IL1 and hypoxic-ischemic insults

描述(申请人提供)：由脑缺血(即中风)引起的脑损伤是一个主要的公共卫生问题。中风造成的脑损伤中，多达50%发生在主要损伤灶之外，组织破坏过程持续数小时至数天。现在很明显，炎症因素导致了这种延迟的病理生理过程。具体地说，研究表明，细胞因子白介素1β(IL-1β)在实验性和临床中风后上调，而其他研究表明它与损伤的进展有关。然而，IL-1β促进神经细胞死亡的细胞和分子途径(S)尚未确定。这在很大程度上是由于缺乏合适的体外模型来评估这些机制。因此，我们利用神经元/星形胶质细胞混合培养建立了一套可靠、重复性好的体外模型系统。在这个模型中，内源性IL-1β的产生通过外源性IL-1β的添加和细胞缺氧诱导的神经元损伤来模拟。我们发现，用这种细胞因子治疗前--但不是同时或治疗后--极大地加强了通过剥夺混合小鼠皮质细胞培养的氧气而导致的神经细胞死亡。IL-1β的作用呈浓度依赖性，并可被重组IL-1受体拮抗剂完全抑制，表明IL-1受体I型(IL1R1)参与了信号转导。此外，我们发现这种IL-1β介导的缺氧性神经元损伤可以被代谢性谷氨酸受体1(但不是mGluR5)的药理拮抗完全阻止。这与不受mGluR1受体拮抗影响的纯缺氧性神经元损伤形成鲜明对比。最后，我们发现IL-1β诱导的损伤的增强依赖于星形胶质细胞IL1R1的表达，而神经元中信号的丢失则没有影响。因此，本五年研究计划的目标是：1)确定IL-1β信号与mGluR1信号协同作用增强缺氧性神经元损伤的分子机制(S)；2)确定介导IL-1β增强效应的一个或多个星形细胞因子；3)评估去除IL-1β信号是否能有效预防/改善直接海马注射N-甲基-D-天冬氨酸和/或大脑中动脉阻塞所致的在体脑损伤。改进对这些事件的定义可能会导致开发新的治疗策略，旨在减缓中风后神经元破坏的进展。

DESCRIPTION (provided by applicant): Injury to the brain caused by cerebral ischemia (i.e. stroke) is a major public health concern. As much as 50% of the brain damage incurred by stroke occurs outside of the primary focus of damage with the process of tissue destruction continuing for hours to days. It is now apparent that inflammatory factors contribute to this delayed pathophysiology. Specifically, studies demonstrate that the cytokine, interleukin 1beta (IL-1beta), is upregulated following experimental and clinical stroke while additional studies implicate it in the progression of injury. However, the cellular and molecular pathway(s) by which IL-1beta contributes to neuronal cell death have yet to be identified. This is largely due to the lack of suitable in vitro models in which to assess these mechanisms. Therefore, we developed a reliable and reproducible in vitro model system utilizing mixed neuronal/astrocyte cortical cell cultures. In this model, endogenous production of IL-1beta is simulated by exogenous addition of IL-1beta and neuronal injury induced by depriving cells of oxygen. We found that pre-treatment - but not concurrent or post-treatment - with this cytokine dramatically potentiated neuronal cell death induced by depriving mixed murine cortical cell cultures of oxygen. The effect of IL-1beta was concentration-dependent and could be completely inhibited by the recombinant IL-1 receptor antagonist, indicating that signaling through the IL-1 receptor type I (IL1R1) was involved. Further, we found this IL-1beta -mediated enhancement of hypoxic-neuronal injury can be completely prevented by pharmacological antagonism of metabotropic glutamate receptor 1 (but not mGluR5). This is in stark contrast to a pure hypoxic neuronal injury which is unaffected by mGluR1 receptor antagonism. Finally, we found that the enhancement of injury induced by IL-1beta was dependent on astrocytic expression of IL1R1 whereas loss of signaling in neurons had no effect. Thus, the objectives of this five year research plan are to 1) determine the molecular mechanism(s) by which IL1beta signaling functionally synergizes with mGluR1 signaling to enhance hypoxic neuronal injury; 2) to determine the astrocytic factor or factors responsible for mediating the IL-1beta enhancing effect; and 3) to assess whether removal of IL-1beta signaling can effectively prevent/ameliorate hippocampal injury in vivo induced by direct hippocampal injection of NMDA and/or middle cerebral artery occlusion. Improved definition of these events could lead to the development of new therapeutic strategies designed to attenuate the progression of neuronal destruction following stroke.