Innate and Adaptive Immunity in Childhood Epilepsy

描述（由申请人提供）：儿童期开始的顽固性癫痫的长期神经系统后遗症确实是毁灭性的。传统的抗癫痫药物不仅不足以控制癫痫发作，而且可能造成有害的副作用，进一步加重癫痫患儿的发展负担。预防癫痫进展和限制癫痫发作相关细胞损伤的替代疗法是迫切需要的。先天免疫反应失调、外周炎症细胞浸润和血脑屏障的破坏与癫痫发作的发生、发展和持续有关。最近在我们实验室进行的一项全基因组搜索，以确定新的药物靶点，显示促炎分子可能是癫痫发生过程中的治疗靶点。我们已经开发了一种新的临床相关的小鼠模型，即延长的热性惊厥（fc），随后是生命后期“第二次袭击”凯尼克酸诱发的癫痫发作。我们使用无偏流式细胞术分析炎性细胞的初步结果发现，在有早期癫痫发作经验的动物中，“第二次袭击”后，中枢神经系统浸润性和中枢神经系统驻留性免疫细胞均显着增加。利用我们的“两击”模型，我们提出验证先天和适应性免疫反应在介导早期癫痫发作的长期致痫效应中起因果作用的假设。具体目的是：(1)为了验证对FCs的免疫反应会导致随后癫痫发作易感增加的假设，我们将确定早期FCs的急性促炎细胞因子产生和长期致癫痫作用是否可以通过抗炎药物米诺扎克或促肾上腺皮质激素治疗后预防；(2)为了区分先天免疫和适应性免疫在早期热性惊厥致敏效应中的作用，我们将利用Cx3cr1EGFP/+转基因小鼠，双色MCP-1::mRFP1；CCR2::CCR2- egfp报告小鼠，以及多色流式细胞术分析来检测和定量先天和适应性免疫细胞。3. 为了直接解决中枢神经系统和外周炎症细胞在癫痫发生过程中的作用，我们将在遗传上不能建立先天（MyD88-/-）或适应性（Rag1-/-）免疫的小鼠中测试我们的癫痫模型。这些研究的结果将揭示适应性免疫和先天免疫对生命早期开始的癫痫发生过程的相对贡献。结果还应确定抗炎药物对早期抗癫痫治疗的潜在有益影响。

DESCRIPTION (provided by applicant): The long-term neurological sequelae of intractable epilepsy that begins in childhood are truly devastating. Conventional antiepileptic drugs are not only inadequate to control seizures, but can cause detrimental side effects to further burden the development of children with epilepsy. Alternative therapies that prevent the progression of epilepsy and limit cellular injury associated with seizures are critically needed. A dysregulated innate immune response, peripheral inflammatory cell infiltration and breakdown of the blood- brain barrier have been implicated in the initiation, progression and perpetuation of seizures. A recent genome-wide search to identify novel drug targets in our laboratory has revealed that proinflammatory molecules may be therapeutic targets in the epileptogenic process. We have developed a new clinically relevant murine model of prolonged febrile convulsions (FCs) followed by "second hit" kainic acid-induced seizures later in life. Our preliminary results using an unbiased flow cytometric analyses of inflammatory cells detected significant increases in both CNS-infiltrating and CNS- resident immune cells after a "second hit" in animals with prior experience of early-life seizures. Using our "two hit" model, we propose to test the hypothesis that innate and adaptive immune responses play a causal role in mediating the long-term epileptogenic effects of early life seizures. The Specific Aims are: (1) To test the hypothesis that immunologic responses to FCs cause heightened susceptibility to subsequent seizures, we will determine whether the acute pro-inflammatory cytokine production and long-term epileptogenic effect of early-life FCs is prevented by post-treatment with an anti- inflammatory drug minozac or ACTH; (2) To distinguish the involvement of innate and adaptive immunity in the sensitizing effects of early-life febrile convulsions, we will utilize Cx3cr1EGFP/+ transgenic mice, dual-color MCP-1::mRFP1;CCR2::CCR2-EGFP reporter mice, and multi-color flow cytometric analysis to detect and quantify innate and adaptive immune cells. 3. To directly address the role of CNS-resident and peripheral inflammatory cells in the epileptogenic process, we will test our epilepsy model in mice genetically incapable of mounting innate (MyD88-/-) or adaptive (Rag1-/-) immunity. Results of these studies will reveal the relative contributions of adaptive and innate immunity to the epileptogenic process that starts in early life. The results should also determine the potential beneficial impact of anti-inflammatory drugs for early anti- epileptic therapy.