title: Ret mouse very large tumors (VLTs) display altered ratios of infiltrating memory to naive T cells: Roles in tumor expansion.
abstract: Melanoma is an aggressive skin cancer, however it is immunogenic. The size of the primary tumor is associated with the nodal metastases. Our goals were to characterize melanoma-associated antigens (MAAs) and tumor-infiltrating T-lymphocytes (TILs) subsets in the few very large tumors (VLTs) developing in ret transgenic mice of melanoma. Tumors >700mg (VLTs) were investigated for MAAs and subsets of TILs. Immunohistochemistry and flow cytometry-based studies were performed to determine the infiltration patterns of T-lymphocytes in VLTs. It was observed that zinc fixative restores the antigenicity of the cell-surface markers of lymphocyte subpopulations without the need of antigen retrieval, whereas formalin-based fixative fails to restore the antigenicity in the presence of antigen retrieval in the immunohistochemistry. VLTs from ret mice express MAAs, such as Tyrosinase, TRP-1, TRP-2 and gp-100. The mean±standard deviation (S.D.) T-cell infiltration per 400 times-high power field in VLTs; CD4(+) (2.33±1.3), CD8(+) (2.00±1.0), and CD4(+) Foxp3(+) (2.5±0.5) regulatory T cells infiltration was exclusively restricted to the tumor stroma. Moreover, our flow cytometry-based data reveal that % mean±S.D. naive CD3(+) CD4(+) T cell infiltration (32.8±4.0%) was significantly larger than effector (25.8±2.8%, p<0.01) and central memory cells (16.1±3.7%, p<0.001) in VLTs. Similarly, between CD3(+) CD8(+) T cells, naive cells infiltrate (57.7±2.3%) in a significantly larger frequency than effector (5.0±0.4%, p<0.0001) and central memory cell (4.8±1.7%, p<0.0001) subsets. These results suggest that the VLTs from ret mice display lowered infiltration ratios between memory and naive T cells, which could be associated with the relatively large growth of VLTs.
authors.affiliation: Skin Cancer Unit, German Cancer Research Center and University Hospital Mannheim, Heidelberg, Germany; Department of Biology, San Diego State University, San Diego, CA-92182, USA. Electronic address: drmwasim@gmail.com.
authors.forename: Mohammad W
authors.initials: MW
authors.lastname: Khan
authors.affiliation: Skin Cancer Unit, German Cancer Research Center and University Hospital Mannheim, Heidelberg, Germany. Electronic address: v.umansky@dkfz-heidelberg.de.
authors.forename: Viktor
authors.initials: V
authors.lastname: Umansky
keywords: Central memory; Effector memory; Melanoma; Ret mice; TILs; Tregs; Very large tumors