Effect of hydrocortisone on immune system of the lizard,Chalcidesocellatus I. Response of lymphoid tissues and cells to in vivo and in vitro hydrocortisone

A single dose of 1 mg/g body weight of hydrocortisone acetate (HC) administered intraperitoneally to adult lizards, $\text{Chalcides}$$\text{ocellatus}$ induced rapidly a reduction of about 85% of thymic lymphocytes. Histological evidence indicated that cortical, as well as, medullary thymocytes are sensitive to HC exposure. Around 40–50% of lymphocytes in peripheral blood (PB) and spleen were depleted at 3–7 days post-HC injection; such depletion durated about 4 weeks for PB but was rather temporary in spleen. Increase in number of bone marrow (BM) lymphocytes was negligible and transient and could by no way account for the dramatic cell losses in the different lymphoid tissues. The findings thus suggested that HC-mediated lymphocyte depletion in lizards is not attributable to redistribution between the different lymphoid compartments but rather to destruction. In direct conformation, lymphocytes were readily lysed in vitro by 10^?3M HC, thymocytes being more vulnerable > PB> spleen >BM lymphocytes.     

Effect of hydrocortisone on immune system of the lizard, II. Differential action on T and B lymphocytes

Lymphocytes of thymus, spleen, peripheral blood (PB) and bone marrow (BM) collected from adult lizards, were cultured for 24 hr in the presence of 10⁻³M hydrocortisone acetate (HC) in order to assess the effect of in vitro HC on lizard T and B cell viability. The results indicated that HC induced stepwise, time-dependent mortality of the majority of thymocytes carrying T cell specific antigen(s) (TSA), 30–50% of T cells of spleen, PB and BM, and of a proportion of splenic B lymphocytes. Administration of 1 mg/g body weight HC to adult . lead to depletion of all TSA⁺ thymocytes. In contrast, T lymphocytes in the peripheral lymphoid compartments revealed both sensitivity and resistance to HC; similarly, B lymphocytes constituted susceptible and resistant subpopulations.     

Thymic and peripheral apoptosis of antigen-specific T cells might cooperate in establishing self tolerance

Aside from CD4⁺CD8⁺ double-positive (DP) thymocytes, the subpopulations of T lineage cells affected by negative selection are unknown. To address whether this process occurs in more mature cell types, we have compared the responses of purified single-positive (SP) murine thymocytes and peripheral T cells to the superantigen staphylococcal enterotoxin B (SEB) utilizing as antigen-presenting cells (APC) a fibroblast cell line expressing transfected I-E^k class II molecules. Whereas ∽ 70% of SEB-reactive SP thymocytes, either CD4⁺ or CD8⁺, undergo programmed cell death (apoptosis) and, therefore, negative selection, CD4⁺ and CD8⁺ antigen-specific peripheral T cells are predominantly activated and proliferate to APC+SEB. Thus, mature thymocytes and peripheral T cells, with identical patterns and levels of expression of CD4, CD8 and T cell receptor (TCR), are programmed to elicit different responses followingTCR stimulation. Unexpectedly, however activation of peripheral T cells was preceded by deletion of a large fraction of Vβ8⁺ T lymphocytes (SEB specific). This surprising phenomenon was also observed in in vivo studies: in fact, administration of SEB to adult mice resulted in depletion of the majority of antigen-specific T cells from the peripheral lymphoid tissues analyzed (lymph nodes and spleen). This depletion is the consequence of deletion as indicated by program cell death of Vβ8⁺ T cells and is followed by proliferation of the remaining SEB-reactive T cells. Clonal elimination of peripheral T cells may represent a mechanism by which tolerance to self antigens never expressed in and/or exported to the thymus is achieved.     

Development of lymphocytes in interleukin 7-transgenic mice

We have developed and established mouse transgenic lines in which the mouse interleukin 7 gene was targeted for expression in the lymphoid cell compartment. Northern blot analysis indicate that the transgene is expressed in bone marrow (BM), spleen and thymus, but not in kidney, liver, brain or heart. Both the frequency and absolute numbers of B cell precursors and mature B lymphocytes are increased in the BM and spleen of the transgenic mice. Although there is no expansion of the pro-T lymphocyte population in the BM, the number of all major subsets of thymocytes and peripheral T lymphocytes is increased in the majority of the transgenic mice analyzed. The B and T cell lymphocytes in the transgenic mice are functionally competent. In contrast, the number of granulocytes and macrophages in the BM of transgenic mice is similar to that in control non-transgenic littermates. Our results indicate that interleukin 7 plays an important role in vivo in the development of B and T lymphocytes.     

Deletion of alloantigen-reactive thymocytes as a mechanism of adult tolerance induction following intrathymic antigen administration

Direct injection of foreign antigen into the adult thymus is a potent route of antigen delivery for the induction of tolerance in vivo. In this report, we demonstrate that tolerance to C57BL/10 (H2^b/BL10) alloantigens can be induced in CBA/Ca (H2^k/CBA) mice by intrathymic (IT) administration of BL10 spleen leukocytes coincident with transient peripheral immunomodulation of CD4⁺ T cells using a depleting anti-CD4 monoclonal antibody. T cell receptor (TCR) transgenic mice (BM3.6; H2^k) expressing a CD8-independent TCR specific for H2K^b were used as recipients to facilitate investigation of the mechanisms responsible for tolerance induction by allowing visualization of events in the thymus following IT injection. IT administration of 5 × 10⁷ BL10 spleen leukocytes and concomitant transient peripheral T cell depletion in BM3.6 mice resulted in a substantial H2K^b-specific deletion of transgenic-TCR⁺ (tg-TCR) thymocytes which was dependent on the level of tg-TCR expression. IT deletion and the failure to export CD8⁺ T cells to the peripheral lymphoid organs correlated with the induction of tolerance to H2K^b; TCR transgenic mice that had received IT injection of BL10 splenocytes and peripheral T cell depletion accepted a H2K^b+ cardiac allograft indefinitely. Analysis of tolerant BM3.6 mice revealed that there were low numbers of CD8⁺ T cells in the periphery giving rise to a substantially reduced reactivity in vitro despite the fact that no donor cells or IT deletion were observed in the thymi of the majority of tolerant mice. These results demonstrate for the first time that IT injection of foreign alloantigen into an adult thymus results in the deletion of thymocytes expressing a TCR specific for the injected alloantigen and suggest that this is an important mechanism of tolerance induction following IT injection of alloantigen in vivo. Furthermore, analysis of tolerant TCR-transgenic mice suggests that IT deletion is not required for the maintenance of tolerance, and that peripheral mechanisms enforce continued hyporesponsiveness to H2K^b following transplantation.     

Isolation of peritoneal precursors of B-1 cells in the adult mouse

Two weeks of daily peritoneopheresis of adult mice result in the selective depletion of B-1 cells, followed by the appearance of a population of B220⁺IgM^?lymphocytes in the peritoneal cavity. These cells share with bone marrow (BM) pre-B cells expression of λ5, V_preB, and RAG-1 genes and a higher fraction of unrearranged V to DJ heavy (H) chain immunoglobulin (Ig) gene segments, when compared with mature B lymphocytes. Upon transfer to SCID recipients, sorted peritoneal B220⁺IgM^? cells fail to colonize the BM, repopulate very few B cells in the spleen, but entirely reconstitute the B-1 cell compartment in the peritoneal and pleuropericardial cavities. In contrast, parallel transfers of sorted BM B220⁺IgM^? cells result in reconstitution of the BM and spleen B lineage cell compartments, but in no coelomic B cell repopulation. Both types of pre-B cells reconstitute splenic plasma cells of donor origin, but with markedly distinct efficiencies: the ratio of IgM-plasma cell/B cell numbers in the spleens of peritoneal pre-B cell recipients is more than 500-fold higher than that of recipients reconstituted by BM pre-B cells. We take these data to indicate that (1) differentiative commitment to the B-1 cell population occurs before selection events on mature cells; (2) B-1 precursors exist or may be locally produced in the adult mouse; (3) there is a lineage-related differential ability of mature B cells to undergo terminal differentiation to high-rate Ig secretion.     

B cell activation in chickens induced by Staphylococcusaureus

Conventional mammalian polyclonal B cell activators were evaluated for activity in chicken spleen and peripheral blood lymphocyte (PBL) cultures. Although lipopolysaccharide was found to have a marginal influence on proliferation, two strains of the bacterium $\text{Staphylococcus}$$\text{aureus}$ (Cowan I and Wood 46 strains) induced moderate proliferation in both spleen and PBL cultures. In spleen cell cultures the proliferating cell population was identified as the B cell. The mitogenic response required the presence of adherent cells since their removal eliminated the response. Evidence of $\text{in vitro}$ polyclonal immunoglobulin synthesis could not be obtained. However, when administered intravenously, $\text{S.}$$\text{aureus}$ induced polyclonal immunoglobulin synthesis.     

Natural cytotoxic cell activity in the snake Psammophis sibilans.

Thymocytes, splenocytes and peripheral blood mononuclear cells (PBMC) of the snake Psammophis sibilans consistently killed the human erythroleukemic cells K562 in a 4 h assay as judged by lactate dehydrogenase enzyme release. PBMC and splenocyte natural cytotoxicity (NC) increased proportionally with increase in the effector/target cell ratio. Spontaneous killer cell activity was consistently 2-3 times higher in peripheral blood (PB) than in spleen. On the other hand, thymocytes displayed low, yet detectable, NC. In an attempt to define the cell subpopulation responsible for natural killer (NK) activity, PBMC were depleted of macrophages or B lymphocytes before use in NK cell assays against K562 cells. Depletion of macrophages did not impair NK activity thus suggesting that macrophages do not mediate spontaneous lysis in the present 4 h assay. Conversely, removal of B lymphocytes by panning onto dishes coated with monoclonal antibody against snake Ig significantly reduced, but did not eliminate, PBMC spontaneous cytotoxicity. These data suggest that T, B and perhaps distinct NK cells participate in spontaneous lysis. This suggestion was confirmed by studies of NC in thymus, spleen and PB the year round. Strong NC was detected during spring and autumn when high numbers of leukocytes including T and B cells can be recovered from spleen and PB. Negligible spontaneous cytotoxicity was observed during early and mid-summer and in winter, periods of the year when snakes are thymus-less and contain few T and B cells in peripheral lymphoid organs. These findings, the first to document natural cytotoxic activity in snakes, were discussed in relation to the issue of NK cell identity in vertebrates.     

Characterization of splenic CD21hi T2 B cells

B cell development is a highly regulated process that initiates in the bone marrow (BM) of adult mice. After reaching the IgM⁺ immature stage in the BM, these B cells migrate to the spleen to complete maturation and incorporation into the long-lived peripheral lymphocyte pool. Studies have identified these splenic immature B cells, and have further attempted to delineate the sequence whereby they transition into mature B cells. As such, these immature splenic populations are termed transitional B cells and have been the focus of intense study. The review summarizes the phenotype and currently known functions of the four putative transitional B cell subsets identified to date. Although most appear to represent short-lived transitional B cells, the CD21^hi T2 B cell population exhibits a number of qualities that question its label as a transitional B cell subset.     

Neonatally tolerant rats actively eliminate donor-specific lymphocytes despite persistent chimerism

Rats from the allotype-marked PVG-RT7^b and PVG-RT1^u-RT7^b strains were injected at birth with semi-allogeneic F₁ bone marrow (BM) cells from athymic nude rats (PVG-rnu/rnu x PVG-RT1^u-rnu/rnu) to induce neonatal tolerance. As adults, 97% of the animals accepted donor-specific allogeneic skin grafts and a majority (65%) of rats were chimeric, expressing the major histocompatibility complex class I and allotype marker of the donor strain. Similar results were obtained when PVG-RT1^u-RT7^b rats were injected at birth with fully allogeneic PVG-rnu/rnu nude BM cells: as adults, 94% accepted donor-specific skin allografts and 76% of recipients were chimeric. Donor-derived CD4T cells, CD8T cells and B cells were found in low numbers (< 2%) in peripheral blood of rats made tolerant by F₁ BM cells. A large proportion of T cells bore the phenotype of recent thymic emigrants, suggesting that they were newly produced. All the evidence was consistent with clonal deletion tolerance, induced centrally within the thymus. The thymus was chimeric and thymocytes failed to respond in vitro to alloantigens of the donor-specific haplotype; donor-specific skin allografts survived indefinitely on athymic nude recipients reconstituted with CD4⁺CD8^? thymocytes or peripheral CD4T cells from tolerant animals. The chimeric state was interesting, since the PVG and PVG-RT1^u rat strains contain a natural killer (NK) cell system that rapidly eliminates (within 24 h) intravenously injected allogeneic or semi-allogeneic lymphocytes – a phenomenon known as allogeneic lymphocyte cytotoxicity or ALC. When neonatal tolerant rats were tested, the ALC index (a measure of cell killing) was unchanged in nonchimeric tolerant rats and significantly altered (reduced killing), but not abolished in chimeric animals. Hence, the injection of allogeneic BM cells which induced specific tolerance in the T cell population failed to tolerize the NK cell system, allowing the constant killing of newly produced donor-derived lymphocytes and putting at risk the very survival of the allogeneic BM cells. This has interesting implications for clinical transplantation.     

Flow Cytometric Identification of CD93 Expression on Naive T Lymphocytes (CD4<Superscript>+</Superscript>CD45RA<Superscript>+</Superscript> Cells) in Human Neonatal Umbilical Cord Blood

Human CD93 has a molecular weight of about 100 kDa and is selectively expressed by myeloid cell lineages in peripheral blood (PB) mononuclear cells. Although CD93 was initially identified as a receptor for complement component 1, subcomponent q phagocytosis (C1qRp) involved in the C1q-mediated enhancement of the phagocytosis of various antigens, several recent studies have reported that CD93 is not a receptor for the C1q-mediated enhancement of phagocytosis. The expression patterns of CD93 have been previously investigated in PB mononuclear cells (lymphocytes, monocytes, and granulocytes) from adult PB and neonatal umbilical cord blood (UCB), and the expression of CD93 was not found on lymphocytes from either normal adult PB or neonatal UCB. However, the detection of CD93 expression in neonatal UCB using CD93 monoclonal antibodies (mAbs) that recognize different antigenic epitopes remains poorly understood. In this study, we examined the expression of CD93 on lymphocytes, monocytes, and granulocytes from neonatal UCB using four different types of CD93 mAb detection probes, mNI-11, R139, R3, and X-2, using flow cytometric and western blot analyses. We found that CD93, as defined using all four mAbs, was expressed on monocytes and granulocytes in PB mononuclear cells from adult PB and neonatal UCB. On the other hand, we observed for the first time that the expression of CD93 on lymphocytes in neonatal UCB can only be detected using the mNI-11 mAb, established in our laboratory, and not with commercially available CD93 mAbs (R139, R3, and X-2). However, CD93 expression on lymphocytes from normal adults was not detected using any of the four CD93 mAbs. Two-color flow cytometric analyses showed that the CD93 recognized by mNI-11 mAb was expressed on CD3⁺ T lymphocytes (mainly CD4⁺ helper T lymphocytes), but not on CD19⁺ B lymphocytes or on CD8⁺ suppressor/cytotoxic T lymphocytes from neonatal UCB. In addition, CD93 was expressed on CD45RA⁺ (naive antigen) lymphocytes from neonatal UCB, but not on CD45RO⁺ (memory antigen) lymphocytes from neonatal UCB or on CD45RA⁺ and CD45RO⁺ lymphocytes from normal adult PB. Three-color flow cytometric analysis showed that CD93 was co-expressed on naive T lymphocytes (CD4⁺CD45RA⁺) from neonatal UCB. In a western blot analysis, the CD93 mAb (mNI-11) immunoprecipitated at a molecular weight of 98 kDa, identified as a CD93 molecule, in the CD4⁺CD45RA⁺ cells from neonatal UCB but not from adult PB, similar to the results in the human monocyte-like cell line U937 (human CD93-positive cells). Taken together, these results provide the first direct evidence of a novel/naive cell population (CD4⁺CD45RA⁺CD93⁺) in neonatal UCB that may have an important role in cell biology, transplantation, and immature/mature immune responses.     

Peripheral blood CD4+CD8+ lymphocytes in cynomolgus monkeys are of resting memory T lineage

In this study, we analyzed peripheral blood CD4+CD8+ double-positive (DP) lymphocytes in adult cynomolgus monkeys (Macaca fascicularis). Forty of 55 monkeys had > 5% of the peripheral blood DP subpopulation (9.3 +/- 5.9%; mean +/- SD) in peripheral blood lymphocytes (PBL) in contrast to a low percentage of peripheral blood DP cells in humans and mice. In a cross-sectional study, the peripheral blood DP cells were found to increase in proportion with age. To clarify whether peripheral blood DP lymphocytes were immature precursors released from thymus without prior differentiation, the expressions of CD8 chains and CD1b on peripheral blood DP lymphocytes were compared with those on thymocytes. The peripheral blood DP lymphocytes were CD8 alpha + beta- and CD1b-, while thymic DP lymphocytes were CD8 alpha + beta + and CD1b +, suggesting that the peripheral blood DP cells are extrathymic T lymphocytes. Furthermore, the peripheral blood DP lymphocytes exhibited a resting memory T cell phenotype with CD2hiCD3+CD28-CD29hiCD49dhiCD69- CD80lo. Taken together, adult cynomolgus monkeys possess a unique peripheral blood DP T cell subpopulation which expresses a resting memory T cell phenotype. In addition, similar phenotypic properties of DP lymphocytes were distributed in the spleen and lymph nodes, although the proportion was less in the spleen and much less in lymph nodes than in PBL.      

FTY720 suppresses the development of colitis in lymphoid‐null mice by modulating the trafficking of colitogenic CD4+ T cells in bone marrow

2‐Amino‐2‐(2‐[4‐octylphenyl]ethyl)‐1,3‐propanediol hydrochloride (FTY720) suppresses T‐cell egress from LN, thereby preventing pathogenic T cells from migrating toward disease sites. However, little is known about whether FTY720 could control the trafficking of T cells without the presence of lymphoid tissues. Here we demonstrate that FTY720 treatment suppresses the recirculation of CD4⁺ T cells in splenectomized (SPX) lymphotoxin‐α^?/? (LT‐α^?/?) mice that lack LN and spleen, as shown by peripheral blood (PB) lymphopenia in FTY720‐treated SPX LT‐α^?/? mice. In a short‐term transfer experiment, the cell number of transferred Ly5.1⁺CD4⁺ T cells recovered from host FTY720‐treated SPX LT‐α^?/? mice (Ly5.2⁺) was markedly decreased in PB, but conversely increased in BM. Notably, FTY720 treatment prevented the development of colitis that is otherwise induced in untreated SPX LT‐α^?/?×RAG‐2^?/? mice upon transfer of colitic lamina propria CD4⁺ T cells. In such mice, the number of CD4⁺ T cells in PB or lamina propria of FTY720‐treated SPX LT‐α^?/?×RAG‐2^?/? recipients was significantly reduced, but that in the BM was significantly increased as compared with untreated control mice. Altogether, the present results indicate that FTY720 treatment may offer an additional role to direct trafficking of CD4⁺ T cells in BM, resulting in the prevention of colitis.     

Functional studies on T cells in adult human bone marrow.

Bone marrow (BM) lymphocytes were obtained by sucrose density gradient centrifugation of the nucleated cells from adult human BM. BM was obtained from rib sections removed routinely during thoracotomy from thirteen patients with a localized lung tumour and from two other patients without tumour (mean age 47 years). The percentage of T cells in BM was high (mean +/- s.d. 27% +/- 17) and increased with age. In eight cases, the function of isolated BM T cells was studied and compared to that of peripheral blood (PB) T cells. BM t cells showed poor helper activity for pokeweed mitogen (PWM) induced Ig production by PB non-T cells, which did not appear to be due to excessive suppressor cell activity. Phytohaemagglutinin (PHA) induced thymidine incorporation was only slightly decreased but peak values were only reached after 6 years, in contrast to 4 days for PB T cells. This delay did not seem to be due to a lack of monocytes. PHA, however, failed to induce cytotoxic activity in BM T cells. PWM-induced thymidine incorporation and responder capacity in the mixed lymphocyte reaction were also very poor. These results are interpreted as suggesting that many of the T cells in adult human marrow are immature.     

Thymus-dependent immune functions in chickens bursectomized with colchicine applied to the anal lips

Thymus-dependent immune functions were investigated in chickens bursectomized neonatally with colchicine solution given $\text{per anum}$. Antibody responses to thymus-dependent antigens sheep red blood cells (SRBC) and human gammaglobulin (HGG) were delayed, reaching the normal level after the third antigen stimulation. Also the mitogenic responses of peripheral blood lymphocytes were preserved, and no changes in the thymic morphology were found. In contrast, antibody responses to bursa-dependent antigen $\text{Brucella abortus}$ were low and the switch of immunoglobulin isotypes from IgM to IgA and IgG was disturbed. It can be concluded that neonatal bursectomy with cloacal administration of colchicine does not significantly affect T celi functions, whereas B cell functions are partially deficient.     

Long-Term Persistence of Limited HTLV-I Tax-specific Cytotoxic T Cell Clones in a Patient with Adult T Cell Leukemia/Lymphoma after Allogeneic Stem Cell Transplantation

Purpose

Adult T cell leukemia/lymphoma (ATL) is a highly aggressive malignancy of T cells caused by human T cell lymphotropic virus type 1 (HTLV-1). Recent clinical studies have suggested that allogeneic stem cell transplantation (HSCT) improves the clinical course of ATL by harnessing a graft-versus-ATL effect, and that donor-derived HTLV-1 Tax-specific CD8⁺ cytotoxic T cells (CTLs) contribute to the graft-versus-ATL effect after HSCT. However, little is known about the immunological characteristics of Tax-specific CTLs in ATL patients who underwent HSCT.

Methods

We serially analyzed frequencies, differentiation, functions and clonal dynamics of Tax-specific CTLs in paired samples of peripheral blood (PB) and bone marrow (BM) from an ATL patient after HSCT at the single-cell level. We used flowcytometric and single-cell T cell receptor (TCR) repertoire analysis methods without culture steps.

Results

Donor-derived Tax-specific CTLs effectively suppressed HTLV-1 replication in both PB and BM at least during chronic graft-versus-host disease after HSCT. Furthermore, Tax-specific CTLs had comparable properties between BM and PB, except for preferential accumulation in BM rather than PB. Tax-specific CTLs persistently existed as less-differentiated CD45RA^-CCR7^- effector memory CTLs based on predominant phenotypes of CD27⁺, CD28^+/? and CD57^+/?. Our approach using single-cell TCR repertoire analysis method showed highly restricted oligoclonal responses of Tax-specific CTLs, and TCR BV7- or BV30- expressing two predominant CTL clones persistently existed and maintained strong cytotoxic activities against HTLV-1 in both PB and BM over three years after HSCT.

Conclusions

These findings about Tax-specific CTLs provide insights into future directions for studies on immunotherapy against ATL.     

Linomide inhibits programmed cell death of peripheral T cells in vivo

Programmed cell death (PCD) is involved in the physiological regulation of lymphocyte turnover, as well in the antigen-driven selection of T and B cells. Here it is shown that the immunomodulator linomide (quinoline-3-carboxamide) inhibits the apoptotic decay of peripheral T lymphocytes in response to three different stimuli. First, linomide reduces the superantigen-mediated apoptosis and deletion of specific T lymphocytes of both the CD4⁺ and the CD8⁺ subsets without affecting other superantigen-triggered phenomena such as T cell expansion and anergy. Second, linomide abolishes the T lymphopenia and inhibits PCD of splenic CD4⁺ and CD8⁺ T cells induced by exogenous glucocorticoids. This effect is restricted to peripheral T lymphocytes and does not concern thymocytes. Finally, linomide abolishes the development of lymphopenia that follows infection with vaccinia virus, while reducing PCD of CD4⁺ and CD8⁺ peripheral T cells. The anti-apoptotic effect of linomide could account for its immunostimulatory properties and might be relevant to the treatment of immunodeficiencies associated with an increased apoptotic decay of T lymphocytes.     

In vitro induction of tumor specific immunity: requirements for T lymphocytes and tumor growth inhibition in vivo

Cortisone-resistant thymocytes, spleen cells, thoracic duct lymphocytes, peritoneal exudate cells and peripheral blood lymphocytes of BALB/c mice were immunized in vitro against syngeneic HPC-108 plasma cell tumor cells. Cocultivation of spleen lymphocytes together with HPC-108 cells generated the highest cytotoxic immune response in comparison to other lymphocyte sources. Cytotoxicity was tested in a 6 hour ⁵¹Cr release assay using HPC-108 cells as target cells. The use of AKR anti-θ C3H serum indicated that thymus-derived (T) lymphocytes are essential to the initiation phase of the immune response to plasma cell tumor cells. Furthermore, evidence is presented that the cytotoxic effector cells in the in vitro tumor immune response are T lymphocytes. Spleen cells activated in vitro against HPC-108 tumor cells were shown to specifically prevent tumor growth from simultaneously injected HPC-108 cells in irradiated recipient mice.     

Janus kinase 3-deficient T lymphocytes have an intrinsic defect in CCR7-mediated homing to peripheral lymphoid organs

Chemokine-mediated signalling involves the activation of a Janus kinase (Jak) pathway. We have previously shown that Jak3 mediates CCR9 and CXCR4 signalling in response to CCL25 and CXCL12 in BM progenitors and thymocytes. The lack of peripheral lymph nodes and Peyer''s patches observed in Jak3^–/– mice suggested a possible role of Jak3 in CCR7-mediated homing to these organs. Here, we demonstrate phosphorylation of Jak3 in peripheral lymphocytes in response CCL19 and CCL21. In addition, Jak3^–/– naïve T cells and pharmacologically inhibited Jak3^+/+ T lymphocytes have impaired chemotactic responses towards these ligands. Interestingly, CCR7 expression was higher in Jak3^–/– thymocytes compared to their Jak3^+/– littermates, indicating that the impaired migration must be caused by impaired CCR7-mediated signalling, in the absence of Jak3. In addition, adoptive transfer experiments showed that Jak3^+/+ mice reconstituted with Jak3^–/– green fluorescent protein (GFP)⁺ bone marrow progenitors had reduced T-lymphocyte homing to peripheral and mesenteric lymph nodes, compared to reconstitution with Jak3^+/+ GFP⁺ progenitors. Furthermore, reciprocal transfer experiments indicated that Jak3^–/– stromal cells were not responsible for the deficient T-cell homing. Finally, we performed direct competitive homing assays and demonstrated that Jak3^–/– T lymphocytes have a clear defect in homing to peripheral and mesenteric lymph nodes, while migration to spleen was moderately impaired. Our data demonstrates that Jak3^–/– T lymphocytes have an intrinsic defect in CCR7-mediated homing to peripheral lymphoid organs.