The biological role and potential therapeutic application of interleukin 7

Interleukin (IL)-7 is a pleiotropic, non-redundant cytokine necessary for the development of B and T lymphocytes, in particular gammadelta T cell receptor-positive cell differentiation. The cytokine can function as a cofactor during myelopoiesis and the generation of cytotoxic T cells and natural killer cells, can activate monocytes/macrophages, and support the survival of mature T cells. A role for IL-7 in promoting the formation of Peyer's patch anlage has also been demonstrated. IL-7 is constitutively expressed in the thymus, bone marrow stromal cells, epithelial and dendritic cells, keratinocytes, as well as in fetal and adult liver. IL-7 acts on various cells through its receptor (IL-7R), a heterodimer consisting of an alpha chain (CD127) that specifically binds IL-7 and a common gamma(c) chain (CD132) shared by other cytokine receptors. The receptor is expressed on bone marrow progenitor cells, lymphoid T and B precursors, and mature T cells. IL-7 activity towards murine endothelial cells has been recently described. The presence of IL-7R on human endothelial cells has also been demonstrated. Several therapeutic applications of recombinant IL-7 have been proposed. These have focused on the enhancement of lymphopoiesis, promotion of stem cell engraftment, and the anti-tumor activity of the cytokine.     

Interleukin-7 inhibits pre-T-cell differentiation induced by the pre-T-cell receptor signal and the effect is mimicked by hGM-CSF in hGM-CSF receptor transgenic mice

We have previously reported that human granulocyte-macrophage colony-stimulating factor (hGM-CSF) causes a stage-specific inhibition of T-cell receptor (TCR) alphabeta cell development in the thymus of transgenic mice constitutively expressing the hGM-CSF receptor. Since it has been reported that the addition of interleukin-7 (IL-7) to fetal thymic organ culture (FTOC) has similar effects, we compared the effects of IL-7 and hGM-CSF on TCR(alphabeta) cell development in hGM-CSF receptor transgenic mice. We reconstituted fetal lobes with sorted pre-T, or post pre-T CD4(-)CD8(-) precursor cells. The addition of either IL-7 or hGM-CSF to these cultures suppressed further differentiation of pre-T cells but not post pre-T cells. At the same time, the cell number was increased, suggesting that pre-T-cell proliferation is stimulated by these cytokines. Furthermore, the differentiation of recombination-activating gene-1 (RAG-1)-deficient pre-T cells in response to anti-CD3 antibody stimulation was suppressed by either IL-7 or hGM-CSF, suggesting that these cytokines inhibit the pre-T-cell receptor (pre-TCR) signal. This inhibition is unexpected because the pre-TCR signal and the IL-7 signal have previously been considered to be co-operative. Recent analysis of the downstream events of IL-7 receptor and GM-CSF receptor revealed that they share common signal transduction molecules. Our results show that IL-7 is able to promote pre-T cell proliferation and to suppress differentiation induced by the pre-TCR signal. GM-CSF can mimic these biological activities of IL-7 when the pre-T cells express GM-CSF receptors. Our data suggest that both timing and level of activation of the IL-7 signalling pathway must be precisely regulated to facilitate the differentiation of thymocytes.     

IL-21的生物学功能及其在病毒感染中的作用

IL-21是最近发现的一种Ⅰ型细胞因子,由活化的CD4^＋T细胞产生,特别是Th2细胞。IL-21R存正常的淋巴组织包括脾、胸腺、淋巴结、外周血淋巴细胞、纤维化肺组织中都有表达。IL-21特异性结合IL-21R介导多种生物学效应,对不同阶段B、T淋巴细胞的分化和自然杀伤（NK）细胞的增殖均起重要作用。因此IL-21有着广泛的生物学功能,在各种疾病包括病毒感染中扮演着重要的角色。     

Identification of an IL-4-inducible gene expressed in differentiating lymphocytes and male germ cells

Interleukin 4 (IL-4) is a cytokine that is involved in the differentiation of B and T lymphocytes. In this report, we describe the identification of a novel gene, N.52, which was cloned from the murine pre-B cell line R8205 grown in the presence of IL-4 for 48 hr. Although N.52 expression is detectable at low levels in unstimulated R8205 cells, the level of N.52 dramatically increases after only 4 hr exposure to IL-4 and remains at a high level up to 48 hr. Although N.52 expression is low or absent in normal spleen B and T cells, its expression can be induced by the differentiation signals delivered by LPS in B cells and by Con A in T-cell hybrids. While N.52 mRNA is absent in all highly differentiated organs, it is detectable in stem cell harboring lymphoid tissues such as bone marrow, fetal liver, and thymus. Furthermore, N.52 mRNA is expressed at strikingly high levels in the testis, specifically in differentiating male germ cells. It is induced by differentiation signals triggered by the combination of cyclic AMP and retinoic acid in teratocarcinoma F9 cells. Taken together, these data suggest that N.52 is a developmentally regulated gene whose expression in cells of the immune and reproductive systems may be controlled by stimuli that induce differentiation.     

Pre-TCR signaling regulates IL-7 receptor alpha expression promoting thymocyte survival at the transition from the double-negative to double-positive stage

The pre-T cell receptor (pre-TCR) and IL-7 receptor (IL-7R) are critical mediators of survival, proliferation and differentiation in immature thymocytes. Here we show that pre-TCR signaling directly maintains IL-7Ralpha expression as developing thymocytes undergo beta-selection. Inhibition of IL-7/IL-7R signaling in (CD44-CD25-) DN4 cells results in decreased generation of double-positive thymocytes due to increased death of rapidly proliferating beta-selected cells. Thus, we identify a mechanism by which pre-TCR signaling controls the selective survival of TCRbeta+ thymocytes, and define a further stage of T cell differentiation in which signaling from a TCR regulates the ability of that cell to respond to cytokine.     

Intra- and extra-thymic expression of the pre-T cell receptor α gene

We have analyzed pre-T cell receptor α (pTα) gene expression in cells from various anatomical sites to investigate the lineage specificity of pTα RNA as well as its presence in pro-T cells and in sites of extrathymic T cell development. pTα RNA is found in precursors of αβ T cells but is absent from mature αβ T cells as well as T cells that express the γδ T cell receptor on the cell surface. pTα expression is exquisitely T lineage specific in that mature and immature B cells, myeloid cells, NK cells and pluripotent stem cells are pTα negative. On the other hand, pTα expression is found in pro-T cells outside the thymus as well as in intra- and extra-thymic sites of T cell development. The latter finding is consistent with the notion that early steps of T cell development within and outside the thymus may be similar.     

Interplay between IL-2 and IL-4 in human thymocyte differentiation: antagonism or agonism

The effect of recombinant interleukin 2 (IL-2) and IL-4, as well as a combination of both lymphokines on human post-natal thymocytes at different maturation stages, was analyzed by culturing highly purified pro-T cells, pre-T cells, double-negative and double-positive thymocyte subsets in the presence of IL-2 and/or IL-4. Both IL-2 and IL-4 responsiveness are developmentally regulated in human thymocytes, since IL-2 and IL-4 responses decline with increasing thymocyte differentiation, double-positive T cells displaying far less proliferation than immature thymocytes. IL-2 and IL-4 may influence pro-T cell growth in both an antagonistic and additive fashion. At low doses, IL-4 inhibits IL-2-supported growth of pro-T cells, whereas, at higher concentrations, this inhibitory effect is masked by the ability of IL-4 to stimulate pro-T cell proliferation. In contrast to peripheral lymphocytes, IL-4 does not down-regulate the expression of the IL-2 receptor light chain on thymocytes. In pro-T cell cultures, IL-2 and IL-4 favour the differentiation of distinct cell populations, namely lymphocytes displaying preferentially a TCR alpha/beta+ and CD4+CD8- phenotype versus predominantly TCR gamma/delta+ and CD4-CD8+ cells, respectively. The effect of IL-2 dominates over that of IL-4, since the composition of cultures set up in the presence of IL-2 plus IL-4 resembles that of cells cultured with IL-2 alone. In synthesis, IL-2 and IL-4 exhibit reciprocal inter-relations in human thymocyte cultures, thus supporting the notion that these lymphokines are implicated in the complex regulation of a local cytokine network.     

Mechanisms of tolerance to self

Several mechanisms exist to prevent lymphocytes from reacting against self-antigens. As T cells develop in the thymus and express antigen-specific receptors, those with high-affinity to self-antigens existing within the thymus are deleted. Low-affinity self-reactive T cells and T cells with receptors against antigens not represented intrathymically will mature and join the peripheral T cell pool. They may either ignore self-antigens expressed by tissues unable to activate T cells through a lack of the appropriate costimulator signals, or they may, under certain conditions, be deleted or rendered anergic and unable to respond. Likewise, B cells that express surface Ig receptors with high binding affinity to membrane-bound self-antigens present in the bone marrow will be rescued by receptor editing or will be deleted, whereas those of lower affinity will migrate to the periphery in either an anergic or indifferent state depending on the degree of receptor engagement by antigen. Once there, their ultimate fate is determined by the availability of T cell help.     

Analysis of gammac-dependent cytokines-mediated immunoregulation

In the study of interleukin-2 (IL-2) -induced signal transduction system, we identified and cloned the third component of IL-2 receptor, IL-2Rgamma chain. Functional high affinity IL-2 receptor consists of three subunits, alpha, beta and gamma chains. Interestingly not only IL-2 but also IL-4, IL-7, IL-9, IL-15 and IL-21 utilize the gamma chain as an essential component of each receptors. Therefore the gamma chain is now called as a common gamma chain (gammac). Moreover the gene of gammac is on the X chromosome, and mutations of gammac gene cause human X-linked severe combined immunodeficiency (X-SCID) characterized by a complete or profound defect of T cells and NK cells, and by the presence of dysfunctional B cells. The dysfunctions in IL-7- and IL-15-induced signal transduction cause the T cell and NK cell defect, respectively and dysfunctions in both IL-4- and IL-21-induced signal transduction cause the B cell dysfunction in X-SCID patients. Gene therapy is a good candidate for X-SCID treatment because only the HLA-matched bone marrow transplantation is an effective therapy. Unfortunately because of an unexpected adverse effect, such gene therapy using retrovirus vector is now aborted. IL-21 is a recently identified cytokine, which shares the gammac. IL-21 regulates the proliferation of T cells, the proliferation and differentiation of B cells, and the activation and expansion of NK cells. We demonstrated that human IL-21 was produced from activated CD4+ central and effector memory T cells but not from naive CD4+ T cells nor CD8+ T cells. Furthermore we found that IL-21 supported cytokine-driven proliferation of CD4+ helper T cells cooperatively with IL-7 and IL-15.     

Partial restoration of B cell development in Jak-3(-/-) mice achieved by co-expression of IgH and E(mu)-myc transgenes

Jak-3 is a non-receptor tyrosine kinase that plays an important role in coordinating signals received through a wide range of cytokine receptors, including the IL-7 receptor (IL-7R). Jak-3-deficient mice have a profound block in B cell development at the pro-to-pre-B cell transition and have very few peripheral B cells. This block has been postulated to reflect the inability of Jak-3(-/-) pro-B cells to respond to IL-7. Here we demonstrate that B cell development can be partially restored in Jak-3-deficient mice when they are bred to mice carrying both a rearranged Ig heavy chain (IgH/Igmu) transgene and a c-myc transgene expressed in the B cell lineage. Jak-3(-/-) mice expressing both of these transgenes exhibit significant increases in the number of B cells in the bone marrow and, to a lesser extent, in the spleen. However, very few rescued B cells were detectable in mice greater than 4 months of age. To determine whether resident hyperactivated Jak-3(-/-) peripheral T cells are responsible for the elimination of the rescued B cells in older mice, we bred IgH transgenic (Igmu Tg)/myc Tg/Jak-3(-/-) mice to T cell-deficient (TCRalpha(-/-)) mice. Data from these experiments suggest that the paucity of B cells in older Jak-3(-/-) mice is largely attributable to the lack of Jak-3 in the B cells themselves. Thus, Jak-3 seems to play several important roles in B cells: during development, to enable cell division, Ig gene rearrangement and cell differentiation, and in mature cells, to promote B cell survival in the periphery.     

Induction of T cell development from hematopoietic progenitor cells by delta-like-1 in vitro

The molecular interactions provided by the thymic microenvironment that predicate T cell development remain obscure. Here, we show that a bone marrow stromal cell line ectopically expressing the Notch ligand Delta-like-1 loses its ability to support B cell lymphopoiesis, but acquires the capacity to induce the differentiation of hematopoietic progenitors into CD4 CD8 double- and single-positive T cells. Both gammadelta-TCR(+) and alphabeta-TCR(+) T cells are generated, and CD8(+) TCR(hi) cells produce gamma-interferon following CD3/TCR stimulation. These results establish that expression of Delta-like-1 on stromal cells provides key signals for the induction of T cell lineage commitment, stage-specific progenitor expansion, TCR gene rearrangement, and T cell differentiation in the absence of a thymus. Thus, it is likely that Delta-like-1/Notch interactions by the thymus underpin its unique ability to promote lineage commitment and differentiation of T cells.     

Deletion of beta-catenin impairs T cell development

T cells encounter two main checkpoints during development in the thymus. These checkpoints are critically dependent on signals derived from the thymic microenvironment as well as from the pre-T cell receptor (pre-TCR) and the alphabeta TCR. Here we show that T cell-specific deletion of beta-catenin impaired T cell development at the beta-selection checkpoint, leading to a substantial decrease in splenic T cells. In addition, beta-catenin also seemed to be a target of TCR-CD3 signals in thymocytes and mature T cells. These data indicate that beta-catenin-mediated signals are required for normal T cell development.     

Antigen and cytokine receptor signals guide the development of the naïve mature B cell repertoire

Immature B cells are generated daily in the bone marrow tissue. More than half of the newly generated immature B cells are autoreactive and bind a self-antigen, while the others are nonautoreactive. A selection process has evolved on the one hand to thwart development of autoreactive immature B cells and, on the other hand, to promote further differentiation of nonautoreactive immature B cells into transitional and mature B cells. These negative and positive selection events are carefully regulated by signals that emanate from the antigen receptor, whether antigen-mediated or tonic, and are influenced by signals that are generated by receptors that bind cytokines, chemokines, and other factors produced in the bone marrow tissue. These signals, therefore, are the predominant driving forces for the generation of a B cell population that is capable of protecting the body from infections while maintaining self-tolerance. Here, we review recent findings from our group and others that describe how tonic antigen receptor signaling and bone marrow cytokines regulate the selection of immature B cells.     

γδTCR ligands and lineage commitment

Two major T lymphocyte lineages – αβ and γδ T cells – develop in the thymus from common precursors. Differentiation of both lineages requires signals coming from TCRs. Development of αβ T cells is driven at early stages by signaling from the pre-TCR, most likely in a ligand-independent fashion, and later – by signals delivered by αβTCRs binding to their ligands – classical or non-classical MHC molecules. γδ lineage cells likewise require TCR signaling for their differentiation. Recent work from several groups suggests that TCR signaling not only ensures the developmental progression towards αβ and γδ lineages but that signal strength instructs lineage fate: weaker TCR signal results in αβ and stronger—in γδ lineage commitment. However, as most γδTCRs remain orphan receptors, it is still debated whether strong signals from γδTCRs in development are generated in a ligand-dependent manner (as in the case of αβTCRs), ligand-independent manner (as for pre-TCR) or both. Here we summarize evidence supporting a possible role for ligands in γδ T cell lineage commitment and the generation of γδ sublineages.