Transient Retention of Photoreceptor Outer Segments in Matrigel-Embedded Retinal Organoids.

Retinal organoids (ROs) are three-dimensional retinal tissues, which are differentiated in vitro from induced pluripotent stem cells (iPSC), ultimately forming all main retinal cell types under defined culture conditions. ROs show several highly specialized retinal features, including the outgrowth of photoreceptor outer segments (OSs). In vivo, the photoreceptor OSs are enveloped and maintained by protrusions of retinal pigment epithelium (RPE) cells, the so-called apical microvilli, while ROs fail to recapitulate this critical interaction in culture development. Here, we define specific co-culture conditions aiming to compensate for the missing physical proximity of RPE and OSs in RO development. Accordingly, functional RPE cells and ROs were differentiated simultaneously from the same iPSC clone, the former resulting in byproduct RPE or bRPE cells. While some co-culture approaches indicated a temporary functional interaction between bRPE and RO photoreceptors, they did not improve the photoreceptor histoarchitecture. In contrast, embedding ROs in a basement membrane extract without bRPE cells showed a robust improvement in the rate of photoreceptor OS retention. RO embedding is a quick and easy method that greatly enhances the preservation of photoreceptor OSs, an important structure for modelling retinal diseases with the involvement of photoreceptors.

A Circulating MicroRNA Profile in a Laser-Induced Mouse Model of Choroidal Neovascularization.

Choroidal neovascularization (CNV) is a pathological process in which aberrant blood vessels invade the subretinal space of the mammalian eye. It is a characteristic feature of the prevalent neovascular age-related macular degeneration (nAMD). Circulating microRNAs (cmiRNAs) are regarded as potentially valuable biomarkers for various age-related diseases, including nAMD. Here, we investigated cmiRNA expression in an established laser-induced CNV mouse model. Upon CNV induction in C57Bl/6 mice, blood-derived cmiRNAs were initially determined globally by RNA next generation sequencing, and the most strongly dysregulated cmiRNAs were independently replicated by quantitative reverse transcription PCR (RT-qPCR) in blood, retinal, and retinal pigment epithelium (RPE)/choroidal tissue. Our findings suggest that two miRNAs, mmu-mir-486a-5p and mmur-mir-92a-3p, are consistently dysregulated during CNV formation. Furthermore, in functional in vitro assays, a significant impact of mmu-mir-486a-5p and mmu-mir-92a-3p on murine microglial cell viability was observed, while mmu-mir-92a-3p also showed an impact on microglial mobility. Taken together, we report a robust dysregulation of two miRNAs in blood and RPE/choroid after laser-induced initiation of CNV lesions in mice, highlighting their potential role in pathology and eventual therapy of CNV-associated complications.

An Eye on Age-Related Macular Degeneration: The Role of MicroRNAs in Disease Pathology.

Age-related macular degeneration (AMD) is the primary cause of blindness in developed countries, and is the third leading cause worldwide. Emerging evidence suggests that beside environmental and genetic factors, epigenetic mechanisms, such as microRNA (miRNA) regulation of gene expression, are relevant to AMD providing an exciting new avenue for research and therapy. MiRNAs are short, non-coding RNAs thought to be imperative for coping with cellular stress. Numerous studies have analyzed miRNA dysregulation in AMD patients, although with varying outcomes. Four studies which profiled dysregulated circulating miRNAs in AMD yielded unique sets, and there is only minimal overlap in ocular miRNA profiling of AMD. Mouse models of AMD, including oxygen-induced retinopathy and laser-induced choroidal neovascularization, showed similarities to some extent with miRNA patterns in AMD. For example, miR-146a is an extensively researched miRNA thought to modulate inflammation, and was found to be upregulated in AMD mice and cellular systems, but also in human AMD retinae and vitreous humor. Similarly, mir-17, miR-125b and miR-155 were dysregulated in multiple AMD mouse models as well as in human AMD plasma or retinae. These miRNAs are thought to regulate angiogenesis, apoptosis, phagocytosis, and inflammation. A promising avenue of research is the modulation of such miRNAs, as the phenotype of AMD mice could be ameliorated with antagomirs or miRNA-mimic treatment. However, before meaningful strides can be made to develop miRNAs as a diagnostic or therapeutic tool, reproducible miRNA profiles need to be established for the various clinical outcomes of AMD.