The stable ascorbic acid derivative 2-O-alpha-D-glucopyranosyl-L-ascorbic acid (AA-2G) was used to investigate the role of ascorbic acid (AA) in B cell differentiation in vitro. AA-2G is stable in a solution unlike AA but is hydrolyzed by cellular alpha-glucosidase to release AA. Mouse spleen B cells were primed for 2 days with an anti-mu antibody in the presence of interleukin (IL)-4 and IL-5 and then washed and recultured with AA-2G in the presence of IL-4 and IL-5. AA-2G, but not AA, dose-dependently increased IgM production, the greatest enhancement being 150% at concentrations of more than 0.5mM. In the absence of IL-4 and IL-5, primed B cells produced a negligible amount of IgM, and AA-2G had no effect. AA-2G-induced IgM production in the presence of IL-4 and IL-5 was inhibited by the alpha-glucosidase inhibitor castanospermine. Intracellular AA content, depleted during the priming period, increased by adding AA-2G at the start of reculture. Treatment of B cells with AA-2G resulted in an increase in the number of IgM-secreting cells, CD138-positive cells and CD45R/B220-negative cells. The number of viable cells in untreated cultures decreased gradually, but the decrease was significantly attenuated by AA-2G, resulting in about 70% more viable cells in AA-2G-treated cultures. AA-2G caused a slight but reproducible enhancement of DNA synthesis and a slight decrease in the number of cells with a sub-G1 DNA content. These results demonstrated that AA released from AA-2G enhanced cytokine-dependent IgM production in anti-mu-primed B cells and suggest that its effect is caused through promoting the differentiation of B cells to plasma cells and attenuating the gradual decrease in the number of viable cells.

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A reduced number and/or reduced activity of natural killer (NK) cells, which are important for defense against a variety of cancers and viral infections, occur under various stress conditions and in patients with various diseases. In this article, we report that the 30% to 50% ethanol precipitate of oyster extract (EPOE50) dose-dependently enhanced the activity of mouse spleen NK cells in vitro and in vivo. The activity of EPOE50 was eluted with a molecular weight of about 2000 by gel filtration and was inactivated by periodate but not by proteinase K. The activity of highly purified NK cells was also augmented by EPOE50 but not by oligodeoxyribonucleotide 1585, which mimics bacterial DNA. Administration of EPOE50 to mice stimulated splenic NK cell activity without a change in splenic NK cell populations. Although the proliferation of B16 tumor cells in vitro was slightly stimulated by EPOE50, the growth of B16 melanoma in vivo was dose-dependently suppressed by administration of EPOE50. Taken together, our results indicate that EPOE50 augmented NK cell activity and that its administration to mice inhibited tumor growth presumably through the activation of NK cells and also suggest that the active substance is a sugar-containing oligomer or polymer and is not of bacterial origin.