馬索羅酚（NDGA）對實驗性自身免疫性腦脊髓炎小鼠炎癥因子表達的影響
發(fā)布時間：2019-06-14 06:58
【摘要】：目的:多發(fā)性硬化(MS)是一種以炎性脫髓鞘和軸索損傷為特征的免疫介導(dǎo)的中樞神經(jīng)系統(tǒng)疾病。多發(fā)性硬化的病因和機制尚不明確,但炎癥是這個疾病早期最重要的環(huán)節(jié)。各種細胞因子以自分泌或旁分泌方式,組成了一個復(fù)雜的動態(tài)網(wǎng)絡(luò)結(jié)構(gòu),參與了免疫反應(yīng)的所有過程及階段,在調(diào)節(jié)免疫反應(yīng)和組織修復(fù)中起著關(guān)鍵性作用,決定著多發(fā)性硬化的病情嚴重程度及發(fā)展趨勢。我們課題組前期對Nrf2通路激活劑萊菔硫烷的研究中發(fā)現(xiàn)其增強小鼠抗氧化應(yīng)激能力和調(diào)節(jié)免疫機制,在實驗性自身免疫性腦脊髓炎(EAE)小鼠中起到神經(jīng)保護作用。然而萊菔硫烷尚未應(yīng)用于臨床,通過文獻查閱,我們發(fā)現(xiàn)了可應(yīng)用于人體的Nrf2通路激活劑——馬索羅酚。馬索羅酚在MS的免疫調(diào)節(jié)機制中的作用尚未明確,需要進一步的研究。本實驗以馬索羅酚作為干預(yù)藥物,觀察EAE小鼠及馬索羅酚治療小鼠發(fā)病嚴重程度及病程的影響。研究馬索羅酚對EAE小鼠IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β表達的調(diào)節(jié)作用。進而探索相關(guān)細胞因子在EAE病程中的作用,并研究馬索羅酚在EAE免疫學(xué)機制中是否發(fā)揮神經(jīng)保護作用。方法:將54只8至10周雌性C57BL/6小鼠,體重18g至20g,隨機分成control組、EAE組和NDGA組,各組分別為18只。每組分為2個亞組,10天組及20天組,各亞組為9只。建立EAE動物模型。NDGA組小鼠自發(fā)病(weaver評分≥1分)當天(記為第0天),每只每日腹腔注射NDGA溶液10mg/kg。EAE組及control組小鼠自第0天,每只每日腹腔注射5%DMSO 10ml/kg。每日一次,直至處死。每天2次對其進行稱重和神經(jīng)功能的評分。利用實時定量PCR方法檢測脊髓及脾中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β表達水平,用ELISA方法檢測腦組織中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β表達水平。結(jié)果:1在發(fā)病第10天及第20天,NDGA組平均神經(jīng)功能評分均較EAE組顯著降低(P0.05)。2各組小鼠脊髓及脾組織中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β的m RNA水平:在發(fā)病10天時,與Control組對比,EAE組IL-6、IL-12、IL-17、IFN-γm RNA水平均增高,NDGA組IL-6、IL-12、IL-17、IFN-γm RNA水平較EAE組低,并有統(tǒng)計差異(P0.05)。EAE組TGF-β、IL-4 m RNA水平降低,NDGA組TGF-β、IL-4 m RNA水平較EAE組高,并有統(tǒng)計差異(P0.05)。在發(fā)病20天時,與Control組對比,EAE組IL-6、IL-12、IL-17、IFN-γm RNA水平均增高,NDGA組IL-12、IL-17、IFN-γm RNA表達較EAE組低,并有統(tǒng)計差異(P0.05)。EAE組TGF-β、IL-4 m RNA水平降低,NDGA組TGF-βm RNA水平較EAE組高,并有統(tǒng)計差異(P0.05)。3各組小鼠腦組織中IL-4、IL-6、IL-12、IL-17、IFN-γ、TGF-β含量變化情況:在發(fā)病10天時,與Control組對比,EAE組IL-6、IL-12、IL-17、IFN-γ的表達均增高,NDGA組IL-6、IL-12、IL-17、IFN-γ的表達較EAE組低,并有統(tǒng)計差異(P0.05)。EAE組TGF-β、IL-4的表達降低,NDGA組TGF-β、IL-4的表達較EAE組高,并有統(tǒng)計差異(P0.05)。在發(fā)病20天時,與Control組對比,EAE組IL-17、IFN-γ的表達均增高,NDGA組IL-17、IFN-γ表達較EAE組低,并有統(tǒng)計差異(P0.05)。NDGA組TGF-β的表達較EAE組高,并有統(tǒng)計差異(P0.05)。結(jié)論:1 EAE的發(fā)病和緩解與炎癥因子的動態(tài)變化相關(guān)。2 NDGA可以減輕EAE小鼠神經(jīng)功能損害嚴重程度,促進神經(jīng)功能恢復(fù),對EAE小鼠具有保護作用。3 NDGA可以降低EAE腦、脊髓及脾組織中促炎癥細胞因子IL-6、IL-12、IL-17、IFN-γ水平,增加抗炎癥細胞因子IL-4和TGF-β水平,具有免疫抗炎作用。4調(diào)節(jié)免疫炎癥平衡可能是NDGA對EAE發(fā)揮保護作用的機制之一。
[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.
【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R744.51
本文編號：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.
【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R744.51
【參考文獻】
相關(guān)碩士學(xué)位論文 前1條
1 王玨瓊;馬索羅酚和丹參酮ⅡA對實驗性自身免疫性腦脊髓炎小鼠治療作用的病理學(xué)研究[D];河北醫(yī)科大學(xué);2014年
，本文編號：2499205
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