PURPOSE: To determine the dose response of human recombinant basic fibroblast growth factor (bFGF) on mitogenic activity, and the supplementary role of serum in cultured bovine and human corneal endothelial cells (BCECs, HCECs). To investigate the effect of bFGF on endothelial wound healing of human corneas in vitro. METHODS: In cell culture, DNA synthesis was assessed by 3H-thymidine incorporation. Wound healing was studied using paired human corneas after mechanical damaging of the endothelium. One cornea was treated with bFGF, and the mate served as control. Wound closure was determined after staining with trypan blue. Endothelial cell density (ECD) was assessed in the closed wound area after alizarin red staining. DNA synthesis was assessed using 3H-thymidine autoradiography. RESULTS: In cell culture, bFGF induced a dose-dependent mitogenic response on BCECs and HCECs. Addition of serum to the culture medium shifted the dose-response curve to considerably lower bFGF concentrations. In organ culture, the time of complete wound closure shortened only marginally (0.5 day) after bFGF treatment (P < 0.01). In the closed wound center, ECD was significantly higher in 1 ng/ml bFGF-treated corneas (686 +/- 134 cells/mm2) than in controls (554 +/- 117 cells/mm2), an increase of +25%. Doses of 0.1 and 10 ng/ml also were effective, but less so than with 1 ng/ml (+11% and +15%, respectively), whereas a dose of 100 ng/ml even had a negative effect (-11%). DNA synthesis was marginally enhanced in bFGF-treated (1 ng/ml) corneas. CONCLUSIONS: The maximal effective dose of bFGF producing a BCEC mitogenic response is dependent on serum. In human senior donor corneas, bFGF promotes endothelial wound healing predominantly by stimulation of cell migration.
PURPOSE: To determine the dose response of human recombinant basic fibroblast growth factor (bFGF) on mitogenic activity, and the supplementary role of serum in cultured bovine and human corneal endothelial cells (BCECs, HCECs). To investigate the effect of bFGF on endothelial wound healing of human corneas in vitro. METHODS: In cell culture, DNA synthesis was assessed by 3H-thymidine incorporation. Wound healing was studied using paired human corneas after mechanical damaging of the endothelium. One cornea was treated with bFGF, and the mate served as control. Wound closure was determined after staining with trypan blue. Endothelial cell density (ECD) was assessed in the closed wound area after alizarin red staining. DNA synthesis was assessed using 3H-thymidine autoradiography. RESULTS: In cell culture, bFGF induced a dose-dependent mitogenic response on BCECs and HCECs. Addition of serum to the culture medium shifted the dose-response curve to considerably lower bFGF concentrations. In organ culture, the time of complete wound closure shortened only marginally (0.5 day) after bFGF treatment (P < 0.01). In the closed wound center, ECD was significantly higher in 1 ng/ml bFGF-treated corneas (686 +/- 134 cells/mm2) than in controls (554 +/- 117 cells/mm2), an increase of +25%. Doses of 0.1 and 10 ng/ml also were effective, but less so than with 1 ng/ml (+11% and +15%, respectively), whereas a dose of 100 ng/ml even had a negative effect (-11%). DNA synthesis was marginally enhanced in bFGF-treated (1 ng/ml) corneas. CONCLUSIONS: The maximal effective dose of bFGF producing a BCEC mitogenic response is dependent on serum. In human senior donor corneas, bFGF promotes endothelial wound healing predominantly by stimulation of cell migration.
Authors: Peter W Rieck; Ruth M von Stockhausen; Sylvia Metzner; Christian Hartmann; Yves Courtois Journal: Graefes Arch Clin Exp Ophthalmol Date: 2003-09-06 Impact factor: 3.117