Combined NSC Grafting and Neurogenic Drug Therapy for Temporal Lobe Epilepsy

描述(由申请人提供)： 细胞移植策略能够减少慢性颞叶癫痫(TLE)患者的自发性复发癫痫(SRS)、学习记忆功能障碍和抑郁具有重要意义，因为约35%的TLE患者存在对抗癫痫药物(AED)耐药的SRS，而AED治疗不能减少认知和情绪功能障碍。由于TLE与分泌抗惊厥蛋白胶质细胞源性神经营养因子(GDNF)的抑制性GABA能神经元和星形胶质细胞有关， 有能力同时产生GABA能神经元和GDNF+星形胶质细胞的细胞移植似乎在抑制SRS方面具有吸引力。事实上，我们最近的研究表明，将内侧神经节隆起(MGE)来源的神经干细胞(NSCs)移植到慢性TLE大鼠的海马区，可以显著降低SRS的频率和强度，增加新的GABA能神经元和GDNF+星形胶质细胞。然而，在海马神经发生或认知功能方面没有看到改善。对移植物的分析表明，移植物来源的细胞的总产率仅为注射细胞的28%，移植到TH神经源性颗粒下带(SGZ)的细胞很少。这一发现表明，MGE-NSC移植后在神经发生和认知方面缺乏改善是移植物来源细胞产量较低和迁移减少的结果。我们推测，将MGE-NSCs移植到海马区和外周应用神经源性化合物将极大地减轻SRS，并逆转慢性TLE中出现的记忆障碍和抑郁。我们认为，这种功能恢复的改善将与大量移植物衍生细胞移植到SGZ，从移植物中产生更多新的GABA能神经元和GDNF+星形胶质细胞，恢复宿主海马GDNF浓度，增加海马神经发生，以及对认知和情绪功能以及神经发生重要的蛋白质/基因的浓度/表达正常化有关。在这个项目中，我们将研究联合MGE-NSCs海马区移植和神经源性药物治疗策略是否能显著减轻慢性TLE大鼠的SRS、认知功能障碍和抑郁。在具体目标1中，我们将确定MGE-NSCs和皮下(S.C.)联合移植的效果。成纤维细胞生长因子2治疗14天。在特定的目标2中，我们将检测MGE-NSCs海马区内移植和sc神经类固醇类似物甘纳松酮的联合作用。我们将测试这些联合疗法所介导的癫痫抑制和认知和情绪功能的改善是否与移植物衍生的GABA能神经元和GDNF阳性星形胶质细胞的产量增加、大量移植物衍生的神经干细胞植入神经源性SGZ、内源性神经干细胞和海马神经发生的活性增加以及认知和情绪功能至关重要的蛋白质/基因的表达增强有关。我们还将把这些结果与MGE来源的GABA能细胞和MGE来源的NSC来源的星形胶质细胞移植的数据进行比较，这些细胞移植有或没有使用成纤维细胞生长因子-2/加那索龙治疗。建议的研究有望为考虑神经干细胞移植策略治疗慢性TLE的临床应用提供必要的关键基准。这些研究也与退伍军人的医疗保健需求相关，因为在伊拉克和阿富汗战争中服役的士兵中，与战斗有关的中到重度头部损伤可能会在未来几年导致慢性TLE。

DESCRIPTION (provided by applicant): PROJECT SUMMARY Cell transplantation strategies capable of diminishing spontaneous recurrent seizures (SRS), learning and memory dysfunction and depression in chronic temporal lobe epilepsy (TLE) have great significance because ~35% of patients with TLE have SRS that are resistant to antiepileptic drugs (AEDs) and AED therapy does not reduce cognitive and mood dysfunction. As TLE is associated with a paucity of inhibitory GABA-ergic neurons and astrocytes secreting the anticonvulsant protein glial cell line-derived neurotrophic factor (GDNF), cell grafts that have the ability to give rise to both GABA-ergic neurons and GDNF+ astrocytes appear attractive for restraining SRS. Indeed, our recent study has shown that grafting of the medial ganglionic eminence (MGE) derived neural stem cells (NSCs) into the hippocampus of rats exhibiting chronic TLE substantially reduces the frequency and intensity of SRS with additions of new GABA-ergic neurons and GDNF+ astrocytes. However, no improvements were seen in the hippocampal neurogenesis or cognitive function. Analyses of grafts revealed that the overall yield of graft-derived cells was only 28% of injected cells and engrafting of cells into th neurogenic subgranular zone (SGZ) was minimal. This finding suggested that lack of improvements in neurogenesis and cognition after MGE-NSC grafting is a consequence of both lower yield and reduced migration of graft- derived cells. We hypothesize that combined grafting of MGE-NSCs into the hippocampus and peripheral administration of neurogenic compounds will greatly diminish SRS as well as reverse memory dysfunction and depression seen in chronic TLE. We propose that such improved functional recovery will be associated with engrafting of significant numbers of graft-derived cells into the SGZ, generation of greater numbers of new GABA-ergic neurons and GDNF+ astrocytes from grafts, restoration of the host hippocampal GDNF concentration, increased hippocampal neurogenesis, and normalization in the concentration/expression of proteins/genes important for cognitive and mood function and neurogenesis. In this project, we will investigate whether combined intrahippocampal grafting of MGE-NSCs and neurogenic drug treatment strategies would greatly diminish SRS, cognitive dysfunction and depression in a rat model of chronic TLE. In Specific Aim 1, we will ascertain the effects of combined intrahippocampal grafting of MGE-NSCs and subcutaneous (S.C.) FGF-2 treatment for 14 days. In Specific Aim 2, we will examine the effects of combined intrahippocampal grafting of MGE-NSCs with S.C. administration of neurosteroid analog ganaxolone. We will test whether seizure-suppression and improved cognitive and mood function mediated by these combination therapies are linked to an increased yield of graft-derived GABA-ergic neurons and GDNF-positive astrocytes, engrafting of greater numbers of graft-derived NSCs into the neurogenic SGZ, increased activity of endogenous NSCs and hippocampal neurogenesis, and enhanced expression of proteins/genes vital for cognitive and mood functions. We will also compare these results with data obtained from grafts of MGE-derived GABA-ergic cells and MGE-NSC derived astrocyte grafts in the presence or absence of FGF-2/Ganaxolone treatment. The proposed studies have promise for providing the critical benchmarks essential for considering clinical application of NSC grafting strategies for treating chronic TLE. The studies are also relevant to the health care needs of Veterans, as combat related moderate to severe head injuries among soldiers who served in Iraq and Afghanistan wars might lead to chronic TLE in the coming years.