Two Distinct Populations of Primary Cytotoxic Cells Infiltrating into Allografted Tumor Rejection Sites: Infiltration of Macrophages Cytotoxic against Allografted Tumor Precedes That of Multiple Sets of Cytotoxic T Lymphocytes with Distinct Specificity to Alloantigens

Abstract

It has been reported that the rejection of tumor allografts is mainly mediated by cytotoxic T lymphocytes (CTLs). Here, we characterized the cytotoxic effector cells of C57BL/6 (B6; H‐2b) mice infiltrating into the rejection site of the i.p. allografted Meth A fibrosarcoma (or P815 mastocytoma) cells of H‐2d origin. Two types of cytotoxic cells (i.e., CD8+ CTLs and macrophages (Mφs)) were identified by flow cytometric fractionation of the infiltrates or by specific in vitro elimination of cells either with antibody (Ab)‐coated beads or with an Ab‐plus complement. Of particular interest, these effector cells showed distinct and unique target specificities. First, the CTLs were inactive against transplanted tumor (e.g., Meth A) cells, whereas they were cytotoxic against donor‐related concanavalin A (Con A) blasts as well as CTLL‐2 (H‐2b) cells transfected with a class I gene of H‐2d origin. A cold target competition assay suggested that the CTLs were composed of multiple sets of T cells, each of which specifically recognized different allo‐antigens. Second, the Mφs lysed the allografted tumor cells but were inert toward the Con A blasts and the CTLL‐2 transfectants. Unexpectedly, the infiltration of Mφs preceded the infiltration of CTLs by several days during the course of rejection. These results indicate that two distinct populations of unique cytotoxic cells (i.e., CTLs and Mφs) are induced in the allografted tumor rejection site, and that the infiltration of cytotoxic Mφs responsible for rejection precedes that of the CTLs cytotoxic against cells expressing donor‐related allo‐antigens.

A Novel Receptor on Allograft (H‐2d)‐Induced Macrophage (H‐2b) toward an Allogeneic Major Histocompatibility Complex Class I Molecule, H‐2Dd, in Mice

“…We showed earlier that TCRαβ, TCRγδ, CD3, and NK-1.1 antigens were not expressed on AIM (27) and that AIM lacking cytophilic Abs on their cell surface were cytotoxic against allografts with MHC haplotype specificity (21,28). These observations led us to propose that AIM should have a unique surface molecule(s) distinct from TCRαβ, TCRγδ, NK receptor or cytophilic Ab, one that is able to recognize MHC class I antigens on allogeneic cells as nonself.…”

“…The allograft (H-2 d )-induced macrophages (AIM; H-2 b ) resulted in the apoptotic death of allograft (e.g., BALB/c skin and Meth A fibrosarcoma cells), but not H-2 k or H-2 b cells (21)(22)(23)(27)(28)(29), suggesting that the AIM-mediated cytotoxicity was allo-, but not thirdparty-or self-, specific. Surprisingly, however, AIM were inactive toward donor-type Con A blasts (23,27). In contrast, allograft-induced CTLs were found to be highly cytotoxic against donor (H-2 d )-type Con A blasts, mastocytoma (e.g., P815) cells, lymphoid (e.g., P388D1 and L1210) tumor cells, and some fibroblastic (e.g., Balb/3T3 and Colon26) cells.…”

“…AIM are cytotoxic against the CTL-resistant allografts with MHC haplotype specificity; and the infiltration of AIM into the transplantation site precedes that of CD8 ϩ T cells (27), implying recognition of MHC class I molecules on allografts by AIM. To isolate cDNAs encoding receptors on AIM toward the allogeneic MHC class I molecules, we established 4 kinds of monoclonal (m) antibodies (Abs) against macrophages infiltrating into allografts.…”

“…allografted 3-methylcholanthrene-induced ascites-type fibrosarcoma (Meth A) cells (26), a CTL-resistant cell line (17). The allograft (H-2 d )-induced macrophages (AIM; H-2 b ) resulted in the apoptotic death of allograft (e.g., BALB/c skin and Meth A fibrosarcoma cells), but not H-2 k or H-2 b cells (21)(22)(23)(27)(28)(29), suggesting that the AIM-mediated cytotoxicity was allo-, but not thirdparty-or self-, specific. Surprisingly, however, AIM were inactive toward donor-type Con A blasts (23,27).…”

Acute Rejection of Allografted CTL-Susceptible Leukemia Cells from Perforin/Fas Ligand Double-Deficient Mice

“…On days 4 -21 after tumor transplantation, peritoneal cells were harvested by peritoneal lavage with PBS. PEC (10 -20 m in diameter) were separated from the allografted tumor cells (Ͼ20 m in diameter) by FACS, as described previously (9,(15)(16)(17)(18)(19)(20)(21)(22)(23).…”