bluuebunny/crossref_metadata_2025_split huggingface dataset:
Granulocyte colony-stimulating factor (CSF), macrophage-CSF, granulocyte-macrophage CSF, interleukin-3, and interleukin-6 levels in sera from children undergoing blood stem cell autografts
Authors: Kawano Y , Takaue Y , Saito S , Sato J , Shimizu T , Suzue T , Hirao A , Okamoto Y , Abe T , Watanabe T 
Published: October 2019

Abstract
Endogenous production of granulocyte colony-stimulating factor (G-CSF), macrophage CSF (M-CSF), granulocyte-macrophage CSF (GM-CSF), interleukin-3 (IL-3), and interleukin-6 (IL-6) was investigated in 10 children who underwent a total of 12 courses of autologous peripheral blood stem cell transplant (PBSCT) by measuring their serum levels using immunoassay kits. The serum G-CSF level increased immediately following infusion of PBSC graft, peaked between days 3 and 7 posttransplant and then declined by the time the granulocyte count rose. No definitive association was found between the continuous high levels of G-CSF and infective episodes, the number of infused nucleated cells, monocytes, CFU-GM, or the number of days required to achieve greater than 0.5 x 10(9)/L granulocyte, greater than 1.0 x 10(9)/L leukocyte, or greater than 50 x 10(9)/L platelet counts. After PBSCT, IL-6 levels tended to be elevated. No detectable serum level of GM-CSF or IL-3 (< 50 pg/mL) was observed before PBSCT and 4 patients showed a transient increase in the GM-CSF level after PBSCT. No significant change was observed in the post-transplant serum levels of IL-3 or M- CSF. The role of endogenously secreted cytokines in early hematopoietic recovery after PBSCT needs further clarification, but, at present, routine use of exogenous G-CSF therapy is not recommended.
The distribution of erythrocyte phospholipids in hereditary spherocytosis demonstrates a minimal role for erythrocyte spectrin on phospholipid diffusion and asymmetry
Authors: Kuypers FA , Lubin BH , Yee M , Agre P , Devaux PF , Geldwerth D 
Published: October 2019

Abstract
In the human erythrocyte membrane phosphatidylcholine and sphingomyelin reside mainly in the outer leaflet, whereas the aminophospholipids, phosphatidylethanolamine and phosphatidylserine, are mainly found in the inner leaflet. Maintenance of phospholipid asymmetry has been assumed to involve interactions between the aminophospholipids and the membrane skeleton, in particular spectrin. To investigate whether spectrin contributes to maintaining the phospholipid transbilayer distribution and kinetics of redistribution, we studied erythrocytes from hereditary spherocytosis patients whose spectrin levels ranged from 34% to 82% of normal. The phospholipid composition and the accessibility of membrane phospholipids to hydrolysis by phospholipases were in the normal range. Spin-labeled phosphatidylserine and phosphatidylethanolamine analogues that had been introduced into the outer leaflet were rapidly transported at 37 degrees C to the inner leaflet, whereas the redistribution of spin-labeled phosphatidylcholine was slower. The kinetics of transbilayer movement of these spin-labeled phospholipid in all samples was in the normal range and was not affected by the level of spectrin. Although these erythrocyte membranes contained as little as 34% of the normal level of spectrin and were characterized by several physical abnormalities, the composition, distribution, and transbilayer kinetics of the phospholipids were found to be normal. We therefore conclude that spectrin plays, at best, only a minor role in maintaining the distribution of erythrocyte membrane phospholipid.
Functional human transcobalamin II isoproteins are secreted by insect cells using the baculovirus expression system
Authors: Quadros EV , Sai P , Rothenberg SP 
Published: October 2019

Abstract
Transcobalamin II (TCII) is a cobalamin (Cbl, vitamin B12)-binding protein in mammalian plasma that facilitates the cellular uptake of the vitamin. To obtain human TCII in sufficient quantity for analytical studies, the complementary DNA (cDNA) encoding TCII was inserted into the plasmid PVL 1393, and the baculovirus expressing TCII was obtained by homologous recombination in Spodoptera frugiperda (SF9) insect cells by cotransfection with the wildtype virus. Under optimized conditions, SF9 cells infected with the recombinant virus secreted 2 to 4 micrograms of TCII per milliliter of culture medium. TCII did not accumulate in the SF9 cells and seemed to be constitutively secreted as observed previously in cultured human endothelial cells. The purified recombinant TCII has the same molecular weight by SDS-PAGE as purified human TCII. The recombinant TCII cross-reacts with an antiserum to native human TCII, binds Cbl and facilitates the uptake of Cbl in eukaryotic cells by binding to the receptor for TCII-Cbl on the plasma membrane of K562 cells. Amino acid sequence analysis of the purified recombinant TCII identified two polypeptides, one identical to the amino acid sequence deduced from the cDNA and a second lacking the first and second N-terminal residues. These sequences are identical to two TCII polypeptides purified from Cohn fraction III of pooled human plasma. The two forms of recombinant TCII have the same isoelectric points as the two predominant isoprotein forms of TCII in human serum. Since the baculovirus construct contains a single cDNA that can encode only one amino acid sequence, the two isoproteins in recombinant TCII must be generated by a mechanism other than allele specific expression. A plausible mechanism for generating isoproteins of nonglycosylated peptides, such as TCII, may be by splicing of the leader peptide at alternative sites.
Antithrombin III Nagasaki (Ser116-Pro): a heterozygous variant with defective heparin binding associated with thrombosis
Authors: Okajima K , Abe H , Maeda S , Motomura M , Tsujihata M , Nagataki S , Okabe H , Takatsuki K 
Published: October 2019

Abstract
A novel variant of antithrombin III (AT III) that lacks affinity for heparin was found in a 33-year-old man who suffered from recurrent cerebral infarction. The propositus had a history of recurrent ischemic attacks from the age of 28. He was obese and had a smoking habit (30 to 40 cigarettes/day), low high-density lipoprotein cholesterol, and a mild glucose intolerance as the possible risk factors for thrombosis. No family history of thromboembolic episodes was observed. Coagulation studies found low heparin cofactor activity (55%) of AT III, with a normal immunoreactive level (109.7%) and progressive antithrombin activity (117%). Other factors capable of predisposing him to hereditary thrombophilia were within normal ranges. Analysis by crossed immunoelectrophoresis in the presence of heparin and affinity chromatography on heparin-Sepharose demonstrated that the propositus' AT III was composed of two populations, one having no affinity for heparin and the other binding heparin normally. Nucleotide sequencing of 7 exons of the propositus' AT III gene using polymerase chain reaction and subcloning disclosed a transition of thymine to cytosine in exon 3a (codon 116) of the AT III gene leading to a Ser116-Pro conversion. Allele-specific oligonucleotide hybridization procedures confirmed the presence of the mutation in the propositus' genomic DNA. Using the same technique, the mutation was also found in his father's genomic DNA, but not in that of his mother. These findings indicate that Ser116 is an important amino acid residue in heparin binding and that the propositus is heterozygous for the abnormality. Furthermore, the fact that the propositus suffered from recurrent cerebral infarction suggested that being heterozygous for a heparin-binding defect would lead to a predisposition to thrombosis when associated with various risk factors. The name AT III Nagasaki is proposed for this variant AT III containing a novel Ser116-Pro mutation.
A bispecific antibody enhances cytokine-induced killer-mediated cytolysis of autologous acute myeloid leukemia cells
Authors: Kaneko T , Fusauchi Y , Kakui Y , Masuda M , Akahoshi M , Teramura M , Motoji T , Okumura K , Mizoguchi H , Oshimi K 
Published: October 2019

Abstract
An anti-CD3 Fab' x anti-CD13 Fab' bispecific antibody (BsAb) was generated. This BsAb reacted with both CD3+ T cells and CD13+ acute myeloid leukemia (AML) cells. We investigated whether cytokine- stimulated peripheral blood mononuclear cells (PBMC) could lyse patient AML cells after addition of the BsAb. When interleukin-2 (IL-2)- stimulated PBMC were assayed for their cytotoxicity against 51Cr- labeled allogeneic and autologous CD13+ AML cells, their activity was markedly enhanced by the addition of the BsAb. PBMC stimulated with IL- 2 plus anti-CD3 monoclonal antibody (MoAb) showed higher proliferative ability and higher cytotoxicity if this was expressed as lytic units per culture. IL-7-stimulated PBMC also exhibited enhanced cytotoxicity against CD13+ AML cells after addition of the BsAb. Ultrastructurally, CD13+ AML cells incubated with IL-2 plus anti-CD3 MoAb-stimulated PBMC and the BsAb showed apoptotic morphologic changes. A colony assay for AML blast progenitors showed that the colony formation of CD13+ AML cells was inhibited by the addition of autologous IL-2 plus anti-CD3 MoAb-stimulated PBMC, and that this inhibition was further enhanced by the addition of the BsAb. A colony assay for normal bone marrow progenitor cells showed that the addition of autologous IL-2 plus anti- CD3 MoAb-stimulated PBMC and the BsAb inhibited the formation of granulocyte-macrophage colonies and mixed-cell colonies. However, the degree of inhibition was smaller than that for the AML blast colonies. Taken together, these findings suggest that this BsAb may be useful for ex vivo purging of CD13+ AML cells in autologous bone marrow transplantation.
Differential induction of apoptosis in undifferentiated and differentiated HL-60 cells by DNA topoisomerase I and II inhibitors
Authors: Solary E , Bertrand R , Kohn KW , Pommier Y 
Published: October 2019

Abstract
The effects of monocytic/macrophage and granulocytic differentiation induced by phorbol myristate acetate (TPA) and all-trans retinoic acid, respectively, were tested on the induction of apoptosis in human promyelocytic leukemia HL-60 cells treated with topoisomerase I and II inhibitors. Using a filter-binding assay, we observed a strong inhibition of DNA fragmentation induced by 3- and 24-hour continuous exposure to camptothecin, VP-16, VM-26, and m-AMSA in TPA- differentiated cells. The inhibition of the typical internucleosomal DNA fragmentation was confirmed by agarose gel electrophoresis. By contrast, drug-induced DNA fragmentation was not inhibited in retinoic acid-differentiated cells, and apoptosis occurred in these cells after 4 to 5 days in the absence of drug treatment. The TPA inhibitory effect was maximal after 24 hours of treatment and was correlated with differentiation, because phorbol dibutyrate ester was active, whereas 4- alpha-TPA, a nontumor promoter that does not induce differentiation, was not active. Using alkaline elution, we observed that TPA and retinoic acid differentiation were associated with changes in topoisomerase-mediated DNA breaks that were not correlated with their differential effects on drug-induced DNA fragmentation. Moreover, TPA also inhibited DNA fragmentation induced by vinblastine, cycloheximide, calphostin C, and x-rays. Using a cell-free system, we observed that DNA fragmentation was not inhibited in nuclei from TPA-differentiated cells. Rather, inhibition of apoptosis seemed to take place in the cytoplasm. We conclude that phenotypic changes associated with TPA- induced differentiation include inactivation of a cytoplasmic activity that can induce DNA fragmentation associated with apoptosis.
A human beta-globin gene fused to the human beta-globin locus control region is expressed at high levels in erythroid cells of mice engrafted with retrovirus-transduced hematopoietic stem cells
Authors: Plavec I , Papayannopoulou T , Maury C , Meyer F 
Published: October 2019

Abstract
Retroviral-mediated gene transfer of human beta-globin provides a model system for the development of somatic gene therapy for hemoglobinopathies. Previous work has shown that mice receiving a transplant of bone marrow cells infected with a retroviral vector containing the human beta-globin gene can express human beta-globin specifically in erythroid cells; however, the level of expression of the transduced globin gene was low (1% to 2% per gene copy as compared with that of the endogenous mouse beta-globin gene). We report here the construction of a recombinant retrovirus vector encoding a human beta- globin gene fused to the 4 major regulatory elements of the human beta- globin locus control region (LCR). The LCR cassette increases the level of expression of the globin gene in murine erythroleukemia cells by 10- fold. To study the level of expression in vivo, mouse bone marrow cells were infected with virus-producing cells and the transduced cells were injected into lethally irradiated recipients. In the majority of provirus-containing mice (up to 75%), expression of human beta-globin in peripheral blood was detected at least 3 to 6 months after transplantation. Twelve animals representative of the level of expression of the transduced gene in blood (0.04% to 3.2% of the endogenous mouse beta-globin RNA) were selected for further analysis. A range of 0.4% to 12% of circulating erythrocytes stained positive for human beta-globin protein. Based on these values, the level of expression of the transduced gene per cell was estimated to be 10% to 39% of the endogenous mouse beta-globin gene. These data demonstrate that fusion of the LCR to the beta-globin gene in a retroviral vector increases the level of beta-globin expression in murine erythroleukemia cells and suggest that high-level expression can be obtained in erythroid cells in vivo after transduction into hematopoietic stem cells.
Synergism of interferon-gamma and stem cell factor on the development of murine hematopoietic progenitors in serum-free culture
Authors: Shiohara M , Koike K , Nakahata T 
Published: October 2019

Abstract
We examined the effects of interferon-gamma (IFN-gamma) on the growth of murine hematopoietic progenitors supported by interleukin-3 (IL-3) or stem cell factor (SCF) in a serum-free culture system. IFN-gamma inhibited IL-3-dependent granulocyte-macrophage colony growth by normal bone marrow cells, but increased the number of pure and mixed megakaryocyte colonies by post-5-fluorouracil bone marrow cells. The addition of IFN-gamma to the culture containing SCF resulted in a synergistic action on the development of primitive hematopoietic progenitors as well as on the development of mature populations. Primitive progenitors responding to SCF + IFN-gamma were suggested to be supported by SCF in the early stage of development and require IFN- gamma for subsequent growth. Replating experiments of blast cell colonies and comparison of total colony growth among SCF + IFN-gamma, SCF + IL-3, and SCF + IFN-gamma + IL-3 suggest that multipotential progenitors supported by SCF + IFN-gamma are a part of those reactive to SCF + IL-3. These findings suggest that IFN-gamma has bifunctional activity on murine hematopoiesis.
Alternative mechanisms with and without steel factor support primitive human hematopoiesis
Authors: Sutherland HJ , Hogge DE , Cook D , Eaves CJ 
Published: October 2019

Abstract
As a first approach to defining the molecular requirements for supporting human hematopoietic stem cell maintenance and differentiation in vitro, we have analyzed and compared the ability of various factors to support the maintenance and initial differentiation of human long-term culture-initiating cells (LTC-ICs), a distinct, rare primitive hematopoietic cell type whose progeny after 5 weeks include cells detectable as colony-forming cells. Normal human marrow cells highly enriched in LTC-ICs (approximately 1% pure) were placed in cultures containing either preestablished, irradiated human marrow adherent feeder layers, or feeders consisting of Steel factor-deficient SI/SI, or normal +/+ murine fibroblasts, or no feeders. In some groups, either Steel factor alone, granulocyte colony-stimulating factor (G- CSF) and interleukin-3 (IL-3), or all three factors combined were also added repeatedly. SI/SI murine fibroblasts were equivalent to +/+ controls and to normal human marrow feeders in supporting both LTC-IC maintenance and clonogenic cell output over a 5-week period. Soluble Steel factor alone could, however, effectively substitute for human marrow feeders to support LTC-IC maintenance, although clonogenic cell output was markedly reduced under these conditions. Conversely, soluble Steel factor with G-CSF and IL-3 or with feeders (or all together) did not further enhance (or depress) LTC-IC maintenance, although under these conditions clonogenic cell output was markedly increased. These findings confirm previous evidence that LTC-IC maintenance and clonogenic cell production are differentially regulated and show for the first time that LTC-IC maintenance can be supported by different nonsynergizing factors that may, but need not, include Steel factor.
Acute infectious mononucleosis stimulates the selective expression/expansion of V beta 6.1-3 and V beta 7 T cells
Authors: Smith TJ , Terada N , Robinson CC , Gelfand EW 
Published: October 2019

Abstract
Acute infectious mononucleosis (AIM) is caused by the Epstein-Barr virus (EBV) and is characterized by a proliferation of atypical lymphocytes, predominantly CD8+ T cells. Various diseases associated with T-cell activation have been shown to stimulate the selective expansion of certain V beta (variable region of the T-cell receptor beta chain) expressing T-cell populations. The purpose of this investigation was to determine if the proliferation of T cells accompanying AIM is associated with selective expression/expansion of distinct populations of V beta T cells. We determined V beta expression in eight patients with clinical and laboratory evidence of AIM, including an atypical lymphocytosis. Gel electrophoresis and quantitative analysis were performed on cDNA amplified by the polymerase chain reaction (PCR) using different V beta region primers. Gel electrophoresis analysis showed prominent V beta 6.1–3 and V beta 7 bands in all eight patients with AIM but not in the controls. Quantitative PCR analysis showed that the V beta 6.1–3 and V beta 7 mean PCR ratios increased, respectively, from 163.0 +/- 22.5 and 142.3 +/- 5.5 in controls to 339.9 +/- 38.8 (P < .03) and 396.1 +/- 45.6 (P < .01) in the eight patients with AIM. Two of the eight patients who had increased V beta 6.1–3 and V beta 7 expression were retested after clinical resolution of AIM and no longer had evidence of increased V beta 6.1–3 and V beta 7 T-cell expression. AIM is associated with a selective increased expression of V beta 6.1–3 and V beta 7 T cells present at the time of initial clinical symptoms and atypical lymphocytosis. This increased expression resolves following recovery from AIM. This V beta-specific selective expression resembles the super- antigen response seen after staphylococcal toxin stimulation and may be caused by EBV triggering of selective expansion of V beta 6.1–3 and V beta 7 T-cell subsets.
Regulation of c-jun gene expression in human T lymphocytes
Authors: Chauhan D , Kharbanda SM , Rubin E , Barut BA , Mohrbacher A , Kufe DW , Anderson KC 
Published: October 2019

Abstract
The present studies have examined the effects of mitogens on induction of early response gene expression in normal peripheral blood T and Jurkat cells. Pokeweed mitogen (PWM) or anti-CD3 significantly increases c-jun messenger RNA (mRNA) levels in T cells. This transient PWM-related increase in c-jun transcripts is maximal after 15 to 30 minutes of exposure of T cells to PWM. PWM induces c-jun gene expression in a concentration-dependent manner. Moreover, PWM similarly induces expression of other genes coding for leucine zipper transcription factors, ie, jun-B and c-fos. Nuclear run on assays demonstrate that PWM treatment is associated with an increased rate of c-jun gene transcription. Transient expression assays with c-jun promoter fragments linked to the chloramphenicol acetyltransferase gene suggest that the PWM-induced increase in transcription is mediated by the AP-1 transcription factor complex. Moreover, treatment of T cells with actinomycin D to block further transcription before their culture with PWM suggests that the increase in c-jun gene expression by PWM is also regulated at least in part by a posttranscriptional mechanism. Cycloheximide does not alter c-jun mRNA induction by PWM. Finally, given that PWM induces B-cell differentiation in an interleukin-6 (IL- 6)-mediated, T-cell-dependent manner, the relationship of c-jun and IL- 6 gene expression in PWM-stimulated T cells was examined. The induction of IL-6 mRNA in T cells stimulated by PWM occurs after maximal induction of c-jun mRNA, at a time when the latter is no longer detectable. These findings suggest that PWM induces c-jun gene expression in T cells by a transcriptional and posttranscriptional mechanism and that AP-1 confers PWM inducibility of this gene. Because the IL-6 promoter has several potential transcriptional control elements, one of which is an AP-1-binding site, future experiments will evaluate the role of c-jun in the regulation of PWM-induced IL-6 synthesis by T cells.
Duplication of small segments within the major breakpoint cluster region in chronic myelogenous leukemia
Authors: Litz CE , McClure JS , Copenhaver CM , Brunning RD 
Published: October 2019

Abstract
The t(9;22) in chronic myelogenous leukemia (CML) may be reciprocal or, in a minority of cases, may result in an extensive deletion of a portion of the major breakpoint cluster region (M-bcr) of the BCR. This report provides evidence of the duplication of small segments within the M-bcr in a small group of patients with CML. Southern blots of Bgl II and Bgl II/BamHI double-digested DNA from the blood or bone marrow of 46 patients with CML were probed with a 5' 1.4-kb Taq I/HindIII M- bcr probe and a 3' 2-kb HindIII/BamHI M-bcr probe. In three patients, rearrangements were noted with both probes in Bgl II-digested DNA, but were not present in Bgl II/BamHI-digested DNA with either probe. Southern analysis of DNA samples double-digested with Bgl II and BspHI from two of these three cases showed no rearrangements with either probe; the M-bcr BspHI site is located 26 bp 3' of the BamHI site in the second intron of the M-bcr. The presence of a rearranged M-bcr with both probes in Bgl II-digested DNA and the lack of rearrangement in Bgl II/BamHI and Bgl II/BspHI double-digested DNA suggest the presence of M- bcr BamHI and BspHI sites on both 9q+ chromosome (9q+) and the Philadelphia chromosome (Ph). This implies a duplication of at least the 26-bp M-bcr BamHI/BspHI fragment in these two samples. Sequence data from one of these two cases confirmed the M-bcr breakpoints to be staggered; the Ph M-bcr breakpoint occurred 258 bp downstream from the 9q+ M-bcr breakpoint. It is concluded that a duplication of small segments within the M-bcr occurs in a small group of patients with CML, which may lead to pseudogermline patterns on Southern blot. Such a duplication may provide insight into the mechanism of some chromosomal translocations in neoplasia.