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Valeska Anja Mueller, Norbert W Schwarz, Steffen Albrecht, Maryam Hatami, Thomas Skutella; Prevention of secondary cataract development with photodynamic therapy with Verteporfin - an in vitro study on human lens epithelial cells (hLEC’s). Invest. Ophthalmol. Vis. Sci. 2014;55(13):3045. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Secondary cataract is a frequent complication after cataract surgery. So far, there are only limited treatment options. The purpose of this in vitro study on human lens epithelial cells (hLECs) in culture, was to define appropriate treatment parameters regarding drug dose, incubation time, laser irradiation settings and applied light dose that lead to a complete cell death.
As a hLEC culture the SV40 immortalized hLEC B-3 (ATCC CRL 11421) line was used. HLECs were cultured in `Eagle`s Minimal Essential Medium` containing 20% fetal bovine serum. After 4-5 days of growing a confluent cell monolayer on 96-black-well platelets, cells were incubated with various Verteporfin concentrations, ranking from 0.25μg-10μg/ml. Incubation time varied from 24hrs to 5min. Laserparameters were: λ=689nm, fluency rate: 1247mW/sqcm and applied light dose between 50-310J/sqcm. For statistics 12 wells have been treated with the same parameters. For qualitative effect control, light microscopy was performed; for quantitative control the colorimetric WST-1 cell proliferation reagent (Roche) was used. The WST-1 test was performed 24h and 72hrs post treatment. Control groups were: drug alone, laser alone and for positive control: untreated cells; negative control: blank/medium only.
Preliminary results show a 100% cell death of the hLECs at an incubation time of 24hrs at all drug doses with an applied light dose of 310J/sqcm at a fluency rate of 1247 mW/sqcm and an irradiation time of 249sec. WST-1 test after 24 hrs. shows values ≤ 0.3 absorbance units (a.u.) and after 72 hrs. still ≤ 0.3 a.u., whereas the positive viability control (no drug, no laser) shows values >0,5 a.u. after 24hrs and after 72hrs > 3,0 a.u..Drug alone and laser alone had the same values as cells only. The results were statistically significant.
For the first time it is demonstrated that PDT with Verteporfin is capable to cause cell death of hLECs in vitro and hence may be a future tool in cataract surgery to prevent the development of secondary cataract. Whether shorter incubation times and a lower light dose may also lead to complete cell death is under investigation at present and more data will be available soon.
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