Abstract
Abstract: :
Purpose: To investigate the existence of slit2, an inhibitory factor for chemokine-induced leukocyte chemotaxis, in human corneas. Methods: Donor corneal rims and primary cultured corneal epithelial, stromal and endothelial cells were used for this experiment. Slit2 mRNA was detected by reverse transcriptase-polymerase chain reaction (RT-PCR) in primary cultured cells and further confirmed by digoxigenin-labeled in situ hybridization (DIG-ISH) in corneal tissue. (1) RT-PCR: Total RNAs were harvested from confluent primary cultured corneal epithelial (n=8), stromal (n=2) and endothelial cells (n=15). RT-PCR was performed for slit2. (2) DIG-ISH: The PCR product of slit2 was cloned and sense/anti-sense DIG labeled RNA probes were made. Corneal rims (n=2) were fixed in 4% paraformaldehyde followed by 30% sucrose cryoprotective treatment and frozen sections were made on silylated glass slides. The hybridization was visualized by colorimetric detection system. Results: Slit2 mRNA was found in all corneal epithelial, stromal and endothelial cells of human donor corneas by both RT-PCR and anti-sense DIG-ISH. Conclusion: This is the first report of slit2 in human corneas. This may provide additional clues to understanding the mechanisms underlying the immune privilege status of the eye.
Keywords: 370 cornea: basic science • 437 inflammation • 433 immune tolerance/privilege