Effective Top-Down LC/MS+ Method for Assessing Actin Isoforms as a Potential Cardiac Disease Marker

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• 作者：Yi-Chen Chen ; Serife Ayaz-Guner ; Ying Peng ; Nicole M. Lane ; Matthew R. Locher ; Takushi Kohmoto ; Lars Larsson ; Richard L. Moss ; Ying Ge
• 刊名：Analytical Chemistry
• 出版年：2015
• 出版时间：August 18, 2015
• 年：2015
• 卷：87
• 期：16
• 页码：8399-8406
• 全文大小：501K
• ISSN：1520-6882

文摘

Actin is the major component of the cytoskeleton, playing an essential role in the structure and motility of both muscle and nonmuscle cells. It is highly conserved and encoded by a multigene family. 伪-Cardiac actin (伪CAA) and 伪-skeletal actin (伪SKA), encoded by two different genes, are the primary actin isoforms expressed in striated muscles. The relative expression levels of 伪SKA and 伪CAA have been shown to vary between species and under pathological conditions. In particular, an increased 伪SKA expression is believed to be a programmed response of a diseased heart. Therefore, it is essential to quantify the relative expression of 伪SKA and 伪CAA, which remains challenging due to the high degree of sequence similarity between these isoforms (98.9%). Herein, we developed a top-down liquid chromatography/mass spectrometry-based (鈥淟C/MS+鈥? method for the rapid purification and comprehensive analysis of 伪-actin extracted from muscle tissues. We thoroughly investigated all of the actin isoforms in healthy human cardiac and skeletal muscles. We found that 伪SKA is the only isoform expressed in skeletal muscle, whereas 伪CAA and 伪SKA are coexpressed in cardiac muscle. We then applied our method to quantify the 伪-actin isoforms in human healthy hearts and failing hearts with dilated cardiomyopathy (DCM). We found that 伪SKA is augmented in DCM compared with healthy controls, 43.1 卤 0.9% versus 23.7 卤 1.7%, respectively. As demonstrated, top-down LC/MS+ provides an effective and comprehensive method for the purification, quantification, and characterization of 伪-actin isoforms, enabling assessment of their clinical potential as cardiac disease markers.