Abstract
Purpose :
TGF-β2 is the predominant TGF-β isoform within the eye and is present in relatively high amounts. TGF-β2 must be under a tight control of a network of different growth factors and matricellular proteins. A disbalance of growth factor was observed in Primary Open Angle Glaucoma (POAG) as TGF-β2 is significantly elevated in the aqueous humor and antagonist like BMP-4 and 7 are reduced. CCN2/CTGF, a downstream mediator of TGF-β2, might play a critical role in the homeostatic balance, so we investigated whether CCN2/CTGF can alter BMP and TGF-β signaling pathways in the outflow tissues.
Methods :
We analyzed the direct effect of CCN2/CTGF on BMP signaling pathways (pSmad 1/5/8) and TGF-β signaling pathways (pSmad 2/3) in a transgenic mouse model with a moderate CCN2/CTGF overexpression in vivo and in immortalized human trabecular meshwork (HTM-N) cells in vitro by immunoblotting, immunohistochemistry and real-time RT-PCR. We investigated whether CCN2/CTGF mediates TGF-β effects via the Rho/RACK and/or ERK signaling pathway. The effect of high CCN2/CTGF expression on the development of the mouse eye was analyzed by immunohistochemistry and scanning electron microscopy. Statistical analysis of data was performed by two-tailed t-test or one-way ANOVA.
Results :
We observed developmental malformations in the ciliary body in the transgenic mice with a high CCN2/CTGF expression accompanied by a marked reduction of BMP signaling pathway. In a mouse model with moderate CCN2/CTGF overexpression, we could detect a dysregulation of the BMP and TGF-β signaling pathways in the 2-month-old mice, with a reduction in BMP activity (pSmad 1/5/8 0.50 ± 0.38, p≤0.05) and an increase in TGF-β signaling (pSmad2 1.85±0.67, p≤0.05; pSmad3 1.86 ± 0.57, p≤0.05) in vivo. CCN2/CTGF (50ng/ml) treatment reduced BMP expression (BMP-4: 0.58 ± 0.5, p≤0.05; BMP-7: 0.5 ± 0.21, p≤0.05) and enhanced TGF-β expression in HTM-N cells in vitro (Tgf-b1: 1.8 ± 0.84, p≤0.05; Tgf-b2: 1.84 ± 0.68, p≤0.05). Inhibition of the Rho/RACK and ERK signaling pathway attenuated the CCN2/CTGF effect on TGF-β in HTM-N cells.
Conclusions :
We conclude that CCN2/CTGF functions as a modulator of the homeostatic balance of BMP and TGF-β signaling pathway, which is shifted in POAG.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.