iBiMED - Institute of Biomedicine, Department of Medical Sciences, University of Aveiro, Aveiro, Portugal; CICECO - Aveiro Institute of Materials, Department of Chemistry, University of Aveiro, Aveiro, Portugal.
iBiMED - Institute of Biomedicine, Department of Medical Sciences, University of Aveiro, Aveiro, Portugal.
Clin Chim Acta. 2020 Mar;502:25-33. doi: 10.1016/j.cca.2019.11.025. Epub 2019 Nov 29.
The loss of proteostasis during aging has been well described using different models, however little is known with respect to protein aggregation levels in biofluids with aging. Therefore, the aim of this study was to assess the pattern of age-related protein aggregation in human plasma using two distinct approaches: analysis with conformation-specific antibodies and FTIR spectroscopy. The latter has been widely used in biomedical research to study protein conformational changes in health and disease. Samples from a primary care based-cohort from the Aveiro region, Portugal, were used for slot-blot analyses followed by immunodetection with conformation-specific antibodies and for the acquisition of FTIR spectra. Immunoblot analyses revealed an age-dependent evolution of the protein conformational profile in human plasma, towards a decrease in prefibrillar oligomers and an increase in fibrillar structures. This finding was also supported by PLS-R multivariate analysis of FTIR data, where a positive correlation between the age of the donors and secondary structure of plasma proteins could be observed. Samples from younger donors are characterized by antiparallel β-sheet-containing structures while intermolecular β-sheets characterized older samples. Exclusion of age-associated co-morbidities improved the correlation between protein conformational profiles and aging. The results reveal structural changes in human plasma proteins from middle to old age, confirming the age-associated changes in protein aggregation, and support the applicability of FTIR as a reliable approach to study proteostasis during aging.
在衰老过程中，蛋白质稳态的丧失已经在不同的模型中得到了很好的描述，然而，关于生物体液中蛋白质聚集水平随年龄的变化知之甚少。因此，本研究的目的是使用两种不同的方法评估人类血浆中与年龄相关的蛋白质聚集模式：使用构象特异性抗体进行分析和傅里叶变换红外（FTIR）光谱分析。后者已广泛用于生物医学研究，以研究健康和疾病中蛋白质构象变化。该研究使用来自葡萄牙阿威罗地区初级保健队列的样本进行斑点印迹分析，然后用构象特异性抗体进行免疫检测，并获取 FTIR 光谱。免疫印迹分析显示，人类血浆中的蛋白质构象谱随年龄的增长而发生变化，向原纤维寡聚物减少和纤维结构增加的方向发展。FTIR 数据分析的偏最小二乘回归（PLS-R）多元分析也支持这一发现，其中可以观察到供体年龄与血浆蛋白质二级结构之间存在正相关。年轻供体的样本特征是含有反平行β-折叠结构的物质，而分子间β-折叠结构则是老年样本的特征。排除与年龄相关的合并症可以提高蛋白质构象谱与衰老之间的相关性。研究结果揭示了从中年到老年人类血浆蛋白质结构的变化，证实了蛋白质聚集的年龄相关性变化，并支持 FTIR 作为研究衰老过程中蛋白质稳态的可靠方法的适用性。
