Generation of B lymphocytes from a single hemopoietic progenitor cell in vitro.

The first stages of the pathway by which lymphocytes differentiate from hemopoietic stem cells were studied at a clonal level. When 211 interleukin 3 (IL-3)-induced blast colonies shown to be capable of differentiating into a variety of hemopoietic cells were individually transferred into wells containing a monolayer of stromal cells, growth in granulocyte, macrophage, megakaryocyte, or mast cell lineages was observed in 192 wells. In seven of these 192 wells, lymphoid cell growth also was seen. The lymphoid cells were proved to be B lymphocytes by phenotype and immunoglobulin gene rearrangement analyses and by demonstration of surface expression of IgM. The clonal origin of myeloid and B lymphocyte lineage cells was further confirmed by the generation of both myeloid and B lymphoid cells in the same well following FACS clone-sorting of IL-3 induced blast cells. These results provide in vitro evidence that cells of B lymphoid and myeloid lineage can originate clonally from single primitive hemopoietic stem cells.

Method of Harvesting the Crude Drug Based on Distribution of Alkaloids in the Hook and in the Stem with Hook of Uncaria rhynchophylla1.

The distribution of the content of the pharmacologically active oxindole alkaloids in the hook and in the stem of the crude drug "Cho-to-ko" (dried stem with hooks of UNCARIA RHYNCHOPHYLLA) prepared from the cultivated plants were investigated. The total oxindole alkaloid content of the stem was similar to that of the hook portion. It was also found that the area closest to the hook showed a slightly higher alkaloid content than the hook, and that the content of the oxindole alkaloid tended to decrease as the distance from the hook increased. Therefore, the presence of the hook does not affect the quality of the crude drug as far as the oxindole alkaloid content is concerned.

Effects of interleukin 3, interleukin 6, and granulocyte colony-stimulating factor on sorted murine splenic progenitor cells.

In order to examine the effect of recombinant growth factors on hemopoietic stem cells, these cells were enriched using wheat germ agglutinin (WGA) and monoclonal antibodies for lineage markers (Lin) such as B220, L3T4, Lyt-2, asialo GM1, Mac-1, and AL-21. Spleen colony-forming units (CFU-S) and in vitro colony-forming units were highly enriched in the fraction of WGA+Lin- spleen cells. To eliminate committed progenitor cells, spleen cells of 5-fluorouracil (5-FU)-treated mice were used. By this treatment, day-8 CFU-S disappeared but day-14 CFU-S were preserved. Day-14 CFU-S were also contained in the fraction of WGA+Lin- cells, which made up about 0.5% of total nucleated spleen cells. Moreover, this fraction contained primitive stem cells that could reconstitute the hemopoiesis of irradiated mice. Sorted WGA+Lin- spleen cells obtained from male 5-FU-treated mice were injected into lethally irradiated female mice. Southern hybridization using a mouse Y chromosome-specific probe showed that the bone marrow, spleen, and thymus of the recipients was reconstituted by male mouse-derived cells. When sorted WGA+Lin- spleen cells of the 5-FU-treated mice were cultured in vitro in the presence of recombinant interleukin 3 (IL-3), interleukin 6 (IL-6), and granulocyte colony-stimulating factor (G-CSF), colony formation was observed only in wells with IL-3, whereas unfractionated spleen cells formed colonies in the presence of IL-3, IL-6, or G-CSF. However, IL-6 but not G-CSF acted synergistically on enriched hemopoietic stem cells in the presence of IL-3. These data suggest that G-CSF or IL-6 did not affect primitive stem cells independently but showed the effect on these cells indirectly or synergistically with IL-3.

[Eight cases of intracranial arteriovenous malformations treated by conventional radiation therapy].

We conducted radiotherapy in 8 cases of intracranial arteriovenous malformation that had difficult embolectomy and other surgery from 1983 to October, 1988. The results in 6 cases of dural arteriovenous malformation at the cavernous sinus were "marked effect" in 4 cases, "relapse" in 1 case, and "no effect" in 1 case. One case of dural arteriovenous malformation at the sigmoid venous sinus demonstrated only slight irradiation effect. No irradiation effect was observed in 1 case of cerebral arteriovenous malformation.

Growth and differentiation of a human megakaryoblastic cell line, CMK.

Recently, a human megakaryoblastic cell line, CMK, was established from the peripheral blood of a megakaryoblastic leukemia patient with Down syndrome. Using this cell line, we studied the proliferation and differentiation of megakaryocytic cells in the presence of highly purified human hematopoietic factors and phorbol 12-myristate-13-acetate (PMA). In a methylcellulose culture system, interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) facilitated colony formation by CMK cells in a dose-dependent manner. The maximum stimulating doses of these factors were 10 and 200 U/mL, respectively. These concentrations were comparable to those that stimulate activity in normal hematopoietic cells. In contrast, granulocyte-colony stimulating factor (G-CSF), macrophage-colony stimulating factor (M-CSF), and erythropoietin (EPO) had no effects on the colony formation of CMK cells. In a liquid culture system, 20% of the CMK cells expressed glycoprotein IIb/IIIa (GPIIb/IIIa) antigen without hematopoietic factors, whereas 40% of the cells expressed GPIIb/IIIa with the addition of IL-3 and GM-CSF. EPO also slightly enhanced expression of GPIIb/IIIa. On the other hand, PMA inhibited growth of CMK cells and induced most of them to express the GPIIb/IIIa antigen. Furthermore, PMA induced CMK cells to produce growth activity toward new inocula of CMK cells. This growth factor (GF) contained colony-stimulating activity (CSA) in normal bone marrow (BM) cells. The activity was believed to be attributable mainly to GM-CSF, since 64% of this activity was neutralized by anti-GM-CSF antibodies and a transcript of GM-CSF was detected in mRNA from PMA-treated CMK cells by Northern blot analysis. These observations suggest that GM-CSF, as well as IL-3, should play an important role in megakaryocytopoiesis.

Congenital saccular aneurysm of the superior vena cava.

A 69-year-old woman was admitted to our hospital for surgical treatment of a mediastinal mass. Midsternotomy was performed which revealed a saccular aneurysm of the superior vena cava, 6 cm in diameter, in the right superior mediastinum. Aneurysmectomy was done by dividing a 15 X 10 mm venous connection with the superior vena cava and the patient's postoperative course was uneventful.