Activated Caspase-6 (Casp6) is associated with age-dependent cognitive impairment and Alzheimer disease (AD). Mice expressing human Caspase-6 in hippocampal CA1 neurons develop age-dependent cognitive deficits, neurodegeneration and neuroinflammation. This study assessed if methylene blue (MB), a phenothiazine that inhibits caspases, alters Caspase-6-induced neurodegeneration and cognitive impairment in mice. Aged cognitively impaired Casp6-overexpressing mice were treated with methylene blue in drinking water for 1 month. Methylene blue treatment did not alter Caspase-6 levels, assessed by RT-PCR, western blot and immunohistochemistry, but inhibited fluorescently-labelled Caspase-6 activity in acute brain slice intact neurons. Methylene blue treatment rescued Caspase-6-induced episodic and spatial memory deficits measured by novel object recognition and Barnes maze, respectively. Methylene blue improved synaptic function of hippocampal CA1 neurons since theta-burst long-term potentiation (LTP), measured by field excitatory postsynaptic potentials (fEPSPs) in acute brain slices, was successfully induced in the Schaffer collateral-CA1 pathway in methylene blue-treated, but not in vehicle-treated, Caspase-6 mice. Increased neuroinflammation, measured by ionized calcium binding adaptor molecule 1 (Iba1)-positive microglia numbers and subtypes, and glial fibrillary acidic protein (GFAP)-positive astrocytes, were decreased by methylene blue treatment. Therefore, methylene blue reverses Caspase-6-induced cognitive deficits by inhibiting Caspase-6, and Caspase-6-mediated neurodegeneration and neuroinflammation. Our results indicate that Caspase-6-mediated damage is reversible months after the onset of cognitive deficits and suggest that methylene blue could benefit Alzheimer disease patients by reversing Caspase-6-mediated cognitive decline.

中文翻译：

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亚甲蓝抑制Caspase-6活性，并逆转Caspase-6诱导的老年小鼠认知障碍和神经炎症。

活化的Caspase-6（Casp6）与年龄依赖性认知障碍和阿尔茨海默氏病（AD）相关。在海马CA1神经元中表达人Caspase-6的小鼠发展出年龄依赖性的认知缺陷，神经退行性变和神经炎症。这项研究评估了抑制胱天蛋白酶的吩噻嗪亚甲蓝（MB）是否能改变Caspase-6诱导的小鼠神经变性和认知障碍。老年认知功能受损的Casp6过表达小鼠在饮用水中用亚甲基蓝处理1个月。亚甲基蓝处理不会改变RT-PCR，Western印迹和免疫组化评估的Caspase-6水平，但会抑制急性脑切片完整神经元中的荧光标记的Caspase-6活性。亚甲基蓝处理分别挽救了由Caspase-6诱导的情景和空间记忆缺陷，分别通过新颖的物体识别和Barnes迷宫测量。亚甲蓝改善了海马CA1神经元的突触功能，因为通过急性脑切片中的场兴奋性突触后突触电位（fEPSPs）测量的Theta-burst长期增强（LTP）成功地在亚甲基蓝处理的Schaffer侧支CA1途径中诱导，但在媒介物治疗的Caspase-6小鼠中却没有。亚甲基蓝处理可减少由离子化钙结合衔接子分子1（Iba1）阳性小胶质细胞数量和亚型以及神经胶质原纤维酸性蛋白（GFAP）阳性星形胶质细胞引起的神经炎症的增加。因此，亚甲蓝通过抑制Caspase-6来逆转Caspase-6诱导的认知功能障碍，和Caspase-6介导的神经变性和神经炎症。我们的结果表明，Caspase-6介导的损伤在认知缺陷发作后数月是可逆的，并且表明亚甲基蓝可以通过逆转Caspase-6介导的认知下降来使阿尔茨海默病患者受益。