A2E induces IL-1ß production in retinal pigment epithelial cells via the NLRP3 inflammasome.

AIMS: With ageing extracellular material is deposited in Bruch's membrane, as drusen. Lipofuscin is deposited in retinal pigment epithelial cells. Both of these changes are associated with age related macular degeneration, a disease now believed to involve chronic inflammation at the retinal-choroidal interface. We hypothesise that these molecules may act as danger signals, causing the production of inflammatory chemokines and cytokines by the retinal pigment epithelium, via activation of pattern recognition receptors. METHODS: ARPE-19 cells were stimulated in vitro with the following reported components of drusen: amyloid-ß (1-42), Carboxyethylpyrrole (CEP) modified proteins (CEP-HSA), Nε-(Carboxymethyl)lysine (CML) modified proteins and aggregated vitronectin. The cells were also stimulated with the major fluorophore of lipofuscin: N-retinylidene-N-retinylethanolamine (A2E). Inflammatory chemokine and cytokine production was assessed using Multiplex assays and ELISA. The mechanistic evaluation of the NLRP3 inflammasome pathway was assessed in a stepwise fashion. RESULTS: Of all the molecules tested only A2E induced inflammatory chemokine and cytokine production. 25 µM A2E induced the production of significantly increased levels of the chemokines IL-8, MCP-1, MCG and MIP-1α, the cytokines IL-1ß, IL-2, IL-6, and TNF-α, and the protein VEGF-A. The release of IL-1ß was studied further, and was determined to be due to NLRP3 inflammasome activation. The pathway of activation involved endocytosis of A2E, and the three inflammasome components NLRP3, ASC and activated caspase-1. Immunohistochemical staining of ABCA4 knockout mice, which show progressive accumulation of A2E levels with age, showed increased amounts of IL-1ß proximal to the retinal pigment epithelium. CONCLUSIONS: A2E has the ability to stimulate inflammatory chemokine and cytokine production by RPE cells. The pattern recognition receptor NLRP3 is involved in this process. This provides further evidence for the link between A2E, inflammation, and the pathogenesis of AMD. It also supports the recent discovery of NLRP3 inflammasome activation in AMD.

Delivery of anti-angiogenic molecular therapies for retinal disease.

Angiogenic diseases of the retina are the leading cause of blindness in the developed world. The development of anti-angiogenic molecular therapies has transformed the prognosis of these conditions, especially age-related macular degeneration. With these new treatments comes the new challenge of delivering an effective dosage to the retina, over a prolonged period of time and in a safe and cost-effective manner. A range of new anti-angiogenics are on the horizon, offering new and varied modes of drug delivery. In addition, a range of new sustained-release drug delivery technologies are being developed.

Periocular autologous Coleman fat graft survival and histopathology.

PURPOSE: Autologous Coleman fat transfer is used in the periocular area and intraorbitally for soft-tissue volume augmentation in postenucleation socket syndrome. This article aims to identify areas of low fat absorption and the histopathology of excised fat. METHODS: Noncomparative case series of patients who underwent excision/debulking of injected autologous fat in the inferior sulcus/lower eyelid. RESULTS: Five of 20 patients with postenucleation socket syndrome who had received intraorbital and sulcus Coleman fat transfer, required fat reduction (sculpting) from the inferior sulcus and lower eyelid due to persistent "overfill" 8 months to 17 months after fat transfer. Between 0.2 ml and 1.4 ml of fat was removed. The excised fat was pale, rubbery, and compact in comparison with the normal lower eyelid fat pad. Histopathology showed healthy adipose cells but with chronic inflammation and fibrous septa. CONCLUSIONS: Fat survival is greater than anticipated in the lower eyelid and inferior sulcus. Therefore, judicious use in these areas is suggested. The grafted fat is distinguishable from the fat in the lower eyelid both clinically and histopathologically.

Human transscleral albumin permeability and the effect of topographical location and donor age.

PURPOSE: To quantify the permeability coefficient of albumin across human sclera and to assess topographical and age-related variation. METHODS: Equatorial superotemporal scleral tissue from 15 donor eyes (mean age 60 years; range 39-84) was mounted in a modified Ussing chamber. Additional tissue was taken from the anterior and posterior superotemporal regions of six eyes, and equatorial superonasal, and inferotemporal regions of a further six eyes. Fluorescein isothiocyanate (FITC)-labeled, 0.412 mM, bovine albumin was placed in one hemichamber facing the internal scleral surface, and the rate of transscleral flux was determined over 24 hours, at 25 degrees C, with a spectrophotometer. RESULTS: Permeability coefficient for equatorial superotemporal scleral tissue at 25 degrees C (+/-SD) was 0.83 +/- 0.50 x 10(-6) cm . s(-1). The permeability coefficient adjusted for 37 degrees C (+/-SD) was 1.43 +/- 0.86 x 10(-6) cm . s(-1). The effect of donor age was assessed for the 15 equatorial superotemporal samples. Regression analysis showed a significant decline in scleral diffusion of albumin with increasing donor age (P = 0.0166). There was no significant difference in diffusion over the different topographical regions tested. The partition coefficient of permeability to albumin also showed a decline with increasing donor age (P = 0.001). CONCLUSIONS: The permeability and partition coefficients of human sclera both significantly decline with increasing donor age. Permeability coefficient shows no significant variation over the different topographical regions tested. The decrease in albumin permeability with increasing donor age may have pharmacokinetic implications when considering transscleral diffusion of high-molecular-weight compounds.