[Experimental study of repairing damaged human amniotic epithelial cells with formulated fibrin clot and cell growth factor].
OBJECTIVE
To investigate whether damaged human amniotic epithelial cells (HAEC) could be repaired on the matrix of formulated fibrin clot in vitro and the effects of epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and transforming growth factor beta(1) (TGF-beta(1)) on the proliferation of HAEC.


METHODS
Ring drill was used to drill the HAEC layer on culture sheets to make quantified models of damaged HAEC, on which the lacks were then covered with fibrin clot. Subsequently, EGF (EGF group), bFGF (bFGF group) and TGF-beta(1) (TGF-beta(1) group) of different concentration were added into the sheets respectively. After the predesigned culturing time, the growing and transiting conditions of HAEC were observed under inverted microscope after Giemsa stain. Also, the proliferating conditions of HAEC were detected by using 5-bromodeoxyuridine (BrdU).


RESULTS
In all groups, HAEC could transit toward damaged area on fibrin clot and grow there. Higher transiting speed and larger cell numbers were observed in the EGF and bFGF groups followed by the control group, while the TGF-beta(1) group showed the relatively poorer results. Proliferating rates of HAEC were 17.8%, 28.0%, 35.3%, 51.6%, 34.1%, 34.2% and 26.0% respectively by EGF of different cultured concentration (1.0 ng/ml, 5.0 ng/ml, 10.0 ng/ml, 20.0 ng/ml, 40.0 ng/ml, 80.0 ng/ml and 160.0 ng/ml). Proliferating rates of HAEC were 18.0%, 35.7%, 43.0%, 52.7%, 67.4%, 43.6% and 30.5% respectively by bFGF of different cultured concentration (1.0 ng/ml, 5.0 ng/ml, 10.0 ng/ml, 20.0 ng/ml, 40.0 ng/ml, 80.0 ng/ml and 160.0 ng/ml). Compared with the control group, EGF groups (EGF concentration ranging from 10 ng/ml to 80 ng/ml) and bFGF groups (bFGF concentration ranging from 5 ng/ml to 80 ng/ml) showed better proliferating effects of HAEC (P < 0.05), especially the 20 ng/ml EGF group and 40 ng/ml bFGF group had the best proliferating results among their own respective groups (P < 0.05). Proliferating rates of HAEC were 17.1%, 15.1%, 9.3%, 6.2%, 4.8%, 3.6%, 2.0% and 1.2% respectively by TGF-beta(1) of different cultured concentration (0.1 ng/ml, 0.2 ng/ml, 0.4 ng/ml, 0.8 ng/ml, 1.6 ng/ml, 3.2 ng/ml, 6.4 ng/ml and 12.8 ng/ml). Proliferating rates of HAEC in TGF-beta(1) groups (TGF-beta(1) concentration ranging from 0.8 ng/ml to 12.8 ng/ml) were significantly lower than that in the control group (P < 0.05).


CONCLUSIONS
HAEC could transit and grow on the matrix of fibrin clot and repair the damaged area. EGF and bFGF could obviously stimulate HAEC proliferation, while TGF-beta(1) might have the inhibitive effects.
Qi
Hong-bo
HB
Department of Obstetrics and Gynecology, The First Affiliated Hospital of Chongqing University of Medical Sciences, Chongqing 400016, China.
Li
Wei
W
Bian
Du-hong
DH
D000650:Amnion / Q000473:pathology*; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D016207:Cytokines / Q000008:administration & dosage / Q000494:pharmacology*; D004815:Epidermal Growth Factor / Q000008:administration & dosage / Q000494:pharmacology; D004847:Epithelial Cells / Q000473:pathology* / Q000648:ultrastructure; D005260:Female; D005322:Premature Rupture of Fetal Membranes / Q000473:pathology / Q000628:therapy*; D005337:Fibrin; D016222:Fibroblast Growth Factor 2 / Q000008:administration & dosage / Q000494:pharmacology; D006801:Humans; D011247:Pregnancy; D016212:Transforming Growth Factor beta / Q000008:administration & dosage / Q000494:pharmacology
D016207:Cytokines; D016212:Transforming Growth Factor beta; D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor; D005337:Fibrin