bluuebunny/crossref_metadata_2025_split huggingface dataset:
Impaired Maturation and Survival of T Lymphocytes, B Lymphocytes, and NK Cells in Mice Lacking Gimap5/Ian5.
Authors: Schulteis Ryan D. , Dai Xuezhi , Chu Haiyan , Edwards Brandon , Haribhai Dipica , Williams Calvin B. , Malarkannan Subramanium , Martin Hessner J. , Anne Kwitek E. , Lernmark Åke , Gorski Jack , Wang Demin , Weiler Hartmut 
Published: April 2020

Abstract
In biobreeding rats (BB), the lyp mutation in Gimap5/Ian5 is associated with T-cell lymphopenia and autoimmune insulinitis. To delineate the role of Gimap5 in peripheral T cell survival, we generated Gimap5-deficient mice by targeted disruption. Gimap5−/− mice are viable to adulthood, but develop fulminant autoimmune hepatitis, become moribund, and die at a median age of 12 weeks. Gimap5−/− mice are lymphopenic, displaying an overall 7-fold reduction in peripheral T cells and a 43-fold reduction in peripheral CD8+ T cells. Additionally, Gimap5−/− mice exhibit an early, complete block in both NK and NKT lineage development and a partial block in B cell commitment to T1, T2, and mature subsets. Data from lethally irradiated congenic strains reconstituted with Gimap5−/− bone marrow will also be presented. Taken together, these data indicate that Gimap5 is necessary for peripheral survival of T lymphocytes and development of B lymphocytes and NK cells. Furthermore, these data suggest that Gimap5 deficiency plays a role in the development of autoimmune pathology in the mouse.
In Vitro and In Vivo Evidence That the PP2A Inhibitor SET Regulates IFN-γ Production in Monokine-Stimulated Natural Killer Cells.
Authors: Trotta Rossana , Col Jessica Dal , Allard Jeffrey , Neviani Paolo , Santhanam Ramasamy , Mao Hsiaoyin , Becknell Brian , Yu Jianhua , Modi Aalok , Blaser Bradley W. , Perrotti Danilo , Caligiuri Michael A. 
Published: April 2020

Abstract
Monokines (i.e. IL-12, IL-18 and IL-15) induce natural killer (NK) cells to produce interferon-γ (IFN-γ), which is critical for monocyte clearance of infectious pathogens and tumor surveillance. To identify new regulators of IFN-γ production we performed oligonucleotide array analysis of unstimulated and IL-12- and IL-18-stimulated NK92 cells. Among the subset of mRNAs differentially regulated in monokine-stimulated cells, we found SET, a potent inhibitor of the protein phosphatase type 2A (PP2A). SET mRNA and/or protein levels were upregulated in IL-12/IL-18- and IL-12/IL-15-stimulated primary human NK cells. Interestingly, the SET protein is also selectively increased in the resting CD56bright NK subset, which is a potent producer of IFN-γ relative to the CD56dim NK subset. To determine whether SET positively regulates IFN-γ production by inhibiting PP2A activity, we employed RNAi and interfered with SET expression in NK92 cells. SET downregulation inhibited IFN-γ secretion by IL-12/IL-18, IL-12/IL-15- or IL-15/IL-18-stimulated NK92 cells. By contrast, ectopic SET expression increased IFN-γ production in monokine-stimulated NK92 and primary human NK cells. Because downregulation of SET augmented PP2A activity in NK92 cells, we sought to investigate whether pharmacologic activation of PP2A inhibits the ability of NK cells to produce IFN-γ. Indeed, suppression of IFN-γ expression and secretion was also observed upon treatment of NK92 and primary NK cells with 1,9-dideoxy-forskolin, a known inducer of PP2A activity. Accordingly, NK cells from mice treated with 1.9-dideoxy-forskolin produced less IFN-γ in response to in vivo monokine stimulation than did NK cells from vehicle-treated mice. Mechanistically, activation of PP2A by SET knock-down or 1,9-dideoxy-forskolin treatment leads to inhibition of ERK1/2, p65RelA and STAT5 activity in monokine-stimulated NK cells. Because these signaling molecules are important for IFN-γ production by monokine-stimulated NK cells, our results strongly suggest that monokine induction of SET expression in NK cells is essential for limiting PP2A activity that, otherwise, would negatively impact the ability of NK cells to produce and release optimal levels of IFN-γ.
Protein Tyrosine Phosphatase Receptor-Type O Truncated (PTPROT) Regulates SYK Phosphorylation, Proximal B-Cell Receptor Signaling and Cellular Proliferation.
Authors: Chen Linfeng , Juszczynski Przemyslaw , Takeyama Kunihiko , Aguiar Ricardo C.T. , Shipp Margaret A. 
Published: April 2020

Abstract
The strength and duration of B-cell receptor (BCR) signaling depends upon the balance between protein tyrosine kinase (PTK) activation and protein tyrosine phosphatase (PTP) inhibition. BCR-dependent activation of the SYK PTK initiates downstream signaling events and amplifies the original BCR signal. Although BCR-associated SYK phosphorylation is clearly regulated by PTPs, SYK has not been identified as a direct PTP substrate. In an earlier screen for genes that might contribute to the pathogenesis of diffuse large B-cell lymphoma (DLBCL), we identified and preliminarily characterized a lymphoid PTP termed PTPROt (truncated). PTPROt is a member of the PTPRO family (also designated GLEPP, PTP-ϕ, PTP-oc and PTPu2), a group of highly conserved receptor-type PTPs that are thought to function as tumor suppressor genes. In earlier functional assays, lymphoma cells overexpressing PTPROt exhibited markedly increased G0/G1 arrest, providing the first functional evidence that this PTP regulated tumor growth. We have now used PTPROt substrate trapping mutants to identify SYK as a major substrate of this tissue-specific and developmentally regulated PTP. Co-immunoprecipitation studies confirmed that SYK is also an in vivo PTPROt substrate. To examine the effects of PTPROt on SYK-mediated BCR signaling, we generated B-lymphoma cell lines that could be induced to express either wild type PTPROt (PTPROt-wt) or mutant forms of the phosphatase (D/A, C/S) following doxycycline treatment. Dox-induced overexpression of PTPROt-wt inhibited baseline and BCR-triggered SYK phosphorylation whereas the inactive PTPROt mutants had no effect. More specifically, PTPROt-wt overexpression inhibited the phosphorylation of SYK TYR352, the initial TYR residue phosphorylated upon BCR signaling. Thereafter, we showed that overexpression of PTPROt-wt inhibited BCR-induced activation of two direct SYK targets, the SHC and BLNK adaptor proteins, and downstream signaling events including calcium mobilization and MAPK/ERK activation. BCRs also transmit low-level survival signals in the absence of receptor engagement and BCR-proximal PTPs and SYK likely modulate this “tonic” BCR signaling. These observations and the basal phosphorylation of SYK TYR352 in certain B-lymphoma cell lines prompted us to examine the effects of PTPROt-wt and the PTPROt D/A and C/S mutants on B-cell lymphoma proliferation and apoptosis. PTPROt-wt overexpression, but not that of PTPROt D/A or C/S, dramatically inhibited lymphoma cell proliferation and induced apoptosis in the absence of BCR crosslinking. Taken together, these data indicate that PTPROt modulates SYK phosphorylation and tonic BCR signaling and highlights the importance of this pathway for tumor cell survival.
Hierarchy of Human B Cell Regeneration after Rituximab Therapy.
Authors: Wirths Stefan , Ruprecht Claudia , Bertoni Francesco , Kanz Lothar , Lanzavecchia Antonio 
Published: April 2020

Abstract
Constant immunoglobulin levels throughout life-time may be maintained by either long lived plasma cells or continuous formation of immunoglobulin secreting cells (ISC). Rituximab therapy targets CD20+ B cells, thus selectively spares B progenitor cells and plasma cells, providing an in vivo model to investigate human B cell homeostasis.
After weekly administration of rituximab in patients with follicular lymphoma, peripheral blood B cells are virtually absent (&lt;0,01%), and no ISC are detected. In contrast, during rituximab maintenance therapy (administered in two months intervals) small numbers of B cells can be detected. These carry the phenotype of new formed plasma blasts (CD27++/CD38++/icIg+++ and Ki67+) and are identified as ISC in ex vivo ELISPOT assay.
Surprisingly, ISC detected in treated patients were the same as in healthy donors in absolute numbers, while B cells were still reduced by 100-fold. Only 4 to 6 months after therapy human naive and memory B cells were detected in peripheral blood again. Thus, regeneration of human peripheral blood B lymphocytes follows a hierarchy: Recirculating ISC recover rapidly to normal steady state levels, while B cells with naive or memory phenotype require longer time periods. This hierarchy may represent two elements within the B cells system: a primary system sustaining constant antibody levels and a secondary system providing a diverse repertoire for adaptive immune responses.
Stimulation of TNF-α Production from Monocytes by Anti-Moesin Antibodies in the Serum of Patients with Aplastic Anemia: A Novel Mechanism of Myelosuppression by Autoantibodies.
Authors: Takamatsu Hiroyuki , Feng Xingmin , Lu Xuzhang , Chuhjo Tatsuya , Okawa Katsuya , Nakao Shinji 
Published: April 2020

Abstract
Although aplastic anemia (AA) is a T-cell mediated disease, recent studies have revealed the presence of antibodies (Abs) specific to proteins derived from hematopoietic progenitor cells in the serum of AA patients. It is as yet unclear whether these auto-Abs play some roles in the pathophysiology of AA. We previously demonstrated that Abs specific to moesin, a membrane-cytoskeleton linker protein in the cytoplasm, were detectable in approximately 37% of AA patients. Some reports identified moesin-like molecules on the surface of blood cells such as T cells and macrophages. It is therefore conceivable that anti-moesin Ab in AA patients may bind these immune cells and modulate hematopoietic function of AA patients. To test these hypotheses, we first studied the expression of moesin on various types of blood cells using monoclonal Ab specific to moesin (clone 38/87). Flow cytometry detected the expression of the protein recognized by anti-moesin Ab on T cells and monocytes from healthy individuals, acute monocytic leukemia cells lines including U937 and THP-1, and an acute T-lymphoblastic leukemia cell line, Molt-4, but failed to detect the molecule on CD34+ cells from healthy individuals and myeloid leukemia cell lines as well as B-lymphoblastic leukemia cell lines. Treatment of THP-1 cells with phorbol 12-myristate 13-acetate (PMA)/lipopolysaccharide (LPS) augmented the expression level of moesin. To confirm the expression of the moesin-like protein on the surface of monocytic leukemia cell lines, Molt-4 and THP-1 were treated with sulfo-NHS-SS-biotin, and the cell surface proteins were isolated with avidin-fixed column, and were subjected to Western blotting and peptide mass fingerprinting. Western blotting with anti-moesin monoclonal Abs showed two clear bands of proteins (75 kD and 80 kD); an amino acid sequence compatible with moesin was confirmed in the protein eluted from the 80 kD band. Next, we purified anti-moesin Abs from AA patients’ sera using affinity chromatography with recombinant moesin protein. Western blotting showed binding of the serum-derived Abs to a fraction of surface proteins of Molt-4, U937 and THP-1. When THP-1 cells were incubated in the presence of PMA and LPS with 5 αg/ml of control IgG or anti-moesin Abs derived from an AA patient’s serum, TNF-α production from THP-1 cells stimulated by anti-moesin Abs was 1.9–2.3 times as much as that from the control culture depending on the concentration of LPS. Incubation of THP-1 cells in the presence of monoclonal anti-moesin Abs showed the similar augmentation of TNF-α production. These results indicate that anti-moesin Abs may be involved in the suppression of hematopoiesis of AA patients by stimulating TNF-α production from monocytes.
Regulation of Erythrocyte Volume by Cell Surface Associated Protein Disulfide Isomerase.
Authors: Romero Jose R. , Bicho Manuel D. , Pong Lin-Chien , Greenbowe Joel , Rivera Alicia 
Published: April 2020

Abstract
Acute and chronic clinical manifestations of sickle cell disease (SCD) are based on vaso-occlusion and impaired blood flow. Dense erythrocytes are believed to be important contributors to the vaso-occlusive manifestations of SCD. However, the physiological regulation of erythrocyte hydration status in SCD is not entirely clear. The Gardos channel and K/Cl cotransport are major contributors to sickle erythrocyte dehydration. Protein disulfide isomerase (PDI) on the cell surface catalyzes disulfide formation, causes redox modifications and has been observed to be up-regulated under hypoxic conditions. We now report the detection of PDI in red cells. Western blot analysis revealed a prominent band, migrating at 55 kDa, in ghost preparation from both sickle and normal erythrocytes. To evaluate the role of PDI in Gardos channel activation, we measured charybdotoxin-sensitive K+ influx in the presence of bacitracin, a well-known blocker of PDI activity. When sickle erythrocytes are exposed to bacitracin, Gardos channel activity is significantly reduced (1.4 ±0.2 to 0.8± 0.05 mmol/L cell x min, n=6, P&lt;0.01). We also observed that Gardos channel activity was attenuated by 0.25 mM bacitracin and was maximally inhibited by 3 mM in sickle and normal erythrocytes. Similar results were observed with phenyl arsine oxide and acetyl-thyroxin, two other well-known inhibitors of cell surface PDI activity. We then studied the effects of PDI inhibition on red cell density profiles of sickle and normal human red cells. Analysis of the baseline density profile indicates that erythrocytes have a median density of 1.10 g/mL. This value significantly increased to 1.125 g/mL (n=2) after 3 h of oxygenation/de-oxygenation cycles. However, in the presence of 3 mM bacitracin, the cellular density profile markedly shifted to the left (1.098 g/mL) following deoxygenation/oxygenation. We also investigated the reductive capacity of freshly isolated human erythrocytes by the ferrocyanide-production method. The reductive capacity of normal Hb A red cells was significantly lower than in Hb S containing cells (2.8 ± 1.1 vs 5.1 ± 1.3 mmol/L cell x h, n=18, p&lt;0.0001). Similar results were observed in Santillies and NYKO1, two transgenic mouse models of sickle cell disease, when compared to C57 mice. These results strongly implicate cell surface associated PDI in cellular dehydration and formation of dense sickle erythrocytes by activating K+ efflux via the Gardos channel and suggest that aberrant regulation of PDI activity and/or its expression may contribute to the pathophysiology of Sickle Cell Disease.
p19Arf Cooperation in Shp2-Induced Leukemogenesis.
Authors: Chen Xiaolan , Huang Yuhui , Yan Ling , Wang Zhizhang , Brault Marie , Thompson Maryann , Yang Elizabeth 
Published: April 2020

Abstract
Shp2 is a protein-tyrosine phosphatase which has both proliferation and apoptosis functions in hematopoietic cells. Gain-of-function (GOF) Shp2 mutations are found in Juvenile Myelomonocytic Leukemia (JMML) and Noonan syndrome patients predisposed to JMML, as well as in common childhood leukemias. Genetic data suggest that Shp2 alone is insufficient for leukemogenesis, and other genetic alterations are required, such as disruption of tumor suppressor pathways. We hypothesized that Arf deletion will enhance GOF Shp2-induced leukemogenesis. In colony forming cell assays (CFC), E76K Shp2 expression in arf−/− bone marrow cells led to increased colony number and size compared to E76K Shp2 expression in arf+/− cells or arf−/− cells with vector alone. To assay leukemia induction, arf+/− or arf−/− bone marrow cells transduced with E76K Shp2 mutant were transplanted into lethally irradiated WT C57BL/6 mice. We found that mice transplanted with E76K Shp2/arf−/− bone marrow cells developed leukemia/lymphoma at an accelerated rate compared to mice transplanted with E76K Shp2/arf+/− or vector/arf−/− bone marrow. While the most common hematological disease in mice transplanted with either E76KShp2/arf+/− or vector/arf−/− bone marrow was myeloproliferation, mice with E76K Shp2/arf−/− bone marrow developed predominantly B220+ leukemia. Compared to leukemia in E76K Shp2/arf+/− reconstituted mice, leukemias arising from E76K Shp2/arf−/− bone marrow were much more aggressive, effacing the bone marrow and spleen, infiltrating the liver diffusely and into surrounding tissue. Diseases arising from vector/arf−/− bone marrow were either localized lymphoma or mild myeloproliferation. Both E76K Shp2 expression and arf deletion were associated with the presence of BrdU-positive immature hematopoietic progenitors in the spleens of pre-leukemic mice, consistent with extramedullary hematopoiesis. These findings suggest that arf deletion cooperated with E76K Shp2’s proliferative function to accelerate leukemogenesis.
The Various Procoagulant Effects of PolyP Require Different Minimal Polymer Lengths.
Authors: Smith Stephanie A. , Huyck Jillian , Choi Sharon H. , Morrissey James H. 
Published: April 2020

Abstract
Introduction: Polyphosphates (polyP) are negatively charged, linear phosphate polymers found throughout biology. PolyP in prokaryotes and unicellular eukaryotes is typically hundreds of phosphate units long, while polyP in dense granules of human platelets is ∼75 phosphates long. PolyP is secreted upon platelet activation, and we recently reported that it is a potent hemostatic regulator, acting at multiple points in the clotting cascade: PolyP triggers the contact pathway, accelerates factor V activation, and abrogates the anticoagulant function of tissue factor pathway inhibitor (TFPI). Our previous experiments utilized relatively heterodisperse polyP preparations. We have now isolated polyP fractions of defined polymer lengths and here report the size dependence of the various procoagulant effects of polyP.
Methods: PolyP preparations were carefully size-fractionated by gel electrophoresis (Clark and Wood, Anal. Biochem. 1987, 161:280–290). Clotting assays were performed using pooled normal plasma with or without added recombinant TFPI or 40–50 μM polyP. Clotting was initiated by tissue factor (TF), factor Xa (FXa), or CaCl2 alone (contact pathway).
Results: Of the polyP procoagulant activities tested, triggering the contact pathway required the longest polymers: We observed weak triggering of the contact pathway with ∼75mers and increasing procoagulant activity as polymer lengths increased, with maximal contact pathway activation requiring 450mers or longer. On the other hand, the downstream procoagulant effects of polyP required significantly shorter polyP polymers. (When clotting is triggered by either TF or FXa, further shortening of clotting times by polyP is attributed to accelerated factor V activation.) Minimum polymer lengths required to shorten TF or FXa clot times were ∼37mers, with maximal activity requiring 75mers or longer. Finally, the ability of polyP to abrogate TFPI anticoagulant function required the shortest minimum polymer lengths: Detectable anti-TFPI activity was observed with 11mers, with maximal activity requiring 49mers or longer.
Conclusions: PolyP exhibited different minimum length requirements for its various procoagulant effects, consistent with the idea that polyP interacts with different proteins at various points in the clotting cascade. Contact pathway activation required very long polymers that were similar in size to those in infectious microorganisms. Therefore, triggering of the contact pathway by polyP may contribute to procoagulant/inflammatory responses to infection. In contrast, acceleration of factor V activation by polyP was maximal with polymers of the size released from human platelets. Even shorter polyP polymers abrogated TFPI function, suggesting that at least some of polyP’s anti-TFPI effect may be mediated by mechanisms other than (or in addition to) its ability to accelerate factor V activation.
C3435T Polymorphism of MDR1 Gene Is Not Associated with p-Glycoprotein Function of Leukemic Blasts and Clinical Outcomes in Patients with Acute Myeloid Leukemia.
Authors: Lee Michael Jinpyo , Hur Eun-Hye , Lee Je-Hwan , Choi Seong-Jun , Lee Jung-Hee , Seol Miee , Kim Keun-Hee , Lee Hee-Jung , Kang Ip-Sol , Shim Kyung-Soo , Ryu Seong-Gil , Kang Young-Ah , Lee Young-Shin , Park Chan-Jeoung , Chi Hyung-Sook , Lee Kyoo-Hyung 
Published: April 2020

Abstract
P-glycoprotein (P-gp) function of leukemic blasts is associated with chemotherapy resistance in acute myeloid leukemia (AML). The C3435T polymorphism in the human MDR1 gene has been studied in AML and contradictory results were reported regarding clinical implications of the polymorphism. We investigated the association of C3435T polymorphism of MDR1 gene with P-gp function of leukemic blasts and clinical outcomes in patients with AML excluding M3 subtype. A total of 200 patients, 127 males and 73 females, were included in this study. Median age was 44 years (range, 14–75). All patients were newly diagnosed and were treated with standard ‘7+3’ induction chemotherapy regimens at the Asan Medical Center, Seoul, Korea between May 1999 and November 2006. The C3435T polymorphism was analyzed with PCR/RFLP method. P-gp function of leukemic blasts was measured at diagnosis by the rhodamine-123 efflux assay. The clinico-laboratory data at diagnosis and data on clinical outcomes were obtained from the AMC Leukemia Registry. Genotype frequency of C3435T polymorphism was 71 (35.5%) for CC, 93 (46.5%) for CT, and 36 (18.0%) for TT. There were no significant differences between different genotypes of C3435T polymorphism regarding age, sex, FAB subtypes, initial leukocyte counts, percentages of blasts in bone marrow or blood at diagnosis, and cytogenetic risk groups. P-gp function of leukemic blasts was not significantly different according to the genotype of C3435T polymorphism: 32.2 ± 24.0% for CC, 35.8 ± 21.4% for CT, and 29.1 ± 24.0% for TT (P=0.370). Complete remission was induced in 79.2% for CC, 79.8% for CT, and 77.8% for TT (P=0.969). Overall, relapse-free and event-free survival probabilities at 5 years were 47.6%, 55.4% and 51.0% for CC, 51.3%, 51.0% and 42.5% for CT, and 53.1%, 60.0% and 55.0% for TT (P= 0.594, 0.932 and 0.764, respectively). In conclusion, C3435T polymorphism does not have significant clinical implications in AML regarding P-gp function of leukemic blasts and clinical outcomes.