAE group was lower, and the level of TGF-5 mRNA in NDGA group was higher than that of EAE group. The levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-2 in the brain of each group were changed. The expression of IL-6, IL-12, IL-17 and IFN-1 in the EAE group was higher than that of the control group at 10 days. The expression of IL-6, IL-12, IL-17 and IFN-1 in the NDGA group was lower than that of the EAE group. There was statistical difference (P0.05). The expression of TGF-1 and IL-4 in EAE group decreased, and the expression of TGF-1 and IL-4 in NDGA group was higher than that of EAE group, and there was statistical difference (P0.05). The expression of IL-17 and IFN-1 in EAE group was higher in EAE group than in control group at 20 days. The expression of IL-17 and IFN-1 in NDGA group was lower than that of EAE group, and there was statistical difference (P0.05). The expression of TGF-1 in NDGA group was higher than that of EAE group, and there was statistical difference (P0.05). Conclusion: The incidence and the response of 1EAE are related to the dynamic changes of the inflammatory factors. The NDGA can reduce the severity of the neurological function damage in the EAE mice, promote the recovery of the nerve function, and have a protective effect on the EAE mice. The 3NDGA can reduce the pro-inflammatory cytokines IL-6, IL-12, IL-17 in the EAE brain, the spinal cord and the spleen tissue. The anti-inflammatory cytokines IL-4 and TGF-1 levels in the anti-inflammatory cytokine IL-4 and TGF-1 levels have an anti-inflammatory effect, and the regulation of the balance of immune inflammation may be one of the mechanisms of NDGA to play a protective role in EAE.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R744.51
本文编号：2499205
[Abstract]:Objective: Multiple sclerosis (MS) is an immune-mediated central nervous system disease characterized by inflammatory defibrination and axonal injury. The cause and mechanism of multiple sclerosis is not clear, but inflammation is the most important step in the early stage of this disease. The various cytokines form a complex dynamic network structure in the form of autocrine or paracrine, and participate in all the processes and stages of the immune response, play a key role in regulating immune response and tissue repair, and determine the severity and development trend of multiple sclerosis. In the early stage of our research group, the anti-oxidative stress ability and the regulation of the immune mechanism of the mice were found in the study of the Nrf2 pathway activator, Lepithionane, and played a role in neuroprotection in the experimental autoimmune encephalomyelitis (EAE) mice. However, that levonthiane has not been applied to clinical use, and it has been found by the literature that the Nrf2 pathway activator of the human body, the assorophenol, can be found. The role of Massorophenol in the immunoregulation mechanism of MS is not clear, and further research is needed. The effect of EAE on the severity of the disease and the course of the course of the treatment of mice with EAE and Moxinol was observed in this experiment as an intervention drug. The regulation of the expression of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in EAE mice was studied. In order to explore the role of relevant cytokines in the course of EAE, and to study whether the neuroprotective effect of the Moxinol in the EAE immunological mechanism was used. Methods:54 female C57BL/6 mice from 8 to 10 weeks were divided into control group, EAE group and NDGA group, and 18 rats were randomly divided into control group, EAE group and NDGA group. Each component was 2 subgroups,10 days and 20 days, and 9 in each subgroup. An EAE animal model was established. In the NDGA group, the mice were intraperitoneally injected with an NDGA solution of 10 mg/ kg (day 0) on the same day (as day 0), each of which was injected with 5% DMSO at 10 ml/ kg per day on the day 0 of each daily intraperitoneal injection of the NDGA solution. Once daily, until termination. It was scored twice a day for weighing and neurological function. The expression levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in the spinal cord and the spleen were detected by real-time quantitative PCR. The levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in the brain tissue were detected by ELISA. Results:1 In the 10 and 20 days of the attack, the mean neurological score of the NDGA group was significantly lower than that of the EAE group (P0.05). The levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-1 in the spinal and spleen tissues of the mice were significantly lower than that in the EAE group (P0.05). The levels of IL-6, IL-12, IL-17 and IFN-were lower in the NDGA group than in the EAE group, and there was a statistical difference (P0.05). The levels of TGF-1 and IL-4 mRNA in the EAE group were lower than that of the EAE group, and there was statistical difference (P0.05). Compared with the control group, the levels of IL-6, IL-12, IL-17 and IFN-mRNA in EAE group were higher in EAE group than in control group, and the level of IL-12, IL-17 and IFN-mRNA in NDGA group was lower than that of EAE group, and there was statistical difference (P0.05). The level of TGF-1 and IL-4 mRNA in the EAE group was lower, and the level of TGF-5 mRNA in NDGA group was higher than that of EAE group. The levels of IL-4, IL-6, IL-12, IL-17, IFN-1 and TGF-2 in the brain of each group were changed. The expression of IL-6, IL-12, IL-17 and IFN-1 in the EAE group was higher than that of the control group at 10 days. The expression of IL-6, IL-12, IL-17 and IFN-1 in the NDGA group was lower than that of the EAE group. There was statistical difference (P0.05). The expression of TGF-1 and IL-4 in EAE group decreased, and the expression of TGF-1 and IL-4 in NDGA group was higher than that of EAE group, and there was statistical difference (P0.05). The expression of IL-17 and IFN-1 in EAE group was higher in EAE group than in control group at 20 days. The expression of IL-17 and IFN-1 in NDGA group was lower than that of EAE group, and there was statistical difference (P0.05). The expression of TGF-1 in NDGA group was higher than that of EAE group, and there was statistical difference (P0.05). Conclusion: The incidence and the response of 1EAE are related to the dynamic changes of the inflammatory factors. The NDGA can reduce the severity of the neurological function damage in the EAE mice, promote the recovery of the nerve function, and have a protective effect on the EAE mice. The 3NDGA can reduce the pro-inflammatory cytokines IL-6, IL-12, IL-17 in the EAE brain, the spinal cord and the spleen tissue. The anti-inflammatory cytokines IL-4 and TGF-1 levels in the anti-inflammatory cytokine IL-4 and TGF-1 levels have an anti-inflammatory effect, and the regulation of the balance of immune inflammation may be one of the mechanisms of NDGA to play a protective role in EAE.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R744.51
【参考文献】
相关硕士学位论文 前1条
1 王珏琼;马索罗酚和丹参酮ⅡA对实验性自身免疫性脑脊髓炎小鼠治疗作用的病理学研究[D];河北医科大学;2014年
，本文编号：2499205
本文链接：https://www.wllwen.com/yixuelunwen/nfm/2499205.html
最近更新
教材专著
