TIM-3 Is Expressed in Melanoma Cells and Is Upregulated in TGF-Beta Stimulated Mast Cells (1)
Though the function of mast cells was mostly known in allergic immune responses and local hypersensitivity, recently their importance has been shown in multiple immunological, inflammatory, developmental, and malignant processes. Interestingly, murine and human mast cells differ in many features such as in their differentiation program, cytokine producing capabilities, or protease content. Although murine mast cells serve as a useful model for several processes, data obtained from these studies cannot be directly transferred to the human system.
Transforming growth factor (TGF)-βI inhibits T-cell proliferation, acquisition of Th cell functions, or differentiation of CD8+ T-cells. TGF-βI also influences macrophages, B-cells, dendritic cells, and contributes to immunosuppression and maintenance of immunological tolerance. Melanomas are also able to constitutively produce TGF-βI and melanoma progression correlates with the overexpression of TGF-β growth factors.
Mucin domain-containing protein (TIM)-3 has been suggested to be a negative regulator of immune responses with limiting the expansion of activated Th1 and T cytotoxic 1 cell populations and was thought to be highly specific for the above-mentioned cells.
Mast cells differentiated for 5 weeks in culture were magnetically separated and then further cultured for an additional 5 days in the presence or absence of TGF-βI. Real-time PCR showed that human mast cells expressed both components (TGF-βRI and TGF-βRII) of the functional receptor for TGF-βI.
Real-time PCR experiments with TaqMan probes fitting to either exon2/3 or exon 6/7 were carried out and results confirmed the upregulation of TIM-3 gene. An antibody against the extracellular domains of TIM-3 found different protein levels between control and TGF-β-treated human mast cells with Western blotting and flow cytometry showed the increased cell surface TIM-3 expression when adding TGF-βI to the culture medium.
An about ∼3-fold order of magnitude higher level of galectin-9, a ligand of TIM-3, was detected in mast cells compared to melanoma cell lines. Furthermore, TGF-βI did not significantly modify galectin-9 expression in mast cells.
Anti-TIM-3 antibody decreased TIM-3 and CD80 expression on murine mast cells during acute myocarditis and reduced Treg populations. The higher TIM-3 level may result in an elevated CD80 expression of human mast cells and CD80 can interact preferentially with the inhibitory molecule CTLA-4 leading to local immunosuppression. human melanoma cell lines WM35 and HT168-M1 were also positive for TIM-3. The presence of TIM-3 on melanoma cells can explain some known effects of galectin-9 in tumors. galectin-9 induces aggregation and reduces the adhesion of breast tumor cells to extracellular matrix proteins.