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Characterization of an antigen-specific T helper cell clone and its products

University of British Columbia

A T helper cell clone, referred to as clone 9, was derived from an allogeneic mixed lymphocyte culture. Clone 9, as well as supernatant factor(s) derived from it, could help the cytotoxic T lymphocyte (CTL) responses of H-2 Db (Db) responder cells to alloantigens, or they could help the CTL responses of non- Db responder cells to Db alloantigens. Clone 9 cells or their factor(s) were active only when added during the first 24 hours of a five-day culture period. Clone 9 or its factor(s) could also synergize with interleukin-2 (IL-2)-containing medium in mounting cytotoxic responses to alloantigens. The helper activity in clone 9 supernatant was not due to IL-2 and it was specifically absorbed out by Db -spleen cells. The characterization of the Db -specific helper factor(ASHF) was facilitated by the isolation of a T hybridoma clone (clone 25), obtained from fusion of clone 9 cells with the T cell lymphoma, BW5147, and a B cell hybridoma that produced an IgM monoclonal antibody (clone 30 IgM) which bound ASHF. An additional monoclonal antibody (F23.1), which recognizes a determinant of the Vβ8 family of the T cell receptor, was also particularly useful for the characterization of ASHF. Analysis with these reagents showed that both clone 30 IgM and F23.1 immunoadsorbents could retain ASHF activity. Preabsorption of the ASHF with Db spleen cells prior to affinity purification over a clone 30 IgM column resulted in the absorption of Db-specific helper activity as well as the loss of a 50,000 molecular weight (MW) band on SDS-PAGE under reducing conditions. Furthermore, affinity purification of ASHF over the F23.1 immunoadsorbent, but not an irrelevant monoclonal antibody (mAb) column, also yielded a 50,000 MW molecule. Taken together, these findings suggest that the 50,000 MW molecule is a component of the ASHF and it is intimately related to the B chain of the T-cell receptor. The mode of action of clone 9 and its products in the induction bfCTL responses was also investigated. It was found that clone 9 and ASHF could help CTL responses by inducing IL-2 production in B6-stimulated cultures. In addition to ASHF, clone 9 cells also produced an additional factor(s) which participated in the induction of CTL responses. This additional factor(s) was referred to as IL-X. IL-X synergized with excess human recombinant IL-2 in the activation of CTL precursors (CTL-P) in the absence of antigenic stimulation. A model which involves the participation of ASHF, T helper cells, IL-2 and IL-X in the induction of CTL responses is proposed.

Text

2010-08-13

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http://hdl.handle.net/2429/27366

Microbiology and Immunology

University of British Columbia

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