mart-1-antigen and Necrosis

Nonocular melanocytic neoplasia is considered uncommon in cats yet is routinely encountered in diagnostic pathology and recognized to exhibit a wide variation in biological behavior. Accurate prediction of clinical outcomes is challenging with no widely recognized prognostic criteria. Signalment and tumor location were retrospectively evaluated in 324 cats diagnosed with nonocular melanocytic neoplasia. Histologic features were described in 141 neoplasms and outcome data were available in 79 cases. Immunohistochemistry using Melan-A, PNL-2, cyclooxygenase 2 (COX-2), and E-cadherin was performed in a subset (

Topics: Animals; Biomarkers, Tumor; Cat Diseases; Cats; Female; Immunohistochemistry; Male; MART-1 Antigen; Melanocytes; Mitosis; Necrosis; Neoplasm Grading; Neoplasms; Prognosis; Retrospective Studies; Sensitivity and Specificity

Dendritic cells (DCs) represent a heterogeneous population of antigen-presenting cells that initiate and orient immune responses in secondary lymphoid organs. In mice, lymphoid organ-resident CD8(+) DCs are specialized at cross-presentation and have developed specific adaptations of their endocytic pathway (high pH, low degradation, and high export to the cytosol). In humans, blood BDCA3(+) DCs were recently shown to be the homologues of mouse CD8(+) DCs. They were also proposed to cross-present antigens more efficiently than other blood DC subsets after in vitro activation, suggesting that in humans cross-presentation is restricted to certain DC subsets. The DCs that cross-present antigen physiologically, however, are the ones present in lymphoid organs. Here, we show that freshly isolated tonsil-resident BDCA1(+) DCs, BDCA3(+) DCs, and pDCs all cross-present soluble antigen efficiently, as compared to macrophages, in the absence of activation. In addition, BDCA1(+) and BDCA3(+) DCs display similar phagosomal pH and similar production of reactive oxygen species in their phagosomes. All three DC subsets, in contrast to macrophages, also efficiently export internalized proteins to the cytosol. We conclude that all freshly isolated lymphoid organ-resident human DCs, but not macrophages, display high intrinsic cross-presentation capacity.

Topics: Animals; Antigen Presentation; Antigens, CD1; Cell Separation; Cross-Priming; Cytosol; Dendritic Cells; Humans; Hydrogen-Ion Concentration; Ligands; Lymphoid Tissue; MART-1 Antigen; Mice; Necrosis; Palatine Tonsil; Phagocytes; Phagosomes; Reactive Oxygen Species; Solubility; Toll-Like Receptors

We evaluated 35 cases of malignant melanomas with substantial necrosis immunostained with S-100, HMB-45, Melan-A, tyrosinase, PNL2, and microphthalmia transcription factor (MITF). Staining patterns were evaluated in viable and necrotic areas of the tumors. S-100 was the most sensitive marker (97%) in the viable tumors, but necrotic areas demonstrated nonspecific staining. Viable tumors stained variably for HMB-45 (25 [71%]), Melan-A (28 [80%]), tyrosinase (30 [86%]), and PNL2 (23 [66%]). Necrotic areas focally reacted to the same antibodies. The necrotic areas that retained immunoreactivity for these markers corresponded to areas where the outline of the tumor cells could still be recognized as ghost cells on the H&E-stained section. Areas that showed complete coagulative necrosis were negative for melanoma markers. MITF variably stained in the viable tumors but was completely negative in necrotic areas. Our study demonstrated that a combination of antibodies to HMB-45, tyrosinase, and PNL2 detected melanocytic differentiation in necrotic areas in 80% of cases.

Topics: Antibodies, Monoclonal; Antigens, Neoplasm; Humans; Immunohistochemistry; MART-1 Antigen; Melanoma; Melanoma-Specific Antigens; Microphthalmia-Associated Transcription Factor; Monophenol Monooxygenase; Necrosis; Neoplasm Proteins; S100 Proteins

In the present study, we demonstrate, in rigorous fashion, that human monocyte-derived immature dendritic cells (DCs) can efficiently cross-present tumor-associated antigens when co-cultured with a mixture of human melanoma cells rendered apoptotic/necrotic by gamma irradiation (Apo-Nec cells).. We evaluated the phagocytosis of Apo-Nec cells by FACS after PKH26 and PKH67 staining of DCs and Apo-Nec cells at different times of coculture. The kinetics of the process was also followed by electron microscopy. DCs maturation was also studied monitoring the expression of specific markers, migration towards specific chemokines and the ability to cross-present in vitro the native melanoma-associated Ags MelanA/MART-1 and gp100.. Apo-Nec cells were efficiently phagocytosed by immature DCs (iDC) (55 +/- 10.5%) at 12 hs of coculture. By 12-24 hs we observed digested Apo-Nec cells inside DCs and large empty vacuoles as part of the cellular processing. Loading with Apo-Nec cells induced DCs maturation to levels achieved using LPS treatment, as measured by: i) the decrease in FITC-Dextran uptake (iDC: 81 +/- 5%; DC/Apo-Nec 33 +/- 12%); ii) the cell surface up-regulation of CD80, CD86, CD83, CCR7, CD40, HLA-I and HLA-II and iii) an increased in vitro migration towards MIP-3beta. DC/Apo-Nec isolated from HLA-A*0201 donors were able to induce >600 pg/ml IFN-gamma secretion of CTL clones specific for MelanA/MART-1 and gp100 Ags after 6 hs and up to 48 hs of coculture, demonstrating efficient cross-presentation of the native Ags. Intracellular IL-12 was detected in DC/Apo-Nec 24 hs post-coculture while IL-10 did not change.. We conclude that the use of a mixture of four apoptotic/necrotic melanoma cell lines is a suitable source of native melanoma Ags that provides maturation signals for DCs, increases migration to MIP-3beta and allows Ag cross-presentation. This strategy could be exploited for vaccination of melanoma patients.

Topics: Antigens, Neoplasm; Apoptosis; CD8-Positive T-Lymphocytes; Cell Differentiation; Cell Line, Tumor; Cell Movement; Chemokine CCL19; Coculture Techniques; Cross-Priming; Dendritic Cells; Epitopes; Gamma Rays; gp100 Melanoma Antigen; Humans; Interleukin-10; Interleukin-12; MART-1 Antigen; Melanoma; Membrane Glycoproteins; Monocytes; Necrosis; Neoplasm Proteins; Phagocytosis; Receptors, CCR7; Time Factors