In an effort to elucidate the role of intrathymic Ia-bearing antigen-presenting cells (APC) on the development of the class II-restricted T cell repertoire, we examined the effect of neonatal ...anti-I-A treatment on both intrathymic and splenic APC function; on the generation of Lyt-2-,L3T4+, Lyt-2+,L3T4-, and Lyt-2+,L3T4+ T cells; and on the development of class I- and class II-specific T cell functions. Both the thymus and the spleen are completely devoid of Lyt-2-,L3T4+ T cells in young mice treated from birth with anti-I-A, and also lack functions associated with this subset, i.e., alloantigen-specific interleukin 2 production (present report), allo-class II-specific and self-class II-restricted T cell proliferative responses, and helper cell function for the generation of cytotoxic T lymphocyte responses (18). Development of the Lyt-2+,L3T4- subset proceeds undisturbed in these mice, in accord with the previously reported normal levels of cytotoxic T lymphocyte precursors (18). The thymus contains normal numbers of the immature cortical Lyt-2+,L3T4+ cells, indicating that acquisition of the L3T4 marker, in and of itself, is not influenced by anti-I-A treatment. This striking absence of the lineage of T cells responsible for class II-specific T cell functions is correlated with absence of thymic APC function for class II-restricted T cell clones. When anti-I-A-treated mice are allowed to recover from the antibody treatment, splenic and thymic APC function return to normal in 2-3 wk, and thymic Lyt-2-,L3T4+ T cell numbers and functions reappear before such cells are detectable in the spleen. Collectively, these findings suggest that development of the Lyt-2-,L3T4+ lineage of class II-specific T cells is entirely dependent on functional I-A-bearing APC cells in the thymus. In addition, the presence of normal levels of Lyt-2+,L3T4-T cells argues that generation of the two major subsets of T cells (i.e., Lyt-2+,L3T4- and Lyt-2-,L3T4+) occurs through separate events, involving unique sites of interactions between precursor T cells and nonlymphoid major histocompatibility complex-bearing thymus cells.
Interactions between self-MHC molecules and T cells are necessary for the proper development of mature T cells, in part due to an absolute requirement for self-MHC-TCR interactions. Recently, we ...showed that CD4-mediated interactions also participate in shaping the T cell repertoire during thymic maturation. We now examine the possible role of the CD8 molecule during in vivo T cell development. Our results demonstrate that perinatal thymi treated with intact anti-CD8 mAb fail to generate CD8 single-positive T cells, while the generation of the other main phenotypes remains unchanged. Most importantly, the use of F(ab')2 anti-CD8 mAb fragments gave identical results, i.e., lack of generation of CD4-/CD8+ cells, with no effect on the generation of CD4+/CD8+. Furthermore, selective blocking of one CD8 allele with F(ab')2 mAbs in F1 mice expressing both CD8 alleles did not interfere with the development of CD4-/CD8+ cells, demonstrating that the absence of CD8+ T cells in homozygous mice is not due to depletion, but rather is caused by a lack of positive selection. This is most likely attributable to a deficient CD8-MHC class I interaction. Our findings strongly advocate that CD8 molecules are vital to the selection process that leads to the development of mature single-positive CD8 T cells.
Severe combined immune deficiency (SCID) mice have a defect in their recombinase system and cannot productively rearrange their immune receptor genes. Thus, SCID thymocytes are arrested at the ...immature "triple negative" phase, not expressing CD3, CD4, or CD8 surface markers. Whole body irradiation of SCID mice induced maturation of their thymocytes to the CD4+/CD8+ double positive, CD3+low stage of differentiation, and resulted in the generation of a thymic cortical region on histologic examination. No mature single positive T cells were detected in the thymus or the periphery. VDJ rearrangements of TCR-beta with restricted clonality were observed in the double positive cells from a given individual. The CD3 complex was expressed on some of these cells, but the cells failed to mobilize intracellular calcium after cross-linking with CD3 Abs. The double positive cells appeared several weeks after irradiation, persisted for many months in the thymus, and by 6 mo generally developed into metastatic lymphoma. Retroviral activation was undetectable in both the preneoplastic and transformed thymocytes. Thus, it appears that the earliest steps in T cell development can be induced in SCID mice by inducing DNA breaks with radiation. This system represents a model of early thymic development, preneoplasia, and neoplasia.
Signaling through the CD28 molecule on T cells by its natural ligand, B7, on APCs has recently been shown to require the presence of an active phosphatidylinositol 3-kinase pathway to mediate some of ...its costimulatory activities (1-7). Using the phosphatidylinositol 3-kinase inhibitor, wortmannin (WN) (8), on human and murine T cells, we have inhibited B7-1-mediated T cell activation and induced Ag-specific tolerance. The addition of WN and/or the B7-1 antagonist, CTLA4Ig, to primary human T cell cultures stimulated with B7-1-transfected allogeneic melanoma cell lines inhibited the generation of alloantigen-specific proliferative and cytolytic responses in vitro. Subsequent examination of these WN- and CTLA4Ig-treated primary T cell cultures revealed that these lymphocyte populations were tolerized to rechallenge with the priming alloantigens in secondary cultures in the absence of additional inhibitor(s). However, reactivity to a third party allogeneic stimulator remained intact. This WN-induced tolerance was reversed by the addition of high dose IL-2, but not IL-4 or IL-7, to the primary cultures, indicating that T cell anergy, not deletion, was responsible for this phenomenon. In vivo studies using a murine graft-vs-host disease (GVHD) model demonstrated that WN treatment of allogeneic donor lymphocytes in vitro failed to generate a significant GVHD in irradiated mouse recipients compared with control allogeneic donor lymphocytes. These findings suggest potentially novel therapeutic strategies for the prevention of GVHD.
One strategy to overcome multidrug resistance in neoplasia is to inhibit the gp170 glycoprotein (relative molecular mass, 170,000) that functions as a plasma membrane, energy-dependent, drug-efflux ...pump. The human colon cancer cell line HT-29, which grows as an ascitic tumor in athymic NCr-nu/nu nude mice, was made multidrug resistant by infection with an MDR1 (also known as PGY1) retrovirus. Referred to as HT-29mdr1, it was used to study reversal of drug resistance in vivo by the anti-P-glycoprotein monoclonal antibody MRK-16. Flow cytometry and radioimmunoassay demonstrated a marked increase in MRK-16 reactivity on HT-29mdr1 cells as compared with its reactivity on the parental, uninfected cell line (HT-29par). The 50% inhibitory concentrations (IC50) of vincristine on HT-29par and HT-29mdr1 cells were 2.5 and 15 ng/mL, respectively. The MRK-16 monoclonal antibody did not affect the vincristine sensitivity of the HT-29par cells. Pretreatment of HT-29mdr1 cells with 10 micrograms/mL MRK-16 in tissue culture partially restored the vincristine sensitivity (IC50 = 7 ng/mL). This modulation of vincristine sensitivity by MRK-16 was then tested in vivo. The median survival times of mice given intraperitoneal transplants of 5 x 10(6) HT-29par or HT-29mdr1 were 37 and 39 days, respectively. Treatment of mice with 1 mg/kg vincristine weekly for 3 weeks, beginning 10 days after tumor injection, resulted in a significant increase in the median survival time of the HT-29par tumor-bearing mice (68 days, P less than .0001), but it had no effect on the HT-29mdr1 tumor-bearing mice. However, treatment of mice bearing the HT-29mdr1 tumor with MRK-16 before vincristine therapy reversed the resistance to the drug (median survival time = 64 days, P less than .0001). The MRK-16 monoclonal antibody alone had no effect on the median survival time of mice given an injection of either HT-29par or HT-29mdr1 cells. These results suggest that strategies employing monoclonal antibody against gp170 may be clinically useful to reverse multidrug resistance.
In the initial series of 198 patients treated at the National Cancer Institute (NCI) with mechlorethamine, vincristine, procarbazine, and prednisone (MOPP) chemotherapy for Hodgkin's disease, a ...review of presenting chest radiographs available on 192 of these patients showed 49 patients with mediastinal masses greater than one third the greatest posteroanterior chest diameter. Five patients had stage IIB disease, and 44 had stage III or IV disease. Thirty-five (71%) patients achieved a complete remission with MOPP chemotherapy. Fourteen (40%) of the complete responders relapsed, but four of these achieved durable remissions in response to subsequent therapy. Thirty (61%) patients have died (14 induction failures, nine relapsed patients, seven complete responders in remission). Thus, with a median follow-up of 20 years (range, 15 to 23), the overall survival for the group is 39%, and the disease-free survival for the complete responders is 60%. A subset of 10 patients received mantle radiation therapy after maximal response to MOPP. One of these patients failed to achieve complete remission, but among the nine complete responders only one has relapsed. In contrast, 13 of 26 (50%) patients achieving a complete response to MOPP alone have relapsed (P2 = .0536). Although MOPP alone was not prospectively compared with MOPP plus radiation therapy in the treatment of advanced-stage massive mediastinal Hodgkin's disease in this series, the retrospective analysis shows a nearly significant difference in disease-free survival favoring combined modality treatment. The difference in tumor mortality between MOPP-treated (44%) and combined modality-treated patients (80%) was also nearly significant (P2 = .055). However, overall survival differences between patients treated with MOPP alone and those treated with combined modality therapy were not significantly different (P2 = 0.23) because of the mortality related to late complications of combined modality treatment.
Recent work has demonstrated the importance of Janus family kinases (JAKs) and signal transducers and activators of transcription (STATs) in the stimulus-response coupling of receptors lacking ...intrinsic tyrosine kinase activity. In particular, the JAK-STAT pathway appears critical in signal transduction by interferon as well as numerous hematopoietic growth factors interacting with members of the hemapoietin receptor superfamily. Although ligands that interact with receptor tyrosine kinases (RTK), such as epidermal growth factor (EGF), platelet-derived growth factor (PDGF) and colony stimulating factor-1 (CSF-1), have been shown to induce increases in phosphorylation of both JAKs and STATs, little is known about activation of this pathway by stem cell factor (SCF). This review will summarize what is known about the JAK/STAT pathway in relation to SCF signal transduction.
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