Akihiko Yabuhara

Development of Natural Killer Cytotoxicity during Childhood: Marked Increases in Number of Natural Killer Cells with Adequate Cytotoxic Abilities during Infancy to Early Childhood

ABSTRACT: The cytotoxicity of natural killer (NK) cells against X562 cells and their responsiveness to interferonα and interleukin 2 (IL-2) were studied throughout child-hood using 51Cr-release and single-cell assays. Although NK activity was extremely low in the neonatal period, it almost reached the adult level during 1 to 5 mo of age and remained at that level thereafter. At the single-cell level, the binding, lytic, and recycling abilities were also depressed in the neonatal period, but these abilities improved conspicuously after this period; in particular, the lysis and recycling were at higher levels during 6 mo to 4 y of age. The absolute numbers of circulating cytotoxic NK cells were high during infancy to early childhood: they were 54 ± 24 (mean ± SD/mm3) in neonates, 115 ± 48 in 1− to 5-mo-old infants, 121 ± 42 in 6− to 12-mo-old infants, 93 ± 26 in 1− to 4-y-old children, and 42 ± 16 in adults. Inter feron-α and IL-2 could enhance NK activity throughout childhood. The IL-2 enhancement was prominent especially in the neonatal period; IL-2 yielded a 2.5-fold in crease in the number of cytotoxic cells and improved the recycling to the adult level. At older ages, interferon-α and IL-2 yielded 1.4-and 1.9-fold increases in the number of eytotoxic cells, respecively, but did not enhance the recycling. The iacreased number of NK cells with adequate cytotoxic abilities during infancy to early childhood indicates the predominance of NK immunity during these periods. IL-2 is a cytokine that induces high levels of NK cytotoxicity even in neonates.

Giant lymph node hyperplasia (Castleman's disease) with spontaneous production of high levels of B-cell differentiation factor activity.

A 13‐year‐old girl presented with general fatigue, back pain, anemia, hyperimmunoglobulinemia, and a mediastinal mass on chest radiograph. A mass was surgically removed, and its histologic examination determined the diagnosis of giant lymph node hyperplasia (Castlemans disease). With removal of the hyperplastic lymph node, the clinical symptoms soon disappeared and the abnormal laboratory findings were markedly improved within 1 month: serum IgG levels decreased from 4350 mg/dl to 1829 mg/dl. Immunostaining on the lymph node sections revealed polyclonal B‐lymphocyte and T‐lymphocyte populations. The patients lymph node cells were cultured without any mitogenic stimulation, and the culture supernatants were assayed for their B‐cell differentiation factor (BCDF) activity to induce IgG production by our Epstein‐Barr virus‐transformed cell line. The patients lymph node cells produced high levels of BCDF activity: the supernatants could increase the IgG production from 140 ng/ml to 410 ng/ml when the values became from 140 ng/ml to 142 ng/ml or 148 ng/ml with those of the control lymph node cells. These results suggest that the hyperimmunoglobulinemia and its prompt improvement with removal of the hyperplastic lymph node may have been related to the spontaneous production of high levels of BCDF activity by the lymph node cells in the patient.

Induction of lymphokine-activated killer and natural killer cell activities from cryopreserved lymphocytes

Lymphokine‐activated killer (LAK) and natural killer (NK) cells were studied for their capacity to retain cytotoxicity after cryopreservation. LAK cells were generated by a 4‐day culture of lymphocytes with recombinant interleukin‐2 (rIL‐2). Cytotoxicity was measured by 51Cr‐release assay at effector:target ratios of 10:1 to 80:1. Cryopreserved LAK cells retained 58.8 to 87.4 percent of cytotoxicity, as compared with that in fresh control cells. Cryopreserved NK cell activity against K562 and Molt‐4 targets was 45.7 to 67.9 percent of the respective values of the fresh control cells. The responsiveness of NK cells to polyinosinic‐polycytidilic acid (poly I:C), interferon‐α (IFN‐α), or rIL‐2 remained intact. Activated NK cell activity after poly I:C or IFN‐α stimulation and that after rIL‐2 were, respectively, comparable to and higher than the endogenous NK cell activity of the fresh cells. The composition of lymphocyte subsets as determined by flow cytometry using monoclonal antibodies did not change after cryopreservation, indicating that cellular loss of the given subsets did not occur during the procedure. The retention of substantial levels of cytotoxicity in cryopreserved LAK and NK cells may make them promising candidates as cytotoxic effector cells.

Frequently relapsing minimal change nephrotic syndrome with natural killer cell deficiency prior to the overt relapse of Hodgkin's disease

A 15-year-old boy developed minimal change nephrotic syndrome (MCNS) during remission of Hodgkins disease. Natural killer (NK) cell activity was practically absent at the onset of MCNS, with a value of 3% compared with the normal value of 44.1%±7.8% (mean ± SD). Treatment with prednisolone resulted in transient remission of MCNs and partial improvement of NK cell activity. Extensive investigations for Hodgkins disease were performed at 1- to 3-month intervals; a relapse finally became apparent 25 months after the diagnosis of MCNS. Successful treatment of Hodgkins disease resulted in complete disappearance of proteinuria and normalisation of NK cell activity. Frequently relapsing MCNS with NK cells deficiency during remission of Hodgkins disease appears to imply its subclinical relapse.

Neurologically normal development of a patient with severe methionine adenosyltransferase I/III deficiency after continuing dietary methionine restriction.

BACKGROUND There is not much information on established standard therapy for patients with severe methionine adenosyltransferase (MAT) I/III deficiency. CASE PRESENTATION We report a boy with MAT I/III deficiency, in whom plasma methionine and total homocysteine, and urinary homocystine were elevated. Molecular genetic studies showed him to have novel compound heterozygous mutations of the MAT1A gene: c.191T>A (p.M64K) and c.589delC (p.P197LfsX26). A low methionine milk diet was started at 31 days of age, and during continuing dietary methionine restriction plasma methionine levels have been maintained at less than 750 μmol/L. He is now 5 years old, and has had entirely normal physical growth and psychomotor development. CONCLUSIONS Although some severely MAT I/III deficient patients have developed neurologic abnormalities, we report here the case of a boy who has remained neurologically and otherwise normal for 5 years during methionine restriction, suggesting that perhaps such management, started in early infancy, may help prevent neurological complications.

A Recycling Defect as a Characteristic of Natural Killer Cells in Childhood Acute Lymphoblastic Leukemia

ABSTRACT: The cytolytic function of natural killer (NK) cells and their responsiveness to interferon-α and IL-2 were investigated in children with acute lymphoblastic leukemia (ALL) using 51Cr-release and single-cell assays. For comparison, such NK cell functions were similarly assayed in neuroblastoma. NK activity in ALL children was extremely low at onset, but it increased gradually during remission and finally reached normal levels. At the single-cell level, their NK cells at onset were defective in the binding, lytic, and recycling abilities. Although the binding and lytic defects improved to normal levels during remission, the recycling, which increased gradually during remission, was still low even after the long-term remission in ALL: the maximal recycling capacity values were 1.9 ± 0.4 (p < 0.001) at onset and 4.6 ± 0.6 (p < 0.05) after 5 y of complete remission, as compared to the value in control children of 5.4 ± 0.7. On the other hand, children with neuroblastoma had no recycling defect after completing the therapy: their maximal recycling capacity value was 5.6 ± 0.7. Bone marrow cells in ALL were also depressed in their recycling ability at all stages. Interferon-α and IL-2 could enhance NK activity and IL-2 could generate lymphokine- activated killer activity at all stages of ALL; however, the recycling defect hardly improved with these treatments. Thus, NK cells in childhood ALL have a recycling defect as a functional characteristic.

Absence of responsiveness to interferon-γ but not to interleukin 2 and depressed recycling as natural killer cell abnormalities in childhood lymphohistiocytic syndrome

To characterize the abnormalities of natural killer (NK) cells in childhood lymphohistiocytic syndrome (LHS), we investigated the number and cytolytic functions of circulating NK cells in 10 LHS children using flow cytometry, 51Cr-release and single-cell assays. In the active phase, the numbers of CD16+ or CD56+ cells and NK activity were normal in more than half of the patients or otherwise decreased. Despite the treatment with interferon-gamma (IFN-gamma) there was no significant increase in NK activity in the children with LHS: the values (mean +/- S.D.) of 32.2% +/- 14.2% became 35.0% +/- 13.3% (P > 0.05) when the control values changed from 45.5% +/- 8.5% to 54.2% +/- 10.1% after the stimulation. However, the NK cells normally responded to interleukin 2 (IL-2). In contrast, NK cells from 9 patients with infectious mononucleosis (IM) responded well to both IFN-gamma and IL-1 (P < 0.01). At the single-cell level, their NK cells had defective recycling capacity with normal killing capacity. The maximal recycling capacity (MRC) values (mean +/- S.D.) were 3.6 +/- 0.8 as compared to the control value of 5.5 +/- 0.9 (P < 0.01). Two of the patients studied had extremely high levels of serum IFN-gamma (9.8 U/ml and 158.0 U/ml) as compared to the control value of < 0.4 U/ml. NK cells may have been strongly stimulated by IFN-gamma in vivo, which probably yields superficially normal NK cell activity in the face of the absence of responsiveness to IFN-gamma but not to IL-2. The defective recycling may be another abnormality of NK cells in LHS.

Beneficial effect of granulocyte colony-stimulating factor in an infant with Pasteurella multocida brain abscess.

Sir: A 2-month-old boy was hospitalized with fever and irritability on 9 October 1990. Six weeks earlier, he had been scratched on the anterior fontanelle by a domestic cat. Computed tomography (CT) scans revealed a large multilocular abscess in the left anterior part of the brain which communicated with the left lateral ventricle. Cultures of CSF grew Paswurella multocida, which was also isolated f rom the oral cavity of the family cat. Intravenous injection of large doses of antibiotics and continuous irrigation of the abscess cavity with antibiotics resulted in a temporary improvement in the CSF findings, fol lowed by a further exacerbat ion (cell count in CSF, 7300/gl) and by absence of increase in the neutrophit count (1600/111), As presented in Fig. 1, random migration and chemotaxis of the patient s neutrophils showed lower levels as compared with those of age-matched controls, but their 0 2 production did not [1, 2]. Accordingly, on 27 October , we injected