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TEM of amacrine and müller cells in the inner plexiform layer. High magnification TEM of an amacrine cell (A) and a Müller cell (B) located in the middle aspect of the inner plexiform layer. Note the lucent cytoplasm (asterisks), dispersed nucleoplasm, and small nucleolus (arrow) in the amacrine cell (A) and the extensive network of relatively electron dense cytoplasmic processes (arrowhead) of the Müller cell wrapping around other cytoplasmic processes in the inner plexiform layer. These cell types, in addition to endothelial cells and fibrous astrocytes, appeared to correspond to a discontinuous relative hypo-reflective band in the inner plexiform layer. Scale bars: A, 2 microns; B, 5 microns.
Source publication
The data presented in this article are related to the research paper entitled “Correlation of Spectral Domain Optical Coherence Tomography with Histology and Electron Microscopy in the Porcine Retina” (Xie et al., 2018) [2]. This research data highlights our technique for retinal fundus image acquisition during spectral domain optical coherence tom...
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Citations
Purpose
To investigate the effect of stanniocalcin-1 (STC-1), a secreted polypeptide exhibiting multiple functions in cell survival and death, on photoreceptor degeneration in a porcine model of retinitis pigmentosa (RP).
Methods
P23H transgenic pigs (TG P23H) and wild-type hybrid littermates were obtained from the National Swine Resource and Research Center. Human recombinant STC-1 was injected intravitreally every 2 weeks from postnatal day 15 (P15) to P75. The contralateral eye was injected with balanced salt solution as a control. Electroretinography (ERG) and spectral domain optical coherence tomography (SD-OCT) were performed to evaluate retinal function and morphology in vivo at P90. Retinal tissue was collected for histologic analysis and molecular assays to evaluate the anti-oxidative and anti-inflammatory mechanisms by which STC-1 may rescue photoreceptor degeneration.
Results
Intravitreal injection of STC-1 improved retinal function in TG P23H pigs with increased photopic and flicker ERG a- and b-wave amplitudes. Greater integrity of the ellipsoid zone (EZ) band on SD-OCT and morphologic rescue with preservation of cone photoreceptors were observed in STC-1 treated TG P23H pigs. STC-1 altered gene expression in TG P23H pig retina on microarray analysis and increased photoreceptor specific gene expression by RT-PCR analysis. STC-1 significantly decreased oxidative stress and the expressions of NLRP3 inflammasome, cleaved caspase-1, and IL-1β in TG P23H pig retina.
Conclusions
Intravitreal administration of STC-1 enhances cone photoreceptor function, improves EZ integrity, and reduces retinal degeneration through anti-oxidative and anti-inflammatory effects in a large animal (pig) model of the most common form of autosomal dominant RP in the United States.
The laser-induced choroidal neovascularization (CNV) model has been widely used for research on wet age-related macular degeneration (wet-AMD) and other ocular neovascular diseases. In this model, the Bruch membrane is perforated by laser injury, resulting in neovascularization formed from the choroidal capillaries. It has become a standard method to evaluate the effect of different treatments on CNV progression in preclinical studies. This protocol can be used in various species, including rat, mouse, pig, and monkey. The rodent laser-induced CNV model is the most commonly used because of the advantages in both cost- and time-efficiency. It takes only 10–15 min to complete the whole laser procedure after adequate training and practicing the technique. Peak CNV formation occurs at approximately 2 weeks after laser application. The entire protocol may require up to 3 weeks to complete the treatment, fundus image acquisition, and tissue collection for histologic analysis. This chapter describes the detailed procedures, protocols, and useful notes on how to induce CNV by laser.
Purpose:
Endothelin-1 (ET-1) is a potent vasoactive factor implicated in development of diabetic retinopathy, which is commonly associated with retinal edema and hyperglycemia. Although the vasomotor activity of venules contributes to the regulation of tissue fluid homeostasis, responses of human retinal venules to ET-1 under euglycemia and hyperglycemia remain unknown and the ET-1 receptor subtype corresponding to vasomotor function has not been determined. Herein, we addressed these issues by examining the reactivity of isolated human retinal venules to ET-1, and results from porcine retinal venules were compared.
Methods:
Retinal tissues were obtained from patients undergoing enucleation. Human and porcine retinal venules were isolated and pressurized to assess diameter changes in response to ET-1 after exposure to 5 mM control glucose or 25 mM high glucose for 2 hours.
Results:
Both human and porcine retinal venules exposed to control glucose developed similar basal tone and constricted comparably to ET-1 in a concentration-dependent manner. ET-1-induced constrictions of human and porcine retinal venules were abolished by ETA receptor antagonist BQ123. During high glucose exposure, basal tone of human and porcine retinal venules was unaltered but ET-1-induced vasoconstrictions were enhanced.
Conclusions:
ET-1 elicits comparable constriction of human and porcine retinal venules by activation of ETA receptors. In vitro hyperglycemia augments human and porcine retinal venular responses to ET-1.
Translational relevance:
Similarities in vasoconstriction to ET-1 between human and porcine retinal venules support the latter as an effective model of the human retinal microcirculation to help identify vascular targets for the treatment of retinal complications in patients with diabetes.
Purpose:
To evaluate different methods of studying cone photoreceptor structure in wild-type (WT) and transgenic pigs carrying the human rhodopsin P23H mutant gene (TgP23H).
Methods:
For in vivo imaging, pigs were anesthetized with tiletamine-zolazepam and isoflurane and given lidocaine-bupivacaine retrobulbar injections. Stay sutures and a custom head mount were used to hold and steer the head for adaptive optics scanning light ophthalmoscopy (AOSLO). Six WT and TgP23H littermates were imaged at postnatal day 30 (P30), P90, and P180 with AOSLO and optical coherence tomography (OCT), and two additional sets of littermates were imaged at P3 and P15 with OCT only. AOSLO imaging and correlative differential interference contrast microscopy were performed on a P240 WT pig and on WT and TgP23H littermates at P30 and P180.
Results:
AOSLO cone density generally underestimates histology density (mean difference ± SD = 24.8% ± 21.4%). The intensity of the outer retinal hyperreflective OCT band attributed to photoreceptors is attenuated in TgP23H pigs at all ages. In contrast, AOSLO images show cones that retain inner and outer segments through P180. At retinal locations outside the visual streak, TgP23H pigs show a heterogeneous degenerating cone mosaic by using two criteria: variable contrast on a split detector AOSLO and high reflectivity on a confocal AOSLO.
Conclusions:
AOSLO reveals that the cone mosaic is similar to ex vivo histology. Its use as a noninvasive tool will enable observation of morphologic changes that arise in the cone mosaic of TgP23H pigs over time.
Translational relevance:
Pigs are widely used for translational studies, and the ability to noninvasively assess cellular changes in the cone mosaic will facilitate more detailed investigations of new retinal disease models as well as outcomes of potential therapies.
Spectral domain optical coherence tomography (SD-OCT) is used as a non-invasive tool for retinal morphological assessment in vivo. Information on the correlation of SD-OCT with retinal histology in the porcine retina, a model resembling the human retina, is limited. Herein, we correlated the hypo- and hyper-reflective bands on SD-OCT with histology of the lamellar architecture and cellular constituents of the porcine retina. SD-OCT images were acquired with the Heidelberg Spectralis HRA + OCT. Histological analysis was performed using epoxy resin embedded tissue and transmission electron microscopy. Photomicrographs from the histologic sections were linearly scaled to correct for tissue shrinkage and correlated with SD-OCT images. SD-OCT images correlated well with histomorphometric data. A hyper-reflective band in the mid-to-outer inner nuclear layer correlated with the presence of abundant mitochondria in horizontal cell processes and adjacent bipolar cells. A concentration of cone nuclei corresponded to a relative hypo-reflective band in the outer portion of the outer nuclear layer. The presence of 3 hyper-reflective bands in the outer retina corresponded to: 1) the external limiting membrane; 2) the cone and rod ellipsoid zones; and 3) the interdigitation zone of photoreceptor outer segments/retinal pigment epithelium (RPE) apical cell processes and the RPE. These correlative and normative SD-OCT data may be employed to characterize and assess the in vivo histologic changes in retinal vascular and degenerative diseases and the responses to novel therapeutic interventions in this large animal model.
