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Abstract #73871 Published in IGR 19-1
Overexpression of ALK5 Induces Human Tenon's Capsule Fibroblasts Transdifferentiation and Fibrosis In Vitro
Fu SH; Wang HY; Zhang JY; Shi HM; Zhang N; Ye W; Xiao YQCurrent Eye Research 2017; 42: 1018-1028
PURPOSE: To investigate the involvement of activin receptor-like kinase 5 (ALK5) in human Tenon's capsule fibroblasts (HTFs) transdifferentiation and fibrosis. METHODS: (1) Cultured HTFs were treated with transforming growth factor beta 1 (TGF-β1) at different concentrations for different durations, mRNA expression of ALK5 and plasminogen activator inhibitor-1 (PAI-1) was measured by quantitative polymerase chain reaction (PCR) while protein expression of ALK5, α-smooth muscle actin (α-SMA), and extracellular matrix deposition including fibronectin (FN) and collagen I (Col1) was assessed by western blot. HTFs with or without TGF-β1 were also treated with an ALK5 activity inhibitor, SB-431542, and fibrosis-related genes were assessed. (2) HTFs were transduced with ALK5 lentivirus (ALK5-OE group) or empty lentivirus (NC-OE) with or without the treatment of SB-431542. Protein expression of ALK5, α-SMA, FN, and Col1 was evaluated. (3) HTFs in the ALK5-OE group and NC-OE group were subjected to a scratch-wound assay and their migratory activities assessed. RESULTS: (1) TGF-β1, in a concentration-dependent manner, upregulated ALK5 and PAI-1 expressions in the HTFs, which peaked between 24 and 36 h. These changes were associated with increases in protein levels of FN, Col1, and α-SMA. These TGF-β1 effects were blocked by the ALK5 inhibitor SB-431542. (2) Similarly, overexpression of ALK5 by lentiviral vector significantly increased protein expression of α-SMA, FN, and Col1. Addition of TGF-β1 to the ALK5-OE cells did not produce additional expression of any of the marker proteins. The upregulation of extracellular matrix and α-SMA can be reduced by SB-431542. (3) In ALK5-OE group, HTFs migration was significantly increased compared with normal control and TGF-β1 could still promote ALK5-OE cells migration. CONCLUSIONS: Our findings suggest that ALK5 is an important mediator of HTFs fibrosis. ALK5 is a potential therapeutic target to suppress scar formation after filtration surgery.
a Department of Ophthalmology , Huashan Hospital, Fudan University , Shanghai , China.
Full articleClassification:
3.6 Cellular biology (Part of: 3 Laboratory methods)
3.5 Molecular biology incl. SiRNA (Part of: 3 Laboratory methods)
2.3 Sclera (Part of: 2 Anatomical structures in glaucoma)
12.8.10 Woundhealing antifibrosis (Part of: 12 Surgical treatment > 12.8 Filtering surgery)
