Purpose

To evaluate the role of endogenously produced laminin and fibronectin as well as the effect of exogenous laminin and fibronectin in the attachment of human corneal epithelial cells in vitro.

Methods

Primary cultured human corneal epithelial cells labeled with 3H-thymidine were seeded onto plates coated with laminin or fibronectin, or onto uncoated bacteriologic plates. Attachment of cells was measured in the presence or absence of antisera against laminin or fibronectin, by counting radioactivity.

Results

Human corneal epithelial cells attached to plates coated with human laminin or human fibronectin in a dose-dependent manner, with 69% and 50% of cells attached to the wells coated with 40 micrograms/ml of laminin and fibronectin, respectively (P < 0.001). The percentage of attachment to uncoated bacteriologic plates increased from 1.2% at 45 min of incubation to 6.7% at 90 min, 22.2% at 3 hr, and 40.1% at 6 hr of incubation. Cycloheximide, a protein synthesis inhibitor, completely inhibited cell attachment. Rabbit antiserum against human fibronectin reduced cell attachment to the uncoated plates to 67% of the control value (P < 0.01), whereas rabbit antiserum against human laminin decreased the attachment to 52% of the control (P < 0.01). A combination of these two antisera reduced cell attachment to 46% of the control (P < 0.01).

Conclusions

Endogenous laminin and fibronectin as well as exogenous laminin and fibronectin play significant roles in the attachment of human corneal epithelial cells in culture.