Related Experiment Videos
Mouse 3T3 fibroblasts induce rat basophilic leukemia (RBL-2H3) cells to acquire responsiveness to compound 48/80
M Swieter1, R J Midura, H Nishikata
1Laboratory of Immunology, National Institute of Dental Research, National Institutes of Health, Bethesda, MD 20892.
Abstract:
2H3 subline of rat basophilic leukemia (RBL-2H3) cells are mast cell analogs that lack responsiveness to nonimmunologic stimuli such as compound 48/80 and substance P. To determine if fibroblasts can influence this responsiveness, RBL-2H3 cells were cocultured with confluent monolayers of mouse 3T3 fibroblasts and assayed for secretagogue-induced histamine release. After 1 wk in coculture, RBL-2H3 cells began to respond to compound 48/80. Responsiveness reached a maximum at 2 wk in coculture and remained at this level for an additional 2 wk. Histamine release was specific, noncytotoxic, dose-dependent, and occurred even in the absence of extracellular Ca2+. No soluble factor from 3T3 cells was found that induced these alterations. Moreover, neither recombinant rat or mouse steel factor, at concentrations up to 250 ng/ml, was able to alter RBL-2H3 cell reactivity to compound 48/80. By 2 wk in coculture, RBL-2H3 cells also became responsive to substance P, although no changes in histamine content, Alcian blue+/safranin- staining or type of serine protease were detected. These results show that 3T3 fibroblasts cause an alteration in the functional repertoire of RBL-2H3 cells and that soluble steel factor cannot duplicate the effect.
Insights
Fibroblasts enhance the responsiveness of rat basophilic leukemia (RBL-2H3) cells to secretagogues like compound 48/80 and substance P. This study demonstrates fibroblast-induced changes in mast cell analog function, independent of soluble factors.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Rat basophilic leukemia (RBL-2H3) cells are mast cell analogs.
- RBL-2H3 cells typically lack responsiveness to non-immunologic stimuli like compound 48/80 and substance P.
Purpose of the Study:
- To investigate if co-culturing with fibroblasts can alter RBL-2H3 cell responsiveness.
- To determine the mechanism behind fibroblast-induced changes in RBL-2H3 cell function.
Main Methods:
- RBL-2H3 cells were co-cultured with mouse 3T3 fibroblasts for up to 4 weeks.
- Secretagogue-induced histamine release assays were performed.
- Analysis included testing for soluble factors and steel factor's effect.
Main Results:
- Co-culture with 3T3 fibroblasts induced responsiveness to compound 48/80 within 1 week, peaking at 2 weeks.
- Responsiveness to substance P was also observed after 2 weeks of co-culture.
- No soluble factor from 3T3 cells or steel factor replicated these functional changes.
Conclusions:
- 3T3 fibroblasts significantly alter the functional repertoire of RBL-2H3 cells.
- Fibroblast-induced changes in mast cell analog responsiveness are not mediated by soluble factors or steel factor.
- Direct cell-to-cell contact or matrix interactions likely play a role in modulating RBL-2H3 cell function.