Abstract
Purpose: To determine the activity of metformin in an in vitro model of posterior capsule opacification (PCO).Design: Experimental laboratory research. Methods: The HLE-B3 lens epithelial cell line was treated with PCO induction media (PCOM) supplemented with transforming growth factor-beta (TGF-β) and fibroblast growth factor (FGF). Different metformin concentrations (0-100 mM) were used. The following cellular parameters were assessed: (1) survival, using a viability assay; (2) morphology, via microscopy and image analysis; (3) migration, using the wound assay; (4) and expression of epithelial (Pax6, E-cadherin) and mesenchymal (α-smooth muscle actin or α-SMA, fibronectin) markers via Western blot. Expression of the uptake receptor SLC22A1 was evaluated in HLE-B3 and in human donor eyes with Western blot and immunohistochemistry, respectively. Statistical analysis of variance (ANOVA) with Tukey post-hoc test was done for analysis of cytotoxicity, morphology and migration data. Results: Metformin was lethal to half (LC50) of the cells at 30 mM, and a decrease in viability (P<0.05) was noted at 5 mM. LECs in PCOM treated with 1 mM metformin showed increased Pax6 and E-cadherin and decreased α-SMA and fibronectin expression. LECs in PCOM treated with metformin also maintained epithelial morphology. Migration was inhibited with 0.5 mM metformin (P<0.05). Both HLE-B3 and the lens epithelium in donor eyes were found to express SLC22A1.Conclusion: Metformin decreased survival and migration in LECs, maintaining epithelial phenotype and reducing mesenchymal marker expression. Metformin therefore has potential as an adjunct in PCO prevention.Financial Disclosures: This work was partially funded by Mitacs Canada.
Highlights
Posterior capsule opacification (PCO) causes a recurrence in visual decline in 20-40% of patients 2-5 years after cataract surgery.[1]
Cells were starved in serum-free (SF) media for 4 hours before addition of any treatment. This in vitro model of PCO utilized the addition of transforming growth factor-beta (TGF-β) and fibroblast growth factor (FGF) to the culture media to induce proliferation and epithelial-to-mesenchymal transition (EMT)
Increasing concentrations of metformin were cytotoxic to lens epithelial cells (LECs) (Figure 1), whether grown in SF or in PCO media
Summary
Posterior capsule opacification (PCO) causes a recurrence in visual decline in 20-40% of patients 2-5 years after cataract surgery.[1] PCO results from the proliferation, migration and secretion of extracellular matrix proteins by residual lens epithelial cells (LECs).[2] This collective response is called epithelial-to-mesenchymal transition (EMT).[3]. EMT is a key process in cancer proliferation and metastasis.[4] The activity of metformin, a biguanide drug used primarily in the treatment of diabetes, has been observed against different cancers in vitro and in vivo. It requires an organic cation transporter for cellular uptake, the main receptor being the solute carrier family 22 member 1 (SLC22A1).[5] Metformin has been noted to inhibit proliferation and EMT6 through a myriad of mechanisms Among these are pathways that are implicated in the development of PCO, such as PI3/Akt (mTOR), MAPK and Wnt.[3]
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