Cell density-dependent regulation of fibronectin splicing at the EDA region in fibroblasts: cell density also modulates the responses of fibroblasts to TGF-beta and cancer cell-conditioned medium.
Recently we reported that cancer cell-fibroblast interactions can modulate the expression of fibronectin (FN) isoforms in vitro, i.e. conditioned medium of human rectal adenocarcinoma cell line RCM-1 (RCM-1 CM) stimulated the expression of EDA-containing FN (EDA(+)FN) mRNA by fibroblasts and this stimulation was partly mediated by transforming growth factor-beta (TGF-beta) included in RCM-1 CM. In the present study, cell density was shown to regulate FN splicing at the EDA region in fibroblasts. Fibroblasts plated at a low cell density expressed a significantly higher percentage of EDA(+)FN mRNA than those plated at a high cell density. Moreover, fibroblast cell density modulated the effects of TGF-beta and RCM-1 CM on FN splicing at the EDA region differently. The time courses of their effects were similar to each other at a high cell density. At a low cell density, however, they were different. TGF-beta showed a relatively short-lived stimulation of EDA(+)FN mRNA, with the peak response 24 h after treatment, followed by a decline to the base line by 72 h. On the other hand, RCM-1 CM caused a prolonged stimulation, maintaining almost the maximum responses from 24 to 72 h. Thus, these results at a low cell density indicated the presence of a factor(s) other than TGF-beta in RCM-1 CM that stimulates the expression of EDA(+)FN mRNA directly or modulates the effect of TGF-beta. The use of several different cell densities might help in the search for new factors affecting FN splicing.
Second Department of Pathology, Miyazaki Medical College, Kiyotake, Japan.
T
T
Inoue
K
K
Nabeshima
Y
Y
Shimao
H
H
Kataoka
M
M
Koono
D000230:Adenocarcinoma / Q000378:metabolism / Q000473:pathology*; D017398:Alternative Splicing; D001483:Base Sequence; D017077:Culture Media, Conditioned; D017931:DNA Primers; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects; D005353:Fibronectins / Q000235:genetics*; D005786:Gene Expression Regulation; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics; D012004:Rectal Neoplasms / Q000378:metabolism / Q000473:pathology*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D017077:Culture Media, Conditioned; D017931:DNA Primers; D005353:Fibronectins; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta