Optimal reaction conditions for the in vitro blast transformation of rat lymph node cells (LNC) by neuraminidase (N) and galactose oxidase (GO) were determined. Treatment with either enzyme alone, or with GO before N, did not produce significant stimulation. The kinetics and magnitude of the response to NGO, as measured by [3H]TdR incorporation, were compared with those induced by periodate treatment and by concanavalin A (ConA). Rat LNC treated with NGO (but blocked by mitomycin C) caused blast transformation of untreated syngeneic LNC. The magnitude of this indirect response was about one-quarter of that attained for NGO-treated LNC, and the kinetics were somewhat slower. Supernatants from cultures of NGO-stimulated cells were not able to activate untreated cells, and, using Marbrook culture vessels, no evidence was found for the release of soluble factors that could either activate untreated cells or enhance the magnitude of indirect stimulation. Indirect stimulation, therefore, appears to require cell-cell contact. A second kind of cellular interaction was observed in experiments in which NGO-treated LNC were cultured at different cell densities in the presence or absence of mitomycin C-blocked filler LNC. In the absence of filler cells, the incorporation of [3H]TdR was a linear function of the number of NGO-treated cells in each culture over a wide range of cell densities. However, if filler cells were present, [3H]TdR-incorporation was substantially enhanced. Thus the filler cell population contains cells that assist or potentiate the blast response to NGO, probably by increasing the number of lymphocytes that are stimulated.
ASJC Scopus subject areas
- Cell Biology