Abstract
Tumor progression is often influenced by infiltration of myeloid cells; depending on the M1- or M2-like activation status, these cells may have either inhibitory or promoting effects on tumor growth. We investigated the properties of tumor-associated myeloid cells in a previously established homotransplantable amelanotic melanoma (RMM tumor line) in F344 rats. RMM tumor nodules were allowed to reach the sizes of 0.5, 1, 2 and 3 cm, respectively. Immunohistochemistry and flow cytometry was performed for macrophage markers CD68 and CD163, and for the antigen-presenting cell marker, MHC class II. Although no significant change was observed in the number of CD68+ and CD163+ macrophages during RMM progression, the number of MHC class II+ antigen-presenting cells was reduced in 3 cm nodules. Real-time RT-PCR of laser microdissection samples obtained from RMM regions rich in MHC class II+ cells demonstrated high expressions of M1-like factors: IFN-γ, GM-CSF and IL-12a. Furthermore, fluorescence-activated cell sorting, followed by real-time RT-PCR for CD11b+ MHC class II+ (myeloid antigen-presenting cells), CD11b+ CD163+ (M2 type myeloid cells), CD11b+ CD80+ (M1 type myeloid cells) and CD11b+ CD11c+ (dendritic cells) cells was performed. Based on the levels of inflammation- and tumor progression-related factors, MHC class II+ antigen-presenting cells showed polarization towards M1, while CD163+ macrophages, towards M2. CD80+ and CD11c+ myeloid cells did not show clear functional polarization. Our results provide novel information on tumor-associated myeloid cells in amelanotic melanoma, and may become useful in further research on melanoma immunity.
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Abbreviations
- CCL:
-
chemokine (C-C motif) ligand
- CD:
-
cluster of differentiation
- CXCL:
-
chemokine (C-X-C motif) ligand
- FACS:
-
fluorescence-activated cell sorting
- FBS:
-
fetal bovine serum
- Flt:
-
FMS-related tyrosine kinase
- GM-CSF:
-
granulocyte-macrophage colony-stimulating factor
- HE:
-
hematoxylin and eosin
- HIF:
-
hypoxia-inducible factor
- IFN:
-
interferon
- IL:
-
interleukin
- LMD:
-
laser microdissection
- MDSC:
-
myeloid-derived suppressor cell
- MHC:
-
major histocompatibility complex
- MMP:
-
matrix metalloproteinase
- NBF:
-
neutral buffered formalin
- PBS:
-
phosphate buffered saline
- PLP:
-
periodate-lysine-paraformaldehyde
- RT-PCR:
-
reverse transcriptase polymerase chain reaction
- TAM:
-
tumor-associated macrophage
- TGF:
-
transforming growth factor
- TIMP:
-
tissue inhibitor of metalloproteinase
- TNF:
-
tumor necrosis factor
- VEGF:
-
vascular endothelial growth factor
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This work was supported in part by JSPS KAKENHI grant number 22380173 to Yamate.
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Bondoc, A., Golbar, H.M., Pervin, M. et al. Participation of Tumor-Associated Myeloid Cells in Progression of Amelanotic Melanoma (RMM Tumor Line) in F344 Rats, with Particular Reference to MHC Class II- and CD163-Expressing Cells. Cancer Microenvironment 10, 9–24 (2017). https://doi.org/10.1007/s12307-017-0193-x
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DOI: https://doi.org/10.1007/s12307-017-0193-x