1] Institute of Biostructures and Bioimaging (IBB)-CNR, CIRPEB, Via Mezzocannone, 16, Naples 80134, Italy [2] Institute of Pharmacology and Clinical Pharmacy, Philipps University of Marburg, Marburg 35032, Germany [3] Department of Neurodegeneration, Royal College of Surgeons in Ireland, Dublin 2, Ireland.
Institute of Pharmacology and Clinical Pharmacy, Philipps University of Marburg, Marburg 35032, Germany.
Cell Death Dis. 2014 Jan 16;5(1):e993. doi: 10.1038/cddis.2013.518.
Delayed neuronal cell death largely contributes to the progressive infarct development and associated functional impairments after cerebral ischemia or brain trauma. Previous studies exposed a key role for the interaction of the mitochondrial protein apoptosis-inducing factor (AIF) and cytosolic cyclophilin A (CypA) in pathways of programmed cell death in neurons in vitro and in vivo. These studies suggested that pro-apoptotic activities of AIF, such as its translocation to the nucleus and subsequent DNA degradation, depend on the physical interaction of AIF with CypA. Hence, this protein complex may represent a new pharmacological target for inhibiting the lethal action of AIF on the brain tissue. In this study, we show that the AIF amino-acid residues 370-394 mediate the protein complex formation of AIF with CypA. The synthetic AIF(370-394) peptide inhibited AIF/CypA complex formation in vitro by binding CypA with a K(D) of 12 μM. Further, the peptide exerted pronounced neuroprotective effects in a model of glutamate-induced oxidative stress in cultured HT-22 cells. In this model system of AIF-dependent cell death, the AIF(370-394) peptide preserved mitochondrial integrity, as detected by measurements of the mitochondrial membrane potential and quantification of mitochondrial fragmentation. Further, the AIF(370-394) peptide inhibited perinuclear accumulation of fragmented mitochondria, mitochondrial release of AIF to the nucleus and glutamate-induced cell death to a similar extent as CypA-siRNA. These data indicate that the targeting of the AIF-CypA axis is an effective strategy of neuroprotection.
迟发性神经元细胞死亡是导致脑缺血或脑创伤后进行性梗死发展和相关功能障碍的主要原因。先前的研究表明，线粒体蛋白凋亡诱导因子（AIF）与细胞质亲环蛋白 A（CypA）之间的相互作用在体外和体内神经元程序性细胞死亡途径中起着关键作用。这些研究表明，AIF 的促凋亡活性，如向核内易位和随后的 DNA 降解，依赖于 AIF 与 CypA 的物理相互作用。因此，该蛋白复合物可能代表抑制 AIF 对脑组织致死作用的新的药理学靶点。在这项研究中，我们表明 AIF 的氨基酸残基 370-394 介导了 AIF 与 CypA 的蛋白复合物形成。合成的 AIF（370-394）肽通过与 CypA 结合以 K(D)为 12 μM 抑制了 AIF/CypA 复合物的形成。此外，该肽在谷氨酸诱导的培养 HT-22 细胞氧化应激模型中表现出明显的神经保护作用。在这个依赖于 AIF 的细胞死亡模型系统中，AIF（370-394）肽通过测量线粒体膜电位和量化线粒体片段化来维持线粒体的完整性。此外，AIF（370-394）肽抑制了核周聚集的碎片化线粒体、线粒体向核内释放 AIF 以及谷氨酸诱导的细胞死亡，其效果与 CypA-siRNA 相似。这些数据表明，靶向 AIF-CypA 轴是一种有效的神经保护策略。
