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Abstract Human mesenchymal stem cells (hMSCs) have gained traction in transplantation therapy due to their immunomodulatory, paracrine, immune‐evasive, and multipotent differentiation potential. The inherent heterogeneity of hMSCs poses a challenge for therapeutic treatments and necessitates the identification of robust biomarkers to ensure reproducibility in both in vivo and in vitro experiments. In this study, we utilized dielectrophoresis (DEP), a label‐free electrokinetic phenomenon, to investigate the heterogeneity of hMSCs derived from bone marrow (BM) and adipose tissue (AD). The electrical properties of BM‐hMSCs were compared to homogeneous mouse fibroblasts (NIH‐3T3), human fibroblasts (WS1), and human embryonic kidney cells (HEK‐293). The DEP profile of BM‐hMSCs differed most from HEK‐293 cells. We compared the DEP profiles of BM‐hMSCs and AD‐hMSCs and found that they have similar membrane capacitances, differing cytoplasm conductivity, and transient slopes. Inducing both populations to differentiate into adipocyte and osteoblast cells revealed that they behave differently in response to differentiation‐inducing cytokines. Histology and reverse transcription‐quantitative polymerase chain reaction (RT‐qPCR) analyses of the differentiation‐related genes revealed differences in heterogeneity between BM‐hMSCs and AD‐hMSCs. The differentiation profiles correlate well with the DEP profiles developed and indicate differences in the heterogeneity of BM‐hMSCs and AD‐hMSCs. Our results demonstrate that using DEP, membrane capacitance, cytoplasm conductivity, and transient slope can uniquely characterize the inherent heterogeneity of hMSCs to guide robust and reproducible stem cell transplantation therapies.
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This content will become publicly available on December 1, 2025
Transient Slope: A Metric for Assessing Heterogeneity from the Dielectrophoresis Spectrum
Cellular heterogeneity, an inherent feature of biological systems, plays a critical role in processes such as development, immune response, and disease progression. Human mesenchymal stem cells (hMSCs) exemplify this heterogeneity due to their multi-lineage differentiation potential. However, their inherent variability complicates clinical use, and there is no universally accepted method for detecting and quantifying cell population heterogeneity. Dielectrophoresis (DEP) has emerged as a powerful electrokinetic technique for characterizing and manipulating cells based on their dielectric properties, offering label-free analysis capabilities. Quantitative information from the DEP spectrum, such as transient slope, measure cells’ transition between negative and positive DEP behaviors. In this study, we employed DEP to estimate transient slope of various cell populations, including relatively homogeneous HEK-293 cells, heterogeneous hMSCs, and cancer cells (PC3 and DU145). Our analysis encompassed hMSCs derived from bone marrow, adipose, and umbilical cord tissue, to capture tissue-specific heterogeneity. Transient slope was assessed using two methods, involving linear trendline fitting to different low-frequency regions of the DEP spectrum. We found that transient slope serves as a reliable indicator of cell population heterogeneity, with more heterogeneous populations exhibiting lower transient slopes and higher standard deviations. Validation using cell morphology, size, and stemness further supported the utility of transient slope as a heterogeneity metric. This label-free approach holds promise for advancing cell sorting, biomanufacturing, and personalized medicine.
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- Award ID(s):
- 2048221
- PAR ID:
- 10577604
- Publisher / Repository:
- MDPI
- Date Published:
- Journal Name:
- Biophysica
- Volume:
- 4
- Issue:
- 4
- ISSN:
- 2673-4125
- Page Range / eLocation ID:
- 695 to 710
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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