RESUMO
A 6-y retrospective case note review was performed to determine the causes of ketoacidosis. 135 patients and 463 diabetic years were involved. Fifty-two ketoacidosis episodes occurred: 19 episodes in new patients and 33 episodes in 19 patients with established diabetes. 27% of newly diagnosed patients presented in ketoacidosis. They were similar in terms of age, sex and proportion living in single parent families to those presenting without ketoacidosis. The 33 ketoacidosis episodes occurring in established patients included 12 episodes in 3 children who were transferred to our care because of uncontrolled diabetes. Insulin omission was the cause of ketoacidosis in 9/19 (47%) patients, and was suspected in a further 5/19 (26%). Family and school problems were common and 14/19 patients came from single parent families. Established patients aged > or = 11 y were predominantly female (10F, 2M), whereas patients aged < or = 10 y were predominantly male (6M, 1F). 7 patients with multiple ketoacidosis episodes were all > or = 11 y and 6 were female. Families with > or = 2 diabetic children appeared vulnerable, 4 cases coming from 3/7 such families.
Assuntos
Cetoacidose Diabética/epidemiologia , Adolescente , Criança , Pré-Escolar , Características da Família , Feminino , Humanos , Lactente , Cetose/epidemiologia , Masculino , Estudos RetrospectivosRESUMO
Tumor necrosis factor alpha (TNFalpha) is a cytokine implicated in the pathogenesis of numerous chronic and acute inflammatory conditions. In the present study, we have characterized the ability of TNFalpha in inducing eosinophil accumulation in rat skin and have shown the inhibitory effects of anti-alpha4 integrin and anti-vascular cell adhesion molecule-1 (VCAM-1) antibodies on this response. The intradermal injection of recombinant human TNFalpha induced a slowly developing, dose-dependent accumulation of 111In-eosinophils in rat skin that was maximal at the dose of 10(-11) mol/site. Coadministration of TNFalpha with the soluble TNFalpha receptor (p55)-IgG fusion protein (TNFR-IgG) totally inhibited the 111In-eosinophil accumulation induced by the cytokine. The TNFalpha-induced 111In-eosinophil accumulation was not affected after pretreatment of rats with the platelet-activating factor (PAF) receptor antagonist UK-74,505 or the antihuman interleukin-8 monoclonal antibody (MoAb) DM/C7. In contrast, the intravenous administration of an anti-alpha4 integrin MoAb, HP2/1 (3.5 mg/kg), or an anti-VCAM-1 MoAb, 5F10 (2 mg/kg), greatly inhibited the 111In-eosinophil accumulation induced by TNFalpha (the responses detected at 10(-11) mol/site were inhibited by 78% and 50%, respectively). These results show that TNFalpha is an effective inducer of eosinophil accumulation in vivo, with this response being dependent on an interaction between alpha4 integrins and VCAM-1.
Assuntos
Antígenos CD/imunologia , Eosinófilos/imunologia , Transdução de Sinais/imunologia , Pele/imunologia , Fator de Necrose Tumoral alfa/farmacologia , Molécula 1 de Adesão de Célula Vascular/imunologia , Animais , Eosinófilos/efeitos dos fármacos , Eosinófilos/patologia , Humanos , Integrina alfa4 , Ratos , Ratos Sprague-Dawley , Receptores de Retorno de Linfócitos/imunologia , Pele/efeitos dos fármacos , Pele/patologiaRESUMO
The interaction of T cell costimulatory molecules with their ligands is required for optimal T cell activation. Interference with such interactions can induce antigen unresponsiveness and delay xeno- and allograft rejection. We have previously shown that LFA3TIP, a soluble human lymphocyte function-associated antigen (LFA)-3 construct, binds CD2 and inhibits responses of human T cells in vitro. This study reports the first use of a human fusion protein, LFA3TIP, to significantly prolong primate cardiac allograft survival. Based on our observations that LFA3TIP inhibits baboon allogeneic mixed lymphocyte reactions, we gave baboon recipients of heterotopic cardiac allografts injections of LFA3TIP, 3 mg/kg i.v., for 12 consecutive days, starting 2 days before transplantation. This regimen delayed graft rejection from an average of 10.6 +/- 2.3 days for human IgG-treated controls (n = 5) to an average of 18.0 +/- 5.3 days for LFA3TIP-injected animals (n = 7; P < or = 0.01). Grafts from LFA3TIP-treated animals showed markedly diminished coronary endothelialitis as compared with control animals. LFA3TIP reached peak serum levels of approximately 100 micrograms/ml after 7-9 injections and persisted in the 10-micrograms/ml range for 1 to 2 weeks after the final injection. Despite these blood levels, circulating antibodies to LFA3TIP were not detected in the serum. No renal or hepatic toxicity was noted. The possible mechanism by which LFA3TIP acts to inhibit graft rejection is discussed; success in prolonging graft survival when LFA3TIP is used as a single-agent therapy suggests great potential for this novel therapeutic agent.
Assuntos
Antígenos CD58/uso terapêutico , Transplante de Coração/imunologia , Imunoglobulina G/uso terapêutico , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/sangue , Anticorpos Monoclonais/uso terapêutico , Antígenos CD2/metabolismo , Antígenos CD58/administração & dosagem , Antígenos CD58/sangue , Rejeição de Enxerto/prevenção & controle , Sobrevivência de Enxerto , Transplante de Coração/efeitos adversos , Transplante de Coração/patologia , Humanos , Imunoglobulina G/administração & dosagem , Imunoglobulina G/sangue , Imunoterapia , Técnicas In Vitro , Teste de Cultura Mista de Linfócitos , Papio , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/sangue , Proteínas Recombinantes de Fusão/uso terapêutico , Fatores de Tempo , Transplante Heterotópico , Transplante HomólogoRESUMO
LFA3TIP, a fusion protein comprised of the first extracellular domain of LFA-3 fused to the hinge, CH2 and CH3 domains of human IgG1, inhibits proliferation of human T cells in vitro. LFA3TIP also inhibits responses of human CD2 transgenic mice by rapidly and totally depleting peripheral T cells. These effects require binding of the LFA-3 and CH2 domains of LFA3TIP to CD2+ T cells and Fc gamma R+ accessory cells, respectively. As CD2 is well conserved in primate species, we evaluated the effects of LFA3TIP in nonhuman primates. We report in vitro results leading to the selection of the baboon as a model for analysis of LFA3TIP, and in vivo effects of single and multidose regimens of LFA3TIP administration. This is the first report of the in vivo administration of an immunomodulatory fusion protein to primates. LFA3TIP is shown to mediate effects on primate T lymphocytes without apparent related toxicities or immunogenicity. Results are discussed in context of potential mechanisms of LFA3TIP immunotherapy.
Assuntos
Adjuvantes Imunológicos/química , Antígenos CD58/administração & dosagem , Proteínas Recombinantes de Fusão/farmacologia , Animais , Antígenos CD2/metabolismo , Antígenos CD58/química , Relação Dose-Resposta a Droga , Feminino , Imunoglobulina G/química , Imunossupressores , Contagem de Leucócitos/efeitos dos fármacos , Ativação Linfocitária , Depleção Linfocítica , Macaca fascicularis , Macaca mulatta , Papio , Ligação Proteica , Proteínas Recombinantes de Fusão/farmacocinéticaRESUMO
Lymphocyte-endothelial cell recognition is an active multistep process central to the pathophysiology of inflammation. In vitro models of lymphocyte adhesion predict that the beta 1 integrin very late antigen-4 (VLA-4), an activation-dependent adhesion receptor, can mediate the firm sustained attachment required for the extravasation of memory lymphocytes. We have used murine contact hypersensitivity as an in vivo model in which to evaluate the role of alpha-4 integrins in an evolving inflammatory response. We demonstrate that the intravenous administration of 75 micrograms of the anti-alpha-4 specific monoclonal antibodies R1-2 or PS/2 4-6 h prior to challenge significantly inhibits the efferent response of 2,4 dinitrofluorobenzene, or oxazolone-sensitized mice. The disease-modifying effect of anti-alpha 4 treatment was evident as a 50-60% reduction in the ear swelling response. By histological analysis treated animals scored lower for edema, number of epidermal lesions and degree of leukocyte infiltration. Antibody-treated animals have elevated numbers of circulating mononuclear leukocytes present in the same relative ratio as untreated control animals, suggesting that the inhibitory effect was not due to antibody-dependent cellular depletion of effector lymphocytes. These data are consistent with a central role for VLA-4 in the pathophysiologic process of inflammation.
Assuntos
Dermatite de Contato/imunologia , Imunidade Celular , Receptores de Antígeno muito Tardio/imunologia , Animais , Anticorpos Monoclonais/imunologia , Dermatite de Contato/patologia , Feminino , Haptenos , Hipersensibilidade Tardia/imunologia , Hipersensibilidade Tardia/patologia , Camundongos , Camundongos Endogâmicos BALB C , Fatores de TempoRESUMO
HIV use the CD4 molecule as their primary cellular receptor. Residues in the N-terminal domain (D1) of CD4 are crucial to HIV attachment through the gp120 envelope component. However, other regions of CD4 appear to be required subsequently for virus- and cell-cell fusion. Little is understood of the post-binding steps which may differ between HIV variants. We report a novel anti-CD4 mAb that does not block CD4/gp120 binding, but that does efficiently block both viral infection and cell-cell syncytia formation, and define its contact site as residues in CD4 D2 using both mouse/human CD4 chimeras and CD4 substitution mutants. We also investigated the basis for its antiviral effect. Using the CD4 D2 specific mAb, we identify another conserved step in HIV infection, as evidenced by its ability to neutralize a broad range of primary isolates and T cell-line passaged strains. Monovalent forms of the mAb were used to determine if its activity was due to masking of the D2 epitope, to steric inhibition, or bivalency. Our data indicate that both binding site and bivalency of the mAb underlie its potency. The need for bivalency is not simply explained by affinity, because monovalent forms can displace the intact mAb and reverse its protective effect. These results provide evidence that binding of the D2-specific mAb prevents structural alterations necessary for membrane fusion.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos CD4/fisiologia , Infecções por HIV/prevenção & controle , Animais , Fusão Celular , Linhagem Celular , Epitopos/análise , HIV/patogenicidade , Proteína gp120 do Envelope de HIV/metabolismo , Fragmentos Fab das Imunoglobulinas/imunologia , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Vascular cell adhesion molecule-1 (VCAM1) is a member of the immunoglobulin (Ig) superfamily which interacts with the integrin very late antigen 4 (VLA4). We have cloned the cDNAs for both murine and rat VCAM1 from endotoxin-treated lung libraries. Both sequences encode proteins with seven extracellular Ig-like domains, which show 75.9% and 76.9% identity, respectively, with human VCAM1. Both murine and human cell lines show VLA4-dependent binding to COS cells transiently expressing murine and rat VCAM1. Two mAbs, M-K/1 and M-K/2, which recognize an antigen on murine bone marrow stromal cell lines, bind to murine VCAM1 expressed in COS cells and block VCAM1-dependent adhesion, confirming that these mAbs recognize murine VCAM1.
Assuntos
Moléculas de Adesão Celular/genética , Regulação da Expressão Gênica , Sequência de Aminoácidos , Animais , Adesão Celular , Linhagem Celular , Clonagem Molecular , Biblioteca Gênica , Inflamação/metabolismo , Pulmão/fisiologia , Camundongos , Dados de Sequência Molecular , Ratos , Homologia de Sequência do Ácido Nucleico , Transfecção , Molécula 1 de Adesão de Célula VascularRESUMO
In a prospective, randomized trial in 100 febrile neutropenic children, aztreonam plus flucloxacillin was compared with piperacillin plus gentamicin. At the 72 h clinical assessment there was no statistically significant difference between the two groups. However, in microbiologically documented infections there was a higher response rate in the piperacillin/gentamicin group (57%) than in the aztreonam/flucoxacillin group (41%). This was contributed to by the poorer Gram-positive cover of the aztreonam/flucloxacillin combination. In clinically documented infections and unexplained fevers the response rate of the two antibiotic regimens was identical. There were two deaths; one early death (in the piperacillin/gentamicin arm) and one late death. At the final assessment a successful outcome was obtained in the remaining patients. In the aztreonam/flucloxacillin group 75% of the episodes required modification compared with 59% in the piperacillin/gentamicin group.
Assuntos
Aztreonam/uso terapêutico , Febre/tratamento farmacológico , Floxacilina/uso terapêutico , Neutropenia/complicações , Adolescente , Bacteriemia/complicações , Bacteriemia/tratamento farmacológico , Criança , Pré-Escolar , Quimioterapia Combinada , Feminino , Febre/complicações , Gentamicinas/uso terapêutico , Bactérias Gram-Negativas/efeitos dos fármacos , Humanos , Lactente , Masculino , Neutropenia/tratamento farmacológico , Piperacilina/uso terapêutico , Estudos ProspectivosRESUMO
Cyclosporine (CS) is a relatively new immunosuppressive agent which is selective in its action in that it acts on lymphocytes, T and B cells, and only indirectly if at all on other haemopoietic cells. Numerous studies over the last decade have shown quite clearly that, in vitro, CS inhibits lymphocyte activation at a relatively early stage by preventing the production and release of lymphokines, probably by inhibition of lymphokine gene transcription. However, studies in vivo have produced clearly different findings--although CS in vivo is completely effective in preventing the development of mature effector function (i.e., T-dependent antibody synthesis by B cells or T cell-mediated cytotoxicity) and the consequences of that effector function (e.g., transplant rejection), there is clear evidence that lymphocytes can be activated to at least the stages of cell division and clonal expansion under cover of therapeutic levels of CS. The contradictions between in vitro and in vivo findings are discussed in terms of the possible mechanism of action of CS in vivo.
Assuntos
Ciclosporinas/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Linfócitos T/imunologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Reação Enxerto-Hospedeiro/efeitos dos fármacos , Reação Hospedeiro-Enxerto/efeitos dos fármacos , Humanos , Linfocinas/antagonistas & inibidores , Transplante de ÓrgãosRESUMO
An indium 111-labelled mouse anti-rat T cell monoclonal antibody, MRC OX-19, was injected intravenously into rats to establish the usefulness of radiolabelled anti-lymphocyte antibodies in imaging lymphoid tissues. Antibody binding in vivo, measured by immunofluorescence analysis of cell suspensions made from lymphoid tissues, was detectable on lymphocytes in blood, spleen and lymph nodes. The extent of binding was time and antibody-dose dependent. Doses of antibody above 80 micrograms/kg body weight resulted in modulation, i.e. loss of CD 5 (T1) molecules from the cell surface, although the cells remained in the circulation. Modulation was demonstrable within 2 h and for at least 24 h after a single injection of antibody. Intravenous injection of 111In-MRC OX-19 resulted in levels of in vivo binding comparable with those seen with unlabelled antibody. Scintillation imaging showed early splenic localisation persisting over 48 h, a more gradual localisation in the lymph nodes seen clearly at 24 h and a steady background. Comparison of the in vivo distribution of labelled antibody and 111In-tropolone-labelled lymphocytes showed that both could be used for external imaging of lymphocytes by scintillation camera.
Assuntos
Anticorpos Monoclonais , Radioisótopos de Índio , Linfócitos/diagnóstico por imagem , Linfócitos T/imunologia , Animais , Linfócitos/imunologia , Linfocintigrafia , Ratos , Ratos EndogâmicosRESUMO
A gene encoding a 113-amino acid, NH2-terminal fragment of CD4, rsT4.113, was constructed and expressed in Escherichia coli under the control of the tryptophan operon promoter. Following induction, rsT4.113 is produced at 5-10% of total E. coli protein, and it is found in inclusion bodies. The protein is purified in two steps under denaturing and reducing conditions. Solubilized rsT4.113 is first purified on a column of Q-Sepharose to remove low molecular weight contaminants and then purified to greater than 95% homogeneity by gel filtration. Renaturation of rsT4.113 is achieved at approximately 20% yield by dilution and dialysis. High performance liquid chromatography analysis of renatured rsT4.113 reveals a less than 15% contaminant of reduced protein. Purified and renatured rsT4.113 contains epitopes for both OKT4a and Leu3a, anti-CD4 monoclonal antibodies which block CD4-gp 120 association, but lacks measurable affinity toward a nonblocking anti-CD4 monoclonal antibody, OKT4. By comparison to a longer form (375 amino acids) of recombinant soluble T4 produced in mammalian cells that contains the entire extracellular domain, rsT4.113 has a comparable affinity for binding to OKT4a and Leu3a in a radioimmunoassay. Analysis of antiviral activity of rsT4.113 demonstrates that the E. coli-derived protein inhibits human immunodeficiency virus-induced syncytium formation with an IC50 of 5-10 micrograms/ml. These data demonstrate that the human immunodeficiency virus-binding domain of CD4 is localized within the NH2-terminal 113 amino acids of CD4 and is contained within a structure homologous to the kappa variable-like domain of immunoglobulins.
Assuntos
Antígenos de Superfície/genética , Genes , HIV/fisiologia , Receptores Virais/genética , Sequência de Aminoácidos , Anticorpos Monoclonais , Fusão Celular , Escherichia coli/genética , Humanos , Dados de Sequência Molecular , Óperon , Fragmentos de Peptídeos/isolamento & purificação , Plasmídeos , Regiões Promotoras Genéticas , Conformação Proteica , Receptores de HIV , Receptores Virais/imunologia , Receptores Virais/isolamento & purificaçãoRESUMO
In this paper, we emphasize the rationale and work-up studies for using two radiolabelled anti-lymphocyte monoclonal antibodies for in vivo application as radiolabelling agents for T and B cells. In vitro experimental work involved radioimmunoassays on human lymphoid cell lines and anticoagulated whole blood with identification of relevant binding kinetics in terms of antibody internalization and elution. We tested also for the effect of the radiolabelled monoclonal antibodies on in vitro cell function defined as mitogen-induced proliferation in whole blood. As a final work-up in an animal model, the distribution of both unlabelled and 111In-labelled anti-Pan T cell monoclonal antibody was studied in the rat. Results from the in vitro experiments pointed to the possibility of using the described technique for specific lymphocyte radiolabelling. The in vivo application enabled us to identify optimal doses of antibody and radioactivity which showed agreement with the in vitro data.
Assuntos
Anticorpos Monoclonais , Radioisótopos de Índio , Marcação por Isótopo/métodos , Linfócitos , Animais , Técnicas In Vitro , Linfócitos/imunologia , RatosRESUMO
The effect of cyclosporine on a systemic graft-versus-host reaction, cardiac allograft rejection, and a local host-versus-graft reaction in the rat were examined in detail. Therapeutic levels of CsA did not inhibit the early stages of lymphocyte activation but did prevent maturation of the immune response to full effector function--viz., graft rejection or clinical GVH disease. In all three models the phenotype changes in T cells associated with the early stages of activation--i.e., induction of receptors for interleukin 2 (IL-R), induction of MHC class II expression, and coexpression of CD4 and CD8 glycoproteins--were not inhibited by CsA. In the GVH and HVG reactions lymphocyte activation proceeded as far as DNA synthesis. In the systemic GVH model animals showed no sign of GVHD for as long as CsA was administered, but withdrawal of the drug resulted in accelerated lethal GVHD.
Assuntos
Ciclosporinas/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Linfócitos T/imunologia , Animais , Antígenos de Diferenciação de Linfócitos T/análise , Rejeição de Enxerto/efeitos dos fármacos , Doença Enxerto-Hospedeiro/imunologia , Transplante de Coração , Reação Hospedeiro-Enxerto/efeitos dos fármacos , Ratos , Ratos Endogâmicos , Linfócitos T/classificaçãoRESUMO
The rate of entry of 3H-leucine-labeled lymphocytes was monitored in cyclosporine (CsA) treated or untreated rats that had received a cardiac allograft 5 days previously. In the untreated recipients there was a preferential localization of labeled cells in the graft heart compared with the native heart, which was evident within the first hour as well as at 3 and 24 hr after injection. In CsA-treated recipients, the rate and extent of entry of lymphocytes in the graft heart was not substantially different from the native heart. Fibrin deposition within allografts, thought to be a consequence of T cell activation, was substantially reduced by CsA treatment of the recipients. A double immunoenzyme staining technique was used to identify Ia-positive macrophages and Ia-positive T cells in cryostat sections of grafts: although the number of macrophages and T cells was reduced in CsA treated recipients, there was no difference in the extent to which these cells were Ia-positive.
Assuntos
Ciclosporinas/farmacologia , Rejeição de Enxerto/efeitos dos fármacos , Transplante de Coração , Animais , Quimiotaxia de Leucócito/efeitos dos fármacos , Feminino , Fibrina/análise , Antígenos de Histocompatibilidade Classe II/análise , Ativação Linfocitária/efeitos dos fármacos , Linfócitos/efeitos dos fármacos , Linfócitos/imunologia , Masculino , Miocárdio/ultraestrutura , Ratos , Ratos Endogâmicos/imunologia , Transplante HomólogoRESUMO
Four cyclo-oxygenase products (COP) of arachidonate were measured in tissue homogenates of rat cardiac allografts at intervals after transplantation. In rejecting graft tissue, there was a progressive decrease in the levels of prostaglandin E2 (PGE2) and PGF2 alpha from the time of grafting, which was accompanied by a rise in the levels of prostacyclin (PGI2, measured as its stable hydrolysis product 6-oxo-PGF1 alpha) and thromboxane A2 (TxA2, measured as its stable hydrolysis product TxB2). When the level of each individual COP in rejecting graft tissue was expressed as a percentage of the total COP measured, PGI2 and TxA2 increased from 9% to 36% and from 7% to 25%, respectively, from day three after grafting until the time of graft rejection. In contrast to these changes in COP levels in rejecting grafts, the levels of all four COP in nonrejecting allografts maintained by treatment of the graft recipients with cyclosporine remained normal--i.e., comparable to the levels in the recipients' own hearts.
Assuntos
Ciclosporinas/farmacologia , Epoprostenol/biossíntese , Rejeição de Enxerto/efeitos dos fármacos , Transplante de Coração , Prostaglandinas E/biossíntese , Prostaglandinas F/biossíntese , Tromboxano B2/biossíntese , 6-Cetoprostaglandina F1 alfa/análise , Animais , Dinoprosta , Dinoprostona , Feminino , Masculino , Ratos , Ratos Endogâmicos/imunologia , Transplante HomólogoRESUMO
Rat T-cell populations were analysed for their surface expression of CD4, CD8 and class II MHC antigens using monoclonal antibodies (McAbs) W3/25, Ox8 and Ox6 in immunofluorescence and immunoenzyme cytochemistry. Resting T cells expressed either CD4 (W3/25) or CD8 (Ox8) molecules, but not both, and did not express class II MHC antigens. In contrast, when purified T cells were stimulated by Con A or by alloantigens in a mixed lymphocyte reaction (MLR), over 60% of the cells expressed class II MHC antigens after 48-72 hr. The expression of class II MHC antigens by the T-cell blasts was transient and no longer evident after 96 hr of activation. In addition, T cells activated by either alloantigen or lectin expressed CD4 (W3/25) and CD8 (Ox8) molecules simultaneously: 30% of cells activated in MLR and 50% of Con A-activated cells expressed both markers within 48 hr of activation. The overlapping of the CD4- and CD8-positive subpopulations within the T-cell population was inferred from the results of staining for each marker separately and confirmed using a double-immunoenzyme staining method.
Assuntos
Antígenos de Superfície/análise , Antígenos HLA-D/análise , Ativação Linfocitária , Linfócitos T/imunologia , Animais , Antígenos de Diferenciação de Linfócitos T , Concanavalina A/farmacologia , Isoantígenos/imunologia , Teste de Cultura Mista de Linfócitos , Ratos , Ratos EndogâmicosRESUMO
Cyclosporine aborts the activation of lymphocytes in vitro before DNA synthesis begins, yet there is also compelling evidence that lymphocytes can become primed, i.e. presumably proliferate in vivo, under the cover of immunosuppressive levels of the drug. Here, Gerry Klaus and Patricia Chisholm discuss the possibility that cyclosporine has a different mode of action in vivo and in vitro.
RESUMO
We have investigated the effects of cyclosporin (CsA) on each of three stages of lymphocyte activation in vivo viz. sequestration of alloantigen-reactive lymphocytes from the circulation into the spleen and lymph nodes, blast transformation and induction of DNA synthesis in the activated cells and release of these cells and their progeny into the circulation. Parental strain lymphocytes injected i.v. into semi-allogeneic rats and recovered from the thoracic duct within 36 h are profoundly unresponsive in a local graft-vs.-host assay to the alloantigens of the F1 hybrid but have normal activity against unrelated alloantigens (negative selection). CsA treatment of the F1 hybrid recipients did not prevent this selective sequestration of antigen-reactive cells. In the untreated F1 hybrid, from 36 h after injection, large numbers of dividing blast cells were released into the lymph. These cells did not appear in the lymph of recipients treated with CsA. However, CsA did not prevent the activation of cells sequestered in the spleen or lymph nodes as assessed by [3H] thymidine incorporation and autoradiography. This unexpected finding suggests that CsA inhibits lymphocyte responses to alloantigens in vivo after DNA synthesis which is a later stage than the in vitro studies have shown.