Generation of monocytes-derived dendritic cells from chicken blood for in vitro studies

Commonly used acronym: DENDRICHICK

Scope of the method

The Method relates to
  • Animal health
The Method is situated in
  • Regulatory use - Routine production
  • Translational - Applied Research
Type of method
  • In vitro - Ex vivo
This method makes use of
  • Animal derived cells / tissues / organs
Species from which cells/tissues/organs are derived
Gallus gallus domesticus
Type of cells/tissues/organs
Blood

Description

Method keywords
  • Monocytes
  • dendritic cells
  • chicken
  • PBMC
Scientific area keywords
  • Dendritic cells isolation
  • Immunology
  • vaccination
  • immunogenicity
Method description

The purpose of the present development is to use avian MoDCs to implement a cellular platform to increase understanding of the immune responses induced by various antigens of interest (e.g. vaccine candidates) and evaluate their immunogenic potential. Considering the difficulty to work on dendritic cells (DCs) isolated ex vivo, we developed a protocol for using in vitro-generated chicken monocyte-derived dendritic cells (MoDCs). Plastic-adherent peripheral blood mononuclear cells (PBMCs) isolated from specific-pathogen free (SPF) chickens are cultured with GM-CSF and IL-4 for 5 days at 41°C in 5% CO2 atmosphere. Following antigen-induced maturation (e.g. LPS, polyinosinic-polycytidylic acid, viral antigens, etc.), typical DC morphology is confirmed by microscope. The endocytic capacity and the expression of surface expression of mannose receptor C-type 1-like B, MHC-II, myeloid markers CD11c, DEC205, and co-stimulatory molecules CD40, CD80, CD86 by MoDCs at different maturation states are assessed by flow cytometry.

Lab equipment
  • Biosafety cabinet,
  • Cell incubator,
  • Microscope,
  • Centrifuge,
  • Flow cytometer.
Method status
  • Still in development

Pros, cons & Future potential

Advantages
  • - Poorly invasive (MoDCs used in place of bone marrow-derived DCs);
  • - Makes it possible to assess the response to an antigen in a large number of individuals and thus evaluate individual variability.
Challenges

Antibodies for cell characterization are not all commercially available.

Future & Other applications
  • - Selection of immunopotentiating agents (e.g. adjuvants) during vaccine development;
  • - Antigen-pulsed MoDCs will be co-cultured with naïve purified T cells to characterize cell-mediated responses triggered by vaccine candidates in vitro.

References, associated documents and other information

References
  • - Tapia-Calle G, Stoel M, de Vries-Idema J, Huckriede A. Distinctive Responses in an In Vitro Human Dendritic Cell-Based System upon Stimulation with Different Influenza Vaccine Formulations. Vaccines (Basel) 2017;5(3):21. https://doi.org/10.3390/vaccines5030021.
  • - Kangethe, R. T., Pichler, R., Chuma, F. N. J., Cattoli, G., & Wijewardana, V. Bovine monocyte derived dendritic cell based assay for measuring vaccine immunogenicity in vitro. Veterinary Immunology and Immunopathology 2018;197,39–48. https://doi.org/10.1016/j.vetimm.2018.01.009.
  • - Wu, Z., Rothwell, L., Young, J. R., Kaufman, J., Butter, C., & Kaiser, P. Generation and characterization of chicken bone marrow-derived dendritic cells. Immunology 2010;129(1),133–145. https://doi.org/10.1111/j.1365-2567.2009.03129.x.
  • - Kamble NM, Jawale CV, Lee JH. Activation of chicken bone marrow-derived dendritic cells induced by a Salmonella Enteritidis ghost vaccine candidate. Poultry Science 2016;95:2274–80. https://doi.org/10.3382/ps/pew158.
  • - Larsen FT, Guldbrandtsen B, Christensen D, Pitcovski J, Kjærup RB, Dalgaard TS. Pustulan Activates Chicken Bone Marrow-Derived Dendritic Cells In Vitro and Promotes Ex Vivo CD4+ T Cell Recall Response to Infectious Bronchitis Virus. Vaccines (Basel) 2020;8:E226. https://doi.org/10.3390/vaccines8020226.
  • - Van den Biggelaar RHGA, van der Maas L, Meiring HD, Pennings JLA, van Eden W, Rutten VPMG, et al. Proteomic analysis of chicken bone marrow-derived dendritic cells in response to an inactivated IBV + NDV poultry vaccine. Sci Rep 2021;11:12666. https://doi.org/10.1038/s41598-021-89810-3.
Associated documents
2022_RE-Place_protocol.pdf

Contact person

Fiona Ingrao

Organisations

Sciensano
Avian Virology and Immunology
Belgium