Immunophenotypic characterization of tumor infiltrating lymphocytes and peripheral blood lymphocytes isolated from melanomatous and non-melanomatous Sinclair miniature swine

Selectively-bred Sinclair miniature swine exhibit a high incidence of congenital malignant melanoma which either proves fatal (10–15% of tumor-bearing piglets) or spontaneously regresses with a biphasic immunological phenomenon (85–90%) and no recurrence of malignancy. Mononuclear leukocytes were isolated from cutaneous melanomas and peripheral blood specimens collected from melanomatous (tumor-bearing) Sinclair swine during second-phase regression, and from peripheral blood specimens collected from non-melanomatous (tumor-free) Sinclair swine and control Hanford swine. Leukocyte identities were determined with single- and dual-parameter indirect immunofluoresence assays via flow cytometry. Assays for the specific surface antigens CD45, CD2, CD4, CD8, CD1, MHC class II, and N1 were employed to develop immunophenotypic profiles within the gated lymphocyte clusters from each TIL and PBL suspension. Significantly more CD8 + T-lymphocytes were identified in TIL suspensions than in peripheral blood leukocyte (PBL) suspensions (P ≤ 0.05), regardless of breed or tumor status. Conversely, PBL suspensions contained significantly higher percentages of CD4 + T-lymphocytes than the levels found in TIL suspensions (P ≤ 0.05). Virtually all TIL were MHC class II +, whereas the percentages of PBL expressing this antigen were markedly lower (P ≤ 0.05). The percentages of T-lymphocytes co-expressing CD4 and CD8, a normal subset unique to swine, were generally consistent in all TIL and PBL suspensions examined. The results of this study have firmly established the immunophenotypic identities of cells associated with the second-phase regression phenomenon of this melanoma and have identified specific variations in the leukocyte profiles of the respective TIL and PBL suspensions.