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An Optimized Method for Isolating and Expanding Invariant Natural Killer T Cells from Mouse Spleen
Published on: October 29, 2015
Optimized ex vivo expansion and functional characterization of canine natural killer cells using genetically modified
Yurim An1, Geun-Seop Kim2,3, YeonJu Baek1
1Department of Applied Biotechnology, College of Industrial Science, Kongju National University, Yesan 32439, Korea.
Importance:
Natural killer (NK) cells are critical effectors of innate immune surveillance and can eliminate malignant cells without prior sensitization. Given the biological and genetic parallels between canine and human cancers, NK cell-based immunotherapies have strong translational potential in veterinary oncology. However, the lack of an efficient and reproducible ex vivo expansion protocol for canine NK cells remains a major barrier to clinical application. Establishing culture conditions for expansion of functional canine NK cells would support adoptive NK cell therapy and translational research.
Objective:
This study aimed to evaluate the effectiveness of genetically modified feeder cells and optimized culture media in expanding canine NK cells.
Methods:
Canine peripheral blood mononuclear cells were co-cultured with irradiated genetically modified K562 or ARH77 cells in either RPMI-1640 or Dulbecco's Modified Eagle Medium (DMEM)/F12-based media. NK cell expansion, purity, cytotoxicity and expression of NK cell-associated receptors were evaluated using flow cytometry, cytotoxicity assays, and quantitative reverse transcription polymerase chain reaction.
Results:
The combination of DMEM/F12-based medium and genetically modified ARH77 feeder cells yielded the highest NK cell expansion rate and purity among the tested conditions. Cytotoxicity differed significantly according to medium type, whereas no significant difference was observed between feeder cell types. In addition, the NK cell-associated receptors were significantly upregulated in expanded NK cells.
Conclusions And Relevance:
Genetically modified ARH77 feeder cells with DMEM/F12-based culture media demonstrated superior efficiency in promoting the ex vivo expansion of canine NK cells, achieving high purity and enhanced cytotoxic potential. These findings highlight a promising and reproducible platform for generating functional NK cells, offering a practical and scalable strategy to support the development of NK cell-based veterinary immunotherapy.
