Mermagya Dhakal

FoxP3+RORγt+ T Helper Intermediates Display Suppressive Function against Autoimmune Diabetes

Recently, traces of double-positive FoxP3+RORγt+ T cells were identified and viewed as dual programming differentiation intermediates geared toward development into T regulatory or Th17 cells. In this study, we report that FoxP3+RORγt+ intermediates arise in the NOD mouse T cell repertoire prior to inflammation and can be expanded with tolerogen without further differentiation. Furthermore, FoxP3+RORγt+ cells express both CD62L and membrane-bound TGFβ and use the former to traffic to the pancreas and the latter to suppress effector T cells both in vitro and in vivo. The cells perform these functions as FoxP3+RORγt+ intermediates, despite being able to terminally differentiate into either FoxP3+RORγt− T regulatory or FoxP3−RORγt+ Th17 cells on polarization. These previously unrecognized observations extend plasticity to both differentiation and function and indicate that the intermediates are poised to traffic to sites of inflammation and target diverse pathogenic T cells, likely without prior conditioning by effector T cells, thus broadening efficacy against autoimmunity.

IL‐13Rα1 is a surface marker for M2 macrophages influencing their differentiation and function

In this study, we examined the role IL‐13 receptor alpha 1 (IL‐13Rα1) plays in macrophage differentiation and function. The findings indicate that IL‐13Rα1 is expressed on the M2 but not on the M1 subset of macrophages and specifically heterodimerizes with the IL‐4Rα chain to form a type II receptor, which controls the differentiation and function of these cells. Indeed, BM cells from IL‐13Rα1+/+ and IL‐13Rα1−/− mice yield equivalent numbers of macrophages when cultured under M2 polarizing conditions. However, IL‐13Rα1−/− BM cells yield a much higher number of macrophages than IL‐13Rα1+/+ BM cells when the differentiation is carried out under M1‐polarizing conditions. Further analyses indicated that macrophages that express IL‐13Rα1 also display surface markers associated with an M2 phenotype. In addition, the IL‐13Rα1+ macrophages were highly efficient in phagocytizing zymosan bioparticles both in vitro and in vivo, and supported differentiation of naïve T cells to a Th2 phenotype. Finally, when stimulated by IL‐13, a cytokine that uses the heteroreceptor, the cells were able to phosphorylate STAT6 efficiently. These previously unrecognized findings indicate that IL‐13Rα1 serves as a marker for M2 macrophages and the resulting heteroreceptor influences both their differentiation and function.

Mechanisms Underlying Antigen-Specific Tolerance of Stable and Convertible Th17 Cells During Suppression of Autoimmune Diabetes

Type 1 diabetes involves both T helper (Th)1 and Th17 cells. While the mechanisms underlying the control of Th1 cells are relatively well defined, those operating modulation of Th17 cells remain unknown. Moreover, given that Th17 cells are plastic and can drive disease as stable or convertible T cells, effective approaches to counter type 1 diabetes would have to alter Th17 function under both circumstances. Herein, we genetically incorporated the BDC2.5-reactive p79 mimotope into an Ig molecule, and the resulting Ig-p79 was used to investigate Th17 tolerance. Accordingly, diabetogenic BDC2.5 Th17 cells were transferred into NOD mice under convertible or stable conditions and their fate was evaluated upon induction of tolerance and disease suppression by Ig-p79. The findings show that convertible (Th17 to Th1) cells display downregulation of the chemokine (C-X-C motif) receptor 3 that was associated with diminished T-box transcription factor T-bet expression, retention in the spleen, and inhibition of trafficking to the pancreas. In contrast, stable Th17 cells downregulated orphan nuclear receptor ROR-γt but increased Fas ligand expression and died by apoptosis. Thus, the final signature transcription factor shapes the mechanism of tolerance in plastic Th17 cells. These findings suggest that effective strategies against type 1 diabetes will require regimens that could drive both mechanisms of tolerance to overcome the disease.

Bone Marrow-Derived IL-13Ra1-Positive Thymic Progenitors Are Restricted to the Myeloid Lineage

The earliest thymic progenitors (ETPs) were recently shown to give rise to both lymphoid and myeloid cells. Whereas the majority of ETPs are derived from IL-7Rα–positive cells and give rise exclusively to T cells, the origin of the myeloid cells remains undefined. In this study, we show both in vitro and in vivo that IL-13Rα1+ ETPs yield myeloid cells with no potential for maturation into T cells, whereas IL-13Rα1− ETPs lack myeloid potential. Moreover, transfer of lineage-negative IL-13Rα1+ bone marrow stem cells into IL-13Rα1–deficient mice reconstituted thymic IL-13Rα1+ myeloid ETPs. Myeloid cells or macrophages in the thymus are regarded as phagocytic cells whose function is to clear apoptotic debris generated during T cell development. However, the myeloid cells derived from IL-13Rα1+ ETPs were found to perform Ag-presenting functions. Thus, IL-13Rα1 defines a new class of myeloid restricted ETPs yielding APCs that could contribute to development of T cells and the control of immunity and autoimmunity.

Neonatal Basophils Stifle the Function of Early-Life Dendritic Cells To Curtail Th1 Immunity in Newborn Mice

Neonatal immunity exhibits weak Th1 but excessive Th2 responses, and the underlying mechanisms remain elusive. In this article, we show that neonatal basophils readily produce IL-4, a cytokine that proved to be pivotal in shaping the programs of both lymphocyte subsets. Besides promoting Th2 programs, IL-4 is captured by the IL-4 heteroreceptor (IL-4Rα/IL-13Rα1) expressed on dendritic cells and instigates IL-12 downregulation. Under these circumstances, differentiating Th1 cells upregulate IL-13Rα1, leading to an unusual expression of the heteroreceptor, which will serve as a death marker for these Th1 cells during rechallenge with Ag. The resulting Th1/Th2 imbalance impacts childhood immunity culminating in sensitivity to allergic reactions, susceptibility to microbial infection and perhaps poor efficacy of pediatric vaccines.

Developmental Expression of IL-12Rβ2 on Murine Naive Neonatal T Cells Counters the Upregulation of IL-13Rα1 on Primary Th1 Cells and Balances Immunity in the Newborn

Upon exposure to Ag on the day of birth, neonatal mice mount balanced primary Th1 and Th2 responses, with the former displaying upregulated IL-13Rα1 expression. This chain associates with IL-4Rα to form a heteroreceptor (IL-4Rα/IL-13Rα1) that marks the Th1 cells for death by IL-4 produced by Th2 cells during rechallenge with Ag, hence the Th2 bias of murine neonatal immunity. The upregulation of IL-13Rα1 on neonatal Th1 cells was due to the paucity of IL-12 in the neonatal environment. In this study, we show that by day 8 after birth, naive splenic T cells are no longer susceptible to IL-13Rα1 upregulation even when exposed to Ag within the neonatal environment. Furthermore, during the 8-d lapse, the naive splenic T cells spontaneously and progressively upregulate the IL-12Rβ2 chain, perhaps due to colonization by commensals, which induce production of IL-12 by cells of the innate immune system such as dendritic cells. In fact, mature T cells from the thymus, a sterile environment not accessible to microbes, did not upregulate IL-12Rβ2 and were unable to counter IL-13Rα1 upregulation. Finally, the 8-d naive T cells were able to differentiate into Th1 cells even independently of IL-12 but required the cytokine to counter upregulation of IL-13Rα1. Thus, in neonatal mice, IL-12, which accumulates in the environment progressively, uses IL-12Rβ2 to counter IL-13Rα1 expression in addition to promoting Th1 differentiation.

Recovery From Overt Type 1 Diabetes Ensues When Immune Tolerance and β-Cell Formation Are Coupled With Regeneration of Endothelial Cells in the Pancreatic Islets

Immune modulation of pancreatic inflammation induces recovery from type 1 diabetes (T1D), but remission was not durable, perhaps because of an inability to sustain the formation and function of new pancreatic β-cells. We have previously shown that Ig-GAD2, carrying GAD 206–220 peptide, induced in hyperglycemic mice immune modulation that was able to control pancreatic inflammation, stimulate β-cell regeneration, and prevent T1D progression. Herein, we show that the same Ig-GAD2 regimen given to mice with overt T1D was unable to reverse the course of disease despite eradication of Th1 and Th17 cells from the pancreas. However, the regimen was able to sustain recovery from T1D when Ig-GAD2 was accompanied with transfer of bone marrow (BM) cells from healthy donors. Interestingly, alongside immune modulation, there was concomitant formation of new β-cells and endothelial cells (ECs) in the pancreas. The new β-cells were of host origin while the donor BM cells gave rise to the ECs. Moreover, transfer of purified BM endothelial progenitors instead of whole BM cells sustained both β-cell and EC formation and reversal of diabetes. Thus, overcoming T1D requires both immune modulation and repair of the islet vascular niche to preserve newly formed β-cells.

APCs expressing high levels of programmed death ligand 2 sustain the development of CD4 T cell memory

The role APCs play in the transition of T cells from effector to memory remains largely undefined. This is likely due to the low frequency at which long-lived T cells arise, which hinders analysis of the events involved in memory development. In this study, we used TCR transgenic T cells to increase the frequency of long-lived T cells and developed a transfer model suitable for defining the contribution of APCs to the development of CD4 T cell memory. Accordingly, naive TCR transgenic T cells were stimulated in vitro with Ag presented by different types of APCs and transferred into MHC class II-deficient mice for parking, and the hosts were later analyzed for long-lived T cell frequency or challenged with suboptimal dose of Ag, and the long-lived cells-driven memory responses were measured. The findings indicate that B cells and CD8α+ dendritic cells sustained elevated frequencies of long-lived T cells that yielded rapid and robust memory responses upon rechallenge with suboptimal dose of Ag. Furthermore, both types of APCs had significant programmed death (PD) ligand 2 expression prior to Ag stimulation, which was maintained at a high level during presentation of Ag to T cells. Blockade of PD ligand 2 interaction with its receptor PD-1 nullified the development of memory responses. These previously unrecognized findings suggest that targeting specific APCs for Ag presentation during vaccination could prove effective against microbial infections.

Antigen-Specific Effector CD4 T Lymphocytes School Lamina Propria Dendritic Cells To Transfer Innate Tolerance

Dendritic cells (DCs) have been shown to play a major role in oral tolerance, and this function has been associated with their ability to produce anti-inflammatory cytokines and to induce suppressive regulatory T cells. In this study, we demonstrate that upon oral administration of Ag, lamina propia (LP) DCs engage specific T cells and acquire a novel mechanism by which they transfer tolerance against diverse T cell specificities. Indeed, when Ig-myelin oligodendrocyte glycoprotein (MOG) carrying the MOG35–55 epitope was orally administered into either T cell–sufficient or –deficient mice, only the T cell–sufficient hosts yielded CD8α+ and CD8α− LP DCs that were able to transfer tolerance to a variety of MHC class II–restricted effector T cells. Surprisingly, these LP DCs upregulated programmed cell death ligand 1 during the initial interaction with MOG-specific T cells and used this inhibitory molecule to suppress activation of T cells regardless of Ag specificity. Furthermore, oral Ig-MOG was able to overcome experimental autoimmune encephalomyelitis induced with CNS homogenate, indicating that the DCs are able to modulate disease involving diverse T cell specificities. This previously unrecognized attribute potentiates DCs against autoimmunity.

Antigen-Free Adjuvant Assists Late Effector CD4 T Cells to Transit to Memory in Lymphopenic Hosts

The events controlling the transition of T cells from effector to memory remain largely undefined. Many models have been put forth to account for the origin of memory precursors, but for CD4 T cells initial studies reported that memory T cells derive from IFN-γ–nonproducing effectors, whereas others suggested that memory emanates from highly activated IFN-γ–producing effectors. In this study, using cell proliferation, expression of activation markers, and production of IFN-γ as a measure of activation, we defined two types of effector CD4 T cells and investigated memory generation. The moderately activated early effectors readily transit to memory, whereas the highly activated late effectors, regardless of their IFN-γ production, develop minimal memory. Boosting with Ag-free adjuvant, however, rescues late effectors from cell death and sustains both survival and IFN-γ cytokine responses in lymphopenic hosts. The adjuvant-mediated memory transition of late effectors involves the function of TLRs, most notably TLR9. These findings uncover the mechanism by which late effector CD4 T cells are driven to transit to memory and suggest that timely boosts with adjuvant may enhance vaccine efficacy.