bluuebunny/crossref_metadata_2025_split huggingface dataset:
p38-MAP Kinase Activation Followed by BIM Induction Is Critical for Glucocorticoid-Induced Apoptosis in Lymphoblastic Leukemia Cells.
Authors: Harada Hisashi , Lu Jianghua , Quearry Bonnie 
Published: April 2020

Abstract
Glucocorticoids (GC) are common components in chemotherapeutic protocols for lymphoid malignancies. GC-induced apoptosis requires the BCL-2 family-regulated pathway, but it remains largely undefined how the signaling pathways and their downstream target molecules, including the BCL-2 family members, participate in GC-induced cell death. BIM (BCL-2-interacting mediator of cell death) identified as a BH3-only protein induces apoptosis and its activity can be modulated at the transcriptional and post-translational levels. BIM is required for apoptosis induced by Taxol, dexamethasone, Gleevec, and HDAC inhibitors, indicating that this protein has a role in anticancer therapy against human malignancies. However, the molecular mechanisms how BIM is regulated and how BIM regulates downstream cell death machinery are not entirely clear.
In this study, we used a T cell acute lymphoblastic leukemia (T-ALL) cell line, CCRF-CEM to examine the molecular mechanisms of Dexamethasone (Dex)-induced cell death. Dex treatment induces the mRNA and protein of BIM in CCRF-CEM cells before the onset of apoptosis. Bim mRNA induction is completely inhibited by a transcription inhibitor, actinomycin D. However, a protein synthesis inhibitor, cycloheximide treatment augments Dex-induced Bim mRNA accumulation (super-induction), suggesting that de novo protein synthesis is not required for the induction of Bim mRNA. BIM protein induction is inhibited by actinomycin D or cycloheximide, thus new BIM protein synthesis is required for its accumulation. These data indicate that Dex treatment increases BIM levels mainly by new mRNA synthesis (transcription). In contrast, the overall expression of the multi-domain pro-apoptotic effectors, BAX and BAK, are constant. The anti-apoptotic members, BCL-2, MCL-1 and BCL-XL, also show constant expression. Down-regulation of BIM by shRNA strongly inhibits Dex-induced apoptosis. Thus, BIM is an essential molecule in Dex-induced apoptosis in CCRF-CEM cells. We then investigated the upstream signaling pathways to induce BIM. p38-mitogen activated protein kinase (p38-MAPK) is activated by Dex treatment prior to BIM induction. Treatment with a p38-MAPK inhibitor, SB203580, dramatically reduces Dex-induced apoptosis as well as BIM induction.
These findings indicate that BIM induction through p38-MAPK activation is a critical pathway in GC-induced cell death.
Novel Functions for a Fibrinolytic Pathway in Controlling the Stem Cell Niche.
Authors: Akiyama Haruyo , Lund Leif R. , Morita Yohei , Nakauchi Hiromitsu , Ogawa Hideoki , Okumura Ko , Dano Keld , Werb Zena , Heissig Beate , Hattori Koichi 
Published: April 2020

Abstract
Hematopoietic stem cells (HSCs) reside within specific niches, where they are maintained by self-renewal and can be mobilized into circulation, but the underlying mechanisms are still unknown. The hematopoietic niche comprises of a local network of stromal cells (like fibroblasts, endothelial cells), accessory cells (T lymphocytes, monocytes), their products (extracellular matrix (ECM) and cytokines, capable of influencing self-renewal, proliferation and differentiation of HSCs. One such ECM molecule is fibrin, which can be found along surfaces in the bone marrow. Its precursor fibrinogen can maintain hematopoiesis in long-term bone marrow liquid cultures and increase the colony size of bone marrow cells cultured in semisolid medium containing serum. Plasmin, a serine protease, proteolytically degrades fibrin and fibrinogen. Plasminogen (Plg), its inactive proenzyme is present in the plasma, interstitial fluid or adherent to ECM proteins and can be activated by tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator (uPA) and other serine proteasesWe demonstrated previously that matrix metalloproteinase-9 (MMP-9) controls stem cell recruitment from the bone marrow niche. During fibrinolysis plasmin participates in the consecutive activation of MMPs. We initially wished to determine if this fibrinolytic cascade might be involved in regulating the stem cell niche. In this study we provide novel mechanistic data demonstrating that classical fibrinolytic factors-Plg/Plasmin are involved in recruitment of HSCs into the cell cycle following myelosuppression and their mobilization from the BM niche influencing cell differentiation, proliferation and migration by activating niche cells to secrete typical niche factors, including Kit-ligand and stromal cell-derived factor-1 (SDF-1, CXCL12). Deletion of Plg in mice prevented hematopoietic stem cells from entering the cell cycle and undergoing differentiation after myelosuppression, leading to the death of the mice. By Plg-dependent and Plg-independent pathways, tissue-type plasminogen activator activated matrix metalloproteinases and released Kit ligand and stromal cell-derived factor-1 from niche cells, thereby promoting stem cell recruitment and proliferation. We conclude that the fibrinolytic cascade plays a critical role in controlling the exit from the stem cell niche and that this represents a general stem cell regulatory axis.
The Hematopoietic Supportive Potential of Human Mesenchymal Stromal Cells Is Associated with Expression of Cadherins.
Authors: Roderburg Christoph , Diehlmann Anke , Wein Frederik , Faber Anne , Krause Ulf , Frankhauser Maria , Seckinger Anja , Ho Anthony D. , Wagner Wolfgang 
Published: April 2020

Abstract
Self renewal and differentiation of hematopoietic stem cells (HSC) are governed by interaction with the supportive microenvironment of the bone marrow. Secreted factors as well as specific cell adhesion proteins are involved in this interaction. As an in vitro model system, the hematopoietic microenvironment can be mimicked by supportive mesenchymal stromal cells (MSC).
We have compared the supportive potential of human MSC from bone marrow (BM) isolated under two different culture conditions (BM-MSC-M1 and BM-MSC-M2), from adipose tissue (AT) and umbilical cord blood (CB) that were all cultivated as described before (Wagner et al. Exp Hematol.2005;11:1402–1416.). As controls we have used the human fibroblast cell line HS68 and the murine fetal liver cell line AFT024. CD34+ cells were isolated from human cord blood and cultured in direct contact with irradiated stromal cells. After four, seven and twelve days the immunophenotype of the hematopoietic cells was analyzed by flow cytometry. Many progenitor cells cultured on BM-MSC or AFT024 maintained a primitive phenotype of CD34+/CD38- cells whereas the proportion of these cells was reduced upon cultivation with CB-MSC and cells cultured on AT-MSC and HS68 displayed a significantly higher expression of CD38 and lower expression of CD34. Furthermore, long term culture initiating-cell (LTC-IC) assays were performed on the different feeder layer. LTC-IC frequency was significantly higher on BM-MSC that were isolated under the two different culture conditions (BM-MSC-M1 1,15 ±0.11%; BM-MSC-M2 1.14±0.08%) and on CB-MSC (1.10±0.13%) compared to AT-MSC (0.32±0.09%) and HS68 (0.67±0.12%). We have compared gene expression profiles of BM-MSC-M1, BM-MSC-M2, CB-MSC, AT-MSC and HS68 by cDNA microarray analysis (51,144 different cDNA clones of the RZPD3 Unigene Set). Differential expression of various genes correlated with the observed differences in supportive potential. Among these were adhesion proteins like N-cadherin, cadherin11, fibronectin1, various integrins (ITGA1, ITGA5 and ITGB1) and VCAM1 as well as secreted proteins including osteonectin, CTGF and SDF1. Westerblot analysis verified on protein level that cadherin11, N-cadherin, and ITGB1 were highly expressed on BM-MSC as compared to AT-MSC and HS68 fibroblasts.
In conclusion MSC from human bone marrow or from umbilical cord blood support to a significantly higher degree the maintenance and proliferation of primitive hematopoietic progenitors than MSC derived from adipose tissue. This affinity correlated with up-regulation of cadherin11, N-cadherin and intergrin-beta1 on BM-MSC and CB-MSC.
Identification of Meis1 Target Genes Involved in the Induction of AML in Collaboration with NUP98-HOXD13.
Authors: Palmqvist Lars , Argiropoulos Bob , Humphries R. Keith 
Published: April 2020

Abstract
Meis1, a homeodomain-containing HOX cofactor, cooperates with multiple native and NUP98-HOX fusion proteins to accelerate the onset of AML. We have recently shown that enforced overexpression of Meis1 in NUP98-HOX transduced bone marrow (bm) cells leads to a marked elevation in the tyrosine receptor molecule, Flt3 transcription and subsequent activation of Flt3 signaling pathways (Blood. Aug 1, 2006). Furthermore, overexpression of wildtype Flt3 is sufficient to collaborate with NUP98-HOX fusions to induce AML in mice, albeit with a longer disease latency than NUP98-HOX+Meis1-mediated AML. To further study the molecular mechanisms underlying Meis1’s strong pro-leukemic effect together with HOX proteins, we conducted a structure-function analysis of Meis1 in the context of NUP98-HOXD13 (ND13) leukemogenesis. We show that the homeodomain (ΔHD, amino acids 272–335) is required for Meis1 collaboration with ND13 while the N-terminal domain (ΔN, amino acids 1–67) is dispensable for leukemogenesis but decelerates the disease onset. Interestingly, primary bm cells transduced with ND13-Meis1ΔHD or ND13-Meis1ΔN revealed no significant upregulation of Flt3 mRNA levels in either case. These results suggest that Meis1 triggers additional Flt3-independent pathways to accelerate leukemia development. Thus, the retained leukemogenic properties of ND13-Meis1ΔN and the longer latency observed for NUP98-HOX fusions and Flt3 overexpression to induce leukemia point to additional roles for Meis1 in induction of leukemia independent of its ability to upregulate Flt3. To search for these additional Meis1-induced leukemic pathway(s), we used the Affymetrix GeneChip MOE430 to compare the gene expression profiles of bm cells transduced with ND13, ND13+Meis1, ND13+Meis1ΔHD and Meis1ΔN. Interestingly, the gene array results indicate that only a relatively small number of genes (72 increased and 64 decreased) differed significantly between the non-leukemic Meis1ΔHD and the leukemic Meis1 bm cells and these genes are all strong candidates as to be direct Meis1 target genes. A significant number of these genes are involved in cytokine receptor pathways as revealed by gene ontology analysis and, furthermore, promoter analysis revealed the presence of putative Hox, Meis1 and Pbx binding sites in several of these. The gene array results also indicate that only a small fraction of genes differed significantly between the non-leukemic Meis1ΔHD and the leukemic Meis1ΔN bm cells suggesting that these genes could be involved in Flt3-independent Meis1 leukemogenic activity.
The Groucho Corepressors, TLE1 and TLE4, Are Candidate Del(9q) AML Tumor Suppressor Genes That Can Complement AML1-ETO.
Authors: Sweetser David A. , Dayyani Farshid , Wang Jianfeng , Zhang Yuntian , Zaidi Tanweer 
Published: April 2020

Abstract
Deletion of the long arm of chromosome 9, del(9q), is one of the most common cytogenetic abnormalities seen with t(8;21) AML. This close association suggests that that the loss of a critical gene(s) on chr9q cooperates with the AML1-ETO (RUNX1-MTG8) fusion gene produced by t(8;21) in leukemogenesis. We recently mapped the commonly deleted region of del(9q) AML to less than 2.4 Mb and identified all the genes mapping to this region. Extensive sequence analysis of these genes failed to reveal obvious mutations in del(9q) AML samples although the expression of a number of these genes appeared low specifically in del(9q) samples (Genes Chr and Ca 44, 279–291). To help determine the critical affected genes in the del(9q) CDR we used an siRNA expression library directed against all del(9q) CDR gene transcripts in an in vitro complementation assay with AML1-ETO. Inducible expression of AML1-ETO in U937 cells (U937T-A/E, Mol Cell Biol 21, 5577–5590) leads to cell cycle arrest and apoptosis. We introduced the del(9q) CDR siRNA library into U937T-A/E cells with the goal of finding siRNAs that rescued these cells after induction of AML1-ETO expression. The most commonly recovered siRNA using flanking PCR primers was directed against TLE1. Transducin-like enhancer of split (TLE) 1 and TLE4, are two members of the Groucho family of co-repressors that map adjacent to the del(9q) CDR. We subsequently demonstrated that knockdown of either TLE1 or TLE4 with specific siRNAs were able to overcome the induced cell cycle arrest in U937T-A/E cells. We further demonstrate that knockdown of either TLE in the AML1-ETO+ Kasumi-1 cell line increased cell division, increased expression of cyclin D1 and Ki67 and reduced the G0/G1 fraction, while expression of TLE1 or TLE4 in Kasumi-1 cells induced cell cycle arrest and apoptosis. Understanding how various mutations work cooperatively to produce a malignant phenotype is one of the great challenges in oncology. The TLEs are known to inhibit NFKB as well as Wnt signaling, two pathways implicated leukemic stem cell expansion. TLE inhibition may represent an important cooperating mutation with AML11-ETO in AML that links Wnt signaling and core binding protein transcription factors. Studies are currently underway to demonstrate this cooperativity and more directly evaluate the role of TLE inhibition in leukemogenesis.
Ceramide Kinase Promotes Store Dependent Ca2+ Signals and Enhances Phagolysosomal Fusion in COS-1 Cells.
Authors: Hinkovska-Galcheva Vania T. , Clark Andrea J. , Kindzelskii Andrei L. , VanWay Susan M. , Huang JiBiao , Shanley Thomas P. , Shayman James A. , Petty Howard R. , Boxer Laurence A. 
Published: April 2020

Abstract
We showed that ceramide-1-phosphate (C1P) is formed by ceramide kinase (CERK). In this study the mechanism by which C1P enhanced phagocytosis and phagolysosomal formation in COS-1 cells expressing hCERK was evaluated. We hypothesized that CERK has a unique role in inducing Ca2+ signaling during Fc-mediated phagocytosis via its generation of C1P. Furthermore, we also hypothesize that CERK mediated changes in Ca2+ levels were not direct but required the participation of a store operated channels (SOC). In some systems, evidence suggests that SOC might be related to transient receptor potential channel (TRP) homologues. This hypothesis was tested using cell lines with different levels of expression of CERK and by using series of pharmacological inhibitors. To monitor subcellular localization of hCERK, red fluorescent protein (RFP)-tagged hCERK was created and confocal microscopy was performed. To study dynamic changes in Ca2+ during phagocytosis, imaging microscopy was used. Phagolysosome formation was evaluated by labeling the cells with lysotracker red DND-99 at 37oC and followed by imaging. When challenged with EIgG, hCERK transfected cells increased phagocytosis and simultaneously increased C1P. During subcellular fractionation of hCERK transfected cells, CERK translocates during activation from the cytosol to a lipid raft fraction. We also observed that TRP-1 accumulated at the site of CERK translocation in lipid rafts. Microfluorimetry of indo-1 revealed that Ca2+ oscillations in COS-1 cells, stably transfected with Fcγ RIIA/hCERK, were much higher compared to Fcγ RIIA transfected cells. The enhanced Ca2+ signals were accompanied by enhanced phagolysosome formation. hCERK transfected cells had a mean rate of fusion of 90%, compare to 76% ( catalytic inactive mutant of CERK-G198DhCERK), 65% (Fcγ RIIA), and 70% (Vector) transfected cells. The indo-1 intensity spikes demonstrated significantly higher Ca2+ intensities for phagosomes associated with the hCERK transfectants. The SOC inhibitor SKF96365 blocked enhanced Ca2+ signaling in the hCERK transfected cells. Similarly SOC inhibitors reduced the phagocytic index and phagolysosomal fusion in hCERK transfected cells. Our data showed that there is a co-localization of EIgG and CERK as well as caveolin-1 and CERK on the surface of the phagocytic cells. Site directed mutagenesis demonstrated that the kinase activity of hCERK was required for higher Ca2+ signals. Pharmacological inhibitors revealed that this Ca2+ signal operates through SOC. Our results show that introduction of the hCERK gene in cells alters the functional behavior of intracellular signals and supports the role of C1P in promoting membrane fusion by modulating Ca2+ signaling through activation of SOC
The Gene Locus Distribution of HLA in the Chinese Population by High Resolution Typing of HLA-A,B,C,DR,DQ and Its Effect on Stem Cell Transplantation.
Authors: Hou Jian Q. , He Jun , Chen Zi X. , Bao Xiao J. , Yao Li , Di Wen Y. , Zhang Hui , Zhang Jian 
Published: April 2020

Abstract
Allogeneic hematopoietic stem cell transplantation (AHSCT)can potentially cure a variety of hematopoietic and congenital metabolic disorders. Ideally, one would hope to find identical unrelated donors who are genotypically identical to the patients at all HLA loci. To explore distribution of HLA gene subsites in the Chinese population and its effect on AHSCT. High resolution typing of HLA-A,B,Cw,DR,DQ were performed on 101 cases from the data base of China Marrow Donor Program (CMDP) donors and 76 patients (ALL 23cases, CML 32cases, AML 23cases)by using PCR-SSP and PCR-SSO. The HLA subsites with highest gene frequency were A*0201, A*0207, A*1101, A*2402, B*4001,B*4601, C*0102, C*0702, DR*0901, DR*1202,DQ*0301and DQ*0303(PF&gt;0.2). The subsites with secondary high frequency were A*3101,A*3303,B*1301,B*1302,B*1501, B*5101,B*5801,C*0303,C*0304,C*0602,C*0801, DR*0405, DR*0406,DR*0701,DR*0803, DR*1501,DQ*0202,DQ*0302,DQ*0401, DQ*0502, DQ*0601(PF&gt;0.1). The phenotypic frequency of A*1101,B*4601,C*0102,DR*0901, DQ*0303 in the Chinese population was as high as 0.38, Among which B*4601and DR*0901 were the single allele with highest frequency. The gene locus in HLA with linkage disequilibrium were A*0207-B*4601- DR*0901-C*0102-DQ*0301or0303;A*0201-B*4601-DR*0405 or DR*0803- C*0102-DQ*0302orDQ*0502;A*3001-B*1302-DR*0701-C*0602-DQ*0202 and A*3303- B*5801-DR*0901-C*0302-DQ*0303 or DQ*0302, respectively. CMDP demonstrated the effect of HLA-A and B allele matching on the development of severe acute GVHD and on the overall survival rate. HLA-DR matching are less effective than classIfor clinical outcomes. Additionally, the HLA-C mismatch was found to have a synergistic effect on the acute GVHD and survival rate when another HLA locus mismatch coexisted. Our results demonstrated that the frequency distribution of HLA gene subsites is important for selecting more suitably matched donor for HSCT and is helpful for making donor recruitment strategy. As the HLA-Cw played important role on aGVHD occurrence and the failure of BMT, more attention should be paied on HLA-Cw allele in clinical HSCT.
The Role of Rab38 in Rat and Human Platelet Function Defects.
Authors: Datta Yvonne H. , Rangel-Filho Artur , Wu Francisca C. , Stephens Kenyatta W. 
Published: April 2020

Abstract
The fawn-hooded hypertensive (FHH) rat is a model of a moderate platelet function defect manifest by severely prolonged tail bleeding times, decreased platelet aggregation to ADP and collagen, and decreased ADP release from dense granules. The FHH rat has a mutation at the ATG start site of the gene encoding the small GTP-binding protein Rab38, which we have identified as being strongly associated with the bleeding phenotype. We have further characterized expression of Rab38 in the FHH rat and in humans. As assessed by reverse transcriptase PCR, we found that Rab38 is expressed in a wide variety of tissues from FHH and control Brown-Norway (BN) rats. However, by western blotting there was no protein expression of Rab38 in the FHH tissues, confirming that the mutation results in a severe defect at the protein level. Confocal immunofluorescence microscopy demonstrated Rab38 localized in punctuate cytoplasmic granules in human megakaryocytic cell lines. The Rab38 staining colocalized with von Willebrand factor (VWF) found in alpha-granules. However, we did not demonstrate any significant colocalization of Rab38 with serotonin, a marker of dense granules or Lamp-1, a marker of lysosomes, in the megakaryocytic cell lines. We also demonstrated colocalization of Rab38 with VWF in normal human platelets. We hypothesized that some humans with mild to moderate platelet dysfunction may have abnormalities of Rab38. We sequenced the entire coding region and intron-exon boundaries of the Rab38 gene in six patient samples collected at Emory University for the CDC Women with Bleeding Disorders and Menorrhagia Study. Four of these patients had detectable platelet aggregation abnormalities and two did not. In this limited initial study, no obvious mutations or new polymorphisms were detected in the Rab38 gene. Numerous known polymorphisms were detected, but no clear pattern was associated with the platelet aggregation abnormalities as of yet. These results along with our previous findings in the FHH rat suggest that Rab38 may be involved in a general secretion mechanism in platelets. It remains to be seen whether mutations in the Rab38 gene account for some of the platelet function defects seen in humans, but we are actively pursuing this hypothesis.
Blood Platelets Organize Pro- and Anti-Angiogenic Factors into Separate, Distinct Alpha Granules: Implications for the Regulation of Angiogenesis.
Authors: Italiano Joseph , Richardson Jennifer L. , Folkman Judah , Klement Giannoula 
Published: April 2020

Abstract
In addition to contributing to hemostasis, platelets are increasingly being viewed as major contributors to wound healing and tumor growth. This can be attributed to the many angiogenesis regulatory proteins selectively taken up and stored within the alpha granules of platelets and deposited in the local environment of a tumor or wound upon platelet aggregation and adhesion. Platelets contain both pro and anti-angiogenesis regulators, and the mechanisms by which they modulate angiogenesis are unclear. To address this question, we examined the localization of both pro-and anti-angiogenic factors in platelets and megakaryocytes using immunostaining and confocal microscopy. Double immunofluorescence microscopy revealed that VEGF (an angiogenesis stimulator) and endostatin (an inhibitor), are localized in separate and distinct alpha granules. Immunofluorescence for thrombospondin 1, basic FGF, platelet factor 4, and placental growth factor subsequently confirmed the separate and distinct alpha granules. These observations, in combination with a recent report that proteinase-activated receptors (PARs) selectively modulate the release of angiogenesis regulators from platelets (Ma et al., PNAS 102:216), motivated the hypothesis that these distinct populations of alpha granules could undergo selective release as well. To investigate whether PARs facilitate differential release of alpha granules, we treated human platelets with PAR agonists or antagonists and assayed for differential alpha granule release. The selective PAR1 agonist stimulated the release of endostatin-containing alpha granules and suppressed the release of VEGF-containing alpha granules. Furthermore, confocal microscopy studies revealed that in the presence of the PAR1 antagonist (SCH79797), thrombin stimulated the release of endostatin-containing alpha granules, but not VEGF-containing granules. These observations suggest that platelets may be organizing angiogenesis regulatory proteins into pharmacologically and morphologically distinct populations of alpha granules, which are then susceptible to differential regulation upon platelet activation.