Pharmaceutical Research

, Volume 19, Issue 10, pp 1480–1487 | Cite as

Analysis of Poly(D,L-Lactic-Co-Glycolic Acid) Nanosphere Uptake by Human Dendritic Cells and Macrophages In Vitro

  • M. E. Christine Lutsiak
  • Deborah R. Robinson
  • Conrad Coester
  • Glen S. Kwon
  • John SamuelEmail author


Purpose. The purpose of this study was to demonstrate and characterize phagocytosis of poly(D,L-lactic-co-glycolic acid) (PLGA) nanospheres by human dendritic cells (DCs).

Methods. Parallel cultures of DCs and macrophages (Mφ) were established from peripheral blood leukocytes using media supplemented with granulocyte-macrophage colony stimulator factor and interleukin-4 (for DC) or granulocyte-macrophage colony stimulator factor alone (for Mφ). PLGA nanospheres containing tetramethylrhodamine-labeled dextran with or without an adjuvant, monophosphoryl lipid A, were prepared using a water/oil/water solvent evaporation technique. Cells were incubated with the nanospheres for 24 h. Confocal laser scanning microscopy was used to determine the intracellular location of the nanospheres and flow cytometry to measure the fraction of phagocytic cells in the culture and the amount of uptake per cell. After phagocytosis, cells were stained for MHC class II molecules, CD14, CD80, and CD86 to identify the phagocytic population.

Results. DCs phagocytosed PLGA nanospheres as efficiently as Mφ. Cell-surface marker expression conclusively established that the phagocytic cells were DC.

Conclusions. DCs can take up PLGA nanospheres. Because DCs are the key professional antigen-presenting cells capable of stimulating naive T cells, our data suggest that PLGA nanospheres can be used as an efficient delivery system for vaccines designed to activate T cell-mediated immune responses.

PLGA nanospheres dendritic cells antigen-presenting cells phagocytosis 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    J. Banchereau and R. M. Steinman. Dendritic cells and the control of immunity. Nature 392:245-252 (1998).Google Scholar
  2. 2.
    M. Croft, D. Duncan, and S. Swain. Response of naive antigenspecific CD4+ T cells in vitro: Characteristics and antigenpresenting cell requirements. J.Exp.Med. 176:1431-1437 (1992).Google Scholar
  3. 3.
    F. Fagnoni, M. Takamizawa, W. Godfrey, A. Rivas, M. Azuma, K. Okumura, and E. Engleman. Role of B70/B7-2 in CD4+ T-cell immune responses induced by dendritic cells. Immunology 85:467-474 (1995).Google Scholar
  4. 4.
    J. Guery and L. Adorini. Dendritic cells are the most efficient in presenting endogenous naturally processed self-epitopes to class II-restricted T cells. J.Immunol. 154:536-544 (1995).Google Scholar
  5. 5.
    R. Thomas, L. Davis, and P. Lipsky. Comparative accessory cell function of human peripheral blood dendritic cells and monocytes. J.Immunol. 151:6840-6852 (1993).Google Scholar
  6. 6.
    S. Macatonia, N. Hosken, M. Litton, P. Vieira, C. Hsieh, J. Culpepper, M. Wysocka, G. Trinchieri, K. Murphy, and A. O'Garra. Dendritic cells produce IL-12 and direct the development of Th1 cells from naive CD4+ T cells. J.Immunol. 154:5071-5079 (1995).Google Scholar
  7. 7.
    C. Heufler, F. Koch, U. Stanzl, G. Topar, M. Wysocka, G. Trinchieri, A. Enk, R. Steinman, N. Romani, and G. Schuler. Interleukin-12 is produced by dendritic cells and mediates T helper 1 development as well as interferon-gamma production by T helper 1 cells. Eur.J.Immunol. 26:659-668 (1996).Google Scholar
  8. 8.
    L. Fong and E. Engleman. Dendritic cells in Cancer Immunotherapy. Annu.Rev.Immunol. 18:245-273 (2000).Google Scholar
  9. 9.
    J. T. Timmerman and R. Levy. Dendritic cell vaccines for cancer immunotherapy. Annu.Rev.Med. 50:507-529 (1999).Google Scholar
  10. 10.
    M. D. Witmer-Pack, M. T. Crowley, K. Inaba, and R. M. Steinman. Macrophages, but not dendritic cells, accumulate colloidal carbon following administration in situ. J.Cell Sci. 105:965-973 (1993).Google Scholar
  11. 11.
    C. Reise Sousa and J. M. Austyn. Phagocytosis of antigens by Langerhans cells. In Dendritic Cells in Fundamental and Clinical Immunology, 1993, pp. 199-204.Google Scholar
  12. 12.
    L. Filgueira, F. O. Nestle, M. Rittig, H. I. Joller, and P. Groscurth. Human dendritic cells phagocytose and process Borrelia burgdorferi. J.Immunol. 157:2998-3005 (1996).Google Scholar
  13. 13.
    K. Inaba, M. Inaba, M. Naito, and R. M. Steinman. Dendritic cell progenitors phagocytose particulates, including Bacillus calmetteguerin organisms, and sensitize mice to mycobacterial antigens in vivo. J.Exp.Med. 178:479-488 (1993).Google Scholar
  14. 14.
    J. Ma, D. Luo, G. S. Kwon, J. Samuel, A. A. Noujaim, and R. Madiyalakan. Use of encapsulated single chain antibodies for induction of anti-idiotypic humoral and cellular immune responses. J.Pharm.Sci. 87:1375-1378 (1998).Google Scholar
  15. 15.
    J. Ma, J. Samuel, G. S. Kwon, A. A. Noujaim, and R. Madiyalakan. Induction of anti-idiotypic humoral and cellular immune responses by a murine monoclonal antibody recognizing the ovarian carcinoma antigen CA125 encapsulated in biodegradable microspheres. Cancer Immunol.Immunother. 47:13-20 (1998).Google Scholar
  16. 16.
    K. D. Newman, J. Samuel, and G. Kwon. Ovalbumin peptide encapsulated in poly(d,l lactic-co-glycolic acid) microspheres is capable of inducing a T helper type 1 immune response. J.Control.Release 54:49-59 (1998).Google Scholar
  17. 17.
    K. D. Newman, D. L. Sosnowski, G. S. Kwon, and J. Samuel. Delivery of MUC1 mucin peptide by Poly(d,l-lactic-co-glycolic acid) microspheres induces type 1 T helper immune responses. J.Pharm.Sci. 87:1421-1427 (1998).Google Scholar
  18. 18.
    D. Wang, D. R. Robinson, G. S. Kwon, and J. Samuel. Encapsulation of plasmid DNA in biodegradable poly(D, L-lactic-coglycolic acid) microspheres as a novel approach for immunogene delivery. J.Control.Release 57:9-18 (1999).Google Scholar
  19. 19.
    C. D. Partidos, P. Vohra, C. Anagnostopoulou, D. H. Jones, G. H. Farrar, and M. W. Steward. Biodegradable microparticles as a delivery system for measles virus cytotoxic T cell epitopes. Mol.Immunol. 33:485-491 (1996).Google Scholar
  20. 20.
    C. D. Partidos, P. Vohra, D. H. Jones, G. H. Farrar, and M. W. Steward. Mucosal immunization with a measles virus CTL epitope encapsulated in biodegradable PLG microparticles. J.Immunol.Methods 195:135-138 (1996).Google Scholar
  21. 21.
    D. F. Nixon, C. Hioe, P. D. Chen, Z. Bian, P. Kuebler, M. L. Li, H. Qiu, X. M. Li, M. Singh, J. Richardson, P. McGee, T. Zamb, W. Koff, C. Y. Wang, and D. O'Hagan. Synthetic peptides entrapped in microparticles can elicit cytotoxic T cell activity. Vaccine 14:1523-1530 (1996).Google Scholar
  22. 22.
    G. D. Moore and H. B. Croxatto. Synthetic microspheres transferred to the rat oviduct on day 1 of pregnancy mimic the transport of native ova. J.Reprod.Fertil. 82:735-742 (1988).Google Scholar
  23. 23.
    F. Sallusto and A. Lanzavecchia. Efficient presentation of soluble antigen by cultured human dendritic cells is maintained by granulocyte/macrophage colony-stimulating factor plus interleukin-4 and downregulated by tumor necrosis factor alpha. J.Exp.Med. 179:1109-1118 (1994).Google Scholar
  24. 24.
    F. Chapuis, M. Rosenzwajg, M. Yagello, M. Ekman, P. Biberfeld, and J. C. Gluckman. Differentiation of human dendritic cells from monocytes in vitro. Eur.J.Immunol. 27:431-441 (1997).Google Scholar
  25. 25.
    S. M. Kiertscher and M. D. Roth. Human CD14+ leukocytes acquire the phenotype and function of antigen-presenting dendritic cells when cultured in GM-CSF and IL-4. J.Leukoc.Biol. 59:208-218 (1996).Google Scholar
  26. 26.
    Y. Ogawa, M. Tamamoto, H. Okada, T. Yashiki, and T. Shimamoto. A new technique to efficiently entrap leuprolide acetate into microcapsules of polylactic acid or copoly(lactic/glycolic) acid. Chem.Pharm.Bull. 36:1095-1103 (1988).Google Scholar
  27. 27.
    A. T. Davis, R. Estensen, and P. G. Quie. Cytochalasin B. 3. Inhibition of human polymorphonuclear leukocyte phagocytosis. Proc.Exp.Biol.Med. 137:161-164 (1971).Google Scholar
  28. 28.
    H. Xu, M. Kramer, H. P. Spengler, and J. H. Peters. Dendritic cells differentiated from human monocytes through a combination of IL-4, GM-CSF and IFN-gamma exhibit phenotype and function of blood dendritic cells. Adv.Exp.Med.Biol. 378:75-78 (1995).Google Scholar
  29. 29.
    C. Schutt. CD14. Int.J.Biochem.Cell Biol. 31:545-549 (1999).Google Scholar
  30. 30.
    A. Aderem and D. Underhill. Mechanisms of phagocytosis in macrophages. Annu.Rev.Immunol. 17:593-623 (1999).Google Scholar

Copyright information

© Plenum Publishing Corporation 2002

Authors and Affiliations

  • M. E. Christine Lutsiak
    • 1
  • Deborah R. Robinson
    • 1
  • Conrad Coester
    • 1
  • Glen S. Kwon
    • 1
  • John Samuel
    • 1
    Email author
  1. 1.Faculty of Pharmacy and Pharmaceutical SciencesUniversity of AlbertaEdmonton, ABCanada

Personalised recommendations