bluuebunny/crossref_metadata_2025_split huggingface dataset:
Amplification of Sca-1+ Lin-WGA+ cells in serum-free cultures containing steel factor, interleukin-6, and erythropoietin with maintenance of cells with long-term in vivo reconstituting potential
Authors: Rebel VI , Dragowska W , Eaves CJ , Humphries RK , Lansdorp PM 
Published: October 2019

Abstract
Normal murine bone marrow (BM) cells were sorted on the basis of low forward and orthogonal light scatter properties, Sca-1 expression (Sca-1+), lack of staining with a cocktail of mature hematopoietic lineage markers (Lin-), and binding of wheat germ agglutinin (WGA+). This approach allowed the reproducible isolation of a very small subpopulation (0.037% +/-0.023% of all nucleated BM cells) that was approximately 400-fold enriched in cells capable of reconstituting both lymphoid and myeloid lineages in lethally irradiated recipients. Transplantation of 30 or 10 of these Sca-1+Lin-WGA+ cells resulted in > or = to 20% donor-derived nucleated peripheral blood cells 3 months posttransplantation in 100% and 22% of the recipients, respectively. When Sca-1+Lin-WGA+ cells were cultured in serum-free medium supplemented with Steel factor, interleukin-6 (IL-6), and erythropoietin (with or without IL-3), a large increase in total cell number, including cells with an Sca-1+Lin-WGA+ phenotype was observed. Single cell cultures showed that 90% to 95% of the input cells underwent at least one division during the first 2 weeks and the remainder died. Interestingly, this proliferative response was not accompanied by a parallel increase in the number of cells with both lymphoid and myeloid repopulating potential in vivo, as quantitation of these by limiting dilution analysis showed they had decreased slightly (1.3-fold) but not significantly below the number initially present. These results demonstrate that Sca-1+Lin-WGA+ cells with long-term repopulating potential can be maintained for 2 weeks in a serum-and stroma cell-free culture, providing a simple in vitro system to study their behavior under well-defined conditions. The observed expansion of Sca-1+Lin-WGA+ cells in vitro without a concomitant increase in reconstituting cells also shows that extensive functional heterogeneity exists within populations of cells with this surface phenotype.
Interleukin-12 induces tyrosine phosphorylation and activation of 44-kD mitogen-activated protein kinase in human T cells
Authors: Pignata C , Sanghera JS , Cossette L , Pelech SL , Ritz J 
Published: October 2019

Abstract
Interleukin-12 (IL-12) is a novel cytokine that enhances numerous functional activities of human T cells and natural killer (NK) cells. The present studies were undertaken to characterize some of the early signaling events following IL-12 stimulation of mitogen-activated normal T cells. In these cells, IL-12 induces rapid tyrosine phosphorylation of proteins of 21, 44, and 54 kD. However, IL-12 does not induce tyrosine phosphorylation in normal resting T cells. In conjunction with increased tyrosine phosphorylation of several substrates, IL-12 stimulation resulted in increased in vitro kinase activity of immunoprecipitated tyrosine phosphorylated proteins. The 44- kD protein has been characterized as one isoform of the mitogen- activated protein (MAP) kinase family. Increased tyrosine phosphorylation of MAP kinase following IL-12 stimulation was also associated with enhanced enzymatic activity of this protein in vitro as measured by myelin basic protein phosphotransferase assay. These studies identify MAP kinase as one of the intracellular elements of the IL-12 signaling pathway in human T cells.
The 47-kD protein increased in neutrophil actin dysfunction with 47- and 89-kD protein abnormalities is lymphocyte-specific protein
Authors: Howard T , Li Y , Torres M , Guerrero A , Coates T 
Published: October 2019

Abstract
A male child born of related parents suffered recurrent infections because of neutrophil actin dysfunction with increased amounts of a 47- kD protein and decreased amounts of an 89-kD protein (NAD 47/89). The patient and family members were studied to define the nature of the abnormal proteins and to examine their role in the functional defects of neutrophil actin dysfunction (NAD) 47/89 polymorphonuclear neutrophils (PMNs). NAD 47/89 PMNs are defective in motility, microfilamentous cytoskeletal structure, and formyl peptide-induced actin polymerization and express increased amounts of a 47-kD protein and decreased amounts of an 89-kD proteins intermediate abnormality in amount of 47-kD and 89-kD proteins in PMNs from parents and a female sibling suggest the disease is an autosomal recessive disorder. Immunoblots with monoclonal antibody (MoAb1) and polyclonal antibody raised to 47-kD protein showed the 89-kD protein is antigenically distinct from the 47-kD protein and the 89-kD protein is not gelsolin. 125I-actin binding to one-dimensional (1 D) and 2 D gels of PMN proteins from NAD 47/89 proband, family members, and controls showed the 47-kD protein binds actin, is acidic (pl = 4.5 to 4.7), is recognized by the MoAb1, exists on 2-D gels as three distinct actin binding species (MWapp 52 kD, 47-kD, and 44-kD), and is present in control PMNs in lesser amount than in PMNs of NAD 47/89 proband or parents. Immunoaffinity purification of the 47 kD actin binding protein on MoAb1 matrix yielded a multimolecular complex with proteins of MWapp 180 kD, 71 kD, 47 kD and actin. Cloning, sequencing, and expression of a 1.58-kb cDNA selected for MoAb1 reactivity from a HL60 expression library and microsequence of native PMNs, 47-kD actin binding protein showed the overexpressed 47-kD protein is lymphocyte-specific protein 1 (LSP1), which is a known actin binding protein. The results show LSP1 is expressed in PMNs and suggest overexpression of LSP1 is related to the motility and cytoskeletal abnormalities in NAD 47/89 PMNs.
Expression of sialosyl-Tn in colony-forming unit-erythroid, erythroblasts, B cells, and a subset of CD4+ cells
Authors: Muroi K , Suda T , Nakamura M , Okada S , Nojiri H , Amemiya Y , Miura Y , Hakomori S 
Published: October 2019

Abstract
The epitopes Tn and sialosyl-Tn are expressed on erythrocytes of individuals with a very rare blood group, who often suffer from “Tn syndrome.” We surveyed expression of Tn and sialosyl-Tn in normal blood cells, malignant transformed cells, and progenitor stem cells from bone marrow (BM). An anti-Tn antibody, IE3, and an anti-sialosyl-Tn antibody, TKH2, were used in this study. TKH2 reacted with erythroblasts, B cells, and a subset of CD4+ cells; but not with erythrocytes. Erythroblastic cell lines (K562, HEL, and UT7/EPO) and B- cell lines (Daudi, Raji, and B-cell lines transformed by Epstein-Barr virus) showed reactivity to TKH2. Similar results from the reactivity of TKH2 with transformed cells from leukemia patients and lymphoma patients were obtained; TKH2 reacted with blasts from erythroleukemia (M6; for 4 of 4 cases) and with lymphocytes from B-cell chronic lymphocytic leukemia (3 of 3), B-cell lymphoma (5 of 5), and CD4+ adult T-cell leukemia (4 of 4), but did not react with blasts from acute myeloid leukemia (M0 to M5; 0 of 22) or acute lymphoid leukemia (B- lymphoid leukemia, 0 of 11; T-lymphoid leukemia, 0 of 2; undifferentiated leukemia, 0 of 1). IE3 did not react with all of the tested cells. CD2-CD19-TKH2+ normal BM cells (BMC) contained blasts and various maturation stages of erythroblasts. The TKH2+ cells produced a large number of colony-forming unit-erythroid (CFU-E) colonies, whereas they produced a small number of burst-forming unit-erythroid colonies and CFU-granulocyte-macrophage colonies. CD34+ normal BMC did not express Tn and sialosyl-Tn. These findings suggest that sialosyl-Tn expresses in CFU-E to erythroblasts.
von Willebrand disease in the RIIIS/J mouse is caused by a defect outside of the von Willebrand factor gene [published erratum appears in Blood 1995 Sep 15;86(6):2461]
Authors: Nichols WC , Cooney KA , Mohlke KL , Ballew JD , Yang A , Bruck ME , Reddington M , Novak EK , Swank RT , Ginsburg D 
Published: October 2019

Abstract
An animal model for human type I von Willebrand disease (vWD) has been previously described in the inbred mouse strain RIIIS/J. Murine vWD is characterized by a prolonged bleeding time, normal von Willebrand factor (vWF) multimer distribution, autosomal dominant inheritance, and proportionately decreased plasma vWF antigen, ristocetin cofactor, and factor VIII (FVIII) activities. To study the molecular genetics of murine vWD, a portion of the vWF gene surrounding exon 28 was cloned, sequenced, and used to develop two informative DNA sequence polymorphisms for rapid genotyping by DNA polymerase chain reaction. RIIIS/J mice were crossed with PWK/Ph mice, an inbred line of Mus musculus musculus, and the F1 progeny backcrossed to the parental PWK/Ph strain. vWF antigen levels in F1 mice were not significantly different from the parental RIIIS/J strain but were markedly decreased compared with the parental PWK/Ph mice. Genetic linkage analysis of 104 backcross progeny showed no correlation between vWF antigen level and vWF genotype. These data indicate that murine vWD is caused by a defect at a novel genetic locus, distinct from the murine vWF gene. The distribution of vWF antigen levels among backcross progeny suggests the presence of one major dominant vWD gene in the RIIIS/J mouse with possible modifying contributions from one or more additional minor loci. These observations may provide new insights into the molecular basis and variable expressivity of human vWD.
Novel retinoic acid, 9-cis retinoic acid, in combination with all-trans retinoic acid is an effective inducer of differentiation of retinoic acid-resistant HL-60 cells
Authors: Kizaki M , Nakajima H , Mori S , Koike T , Morikawa M , Ohta M , Saito M , Koeffler HP , Ikeda Y 
Published: October 2019

Abstract
Recent studies have shown that a high proportion of patients with acute promyelocytic leukemia (APL) achieve complete remission after treatment with all-trans retinoic acid (RA). Nevertheless, despite an initial good response, most patients that received continuous treatment with all-trans RA relapse and develop RA-resistant disease. The 9-cis RA is a high-affinity ligand for retinoid X receptors (RXRs) and also binds efficiently to retinoic acid receptors (RARs); all-trans RA is a ligand for RARs. Both alone are able to induce differentiation of wild-type HL- 60 cells. We found that neither all-trans RA nor 9-cis RA (< 2 x 10(-6) mol/L) induced differentiation of RA-resistant HL-60 cells into either mature granulocytes or monocytes. However, morphologic differentiation of the RA-resistant HL-60 cells was induced by 10(-6) mol/L all-trans RA combined with various concentrations (10(-12) to 10(-6) mol/L) of 9- cis RA. Electron microscopic examination also confirmed that the combination of both retinoids induced RA-resistant HL-60 cells to differentiate to mature granulocytes. Functional analysis of differentiation (NBT reduction activity) confirmed the necessity of both analogs to induce differentiation. Also, expression of myeloid- specific differentiation antigens (CD11b and CD14) as well as migration inhibitory factor-related protein (MRP)-8/14 mRNAs were upregulated only in the presence of both retinoids in a dose-dependent manner. In these conditions 3H-thymidine incorporation was inhibited and numbers of viable cells were decreased, suggesting that all-trans RA with 9-cis RA may inhibit cell growth and induce differentiation of RA-resistant HL-60 cells into mature granulocytes. These studies suggest that 9-cis RA in combination with all-trans RA is an effective inducer of RA- resistant HL-60 cells and may have implications for both the biology of retinoids and clinical treatment of RA-resistant acute myelogenous leukemia, including APL patients.
Retinoic acid-resistant HL-60 cells exclusively contain mutant retinoic acid receptor-alpha
Authors: Li YP , Said F , Gallagher RE 
Published: October 2019

Abstract
Sequence analysis of the retinoic acid receptor-alpha (RAR alpha) gene from a subline of HL-60 cells (RA-res) stably resistant to all-trans retinoic acid (RA) disclosed a single-base change in codon number 411, the same C to T transition previously reported in an independently selected HL-60 RA resistant clone by Robertson et al (Blood 80:1885, 1992). This mutation eliminates a FokI restriction endonuclease site. Using primers framing this mutation in exon 9 of the RAR alpha gene, we showed that polymerase chain reaction products amplified from either mRNA or genomic DNA templates from the RA-res subline were completely resistant to FokI digestion whereas those from wild-type (wt) HL-60 cells could be digested to completion. The lack of a normal allele in the RA-res cells was confirmed by mixing experiments and hybridization analyses. Southern blot analysis of DNA from the RA-res and wt cells versus control placental DNA indicated that the RAR alpha gene is not haploid. The independent isolation of the same RAR alpha mutation in different laboratories suggests either that the mutation exits in a small subpopulation in the wt line or that this is a mutational “hot spot.” Furthermore, the results indicate that if a dominant negative mode of resistance is involved in the RA-res subline, this must involve interference with the function of heterologous receptor proteins such as the retinoid X receptors. The lack of any normal RAR alpha in this subline may facilitate studies of the mode of action of retinoids.
C3a activates the respiratory burst in human polymorphonuclear neutrophilic leukocytes via pertussis toxin-sensitive G-proteins
Authors: Elsner J , Oppermann M , Czech W , Kapp A 
Published: October 2019

Abstract
In contrast to C5a, which represents a well-established potent activator of the respiratory burst in polymorphonuclear neutrophilic granulocytes (PMN), the functional role of C3a in the activation of PMN is, so far, poorly understood. Herein, the potential role of human C3a in the activation of the respiratory burst in human PMN was investigated. The release of reactive oxygen species (ROS) of PMN from healthy donors was measured by lucigenin-dependent chemiluminescence. C3a dose-dependently induced the production of ROS in human PMN in the range between 10 ng/mL and 1,000 ng/mL, whereas C3a-desArg was inactive. Flow cytometric measurement of H2O2 by dihydrorhodamine-123 labeling of anti-CD16-stained PMN showed that predominantly neutrophilic PMN are responsible for the C3a-induced activation of the respiratory burst. To exclude that C3a stimulation was caused by contamination with C5a, the specificity of C3a-induced activation of PMN was shown using monoclonal antibodies (MoAbs). Accordingly, the effect of C3a was completely abolished in the presence of Fab fragments of a blocking anti-C3a MoAb. In addition, blockade of the C5a receptor by the anti-C5a receptor (anti-C5aR) MoAb, S5/1, totally inhibited the C5a-induced production of ROS, whereas the C3a response in the presence of the anti-C5aR MoAb was unaffected. The specificity of the response was further confirmed by homologous desensitization after restimulation with C3a. In contrast, no cross-desensitization was observed upon stimulation with C5a. The C3a-induced ROS production by PMN was inhibited by pertussis toxin, indicating the involvement of guanine nucleotide-binding proteins (Gi proteins) in the signal transduction process initiated by C3a. In addition, stimulation of PMN by C3a resulted in a transient increase in the cytosolic free calcium concentration ([Ca2+]i) in a dose-dependent manner. In contrast to C3a- induced ROS production, C3a did not induce a chemotactic response in PMN, indicating functional qualitative differences as compared with C5a. In summary, these results show that C3a is a potent activator of the respiratory burst in human PMN. Therefore, these findings point to a novel role of C3a in the pathogenesis of inflammatory diseases associated with increased C3a levels and PMN activation.
Mild elliptocytosis associated with the alpha 34 Arg-->Trp mutation in spectrin Genova (alpha I/74)
Authors: Perrotta S , Miraglia del Giudice E , Alloisio N , Sciarratta G , Pinto L , Delaunay J , Cutillo S , Iolascon A 
Published: October 2019

Abstract
We report a new mutation responsible for nonhemolytic hereditary elliptocytosis (HE). The proband displayed an impaired spectrin self- association and an increase of the alpha I 74-kD fragment (alpha I/74 abnormality). The responsible mutation occurred in exon 2 of spectrin alpha-gene: alpha 34 Arg-->Trp (CGG-->TGG), defining spectrin Genova. In Trans to allele alpha Genova, the proband disclosed allele alpha LELY, a common low-expression allele of spectrin alpha-gene. It was recognized through particular peptide maps as well as characteristic mutations in exon 40 and intron 45, respectively. The father, who carried allele alpha Genova, but not allele alpha LELY, had a milder presentation. The sensitization of allele alpha Genova by allele alpha LELY was noticeable in the proband as compared with his father. Nevertheless, it was not as sharp as that observed with many other alpha I/74 HE alleles. Therefore, each alpha I/74 HE allele has a distinct intrinsic severity.
Paternal origin of the rearranged major breakpoint cluster region in chronic myeloid leukemia
Authors: Litz CE , Copenhaver CM 
Published: October 2019

Abstract
The Philadelphia chromosome, t(9;22), is present in virtually all cases of chronic myeloid leukemia (CML). It has previously been shown by cytogenetic studies that the rearranged chromosome 22 in patients with CML is exclusively maternal in origin. To address this issue at a molecular level, the major breakpoint cluster region (M-bcr) on chromosome 22 was examined using Southern blot assays and M-bcr Pvu II and Mae II restriction site polymorphisms in three CML patients. In all three cases, the rearranged allele was paternal in origin. These results indicate that the paternally derived M-bcr allele may also be involved in the M-bcr rearrangement.
Phase I study of recombinant interleukin-1 beta in patients undergoing autologous bone marrow transplant for acute myelogenous leukemia
Authors: Nemunaitis J , Appelbaum FR , Lilleby K , Buhles WC , Rosenfeld C , Zeigler ZR , Shadduck RK , Singer JW , Meyer W , Buckner CD 
Published: October 2019

Abstract
A phase I trial was conducted with recombinant human interleukin-1 beta (rhIL-1 beta) in patients undergoing autologous bone marrow (BM) transplantation for acute myelogenous leukemia. rhIL-1 beta was administered at 3 dose levels (0.01, 0.02, 0.05 microgram/kg) by 30 minute intravenous infusion once a day beginning on the day of BM infusion and continuing for a total of 5 doses. A total of 17 patients were entered on the trial, and their results were compared with those of 74 consecutive historical control patients that did not receive colony stimulating factors. Moderate toxicity was observed in all patients. All 17 patients developed fever and chills within 30 minutes after initiation of rhIL-1 beta, and hypotension was observed in 14 of 17 patients 5 to 8 hours after the infusion. A total of 30% of patients required therapy (normal saline or dopamine) for treatment of hypotension. Therefore, dose escalation was discontinued at the 0.05 microgram/kg dose level. The number of days required to achieve an absolute neutrophil count greater than 500 mL in patients who received rhIL-1 beta was less than in historical patients (25 v 34; P = .02). This appeared to correlate with a reduced incidence of infection between days 0 and 28 after BM infusion (12% v 23%; P = .049). Median bilirubin, median creatinine, platelet recovery, and days in the hospital were not different between study patients and historical controls. Survival of patients who received rhIL-1 beta compared with that of historical patients was improved (30% v 20%; P = .04). These possible benefits were achieved at the cost of moderate toxicity during rhIL-1 beta administration.
Influence of mutations and size of multimers in type II von Willebrand disease upon the function of von Willebrand factor
Authors: Christophe O , Ribba AS , Baruch D , Obert B , Rouault C , Niinomi K , Pietu G , Meyer D , Girma JP 
Published: October 2019

Abstract
We compared the properties of plasma von Willebrand factor (vWF) from normal individuals and from two patients with type IIA (Glu875Lys) and type IIB (duplication of Met 540) von Willebrand disease (vWD) with the corresponding fully multimerized recombinant proteins. We included cryosupernatant from normal human plasma and type IIA plasma (Cys509Arg). Functions of vWF were analyzed by binding assays to platelets in the presence of ristocetin or botrocetin. Parameters of binding (number of binding sites per vWF subunit, and dissociation constant Kd) were quantitatively estimated from the binding isotherms of 125I-botrocetin or glycocalicin to vWF, independently of the size of the multimers. We found that ristocetin- or botrocetin-induced binding to platelets was correlated in all cases with the size of vWF multimers. In the absence of inducer, only type IIB rvWF Met-Met540 spontaneously bound to platelets. No significant difference of binding of purified botrocetin to vWF was found between normal and patients' plasma, or between wild-type rvWF (rvWF-WT) and rvWF-Lys875. In contrast, affinity of botrocetin for type IIB rvWF Met-Met540 was decreased. Botrocetin-induced binding of glycocalicin to vWF from all plasma and cryosupernatant was similar. Compared with rvWF-WT, binding of glycocalicin to rvWF-Lys875 was normal. In contrast, the affinity for type IIB rvWF Met-Met540 was 10-fold greater. Thus, our data suggest that, in the patients tested, the abnormal IIA phenotype results from the lack of large-sized multimers and is independent of the point mutations. In contrast, the type IIB mutation is directly involved by providing a conformation to the vWF subunits that allows the high molecular weight multimers to spontaneously interact with platelet glycoprotein Ib.
Human cytomegalovirus alters interleukin-6 production by endothelial cells
Authors: Almeida GD , Porada CD , St Jeor S , Ascensao JL 
Published: October 2019

Abstract
In an effort to study whether human cytomegalovirus (HCMV) can disrupt the balanced cytokine network that controls human hematopoiesis, we investigated the ability of a laboratory strain HCMV (AD169) to alter the production of interleukin-6 (IL-6) by cultured endothelial cells (HUVECs). ECs are important components of human bone marrow stroma and produce factors that stimulate the proliferation and differentiation of human hematopoietic progenitors. HCMV was able to greatly increase production of both mRNA and protein for IL-6 in unprimed HUVECs. When we discriminated between viral pellet and cleared viral supernatants, the supernatants induced an increase in mRNA at 30 minutes and protein by 2 hours, whereas an increase in IL-6 caused by virus itself did not become evident until 12 hours. The possibility that IL-6 induction was simply caused by the presence in the viral stock of endotoxin, IL-1 alpha, IL-1 beta, tumor necrosis factor alpha, or IL-4, all known inducers of IL-6 in HUVECs, was ruled out by the addition of polymyxin B and appropriate neutralizing antibodies. These findings show that HCMV is capable of directly and indirectly modulating the production by HUVECs of IL-6, one of the cytokines involved in the process of hematopoiesis.