FGF-2 Differentially Regulates Lens Epithelial Cell Behaviour during TGF-β-Induced EMT

Abstract

Fibroblast growth factor (FGF) and transforming growth factor-beta (TGF-β) can regulate and/or dysregulate lens epithelial cell (LEC) behaviour, including proliferation, fibre differentiation, and epithelial–mesenchymal transition (EMT). Earlier studies have investigated the crosstalk between FGF and TGF-β in dictating lens cell fate, that appears to be dose dependent. Here, we tested the hypothesis that a fibre-differentiating dose of FGF differentially regulates the behaviour of lens epithelial cells undergoing TGF-β-induced EMT. Postnatal 21-day-old rat lens epithelial explants were treated with a fibre-differentiating dose of FGF-2 (200 ng/mL) and/or TGF-β2 (50 pg/mL) over a 7-day culture period. We compared central LECs (CLECs) and peripheral LECs (PLECs) using immunolabelling for changes in markers for EMT (α-SMA), lens fibre differentiation (β-crystallin), epithelial cell adhesion (β-catenin), and the cytoskeleton (alpha-tropomyosin), as well as Smad2/3- and MAPK/ERK1/2-signalling. Lens epithelial explants cotreated with FGF-2 and TGF-β2 exhibited a differential response, with CLECs undergoing EMT while PLECs favoured more of a lens fibre differentiation response, compared to the TGF-β-only-treated explants where all cells in the explants underwent EMT. The CLECs cotreated with FGF and TGF-β immunolabelled for α-SMA, with minimal β-crystallin, whereas the PLECs demonstrated strong β-crystallin reactivity and little α-SMA. Interestingly, compared to the TGF-β-only-treated explants, α-SMA was significantly decreased in the CLECs cotreated with FGF/TGF-β. Smad-dependent and independent signalling was increased in the FGF-2/TGF-β2 co-treated CLECs, that had a heightened number of cells with nuclear localisation of Smad2/3 compared to the PLECs, that in contrast had more pronounced ERK1/2-signalling over Smad2/3 activation. The current study has confirmed that FGF-2 is influential in differentially regulating the behaviour of LECs during TGF-β-induced EMT, leading to a heterogenous cell population, typical of that observed in the development of post-surgical, posterior capsular opacification (PCO). This highlights the cooperative relationship between FGF and TGF-β leading to lens pathology, providing a different perspective when considering preventative measures for controlling PCO.

Biomarker Signature in Aqueous Humor Mirrors Lens Epithelial Cell Activation: New Biomolecular Aspects from Cataractogenic Myopia

“…In vitro studies on cultured LECs might elucidate these aspects using pathological LECs and neutralizing factors (mediators) or control LECs and pathological AH (differentiation studies). Regarding the appearance of the EMT phenotype, the participation of other mediators such as FGF, EGF, NGF, and BDNF, which are known to be increased in AH and able to work as modulators of cell differentiation/contractile activity, cannot be excluded [ 41 , 42 , 43 , 44 , 45 , 46 , 47 ]. In previous studies, these growth factors have been reported to be profibrogenic and anti-inflammatory [ 46 , 47 ].…”

“…Regarding the appearance of the EMT phenotype, the participation of other mediators such as FGF, EGF, NGF, and BDNF, which are known to be increased in AH and able to work as modulators of cell differentiation/contractile activity, cannot be excluded [ 41 , 42 , 43 , 44 , 45 , 46 , 47 ]. In previous studies, these growth factors have been reported to be profibrogenic and anti-inflammatory [ 46 , 47 ]. In particular, NGF has been reported to trigger the EMT phenotype and TGFβ and FGF, but the presence of two specific receptors (trkA NGFR and p75 NTR ) allows NGF to mediate a selective response in EMT cells from survival to apoptosis, suggesting additional homeostatic considerations [ 46 , 47 ].…”

“…In previous studies, these growth factors have been reported to be profibrogenic and anti-inflammatory [ 46 , 47 ]. In particular, NGF has been reported to trigger the EMT phenotype and TGFβ and FGF, but the presence of two specific receptors (trkA NGFR and p75 NTR ) allows NGF to mediate a selective response in EMT cells from survival to apoptosis, suggesting additional homeostatic considerations [ 46 , 47 ].…”

Deciphering the fibrotic process: mechanism of chronic radiation skin injury fibrosis

“…The mammalian target of rapamycin (mTOR) pathway plays a crucial role in this process, enhancing protein synthesis and myofibroblast activation ( 40 ). Further to its direct effects on fibroblasts, TGF-β can induce EMT, contributing to the pool of myofibroblasts secreting fibrotic material ( 41 ).…”

Myo/Nog Cells: The Jekylls and Hydes of the Lens

“…Expression of α-SMA was detected in LECs lacking BAI1 in explants depleted of Myo/Nog cells and treated with transforming growth factor beta 1 (TGF-β1) [ 62 ] ( Figure 5 ). TGF-β2, a known inducer of epithelial to mesenchymal transition and expression of α-SMA in the lens [ 58 , 77 , 78 , 79 , 80 , 81 , 82 , 83 , 84 , 85 , 86 , 87 , 88 ], was added to explant cultures in an effort to stimulate the migration and transdifferentiation of LECS to myofibroblasts in the absence of Myo/Nog cells. TGF-β2, but not TGF-β1, resulted in the loss of the majority of LECs within 48 h [ 62 ] ( Figure 5 H).…”