Koichi Ishimoto

Problems of mass screening for neuroblastoma: Analysis of false-negative cases

The Japanese mass screening (MS) system for neuroblastoma at 6 months of age has resulted in the earlier diagnosis of the tumor with excellent therapeutic results. However, some problems are involved in the present MS system. We present six false-negative cases, ages ranging from 1 year 11 months to 3 years 11 months. Neuroblastoma cell taken from four of these patients were studied biologically. These patients had advanced disease (one was stage III; five were stage IV). Three of the patients have died and one is terminally ill despite undergoing surgery combined with intensive chemotherapy. Cytogenetic analysis performed in three cases showed that all the cases had diploid chromosome mode associated with 1P-, double minutes (DMs), or marker chromosomes. N-myc oncogene analysis, performed in four cases, showed amplification in two; one patient had diploid chromosomes, but the other was not examined cytogenetically. These findings were strikingly different biologically from those of cases found by MS. The majority of neuroblastomas detected by MS were found to be triploid tumors without N-myc amplification. These findings suggest that the main reason for the false-negative results in the patients we examined is that they were tumor-free or the tumors were so small in size that they were unable to produce urinary vanillylmandelic acid and or homovanillic acid levels high enough to be detected at the time of MS. Therefore, we conclude that MS at 6 months of age is too early to detect neuroblastoma with a diploid chromosome mode and/or amplified N-myc oncogene. We propose that MS at the age of 1 year 6 months would be more effective to pick up these cases, because treatment strategies depend on the different biological characteristics of tumor cells.

Characterization of the common acute lymphoblastic leukaemia antigen (CD10) as an activation molecule on mature human B cells.

Distinct expression pattern of CD10 molecules during B cell activation was analysed using in vivo and in vitro systems. By two‐colour flowcytometrical analysis, CD10 was found to be expressed at a specificstage of in vivo activating B cells. The expression of CD10 during B cell activation appeared to be unique from that of other activation‐related B cell antigens including L29, MA6, OKT9 and OKT10. Although the expression of CD10 was associated with that of the activation‐related B cell antigens, CD 10+ B cells could be separated in the distinct fractions to those expressing other activation‐related B cell antigens when fractionated by cell gravity. In particular, certain CD10+ B cells were detected positive for the resting B cell antigen, L30. In vitro studies revealed that CD10+ B cells arose from CD10‐ B cells at an early step of B cell activation, and disappeared lately when activated by Staphylococcus aureus Cowan I. Collectively, CD10 was an antigen transiently expressed at an early phase of B cell activation process. Expression of CD10 and other antigens on Burkitts lymphomas (15 cases) was studied next. All cases were CD10+, and 87% (13 cases) were also L30+. In addition, six of CD10+ L30+ cases were L29+. This observation suggested that Burkitts lymphomas were phenotypically similar to the B cells at an early phase of activation, those expressing CD10 and L30, simultaneously. The present study has dissected a precise expression pattern of CD10 on mature B cell activation in vitro and in vivo, and could be implicated for the histogenesis of one of the poorly characterized B cell lymphoma, namely Burkitts lymphoma.

Granulocyte colony-stimulating factor directly affects human monocytes and modulates cytokine secretion

OBJECTIVE Recent reports have indicated that monocytes express receptors for the granulocyte colony-stimulating factor (G-CSF). The direct effects of G-CSF on cytokine secretion in monocytes were examined. MATERIALS AND METHODS A monocytic cell line NOMO-1 that secretes multiple cytokines upon stimulation with lipopolysaccharide (LPS) was used. Normal human monocytes were purified by negative selection using magnetic beads. Cells pretreated with or without G-CSF were stimulated with LPS, and the subsequent concentrations of cytokines and chemokines in supernatants were determined by sandwich enzyme-linked immunosorbent assay. RESULTS NOMO-1 cells were found to express receptors for G-CSF. Although G-CSF stimulation did not induce cytokine secretion, pretreatment with G-CSF significantly attenuated LPS-stimulated secretion of the proinflammatory cytokines tumor necrosis factor-alpha and interleukin (IL)-12 in NOMO-1 cells. Simultaneously, however, G-CSF pretreatment apparently enhanced LPS-induced secretion of IL-10 and monocyte chemoattractant protein-1, whereas secretions of IL-1beta, IL-6, and IL-8 were unaffected. When normal human monocytes from healthy volunteers were similarly examined, marked individual variations in LPS-induced secretion of cytokines were observed. Although some exceptions exist, a similar tendency as to the effects of G-CSF treatment on cytokine secretions as that in NOMO-1 cells was observed in human monocytes. CONCLUSIONS Our data suggest that G-CSF directly affects monocytes and modulates their cytokine secretion. NOMO-1 cells can provide an alternate model for in vitro culture of monocytes to investigate the effects of G-CSF on cytokine secretion by these cells.

Granulocyte colony-stimulating factor down-regulates the surface expression of the human leucocyte adhesion molecule-1 on human neutrophils in vitro and in vivo

Summary. The leucocyte adhesion molecule‐1 (LAM‐1) is the human homologue of the murine peripheral lymph node homing receptor, MEL‐14, and might play a crucial role in neutrophil localization at inflammatory sites. We have reported previously that recombinant human granulocyte colony‐stimulating factor (rhG‐CSF) stimulates or enhances several neutrophil functions in vivo, as well as in vitro. To further explore the possible role of G‐CSF in inflammation we studied the effect of rhG‐CSF on the surface expression of LAM‐1 on human neutrophils, both in vitro and in vivo. The expression of LAM‐1 by human neutrophils was investigated by indirect immunofluorescence using flow cytometry and monoclonal antibodies anti‐Leu‐8 and TQ1. A whole blood analysis was performed to minimize in vitro manipulation. Most circulating human neutrophils expressed LAM‐1 on the cell surface. Brief exposure of neutrophils to rhG‐CSF in vitro decreased the surface expression of LAM‐1. rhG‐CSF down‐regulated neutrophil LAM‐1 expression in a time‐ and dose‐dependent manner. Neutrophils from healthy volunteers and from patients who were receiving rhG‐CSF exhibited a decreased expression of LAM‐1 after rhG‐CSF administration, and the expression thereafter returned or overshot the pretreatment level after stopping rhG‐CSF administration. These findings indicate that rhG‐CSF down‐regulates the surface expression of LAM‐1 on human neutrophils in vivo, as well as in vitro, and G‐CSF might participate in neutrophil‐endothelial cell interaction in inflamed tissue.

Characteristics and management of patients with fetal neuroblastoma.

At 35 weeks 6 days of gestational age, ultrasound evaluation of the fetal abdomen showed a mixed cystic mass in the superior pole of the left kidney. The mass was suspected to be an adrenal hemorrhage or neuroblastoma. The diagnosis was fetal neuroblastoma. Differential diagnosis enabled the fetal neuroblastoma to be distinguished from adrenal hemorrhage. The parameters of diagnosis of fetal neuroblastoma include no specific ultrasonographic pattern, lack of palpability, and no tumor markers. However, certain features do characterize fetal neuroblastoma, such as little metastases, complete resection at operation, and excellent prognosis. In cases of suspected neuroblastoma, a laparotomy performed as soon as possible is generally regarded as the best course of treatment. Nonetheless, biological analyses of the tumor may prove in the future to be necessary for determining whether or not laparotomy is the best treatment.

Electromagnetic wave emitting products and “Kikoh” potentiate human leukocyte functions

Tourmaline (electric stone, a type of granite stone), common granite stone, ceramic disks, hot spring water and human palmar energy (called “Kikoh” in Japan and China), all which emit electromagnetic radiation in the far infrared region (wavelength 4–14 µm). These materials were thus examined for effects on human leukocyte activity and on lipid peroxidation of unsaturated fatty acids. It was revealed that these materials significantly increased intracellular calcium ion concentration, phagocytosis, and generation of reactive oxygen species in neutrophils, and the blastogenetic response of lymphocytes to mitogens. Chemotactic activity by neutrophils was also enhanced by exposure to tourmaline and the palm of “Kikohshi” i.e., a person who heals professionally by the laying on of hands. Despite the increase in reactive oxygen species generated by neutrophils, lipid peroxidation from unsaturated fatty acid was markedly inhibited by these four materials. The results suggest that materials emitting electromagnetic radiation in the far infrared range, which are widely used in Japan for cosmetic, therapeutic, and preservative purposes, appear capable of potentiating leukocyte functions without promoting oxidative injury.

Mass Screening for Neuroblastoma

Neuroblastoma is the most common solid tumour in infancy. The introduction of multimodality therapy has improved treatment, but the prognosis of advanced neuroblastoma remains dismal. In 1974, Sawada and colleagues started experimental screening for neuroblastoma at 6 months of age using the vanillylmandelic acid (VMA) spot test1,2. The excellent results of treatment in these patients led to the introduction in 1985 of a nationwide mass screening programme in Japan3,4. The idea of mass screening infants at 6 months is largely based on the following three findings: 1. Over 90% of children with neuroblastoma excrete increased quantities of catecholamine metabolites, which can be measured in random urine samples obtained from diapers: it is therefore possible to screen and detect these tumours before symptoms become manifest. 2. Children who present before 1 year of age have a better outlook than older children. 3. Patients with localized disease fare better than those with advanced disease.