Abstract
Purpose:
Fibronectin is an extracellular matrix (ECM) protein produced by the lens throughout its development. Fibronectin deposition in ECM is required for lens placode thickening and invagination. Further fibronectin expression increases during progression of posterior capsule opacification and is a marker of lens epithelium-epithelial to mesenchymal transition (EMT). Fibrotic PCO development also involves activation of TGFβ signaling. TGFβ is secreted in a latent form bound to latent TGFβ binding proteins (LTBP). TGFβ/LTBP complexes interact with ECM to form an extracellular inactive TGFβ depot. LTBP1, the most abundant LTBP in the lens requires cell associated fibronectin to tether latent TGFβ to the ECM and participate in interactions necessary for TGFβ activation. Thus we hypothesize that fibronectin is crucial for TGFβ activation critical for development of fibrotic PCO.
Methods:
Mice were created homozygous for fibronectin floxed allele and MLR10-Cre transgene (cFN). MLR10-Cre is expressed in all lens cells starting at lens vesicle stage. Loss of fibronectin expression was determined by PCR and immunostaining. Histology and scanning electron microscopy were used to analyze lens morphology. To determine role of fibronectin in regulation of fibrotic PCO, cFN mice were subjected to surgery modeling human cataract surgery. Capsular bags were then analyzed at 12 hrs, 48 hrs and 5 days post surgery for PCO development by analyzing multiple EMT and fiber differentiation markers.
Results:
cFN lenses are transparent, with no morphological abnormalities when compared to control lenses. Analysis of cFN capsular bags 5 days post surgery shows a down regulation of markers of EMT such as αSMA, however no differences were observed in levels of fiber cell markers such as cMaf. Analysis of TGFβ signaling 5 days post surgery in cFN capsular bags shows a down regulation of pSmad3 as compared to controls. Levels of LTBP1 were found to be up regulated in cFN lenses as compared to controls, further cFN capsular bags showed a down regulation of LTBP1 as compared to controls 5 days post surgery.
Conclusions:
Our data suggests that loss of fibronectin after lens vesicle formation does not affect lens development and morphology. However fibronectin appears to be crucial for TGFβ activation critical for the development of fibrotic PCO. Overall these data demonstrate a key role of fibronectin in EMT during PCO development but not lens fiber differentiation.
Keywords: 512 EMT (epithelial mesenchymal transition) •
519 extracellular matrix •
445 cataract