Histological Changes After Photodynamic Therapy (PDT) Combined With Anti-vascular Endothelial Growth Factor (VEGF) Therapy In The Mouse Retina

Abstract

Purpose: : Photodynamic therapy (PDT) and/or anti-vascular endothelial growth factor (VEGF) drugs constitute current treatments targeting choroidal neovascularization in age-related macular degeneration. Concern that PDT might induce VEGF and exacerbate the disease has led us to current practice of using anti-VEGF drugs with PDT simultaneously. However, the underlying molecular mechanisms of these therapies are not well understood. The influence of the combined therapy still remains obscure. In this study, we analyze histological influence of PDT simultaneously combined with anti-VEGF therapy.

Methods: : Six week-old C57/B6 male mice were prepared. Verteporfin was injected from their tail veins and the intact retinas were exposed to the laser to activate the drug 15 minutes after the injection. Histological changes were evaluated by hematoxylin eosin (HE) staining, TUNEL assay to detect apoptosis, and immunohistochemistry. Measurement of the VEGF level in the retina or retinal pigment epithelium (RPE)-choroid complex was performed by ELISA in each time point after PDT. A VEGF inhibitor, VEGFRc/fusion protein, or control vehicle was injected intravitreally immediately after PDT.

Results: : HE staining and TUNEL assay showed no obvious histological change after twenty-second irradiation during PDT. However, immunohistochemical analysis showed that glial fibrillary acidic protein, which represents reactivation of Müller glial cells, is upregulated in whole area of the retina after PDT. Level of VEGF protein in the retina was upregulated immediately and transiently after PDT, whereas in the RPE-choroid complex, it was reduced, transiently. VEGF suppression after PDT resulted in apoptotic destruction of the RPE as well as photoreceptor cell layer only in the irradiated area during PDT.

Conclusions: : Reactivation of Müller glial cells was induced in the whole retina after PDT, although overt tissue damage was not detected morphologically. Immediately after PDT, VEGF was required to protect the irradiated tissue including photoreceptor cells and RPE.

Keywords: Muller cells • retinal pigment epithelium • retinal glia