Dose-related structural effects of photodynamic therapy on choroidal and retinal structures of human eyes

PURPOSE: To determine the effects of photodynamic therapy (PDT) on choroidal and retinal structures of human eyes. METHODS: One eye from each of three patients with large malignant melanomas of the uvea destined for enucleation received PDT using verteporfin according to the approved treatment recommendations for patients with age-related macular degeneration. Two laser spots and two light doses (50 J/cm(2) and 100 J/cm(2)) were applied in unaffected chorioretinal areas. The effects of PDT were assessed by fluorescein and indocyanine-green angiography. The eyes were enucleated 1 week later, fixed in buffered paraformaldehyde/glutaraldehyde solution, bisected along the laser spots, and processed for light and electron microscopy. RESULTS: In agreement with the clinical angiographic findings of hypofluorescence, a rather selective occlusion of the choriocapillary layer was observed in the 50-J/cm(2) PDT areas, whereas the 100-J/cm(2) PDT areas additionally revealed closure of deeper choroidal vessels and focal alterations of the retinal pigment epithelium. The overlying neurosensory retina, including photoreceptors and retinal capillaries, was well preserved in all PDT areas. Electron microscopy showed that alterations of the choriocapillary endothelium comprised swelling, shrinkage and fragmentation of endothelial cells, detachment from their basement membrane up to complete degeneration of the endothelial lining, leading to platelet aggregation, degranulation, and thrombus formation. Complete occlusion of capillary lumina by fibrin, thrombocytes, and cellular debris was observed. Remaining intact endothelial cells appeared to be reorganized into novel smaller vascular channels within occluded lumina. CONCLUSIONS: PDT with verteporfin at a dosage used clinically induces selective occlusion of the physiological choriocapillaris without affecting deeper choroidal, retinal, and optic nerve vessels or the overlying retinal pigment epithelium and neurosensory retina. The main mechanism of action appears to be vascular thrombosis induced by cytotoxic damage of endothelial cells and platelet activation. An increase in light dose enhances the occlusive effect with thrombosis within deeper choroidal layers and damage to the retinal pigment epithelium. However, photoreceptors remained intact at all light doses used.     

Mechanism of photodynamic occlusion using liposomal Zn(II)-phtalocyanine

PURPOSE: To evaluate the potential of liposomal Zinc(II)-phthalocyanine (ZnPc) to selectively target subretinal vasculature. METHODS: Photodynamic therapy (PDT) with liposomal Zinc(II)-phtalocyanine was used to induce choroidal occlusion in eyes of pigmented rabbits. Drug doses of 0.16, 0.24, 0.32, and 0.4 mg/kg body weight were administered. Photosensitization was performed at a wavelength of 671 nm and an irradiance of 100 mW/cm2 applying fluences of 5, 10, 20, and 50 J/cm2. RESULTS: Using liposomal ZnPc, occlusion of choroidal vessels was achieved without damage to the overlying neurosensory retina. A tight dose correlation was found with a drug dose of 0.32 mg/kg and a light dose of 10 J/cm2 inducing a selective thrombosis of the subretinal capillary layer. Histology revealed a selective intravascular alteration of the endothelial cells. CONCLUSIONS: PDT using liposomal ZnPc allows occlusion of subretinal vasculature with maintenance of neuroretina and RPE. The destructive effect on choroidal vascular endothelium is intensive.     

Influence of photodynamic therapy on expression of vascular endothelial growth factor (VEGF), VEGF receptor 3, and pigment epithelium-derived factor

PURPOSE. To evaluate the impact of photodynamic therapy (PDT) on expression and distribution of vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR)-3, and pigment epithelium-derived factor (PEDF). METHODS. Eyes of patients scheduled for enucleation due to untreatable malignancy served as study eyes (n = 4), age-matched donor eyes were used as the control (n = 4). PDT using verteporfin with the recommended standard parameters was applied to intact areas of the perimacular region. Lesions were classified by ophthalmoscopy, fluorescein angiography (FA), and indocyanine green angiography (ICGA), as well as light and electron microscopic (LM/EM) histology. Immunolabeling using specific antibodies against VEGF, VEGFR-3, and PEDF was performed in PDT-treated areas, untreated collateral areas in study eyes, and untreated areas of control eyes. Specimens were fixed in 4% paraformaldehyde and 1% glutaraldehyde and embedded in paraffin. Four-micrometer-thick sections were stained using the peroxidase-labeled streptavidin-biotin method. RESULTS. All PDT-treated areas demonstrated characteristic choroidal hypofluorescence by FA and ICGA. LM/EM histology revealed selective damage of choriocapillary endothelial cells. VEGF was expressed in the endothelial layer of choriocapillaries and focally within larger choroidal vessels in treated areas, but not in untreated areas. Sites with positive VEGF labeling also demonstrated upregulation of VEGFR-3. PEDF expression was localized to retinas in all eyes; however, PEDF staining of choroidal endothelial cells was specific for treated areas of study eyes. CONCLUSIONS. PDT using verteporfin induces a reproducible angiogenic response in elderly human eyes. VEGF, VEGFR-3, and PEDF expression is enhanced after PDT. Choroidal endothelial cells appear to be the primary site of angiogenic stimulation.     

Influence of treatment parameters on selectivity of verteporfin therapy

PURPOSE: To improve selectivity of verteporfin therapy (PDT) in neovascular age-related macular degeneration (AMD) using modified treatment parameters. METHODS: Nineteen consecutive patients with predominantly classic choroidal neovascularization (CNV) in AMD were treated with 6 mg/m2 verteporfin given as bolus infusion. Patients received PDT with a fluence of either 25 or 50 J/cm2. Choroidal perfusion changes were evaluated by indocyanine green angiography (ICGA) at baseline, day 1, week 1, week 4, and month 3. Secondary outcomes were CNV closure rate and therapy-induced leakage documented by fluorescein angiography (FA). The safety of the treatment was assessed with ETDRS visual acuity. RESULTS: Complete CNV closure was achieved in all patients at day 1. Choroidal hypoperfusion was minimal in eyes treated with a reduced fluence of 25 J/cm2. Most patients treated with 50 J/cm2 showed significant choriocapillary nonperfusion at week 1, lasting as long as 3 months. A transient PDT-induced increase in leakage area in FA at day 1 was found to be more extensive in the 50-J/cm2 group. CONCLUSIONS: Bolus administration of verteporfin combined with a reduced light dose achieved improved selectivity of photodynamic effects, avoiding collateral alteration of the physiologic choroid while obtaining complete CNV closure. An increased selectivity with decreased effect on the surrounding choroid should be of advantage in verteporfin monotherapy as well as in combination strategies.     

Photodynamic therapy with verteporfin for choroidal neovascularization in patients with diabetic retinopathy.

PURPOSE: To report the use of photodynamic therapy (PDT) with verteporfin in three patients with choroidal neovascularization (CNV) from age-related macular degeneration and underlying diabetic retinopathy. The level of diabetic retinopathy would have excluded these patients from participation in previously reported randomized clinical trials evaluating PDT with verteporfin due to a theoretic concern of damage to the overlying retinal vasculature. DESIGN: Retrospective interventional case series. METHODS: Three patients from a referral practice with at least severe nonproliferative diabetic retinopathy and a history of clinically significant macular edema developed loss of vision from concurrent choroidal neovascularization evaluated with fundus photography and fluorescein angiography before and after PDT with verteporfin to identify adverse retinal vascular events. RESULTS: Four eyes in three patients had PDT using verteporfin. Three eyes received two treatments. With short follow-up, visual acuity remained stable in two eyes, improved from 20/400 to 20/320 in one eye, and decreased from 20/200 to 20/400 in one eye. Fluorescein angiograms at intervals from 2 weeks to 3 months after PDT showed no damage to the retinal vasculature or progression of the diabetic retinopathy, but did show a decreased area of fluorescein leakage from CNV. One eye that had new subretinal hemorrhage following treatment appeared to show new vasculopathy on initial evaluation of the post-treatment angiogram. Retrospective review suggested that the subretinal hemorrhage provided increased contrast to more easily visualize vasculopathy that was present before the PDT. CONCLUSIONS: Three patients with diabetic retinopathy undergoing a total of seven PDT treatments with verteporfin in four eyes had no new retinal vascular abnormalities develop. No other atypical responses of CNV to PDT were noted except new subretinal hemorrhage, providing increased contrast of the overlying vasculature, which gave the false impression of the development of new vasculopathy in one eye. Patients with diabetic retinopathy who have concurrent CNV for which PDT with verteporfin is recommended should be cautioned regarding the theoretical concerns of harming the retinal vasculature. Periodic surveillance for such concerns seems warranted until more experience is obtained.     

Expression of angiogenic factors by photodynamic therapy

PURPOSE: The purpose of this study was to evaluate the impact of photodynamic therapy (PDT) on the regulation of angiogenic factors such as vascular endothelial growth factor (VEGF) and pigment epithelium derived factor (PEDF) in human eyes. METHODS: Eyes of patients with untreatable malignancy served as the study eyes (n = 2), age-matched donor eyes were used as controls (n = 2). Standard Verteporfin-PDT was applied to intact areas of the posterior pole. One week after PDT the eyes were examined by ophthalmoscopy as well as fluorescein (FA) and indocyanine green angiography (ICGA). After enucleation the eyes were processed for LM/EM histology. Immunolabeling using specific antibodies against VEGF and PEDF was performed in PDT-treated areas, untreated collateral areas of study eyes and untreated areas of control eyes. RESULTS: All PDT-treated areas demonstrated a typical choroidal hypofluorescence on FA/ICGA. In LM/EM histology a selective damage of choriocapillary endothelial cells was found. VEGF expression was localized only to choriocapillary endothelial cells and focally in larger choroidal vessels of PDT-treated areas. Untreated areas of study eyes and controls were VEGF-negative. PEDF staining was observed in retinas and RPE of all eyes, but only choroidal endothelial cells of PDT-treated areas showed a PEDF-positive reactivity. CONCLUSION: PDT not only induces structural and angiographic, but also biological effects in human eyes. VEGF and PEDF expression can be documented in choroidal endothelial cells following PDT and could have an impact on the recovery process after treatment.     

Verteporfin photodynamic therapy in the rat model of choroidal neovascularization: angiographic and histologic characterization

PURPOSE: To develop a model of verteporfin photodynamic therapy (PDT) for experimental choroidal neovascularization CNV in the rat. METHODS: A laser injury model was used to induce experimental CNV in rats. The transit and accumulation of the photosensitizer verteporfin was assessed angiographically in CNV lesions, to determine the optimal time for delivery of light energy. The CNV lesions were then treated with verteporfin PDT, with two doses of verteporfin (3.0 and 6.0 mg/m(2)) and four activating doses of light energy (10, 25, 50, and 100 J/cm(2)). Closure of the CNV was assessed both angiographically and histologically. Verteporfin PDT was also performed on areas of normal choroid and retina at the two verteporfin doses and four light energy doses. The effect of these treatments on these structures was also assessed angiographically and histologically. RESULTS: Peak verteporfin intensities in the CNV were detected at 15 to 20 minutes after intravenous injection. Rates of closure of the CNV varied as a function of the dose of verteporfin and of the activating light energy. Angiographic closure of the CNV correlated with damage to the neovascular complex, as seen with light and electron microscopy. Damage to areas of normal choroid and retina treated with verteporfin PDT also varied as a function of the verteporfin and light energy doses. CONCLUSIONS: Verteporfin PDT for experimental CNV in the rat is a feasible, effective, and reproducible model that can be used for testing the efficacy of adjunctive therapy to verteporfin PDT.     

Effect of photodynamic therapy on the function of the outer blood-retinal barrier in an in vitro model

Background Photodynamic therapy (PDT) is a well established clinical treatment for age-related macular degeneration (AMD), and comprises intravenous injection of verteporfin and subsequent application of a non-thermal laser beam to the area of AMD to induce selective vascular occlusion. Since there is evidence that PDT may cause outer blood-retinal barrier (BRB) breakdown and possibly RPE cell alteration, we investigated the effect of PDT on the BRB function of the RPE in an in vitro model.Methods Twenty-one monolayers of human RPE cells were cultured on semipermeable membranes until a stable barrier function was achieved as determined by transepithelial electrical resistance (TER) and sodium fluorescein permeability. To test the effect of PDT on the outer BRB function, non-thermal laser (692 nm), verteporfin or a combination of both were applied. TER assessment prior to and after PDT was utilized to identify changes in barrier function of the RPE in this in vitro model. Finally, monolayers of RPE cells were evaluated by transmission electron microscopy (TEM).Results No significant TER decrease was observed after application of non-thermal laser alone or after administration of verteporfin in therapeutic concentrations, but combination of these modalities resulted in significantly decreased TER within 4 h. Except for intercellular blisters, no damage to the RPE was evident in TEM. Verteporfin added at concentrations higher than therapeutic doses (2 mg/ml) resulted in an immediate decrease in TER and damage to the RPE cells.Conclusion The combination of a therapeutic concentration of verteporfin and application of non-thermal laser resulted in a morphologically and functionally detectable breakdown of the outer BRB function of the RPE without any damage to the RPE cells themselves in vitro. However, increasing the concentration of verteporfin can result in RPE cell damage.     

Choroidal vascular remodelling in central serous chorioretinopathy after indocyanine green guided photodynamic therapy with verteporfin: a novel treatment at the primary disease level

AIMS: To evaluate the changes in the choroidal vasculature in central serous chorioretinopathy (CSC) after photodynamic therapy (PDT) with verteporfin and to assess its potential role as a treatment option. METHODS: A prospective, non-comparative, interventional study was performed in eyes with persistent CSC or chronic CSC that had fluorescein leakage at the fovea. All eyes received one single session of PDT with verteporfin (6 mg/m2 body surface area) followed by application of 50 J/cm2 laser at 689 nm. The laser spot size was guided by findings in ICG-A. RESULTS: Six eyes from six patients with a mean follow up of 12.7 months were analysed. Narrowing of the original dilated choroidal vessels and decrease in extravascular leakage could be demonstrated in all (100%) PDT treated eyes. 3 months after PDT, the mean diameter of the dilated choroidal vessel reduced from 546 microm to 371 microm (p=0.028). Five (83%) patients had improvement in visual symptoms and best corrected visual acuity. Fluorescence leakage stopped at the 1 month follow up in five eyes (83%) and at 3 months in all six eyes (100%). One eye developed choroidal neovascularisation at 3 month follow up. There was no other serious ocular or systemic complication. CONCLUSIONS: PDT is successful in stopping the fluorescein leakage in all six patients without recurrence of CSC. The ICG-A findings of choroidal vascular remodelling and decreased choroidal permeability after PDT are encouraging. As the sample size is small and the mean follow up period is short, further trials of PDT with verteporfin for CSC are required to address the optimal parameters in ensuring longer term safety and efficacy outcome.     

Photodynamic therapy with verteporfin in a rabbit model of corneal neovascularization

PURPOSE: To determine the efficacy of photodynamic therapy (PDT) with verteporfin (Visudyne; Novartis AG, Basel, Switzerland) for treatment of corneal neovascularization in a rabbit eye model. METHODS: Corneal neovascularization was induced in Dutch belted rabbits by placing an intrastromal silk suture near the limbus. Verteporfin was administered by intravenous injection at a dose of 1.5 mg/kg, and the pharmacokinetics of verteporfin distribution in the anterior segment or PDT-induced (laser energy levels 17, 50, and 150 J/cm(2)) regression of corneal blood vessels were then determined. To assess PDT-induced toxicity of the anterior segment, corneal and iris/ciliary body histology, and IOP were evaluated after PDT. RESULTS: Verteporfin accumulation in vascularized regions of the cornea and the iris/ciliary body tissue were time dependent and maximum levels achieved at 60 minutes after injection. In rabbits, PDT of corneal vessels using laser energy of 17 or 50 J/cm(2) resulted in 30% to 50% regression of corneal neovascularization; however, in these animals, a rapid regrowth of new blood vessels occurred between 3 and 5 days. In the rabbits receiving PDT using laser energies of 150 J/cm(2), the mean vessel regression was 56%. During the nine days of the laser therapy follow-up period, no vessel regrowth was observed in these rabbits. Histologic examination of the anterior segment after PDT (150 J/cm(2)) showed localized degeneration of the corneal blood vessels without observable change in other anterior segment structures. CONCLUSIONS: These results provide evidence that PDT can produce significant regression of neovascular corneal vessels with no observable toxicity to the anterior segments. However, the optimal laser energy necessary to induce long-term regression (150 J/cm(2)) was three times that used to treat choroidal neovascularization.     

The segmental characteristic of the human choroidal vessels and its clinical significance

The segmental vascular supply and anastomosis of human choroidal vessels were studied in 10 subjects (20 eyes) by the corrosion cast technique and scanning electron microscopy. The distribution pattern of the choroidal vessels was characterized by distinct segmental arrangement of large vessels and lobular arrangement of choriocapillaries. Maximal choriocapillary density and caliber were observed in the posterior pole. The mean diameters of the choriocapillaries in the macular, the equatorial, and the peripheral areas were 25.19 + 0.99 microns, 22.42 + 1.07 microns, and 28.1 + 1.29 microns respectively. It was also revealed that interarterial anastomosis existed among large arteries, precapillary arterioles, and even choriocapillaries. Veno-venous anastomotic communications were a general phenomenon. Therefore, the choroidal vessels are not true end-arteries anatomically. Obstructions at different levels of the choroidal arteries show corresponding clinical features. Occlusion in a posterior ciliary artery or a large choroidal artery is known as the triangular syndrome. Choriocapillary obstruction can be caused by many diseases. The relationship between the RPE and the choriocapillaries was discussed.     

Ocular changes after photodynamic therapy

PURPOSE: The aim of this study was to identify the changes in the primate visual system after a single session of photodynamic therapy (PDT) in an intact nonhuman primate retina. METHODS: As part of a larger study, PDT (wavelength 689 nm, 50 J/cm2, 600 mW/cm2, 83 seconds, 4-mm spot size) with verteporfin (6 mg/m2 intravenous infusion) was performed in one eye each of two cynomolgus monkeys. Fundus photography, fluorescein angiography (FA), indocyanine green angiography (ICG), optical coherence tomography (OCT), and multifocal electroretinography (mfERG) were performed at baseline and 12 time points (1-283 days) after PDT. In addition, retinal histopathologic findings were evaluated at 9 months. RESULTS: Various morphologic changes, including whitening of the treated area, RPE proliferation, closure of the choroidal vasculature, and subretinal edema (followed by foveolar thinning) were observed. Most of the changes persisted and were detectable in histopathologic evaluation at 9 months. Reductions of the mfERG amplitude, followed by varying degrees of recovery from the treated and the border regions, were observed. This was accompanied by progressive delay of P1 peak time up to 3 months after treatment, followed by complete recovery at 9 months. In addition, the nontreated area showed amplitude and timing mfERG deficits, which underwent gradual (but not complete) recovery. CONCLUSIONS: In a primate model, under standard clinical parameters, a single PDT treatment resulted in various dynamic morphologic and functional retinal changes detectable for up to 9 months after treatment. The significance of the observed changes and possible ways of pharmacologic interference with PDT adverse effects are discussed.     

Changes in confocal indocyanine green angiography through two years after photodynamic therapy with verteporfin

PURPOSE: To evaluate vascular changes documented by confocal indocyanine green angiography (ICGA) through 2 years after photodynamic therapy (PDT) with verteporfin of neovascular age-related macular degeneration (AMD). DESIGN: Single-center, 2-year, randomized, double-masked, interventional, placebo-controlled trial (subset from Treatment of AMD with PDT Study [TAP]). PARTICIPANTS: Sixty patients with subfoveal choroidal neovascularization (CNV) resulting from AMD. INTERVENTION: Patients were randomized in a ratio of 2:1 to a standard regimen using verteporfin therapy at a drug dose of 6 mg/m(2) body surface area and a light dose of 50 J/cm(2) or a sham treatment with placebo infusion and light exposure. Retreatments, if persistent fluorescein leakage from CNV was documented, were scheduled at 3-month intervals for up to 2 years. Confocal ICGA with tomographic sections was performed at baseline and continuously at the month 3, 6, 12, and 24 examinations using a standardized protocol. MAIN OUTCOME MEASURES: Analysis included the size of the neovascular net, the area of late hyperfluorescence, and choroidal hypofluorescence during early- and late-phase imaging. RESULTS: In the verteporfin-treated group, the mean size of the CNV and the mean area of late leakage consistent with active leakage or staining showed no further enlargement at month 12 and were reduced at month 24. In the placebo-treated group, new vessels grew threefold compared with baseline and exhibited persistent late hyperfluorescence resulting from leakage at 24 months. Associated choroidal hypofluorescence within the treated area was significantly increased in eyes treated with verteporfin PDT compared with the control group during the first year, persisted during all ICGA phases, and was irreversible during follow-up. Image analysis revealed choroidal hypoperfusion with choriocapillary dropout, which correlated with chorioretinal atrophy clinically. Progressive destruction of choroidal integrity by fibrosis in control eyes led to a similar extent of collateral hypofluorescence in both groups through the 24-month examination. CONCLUSIONS: Indocyanine green angiography is an important adjunct in the identification of vascular effects associated with verteporfin PDT. Repeated treatments effectively arrested CNV growth and reduced leakage activity. The collateral impairment of choroidal perfusion appears to influence the visual outcome of the treatment.     

Threshold and retreatment parameters of NPe6 photodynamic therapy in retinal and choroidal vessels

BACKGROUND AND OBJECTIVE: To determine the threshold fluence for producing choroidal and retinal vascular occlusion with mono-L-aspartyl chlorin e6 (NPe6) photodynamic therapy (PDT) during primary treatment and the effect of retreatment. METHODS: Primary treatment: Rats, rabbits, and monkeys underwent NPe6 PDT to determine the threshold fluences for choroidal and retinal vessel occlusion. The threshold was determined by analyzing fluorescein angiograms for areas of nonperfusion. Retreatment: Dutch-belted rabbits underwent NPe6 PDT followed by fluorescein angiography. Rabbits were retreated one week later at the same parameters. RESULTS: Fluence levels and vascular damage thresholds were always higher for retinal than for choroidal vascular occlusion. Retreatment caused choroidal vessel closure at all tested fluences but retinal capillaries closed only at a fluence > 17.7 J/cm2. CONCLUSION: NPe6 PDT has a lower threshold to occlude choroidal vessels than retinal vessels. The cumulative effect of retreatment does not damage retinal vessels unless the threshold is exceeded during a single retreatment session.     

BDNF reduces the retinal toxicity of verteporfin photodynamic therapy

PURPOSE: Verteporfin photodynamic therapy (PDT) is the most effective treatment for age-related macular degeneration, using laser activation of a photosensitizing dye to achieve closure of choroidal neovascularization. Although PDT preferentially affects pathologic vessels, it can also cause collateral damage to the overlying retina. In the current study, it was found that the neuroprotective agent brain-derived neurotrophic factor (BDNF) reduces this retinal damage. METHODS: Normal adult rats received intravitreal BDNF in one eye and PBS or no injection in the other eye 2 days before PDT. RESULTS: Control eyes exhibited choroidal hypofluorescence, moderate to severe photoreceptor loss, and depression of local retinal function measured using multifocal ERG in the laser-treated area. BDNF-injected eyes had more surviving photoreceptors and improved multifocal ERG responses 1 week after PDT. BDNF did not diminish the effect of PDT on the choroidal circulation as assessed by fluorescein angiography, and there was no evidence of retinal toxicity due to BDNF treatment. CONCLUSIONS: These results suggest that adjunctive neuroprotective therapy may reduce collateral damage to photoreceptors and improve visual outcome after PDT.     

Comparison of reduced and standard light application in photodynamic therapy of the eye in two rabbit models

Background Current PDT treatment for age-related macular degeneration uses a standard radiant exposure of 50 J/cm² at an irradiance of 600 mW/cm². However; there is a general problem with the unusually high irradiance; in fact, the rate of photochemical production of singlet oxygen may be limited by insufficiently oxygenized neovascular tissue. It was the aim of this study to evaluate the efficacy of verteporfin (Visudyne) photoactivation to induce thrombosis of choriocapillaries and in experimentally induced corneal neovascularizations in rabbits by varying irradiance and retinal radiant exposure.Materials and methods The light-dose threshold to induce micro-thrombosis in the choriocapillaries (seven eyes) and in corneal neovascularizations (eight eyes) of Chinchilla-Bastard rabbits using different retinal irradiances (100 and 600 mW/cm²) at different radiant exposures (20, 10, 5, 2.5, 1.25, 0.62, and 0.3 J/cm²) was evaluated. Induction of neovascularizations was performed 7 days prior to PDT treatment using intracorneal silk sutures. A dose of 2 mg/kg verteporfin was intravenously infused 10 min before standard PDT. The criterion for vascular thrombosis was vessel closure as determined by fluorescein angiography 1 h and 1 day post exposure.Results Experiments on the choroid revealed vessel closure 1 h after irradiation at ED₅₀ = 10.8 J/cm² (both 600 and 100 mW/cm²) and after 24 h at ED₅₀ = 2.4 J/cm² (600 mW/cm²) versus 1.8 J/cm² (100 mW/cm²). Vessel closure was enhanced at irradiation with 100 mW/cm². Regarding corneal neovascularizations, vessel thrombosis was observable by dark appearance of irradiated clotted neovascular tissue and angiographically by a lack of leakage at ED₅₀ thresholds of 0.62 J/cm² (1 h) and 0.41 J/cm² (1 day) for 100 mW/cm² and of 0.99 J/cm² (1 h), and 0.67 J/cm² (1 day) for 600 mW/cm². Thus in both experiments thresholds for vessel closure were reduced by a factor of 1.5 for the lower intensity. Histology revealed more selective vessel occlusion without RPE and photoreceptor damage for 100 mW/cm² rather than 600 mW/cm² intensity at threshold irradiation.Conclusion Low-intensity PDT with verteporfin for neovascular tissue seems to be more effective than regular high-intensity PDT. Future preclinical trials should address the issue of proper dosimetry for effective PDT in age-related macular degeneration.None of the authors have any proprietary interest in any material or method used in the study     

Evaluation of the new photosensitizer Tookad (WST09) for photodynamic vessel occlusion of the choroidal tissue in rabbits

PURPOSE: To determine the efficacy of Tookad (WST09; Negma-Lerads, Magny-Les-Hameaux, France) photodynamic therapy (T-PDT) by evaluating the angiographic and histologic closure of choroidal vessels at different radiance exposures, drug dosages, and intervals between photosensitizer injection and laser application in a rabbit model. METHODS: Chinchilla Bastard rabbits were injected intravenously with three different dye concentrations (2.5, 5, and 10 mg/kg) before application of light. In every group T-PDT was performed at four different times after injection: 5, 15, 30, and 60 minutes with different radiance exposures ranging from 200 to 3 J/cm2. Fundus photographs and fluorescein angiograms were obtained 90 minutes after injection. Follow-up angiographies were performed at days 1, 3, 7, and 14 after initial treatment. Histology was performed in selected cases immediately after treatment and on days 1, 3, and 7. RESULTS: Immediately after irradiation, most of the visible lesions were angiographically hyperfluorescent due to damaged vessel endothelium and associated RPE damage. Lesions from high-radiance exposures revealed immediate hypofluorescence, indicating vessel closure. Hypofluorescent lesions appeared mainly during day 1 (all lesions angiographically visible, some hypofluorescent) to day 3 (all lesions hypofluorescent) after treatment. At day 7, ophthalmoscopically visible hyperpigmentation took place in all lesions. ED50 thresholds for angiographic hypofluorescence determined at day 3 after treatment with 2.5 mg/kg were 18.8 J/cm2 (5 minutes), 62.0 J/cm2 (15 minutes), and >100 J/cm2 (30 minutes); with 5 mg/kg, 8.4 J/cm2 (5 minutes), 22.8 J/cm2 (15 minutes), 54.5 J/cm2 (30 minutes), and >100 J/cm2 (60 minutes); and with 10 mg/kg, 11.7 J/cm2 (30 minutes) and 54.1 J/cm2 (60 minutes). Histology of the angiographically hypofluorescent lesions revealed vessel thrombosis in all groups 1 hour after PDT up to 7 days after treatment. Sparing of photoreceptors indicated selectivity of T-PDT; however, slight damage was partly observable. After 7 days, localized proliferation of the RPE cells was noted and was enhanced 14 days after treatment. CONCLUSIONS: T-PDT has the potential to achieve selective choroidal vessel occlusion with proper parameter selection, such as (1) 2.5 mg/kg, 5 minutes, 100 J/cm2; (2) 5 mg/kg, 5 minutes, 25 J/cm2; or (3) 5 mg/kg, 15 minutes, 50 J/cm2; however, slight damage to the photoreceptors cannot be ruled out. RPE proliferation indicates primary RPE damage due to PDT, also described with the use of all other photosensitizers.     

Transpupillary thermotherapy: effect of wavelength on normal primate retina

PURPOSE: To correlate changes in primate fundus after transpupillary thermotherapy (TTT) at two wavelengths. METHODS: Twelve primate eyes were treated with TTT using a wavelength of 635 nm (n=7) or 810 nm (n=5). Laser parameters were as follows: 635 nm (spot size, 1 mm; duration, 30-8 seconds; and fluence [power over time], 20-91.4 J/cm) and 810 nm (spot size, 2 mm; duration, 60 seconds; and fluence, 96-436 J/cm). Fundus photography, fluorescein and indocyanine green angiography, and enucleation were performed at time 0 or 2 weeks after TTT for histologic analysis. RESULTS: Threshold for fundus lesions (91.4 J/cm at 635 nm and 191 J/cm at 810 nm), acute and chronic retinal damage shown by histologic analysis (79.2 J/cm at 635 nm and 96 J/cm at 810 nm), and choroidal vessel occlusion (50 J/cm at 635 nm and 96 J/cm at 810 nm) were lower at 635 nm. Disorganization of the retina and retinal pigment epithelium was seen for both wavelengths at time 0 and 2 weeks after TTT. Occlusion of the choriocapillaris and choroidal stromal vessels was noted only in specimens obtained 2 weeks after TTT. CONCLUSIONS: TTT resulted in acute and delayed damage to the neurosensory retina that persisted at 2 weeks. The 635-nm wavelength demonstrated a lower threshold fluence for visible fundus lesions, retinal damage, and choroidal vascular occlusion than the 810-nm laser.     

Photodynamic therapy for bilateral idiopathic detachment of the RPE

BACKGROUND: A bilateral monofocal detachment of the pigment epithelium (RPE) without any signs of ARMD or other retinal pathology represents a therapeutic challenge. PATIENT: An otherwise healthy 51-year-old woman presented first with a monocular, later a binocular decrease of visual acuity and metamorphopsia. Optical coherence tomography (OCT) showed a bilateral dome-shaped detachment of the RPE. Pooling beneath the detachment was documented using fluorescein angiography. A choroidal neovascularisation could be excluded by means of indocyanine green angiography (ICG). Due to the binocular decrease of visual acuity from 20/20 to 20/63 on the right eye and 20/100 on the left eye in spite of intensive systemic acetazolamide therapy both eyes were treated with photodynamic therapy (PDT) using verteporfin. During monthly controls, the visual acuity increased up to 20/20 on the right eye and 20/25 on the left eye. Metamorphopsia was also reduced. CONCLUSION: An idiopathic detachment of the pigment epithelium can effectively be treated using PDT whereas the underlying pathology remains unclear.     

Choriocapillaris photodynamic therapy using indocyanine green

PURPOSE: To evaluate the potential of photodynamic therapy using indocyanine green for occlusion of choroidal neovascularization, the authors studied efficiency and collateral damage of photodynamic therapy-induced photothrombosis in the rabbit choriocapillary layer. METHODS: Fundus photography, fluorescein angiography, and light and transmission electron microscopy were used to study the efficiency of photodynamic therapy-induced photothrombosis using indocyanine green as the photosensitizer, and to assess the resultant collateral damage. The delivery system consisted of a modified infrared diode laser tuned to 810 nm, near the maximum absorption peak of indocyanine green. RESULTS: Choriocapillary occlusion was achieved at indocyanine green doses of 10 and 20 mg/kg and a radiant as low as 6.3 J/cm(2). When photodynamic therapy was performed with indocyanine green doses of 10 mg/kg, damage to the neural retina was minimal. Only inner photoreceptor segments showed degeneration, probably secondary to choroidal ischemia. Bruch membrane remained intact. Retinal pigment epithelium was invariably damaged, as seen with other photosensitizers. Temporary occlusion of large choroidal vessels occurred at both dye doses. CONCLUSIONS: In this experimental study, photodynamic therapy using indocyanine green and 810-nm light irradiation produced endothelium-bound intraluminal photothrombosis, with preservation of the retinal architecture and minimal loss of visual cells. Membrane targetability, hydrophilic and fluorescent properties, and activation at 805 nm suggest indocyanine green as a potential photosensitizer for choroidal neovascularization. These combined considerations point toward further study of photodynamic therapy using indocyanine green for the treatment of choroidal vascular disease.