Abstract
Purpose :
Cataract surgery (CS) has reduced the burden of cataract-associated blindness, but its outcomes are compromised by inflammation and the later development of posterior capsular opacification. Previously we have shown that the lens epithelium drastically remodels its transcriptome within 6H of surgery with differentially expressed genes highly enriched in immediate early transcription factors (IETFs) like FOSB and EGR1 as well as proinflammatory markers. Bioinformatics and wet lab experiments predict that the ERK pathway may acutely regulate IETF and inflammatory marker expression in injured LECs. However, the molecular mechanisms by which surgery acutely activates ERK and later inflammation and fibrosis post CS is unknown. As CS would cause physical deformation of remnant LECs, this project seeks to explore whether mechanical strain triggers acute changes in LEC gene expression.
Methods :
Wildtype (WT) mice were subjected to lens fiber cell removal to model CS and semiquantitative immunostaining for IETF and immune markers was performed. An ERK inhibitor, U0126, was introduced intraocularly immediately following lens fiber cell removal to assess effect of ERK signaling on IETF expression in WT mice. Mechanical sensitive channels (MSC) were generally inhibited with Gadolinium (Gd) during the preparation of LEC explants and IETFs/inflammatory cytokine expression evaluated by Qrt-PCR. Yoda1, a Piezo1 agonist, was introduced to WT mice via intraperitoneal injection to assess effect of activating this MSC on IETF expression.
Results :
Immunostaining and pathway analysis revealed that ERK pathway signaling as well as FOSB and EGR1 protein levels acutely upregulate in LECs PCS; and administration of U0126 immediately after fiber cell removal attenuated this IETF upregulation. Further, we found that IETF upregulation in LECs also rapidly occurs in our LEC explant culture model – indicating cell autonomous regulation. Gd seems to inhibit some IETFs in LECs post 6H culture – implying biomechanical stress as a regulator. Activation of piezo1 with yoda1 appears to regulate a portion of the immediate early response – suggesting its role in mediating IETFs.
Conclusions :
We show that activation of ERK acutely upregulates IETFs and some proinflammatory markers in injured LECs and suggests that MSC inhibition can attenuate these acute LEC responses. Work to explore mechanical stress’ effect on other acute LEC responses is ongoing.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.