Development of novel therapy for immune-mediated hematological diseases targeted for phagocytosis and complement regulatory protein

主要研究结果如下：克隆了免疫介导的溶血性贫血（immunemodional hemolytic anemia，IMHA）中最常见的自身抗原带3和血型糖蛋白（glycophorin）的cDNA，并研究了它们与犬疾病的关系。我们还克隆了被认为是噬血细胞作用抑制剂的犬CD 47分子的cDNA，并检测了其在IMHA犬的红细胞和白色血细胞上的表达。正常犬红细胞和淋巴细胞上均有CD 47表达，再生红细胞中CD 47表达水平升高。这些结果表明，再生的红细胞中含有大量的CD 47分子，可能具有抗噬血作用的能力。此外，随着红细胞贮存时间的延长，犬红细胞表面CD 47的表达水平逐渐降低，表明CD 47的表达可能与红细胞贮存性疾病有关。P-gP与细胞裂解的关系也有报道。本文对犬P-gP及编码P-gP蛋白的mdr 1基因进行了基础研究。我们实验性地将mdr 1基因导入犬细胞，并证实其表达水平和功能。此外，我们检测了牧羊犬细胞P-gP的表达水平，并对犬IMHA和IMTP的临床病例进行了临床病理学研究，以了解其预后因素并建立评分系统。然而，在该项目中获得的结果包含有价值的信息，这可能是有用的狗和猫的免疫介导的血液病的理解。

The summary of our research results are as follows:We cloned cDNA of Band 3 and glycophorin, which are the most known self antigen in immune-mediated hemolytic anemia (IMHA), and examine its relationship with canine diseases. We also cloned cDNA of canine CD47 molecule, which considered as inhibitor for hemophagocytosis, and examine its expression on red and white blood cells of dogs with IMHA. CD47 was expressed on RBC and lymphocytes in healthy dogs and its expression level was increased in regenerated RBC. These results indicated that regenerated RBC with high amount of CD47 molecules may resistant to hemophagocytosis. Furthermore, the expression level of CD47 on canine RBC was gradually decreased by RBC storage, indicating that the CD47 expression may related with RBC storage diseases.P-glycoprotein (P-gP) is a membrane protein which considered as chemoresistance. P-gP also was reported its relationship to cell lysis. We investigated several basic research on canine P-gP and mdr1 gene which codes P-gP protein. We experimentally introduce mdr1 gene into canine cells and confirm its expression level and function. Furthermore, we examine the expression level of P-gP on cells of Collie dog.We also performed clinicopathological study on clinical cases of canine IMHA and IMTP to know the prognostic factors and to develop scoring system.We can not develop novel therapy for immune-mediated hematological diseased of dogs and cats during the project term (2006-2007). However, the results obtained in the project contain valuable information which might be useful for understanding of immune-mediated hematological diseases of dogs and cats.