【作者】 李娟；

【导师】 刘洪波；

【作者基本信息】 郑州大学 ， 神经病学， 2021， 硕士

【摘要】 背景和目的视神经脊髓炎谱系疾病(neuromyelitis optica spectrum disorders,NMOSD)是一种区别于多发性硬化(multiple sclerosis,MS)的中枢神经系统脱髓鞘性疾病,特点是严重的视神经炎或者纵向广泛的横贯性脊髓炎,二者也可同时存在。致病性血清抗水通道蛋白-4(aquaporin4,AQP-4)抗体的发现促进了 NMOSD的研究,国际 NMO 诊断小组(International Panel for NMO diagnosis,IPND)提出了基于AQP-4抗体的NMSOD分层诊断。然而,研究表明,单独转移抗AQP-4抗体不能在实验动物中诱导NMO病理改变,与特异性T细胞共转移可诱导中枢神经系统炎性病变,这表明T细胞在NMOSD的潜在发病机制中可能扮演着重要角色。越来越多的免疫学研究报道细胞因子和趋化因子在NMOSD的发病机制中起重要作用,并表明NMOSD是一种2型辅助性T细胞(T helper cells,Th2)和Th17相关疾病。白细胞介素19(interleukin-19,IL-19)是IL-10家族的成员之一,是一种具有多功能的细胞因子,主要由免疫细胞如活化的淋巴细胞、巨噬细胞和单核细胞分泌。细菌脂多糖(LPS)可刺激这些细胞中IL-19的表达,并且这种作用通过Th2细胞因子IL-4和IL-13的处理而增强;另外,IL-19与靶细胞上IL-20R1/IL-20R2结合,激活参与Th17细胞分化的信号转导子和转录激活子(signal transducer and activator of transcription,STAT)3 磷酸化,因此,IL-19的产生与炎症引起Th2和Th17细胞应答有关,暗示了从过敏性疾病到自身免疫性疾病等广泛的免疫系统的影响。白细胞介素26(IL-26)也属于IL-10家族,主要来源于上皮细胞和免疫细胞,包括肺泡巨噬细胞、Th1和Th17细胞、NK细胞和巨噬细胞样滑膜细胞。IL-26是一种与Th17相关的细胞因子,在Th17细胞中与IL-22高水平共表达,受Th17细胞相关细胞因子IL-1β、IL-23和维甲酸相关孤儿受体(retinoid-related orphan receptor,ROR)γ t 调节,它通过由 IL-20R1 和 IL-10R2 链组成的异二聚体受体复合物发出信号,并诱导Janus激酶和信号转导子与转录激活子(Janus kinase-signal transducer and activator of transcription,JAK-STAT)激活,导致STAT1和STAT3磷酸化。二者均在炎症性肠病、类风湿关节炎、银屑病等多种自身免疫性疾病中发挥重要作用,但在NMOSD的发生发展中的作用尚不明确,因此,本研究通过检测NMOSD患者血清和脑脊液中IL-19和IL-26的浓度并分析二者与其他临床指标之间的相关性,初步探索其在NMOSD不同阶段的作用,以期为NMOSD的诊断和治疗提供进一步的理论基础。方法本研究为病例对照研究。收集2019年9月-2020年12月在郑州大学第一附属医院神经内科就诊的视神经脊髓炎谱系疾病患者25例(男性6例,女性19例),分别于急性期和缓解期抽取肘静脉血标本。同时于住院期间获得急性期脑脊液标本12例。详细记录患者病历资料,包括姓名、性别、年龄、病史、入院扩展残疾状态量表(Expanded Disability Status Scale,EDSS)评分、实验室检查结果、影像学检查结果、神经电生理检查结果等。选取同时期于我院门诊就诊的18例健康志愿者作为对照组,其中男性6例,女性12例;另外,收集同时期于我院就诊的8例其他非炎性神经系统疾病(other non-inflammatory neurological diseases,ONND)患者(包括:原发性头痛、精神心理性疾病)的脑脊液作为ONND组。应用酶联免疫吸附法(Enzyme-linked Immunosorbent Assay,ELISA)检测所有标本中IL-19和IL-26的浓度,分析二者在各组中的差异,并分析与其他临床指标之间的相关性。所有数据采用SPSS26.0统计软件进行分析。定量资料均用均数±标准差表示;正态分布资料组间比较采用配对t检验或者两独立样本t检验;非正态分布资料组间比较采用非参数检验;两组定量资料的相关性采用Pearson或Spearman相关分析;卡方检验或Fisher精确概率法比较两组二分类资料;P<0.05表示差异有统计学意义。结果1.血清IL-19水平在NMOSD患者急性期为10.70±3.63pg/ml,缓解期为15.22±4.44pg/ml,对照组为7.68±2.18pg/ml。急性期IL-19的水平高于健康对照组(P=0.003);缓解期IL-19的水平高于健康对照组(P<0.001);缓解期IL-19的水平高于急性期(P<0.001)。2.血清IL-26水平NMOSD患者急性期为42.87±10.55pg/ml,缓解期为30.99±5.12pg/ml,对照组为27.84±5.72pg/ml。急性期IL-26的水平高于缓解期和健康对照组(P<0.001);缓解期IL-26的水平略高于健康对照组,但差异无统计学意义(P=0.066);3.急性期血清IL-19的水平与患者EDSS评分呈负相关(r=0.601,P=0.001)。与脊髓病变长度无明显相关性(r=0.014,P=0.952)。急性期血清IL-26的水平与患者EDSS评分和脊髓病变长度均无明显相关性(P>0.05)。4.急性期血清IL-19和IL-26的水平与血清AQP-4状态无关(P>0.05)。5.急性期、缓解期血清IL-19与IL-26的水平均不相关(P>0.1)。6.急性期脑脊液IL-19的水平为31.59±3.84pg/ml,ONND组为24.70±5.69pg/ml,急性期高于ONND组(P=0.017)。脑脊液IL-26的水平为78.54±17.74pg/ml,ONND组为29.05±7.45pg/ml,急性期高于ONND组(P=0.001)。结论1.血清IL-19的水平在缓解期升高更显著,提示IL-19主要在缓解期起抗炎作用。急性期血清IL-19的水平与EDSS评分呈负相关,提示IL-19在急性期也可发挥保护作用,并可限制炎症的严重程度,有望成为潜在的生物学标志物和治疗靶点。2.血清和脑脊液IL-26的水平在急性期均升高,提示IL-26主要在疾病急性期发挥作用。但与EDSS评分及累及脊髓节段的长度无明显相关,推测其与疾病严重程度无关,可能通过调控炎症反应中的其他途径参与疾病的进展。更多还原

【Abstract】 Background and objectiveNeuromyelitis optica spectrum disorders(NMOSD)is a demyelinating disease of the central nervous system that is different from multiple sclerosis(MS).It is characterized by severe optic neuritis or extensive longitudinal transverse spinal cord Inflammation,the two can also exist at the same time.The discovery of pathogenic serum anti-aquaporin 4(AQP-4)antibodies promoted the research of NMOSD.The International Panel for NMO diagnosis(IPND)proposed a NMSOD score based on AQP-4 antibodies.Layer diagnosis.However,studies have shown that transfer of anti-AQP-4 antibody alone cannot induce NMO pathological changes in experimental animals,and co-transfer of specific T cells can induce central nervous system inflammatory lesions,which indicates that T cells may be involved in the potential pathogenesis of NMOSD.Plays an important role.More and more immunological studies have reported that cytokines and chemokines play an important role in the pathogenesis of NMOSD,and indicate that NMOSD is a type 2 T helper cells(Th2)and Th17 related diseases.Interleukin-19(IL-19)is a member of the IL-10 family.It is a multifunctional cytokine,mainly composed of immune cells such as activated lymphocytes,macrophages and monocytes.secretion.Bacterial lipopolysaccharide(LPS)can stimulate the expression of IL-19 in these cells,and this effect is enhanced by the treatment of Th2 cytokines IL-4 and IL-13;in addition,IL-19 interacts with IL-20R1/IL-20R2 binds to activate signal transducer and activator of transcription(STAT)3 phosphorylation involved in Th17 cell differentiation.Therefore,the production of IL-19 is related to the response of Th2 and Th17 cells caused by inflammation.It implies a wide range of immune system influences from allergic diseases to autoimmune diseases.Interleukin 26(IL-26)also belongs to the IL-10 family,mainly derived from epithelial cells and immune cells,including alveolar macrophages,Thl and Th17 cells,NK cells and macrophage-like synovial cells.IL-26 is a Thl7-related cytokine.It is co-expressed with IL-22 in Th17 cells at high levels.It is affected by Th17 cell-related cytokines IL-1β,IL-23,and retinoic acid-related orphan receptors(retinoid-related receptors).orphan receptor(ROR)yt regulation,which sends signals through the heterodimeric receptor complex composed of IL-20R1 and IL-10R2 chains,and induces Janus kinase,signal transducer and transcription activator(Janus kinase-signal).transducer and activator of transcription,JAK-STAT)activates,leading to phosphorylation of STAT1 and STAT3.Both of them play an important role in various autoimmune diseases such as inflammatory bowel disease,rheumatoid arthritis,psoriasis,etc.,but their role in the occurrence and development of NMOSD is still unclear.Therefore,this study tested NMOSD patients Concentrations of IL-19 and IL-26 in serum and cerebrospinal fluid and analyze the correlation between the two and other clinical indicators,preliminary explore their role in different stages of NMOSD,in order to provide a further theoretical basis for the diagnosis and treatment of NMOSD.MethodsThis study is a case-control study.Collected 25 patients with neuromyelitis optica spectrum disease(6 males and 19 females)who attended the Department of Neurology of the First Affiliated Hospital of Zhengzhou University from September 2019 to December 2020,and collected blood from the cubital veins in the acute and remission phases.specimen.At the same time,12 cases of acute cerebrospinal fluid specimens were obtained during hospitalization.Record patient medical history data in detail,including name,gender,age,medical history,admission to the Hospital Expanded Disability Status Scale(EDSS)score,laboratory test results,imaging test results,neuroelectrophysiological test results,etc.Eighteen healthy volunteers who visited our hospital during the same period were selected as the control group,including 6 males and 12 women.In addition,8 cases of other non-inflammatory neurological diseases(other non-inflammatory neurological diseases)who visited our hospital during the same period were collected.The cerebrospinal fluid of patients with inflammatory neurological diseases(ONND)(including:primary headache,mental illness)was regarded as the ONND group.Use Enzyme-linked Immunosorbent Assay(ELISA)to detect the concentration of IL-19 and IL-26 in all samples,analyze the difference between the two in each group,and analyze the correlation with other clinical indicators.All data were analyzed using SPSS26.0 statistical software.Quantitative data are expressed as mean ± standard deviation;paired t test or two independent sample t test is used for comparison between normal distribution data groups;nonparametric test is used for non-normal distribution data comparison between groups;the correlation between two groups of quantitative data is used Pearson or Spearman correlation analysis;Chi-square test or Fisher’s exact probability method to compare the two groups of binary data;P<0.05 indicates that the difference is statistically significant.Results 1.Serum IL-19 level in the acute phase of NMOSD patients is 10.70±3.63pg/ml,the remission phase is 15.22±4.44pg/ml,and the control group is 7.68±2.18pg/ml.The level of IL-19 in the acute phase was higher than that of the healthy control group,and the difference was statistically significant(P=0.003);the level of IL-19 in the remission phase was higher than that of the healthy control group,and the difference was statistically significant(P<0.001);The level of IL-19 was higher than that in the acute phase,and the difference was statistically significant(P<0.001).2.Serum IL-26 level in NMOSD patients is 42.87±10.55pg/ml in acute phase,30.99±5.12pg/ml in remission phase,and 27.84±5.72pg/ml in control group.The level of IL-26 in the acute phase was higher than that in the remission period and the healthy control group,and the difference was statistically significant(P<0.001);the level of IL-26 in the remission period was slightly higher than that of the healthy control group,but the difference was not statistically significant(P=0.066);3.The level of serum IL-19 in the acute phase was negatively correlated with the patient’s EDSS score(r=0.601,P=0.001),There is no significant correlation with the length of spinal cord lesions(r=0.014,P=0.952).The level of serum IL-26 in the acute phase was not significantly correlated with the patient’s EDSS score and the length of spinal cord lesions(P>0.05).4.The levels of serum IL-19 and IL-26 in the acute phase have nothing to do with the status of serum AQP-4(P>0.05).5.The levels of serum IL-19 and IL-26 in the acute and remission phases are not correlated(P>0.1).6.The level of IL-19 in the cerebrospinal fluid in the acute phase was 31.59±3.84pg/ml,and that in the ONND group was 24.70±5.69pg/ml,which was higher in the acute phase than in the ONND group,and there was a statistical difference(P=0.017).The level of IL-26 in the cerebrospinal fluid was 78.54±17.74pg/ml,and it was 29.05±7.45pg/ml in the ONND group,which was higher in the acute phase than in the ONND group,and there was a statistical difference(P=0.001).Conclusion1.The level of serum IL-19 increased significantly in the remission period,suggesting that IL-19 mainly plays an anti-inflammatory effect in the remission period.The level of serum IL-19 in the acute phase is negatively correlated with the EDSS score,suggesting that IL-19 can also play a protective role in the acute phase and limit the severity of inflammation.It is expected to become a potential biological marker and therapeutic target.2.The levels of IL-26 in serum and cerebrospinal fluid both increase in the acute phase,suggesting that IL-26 mainly plays a role in the acute phase of the disease.However,it is not significantly related to the EDSS score and the length of the spinal cord segment involved.It is speculated that it has nothing to do with the severity of the disease,and may participate in the progression of the disease through other ways of regulating the inflammatory response.更多还原