Expression of cytokine genes, including IL-6, in human malignant melanoma cell lines.
As a preliminary to transducing human melanoma cells with lymphokine genes, we sought for constitutive gene expression and production of eight interleukins, tumour necrosis factors and granulocyte-colony stimulating factor in 19 human melanoma cell lines. Conversion of RNA into cDNA by reverse transcriptase and polymerase chain reaction (RT-PCR) were employed to evaluate gene expression while enzyme-linked immunosorbent assays (ELISA) or biological assays were used to assess the presence of proteins. No expression of interleukins (IL) 3, 4, and 5 or interferon-gamma RNA was found, while the other cytokines were variably expressed in melanoma lines, with IL-1 alpha, IL-1 beta, IL-6, IL-8, being detectable in most of the lines. At protein level, 10 melanoma cells were tested with ELISA and all were found to produce IL-8, five produced IL-6, two tumour necrosis factor (TNF)-alpha, one IL-1 alpha and two TNF beta. The levels of TNF beta were at the limit of test sensitivity. The amount of various cytokines released by the different lines varied widely. Biological assay with the D10-G4 clone confirmed the presence of IL-1 alpha in the supernatant of melanoma (ME) 10221 and revealed an IL-1 activity in the supernatant of Me 4024/1. The proliferating activity of melanoma supernatants on D10-G4 was inhibited by treatment with polyclonal antibodies against IL-1 alpha but not with antibodies against IL-1 beta. TNF biological activity was tested against the TNF-susceptible fibrosarcoma WEHI 164 clone 13.(ABSTRACT TRUNCATED AT 250 WORDS)
Colombo
M P
MP
Division of Experimental Oncology D, Istituto Nazionale Tumori, Milano, Italy.
Maccalli
C
C
Mattei
S
S
Melani
C
C
Radrizzani
M
M
Parmiani
G
G
D001483:Base Sequence; D017077:Culture Media, Conditioned / Q000494:pharmacology; D016207:Cytokines / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005354:Fibrosarcoma; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D008545:Melanoma / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D017077:Culture Media, Conditioned; D016207:Cytokines; D015850:Interleukin-6; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha