Transcriptome Analysis of TGFBI Knockdown vs Normal Corneal Epithelial Cells: Implications for TGFBI Corneal Dystrophy Treatment.

Abstract

This study aimed to clarify the role Transforming Growth Factor Beta Induced (TGFBI) protein plays in corneal epithelial homeostasis by using RNA interference and to explore the possibility of gene therapy as a treatment modality for the visually debilitating TGFBI Corneal Dystrophies (CDs). TGFBI knockdown (KD) in Human Corneal Epithelial Cells (HCECs) was performed by using shRNA lentiviral vectors. RNA sequencing and comprehensive transcriptome analysis were performed to investigate the differential expression between control HCECs and TGFBI KD HCECs. Over Representation Analysis of the differentially expressed (DE) genes delineated the effect inhibition of TGFBI would have on molecular pathways, corneal structure and function. An effective KD of 70.5% was achieved. The functions of downregulated genes in TGFBI KD HCECs indicate decreased inflammation (MTPN, IL1B, IL6, JAK2), decreased angiogenesis (CD24, IL6, JAK2), and decreased corneal scarring (AREG, ITGA11). The functions of upregulated genes indicate increased ECM remodeling, fibrosis, and neovascularisation (MMP2, AKT1, COL6A1, COL6A2), increased integrin signaling (ICAM, ITGA6), and increased cell proliferation (AKT1, ITGA6). Enriched associations of the DE genes included cell adhesion molecules, ECM structural constituents, RNA transport & metabolism, SMAD2/SMAD3:SMAD4 modulation, JAK-STAT and PI3K-Akt signaling pathways. This proof-of-concept study shows that it is possible to effectively silence TGFBI with shRNA in HCECs and provides valuable insight into how TGFBI dysfunction might impact corneal epithelial function. In view of the lack of targeted treatment available, the therapeutic potential of shRNA targeting TGFBI should be explored further since it can potentially revolutionize the future management of TGFBI CDs.