No abstract

GATA transcription factors interact with FOG proteins to regulate tissue development by activating and repressing transcription. FOG-1 (ZFPM1), a co-factor for the haematopoietic factor GATA-1, binds to the NuRD corepressor complex through a conserved N-terminal motif. Surprisingly, we detected NuRD components at both repressed and active GATA-1/FOG-1 target genes in vivo. In addition, while NuRD is required for transcriptional repression in certain contexts, we show a direct requirement of NuRD also for FOG-1-dependent transcriptional activation. Mice in which the FOG-1/NuRD interaction is disrupted display defects similar to germline mutations in the Gata1 and Fog1 genes, including anaemia and macrothrombocytopaenia. Gene expression analysis in primary mutant erythroid cells and megakaryocytes (MKs) revealed an essential function for NuRD during both the repression and activation of select GATA-1/FOG-1 target genes. These results show that NuRD is a critical co-factor for FOG-1 and underscore the versatile use of NuRD by lineage-specific transcription factors to activate and repress gene transcription in the appropriate cellular and genetic context.

Recent reports describe hematopoietic abnormalities in mice with targeted instability of the mitochondrial genome. However, these abnormalities have not been fully described. We demonstrate that mutant animals develop an age-dependent, macrocytic anemia with abnormal erythroid maturation and megaloblastic changes, as well as profound defects in lymphopoiesis. Mice die of severe fatal anemia at 15 months of age. Bonemarrow transplantation studies demonstrate that these abnormalities are intrinsic to the hematopoietic compartment and dependent upon the age of donor hematopoietic stem cells. These abnormalities are phenotypically similar to those found in patients with refractory anemia, suggesting that, in some cases, the myelodysplastic syndromes are caused by abnormalities of mitochondrial function. (Blood. 2009;114:4045-4053)

To cite this article: Shelat SG, Smith P, Ai J, Long Zheng X. Inhibitory autoantibodies against ADAMTS-13 in patients with thrombotic thrombocytopenic purpura bind ADAMTS-13 protease and may accelerate its clearance in vivo. J Thromb Haemost 2006; 4: 1707-17 Summary. Background: Many patients with acquired thrombotic thrombocytopenic purpura (TTP) harbor autoantibodies that may bind and/or inhibit ADAMTS-13 proteolytic activity and accelerate its clearance in vivo. Methods: To test this hypothesis, we determined ADAMTS-13 activity and antigen levels in parallel plasma samples from patients clinically diagnosed with TTP. Collagen binding, GST-VWF73 and FRETS-VWF73 assays were used to determine ADAMTS-13 activity and to detect inhibitory autoantibodies. Enzyme-linked immunosorbent assay (ELISA) and immunoprecipitation plus Western blotting (IP/WB) were used to detect total anti-ADAMTS-13 IgG (inhibitory and non-inhibitory). Results: Among 40 patients with TTP (21 idiopathic and 19 nonidiopathic), inhibitory autoantibodies were detected (by FRETS-VWF73) in 52% of idiopathic and 0% of nonidiopathic TTP patients. In contrast, non-inhibitory IgG autoantibodies were detected in 29% of idiopathic and 50% of non-idiopathic TTP patients. The concentration of inhibitory IgG autoantibody in idiopathic TTP patients was significantly higher than that of non-inhibitory IgG in either idiopathic or non-idiopathic TTP patients. Idiopathic TTP patients demonstrated significantly reduced ADAMTS-13 activity compared with non-idiopathic patients, but only slightly lower ADAM-TS-13 antigen levels. Interestingly, patients with inhibitory autoantibodies exhibited significantly lower ADAMTS-13 antigen levels than those with only non-inhibitory IgG autoantibodies or no autoantibody. Serial plasma exchanges increased levels of ADAMTS-13 activity and antigen concurrently in patients with inhibitory autoantibodies. Conclusion:The identification of severe ADAMTS-13 deficiency and autoantibodies or inhibitors appears to be assay-dependent; the inhibitory IgG autoantibodies, in addition to binding and inhibiting ADAMTS-13 proteolytic activity, may accelerate ADAMTS-13 clearance in vivo.Keywords: a disintegrin and metalloprotease with thrombospondin type 1 repeats, autoimmune disorder, thrombotic microangiopathy, von Willebrand factor.

IntroductionHematopoiesis is the process whereby the blood cell lineages are generated from a multipotent hematopoietic stem cell (HSC). HSCs possess the unique ability to self-renew but also differentiate to give rise to mature blood cells. Long-term HSCs (LT-HSCs) are the most primitive HSCs and can fully reconstitute the hematopoietic compartment of lethally irradiated animals. They reside in a hypoxic niche and are generally quiescent. 1,2 In comparison, short-term HSCs (ST-HSCs) are temporally limited in their ability to reconstitute lethally irradiated animals and more actively cycle. HSCs are regulated through their environmental niche, cytokine signaling, and the orchestrated activities of various transcription factors. 3 However, there is a relative paucity of information about the signal transduction events that regulate HSC function. In particular, the effects of fatty acid (FA) metabolism and lipid mediators on HSC function are not well understood.The function of HSCs is dependent on their unique capacity to both differentiate and self-renew, which is related to their proliferative capacity. 4,5 Highly proliferative HSCs are less efficient at reconstituting lethally irradiated mice. 6 Thus, HSCs that lack cell-cycle inhibitors required to maintain quiescence, such as p21cip/WAF, are defective. 7 HSC function is also regulated by transcription factors including Gfi-1, Bmi, Hox4b, and PU.1. [8][9][10][11] Furthermore, canonical Wnt signaling was shown to regulate HSC function by maintaining quiescence and initiating a self-renewal program of gene transcription. [12][13][14] 12/15-Lipoxygenase (12/15-LOX) is a lipid-peroxidizing enzyme that mediates unsaturated FA metabolism. 12/15-LOX introduces molecular oxygen into arachidonic acid (AA) and linoleic acid to produce bioactive labile lipid intermediates such as 12(S)-hydroperoxyeicosatetraenoic acid, 15(S)-hydroperoxyeicosatetraenoic acid, and 13(S)-hydroperoxyoctadecadienoic acid. These intermediates are rapidly reduced by glutathione reductase, which releases reactive oxygen species (ROS) and produces additional bioactive lipid metabolites, including 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE), 15(S)-hydroxyeicosatetraenoic acid (15(S)-HETE), 13(S)-hydroxyoctadecadienoic (13(S)-HODE), lipoxins, and hepoxilins. 15 Eicosanoid products activate multiple signaling pathways and transcription factors such as peroxisome proliferator-activated receptor-␥, interferon consensus sequencebinding protein/interferon regulatory factor-8 (ICSBP/IRF-8), and nuclear factor-B to modulate gene transcription. 16,17 These lipid mediators have pleiotropic affects on myriad cell types, 18 but our knowledge of their role in HSCs is limited. 19,20 Notably, in leukemia that results from abnormal hematopoiesis, lipid mediators are reduced. [21][22][23] Previously, we showed that 12/15-LOX is a novel suppressor of myeloproliferative disease (MPD). 24 Although less than 15% of 12/15-LOX-deficient mice (Alox15 mice) develop a severe MPD over the course of a year, the majority ...

Mineralocorticoids, such as deoxycorticosterone acetate (DOCA), and angiotensin II (AngII) act synergistically in the brain to elicit salt appetite. Glucocorticoids, such as dexamethasone (DEX), also may enhance the behavioral effects of DOCA and AngII. However, the brain regions involved in these behavioral interactions have not been elucidated. This study tested the hypothesis that DEX potentiates the effects of DOCA on AngII binding, especially at the AT1 receptor. We confirmed that DEX potentiated the effects of DOCA on salt appetite. Concomitantly, steroid-specific and region-specific changes in AT1 binding were noted. Specifically, in the hypothalamic paraventricular nucleus, treatment with DEX or DOCA + DEX increased AT1 binding. In the subfornical organ (SFO) and area postrema, there was an increase in AT1 binding when both steroids were combined, but not when given individually. However, there was no change in AT2 binding in any brain region studied and no change in AT1 or AT2 binding to either receptor subtype in the pituitary. The results indicate that DOCA and DEX may increase the sensitivity of the brain to the behavioral and physiological actions of AngII by upregulating AT1 receptors in the SFO and area postrema.

Hemoglobin production during erythropoiesis is mechanistically coupled to the acquisition and metabolism of iron. We discovered that iron regulates the expression of ␣-hemoglobinstabilizing protein (AHSP), a molecular chaperone that binds and stabilizes free ␣-globin during hemoglobin synthesis. In primates, the 3-untranslated region (UTR) of AHSP mRNA contains a nucleotide sequence resembling iron responsive elements (IREs), stem-loop structures that regulate gene expression post-transcriptionally by binding iron regulatory proteins (IRPs). The AHSP IRE-like stem-loop deviates from classical consensus sequences and binds IRPs poorly in electrophoretic mobility shift assays. However, in cytoplasmic extracts, AHSP mRNA co-immunoprecipitates with IRPs in a fashion that is dependent on the stem-loop structure and inhibited by iron. Moreover, this interaction enhances AHSP mRNA stability in erythroid and heterologous cells. Our findings demonstrate that IRPs can regulate mRNA expression through non-canonical IREs and extend the repertoire of known iron-regulated genes. In addition, we illustrate a new mechanism through which hemoglobin may be modulated according to iron status.

The CXCR4 receptor (Chemokine C-X-C motif receptor 4) is highly expressed in different hematological malignancies including chronic lymphocytic leukemia (CLL). The CXCR4 ligand (CXCL12) stimulates CXCR4 promoting cell survival and proliferation, and may contribute to the tropism of leukemia cells towards lymphoid tissues. Therefore, strategies targeting CXCR4 may constitute an effective therapeutic approach for CLL. To address that question, we studied the effect of Ulocuplumab (BMS-936564), a fully human IgG4 anti-CXCR4 antibody, using a stroma – CLL cells co-culture model. We found that Ulocuplumab (BMS-936564) inhibited CXCL12 mediated CXCR4 activation-migration of CLL cells at nanomolar concentrations. This effect was comparable to AMD3100 (Plerixafor - Mozobil), a small molecule CXCR4 inhibitor. However, Ulocuplumab (BMS-936564) but not AMD3100 induced apoptosis in CLL at nanomolar concentrations in the presence or absence of stromal cell support. This pro-apoptotic effect was independent of CLL high-risk prognostic markers, was associated with production of reactive oxygen species and did not require caspase activation. Overall, these findings are evidence that Ulocuplumab (BMS-936564) has biological activity in CLL, highlight the relevance of the CXCR4-CXCL12 pathway as a therapeutic target in CLL, and provide biological rationale for ongoing clinical trials in CLL and other hematological malignancies.