The in vivo requirements for human natural killer (NK) cell development and differentiation into cytotoxic effectors expressing inhibitory receptors for self–major histocompatability complex class I (MHC-I; killer Ig-like receptors [KIRs]) remain undefined. Here, we dissect the role of interleukin (IL)-15 in human NK cell development using Rag2−/−γc−/− mice transplanted with human hematopoietic stem cells. Human NK cell reconstitution was intrinsically low in this model because of the poor reactivity to mouse IL-15. Although exogenous human IL-15 (hIL-15) alone made little improvement, IL-15 coupled to IL-15 receptor α (IL-15Rα) significantly augmented human NK cells. IL-15–IL-15Rα complexes induced extensive NK cell proliferation and differentiation, resulting in accumulation of CD16+KIR+ NK cells, which was not uniquely dependent on enhanced survival or preferential responsiveness of this subset to IL-15. Human NK cell differentiation in vivo required hIL-15 and progressed in a linear fashion from CD56hiCD16−KIR− to CD56loCD16+KIR−, and finally to CD56loCD16+KIR+. These data provide the first evidence that IL-15 trans-presentation regulates human NK cell homeostasis. Use of hIL-15 receptor agonists generates a robust humanized immune system model to study human NK cells in vivo. IL-15 receptor agonists may provide therapeutic tools to improve NK cell reconstitution after bone marrow transplants, enhance graft versus leukemia effects, and increase the pool of IL-15–responsive cells during immunotherapy strategies.

Consistent with their function in immune surveillance, natural killer (NK) cells are distributed throughout lymphoid and nonlymphoid tissues. However, the mechanisms governing the steady-state trafficking of NK cells remain unknown. The lysophospholipid sphingosine 1-phosphate (S1P), by binding to its receptor S1P1, regulates the recirculation of T and B lymphocytes. In contrast, S1P5 is detected in the brain and regulates oligodendrocyte migration and survival in vitro. Here we show that S1P5 was also expressed in NK cells in mice and humans and that S1P5-deficient mice had aberrant NK cell homing during steady-state conditions. In addition, we found that S1P5 was required for the mobilization of NK cells to inflamed organs. Our data emphasize distinct mechanisms regulating the circulation of various lymphocyte subsets and raise the possibility that NK cell trafficking may be manipulated by therapies specifically targeting S1P5.

Interleukin-15 (IL-15) is crucial for the generation of multiple lymphocyte subsets (natural killer (NK), NK-T cells, and memory CD8 T cells), and transpresentation of IL-15 by monocytes and dendritic cells has been IL-153 is a cytokine that was originally described, like IL-2, as a T cell growth factor (1). Both cytokines belong to the four ␣-helix bundle family, and their membrane receptors share two subunits (the IL-2R/ IL-15R ␤ and ␥ chains) responsible for signal transduction (2). The IL-2R␤/␥ complex is an intermediate affinity receptor for both cytokines that is expressed by most NK cells and can be activated in vitro by nanomolar concentrations of IL-2 or IL-15. The high affinity IL-2 and IL-15 receptors, such as those expressed on activated T cells, can be activated with picomolar concentrations of either cytokine, and additionally contain their own private ␣ chain (IL-2R␣ and IL-15R␣) that confers cytokine specificity and enhances the affinity of cytokine binding (3).Both cytokines play pivotal roles in innate and adaptative immunity. Whereas initial in vitro experiments have shown a large functional overlap in the effects of the two cytokines (induction of the proliferation and cytotoxicity of activated lymphocytes and NK cells, co-stimulation of B cell proliferation and immunoglobulin synthesis, and chemoattraction of T cells) (1, 4 -6), more recent experiments have indicated that they can exert complementary or even contrasting actions in vivo. Whereas IL-2 or IL-2R␣ knock-out in mice was associated with autoimmune phenotypes with increased populations of activated T and B cells, IL-15 and IL-15R␣ knock-out resulted in specific defects in NK, NK-T, intraepithelial lymphocytes, and memory CD8 T cells (7,8). Furthermore, IL-2 promotes peripheral tolerance by inducing activation-induced cell death, whereas IL-15 inhibits IL-2-mediated activation-induced cell death (9), and, unlike IL-2, IL-15 is a survival factor for CD8 memory T cells (10). In line with these observations, it has been suggested that the major role of IL-2 is to limit continuous expansion of activated T cells, whereas IL-15 is critical for initiation of T cell division and survival of memory T cells (11). A novel mechanism of IL-15 action described recently is that of transpresentation in which IL-15 and IL-15R␣ are coordinately expressed by antigen-presenting cells (monocytes and dendritic cells), and IL-15 bound to IL-15R␣ is presented in trans to neighboring NK or CD8 T cells expressing only the IL-15R␤/␥ receptor (12). As a co-stimulatory event occurring at the immunological synapse, IL-15 transpresentation now appears to be a dominant mechanism for IL-15 action in vivo (13, 14) and appears to play a major role in tumor immunosurveillance (15).The IL-15R␣ and IL-2R␣ subunits form a sub-family of cytokine receptors in that they comprise extracellular parts, so called "sushi" structural domains (one in IL-15R␣ and two in IL-2R␣), at their N terminus, which are also found in complement or adhesion molecules (16). In both cases, these ...

Adsorption of plasma protein decreases urinary protein excretion in patients with recurrence of the nephrotic syndrome after renal transplantation. Studies of the adsorbed proteins should provide information about the mechanism of this disease.

The interaction of interleukin-2 (IL-2) and IL-2 receptors critically regulates the T-cell immune response following antigen activation. IL-2 can signal through high or intermediate affinity receptors which contain IL-2R alpha (refs 3, 4) +beta (refs 5-8) +gamma (ref. 9) or beta+gamma chains, respectively. IL-2R gamma is a common gamma chain, gamma c, also shared by the IL-7 (ref. 10) and IL-4 (refs 11, 12) receptors, which when mutated results in X-linked severe combined immunodeficiency. Using chimaeric receptor constructs together with monoclonal or bispecific antibodies we demonstrate here that IL-2 signalling requires ligand-induced extracellular-domain-mediated heterodimerization of the beta- and gamma c-chain cytoplasmic domains. Anti-IL-2R alpha monoclonal antibodies trigger proliferation of cells transfected with chimaeric constructs in which the extracellular domains of IL-2R beta and gamma c are replaced by that of IL-2R alpha. Other experiments using chimaeric constructs indicated that IL-2 binds monomerically and monovalently to IL-2R alpha and that the beta-transmembrane domain is not required for receptor chain interactions. Finally, we provide a method for mapping residues in the gamma c cytoplasmic domain even in cells that constitutively express gamma c.

IL-15 and IL-2 are two structurally and functionally related cytokines whose high affinity receptors share the IL-2R β-chain and γ-chain in association with IL-15R α-chain (IL-15Rα) or IL-2R α-chain, respectively. Whereas IL-2 action seems restricted to the adaptative T cells, IL-15 appears to be crucial for the function of the innate immune responses, and the pleiotropic expression of IL-15 and IL-15Rα hints at a much broader role for the IL-15 system in multiple cell types and tissues. In this report, using a highly sensitive radioimmunoassay, we show the existence of a soluble form of human IL-15Rα (sIL-15Rα) that arises from proteolytic shedding of the membrane-anchored receptor. This soluble receptor is spontaneously released from IL-15Rα-expressing human cell lines as well as from IL-15Rα transfected COS-7 cells. This release is strongly induced by PMA and ionomycin, and to a lesser extent by IL-1β and TNF-α. The size of sIL-15Rα (42 kDa), together with the analysis of deletion mutants in the ectodomain of IL-15Rα, indicates the existence of cleavage sites that are proximal to the plasma membrane. Whereas shedding induced by PMA was abrogated by the synthetic matrix metalloproteinases inhibitor GM6001, the spontaneous shedding was not, indicating the occurrence of at least two distinct proteolytic mechanisms. The sIL-15Rα displayed high affinity for IL-15 and behaved as a potent and specific inhibitor of IL-15 binding to the membrane receptor, and of IL-15-induced cell proliferation (IC50 in the range from 3 to 20 pM). These results suggest that IL-15Rα shedding may play important immunoregulatory functions.

Interleukin (IL)-15 has an important role in tumor immunosurveillance and has a contemplated use in tumor immunotherapy. We have previously engineered the fusion protein RLI, composed of the NH 2 -terminal (amino acids 1-77, sushi+) domain of IL-15 receptor α coupled via a linker to IL-15, and shown that it displayed far better efficacy than IL-15 in vitro. In this report, we investigated in vivo whether RLI would be a better alternative than IL-15 and IL-2 for cancer treatment using two distinct animal models. B16F10 mouse melanoma cells were injected in C57BL/6 mice either i.v. or intrasplenically for lung or liver metastasis, respectively. HCT-116 human colorectal cancer cells were injected in the cecum of nude mice. We show that RLI has a higher efficiency than IL-15 or IL-2 to reduce lung and liver metastasis and enhance survival in the mouse B16F10 melanoma model, a result that was associated with a higher half-life in vivo. We also found that the antitumoral effect of RLI was completely abolished by in vivo depletion of natural killer cells using anti-asialoGM1 antibody. Moreover, RLI was also efficient to reduce by 50% tumor growth and the progression of metastasis of human colon carcinoma cells in an orthotopic nude mouse model. The fusion protein RLI has revealed strong anticancer effect in two different cancer models overcoming the limited effect of IL-15 by increasing its bioavailability and efficiency. These findings hold significant importance for the use of RLI as a potential adjuvant/therapeutic. [Mol Cancer Ther 2009;8(9):2736-45]

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