Yoshihisa Wakabayashi

Immunodeficiency and IL‐6 production by peripheral blood monocytes in multicentric Castleman's disease

Summary. To study the pathogenesis of multicentric Castlemans disease (MCD), IL‐6 producing cells and immune function were investigated in four MCD patients. The expression of IL‐6 mRNA in one MCD lymph node was analysed by in situ hybridization. IL‐6 mRNA expressing cells were scattered in the interfollicular areas and did not resemble plasma cells. Spontaneous IL‐6 production was detected in the culture supernatants of peripheral blood mononuclear cells (PBMNC) from four patients. The IL‐6 producing cells among the PBMNC were found to be monocytes by both in situ hybridization and immunohistochemistry. We evaluated immune function in four MCD patients. These studies show: (1) a negative PPD skin test in 3/4 patients. (2) decreased IL‐2 production in 3/4 patients, (3) decreased T cell colony formation in 3/4 patients, (4) decreased NK activity and NK cell number in 2/4 patients, (5) increased soluble IL‐2 receptor in 4/4 patients, and (6) decreased CD4/CD8 ratio in 3/4 patients. These results show that MCD resembles, in several ways, acquired immunodeficiency syndrome (AIDS).

Effects of bestatin (Ubenimex) on human T-cell colony formation.

The antitumor action of bestatin is considered to be an indirect action mediated by T-cells. Therefore, we investigated the effects of bestatin on the differentiation and proliferation of human precursor T-cells using a colony formation technique. Bestatin did not increase the overall number of T-cell colonies, but it significantly increased in CD4+ cell and significantly decreased in CD8+ cell subpopulations. It also induced CD4+ 8+ cells.These findings indicated that bestatin acts on precursor T-cells to induce the differentiation of these cells into CD4+ cells.

Colony-Stimulating Factors in B-Cell Colony Formation in Patients with B-Cell Chronic Lymphocytic Leukemia

A new in vitro colony growth assay system method was found to be reliable in its use for evaluating B-cell proliferation in normal subjects and in 9 patients with B-cell chronic lymphocytic leukemia (B-CLL). The method is based on a phytohemagglutinin (PHA)-stimulated monocyte and T-cell-conditioned medium (PHA-MTCM) composed of PHA, silica, normal monocytes and normal T cells. The number of colonies proliferated was significantly greater in 5 patients who had not undergone treatment than in normal subjects (1,417 +/- 660 vs. 661 +/- 119) (p less than 0.002). Normal cultured B-cell colonies were shown to be 71% surface IgM colonies, and 6% cytoplasmic IgA colonies with the appearance of blastic cells. B-CLL colonies, on the other hand, were demonstrated to be monoclonal with the same CLL circulating cells being retained. We also studied the effect of interleukin-2 (IL-2) on B-cell colony growth assay in 4 patients with B-CLL cells. Only 1 patient with M protein responded to IL-2, proliferated and expressed IL-2 receptors. Although 3 patients without M protein did not respond to IL-2, they did respond to the supernatant, and they proliferated but expressed no IL-2 receptors.

Megakaryocyte Colony-Stimulating Factor and Burst-Promoting Activity in LPS-Treated Mouse Spleen Cell-Conditioned Medium

In order to clarify the mechanism(s) of increased splenic hematopoiesis noted in lipopolysaccharide (LPS)-injected mice, the effects of spleen cell-conditioned medium (SPCM) on megakaryocyte colony (CFU-meg) formation and early erythroid (BFU-e) differentiation were investigated. After spleen cells from LPS-injected mice were incubated for 3 days, the SPCM was assayed for megakaryocyte colony-stimulating factor (Meg-CSF) in CFU-meg assay and for burst-promoting activity (BPA) and erythropoietin (Epo) in erythroid colony assays (i.e., CFU-e, BFU-e). Colony formation of CFU-meg and BFU-e peaked with the addition of 30 and 10-15% SPCM, respectively. Spleen cells from LPS-injected mice produced Meg-CSF and BPA when compared with controls. However, conditioned medium from spleen cells depleted of phagocytic cells had low Meg-CSF and BPA. SPCM did not contain detectable quantities of Epo. It appears likely that local splenic production of Meg-CSF and BPA may affect proliferation of CFU-meg and erythroid progenitor cells in the spleen.

T-cell colony formation in healthy homosexual men without anti-human immunodeficiency virus antibody.

In order to study the abnormal differentiation and proliferation of precursor T cells in homosexual men, a study was made of T-cell colony formation in 15 healthy homosexual men without anti-human immunodeficiency virus (HIV) antibody. The colony forming method used was the one-step monolayer technique using phytohemagglutinin as an inducer. Our results revealed that colony formation was extremely reduced and 3 cases showed no colony growth. However, with the addition of recombinant interleukin 2 to the culture system, the number of colonies increased up to the same level as in heterosexual controls. In addition, the analysis of CD3+, 4+, and 8+ cells of colony component cells revealed a decrease in CD3+ and 4+ cells. However, with the addition of recombinant interleukin 2, the numbers of CD3+ cells distinctly increased. The above results suggest that a disorder in the differentiation and proliferation of precursor T cells in homosexual men might be one of the causes of immunodeficiency.

Mouse peritoneal macrophages produce CFU-gm and CFU-meg growth-promoting factors

The effect of peritoneal macrophage-conditioned medium (macrophage CM) from lipopolysaccharide-injected mice on granulocyte-macrophage colony (CFU-gm) and megakaryocyte colony (CFU-meg) formation was examined using a plasma clot culture system. Macrophage CM stimulated mouse bone marrow cells to form CFU-gm and CFU-meg in the presence of pokeweed mitogen-stimulated spleen cell-conditioned medium, but it had no effect on colony formation in the absence of exogenous colony-stimulating factors (CSF). CFU-gm and CFU-meg colony formation was enhanced by low concentrations of exogenous GM-CSF or Meg-CSF alone. These data demonstrate that macrophage CM contains an activity that sensitized CFU-gm and CFU-meg to exogenous CSF.