Dexamethasone Induces Senescence-Associated Changes in Trabecular Meshwork Cells by Increasing ROS Levels Via the
Haijun Li1, Jing Ren1, Huiling Cui1
1Henan Provincial People's Hospital, Henan Eye Institute, Henan Eye Hospital, Zhengzhou University People's Hospital, Henan University People's Hospital, Zhengzhou, China.
Abstract:
Glaucoma is a serious complication of glucocorticoid (GC) therapy arising through elevations in intraocular pressure (IOP). Dexamethasone (DEX) is reported to contribute to elevated IOP through different effects on the trabecular meshwork but whether DEX contributes to glaucoma development through the induction of cellular senescence is still unclear. We explored the actions of DEX on transformed human trabecular meshwork cells (HTMCs) using RNA-seq and conducted bioinformatic analyses to determine the affected pathways. Among the 4,103 differentially expressed genes identified in transformed HTMCs treated with 400 nM DEX (2,036 upregulated and 2,067 downregulated genes, respectively), bioinformatic analyses revealed significant enrichment and potential interplay between the transforming growth factor beta (TGFβ)41; signaling and cellular senescence pathways. DEX treatment induced senescence changes in primary and transformed HTMCs as indicated by increases in SA-β-gal positivity, interleukin (IL)-6 secretion, and senescence-associated heterochromatin foci (SAHF) along with selective accumulation of senescence marker p15 and elevations in reactive oxygen species (ROS) levels. Notably, the DEX-induced senescence changes were rescued by treatment with the TGFβ/Smad3 pathway inhibitor SIS3. Furthermore, we show that DEX increases cellular ROS levels via upregulation of NADPH oxidase 4 (NOX4) through activation of Smad3, and that SIS3 decreases ROS levels by downregulating NOX4. Instructively, inhibiting NOX4 with GLX351322 and scavenging ROS with NAC were both effective in preventing DEX-induced senescence changes. Similarly, we found in the mouse model that DEX-ac upregulated p15 and NOX4 expression in the trabecular meshwork, with cotreatment with GLX351322 alleviating elevations in IOP. We establish that DEX induces senescence changes in HTMCs by increasing ROS levels via the TGFβ/Smad3/NOX4 axis, increasing IOP and contributing to glaucoma development.
Insights
Glucocorticoid therapy can cause glaucoma by inducing cellular senescence in the eye's trabecular meshwork. This study reveals dexamethasone triggers senescence via the TGFβ/Smad3/NOX4 pathway, increasing intraocular pressure.
Area of Science:
- Ophthalmology
- Cell Biology
- Pharmacology
Background:
- Glaucoma is a severe complication of glucocorticoid (GC) therapy, often linked to elevated intraocular pressure (IOP).
- The precise mechanisms by which GCs, like dexamethasone (DEX), induce glaucoma, particularly through cellular senescence, remain unclear.
- DEX affects the trabecular meshwork, but its role in promoting glaucoma via senescence requires further investigation.
Purpose of the Study:
- To investigate whether dexamethasone (DEX) induces cellular senescence in human trabecular meshwork cells (HTMCs).
- To elucidate the molecular pathways involved in DEX-induced senescence and elevated IOP.
- To explore potential therapeutic interventions targeting this pathway.
Main Methods:
- RNA sequencing (RNA-seq) and bioinformatic analyses were performed on HTMCs treated with DEX.
- Cellular senescence was assessed using SA-β-gal staining, IL-6 secretion, SAHF formation, p15 expression, and ROS levels.
- In vivo studies utilized a mouse model to evaluate DEX effects on IOP and trabecular meshwork gene expression.
Main Results:
- DEX treatment significantly altered gene expression in HTMCs, with notable enrichment in transforming growth factor beta (TGFβ) signaling and cellular senescence pathways.
- DEX induced clear signs of cellular senescence, including increased SA-β-gal positivity, IL-6, SAHF, p15, and ROS levels.
- Inhibition of the TGFβ/Smad3 pathway (using SIS3), NADPH oxidase 4 (NOX4) (using GLX351322), or ROS (using NAC) effectively prevented DEX-induced senescence. In mice, GLX351322 mitigated DEX-induced IOP elevation.
Conclusions:
- Dexamethasone induces cellular senescence in human trabecular meshwork cells by upregulating ROS via the TGFβ/Smad3/NOX4 signaling axis.
- This DEX-induced senescence contributes to elevated IOP and glaucoma development.
- Targeting the TGFβ/Smad3/NOX4 pathway presents a potential therapeutic strategy for preventing GC-induced glaucoma.
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