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Antigen-specific T-cell factors (TCF) play a role in the initiation of cellular immune responses. In allogeneic mouse-tumor models lymphocytes from the direct tumor surroundings of both euthymic and nude mice produce TCF. These lymphocytes produce TCF when collected already 1 day after subcutaneous (sc) injection of tumor cells. In contrast to euthymic mice, draining lymph nodes and spleen of nude mice did not contain TCF-producing lymphocytes at any stage after sc tumor cell injection. In sensitized euthymic mice TCF production by lymphocytes is significantly higher in the direct tumor surroundings than in draining lymph nodes or spleen. At 2 and 5 days after tumor cell injection, the mononuclear cell infiltrate of the tissue surrounding the tumor in euthymic mice showed low expression of Thy 1, CD3, TCR alpha beta, TCR gamma delta, CD4, CD8, and asialo GM1, whereas several lymphocytes and mast cells were positive for monoclonal antibody (mAb) 14-30 (directed against TCF). In both euthymic and nude mice, sc injected tumor cells showed apoptosis. In conclusion, the direct tumor surroundings are the first (and, for nude mice, the only) site of TCF production, sc injection of tumor cells attracts mAb 14-30-positive lymphocytes and renders mast cells positive for mAb 14-30.

Sensitized spleen and peripheral lymph node lymphocytes were tested after different types of immunization with allogeneic tumour cells for their capacity to induce macrophage cytotoxicity in vitro. The macrophages were rendered cytotoxic either by direct contact with lymphocytes and tumour cells (activation of macrophages) or by a factor (macrophage arming factor, MAF), released by the sensitized lymphocytes incubated with tumour cells (arming of macrophages). Both types of reactions are T-cell dependent. Macrophage activation is a more sensitive way to detect lymphocytes with the capacity to render macrophages cytotoxic than arming of macrophages. The route of immunization subcutaneously (s.c.) or intraperitoneally (i.p.) with allogeneic cells did not influence the induction of lymphocytes with the capacity to render macrophages cytotoxic. However, the tumour cells had to be intact as disrupted cells (suspended in Freund's complete adjuvant, FCA) did not induce macrophages activating lymphocytes. The adjuvant dimethyl dioctadecyl ammonium bromide (DDA) did not increase the lymphocyte response. Intact allogeneic tumour cells were needed in vitro when used for secondary antigenic stimulation. This secondary stimulation was independent of antigen presentation by macrophages. This suggests that also in vivo the primary response is independent of macrophage antigen presentation. Delayed-type hypersensitivity and antibody responses against the allogeneic tumour cells were comparable after s.c. and i.p. immunization and after immunization with FCA and DDA.

The phenotype of lymphocytes, obtained from mice immunized with allogeneic tumor cells, with the capacity to induce macrophage cytotoxicity was determined. Macrophage cytotoxicity was induced, either by incubating the macrophages with Macrophage Arming Factor (MAF) containing supernatants of cultures of sensitized lymphocytes and tumor cells (arming) or by incubating the macrophages directly with sensitized lymphocytes and tumor cells (activation). The MAF producing or activating capacity of the lymphocytes was not only "triggered" by the sensitizing tumor cells but also by normal cells and other tumor cells bearing the H-2 determinants of the sensitizing tumor cell. The capacity to render macrophages cytotoxic was not reduced after treatment of the lymphocytes with mitomycin-C or treatment with anti-murine Ig and complement. This capacity of the lymphocytes was abrogated after treatment with anti-T-cell serum or anti-Thy 1.2 serum and complement. After treatment with anti-Lyt 1 or anti-Lyt 2 serum and complement, the activating capacity was significantly reduced and the MAF producing capacity of the lymphocytes abrogated. Mixing the Lyt 1 depleted and Lyt 2 depleted lymphocytes or addition of normal lymphocytes to the Lyt 1 depleted or Lyt 2 depleted populations did not restore the MAF producing and activating capacities. This indicated that the lymphocytes inducing macrophage cytotoxicity in this allogeneic system are Lyt-1+2+ T-lymphocytes, which do not need to divide prior to perform their action.

In a delayed-type hypersensitivity reaction serotonin released from mast cells plays an important role in the induction of a cellular infiltrate at the site of antigen challenge. In analogy, we have studied whether it is possible to enhance the number of intratumoral macrophages by injecting serotonin into a s.c. SL2 lymphosarcoma. The vessels in the tissue surrounding the tumor responded well to serotonin, as there was an influx of i.v. injected 51Cr-labeled sensitized spleen cells in this tissue during the first 4 h after intratumoral injection of serotonin. At 24 h after serotonin injection there was an influx of macrophages into this tumor-surrounding tissue. No influx of cells was detected in the tumor itself during the first hours after injection of serotonin. In the tumor, similar phenomena occurred as in the surrounding tissue, but with a delay of about 24 h. This suggests that lymphocytes leave the blood circulation in the tumor-surrounding tissue and migrate to the tumor. The influx of macrophages into the tumor after intratumoral injection of serotonin is probably due to an immunological reaction as the lymphocyte influx preceeds the macrophage influx into tumors. In addition, transfer of sensitized lymphocytes, as well as lymphocytes from a tumor-bearing host caused an enhanced influx of macrophages into the tumor. To test the specificity and serotonin dependency of the phenomenon of infiltrating cells in tumors we have used a footpad swelling assay in which the serotonin dependency and the antigen specificity of the response against syngeneic tumor cells was shown. The following picture emerged: an intratumoral serotonin injection enables lymphocytes to leave blood vessels in the tumor-surrounding tissue. These lymphocytes with specificity for tumor antigens migrate to the tumor. After contact with the antigenic tumor cells, these lymphocytes secrete chemoattractive factors for monocytes/macrophages. Also these monocytes/macrophages leave the circulation in the tumor-surrounding tissue. Subsequently the macrophages invade the tumor. We conclude that the number of intratumoral macrophages can be enhanced by serotonin.

Summary:An X-linked severe combined immunodeficient (SCID) patient received a nonirradiated erythrocyte transfusion and developed transfusion-associated graft-versus-host disease (TAGVHD), which was controllable with high-dose corticosteroids. Haplo-identical SCT was performed, after a myeloablative conditioning regimen. At day +26, he developed GVHD. Chimerism studies revealed DNA of the erythrocyte transfusion donor (ETD) and recipient only. Because of early nonengraftment and the presence of alloreactive T cells of ETD origin, the patient was treated with an immunosuppressive conditioning regimen followed by a second SCT from the same donor. While tapering immunosuppression, he again developed mild GVHD, and DNA of ETD and bone marrow donor origin were both present. On cyclosporin, the ETD-DNA signal finally disappeared. High-resolution HLA typing revealed haplo-identity between BMD, ETD and the patient, which might have contributed to the relative mild course of the TAGVHD.Bone Marrow Transplantation (2003) 32, 1027-1030. doi:10.1038/sj.bmt.1704266 [ABSTRACT FROM AUTHOR]

A polymerase chain reaction (PCR) was developed, using combinations of an oligonucleotide primer for a T cell receptor Vβ gene family and one for the constant Cβ gene segments, to assess the expression of each of 20 Vβ gene families in RNA after reverse transcription into cDNA. The detection was done after agarose gel electrophoresis of PCR products and ethidium bromide staining. The positive identification of the PCR products was done by hybridization with a Jβ oligonucleotide probe. For T cell lines, a signal was observed in the Vβ2 combination for Jurkat cells. Vβ5a in HSB cells, Vβ2 and Vβ2a in Molt-3 cells and Vβ2. Vβ5a and Vβ2a in Moit-4 cells. Using mixtures of RNA from different cell lines, the sensitivity of the method was in the range of 0.1-0.5%. In peripheral blood mononuclear cells from four donors, taken at three different occasions, all Vβ families were detectable. The intensity of the PCR product varied between various Vβ gene families. Flow cytometric analysis of blood mononuclear cells from the same donors with a restricted series of Vβ gene family-specific antibodies also revealed the presence of all families. The approach to assess Vβ gene family expression in heterogeneous populations opens the possibility to study T cell receptor variable gene expression in relation to physiology and pathologic processes. [ABSTRACT FROM AUTHOR]

In different murine systems, delayed-type hypersensitivity (DTH) swelling responses at 24–48 hr after antigen challenge were preceeded by an early 2-hr swelling response. The 24-hr DTH response is thought to depend on this early (DTH-initiating) hypersensitivity response. In this paper we show that in the syngeneic DBA/2-SL2 routine turnout system only an early 2-hr swelling response can be evoked. This early hypersensitivity response was tumour specific and serotonin dependent. The early hypersensitivity response in contact hypersensitivity has been ascribed to antigen-specific T-cell factors. To test whether similar T-cell factors were involved in the early hypersensitivity response in this syngencic turnout system, we have transferred lymph node, spleen lymphocytes and scram from immunized mice into naive recipients. The serum was fractionated in two fractions, a 50,000–80,000 MW fraction, and a 120,000–190,000 MW fraction. In recipients of lymphocytes, total serum and the 50,000–80,000 MW fraction of the serum, an early hypersensitivity response can be evoked. So, these data suggest the involvement of specific T-cell factors in the development of an early hypersensitivity response against syngeneic tumour cells. Despite the development of an early (DTH initiating) hypersensitivity swelling response these immunized animals cannot develop a classical 24-hr swelling response. This absence of the 24-hr response in the presence of the 2-hr response is discussed in relation to the frequently observed immune suppression in tumour-bearing mice. [ABSTRACT FROM AUTHOR]