Three-dimensional two-layer collagen matrix gel culture model for evaluating complex biological functions of monocyte-derived dendritic cells.
Dendritic cell-like cells (Mo-DCs) generated from peripheral blood monocytes with interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) have been used as tools to treat cancer patients (DC-vaccines). Because Mo-DCs have multiple antigen presentation-related functions, including phagocytosis, migration, cytokine production, and T cell stimulation, establishment of a method for simultaneously evaluating the various functions of Mo-DCs is important. We developed a new in vitro three-dimensional two-layer collagen matrix culture model that consists of a collagen gel containing Mo-DCs as the lower layer and a collagen gel containing necrotic GCTM-1 tumor cells and/or T cells as the upper layer. We used this system to observe simultaneously multiple functions of Mo-DCs by phase-contrast or fluorescence microscopy and to assess IL-12 secretion during more than 2 weeks of culture. We also observed interactions between Mo-DCs and necrotic GCTM-1 or T cells on an individual cell basis by time-lapse videomicroscopy. In addition, we collected Mo-DCs from the collagen gels by collagenase treatment and analyzed the expression of antigen presentation-related molecules such as HLA-DR, CD80, CD83, and CD86 on Mo-DCs. This model may be a useful tool for evaluation of the various functions of Mo-DCs used as DC vaccines and for studies of the complex behaviors of Mo-DCs in vivo.
Tasaki
Akira
A
Department of Cancer Therapy and Research, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8525, Japan.
Yamanaka
Naoki
N
Kubo
Makoto
M
Matsumoto
Kotaro
K
Kuroki
Hideo
H
Nakamura
Katsuya
K
Nakahara
Chihiro
C
Onishi
Hideya
H
Kuga
Hirotaka
H
Baba
Eishi
E
Tanaka
Masao
M
Morisaki
Takashi
T
Katano
Mitsuo
M
D017951:Antigen Presentation / Q000276:immunology; D018929:Cell Culture Techniques / Q000379:methods*; D045744:Cell Line, Tumor; D019070:Cell Lineage; D002465:Cell Movement; D002470:Cell Survival; D018920:Coculture Techniques; D003094:Collagen; D003713:Dendritic Cells / Q000502:physiology*; D005782:Gels; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D018664:Interleukin-12 / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008858:Microscopy, Phase-Contrast; D018715:Microscopy, Video; D009000:Monocytes / Q000166:cytology; D010587:Phagocytosis; D013601:T-Lymphocytes / Q000276:immunology
D005782:Gels; D018664:Interleukin-12; D007371:Interferon-gamma; D003094:Collagen