Use of a modified Transwell culture to quantitate the contribution of non-IL-7 substances to growth of an IL-7-dependent pre-B cell clone.
Clone 3 is a mouse pre-B cell line that cannot grow in standard tissue culture media but is immortalized by coculture with a bone marrow-derived feeder layer. In addition, clone 3 cells can be passaged indefinitely in recombinant interleukin-7 (IL-7) in the absence of a feeder layer if maintained at high cell density. Using monoclonal antibody to IL-7 and Transwells ligated to dialysis membranes, we have examined the relative contributions of IL-7, both endogenous and exogenous, and of "non-IL-7" feeder layer factors to growth promotion of clone 3 cells. There is synergy between feeder layers and exogenous IL-7 that is most marked when the latter is present in suboptimal concentrations. The synergizing activity is not neutralized by antibody to IL-7 and appears to be freely dialyzable. This "non-IL-7" effect is common to two different feeder layers, the one derived from bone marrow (3E) being an IL-7 producer, and the other, 3T3 fibroblasts, making no detectable IL-7. These experiments reveal a substantial contribution of the dialyzable moiety to the total feeder layer effect, and are the first to demonstrate cytokine-dependent low-molecular-weight synergy in a coculture. This demonstration is possible because the synergizing cofactor(s) can cross a semipermeable membrane whereas the cytokine and its neutralizing antibody cannot.
Muirhead
M
M
Veterans Administration Medical Center, Little Rock, AR.
Schwartz
R C
RC
Waldschmidt
T J
TJ
D016475:3T3 Cells; D000818:Animals; D000911:Antibodies, Monoclonal; D001402:B-Lymphocytes / Q000166:cytology* / Q000187:drug effects / Q000276:immunology; D015152:Blotting, Northern; D001854:Bone Marrow Cells; D002455:Cell Division / Q000187:drug effects; D002999:Clone Cells; D046508:Culture Techniques / Q000379:methods; D003956:Dialysis; D015851:Interleukin-7 / Q000235:genetics / Q000276:immunology / Q000494:pharmacology*; D051379:Mice; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger; D011994:Recombinant Proteins / Q000494:pharmacology; D014158:Transcription, Genetic
D000911:Antibodies, Monoclonal; D015851:Interleukin-7; D012333:RNA, Messenger; D011994:Recombinant Proteins; D011061:Poly A; D012313:RNA