CD47 promotes peripheral T cell survival by preventing dendritic cell-mediated T cell necroptosis.

Conventional dendritic cells (cDCs) are required for peripheral T cell homeostasis in lymphoid organs, but the molecular mechanism underlying this requirement has remained unclear. We here show that T cell-specific CD47-deficient (Cd47 ΔT) mice have a markedly reduced number of T cells in peripheral tissues. Direct interaction of CD47-deficient T cells with cDCs resulted in activation of the latter cells, which in turn induced necroptosis of the former cells. The deficiency and cell death of T cells in Cd47 ΔT mice required expression of its receptor signal regulatory protein α on cDCs. The development of CD4+ T helper cell-dependent contact hypersensitivity and inhibition of tumor growth by cytotoxic CD8+ T cells were both markedly impaired in Cd47 ΔT mice. CD47 on T cells thus likely prevents their necroptotic cell death initiated by cDCs and thereby promotes T cell survival and function.

Targeting of SIRPα as a potential therapy for Langerhans cell histiocytosis.

Langerhans cell histiocytosis (LCH) is a rare neoplastic disorder characterized by inflammatory lesions arising from the anomalous accumulation of pathogenic CD1a+ CD207+ dendritic cells (DCs). SIRPα is a transmembrane protein highly expressed in myeloid cells such as DCs and macrophages. Here we show that SIRPα is a potential therapeutic target for LCH. We found that SIRPα is expressed in CD1a+ cells of human LCH lesions as well as in CD11c+ DCs in the spleen, liver, and lung of a mouse model of LCH (BRAFV600ECD11c mouse), in which an LCH-associated active form of human BRAF is expressed in a manner dependent on the mouse Cd11c promoter. BRAFV600ECD11c mice manifested markedly increased numbers of CD4+ T cells, regulatory T cells, and macrophages as well as of CD11c+ MHCII+ DCs in the spleen. Monotherapy with a mAb to SIRPα greatly reduced the percentage of CD11c+ MHCII+ DCs in peripheral blood, LCH-like lesion size in the liver, and the number of CD11c+ MHCII+ DCs in the spleen of the mutant mice. Moreover, this mAb promoted macrophage-mediated phagocytosis of CD11c+ DCs from BRAFV600ECD11c mice, whereas it had no effects on the viability or CCL19-dependent migration of such CD11c+ DCs or on their expression of the chemokine genes Ccl5, Ccl20, Cxcl11, and Cxcl12. Our results thus suggest that anti-SIRPα monotherapy is a promising approach to the treatment of LCH that is dependent in part on the promotion of the macrophage-mediated killing of LCH cells.

Preclinical evaluation of the efficacy of an antibody to human SIRPα for cancer immunotherapy in humanized mouse models.

Tumor-associated macrophages (TAMs) are abundant in the tumor microenvironment and are considered potential targets for cancer immunotherapy. To examine the antitumor effects of agents targeting human TAMs in vivo, we here established preclinical tumor xenograft models based on immunodeficient mice that express multiple human cytokines and have been reconstituted with a human immune system by transplantation of human CD34+ hematopoietic stem and progenitor cells (HIS-MITRG mice). HIS-MITRG mice supported the growth of both human cell line (Raji)- and patient-derived B cell lymphoma as well as the infiltration of human macrophages into their tumors. We examined the potential antitumor action of an antibody to human SIRPα (SE12C3) that inhibits the interaction of CD47 on tumor cells with SIRPα on human macrophages and thereby promotes Fcγ receptor-mediated phagocytosis of the former cells by the latter. Treatment with the combination of rituximab (antibody to human CD20) and SE12C3 inhibited Raji tumor growth in HIS-MITRG mice to a markedly greater extent than did rituximab monotherapy. This enhanced antitumor effect was dependent on human macrophages and attributable to enhanced rituximab-dependent phagocytosis of lymphoma cells by human macrophages. Treatment with rituximab and SE12C3 also induced reprogramming of human TAMs toward a proinflammatory phenotype. Furthermore, the combination treatment essentially prevented the growth of patient-derived diffuse large B cell lymphoma in HIS-MITRG mice. Our findings thus support the study of HIS-MITRG mice as a model for the preclinical evaluation in vivo of potential therapeutics, such as antibodies to human SIRPα, that target human TAMs.

GCN2 regulates pancreatic ß cell mass by sensing intracellular amino acid levels.

EIF2AK4, which encodes the amino acid deficiency-sensing protein GCN2, has been implicated as a susceptibility gene for type 2 diabetes in the Japanese population. However, the mechanism by which GCN2 affects glucose homeostasis is unclear. Here, we show that insulin secretion is reduced in individuals harboring the risk allele of EIF2AK4 and that maintenance of GCN2-deficient mice on a high-fat diet results in a loss of pancreatic ß cell mass. Our data suggest that GCN2 senses amino acid deficiency in ß cells and limits signaling by mechanistic target of rapamycin complex 1 to prevent ß cell failure during the consumption of a high-fat diet.

Histone deacetylase regulates insulin signaling via two pathways in pancreatic ß cells.

Recent studies demonstrated that insulin signaling plays important roles in the regulation of pancreatic ß cell mass, the reduction of which is known to be involved in the development of diabetes. However, the mechanism underlying the alteration of insulin signaling in pancreatic ß cells remains unclear. The involvement of epigenetic control in the onset of diabetes has also been reported. Thus, we analyzed the epigenetic control of insulin receptor substrate 2 (IRS2) expression in the MIN6 mouse insulinoma cell line. We found concomitant IRS2 up-regulation and enhanced insulin signaling in MIN6 cells, which resulted in an increase in cell proliferation. The H3K9 acetylation status of the Irs2 promoter was positively associated with IRS2 expression. Treatment of MIN6 cells with histone deacetylase inhibitors led to increased IRS2 expression, but this occurred in concert with low insulin signaling. We observed increased IRS2 lysine acetylation as a consequence of histone deacetylase inhibition, a modification that was coupled with a decrease in IRS2 tyrosine phosphorylation. These results suggest that insulin signaling in pancreatic ß cells is regulated by histone deacetylases through two novel pathways affecting IRS2: the epigenetic control of IRS2 expression by H3K9 promoter acetylation, and the regulation of IRS2 activity through protein modification. The identification of the histone deacetylase isoform(s) involved in these mechanisms would be a valuable approach for the treatment of type 2 diabetes.

The Presence of Venous Gas Does Not Affect the Prognosis in Emphysematous Cystitis.

Objective Emphysematous cystitis (EC) has a high mortality rate compared with urinary tract infection without emphysema. However, its prognostic factors have yet to be determined. The presence of venous gas is suspected to be a rare, adverse prognostic factor of EC. However, all four previously reported cases improved. We hypothesized that venous gas is not an adverse prognostic factor of EC and aimed to assess the effect of venous gas on the EC prognosis. Methods Medical records were reviewed retrospectively. Patients The patients diagnosed with EC at Yodogawa Christian Hospital between April 2004 and September 2014 were included. Results Venous gas was present in 15 of 23 patients with EC. There was no significant difference in the background or clinical presentation between patients with or without venous gas. All patients with venous gas survived without invasive measures, whereas 50% of patients without venous gas died. Conclusion There was no marked difference in the mortality rate due to EC between the patients with and without venous gas. Venous gas may be a more common and less worrying finding in EC than assumed. It does not reflect the severity of infection, and air embolisms have not been reported so far. Venous gas may not affect the prognosis. This may be due to the differences in the mechanism of venous gas production. Gas in EC may develop due to glucose fermentation and intravesical pressurization, in contrast to the necrotizing infection seen in other emphysematous infections. This is the first study to assess the effect of venous gas on EC prognosis.

Luman is involved in osteoclastogenesis through the regulation of DC-STAMP expression, stability and localization.

Luman (also known as CREB3) is a type-II transmembrane transcription factor belonging to the OASIS family that localizes to the endoplasmic reticulum (ER) membrane under normal conditions. In response to ER stress, OASIS-family members are subjected to regulated intramembrane proteolysis (RIP), following which the cleaved N-terminal fragments translocate to the nucleus. In this study, we show that treatment of bone marrow macrophages (BMMs) with cytokines - macrophage colony-stimulating factor (M-CSF) and RANKL (also known as TNFSF11) - causes a time-dependent increase in Luman expression, and that Luman undergoes RIP and becomes activated during osteoclast differentiation. Small hairpin (sh)RNA-mediated knockdown of Luman in BMMs prevented the formation of multinucleated osteoclasts, concomitant with the suppression of DC-STAMP, a protein that is essential for cell-cell fusion in osteoclastogenesis. The N-terminus of Luman facilitates promoter activity of DC-STAMP, resulting in upregulation of DC-STAMP expression. Furthermore, Luman interacts with DC-STAMP, and controls its stability and localization. These results suggest that Luman regulates the multinucleation of osteoclasts by promoting cell fusion of mononuclear osteoclasts through DC-STAMP induction and intracellular distribution during osteoclastogenesis.

Promotion of Cancer Cell Proliferation by Cleaved and Secreted Luminal Domains of ER Stress Transducer BBF2H7.

BBF2H7 is an endoplasmic reticulum (ER)-resident transmembrane basic leucine zipper (bZIP) transcription factor that is cleaved at the transmembrane domain by regulated intramembrane proteolysis in response to ER stress. The cleaved cytoplasmic N-terminus containing transcription activation and bZIP domains translocates into the nucleus to promote the expression of target genes. In chondrocytes, the cleaved luminal C-terminus is extracellularly secreted and facilitates proliferation of neighboring cells through activation of Hedgehog signaling. In the present study, we found that Bbf2h7 expression levels significantly increased by 1.070-2.567-fold in several tumor types including glioblastoma compared with those in respective normal tissues, using the ONCOMINE Cancer Profiling Database. In some Hedgehog ligand-dependent cancer cell lines including glioblastoma U251MG cells, the BBF2H7 C-terminus was secreted from cells into the culture media and promoted cancer cell proliferation through activation of Hedgehog signaling. Knockdown of Bbf2h7 expression suppressed the proliferation of U251MG cells by downregulating Hedgehog signaling. The impaired cell proliferation and Hedgehog signaling were recovered by addition of BBF2H7 C-terminus to the culture medium of Bbf2h7-knockdown U251MG cells. These data suggest that the secreted luminal BBF2H7 C-terminus is involved in Hedgehog ligand-dependent cancer cell proliferation through activation of Hedgehog signaling. Thus, the BBF2H7 C-terminus may be a novel target for the development of anticancer drugs.

Lysosomal accumulation of Trk protein in brain of GM1 -gangliosidosis mouse and its restoration by chemical chaperone.

G(M1) -gangliosidosis is a fatal neurodegenerative disorder caused by deficiency of lysosomal acid ß-galactosidase (ß-gal). Accumulation of its substrate ganglioside G(M1) (G(M1) ) in lysosomes and other parts of the cell leads to progressive neurodegeneration, but underlying mechanisms remain unclear. Previous studies demonstrated an essential role for interaction of G(M1) with tropomyosin receptor kinase (Trk) receptors in neuronal growth, survival and differentiation. In this study we demonstrate accumulation of G(M1) in the cell-surface rafts and lysosomes of the ß-gal knockout (ß-gal-/-) mouse brain association with accumulation of Trk receptors and enhancement of its downstream signaling. Immunofluorescence and subcellular fractionation analysis revealed accumulation of Trk receptors in the late endosomes/lysosomes of the ß-gal-/- mouse brain and their association with ubiquitin and p62. Administration of a chemical chaperone to ß-gal-/- mouse expressing human mutant R201C protein resulted in a marked reduction of intracellular storage of G(M1) and phosphorylated Trk. These findings indicate that G(M1) accumulation in rafts causes activation of Trk signaling, which may participate in the pathogenesis of G(M1) -gangliosidosis.