[Cartilage-derived morphogenetic protein 1 initiates chondrogenic differentiation of human dermal fibroblasts in vitro].
OBJECTIVE
To explore the factors influencing the differentiation of fibroblasts into chondrocyte phenotype induced by a growth factor, cartilage-derived morphogenetic protein 1 (CDMP1).


METHODS
Fibroblasts isolated from foreskin obtained during circumcision were cultured in the forms of micromass and monolayer culture. The culture fluid of the fibroblasts at the passage 2, 7, and 10 was added with CDMP1 of the concentrations at the concentrations of 10, 30, 100, and 300 ng/ml respectively and co-cultured for 7 days. RT-PCR was used to detect the expression of bone morphogenetic protein receptor (BMPR), activin receptor-like kinase (ALK) receptor, collagen types II, IV, and X before and after CDMP1 induction. Western blotting was used to detect the protein expression of collagen types II, a transcriptional factor Sox9, and aggrcan before and after the induction. Flow cytometry was used to detect the superficial markers CD29, CD105, CD106, and CD166, and the expression of collagen types I and II.


RESULTS
Western blotting showed that the collagen type II positive cell rate in the passage 5 cells was 74.3% +/- 0.4%, not significantly different from that of the passage 2 cells (73.4% + 0.5%). When the concentrations of CDMP1 were 10 and 30 ng/ml no expression of aggrecan and collagen type II was detected, When the concentrations of CDMP1 was 100 and 300 ng/ml, the expression of aggrecan and collagen type II could be detected and without significant differences between these 2 concentrations. The expression of aggrcan and collagen type II mRNA disappeared in the monolayer cultured P2 and P5 fibroblasts induced by CDMP1 for 14 days, However, RT-PCR and Western blotting showed expression of collagen type II, aggrcan and SOX9 in the micromass cultured fibroblasts. RT-PCR showed that all fibroblasts cultured in vitro expressed ActR-I/ALK-2, BMPR-IA/ALK-3, and BMPR-IB/ALK-6 genes, and the expression of these genes significantly increased after CDMP1 for 7 days.


CONCLUSIONS
CDMP1 stimulates the human dermal fibroblasts expanded in vitro to differentiate into chondrogenic phenotype in a dose dependent manner. Three-dimensional culture environments accelerate the chondrogenic differentiation. The expression of ALK receptors may involve the CDMP1 stimulated differentiation of fibroblasts.
Cui
Lei
L
Department of Plastic and Reconstructive Surgery, Shanghai Ninth People's Hospital, Shanghai Second Medical University, Shanghai 200011, China.
Yin
Shuo
S
Deng
Chen-liang
CL
Li
Yu-lin
YL
Yang
Guang-hui
GH
Chen
Fu-guo
FG
Liu
Wei
W
Liu
De-li
DL
Cao
Yi-lin
YL
D000293:Adolescent; D000328:Adult; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D002648:Child; D019902:Chondrocytes / Q000166:cytology*; D020405:Dermis / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology*; D055428:Growth Differentiation Factor 5 / Q000494:pharmacology*; D006801:Humans; D008297:Male; D055815:Young Adult
D055428:Growth Differentiation Factor 5