Quantification of decellularized human myocardial matrix: A comparison of six patients.
Quantification of decellularized human myocardial matrix: A comparison of six patients.
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DOI：
10.1002/prca.201500048
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发表时间：
2016-01
期刊：
影响因子：
--
通讯作者：
Hansen KC
中科院分区：
文献类型：
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作者：
Johnson TD;Hill RC;Dzieciatkowska M;Nigam V;Behfar A;Christman KL;Hansen KC
本研究的目的是对从6个不同的身体供体心脏分离的人心脏细胞外基质(ECM)进行表征和定量分析。以6例人身体供体心脏为材料，采用去细胞方法分离细胞外基质，并用硫酸糖胺多聚糖含量(SGAG)和聚丙烯酰胺凝胶电泳法(PAGE)进行鉴定。然后使用ECM靶向定量conCATamers(QconCAT)通过液色谱选择反应监测(LC-SRM)分析来定量蛋白质含量，该分析使用83个稳定同位素标记(SIL)肽代表48种不同的蛋白质。采用LC-MS/MS(LC-MS/MS)进行非靶向全局分析。SGAG含量、PAGE和QconCAT蛋白质组学分析显示，六个患者样本之间存在显著差异。定量蛋白质组学分析表明，蛋白质含量主要由多种纤维状胶原组成。此外，难以去除的细胞蛋白质的量化只占总蛋白质含量的不到1%，对于脱细胞生物材料来说，这是非常低的。全球蛋白质组学鉴定出200多种不同的蛋白质存在于人类心脏细胞外基质中。总而言之，人类心肌细胞外基质的量化和表征在六个被调查的患者之间显示了显著的患者之间的差异。这对于开发同种异体衍生生物材料和增加我们对人类心肌细胞外基质成分的了解是一个重要的成果。
The purpose of this study was to characterize and quantitatively analyze human cardiac extracellular matrix (ECM) isolated from six different cadaveric donor hearts. ECM was isolated by decellularization of six human cadaveric donor hearts and characterized by quantifying sulfated glycosaminoglycan content (sGAG) and via polyacrylamide gel electrophoresis (PAGE). The protein content was then quantified using ECM-targeted Quantitative conCATamers (QconCAT) by Liquid Chromatography - Selected Reaction Monitoring (LC-SRM) analysis using 83 stable isotope labeled (SIL) peptides representing 48 different proteins. Non-targeted global analysis was also implemented using liquid chromatography tandem mass spectrometry (LC-MS/MS). The sGAG content, PAGE, and QconCAT proteomics analysis showed significant variation between each of the six patient samples. The quantitative proteomics indicated that the majority of the protein content was composed of various fibrillar collagen components. Also, quantification of difficult to remove cellular proteins represented less than 1% of total protein content, which is very low for a decellularized biomaterial. Global proteomics identified over 200 distinct proteins present in the human cardiac ECM. In conclusion, quantification and characterization of human myocardial ECM showed significant patient-to-patient variability between the six investigated patients. This is an important outcome for the development of allogeneic derived biomaterials and for increasing our understanding of human myocardial ECM composition.