Establishment of an animal model for P-ANCA positive crescentic glomerulonephritis

P-ANCA阳性新月体肾小球肾炎动物模型的建立

基本信息

批准号：
10671016
负责人：
KANEOKA Hidetoshi
金额：
$ 1.92万
依托单位：
Fukuoka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10671016/
关键词：
RPGN P-ANCA HLA SCID mouse Myeloperoxidase Animal model T cell epitope Autoimmune disease

项目摘要

Primary crescentic glomerulonephritis is rapidly progressive and poorly prognostic. The treatment depends mainly on glucocorticoid hormone and imuunosuppressants. Animal models play pivotal roles to establish new strategy for therapy of intractable of intractable diseases. In this study, I tried to raise severe combined immunodeficiency (SCID) mice carrying perinuclear anti-neutrophil cytoplasmic antibody (P-ANCA) and crescentic glomerulonephritis (GN).1. Association between P-ANCAGN and HLA class II genotypes.Utilizing PCR-RFLP and PCR-SSO, I determined HLA class II genotypes of 34 Japanese patients with P-ANCAGN. I found significant of P-ANCAGN with HLA-DRB1ィイD1*ィエD10803 (Odd's ratio 2.5 p<0.01) and with HLA-DQB1ィイD1*ィエD10601 (Odd's ratio 26 p<0.01), DRB1ィイD1*ィエD10803 and DQB1ィイD1*ィエD10601 are in linkage disequiribrium in Japanese population.2. Detection of human P-ANCA in patients and mice.Solid phase enzyme linked immunoadsorbent assay was used to detect human P-ANCA. In brief, pur … More ified myeloperoxidase (MPO) was immobilized on a plastic microtiter plate, sample serum was applied, wells were washed extensively, alkaline phosphatase labeled goat anti-human IgG or IgA antibody was added, and PNPP was added as substrate. The titers determined at my lab were well correlated with those from SRL laboratory.3. MPO specific T cell proliferation.Peripheral lymphocytes were cocultured with purified MPO for 7 days, and the proliferation was assayed by uptake of tritiated thymidine. No T cell specific response was demonstrated.4. SCID mouse producing P-ANCA.Peripheral lymphocytes from P-ANCAGN were transferred to 8-week-age SCID mice. The body weight, systolic and diastolic blood pressure, urine proteinuria, human IgG and IgM, and human P-ANCA were monitored every two weeks. The mice were sacrificed at 12 weeks of follow-up. Human IgG and IgM were detected in mice with human lymphocytes, but P-ANCA only in those with patient lymphocytes. The mice died before end point, and all were mice transferred patient lymphocyte. No mice showed proteinuria or hypertension suggesting P-ANCAGN, Pathological examination is under investigation. Less

原发性新月肾小球肾炎是迅速进行的，预后不佳。该治疗主要取决于糖皮质激素和肌抑制剂。动物模型扮演着关键角色，以建立治疗顽固性疾病的新策略。在这项研究中，我试图提高携带核周抗中性嗜性胞质抗体（P-ANCA）和新月肾小球肾炎（GN）的严重组合免疫缺陷（SCID）小鼠。1。 P-cancagn和HLA II类基因型之间的关联。利用PCR-RFLP和PCR-SSO，我确定了34例日本P-cancagn患者的HLA II类基因型。我发现与HLA-DRB1II D1*IE D10803（奇数比2.5 p <0.01）以及HLA-DQB1II D1*IE D10601（奇数比26 p <0.01），DRB1II D1 D1*II D10803和DQB1II D101*II d10601 II d10601，我发现了p-cancagn的意义。人口2。检测患者和小鼠的人类p-anca。固相酶连接的免疫吸附测定法用于检测人类p-anca。简而言之，将更多的骨过氧化物酶（MPO）固定在塑料微量盘板上，施加样品血清，对井进行广泛洗涤，碱性磷酸酶标记为山羊抗人类IgG或IgA抗体，并加入PNPP，并添加PNPP为substrate。在我的实验室确定的滴度与SRL实验室的滴度相关。3。将MPO特异性T细胞增殖。将外淋巴细胞与纯化的MPO共培养7天，并通过摄取tri的胸苷的摄取来分配增殖。未证明T细胞特异性响应4。 SCID小鼠产生的P-ANCA。来自P-Cancagn的外膜淋巴细胞转移到8周时代的SCID小鼠中。每两周监测体重，收缩压和舒张压，尿液蛋白尿，人IgG和IgM以及人P-anca。在随访的12周时处死小鼠。在患有人淋巴细胞的小鼠中检测到人IgG和IgM，但仅在患有患者淋巴细胞的人中P-ANCA。小鼠在终点前死亡，所有小鼠均转移患者淋巴细胞。没有小鼠表现出蛋白质症或高血压表明p- cancagn，正在研究病理检查。较少的