Daudt Donald Raymond, Yorio Thomas
Department of Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, TX 76107, USA.
Mol Vis. 2011;17:1172-81. Epub 2011 May 4.
Neurodegenerative diseases and neurotraumas typically result in apoptosis of specific neurons leading to the pathology observed during the disease state. Existing treatments target the symptoms instead of preventing the death of these neurons. Although neuroprotective drugs should be useful as a treatment to prevent further loss of neurons, efficacious molecules are lacking. FK506 (tacrolimus), a widely used immunosuppressant drug, has significant neuroprotective and neuroregenerative properties throughout the central nervous system, including the eye. FK506 achieves these properties through interaction with FK506 binding proteins (FKBP), including FK506 binding protein 51 (FKBP51). In this study, we examine the effects of FKBP51 as a neuroprotective agent on a neuronal cell line.
We cultured 661w cell cultures with or without FK506, or stably transfected them with an FKBP51 expression vector. These cells were then exposed to the apoptosis-inducing agent staurosporine. Cell viability was determined using a calcein AM/propidium iodide assay. Protein levels and activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) were determined by western immunoblot analysis.
FKBP51 overexpression significantly protected 661w cell cultures from staurosporine-induced apoptosis. FKBP51 overexpression also significantly increased NF-κB p65 protein levels and activated NF-κB p65. FK506 treatment significantly protected 661w neuronal cultures from staurosporine-induced apoptosis. FK506 increased FKBP51, NF-κB p65, and levels of activated NF-κB p65 protein.
These results suggest that FKBP51 protects 661w cell cultures from apoptosis induced by staurosporine. Additionally, FK506 protected 661w cell cultures from apoptosis and displayed a mechanism similar to that of FKBP51 overexpression. Both FK506 and FKBP51 appear to act through activation of NF-κB p65 protein, suggesting a common pathway for neuroprotection. These findings implicate FKBP51 as a protein important to neuronal cell culture survival. FKBP51 may be a potential therapeutic drug target for preventing the neurodegeneration and neurotrauma that occur during neurodegenerative diseases.
神经退行性疾病和神经创伤通常会导致特定神经元凋亡，进而引发疾病状态下所观察到的病理变化。现有的治疗方法针对的是症状，而非预防这些神经元的死亡。尽管神经保护药物作为预防神经元进一步丧失的治疗手段应该是有用的，但目前缺乏有效的分子。FK506（他克莫司）是一种广泛使用的免疫抑制药物，在包括眼睛在内的整个中枢神经系统中具有显著的神经保护和神经再生特性。FK506通过与FK506结合蛋白（FKBP）相互作用来实现这些特性，其中包括FK506结合蛋白51（FKBP51）。在本研究中，我们检测了FKBP51作为一种神经保护剂对一种神经元细胞系的影响。
我们用或不用FK506培养661w细胞培养物，或者用FKBP51表达载体对其进行稳定转染。然后将这些细胞暴露于凋亡诱导剂星形孢菌素。使用钙黄绿素AM/碘化丙啶检测法测定细胞活力。通过蛋白质免疫印迹分析测定活化B细胞核因子κB（NF-κB）的蛋白质水平和活化情况。
FKBP51过表达显著保护661w细胞培养物免受星形孢菌素诱导的凋亡。FKBP51过表达还显著提高了NF-κB p65蛋白水平并激活了NF-κB p65。FK506处理显著保护661w神经元培养物免受星形孢菌素诱导的凋亡。FK506增加了FKBP51、NF-κB p65以及活化的NF-κB p65蛋白水平。
这些结果表明FKBP51保护661w细胞培养物免受星形孢菌素诱导的凋亡。此外，FK506保护661w细胞培养物免受凋亡，并且显示出与FKBP51过表达类似的机制。FK506和FKBP51似乎都通过激活NF-κB p65蛋白发挥作用，提示存在一条共同的神经保护途径。这些发现表明FKBP51是对神经元细胞培养存活至关重要的一种蛋白质。FKBP51可能是预防神经退行性疾病期间发生的神经退行性变和神经创伤的潜在治疗药物靶点。
