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Reem Z. Renno, Francois C. Delori, Robin A. Holzer, Evangelos S. Gragoudas, Joan W. Miller; Photodynamic Therapy Using Lu-Tex Induces Apoptosis In Vitro, and Its Effect Is Potentiated by Angiostatin in Retinal Capillary Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2000;41(12):3963-3971.
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purpose. To examine the effect of combining angiostatin with photodynamic
therapy (PDT) using Lutetium Texaphyrin (Lu-Tex; Alcon, Fort Worth, TX)
as a photosensitizer in bovine retinal capillary endothelial (BRCE) and
retinal pigment epithelial (RPE) cells and to determine the mode of
PDT-induced cell death in these cell lines.
methods. Cultured BRCE and RPE cells were incubated with angiostatin (500 ng/ml)
for 18 hours and subjected to Lu-Tex/PDT, using treatment parameters
previously optimized (3 μg/ml Lu-Tex for 30 minutes followed by timed
irradiation at 732 nm). Cellular survival was assessed after a 1-week
cellular proliferation. Data were analyzed using Student’s t-test. Caspase 3 activity was monitored in cells after
PDT using a fluorogenic substrate, (Asp-Glu-Val-Asp)-AFC
(7-amino-4-trifluoromethyl coumarin) [DEVD-AFC], of caspase
3. After PDT, expression of Bcl-2, Bcl-xL, Bax, and Bak was
also examined in cell lysates by Western blot analysis.
results. A synergistic cytotoxic effect of angiostatin and Lu-Tex/PDT was
observed in BRCE cells at all fluences used (5, 10, and 20
J/cm2; P ≤ 0.05). These findings
applied only if angiostatin was delivered before PDT. No such
interactive killing effect was observed in RPE cells. Caspase 3
activity was elevated within 10 minutes of PDT in BRCE and RPE cells
and was fluence dependent. Differential modulation of Bcl-2 family
members was observed after PDT in BRCE and RPE cells.
conclusions. The combination of angiostatin and Lu-Tex/PDT potentiates the cytotoxic
effect of Lu-Tex/PDT on BRCE but not on RPE cells. This may provide a
strategy to increase the selectivity of PDT in damaging capillary
endothelial cells with less damage to RPE cells. Lu-Tex/PDT induces
rapid caspase-dependent apoptosis in BRCE and RPE cells. Furthermore,
Lu-Tex/PDT induces apoptosis through selective modulation of members of
the Bcl-2 family and differs between BRCE and RPE
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