Dendritic cells stimulate the expansion of bcr-abl specific CD8+ T cells with cytotoxic activity against leukemic cells from patients with chronic myeloid leukemia

The role of T lymphocytes in the control of chronic myeloid leukemia (CML) after bone marrow transplantations has been clearly shown. This effect closely correlates with graft-versus-host disease (GVHD). A specific graft-versus-leukemia (GVL) effect separate from GVHD has been postulated but has been difficult to show. One possible target for specific GVL activity is the bcr-abl fusion protein characteristic of CML. We have investigated the use of normal peptide-pulsed dendritic cells for the generation of cytotoxic, bcr-abl-specific T cells from normal donors. T cells (CD3+, CD8+, TCR alpha beta+, and NK receptor-negative) generated from a normal donor (HLA A24, B52, B59, Cw1) after stimulation with autologous dendritic cells, primed with a 16 mer peptide spanning the b3a2 breakpoint of bcr-abl, lysed CML cells from the peripheral blood of seven patients with CML with the b3a2 breakpoint. CML cells from four patients with only the b2a2 breakpoint were not lysed. Phytohemagglutinin (PHA) blasts derived from peripheral blood of patients with CML were not lysed, suggesting that cytotoxicity was not due to alloreactivity. Blocking experiments with anti-HLA-A,B,C indicated that cytotoxicity was dependent on recognition of major histocompatibility complex (MHC) class I molecules, although cytotoxicity was not MHC-restricted because not all patients shared HLA types with the T-cell donor. Specificity for bcr-abl and absence of alloreactivity was confirmed by the presence of lytic activity against autologous and allogeneic class I HLA-A matched monocytes pulsed with the 16 mer bcr-abl fusion peptide, but not against unpulsed monocytes or monocytes pulsed with other peptides. These results show that bcr-abl-specific T cells with marked cytotoxic activity against CML cells can be generated and amplified from normal donor peripheral blood. Recognition of HLA molecules is essential for cytotoxicity but strict HLA identity is not required.     

Cellular immune responses of human cadaver donor bone marrow cells and their susceptibility to commonly used immunosuppressive drugs in transplantation

BACKGROUND: The cascade of immunological effects brought about by donor bone marrow cell (DBMC) infusions in human organ transplantation, especially in the context of continuous pharmacologic immunosuppression, is not fully understood. Yet, in inbred rodents and even primates, administration of specific bone marrow cells has caused a state of acquired immunologic tolerance. METHODS: In vitro mixed lymphocyte culture (MLC) and cell-mediated lympholysis (CML) culture systems were used to compare the responding and regulatory properties of DBMC and individual bone marrow cell subsets versus spleen cells in the presence or absence of pharmacologic immunosuppression. RESULTS: In the absence of immunosuppressive drugs, the DBMC proliferated in MLC and in response to phytohemagglutinin, but to a lower magnitude than donor spleen cells. In CML assays, DBMC failed to function as cytotoxic cells. Removal of both CD3+ and CD34+ cells together (not just singly) had to occur for complete abrogation of the proliferative response of DBMC evoked in the presence of allogeneic stimulating cells. Testing several experimental variables using flow cytometric analysis led to the conclusion that when purified DBMC CD34+ cells were placed in coculture with irradiated allogeneic peripheral blood mononuclear cells, such CD34+ cells give rise both to CD3- TCRalphabeta+ as well as to dimly staining CD3+ TCRalphabeta+ cells. Low pharmacologic concentrations of tacrolimus/cyclosporine (CsA) and mycophenolic acid (MPA) singly or in combination had no effect on the spontaneous proliferation of DBMC and had significantly less inhibitory activity on MLC responses of DBMC and its purified CD3+ or CD34+ subpopulations, compared with the responses of spleen cells. Moreover, the previously described regulatory effects of DBMC on the MLC responses of peripheral blood or splenic responding cells were not inhibited by these immunosuppressive drugs. CONCLUSIONS: Taken together, these results support the notion that in vitro DBMC subpopulations, which proliferate as responding cells in co-culture with x-irradiated allogeneic cells and which cause regulatory effects when added as a third component to MLC reactions, seem to be culture-generated lymphoid cell lineage(s) progeny of CD34+ cells. This possibly includes unique CD3+ "primitive" (dimly staining) T cells, which are not as inhibited in their function by tacrolimus/CsA and MPA, as are postthymic (splenic) T cells.     

Generation of in vitro natural cytotoxicity of horse lymphocytes against sarcoid-derived tumor cells not expressing major histocompatibility complex antigens

OBJECTIVE: To analyze in vitro lymphocyte-mediated immune responses of horses with sarcoids against allogeneic sarcoid cells containing endogenous retrovirus but not expressing major histocompatibility complex antigens. DESIGN: Lymphocyte-mediated immune reactions were assessed by means of proliferative responses in mixed lymphocyte tumor cell culture (MLTC) assay and lymphocyte-mediated cytotoxicity against various equine target cells. ANIMALS: 12 horses with sarcoid tumors and 15 control horses. PROCEDURE: Blood lymphocytes were cocultured in MLTC with allogeneic sarcoid cells (Mc-1, BayMc-1), equine testis cells, or normal equine dermal fibroblasts. Lymphocytes were assayed for proliferative responses by [3H]thymidine uptake and for cytotoxicity against the same targets by 51Cr release assay. The lymphocyte populations were analyzed for some common surface markers. RESULTS: Lymphocytes from horses with sarcoids exerted an anamnestic proliferative response in MLTC against Mc-1 cells, but this procedure never generated cytotoxic lymphocytes. However, lymphocytes from all horses cultured in medium with 10% allogeneic serum only had selective. natural cytotoxicity against Mc-1 that was generated without DNA synthesis. Approximately 80% of the lymphocytes disappeared during culture; however the remaining population of small, viable lymphocytes indicated a decrease of CD4+ T lymphocytes, but numbers of T cells with receptors for Helix pomatia A hemagglutinin were unaffected. Few lymphocytes had Fc-receptors for IgG, were complement-reactive positive cells or were B cells expressing surface immunoglobulin. CONCLUSIONS: Results may indicate a natural defense system, which preferentially recognizes and lyses tumor cells that are deficient in surface expression of major histocompatibility complex antigens, without intervention of conventional T-cell receptors or antibodies.     

4-Hydroxynonenal and cell signalling

Donor leukocyte transfusions (DLT) have an anti-leukemic effect in most patients with a relapse of chronic myeloid leukemia (CML) after allogeneic stem cell transplantation. However, DLT are often complicated by graft-versus-host disease. Selection of donor lymphocytes with a relative specificity for leukemic cells is desirable. The generation of leukemia-reactive cytotoxic T lymphocyte (CTL) responses between HLA-identical donors and patients in bulk cultures showed major variations, and false negative results were observed. In a modification of a limiting dilution analysis (LDA) two-fold serial dilutions of HLA-identical donor mononuclear cells (MNC) were cultured in the presence of CML cells. The anti-leukemic CTL precursor frequencies in these donors varied between <1 and 9 per 106 MNC. HLA-restricted CD4+ or CD8+ lymphocytes as well as MHC non-restricted gammadelta T cells were responsible for the anti-leukemic responses. A positive correlation between cytotoxicity in the various wells after 3, 4 and 5 weeks of culture could be found. The LDA may be superior to bulk cultures in selecting stable immune responses and in separating multiple different anti-leukemic T cell responses in each donor-patient combination.     

Urocanic acid enhances IL-10 production in activated CD4+ T cells

The immunosuppressive effects of UV radiation have been well documented. This suppression has been attributed to the action of the cis form of urocanic acid (UCA), a photoproduct of trans-UCA, a natural constituent of the skin. Here, we show that mouse spleen cells preincubated with cis-UCA have a diminished proliferative response to allogeneic cells in MLC and to stimulation with anti-CD3 mAb. Cells preincubated with cis-UCA also had a decreased ability to serve as APC and to stimulate the proliferation of allogeneic lymphocytes in MLC. Simultaneously, the production of IL-2 and IFN-gamma by cells preincubated with cis-UCA was decreased. However, IL-10 gene expression and IL-10 protein secretion by spleen cells stimulated in the presence of cis-UCA were significantly enhanced. The principal cell population displaying the cis-UCA-induced elevated production of IL-10 was CD4+ T cells, which were shown to be a direct target of cis-UCA action. This was also supported by the observation that production of IL-10 by stimulated splenic non-T cells or by macrophages was not altered by cis-UCA. The enhanced production of IL-10 by activated CD4+ T cells may represent a novel pathway of UVB radiation-induced, cis-UCA-mediated immunosuppression. We suggest that the elevated production of IL-10 by activated CD4+ T cells may account for the suppressor T cell phenomena described in UV-irradiated recipients.     

Generation of cytotoxic immune responses during the progression of a rat glioma

Cytotoxic T lymphocytes specific for tumor-associated antigens are produced by exposing animals to tumor cells and stimulating lymphocytes from animals immunized in vitro with tumor cells and small amounts of interleukin-2 (IL-2). This study was designed to determine whether a fast-growing immunogenic avian sarcoma virus-induced glioma produces primed cytotoxic T lymphocyte precursors during its progression. Lymphocytes from intracerebral glioma-bearing rats generally failed to proliferate in vitro in response to immunization with tumor cells and IL-2 and, when proliferative responses were observed, the lymphocytes were not cytotoxic for glioma cells. However, when the same tumor was growing subcutaneously, lymphocytes proliferated and exhibited glioma-specific cytotoxicity when stimulated in vitro with autologous tumor cells and IL-2. Subcutaneous immunization of intracerebral glioma-bearing rats with tumor cells and adjuvant induced strong cytotoxic T lymphocyte responses. The results demonstrated that, while intracerebral tumor progression itself does not induce an anti-glioma immune response, immune responses to tumor-associated antigens may be induced by systemic immunization of tumor-bearing animals. The results suggest that the immunogenicity of brain tumors is masked by the immunologically privileged status of the brain, not by the induction of generalized immune suppression during tumor progression.     

Detection of Borrelia burgdorferi-specific CD8+ cytotoxic T cells in patients with Lyme arthritis

T cells are believed to play an important role in the pathogenesis of Lyme arthritis (LA), an inflammatory joint disease caused by the spirochete Borrelia burgdorferi (Bb). The presence or absence of certain Bb-specific CD4+ T helper cells has been associated with prognosis. Since recent observations suggested the activation of CD8+ T cells during infection with Bb, we searched for CD8+ cytotoxic T lymphocytes in patients with LA. CD8+ T cell lines were generated from peripheral blood and synovial fluid of five patients with LA. In addition, CD8+ T cells were expanded by Ag-specific stimulation in bulk cultures. A cytotoxicity assay was established using target cells infected with recombinant vaccinia viruses expressing the borrelial proteins outer surface protein (Osp) A, OspB, or flagellin. We found Bb-specific CTL lines derived from the peripheral blood of three patients with LA with specificity for flagellin, OspA, and OspB. All Bb-specific CTL lines were CD3+, CD8+, and TCRalphabeta, and cytotoxic activity was HLA class I restricted. Moreover, CD8+ T cells expanded by Ag-specific stimulation in vitro demonstrated Bb-specific and HLA class I-restricted lysis toward individual borrelial proteins. Interestingly, Bb-specific lytic activity was only detected in patient samples obtained after the disappearance of arthritis. We report the detection of Bb-specific cytotoxic CD8+ T cells in patients with LA. The induction of specific CD8+ T cells may play an important role in disease control and may have important bearings for the development of effective vaccines against Lyme borreliosis.     

Teratogenic effects and distribution of cadmium (Cd2+) administered via osmotic minipumps to gravid CF-1 mice

Cytotoxic T lymphocytes (CTL) specific against autologous human cervical cancer cells were generated in vitro from peripheral blood leukocytes (PBL) from four patients with non-keratinized epidermoid carcinoma. For this purpose, these patients' PBL were co-cultured for 28 days either with IL-2 or a mixture of IL-2, IFN-gamma and TNF-alpha in the presence of autologous tumour cells (ATC). Our results showed that these CTL were highly cytotoxic for ATC, weakly cytotoxic for heterologous cervical cancer tumour cells, and not cytotoxic for carcinoma cell lines, normal cervix cells nor autologous PBL. Proliferation and cytotoxicity against ATC were greater when the PBL were activated with the three cytokines. These CTL had a CD4:CD8 ratio of 1:1, were CD16- and CD45RO+ and their killing activity was inhibited by antibodies against CD3, CD8 and MHC-class I but not by antibodies against CD4, CD16 or HLA-class II. The possibility of generating specific CTL in long term cultures for cervical cancer therapy is also discussed.     

HIV-1 variation diminishes CD4 T lymphocyte recognition

Effective long-term antiviral immunity requires specific cytotoxic T lymphocytes and CD4(+) T lymphocyte help. Failure of these helper responses can be a principle cause of viral persistence. We sought evidence that variation in HIV-1 CD4(+) T helper epitopes might contribute to this phenomenon. To determine this, we assayed fresh peripheral blood mononuclear cells from 43 asymptomatic HIV-1(+) patients for proliferative responses to HIV-1 antigens. 12 (28%) showed a positive response, and we went on to map dominant epitopes in two individuals, to p24 Gag restricted by human histocompatibility leukocyte antigen (HLA)-DR1 and to p17 Gag restricted by HLA-DRB52c. Nine naturally occurring variants of the p24 Gag epitope were found in the proviral DNA of the individual in whom this response was detected. All variants bound to HLA-DR1, but three of these peptides failed to stimulate a CD4(+) T lymphocyte line which recognized the index sequence. Antigenic variation was also detected in the p17 Gag epitope; a dominant viral variant present in the patient was well recognized by a specific CD4(+) T lymphocyte line, whereas several natural mutants were not. Importantly, variants detected at both epitopes also failed to stimulate fresh uncultured cells while index peptide stimulated successfully. These results demonstrate that variant antigens arise in HIV-1(+) patients which fail to stimulate the T cell antigen receptor of HLA class II-restricted lymphocytes, although the peptide epitopes are capable of being presented on the cell surface. In HIV-1 infection, naturally occurring HLA class II-restricted altered peptide ligands that fail to stimulate the circulating T lymphocyte repertoire may curtail helper responses at sites where variant viruses predominate.     

Rapid induction of primary human CD4+ and CD8+ T cell responses against cancer-associated MUC1 peptide epitopes

Antigen-specific MHC class II- and class I-restricted helper and cytotoxic T cell responses are important anti-cancer immune responses. MUC1 mucin is a potentially important target for immunotherapy because of its high expression on most human adenocarcinomas. MUC1 peptide-specific type 1 T cell responses were generated in vitro using human peripheral blood lymphocytes (PBL), incubated with liposomes containing synthetic MUC1 lipopeptide antigen. Only two weekly stimulations with the liposomal MUC1 formulation led to the generation of potent anti-MUC1-specific T cell proliferation as well as class I-restricted cytotoxic responses. Thus the use of PBL pulsed with liposome-encapsulated antigen provides an effective approach of rapidly generating effective antigen-presenting cell (APC) function as well as antigen specific T cells in vitro. It may be feasible to use this technology for the rapid and effective generation of APC and/or T cells as cellular vaccines for adenocarcinomas.     

Lack of intrathyroidal tumor necrosis factor alpha in Graves'' disease

Human colonic intraepithelial lymphocytes from control subjects down-regulate the proliferative responses of primed allogeneic peripheral blood mononuclear cells on rechallenge with antigens or phytohaemagglutinin (PHA). In contrast, human colonic intraepithelial lymphocytes from patients with inflammatory bowel disease fail to down-regulate the proliferative responses of primed allogeneic peripheral blood mononuclear cells on rechallenge with antigens. These findings may be important in the development and maintenance of the mucosal immunological activation of inflammatory bowel disease.     

Swedish gynecologists'' and general practitioners'' views on the climacteric period: knowledge, attitudes and management strategies

The cytolytic T lymphocyte (CTL) response has often been used to assess the reconstitution of T cell function after allogeneic or autologous bone marrow transplantation (BMT). Less is known, however, about the reconstitution of the CTL response after peripheral blood stem cell transplantation (PBSCT). Therefore, we investigated the CTL response against Epstein-Barr virus (EBV) of patients undergoing autologous PBSCT. CTLs of six patients with relapsed non-Hodgkin's lymphoma and multiple myeloma were established before and at different times after PBSCT by in vitro stimulation of peripheral blood lymphocytes with autologous EBV-transformed lymphoblastoid cell lines (LCLs). The efficiency of T cell priming by LCLs was assessed at the time of initiation of CTL lines; the proliferative response was strongly reduced during the first 4 months and increased 5 months or more following PBSCT. Cytolytic activity was measured after three or four restimulations of CTLs. All patients investigated had a detectable EBV-specific CTL response which was poor during the first weeks after transplantation, accompanied by a strong non-MHC-restricted cytotoxic activity and a high proportion of CD56-positive T cells. Five or more months after PBSCT, a specific CTL response against EBV was seen which was similar to the situation prior to PBSCT, while the unspecific cytotoxic response decreased. Blocking experiments with monoclonal anti-CD3, anti-CD8 or anti-MHC I antibodies resulted in substantial inhibition of autologous LCL lysis, whereas anti-CD4 or anti-MHC II antibodies had no effect. Finally, autologous PHA blasts of a patient with the HLA haplotype A1/9+, B5/8+, Cw4/7+, were loaded with various EBNA-derived nonapeptides known to be presented by HLA B8 or A11, and exposed to autologous, EBV-directed CTLs. Specific lysis by CTLs only occurred with HLA B8-, but not with HLA A11-restricted nonapeptides. This demonstrated the existence of an MHC I-restricted anti-EBV CTL response after PBSCT. Taken together, the results show that the anlaysis of the EBV-directed CTL activity may serve as a surrogate marker to assess the reconstitution of the cellular immune response in patients undergoing autologous PBSCT.     

Allogeneic and semiallogeneic immunizations with a strong transplantation antigen (Ag-B)

A comparison has been made between the alloantibody response evoked by graded doses of cells being allogeneic and semiallogeneic with the host. The responses were measured by hemagglutination and complement-dependent lymphocytotoxicity. In the strain combination of rats used (DA leads to HO), a marked difference in the optimal dose was found after i.v. injection of lymphocytes. Although semiallogeneic cells gave the better antigenic stimulus at lower cell doses (0.3 and 3 X 10(6) cells), allogeneic cells were better at higher cell doses (30 and 150 X 10(6) cells). Immunization experiments with allogeneic and semiallogeneic erythrocytes or mitomycin- or heat-treated lymphocytes indicated that the antigenicity of the cells was not directly related to the cell surface concentration of antigen and was independent on the proliferative capacity of the lymphocytes; it was, however, curtailed by heat treatment of these cells. The 3 X 10(6), but not the 0.3 or 30 X 10(6) dose of semiallogeneic cells primed efficiently for a secondary hemagglutinin and cytotoxic response. I.v. injected 51Cr-labelled allogeneic and semiallogeneic lymphocytes showed different localized patterns. Between 4 and 24 hr after injection allogeneic lymphocytes were apparently more rapidly lost from the recipient lymph nodes. Collectively, these data indicate that the different immunizing properties of i.v. injected allogeneic and semiallogeneic lymphocytes are not simple consequences of the antigen dose transferred, but suggest that different localization patterns might influence the ability of these cells to induce cytotoxic and hemagglutinating alloantibody formation.     

Tuberculous Addison''s disease. Utility of CT in diagnosis and follow-up

A patient with renal cell cancer developed acute renal failure due to biopsy-proven acute tubulo-interstitial nephritis (AIN) in the 6th week of continuous infusion of 9 x 10(6) IU m-2 day-1 recombinant interleukin-2 (rIL-2). We investigated whether the AIN was the result of a cellular cytotoxic reaction induced by the rIL-2 treatment. The cytolytic activity of cryopreserved peripheral blood lymphocytes (PBL), isolated before and at the end of the rIL-2 treatment (at the time of AIN), was studied after 5 days of culture with or without rIL-2 or anti-CD28 and immobilized anti-CD3 antibodies. The PBL isolated before and at the end of the rIL-2 treatment showed cytolytic activity towards a number of allogeneic targets. However, only the PBL isolated at the end of the rIL-2 treatment showed, when stimulated with rIL-2 in vitro, significant cytolytic activity against an autologous renal cell line cultured from the AIN biopsy specimen and against an allogeneic renal cell cancer cell line. These PBL displayed no enhanced killing capacity towards autologous PBL and the melanoma cell line M14. These observations suggest that the AIN may be the result of a cytotoxic lymphocyte-mediated reaction induced by the rIL-2 treatment.     

Graft-versus-host-disease-associated lymphoid hypoplasia and B cell dysfunction is dependent upon donor T cell-mediated Fas-ligand function, but not perforin function

Allogeneic bone marrow transplant recipients often exhibit a graft-versus-host-disease (GVHD)-associated immune deficiency that can be prolonged and lead to life-threatening infections. We have examined the role of donor T cell-mediated cytotoxic function in the development of GVHD-associated immune deficiency. A major histocompatibility complex-matched model of allogeneic bone marrow transplantation was employed in which lethally irradiated C3H.SW mice received a nonlethal dose of T cells from either perforin-deficient (B6-perforin 0/0), Fas-ligand (FasL)-defective (B6-gld), or normal (B6) allogeneic donor mice. T cell-depleted marrow from B6-Ly-5.1 congenic donor mice was transplanted along with the donor T cell populations to determine the effects of donor T cell-mediated cytotoxicity on engraftment. Our results demonstrate that recipients of perforin-deficient or normal allogeneic T cells exhibit profound lymphoid hypoplasia and severely reduced splenic proliferative responses to lipopolysaccharide in vitro. In contrast, GVHD-associated lymphoid hypoplasia is dramatically reduced and in vitro B cell function is intact in recipients of FasL-defective allogeneic T cells. Engraftment of myeloid and erythroid lineage cells occurs irrespective of donor T cell cytotoxic function. Although recipients of perforin-deficient or normal allogeneic T cells exhibited hematopoietic engraftment exclusively of donor origin, recipients of FasL-defective donor T cells exhibited significant mixed chimerism (Ly-5.1/Ly-5.2). Because only marrow of donor origin was transplanted, this finding suggests that Fas-mediated antirecipient cytotoxicity is required for clearance of residual hematopoietic stem cells of host origin that persist following lethal irradiation.     

Differential expression of natural killer cell markers: human versus baboon

The use of baboons as a model for the study of allo- and xenotransplantation has become increasingly important, but there are few studies on the basic immunological responses in baboons that might be relevant for a rejection reaction. In present study, the cell-surface phenotype, cytokine-induced activation and growth, and cytotoxicity of baboon and human natural killer (NK) and lymphokine-activated killer (LAK) cells were compared. A panel of murine monoclonal antibodies specific for human cell-surface markers expressed on lymphocytes was used to compare relevant baboon and human peripheral blood lymphocytes (PBL). Baboon PBL were 52.1+/-2.9% CD8+, 18.5+/-2.2% CD16+, 3.0+/-0.5% CD25+, and 5.5+/-1.8% CD69+. The corresponding proportions in humans were 23.8+/-7.1%, 12.8+/-3.2%, 4.5+/-1.0%, and 2.3+/-1.1%. In contrast to human PBL, less than 1% of baboon lymphocytes expressed CD56, CD57, and CD122 (interleukin [IL]-2Rbeta). Baboon lymphocytes showed NK cytotoxic activity against the human K562 and CEM cell lines, which was comparable to human NK activity. Depletion of baboon CD16+ or CD8+ cells led to dramatic decreases in NK cytotoxicity, and removal of both subsets completely abrogated NK activity. Incubation of baboon lymphocytes with human recombinant IL-2 for 1 week led to the appearance of CD56+ cells (11.3+/-2.8%). Most of the baboon CD56+ cells induced in culture were in S and G2 phases of cell cycle. Both baboon and human IL-2-activated lymphocytes were highly cytotoxic against the human LAK-sensitive cell line Daudi. Depletion of baboon CD8+ but not CD56+ cells significantly decreased LAK activity. These studies revealed differences in the NK system of humans and baboons that should be taken into consideration when analyzing immune responses to allo- and xenotransplantation in baboons.     

CD94/NKG2 inhibitory receptor complex modulates both anti-viral and anti-tumoral responses of polyclonal phosphoantigen-reactive V gamma 9V delta 2 T lymphocytes

Viral, bacterial, protozoal, and cancer-associated Ags elicit strong responses in human gammadelta T lymphocytes. The majority of these cells in the peripheral blood express the Vgamma9Vdelta2-encoded TCR and recognize nonpeptidic phosphoantigens without an apparent MHC restriction. We have shown that Vgamma9Vdelta2 T cells express the inhibitory CD94/NKG2 receptor for HLA class I molecules. The anti-CD94 mAb inhibits 1) the Vgamma9Vdelta2 T cell proliferation in response mycobacterial phosphoantigens and 2) the HIV-induced Vgamma9Vdelta2 T cell expansion. Vgamma9Vdelta2 T cells stimulated with nonpeptidic mycobacterial antigens produce IFN-gamma and TNF-alpha. Signaling through the CD94/NKG2 receptor interferes with the synthesis of these cytokines. The CD94/HLA class I interaction is also involved in the cytotoxic activity of Vgamma9Vdelta2 T cells. The Vgamma9Vdelta2 T cell regulation through the CD94 receptor may be important for the potentially dual function in innate immunity, i.e., 1) NK-like and 2) TCR ligand-induced cytolytic activities.     

Major differences in response to graded hypoxia between hypoglossal and neocortical neurons

Allogeneic T cells are capable of discriminating between leukemia cells and non-malignant hematopoetic cells. This has been concluded from clinical BMT data and demonstrated by in vitro experiments. We analyzed the frequency and specificity of leukemia-reactive T cells from syngeneic and allogeneic blood donors by limiting dilution assays for interleukin-2-producing (TH1) and cytotoxic (CTLp) T cells. Target cells were leukemia blasts obtained from a patient with common ALL. Control targets were generated by EBV transformation. Effector cells were generated from the peripheral blood of the patient in remission, from his syngeneic brother and from eight healthy, HLA-mismatched volunteers. The effector cells were stimulated with leukemia cells and interleukin-2. Neither the patient nor his brother were able to generate anti-leukemic CTLp or TH1. The HLA-mismatched allogeneic donors displayed anti-leukemic CTLp frequencies with a range from 0 to 68 million. TH1 cells with anti-leukemic specificity were not detectable. We conclude that there are great inter-individual differences in the GVL potential of fully allogeneic peripheral T lymphocytes. It is possible to identify T cell lines with reactivity against leukemia blasts and non-reactivity against normal hematological cells from the same individual. These cell lines are potential effector cells for immunotherapy of human leukemia.     

Enhancing effect of natural killer cell stimulatory factor (NKSF/interleukin-12) on cell-mediated cytotoxicity against tumor-derived and virus-infected cells

Natural killer cell stimulatory factor (NKSF) or interleukin-12 (IL-12) is a heterodimeric cytokine with pleiomorphic effects on T and NK cells, including induction of lymphokine production, mitogenesis, and enhancement of spontaneous cytotoxic activity. Similarly to IL-2, NKSF/IL-12 enhances NK cell-mediated cytotoxicity within a few hours and independently from induced proliferation. This effect is independent from other induced cytokines, because it is not prevented by antibodies neutralizing interferon (IFN)-alpha, IFN-beta, IFN-gamma, IL-2 or tumor necrosis factor (TNF)-alpha and, unlike the induction of IFN-gamma production by peripheral blood lymphocytes, it does not require HLA class II-positive accessory cells. Enhanced cytotoxicity is accompanied by morphologic changes in NK cells, including a significant increase in the number of cytoplasmic granules. In addition to the previously described ability to enhance the cytotoxic activity of NK cells against tumor-derived target cells, NKSF/IL-12 is also a potent stimulator of cytotoxicity against virus-infected cells, either fibroblasts acutely infected with herpes viruses or T cell lines chronically infected with human immunodeficiency virus-1. NK cell-mediated antibody-dependent cytotoxicity or anti-CD16 antibody-redirected lysis is not significantly enhanced by NKSF/IL-12. However, the ability of resting peripheral blood T cells to mediate anti-CD3 antibody-redirected lysis is enhanced by 18-h incubation with NKSF/IL-12, indicating that this lymphokine can modulate the cytotoxic capability of both NK and T cells.