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Nox4-mediated ROS production is involved, but not essential for TGFβ-induced lens EMT leading to cataract
S J Das1, T F L Wishart1, K Jandeleit-Dahm2
1Discipline of Anatomy and Histology, Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
Abstract:
The reactive oxygen species (ROS) producing enzyme, NADPH oxidase 4 (Nox4), is upregulated in response to TGFβ in lens epithelial cells in vitro, and its selective inhibition was shown to block aspects of TGFβ-induced epithelial-mesenchymal transition (EMT). In the present in situ study we validate the role(s) of Nox4 in TGFβ-induced lens EMT leading to anterior subcapsular cataract (ASC) formation. Mice overexpressing TGFβ in the lens, that develop ASC, were crossed to Nox4-deficient mice. When comparing mice overexpressing TGFβ 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 TGFβ-induced fibrotic cataracts. In the absence of Nox4, we also see elevated levels of ERK1/2 activity that was shown to be required for TGFβ/Smad2/3-signaling. qRT-PCR revealed upregulation of Nox2 and its regulatory subunit in TGFβ-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.
Insights
NADPH oxidase 4 (Nox4) deficiency delays cataract formation and epithelial-mesenchymal transition (EMT) in mice. This suggests Nox4 plays a key role in transforming growth factor beta (TGFβ)-induced fibrotic cataracts.
Area of Science:
- Ophthalmology
- Cell Biology
- Biochemistry
Background:
- NADPH oxidase 4 (Nox4) produces reactive oxygen species (ROS) and is upregulated by TGFβ in lens epithelial cells.
- Nox4 inhibition can block TGFβ-induced epithelial-mesenchymal transition (EMT), a process implicated in cataract formation.
Purpose of the Study:
- To investigate the role of Nox4 in TGFβ-induced lens EMT and anterior subcapsular cataract (ASC) formation in vivo.
- To elucidate the molecular mechanisms linking Nox4, ROS, and TGFβ signaling in cataract development.
Main Methods:
- Utilized a mouse model overexpressing TGFβ in the lens, crossed with Nox4-deficient mice.
- Analyzed cataract onset, EMT protein markers, and ERK1/2 activity.
- Performed qRT-PCR to assess Nox2 and its regulatory subunit expression.
Main Results:
- Nox4 deficiency delayed the onset of ASC and EMT markers in TGFβ-overexpressing mice.
- Absence of Nox4 led to elevated ERK1/2 activity, which is crucial for TGFβ/Smad2/3 signaling.
- Upregulation of Nox2 was observed in TGFβ-overexpressing lens epithelial cells lacking Nox4.
Conclusions:
- Nox4 is a critical mediator of TGFβ-induced lens EMT and fibrotic cataract formation.
- Nox4 interacts with Smad2/3 and/or ERK1/2 signaling pathways in the development of fibrotic diseases.
- These findings offer insights into potential therapeutic targets for specific forms of cataract.
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