bluuebunny/crossref_metadata_2025 huggingface dataset:
The expression of the p75 subunit of interleukin 2 receptor in Tac negative leukemic cells of two patients with large granular lymphocytic leukemia
Authors: Tagawa S , Hatakeyama M , Shibano M , Taniguchi T , Kitani T
Published: October 2019

The expression of interleukin 2 receptor (IL-2R) on leukemic cells of natural killer (NK) and T cell lineages in two patients with large granular lymphocytic (LGL) leukemia was examined. The p55 Tac IL-2R was not detected by the indirect immunofluorescence method and it did not participate in the IL-2 binding to the surface of these cells. However, these leukemic cells proliferated in a IL-2 dose-dependent manner and expressed p55. A p75 IL-2 receptor (IL-2-R) subunit was detected on the LGL leukemic cells of both NK and T lineages in a crosslink assay. Thus, it is suggested that the primary signal of IL-2 is mediated by the p75 alone. A study of the inhibitions of the proliferative response of LGL leukemia cells by anti-Tac revealed that both p75 and secondarily induced p55 are required for the cell proliferation.
Phenotypic heterogeneity in aneuploid multiple myeloma indicates pre-B cell involvement
Authors: Epstein J , Barlogie B , Katzmann J , Alexanian R
Published: October 2019

The expression of early and mature B cell markers, surface beta 2- microglobulin (B2M) and cytoplasmic immunoglobulin (clg) by aneuploid tumor cells in bone marrow aspirates from 44 patients with multiple myeloma was evaluated by correlated DNA immunofluorescence flow cytometry. Myeloma tumor cells of almost 90% of the patients contained monoclonal clg and expressed the mature plasma cell antigen R1–3 as well as surface B2M; common acute lymphoblastic leukemia antigen (CALLA) was present in 55%, B2 in 17%, and B4 in 23% of samples studied. Coexpression of CALLA and clg in 46% of all patients identified a novel myeloma phenotype without known counterpart in the normal differentiation of B cells. CALLA and clg were independently expressed and gave rise to CALLA+/clg-, CALLA+/clg+, and CALLA-/clg+ cells. The association of CALLA and mature plasma cell markers may define discrete stages of neoplastic plasma cell differentiation.
Identification of Bakb, a new platelet-specific antigen associated with posttransfusion purpura
Authors: Kickler TS , Herman JH , Furihata K , Kunicki TJ , Aster RH
Published: October 2019

Baka is a platelet alloantigen whose putative allele, Bakb, has not been identified previously. By using a serum, “Har,” obtained from a patient with posttransfusion purpura, we describe the platelet alloantigen Bakb. The Har serum reacted with an NP-40-extractable platelet membrane protein of 142 kd with mobility similar to platelet glycoprotein IIb alpha. We found that the antigen recognized by the Har serum is inherited in an autosomal dominant mode with an apparent gene frequency of .39. Chi-square analysis of observed and expected phenotype frequencies indicated that serum Har recognizes Bakb, the anticipated allele of Baka. Our findings provide new evidence for polymorphism of glycoprotein IIb and for the association of posttransfusion purpura with alloimmunization to determinants on this glycoprotein.
Ti (WT31)-negative, CD3-positive, large granular lymphocyte leukemia with nonspecific cytotoxicity
Authors: Oshimi K , Hoshino S , Takahashi M , Akahoshi M , Saito H , Kobayashi Y , Hirai H , Takaku F , Yahagi N , Oshimi Y
Published: October 2019

A case of WT31-, CD3+ large granular lymphocyte leukemia is reported. On surface marker analysis, the proliferating cells were found to be CD3+4–8–16+ and WT31-. By two-color immunofluorescence staining, CD3+4- 8- cells were found to be WT31-, and a small population of WT31+ cells expressed either CD4 or CD8. WT31-, CD3+ cells were also identified in a bulk culture of lymphocytes expanded in vitro. Because WT31 monoclonal antibody (MoAb) reacts with the nonpolymorphic epitope of the disulfide-linked heterodimer of the T cell antigen receptor (Ti), the absence of the WT31-reactive Ti determinant may represent an expression of different CD3-associated polypeptides. The rearrangement of the Ti-beta and Ti-gamma genes but not the immunoglobulin gene was demonstrated, and the single pattern of rearrangement indicated the monoclonal origin of the lymphocytes. When the lymphocytes were assayed for their cytotoxicity against K562, MOLT-4, Daudi, and Raji tumor cell lines, a broad spectrum of cytotoxicity for these tumor cells was observed, and the lymphocytes also exhibited antibody- and lectin- dependent cellular cytotoxicity and lymphokine-activated killer activity. Treatment with anti-CD2 and anti-CD3 MoAbs inhibited their nonspecific cytotoxicity. The anti-CD3-mediated inhibition of nonspecific cytotoxicity suggested that an as yet unidentified Ti, present in association with the CD3 molecule on these lymphocytes, serves as a specific receptor for target tumor cell recognition.
Mechanism of transient adsorption of fibrinogen from plasma to solid surfaces: role of the contact and fibrinolytic systems
Authors: Brash JL , Scott CF , ten Hove P , Wojciechowski P , Colman RW
Published: October 2019

The transient detection of fibrinogen on surfaces has been described (Vroman effect) and high-mol-wt kininogen (HK) has been shown to play a role in this reaction. In this study, we attempted to identify the form of HK responsible for preventing detection of the fibrinogen initially adsorbed from plasma to various artificial surfaces and to determine if other plasma components were involved. We compared 125I-fibrinogen adsorption in the presence of normal plasma to plasma deficient in specific proteins. On all surfaces tested, we found that fibrinogen was displaced from the surface. The extent of displacement was greatly reduced, however, but not eliminated in HK-deficient plasma. Factor XII- deficient plasma also showed reduced fibrinogen displacement. These data indicate that HK can actually displace fibrinogen; however, factor XII, or a factor XII-mediated reaction also appears to be necessary for this displacement to occur. Furthermore, when normal plasma was first subjected to extensive contact activation by dextran sulfate, during which the HK was extensively degraded to components smaller than the light chain (as assessed by Western blotting), we observed greatly reduced displacement of fibrinogen. Extensive contact activation of Factor XI-deficient plasma failed to show low-mol-wt derivatives, however, and displacement of fibrinogen was similar to normal plasma that had not undergone extensive activation. These data indicate that HKa (active cofactor produced during contact activation by factor XIIa or kallikrein) is primarily responsible for displacing fibrinogen, and that HKi (inactive cofactor generated by factor XIa) cannot displace fibrinogen. The fibrinogen from all plasma samples looked similar by Western blot analysis, suggesting that fibrinogenolysis was not a component of the Vroman effect. In addition, experiments performed with plasma prechromatographed on lysine agarose showed that a lysine- agarose adsorbable protein may be minimally involved in fibrinogen desorption and a synergism may exist between HK and that protein.
Adhesive properties of the carbohydrate-modified von Willebrand factor (CHO-vWF)
Authors: Federici AB , De Romeuf C , De Groot PG , Samor B , Lombardi R , D'Alessio P , Mazurier C , Mannucci PM , Sixma JJ
Published: October 2019

In this cooperative study, we explored the role of the carbohydrate moiety (CHO) of von Willebrand factor (vWF) in supporting platelet adhesion. Because of previous discrepant results, all purification steps and CHO modifications by various enzymes were critically evaluated. Under our conditions, CHO-modified vWF preparations contained less than 5% of the initial sialic acid ([Neu]-ase-vWF) and less than 45% ([Neu-Gal]-ase-vWF) or 21% ([Neu-Gal-eF]-ase-vWF) of the D-galactose. These preparations usually showed increased electrophoretic mobility but no significant loss of high-mol-wt multimers when proteolysis had been prevented. Some degree of proteolysis was noted in some carbohydrate-modified vWFs, but the degree of degradation observed did not correlate with the removal of D- galactose. Platelet adhesion to various matrices increased after removal of the terminal sialic acid ([Neu]-ase-vWF) and approximately 45% of the D-galactose ([Neu-Gal]-ase-vWF), but returned to normal values when greater than 70% of the total carbohydrate had been removed by endoglycosidase F [Neu-Gal-ef]-ase-vWF). These changes in reactivity were also reflected in the spontaneous aggregation in normal platelet- rich plasma (PRP) after CHO removal.
Effects of hematopoietin-1 and interleukin 1 activities on early hematopoietic cells of the bone marrow
Authors: Zsebo KM , Wypych J , Yuschenkoff VN , Lu H , Hunt P , Dukes PP , Langley KE
Published: October 2019

Hematopoietin-1 (H-1) was purified from the human cell line 5637 and two amino acid sequences were observed in the preparation. One sequence was identical to that of interleukin 1 alpha (IL 1 alpha) and the other to that of IL 1 beta. The action of recombinant IL 1 alpha and other hematopoietic growth factors was studied using (a) a high proliferative potential colony-forming cell assay that uses primitive hematopoietic precursors from bone marrow, and (b) a spleen colony-forming unit assay. The results indicate that the IL 1 alpha target cell population is different than the target cell populations of IL 3, granulocyte- macrophage colony-stimulating factor; that IL 1 alpha in combination with mononuclear phagocyte colony-stimulating factor provides a proliferative stimulus; and that IL 1 alpha has at least a survival- enhancing and possibly proliferation-inducing effect on primitive hematopoietic stem cells.
Formation of factor-independent hematopoietic multilineage colonies after Abelson virus infection
Authors: Chung SW , Ruscetti S , Wong PM
Published: October 2019

Yolk-sac-derived hematopoietic cells were infected with a helper-free stock of Abelson virus (A-MuLV). After infection, cells were plated in a clonogenic methylcellulose culture in the absence of exogenous growth factors such as interleukin 3 (IL 3) and erythropoietin (Epo). No colonies were observed in cultures without viral infection, whereas factor-independent colonies were consistently observed with virus- infected cultures. The number of colonies was linearly correlated with the number of cells plated. Erythroid-mix colonies consisting mostly of erythroblasts, macrophages, and mast cells could be observed. Tumorigenic, continuously growing mast cell lines could be generated at high frequency from these erythroid-mix colonies after they were initially passaged in the presence of an irradiated feeder layer for 4 to 8 weeks. Southern blot analysis of the DNA from five of these lines examined were all shown to contain integrated A-MuLV proviral DNA. These data are evidence that A-MuLV can directly infect embryonic multipotent hematopoietic progenitor cells and drive them to differentiate to various progeny cells without exogenous growth factors.
Proliferation of normal and malignant human immature lymphoid cells
Authors: Campana D , Janossy G
Published: October 2019

In this study, the proliferative activity of human B and T cell precursors in central lymphoid organs, acute lymphoblastic leukemia (ALL) cells, and permanent cell lines was investigated with double- and triple-color-labeling methods for the analysis of cell cycle-associated features such as 5-bromo-2′-deoxyuridine (BrdU) incorporation and the expression of a nuclear proliferation-associated antigen, Ki67, together with the phenotypic profile of the cells. In infant and regenerating bone marrow (BM), 41.5% +/- 4.0% of terminal deoxynucleotidyl transferase (TdT+) cells were Ki67+, and 30.0% +/- 4.0% incorporated BrdU. A similar proportion of TdT+ dividing cells was observed in adult BM. The proliferative activity of the B cell progenitors reached the peak at the pre-B stage: 80.8% +/- 7.6% and 35.3% +/- 6.1% of c mu +, RFB7- cells were Ki67+ and BrdU+, respectively. In contrast, greater than 95% of surface immunoglobulin- positive BM lymphocytes were resting cells. In infant thymus the highest dividing capacity (95% Ki67+, 60% to 90% BrdU+) was observed in large cortical thymocytes (TdT+, CD1-, cCD3+), and TdT+, CD1+ cortical thymocytes also showed a high proliferative activity (74.3% +/- 2.3% Ki67+, 22.0% +/- 1.0% BrdU+), but TdT-, mCD3+ thymic lymphocytes were mainly resting cells (less than 5% Ki67+, less than 1% BrdU+). The proliferative activity of null and common ALL blasts was significantly lower than that of normal BM TdT+ cells (15.5% +/- 4.2% Ki67+, 6.2% +/- 2.1% BrdU+; P less than .001). Dividing ALL blasts were TdT+ and expressed surface antigens detected by CD10 and/or CD19 antibodies. In T cell-ALL, the percentages of Ki67+ and BrdU+ blasts were also lower than those found in the corresponding normal immature thymocytes (13.0% +/- 3.1% and 2.4% +/- 1.3%, respectively; P less than .001). Thus, ALLs derive from actively proliferating lymphoid precursors but have a lower dividing capacity than the corresponding normal cell types. In ALL cases with heterogeneous expression of markers such as cmu and CD1, dividing blasts were distributed among both negative and positive populations, thus indicating that blasts with signs of differentiation also remain within the dividing pool.
Introduction of a human colony stimulating factor-1 gene into a mouse macrophage cell line induces CSF-1 independence but not tumorigenicity
Authors: Roussel MF , Rettenmier CW , Sherr CJ
Published: October 2019

The SV40-immortalized mouse macrophage cell line, BAC1.2F5, is strictly dependent on CSF-1 for its survival and proliferation in culture. Introduction of a retroviral vector containing a 1.6 kilobase (kb) pair human CSF-1 cDNA into these cells abrogated their growth factor dependence but did not render the cells tumorigenic in nude mice. The infected macrophages contained multiple copies of the vector provirus, expressed both membrane-bound and secreted forms of CSF-1, and exhibited constitutive down modulation of the murine CSF-1 receptor. Because insertion of the v-fms gene has previously been shown to abrogate factor dependence and induce tumorigenicity in BAC1.2F5 macrophages, the failure of these cells to express a fully transformed phenotype after persistent stimulation by endogenous CSF-1 suggests that the v-fms and c-fms gene products provide different signals for cell proliferation.
Thrombin decreases von Willebrand factor binding to platelet glycoprotein Ib
Authors: George JN , Torres MM
Published: October 2019

Thrombin is a physiological agonist that promotes platelet aggregation and secretion. In this study we observed that thrombin can also inhibit a function of platelets related to primary hemostasis. Platelet stimulation by thrombin decreased the binding of von Willebrand factor (vWF) to glycoprotein (GP) Ib and decreased ristocetin-induced agglutination, in vitro reactions that correlate with initial platelet adhesion to the vessel wall. Binding of the monoclonal antibody API to GP Ib was also decreased. Cytoskeletal participation in the change of GP Ib was suggested because pretreatment of platelets with cytochalasin to prevent actin filament formation prevented the thrombin-induced decreases in vWF binding. API binding, and ristocetin-induced agglutination. Measurement of GP Ib in detergent extracts by electroimmunoassay demonstrated no loss after thrombin stimulation. Electroimmunoassay also demonstrated that the API epitope of GP Ib on intact thrombin-treated platelets was accessible for complete digestion by chymotrypsin. Therefore GP Ib was neither released from the platelet surface nor internalized by thrombin treatment. A previously recognized effect of thrombin is its induction of receptor sites on platelet surface GP IIb-IIIa for contact-promoting proteins, including vWF that are involved in the platelet spreading and aggregation that follow adhesion. Therefore the action on GP Ib may combine with the effect on GP IIb-IIIa to shift platelet reactivity from GP Ib-vWF-mediated initial contact with the vessel wall to GP IIb-IIIa-mediated spreading and aggregation.
S-100 beta positive T cell leukemia
Authors: Takahashi K , Ohtsuki Y , Sonobe H , Hayashi K , Nakamura S , Kotani S , Kubonishi I , Miyoshi I , Isobe T , Kita K
Published: October 2019

We reported a peculiar case with T cell leukemia. The patient was a 34- year-old woman showing extensive splenomegaly and marked leukemic cell proliferation and running a rapid fatal clinical course. The leukemic cells were morphologically ordinary lymphocytes showing suppressor/cytotoxic(s/c) T cell phenotypes and containing S-100b protein. Southern blot analysis revealed rearrangement of the beta chain genes of the T cell receptor (TcR) of the leukemic cells. Because these phenotypic and morphologic features were identical with those of S-100 beta+T lymphocytes (S-100 beta +TL) in normal human peripheral blood, we regarded this case as S-100 beta +T cell leukemia. We discussed clinicopathological features of S-100 beta +T cell leukemia/lymphoma by assessing similar cases reported so far. S-100 beta +T cell leukemia/lymphoma is a new type of s/c T lymphocytic leukemia/lymphoma with aggressive features.
The role of endothelium in factor Xa regulation: the effect of plasma proteinase inhibitors and hirudin
Authors: Friedberg RC , Hagen PO , Pizzo SV
Published: October 2019

The role of endothelium in the inhibition of human factor Xa was studied in a plasma environment. Human factor Xa can bind to and function on bovine aortic endothelium in a manner similar to that of bovine factor Xa. Approximately 70% of the bound factor Xa is subject to inhibition by plasma proteinase inhibitors, and the remaining 30% is irreversibly bound as part of a 125 Kd membrane-associated complex not subject to proteolytic degradation. The proportion reversibly bound and its rate of release do not alter with changes in calcium, citrate, heparin, or active proteinase inhibitor concentrations. The principal plasma proteinase inhibitor of human factor Xa was antithrombin III, which accounted for 60% to 65% of factor Xa released from endothelium, with alpha 1-proteinase inhibitor inactivating 20% to 25% and alpha 2- macroglobulin approximately 15%. All of the reversibly bound factor Xa was identified in complex with one of these three proteinase inhibitors. The thrombin active-site inhibitor hirudin was found to markedly accelerate the displacement of reversibly bound factor Xa from the endothelium and to associate specifically with factor Xa without a loss of activity toward chromogenic substrates, perhaps accounting for a novel mechanism of anticoagulation.
Multiple polymorphic sites in factor X locus
Authors: Hassan HJ , Guerriero R , Chelucci C , Leonardi A , Mattia G , Leone G , Mariani G , Mannucci PM , Peschle C
Published: October 2019

The structure of factor X (FX) gene was analyzed in five FX deficient pedigrees with four different variants of the disease, as well as in 50 normal subjects. Genomic DNA from the deficient patients and the normal controls was digested with 12 restriction endonucleases and hybridized with a FX cDNA probe. The results seemingly exclude gross gene deletions or rearrangements in the deficient patients. A variety of polymorphic sites (ie, EcoRI, HindIII, PstI, PvuII, TaqI) was observed within the FX locus and their relative frequency was established. Intriguingly, a highly polymorphic region for the PvuII endonuclease was identified and located approximately 3 kilobases (kb) from the last 3′ exon. These polymorphisms allowed us to analyze the allelic segregation in a FX deficient family and to identify a homozygous subject.
Increased resistance to plasmic degradation of fibrin with highly crosslinked alpha-polymer chains formed at high factor XIII concentrations
Authors: Francis CW , Marder VJ
Published: October 2019

We have previously demonstrated that increasing factor XIII concentrations above that present in plasma (1 U/mL) results in the formation of very high molecular weight alpha fate polyacrylamide and agarose gel electrophoresis (SDS-PAGE). In this report, we have examined the effect of such crosslinking on plasmic susceptibility of fibrin prepared from purified fibrinogen and from plasma in the presence of factor XIII concentrations between 0 and 10 U/mL. The crosslinking achieved with purified fibrinogen at 1 U/mL factor XIII increased resistance to plasmic degradation by 32% as measured in a radiolabeled clot lysis system. However, further increases in plasmic resistance occurred at factor XIII concentrations of 2 and 10 U/mL, the latter decreasing the lysis rate to 45% of that which occurred in the absence of factor XIII. To achieve the same rate of clot lysis with fibrin formed using 10 U/mL rather than 1 U/mL of factor XIII, an increase in plasmin concentration of up to 4.2-fold was required. Similar results were obtained using clots prepared from plasma in the presence of factor XIII concentrations greater than 1 U/mL. Since the alpha 2-plasmin inhibitor content was the same for fibrin at 1 or 10 U/mL factor XIII, the increasing plasmic resistance could not be attributed to increased binding of the inhibitor. We conclude that fibrin prepared in the presence of factor XIII at concentrations exceeding that in plasma shows increased resistance to plasmic degradation, which is likely explained by the formation of very high molecular weight alpha polymer chains.
High titer anti-HIV antibody reactivity associated with a paraprotein spike in a homosexual male with AIDS related complex
Authors: Ng VL , Hwang KM , Reyes GR , Kaplan LD , Khayam-Bashi H , Hadley WK , McGrath MS
Published: October 2019

We observed a human immunodeficiency virus (HIV)-infected homosexual male with AIDS related complex (ARC) who had a serum globulin level of 80 g/L. Serum protein electrophoresis revealed a gamma globulin fraction of 40 g/L, of which 50% (20 g/L) was contained within a paraprotein spike, comprised predominantly of IgG kappa. This patient also had high titer anti-HIV antibodies in his serum, which were Western blot reactive at a final dilution of 1:500,000, and recognized gp120env, p66pol, p55gag, p53pol, p41gag, and p24gag. Because paraproteins in the past have been shown to be directed against specific antigens, we purified this patient's paraprotein using a modified high performance liquid chromatography (HPLC)-hydroxylapatite procedure and tested the purified paraprotein for anti-HIV antibody activity. The purified paraprotein retained anti-HIV antibody activity to a final dilution of 1:100,000, and recognized p66pol, p55gag, p53pol, p41gag, and p24gag. The recognition of both “gag” and “pol” gene products suggested that the purified paraprotein might not be monoclonal in origin. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that the purified paraprotein contained at least two immunoglobulin light chain species (Mol wt 30 to 33 Kd). Affinity chromatography of the purified paraprotein using a p24- Sepharose 4B matrix separated the “gag” and “pol” antibody activities. Immunoglobulin gene rearrangement analysis of a bone marrow aspirate (which contained 15% plasma cells) failed to reveal a clonal population of immunoglobulin producing cells. We conclude that this patient's paraprotein accounted for most of the anti-HIV activity present in whole serum, and that this paraprotein was not monoclonal in origin.
The use of enzymopathic human red cells in the study of malarial parasite glucose metabolism
Authors: Roth E Jr , Joulin V , Miwa S , Yoshida A , Akatsuka J , Cohen-Solal M , Rosa R
Published: October 2019

The in vitro growth of Plasmodium falciparum malaria parasites was assayed in mutant red cells deficient in either diphosphoglycerate mutase (DPGM) or phosphoglycerate kinase (PGK). In addition, cDNA probes developed for human DNA sequences coding for these enzymes were used to examine the parasite genome by means of restriction endonuclease digestion and Southern blot analysis of parasite DNA. In both types of enzymopathic red cells, parasite growth was normal. In infected DPGM deficient red cells, no DPGM activity could be detected, and in normal red cells, DPGM activity declined slightly in a manner suggestive of parasite catabolism of host protein. However, in infected PGK deficient red cells, there was a 100-fold increase in PGK activity, and in normal red cells, a threefold increase in PGK activity was observed. Parasite PGK could be recovered from isolated parasites, and a marked increase in heat instability of parasite PGK as compared with the host cell enzyme was noted. Neither cDNA probe was found to cross- react with DNA sequences in the parasite genome. It is concluded that the parasite has no requirement for DPGM, and probably has no gene for this enzyme. On the other hand, the parasite does require PGK, (an adenosine triphosphate [ATP] generating enzyme) and synthesizes its own enzyme, which must have been encoded in the parasite genome. The parasite PGK gene most likely lacks sufficient homology to be detected by a human cDNA probe. Enzymopathic red cells are useful tools for elucidating the glycolytic enzymology of parasites and their co- evolution with their human hosts.
Identification of base substitutions in the promoter regions of the A gamma- and G gamma-globin genes in A gamma- (or G gamma-) beta+-HPFH heterozygotes using the DNA-amplification-synthetic oligonucleotide procedure
Authors: Yang KG , Stoming TA , Fei YJ , Liang S , Wong SC , Masala B , Huang RB , Wei ZP , Huisman TH
Published: October 2019

DNA amplification combined with hybridization with 32P-labeled synthetic oligonucleotide probes has been used to identify base substitutions in the 5′ promoter region of the A gamma globin gene in members of eleven families from China, Sardinia, Canada, and the United States who had a heterozygosity for the A gamma-beta+-hereditary persistence of fetal hemoglobin (HPFH), and in members of six black families with a possible G gamma-beta+-HPFH heterozygosity. All three known A gamma types were observed, ie, the British type (-198, T----C), the Chinese type (-196, C----T), and the Green type (-117, G----A); the latter has been found in a black family. Of the six families with G gamma-beta+-HPFH, three had C----G at -202 and none T----C -175. Conditions for hybridization of amplified DNA with the specific probes are provided and the usefulness of the technique is discussed. The increase in numbers of A gamma(G gamma)-beta+-HPFH heterozygotes with specific base substitutions greatly enhances the probability of a direct correlation between these substitutions and the increase in the production of a specific gamma chain.
High numbers of CD4+ T cells showing abnormal recognition of DR antigens in lymphoid organs involved by Hodgkin's disease
Authors: Maggi E , Parronchi P , Macchia D , Bellesi G , Romagnani S
Published: October 2019

Purified T lymphocytes (E rosetting cells) isolated from the involved lymphoid organs (lymph nodes and spleen) of five patients with Hodgkin's disease (HD) were cloned under culture conditions (phytohemagglutinin plus interleukin-2) that allow clonal expansion of most T lymphocytes. A total number of 104 CD4+ T cell clones so obtained were tested for their ability to proliferate in response to autologous mitomycin-treated non-T cells. About half of these clones but none of 234 CD4+ T cell clones derived from normal lymphoid tissues or peripheral blood displayed a proliferative response to autologous stimulators. When clones proliferating in autologous mixed lymphocyte reaction (AMLR) were assessed for their ability to respond in allogeneic MLR (allo-MLR), most of them were found to exhibit consistent proliferation in response to more than one haplotype. Both the AMLR and the allo-MLR by HD clones were inhibited by adding monoclonal antibodies (MoAbs) reactive with monomorphic determinants of major histocompatibility complex (MHC) class II (DR) antigens to the cultures, whereas MoAbs reactive with MHC class I antigens were without effect. These studies suggest that lymphoid organs involved by HD contain high proportions of CD4 T cells showing abnormal recognition of DR antigens. These unusual cells may play an important role in the pathogenetic mechanisms occurring in HD.
Transplantation of human hairy cell leukemia in radiation- preconditioned nude mice: characterization of the model by histological, histochemical, phenotypic, and tumor kinetic studies
Authors: Faguet GB , Agee JF
Published: October 2019

Two cell lines (EH and HK) with hairy cell leukemia (HCL) immunophenotypes were recently derived from two HCL patients. Both cell lines were transplanted subcutaneously (2 x 10(5) or 2 x 10(6)/mouse) in male BALB/c nu/nu mice (n = 128) with a 97% success rate when coimplanted with nonproliferative HT-1080 fibrosarcoma cells (2 x 10(6)/mouse) in recipients preconditioned with total-body irradiation (200 R weekly for 3 weeks). Tumors appeared five to ten days postimplant and reached up to 25% of body weight after a mean survival of 8 weeks (range, 30 to 90 days). Tumor histology suggested large cell lymphoma. Cytochemically and immunophenotypically, tumor cells were indistinguishable from their parent cells. Species and lineage derivation of tumor cells was confirmed by antibody probes against the mouse histocompatibility antigen H-2, human T and B lymphocyte antigens, and the HCL-associated common chronic lymphocytic leukemia antigen (cCLLa). In order of decreasing frequency, metastases occurred in the spleen, lungs, pleura, lymph nodes, bone marrow, and kidneys. Up to 12% of circulating lymphoid cells in mice were cCLLa-positive, which suggested hematogenous tumor dissemination. This HCL xenotransplantation model might be useful in preclinical studies for exploring novel experimental therapies for the management of human HCL.
Subsecond calcium dynamics in ADP- and thrombin-stimulated platelets: a continuous-flow approach using indo-1
Authors: Jones GD , Gear AR
Published: October 2019

The regulation and kinetics (less than 5 seconds) of cytosolic calcium changes ([Ca2+]i) in stimulated blood platelets have been investigated under physiological blood flow conditions. Using a newly-developed continuous-flow approach with indo-1-loaded human platelets, adenosine diphosphate (ADP, 10 mumol/L) and thrombin (5 U/mL) were equally effective in significantly increasing [Ca2+]i by 0.5 seconds. ADP induced a transient [Ca2+]i peak of 1 to 2 mumol/L near 2 seconds, whereas thrombin caused a sustained and larger response. The first phase (less than 2 seconds) was not influenced by a lack of extracellular Ca2+, in contrast to the subsequent [Ca2+]i increase that only reached about 0.7 mumol/L for either ADP or thrombin. The shear rates used in our continuous-flow apparatus were physiological (less than 1,258 sec-1) and only slightly increased the basal [Ca2+]i of 0.1 mumol/L. Platelet aggregation (less than 5 seconds), assessed by single- particle counting, was not altered in platelets loaded with indo-1/AM (2.5 mumol/L).
Defective helper function of purified T4 cells and excessive suppressor activity of purified T8 cells in patients with B-cell chronic lymphocytic leukemia. T4 suppressor effector cells are present in certain patients
Authors: Kunicka JE , Platsoucas CD
Published: October 2019

We investigated helper and suppressor functions to B-cell responses and T-T cell interactions of purified T4 and T8 cells from 20 untreated patients with B-cell chronic lymphocytic leukemia (CLL). Appropriate mixtures of purified T4 or T8 cells from patients with CLL were cultured with purified B cells or T4 and B cells from normal donors for 7 days with pokeweed mitogen (PWM). IgM, IgA, and IgG produced were determined in the supernatants of these cultures by a heavy chain- specific enzyme-linked immunosorbent assay (ELISA) and compared to those obtained by the corresponding mixtures of T4, T8, and B cells from normal donors. Purified T4 cells from 14 of 20 patients with CLL exhibited defective helper function (P less than .001) to immunoglobulin (Ig) production by purified B cells from normal donors. Purified T4 cells from 6 of these 14 patients were able to suppress significantly (P less than .001) and in a concentration-dependent manner Ig production by mixtures of T4 and B cells from normal donors, in the absence of T8 cells. These suppressor effector T4 cells from certain patients were partially radiosensitive. Purified T8 cells from 8 of 20 patients with CLL exhibited excessive suppressor activity. These cells significantly suppressed (P less than .001), Ig production by mixtures of T4 and B cells from normal donors to a degree significantly higher (P less than .005) than that observed by equal numbers of T8 cells from normal donors. This inhibition was dependent on the numbers of the T8 CLL cells added to the cultures. Excessive suppressor activity by T8 CLL cells was at least in part radiosensitive in four of eight patients. These results demonstrate a wide range of immunoregulatory T-cell abnormalities in patients with CLL. Naturally occurring T4 suppressor effector cells, directly inhibiting Ig production by mixtures of T4 and B cells, in the absence of T8 cells, are present in certain patients with CLL.
Karyotype evolution and multilineage involvement of Philadelphia chromosome-positive clones in blastic transformation of two patients with chronic myelocytic leukemia
Authors: Sato Y , Kitano K , Tsunoda S , Yoshida M , Kajii E , Suda T , Sakamoto S , Motoyoshi K , Saito M , Miura Y
Published: October 2019

Simultaneous analysis of the cell morphology and karyotypes on single colonies was carried out in two patients with Philadelphia chromosome (Ph1)-positive chronic myelocytic leukemia in blastic transformation in order to clarify the origin of leukemic cells involved. Patient no. 1 was in a typical myeloblastic transformation and patient no. 2 in “basophilic transformation.” Both patients exhibited karyotype evolution in blastic phase (BP), so that we could differentiate BP clones with additional chromosomal abnormalities from chronic phase (CP) clones with only Ph1 among single colonies. The number of single colonies yielding two or more analyzable metaphases was 18 in patient no. 1, and 19 in patient no. 2. Among these colonies, only three in patient no. 1 and none in patient no. 2 were from CP clones and 15 in patient no. 1 and 19 in patient no. 2 were from BP clones. Morphological examination revealed that not only blasts but also mature neutrophils, eosinophils, basophils, macrophages, and erythroblasts were derived from BP clones. These results suggested that (1) BP clones developed at the pluripotent stem cell level, (2) additional chromosomal abnormalities were not restricted to occur in a specific cell line representative in BP; and (3) BP clones, if not all, may retain capacity for maturation and differentiation.
Cytogenetic features of Hodgkin's disease suggest possible origin from a lymphocyte
Authors: Cabanillas F , Pathak S , Trujillo J , Grant G , Cork A , Hagemeister FB , Velasquez WS , McLaughlin P , Redman J , Katz R
Published: October 2019

Surface marker and gene rearrangement data have supported various hypotheses about the origin of the malignant cell in Hodgkin's disease. Cytogenetic data about this disorder, however, are very scanty. To determine if any chromosomal abnormalities that could add further information to this controversial point are present, we studied tumor samples from 49 patients. Abnormal metaphases were obtained in 18 cases. The most common breakpoints were in 11q23, 14q32, 6q11–21, and 8q22–24. These are common breakpoints in lymphoma and raise the possibility that the malignant cell in Hodgkin's disease may be derived from a lymphocyte. The 11q23 breakpoint is also seen in t(4;11) and t(9;11), which is typical of a type of childhood B-cell acute lymphoblastic leukemia characterized by the presence of aberrant myeloid and monocytic markers. Myeloid and monocytic markers are common in Reed-Sternberg cells.
Sizing of the human T cell receptor alpha locus and detection of a large deletion in the Molt-4 cell line
Authors: Roth MS , Collins FS , Ginsburg D
Published: October 2019

The human T cell receptor alpha (TCR-alpha) chain gene consists of discontinuous DNA segments encoding multiple variable (V), multiple joining (J), and one constant (C) region. Unlike other immunoglobulin or TCR genes, however, the TCR-alpha locus exhibits an unusual dispersal of J segments that occupy a region of greater than 50 kilobases (kb) upstream of the C region, with the exact size still unknown. We report here the study of the TCR-alpha genetic locus by using pulsed-field gel electrophoresis (PFGE), which permits the separation of large DNA fragments. Analysis of DNA prepared from normal peripheral blood mononuclear cells, human endothelial cells, and a B cell line demonstrates that both V and C sequences are contained within a single 400-kb SfiI restriction fragment. PFGE analysis of the T cell line Molt 4 suggests a greater than 600-kb deletion involving the TCR- alpha gene.
Significance of extra 18q- chromosome in Japanese t(14;18)-positive lymphoma
Authors: Fukuhara S , Ohno H , Amakawa R , Edamura S , Tomono N , Nasu K , Doi S , Yamabe H , Abe M , Wakasa H
Published: October 2019

Karyotype evolution of t(14;18)-positive lymphoma was studied in 13 Japanese patients. The extra 18q- chromosome, found in six of ten patients with complex karyotypes, was the most common change subsequent to a t(14;18)(q32;q21) chromosome translocation. The additional change was interpreted as being a duplication of an 18q- derived from a t(14;18). The six patients had transformed histology of follicular small cleaved cell lymphoma or diffuse large cell lymphoma, and five of them had extranodal expansion associated with a poor prognosis. These findings indicate that the extra 18q-, together with other chromosome abnormalities, is closely associated with the advanced grade disease of t(14;18)-positive lymphoma, and the extra chromosome is evolutionally comparable with the second Philadelphia (Ph1) chromosome often found in the blastic phase of chronic myelocytic leukemia carrying a t(9;22)(q34;q11). In addition, since the extra 18q- is rarely found in American patients with t(14;18)-positive lymphoma, there appears to be a difference in the karyotype evolution between Japanese and American patients.
The pathophysiology and clinical relevance of platelet heterogeneity
Authors: Thompson CB , Jakubowski JA
Published: October 2019

Recent studies on platelet heterogeneity support the hypothesis that platelet production is regulated to maintain a constant functional platelet mass. In concept this form of regulation is analogous to the manner by which RBC production is controlled to maintain the oxygen- carrying capacity of blood. The platelet mass appears to correlate more closely with platelet function than the platelet count alone, since several factors in addition to the platelet count have been shown to influence the platelets' hemostatic function. These factors include platelet size, density, age, and previous hemostatic interactions. Application of these concepts to clinical problems has provided important insights into platelet physiology and reactivity. Failure to account for differences in platelet heterogeneity among individuals may introduce significant errors in the interpretation of data from laboratory and clinical investigations. However, despite advances, a number of practical issues remain to be resolved before measurements of platelet heterogeneity become accepted as routine clinical tests and are used in the diagnosis of pathologic states.
Decreased plasma gelsolin levels in patients with Plasmodium falciparum malaria: a consequence of hemolysis?
Authors: Smith DB , Janmey PA , Sherwood JA , Howard RJ , Lind SE
Published: October 2019

Mammalian plasma contains a high-affinity actin-binding protein, plasma gelsolin, that severs actin filaments. Destruction of erythrocytes could result in the release of erythrocyte cytoskeletal actin into the plasma where it could bind to gelsolin. If the clearance of actin- gelsolin complexes exceeds its synthesis, lowering of the plasma gelsolin concentration might follow. To test this hypothesis, we measured plasma gelsolin levels in patients with falciparum malaria, a disease where at least part of the hemolysis takes place in the intravascular space and that is usually not accompanied by dysfunction of other organs. Two functional gelsolin assays showed that the mean plasma gelsolin concentration of 18 Nigerian children with Plasmodium falciparum malaria was less than 50% (P less than .001) of healthy Nigerian control subjects tested at the same time. Patients with pneumonia and febrile seizures also had depressed gelsolin levels, which indicates that factors other than hemolysis can lower gelsolin concentrations. Gelsolin levels were measured in 11 patients from The Gambia with P falciparum malaria before and approximately 3 weeks after treatment. In all cases the gelsolin level increased after treatment. To confirm the hypothesis that hemolysis can result in a lowering of plasma gelsolin levels, hemolysis was induced in rabbits, either acutely (by the injection of human serum) or subacutely (by the administration of phenylhydrazine). A fall in plasma gelsolin levels was seen, the rate of fall differing with the extent of hemolysis. Affinity adsorption of plasma from animals undergoing acute hemolysis with Sepharose beads coupled to the actin-binding protein DNase I, followed by immunoblotting of adherent proteins with antiactin antiserum demonstrated the presence of actin in circulating rabbit plasma. These studies suggest that under some conditions components of the red cell cytoskeleton are exposed to plasma proteins and that accelerated clearance of actin-gelsolin complexes may explain in part the depressed plasma gelsolin levels seen in patients with falciparum malaria.
Bivariate flow karyotyping in human Philadelphia-positive chronic myelocytic leukemia
Authors: Arkesteijn GJ , Martens AC , Hagenbeek A
Published: October 2019

Chromosome analysis on clinical leukemia material was done by means of flow cytometry (flow karyotyping) to investigate the applicability of this technique in the detection of leukemia-associated abnormalities. Flow karyotyping was performed on blood or bone marrow samples from eight patients with chronic myelocytic leukemia (CML) after a culture period of four days and arresting the cells in metaphase during the last 16 hours. Discontinuous density gradient centrifugation proved to be essential in removing debris and dead cells from the cell suspensions. By this procedure the mitotic index increase ranged from 2 to 80 times initial values. Chromosomes were isolated and stained with two base pair-specific fluorochromes, ie, chromomycin A3 and Hoechst 33258, and run through a specially designed dual-laser beam flow cytometer. Generally, 20,000 chromosomes or more were measured. The data were computer stored in list mode. Besides the clear detection of the specific Philadelphia chromosome, trisomies and other additional chromosomal aberrations [like an i(17q)] were visualized. Quantitative analysis revealed the percentage of subclones containing a certain chromosomal anomaly. Conventional cytogenetic analysis confirmed these findings. In seven of eight cases, CML could be diagnosed on the basis of the presence of a Philadelphia chromosome in the flow karyogram. In one of these seven, the conventional cytogenetic analysis was unknown at that time. The remaining six all matched the standard cytogenetics. The one failure out of eight could be attributed to the specific stimulating conditions in the culture. Although it is impossible by this technique to determine the position of the breakpoint, the involved chromosomes in the translocation event could be identified. In some cases, low percentages of aberrations could not be detected. This study shows that CML can be diagnosed on the basis of flow karyotypic results. Additional chromosomal aberrations can be detected provided that changes in the amount of DNA per chromosome have occurred. Exact quantification of the composition of subclones in the case of mosaicism appear difficult.
Distinct variants of erythrocyte protein 4.1 inherited in linkage with elliptocytosis and Rh type in three white families
Authors: McGuire M , Smith BL , Agre P
Published: October 2019

Hereditary elliptocytosis is a heterogeneous disorder resulting from defects in the erythrocyte membrane skeleton. Although some cases of elliptocytosis result from defects in spectrin, the specific structural abnormality has yet to be identified in the majority of cases. Protein 4.1 plays an essential role in erythrocyte membrane physiology, and deficiencies have been implicated in only a few rare cases of elliptocytosis. By using 4.1 immunoblots and a 4.1 radioimmunoassay we identified distinct variants of protein 4.1 in 15 elliptocytic members of three US white families with the Rh-linked form of elliptocytosis. Elliptocytic members of family G were heterozygotes for a low-molecular weight (mol wt) 4.1 variant (65,000 to 68,000 daltons; normal, 80,000) inherited in linkage with the Rz phenotype. Elliptocytic members of family C expressed a simple partial deficiency of protein 4.1 (63% of the normal level) that was inherited in linkage with the r phenotype. Elliptocytic members of family N were heterozygotes for a high-mol wt 4.1 variant (100,000 daltons) also inherited in linkage with the r phenotype. These studies indicate that mutant forms of protein 4.1 are not uncommon in elliptocytosis among whites and that different kindreds probably express different mutations. The observed linkage of elliptocytosis and Rh blood type most likely results from the close proximities of the 4.1 gene (site of the mutation) and the Rh gene, which is located nearby on the short arm of chromosome 1.
Glycoprotein IIb-IIIa and RGD(S) are not important for fibronectin- dependent platelet adhesion under flow conditions
Authors: Nievelstein PF , Sixma JJ
Published: October 2019

Previous studies have indicated that activated blood platelets interact with fibronectin through binding of fibronectin to the glycoprotein IIb- IIIa complex (GPIIb-IIIa). The cell attachment site of fibronectin with its crucial arg-gly-asp(-ser) [RGD(S)]sequence is involved in these bindings. We studied the importance of these interactions for the fibronectin dependence of platelet adhesion under flow conditions. An RGDS-containing hexapeptide (GRGDSP) was compared with a nonreactive control peptide (GRGESP). The GRGDSP-peptide inhibited thrombin-induced aggregation and adhesion under static conditions at 0.1 mmol/L. This concentration had no effect on platelet adhesion to nonfibrillar collagen type I in flow. GRGDSP at 1 mmol/L had a significant inhibitory effect at 1,500 s-1, but not at the lower shear rates of 800 and 300 s-1 where platelet adhesion is also fibronectin dependent. On the matrix of cultured human umbilical vein endothelial cells, 1 mmol/L GRGDSP had no effect on platelet adhesion. The relation between GPIIb- IIIa and fibronectin dependence was investigated with platelets of a patient with Glanzmann's thrombasthenia and monoclonal antibodies to GPIIb-IIIa using endothelial cell matrix (ECM) as a surface. Platelets of normal controls or a patient with Glanzmann's thrombasthenia showed a similar inhibition of adhesion in the presence of fibronectin-free plasma after the ECMs had been preincubated with antifibronectin F(ab')2 fragments. Incubation of platelets with anti-GPIIb-IIIa showed inhibition of platelet adhesion at high shear rates. Dependence on fibronectin for platelet adhesion was still observed even though separate experiments had shown that these anti-GPIIb-IIIa antibodies could block binding of radiolabeled fibronectin to thrombin-activated platelets. These data suggest the existence of another binding system for the interaction of platelets with fibronectin that may only appear when fibronectin is present on a surface.