Cofilin, a cytoskeletal actin severing protein, is essential for the initiation phase of apoptosis. The formation of cofilin rods (containing 1:1 cofilin:actin) has been studied in cultured mammalian neurons under conditions of excessive glutamate, ATP depletion (ATP-D) or oxidative stress. These conditions simulate the pathologies occurring during ischemic stroke. In this study, we investigated the potential involvement of cofilin during ischemic-stroke induced apoptosis. Transient middle cerebral artery occlusion (tMCAO) was performed to establish an experimental model of ischemic stroke. We used 2,3,5-Triphenyltetrazolium Chloride (TTC) and immunostaining of the neuronal marker neuronal nuclei (NeuN) to evaluate the evolving phases of infarction in rats subjected tMCAO. Immunostaining and TdT-mediated dUTP Nick-End Labeling (TUNEL) apoptosis staining were collaboratively used to examine cofilin rod formation and cell apoptosis in response to ischemia at different time points (2 h, 8 h, 24 h and 7 d). Our results showed that infarct volume increased initially, between the first 2 h to 24 h and became stabilized 24 h to 7 d after tMCAO. The formation of cofilin rods significantly increased in the cortical core (from 2 h) and penumbra (from 8 h), peaking at 24 h and gradually diminishing 7 d after tMCAO. Progressive accumulation of cofilin rods subsequently induced microtubule-associated protein-2 (MAP2) degradation and ischemic cell apoptosis in the infarct cortex after stroke. To further corroborate the role of activated cofilin in ischemic stroke, inhibition of cofilin by LIM kinase (Limk1) over-expression was performed. Lmik1 reduced cofilin rod formation and MAP2 degradation, and consequently, attenuated cofilin mediated-apoptosis 24 h after tMCAO. From this evidence we conclude that cofilin plays a role in the onset of ischemic-induced apoptosis and may be efficacious in future studies as a drug target for ischemic stroke.

Cofilin 是一种细胞骨架肌动蛋白切断蛋白，对于细胞凋亡的起始阶段至关重要。在谷氨酸过量、ATP 耗竭 (ATP-D) 或氧化应激条件下，已在培养的哺乳动物神经元中研究了 cofilin 棒（含有 1:1 cofilin:actin）的形成。这些条件模拟缺血性中风期间发生的病理。在这项研究中，我们研究了 cofilin 在缺血性中风诱导的细胞凋亡过程中的潜在参与。进行短暂性大脑中动脉闭塞（tMCAO）以建立缺血性脑卒中的实验模型。我们使用 2,3,5-三苯基四唑氯化物 (TTC) 和神经元标记神经元核 (NeuN) 的免疫染色来评估接受 tMCAO 的大鼠梗死的演变阶段。免疫染色和 TdT 介导的 dUTP 缺口末端标记 (TUNEL) 细胞凋亡染色共同用于检查不同时间点（2 小时、8 小时、24 小时和 7 天）缺血时 cofilin 棒的形成和细胞凋亡。我们的结果表明，梗死体积最初在最初的 2 小时至 24 小时之间增加，并在 tMCAO 后的 24 小时至 7 天稳定下来。cofilin 棒的形成在皮质核心（从 2 小时开始）和半影区（从 8 小时开始）显着增加，在 24 小时达到峰值，并在 tMCAO 后 7 天逐渐减少。cofilin 棒的逐渐积累随后在中风后诱导梗塞皮层中的微管相关蛋白 2 (MAP2) 降解和缺血性细胞凋亡。为了进一步证实活化的 cofilin 在缺血性中风中的作用，进行了 LIM 激酶 (Limk1) 过表达对 cofilin 的抑制。Lmik1 减少了 cofilin 棒的形成和 MAP2 降解，因此在 tMCAO 后 24 小时减弱了 cofilin 介导的细胞凋亡。根据这一证据，我们得出结论，cofilin 在缺血性诱导的细胞凋亡的发生中起作用，并且可能在未来的研究中作为缺血性中风的药物靶点有效。