CD4 memory T cells survive and proliferate but fail to differentiate in the absence of CD40

Secondary T cell responses are enhanced because of an expansion in numbers of antigen-specific (memory) cells. Using major histocompatibility complex class II tetramers we have tracked peptide-specific endogenous (non-T cell receptor transgenic) CD4 memory T cells in normal and in costimulation-deficient mice. CD4 memory T cells were detectable after immunization for more than 200 days, although decay was apparent. Memory cells generated in CD40 knockout mice by immunization with peptide-pulsed wild-type dendritic cells survived in the absence of CD40 and proliferated when boosted with peptide (plus adjuvant) in a CD40-independent fashion. However, differentiation of the memory cells into cytokine-producing effector cells did not occur in the absence of CD40. The data indicate that memory cells can be generated without passing through the effector cell stage.     

早期康复治疗对脑卒中后偏瘫患者短期和长期功能评定的影响

目的:观察脑卒中偏瘫患者早期康复治疗1,3,6个月后功能综合评定量表(functionalcomprehensiveassessment,FCA)评分的变化。方法:42例患者随机分为治疗组(23例)和对照组(19例)。治疗组给予综合康复治疗,包括:①偏瘫医疗体操。②神经肌肉促进技术。③肌电反馈治疗。④日常生活活动(ADL)训练。⑤康复护理。⑥针灸治疗。对照组只给予常规康复指导。治疗前、治疗1,3,6个月后分别进行FCA评定。并将评定数据加以比较。结果:治疗前两组FCA评分比较差异无显著性意义(P>0.05);治疗1个月后治疗组(67.53±16.81)分,优于对照组(59.96±20.97)分,差异有显著性意义(t=1.6648,P<0.05);治疗3个月后,治疗组(85.50±16.52)分,高于对照组(77.84±16.74)分,差异有显著性意义(t=1.7048,P<0.05);治疗6个月后治疗组(94.73±13.54)分,对照组(85.34±12.81)分,差异有显著性义(t=2.3195,P<0.05)结论:早期康复治疗对脑卒中后偏瘫患者短期及长期功能恢复均有明显疗效。     

Leukemia inhibitory factor improves survival of retroviral vector- infected hematopoietic stem cells in vitro, allowing efficient long- term expression of vector-encoded human adenosine deaminase in vivo

Low recovery and poor retroviral vector infection efficiency of hematopoietic stem cells has hindered application of gene therapy for disease affecting blood-forming tissues. Developmental restriction (or death) of stem cells during ex vivo infection has contributed to these difficulties. In these studies we report that the cytokine leukemia inhibitory factor (LIF) directly or indirectly supported the survival of hematopoietic stem cells during culture of bone marrow with vector-producing fibroblasts, resulting in efficient recovery of stem cells able to compete for engraftment in irradiated recipient animals. The infection efficiency of hematopoietic stem cells recovered from these cultures was approximately 80%; and all recipients (20/20) of the LIF-treated marrow were stably engrafted with the progeny of provirus-bearing stem cells. Expression of vector-encoded human adenosine deaminase (hADA) was detected in all recipients at levels averaging 15-50% of endogenous murine ADA in all their hematolymphoid tissues. Survival of stem cells in untreated cultures was approximately 10% of that observed from LIF-treated cultures, resulting in poor engraftment of recipient animals with transplanted cells. The infection efficiency of the few stem cells recovered from untreated cultures, however, was high (approximately 80%), suggesting that LIF did not have an effect on infection efficiency per se, but acted at the level of stem cell survival. Consistent with the poor engraftment observed in the control animals, expression of vector-encoded ADA was only approximately 4-20% of the endogenous levels. These results support the postulated role of LIF as a regulator of hematopoiesis and suggest that cytokine stimulation can positively affect inefficient retroviral vector transduction in hematopoietic stem cells.     

T cells expressing specific V beta elements regulate immunoglobulin E production and airways responsiveness in vivo

The role of T cells expressing specific V beta elements was examined in the regulation of allergen-specific immunoglobulin (Ig)E production and airways responsiveness (AR). In BALB/c mice, inhalation of the allergen ovalbumin (OVA) induced an IgE anti-OVA response, immediate cutaneous reactivity, and increased AR. These results were associated with an expansion of V beta 8.1/8.2 T cells in local draining lymph nodes of the airways and the lung. Transfer of V beta 8.1/8.2 T cells from sensitized mice stimulated an IgE anti-OVA response, immediate cutaneous hypersensitivity, and increased AR in naive syngeneic recipients. In contrast, OVA-reactive V beta 2 T cells inhibited these effects. These data demonstrate for the first time that T cells with different V beta specificities play a critical role in the in vivo regulation of allergen-specific IgE production and AR.     

Interleukin-15 favors the expansion of central memory CD8+ T cells in ex vivo generated, antileukemia human cytotoxic T lymphocyte lines

We demonstrated in previous studies that interleukin (IL) -2 supports in vitro cell proliferation of donor-derived cytotoxic T lymphocyte (CTL) lines directed against different types of leukemia blasts. The aim of this study was to compare the capacity of IL-15 with that of IL-2 in supporting the proliferation and cytotoxic activity of antileukemia CTL cultures, and their influence on T-cell memory compartment differentiation. Antileukemia CTL lines were generated using donor-derived dendritic cells pulsed with apoptotic leukemia blasts, in the presence of IL-12 and IL-7, during the primary culture, and expanded through 2 rounds of leukemia-specific stimulation and 1 round of antigen-independent expansion, each supplemented with either IL-2 or IL-15. Both IL-2-supplemented (IL-2-CTLs) and IL-15-supplemented (IL-15-CTLs) lines contained predominant numbers of CD45RA/CCR7 effector memory (TEM) and CD45RA/CCR7 (TEMRA+) T cells. Significantly higher numbers (P<0.05) of CD8-positive central memory T cells (TCM), and higher expansion rate, together with comparable cytotoxic activity, were observed in IL-15-CTLs compared with IL-2-CTLs. Altogether, these results demonstrate that IL-15 enhances recovery of CTL activity, without loss of leukemia-directed specificity, and favors expansion of TCM CD8-positive cells, expected to exhibit long-term survival and differentiation capacity in vivo in the presence of a limited amount of antigen.     

Monitoring the trafficking of adoptively transferred antigen- specific CD8-positive T cells in vivo, using noninvasive luminescence imaging

Understanding of the trafficking of antigen-specific CD8(+) T cells in vivo will provide insight about how our immune system controls infectious diseases and cancers. In the current study we used a luciferase-expressing human papillomavirus type 16 (HPV-16) E7-specific CD8(+) T cell for adoptive transfer to control E7-expressing TC-1 tumor cells. We used noninvasive luminescence imaging to monitor the trafficking of E7-specific CD8(+) T cells over time. We also boosted the luciferase-expressing E7-specific CD8(+) T cells in vivo, using E7-expressing vaccinia. We found that injected E7-specific T cells preferentially migrated to the E7-expressing tumor site but not to the E7-negative control tumor site, and increased in number at the tumor site over time. In addition, vaccination with E7-expressing vaccinia led to a significant increase in the number of E7-specific CD8(+) T cells at the tumor site, resulting in a significant antitumor effect compared with vaccination with wild-type vaccinia. Thus, our data suggest that the antitumor effects generated by adoptive transfer of E7-specific CD8(+) T cells can be significantly enhanced by vaccination with E7-expressing vaccinia and that our system represents a plausible approach to investigate the trafficking and biology of antigen-specific T cells in vivo.     

Cytotoxic T lymphocytes reactive against a syngeneic murine tumor and their specific suppressor T cells are both elicited by in vitro allosensitization

Sensitization of C57BL/6 (B6, H-2b) splenocytes against normal BALB/c (H-2d) leukocytes (B6 a/BALB) in bulk MLC induced CTL reactive against the syngeneic (H-2b) nonimmunogenic lymphoma PIR-2, in addition to the CTL directed against the corresponding (H-2d) allotargets. However, MLC-derived lymphocytes did not directly exhibit anti-PIR-2 cytotoxicity in spite of the high anti-PIR-2 CTL frequency (up to 1/20) among them, as demonstrated by the limiting dilution culture (LDC) technique. The present study was undertaken to resolve this contradiction. We found that anti-PIR-2 cytotoxicity could be detected only when B6 a/BALB MLC-derived responding cells were plated in LDC at low numbers (less than 200) of cells/well. In contrast, increasing the number of the plated cells to 500-5,000 resulted in a gradual decrease in the percentage of wells cytotoxically reactive against PIR-2, whereas the percentage of wells exhibiting cytotoxicity against the allotargets remained unchanged (100%). This decrease of anti-PIR-2 cytotoxicity in LDC and the lack of anti-PIR-2 reactivity among MLC-derived lymphocytes were shown by mixing experiments to result from the activity of radioresistant Thy-1+, Lyt-2+, L3T4- suppressor cells, blocking the anti-PIR-2 cytotoxicity at the effector phase. The suppression was specific as indicated by the following observations: (a) freshly obtained B6 splenocytes, cultured unsensitized B6 splenocytes, mitogen-induced B6 lymphoblasts, B6 LAK cells, or B6 a/B6 MLC-derived lymphocytes were not suppressive; (b) anti-PIR-2 cytotoxicity elicited in B6 a/BALB LDC was suppressed only by lymphocytes derived from B6 a/BALB MLC and not from B6 a/C3H (H-2k) MLC; and (c) B6 a/BALB MLC-induced suppressor cells could be adsorbed on monolayers of BALB/c but not of C3H lymphoblasts. Since both syngeneic tumor and allogeneic target cells were lysed by the same clonal cell population but only the antisyngeneic activity was suppressed, we suggest that a single CTL can exhibit two cytotoxic activities that are differentially affected by the described suppressor cells. This mode of suppression may play a role in controlling autoimmune reactivity.     

IL-2 and IL-15 each mediate de novo induction of FOXP3 expression in human tumor antigen-specific CD8 T cells

Although FOXP3 is primarily expressed by regulatory CD4 T cells (Treg) in vivo, polyclonal activation of human CD8 T cells can result in the expression of FOXP3 in a fraction of CD8 T cells. However, the cellular lineage and mechanism of FOXP3 induction in CD8 T cells remain unclear. Here, we demonstrate that interleukin-2 (IL-2) induces FOXP3 expression in OKT3-stimulated or antigen-stimulated CD8 T cells, indicating that FOXP3 expression is neither limited to a unique subset of CD8 T cells nor dependent on the mode of T-cell receptor stimulation. In the absence of IL-2, antigen stimulation resulted in T-cell activation and acquisition of effector function without induction of FOXP3, indicating that acquisition of effector function is independent of induction of FOXP3 expression in CD8 T cells. Interestingly, IL-15, but not IL-7 or IL-21, also led to de novo induction of FOXP3 in antigen-specific CD8 T cells, suggesting that signaling by IL-2/IL-15Rbeta chain is pivotal for induction of FOXP3 in human CD8 T cells. These findings indicate that induction of FOXP3 is intrinsic to CD8 T cells that are activated in the presence of IL-2 or IL-15, and in vitro-induced expression of FOXP3 cannot be simply interpreted as an indicator of Treg activity or activation marker.     

Visualization, characterization, and turnover of CD8+ memory T cells in virus-infected hosts

The cellular basis of T cell memory is a controversial issue and progress has been hampered by the inability to induce and to trace long- term memory T cells specific for a defined antigen in vivo. By using the murine model of lymphocytic choriomeningitis virus (LCMV) infection and an adoptive transfer system with CD8+ T cells from transgenic mice expressing an LCMV-specific T cell receptor, a population of authentic memory T cells specific for LCMV was generated and analyzed in vivo. The transgenic T cells that have expanded (1,000-fold) and then decreased (10-fold) in LCMV-infected C57BL/6 recipient mice exhibited the following characteristics: they were (a) of larger average cell size than their naive counterparts but smaller than day 8 effector cells; (b) heterogeneous with respect to expression of cell surface "memory" markers; and (c) directly cytolytic when isolated from recipient spleens. The time-dependent proliferative activity of these LCMV-specific memory T cells was analyzed in the recipients by bromodeoxyuridine labeling experiments in vivo. The experiments revealed that LCMV-specific CD8+ memory T cells can persist in LCMV- immune mice for extended periods of time (>2 mo) in the absence of cell division; the memory population as a whole survived beyond 11 mo.     

B7H costimulates clonal expansion of, and cognate destruction of tumor cells by, CD8(+) T lymphocytes in vivo.

B7H/B7RP (hereby called B7H) is a new member of the B7 family of costimulatory molecules and interacts with inducible costimulatory molecule (ICOS). Its function for CD8 T cells has not been reported. We report here that expression of B7H on the tumor cells reduced tumorigenicity and induced immunity to subsequent challenge with parental tumor cells. The immune protection correlates with an enhanced cytotoxic T lymphocyte (CTL) response against P1A, the major tumor antigen expressed in the J558 tumor. To understand the mechanism of immune protection, we adoptively transferred transgenic T cells specific for tumor antigen P1A into mice that bore P1A-expressing tumors. We found that while the transgenic T cells divided faster in mice bearing the B7H(+) tumors, optimal B7H-induced clonal expansion of P1CTL required costimulation by B7-1 and B7-2 on the endogenous host antigen-presenting cells (APCs). Interestingly, when B7H(+) and B7H(-) tumors were coinjected, P1CTL selectively eliminated the B7H(+) tumor cells. Moreover, B7H expressed on the tumor cells made them highly susceptible to destruction by CTL in vivo, even if the CTL was administrated into mice with large tumor burdens. Tumors that recurred in the P1CTL-treated mice lost transfected B7H and/or H-2L(d), the class I molecule that presents the P1A peptide. Taken together, our results reveal that B7H costimulates clonal expansion of, and cognate destruction by CD8(+) T lymphocytes in vivo.     

Efficacy of cancer gene therapy in aging: adenocarcinoma cells engineered to release IL-2 are rejected but do not induce tumor specific immune memory in old mice

Numerous studies have demonstrated the efficacy of cytokine gene-engineered tumor cells to induce tumor rejection and specific memory acquisition into syngeneic immunocompetent mice by activation of host-dependent antitumor responses. A progressive immune dysfunction, mainly involving thymus-dependent specific immunity, occurs during aging. In this study we evaluated whether the injection of IL-2 gene-transfected tumor cells in old mice causes an immune activation which results in tumor rejection and induction of specific immune memory as occurs in young animals. Young and old mice were inoculated with syngeneic parental mammary adenocarcinoma cells (TS/A p.c.) or with TS/A cells engineered to release IL-2 (TS/A-IL2). Three clones of TS/A-IL-2 cells were used producing low (30 U, B1.30), intermediate (3600 U, B6.3600), or high (6000 U, B4.6000) IL-2. While the B1.30 clone grew in 100% of mice, the B6.3600 and B4.6000 clones were promptly rejected in both young and old animals. In young mice, rejection was associated with a large neutrophil and macrophage infiltration, with a minor number of CD4+ and CD8+ lymphocytes. In old mice, neutrophils and macrophages were the main cells involved in tumor rejection whereas both CD4+ and CD8+ lymphocytes were scarcely present in tumoral infiltrate. A lower number of apoptotic tumor cells was found in TS/A-IL2-challenged old mice in comparison with young animals. To test whether the injection of TS/A-IL2 cells induced a specific immune memory, mice with no tumors after the challenge with B6.3600 and B4.6000 clones received a lethal challenge of TS/A p.c. 90% and 30% of young mice previously injected with B4.6000 or B6.3600 clones, respectively, rejected TS/A p.c. In old mice, B4.6000 cells did not confer protection, whereas only 10% of mice which received B6.3600 cells were able to reject TS/A p.c. Neither the graft of a young thymus or the adoptive transfer of young T lymphocytes to old mice induced specific immune memory for TS/A p.c. in old animals. These data suggest the necessity to refine antitumor vaccination procedures in aging.     

Plasmacytoid dendritic cells induce NK cell-dependent, tumor antigen-specific T cell cross-priming and tumor regression in mice

A prerequisite for strong adaptive antiviral immunity is the robust initial activation of the innate immune system, which is frequently mediated by TLR-activated plasmacytoid DCs (pDCs). Natural antitumor immunity is often comparatively weak, potentially due to the lack of TLR-mediated activation signals within the tumor microenvironment. To assess whether pDCs are capable of directly facilitating effective antitumor immune responses, mice bearing established subcutaneous B16 melanoma tumors were administered TLR9-activated pDCs directly into the tumor. We found that TLR9-activated pDCs induced robust, spontaneous CTL cross-priming against multiple B16 tumor antigens, leading to the regression of both treated tumors and untreated tumors at distant contralateral sites. This T cell cross-priming was mediated by conventional DCs (cDCs) and was completely dependent upon the early recruitment and activation of NK cells at the tumor site. NK cell recruitment was mediated by CCR5 via chemokines secreted by pDCs, and optimal IFN-gamma production by NK cells was mediated by OX40L expressed by pDCs. Our data thus demonstrated that activated pDCs are capable of initiating effective and systemic antitumor immunity through the orchestration of an immune cascade involving the sequential activation of NK cells, cDCs, and CD8(+) T cells.     

Rearrangement and expression of the alpha, beta, and gamma chain T cell receptor genes in human thymic leukemia cells and functional T cells

Using cDNA and genomic probes representing the alpha, beta, and gamma chain of the human T cell receptor genes, we have examined the structure and expression of these genes in 14 human leukemic T cell lines, representing different stages of thymic differentiation, and 15 functional human T cell clones. Rearrangement of the gamma and beta chain genes was found in all of the functional T cell clones and all but one (P30/OKUBO) thymic leukemia cell line; all of the lines that had rearrangement of the beta chain expressed beta mRNA. Expression of the alpha chain was found in all of the functional T cell clones examined, while rearrangement of the alpha chain gene, using currently available probes to the J region, could be shown in 10 of 13 functional clones. In contrast, expression of the alpha chain was found in 6 of 10 leukemic T cell lines, while rearrangement was found in six of these nine cell lines. Of the 14 leukemic cell lines studied for rearrangement of the alpha chain, rearrangement was found in six cases. The data obtained with the cell lines are consistent with an ordered rearrangement and expression of the gamma, beta, and alpha chains of the T cell antigen receptor (TcR) genes. The leukemic cell lines used in the present study have previously been characterized with regard to cell surface antigens and intracellular enzymes. Based on those results a scheme of thymic development was proposed. The developmental stages identified by those studies are not in complete agreement with stages of T cell development, as determined in the present study using molecular probes.     

T lymphocytes specific for immunoglobulin allotype. I. Igh- 1(b)-specific T cells demonstrated by suppression in vivo and cytotoxicity in vitro

We show that determinants of IgG(2a) of C57BL/6 mice (Igh-1(b)) stimulate allotypespecific T cells in BALB/c mice. Such cells are detected in two different functional assays; chronic allotype suppression and T cell-mediated cytotoxicity. A population of suppressor T cells capable of inducing chronic Igh-1(b) suppression was demonstrated by rosetting procedures to possess Igh-1(b)-specific receptors, a result interpreted as indicating that suppressor T cells may act directly upon allotype-bearing B cells. From similar populations we were also able to demonstrate Igh-1(b)-specific cytotoxic T cells. Such cells were lytic for target myeloma cells expressing the Igh-1(b) allotype of IgG28, and were ineffective against a variant cell line failing to express Igh-1(b), and other target cell lines expressing different allotypes or isotypes. The similar specificity of suppressor T cells and cytotoxic T lymphocytes for Igh-1(b) allotype raises the possibility that the target in allotype suppression is a B cell, and that allotype-specific cytotoxic T cells may play some role in regulation of allotype expression in the suppressed state.     

Synthetic peptides as antigens and competitors in recognition by H-2-restricted cytolytic T cells specific for HLA

The specificity of peptide recognition by a number of Kd-restricted CTL clones specific for HLA-CW3 or HLA-A24 was investigated. The CTL clones were derived from DBA/2 (H-2d) mice immunized with syngeneic P815 mouse cells transfected with genes encoding HLA-CW3 or HLA-A24 class I molecules. We had previously shown that CTL clones that lysed P815-CW3 transfectant target cells could lyse P815 (HLA-) target cells incubated with synthetic CW3 peptides corresponding to the COOH-terminal end of the alpha 2 domain. In the present study, we found that Kd-restricted CTL clones that lysed P815-A24 transfectant target cells recognized a synthetic peptide from the same region (residues 170-182) of the A24 molecule. CW3 and A24 differ by only one amino acid within this region. Recognition of CW3 or A24 peptides corresponded exactly with lysis of P815-HLA transfectants both for clones that mutually exclusively lysed CW3 or A24 transfectant target cells and for CW3/A24 crossreactive CTL clones. The latter CTL clones that lysed both CW3 and A24 transfectant target cells showed a clear preference for the peptide corresponding to the immunizing HLA allele. The homologous CW3 and A24 peptides could compete with each other for recognition, in contrast to a peptide from the same region of HLA-B7. Peptides from the corresponding region of the endogenous Kd and Dd/Ld molecules could also inhibit recognition of CW3 and A24 peptides. Competition with peptides apparently occurred at the level of the target cell. These results are consistent with a model whereby MHC class I molecules position protein fragments or peptides for specific recognition by T cells.     

MicroRNA-31 functions as an oncogenic microRNA in mouse and human lung cancer cells by repressing specific tumor suppressors

MicroRNAs (miRNAs) regulate gene expression. It has been suggested that obtaining miRNA expression profiles can improve classification, diagnostic, and prognostic information in oncology. Here, we sought to comprehensively identify the miRNAs that are overexpressed in lung cancer by conducting miRNA microarray expression profiling on normal lung versus adjacent lung cancers from transgenic mice. We found that miR-136, miR-376a, and miR-31 were each prominently overexpressed in murine lung cancers. Real-time RT-PCR and in situ hybridization (ISH) assays confirmed these miRNA expression profiles in paired normal-malignant lung tissues from mice and humans. Engineered knockdown of miR-31, but not other highlighted miRNAs, substantially repressed lung cancer cell growth and tumorigenicity in a dose-dependent manner. Using a bioinformatics approach, we identified miR-31 target mRNAs and independently confirmed them as direct targets in human and mouse lung cancer cell lines. These targets included the tumor-suppressive genes large tumor suppressor 2 (LATS2) and PP2A regulatory subunit B alpha isoform (PPP2R2A), and expression of each was augmented by miR-31 knockdown. Their engineered repression antagonized miR-31–mediated growth inhibition. Notably, miR-31 and these target mRNAs were inversely expressed in mouse and human lung cancers, underscoring their biologic relevance. The clinical relevance of miR-31 expression was further independently and comprehensively validated using an array containing normal and malignant human lung tissues. Together, these findings revealed that miR-31 acts as an oncogenic miRNA (oncomir) in lung cancer by targeting specific tumor suppressors for repression.     

Lineage relationships, homeostasis, and recall capacities of central- and effector-memory CD8 T cells in vivo

The lineage relationships of central-memory T cells (T(CM)) cells and effector-memory T cells (T(EM)), as well as their homeostasis and recall capacities, are still controversial. We investigated these issues in a murine model using two complementary approaches: T cell receptor repertoire analysis and adoptive transfer experiments of purified H-Y-specific T(CM) and T(EM) populations. Repertoire studies showed that approximately two thirds of T(CM) and T(EM) clones derived from a common naive precursor, whereas the other third was distinct. Both approaches highlighted that T(CM) and T(EM) had drastically distinct behaviors in vivo, both in the absence of antigen or upon restimulation. T(CM) clones were stable in the absence of restimulation and mounted a potent and sustained recall response upon secondary challenge, giving rise to both T(CM) and T(EM), although only a fraction of T(CM) generated T(EM). In contrast, T(EM) persisted for only a short time in the absence of antigen and, although a fraction of them were able to express CD62L, they were unable to mount a proliferative response upon secondary challenge in this model.     

Effect of in vivo administration of Lyt antibodies. Lyt phenotype of T cells in lymphoid tissues and blocking of tumor rejection

After transplantation of B6RV2 leukemia, initial tumor growth was followed by tumor regression in B6 (CB6F1) female, but not male, mice. This indicated that H-Y antigen is involved in B6RV2 rejection by syngeneic female recipient mice. In the case of another leukemia, BALB.RL male 1, and Ir gene, probably identical to the Rgv-1 gene, is responsible for RL male 1 rejection. Thus, F1 hybrids of BALB/c with certain other strains of mice can reject RL male 1. Using these two different systems of tumor rejection, we investigated the effects of in vivo administration of Lyt and Thy-1 monoclonal antibodies (mAb). Results showed that Lyt-2 and -3 mAb blocked both B6RV2 rejection by B6 female mice and BALB.RL male 1 rejection by CB6F1 mice. The specificity of blocking was confirmed by use of Lyt-2 and -3 mAb to reciprocal alleles and mice from B6 Lyt-congeneic stocks. No blocking was observed with Lyt-1 and Thy-1 mAb. The Lyt phenotype of T cells in lymphoid tissues from mice treated with mAb was then studied. Blocking of the Lyt-2+3+ population was observed in the lymph node and spleen, but not in the thymus. These results indicate the involvement of Lyt-2+3+ cells (or Lyt-2,3 antigen) in tumor rejection. The precise mechanism of blocking is unknown, but it was observed after even a single injection of Lyt-2,3 mAb on day 9 after tumor transplantation, suggesting that effector cells were functionally blocked, rather than that the generation of these cells was inhibited.