Single-cell mass cytometry reveals cell type- and cluster-specific heterogeneity in silver nanoparticle responses in a 3D alveolar tetra-culture model

Abstract

Silver nanoparticles (AgNPs) are widely used in medicine, environmental science, and industry. However, their heterogeneous interactions with complex biological systems, especially at the single-cell level, are not fully understood yet. Conventional toxicity assessment methods are typically conducted on oversimplified in vitro models that fail to replicate actual physiological conditions, and measure collective responses across cell populations, obscuring differences among individual cells. To overcome these limitations, we utilized single-cell mass cytometry (CyTOF) to investigate individual cell responses to AgNP-induced stress, combined with a 3D alveolar tetra-culture model designed to better reflect the complexity of biological systems. Single-cell mass cytometry of a 3D alveolar model revealed heterogeneous, cell type–specific responses to AgNP exposure. Specifically, PMA-differentiated THP-1, A549 and EA.hy926 cells exhibited high AgNP association but limited cytotoxicity, indicating activation of stress-mitigation pathways, while THP-1 cells showed early inflammatory activation despite minimal AgNP association, suggesting an indirect mechanism. Single-cell analysis and FlowSOM clustering revealed distinct subpopulations exhibiting diverse intracellular signaling profiles of inflammatory cytokines, anti-inflammatory mediators, and stress-response proteins, which unveiled common cellular responses and unique cell-type specific pathways determining cell fate (survival, transitional states, or apoptosis) upon AgNP exposure. This study introduces a novel framework for studying heterogeneous interactions of nanoparticles with complex biological systems by integrating a 3D alveolar tetra-culture model with single-cell mass cytometry analysis, enabling the dissection of nanoparticle-induced stress responses at an unprecedented level of detail. These insights have broad implications for nanotoxicology and nanomedicine, underscoring the need to account for cellular heterogeneity when evaluating nanoparticle-induced toxicity.

Graphical abstract: Single-cell mass cytometry reveals cell type- and cluster-specific heterogeneity in silver nanoparticle responses in a 3D alveolar tetra-culture model

Supplementary files

Article information

Article type
Paper
Submitted
02 5月 2025
Accepted
08 7月 2025
First published
15 7月 2025
This article is Open Access
Creative Commons BY-NC license

Environ. Sci.: Nano, 2025, Advance Article

Single-cell mass cytometry reveals cell type- and cluster-specific heterogeneity in silver nanoparticle responses in a 3D alveolar tetra-culture model

E. Lee, S. Park, S. M. Ha, M. Kim, S. Park, A. Chary, T. Serchi and T. H. Yoon, Environ. Sci.: Nano, 2025, Advance Article , DOI: 10.1039/D5EN00439J

This article is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported Licence. You can use material from this article in other publications, without requesting further permission from the RSC, provided that the correct acknowledgement is given and it is not used for commercial purposes.

To request permission to reproduce material from this article in a commercial publication, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party commercial publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Social activity

Spotlight

Advertisements