The transcription factor STAT5 is critical in dendritic cells for the development of TH2 but not TH1 responses.

Dendritic cells (DCs) are critical in immune responses, linking innate and adaptive immunity. We found here that DC-specific deletion of the transcription factor STAT5 was not critical for development but was required for T helper type 2 (TH2), but not TH1, allergic responses in both the skin and lungs. Loss of STAT5 in DCs led to the inability to respond to thymic stromal lymphopoietin (TSLP). STAT5 was required for TSLP-dependent DC activation, including upregulation of the expression of costimulatory molecules and chemokine production. Furthermore, TH2 responses in mice with DC-specific loss of STAT5 resembled those seen in mice deficient in the receptor for TSLP. Our results show that the TSLP-STAT5 axis in DCs is a critical component for the promotion of type 2 immunity at barrier surfaces.

Targeting myostatin/activin A protects against skeletal muscle and bone loss during spaceflight.

Among the physiological consequences of extended spaceflight are loss of skeletal muscle and bone mass. One signaling pathway that plays an important role in maintaining muscle and bone homeostasis is that regulated by the secreted signaling proteins, myostatin (MSTN) and activin A. Here, we used both genetic and pharmacological approaches to investigate the effect of targeting MSTN/activin A signaling in mice that were sent to the International Space Station. Wild type mice lost significant muscle and bone mass during the 33 d spent in microgravity. Muscle weights of Mstn-/- mice, which are about twice those of wild type mice, were largely maintained during spaceflight. Systemic inhibition of MSTN/activin A signaling using a soluble form of the activin type IIB receptor (ACVR2B), which can bind each of these ligands, led to dramatic increases in both muscle and bone mass, with effects being comparable in ground and flight mice. Exposure to microgravity and treatment with the soluble receptor each led to alterations in numerous signaling pathways, which were reflected in changes in levels of key signaling components in the blood as well as their RNA expression levels in muscle and bone. These findings have implications for therapeutic strategies to combat the concomitant muscle and bone loss occurring in people afflicted with disuse atrophy on Earth as well as in astronauts in space, especially during prolonged missions.

CD4+ Group 1 Innate Lymphoid Cells (ILC) Form a Functionally Distinct ILC Subset That Is Increased in Systemic Sclerosis.

Innate lymphoid cells (ILC) are a heterogeneous group of cellular subsets that produce large amounts of T cell-associated cytokines in response to innate stimulation in the absence of Ag. In this study, we define distinct patterns of surface marker and cytokine expression among the ILC subsets that may further delineate their migration and function. Most notably, we found that the subset previously defined as group 1 ILC (ILC1) contains CD4(+) CD8(-), CD4(-) CD8(+), and CD4(-) CD8(-) populations. Although all ILC1 subsets shared characteristics with Th1 cells, CD4(+) ILC1 also demonstrated significant phenotypic and functional heterogeneity. We also show that the frequencies of CD4(+) ILC1 and NKp44(+) group 3 ILC, but not CD4(-) ILC1 or group 2 ILC, are increased in the peripheral blood of individuals with systemic sclerosis (SSc), a disease characterized by fibrotic and vascular pathology, as well as immune dysregulation. Furthermore, we demonstrate that CD4(+) and CD4(-) ILC1 are functionally divergent based on their IL-6Rα expression and that the frequency of IL-6Rα expression on ILC is altered in SSc. The distinct phenotypic and functional features of CD4(+) and CD4(-) ILC1 suggest that they may have differing roles in the pathogenesis of immune-mediated diseases, such as SSc.

Transcriptional regulation of IL-15 expression during hematopoiesis.

Dendritic cells (DCs) are the most commonly studied source of the cytokine IL-15. Using an IL-15 reporter transgenic mouse, we have recently shown previously unappreciated differences in the levels of IL-15 expressed by subsets of conventional DCs (CD8⁺ and CD8⁻). In this study, we show that IL-15 promoter activity was differentially regulated in subsets of hematopoietically derived cells with IL-15 expression largely limited to myeloid lineages. In contrast, mature cells of the lymphoid lineages expressed little to no IL-15 activity. Surprisingly, we discovered that hematopoietic stem cells (lineage⁻Sca-1⁺c-Kit⁺) expressed high levels of IL-15, suggesting that IL-15 expression was extinguished during lymphoid development. In the case of T cells, this downregulation was Notch-dependent and occurred in a stepwise pattern coincident with increasing maturation and commitment to a T cell fate. Finally, we further demonstrate that IL-15 expression was also controlled throughout DC development, with key regulatory activity of IL-15 production occurring at the pre-DC branch point, leading to the generation of both IL-15⁺CD8⁺ and IL-15(⁻/low)CD8⁻ DC subsets. Thus, IL-15 expression is coordinated with cellular fate in myeloid versus lymphoid immune cells.

Cutting edge: the role of IFN-α receptor and MyD88 signaling in induction of IL-15 expression in vivo.

IL-15 plays a multifaceted role in immune homeostasis, but the unreliability of IL-15 detection has stymied exploration of IL-15 regulation in vivo. To visualize IL-15 expression, we created a transgenic mouse expressing emerald-GFP (EmGFP) under IL-15 promoter control. EmGFP/IL-15 was prevalent in innate cells including dendritic cells (DCs), macrophages, and monocytes. However, DC subsets expressed varying levels of EmGFP/IL-15 with CD8(+) DCs constitutively expressing EmGFP/IL-15 and CD8(-) DCs expressing low EmGFP/IL-15 levels. Virus infection resulted in IL-15 upregulation in both subsets. By crossing the transgenic mice to mice deficient in specific elements of innate signaling, we found a cell-intrinsic dependency of DCs and Ly6C(+) monocytes on IFN-α receptor expression for EmGFP/IL-15 upregulation after vesicular stomatitis virus infection. In contrast, myeloid cells did not require the expression of MyD88 to upregulate EmGFP/IL-15 expression. These findings provide evidence of previously unappreciated regulation of IL-15 expression in myeloid lineages during homeostasis and following infection.

Mapping the genetic landscape establishing a tumor immune microenvironment favorable for anti-PD-1 response in mice and humans.

Identifying host genetic factors modulating immune checkpoint inhibitor (ICI) efficacy has been experimentally challenging because of variations in both host and tumor genomes, differences in the microbiome, and patient life exposures. Utilizing the Collaborative Cross (CC) multi-parent mouse genetic resource population, we developed an approach that fixes the tumor genomic configuration while varying host genetics. With this approach, we discovered that response to anti-PD-1 (aPD1) immunotherapy was significantly heritable in four distinct murine tumor models (H2 between 0.18-0.40). For the MC38 colorectal carcinoma system (H2 = 0.40), we mapped four significant ICI response quantitative trait loci (QTL) localized to mouse chromosomes (mChr) 5, 9, 15 and 17, and identified significant epistatic interactions between specific QTL pairs. Differentially expressed genes within these QTL were highly enriched for immune genes and pathways mediating allograft rejection and graft vs host disease. Using a cross species analytical approach, we found a core network of 48 genes within the four QTLs that showed significant prognostic value for overall survival in aPD1 treated human cohorts that outperformed all other existing validated immunotherapy biomarkers, especially in human tumors of the previously defined immune subtype 4. Functional blockade of two top candidate immune targets within the 48 gene network, GM-CSF and high affinity IL-2/IL-15 signaling, completely abrogated the MC38 tumor transcriptional response to aPD1 therapy in vivo. Thus, we have established a powerful cross species in vivo platform capable of uncovering host genetic factors that establish the tumor immune microenvironment configuration propitious for ICI response.

CD4+ T cell regulation of CD25 expression controls development of short-lived effector CD8+ T cells in primary and secondary responses.

Both CD4(+) T cell help and IL-2 have been postulated to "program" activated CD8(+) T cells for memory cell development. However, the linkage between these two signals has not been well elucidated. Here we have studied effector and memory CD8(+) T cell differentiation following infection with three pathogens (Listeria monocytogenes, vesicular stomatitis virus, and vaccinia virus) in the absence of both CD4(+) T cells and IL-2 signaling. We found that expression of CD25 on antigen-specific CD8(+) T cells peaked 3-4 days after initial priming and was dependent on CD4(+) T cell help, likely through a CD28:CD80/86 mediated pathway. CD4(+) T cell or CD25-deficiency led to normal early effector CD8(+) T cell differentiation, but a subsequent lack of accumulation of CD8(+) T cells resulting in overall decreased memory cell generation. Interestingly, in both primary and recall responses KLRG1(high) CD127(low) short-lived effector cells were drastically diminished in the absence of IL-2 signaling, although memory precursors remained intact. In contrast to previous reports, upon secondary antigen encounter CD25-deficient CD8(+) T cells were capable of undergoing robust expansion, but short-lived effector development was again impaired. Thus, these results demonstrated that CD4(+) T cell help and IL-2 signaling were linked via CD25 up-regulation, which controls the expansion and differentiation of antigen-specific effector CD8(+) T cells, rather than "programming" memory cell traits.

MHC class I and TCR avidity control the CD8 T cell response to IL-15/IL-15Rα complex.

IL-15 operates via a unique mechanism termed transpresentation. In this system, IL-15 produced by one cell type is bound to IL-15Rα expressed by the same cell and is presented to apposing cells expressing the IL-15Rß/γC complex. We have shown that administering soluble IL-15Rα complexed with IL-15 can greatly enhance IL-15 activity. We now show that the naive CD8 T cell response to exogenous IL-15/IL-15Rα complex is MHC class I dependent. In the absence of ß2 microglobulin, naive CD8 T cells scarcely proliferated in response to IL-15/IL-15Rα complex, whereas memory cells proliferated, although to a lesser extent, compared with levels in control mice. The loss of ß2m or FcRn slightly reduced the extended half-life of IL-15/IL-15Rα complex, whereas FcRn deficiency only partially reduced the naive CD8 T cell proliferative response to IL-15/IL-15Rα complex. In addition, we demonstrated a link between TCR avidity and the ability of a T cell to respond to IL-15/IL-15Rα complex. Thus, T cells expressing low-avidity TCR responded poorly to IL-15/IL-15Rα complex, which correlated with a poor homeostatic proliferative response to lymphopenia. The inclusion of cognate peptide along with complex resulted in enhanced proliferation, even when TCR avidity was low. IL-15/IL-15Rα complex treatment, along with peptide immunization, also enhanced activation and the migratory ability of responding T cells. These data suggest that IL-15/IL-15Rα complex has selective effects on Ag-activated CD8 T cells. Our findings have important implications for directing IL-15/IL-15Rα complex-based therapy to specific Ag targets and illustrate the possible adjuvant uses of IL-15/IL-15Rα complex.

The biology of thymic stromal lymphopoietin (TSLP).

Originally shown to promote the growth and activation of B cells, thymic stromal lymphopoietin (TSLP) is now known to have wide-ranging impacts on both hematopoietic and nonhematopoietic cell lineages, including dendritic cells, basophils, eosinophils, mast cells, CD4⁺, CD8⁺ and natural killer T cells, B cells and epithelial cells. While TSLP's role in the promotion of TH2 responses has been extensively studied in the context of lung- and skin-specific allergic disorders, it is becoming increasingly clear that TSLP may impact multiple disease states within multiple organ systems, including the blockade of TH1/TH17 responses and the promotion of cancer and autoimmunity. This chapter will highlight recent advances in the understanding of TSLP signal transduction, as well as the role of TSLP in allergy, autoimmunity and cancer. Importantly, these insights into TSLP's multifaceted roles could potentially allow for novel therapeutic manipulations of these disorders.

The multiple facets of thymic stromal lymphopoietin (TSLP) during allergic inflammation and beyond.

Originally shown to promote the growth and activation of B cells, TSLP is now known to have wide-ranging impacts on hematopoietic and nonhematopoietic cell lineages, including DCs, basophils, eosinophils, mast cells, CD4(+), CD8(+), and NK T cells, B cells, and epithelial cells. Whereas the role of TSLP in the promotion of TH2 responses has been studied extensively in the context of lung- and skin-specific allergic disorders, it is becoming increasingly clear that TSLP may impact multiple disease states within multiple organ systems, including the blockade of TH1/TH17 responses and the promotion of cancer and autoimmunity. This review will highlight recent advances in the understanding of TSLP signal transduction, as well as the role of TSLP in allergy, autoimmunity, and cancer. Importantly, these insights into the multifaceted roles of TSLP could potentially allow for novel, therapeutic manipulations of these disorders.

Combined IL-15/IL-15Ralpha immunotherapy maximizes IL-15 activity in vivo.

IL-15 has substantial potential as an immunotherapeutic agent for augmenting immune responses. However, the activity of IL-15 is mediated by a unique mechanism in which the cytokine is transpresented by cell-bound high-affinity IL-15Ralpha to target cells expressing the IL-15Rbeta and the common gamma-chain. Thus, the efficacy of administered IL-15 alone may be limited by the availability of free IL-15Ralpha. We now show that administration of soluble IL-15/IL-15Ralpha complexes greatly enhanced IL-15 half-life and bioavailability in vivo. Treatment of mice with this complex, but not with IL-15 alone, resulted in robust proliferation of memory CD8 T cells, NK cells, and NK T cells. The activity of the complex required IL-15Rbeta, but not IL-15Ralpha, expression by the responding cells and was IL-7-independent. Interestingly, IL-15/IL-15Ralpha immunotherapy also caused naive CD8 T cell activation and development into effector cells and long-term memory T cells. Lastly, complexed IL-15, as compared with IL-15 alone, dramatically reduced tumor burden in a model of B16 melanoma. These findings hold significant importance for the use of IL-15 as a potential adjuvant/therapeutic and inducer of homeostatic proliferation, without the necessity for prior immunodepletion.