Molecular cloning and biological characterization of a novel murine lymphoid growth factor.

Using a bioassay consisting of the proliferation of a murine B cell line, a cDNA of a gene whose product supports the growth of that cell line was isolated from a thymic stromal cell line. This factor, termed thymic stromal lymphopoietin (TSLP), is a protein of 140 amino acids. The gene encoding TSLP was mapped to murine chromosome 18. Purified recombinant TSLP supported the growth of pre-B cell colonies in vitro, but had no myelopoietic activity. TSLP had comitogenic activity for fetal thymocytes, but was not as potent as interleukin 7 in lobe submersion cultures. Injection of TSLP into neonatal mice induced the expansion of B220(+)BP-1(+) pre-B cells.

A thymic stromal cell line supports in vitro development of surface IgM+ B cells and produces a novel growth factor affecting B and T lineage cells.

A thymic stromal cell line with a medullary phenotype (Z210R.1) supported the differentiation of surface IgM+ B cells when cocultured with fetal liver cells in vitro. Conditioned medium (CM) from this cell line supported the long-term growth of a B cell line (NAG8/7) isolated from cocultures and enhanced the proliferation of unfractionated thymocytes to suboptimal concentrations of anti-CD3 antibodies in vitro. Biological assays of the CM detected interleukin-7 (IL-7) but not IL-1, IL-2, IL-3, IL-4, IL-6, Steel factor (SCF), leukemia inhibitory factor (LIF), or macrophage or granulocyte colony-stimulating factors (M-CSF or G-CSF). The failure of recombinant IL-7 to maintain the long-term growth of NAG8/7 cells and the inability of anti-IL-7 antibodies to significantly affect the response of either NAG8/7 cells or thymocytes to CM suggested the presence of one or more other cytokines in the CM. Analysis of concentrated CM fractionated by anion exchange chromatography revealed a single peak of activity in the NAG8/7 assay with an elution profile that was distinct from IL-7. Two peaks of activity were detected in the thymocyte response to anti-CD3 antibodies; one corresponded to IL-7 and the other corresponded to the same fractions that stimulated NAG8/7 cells. The second peak of thymocyte stimulatory activity could not be inhibited by neutralizing anti-IL-7 antibodies. In addition to producing a cytokine with unique properties, this thymic stromal cell exhibits a functional homology to bone marrow or fetal liver stromal cells not previously appreciated.

Oncostatin M transforms lymphoid tissue function in transgenic mice by stimulating lymph node T-cell development and thymus autoantibody production.

Oncostatin M (OM) is a member of the IL-6 subfamily of cytokines that is expressed in primary lymphoid tissues such as bone marrow and thymus, as well as in secondary lymphoid tissues and activated leukocytes. We produced transgenic mice that overexpressed the human, bovine, or mouse OM genes and compared their relative ability to modulate lymphopoiesis. Each species of cytokine induced a similar extrathymic pathway of T-cell development involving the accumulation of immature T cells within lymph nodes. Reconstitution experiments utilizing lethally irradiated athymic mice indicated that OM had caused hematopoietic precursors within fetal liver and bone marrow to initiate lymph node T-cell development in the absence of a thymic environment. Breeding experiments with IL6-/- and IL-7r(alpha)-/- deficient mice, indicated that induction of this extrathymic pathway by the OM transgene occurred in the absence of IL-6, but was strictly dependent on IL-7 receptor signaling. Separately, OM stimulated the accumulation of immature B cells within the transgenic thymus and caused the subcapsular regions of the thymus to expand with mature B and T cells. This thymus conversion to secondary lymphoid tissue was responsible for a lethal autoimmune-like disease marked by high titers of circulating autoantibodies, proteinuria, and glomerulonephritis. The conserved phenotypes elicited by these three forms of OM indicate that this potent hematopoietic cytokine can regulate lymphoid tissue function and morphogenesis.

Thymic stromal lymphopoietin: a cytokine that promotes the development of IgM+ B cells in vitro and signals via a novel mechanism.

A novel cytokine from a thymic stromal cell line (thymic stromal lymphopoietin (TSLP)) promotes the development of B220+/IgM+ immature B cells when added to fetal liver cultures, long term bone marrow cultures, or bone marrow cells plated in semisolid medium. Because the activities of TSLP overlap with those of IL-7 in some in vitro assays, we compared the signaling mechanisms employed by TSLP and IL-7. Proliferation of a factor-dependent pre-B cell line (NAG8/7) in response to either TSLP or IL-7 was inhibited by anti-IL-7R alpha mAbs, suggesting that the functional TSLP receptor complex uses IL-7R alpha. In contrast, three different Abs to the common cytokine receptor gamma-chain had no effect on the response of these cells to TSLP, indicating that the functional TSLP receptor complex does not use the common cytokine receptor gamma-chain. Both cytokines induced activation of Stat5, but only IL-7 induced activation of the Janus family kinases Jak1 and Jak3. In fact, TSLP failed to activate any of the four known Janus family kinases, suggesting that Stat5 phosphorylation is mediated by a novel mechanism. Taken together, these data support the idea that TSLP can make unique contributions to B lymphopoiesis and indicate that it does so by mechanisms distinct from IL-7.