Abstract
Purpose: :
To investigate whether matrix metalloprotease–2 (MMP–2) and its reported stimulants, concanavalin A (ConA) and 17ß–estradiol (E2) are involved in the migratory potential of retinal progenitor cells in ex vivo transplantation studies.
Methods: :
Retinal explant cultures were performed on a Millicell–CM chamber filter in six–well culture plate, each containing 1–ml culture medium with the following additives when indicated: active MMP–2, 20 µg/ml Con A, 20 µg/ml Con A and MMP–2 inhibitor, 10–10 M E2, 10–10 M E2 and MMP–2 inhibitor, 20 µg/ml Con A and 10–10 M E2, 20 µg/ml Con A, 10–10 M E2, and MMP–2 inhibitor, 20 µg/ml MMP–2 inhibitor, or no additional substances. Retinal cells from P0–P2 green fluorescent protein (GFP) transgenic mice were dissociated with the Papain Dissociation System and 2 microliters of cell suspension (1.0×105 cells/µl) were placed between the host retina and chamber filter (subretinal space). Explants were the cultured at 34°C in 5% CO2, with a change of media every other day; samples were harvested and processed for histology approximately one week later.
Results: :
In the ex vivo MMP–2 group, 18.43 ± 1.60% of the total GFP+ cells migrated into host retina, a significantly greater proportion than observed in the ex vivo control group (2.46 ± 0.54%, p < 0.01, Student's t–test). The application of Con A, E2, or both increased the percentages of migrating cells (15.68 ± 2.86%, 12.88 ± 2.29%, and 17.72 ± 2.43%, respectively). These increases were statistically significant (p < 0.05, ex vivo Con A group versus ex vivo control group; p < 0.01, ex vivo E2 group versus ex vivo control group; p < 0.05, ex vivo Con A/ E2 group versus ex vivo control group). Although there was no additive effect on the percentage of the migrated cells when the Con A and E2 treatments were given together, the cells increasingly migrated into the inner retina following combined treatment. The ratio of migrated cells in each condition decreased (7.54 ± 0.55%, 5.82 ± 1.06%, and 11.71 ±1.66% for the ex vivo Con A/MMP–2 inhibitor, ex vivo E2/MMP–2 inhibitor, and ex vivo Con A/E2/ MMP–2 inhibitor groups, respectively).
Conclusions: :
These results suggest that Con A and E2 might facilitate grafted cell migration by activating MMP–2 in situ, providing a clue to the successful outcome of transplantation therapy in retinal degenerative diseases.
Keywords: transplantation