Zika Virus Infection of Human Iris Pigment Epithelial Cells

During recent Zika epidemics, adults infected with Zika virus (ZIKV) have developed organ-specific inflammatory complications. The most serious Zika-associated inflammatory eye disease is uveitis, which is commonly anterior in type, affecting both eyes and responding to corticosteroid eye drops. Mechanisms of Zika-associated anterior uveitis are unknown, but ZIKV has been identified in the aqueous humor of affected individuals. The iris pigment epithelium is a target cell population in viral anterior uveitis, and it acts to maintain immune privilege within the anterior eye. Interactions between ZIKV and human iris pigment epithelial cells were investigated with infectivity assays and RNA-sequencing. Primary cell isolates were prepared from eyes of 20 cadaveric donors, and infected for 24 hours with PRVABC59 strain ZIKV or incubated uninfected as control. Cytoimmunofluorescence, RT-qPCR on total cellular RNA, and focus-forming assays of culture supernatant showed cell isolates were permissive to infection, and supported replication and release of infectious ZIKV. To explore molecular responses of cell isolates to ZIKV infection at the whole transcriptome level, RNA was sequenced on the Illumina NextSeq 500 platform, and results were aligned to the human GRCh38 genome. Multidimensional scaling showed clear separation between transcriptomes of infected and uninfected cell isolates. Differential expression analysis indicated a vigorous molecular response of the cell to ZIKV: 7,935 genes were differentially expressed between ZIKV-infected and uninfected cells (FDR < 0.05), and 99% of 613 genes that changed at least two-fold were up-regulated. Reactome and KEGG pathway and Gene Ontology enrichment analyses indicated strong activation of viral recognition and defense, in addition to biosynthesis processes. A CHAT network included 6275 molecular nodes and 24 contextual hubs in the cell response to ZIKV infection. Receptor-interacting serine/threonine kinase 1 (RIPK1) was the most significantly connected contextual hub. Correlation of gene expression with read counts assigned to the ZIKV genome identified a negative correlation between interferon signaling and viral load across isolates. This work represents the first investigation of mechanisms of Zika-associated anterior uveitis using an in vitro human cell model. The results suggest the iris pigment epithelium mounts a molecular response that limits intraocular pathology in most individuals.

Use of automatic refractometer infrared images to screen pigment dispersion syndrome: a cross-sectional observational study from a preliminary hypothesis

Background: This study was performed to evaluate the use of anterior segment images, obtained with an automatic refractometer, to identify early defects of the iris pigment epithelium in patients with pigment dispersion syndrome (PDS) or pigmentary glaucoma (PG) without observable alterations at the slit lamp. Methods: In this cross-sectional observational study, carried out from January 2018 to December 2019, in Policlinico Citta di Udine Health Center, Udine, Italy, we observed anterior segment infrared images of 1700 subjects who were undergoing routine ophthalmological examination using an automatic refractometer. We selected infrared images of subjects who fulfilled the inclusion and exclusion criteria and looked for a focal defect in the iris pigment epithelium. Results: Twenty patients with focal iris pigment epithelial defect were identified and none of them showed evident signs of PDS. After the necessary explanations, they agreed to have further examinations to verify the possibility of PDS. An in-depth evaluation of ocular structures, including gonioscopy, demonstrated the presence of PDS in all subjects with iris defects. Conclusions: The use of infrared images obtained by an automatic refractometer could provide early and easy identification of PDS in crowded ophthalmology clinics or mass screening programs; yet, more well-designed studies are necessary to confirm these preliminary findings and prove this proposed screening tool.

Time Course Changes of the Mechanical Properties of the Iris Pigment Epithelium in a Rat Chronic Ocular Hypertension Model

Background. The flow field of aqueous humor correlates to the stiffness of iris pigment epithelium (IPE) which acts as a wall of posterior chamber. We focus on the variations of IPE stiffness in a rat ocular hypertension (OHT) model, so as to prepare for exploring the mechanism of duration of OHT. Methods. Episcleral venous cauterization (EVC) was applied on one eye of male adult Sprague-Dawley rats to induce chronic high intraocular pressure. According to the duration of OHT (0, 1, 2, 4, and 8 weeks), rats were randomly divided into Gw0, Gw1, Gw2, Gw4, and Gw8. Atomic force microscope (AFM) analysis was applied to test IPE stiffness in three regions: iris root, mid-periphery, and pupillary-margin in each group. Histological changes of IPE were also examined in Gw4 and Gw8. Results. There was an overall growing tendency of IPE stiffness in EVC eye. IPE in EVC eye was significantly stiffer than fellow eye in Gw2, Gw4, and Gw8 (in iris root, mid-periphery, and pupillary-margin, p<0.05). IPE in EVC eye in pupillary-margin was significantly stiffer than iris root in Gw4 and Gw8 (p<0.05). In EVC eye, IPE becomes thinner and IPE cell density decreases. Conclusion. IPE stiffness increases gradually with the duration of chronic high intraocular pressure.