Rapid Cytotoxicity Screening Platform for Amyloid Inhibitors Using a Membrane-Potential Sensitive Fluorescent Probe

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• 作者：Jihoon Kim ; Yasuhiko Sasaki ; Wataru Yoshida ; Natsuki Kobayashi ; Anthony J. Veloso ; Kagan Kerman ; Kazunori Ikebukuro ; Koji Sode
• 刊名：Analytical Chemistry
• 出版年：2013
• 出版时间：January 2, 2013
• 年：2013
• 卷：85
• 期：1
• 页码：185-192
• 全文大小：435K
• 年卷期：v.85,no.1(January 2, 2013)
• ISSN：1520-6882

文摘

The growing interest in membrane interactions of amyloidogenic proteins indicates that lipid binding and the regulation of membrane potential are critical to the onset and progression of neurodegenerative diseases such as Parkinson鈥檚 (PD), Alzheimer鈥檚 (AD), and prion diseases. Advancing the understanding of this field requires the application of varied biophysical and biological techniques designed to probe the characteristics and underlying mechanisms of membrane-peptide interactions. Therefore, the development of a rapid cytotoxicity evaluation system using a membrane potential-sensitive bis-oxonol fluorescent dye, DiBAC4(3) is reported here. The exposure of C-terminal truncated 伪-synuclein 119 (伪-Syn119) and amyloid-尾_1鈥?2 (A尾_1鈥?2) to U2-OS cell cultures resulted in an immediate, significant, and concentration-dependent increase in fluorescence response of DiBAC4(3). This response was strongly correlated with the cytotoxicity of 伪-Syn119 and A尾_1鈥?2 as determined by conventional CC8 and ATP assays. Furthermore, the capacity of well-defined polyphenolic antioxidants (i.e., pyrroloquinoline quinone (PQQ), baicalein, (鈭?-epigallocatechin-3-gallate (EGCG), and myricetin) to mitigate amyloid-induced cytotoxicity was evaluated using the developed biosensing system. We envisage that this work would accelerate the development of a rapid and cost-effective high-throughput screening platform in drug discovery for AD and PD.