Assay Ready Cells and Extracellular Vesicles

The human monocytic cell line THP-1 (ATCC TIB-202), which was originally derived from peripheral blood of an acute monocytic leukemia patient, is a versatile and essential tool for the biomedical research community.¹ In addition to its ability to model monocyte function and maturation, THP-1 cells also have widespread applications in immuno-oncology and cancer research, drug development and toxicology screening, and studies relating to infectious diseases. For these reasons, THP-1 cells are an attractive candidate for in vitro bioassays. We previously highlighted ATCC’s ThawReady™ portfolio, which includes an assay-ready, luciferase reporter cell line derived from THP-1 parental cells, as a novel solution for standardizing and streamlining bioassay workflow.² ThawReady™ cells offer high viability, high cell number, and functionality in a reproducible manner. Importantly, these cells are prepared in a novel cryopreserved format that facilitates their immediate use post-thaw. This feature, coupled with their ability to express firefly luciferase gene (luc2) under control of a NFκB promoter, further enables their utility across a variety of in vitro assay platforms.

In recent years, extracellular vesicles (EVs), particularly those derived from mesenchymal stem cells (MSCs), have been recognized for playing a critical role in immunomodulation and tissue regeneration.^3-5 Due to their nano-scale size, EVs can effectively interact with target cells to transfer their bioactive molecular cargo, which includes various nucleic acids, cytokines, proteins, and lipids. Therefore, MSC EVs are emerging as a cell-free alternative to traditional cell-based therapies. We have previously reported multi-functional effects (e.g., pro-migratory, anti-inflammatory, anti-apoptotic, pro-angiogenic) of MSC EVs across different cell types, including primary cells, CNS-relevant cells, and retinal epithelial cells.^6-8 More recently, we have leveraged ThawReady™ THP-1 NF-κB-Luc2 cells (ATCC TIB-202-NFkB-LUC2-AR) to directly assess the effects of MSC EVs (ATCC SCRC-4000-EXM) on monocyte activation.

In these preliminary studies, we demonstrated that lipopolysaccharide (LPS) from E. coli can elicit a potent pro-inflammatory response correlating to NFκB activation, as measured by luciferase expression, in ThawReady™ cells. These studies have also allowed us to rapidly and reproducibly screen the reparative properties of MSC EVs in LPS-stimulated cells. Our data confirm that MSC EVs can significantly decrease expression of luciferase under control of an NFκB promoter in a time and dose-response manner, thereby highlighting their anti-inflammatory properties. Interestingly, we observed that the anti-inflammatory effects of MSC EVs were able to outlast those of the positive control drug, suggesting that MSC EVs may have an increased half-life which further enhances their reparative properties. These results also suggest that MSC EVs may partially exert their functional effects through the modulation of NFκB signaling, an observation that paves the way for future mechanistic studies. Additionally, since LPS primarily activates NFκB through toll-like receptors (TLRs) and MyD88 signaling, future experiments can include stimulators (e.g., TNFα) that activate NFkB through other signaling pathways. Therefore, ThawReady™ cells have the potential to be utilized for studying multiple mechanisms of NFκB activation not only for EV-mediated cellular repair but also for studies investigating other anti-inflammatory or therapeutic compounds. Collectively, this highlights ThawReady™ as a convenient, consistent, and effective platform that can be leveraged to accelerate innovative and impactful research that has widespread application not only to the field of immunology but also to regenerative medicine.

Did you know?

ATCC now offers ThawReady™ 3-D Spheroid Kits for effortless, consistent high-throughput screening.

References

1. Bosshart H, Heinzelmann M. THP-1 cells as a model for human monocytes. Ann Transl Med 4(21): 438, 2016. PubMed: 27942529
2. ATCC. ThawReady™ Cells: A Consumable Cell Format to Streamline and Standardize Bioassay Workflows. ATCC. https://www.atcc.org/blogs/2024/thawready-cells-a-consumable-cell-format-to-streamline-and-standardize-bioassay-workflows. Published November 6, 2024. Accessed February 4, 2025.
3. Ayala-Cuellar AP, et al. Roles of Mesenchymal Stem Cells in Tissue Regeneration and Immunomodulation. Biomol Ther (Seoul) 27(1): 25-33, 2019. PubMed: 29902862
4. Kou M, et al. Mesenchymal stem cell-derived extracellular vesicles for immunomodulation and regeneration: a next generation therapeutic tool? Cell Death Dis 13(7): 580, 2022. PubMed: 35787632
5. Patel DM, Shah J, Srivastava AS. Therapeutic potential of mesenchymal stem cells in regenerative medicine. Stem Cells Int 2013: 496218, 2013. PubMed: 23577036
6. Branscome H, et al. Stem Cell Extracellular Vesicles and their Potential to Contribute to the Repair of Damaged CNS Cells. J Neuroimmune Pharmacol 15(3): 520–537, 2020. PubMed: 31338754
7. Branscome H, et al. Retroviral infection of human neurospheres and use of stem Cell EVs to repair cellular damage. Sci Rep 12(1): 2019, 2022. PubMed: 35132117
8. Hindle J, et al. hTERT-Immortalized Mesenchymal Stem Cell-Derived Extracellular Vesicles: Large-Scale Manufacturing, Cargo Profiling, and Functional Effects in Retinal Epithelial Cells. Cells 13(10): 861, 2024. PubMed: 38786083