Hiromi Ogura

Expression of granulocyte and granulocyte-macrophage colony-stimulating factors by human non-hematopoietic tumor cells

The expression of granulocyte colony-stimulating factor (G-CSF) mRNA was studied in human non-hematopoietic tumors, including 18 cases of lung cancers 10 cases of stomach cancers, three cases of glioblastomas, and one case each of breast phyllode sarcoma, thyroid cancer, and hepatocellular carcinoma. Northern blot analysis detected G-CSF mRNA in two of the lung cancer cases, in one of the glioblastoma cases, and in both the breast phyllode sarcoma and hepatocellular carcinoma cases. Since G-CSF receptors were not detected on the tumor cells by 125I-G-CSF binding assay, G-CSF autocrine loop are probably not involved in the growth of these G-CSF-producing tumors. Interestingly, granulocyte-macrophage colony-stimulating factor (GM-CSF) mRNA was concomitantly expressed in most of these G-CSF-producing tumors. No major gene deletions or rearrangements of G-CSF and GM-CSF genes were demonstrated by Southern blot analysis in the tumors expressing G-CSF and GM-CSF mRNAs except for one of the glioblastomas (G3) in which one chromosome 17 allele was deleted. Although the mechanism of the concomitant expression of G-CSF and GM-CSF mRNA is unknown, relatively high frequency of this phenomenon suggests the presence of common transcriptional factors acting on regulatory regions of G-CSF and GM-CSF genomes.

ATP11C is a major flippase in human erythrocytes and its defect causes congenital hemolytic anemia

Phosphatidylserine is localized exclusively to the inner leaflet of the membrane lipid bilayer of most cells, including erythrocytes. This asymmetric distribution is critical for the survival of erythrocytes in circulation since externalized phosphatidylserine is a phagocytic signal for splenic macrophages. Flippases are P-IV ATPase family proteins that actively transport phosphatidylserine from the outer to inner leaflet. It has not yet been determined which of the 14 members of this family of proteins is the flippase in human erythrocytes. Herein, we report that ATP11C encodes a major flippase in human erythrocytes, and a genetic mutation identified in a male patient caused congenital hemolytic anemia inherited as an X-linked recessive trait. Phosphatidylserine internalization in erythrocytes with the mutant ATP11C was decreased 10-fold compared to that of the control, functionally establishing that ATP11C is a major flippase in human erythrocytes. Contrary to our expectations phosphatidylserine was retained in the inner leaflet of the majority of mature erythrocytes from both controls and the patient, suggesting that phosphatidylserine cannot be externalized as long as scramblase is inactive. Phosphatidylserine-exposing cells were found only in the densest senescent cells (0.1% of total) in which scramblase was activated by increased Ca2+ concentration: the percentage of these phosphatidylserine-exposing cells was increased in the patient’s senescent cells accounting for his mild anemia. Furthermore, the finding of similar extents of phosphatidylserine exposure by exogenous Ca2+-activated scrambling in both control erythrocytes and the patient’s erythrocytes implies that suppressed scramblase activity rather than flippase activity contributes to the maintenance of phosphatidylserine in the inner leaflet of human erythrocytes.

A new glucose-6-phosphate dehydrogenase variant (G6PD Tsukui) associated with congenital hemolytic anemia

SummaryA new glucose-6-phosphate dehydrogenase (G6PD) variant associated with chronic nonspherocytic hemolytic anemia was found in a 20-year-old Japanese male who showed mild hemolysis after an upper respiratory tract infection. The patient had been noted to have jaundice and reticulocytosis several times before this episode. The enzyme activity of the variant was 1.5% of normal. The enzymatic characteristics were slow anodal electrophoretic mobility, high Km G6P, normal Km NADP, decreased heat stability, and a normal pH optimum. From these results, the enzyme was considered to be a new class 1 variant and was designated G6PD Tsukui.

Several new approaches to improvement of alpha interferon therapy in chronic myelogenous leukaemia.

Several basic experimental and clinical studies were carried out in an attempt to improve the efficacy of alpha interferon therapy for chronic myelogenous leukaemia (CML). First, the combined use of hydroxyurea (HU) and interferon (500-1000 mg daily) in interferon-resistant cases facilitated maintenance of reduced leucocyte production, or a reduction in the dose of interferon, although suppression of Philadelphia chromosome (Ph1)-positive clones was not observed in most cases. In order to try and decrease the rate of lymphoblastic crisis during the course of interferon therapy, we recently added methotrexate (MTX) (10-15 mg, weekly) to the treatment protocol. Since then, no lymphoblastic crisis has been observed. Second, the in vitro expression of alpha interferon-stimulated gene (ISG) mRNA was shown to be markedly decreased in granulocytes of one representative interferon-resistant case, compared to that in granulocytes of the three interferon-sensitive cases. Interestingly, it was found that the transcriptional activity in this case became almost normal when the blood granulocytes were controlled by the addition of HU. These findings suggest that the in vitro transcriptional assay of ISG mRNA may be clinically useful for predicting alpha interferon efficacy. Third, when genetically manipulated, alpha interferon-producing NIH/3T3 cells were co-transplanted using diffusion chambers into nude mice bearing a CML cell line, KU812, the CML tumour growth was shown to be markedly suppressed. This experimental model for alpha interferon replacement gene therapy suggests some directions for future studies on interferon therapy.

A novel PGK1 mutation associated with neurological dysfunction and the absence of episodes of hemolytic anemia or myoglobinuria

Phosphoglycerate kinase (PGK) deficiency affects three different organs: red blood cells (RBC), the central nervous system, and muscles. Next-generation sequencing identified a hemizygous PGK1 mutation (p.V217I) in a 16-year-old Japanese male patient presenting with intellectual disability and episodes of muscle weakness of unknown etiology. Enzymatic analysis demonstrated slightly lower RBC-PGK activity and compensatory increases of other glycolysis enzymes. This is the first PGK1 mutation found through next-generation sequencing.

Case Report: Alternation Therapy with Antileukemic Agents and Recombinant Human Granulocyte Colony-Stimulating Factor for RAEB in Transformation

Under the assumption that in some patients with refractory anemia with excess of blasts (RAEB), the abnormal clones might be less responsive to granulocyte colony-stimulating factor than normal clones, the authors tried alternation therapy with a recombinant form of this factor (rhG-CSF) and antileukemic agents in the treatment of two patients with RAEB in transformation. After repetition of the short-cycled alternation therapy, the hematologic findings of both patients were completely normalized and have remained so without any adverse side effects under the continuation of this therapy for more than 5 months. Judging from our clinical experience, the alternation therapy may be a new efficient therapeutic strategy for RAEB and some types of slowly progressive leukemia.