Abstract
Purpose :
The aim of this research is to investigate the effect of sublethal thermal laser irradiation on junctions and related cell functions of RPE cells under normal and reduced serum concentrations
Methods :
ARPE-19 cells were cultured for 2 weeks on trans-well permeable inserts in medium with 1% or 10% fetal calf serum (FCS) concentrations. Cells were irradiated with a 1.94 µm continuous-wave thulium laser for 10s to achieve temperatures of 44°C or 48°C. Cell viability was verified by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test. Functional integrity of tight junctions was assessed by measuring the transepithelial resistance (TER) over 2 weeks following irradiation. The apical and basolateral secretions of vascular endothelial growth factor (VEGF) were assessed using enzyme-linked immunosorbent assay (Elisa)
Results :
MTT assays confirmed that irradiation at 44 and 48°C did not significantly increase RPE cell death under both FCS concentrations. After seeding, TER of RPE cell culture increased gradually reaching a maximum after 2 weeks under both FCS concentrations. After laser irradiation, TER of RPE in 10% FCS gradually increased, becoming significantly higher than that of non-irradiated control cells from 7 days for 44°C and from 3 days for 48°C, reaching 1.08- and 1.11-fold, respectively, 9 days after irradiation. In contrast, RPE cells cultured in 1% FCS showed a gradual TER increase only after irradiation at 48°C, reaching a significant 1.06-fold within 7 days after irradiation. Before irradiation, VEGF was secreted more basolaterally under both 10% and 1% FCS. Cells cultured under 10% FCS showed a decrease in the ratio of VEGF secretion (basolateral/apical) from day 2 to 9 after irradiation. In cells cultured under 1% FCS, this reduction in VEGF secretion ratio was limited to 48°C and was only significant on day 9 post-irradiation. At both serum concentrations, the original polarization of VEGF secretion was maintained in the non-irradiated controls
Conclusions :
Results suggest that sublethal thermal stimulation may improve RPE junction functionality, strengthening the outer blood-retinal barrier, and alter VEGF secretion polarity. This might explain certain advantages of sublethal thermal retinal laser therapy. Variations observed under reduced FCS imply differences in cell responses depending on the RPE conditions
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.