Suppression of corneal neovascularization by culture supernatant of human amniotic cells.
OBJECTIVE
To examine the applicability of culture supernatant of human amniotic cells on basic fibroblast growth factor (bFGF)-induced corneal neovascularization.


METHODS
Human amniotic epithelial and mesenchymal cells (AC) were obtained from human amniotic membranes by digesting with collagenase and maintained in serum-containing medium. The AC preparations predominantly contained cytokeratin-positive cells (91.2 +/- 3.1%, n = 4). The culture supernatant was prepared by cultivating AC in serum-free medium for 24 hours. Neovascularization was obtained by a micropocket assay with Hydron pellets containing bFGF. Migration assay was carried out by a double-chamber method using human umbilical vein endothelial cells (HUVEC). Cell growth assay was done by MTT assay using HUVEC.


RESULTS
Basic fibroblast growth factor-induced corneal neovascularization was significantly reduced by administration of AC culture supernatant. When either control or AC culture supernatant was administered three times per day for 10 days, the area with neovascularization was 13.2 +/- 3.2 mm2 and 4.0 +/- 1.4 mm2 for the control and AC culture supernatant-treated eyes, respectively (n = 7, p = 0.021). AC culture supernatant strongly inhibited bFGF-induced migration and cell growth of HUVEC.


CONCLUSIONS
Amniotic cell culture supernatant contains potent inhibitors of neovascularization. This effect is explained in part by inhibition of migration and cell growth of vascular endothelial cells. AC culture supernatant may be applicable for treatment of corneal diseases with neovascularization.
Kobayashi
Namie
N
Department of Biomolecular Science, Fukushima Medical University School of Medicine, Japan.
Kabuyama
Yukihito
Y
Sasaki
Shinji
S
Kato
Kei-ichiro
K
Homma
Yoshimi
Y
D000650:Amnion / Q000166:cytology*; D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D018929:Cell Culture Techniques; D002455:Cell Division / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D003315:Cornea / Q000187:drug effects* / Q000473:pathology; D016510:Corneal Neovascularization / Q000139:chemically induced / Q000473:pathology / Q000517:prevention & control*; D003470:Culture Media / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000633:toxicity; D006801:Humans; D011817:Rabbits
D020533:Angiogenesis Inhibitors; D003470:Culture Media; D016222:Fibroblast Growth Factor 2