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Does Metalloproteinases Play an Important Role in the Wound Healing Process of the Conjunctiva following Filtering Procedures for Glaucoma?
1Welsandt G., 2Hueber A., 1Kociok N., 2Esser J., 1Joussen A. M., 2Krieglstein G. K., 2Mietz H.,
1Universität zu Köln, Zentrum für Augenheilkunde, Abteilung für Netzhaut- und Glaskörperchirurgie (Köln)
2Universität zu Köln, Zentrum für Augenheilkunde (Köln)
Purpose: Latanoprost does not change the composition of the extracellular matrix of the conjunctiva as compared to e.g. betablockers. Latanoprost causes an up-regulation of metalloproteinases (MMP) and their inhibitors (TIMP). We aimed to detect different iso-types of MMPs and TIMPs in Tenon´s fibroblasts and their activity influenced by latanoprost performing in vitro and in vivo studies.
Method: For detection of MMPs and TIMPs immunocytochemistry was perforemd. for specific RNA analysis, RT-PCR (polymerase chain reaction) was done. In addition, western blot analysis and flow cytometry were carried out using latanoprost treated cells. To evaluate the presence of an activated form of MMP, the zymography technique was performed. Real time RT-PCR was used to determine different types of MMP and TIMP. After treatment with latanoprost for 1 and 4 weeks, MMP and TIMP were detected in rabbit conjunctiva by immunohistochemistry.
Results: The presence of the proteins MMP-3 and TIMP-2 in Tenon fibroblasts was confirmed by immunohistochemistry. RT-PCR showed the presence of MMP-3 and TIMP-2 at the RNA-level. Using the western blot analysis, similar results were obtained. Flow cytometry showed an up-regulation of MMP-3 and TIMP-2 in latanoprost-treated cells (from 0.97 in control up to 1.20 after latanoprost-treatment for TIMP-2 and from 1.04 in control up to 1.24 following latanoprost-treatment for MMP-3). With the zymography technique, an increase of the activated protein of MMP-3 was detected following latanoprost-treatment. With the Real time RT-PCR, the presence of MMPs 1, 2, 3, 7, 9, and 14 as well TIMPs 1 and 2 were confirmed in Tenon´s fibroblasts. The animal experiment showed an increase of immunreaction for MMP-3 and TIMP-2 after latanoprost-treatment.
Conclusions: Many different types of MMPs and TIMPs could be detected, all of which are involved with the turn-over of extracellular matrix components. An increase of MMP-3 activity was shown after treatment with latanoprost. It may be reasonable to speculate, that MMPs and TIMPs play an important role in morphologic alterations of the conjunctiva following topical therapy for glaucoma.