Search

Regulatory T cells (Tregs) are produced in the thymus to establish self-tolerance, and agonistic stimuli by self-Ags play a pivotal role in this process. Although two types of APCs, medullary thymic epithelial cells (mTECs) and dendritic cells (DCs), are responsible for presenting self-Ags together with costimulatory/cytokine signals, the distinct role of each APC in producing Tregs remains enigmatic. We have approached this issue by depleting the mTECs and DCs using mice expressing diphtheria toxin receptors driven by Aire and CD11c promoters, respectively. Depletion of mTECs showed an effect on Treg production quantitatively and qualitatively more profound than that of DCs followed by the development of distinct organ-specific autoimmune lesions in the hosts. Because self-Ags produced by mTECs are transferable to DCs through a process known as Ag transfer, we monitored the process of Ag transfer using mice expressing GFP from TECs. Although GFP expressed from total TECs was effectively transferred to DCs, GFP expressed from cortical TECs was not, suggesting that mTECs are the predominant source of self-Ags. We also found that GFP expressed not only from mature mTECs but also from immature mTECs was transferred to DCs, suggesting that a broad spectrum of molecules were subjected to Ag transfer during mTEC development. Interestingly, the numbers of recirculating non-Tregs producing IL-2, an important source for Treg expansion in the thymus, were reduced only in the mTEC-depleted mice. These results suggested the cooperative but distinct role of mTECs and DCs in the production of Tregs to avoid autoimmunity.

The conventional wet impregnation method yields nanoparticles of BaTi02.5H0.5 perovskite hydride on a TiH2 surface that works as a support for ammonia synthesis catalysts.

Aire, the defect of which is responsible for the development of autoimmunity, is predominantly expressed in medullary thymic epithelial cells, and it controls a wide variety of genes, including those of tissue-restricted Ags, for establishing thymic tolerance. Aire is also expressed from APCs in the periphery, called extrathymic Aire-expressing cells (eTACs), and their complementing role to thymic tolerance has been suggested. eTACs are composed of two distinct classes of APCs, conventional dendritic cell (cDC)–type and group 3 innate lymphoid cell (ILC3)-like–type expressing retinoic acid receptor–related orphan receptor γt (RORγt). Although the essential role of Aire in the latter in the Th17-mediated immune response against Candida albicans has been reported, the role of Aire in the cDC-type eTACs for this action has not been examined. Furthermore, the significance of Aire in the production of the transcriptome of the cDC-type eTACs remains unknown. We have approached these issues using a high-fidelity Aire-reporter mouse strain. We found that although the cDC-type eTACs dominated ILC3-like–type eTACs in number and they served as efficient APCs for the immune response against an exogenous Ag as well as for the C. albicans–specific Th17 immune response, loss of Aire in cDC-type eTACs showed no clear effect on these functions. Furthermore, loss of Aire showed no major impact on the transcriptome from cDC-type eTACs. These results suggested that Aire in cDC-type eTACs may not have a cell-intrinsic role in the immune response in contrast to the role of Aire in ILC3-like–type eTACs.

Ly6C comprises two homologous components of Ly6C1 and Ly6C2, and the expression of either of the Ly6C molecules defines unique functional subsets of monocytes. Ly6C is also expressed by other immune cell types, including Aire-expressing medullary thymic epithelial cells. Because the role of Ly6C expression in determining the functional subsets remains unclear, we generated mice deficient for both Ly6C1 and Ly6C2 with CRISPR-Cas9–mediated deletion. Mice deficient for Ly6C1/Ly6C2 showed no major alterations in the subsets and function of monocyte and other immune cells, including the cells involved in the dextran sulfate sodium salt–induced colitis model. By generating the mice deficient for Ly6C1 alone, we have also investigated the expression pattern of Ly6C1 and Ly6C2 in immune cells. Except for medullary thymic epithelial cells and CD4 single-positive T cells, immune cells predominantly expressed Ly6C2. Thus, despite the importance as a marker with a unique differential expression pattern, the Ly6C molecules have no major impact on determining the functional subsets and maintaining immune homeostasis.

Ru/Ce0.5La0.5−xTixO1.75+0.5x solid solutions with high Ce4+ reducibility and a large number of mesopores were synthesized and their ammonia synthesis activity was investigated.

Deficiency for AIRE/Aire in both humans and mice results in the development of organ-specific autoimmune disease. We tested whether augmented and/or dysregulated AIRE/Aire expression might be also prone to the breakdown of self-tolerance. To define the effect of augmented Aire expression on the development of autoimmunity, antigen-specific clonal deletion and production of clonotypic regulatory T cells (Tregs) in the thymus were examined using mice expressing two additional copies of Aire in a heterozygous state (3xAire-knockin mice: 3xAire-KI). We found that both clonal deletion of autoreactive T cells and production of clonotypic Tregs in the thymus from 3xAire-KI were impaired in a T-cell receptor-transgenic system. Furthermore, 3xAire-KI females showed higher scores of experimental autoimmune encephalomyelitis induced by myelin oligodendrocyte glycoprotein than wild-type littermates, suggesting that augmented Aire expression exacerbates organ-specific autoimmunity under disease-prone conditions. In humans, we found that one patient with amyopathic dermatomyositis showed CD3–CD19– cells expressing AIRE in the peripheral blood before the treatment but not during the remission phase treated with immunosuppressive drugs. Thus, not only loss of function of AIRE/Aire but also augmented and/or dysregulated expression of AIRE/Aire should be considered for the pathogenesis of organ-specific autoimmunity. We suggest that further analyses should be pursued to establish a novel link between organ-specific autoimmune disease and dysregulated AIRE expression in clinical settings.

Vγ6

Puel and Casanova and Kisand et al . challenge our conclusions that interferonopathy and not IL-17/IL-22 autoantibodies promote candidiasis in autoimmune polyendocrinopathy–candidiasis–ectodermal dystrophy. We acknowledge that conclusive evidence for causation is difficult to obtain in complex human diseases. However, our studies clearly document interferonopathy driving mucosal candidiasis with intact IL-17/IL-22 responses in Aire -deficient mice, with strong corroborative evidence in patients.

Hydrogen adsorption/desorption properties of magnesium oxalate and calcium oxalate (M(ox); M = Mg2+ and Ca2+) were measured at selected temperatures between 77 K and 200 K. The hydrogen uptake amou...

Medullary thymic epithelial cells (mTECs), which express a wide range of tissue-restricted Ags (TRAs), contribute to the establishment of self-tolerance by eliminating autoreactive T cells and/or inducing regulatory T cells. Aire controls a diverse set of TRAs within Aire-expressing cells by employing various transcriptional pathways. As Aire has a profound effect on transcriptomes of mTECs, including TRAs not only at the single-cell but also the population level, we suspected that Aire (Aire+ mTECs) might control the cellular composition of the thymic microenvironment. In this study, we confirmed that this is indeed the case by identifying a novel mTEC subset expressing Ly-6 family protein whose production was defective in Aire-deficient thymi. Reaggregated thymic organ culture experiments demonstrated that Aire did not induce the expression of Ly-6C/Ly-6G molecules from mTECs as Aire-dependent TRAs in a cell-intrinsic manner. Instead, Aire+ mTECs functioned in trans to maintain Ly-6C/Ly-6G+ mTECs. Thus, Aire not only controls TRA expression transcriptionally within the cell but also controls the overall composition of mTECs in a cell-extrinsic manner, thereby regulating the transcriptome from mTECs on a global scale.

Autoimmune regulator+ (Aire) medullary thymic epithelial cells (mTECs) play a critical role in tolerance induction. Several studies demonstrated that Aire+mTECs differentiate further into Post‐Aire cells. Yet, the identification of terminal stages of mTEC maturation depends on unique fate‐mapping mouse models. Herein, we resolve this limitation by segmenting the mTEChi(MHCIIhiCD80hi) compartment into mTECA/hi (CD24−Sca1−), mTECB/hi (CD24+Sca1−), and mTECC/hi (CD24+Sca1+). While mTECA/hi included mostly Aire‐expressing cells, mTECB/hi contained Aire+ and Aire− cells and mTECC/hi were mainly composed of cells lacking Aire. The differential expression pattern of Aire led us to investigate the precursor‐product relationship between these subsets. Strikingly, transcriptomic analysis of mTECA/hi, mTECB/hi, and mTECC/hi sequentially mirrored the specific genetic program of Early‐, Late‐ and Post‐Aire mTECs. Corroborating their Post‐Aire nature, mTECC/hi downregulated the expression of tissue‐restricted antigens, acquired traits of differentiated keratinocytes, and were absent in Aire‐deficient mice. Collectively, our findings reveal a new and simple blueprint to survey late stages of mTEC differentiation., Although the expression of Aire was initially considered to mark a terminal stage of mTEC maturation, recent studies have demonstrated that mTECs extend their differentiation beyond Aire. Integrating flow cytometry, functional, and transcriptomic analyses, Ferreirinha et al. provide an effective phenotypic‐based method to resolve Early‐, Late‐ and Post‐Aire mTECs.

Dehydration and rehydration behavior of a layered double hydroxide (LDH), Mg2Al(OH)6.2Cl0.8·nH2O, was investigated. The dehydration proceeds in two steps; at first interlayer water is released from 50 °C, then dehydration condensation of hydroxyl groups starts from 300 °C. Interlayer anion Cl− is also desorbed as HCl during the second dehydration. The mole fraction of water removed by dehydration before the HCl desorption was estimated by thermogravimetric analysis; 1.85 mol-H2O/mol-LDH. The dehydrated material, without HCl loss, was quantitatively hydrated by refluxing in water for 480 min.

Aire controls the fate of autoreactive thymocytes (i.e., clonal deletion or development into regulatory T cells [Tregs]) through transcriptional control of the expression of tissue-restricted self-antigens (TRAs) from medullary thymic epithelial cells (mTECs) and bone marrow (BM)-derived cells. Although TRAs expressed by mTECs and BM-derived cells are suggested to complement each other to generate a full spectrum of TRAs, little is known about the relative contribution of TRAs from each component for establishment of self-tolerance. Furthermore, the precise role of Aire in specific types of Aire-expressing APCs remains elusive. We have approached these issues by generating two different types of transgenic mouse (Tg) model, which express a prefixed model self-antigen driven by the insulin promoter or the Aire promoter. In the insulin-promoter Tg model, mTECs alone were insufficient for clonal deletion, and BM-derived APCs were required for this action by utilizing Ag transferred from mTECs. In contrast, mTECs alone were able to induce Tregs, although at a much lower efficiency in the absence of BM-derived APCs. Importantly, lack of Aire in mTECs, but not in BM-derived APCs, impaired both clonal deletion and production of Tregs. In the Aire-promoter Tg model, both mTECs and BM-derived APCs could independently induce clonal deletion without Aire, and production of Tregs was impaired by the lack of Aire in mTECs, but not in BM-derived APCs. These results suggest that the fate of autoreactive thymocytes together with the requirement for Aire depend on the cell types that express self-antigens and the types of APCs involved in tolerance induction.

Diabetes can be ameliorated by additive AIRE expression in peripheral DCs, Tissue-specific autoimmune diseases are assumed to arise through malfunction of two checkpoints for immune tolerance: defective elimination of autoreactive T cells in the thymus and activation of these T cells by corresponding autoantigens in the periphery. However, evidence for this model and the outcome of such alterations in each or both of the tolerance mechanisms have not been sufficiently investigated. We studied these issues by expressing human AIRE (huAIRE) as a modifier of tolerance function in NOD mice wherein the defects of thymic and peripheral tolerance together cause type I diabetes (T1D). Additive huAIRE expression in the thymic stroma had no major impact on the production of diabetogenic T cells in the thymus. In contrast, huAIRE expression in peripheral antigen-presenting cells (APCs) rendered the mice resistant to T1D, while maintaining other tissue-specific autoimmune responses and antibody production against an exogenous protein antigen, because of the loss of Xcr1+ dendritic cells, an essential component for activating diabetogenic T cells in the periphery. These results contrast with our recent demonstration that huAIRE expression in both the thymic stroma and peripheral APCs resulted in the paradoxical development of muscle-specific autoimmunity. Our results reveal that tissue-specific autoimmunity is differentially controlled by a combination of thymic function and peripheral tolerance, which can be manipulated by expression of huAIRE/Aire in each or both of the tolerance mechanisms.

The thymus is the organ dedicated to the generation of T cells, which are key effector cells in immune clearance of pathogens and tumours. However, T cells can also react to our own tissues or turn into cancer cells, such as in the case of autoimmunity and leukaemia, respectively. Therefore, the development and selection of T cells is a tightly regulated process that proceeds within inductive thymic microenvironments formed by cortical (c) and medullary (m) thymic epithelial cells (TECs). Herein, we critically summarize our current knowledge on the molecular principles underlying the development and diversification of TEC compartments and highlight their specialized roles in specific stages of T cell development. Knowledge in this area is of fundamental and clinical relevance to understand how the immune system reaches the equilibrium between immunity and tolerance induction.

mTEC progenitors differentiate to mature Aire+ mTECs through a pathway that initiates with RANK and LtβR signaling via the nonclassical NF-κB, followed by TRAF-6–driven maturation., Medullary thymic epithelial cells (mTECs) expressing autoimmune regulator (Aire) are critical for preventing the onset of autoimmunity. However, the differentiation program of Aire-expressing mTECs (Aire+ mTECs) is unclear. Here, we describe novel embryonic precursors of Aire+ mTECs. We found the candidate precursors of Aire+ mTECs (pMECs) by monitoring the expression of receptor activator of nuclear factor-κB (RANK), which is required for Aire+ mTEC differentiation. pMECs unexpectedly expressed cortical TEC molecules in addition to the mTEC markers UEA-1 ligand and RANK and differentiated into mTECs in reaggregation thymic organ culture. Introduction of pMECs in the embryonic thymus permitted long-term maintenance of Aire+ mTECs and efficiently suppressed the onset of autoimmunity induced by Aire+ mTEC deficiency. Mechanistically, pMECs differentiated into Aire+ mTECs by tumor necrosis factor receptor-associated factor 6-dependent RANK signaling. Moreover, nonclassical nuclear factor-κB activation triggered by RANK and lymphotoxin-β receptor signaling promoted pMEC induction from progenitors exhibiting lower RANK expression and higher CD24 expression. Thus, our findings identified two novel stages in the differentiation program of Aire+ mTECs.

Aire in medullary thymic epithelial cells (mTECs) plays an important role in the establishment of self-tolerance. Because Aire+ mTECs appear to be a limited subset, they may constitute a unique lineage(s) among mTECs. An alternative possibility is that all mTECs are committed to express Aire in principle, but Aire expression by individual mTECs is conditional. To investigate this issue, we established a novel Aire reporter strain in which endogenous Aire is replaced by the human AIRE-GFP-Flag tag (Aire/hAGF-knockin) fusion gene. The hAGF reporter protein was produced and retained very efficiently within mTECs as authentic Aire nuclear dot protein. Remarkably, snapshot analysis revealed that mTECs expressing hAGF accounted for >95% of mature mTECs, suggesting that Aire expression does not represent a particular mTEC lineage(s). We confirmed this by generating Aire/diphtheria toxin receptor–knockin mice in which long-term ablation of Aire+ mTECs by diphtheria toxin treatment resulted in the loss of most mature mTECs beyond the proportion of those apparently expressing Aire. These results suggest that Aire expression is inherent to all mTECs but may occur at particular stage(s) and/or cellular states during their differentiation, thus accounting for the broad impact of Aire on the promiscuous gene expression of mTECs.

Aire has been cloned as the gene responsible for a hereditary type of organ-specific autoimmune disease. Aire controls the expression of a wide array of tissue-restricted Ags by medullary thymic epithelial cells (mTECs), thereby leading to clonal deletion and Treg-cell production, and ultimately to the establishment of self-tolerance. However, relatively little is known about the mechanism responsible for the control of Aire expression itself. In this issue of the European Journal of Immunology, Haljasorg et al. [Eur. J. Immunol. 2015. 45: 3246-3256] have reported the presence of an enhancer element for Aire that binds with NF-κB components downstream of the TNF receptor family member, RANK (receptor activator of NF-κB). The results suggest that RANK has a dual mode of action in Aire expression: one involving the promotion of mTEC differentiation and the other involving activation of the molecular switch for Aire within mature mTECs.

Cortical thymic epithelial cells (cTECs) and medullary thymic epithelial cells (mTECs) play essential roles in the positive and negative selection of developing thymocytes, respectively. Aire in mTECs plays an essential role in the latter process through expression of broad arrays of tissue-restricted Ags. To determine whether the location of Aire within the medulla is absolutely essential or whether Aire could also function within the cortex for establishment of self-tolerance, we used bacterial artificial chromosome technology to establish a semiknockin strain of NOD-background (β5t/Aire-transgenic) mice expressing Aire under control of the promoter of β5t, a thymoproteasome expressed exclusively in the cortex. Although Aire was expressed in cTECs as typical nuclear dot protein in β5t/Aire-Tg mice, cTECs expressing Aire ectopically did not confer transcriptional expression of either Aire-dependent or Aire-independent tissue-restricted Ag genes. We then crossed β5t/Aire-Tg mice with Aire-deficient NOD mice, generating a strain in which Aire expression was confined to cTECs. Despite the presence of Aire+ cTECs, these mice succumbed to autoimmunity, as did Aire-deficient NOD mice. The thymic microenvironment harboring Aire+ cTECs, within which many Aire-activated genes were present, also showed no obvious alteration of positive selection, suggesting that Aire’s unique property of generating a self-tolerant T cell repertoire is functional only in mTECs.

Aire is predominantly expressed by medullary thymic epithelial cells (mTECs), and it is essential for maintaining self-tolerance by controlling the expression of tissue-restricted self-antigens (TRAs). Besides mTECs, extra-thymic Aire-expressing cells (eTACs) have been reported. However, the exact nature of eTACs has been difficult to study because of the small numbers together with low expression levels of Aire within the cells. We have recently established a novel Aire reporter strain in which endogenous Aire was replaced by the human AIRE-GFP-Flag tag fusion gene (Aire/hAGF-knockin: J. Immunol. 2015). hAGF-reporter protein was efficiently produced and retained within the mTECs as authentic Aire nuclear dot protein. With this high-sensitivity and high-fidelity Aire-reporter strain, we found that DCs in the spleen and lymph nodes expressed Aire although their expression levels were rather low. Aire-expressing DCs showed high levels of MHC-II and CD80/CD86, and half of which expressed CD8. In the thymus, we found that there were two types of DCs expressing Aire: hAGF-low DCs and hAGF-high DCs. Interestingly, hAGF-low DCs had small hAGF dots within the nuclei, whereas hAGF-high DCs retained larger hAGF dots which were mainly present outside the nuclei, suggesting that hAGF-high DCs had acquired hAGF protein from mTECs. Acquisition of hAGF protein by hAGF-high DCs from mTECs was confirmed by the bone-marrow transfer experiment. Mechanisms underlying the transfer of nuclear protein (i.e., hAGF protein) from mTECs to thymic DCs, and immunological consequence of this event are now under investigation.

Hassall’s corpuscles (HCs) are composed of cornifying, terminally differentiated medullary thymic epithelial cells (mTECs) that are developed under the control of Aire. Here, we demonstrated that HC-mTECs show features of cellular senescence and produce inflammatory cytokines and chemokines including CXCL5, thereby recruiting and activating neutrophils to produce IL-23 in the thymic medulla. We further indicated that thymic plasmacytoid dendritic cells (pDCs) expressing IL-23 receptors constitutively produced Ifna, which plays a role in single positive (SP) cell maturation, in an Il23a-dependent manner. Neutrophil depletion with anti-Ly6G antibody injection resulted in a significant decrease of Ifna expression in the thymic pDCs, suggesting that thymic neutrophil activation underlies the Ifna expression in thymic pDCs in steady state conditions. A New Zealand White mouse strain showing HC hyperplasia exhibited greater numbers and activation of thymic neutrophils and pDCs than B6 mice, whereas Aire-deficient B6 mice with defective HC development and SP thymocyte maturation showed significantly compromised numbers and activation of these cells. These results collectively suggested that HC-mTECs with cell-senescence features initiate a unique cell activation cascade including neutrophils and pDCs leading to the constitutive IFNα expression required for SP T-cell maturation in the thymic medulla.

Medullary thymic epithelial cells (mTECs), which express a wide range of tissue-restricted Ags (TRAs), contribute to the establishment of self-tolerance by eliminating autoreactive T cells and/or inducing regulatory T cells. Aire controls a diverse set of TRAs within Aire-expressing cells by employing various transcriptional pathways. As Aire has a profound effect on transcriptomes of mTECs, including TRAs not only at the single-cell but also the population level, we suspected that Aire (Aire

Essential roles of NF-κB–inducing kinase (NIK) for the development of medullary thymic epithelial cells (mTECs) and regulatory T cells have been highlighted by studies using a strain of mouse bearing a natural mutation of the NIK gene (aly mice). However, the exact mechanisms underlying the defect in thymic cross-talk leading to the breakdown of self-tolerance in aly mice remain elusive. In this study, we demonstrated that production of regulatory T cells and the final maturation process of positively selected conventional αβ T cells are impaired in aly mice, partly because of a lack of mature mTECs. Of note, numbers of thymic dendritic cells and their expression of costimulatory molecules were also affected in aly mice in a thymic stroma–dependent manner. The results suggest a pivotal role of NIK in the thymic stroma in establishing self-tolerance by orchestrating cross-talk between mTECs and dendritic cells as well as thymocytes. In addition, we showed that negative selection was impaired in aly mice as a result of the stromal defect, which accounts for the development of organ-specific autoimmunity through a lack of normal NIK.

Objective To show the role of the autoimmune regulator (Aire), a gene expressed in medullary thymic epithelial cells (mTEC), in the tolerance to encephalitogenic myelin epitopes. Methods C57BL/6J Aire-deficient and wild-type mice were immunized with myelin oligodendrocyte glycoprotein peptide (MOG35–55) or proteolipid protein peptide (PLP178–191). PLP178–191 is contained only in PLP/DM20, an isoform derived from a splice variant of PLP. We evaluated the development of experimental autoimmune encephalomyelitis (EAE) and reported the T cell response to these peptides. Results mTEC from wild-type mice expressed PLP/DM20, but those from Aire-deficient mice only expressed it at low levels. Immunization with PLP178–191 induced more severe EAE in Aire-deficient mice than in the wild-type mice. In contrast, MOG35–55 induced EAE of a similar grade in the wild-type and Aire-knockout mice. In recall response assays to PLP178–191, T cells from Aire-deficient mice produced significantly more interleukin (IL)-17 and interferon (IFN)-γ than the wild-type mice did. Adoptive transfer of CD4+ T cells purified from PLP178–191-immunized Aire-deficient mice induced a more severe EAE than a similar transfer from the immunized wild-type mice. In comparison with wild-type mice, we also found that sera from aged, naive Aire-deficient mice showed higher titers of PLP178–191-, but not MOG35–55-specific immunoglobulin G autoantibodies. Conclusion Aire is involved in establishing central tolerance to PLP178–191, but not to MOG35–55, and its deficiency induces spontaneous autoreactivity to PLP178–191.

Understanding the cellular dynamics of Aire-expressing lineage(s) among medullary thymic epithelial cells (AEL-mTECs) is essential for gaining insight into the roles of Aire in establishment of self-tolerance. In this study, we monitored the maturation program of AEL-mTECs by temporal lineage tracing, in which bacterial artificial chromosome transgenic mice expressing tamoxifen-inducible Cre recombinase under control of the Aire regulatory element were crossed with reporter strains. We estimated that the half-life of AEL-mTECs subsequent to Aire expression was ∼7–8 d, which was much longer than that reported previously, owing to the existence of a post-Aire stage. We found that loss of Aire did not alter the overall lifespan of AEL-mTECs, inconsistent with the previous notion that Aire expression in medullary thymic epithelial cells (mTECs) might result in their apoptosis for efficient cross-presentation of self-antigens expressed by AEL-mTECs. In contrast, Aire was required for the full maturation program of AEL-mTECs, as exemplified by the lack of physiological downregulation of CD80 during the post-Aire stage in Aire-deficient mice, thus accounting for the abnormally increased CD80high mTECs seen in such mice. Of interest, increased CD80high mTECs in Aire-deficient mice were not mTEC autonomous and were dependent on cross-talk with thymocytes. These results further support the roles of Aire in the differentiation program of AEL-mTECs.

Dehydrogenation properties and crystal structure of the double anion complex hydride Mg(BH4)(NH2) were studied by thermal analyses and synchrotron X-ray diffraction. The stoichiometric mixture of Mg(BH4)2 and Mg(NH2)2 were ball-milled and then heated to 453 K to form Mg(BH4)(NH2) crystal. The dehydrogenation of Mg(BH4)(NH2) occurs in two-stage at 513 K and 688 K. The following reaction sequence is suggested by the results of thermal analyses; Mg(BH4)(NH2) → MgH2 + BN + 2H2 (7.3 mass% weight loss) → Mg + BN + 3H2 (11.0 mass% weight loss in total). The dehydrogenation temperature of Mg(BH4)(NH2) is approximately 50 K lower than that of the other double anion complex Li2(BH4)(NH2). The crystal structure of Mg(BH4)(NH2) was determined by the measurement at 300 K (crystal system: tetragonal, space group: I41 (No. 80), lattice constants: a = 5.792(1), c = 20.632(4) A at 300 K). In the crystal of Mg(BH4)(NH2), the cation (Mg2+) and the anions ( BH 4 - and NH 2 - ) are stacked alternately along the c-axis direction. The Mg2+ cation is tetrahedrally coordinated with two BH 4 - anions and two NH 2 - anions.

Background— Hypoandrogenemia is associated with an increased risk of ischemic diseases. Because actions of androgens are exerted through androgen receptor (AR) activation, we studied hind-limb ischemia in AR knockout mice to elucidate the role of AR in response to ischemia. Methods and Results— Both male and female AR knockout mice exhibited impaired blood flow recovery, more cellular apoptosis, and a higher incidence of autoamputation after ischemia. In ex vivo and in vivo angiogenesis studies, AR-deficient vascular endothelial cells showed reduced angiogenic capability. In ischemic limbs of AR knockout mice, reductions in the phosphorylation of the Akt protein kinase and endothelial nitric oxide synthase were observed despite a robust increase in hypoxia-inducible factor 1α and vascular endothelial cell growth factor ( VEGF ) gene expression. In in vitro studies, siRNA-mediated ablation of AR in vascular endothelial cells blunted VEGF-stimulated phosphorylation of Akt and endothelial nitric oxide synthase. Immunoprecipitation experiments documented an association between AR and kinase insert domain protein receptor that promoted the recruitment of downstream signaling components. Conclusions— These results document a physiological role of AR in sex-independent angiogenic potency and provide evidence of novel cross-talk between the androgen/AR signaling and VEGF/kinase insert domain protein receptor signaling pathways.

Complex hydride Mg(BH4)(NH2), which consists of double anion BH4− and NH2−, was synthesized and the crystal structure was analyzed by synchrotron X-ray diffraction. The mixture sample of Mg(BH4)2 + Mg(NH2)2 prepared by ball milling was reacted and crystallized to Mg(BH4)(NH2) by heating at about 453 K. This crystal phase transforms into amorphous phase above 473 K and subsequently the dehydrogenation begins. The crystal structure of Mg(BH4)(NH2) was determined from measurement data at 453 K (chemical formula: Mg0.94(BH4)1(NH2)0.88, crystal system: tetragonal, space group: I41 (No.80), Z = 8, lattice constants: a = 5.814(1), c = 20.450(4) A at 453 K). Mg(BH4)(NH2) is ionic crystal which the cation (Mg2+) and the anions (BH4− and NH2−) are stacking alternately along the c-axis direction. Two BH4− and two NH2− tetrahedrally coordinate around Mg2+ ion.

Hassall’s corpuscles are concentric clusters of keratinized epithelial cells located within the thymic medulla of humans and guinea pigs but are scant in mouse and rat. They are considered to be the terminally differentiated stages of medullary thymic epithelial cells (mTECs) but the mechanisms of their origin are unclear. We have previously deleted the TGF-β type II receptor (TGFβRII) specifically in mouse TECs and reported that these mice have mitigated thymic involution and exhibit earlier reconstitution post-irradiation. In this study, we analyzed the differentiation of mTECs in the TGFβRII-knockout mice. Interestingly, the TGFβRII-knockout mice display enhanced development of Hassall’s corpuscles. The expression of Aire, stromal-cell-derived factor 1 and thymic stromal lymphopoietin in the thymi of the TGFβRII-knockout mice was similar to that previously reported for the human thymus. In addition, the putative epithelial progenitor markers MTS20 and MTS24 labeled Hassall’s corpuscles in normal mice, but the extent and intensity of this staining were greatly enhanced in Hassall’s corpuscles of the TGFβRII-knockout mice. The phosphorylated forms of ERK and JNK were also found in Hassall’s corpuscles of the TGFβRII-knockout mice. Taken together, we suggest that TGFβRII-mediated signaling in TECs inhibits their development into Hassall’s corpuscles in mice.

Our body is equipped with an immune system that normally distinguishes between microorganisms (non-self) and components of our body (self), thereby protecting us from the invasion by many pathogens. However, intractable autoimmune disease, in which somehow our immune system attacks our own body, could develop by unknown mechanisms. With aiming toward novel therapeutic approaches to chronic inflammation caused by autoimmunity, we are studying the mechanisms underlying the development of organ-specific autoimmune diseases caused by the abnormal function of antigen-presenting cells in the thymus. We are particularly interested in how the immunological self is presented to developing thymocytes to establish the self-tolerance within the thymic microenvironment. NF-κB activation pathway in medullary thymic epithelial cells (mTECs) and Aire in mTECs, a transcription factor which is responsible for the development of rather rare hereditary type of autoimmune disease, are the main focus of our research, hoping that manipulations of those factors could control the chronic inflammation observed in many organ-specific autoimmune diseases.

Summary Toll-like receptor-7 (TLR7) and 9, innate immune sensors for microbial RNA or DNA, have been implicated in autoimmunity. Upon activation, TLR7 and 9 are transported from the endoplasmic reticulum (ER) to endolysosomes for nucleic acid sensing by an ER-resident protein, Unc93B1. Little is known, however, about a role for sensor transportation in controlling autoimmunity. TLR9 competes with TLR7 for Unc93B1-dependent trafficking and predominates over TLR7. TLR9 skewing is actively maintained by Unc93B1 and reversed to TLR7 if Unc93B1 loses preferential binding via a D34A mutation. We here demonstrate that mice harboring a D34A mutation showed TLR7-dependent, systemic lethal inflammation. CD4 + T cells showed marked differentiation toward T helper 1 (Th1) or Th17 cell subsets. B cell depletion abolished T cell differentiation and systemic inflammation. Thus, Unc93B1 controls homeostatic TLR7 activation by balancing TLR9 to TLR7 trafficking.

The Li–Mg–N–H system has the property of reversible reaction with hydrogen between hydrogenation and dehydrogenation (Mg 3 N 2 + 4Li 3 N + 12H 2 ↔ 3Mg(NH 2 ) 2 + 12LiH). At the several dehydrogenation stages of p–c isotherm measurement at 523 K, the structural change was investigated using the synchrotron X-ray diffraction. There are two regions in p–c isotherm of the Li–Mg–N–H system, i.e. plateau and sloping region. In the plateau region, Mg(NH 2 ) 2 and Li 3 Mg 3 (NH 2 )(NH) 4 coexist. In the sloping region, the intermediate phase Li 3+3 y Mg 3 (NH 2 ) 1− y (NH) 4+2 y changes continuously from Li 3 Mg 3 (NH 2 )(NH) 4 to Li 2 Mg(NH) 2 . The chemical composition of the intermediate phase was estimated from the amount of desorbed hydrogen by p–c isotherm and the atomic ratio of Mg and N by Rietveld analysis. The crystal structure of the intermediate phase, Li 3+3 y Mg 3 (NH 2 ) 1− y (NH) 4+2 y (space group: I 222), was determined. Because all these intermediate structures are similar to anti-CaF 2 -type, it is deduced that the dehydrogenation process are caused by the diffusion of Li + to cation sites of Mg(NH 2 ) 2 . The analysis of structural change clarified the dehydrogenation process that is accomplished by the diffusion of Li + and Mg 2+ without N atom diffusion.

In the mechanism of thymus-induced central tolerance, the transcription factor Aire has been demonstrated to promote the expression of a wide range of peripheral organ-specific antigens (Ags) in the medullary thymic epithelial cells (mTECs), which serve as self-Ags in negative selection. We examined the expression of desmoglein 3 (Dsg3), the autoantigen in pemphigus vulgaris (PV), in mouse thymus and the involvement of Aire in tolerance to Dsg3. Immunofluorescence and in situ hybridization revealed Dsg3 in single cells or in clusters in ∼3% of mTECs near the cortico-medullary junction of the thymus in C57BL/6 mice. Dsg3-expressing mTECs also expressed some Ags of skin-unrelated peripheral organs simultaneously. In contrast, Dsg3-positive mTECs were not detected in the Aire(-/-) thymus. Adoptive transfer of splenocytes from Aire(-/-) mice immunized with Dsg3 did not induce anti-Dsg3 IgG production or PV phenotype in Rag2(-/-) recipient mice. However, Aire(-/-) CD4(+) T cells, but not Aire(+/+) CD4(+) T cells, induced low levels of anti-Dsg3 IgG production when transferred with Dsg3(-/-) B cells. These findings indicate that Aire has an important role in Dsg3 expression as well as in selection of T cells that help B cells to produce anti-Dsg3 IgG in thymus.

The existence of Mg(B12H12) as an intermediate compound during the dehydrogenation process of Mg(BH4)2 has been verified. To elucidate the formation process of Mg(B12H12), the dehydrogenation products of Mg(BH4)2 at different temperatures were characterized via Xray diffraction and B magic angle spinning NMR measurements. The experimental results indicate that several new intermediate compounds such as Mg(B2H6) and Mg(B5H9) or Mg(B5H8)2 are expected to be formed prior to the formation of Mg(B12H12). Thus, it is suggested that the formation of [B12H12] 2 from [BH4] occurs via the gradual evolution of the B-H complex anion, i.e., [BH4] ! [B2H6] ! [B5H9] or [B5H8] ! [B12H12] . [doi:10.2320/matertrans.MA201002]

The current prevailing view regarding the role of Aire in self-tolerance is that it is involved in the transcriptional control of many tissue-restricted self-antigen genes in thymic epithelial cells in the medulla (mTECs); however, accumulating evidence also suggests that Aire has other roles, e.g. in mTEC differentiation, and furthermore that Aire can either promote or inhibit the mTEC differentiation program, i.e. Aire does not play a neutral role in mTEC differentiation. This review discusses when and how Aire plays an important role in controlling the organization of mTECs required for the expression of self-antigen genes.

The mutagenicity of a series of the light element hydrides (containing NaAlH4, Mg(NH2)2, LiBH4, etc.) was examined by evaluating the frequency of mutation in bacterial DNAs. Although some materials were suspected to be slightly mutagenic, their effect was much less malignant than that of well-known potent mutagens. The hydrides exhibited high cytotoxicity, rather than mutagenicity. A Mg(NH2)2-related material was also subjected to a series of toxicity tests on aqueous organisms, i.e., algae, water fleas and fish. The result suggests that the material is as toxic as alkaline metal hydroxides, such as NaOH and KOH.

Ly6 family proteins are mainly expressed by myeloid-cell lineages such as granulocytes (expressing Ly6G) and monocytes (expressing Ly6C). However, exact function of Ly6 family protein in each cell-type remains unknown. For example, although expression of Ly6C has been used to define functionally distinct subset of monocytes (Ly6C+ monocytes vs. Ly6C− monocytes), molecular basis for the expression of Ly6C and functional difference between the two populations are largely unknown. Medullary thymic epithelial cells (mTECs) are composed of heterogeneous populations in terms of the spectrum of tissue-restricted antigens (TRAs) expression for ensuring the elimination of autoreactive T-cell repertoire. We have recently identified mTEC subset(s) that express Ly6C and Ly6G both recognized by monoclonal antibody Gr-1 (i.e., Ly6C/6G+ mTECs). Approximately 10% of total mTECs expressed Ly6C/6G and Ly6C/6G+ mTECs were preferentially localized at the cortico-medullary junction. We found that Ly6C/6G+ mTECs and Ly6C/6G− mTECs do not form distinct lineage(s). Instead, Ly6C/6G− mTECs are able to convert into Ly6C/6G+ mTECs and vice versa in a reaggregated thymic organ culture (RTOC). Interestingly, Aire-deficient mice do not possess Ly6C/6G+ mTECs. RNA-seq analyses have revealed that Ly6C/6G+ mTECs express higher levels of chemokines, integrins and extracellular matrix proteins compared with those from Ly6C/6G− mTECs, suggesting that Ly6C/6G+ mTECs might constitute a unique tolerogenic subset among heterogenous mTECs.

Calcium borohydride Ca(BH4)2 is one of the promising new hydrogen storage materials because of its large amount of hydrogen desorption capability (9.6 mass%). The crystal structures of α-Ca(BH4)2 (space group: Fddd, lattice constants: a = 8.7782(2) A, b = 13.129(1) A, c = 7.4887(9) A) and β-Ca(BH4)2 (P42/m, a = 6.9509(5) A, c = 4.3688(3) A) were refined by synchrotron X-ray diffraction at 300 and 433 K, respectively. The unsolved structures of γ-Ca(BH4)2 (Pbca, a = 7.525(1) A, b = 13.109(2) A, c = 8.403(1) A) and Ca(BH4)2·H2O (Pnma, a = 8.200(1) A, b = 5.8366(7) A, c = 11.851(2) A) were determined. In α-, β- and γ-Ca(BH4)2 structures, six boron atoms around a calcium atom construct CaB6 octahedron. The polymorphism of Ca(BH4)2 is formed by the different connection with adjacent octahedrons sharing vertexes and edges of the CaB6 octahedron. Furthermore, the charge density distribution in α-Ca(BH4)2 was experimentally determined by maximum entropy method. It is clarified that the bonding nature in α-Ca(BH4)2 ionic crystal is constructed from Ca2+ cation and BH4− anion.

AIRE is the gene responsible for a rather rare hereditary type of autoimmune disease. The mechanism underlying the autoimmune pathogenesis caused by AIRE deficiency is a focus of intense research because it could provide clues to the fundamental question of how the immune system discriminates between self and non-self within the thymic microenvironment.

Expression of the p16Ink4a tumor suppressor gene, a sensor of oncogenic stress, is up-regulated by a variety of potentially oncogenic stimuli in cultured primary cells. However, because p16Ink4a expression is also induced by tissue culture stress, physiological mechanisms regulating p16Ink4a expression remain unclear. To eliminate any potential problems arising from tissue culture–imposed stress, we used bioluminescence imaging for noninvasive and real-time analysis of p16Ink4a expression under various physiological conditions in living mice. In this study, we show that oncogenic insults such as ras activation provoke epigenetic derepression of p16Ink4a expression through reduction of DNMT1 (DNA methyl transferase 1) levels as a DNA damage response in vivo. This pathway is accelerated in the absence of p53, indicating that p53 normally holds the p16Ink4a response in check. These results unveil a backup tumor suppressor role for p16Ink4a in the event of p53 inactivation, expanding our understanding of how p16Ink4a expression is regulated in vivo.