Abstract
Purpose :
We have shown previously that MMC induces expression of senescence associated proteins in vitro and in vivo. Treating corneal epithelial cells and corneas with compounds that inhibit ROCK induces expression of mRNAs associated with senescence. The purpose of these experiments is to determine whether MMC and RI impact HCLE cell migration and mitochondrial activation using similar or different mechanisms.
Methods :
Immortalized Human Corneal Epithelial (HCLE) cells were grown and assessed for cell velocity using time lapse imaging of randomly migrating cells or were treated with fluorescently labeled CMXROS for 30 min, fixed and imaged to quantify activated mitochondria. The following conditions were assessed: a) Control cells, b) 5μm RI treated cells, c) 0.002% MMC for 3 hrs (transient) before being washed and incubated in media without MMC, and d) transient MMC treatment followed by addition of RI overnight.
Results :
RI decreased cell migration rates by 11% compared to control cells whereas MMC decreased cell migration rates by 25%. Treating cells with RI after MMC lead to a reduction in cell migration of 29% but the difference between MMC alone (25%) and MMC/RI (29%) was not significant. RI and MMC both reduce CMXROS by 6% compared to controls. Adding RI to MMC treated cells reduces CMXROS further to 13% compared to controls. These differences are significant.
Conclusions :
The reduction in the rate of cell migration caused by RI and MMC are accompanied by a reduction in active mitochondria determined by CMXROS compared to untreated cells. However, the significant difference between the rate of cell migration seen in cells treated with RI compared to those treated with MMC is not due to differences in active mitochondria since CMXROS levels are the same for those 2 variables.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.