Development of an animal model of opticospinal form of multiple sclerosis using the HLA-DP5 transgenic mice.

使用 HLA-DP5 转基因小鼠开发视脊髓形式的多发性硬化症动物模型。

• 批准号: 14370209
• 负责人: KIRA Jun-ichi
• 金额: $ 9.6万
• 依托单位: Kyushu University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14370209/
• 关键词: multiple sclerosis; opticospinal; hsp105; HLA class II; transgenic mouse; SEREX; HSP105

1.Identification of a novel autoantigen hsp105 in patients, with multiple sclerosisTo discover novel autoantigens in Japanese patients with multiple sclerosis(MS), we immunoscreened spinal cord-derived cDNA library with sera from 8 MS patients(5-10×10^5 clones screened in each patient). One of five positive clones was heat shock protein 105(hsp105), which is expressed most abundantly in the brain. We then examined immune responses to hsp105 in MS patients and found that (1)the frequency of anti-hsp105 IgG antibody increased in MS patients than in controls, (2)hsp105 expression was enhanced in the MS lesions and EAE, (3)significant proliferation of CD4^+CD45RO^+T cell to hsp105 was noted only in MS patients, (4)in the ELISPOT assay, IL-10 response to hsp105 was significantly higher than IL-4 and IFN-gamma responses to hsp105 in both MS patients and healthy controls, however the IL-10 response was significantly lower as compared with controls. These findings suggest the immune response to hsp105 in human and that the response is attenuated in MS patients. Furthermore, we studied immunogenicity of hsp105 in mice and found that (5)immunization with either human or autologous mouse hsp105 did not induce EAE, (6)mice vaccinated with pHSP105showed marked exacerbation of EAE. In future, we are going to analyze the mechanism of exacerbation of EAE by pHSP105.2.Animal model of opticospinal form of multiple sclerosisIn Japan, susceptibility to OS-MS is associated with the HLA-DPB1*0501 allele. HLA-DP5(DPA1*02022/DPB1*0501) transgenic mice were generated by co-injection of the HLA-DPA1*02022/pDOI-6 fragment and HLA-DPB1*0501/pDOI-6 fragment into fertilized mouse eggs(C57BL/6). The resulting mice were tested for integration of the transgene by PCR analysis of tail DNA. Now, transgenic lines derived from three independent founders are tested for expression of HLA-DPA1*02022 RNA and HLA-DPB1*0501 RNA in thymus and spleen.

1.多发性硬化症患者中新的自身抗原hsp 105的鉴定为了在日本多发性硬化症（MS）患者中发现新的自身抗原，我们用8名MS患者的血清免疫筛选脊髓来源的cDNA文库（每个患者筛选5 - 10 × 10^5个克隆）。5个阳性克隆之一是热休克蛋白105（hsp 105），它在脑中表达最丰富。然后，我们检测了MS患者对hsp105的免疫应答，发现（1）MS患者中抗hsp105 IgG抗体的频率比对照组增加，（2）MS病变和EAE中hsp105表达增强，（3）仅在MS患者中观察到CD4 ^+ CD45RO ^+ T细胞向hsp105的显著增殖，（4）在ELISPOT测定中，在MS患者和健康对照中，IL-10对hsp105的应答显著高于IL-4和IFN-γ对hsp105的应答，然而与对照相比，IL-10应答显著较低。这些发现表明，在人类中的热休克蛋白105的免疫反应，并在MS患者的反应减弱。此外，我们研究了hsp 105在小鼠体内的免疫原性，发现（5）人或自体小鼠hsp 105免疫均未诱发EAE;（6）pHSP 105免疫小鼠后EAE明显加重。2.视脊髓型多发性硬化症的动物模型在日本，OS-MS的易感性与HLA-DPB 1 * 0501等位基因相关。通过将HLA-DPA1 * 02022/pDOI-6片段和HLA-DPB 1 * 0501/pDOI-6片段共注射到受精的小鼠卵（C57 BL/6）中来产生HLA-DP 5（DPA1 * 02022/DPB 1 * 0501）转基因小鼠。通过尾部DNA的PCR分析来测试所得小鼠的转基因整合情况。现在，测试来自三个独立建立者的转基因系在胸腺和脾脏中HLA-DPA1 * 02022 RNA和HLA-DPB1 * 0501 RNA的表达。

项目成果

吉良潤一: "多発性硬化症の臨床疫学-環境原因と遺伝原因"日本臨床. 61. 1300-1310 (2003)

Junichi Kira：“多发性硬化症的临床流行病学 - 环境原因和遗传原因”日本临床 61. 1300-1310 (2003)

三野原元澄: "多発性硬化症とheat shock protein 105"日本臨床. 61. 1317-1322 (2003)

Motosumi Minohara：“多发性硬化症和热休克蛋白 105”日本临床杂志 61. 1317-1322 (2003)。

Osoegawa M et al.: "Myelitis with atopic diathesis : a nationwide survey of 79 cases in Japan."J Neurol Sci. (In press).

Osoekawa M 等人：“具有特应性素质的脊髓炎：对日本 79 例病例进行的全国性调查。”J Neurol Sci。

越智博文, 吉良潤一: "Annual Review 神経 2003"柳澤信夫、篠原幸人、岩田誠、清水輝夫、寺本明 編 (中外医学社). 360 (2003)

Hirofumi Ochi、Junichi Kira：“Annual Review Neurology 2003”，由 Nobuo Yanagisawa、Yukito Shinohara、Makoto Iwata、Teruo Shimizu 和 Akira Teramoto 编辑（Chugai Igakusha）（2003 年）。

吉良潤一: "日本の多発性硬化症"Current Insights in Neurological Science -神経内科・脳神経外科・精神科の最新トピックス. (In press).

Junichi Kira：“日本的多发性硬化症”神经科学的最新见解 - 神经病学、神经外科和精神病学的最新主题（正在出版）。