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Flow cytometric identification and cell-line establishment of macrophages in naked mole-rats
Haruka Wada1, Yuhei Shibata1,2, Yurika Abe1
1Division of Immunobiology, Institute for Genetic Medicine, Hokkaido University, Sapporo, Japan.
Abstract:
Naked mole rats (NMRs) have extraordinarily long lifespans and anti-tumorigenic capability. Recent studies of humans and mice have shown that many age-related diseases, including cancer, are strongly correlated with immunity, and macrophages play particularly important roles in immune regulation. Therefore, NMR macrophages may contribute to their unique phenotypes. However, studies of the roles of macrophages are limited by material restrictions and the lack of an established experimental strategy. In this study, we developed a flow cytometric strategy to identify NMR macrophages. The NMR macrophages were extractable using an off-the-shelf anti-CD11b antibody, M1/70, and forward/side scatter data obtained by flow cytometry. NMR macrophages proliferated in response to human/mouse recombinant M-CSF and engulfed Escherichia coli particles. Interestingly, the majority of NMR macrophages exhibited co-staining with an anti-NK1.1 antibody, PK136. NK1.1 antigen crosslinking with PK136 results in mouse NK cell stimulation; similarly, NMR macrophages proliferated in response to NK1.1 antibody treatment. Furthermore, we successfully established an NMR macrophage cell line, NPM1, by transduction of Simian virus 40 early region that proliferated indefinitely without cytokines and retained its phagocytotic capacity. The NPM1 would contribute to further studies on the immunity of NMRs.
Insights
Naked mole rat (NMR) macrophages were identified using a novel flow cytometry method. A new cell line, NPM1, was established, aiding future NMR immunity research.
Area of Science:
- Comparative immunology
- Rodent models
- Longevity research
Background:
- Naked mole rats (NMRs) exhibit exceptional longevity and cancer resistance.
- Immune system, particularly macrophages, is linked to aging and disease in mammals.
- Understanding NMR macrophages is key to their unique anti-aging and anti-cancer properties.
Purpose of the Study:
- To develop a reliable method for identifying and isolating NMR macrophages.
- To characterize the functional properties of NMR macrophages.
- To establish a continuous NMR macrophage cell line for further research.
Main Methods:
- Utilized flow cytometry with anti-CD11b antibody (M1/70) and scatter data for NMR macrophage identification.
- Assessed macrophage proliferation in response to M-CSF and phagocytosis of E. coli.
- Investigated NK1.1 antibody (PK136) crosslinking effects on NMR macrophages.
- Established an NMR macrophage cell line (NPM1) using Simian virus 40 early region transduction.
Main Results:
- Successfully identified and isolated NMR macrophages via flow cytometry.
- NMR macrophages demonstrated M-CSF-dependent proliferation and phagocytic activity.
- NK1.1 antibody treatment stimulated NMR macrophage proliferation.
- Established the NPM1 cell line, which proliferates indefinitely and retains phagocytosis.
Conclusions:
- Developed a robust flow cytometry strategy for NMR macrophage research.
- NMR macrophages possess functional characteristics, including proliferation and phagocytosis, and respond to NK1.1 stimulation.
- The NPM1 cell line provides a valuable tool for investigating NMR immunity and longevity mechanisms.
