Interleukin-8 and monocyte chemotactic protein-1 production by a human glioblastoma cell line, T98G in coculture with monocytes: involvement of monocyte-derived interleukin-1alpha.
We have previously demonstrated that a human glioblastoma cell line, T98G cells, produced high levels of interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) when stimulated with IL-1 or tumor necrosis factor-alpha (TNF-alpha). In this study, we found that T98G cells are capable of producing large amounts of IL-8 and MCP-1 when cocultured with human peripheral blood monocytes or a monocytic cell line, U937 cells. Since it is possible that both glioblastoma cells and monocytes are capable of producing chemokines, we determined which type of cells actually produced IL-8 and MCP-1, by the fixation of one or the other cell type with 3% paraformaldehyde (PA). This procedure revealed that T98G cells were the main source and that PA-treated monocytes effectively stimulated IL-8 and MCP-1 production by T98G cells. Both IL-8 and MCP-1 gene expression and protein production by T98G cells were confirmed by northern blot as well as immunohistochemical staining methods. To analyze the molecules on human monocytes responsible for inducing IL-8 and MCP-1 by T98G cells, several antibodies (Abs) as well as IL-1 receptor antagonist (IL-1Ra) were tested. Anti-IL-1alpha Ab and IL-1Ra almost completely abolished the IL-8/MCP-1-inducing capacity of the PA-fixed monocytes, while no inhibition was obtained with anti-IL-1beta, anti-TNF-alpha or Abs against CD11b/18, L-selectin or ICAM-1, indicating that membrane-associated IL-1alpha is involved in the IL-8/MCP-1 induction, while secreted IL-1alpha plays a major role in this cell-to-cell, i.e., juxtacrine interaction in unfixed conditions.
Kasahara
T
T
Department of Biochemistry, Kyoritsu College of Pharmacy, Tokyo, Japan. kasahara-td@kyoritsu-ph.ac.jp
Oda
T
T
Hatake
K
K
Akiyama
M
M
Mukaida
N
N
Matsushima
K
K
D000911:Antibodies, Monoclonal / Q000494:pharmacology; D018932:Chemokine CCL2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D018920:Coculture Techniques; D015870:Gene Expression / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry; D007375:Interleukin-1 / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D016209:Interleukin-8 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D009000:Monocytes / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism
D000911:Antibodies, Monoclonal; D018932:Chemokine CCL2; D007375:Interleukin-1; D016209:Interleukin-8; D012333:RNA, Messenger