Fibroblasts maintain the phenotype and viability of the rat heparin-containing mast cell in vitro.
Rat serosal heparin-containing mast cells (HP-MC) were maintained in vitro for as long as 30 days when co-cultured with mouse skin-derived 3T3 fibroblasts. In contrast, when the mast cells were cultured alone, on fibronectin-, gelatin-, or dermal-collagen-coated dishes, on acid and heat-killed fibroblasts in the presence or absence of 24 hr fibroblast-conditioned medium, or on a monolayer of mouse serosal macrophages, they failed to adhere to the dishes, released significant amounts of their histamine and lactate dehydrogenase, and stained with trypan blue, indicating a loss of viability. The rat serosal HP-MC cultured with the 3T3 fibroblasts became so adherent to the fibroblasts that the two cell types could be separated from one another only by trypsinization. The cultured HP-MC stained with both alcian blue and safranin and continued to synthesize proteoglycan at a rate comparable to that of freshly isolated cells. The 35S-labeled proteoglycan synthesized by these cultured cells, like that produced by freshly isolated rat serosal HP-MC, was a 750,000 to 1,000,000 m.w. proteoglycan containing only heparin glycosaminoglycans of 50,000 to 100,000 m.w. When HP-MC were cultured for 1 wk with the fibroblasts and were then incubated for 5 min with a 1/20 dilution of rabbit anti-rat IgE, they generated and released an average of 22 +/- 10 ng (mean +/- SD, n = 5) of prostaglandin D2 per 10(6) cells and exocytosed a higher net percentage of their total histamine content (44 +/- 11% [mean +/- SD, n = 8]) than did cells just isolated from the animal (6 +/- 4% [mean +/- SD, n = 4]). As assessed by electron microscopy, many of the cultured HP-MC resembled freshly isolated cells except that some secretory granules had fused with one another in some cells. Morphologically, after activation the cultured HP-MC underwent compound exocytosis like freshly isolated cells. These results demonstrate that the in vivo differentiated rat HP-MC maintain their histology, morphology, immunologic responsiveness, histamine content, and ability to synthesize heparin proteoglycan when co-cultured with living fibroblasts.
Levi-Schaffer
F
F
Austen
K F
KF
Caulfield
J P
JP
Hein
A
A
Bloes
W F
WF
Stevens
R L
RL
D000818:Animals; D000888:Antibodies, Anti-Idiotypic; D002470:Cell Survival; D002478:Cells, Cultured; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D005347:Fibroblasts / Q000502:physiology* / Q000648:ultrastructure; D006493:Heparin / Q000378:metabolism*; D006636:Histamine Release; D007073:Immunoglobulin E / Q000276:immunology; D008407:Mast Cells / Q000276:immunology / Q000502:physiology* / Q000648:ultrastructure; D051379:Mice; D010641:Phenotype; D015230:Prostaglandin D2; D011457:Prostaglandins D / Q000096:biosynthesis; D011509:Proteoglycans / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D012704:Serous Membrane / Q000166:cytology
D000888:Antibodies, Anti-Idiotypic; D011457:Prostaglandins D; D011509:Proteoglycans; C026408:anti-IgE antibodies; D007073:Immunoglobulin E; D006493:Heparin; D015230:Prostaglandin D2