Fas expression and Fas monoclonal antibody-induced apoptosis in a human squamous cell carcinoma cell line, SCC-25.
Fas antigen is a cell surface protein that mediates apoptosis via signal transduction from the plasma membrane. Using reverse transcriptase-polymerase chain reaction (RT-PCR), messenger RNA for Fas antigen was detected in the human oral squamous cell carcinoma cell line, SCC-25. In serum-free medium, a monoclonal anti-Fas antibody (CH-11) induced Fas antigen expression in SCC-25 cells, as determined by immunocytochemistry and Western blotting, using an anti-Fas polyclonal antibody (Fas D) as primary antibody. Fas antigen was localized to the cytoplasm and the cell membrane. The molecular weight of the protein recognized by Western blot analysis was 35,000, consistent with the value reported for the Fas antigen. The CH-11 antibody did not induce Fas antigen expression in serum-containing medium. To determine whether CH-11 could induce apoptosis in oral squamous cell carcinoma, we examined its effects on the survival of cultured SCC-25 cells. Anti-Fas monoclonal antibody in serum-free medium induced cytotoxicity in SCC-25 cells in a time-dependent manner up to 8 h, as determined by phase-contrast microscopy and WST-1 assay. Marked nuclear condensation and fragmentation of chromatin were observed in the CH-11-treated cells using Hoechst 33342 staining. This anti-Fas monoclonal antibody also induced DNA ladder formation in SCC-25 cells in a time-dependent manner. The present results indicate that the anti-Fas monoclonal antibody (CH-11) may mediate apoptosis by binding to the Fas antigen expressed in SCC-25 cells.
Seta
C
C
First Department of Oral and Maxillofacial Surgery, Kyushu Dental College, Kitakyushu, Japan.
Fujita
M
M
Muraki
Y
Y
Fukuda
J
J
Kobayashi
S
S
Haneji
T
T
D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology*; D017209:Apoptosis / Q000276:immunology*; D002294:Carcinoma, Squamous Cell / Q000276:immunology* / Q000648:ultrastructure; D002462:Cell Membrane / Q000276:immunology; D002467:Cell Nucleus / Q000648:ultrastructure; D002470:Cell Survival; D002843:Chromatin / Q000648:ultrastructure; D016895:Culture Media, Serum-Free; D003593:Cytoplasm / Q000276:immunology; D003602:Cytotoxicity, Immunologic; D053938:DNA Fragmentation; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008970:Molecular Weight; D009062:Mouth Neoplasms / Q000276:immunology* / Q000648:ultrastructure; D015398:Signal Transduction / Q000276:immunology; D013997:Time Factors; D014407:Tumor Cells, Cultured; D019014:fas Receptor / Q000235:genetics / Q000276:immunology*
D000911:Antibodies, Monoclonal; D002843:Chromatin; D016895:Culture Media, Serum-Free; D019014:fas Receptor