Human dermal fibroblast clones derived from single cells are heterogeneous in the production of mRNAs for alpha 1(I) procollagen and transforming growth factor-beta 1.
Fibroblast clonal heterogeneity has been reported for growth and protein synthesis, but quantitative studies of synthetic phenotype at the pretranslational level have been limited because of difficulty in reliably growing large numbers of clonal cells. We have recently shown a unique stimulatory activity of low oxygen tension in the early phases of clonal growth, which can be used to establish clonal fibroblast cultures suitable for Northern analysis. Using this methodology, we have measured mRNA levels of alpha 1(I) procollagen and transforming growth factor-beta 1 (TGF-beta) both at baseline and after TGF-beta stimulation in a total of 43 clones derived from single cells and from seven different cell strains. We report a remarkable baseline heterogeneity, commonly four- to six-fold, in procollagen mRNA levels among clones and between clones and their parent cultures. Conversely, differences in baseline TGF-beta mRNA levels among clones were either not present or less than onefold. The clonal phenotypic expression of alpha 1(I) procollagen mRNA remained stable after eight additional cell passages. TGF-beta stimulation of itself (autoinduction) was highly variable among clones (range of increases 30% to 150%), and up-regulation of procollagen mRNA levels after TGF-beta stimulation was detected in only 15 (54%) of 28 clonal cultures (range of increases 30% to 353%). A notable lack of correlation was found between baseline mRNA levels of TGF-beta and alpha 1(I) procollagen in clonal cultures. In conclusion, fibroblast clonal populations are remarkably heterogeneous in their baseline procollagen mRNA levels and in their response to TGF-beta.
Falanga
V
V
Department of Dermatology and Cutaneous Surgery, University of Miami School of Medicine, Florida 33136, USA.
Zhou
L H
LH
Takagi
H
H
Murata
H
H
Ochoa
S
S
Martin
T A
TA
Helfman
T
T
D000328:Adult; D002478:Cells, Cultured; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D011347:Procollagen / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis*; D012867:Skin / Q000166:cytology / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000235:genetics*
D011347:Procollagen; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta