Purpose: To determine the influence of serum on transforming growth factor (TGF)-β2 mediated effects on the proliferation of corneal endothelial cells (CE). Methods: Rat CE were grown in explant culture and the proliferation of CE was measured by [3H]thymidine bioassay. Subconfluent cells were synchronized in the GO (quiescent) phase of the cell cycle by serum starvation for 24 hr. Serum and [3H]thymidine were then added to the cells in the presence or absence of a physiological concentration (5 ng ml-1) of exogenous active TGF-β2. Radioactivity was measured at various time points to detect DNA synthesis. These experiments were repeated without adding serum after serum starvation. Preincubation of exogenous TGF-β2 with neutralizing antibody was used to test the cytokine specificity. Results: Without TGF-β2, a linear increase in [3H]thymidine incorporation, indicating S-phase, began approximately 16 hr after serum addition, then plateaued at approximately 24 hr. Serum promoted DNA synthesis of CE in a dose-dependent manner at concentrations of 0.5-10%. In cultures with 10% serum, TGF-β2 (0.5, 1, 5, and 20 ng ml-1) suppressed CE growth dose-dependently. The growth amplitude decreased and the time before S-phase entry, G1 phase, was prolonged to 24 hr. In culture with 1% serum, TGF-β2 (5 ng ml-1) suppressed the CE proliferation by delaying S-phase entry without suppressing growth amplitude. In cultures without serum, TGF-β2 promoted CE growth to a level similar to that of cultures supplemented with 0.5% serum. Conclusions: Responses of cultured CE to exogenous TGF-β2 depended on the concentration of serum in the medium. This result implies a possibility that in vivo serum influx through a compromised bloodocular barrier could influence the CE growth by changing the responses of these cells to TGF-β2 in aqueous humor.
ASJC Scopus subject areas
- Sensory Systems
- Cellular and Molecular Neuroscience