bluuebunny/crossref_metadata_2025 huggingface dataset:
Overexpression of mitogen-activated protein kinase (ERK1) enhances T- cell cytokine gene expression: role of AP1, NF-AT, and NF-KB
Authors: Park JH , Levitt L
Published: October 2019

Transfected Jurkat cells overexpressing extracellular signal-regulated kinase (ERK1), also referred to as mitogen-activated protein (MAP) kinase, were selected by Western blotting assay using anti-ERK1 and antiphosphotyrosine antibodies in combination with a functional MAP kinase assay. We then asked whether enhanced ERK1 expression had any effect on induction of T-cell cytokine genes. The results show that overexpression of ERK1 enhances expression of T-cell interleukin-2 (IL- 2), IL-3, and granulocyte-macrophage colony-stimulating factor mRNA; no change was seen in expression of the alpha-actin gene. DNA-binding activities of the transcription factors AP1, NF-AT, and NF-kB were specifically increased twofold to fourfold in ERK1-overexpressing clones relative to nontransformed or vector-transformed cells, whereas no enhancement of CK1-CK2 protein DNA binding activity was detected after ERK1 overexpression. Additionally, increased NF-AT DNA binding activity was associated with functional enhancement of NF-AT transactivating activity in ERK1-overexpressing cells. These results provide direct evidence for the role of MAP kinase in the regulation of cytokine gene expression and indicate that such regulation is likely mediated through the enhanced DNA binding activity of specific nuclear transcription factors.
Translocation (14;18)-positive cells are present in the circulation of the majority of patients with localized (stage I and II) follicular non- Hodgkin's lymphoma
Authors: Lambrechts AC , Hupkes PE , Dorssers LC , van't Veer MB
Published: October 2019

Stage I and II follicular non-Hodgkin's lymphoma (NHL) is clinically defined as a localized disease. To study the possibility that this disease is in fact disseminated, we used the sensitive polymerase chain reaction (PCR) method using translocation (14;18) as marker. Samples from 21 patients who were clinically diagnosed with stage I or II follicular NHL were analyzed for the presence of t(14;18)-positive cells using PCR. We analyzed (1) the diagnostic lymph node biopsy and (2) the peripheral blood or bone marrow samples from these patients. Translocation (14;18) cells were detected in the diagnostic lymph node biopsies of 12 patients. In 9 of these patients, t(14;18)-positive cells were detected in peripheral blood and/or bone marrow samples at diagnosis and/or after therapy. Thus, in 75% of the follicular NHL patients carrying the t(14;18) as a marker for lymphoma cells, t(14;18)- positive cells were detected in peripheral blood and bone marrow at diagnosis and after therapy. Our results show that t(14;18)-positive cells can be detected in the circulation of patients with stage I and II follicular NHL, indicating that, although diagnosed as localized, the disease is disseminated.
Transfected leukocyte integrin CD11b/CD18 (Mac-1) mediates phorbol ester-activated, homotypic cell:cell adherence in the K562 cell line
Authors: Hickstein DD , Grunvald E , Shumaker G , Baker DM , Back AL , Embree LJ , Yee E , Gollahon KA
Published: October 2019

The CD11b/CD18 leukocyte integrin molecule mediates diverse neutrophil adherence-related functions, including cell:cell and cell:extracellular matrix attachments. To study the individual role of this leukocyte integrin in cell adherence in hematopoietic cells, we expressed the CD11b/CD18 complex on the surface of K562 cells, a cell line derived from an individual with chronic myelogenous leukemia in blast crisis. We used an amphotrophic retroviral vector designated LCD18SN, harboring the complete coding sequence for the CD18 subunit, to transfer the CD18 cDNA into K562 cells and select stable cell lines. The CD11b subunit in the expression plasmid pREP4 was transfected into these K562/CD18 cells by electroporation and stable cell clones were selected. These K562 cells possessed RNA and intracellular protein for each subunit, and they expressed the CD11b/CD18 heterodimer on the cell surface. When CD11b/CD18 expressing K562 cells were stimulated with phorbol myristate acetate (50 ng/mL) for 24 to 48 hours, these K562 cells formed dense cell:cell aggregates. This homotypic aggregation required both activation of the CD11b/CD18 complex and the induction of the counter- receptor for CD11b/CD18 on the conjugate cell. This cell line will (1) enable the structure-function relationships between cell activation and homotypic adherence to be assessed, (2) provide the opportunity to identify accessory molecules required for activation of the CD11b/CD18 complex, and (3) facilitate the identification of novel ligands for the CD11b/CD18 complex.
Enrichment and functional characterization of Sca-1+WGA+, Lin-WGA+, Lin- Sca-1+, and Lin-Sca-1+WGA+ bone marrow cells from mice with an Ly-6a haplotype
Authors: Jurecic R , Van NT , Belmont JW
Published: October 2019

Approximately 4% to 5% of all bone marrow (BM) cells and 8% to 9% of low density BM cells from FVB/N and BALB/c mice (Ly-6a haplotype) show high to intermediate expression of Ly-6E.1 antigen, recognized by the Sca-1 antibody. Functional properties of enriched cells expressing Ly- 6E.1-allelic form of Sca-1 antigen were analyzed and correlated with the properties of cells expressing the carbohydrate binding sites for the lectin wheat-germ agglutinin (WGA). Using equilibrium density centrifugation and fluorescence-activated cell sorting, Sca-1+WGA+, Lin- WGA+, Lin-Sca-1+, and Lin-Sca-1+WGA+ cells were isolated and their splenic colony-forming unit (CFU-S) cell content, radioprotection ability, and long-term reconstitution capacity determined. Enriched Sca- 1+WGA+, Lin-WGA+, Lin-Sca-1+ and Lin-Sca-1+WGA+ cells gave rise to 1 CFU-S12 cell out of 26, 20, 21, and 15 sorted cells, respectively. When transplanted into lethally irradiated recipients (100 to 500 cells/mouse) all populations rescued 70% to 100% of recipients in a 30- day radioprotection assay and mediated survival of 40% to 80% of recipients 6 months after transplantation. Using transgenic mice as cell donors we have shown that 12 to 16 weeks after transplantation of 100 Sca-1+WGA+, Lin-WGA+, Lin-Sca-1+, and Lin-Sca-1+WGA+ cells, 40% to 80% of recipients had donor cells in BM, spleen, thymus, and lymph nodes. These results indicate that the population of cells expressing Ly-6E.1 form of Sca-1 antigen in two analyzed mouse strains with Ly-6a haplotype contains CFU-S and long-term repopulating cells. Furthermore, the data suggest that, at least in FVB/N mice, day-12 CFU-S cells and cells with long-term repopulating capacity simultaneously express Ly- 6E.1 form of Sca-1 antigen and WGA-binding molecules.
Detection of antitrophoblast antibodies in the sera of patients with anticardiolipin antibodies and fetal loss
Authors: McCrae KR , DeMichele AM , Pandhi P , Balsai MJ , Samuels P , Graham C , Lala PK , Cines DB
Published: October 2019

Women with anticardiolipin antibodies (ACLA) are at increased risk for fetal loss. One potential explanation for this outcome is that sera from these individuals contain antibodies reactive with trophoblast cells, which are involved in the establishment of the uteroplacental vasculature and maintenance of placental blood fluidity. To examine this hypothesis, we compared the incidence of trophoblast-reactive antibodies in 27 patients with ACLA and a history of fetal loss with that in 29 normal pregnant women. Sera from 20 patients, but only one control, contained trophoblast-reactive antibodies (P < .001). These antibodies were not directed against major histocompatibility class I antigens, and reacted with both term and first-trimester trophoblast cells. In most cases, sera from which ACLA were adsorbed by cardiolipin- containing liposomes maintained reactivity against cells. In addition, patient Ig fractions immunoprecipitated an approximately 62-kD protein from the trophoblast cell surface, stimulated the release of arachidonic acid and thromboxane A2 by trophoblasts, and inhibited the binding of prourokinase to trophoblast urokinase receptors. These observations show that sera from women with ACLA and a history of fetal loss contain antitrophoblast antibodies. These antibodies may be serologically distinct from ACLA, and may contribute to the pathogenesis of fetal demise.
Consensus AP-1 and CRE motifs upstream from the human cytotoxic serine protease B (CSP-B/CGL-1) gene synergize to activate transcription
Authors: Hanson RD , Grisolano JL , Ley TJ
Published: October 2019

The human CGL-1/cytotoxic serine protease B gene (CSP-B; also known as granzyme B) is transcriptionally activated during cytotoxic T- lymphocyte maturation. Activation can be mimicked in the PEER T-cell leukemia cell line by treatment with 12-O-tetradecanoylphorbol-13- acetate (TPA) and N6–2′-O-dibutyryladenosine 3′,5′-cyclic monophosphate (bt2cAMP). In this report, we show that a consensus AP-1 element and a consensus cAMP response element (CRE) located 5′ to the CSP-B transcriptional start site are both required for transcriptional activation of the CPS-B promoter in TPA + bt2cAMP-stimulated PEER cells. A 94-bp fragment containing both elements activates a heterologous promoter in an orientation-independent fashion. Several single nucleotide substitutions in the AP-1 site abolish activity of the 94-bp fragment. Several point mutations in the consensus CRE substantially reduce promoter activity, but one CRE mutation increases activity fourfold. Replacement of the CRE with a second copy of the AP- 1 site results in a level of transcriptional activity comparable with that of the wild-type sequence, but replacement of the AP-1 site with a CRE abolishes activity. Neither the AP-1 site nor the CRE can be effectively replaced with an SP-1 site. Deletions between the AP-1 site and the CRE retain full activity only if helical spacing is preserved, suggesting that synergism between these two elements is either the result of cooperative binding of factors to the DNA or of cooperative binding of DNA-bound factors to another protein.
Exogenous tat protein activates human endothelial cells [see comments]
Authors: Hofman FM , Wright AD , Dohadwala MM , Wong-Staal F , Walker SM
Published: October 2019

tat protein, a human immunodeficiency virus (HIV) gene product that functions as a transactivator for HIV replication, is known to be secreted extracellularly by infected cells. To determine the potential role of tat in the dissemination of HIV into extravascular tissue, this protein was examined for its ability to activate human endothelial cells. The results show that tat does indeed stimulate endothelial cells. This is evidenced by their expression of the endothelial- leukocyte adhesion molecules, E-selectin, critical for the initial binding of leukocytes to the blood vessel wall, and their increased synthesis of interleukin-6 (IL-6), a cytokine known to enhance endothelial cell permeability. Furthermore, tat acts synergistically with low concentrations of tumor necrosis factor-alpha to enhance IL-6 secretion. These data suggest that extracellular tat protein secreted or released into the microenvironment may contribute significantly to the determination of specific sites of leukocyte binding to blood vessels, to transmigration into tissue, and to eventual dissemination of HIV-infected cells or free virions into tissue.
Hematopoietic progenitor cells from patients with adult T-cell leukemia- lymphoma are not infected with human T-cell leukemia virus type 1
Authors: Nagafuji K , Harada M , Teshima T , Eto T , Takamatsu Y , Okamura T , Murakawa M , Akashi K , Niho Y
Published: October 2019

The in vivo host range of human T-cell leukemia virus type 1 (HTLV-1) has not been definitively established. To determine if hematopoietic stem cells from patients with adult T-cell leukemia-lymphoma (ATL) are infected with HTLV-1, we used a clonogenic progenitor assay followed by the polymerase chain reaction for the detection of HTLV-1 DNA. In vitro growth characteristics of myeloid (CFU-GM) and erythroid (BFU-E) progenitor cells among nonadherent T-cell-depleted bone marrow (BM) mononuclear cells (NA-T-MNCs) from 10 patients with ATL was not significantly different from those of HTLV-1-seronegative controls (P = .20); numbers of colonies per 1 x 10(5) NA-T-MNCs were 34.9 +/- 7.6 for CFU-GM and 39.0 +/- 12.5 for BFU-E in patients with ATL, whereas those were 32.1 +/- 9.5 for CFU-GM and 41.4 +/- 12.7 for BFU-E in normal controls. HTLV-1 DNA was not detected in individual colonies formed by CD34+ cells from any of the patients. Similarly HTLV-1 DNA was not detected in 1 x 10(3) CD34+ cells sorted on a fluorescence-activated cell sorter (FACS) from six patients with ATL studied. In contrast, HTLV-1 DNA was detected in BM mononuclear cells from all patients. These observations clearly indicate that hematopoietic progenitor cells from patients with ATL are normal in their colony-forming capacity and that CD34+ cells from patients with ATL are not infected with HTLV-1 in vivo.
The B7/BB1 antigen is expressed by Reed-Sternberg cells of Hodgkin's disease and contributes to the stimulating capacity of Hodgkin's disease-derived cell lines
Authors: Delabie J , Ceuppens JL , Vandenberghe P , de Boer M , Coorevits L , De Wolf- Peeters C
Published: October 2019

The B7/BB1 molecule has recently been found to be expressed on professional antigen-presenting cells and to be the natural ligand for CD28 and CTLA-4 on T cells. On binding of B7/BB1, CD28 transduces a signal that synergizes with triggering of the T-cell antigen receptor, resulting in enhanced cytokine secretion. In view of the data supporting an antigen-presenting function of Reed-Sternberg cells, we evaluated the expression of B7/BB1 in lymph nodes affected by Hodgkin's disease. B7/BB1 was found to be strongly expressed by the Reed- Sternberg cells in all 47 cases of Hodgkin's disease studied. Moreover, Reed-Sternberg cells were frequently surrounded by CD28-expressing T cells. Evidence for a functional role of B7/BB1 on Reed-Sternberg cells was obtained by our findings that T-cell proliferation and interleukin- 2 (IL-2) production in the primary allogenic mixed lymphocyte reaction (MLR), using the B7/BB1-expressing Hodgkin's disease-derived cell lines L428 and KM-H2 as stimulators, could be partially blocked by adding anti-B7 monoclonal antibody. B7/BB1 expression was also evaluated in a group of non-Hodgkin's lymphomas (n = 46). Whereas B7/BB1 was not expressed by the neoplastic cells of most non-Hodgkin's lymphomas, including T-cell-rich B-cell lymphoma (n = 11), it was present on the neoplastic cells of anaplastic large-cell lymphoma (Ki-1 lymphoma) (n = 5) and follicular lymphoma (n = 4). Our data provide further evidence for an accessory cell function of Reed-Sternberg cells. The accessory cell function of Reed-Sternberg cells might lead to pronounced T-cell activation in vivo, which might contribute to the Hodgkin's syndrome. In addition, our study indicates that B7/BB1 may be a useful marker for differentiating Hodgkin's disease from morphologically similar conditions such as T-cell-rich B-cell lymphoma.
New insights suggesting a possible role of a heat shock protein 70-kD family-related protein in antigen processing/presentation phenomenon in humans
Authors: Manara GC , Sansoni P , Badiali-De Giorgi L , Gallinella G , Ferrari C , Brianti V , Fagnoni FF , Ruegg CL , De Panfilis G , Pasquinelli G
Published: October 2019

A possible role of the peptide binding protein (PBP) 72/74 in antigen processing and presentation has been recently suggested in mice. In order to evaluate a possible analogous role of a PBP72/74-related protein in humans, immunoelectron microscope investigations, functional studies, and immunofluorescence analyses were performed on normal human peripheral antigen-presenting cells. We demonstrated that the determinant recognized by antiheat shock protein (HSP) 72/73 monoclonal antibody (MoAb) is constitutively expressed on the cell surface of monocytes as well as of B cells. Moreover, the capability of monocytes to present a recall antigen to T cells was significantly decreased when preincubated with an anti-HSP72/73 MoAb. These data add further strength to a potential role of a protein related to human PBP72/74 homologue in antigen processing and/or presentation. Finally, the capability of anti-HSP72/73 MoAb to impair the ability of fixed monocytes to present a synthetic peptide demonstrates that cell surface- localized PBP72/74-related protein could play a role in antigen presentation.
Granulocyte colony-stimulating factor receptor mRNA upregulation is an immediate early marker of myeloid differentiation and exhibits dysfunctional regulation in leukemic cells
Authors: Steinman RA , Tweardy DJ
Published: October 2019

The identification of early markers of myeloid differentiation can facilitate an understanding of how differentiation is arrested in leukemogenesis. Using murine bone marrow and the granulocyte-precusor cell line 32Dc13, we show that message for the granulocyte colony- stimulating factor receptor (G-CSFR) is upregulated by G-CSF in an immediate early fashion that is specific to the differentiation pathway and is antagonized by interleukin-3. We further show that G-CSFR message is superinduced by cycloheximide and that these patterns of regulation are altered in leukemic cell lines. In particular, the v-abl oncogene product supresses both ligand-mediated upregulation and superinduction of the G-CSFR gene.
Antithrombin-Gly 424 Arg: a novel point mutation responsible for type 1 antithrombin deficiency and neonatal thrombosis
Authors: Jochmans K , Lissens W , Vervoort R , Peeters S , De Waele M , Liebaers I
Published: October 2019

Inherited type 1 antithrombin (AT) III deficiency is characterized by a decrease of immunoreactive and functional protein levels to about 50%. The disorder is associated with a significantly increased risk of thromboembolism. We have investigated the molecular basis of type 1 AT deficiency in a Belgian family. The diagnosis of the disease was primarily made in a newborn girl with unusually severe thrombotic complications. Using the polymerase chain reaction and single-strand conformation polymorphism analysis, followed by direct sequencing of AT gene fragments, we identified a novel point mutation in exon 6. We detected a G to C substitution in the first position of codon 424 leading to a glycine to arginine substitution. The modification at this highly conserved position in the serine protease inhibitor gene family probably leads to an unstable mutant-gene product. The mutation creates a unique restriction site for the enzyme Hha I in exon 6. This change permitted a rapid and accurate screening of the kindred with identification of the molecular defect in five other family members.
Effect of human recombinant cytokines on the induction of macrophage procoagulant activity
Authors: Schwager I , Jungi TW
Published: October 2019

A panel of human recombinant cytokines was tested for induction of procoagulant activity (PCA) in human monocyte-derived macrophages. Nonadherent culture conditions were used, and PCA was determined with whole cells rather than cell lysates. It was assured by Limulus amebocyte lysate assay that tested cytokines displayed low levels of endotoxin activity within the range of biologic activity. Additional evidence to rule out an endotoxin effect was provided by heat- inactivation experiments. Interferon-gamma (IFN-gamma), interleukin-1 beta (IL-1 beta), and tumor necrosis factor-alpha (TNF-alpha) were strong macrophage PCA inducers. The low level of PCA induced by IL-2, granulocyte-macrophage colony-stimulating factor (GM-CSF), M-CSF, IL-4, IL-6, IL-10, and IFN-alpha could not be distinguished from that induced by traces of endotoxin contaminating the preparations. Transforming growth factor-beta decreased constitutively expressed PCA within 24 hours of exposure. PCA induced by IFN-gamma, IL-1 beta, and TNF-alpha depended largely on tissue factor expression, as evidenced by experiments with factor X-deficient plasma and antitissue factor antibodies. In macrophages subcultured in adherence, IL-1 beta was a strong PCA inducer, whereas IFN-gamma and TNF-alpha promoted little PCA increase. This observation and different kinetics of PCA induction suggested that mechanisms of PCA induction are distinct for the three cytokines. Thus, we showed that well-characterized cytokines critically involved in the promotion of cell-mediated antimicrobial defense/delayed-type hypersensitivity and considered for clinical application promote local fibrin deposition by a direct effect on macrophages.
Synergy between cyclosporin A and a monoclonal antibody to B7 in blocking alloantigen-induced T-cell activation
Authors: Van Gool SW , Ceuppens JL , Walter H , de Boer M
Published: October 2019

Costimulatory signals are absolutely required for T-cell activation after T-cell receptor/major histocompatibility complex-peptide interaction. So far, the best-known candidate essential costimulatory signal is mediated by interaction of CD28 on T cells with B7 on antigen- presenting cells. Using an allogeneic B7+ Epstein-Barr virus- transformed B-cell line as stimulator, we found that addition of a monoclonal antibody (MoAb) to B7 that efficiently blocks B7-CD28 interaction only partially inhibited proliferation and interleukin-2 (IL-2) production in primary and secondary mixed lymphocyte reactions (MLR), whereas the generation of cytotoxic T lymphocytes (CTL) was not affected. Inhibition of primary or secondary MLR-induced T-cell activation with cyclosporin A (CsA) at nontoxic concentrations also was never complete. However, the combination of CsA and anti-B7 MoAb B7–24 synergistically blocked allogeneic B cell-induced T-cell proliferation, IL-2 production, and CTL generation. These data suggest that the mere blockage of B7-CD28 interaction during allotransplantation will be insufficient to prevent rejection or graft-versus-host disease. However, low CsA concentrations, when combined with an agent blocking B7-CD28 interaction, can potentially achieve complete immunosuppression.
Consistent loss of the D5S89 locus mapping telomeric to the interleukin gene cluster and centromeric to EGR-1 in patients with 5q- chromosome
Authors: Nagarajan L , Zavadil J , Claxton D , Lu X , Fairman J , Warrington JA , Wasmuth JJ , Chinault AC , Sever CE , Slovak ML
Published: October 2019

Interstitial deletions of the long arm of chromosome 5 are common in a number of disorders of leukemic and preleukemic myeloid disorders. Although the limits of these deletions vary among patients, a region of cytogenetic overlap that includes band 5q31 is deleted consistently, suggesting loss of 5q31 loci critical for normal myeloid differentiation and leukemogenesis. An anonymous genomic DNA segment D5S89, previously mapped to 5q21–31, detects consistent loss of alleles in cases showing the 5q- chromosome at presentation or relapse. Analysis of a panel of natural-deletion somatic-cell hybrids in conjunction with irradiation hybrids containing fragments of human chromosome 5q shows that the D5S89 locus is telomeric to the interleukin (IL) genes (IL-3, IL-4, IL-5, IL-9, and granulocyte- macrophage colony-stimulating factor [GM-CSF]) and interferon response factor-1 (IRF-1) gene and centromeric to the early response transcription factor (early growth response gene-1 [EGR-1]) on 5q31. To further define the principal region of loss, we have isolated and characterized yeast artificial chromosomes (YACs) spanning D5S89. The presence of several CpG islands within the 300-kb YAC is suggestive of multiple transcription units. However, IL-4, IL-5, IRF-1, IL-3, GM-CSF, and EGR-1 genes were not detected in the YAC clone spanning D5S89, implying that none of these genes are in the vicinity of the D5S89 marker. Further characterization of these YACs should facilitate the isolation of novel candidate genes that may play a role in the evolution of the abnormal phenotype associated with 5q- chromosome.
A bone marrow stromal-derived growth factor, interleukin-11, stimulates recovery of small intestinal mucosal cells after cytoablative therapy
Authors: Du XX , Doerschuk CM , Orazi A , Williams DA
Published: October 2019

The proliferation of epithelial cells lining the small intestinal mucosa may be regulated by microenvironmental signals leading to differentiation of precursor cells in the small intestinal crypts. Proliferation of hematopoietic cells within the hematopoietic microenvironment is known to be regulated by a growing number of glycoprotein growth factors in a hierarchial fashion. We studied the effects of administration of the microenvironment-derived hematopoietic growth factor interleukin-11 (IL-11) on mice given combination radiation/chemotherapy. Treatment of such mice with IL-11 led to significantly increased survival and evidence of rapid recovery of the small intestinal mucosa, which is severely damaged by these cytoxic agents. This recovery was associated with an increase in the mitotic index of crypt cells and an increased frequency of staining of these cells with a monoclonal antibody to proliferating cell nuclear antigen, a member of the cyclin family of nuclear antigens.
Pulmonary fat embolism: a distinct cause of severe acute chest syndrome in sickle cell anemia
Authors: Vichinsky E , Williams R , Das M , Earles AN , Lewis N , Adler A , McQuitty J
Published: October 2019

The etiology of most cases of acute chest syndrome (ACS) in sickle cell disease (SCD) is unknown. Although pulmonary fat embolism (PFE) is frequently found on autopsy, it is rarely considered in the differential diagnosis in pediatric patients. We conducted a study to determine if we could identify PFE in SCD patients with ACS, define the clinical and laboratory course of PFE, and determine if bronchoalveolar lavage is safe and useful in diagnosis of PFE. Twenty-seven SCD patients with ACS were evaluated and compared with 43 control patients. Serial tests (complete blood count, platelet count, nucleated red blood cells [NRBCs], chest x-ray, and oxygen saturations) were compared with steady-state results. Diagnosis of PFE was made by quantitative evaluation of pulmonary macrophages for intracellular fat. No serious complications from bronchoscopy were observed. In the SCD patients with ACS, 12 were PFE+ and 15 were PFE-. The clinical course of the two groups was quite different. All PFE+ patients experienced bone pain and 11 of 12 had chest pain. In contrast, only 6 of 15 of PFE- patients had bone or chest pain. Neurologic symptoms developed in 6 of 12 of the PFE+ group and in none of the PFE- group. Mean hospital days for PFE+ was 13 compared with 7 for PFE-. Laboratory studies in PFE+ showed a significant decrease in hemoglobin (-2 g, P < .05), platelet count (- 293,000, P < .001), and an increase in NRBCs/100 white blood cells (+8.3, P < .001) compared with PFE-. These results indicate that when PFE is associated with ACS, it is characterized by a distinct clinical course, and that bronchial lavage is a safe and useful test in diagnosing PFE in patients with ACS.
Amifostine (WR-2721) shortens the engraftment period of 4- hydroperoxycyclophosphamide-purged bone marrow in breast cancer patients receiving high-dose chemotherapy with autologous bone marrow support
Authors: Shpall EJ , Stemmer SM , Hami L , Franklin WA , Shaw L , Bonner HS , Bearman SI , Peters WP , Bast RC Jr , McCulloch W
Published: October 2019

4-Hydroperoxycyclophosphamide (4-HC), a commonly used marrow-purging agent, is active against many tumors, but is also toxic to normal marrow progenitors. Amifostine (WR-2721) is a sulfhydryl compound with chemoprotectant activity. Preclinical studies using suspensions of bone marrow and breast cancer cells demonstrated that ex vivo treatment with amifostine followed by 4-HC resulted in protection of marrow progenitors, with no compromise in the antitumor effect of 4-HC. This fact stimulated the development of a clinical trial. Bone marrow was harvested from 15 poor-prognosis breast cancer patients and randomly assigned to ex vivo treatment with amifostine followed by 4-HC (amifostine + 4-HC), or treatment with 4-HC alone. High-dose chemotherapy was then administered followed by infusion of the purged autologous bone marrow support (ABMS). Leukocyte engraftment, defined as a white blood cell count > or = 1 x 10(9)/L, was achieved in an average of 26 days for patients whose marrow was purged with amifostine + 4-HC versus 36 days for patients whose marrow was purged with 4-HC alone (P = .032). The average number of platelet transfusions (12 v 29; P = .017) and days of antibiotic therapy (28 v 40; P = .012) were significantly less for patients whose marrow was exposed to amifostine + 4-HC, compared with 4-HC alone. Unpurged backup marrow fractions were infused into three patients whose marrow was purged with 4-HC alone, because of inadequate marrow recovery. None of the patients who received amifostine + 4-HC-purged marrow required a backup marrow fraction. Complete remissions were achieved in 83% of patients with measurable disease, with no difference between the two cohorts. Forty- three percent of patients remained alive and progression-free at a mean of 13 months posttransplant. There was no significant difference in the rate or pattern of relapse for patients whose marrow was purged with amifostine + 4-HC compared with those whose marrow was purged with 4-HC alone. Ex vivo treatment of marrow with amifostine significantly shortens the time to marrow recovery, thereby reducing the risk of myelosuppressive complications in breast cancer patients receiving high- dose chemotherapy and 4-HC-purged ABMS. Since supportive care requirements are also significantly decreased, amifostine may reduce the cost of such therapy.
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

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.
Fractionated versus single-dose total body irradiation at low and high dose rates to condition canine littermates for DLA-identical marrow grafts
Authors: Storb R , Raff RF , Appelbaum FR , Deeg HJ , Graham TC , Schuening FG , Sale G , Bryant E , Seidel K
Published: October 2019

We explored in dogs the immunosuppressive properties of 450 cGy total body irradiation (TBI) delivered from two opposing 60Co sources, as assessed by the criterion of successful engraftment of allogeneic genotypically DLA-identical littermate marrow. Two questions were asked in this study. Firstly, does dose rate affect the immunosuppressive effect of TBI when administered in a single dose? Secondly, does fractionation alter the immunosuppression of TBI when delivered at a very fast dose rate? Dose rates studied included 7 and 70 cGy/min, and fractionation involved four fractions of 112.5 cGy each, with 6-hour minimum interfraction intervals. Six of 7 dogs receiving 450 cGy single- dose TBI at 70 cGy/min showed sustained engraftment of the allogeneic marrow, compared with 1 of 7 dogs receiving single-dose TBI at 7 cGy/min (P = .01). Fractionated TBI at 70 cGy/min resulted in sustained allogeneic engraftment in 3 of 10 dogs, a result that was statistically significantly worse than that with single-dose TBI at 70 cGy/min (P = .03) and not statistically different (P = .24) from that with fractionated TBI delivered at 7 cGy/min (0 of 5 dogs engrafted). A single dose of 450 cGy of TBI delivered at a rate of 70 cGy/min is significantly more immunosuppressive than the same total dose delivered at 7 cGy/min. Fractionated TBI at 70 cGy/min is significantly less immunosuppressive than single-dose TBI at 70 cGy/min and not significantly different from fractionated TBI administered at 7 cGy/min. Results are consistent with the notion that significant DNA repair in lymphoid cells is possible during interfraction intervals at the relatively high dose rate of 70 cGy/min.
Allogeneic T-cell clones able to selectively destroy Philadelphia chromosome-bearing (Ph1+) human leukemia lines can also recognize Ph1- cells from the same patient
Authors: Oettel KR , Wesly OH , Albertini MR , Hank JA , Iliopolis O , Sosman JA , Voelkerding K , Wu SQ , Clark SS , Sondel PM
Published: October 2019

Immunocompetent cells in bone marrow allografts have been associated with a graft-versus-leukemia (GVL) effect. To further characterize effector mechanisms that may be involved in this GVL phenomenon, we have previously established an in vitro model to identify allogeneic T- cell clones that selectively mediate cytotoxicity against a patient's leukemic cells, but not against nonleukemic lymphocytes from the same patient. We have modified this in vitro model to test whether the Ph1 chromosome and the P210 fusion protein it controls have a detectable role in leukemia-specific recognition by allogeneic T-cell clones. In this report, T-cell lines reactive with allogeneic Ph1 chromosome- bearing (Ph1+) chronic myeloid leukemia (CML) cell lines were derived and selected to be minimally reactive with Ph1 negative (Ph1-) lymphoid lines from the same patient. However, after prolonged culture, these same T-cell lines also mediated significant destruction of the Ph1- target cells from the same patients. These T-cell lines specifically recognized cells from the allogeneic CML patient to which they were sensitized, and were not contaminated by an outgrowth of natural killer cells. Furthermore, subclones could be derived from these T-cell lines, and some of these subclones again showed selective killing of the allogeneic Ph1+ leukemia cell lines, and not of the Ph1- cell line from the same patient. Analyses of T-cell receptor (TCR) genes showed the alloreactive T-cell lines and the Ph1+ selective subclones derived from them to be of the same clonal origin. This suggests that the same T cells reacting with antigens expressed on the nonleukemic Ph1- targets can at times selectively and preferentially kill the allogeneic Ph1+ cells. As the same TCR that recognizes Ph1+ cells also can recognize the Ph1- targets, it appears that the Ph1+ chromosome does not play a detectable role in recognition by these allogeneic T-cell clones. This in vitro observation may provide a model for evaluating the relationship between GVL and graft-versus-host disease effects.
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

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.
Molecular characterization of CD30+ anaplastic large-cell lymphoma: high frequency of c-myc proto-oncogene activation
Authors: Inghirami G , Macri L , Cesarman E , Chadburn A , Zhong J , Knowles DM
Published: October 2019

Anaplastic large-cell lymphoma (ALCL) represents a morphologically distinct type of non-Hodgkin's lymphoma (NHL) characterized phenotypically by the expression of the CD30 antigen, a new member of the nerve growth factor gene family. The lymphoid origin of ALCL has been documented using immunohistochemical and molecular genetic analyses. However, very little is known so far regarding the precise pathogenetic mechanisms involved in its development and progression. Therefore, we investigated bcl-2, p53, and retinoblastoma gene (Rb) expression immunohistochemically; the occurrence of bcl-2, c-myc, and Rb gene rearrangements using Southern blotting; and the presence of ras and p53 gene somatic mutations by single-strand conformation polymorphism assay in a panel of 18 well-characterized ALCLs. In addition, the presence of Epstein-Barr (EBV) and human T-cell lymphotropic virus type I (HTLV-I) genomes were investigated using polymerase chain reaction. We identified abnormal c-myc gene products in 6 of 18 cases (33%) of ALCL. On the other hand, the bcl-2 and Rb genes were not rearranged and K-, N-, and H-ras gene somatic mutations were not found. Significant levels of p53 protein expression were found in more than 60% of ALCLs, but only a single ALCL carried a p53 gene mutation (exon 5). Only 3 ALCL cases, all occurring in human immunodeficiency virus-infected patients, were positive for EBV genomes. On the other hand, contrary to previous findings, no HTLV-I products could be identified. Despite the fact that the c-myc proto- oncogene appears to be frequently altered in ALCL, no pathognomonic abnormality could be identified and therefore additional studies and new strategies should be designed to identify the pathogenetic mechanisms involved in the development of ALCL.
Piecemeal degranulation of mast cells in the inflammatory eyelid lesions of interleukin-4 transgenic mice. Evidence of mast cell histamine release in vivo by diamine oxidase-gold enzyme-affinity ultrastructural cytochemistry
Authors: Dvorak AM , Tepper RI , Weller PF , Morgan ES , Estrella P , Monahan-Earley RA , Galli SJ
Published: October 2019

We used light and electron microscopy to analyze the eyelid inflammation that develops in transgenic mice that overexpress interleukin-4 (IL-4; Tepper et al, Cell 62:457, 1990). Analysis of alkaline Giemsa-stained plastic sections examined by light microscopy (Dvorak et al, J Exp Med 132:558, 1970), as well as by routine transmission electron microscopy, indicated that the mast cells in the inflammatory eyelid lesions were undergoing piecemeal degranulation, a form of secretion in which the cells' cytoplasmic granules exhibit characteristic morphologic changes that are thought to be associated with the prolonged, vesicle-mediated release of the granules' constituents. Moreover, by using a newly reported enzyme affinity-gold method, which stains histamine based on binding to diamine oxidase-gold (Dvorak et al, J Histochem Cytochem 41:787, 1993), we show that these activated mast cells had released much of their histamine content. The eyelid lesions also exhibited increased numbers of mast cells; interstitial fibrosis, particularly around cutaneous nerves and blood vessels; activated fibroblasts; focal axonal damage; venules with endothelial cells containing numerous vesiculo-vacuolar organelles; and infiltrates of neutrophils and eosinophils. Our findings illustrate that overexpression of the IL-4 gene in vivo can result in eyelid lesions associated with piecemeal degranulation of mast cells, as well as tissue fibrosis and a variety of other pathologic changes. These results also represent the first direct morphologic evidence for histamine secretion by mast cells in vivo.
Flow cytometry of blood and bone marrow cells from patients with hairy cell leukemia: phenotype of hairy cells and lymphocyte subsets after treatment with 2-chlorodeoxyadenosine
Authors: Juliusson G , Lenkei R , Liliemark J
Published: October 2019

By flow cytometry and an extensive set of markers, we characterized leukemic cells from the blood and bone marrow of 68 symptomatic patients with hairy cell leukemia (HCL). Hairy cells identified in the large cell gate always expressed CD19, CD20, HLA-DR, CD45RA, and B-ly 7. Other markers were occasionally expressed, such as CD38, CD45RO, CD23, CD15, CD4, CD5, and CD10 (expressed on more than 20% of the hairy cells in 44%, 25%, 21%, 18%, 12%, 10%, and 5% of evaluated cases, respectively). During treatment with 2-chlorodeoxyadenosine (CdA), the median lymphocyte counts decreased from 2,000/microL to 300/microL. Flow cytometry was repeated at the nadir (n = 24) of lymphocyte counts, at 3 months (n = 46), at 6 months (n = 50), at 1 year (n = 39), and at 2 years (n = 12) after treatment. The initial decrease of CD8+ and CD20+ cells was greater than that of CD4+ and natural killer (NK) cells, leading to an increasing CD4/CD8 ratio. Median nadir values of CD4+, CD8+, CD20+, and NK cells were 128/microL, 78/microL, 10/microL, and 13/microL, respectively. The subsequent recovery was quicker for CD8+ and NK cells, leading to a normalization within 3 months, whereas CD20+ and CD4+ cells required 1 or 2 years to enter the normal range. The CD4/CD8 ratio thus decreased after the nadir and remained less than 1. CD45RA+ CD4 cells and CD45RA+/CD45RO+ double-positive cells were less affected by CdA. Activated T cells, ie, HLA-DR+ cells, rarely decreased below the normal range and often recovered with an overshoot. CD10+ cells increased in the bone marrow posttreatment as an indication of normal B-cell regeneration in 16 of 36 (44%) patients. The quick regeneration of certain lymphoid subsets might explain the lack of late infections in CdA-treated HCL patients.
RANTES- and interleukin-8-induced responses in normal human eosinophils: effects of priming with interleukin-5
Authors: Schweizer RC , Welmers BA , Raaijmakers JA , Zanen P , Lammers JW , Koenderman L
Published: October 2019

We report that responses of normal human eosinophils toward the chemokines RANTES and interleukin-8 (IL-8) are modulated and upregulated by priming with IL-5. In a modified Boyden chamber assay, we studied migratory responses toward the members of the chemokine family RANTES, monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1 alpha (MIP-1 alpha) (C-C subfamily), and IL-8, platelet factor-4 (PF-4), and neutrophil-activating peptide-2 (NAP-2) (C-x-C subfamily). These chemokines were also studied in terms of actin polymerization and ([Ca2+]i)-mobilizing properties, intracellular signals that are thought to play a role during migratory responses. We found that eosinophils showed significant migratory responses toward RANTES and IL-8 at concentrations of 10(-9) to 10(-7) mol/L only after priming with IL-5 (10 pmol/L). At these concentrations, PF-4, NAP-2, MCP-1, and MIP-1 alpha induced no significant migratory responses after priming. Unprimed eosinophils only showed a significant migratory response toward RANTES (10(-6) mol/L). Changes in [Ca2+]i were found after addition of RANTES, MIP-1 alpha, and NAP-2 (10 nmol/L) to unprimed eosinophils. RANTES (10(-9) to 10(-7) mol/L) significantly induced actin polymerization both in primed and unprimed eosinophils, whereas IL-8 (10(-9) to 10(-8) mol/L) and MIP-1 alpha (10(-8) mol/L) only induced actin polymerization after priming with IL-5. NAP-2, PF-4, and MCP-1 did not affect actin polymerization. These findings are further evidence for the hypothesis that cytokines like IL-5 and locally secreted chemokines like RANTES and IL-8 are both at the basis of specific eosinophil influx into the allergic inflammatory locus.
Detection of residual lymphoma cells by polymerase chain reaction in peripheral blood is significantly less predictive for relapse than detection in bone marrow
Authors: Gribben JG , Neuberg D , Barber M , Moore J , Pesek KW , Freedman AS , Nadler LM
Published: October 2019

Polymerase chain reaction (PCR) amplification of the t(14;18) has been shown to be a highly sensitive method to detect minimal residual disease in patients with non-Hodgkin's lymphoma (NHL) whose tumors bear this translocation. The ideal tissue source to detect residual lymphoma would be from a previously involved lymph node. However, lymphoid tissue is rarely available once patients achieve complete remission. Although PCR amplification has been used to detect residual lymphoma cells in both bone marrow (BM) and peripheral blood (PB) of patients in complete remission, it is presently unknown whether BM and PB are equivalent tissue sources to detect residual disease. In the present study, we compared the clinical utility of the detection of residual lymphoma in both the BM and the PB of patients with advanced-stage non- Hodgkin's lymphoma before, at the time of, and after high-dose therapy and autologous BM transplantation (ABMT). The detection of residual lymphoma in either the BM or PB was associated with decreased disease- free survival. However, in the present study, 44% of patients who relapsed had no evidence of circulating lymphoma cells in their PB. At the time of BM harvest, PCR-detectable residual lymphoma cells were detected in 211 of 212 patients; although, in a subset of these patients analyzed, lymphoma cells were detected in the peripheral blood of only 49% of patients. When residual lymphoma cells within the autologous BM are infused into the patient these cells are rapidly detectable circulating in the PB in the patient. These cells continue to circulate during the immediate posttransplant period and be detectable in the PB in the majority of patients who are infused with marrow containing residual lymphoma. We conclude that BM is a more informative tissue source than PB in detecting minimal residual disease at the time of and after ABMT, and that contamination of PB early after ABMT appears to be the consequence of reinfusion of lymphoma cells within autologous marrow.