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Determining Activation Status of Natural Killer Cells Following Stimulation via Cytokines and Surface Receptors

Lizeth G Meza Guzman1,2, Sandra E Nicholson3,4

  • 1The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia. meza.l@wehi.edu.au.

Methods in Molecular Biology (Clifton, N.J.)
|March 28, 2022
PubMed

Insights

This study explores Natural Killer (NK) cell activation pathways by measuring key markers like interferon gamma (IFNγ) and CD107a. Researchers utilized flow cytometry to analyze NK cell responses to various stimuli, providing insights into immune cell signaling.

Area of Science:

  • Immunology
  • Cellular Biology
  • Molecular Biology

Background:

  • Natural Killer (NK) cells are crucial for innate immunity, responding to cytokines, target cells, and antibody-mediated receptor activation.
  • Understanding NK cell signaling pathways is vital for dissecting immune responses and developing immunotherapies.
  • Standard methods for assessing NK cell activation include measuring interferon gamma (IFNγ) production and CD107a (LAMP-1) expression.

Purpose of the Study:

  • To detail methods for dissecting specific signaling intermediates in distinct NK cell activation pathways.
  • To outline the measurement of NK cell activation status via IFNγ production and LAMP-1 (CD107a) expression.
  • To describe the assessment of cytotoxic potency through perforin, granzyme, and TNFα production.

Main Methods:

  • NK cell activation was induced using cytokines, target cell interactions, or antibody-mediated receptor engagement.
  • Antibodies were immobilized on cell culture plates to facilitate receptor crosslinking for in vitro activation assays.
  • Flow cytometry was employed to measure multiple parameters of NK cell activation and cytotoxicity.

Main Results:

  • The study demonstrates that various stimuli can activate distinct NK cell signaling pathways.
  • Interferon gamma (IFNγ) production and LAMP-1 (CD107a) expression serve as reliable indicators of NK cell activation status.
  • Perforin, granzyme, and tumor necrosis factor alpha (TNFα) levels reflect the cytotoxic potential of activated NK cells.

Conclusions:

  • This work provides a comprehensive framework for studying NK cell activation mechanisms.
  • The described methods allow for detailed analysis of signaling intermediates in different NK cell activation contexts.
  • Flow cytometry is presented as a versatile tool for quantifying NK cell responses and cytotoxic functions.
Keywords:
Activating receptorsAntibody-directed activationCytokineEffector cellFlow cytometryIFNγCD107aNK cellNatural Killer cellTarget cell

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