Nox4-mediated ROS production is involved, but not essential for TGFβ-induced lens EMT leading to cataract

The reactive oxygen species (ROS) producing enzyme, NADPH oxidase 4 (Nox4), is upregulated in response to TGF beta in lens epithelial cells in vitro, and its selective inhibition was shown to block aspects of TGF beta-induced epithelial-mesenchymal transition (EMT). In the present in situ study we validate the role(s) of Nox4 in TGE beta-induced lens EMT leading to anterior subcapsular cataract (ASC) formation. Mice overexpressing TGF beta in the lens, that develop ASC, were crossed to Nox4-deficient mice. When comparing mice overexpressing TGF beta in lens, to mice that were also deficient for Nox4, we see the delayed onset of cataract, along with a delay in EMT protein markers normally associated with TGF beta-induced fibrotic cataracts. In the absence of Nox4, we also see elevated levels of ERK1/2 activity that was shown to be required for TGEWSmad2/3-signaling. qRT-PCR revealed upregulation of Nox2 and its regulatory subunit in TGF beta-overexpressing lens epithelial cells devoid of Nox4. Taken together, these findings provide an improved platform to delineate putative Nox4 (and ROS) interactions with Smad2/3 and/or ERK1/2, in particular in the development of fibrotic diseases, such as specific forms of cataract.