Beta-1 integrin loss leads to partial activation of pro-fibrotic pathways in the lens epithelium

Riggio, Brittany
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University of Delaware
Integrins are heterodimeric adhesion receptors important for the detection of extracellular matrix, adhesion dependent signaling and tissue structure. Previously, the M. Duncan laboratory has shown that loss of ß1-integrin from the E12.5 mouse lens (ß1MLR10) results in cell disorganization of the lens epithelium at E16.5, followed by its apoptosis at birth and ultimately in microphthalmia in adulthood. However, the identity of the cell signaling pathways that ß1-integrin controls in the lens and the mechanisms causing the cellular phenotype of these mice is not known. In order to investigate this, mice lacking ß1-integrin in the lens were obtained by mating mice harboring loxP sites around exon 3 of the ß1-integrin gene, to MLR10-cre mice, which express Cre-recombinase in all lens cells beginning at E10.5. mRNA levels of known genes involved in extracellular matrix and TGF-ß superfamily signaling were assessed by quantitative rtPCR SuperArray. Changes in protein expression and/or activation compared to wildtype C57Bl/6 controls were detected using immunofluorescence, and western blotting. This lead to the findings that loss of ß1-integrin leads to partial epithelial-mesenchymal transition of the lens epithelium, which is distinct from the typical injury induced EMT seen in the lens. In ß1-integrin null lenses, N Cadherin levels are decreased whereas vimentin levels are increased in the lens epithelium, but decreased in the lens overall by western blotting. E Cadherin mRNA levels were increased prior to the onset of the phenotype, while after, protein levels are decreased. The expression of a number of known targets of TGF-ß superfamily signaling was altered and the forms of phosphorylated Smad3 found in the lens were perturbed as a result of loss of ß1-integrin. Finally, other molecules that interact and crosstalk with both integrin and TGF-ß pathways were also affected. Fibronectin and pFAK (Y397) levels were decreased in the ß1MLR10 lenses, whereas pERK levels were unchanged. In conclusion, these data show that loss of ß1-integrin from the lens epithelium results in the mis-regulation of multiple pathways and leads to the differential expression of numerous genes involved in TGF-ß signaling. This reveals the complexity and importance of ß1-integrin function in the lens epithelium.