Silibinin Inhibits Platelet-Derived Growth Factor-Driven Cell Proliferation via Downregulation of N-Glycosylation in Human Tenon's Fibroblasts in a Proteasome-Dependent Manner.

Yi-Hao Chen,Da-Wen Lu,Ming-Cheng Tai,Ching-Long Chen,Jiann-Torng Chen,Chang-Min Liang,Eve-Isabelle Pecheur

doi:10.1371/journal.pone.0168765

Abstract

The objective of this study was to evaluate the effects of silibinin on cell proliferation in platelet-derived growth factor (PDGF)-treated human Tenon's fibroblasts (HTFs). The effect of silibinin on cell proliferation in PDGF-treated HTFs was determined by examining the expression of proliferating cell nuclear antigen (PCNA) and performing WST-1 assays. Cell cycle progression was evaluated using flow cytometry. The related cyclins and cyclin-dependent kinases (CDKs) were also analyzed using western blot. A modified rat trabeculectomy model was established to evaluate the effect of silibinin on cell proliferation in vivo. Western blot analysis was carried out to determine the effect of silibinin on the expression of PDGF receptor and on the downstream signaling pathways regulated by PDGF receptor. PDGF elevated the expression of PCNA in HTFs, and this elevation was inhibited by silibinin. The inhibitory effect of silibinin on cell proliferation was also confirmed via WST-1 assay. PDGF-stimulated cell cycle in HTFs was delayed by silibinin, and the related cyclin D1 and CDK4 were also suppressed by silibinin. In the rat model of trabeculectomy, silibinin reduced the expression of PCNA at the site of blebs in vivo. The effects of silibinin on PDGF-stimulated HTFs were mediated via the downregulation of PDGF receptor-regulated signaling pathways, such as ERKs and STATs, which may be partially caused by the downregulation of N-glycosylation of PDGF receptor beta (PDGFRβ). The effect of silibinin on modulation of N-glycosylation of PDGFRβ was mediated in a proteasome-dependent manner. Silibinin inhibited cell proliferation and delayed cell cycle progression in PDGF-treated HTFs in vitro. PDGF also modulated the process of N-glycosylation of the PDGFRβ in a proteasome-dependent manner. Our findings suggest that silibinin has potential therapeutic applications in glaucoma filtering surgery.

Highlights

Glaucoma is a major cause of irreversible blindness worldwide and presents as a progressive optic atrophy [1,2,3]
The amount of cyclin D1 and cyclin-dependent kinase 4 (CDK4) significantly decreased in human Tenon’s fibroblasts (HTFs) treated with both silibinin and platelet-derived growth factor (PDGF). These findings showed that silibinin delayed PDGFinduced cell cycle progression in HTFs through modulation of cyclin D1 and CDK4
We further examined whether the ligand binding affinity of PDGF receptor on the surface of HTFs is affected by silibinin by using a fluorescence-based technology

Summary

Introduction

Glaucoma is a major cause of irreversible blindness worldwide and presents as a progressive optic atrophy [1,2,3]. Glaucoma filtering surgery is performed to create an artificial route to drain the aqueous humor from the anterior chamber to the subconjunctival space, leaving a bleb that is formed in the subconjunctival space [5]. The bleb formation is similar to the wound healing process of soft tissues, which involves inflammation, proliferation, and wound remodeling [6, 7]. Scar formation in the bleb, which is a manifestation of extensive wound healing process, is the major cause of failure in this surgery [8]. To attenuate scar formation, the inhibition of inflammation, proliferation, or wound remodeling has been proposed as a strategy. Several agents have been studied for adjunctive use [9, 10], but none has produced satisfactory results

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