Feasibility to generate monocyte-derived dendritic cell from coculture with melanoma tumor cells in the presence of granulocyte/macrophage colony-stimulating factor (GM-CSF) and interleukin-4

Young T. Kim, Evan M Hersh, Katrina T. Trevor

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Objective: We investigated how melanoma cells and membrane-bound granulocyte/macrophage colony stimulating factor (mbGM-CSF) melanoma cell lines affect the differentiation of dendritic cells (DC) from CD14+ monocytes. Method of study: The malignant melanoma cell lines (Conley and Jorp) and mbGM-CSF-positive cell lines (Conley B-F8 and Jorp C-E6) were cultured and cell-free supernatants were collected from each cell line cultures to assess the GM-CSF level. Adherent monocytes were cocultured for 6-7 days with irradiated mbGM-CSF and wild type melanoma cells (50 Gy) at each 1 : 1 and 0.1 : 1 ratio in six-well culture plates in ex vivo culture medium containing interleukin (IL)-4. Immunophenotyping was performed by triple color immunofluorescence staining with flow cytometry analysis. Results: GM-CSF was detected at low levels in the culture supernatants of Conley B-F8 (0.48 ng/106 cells/24 hr), whereas there was no detectable GM-CSF in that of Conley melanoma cell line. Monocytes cultured with GM-CSF/IL-4 generated the expression of high levels of HLA DR, CD la and CD86, while the expression of CD 14 and CD83 were in low amounts. However, the addition of GM-CSF to these cultures resulted in an increased expression of these markers and decreased that of CD14. Monocytes cocultured with Jorp C-E6 illustrated similar expression pattern of CD la, HLA DR and CD 14 in the presence or absence of GM-CSF as Conley B-F8 melanoma cell line. Monocytes cocultured with Conley B-F8 melanoma cells at 1 : 1 and 0.1 : 1 ratio showed no significant difference in expression of CD la, CD14 and CD83 between the two ratios. Conclusion: Our results illustrate the feasibility to generate monocyte-derived DC from coculture with melanoma tumor cells in the presence of GM-CSF and IL-4. However, mbGM-CSF tumor cells did not significantly enhance the DC differentiation through juxtacrine stimulation unless soluble GM-CSF was added in the culture media.

Original languageEnglish (US)
Pages (from-to)230-238
Number of pages9
JournalAmerican Journal of Reproductive Immunology
Volume49
Issue number4
DOIs
StatePublished - Apr 1 2003

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Granulocyte-Macrophage Colony-Stimulating Factor
Coculture Techniques
Interleukin-4
Dendritic Cells
Monocytes
Melanoma
Neoplasms
Cell Line
HLA-DR Antigens
Membranes
Culture Media
Immunophenotyping
Fluorescent Antibody Technique
Cell Differentiation
Cultured Cells
Flow Cytometry
Color
Cell Culture Techniques
Cell Membrane
Staining and Labeling

Keywords

  • Coculture
  • Dendritic cell
  • GM-CSF
  • mbGM-CSF

ASJC Scopus subject areas

  • Immunology
  • Obstetrics and Gynecology

Cite this

@article{67f8a88833a84efc8fba9678654fbbdf,
title = "Feasibility to generate monocyte-derived dendritic cell from coculture with melanoma tumor cells in the presence of granulocyte/macrophage colony-stimulating factor (GM-CSF) and interleukin-4",
abstract = "Objective: We investigated how melanoma cells and membrane-bound granulocyte/macrophage colony stimulating factor (mbGM-CSF) melanoma cell lines affect the differentiation of dendritic cells (DC) from CD14+ monocytes. Method of study: The malignant melanoma cell lines (Conley and Jorp) and mbGM-CSF-positive cell lines (Conley B-F8 and Jorp C-E6) were cultured and cell-free supernatants were collected from each cell line cultures to assess the GM-CSF level. Adherent monocytes were cocultured for 6-7 days with irradiated mbGM-CSF and wild type melanoma cells (50 Gy) at each 1 : 1 and 0.1 : 1 ratio in six-well culture plates in ex vivo culture medium containing interleukin (IL)-4. Immunophenotyping was performed by triple color immunofluorescence staining with flow cytometry analysis. Results: GM-CSF was detected at low levels in the culture supernatants of Conley B-F8 (0.48 ng/106 cells/24 hr), whereas there was no detectable GM-CSF in that of Conley melanoma cell line. Monocytes cultured with GM-CSF/IL-4 generated the expression of high levels of HLA DR, CD la and CD86, while the expression of CD 14 and CD83 were in low amounts. However, the addition of GM-CSF to these cultures resulted in an increased expression of these markers and decreased that of CD14. Monocytes cocultured with Jorp C-E6 illustrated similar expression pattern of CD la, HLA DR and CD 14 in the presence or absence of GM-CSF as Conley B-F8 melanoma cell line. Monocytes cocultured with Conley B-F8 melanoma cells at 1 : 1 and 0.1 : 1 ratio showed no significant difference in expression of CD la, CD14 and CD83 between the two ratios. Conclusion: Our results illustrate the feasibility to generate monocyte-derived DC from coculture with melanoma tumor cells in the presence of GM-CSF and IL-4. However, mbGM-CSF tumor cells did not significantly enhance the DC differentiation through juxtacrine stimulation unless soluble GM-CSF was added in the culture media.",
keywords = "Coculture, Dendritic cell, GM-CSF, mbGM-CSF",
author = "Kim, {Young T.} and Hersh, {Evan M} and Trevor, {Katrina T.}",
year = "2003",
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language = "English (US)",
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journal = "American Journal of Reproductive Immunology and Microbiology",
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TY - JOUR

T1 - Feasibility to generate monocyte-derived dendritic cell from coculture with melanoma tumor cells in the presence of granulocyte/macrophage colony-stimulating factor (GM-CSF) and interleukin-4

AU - Kim, Young T.

AU - Hersh, Evan M

AU - Trevor, Katrina T.

PY - 2003/4/1

Y1 - 2003/4/1

N2 - Objective: We investigated how melanoma cells and membrane-bound granulocyte/macrophage colony stimulating factor (mbGM-CSF) melanoma cell lines affect the differentiation of dendritic cells (DC) from CD14+ monocytes. Method of study: The malignant melanoma cell lines (Conley and Jorp) and mbGM-CSF-positive cell lines (Conley B-F8 and Jorp C-E6) were cultured and cell-free supernatants were collected from each cell line cultures to assess the GM-CSF level. Adherent monocytes were cocultured for 6-7 days with irradiated mbGM-CSF and wild type melanoma cells (50 Gy) at each 1 : 1 and 0.1 : 1 ratio in six-well culture plates in ex vivo culture medium containing interleukin (IL)-4. Immunophenotyping was performed by triple color immunofluorescence staining with flow cytometry analysis. Results: GM-CSF was detected at low levels in the culture supernatants of Conley B-F8 (0.48 ng/106 cells/24 hr), whereas there was no detectable GM-CSF in that of Conley melanoma cell line. Monocytes cultured with GM-CSF/IL-4 generated the expression of high levels of HLA DR, CD la and CD86, while the expression of CD 14 and CD83 were in low amounts. However, the addition of GM-CSF to these cultures resulted in an increased expression of these markers and decreased that of CD14. Monocytes cocultured with Jorp C-E6 illustrated similar expression pattern of CD la, HLA DR and CD 14 in the presence or absence of GM-CSF as Conley B-F8 melanoma cell line. Monocytes cocultured with Conley B-F8 melanoma cells at 1 : 1 and 0.1 : 1 ratio showed no significant difference in expression of CD la, CD14 and CD83 between the two ratios. Conclusion: Our results illustrate the feasibility to generate monocyte-derived DC from coculture with melanoma tumor cells in the presence of GM-CSF and IL-4. However, mbGM-CSF tumor cells did not significantly enhance the DC differentiation through juxtacrine stimulation unless soluble GM-CSF was added in the culture media.

AB - Objective: We investigated how melanoma cells and membrane-bound granulocyte/macrophage colony stimulating factor (mbGM-CSF) melanoma cell lines affect the differentiation of dendritic cells (DC) from CD14+ monocytes. Method of study: The malignant melanoma cell lines (Conley and Jorp) and mbGM-CSF-positive cell lines (Conley B-F8 and Jorp C-E6) were cultured and cell-free supernatants were collected from each cell line cultures to assess the GM-CSF level. Adherent monocytes were cocultured for 6-7 days with irradiated mbGM-CSF and wild type melanoma cells (50 Gy) at each 1 : 1 and 0.1 : 1 ratio in six-well culture plates in ex vivo culture medium containing interleukin (IL)-4. Immunophenotyping was performed by triple color immunofluorescence staining with flow cytometry analysis. Results: GM-CSF was detected at low levels in the culture supernatants of Conley B-F8 (0.48 ng/106 cells/24 hr), whereas there was no detectable GM-CSF in that of Conley melanoma cell line. Monocytes cultured with GM-CSF/IL-4 generated the expression of high levels of HLA DR, CD la and CD86, while the expression of CD 14 and CD83 were in low amounts. However, the addition of GM-CSF to these cultures resulted in an increased expression of these markers and decreased that of CD14. Monocytes cocultured with Jorp C-E6 illustrated similar expression pattern of CD la, HLA DR and CD 14 in the presence or absence of GM-CSF as Conley B-F8 melanoma cell line. Monocytes cocultured with Conley B-F8 melanoma cells at 1 : 1 and 0.1 : 1 ratio showed no significant difference in expression of CD la, CD14 and CD83 between the two ratios. Conclusion: Our results illustrate the feasibility to generate monocyte-derived DC from coculture with melanoma tumor cells in the presence of GM-CSF and IL-4. However, mbGM-CSF tumor cells did not significantly enhance the DC differentiation through juxtacrine stimulation unless soluble GM-CSF was added in the culture media.

KW - Coculture

KW - Dendritic cell

KW - GM-CSF

KW - mbGM-CSF

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U2 - 10.1034/j.1600-0897.2003.01200.x

DO - 10.1034/j.1600-0897.2003.01200.x

M3 - Article

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AN - SCOPUS:0038288793

VL - 49

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JO - American Journal of Reproductive Immunology and Microbiology

JF - American Journal of Reproductive Immunology and Microbiology

SN - 8755-8920

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