Graduate School of Biomedical Engineering, University of New South Wales, Sydney, NSW, 2052, Australia.
Mesenchymal Stem Cell Laboratory, School of Biomedicine, Faculty of Health and Medical Sciences, University of Adelaide, Adelaide, South Australia, 5000, Australia.
Stem Cell Rev Rep. 2024 Nov;20(8):2283-2292. doi: 10.1007/s12015-024-10778-4. Epub 2024 Aug 27.
To use autofluorescence multispectral imaging (AFMI) to develop a non-invasive assay for the in-depth characterisation of human bone marrow derived mesenchymal stromal cells (hBM-MSCs).
hBM-MSCs were imaged by AFMI on gridded dishes, stained for endpoints of interest (STRO-1 positivity, alkaline phosphatase, beta galactosidase, DNA content) then relocated and results correlated. Intensity, texture and morphological features were used to characterise the colour distribution of regions of interest, and canonical discriminant analysis was used to separate groups. Additionally, hBM-MSC lines were cultured to arrest, with AFMI images taken after each passage to investigate whether an assay could be developed for growth potential.
STRO-1 positivity could be predicted with a receiver operator characteristic area under the curve (AUC) of 0.67. For spontaneous differentiation this was 0.66, for entry to the cell-cycle it was 0.77 and for senescence it was 0.77. Growth potential (population doublings remaining) was estimated with an RMSPE = 2.296. The Mean Absolute Error of the final prediction model indicated that growth potential could be predicted with an error of ± 1.86 doublings remaining.
This non-invasive methodology enabled the in-depth characterisation of hBM-MSCs from a single assay. This approach is advantageous for clinical applications as well as research and stands out for the characterisation of both present status as well as future behaviour. The use of data from five MSC lines with heterogenous AFMI profiles supports potential generalisability.
利用自发荧光多光谱成像（AFMI）技术，开发一种非侵入性的方法，深入分析人骨髓间充质基质细胞（hBM-MSCs）。
在网格培养皿上对 hBM-MSCs 进行 AFMI 成像，对感兴趣的终点（STRO-1 阳性、碱性磷酸酶、β半乳糖苷酶、DNA 含量）进行染色，然后重新定位并进行结果相关性分析。利用强度、纹理和形态特征来描述感兴趣区域的颜色分布，并采用典型判别分析来分离不同的组别。此外，还对 hBM-MSC 进行培养，使其进入阻滞状态，在每个传代后拍摄 AFMI 图像，以研究是否可以开发一种用于生长潜力的检测方法。
STRO-1 阳性可以通过接受者操作特征曲线（ROC）下面积（AUC）为 0.67 进行预测。对于自发性分化，AUC 为 0.66；对于进入细胞周期，AUC 为 0.77；对于衰老，AUC 为 0.77。生长潜力（剩余倍增数）估计的均方根均方根误差（RMSPE）为 2.296。最终预测模型的平均绝对误差表明，生长潜力可以通过 ± 1.86 个倍增数的误差进行预测。
这种非侵入性的方法可以从单次检测中深入分析 hBM-MSCs。这种方法对于临床应用以及研究都具有优势，突出了对当前状态和未来行为的特征分析。使用 5 条具有不同 AFMI 特征的 MSC 系的数据支持了其潜在的通用性。
