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J Immunol Methods. 2000 Dec 1;246(1-2):51-9.

A method for investigating the role of homotypic adhesion in lymphocyte activation.

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1
Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, WC1E 7HT, London, UK.

Abstract

B cells activated via CD40 in vitro form striking homotypic aggregates, especially in the presence of costimuli such as anti-IgM, whereas those stimulated by anti-IgM alone do not. Blocking aggregation with anti-LFA-1alpha also significantly inhibits CD40-stimulated B cell proliferation, suggesting that homotypic adhesion is important for B cell activation via this receptor. To investigate this we have developed a culture system where murine B cells are stimulated in semi-solid agarose, which prevents cell-cell interactions. B cells respond to various mitogenic stimuli, including anti-CD40, in an essentially normal fashion when cultured in agarose. Furthermore, anti-LFA-1 exerts similar inhibitory effects on B cell proliferation regardless of whether the cells are in liquid, or semi-solid medium. These results indicate that homotypic aggregation is not necessary for CD40-stimulated B cell proliferation and the inhibitory effects of anti-LFA-1 could, therefore, be due to the delivery of a negative signal via this integrin, rather than as a result of inhibition of B cell clustering. Furthermore, reaggregation experiments indicated that anti-IgM-stimulated B cells are attracted into anti-CD40-generated clusters, even though they do not form clusters themselves. Taken together these results indicate that clustering is a consequence of B cell activation via CD40, rather than a necessary prelude to B cell proliferation. We postulate that homotypic aggregation may involve an unknown B cell-derived chemokine.

PMID:
11121546
[Indexed for MEDLINE]
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