Colo205 cells were treated with PBS, HPPH-PDT (0

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Colo205 cells were treated with PBS, HPPH-PDT (0.3 J/cm2) or H2O2 (50 M). cytometry using antibodies specific for components of MHC class I molecules and by quantitative PCR using specific primers. Expression of MHC class I-related molecules following HPPH-based PDT (HPPH-PDT) of murine tumors was monitored using a chimeric NKG2D receptor. Results In vitro HPPH-PDT significantly induces MICA in Colo205 cells, but had no effect on MHC class I molecule expression. PDT also induced expression of NKG2D ligands (NKG2DL) following in vivo HPPH-PDT of a murine tumor. Induction of MICA corresponded to increased NK killing of PDT-treated tumor cells. Conclusions PDT induction of MICA on human tumor cells and increased expression of NKG2DL by murine tumors following PDT may play a role in PDT induction of anti-tumor immunity. This conclusion is supported by our results demonstrating that tumor cells have increased sensitivity to NK cell lysis following PDT. 0.01 when compared to cells treated with PBS alone); H2O2-treated target cells were also significantly more sensitive to lysis by NK-92MI cells at ratios of 25:1 and 50:1. These findings correspond to the degree of MICA induction found following treatment with HPPH-PDT and H2O2 (Fig. 6). Open in a separate window Fig. 6 HPPH-PDT enhances NK killing of Colo205 cells. Colo205 cells were treated with PBS, HPPH-PDT (0.3 J/cm2) or H2O2 (50 M). Following a 24-hour incubation cells were used as target cells in a NK cytotoxicity assay as described in Materials and Methods Section. Results are shown as mean-specific lysis SEM. DISCUSSION We show that PDT of human Schisandrin B colon carcinoma cells in vitro and murine colon tumors in vivo leads to increased expression of non-classical MHC class I molecules. These molecules are recognized by the cytotoxicity receptor NKG2D expressed on CD8+ T cells and NK cells. The induction of NKG2DL expression in human Colo205 colon carcinoma cells by PDT appears to be limited to induction of MICA. We were unable to detect induction of either MICB or any of the ULBP family members. In contrast to the induction of MICA by PDT, treatment does not result in increased expression of either classical MHC class I molecules or molecules associated with antigen presentation by MHC class I molecules. Induction of MICA by HPPH-PDT corresponded to an increase in NK killing of the PDT-treated tumor cells. NKG2DL expression can be induced by oxidative stress [23,40]. Interestingly, induction of MICA by PDT was significantly less than that observed following treatment with H2O2. It is possible that the half-life of PDT-induced ROS is too short to allow for gene induction or that PDT induction of MICA is independent of oxidative stress. This possibility is supported by the results showing that treatment with HPPH alone leads to induction of MICA. HPPH localizes to the mitochondrial membrane [41]; mitochondrial damage or stress can lead to NF-B activation and cytokine secretion [42], which may lead to increases in MICA expression [16,23]. The differential expression of Rabbit Polyclonal to IL4 MICA and MICB in response to HPPH-PDT is somewhat surprising given that these two molecules talk about significant homology and so are encoded by evolutionarily conserved genes [43]. Nevertheless, MICA and MICB are extremely many and polymorphic research show they can become differentially controlled [23,40]. Additionally it is obvious that NKG2DL manifestation and rules varies Schisandrin B between tumor cell lines [16]. Essential to our research are the results displaying that transcriptional rules of NKG2DL can be both cell type and environment reliant [16]. Importantly, earlier studies show that hematoporphyrin-based PDT induces ULBP-1 and -2 genes inside a gastric tumor cell range and MICA/B, ULBP-1, -2, and -3 inside a lung tumor cell range [39]. Other research have also analyzed the consequences of PDT on MHC course I manifestation with mixed outcomes. Verteporfin-based PDT result in a downregulation of traditional MHC course I surface manifestation on murine dendritic cells but got no influence on B cell manifestation of these substances [44]. Blom et al. [45] proven that hematoporphyrin-based PDT of human being ocular melanoma cells resulted in rapid adjustments in manifestation that came back to pre-treatment amounts within 6 hours. Sadly, our study will small to clarify these discrepancies additional to further claim that tumor cell manifestation of MHC course I is apparently unaffected by PDT. This conclusion is supported from the scholarly study of Abdel-Hady et al. [12], which Schisandrin B ultimately shows that 5-aminolevulinic acid-based PDT struggles to induce MHC course I manifestation on tumor cells missing MHC.