Lung Cancer-Derived Galectin-1 Mediates Dendritic Cell Anergy through Inhibitor of DNA Binding 3/IL-10 Signaling Pathway
Journal of Immunology
Material Supplementary 0.DC1 http://www.jimmunol.org/content/suppl/2010/12/29/jimmunol.100294 Subscription http://jimmunol.org/subscription is online at: The Journal of Immunology Information about subscribing to Permissions Lung cancer, one of the leading causes of death worldwide, is often associated with a state of immune suppression, but the molecular and functional basis remains enigmatic. Evidence is provided in this paper supporting the role of lung cancer-derived soluble lectin,
... ble lectin, galectin-1, as a culprit in dendritic cell (DC) anergy. We have shown that galectin-1 is highly expressed in lung cancer cell lines, together with the serum and surgical samples from lung cancer patients. Functionally, lung cancer-derived galectin-1 has been shown to alter the phenotypes of monocyte-derived DCs (MdDCs) and impair alloreactive T cell response, concomitant with the increase of CD4 + CD25 + FOXP3 + regulatory T cells. The regulatory effect of galectin-1 is mediated, in part, through its ability to induce, in an Id3 (inhibitor of DNA binding 3)-dependent manner, the expression of IL-10 in monocytes and MdDCs. This effect is inhibited by the addition of lactose, which normalizes the phenotypic and functional alterations seen in MdDCs. Of note, significant upregulation of IL-10 was seen in tumor-infiltrating CD11c + DCs in human lung cancer samples. This was also noted in mice transplanted with lung cancer cells, but not in those receiving tumor cells with galectin-1 knockdown. Furthermore, a significant reduction was noted in lung cancer incidence and in the levels of IL-10-expressing, tumor-infiltrating DCs, in mice receiving galectin-1-silenced tumor cells. These results thus suggest that the galectin-1/IL-10 functional axis may be crucial in lung cancer-mediated immune suppression, and that galectin-1 may serve as a target in the development of lung cancer immunotherapy. FIGURE 3. Lung cancer-derived CM and galectin-1 increased the expression of IL-10 in CD14 + monocytes. A, Levels of IL-10 mRNA of CD14 + monocytes. B, rhIL-10 changed the phenotypes of MdDC. C, Effect of lactose on IL-10 upregulation. CD14 + monocytes were treated with RPMI 1640, A549-CM (20%), NCI-H460-CM (20%), and galectin-1 (2 mg/ml) or IL-10 (20 ng/ml) presented in GM-CSF and IL-4 with or without lactose (50 mg/ml), for the indicated times (5 d for surface marker assay, 1 d for lactose blocking assay). mRNA levels and various cell surface markers were measured. D, IL-10 knockdown reversed the alteration of lung cancer-CM and galectin-1 on MdDC phenotype. siRNA transfected CD14 + monocytes were treated with RPMI 1640, lung cancer-CM (20%), and galectin-1 (2 mg/ml) presented in GM-CSF and IL-4 for 5 d. Surface markers were assessed by flow cytometry. Results are representative of at least three independent experiments. Asterisk indicates a significant difference between the two test groups, as analyzed by ANOVA with Student t test post hoc (*p , 0.05; **p , 0.01). The Journal of Immunology 5 by guest on September 11, 2017 http://www.jimmunol.org/ Downloaded from FIGURE 7 . Increased amounts of IL-10 on CD11c + DCs in the tumor section of lung cancer-bearing mice. A, IL-10 high CD11c + DCs infiltrated cancer sections. B, The levels of IL-10 CD11c + DCs isolated from the lungs of lung cancer-bearing and control mice. C, Galectin-1 knockdown decreased the production of IL-10 in CD11c + DCs that infiltrated cancer sections. D, Representative lung sections in galectin-1 wild type and knockdown LLC-bearing mice. LLC, scramble shRNA, and galectin-1 shRNA-transfected LLC were injected into mice via the tail vein. After 14 d, nontumor and tumor regions of the lungs were harvested. CD11c + DCs were isolated from fresh lung or tumor tissue and the culture medium collected after 24-h incubation. IL-10 levels in the culture medium were assessed by ELISA, and IL-10 mRNA was determined by quantitative PCR. Nontumor and tumor regions were also cut and stained, and analyzed by confocal microscopy (310 and 340) (red, IL-10; green, CD11c; blue, DAPI). Data are representative of three independent experiments (control group, n = 8; LLC group, n = 6; scramble siRNA group, n = 6-8; and galectin-1 knockdown group, n = 6-8). Asterisk indicates a significant difference with the control, as analyzed by ANOVA with Student t test post hoc (**p , 0.01).