利用反射光进行衰老检测

IF 7.8 1区 医学 Q1 Biochemistry, Genetics and Molecular Biology
Aging Cell Pub Date : 2024-08-05 DOI:10.1111/acel.14295
Benjamin Dedic, Leo Westerberg, Andrea Mosqueda Solís, Kyle D. Dumont, Jorge L. Ruas, Anders Thorell, Erik Näslund, Kirsty L. Spalding
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引用次数: 0

摘要

衰老是发生在发育、组织修复、癌症和衰老过程中的重要细胞程序。衰老的增加也与疾病状态有关,包括肥胖和 2 型糖尿病(T2D)。在单细胞水平表征和量化衰老细胞一直是一项挑战,尤其是在大型原代细胞(如人类脂肪细胞)中更是如此。在这项研究中,我们提出了一种利用反射光精确测量衰老相关的β-半乳糖苷酶(SABG)染色的新方法,该方法可与免疫荧光相结合,并与人类和小鼠的原代成熟脂肪细胞以及分化的 3T3-L1 细胞兼容。这项技术结合了多种标准,包括稳健的样本特异性 pH 值控制,对细胞的衰老状态进行了更全面的分类。我们利用共聚焦显微镜的精确性,通过反射光检测 X-gal 晶体,实现了比传统明视野技术更高的灵敏度。这种策略可以捕捉到 SABG 染色样本中的所有 X-gal 沉淀,揭示出多种 X-gal 染色模式,提高了检测灵敏度。此外,我们还证明了反射光在检测和量化成熟人类脂肪细胞的衰老方面优于 Western 印迹分析,因为它能更准确地反映 SABG 的活性。这种检测策略能更深入地研究衰老细胞的特征,特别是深入研究脂肪细胞衰老与肥胖相关疾病(如 T2D)之间的关系。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Senescence detection using reflected light

Senescence detection using reflected light

Senescence detection using reflected light

Senescence is an important cellular program occurring in development, tissue repair, cancer, and aging. Increased senescence is also associated with disease states, including obesity and Type 2 diabetes (T2D). Characterizing and quantifying senescent cells at a single cell level has been challenging and particularly difficult in large primary cells, such as human adipocytes. In this study, we present a novel approach that utilizes reflected light for accurate senescence-associated beta-galactosidase (SABG) staining measurements, which can be integrated with immunofluorescence and is compatible with primary mature adipocytes from both human and mouse, as well as with differentiated 3T3-L1 cells. This technique provides a more comprehensive classification of a cell's senescent state by incorporating multiple criteria, including robust sample-specific pH controls. By leveraging the precision of confocal microscopy to detect X-gal crystals using reflected light, we achieved superior sensitivity over traditional brightfield techniques. This strategy allows for the capture of all X-gal precipitates in SABG-stained samples, revealing diverse X-gal staining patterns and improved detection sensitivity. Additionally, we demonstrate that reflected light outperforms western blot analysis for the detection and quantification of senescence in mature human adipocytes, as it offers a more accurate representation of SABG activity. This detection strategy enables a more thorough investigation of senescent cell characteristics and specifically a deeper look at the relationship between adipocyte senescence and obesity associated disorders, such as T2D.

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来源期刊
Aging Cell
Aging Cell 生物-老年医学
CiteScore
14.40
自引率
2.60%
发文量
212
审稿时长
8 weeks
期刊介绍: Aging Cell, an Open Access journal, delves into fundamental aspects of aging biology. It comprehensively explores geroscience, emphasizing research on the mechanisms underlying the aging process and the connections between aging and age-related diseases.
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