Quantitative Optical Coherence Tomography for Longitudinal Monitoring of Postnatal Retinal Development

A better study of the postnatal retinal development is not only essential for the in-depth understanding of the nature of the vision system but also may provide insights for treatment developments of eye conditions, such as retinopathy of premature (ROP). To date, quantitative analysis of postnatal retinal development is primarily limited to endpoint histological examination. This study is to validate in vivo optical coherence tomography (OCT) for longitudinal monitoring of postnatal retinal development in developing mouse eyes. Three-dimensional (3D) frame registration and super averaging were adopted to investigate the fine structure of the retina. Interestingly, a hyporeflective layer (HRL) between the nerve fiber layer (NFL) and inner plexiform layer (IPL) was observed in developing eyes and gradually disappeared with aging. To interpret the observed retinal layer kinetics, a model based on eyeball expansion, cell apoptosis, and retinal structural modification was proposed.

Characterization of retinal development in 13-lined ground squirrels

Purpose: The cone-dominant, 13-lined ground squirrel (13-LGS) retina mimics the human foveal region but retinal development in this useful rodent species has not been reported. Here, the embryonic and postnatal development of the 13-LGS retina was studied to further characterize the species as a practical alternative animal model for investigating cone-based vision in health and disease. Methods: The spatiotemporal expression of key progenitor and cell type markers was examined in retinas from defined embryonic and postnatal stages using immunohistochemistry. Changes in the postnatal gene expression were also assessed by qPCR. Results: The 13-LGS neuroblastic layer expressed key progenitor markers (Sox2, Vsx2, Pax6, and Lhx2) at E18. Sequential cell fate determination evidenced by the first appearance of cell type-specific marker labeling was: at E18, ganglion cells (Brn-3A, HuC/D) and microglia (Iba1); at E24-25.5 shortly before birth, photoreceptor progenitor (Otx2, Recoverin), horizontal and amacrine cells (Lhx1, Oc1); and at P15, bipolar cells (Vsx1, CaBP5) and Muller glia cells (GS, Rlbp1). Photoreceptor maturation indicated by opsin+ outer segments and PNA labeling of cone sheaths was completed at the time of eye opening, P21-24. Conclusions: The timeline and order of retinal cell development in the 13-LGS generally matches that recorded from other mammalian models but with a stark variation in the proportion of various cell types due to cone-dense photoreceptors. This provides a baseline for future examinations of developmental, disease model, and stem cell approach studies employing this emerging rodent model of human vision.

Embryonic hyperglycemia perturbs the development of specific retinal cell types, including photoreceptors

Diabetes is linked to various long-term complications in adults, such as neuropathy, nephropathy, and diabetic retinopathy. Diabetes poses additional risks for pregnant women, because glucose passes across the placenta, and excess maternal glucose can result in diabetic embryopathy. While many studies have examined the teratogenic effects of maternal diabetes on fetal heart development, little is known about the consequences of maternal hyperglycemia on the development of the embryonic retina. To address this question, we investigated retinal development in two models of embryonic hyperglycemia in zebrafish. Strikingly, we found that hyperglycemic larvae displayed a significant reduction in photoreceptors and horizontal cells, whereas other retinal neurons were not affected. We also observed reactive gliosis and abnormal optokinetic responses in hyperglycemic larvae. Further analysis revealed delayed retinal cell differentiation in hyperglycemic embryos that coincided with increased reactive oxygen species (ROS). Our results suggest that embryonic hyperglycemia causes abnormal retinal development via altered timing of cell differentiation and ROS production, which is accompanied by visual defects. Further studies using zebrafish models of hyperglycemia will allow us to understand the molecular mechanisms underlying these effects.

Essential functions of MLL1 and MLL2 in retinal development and cone cell maintenance

MLL1 (KMT2A) and MLL2 (KMT2B) are homologous members of the mixed-lineage leukemia (MLL) family of histone methyltransferases involved in epigenomic transcriptional regulation. Their sequence variants have been associated with neurological and psychological disorders, but little is known about their roles and mechanism of action in CNS development. Using mouse retina as a model, we previously reported the roles of MLL1 in retinal neurogenesis and horizontal cell maintenance. Here we determine roles of MLL2 and MLL1/MLL2 together in retinal development using conditional knockout (CKO) mice. Deleting Mll2 from Chx10+ retinal progenitors resulted in a similar phenotype as Mll1 CKO, but removal of both alleles produced much more severe deficits than each single CKO: 1-month double CKO mutants displayed null light responses in electroretinogram; thin retinal layers, including shorter photoreceptor outer segments with impaired phototransduction gene expression; and reduced numbers of M-cones, horizontal and amacrine neurons, followed by fast retinal degeneration. Despite moderately reduced progenitor cell proliferation at P0, the neurogenic capacity was largely maintained in double CKO mutants. However, upregulated apoptosis and reactive gliosis were detected during postnatal retinal development. Finally, the removal of both MLLs in fated rods produced a normal phenotype, but the CKO in M-cones impaired M-cone function and survival, indicating both cell non-autonomous and autonomous mechanisms. Altogether, our results suggest that MLL1/MLL2 play redundant roles in maintaining specific retinal neurons after cell fate specification and are essential for establishing functional neural networks.

The role of interferon regulatory factor 8 for retinal tissue homeostasis and development of choroidal neovascularisation

Background Microglia cells represent the resident innate immune cells of the retina and are important for retinal development and tissue homeostasis. However, dysfunctional microglia can have a negative impact on the structural and functional integrity of the retina under native and pathological conditions. Methods In this study, we examined interferon-regulatory factor 8 (Irf8)–deficient mice to determine the transcriptional profile, morphology, and temporospatial distribution of microglia lacking Irf8 and to explore the effects on retinal development, tissue homeostasis, and formation of choroidal neovascularisation (CNV). Results Our study shows that Irf8-deficient MG exhibit a considerable loss of microglial signature genes accompanied by a severely altered MG morphology. An in-depth characterisation by fundus photography, fluorescein angiography, optical coherence tomography and electroretinography revealed no major retinal abnormalities during steady state. However, in the laser-induced CNV model, Irf8-deficient microglia showed an increased activity of biological processes critical for inflammation and cell adhesion and a reduced MG cell density near the lesions, which was associated with significantly increased CNV lesion size. Conclusions Our results suggest that loss of Irf8 in microglia has negligible effects on retinal homeostasis in the steady state. However, under pathological conditions, Irf8 is crucial for the transformation of resident microglia into a reactive phenotype and thus for the suppression of retinal inflammation and CNV formation.

Widespread translational control regulates retinal development in mouse

Retinal development is tightly regulated to ensure the generation of appropriate cell types and the assembly of functional neuronal circuitry. Despite remarkable advances have been made in understanding regulation of gene expression during retinal development, how translational regulation guides retinogenesis is less understood. Here, we conduct a comprehensive translatome and transcriptome survey to the mouse retinogenesis from the embryonic to the adult stages. We discover thousands of genes that have dynamic changes at the translational level and pervasive translational regulation in a developmental stage-specific manner with specific biological functions. We further identify genes whose translational efficiencies are frequently controlled by changing usage in upstream open reading frame during retinal development. These genes are enriched for biological functions highly important to neurons, such as neuron projection organization and microtubule-based protein transport. Surprisingly, we discover hundreds of previously uncharacterized micropeptides, translated from putative long non-coding RNAs and circular RNAs. We validate their protein products in vitro and in vivo and demonstrate their potentials in regulating retinal development. Together, our study presents a rich and complex landscape of translational regulation and provides novel insights into their roles during retinogenesis.