To test the effects of dedicator of cytokinesis protein 1 (DOCK1) with its binding partner engulfment and cell motility protein 1 (ELMO1)-Rac1 axis on the vitreous-induced biological functions of retinal pigment epithelial (RPE) cells.
Methods:
Rac1 activity in RPE cells after vitreous stimulation was detected via a pull-down assay. The related protein expression levels were examined via western blot analysis. DOCK1 and ELMO1 knockdown cells were generated via CRISPR-Cas9 technology. Cytoskeletal reorganization was detected by immunofluorescent localization of F-actin. Cell proliferation, migration, invasion, and contraction ability were measured via the CCK8 assay, wound healing assay, transwell invasion assay, and collagen contraction assay.
Results:
Rac1 activity was significantly elevated in ARPE-19 cells stimulated with vitreous fluid for 30 min to 3 h. Depletion of either DOCK1 or ELMO1 with CRISPR/Cas9 attenuated vitreous-stimulated Rac1 activity, thus reversing the vitreous-induced cytoskeletal rearrangements. The functional cell biology results revealed that deficiencies of DOCK1 and ELMO1 significantly impeded the migration, invasion, and contraction abilities of vitreous-stimulated human RPE cells.
Conclusion:
This study demonstrated that the DOCK1/ELMO1-Rac1 axis plays an essential role in the pathogenesis of proliferative vitreoretinopathy (PVR), thus suggesting that interruption of this axis has potential for PVR therapy.
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