National Avian Research Facility
National Avian Research Facility

CSF1R-reporter lines

Macrophage reporter lines – CSF1R-eGFP (MacGreen) and CSF1R-mApple (MacRed).

Summary

The CSF1R-eGFP and CSF1R-mApple reporter transgenic chickens can be used to visualise immune cells of the mononuclear phagocyte system (monocytes, macrophages and dendritic cells).  In addition to antigen sampling epithelial cells (also known as M-cells). Colony stimulating factor 1 receptor (CSF1R) acts as the receptor for colony stimulating factor 1, a cytokine that controls the production, differentiation, and function of macrophages in vertebrates. Transgene expression and cell-surface CSF1R are also detectable in heterophil granulocytes, but at a level approximately one-tenth of that in the monocytes1. Cells of the chicken mononuclear phagocyte system can be detected by both flow cytometry and immunofluorescence staining1–5. Transgene expression in heterophils can only be visualised by flow cytometry. The brightness and specificity of transgene gene expression in the CSF1R-reporter transgenic chickens enable visualisation of these lymphoid structures in both embryonic and post-hatch chickens1,2.

 CSF1R-eGFP (green) transgene expression ileum
External views of the ileum in the small intestine of a 7-day old CSF1R-eGFP chick. The high concentration of eGFP+ aggregated cells enables visualisation of the Peyer's Patch. Scale bar = 500 µm.
CSF1R-eGFP chicken
CSF1R-eGFP transgene expression in the follicle-associated epithelium of the bursa of Fabricius. F-actin (red) and jacalin lectin (blue) staining also shown.

 

Utility

The CSF1R-reporter transgenic chickens have improved understanding of the embryonic development of the mononuclear phagocyte system and CSF1R signalling2. The CSF1R-reporter transgenic chickens have been used to investigate how the immune system responds to avian pathogenic Escherichia coli infection6, Avian Influenza Virus7, Infectious Bronchitis Virus and Newcastle Disease Virus (Vervelde et al. In Prep.). Furthermore, CSF1R-reporter transgenic chickens have been used to improve knowledge of the avian respiratory immune system3, and facilitated the development of precision-cut lung slices ex vivo to examine avian host–pathogen interactions and inflammatory responses7,8Both CSF1R-eGFP and CSF1R-mApple reporter transgenic chickens can be used to identify and study chicken M-cells in mucosal tissues9.

 

Line origin

The CSF1R-reporter transgenic chickens were developed by Dr Adam Balic, Professor Helen Sang, and Professor David Hume, and funded by BBSRC. The CSF1R-reporter lines have reporter genes that are expressed in a specific immune cell lineage, based upon control elements of the colony stimulating factor 1 receptor (CSF1R) locus1. The CSF1R-reporter lines were created by use of lentiviral gene transfer. The CSF1R control elements utilised in the construction of the transgene consist of the CSF1R conserved regulatory element, Fms intronic regulatory element (FIRE), and the CSF1R promoter region. The cell lineage specificity of reporter gene expression was initially confirmed by demonstration of coincident expression with the endogenous CSF1R protein1. The CSF1R-reporter lines differ in the use of fluorescent reporter protein, eGFP (green) or mApple (red).

 

If you use the CSF1R-reporter lines for your research, please reference; Balic, A. et al. Visualisation of chicken macrophages using transgenic reporter genes: Insights into the development of the avian macrophage lineage. Development, 141, 3255-3265 (2014), in your publications.

CSF1R-eGFP and CSF1R-mApple Composite
Visualisation of organized lymphoid structures in the respiratory (a-c) and intestinal (d-g) tract of CSF1R-reporter transgenic chickens using whole mount analysis. a. Airsac. b. Trachea. c. BALT region in the intrapulmonary bronchi. d. Peyer’s patch in small intestine. e. Meckel’s diverticulum. f. Caecal tonsil. g. Lymphoid aggregates in the large intestine.

Publications

  1. Balic, A. et al. Visualisation of chicken macrophages using transgenic reporter genes: Insights into the development of the avian macrophage lineage. Development 141, 3255–3265 (2014).
  2. Garceau, V. et al. The development and maintenance of the mononuclear phagocyte system of the chick is controlled by signals from the macrophage colony-stimulating factor receptor. BMC Biol. 13, (2015).
  3. Sutton, K. et al. Visualisation and characterisation of mononuclear phagocytes in the chicken respiratory tract using CSF1R-transgenic chickens. Vet. Res. 49, 1–14 (2018).
  4. Dora, D. et al. Intraganglionic macrophages: a new population of cells in the enteric ganglia. J. Anat. 233, 401–410 (2018).
  5. Garcia-Morales, C. et al. Production and characterisation of a monoclonal antibody that recognises the chicken CSF1 receptor and confirms that expression is restricted to macrophage-lineage cells. Dev. Comp. Immunol. 42, 278–285 (2014).
  6. Alber, A. et al. Avian Pathogenic Escherichia coli (APEC) Strain-Dependent Immunomodulation of Respiratory Granulocytes and Mononuclear Phagocytes in CSF1R-Reporter Transgenic Chickens. Front. Immunol. 10, 3055 (2020).
  7. Bryson, K. J. et al. Precision cut lung slices: A novel versatile tool to examine host-pathogen interaction in the chicken lung. Vet. Res. 51, 1–16 (2020).
  8. Garrido, D. et al. The role of type I interferons (IFNs) in the regulation of chicken macrophage inflammatory response to bacterial challenge. Dev. Comp. Immunol. 86, 156–170 (2018).
  9. Balic, A. et al. Antigen sampling csf1r-expressing epithelial cells are the functional equivalents of mammalian m cells in the avian follicle-associated epithelium. Front. Immunol. 10, 2495 (2019).