Evidence for gelsolin as a corneal crystallin in zebrafish

We have shown that gelsolin is one of the most prevalent water-soluble proteins in the transparent cornea of zebrafish. There are also significant amounts of actin. In contrast to actin, gelsolin is barely detectable in other eye tissues (iris, lens, and remaining eye) of the zebrafish. Gelsolin cDNA hybridized intensely in Northern blots to RNA from the cornea but not from the lens, brain, or headless body. The deduced zebrafish gelsolin is approximately 60% identical to mammalian cytosolic gelsolin and has the characteristic six segmental repeats as well as the binding sites for actin, calcium, and phosphatidylinositides. In situ hybridization tests showed that gelsolin mRNA is concentrated in the zebrafish corneal epithelium. The zebrafish corneal epithelium stains very weakly with rhodamine-phalloidin, indicating little F-actin in the cytoplasm. In contrast, the mouse corneal epithelium contains relatively little gelsolin and stains intensely with rhodamine-phalloidin, as does the zebrafish extraocular muscle. We propose, by analogy with the diverse crystallins of the eye lens and with the putative enzyme-crystallins (aldehyde dehydrogenase class 3 and other enzymes) of the mammalian cornea, that gelsolin and actin-gelsolin complexes act as water-soluble crystallins in the zebrafish cornea and contribute to its optical properties.     

Repeated exposure to benzalkonium chloride in the Ex Vivo Eye Irritation Test (EVEIT): observation of isolated corneal damage and healing

The prediction of side-effects is a key issue in the REACH initiative on chemicals, in the production of cosmetics and in the preclinical testing of drugs. A new ex vivo test for repeated substance application is presented, that is able to identify corrosive and irritant effects on the eye by using crucial endpoints, such as cellular and morphological damage, and healing characteristics. The test is intended to replace the Draize eye test and to improve the preclinical testing of drugs and chemicals that are likely to come into direct contact with the cornea. The Ex Vivo Eye Irritation Test (EVEIT) is a self-healing system, involving living corneas obtained from abattoir rabbit eyes. The corneas are cultured in a similar way to the method used during the transplantation of corneal grafts. The corneas are exposed to multiple small, mechanical abrasions, and then test substances are repeatedly dropped onto the centres of the corneas. The test substances applied in this study were citrate-buffered hyaluronate eye drops and an artificial tear replacement, with increasing concentrations of up to 0.1% benzalkonium chloride. A dose-dependent inhibition of recovery and impairment of the lactate production mechanism in the cornea was observed with benzalkonium chloride treatment.     

Relationships between eye factors and lens-forming transformations in the cornea and pericorneal epidermis of larval Xenopus laevis

Larval Xenopus laevis at stage 56 (Nieuwkoop and Faber, '56) were subjected to various types of lentectomy: (1) simple lentectomy, from the pupillary space after incision of outer and inner cornea; (2) lentectomy from the dorsal region of the eye; (3) lentectomy from the dorsal region of the eye and simultaneous incision of the outer cornea; (4) lentectomy from the dorsal region of the eye and simultaneous incision of the outer and inner cornea. The results obtained show that the outer cornea underwent lens-forming transformations only when the inner cornea had been incised, thus permitting outer cornea (Experiments I-IV). No lens regeneration occurred when the inner cornea was left intact (Experiments II, III). It was concluded that the factor(s) allowing the lens-forming transformations of the outer cornea is not an aspecific nutritional factor(s) but a more specific factor(s) that cannot reach the outer cornea when the inner cornea is intact. Therefore, the absence of the lens and sufficient nutrient available to the outer cornea are not enough to allow lens regeneration from the outer cornea. When lens removal was carried out through the dorsal part of the eye (Experiments III-IV) the lens regenerated from the pericorneal epidermis of this region in a large number of cases.     

不同角膜表面处理方式在全飞秒SMILE手术中的应用效果分析

摘要 目的：探讨不同角膜表面处理方式在全飞秒SMILE手术中的应用效果。方法：选择2021年10月至2022年9月来我院择期行全飞秒SMILE手术的患者50例,根据先右后左手术原则,观察眼与对照眼没有固定左右眼,观察眼25例选择右眼,25例选择左眼,对照眼反之选择另一眼,观察眼使用开睑器开睑后用三角海绵擦拭清洁角膜表。对照眼眼自净（眨眼）后使用开睑器开眼睑。对比观察眼与对照眼的角膜光密度、角膜表面颗粒物质计数、角膜表面湿度、术中负压吸引完成时间、失吸比例、OBL发生率,分析50例患者的舒适度。结果：观察眼的角膜光密度为16.33±2.12,对照组为16.85±2.58,组间对比无统计学意义（t=-1.101,P=0.274）。对照眼的表面颗粒物质计数明显较观察眼低,角膜表面湿度明显较观察眼高（P<0.05）。对照眼与观察组术中负压吸引完成时间、失吸比例对比无统计学意义（P>0.05）。观察眼中OBL发生率为6.00 %,对照眼中OBL发生率为2.00 %,组间对比无统计学意义（P=0.617>0.05）。50例患者中对三角海绵擦拭角膜感觉恐惧者占比32.00%（16/50）,三角海绵擦拭角膜后对注视绿点存在影响者占比20.00 %（10/50）,手术中选择三角海绵擦拭角膜者20例,占比40.00 %（20/50）,选择眼自净（眨眼）者30例,占比60.00 %（30/50）。结论：眼自净（眨眼）可以代替三角海绵擦拭角膜,用于全飞秒SMILE手术,提高患者舒适度。     

APCs in the anterior uveal tract do not migrate to draining lymph nodes

The migration of APCs from sites of infection and their maturation are critical elements in the generation of immune responses. However, the paths by which intraocular Ags migrate to draining lymph nodes are not known because the eye has limited lymphatic vessels. To date, only dendritic cells from the cornea and conjunctiva have been shown to emigrate. We demonstrate that phagocytic APCs in the anterior uveal tissues of the murine eye that ingest fluorescent latex beads do not migrate to regional lymph nodes. The beads are ingested in the uveal tract by cells expressing MHC class II, CD11c, or F4/80. Using intravital time-lapse videomicroscopy to monitor iris APC migration after anterior chamber injection of fluorescent Ag, fluorescently labeled APCs fail to move at multiple observation times, even in the presence of Ag and LPS. Whereas an as yet unidentified ocular nonphagocytic APC subset might migrate from the anterior uveal tissues, it is more probable that immune responses in the draining lymph nodes are engendered by soluble Ag escaping the eye through interstitial spaces. The inability of anterior uveal tissue APCs to migrate to lymph nodes may contribute to deviant immune responses that dominate after Ags are introduced into the anterior chamber.     

Interrelationship of the mediator receptors in the terminals of the afferent nerves of the cornea of the eye]

In local application of substances with a mediator action (adrenaline, acetylcholine, histamine) and blockers of specific receptors (dihydroergotoxin, inderal, atropine and of subcutaneous dimedrol) together with dicaine it was possible to reveal a distinct presence in the cornea of the eye of the mediator receptors capable of influencing one another. Pharmacological analysis permitted a supposition to be made that there were two receptor systems in the sensory nerve endings of the cornea; these systems were in reciprocal inhibitory relations; due to this their regular influence on the processes of excitation and inhibition is effected in the sensory nerve ending.     

Relationships between Presence of the Eye Cup and Maintenance of Lens-Forming Capacity in Larval Xenopus laevis

The lentectomized eye of larval Xenopus laevis can regenerate a lens by a process of lens-transdifferentiation of the cornea and pericorneal epidermis. These tissues can form the lens only when they become in direct communication with the environment of the vitreous chamber (neural retina) indicating that the eye cup plays a fundamental role in this process.
In this work the role of the eye cup in the maintainance of the lens-forming capacity of the cornea and pericorneal epidermis was studied by allowing these tissues to cover the enucleated orbit for different periods, and then implanting them into the vitreous chamber of the contralateral eye. Under these experimental conditions the maintainance of the lens-forming capacity of the cornea and pericorneal epidermis showed no significant correlation with the time from enucleation to implantation.     

Transdifferentiation of ocular tissues in larval Xenopus laevis

Transdifferentiation phenomena offer a useful opportunity to study experimentally the mechanisms on which cell phenotypic stability depends. The capacities of vertebrate eye tissues to reprogram cell differentiation are well known in avian and mammalian embryos, and in larval and adult newt. From research into the capacity of anuran eye tissues to reprogram differentiation into a new pathway, considerable data have accumulated concerning the transdifferentiative capacities of eye tissues in larval Xenopus laevis. This work reviews the data concerning the transdifferentiative phenomena of eye tissues in that species and, based on these, aims to establish the extent of our knowledge about the mechanism controlling these processes. In larval Xenopus laevis the outer cornea can regenerate a lens by a lens-transdifferentiation process triggered and substained by a factor(s), probably of a protein nature, produced by the neural retina. In a normal eye phenotypic stability of the outer cornea is guaranteed by the presence of the inner cornea and lens, which prevent the spread of retinal factor(s). The stimulus for lens transdifferentiation of the outer cornea can be supplied by other tissues as well, but this capacity is not widely distributed. The iris and retinal pigmented epithelium can transdifferentiate into neural retina if isolated from the surrounding tissues and implanted in the vitreous chamber. As for lens transdifferentiation of the outer cornea, retinal transdifferentiation of the iris can be stimulated by certain nonocular tissues as well.     

虻复眼感杆光导的模式与适应状态的关系

利用逆向照明方法,不经角膜中和,通过光学显佩镜快速观察和记录了暗适应,短时间明适应和长时间明适应状态下,虻(Atylotus miser Szilady)复眼感杆光导的模式。同时与角膜中和的实验结果作了比较。本文还讨论了憎黄虻复眼角膜透镜的作用以及该复眼对视觉信憎黄息抽样方式与适应状态的关系。     

Post-embryonic photoreceptor development and dark/light adaptation in the spittle bug Philaenus spumarius (L.) (Homoptera, Cercopidae)

The aims of this paper have been to describe (1) the general structure of the compound eye of the spittle bug Philaenus spumarius, (2) the eye's post-embryonic development, (3) photomechanical changes upon dark/light adaptation in the eye, and (4) how leaving the semi-aquatic foam bubble and turning into an adult affects the organization of the eye. Spittle bugs, irrespective of size or sex, possess apposition type compound eyes. The eye's major components (i.e. facet, cornea, cone and rhabdom) grow rather isometrically from the smallest nymph to the adult. Photomechanical changes can occur during both nymphal and adult phases and manifest themselves through pigment granules and mitochondria migrating to and away from the rhabdom, and rhabdom diameters varying with time of day and ambient light level. When a nymph transforms into an adult, its compound eyes’ dorsoventral axes widen, facet diameters increase, facet shapes turn from circular to pentagonal and hexagonal, the cornea thickens and the rhabdoms become thinner. The agile adults, free from the foam that surrounds the nymphs, can be expected to need their vision more than the nymphs, and the changes in eye structure do, indeed, indicate that the adults have superior visual acuity. A thicker cornea in the adults reduces water loss and protects the compound eye from mechanical and light-induced damage: protection given to the nymphs by their foam bubbles.     

A lattice model for computing the transmissivity of the cornea and sclera.

The method of photonic band structure is used to calculate the frequencies of light that propagate in lattice models of the cornea and sclera of the mammalian eye, providing an explanation for transparency in the cornea that first properly accounts for multiple scattering of light. Each eye tissue is modeled as an ordered array of collagen rods, and photonic band structure methods are used to solve Maxwell's equations exactly for these models, a procedure that automatically effectively includes all orders of multiple scattering. These calculations show that the dispersion relation for the cornea is linear in the visible range, implying that the cornea is transparent. We show that the transmissivity is approximately 97% by using an effective medium approximation derived from the photonic band structure results and applicable in the visible region. In contrast, the dispersion relation for the model in the sclera is not linear in the visible region, and there are band gaps in this region that could play an important role in the transmission of light in the sclera.     

胭脂鱼眼早期形态发生研究

采用组织学方法观察了胭脂鱼(Myxocyprinus asiaticus) 眼的发生过程, 结果显示: 胭脂鱼眼的发育经历了眼原基形成期、眼囊形成期、视杯形成期、晶体板形成期、晶体囊形成期、角膜原基形成期、角膜上皮形成期、视网膜细胞增殖期、晶状体成熟期、眼色素形成期以及眼成型期等11个时期。视网膜发育最早, 起始于眼原基的形成, 直至眼成型期分化完成, 形成了厚度不一的8层细胞, 由内向外依次为神经纤维层、神经细胞层、内网层、内核层、外网层、外核层、视杆视锥层和色素上皮层, 且发育历时最长, 约264h。晶状体的发育在视网膜之后, 始于晶体板的形成, 于出膜前期成熟, 发育历时最短, 约74h。角膜发育最晚, 始于角膜原基的形成, 出膜1 d分化为透明的成熟角膜, 发育历时约96h。出膜4 d仔鱼眼色素沉积明显, 视网膜各层分化明显, 晶状体内部完全纤维化, 眼的形态结构基本发育完全。     

Deeper in the human cornea proteome using nanoLC-Orbitrap MS/MS: An improvement for future studies on cornea homeostasis and pathophysiology

The cornea is a transparent, avascular, and highly specialized connective tissue that provides the majority of light refraction in the optical system of the eye. The human cornea is composed of several layers interacting in a complex manner and possessing specific functions, like eye protection and optical clearness. Only few proteomic studies of mammalian cornea have been performed leading to the identification of less than 200 proteins in human corneas. The present study explores the proteome of the intact normal human cornea using a shot-gun nanoLC-MS/MS strategy and an LTQ Orbitrap mass spectrometer. A total of 2070 distinct corneal proteins were identified from five human cornea samples, which represents a 14-fold improvement in the number of proteins identified so far for human cornea. This enlarged dataset of human corneal proteins represents a valuable reference library for further studies on cornea homeostasis and pathophysiology. Network and gene ontology analyses were used to determine biological pathways specific of the human cornea. They allowed the identification of subnetworks of putative importance for corneal diseases, like a redox regulation and oxidative stress network constituted of aldehyde and alcohol dehydrogenases, most of them being described for the first time in human cornea.     

Experimental analysis of lens-forming capacity in Xenopus borealis larvae

Previously, the only anuran amphibians known to have the capacity to regenerate a lens after lentectomy were Xenopus laevis and Xenopus tropicalis. This regeneration process occurs during the larval life through transdifferentiation of the outer cornea promoted by inductive factors produced by the retina and accumulated inside the vitreous chamber. However, the capacity of X. tropicalis to regenerate a lens is much lower than that of X. laevis. This study demonstrates that Xenopus borealis, a species more closely related to X. laevis than to X. tropicalis, is not able to regenerate a lens after lentectomy. Nevertheless, some morphological modifications corresponding to the first stages of lens regeneration in X. laevis were observed in the outer cornea of X. borealis. This suggested that in X borealis the regeneration process was blocked at early stages. Results from histological analysis of X. borealis and X. laevis lentectomized eyes and from implantation of outer cornea fragments into the vitreous and anterior chambers demonstrated that: (i) in X. borealis eye, the lens-forming competence in the outer cornea and inductive factors in the vitreous chamber are both present, (ii) no inhibiting factors are present in the anterior chamber, the environment where lens regeneration begins, (iii) the inability of X. borealis to regenerate a lens after lentectomy is due to an inhibiting action exerted by the inner cornea on the spreading of the retinal factor from the vitreous chamber towards the outer cornea. This mechanical inhibition is assured by two distinctive features of X. borealis eye in comparison with X. laevis eye: (i) a weaker and slower response to the retinal inducer by the outer cornea; (ii) a stronger and faster healing of the inner cornea. Unlike X. tropicalis and similar to X. laevis, in X. borealis the competence to respond to the retinal factor is not restricted to the corneal epithelium but also extends to the pericorneal epidermis.     

Ultrastructural immuno-localization of tropoelastin in the chick eye

Summary Immuno-gold labeling at the electron-microscopy level was used to investigate the distribution of tropoelastin in the chick eye. Intense staining was found in the amorphous part of mature elastic fibers in different regions of the organ. In elaunin fibers, both the amorphous core and the surrounding microfibrils were clearly labeled. In addition, reactive sites were detected in the oxitalan fibers of the stroma of the cornea and in Descemet's membrane, which showed a gradient of reactive sites increasing from the center toward the periphery. Oxitalan fibers of the stroma often fused with Descemet's membrane; the pattern of immunological staining suggested a continuity between the two structures. In the ciliary zonule, labeling for tropoelastin was observed in discrete areas on the bundles of microfibrils. The results show a complex structural organization of elastic tissue; this may be important in endowing the various parts of the eye with different mechanical properties.     

The transparent lens and cornea in the mouse and zebra fish eye

The lens and cornea combine to form a single optical element in which transparency and refraction are the fundamental biophysical characteristics required for a functional visual system. Although lens and cornea have different cellular and extracellular specializations that contribute to transparency and refraction, their development is closely related. In the embryonic mouse, the developing cornea and lens separate early. In contrast, zebra fish lens and cornea remain connected during early development and the optical properties of the cornea and lens observed by slit lamp and quasielastic laser light scattering spectroscopy (QLS) are more similar in the zebra fish eye than in the mouse eye. Optical similarities between cornea and lens of zebra fish may be the result of similarities in the cellular development of the cornea and lens.     

Detection of photogenotoxicity in skin and eye in rat with the photo comet assay

Photosensitizing drugs increase the sensitivity of the skin and the eye toward normally harmless sunlight conditions and are known to enhance the induction of skin tumors or severe injuries to the eye. The photogenotoxicity of five common drugs (sparfloxacin, dacarbazine, chlorpromazine and 8-methoxypsoralen, promazine) was investigated in the skin as well as in the retina and cornea of Wistar rats. The compounds were administered once orally by gavage and the resulting DNA damage was analyzed in the newly developed in vivo photo comet assay. All drugs except of promazine were clearly photogenotoxic in the skin. In the cornea sparfloxacin and dacarbazine induced an increased DNA damage following irradiation. A photogenotoxic effect in the retina was observed by sparfloxacin, which is the only compound tested that absorbs wavelengths reaching the retina. The drug concentration analysis revealed that the compounds were distributed into plasma, skin and eye at concentrations, which were photogenotoxic in vitro. Additionally, histopathological analysis showed no relevant alterations or inductions of necrosis, apoptosis or inflammation in the skin or eye. In conclusion, we confirmed the photogenotoxic potential of compounds from different chemical classes in the skin. Moreover, it is the first time that photogenotoxicity has been detected in the retina and cornea in an in vivo study. Based on our results it is concluded that the photo comet assay in rat is an easy and reliable method to elucidate drug induced photogenotoxicity under conditions, which are relevant to human exposure.     

The fine structure of the pallial eyes of Laternula truncata (Bivalvia: Anomalodesmata: Pandoracea)

The structure of the pallial eyes of Laternula truncata (Lamarck 1818) has been studied using the light and electron microscopes. The eye is complex and can be- considered to be- the most advanced yet described for a bivalve mollusc. The cornea consists of modified flattened epithelial cells with an external border of microvilli. The cornea covers a large, circular, multinucleate lens. The lens comprises (1) centrally located translucent lens cells, (2) laterally located supporting cells from which cell processes interdigitate with processes from the lens cells. The retina is two layered and inverted. The proximal and distal retinae are made up of concentrically arranged laminae derived from the membranes of ciliary basal bodies. The cilia comprise a base and feet, but no root system and have a 9+0 arrangement of filaments.
The pigment cup or tapetum is bounded by a sclerotic coat and is three layered, each layer possessing characteristic pigment granules. From the base of the eye arises a large optic nerve.
The eye possesses an eye appendage, the epithelium of which is invaginated on its internal border to form a groove within which are found some 28 cilia. The cilia, it is thought, make contact with the microvilli of the epithelium when the appendage is touched. Such an action serves to protect the delicate eye from damage.
The structure of the eye is compared with that of other molluscs, particularly members of the Bivalvia.     

Size-Dependent Diffusion of Dextrans in Excised Porcine Corneal Stroma

Delivery of therapeutic agents to the eye requires efficient transport through cellular and extracellular barriers. We evaluated the rate of diffusive transport in excised porcine corneal stroma using fluorescently labeled dextran molecules with hydrodynamic radii ranging from 1.3 to 34 nm. Fluorescence correlation spectroscopy (FCS) was used to measure diffusion coefficients of dextran molecules in the excised porcine corneal stroma. The preferential sensitivity of FCS to diffusion along two dimensions was used to differentially probe diffusion along the directions parallel to and perpendicular to the collagen lamellae of the corneal stroma. In order to develop an understanding of how size affects diffusion in cornea, diffusion coefficients in cornea were compared to diffusion coefficients measured in a simple buffer solution. Dextran molecules diffuse more slowly in cornea as compared to buffer solution. The reduction in diffusion coefficient is modest however (67% smaller), and is uniform over the range of sizes that we measured. This indicates that, for dextrans in the 1.3 to 34 nm range, the diffusion landscape of corneal stroma can be represented as a simple liquid with a viscosity approximately 1.5 times that of water. Diffusion coefficients measured parallel vs. perpendicular to the collagen lamellae were indistinguishable. This indicates that diffusion in the corneal stroma is not highly anisotropic. Our results support the notion that the corneal stroma is highly permeable and isotropic to transport of hydrophilic molecules and particles with hydrodynamic radii up to at least 34 nm.