Establishment of human embryonic stem (ES) cell lines has created great potential for regenerative medicine, because many types of human cells could be produced by their unlimited growth and pluripotent differentiation in culture. Primate and human ES cell lines have been established from blastocysts of monkey and surplus human blastocysts from fertility clinics. They showed several differences compared to mouse ES cells, including different expression patterns of surface antigens and very weak response to the LIF and gp 130 signals, which are widely used to repress spontaneous differentiation of mouse ES cell colonies. We have established several ES cell lines from blastocysts of the cynomolgus monkey. They can be maintained in culture as stem cell colonies, and they produce several differentiated cell types in culture. When such ES cells were transplanted into SCID mice, they produced teratomas containing many differentiated tissues.

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We investigated activation of MAP kinase subtype cascades in human neutrophils stimulated by cytokines and the role of MAP kinases in neutrophil activation. Distinct MAP kinase subtype cascades were activated in human neutrophils stimulated by granulocyte colonystimulating factor (G-CSF), granulocyte-macrophage colonystimulating factor (GM-CSF), tumor necrosis factor-α (TNF) and interleukin-1 β (IL-1) . G-CSF selectively activated the MEK-ERK cascade, and IL-1 selectively activated the MKK 3/6-p 38 cascade, respectively. GM-CSF activated the MEK-ERK cascade strongly and the MKK 3/6-p 38 cascade weakly. TNF activated the MKK 3/6-p 38 cascade strongly and the MEK-ERK cascade weakly. The potency of these cytokines to activate the MEK-ERK cascade was GM-CSF>G-CSF=TNF, whereas that to activate the MKK 3/6-p 38 cascade was TNF>GM-CSF>IL-1. JNK was not activated by any cytokine. GM-CSF-and TNF-induced superoxide release and adherence were inhibited by PD 98059 (MEK inhibitor) as well as SB 203580 (p 38 inhibitor) . IL-1, a selective activator of p 38 cascade, also induced superoxide release and up-regulation of CD 11 b and CD 15, and both responses were inhibited by SB 203580. The results show that (a) G-CSF, GM-CSF, TNF and IL-1 activated the distinct MAP kinase subtype cascade in human neutrophils; (b) activation of ERK and p 38 is involved in superoxide release and adherence induced by cytokines.

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Glucosamine, an amino monosaccharide occurring naturally in the connective and cartilage tissues, contributes to maintain the strength, flexibility and elasticity of these tissues. In recent years, glucosamine has been widely used to treat osteoarthritis in humans. Neutrophils, which usually function as the primary defenders in acute bacterial infections, are also implicated in the destructive inflammatory responses in arthritis. Recently, we have revealed the inhibitory actions of glucosamine on neutrophil functions. Glucosamine suppressed superoxide generation by neutrophils, and inhibited phagocytosis of opsonized particles. Furthermore, glucosamine inhibited the release of granule enzyme from phagocytosing neutrophils, and repressed chemotaxis. In addition, glucosamine significantly inhibited the upregulation of adhesion molecule CD 11 b, polymerization of actin and phosphorylation of p 38 MAPK, associated with neutrophil activation. Together these observations likely suggest that glucosamine suppresses the neutrophil functions, thereby exhibiting the anti-inflammatory actions in arthritis.

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Neural stem cells are a subtype of progenitor cells in the nervous system that can selfrenew and generate neurons, astrocytes, and oligodendrocytes. Stem cells have been isolated from many regions of the embryonic nervous system. Recent studies reveled that adult neural stem cells exist in the adult neurogenic regions, the hippocampus and the subventricular zone, and in some non -neurogenic regions, including spinal cord. Now, neural stem cells can be isolated and cultured as floating, multicellular neurospheres. This review summarizes isolation methods for neural stem cells and the regulatory mechanisms of neural differentiation. The potential therapeutic uses of neural stem cells are also discussed.

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Imbalance of Th1/Th2 responses is associated with the development of human allergic and autoimmune diseases. We found that Txk expression is restricted to Th 1/Th 0 cells that have IFN-γ producing potential, and that Txk regulates differentiation of naive T cells into Th 1 cells in normal humans.
We first studied expression of Txk in normal T cells cultured with various Th 1 and Th 2 cytokines. We found that Txk expression in the T cells was enhanced by Th 1 cytokines (IL-12, IFN-γ and IL-18) . Th 2 cytokines (IL-4 and IL-13) inhibited Txk expression. We next studied whether Txk expression was modulated in patients with bronchial asthma, atopic dermatitis and rheumatoid arthritis. Txk expression in the peripheral blood T cell is reduced in patients with allergic diseases. In contrast, synovium infiltrating T cells exhibited enhanced Txk expression. Collectively, we found that Th 1 as well as Th 2 cytokines regulated Txk expression. In turn, aberrant Txk expression was involved in the Th 1/Th 2 imbalance in patients with allergic and autoimmune diseases. Txk may become a possible therapeutic target of the diseases with Th1/Th2 imbalance.

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This study was aimed to reveal the stellate cell derived factors that regulate hepatocyte proliferation in culture. Rat hepatocytes and stellate cells were cultured in serum free Williams-E medium. We used hepatocyte monoculture and two different co-cultures of hepatocytes and stellate cells ; 1) co-culture on the same surface (Co-mix.) and 2) co-culture without contact between hepatocytes and stellate cells using a culture insert (Co-sep.) . Changes in the number and the DNA synthesis of hepatocytes was estimated. Although the number of hepatocyte decreased to 76% of the original at 48 h after starting mono-culture, it was kept at 106% in Co-mix and increased to 135% in Co-sep. The hepatocyte DNA synthesis was enhanced by carbenoxolone, gap junction blocker, in Co-mixo and reduced by NK 1 in each coculture. PD 153035, an EGF receptor specific inhibitor, had no effect. HeparitinaseI (20 mU/ml) and sodium chrolate (25 mM) reduced the hepatocyte DNA synthesis in Co-sepo to 71%. Activation of MAPK was induced in hepatocytes stimulated by conditioned mediums. These results indicated that hepatocyte proliferation was stimulated in the presence of stellate cells through HGF, extracellular heparan sulfate, and heparan sulfate proteoglycan, and might be negatively regulated by gap junction dependent mechanism.

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