The role of microglia/macrophages and their therapeutic use in Krabbe's disease

描述（由申请人提供）：克拉伯病，或球状细胞白质营养不良，是由溶酶体酶半乳糖脑苷酶（GALC）基因编码突变引起的，GALC是在髓磷脂形成的少突胶质细胞和雪旺细胞中消化半乳糖脑苷所必需的。这种快速进展的脱髓鞘疾病的病理特征是神经系统中巨噬细胞（球状细胞）的显著积累。然而，这些细胞的功能尚不清楚。迄今为止，骨髓干细胞和脐带血细胞移植是唯一被证明有益的治疗方法，供体来源的小胶质细胞可能提供酶。然而，大多数婴儿发病的患者无法治疗，并在两岁时死亡。我们的长期目标是通过加强酶替代和延长成功治疗的时间窗口，为任何阶段的患者设计一种有效的治疗方法。有GALC基因突变的抽搐鼠（twi）被认为是克拉伯氏病的真正模型。最近，我们通过将twi和巨噬细胞缺陷的成骨小鼠（op）杂交，创造了巨噬细胞缺陷的双胞胎小鼠（twi+op），巨噬细胞缺陷的成骨小鼠（op）是唯一一种消除巨噬细胞的克拉伯病模型。我们对twi+op的初步研究表明，小胶质细胞/巨噬细胞的缺失加剧了twi的脱髓鞘病理。我们还表明，当双少突胶质细胞在体内获得酶时，它们可以维持很长一段时间。指导这一建议的具体假设是，小胶质细胞/巨噬细胞在twi脱髓鞘病理中是有益的，这些细胞可以作为治疗载体，为突变的髓鞘细胞提供GALC。为了验证这些假设，我们提出了三个具体目标。1)确定小胶质细胞/巨噬细胞在双胞胎脱髓鞘病理中的作用。采用多学科方法，将分析twi+op中加剧的脱髓鞘，并探索潜在的分子机制。2)确定局部或全身移植小胶质细胞和巨噬细胞作为酶替代手段是否可以缓解twi脱髓鞘。3)确定过表达galc的骨髓干细胞与感染galc基因编码慢病毒的脐带血细胞移植是否优于常规的双胞骨髓移植。这些综合数据将为设计治疗该疾病所有阶段患者的治疗方法提供必要的基础。

DESCRIPTION (provided by applicant): Krabbe's disease, or globoid cell leukodystrophy, is caused by mutations in the gene coding for the lysosomal enzyme, galactocerebrosidase (GALC), which is required to digest galactocerebroside in myelin forming oligodendrocytes and Schwann cells. A pathological hallmark of this rapidly progressive demyelinating disease is the marked accumulation of macrophages (globoid cells) in the nervous system. However, the function of these cells is unclear. To date, transplantation of bone marrow stem cells and umbilical cord blood cells are the only treatments that have proven beneficial, with donor-derived microglia potentially supplying enzyme. However, the majorities of patients with infantile onset are untreatable and succumb by two years of age. Our long term goal is to devise an efficient therapy for patients at any stage of the disease by enhancing enzyme replacement and by extending the time window for successful therapy. The twitcher mouse (twi), which has a mutation in the GALC gene, is considered a bonafide model of Krabbe's disease. Recently we created macrophage-deficient twi mice (twi+op) by crossing twi and macrophage-deficient osteopetrotic mice (op), the only model of Krabbe's disease from which macrophages are eliminated. Our preliminary study of twi+op indicates that the absence of microglia/macrophages exacerbates the demyelinating pathology of twi. We have also shown that twi oligodendrocytes can be maintained for long periods of time when they have access to enzyme in vivo. The specific hypotheses guiding this proposal are that microglia/macrophages are beneficial in the demyelinating pathology of twi and these cells can be used as a therapeutic vehicle to supply GALC to mutant myelinating cells. To test these hypotheses, we propose three specific aims. 1) Determine the roles of microglia/macrophages in the demyelinating pathology of twi. Using a multidisciplinary approach, the exacerbated demyelination in twi+op will be analyzed and the underlying molecular mechanisms will be explored. 2) Determine whether microglia and macrophages transplanted locally or systemically as a means of enzyme replacement can alleviate the demyelination of twi. 3) Determine whether transplantation of GALC-overexpressing bone marrow stem cells and umbilical cord blood cells infected with a GALC-gene-encoding lentivirus can outperform the conventional bone marrow transplantation in twi. These combined data will provide the foundation required to devise therapeutic approaches to treating patients at all stages of the disease.