Delivery of Topically Applied Calpain Inhibitory Peptide to the Posterior Segment of the Rat Eye

<div><p>We developed an inhibitory peptide that specifically acts against mitochondrial μ-calpain (Tat-μCL, 23 amino acid, 2857.37 Da) and protects photoreceptors in retinal dystrophic rats. In the present study, we topically administered Tat-μCL to the eyes of Sprague-Dawley rats for 7 days to determine both the delivery route of the peptide to the posterior segment of the eye and the kinetics after topical application in adult rats. Distribution of the peptide was determined by immunohistochemical analysis, and enzyme-linked immune-absorbent assay was used to quantify the accumulation in the retina. Peptides were prominently detected in both the anterior and posterior segments of the eye at 1 h after the final eye drop application. Immunohistochemically positive reactions were observed in the retina, optic nerve, choroid, sclera and the retrobulbar tissues, even in the posterior portion of the eye. Immunoactivities gradually diminished at 3 and 6 h after the final eye drop. Quantitative estimations of the amount of peptide in the retina were 15.3, 5.8 and 1.0 pg/μg protein at 1, 3 and 6 h after the final instillation, respectively. Current results suggest that while the topically applied Tat-μCL peptide reaches the posterior segment of the retina and the optic nerve, the sufficient concentration (> IC50) is maintained for at least 6 h in the rat retina. Our findings suggest that delivery of topically applied peptide to the posterior segment and optic nerve occurs through the conjunctiva, periocular connective tissue, sclera and optic nerve sheath.</p></div

Distribution of topically instilled Tat-μCL in the anterior segment of the rat eye at 1 h (upper) and 3 h (lower) after the final eye drop application.

<p>Peptides are detected in the cornea, sclera, iris, ciliary body, choroid and RPE at the 1 h time point. The peptides also accumulated in the retina at the 1 h time point. The immunoreactivity weakened at the 3 h time point. DAPI: 4μ 6-diamidino-2-phenylindole. White bars indicate a 200 μm length. Results are representative of three independent experiments [n = 3 eyes (3 rats)].</p

Chronological histological changes in the photoreceptor layer during the early stages (PN days 18 to 28; hematoxylin and eosin stained) of retinal degeneration in RCS^-/- rats.

<p>Arrows indicate the extracellular lamellar material deposited on the apical surface of the RPE (PN days 18 and 23) and in the photoreceptor OS layer (PN days 25 and 28). Arrowheads indicate the photoreceptor IS layer (PN days 18 to 28). The black bar represents 50μm length.</p

Chronological changes in the photoreceptor IS and OS layers during the early stages (PN days 17 to 26) of retinal degeneration in RCS^-/- rat—observed using optical coherent tomography and the corresponding histological sections (PN days 18, 23, and 28).

<p>A yellow arrow indicates the apical hyperreflective band and a light green arrow indicates the IS ellipsoid zone. Black arrows indicate the extracellular lamellar material. Arrowheads indicate the photoreceptor inner segment. The black and white bars represent 100μm length.</p

Chronological changes in the amplitudes of the ERG a- (a) and b- (b) waves in RCS rats.

<p>Closed circles indicate RCS^-/- rats and open circles indicate RCS^+/+ rats. Statistical significance (with Bonferroni’s <i>post hoc</i> test): * <i>P</i> < 0.05; *** <i>P</i> < 0.001.</p

Chronological changes in the retinal layers during the progressive stage (PN days 33 to 47) of RCS^-/- rat retinal degeneration—observed using optical coherent tomography and the corresponding histological sections (PN days 33 and 46).

<p>Dark green arrows indicate the outer nuclear layer. An orange arrow indicates the photoreceptor IS and OS layers. Yellow arrows indicate the inner retinal layer. White arrows indicate the retinal pigment epithelium and choroid. The black and white bars represent 100μm length.</p

Optical Coherence Tomography of Retinal Degeneration in Royal College of Surgeons Rats and Its Correlation with Morphology and Electroretinography

<div><p>Purpose</p><p>To evaluate the correlation between optical coherence tomography (OCT) and the histological, ultrastructural and electroretinography (ERG) findings of retinal degeneration in Royal College of Surgeons (RCS^-/-) rats.</p><p>Materials and Methods</p><p>Using OCT, we qualitatively and quantitatively observed the continual retinal degeneration in RCS^-/- rats, from postnatal (PN) day 17 until PN day 111. These findings were compared with the corresponding histological, electron microscopic, and ERG findings. We also compared them to OCT findings in wild type RCS^+/+ rats, which were used as controls.</p><p>Results</p><p>After PN day 17, the hyperreflective band at the apical side of the photoreceptor layer became blurred. The inner segment (IS) ellipsoid zone then became obscured, and the photoreceptor IS and outer segment (OS) layers became diffusely hyperreflective after PN day 21. These changes correlated with histological and electron microscopic findings showing extracellular lamellar material that accumulated in the photoreceptor OS layer. After PN day 26, the outer nuclear layer became significantly thinner (<i>P</i> < 0.01) and hyperreflective compared with that in the controls; conversely, the photoreceptor IS and OS layers, as well as the inner retinal layers, became significantly thicker (<i>P</i> < 0.001 and <i>P</i> = 0.05, respectively). The apical hyperreflective band, as well as the IS ellipsoid zone, gradually disappeared between PN day 20 and PN day 30; concurrently, the ERG a- and b-wave amplitudes deteriorated. In contrast, the thicknesses of the combined retinal pigment epithelium and choroid did not differ significantly between RCS^-/- and RCS^+/+ rats.</p><p>Conclusion</p><p>Our results suggest that OCT demonstrates histologically validated photoreceptor degeneration in RCS rats, and that OCT findings partly correlate with ERG findings. We propose that OCT is a less invasive and useful method for evaluating photoreceptor degeneration in animal models of retinitis pigmentosa.</p></div

Macular Hole Caused by Retained Subfoveal Perfluorocarbon that Subsequently Closed After Its Spontaneous Resolution: A Case Report

<p><b>Article full text</b></p> <p><br></p> <p>The full text of this article can be found here<b>.</b> <a href="https://link.springer.com/article/10.1007/s40123-017-0107-5">https://link.springer.com/article/10.1007/s40123-017-0107-5</a></p><p></p> <p><br></p> <p><b>Provide enhanced content for this article</b></p> <p><br></p> <p>If you are an author of this publication and would like to provide additional enhanced content for your article then please contact <a href="http://www.medengine.com/Redeem/”mailto:[email protected]”"><b>[email protected]</b></a>.</p> <p><br></p> <p>The journal offers a range of additional features designed to increase visibility and readership. All features will be thoroughly peer reviewed to ensure the content is of the highest scientific standard and all features are marked as ‘peer reviewed’ to ensure readers are aware that the content has been reviewed to the same level as the articles they are being presented alongside. Moreover, all sponsorship and disclosure information is included to provide complete transparency and adherence to good publication practices. This ensures that however the content is reached the reader has a full understanding of its origin. No fees are charged for hosting additional open access content.</p> <p><br></p> <p>Other enhanced features include, but are not limited to:</p> <p><br></p> <p>• Slide decks</p> <p>• Videos and animations</p> <p>• Audio abstracts</p> <p>• Audio slides</p

Typical OCT image of a RCS^+/+ rat at PN day 33, along with a corresponding histological section of a RCS^+/+ rat at PN day 29 (hematoxylin and eosin stained).

<p>The retinal pigment epithelium (RPE) was detached. The top yellow line indicates the retinal surface, the second line indicates the border between the inner nuclear layer and outer plexiform layer, the third line indicates the upper limit of the IS ellipsoid line, the fourth line indicates the surface of the RPE, and the bottom line indicates the bottom of the choroid. Layer A comprises the nerve fiber layer, the ganglion cell layer, the inner plexiform layer and the inner nuclear layer. Layer B comprises the outer plexiform layer and the outer nuclear layer. Layer C comprises the photoreceptor IS and OS layers. Layer D comprises the RPE and the choroid. The black and white bars represent 100μm length.</p

Characterization of photoreceptor degeneration in the rhodopsin P23H transgenic rat line 2 using optical coherence tomography

<div><p>Purpose</p><p>To characterize the optical coherence tomography (OCT) appearances of photoreceptor degeneration in the rhodopsin P23H transgenic rat (line 2) in relation to the histological, ultrastructural, and electroretinography (ERG) findings.</p><p>Materials and methods</p><p>Homozygous rhodopsin P23H transgenic albino rats (line 2, very-slow degeneration model) were employed. Using OCT (Micron IV^®; Phoenix Research Labs, Pleasanton, CA, USA), the natural course of photoreceptor degeneration was recorded from postnatal day (P) 15 to P 287. The OCT images were qualitatively observed by comparing them to histological and ultrastructural findings at P 62 and P 169. In addition, each retinal layer was quantitatively analyzed longitudinally during degeneration, compared it to that observed in wild type Sprague-Dawley (SD) rats. The relationships between the ERG (full-field combined rod-cone response, 3.0 cds/m² stimulation) findings and OCT images were also analyzed.</p><p>Results</p><p>In the qualitative study, the two layers presumably corresponding to the photoreceptor inner segment ellipsoid zone (EZ) and interdigitation zone (IZ) were identified in the P23H rat until PN day 32. However, the photoreceptor inner and outer segment (IS/OS) layer became diffusely hyperreflective on OCT after P 46, and the EZ and IZ zones could no longer be identified on OCT. In contrast, in the SD rats, the EZ and IZ were clearly distinguished until at least P 247. The ultrastructural study showed partial disarrangements of the photoreceptor outer segment discs in the P23H rats at P 62, although a light-microscopic histological study detected almost no abnormality in the outer segment. In the quantitative study, the outer retinal layer including the outer plexiform layer (OPL) and the outer nuclear layer (ONL) became significantly thinner in the P23H rats than in the SD rats after P 71. The thickness of the IS/OS layer was maintained in the P23H rats until P 130, and it became statistically thinner than in the SD rats at P 237. The longitudinal attenuation in the amplitude of the a- and b-waves of ERG was significantly correlated with the thickness of the combined OPL and ONL but not with that of the IS/OS layer.</p><p>Conclusion</p><p>OCT showed the degenerated photoreceptor IS/OS layer in rhodopsin P23H transgenic rats (line 2) as a diffuse hyperreflective zone, even in the early stage, with the partially disarranged and destabilized OS discs recognizable by ultrastructural assessment but not by a histological study. The amplitude of the a- and b-waves mainly depends on the thickness of the OPL and ONL layer rather than the thickness of the photoreceptor IS/OS layer in P23H rats.</p></div