Divergent Metabolomic Signatures of TGF2 and TNF in the Induction of Retinal Epithelial-Mesenchymal Transition

Epithelial-mesenchymal transition (EMT) is a dedifferentiation program in which polarized, differentiated epithelial cells lose their cell-cell adhesions and transform into matrix-producing mesenchymal cells. EMT of retinal pigment epithelial (RPE) cells plays a crucial role in many retinal diseases, including age-related macular degeneration, proliferative vitreoretinopathy, and diabetic retinopathy. This dynamic process requires complex metabolic reprogramming to accommodate the demands of this dramatic cellular transformation. Both transforming growth factor-beta 2 (TGF beta 2) and tumor necrosis factor-alpha (TNF alpha) have the capacity to induce EMT in RPE cells; however, little is known about their impact on the RPE metabolome. Untargeted metabolomics using high-resolution mass spectrometry was performed to reveal the metabolomic signatures of cellular and secreted metabolites of primary human fetal RPE cells treated with either TGF beta 2 or TNF alpha for 5 days. A total of 638 metabolites were detected in both samples; 188 were annotated as primary metabolites. Metabolomics profiling showed distinct metabolomic signatures associated with TGF beta 2 and TNF alpha treatment. Enrichment pathway network analysis revealed alterations in the pentose phosphate pathway, galactose metabolism, nucleotide and pyrimidine metabolism, purine metabolism, and arginine and proline metabolism in TNF alpha-treated cells compared to untreated control cells, whereas TGF beta 2 treatment induced perturbations in fatty acid biosynthesis metabolism, the linoleic acid pathway, and the Notch signaling pathway. These results provide a broad metabolic understanding of the bioenergetic rewiring processes governing TGF beta 2- and TNF alpha-dependent induction of EMT. Elucidating the contributions of TGF beta 2 and TNF alpha and their mechanistic differences in promoting EMT of RPE will enable the identification of novel biomarkers for diagnosis, management, and tailored drug development for retinal fibrotic diseases.