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Figure S1. Skin exposure to UVB induces splenic erythroid cell proliferation in mice. Figure S2. Skin exposure to UVB results in expansion of splenic cell populations with erythroid colony-forming potential. Figure S3. Administration of an anti-EPO antibody prevents UVB-responsive splenic erythroid expansion. Figure S4. Tumor growth induces splenic erythroid cell proliferation in mice. Figure S5. CD235A immunostaining identifies tumor-associated erythroid cells in human cancer tissues.

Hematopoietic-derived cells are integral components of the tumor microenvironment and serve as critical mediators of tumor–host interactions. Host cells derived from myeloid and lymphoid lineages perform well-established functions linked to cancer development, progression, and response to therapy. It is unclear whether host erythroid cells also contribute to shaping the path that cancer can take, but emerging evidence points to this possibility. Here, we show that tumor-promoting environmental stress and tumor-induced hemodynamic changes trigger renal erythropoietin production and erythropoietin-dependent expansion of splenic erythroid cell populations in mice. These erythroid cells display molecular features indicative of an immature erythroid phenotype, such as the expression of both CD71 and TER119 and the retention of intact nuclei, and express genes encoding immune checkpoint molecules. Nucleated erythroid cells with similar properties are present in mouse and human tumor tissues. Antibody-mediated erythropoietin blockade reduces tumor-responsive erythroid cell induction and tumor growth. These findings reveal the potential of tumor-induced erythropoietin and erythroid cells as targets for cancer treatment.Implications: Our study identifies erythropoietin and erythroid cells as novel players in tumor–host interactions and highlights the involvement of multiorgan signaling events in their induction in response to environmental stress and tumor growth.

Keratinocytes, the epithelial cells of the skin, reprogram their gene expression and produce immune effector molecules when exposed to environmental and endogenous triggers of inflammation. It remains unclear how keratinocytes process physiological signals generated during skin irritation and switch from a homeostatic to an inflammatory state. In this article, we show that the stress-activated protein kinase p38α is crucial for keratinocytes to prompt changes in their transcriptome upon cytokine stimulation and drive inflammation in allergen-exposed skin. p38α serves this function by phosphorylating p63, a transcription factor essential for the lineage identity and stemness of the skin epithelium. Phosphorylation by p38α alters the activity of p63 and redeploys this developmental transcription factor to a gene expression program linked to inflammation. Genetic ablation and pharmacological inhibition of p38α or the p38α–p63 target gene product MMP13 attenuate atopic dermatitis–like disease in mice. Our study reveals an epithelial molecular pathway promoting skin inflammation and actionable through treatment with topical small-molecule therapeutics.

Coordinated induction, but also repression, of genes are key to normal differentiation. Although the role of lineage-specific transcription regulators has been studied extensively, their functional integration with chromatin remodelers, one of the key enzymatic machineries that control chromatin accessibility, remains ill-defined. Here we investigate the role of Mi-2β, a SNF-2-like nucleosome remodeler and key component of the nucleosome remodeling and histone deacetylase (NuRD) complex in early B cells. Inactivation of Mi-2β arrested differentiation at the large pre-B-cell stage and caused derepression of cell adhesion and cell migration signaling factors by increasing chromatin access at poised enhancers and chromosome architectural elements. Mi-2β also supported IL-7R signaling, survival, and proliferation by repressing negative effectors of this pathway. Importantly, overexpression of Bcl2, a mitochondrial prosurvival gene and target of IL-7R signaling, partly rescued the differentiation block caused by Mi-2β loss. Mi-2β stably associated with chromatin sites that harbor binding motifs for IKAROS and EBF1 and physically associated with these transcription factors both on and off chromatin. Notably, Mi-2β shared loss-of-function cellular and molecular phenotypes with IKAROS and EBF1, albeit in a distinct fashion. Thus, the nucleosome remodeler Mi-2β promotes pre-B-cell differentiation by providing repression capabilities to distinct lineage-specific transcription factor-based regulatory networks.

The cohesin complex participates in the organization of 3D genome through generating and maintaining DNA loops. Stromal antigen 2 (STAG2), a core subunit of the cohesin complex, is frequently mutated in various cancers. However, the impact of STAG2 inactivation on 3D genome organization, especially the long-range enhancer-promoter contacts and subsequent gene expression control in cancer, remains poorly understood. Here we show that depletion of STAG2 in melanoma cells leads to expansion of topologically associating domains (TADs) and enhances the formation of acetylated histone H3 lysine 27 (H3K27ac)-associated DNA loops at sites where binding of STAG2 is switched to its paralog STAG1. We further identify Interferon Regulatory Factor 9 (IRF9) as a major direct target of STAG2 in melanoma cells via integrated RNA-seq, STAG2 ChIP-seq and H3K27ac HiChIP analyses. We demonstrate that loss of STAG2 activates IRF9 through modulating the 3D genome organization, which in turn enhances type I interferon signaling and increases the expression of PD-L1. Our findings not only establish a previously unknown role of the STAG2 to STAG1 switch in 3D genome organization, but also reveal a functional link between STAG2 and interferon signaling in cancer cells, which may enhance the immune evasion potential in STAG2-mutant cancer.

Objective There are few reports on the outcomes of 12-week paritaprevir, ombitasvir, and ritonavir (PTV/OBV/r) treatment in real-world clinical settings. We aimed to evaluate the efficacy and safety of 12-week treatment with ritonavir-boosted paritaprevir and ombitasvir in patients with hepatitis C virus (HCV) genotype 1 infection in a real-world setting. Methods Fifty-eight patients with chronic hepatitis or compensated hepatic cirrhosis and genotype-1 HCV infection were treated with PTV/OBV/r and followed for 24 weeks after the completion of treatment in 10 centers in northern Tohoku. The efficacy and safety of this 12-week treatment regimen was analyzed. Results Among the 58 treated patients, 18 (31%) had compensated liver cirrhosis, while 11 (19%) patients had experienced treatment failure with another treatment regimen. NS5A resistance-associated variants (RAVs) were detected at baseline in 3 patients (5.2%), including Y93H in two patients and L31M in two patients. One patient had NS5A RAVs at both positions 93 and 31. The overall sustained virological response (SVR) 24 rate was 96.6%. Three patients with NS5A RAVs also achieved an SVR24. The SVR24 rate was not significantly affected by age, sex, prior treatment, prior history of HCC, or liver stiffness. The mean alanine aminotransferase (ALT) levels decreased significantly during this treatment. Adverse events occurred in 15 patients (26%), 26% of which were grade 1 or 2. No severe adverse events occurred. Conclusion In this real-world study, 12-week PTV/OBV/r treatment was effective and safe for treating patients with HCV-1 infection who had chronic hepatitis or compensated hepatic cirrhosis.

IKAROS is required for the differentiation of highly proliferative pre-B-cell precursors, and loss of IKAROS function indicates poor prognosis in precursor B-cell acute lymphoblastic leukemia (B-ALL). Here we show that IKAROS regulates this developmental stage by positive and negative regulation of superenhancers with distinct lineage affiliations. IKAROS defines superenhancers at pre-B-cell differentiation genes together with B-cell master regulators such as PAX5, EBF1, and IRF4 but is required for a highly permissive chromatin environment, a function that cannot be compensated for by the other transcription factors. IKAROS is also highly enriched at inactive enhancers of genes normally expressed in stem-epithelial cells. Upon IKAROS loss, expression of pre-B-cell differentiation genes is attenuated, while a group of extralineage transcription factors that are directly repressed by IKAROS and depend on EBF1 relocalization at their enhancers for expression is induced. LHX2, LMO2, and TEAD-YAP1, normally kept separate from native B-cell transcription regulators by IKAROS, now cooperate directly with them in a de novo superenhancer network with its own feed-forward transcriptional reinforcement. Induction of de novo superenhancers antagonizes Polycomb repression and superimposes aberrant stem epithelial cell properties in a B-cell precursor. This dual mechanism of IKAROS regulation promotes differentiation while safeguarding against a hybrid stem epithelial B-cell phenotype that underlies high-risk B-ALL. Funding Agencies|National Institutes of Health [R01CA162092, R21AI124326, R01CA090576, R01CA190964]

Purpose of review Loss of IKAROS in committed B cell precursors causes a block in differentiation while at the same time augments aberrant cellular properties, such as bone marrow stromal adhesion, self-renewal and resistance to glucocorticoid-mediated cell death. B cell acute lymphoblastic leukaemias originating from these early stages of B cell differentiation and associated with IKAROS mutations share a high-risk cellular phenotype suggesting that deregulation of IKAROS-based mechanisms cause a highly malignant disease process. Recent studies Recent studies show that IKAROS is critical for the activity of super-enhancers at genes required for pre-B cell receptor (BCR) signalling and differentiation, working either downstream of or in parallel with B cell master regulators such as EBF1 and PAX5. IKAROS also directly represses a cryptic regulatory network of transcription factors prevalent in mesenchymal and epithelial precursors that includes YAP1, TEAD1/2, LHX2 and LMO2, and their targets, which are not normally expressed in lymphocytes. IKAROS prevents not only expression of these 'extra-lineage' transcription factors but also their cooperation with endogenous B cell master regulators, such as EBF1 and PAX5, leading to the formation of a de novo for lymphocytes super-enhancer network. IKAROS coordinates with the Polycomb repression complex (PRC2) to provide stable repression of associated genes during B cell development. However, induction of regulatory factors normally repressed by IKAROS starts a feed-forward loop that activates de-novo enhancers and elevates them to super-enhancer status, thereby diminishing PRC2 repression and awakening aberrant epithelial-like cell properties in B cell precursors. Summary Insight into IKAROS-based transcriptional circuits not only sets new paradigms for cell differentiation but also provides new approaches for classifying and treating high-risk human B-ALL that originates from these early stages of B cell differentiation.

The Ikaros family of DNA-binding proteins are critical regulators of lymphocyte differentiation. In multipotent, hematopoietic progenitors, Ikaros supports transcriptional priming of genes promoting lymphocyte differentiation. Ikaros targets the Nucleosome Remodeling Deacetylase (NuRD) complex to lymphoid lineage genes, thereby increasing chromatin accessibility and transcriptional priming. After lymphoid lineage specification, Ikaros expression is raised to levels characteristic of intermediate B cell and T cell precursors, which is necessary to support maturation and prevent leukemogenesis. Loss of Ikaros in T cell precursors allows the NuRD complex to repress lymphocyte genes and extends its targeting to genes that support growth and proliferation, causing their activation and triggering a cascade of events that leads to leukemogenesis. Loss of Ikaros in B cell precursors blocks differentiation and perpetuates stromal adhesion by enhancing integrin signaling. The combination of integrin and cytokine signaling in Ikaros-deficient pre-B cells promotes their survival and self-renewal. The stages of lymphocyte differentiation that are highly dependent on Ikaros are underscored by changes in Ikaros transcription, supported by a complex network of stage-specific regulatory networks that converge upon the Ikzf1 locus. It is increasingly apparent that understanding the regulatory networks that operate upstream and downstream of Ikaros is critical not only for our understanding of normal lymphopoiesis, but also in placing the right finger on the mechanisms that support hematopoietic malignancies in mouse and human.

Extra-gastrointestinal stromal tumors (E-GISTs) not associated with the alimentary tract in the pelvic cavity are extremely rare. We treated a 49-year-old Japanese man with such an E-GIST in the pelvic cavity who underwent an intrapelvic tumorectomy with a total prostatectomy and partial rectum resection. Gross examination of the specimen revealed an 8.1 × 5 × 4 cm white-grayish mass. Histological findings showed uniform spindle cells with scant atypia that formed interlacing bundles or whorl patterns. These neoplastic cells did not invade adjacent organs, including the gut. Immunohistochemical findings revealed that the neoplastic cells were positive for c-kit, CD34, and vimentin. Molecular analysis showed a c-kit mutation at exon 9 with duplication of Ala and Tyr. Our diagnosis was E-GIST, which belongs to the intermediate group of GIST. Following the operation, we administered imatinib mesylate for 6 months. After stopping for 5 months, it was administered again for local recurrence. We are planning our future strategy for this case including surgical resection as necessary.

The ability of somatic stem cells to self-renew and differentiate into downstream lineages is dependent on specialized chromatin environments that keep stem cell-specific genes active and key differentiation factors repressed but poised for activation. The epigenetic factors that provide this type of regulation remain ill-defined. Here we provide the first evidence that the SNF2-like ATPase Mi-2β of the Nucleosome Remodeling Deacetylase (NuRD) complex is required for maintenance of and multilineage differentiation in the early hematopoietic hierarchy. Shortly after conditional inactivation of Mi-2β, there is an increase in cycling and a decrease in quiescence in an HSC (hematopoietic stem cell)-enriched bone marrow population. These cycling mutant cells readily differentiate into the erythroid lineage but not into the myeloid and lymphoid lineages. Together, these effects result in an initial expansion of mutant HSC and erythroid progenitors that are later depleted as more differentiated proerythroblasts accumulate at hematopoietic sites exhibiting features of erythroid leukemia. Examination of gene expression in the mutant HSC reveals changes in the expression of genes associated with self-renewal and lineage priming and a pivotal role of Mi-2β in their regulation. Thus, Mi-2β provides the hematopoietic system with immune cell capabilities as well as with an extensive regenerative capacity.

Ikaros DNA binding factor plays critical roles in lymphocyte development. Changes in Ikaros expression levels during lymphopoiesis are controlled by redundant but also unique regulatory elements of its locus that are critical for this developmental process. We have recently shown that Ikaros binds its own locus in thymocytes in vivo. Here, we evaluated the role of an Ikaros binding site within its major lympho-myeloid promoter. We identified an Ikaros/Ets binding site within a promoter sub-region that was highly conserved in mouse and human. Deletion of this binding site increased the percentage of the reporter-expressing mouse lines, indicating that its loss provided a more permissive chromatin environment. However, once transcription was established, the lack of this site decreased transcriptional activity. These findings implicate a dual role for Ikaros/Ets1 binding on Ikzf1 expression that is exerted at least through its promoter.

Regulated expression of Ikaros is critical for normal hemopoiesis and lymphocyte development. To elucidate the mechanisms underlying transcription of Ikaros, tissue-specific DNase I-hypersensitive sites (DHS) were mapped throughout the Ikaros locus, and several promoters were identified. The activity of these regulatory regions was elucidated using an enhanced green fluorescent protein (EGFP) reporter in transgenic mice. Two genomic fragments, each containing a distinct promoter and its associated DHS cluster, were found to be active in the myeloid (DHS-C2 and DHS-C3) and B-cell (DHS-C3) lineages. Although neither of these regulatory regions was active within the majority of differentiating thymocytes and mature T cells, the DHS-C3 region was active at the earliest stages (DN1–DN3) of T-cell differentiation. However, when the DHS-C3 region was combined with the downstream intronic DHS-C6 cluster, its activity was maintained and raised to higher levels at subsequent stages of T-cell differentiation. This combination of regulatory elements provided reporter expression that closely resembles that of endogenous Ikaros during hemo-lymphopoiesis, and it decreased (but did not alleviate) position effect variegation within the expressing cell types.

A53-year-old woman was hospitalized with a hepatic tumor that had been detected on health-screening ultrasonography. Laboratory data showed no marked findings: HBs-Ag and HCV-Ab were negative, and levels of serum AFP, PIVKA-II, CEA, and CA 19-0 were within their normal ranges. Ultrasonogram showed a hyperechoic circle concentric with and inside of a hypoechoic mass in the right hepatic lobe. The tumor appeared isodense with the surrounding liver in unenhanced CT, but enhanced CT showed a low density mass. A celiac arteriogram revealed a hypervascular tumor. The resected tumor measured 20×15 mm. Microscopic findings showed a carcinoid tumor that was positive for Grimelius stain. Further examination of the lung, gastrointestinal tract, and other organs found no primary carcinoid tumor. The patient has been well and free from tumor of the liver or elsewhere for the 7 years since her operation. We therefore considered this lesion to have been a primary carcinoid tumor of the liver. Only 14 cases of this disease have ever been reported in Japan. This report includes a review of the literature on the imaging of this rare tumor.

B-cell acute lymphoblastic leukemia (B-ALL) is a malignancy of precursor B-lymphocytes affecting both children and adults. Deletions and dominant-negative mutations in IKZF1, the gene encoding the Ikaros transcription factor, are found in ~85% of Ph+ B-ALL and in some cases of Ph– B-ALL, and are associated with poor prognosis. Genomic studies of high-risk Ph– or “Ph-like” B-ALLs have revealed frequent mutation and activation of TK genes and signaling pathways. While ABL1 tyrosine kinase inhibitors (TKIs) such as dasatinib and imatinib have been added to chemotherapy regimens for Ph+ B-ALL, over half of these patients will still relapse, which correlates with residual disease burden in the bone marrow (BM) following induction therapy. Hence, new therapeutic strategies are needed for patients with Ikaros-mutant, high-risk Ph+ and Ph– B-ALL. Using mice with a conditional Ikzf1 mutation (Ike5fl) where the recombined allele is similar to the dominant-negative Ik6 mutant found in human B-ALL, we demonstrated recently that Ikaros DNA-binding function is required in the B-lymphoid lineage for transition from the large to small pre-B cell stage of differentiation, and that arrest at this stage of development can give rise to B-ALL (Joshi et al., Nat. Immunol. 2014;15:294). The survival and proliferation of Ikaros mutant pre-B cells is dependent on increased integrin-mediated stromal adhesion and activation of focal adhesion kinase (FAK). FAK is a non-receptor TK, downstream of integrins and growth factor receptors, which plays important roles in cancer stem cell biology, the tumor microenvironment and tumorigenesis. VS-4718 and VS-6063 (defactinib) are potent, orally bioavailable FAK inhibitors that inhibit tumor growth and metastasis in preclinical models, and are currently under evaluation in clinical trials in patients with various solid tumors. VS-6063 has demonstrated tolerability and preliminary signs of clinical activity as a single agent and in combination with paclitaxel in phase I trials (ASCO, 2014). Here, we show that BCR-ABL1 cooperates with Ikzf1 mutation to accelerate B-leukemogenesis in mice. BCR-ABL1+ Ikaros-mutant B-ALLs exhibit stroma-mediated resistance to ABL1 TKIs, while the FAK inhibitors VS-4718 and VS-6063 are effective in blocking stromal adhesion and inducing apoptosis in both mouse and human Ikaros-mutant B-ALL samples. To test whether dysregulation of TK signaling cooperates with Ikzf1 mutation in the pathogenesis of high-risk B-ALL, we isolated BM B-lymphoid progenitor cells from wild-type (WT), IkE5fl/+ CD2-Cre, and IkE5fl/fl CD2-Cre donors, transduced them with BCR-ABL1 retrovirus and transplanted the cells into recipient mice. We observed a dramatic acceleration of precursor B-lymphoid leukemia induced by BCR-ABL1 in IkE5Δ/+ and particularly in IkE5Δ/Δ donor cells that correlated with a striking (~30-fold) increase in the frequency of engrafting leukemia-initiating or leukemic stem cells (LSCs). Relative to Ikzf1 WT BCR-ABL1+ leukemic cells, Ikzf1-mutant BCR-ABL1+ blasts showed significant resistance to imatinib and dasatinib that was dependent on the presence of OP9 stroma. The effect of FAK inhibition, using the FAK inhibitors VS-4718, VS-6062, and VS-6063 (Verastem), was first tested on murine B-ALL cells (genotypes Ikzf1 mutant, Ikzf1 mutant BCR-ABL1+, and Ikzf1 WT BCR-ABL1+) grown on OP9 stroma. FAK inhibitor treatment abolished stromal adhesion of Ikzf1-mutant B-ALL and induced apoptosis in non-adherent cells, but had little effect on Ikzf1 WT B-ALL cells. VS-4718 and VS-6063 were each synergistic with dasatinib in reducing the viability of Ikzf1-mutant BCR-ABL1+ B-ALL cells cultured on OP9 stroma. For primary human B-ALL samples grown on OP9 stroma, IKZF1-mutant cells were also more sensitive to FAK inhibitor treatment than WT IKZF1 WT B-ALL, with or without BCR-ABL1 expression. Collectively, these observations suggest a new model to explain the pathogenesis of high-risk B-ALL and its resistance to therapy. B-ALLs with IKZF1 mutations may be resistant to TKIs and to chemotherapy by virtue of their stromal adhesion phenotype, resulting in failure to eliminate BM LSCs. Inhibition of FAK signaling in Ph+ or Ph­–IKZF1-mutant B-ALL may reverse the stromal-mediated resistance to ABL1 TKIs and/or chemotherapy. Therefore, FAK inhibitors warrant further investigation for the treatment of high-risk IKZF1-mutant B-ALL patients. Disclosures Joshi: Verastem: Research Funding. Yoshida:Verastem, Inc.: Research Funding. Paraskevopoulou:Verastem, Inc.: Research Funding. Zhang:Verastem, Inc.: Research Funding. Krause:Glycomimetics. Inc.: Research Funding. Shapiro:Verastem: Employment, Equity Ownership. Weaver:Verastem: Employment, Equity Ownership. Pachter:Verastem Inc.: Employment, Equity Ownership. Georgopoulos:Verastem, Inc.: Research Funding.

Precise control of gene expression plays fundamental roles in brain development, but the roles of chromatin regulators in neuronal connectivity have remained poorly understood. We report that depletion of the NuRD complex by invivo RNAi and conditional knockout of the core NuRD subunit Chd4 profoundly impairs the establishment of granule neuron parallel fiber/Purkinje cell synapses in the rodent cerebellar cortex invivo. By interfacing genome-wide sequencing of transcripts and ChIP-seq analyses, we uncover a network of repressed genes and distinct histone modifications at target gene promoters that are developmentally regulated by the NuRD complex in the cerebellum invivo. Finally, in a targeted invivo RNAi screen of NuRD target genes, we identify a program of NuRD-repressed genes that operate as critical regulators of presynaptic differentiation in the cerebellar cortex. Our findings define NuRD-dependent promoter decommissioning as a developmentally regulated programming mechanism that drives synaptic connectivity in the mammalian brain.

Survivin is expressed in the nucleus and/or cytoplasm of various types of malignant tumor cells. Nuclear survivin is indispensable for complete mitosis, while cytoplasmic survivin functions as an apoptosis inhibitor. We examined the difference in the survivin expression among stromal cells of fibroadenoma, and benign and malignant phyllodes tumors.Tumor sections were immunohistochemically stained with an anti-human survivin antibody and the labeling index of survivin was calculated.In stromal cells of all tumors, survivin was expressed in the nuclei but not in the cytoplasm. The labeling indices of the stromal cells in five malignant phyllodes tumors (20.5±3.0) were significantly greater than those observed in eight fibroadenomas (1.9±0.6) or nine benign phyllodes tumors (3.0±0.9).In the present study it was shown that stromal cells in malignant phyllodes tumors express nuclear survivin more extensively than stromal cells in benign phyllodes tumors or fibroadenomas.

Deletion of the DNA-binding domain of the transcription factor Ikaros generates dominant-negative isoforms that interfere with its activity and correlate with poor prognosis in human precursor B cell acute lymphoblastic leukemia (B-ALL). Here we found that conditional inactivation of the Ikaros DNA-binding domain in early pre-B cells arrested their differentiation at a stage at which integrin-dependent adhesion to niches augmented signaling via mitogen-activated protein kinases, proliferation and self-renewal and attenuated signaling via the pre-B cell signaling complex (pre-BCR) and the differentiation of pre-B cells. Transplantation of polyclonal Ikaros-mutant pre-B cells resulted in long-latency oligoclonal pre-B-ALL, which demonstrates that loss of Ikaros contributes to multistep B cell leukemogenesis. Our results explain how normal pre-B cells transit from a highly proliferative and stroma-dependent phase to a stroma-independent phase during which differentiation is enabled, and suggest potential therapeutic strategies for Ikaros-mutant B-ALL.

A 67-year-old man with bladder cancer who was treated with transurethral resection of bladder tumour(TUR-Bt)and chemotherapy at the age of 59 years was diagnosed as having urothelial cancer by biopsy 8 years later. Detailed examination revealed the presence of synchronous triple cancer, with hepatocellular cancer and gastric cancer. Subsequently, semi-total gastrectomy, partial hepatectomy(S6), radio frequency ablation(S5, S7), and cholecystectomy were performed. Histologically, the gastric tumor was a moderately differentiated tubular adenocarcinoma, the hepatic tumor was a moderately differentiated hepatocellular carcinoma, the bladder tumor was a transitional cell carcinoma, and the ureteral tumor was an urothelial carcinoma.

Deletion of IL-5Rα-chain (IL-5Rα−/−) selectively influenced the mucosal IgA responses in vivo. While levels of IgA in mucosal secretions were more reduced in IL-5Rα−/− mice than in wild-type mice, the levels of IgA in serum were not changed. The frequency of IgA-producing cells was reduced in mucosal effector sites (e.g., intestinal lamina propria and nasal passage), but not in inductive sites such as Payer’s patches and nasal-associated lymphoreticular tissues in IL-5Rα−/− mice. IgA-committed (surface IgA+; sIgA+) B-1 cells mainly resided in mucosal effector tissues, while conventional sIgA+ B (B-2) cells formed in mucosal inductive sites of wild-type mice. In contrast, in the effector tissue of IL-5Rα−/− mice, sIgA+ B-1 cells, but not sIgA+ B-2 cells in the inductive site, were significantly reduced. IL-5Rα was more expressed on sIgA+ B-1 cells than was IL-6R, while both IL-5Rα and IL-6R were expressed on sIgA+ B-2 cells in wild-type mice. sIgA+ B-1 cells produced high levels of IgA with rIL-5 rather than of rIL-6 in vitro. Taken together, the findings suggest that the IL-5/IL-5R signaling pathway is critically important for the development of common mucosal immune system independent sIgA+ B-1 cell in mucosal effector tissues in vivo.

electric impulse単独の正常肝への影響および肝癌への治療効果についてラットを用い検討した. 正常肝と癌周囲に対し2または4本の針状電極を電極間距離10mmで肝葉に刺入し, 100μsecの矩形パルスを8パルス通電した. 通電電圧を500, 1, 000, 1, 500V/cmとし, 通電形態を電極2本通電, 4本4辺通電, 4本4辺と対角線を加えた6辺通電に分け検討した. 正常肝での肝機能は, 通電電圧に相関して上昇し, 壊死範囲を反映していた. 正常肝および肝癌の通電では, いずれも通電電圧に相関して完全壊死頻度が上昇し, 4辺1, 500V/cm, 6辺1, 000V/cmにより完全壊死が得られた. 7日後の組織所見は, 通電部のみ肉芽組織に置換されていた. 副作用は, electric impulseに同調した筋収縮のみで, 他には認められなかった. electric impulse単独療法は, 電圧により目的とする部位の完全壊死が副作用なく得られ, 肝癌の新しい治療法となる可能性が示唆された.

Ikaros is a critical regulator of lymphocyte development and homeostasis; thus, understanding its transcriptional regulation is important from both developmental and clinical perspectives. Using a mouse transgenic reporter approach, we functionally characterized a network of highly conserved cis-acting elements at the Ikzf1 locus. We attribute B-cell and myeloid but not T-cell specificity to the main Ikzf1 promoter. Although this promoter was unable to counter local chromatin silencing effects, each of the 6 highly conserved Ikzf1 intronic enhancers alleviated silencing. Working together, the Ikzf1 enhancers provided locus control region activity, allowing reporter expression in a position and copy-independent manner. Only 1 of the Ikzf1 enhancers was responsible for the progressive upregulation of Ikaros expression from hematopoietic stem cells to lymphoid-primed multipotent progenitors to T-cell precursors, which are stages of differentiation dependent on Ikaros for normal outcome. Thus, Ikzf1 is regulated by both epigenetic and transcriptional factors that target its enhancers in both redundant and specific fashions to provide an expression profile supportive of normal lymphoid lineage progression and homeostasis. Mutations in the Ikzf1 regulatory elements and their interacting factors are likely to have adverse effects on lymphopoiesis and contribute to leukemogenesis., link_to_OA_fulltext

Cell fate depends on the interplay between chromatin regulators and transcription factors. Here we show that activity of the Mi-2β nucleosome-remodeling and histone-deacetylase (NuRD) complex was controlled by the Ikaros family of lymphoid lineage-determining proteins. Ikaros, an integral component of the NuRD complex in lymphocytes, tethered this complex to active genes encoding molecules involved in lymphoid differentiation. Loss of Ikaros DNA-binding activity caused a local increase in chromatin remodeling and histone deacetylation and suppression of lymphoid cell-specific gene expression. Without Ikaros, the NuRD complex also redistributed to transcriptionally poised genes that were not targets of Ikaros (encoding molecules involved in proliferation and metabolism), which induced their reactivation. Thus, release of NuRD from Ikaros regulation blocks lymphocyte maturation and mediates progression to a leukemic state by engaging functionally opposing epigenetic and genetic networks.

In radiation heat transfer evaluation, the hemispherical emittance spectrum is the most important characteristic quantity. This emittance is directly related to the hemispherical reflectance RHH for perfect-diffuse hemispherical irradiation. However, there has been no proper technique for measuring RHH of real surfaces which reflect radiation imperfect-diffusely. In this paper, we present a new reflectometer system for measuring a spectrum of RHH in a wavelength region of 0.35-1.10μm. This system has two ellipsoidal mirrors and a beam splitter to realize the perfect-diffuse irradiation and the hemispherical reflection detection. This system also measures the spectra of normal- and obliqueincident hemispherical reflectances. We apply this technique to the investigation of the spectral and angular characteristics of reflection from rough surfaces of mild steel, and demonstrate the capability of the system in a fundamental study of radiation characteristics of materials.

The distribution of CCK-immunoreactive cells was investigated by the indirect fluorescence method in the cerebral cortex of LEC (Long-Evans Cinnamon) rats which have recently been introduced as a model of jaundice and hepatic encephalopathy. Reduction of CCK-immunoreactive nerve cells was observed in the brains of LEC rats with stage III to V hepatic encephalopathy in comparison with the rats without hepatic encephalopathy. These observations were confirmed by counting the CCK-positive nerve cells at magnification x 125. 3 microscopic fields per animal were immunostained and CCK-immunoreactive nerve cells counted. The number of CCK-immunoreactive cells per field was 28.5 +/- 0.7 (mean +/- SEM, n = 5) in rats without hepatic encephalopathy, and 14.1 +/- 2.0 (n = 3) in rats with stage III to V hepatic encephalopathy. Thus, the number of CCK-immunoreactive nerve cells was significantly less in rats with stage III to V hepatic encephalopathy (p less than 0.05). A decrease in CCK-immunoreactive nerve cells was not observed in the rats with stage I or II hepatic encephalopathy. This study shows that there is a relationship between the severity of hepatic encephalopathy and the number of CCK-immunoreactive nerve cells in the cerebral cortex.

We established a new method of plasma treatment for the removal of interfering factors in the plasma to allow detection of endotoxin by limulus test. The limulus test used was an endotoxin-specific chromogenic test, the Endospecy test. Perchloric acid (PCA) treatment and centrifugation (PCA method) is usually used to remove interfering factors from plasma, with the precipitate being discarded and the supernatant used to detect endotoxin. As the solubilized precipitates of endotoxin-spiked plasma and some patient plasma were found to contain the Endospecy activity, we have devised a new method assaying endotoxin in both the supernatant and precipitate. This study confirmed that the solubilized precipitate of endotoxin-spiked plasma had Endospecy activity and found that the precipitate had other endotoxin activities, such as lethality in galactosamine-sensitized mice and pyrogenicity in rabbits. We also confirmed that interfering factors were completely removed from plasma samples by this new method. The endotoxin level after the new PCA method was found to be about 8 times higher than that determined after PCA treatment and the new PCA method surpasses the conventional PCA method with regard to the positive rate of endotoxin contents in clinical samples. These results indicate that the new PCA method is superior to the PCA method as a plasma pretreatment method for limulus test.

A new reflectometer system is developed for measuring the radiative characteristics of solid materials from a thermal engineering point of view. This system employs an ellipsoidal mirror in a simple optical alignment, and measures the spectrum of the normal-incident hemispherical reflectance in a wavelength region from 0.35 to 1.10μm. A single spectral scanning offers the hemispherical reflectance spectrum which completely covers both of the specular and diffuse components of reflection. The reflectance is directly interpreted into the absorptance for the normal incidence. The system also measures the angular dependence of reflection readily. The system is applied to a systematic study on the reflection characteristics of rough metallic surfaces. The predominant contribution of the specular component in the angular characteristics is demonstrated.

漢方薬が原因と推定された劇症肝炎の1例を経験した.症例は62歳男性.痔核治療のため漢方薬(金鵄丸)の服用を開始したところ,6週間後に倦怠感と尿濃染が出現した.服用中止により一旦症状の消失をみたが,服薬再開後5週間で上記症状が再出現,黄疸の出現をみ入院となった.凝固能低下が著明で(PT 28%, HPT 19%),種々の治療にもかかわらず,4週間後多臓器不全の状態で死亡した.剖検肝は495gと萎縮著明で,肝組織像は広範性肝壊死を示した.金鵄丸による薬剤性肝炎は本例を含め9例が報告されている.その特徴は,金鵄丸が原因との認識が遅れたため反復服用により肝炎の繰り返しをみる例が多いこと,発疹,好酸球増多がみられないことであった.本例は,漢方薬により劇症肝炎を来した初めての報告である.漢方薬の使用が増加しているおり,漢方薬によっても本例のごとき重症肝障害が惹起されうることに注意すべきである.

An 80-year-old woman with a pleomorphic carcinoma (PC) producing hCG was admitted to Nippon Steel Hirohata Hospital because of an abnormal shadow on CT seen during a follow-up examination after surgery for breast cancer. A right upper lobectomy was performed due to rapid growth of the shadow 3 months later. Macroscopically the tumor was a 4.8 x 4.0 cm well-circumscribed grayish-white mass. On histology the tumor consisted mostly of intermingled spindle and polygonal cells, while evidence of poorly differentiated adenocarcinoma was seen in a few areas. A diagnosis of PC was made due to hCG expression in approximately 20% of the spindle and polygonal cells on immunohistology. Six months after the operation metastasis to the liver and adrenal gland was seen on CT. The patient died due to metastases 1 year after the operation, even though the patient had been at stage 1B at the time of the operation and appropriate chemotherapy had been given. PC patients with immunohistochemical hCG expression have elevated risk of local recurrence and metastasis.

Hyperplastic polyps (HPs) of the stomach have been reported to be mostly benign. However, in rare cases, carcinomas have been found in HPs. We treated a 59-year-old Japanese male who underwent a total gastrectomy, and a gross examination of the resected stomach revealed a 4.8 × 3.8-cm polyp on the greater curvature of the antrum and multiple small polyps in the whole gastric mucosa. Histologically, the large polyp consisted mainly of hyperplastic foveolar epithelium, while the presence of variously colored lobules demonstrated a poorly differentiated adenocarcinoma mixed with signet-ring cell carcinoma. Hyperplastic polyps should therefore be carefully examined microscopically as a polypectomy specimen and in resected stomach specimens.

Deletions and dominant-negative mutations in IKZF1, the gene encoding Ikaros transcription factor, are found in ∼85% of Ph+ B-ALL and in some cases of Ph− B-ALL, and are associated with poor prognosis. Genomic studies of high-risk Ph− or “Ph-like” B-ALLs have revealed frequent mutation and activation of TK genes and signaling pathways. While ABL1 tyrosine kinase inhibitors (TKIs) such as dasatinib and ponatinib have been incorporated into chemotherapy regimens for Ph+ B-ALL, the majority of patients still relapse, which correlates with residual bone marrow disease following induction therapy. New therapeutic strategies are needed for patients with Ikaros-mutant, high-risk Ph+ and Ph− B-ALL. Using mice with a conditional Ikzf1 mutation (Ike5fl) that mimics the dominant-negative Ik6 mutant found in human B-ALL, we demonstrated that loss of Ikaros DNA-binding function arrests B-lymphoid development at a large pre-B cell stage that can give rise to B-ALL. Survival and proliferation of Ikaros mutant pre-B cells is dependent on increased integrin-mediated stromal adhesion and activation of focal adhesion kinase (FAK). FAK is a non-receptor TK downstream of integrins and growth factor receptors that plays important roles in cancer stem cell biology and the tumor microenvironment. Here, we show that dysregulated tyrosine kinases, including BCR-ABL1, cooperate with Ikaros mutation to accelerate the development of B-lymphoblastic leukemia in mice, correlated with a striking (∼30-fold) increase in the frequency of engrafting leukemia-initiating or leukemic stem cells (LSCs). Ikzf1-mutant BCR-ABL1+ lymphoblasts exhibit relative resistance to ABL1 TKIs including dasatinib that is dependent on the presence of stroma. VS-4718 is a potent, orally bioavailable FAK inhibitor that inhibits tumor growth and metastasis in preclinical models, and is currently under evaluation in clinical trials in patients with various solid tumors. VS-4718 treatment abolished stromal adhesion and induced apoptosis of Ikzf1-mutant B-ALL and synergized with dasatinib, but had little effect on Ikzf1 WT B-ALL cells. Primary human Ph+ B-ALL samples showed a correlation between IKZF1 mutation status, stromal adherence, and sensitivity to FAK and ABL inhibitors in vitro, and BM tropism in vivo in xenografted NSG mice. Results from ongoing in vivo therapy experiments will be presented. Together, these results suggest a new model for the pathogenesis of high-risk B-ALL and its resistance to conventional therapy. B-ALLs with IKZF1 mutations may be resistant to TKIs and to chemotherapy by virtue of their stromal adhesion phenotype, resulting in failure to eliminate BM leukemic stem cells. Inhibition of FAK signaling in Ph+ or Ph− IKZF1-mutant B-ALL may reverse the stromal-mediated resistance to ABL1 TKIs and/or chemotherapy. FAK inhibitors represent a promising new approach for the treatment of high-risk IKZF1-mutant B-ALL patients. Citation Format: Nilamani Jena, Ila Joshi, Toshimi Yoshida, Zhihong Zhang, Zhong-Ying Liu, Prasanthi Tata, Irina M. Shapiro, Jonathan A. Pachter, David T. Weaver, Katia Georgopoulos, Richard A. Van Etten. Targeting focal adhesion kinase is a novel approach to therapy of high-risk, Ikaros-mutant acute B-cell lymphoblastic leukemia. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4202. doi:10.1158/1538-7445.AM2015-4202

Hematopoiesis is the developmental process by which all blood and immune cells are generated. A decade-old scheme has supported an early and complete separation of the erythro-myeloid from the lymphoid lineages. Recent advances have re-drawn this map, separating lymphoid and myeloid from erythroid programs early in development. Subsequently, the fate restriction of both the lympho-myeloid and the erythro-megakaryocyte progenitors is dependent on Ikaros and its associated chromatin regulators. Genetic studies of this family of nuclear factors are now providing unique insight into the functional molecular signatures that bestow plasticity to the hematopoietic stem cell and its early progeny.

Ikaros is expressed in early hematopoietic progenitors and is required for lymphoid differentiation. Analysis of Ikaros null populations revealed a lack of defining markers for early fate-restricted progenitors, but it was difficult to discern whether Ikaros was required for formation of these populations, or for expression of these markers. Here we use a GFP reporter based on Ikaros regulatory elements to identify the HSC and separate early progenitors in both wild-type and Ikaros-null mice. The presence of lympho-myeloid progenitors is revealed in Ikaros-null mice, which lack the defining factor Flt3 and are capable of myeloid, but not lymphoid differentiation. In contrast, lack of Ikaros in the common myeloid progenitor results in increased formation of erythro-megakaryocyte at the expense of myeloid progenitors and influences their subsequent differentiation. By this approach, pivotal roles for Ikaros in distinct fate decisions in the early hematopoietic hierarchy are revealed.

In many developmental systems, nuclear regulators have been implicated in coupling key events in gene expression with specific cell fate and lineage decisions. In the hemo-lymphoid system, the Ikaros gene family of zinc finger DNA binding factors controls lymphocyte specification and homeostasis from the hemopoietic stem cell (HSC) throughout development. The dependence of hemo-lymphoid differentiation on Ikaros DNA binding activity together with the presence of Ikaros proteins within higher order chromatin remodeling complexes supports the hypothesis that Ikaros plays a key role in the lineage-specific remodeling of chromatin. Association of Ikaros and its remodeling partners with the chromatin of key lineage-specific genes, and the dependence of these genes on Ikaros complexes for their expression supports this hypothesis and provides unique paradigms to study chromatin regulation of differentiation.

The transcription factor GATA-3 is expressed in quiescent hematopoietic stem cells (HSCs) with long-term repopulating ability. Stress-mediated activation of HSCs promotes localization of GATA-3 to the nucleus in a manner dependent on the mitogen-activated protein kinase p38α and restricts self-renewal.