Hematogenous photosensitization. A mechanism for the development of age-related macular degeneration.

Age-related macular degeneration (ARMD) is one of the leading causes of severe visual loss in the United States. Numerous risk factors have been investigated, but the pathogenesis of ARMD has remained elusive. The authors propose that ARMD develops as a direct result of photosensitization of the vascular endothelium of the choriocapillaris, Bruch's membrane, and the retinal pigment epithelium (RPE) by superoxide anion and singlet oxygen generated by photoactive compounds in blood. Using electron-spin resonance spectrometry, the free-radical trap, 5,5-dimethyl-1-pyrroline-N-oxide, and the singlet-oxygen trap, 2-(9,10-dimethoxyanthracentyl)-t-butylhydroxylamine, the authors demonstrate that the photoactive compound, protoporphyrin IX (PP IX), a naturally occurring precursor molecule of hemoglobin found in erythrocytes and plasma, generates superoxide anion and singlet oxygen. The amount of reactive-oxygen species produced by this system is dependent on the concentration of PP IX and the intensity and wavelength of the light delivered. Furthermore, the production of these photooxidants is significantly reduced by filtering the excitatory wavelengths of PP IX. These photogenerated oxidants could damage the vascular endothelium of the choriocapillaris, Bruch's membrane, and the RPE, necessitating a reparative process. This could result in features characteristically seen in ARMD such as a thickened Bruch's membrane, RPE atrophy, and hyperplasia. Prevention of phototoxic damage by this mechanism could involve enhancing protective enzymes, increasing scavenger substances, or supplying appropriate filters to eliminate the exciting wavelengths of light.

An elevated hematogenous photosensitizer in the preterm neonate.

PURPOSE: Human blood contains low levels of protoporphyrin IX (PP IX), a photoactive compound that produces reactive oxygen species when exposed to light. It has been proposed that photoactivation of PP IX and subsequent generation of potentially tissue-damaging reactive oxygen may be a mechanism of retinal injury in retinopathy of prematurity (ROP). The purpose of this study is to determine an association between blood PP IX level and infant birth-weight and gestational age. METHODS: Erythrocyte PP IX levels were measured from the umbilical cord blood of 31 neonates, both full term and preterm. Birthweights and gestational ages were recorded. RESULTS: PP IX levels in infants weighing < 1000 grams (n = 7) average 200 micrograms/dl, which is significantly higher than PP IX levels in infants weighing > 1000 grams (125 micrograms/dl, n = 24; P < 0.02). CONCLUSION: Elevated PP IX levels may place preterm neonates at increased risk for photosensitizing retinal injury.

Light-induced deposits in Bruch's membrane of protoporphyric mice.

Photosensitization of choriocapillary endothelium with blood-borne photosensitizers, such as protoporphyrin IX, has been proposed as a mechanism for the choriocapillary sclerosis and Bruch's membrane deposits seen in age-related macular degeneration. Utilizing a mouse model of protoporphyria with approximately a 10-fold increase in protoporphyrin IX level and exposure to blue light (14 microW/cm2; bandwidth, 390 to 430 nm), a time- and light-dependent increase in choriocapillary and sub-retinal pigment epithelium basal laminar-like deposits could be demonstrated at 7 months by transmission electron microscopy. Thickening of the choriocapillary endothelial basement membrane with a homogeneous electron-dense material was first noted in protoporphyric mice exposed to blue light for 13 weeks. At 28 weeks the experimental animals exhibited a thick band of homogeneous deposits at the level of the choriocapillary basement membrane and electron-dense fibrillogranular deposits of varying sizes along the inner aspect of Bruch's membrane, with fibrils measuring up to 16 nm, with a periodicity of 13 nm. These deposits contributed to an overall thickening of Bruch's membrane with narrowing of the choriocapillaris. The morphologic appearance and localization of these deposits within Bruch's membrane of this animal model are similar to previously described deposits noted in the aging Bruch's membrane prior to the development of age-related macular degeneration.

Light-dependent generation of superoxide from human erythrocytes.

Using the cytochrome c reduction method, we investigated light-dependent erythrocytic superoxide production. After 4 h light and dark exposure of erythrocytes from eight healthy human subjects, an average of 18.6% more superoxide was generated by erythrocytes exposed to light. Pretreatment of erythrocytes with the superoxide dismutase inhibitor N,N-diethyldithiocarbamate increased detection of superoxide while pretreatment with the anion channel blocker 4-acetamido-4'-isothiocyano-2,2'-disulfonic acid stilbene decreased detection. These findings indicate that substantially more spontaneously generated superoxide is produced and escapes from normal erythrocytes at ambient oxygen tensions on exposure to light. This excess generation and escape of superoxide from erythrocytes exposed to light may result in tissue photosensitization, especially in the retina of the eye, where high oxygen tension, blood and chronic light exposure occur simultaneously.

Injection of tissue plasminogen activator into a branch retinal vein in eyes with central retinal vein occlusion.

PURPOSE: Central retinal vein occlusion (CRVO) often produces significant and permanent loss of vision in the affected eye. The purpose of this study was to determine if patients with vision loss secondary to CRVO treated with retinal vein cannulation and infusion of tissue plasminogen activator (t-PA) experienced recovery of visual acuity. DESIGN: Prospective, noncomparative, interventional case series. PARTICIPANTS: Thirty eyes of 30 consecutive patients with CRVO underwent the procedure, but two were subsequently excluded. The remaining 28 eyes of 28 patients with CRVO for an average of 4.9 months before intervention (range, 0.25-30 months) and best-corrected visual acuity 20/63 or worse were included in the study. INTERVENTION: All patients underwent pars plana vitrectomy with cannulation and infusion of t-PA into a branch retinal vein. MAIN OUTCOME MEASURES: Change in visual acuity and the development of complications such as vitreous hemorrhage and neovascular glaucoma were monitored. RESULTS: Twenty-two of 28 patients (79%) experienced at least one line of visual improvement during the follow-up period (average, 11.8 months; range, 3-24 months), and the same number had this level of improvement at the last follow-up examination. Fifteen patients (54%) gained 3 or more lines of acuity within 6 months after the procedure, and 14 (50%) had acuity at last follow-up at least 3 lines better than baseline acuity (average, 6.8 lines). Seven patients had postoperative vitreous hemorrhages ranging from 1 week to 11 months after the procedure; two cleared spontaneously. One patient had a postoperative retinal detachment from a peripheral retinal break that was repaired successfully with pneumatic retinopexy. No other serious intraoperative or early postoperative complications were noted. CONCLUSIONS: Vitrectomy with retinal vein cannulation and infusion of t-PA is a relatively safe procedure that may improve vision in eyes with CRVO.

Photosensitization of retinal pigment epithelium by protoporphyrin IX.

BACKGROUND: Clinical evidence of injury to the retinal pigment epithelium is an important feature of age-related macular degeneration, but the mechanism of this injury is unknown. Blue-light-dependent activation of the blood-borne photosensitizer protoporphyrin IX is known to produce free radicals which may damage cells and tissues. This study was undertaken to determine the effect of blue light and protoporphyrin IX on retinal pigment epithelial cells in vitro. METHODS: Third-passage porcine retinal pigment epithelial cells were plated in six-well culture plates at 100,000 cells/well and grown to confluence. Retinal pigment epithelial cells were then incubated in culture media with and without 35 micrograms/dl protoporphyrin IX and exposed to low intensity (118 microW/cm2) blue, blue-free, or full-spectrum white light in an irradiating incubator for 16 h on/8 h off cycles for 7 days. Some of the wells were shielded from light (dark controls). Retinal pigment epithelial cells were examined by light microscopy and were trypsinized and counted after 7 days. RESULTS: White light with and without protoporphyrin IX and protoporphyrin IX in dark conditions did not decrease the retinal pigment epithelial cell count significantly. Blue light alone and blue light with protoporphyrin IX decreased the cell count by 22 +/- 4% and 35 +/- 3% compared to the controls, respectively. CONCLUSION: Blue wavelength light without exogenous protoporphyrin IX has a cytotoxic effect on confluent cultures of retinal pigment epithelium, suggesting that endogenous photosensitizers may be present in retinal pigment epithelial cells. Protoporphyrin IX has an additive cytotoxic effect in the presence of blue light, suggesting that this photosensitizer is capable of mediating blue-light-induced retinal pigment epithelial damage. Since protoporphyrin IX is present in blood and tissue fluids, and the retina is chronically exposed to light, protoporphyrin IX-mediated free radical formation may occur in vivo and may play a role in retinal pigment epithelial changes that occur early in the pathogenesis of age-related macular degeneration.

Light-dependent spin trapping of hydroxyl radical from human erythrocytes.

The generation of reactive oxygen species from human erythrocytes has previously been demonstrated. Furthermore, erythrocytic protoporphyrin IX has been shown to generate superoxide and singlet oxygen when exposed to light. These findings suggest that a component of erythrocytic reactive oxygen species production may be light-dependent. By inhibiting erythrocyte superoxide dismutase, catalase, and glutathione peroxidase with N,N-diethyldithiocarbamate or sodium cyanide, we demonstrate the light-dependent generation of hydroxyl radical in human erythrocytes using spin trapping/Electron Spin Resonance spectroscopy. This finding may be significant in tissues where blood is exposed to light, such as in the eye.

Histopathologic examination of vascular patterns in subfoveal neovascular membranes.

BACKGROUND: Subfoveal neovascular membranes cause significant visual loss in age-related macular degeneration (AMD) and the ocular histoplasmosis syndrome. The frequency of post-laser treatment persistence or recurrence of subfoveal membranes in AMD is as high as 51%. The reason for the high incidence of failure after laser treatment is unknown. The authors performed a histopathologic study of subfoveal membranes to determine the distribution of blood vessels within the neovascular complex, and to see if the blood vessel pattern would provide insight into the reason for laser treatment failure. METHODS: The authors used light microscopy to examine serial sections of subfoveal membranes from six patients (4 with AMD, 2 with the ocular histoplasmosis syndrome). The data from this examination were used to create detailed two-dimensional vascular maps of each membrane. RESULTS: The authors found that subfoveal membranes from patients with AMD and the ocular histoplasmosis syndrome, whether occurring de novo or after laser treatment, have a nonuniform distribution of blood vessels, and that large areas which include the membrane margin may be avascular. CONCLUSIONS: Using current laser treatment protocols, it is likely that avascular or poorly perfused peripheral areas of the neovascular complex would be left untreated after laser photocoagulation. Partial treatment of the neovascular complex may contribute to the high rate of post-laser treatment persistence or recurrence of subfoveal membranes.