HGF/Met Axis Protects Human Retinal Pigment Epithelial Cells from Cell Death

Abstract

Demise of the retinal pigment epithelium (RPE) caused by etiologies such as age‐related macular degeneration is a major underlying factor leading to vision loss. Cell death can occur through two major pathways: apoptosis and necrosis/necroptosis. In the former, Fas/FasL death receptor signaling induces apoptosis, but its role in human RPE cell death is not thoroughly elucidated. In the latter, reactive oxygen species (ROS) can induce necrosis/necroptosis. Additionally, Hepatocyte Growth Factor (HGF) is a potent growth and survival factor for epithelial cells, but its functional role in RPE is not well characterized, especially its ability to protect RPE cells from death and to induce their growth. Accordingly, we hypothesized that HGF/Met protects human RPE cells from Fas‐promoted apoptosis and H2O2‐induced necrosis/necroptosis. We analyzed whether the HGF receptor (Met) is expressed in human and non‐human primate (NHP) eye tissue via western blot analysis and immunohistochemistry. To determine whether HGF/Met protects RPE cells against death induced by Fas/FasL and H2O2, we utilized gain‐ and loss‐of‐function models, cell survival assays and gene expression analysis (RNAseq). Human ARPE‐19 cell line was treated with various combinations of HGF, Met kinase inhibitor SU11274 (SU), FasL or H2O2. Cell growth and survival was determined via MTT assay. Met, Akt, and Erk activation were analyzed by western blot. Western blot analysis and immunohistochemistry confirmed Met activity and signaling in human and NHP eye tissue, including in the RPE cell layer. We found that HGF elicited Met stimulation and signaling culminating in activation of its downstream effectors Akt and Erk, proteins involved in two major pathways controlling cell survival and proliferation. We also uncovered that HGF inhibited FasL‐ and ROS‐induced killing of RPE. Conversely, inhibiting Met signaling rendered RPE susceptible to cell death induced by FasL and H2O2. Global gene expression analysis using an RNAseq approach revealed that HGF controls expression of numerous genes governing RPE cell growth, metabolism, and survival to name a few. In particular, HGF significantly upregulated 400 and repressed expression of 420 genes six hours post‐treatment; at 24 hours post‐treatment, HGF upregulated 291 and downregulated expression of 260 genes. Expression of 11,000 genes was unchanged. Our studies establish that HGF and Met play a major role in promoting the survival and homeostasis of RPE cells by inhibiting two major pathways of RPE cell death. Our findings suggest HGF/Met is a potential druggable target for therapeutic management of RPE‐related human ocular diseases.