Advertisement

Isolation of Dendritic Cells from Rat Intestinal Lymph and Spleen

  • Gordon MacPherson
  • Michelle Wykes
  • Fang-Ping Huang
  • Chris Jenkins
Part of the Methods in Molecular Medicine™ book series (MIMM, volume 64)

Abstract

Dendritic cells (DC) are rare cells in peripheral tissues, and their isolation from tissues is fraught with problems. Thus, the proportion of DC within a tissue that is extracted is unknown, isolation procedures may select for subpopulations, and the isolation procedure itself may affect their properties. As part of their life history, DC migrate from peripheral tissues, via peripheral, afferent lymph to lymph nodes, even in the absence of exogenous antigenic stimulation. They are extracted within the node and very few, if any, appear in efferent lymph. These lymph DC (L-DC) represent a population that has matured in the periphery, that may have acquired antigen (Ag), and that may be engaged in active Ag transport to lymph nodes. As such they are a physiologically relevant DC population. In large animals such as sheep and cattle, L-DC can be isolated by direct cannulation of peripheral lymphatics, but yields are relatively low. In rodents, direct cannulation of some peripheral lymphatics is possible (1), but yields of cells are minuscule. To get around this problem we and others (1 -7), have utilized lymphadenectomy as a means of collecting pseudo-afferent lymph. When lymph nodes are removed, over a period of weeks, the afferent and efferent lymphatics join as part of the healing process, leaving cells in peripheral lymph free to enter central lymph. Central lymphat ics are relatively easy to cannulate, and cannulation can be maintained for considerable periods of time (see Note 1).

Keywords

Dendritic Cell Hank Balance Salt Solution Tissue Culture Supernatant Efferent Lymphatic Splenic Dendritic Cell 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. 1.
    Pugh, C.W., MacPherson, G. G., and Steer, H. W. (1983) Characterization of nonlymphoid cells derived from rat peripheral lymph. J. Exp. Med. 157, 1758–79.PubMedCrossRefGoogle Scholar
  2. 2.
    Mayrhofer, G., Holt, P. G., and Papdimitriou, J. M. (1986) Functional characteristics of the veiled cells in afferent lymph from the rat intestine. Immunology 58, 379–387.PubMedGoogle Scholar
  3. 3.
    Harkiss, G., Hopkins, J., and McConnell, I. (1990) Uptake of antigen by afferent lymph dendritic cells mediated by antibody. Eur. J. Immunol. 20, 2367–2373.PubMedCrossRefGoogle Scholar
  4. 4.
    Bujdoso, R., Hopkins, J., Dutia, B. M., Young, P., McConnell, I. (1989) Characterisation of sheep afferent lymph dendritic cells and their role in antigen carriage. J. Exp. Med. 170, 1285–1302.PubMedCrossRefGoogle Scholar
  5. 5.
    Bujdoso, R., Harkiss, G., Hopkins, J., and McConnell, I. (1990) Afferent lymph dendritic cells: a model for antigen capture and presentation in vivo. Int. Rev. Immunol. 6, 177–86.PubMedCrossRefGoogle Scholar
  6. 6.
    Howard, C. J., Sopp, P., Brownlie, J., Parsons, K. R., Kwong, L. S., and Collins, R. A. (1996) Afferent lymph veiled cells stimulate proliferative responses in allogeneic CD4+and CD8+T cells but not gamma delta TCR+T cells. Immunology 88, 558–64.PubMedCrossRefGoogle Scholar
  7. 7.
    Howard, C. J., Sopp, P., Brownlie, J., Kwong, L. S., Parsons, K. R., and Taylor, G. (1997) Identification of two distinct populations of dendritic cells in afferent lymph that vary in their ability to stimulate T cells. J. Immunol. 159, 5372–5382.PubMedGoogle Scholar
  8. 8.
    Matsuno, K., Ezaki, T., Kudo, S., and Uehara, Y. (1996) A life stage of particleladen rat dendritic cells in vivo: their terminal division, active phagocytosis, and translocation from the liver to the draining lymph. J. Exp. Med. 183, 1865–1878.PubMedCrossRefGoogle Scholar
  9. 9.
    Kudo, S., Matsuno, K., Ezaki, T., and Ogawa, M. (1997) A novel migration pathway for rat dendritic cells from the blood: hepatic sinusoids-lymph translocation. J. Exp. Med. 185, 777–784.PubMedCrossRefGoogle Scholar
  10. 10.
    Gowans, J. L. (1959) The recirculation of lymphocytes from blood to lymph in the rat. J. Physiol. 146, 54.PubMedGoogle Scholar
  11. 11.
    Bollman, J. L., Cain, J. C., and Grindley, J. H. (1948) Techniques for the collection of lymph from the liver, small intestine or thoracic duct of the rat. J. Lab. Clin. Med. 33, 1349.PubMedGoogle Scholar
  12. 12.
    Steinman, R. M., Cohn, Z. A. (1973) Identification of a novel cell type in peripheral lymphoid organs of mice. I. Morphology, quantitation, tissue distribution. J. Exp. Med. 137, 1142–1162.PubMedCrossRefGoogle Scholar
  13. 13.
    Klinkert, W. E. F., LaBadie, J. H., and Bowers, W. E. (1982) Accessory and stimulating properites of dendritic cells and macrophages isolated from various rat tissues. J. Exp. Med. 156, 1–19.PubMedCrossRefGoogle Scholar
  14. 14.
    Metlay, J. P., Witmer-Pack, M. D., Agger, R., Crowley, M. T., Lawless, D., and Steinman, R. M. (1990) The distinct leukocyte integrins of mouse spleen dendritic cells as identified with new hamster monoclonal antibodies. J. Exp. Med. 171, 1753–1771.PubMedCrossRefGoogle Scholar
  15. 15.
    Inaba, K., Pack, M., Inaba, M., Sakuta, H., Isdell, F., and Steinman, R. M. (1997) High levels of amajorhistocompatibility complex II-self peptide complex on dendritic cells from the T cell areas of lymph nodes. J. Exp. Med. 186, 665–672.PubMedCrossRefGoogle Scholar
  16. 16.
    Schuler, G., and Steinman, R. M. (1985) Murine epidermal Langerhans cells mature into potent immunostimulatory dendritic cells in vitro. J. Exp. Med. 161, 526–546.PubMedCrossRefGoogle Scholar
  17. 17.
    Austyn, J. M., Hankins, D. F., Larsen, C. P., Morris, P. J., Rao, A. S., Roake, J. A. (1994) Isolation and characterization of dendritic cells from mouse heart and kidney. J. Immunol. 152, 2401–2410.PubMedGoogle Scholar
  18. 18.
    MacPherson, G. G. (1989) Properties of lymph-borne (veiled) dendritic cells in culture. I. Modulation of phenotype, survival and function: partial dependence on GM-CSF. Immunology 68, 102–107.PubMedGoogle Scholar
  19. 19.
    Leenen, P. J., Radosevic, K., Voerman, J. S., et al. (1998) Heterogeneity of mouse spleen dendritic cells: in vivo phagocytic activity, expression of macrophage markers, and subpopulation turnover. J. Immunol. 160, 2166–2173.PubMedGoogle Scholar
  20. 20.
    Gyure, L. and Hall, J. (1990) A quick method for obtaining lymph borne dendritic macrophages from rats. J. Immunol. Methods. 131, 49–53.PubMedCrossRefGoogle Scholar
  21. 21.
    Liu, L. M., Zhang, M., Jenkins, C., and MacPherson, G. G. (1998) Dendritic cell heterogeneity in vivo: Two functionally different dendritic cell populations in rat intestinal lymph can be distinguished by CD4 expression. J. Immunol. 161, 1146–1155.PubMedGoogle Scholar

Copyright information

© Humana Press Inc. 2001

Authors and Affiliations

  • Gordon MacPherson
    • 1
  • Michelle Wykes
    • 1
  • Fang-Ping Huang
    • 1
  • Chris Jenkins
    • 1
  1. 1.Sir William Dunn School of PathologyUniversity of OxfordOxfordUK

Personalised recommendations