Multiple Directional Differentiation Difference of Neonatal Rat Fibroblasts from Six Organs.
OBJECTIVE
Fibroblasts are abundantly distributed throughout connective tissues in the body and are very important in maintaining the structural and functional integrity. Recent reports have proved that fibroblasts and mesenchymal stem cells share much more in common than previously recognized. The aim of this study was to investigate comparative studies in fibroblasts on the differences in the expression of molecular markers and differentiation capacity from different organs.


METHODS
Combined trypsin/collagenase enzymes digestion method was used to isolate and culture the fibroblasts derived from heart, liver, spleen, lung, kidney and skin. Cell activity was determined by methyl thiazolyl tetrazolium (MTT) assay. Common molecular markers for fibroblasts such as vimentin, DDR2 and FSP1, stem cell markers nanog, c-kit and sca-1 were detected by RT-PCR, immunofluorescence and western blotting. The osteogenic, adipogenic and cardiogenic differentiations of fibroblasts were performed by inductive culture in special mediums, and analyzed by Alizarin red, Oil red O and immunofluorescence staining of cTnT respectively.


RESULTS
The proliferation rate of fibroblasts in lung was faster than in other five organs. Common molecular markers for fibroblasts were expressed differently in different organs. DDR2 was strongly expressed in fibroblasts in the heart, partly expressed in the heart, skin, liver and spleen. Interestingly, no expression of DDR2 was detected in liver and kidney. However, vimentin and FSP1 were consistently expressed in fibroblasts from skin, liver, kidney, spleen and lung. nanog expression in fibroblasts from lung was less than that from heart, skin, liver and spleen (P < 0.01). c-kit expression in fibroblasts from heart, skin and kidney was higher than that from spleen (P < 0.05), while the c-kit positive fibroblasts from liver was obviously higher than that from spleen (P < 0.01). But sca-1 expression in fibroblasts from lung was the lowest among six organs (P < 0.01). Directed differentiation in vitro had demonstrated that skin fibroblasts had the strongest multiple differentiation potential, and the next was cardiac fibroblasts. And fibroblasts in liver and kidney had the advantage in myocardial differentiation, while fibroblasts in spleen only had the advantage in osteogenic differentiation.


CONCLUSIONS
There are obvious heterogeneity in molecular markers and muti-directional differentiation in fibroblasts from six organs.
Department of Human Anatomy and Embryology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, P.R. China.
Yuqiao
Y
Chang
Kang
K
Guo
Qiong
Q
Li
Cixia
C
Li
Zhikun
Z
Guo
He
H
Li
D050156:Adipogenesis / Q000235:genetics; D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics*; D048430:Cell Shape / Q000235:genetics; D002478:Cells, Cultured; D000070820:Discoidin Domain Receptor 2 / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D007668:Kidney / Q000166:cytology; D008099:Liver / Q000166:cytology; D008168:Lung / Q000166:cytology; D009206:Myocardium / Q000166:cytology; D000071317:Nanog Homeobox Protein / Q000235:genetics / Q000378:metabolism; D010012:Osteogenesis / Q000235:genetics; D019009:Proto-Oncogene Proteins c-kit / Q000235:genetics / Q000378:metabolism; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012867:Skin / Q000166:cytology; D013154:Spleen / Q000166:cytology; D014746:Vimentin / Q000235:genetics / Q000378:metabolism
D002135:Calcium-Binding Proteins; C579666:FSP1 protein, rat; D000071317:Nanog Homeobox Protein; C547030:Nanog protein, rat; D014746:Vimentin; C000603244:Ddr2 protein, rat; D000070820:Discoidin Domain Receptor 2; D019009:Proto-Oncogene Proteins c-kit