Ex vivo demonstration of canine corneal pre-Descemet's anatomy using pneumodissection as for the big bubble technique for deep anterior lamellar keratoplasty.

The recent discovery and characterization of pre-Descemet's layer (PDL; also termed the Dua's layer or the Dua-Fine layer) has advanced the understanding of various posterior corneal pathologies and surgeries in human. This study aimed to characterize the ultrastructure of the posterior stroma and interfacial zone of Descemet's membrane (DM) in canine eyes. Eighteen canine corneo-scleral discs were included. Intrastromal air injection resulted in the formation of type 1 big bubble (BB) in 73% (n = 11/15) of corneas, with a mean diameter of 11.0 ± 1.3 mm. No type 2 BB was created. Anterior segment optical coherence tomography, histology and transmission electron microscopy confirmed that the wall of BB was composed of DM, in contact with remaining stroma (canine PDL; cPDL). The cPDL was populated with keratocytes, of varying thickness of 16.2 ± 4.2 µm in close apposition to the DM, and composed of collagen bundles arranged in transverse, longitudinal and oblique directions. The interfacial zone, between DM and cPDL, showed fibril extension in all three directions, predominantly longitudinal. Irregular extensions of DM material into cPDL stroma were observed. No long-spaced collagen was detected. In conclusion, there exists a well-defined cleavage plane between the posterior stroma and cPDL, with similar but not identical characteristics as in humans, that is revealed by pneumodissection. This adds to our understanding of the anatomy of the posterior most canine cornea, which will have significant clinical impact on posterior corneal surgery and understanding of corneal pathology in dogs.

Morphogenetic events influencing corneal maturation, development, and transparency: Light and electron microscopic study.

The development of the cornea is a fascinating process. Its dual origin involves the differentiation of surface ectoderm cells and the migration of mesenchymal cells of neural crest origin. This research aimed to demonstrate the morphogenesis of the rabbit cornea from fetal to postnatal life using light- and electron microscopy, and immunohistochemical analysis. There were 27 rabbit embryos and nine rabbits used. The rabbit cornea begins its prenatal development on the twelfth day of gestation. The surface ectoderm differentiates into the corneal epithelium on day 13. Intriguingly, telocytes were visible within the epithelium. The secondary stroma develops on the sixteenth day of gestation by differentiation of keratocytes. At the age of 2 weeks, the lamellae of collagenous fibers become highly organized, and the stroma becomes avascular, indicating that the cornea has become transparent. Bowman's membrane appears on day 23 of pregnancy and disappears on day 30. The Descemet's membrane appears at this time and continues to thicken postnatally. The corneal endothelium appears on the twentieth gestational day as double layer of flattened cells and becomes a single layer of cuboidal cells on day 30. The spaces between the endothelial cells resemble craters. VEGF immunohistochemical expression increases over the course of development, reaching its peak in the first week after birth before decreasing in all corneal layers and becoming negative in the stroma. In conclusion, numerous morphogenetic events contribute to corneal maturation and transparency, allowing the cornea to perform its vital functions.

Ultrastructure of the dorsal ocellus of Bittacus planus larvae (Mecoptera: Bittacidae) with evolutionary significance.

The Bittacidae are unique in holometabolous insects in that their larvae bear a dorsal ocellus on the frons. The fine structure of the dorsal ocellus, however, has not been investigated to date. Here, the ultrastructure of the larval dorsal ocellus was studied in the hangingfly Bittacus planus Cheng, 1949 using light, scanning, and transmission electron microscopy. The dorsal ocellus of the larvae comprises a cornea, corneagenous cells, and retinula cells. The cornea is a laminated structure. A layer of corneagenous cells is located below the cornea. Numerous retinula cells are arranged tightly beneath the corneagenous cells. The retinula cells modify their adjacent membranes into numerous linear microvilli, which form an analogue of the rhabdom among adjacent retinula cells. The results show that the dorsal ocellus of larval Bittacidae is a highly vestigial organ and appears to be degenerating during the postembryonic development. The presence of the vestigial dorsal ocellus is likely to represent an ancestral plesiomorphy of holometabolous insects, providing new evidence for exploring the evolutionary origin of holometabolous larvae.

Comparative study of two re-embedding methods on the ultrastructure of corneal tissue.

PURPOSE: To expand the routine of pathological diagnostics of surgical keratoplasty specimens via transmission electron microscopy. The target was to identify the best re-embedding method for optimal structural preservation of formalin fixed paraffin embedded (FFPE) corneal tissue re-embedded into resin for ultrastructural analysis. BASIC PROCEDURES: Bovine FFPE corneal tissue was re-embedded into resin with either a rapid osmium-free four-hour-method or a four-day-routine-method known from nephropathology, compared with primary resin embedded bovine corneal tissue. The analysis involved the ultrastructure of cytoplasm, the intercellular interfaces of superficial epithelial cells, deepest basal epithelial cells and corneal endothelial cells, cell matrix interfaces, Bowman layer, corneal stroma, its microfibril bundles and Descemet membrane. MAIN FINDINGS: The main observation was the equally reduced preservation status of re-embedded FFPE corneal tissue independent of the used re-embedding method. This extends to the intercellular contacts of superficial epithelial cells and the apical tight junctions of corneal endothelial cells. Hemidesmosomal cell matrix contacts showed less demarcation in re-embedded specimens. Cell matrix interfaces of Bowman layer and Descemet membrane were more clearly bordered in primary resin embedded than re-embedded tissue. In contrast, gap junctions in re-embedded tissue were detected in deepest basal epithelial cells and corneal endothelial cells with comparable preservation to primary resin embedding. Bowman layer, corneal stromal extracellular matrix, its microfibril bundles and Descemet membrane showed equal ultrastructural preservation in all evaluated methods. PRINCIPAL CONCLUSION: Corneal tissue can be successfully analysed with transmission electron microscopy after a rapid osmium-free four hour re-embedding procedure from FFPE material. A comparable morphology with primary resin embedded material can be obtained for gap junctions of deepest basal epithelial cells and corneal endothelial cells, further for Bowman layer, corneal stromal extracellular matrix, its microfibril bundles and Descemet membrane.

Morphological adaptations on the eye of the golden gray mullet (Chelon aurata): Using light and scanning electron microscopical study.

The present investigations were designed to describe the ultrastructural properties of the eye of the golden gray mullet (Chelon aurata). For this purpose, the eyes were examined grossly, and by light and electron microscope. The external layer consists of the cornea and the sclera. Three layers compose the cornea; the anterior stratified cuboidal epithelial; the anterior limiting (Bowman) membrane; and the thick dermal layer of the stroma. The mucoidal layer has small collagen fiber bundles embedded in the CT layer and located between the anterior portion of the scleral cornea and the dermal stroma, (or "substancia propria"). The iridescent layer is thin at the center and thick at the periphery. It contains a pigmented layer with many ossicles. SEM analysis reveals that the cornea consists of undetermined shaped cells joined together by numerous thread-like micro-ridges, with several micro-tubercles on the external surface. The photoreceptor layer had two types of cells: the rod-shaped and the cone-shaped cells. The cone cells differentiate into two types of cells: single and double cells. SEM analysis of the retina showed that rod cells appear as thin long uniform rod-like, while the cone cells appear as rod cells with ovoid bases. SEM analysis demonstrates that the inner side of the retinal epithelium appears to be wrapped around itself. The morphological appearance of the eye adapts to life in superficial aquatic conditions. In conclusion, the current findings provide morphologic evidence to better understand the mechanism of fish vision adaptation to environmental conditions. RESEARCH HIGHLIGHTS: The transparent cornea composed of three layers; anterior stratified cuboidal epithelial, Bowman's membrane, and a thick dermal stromal layer. The mucoidal layer is formed from small collagen fibers bundles embedded in the CT layer and located between the anterior portion of the scleral cornea and the dermal stroma. There are two types of photoreceptor cells: rod and cone cells. Cone cells can be single and double cells.

A unique pre-endothelial layer at the posterior peripheral cornea: ultrastructural study.

This study was conducted to investigate the ultrastructure of a unique structures at the anterior side of the endothelium of the posterior peripheral cornea and compare their inner fibers to those of the limbus and sclera. The unique structures at the anterior side of endothelium was referred as a pre-endothelial (PENL) structures in the present manuscript. Ten anonymous-donor human corneoscleral rims (leftover after corneal transplants) were processed for electron microscopy. Semi-thin sections were examined using an Olympus BX53 microscope, and ultrathin sections were studied using a JOEL 1400 transmission electron microscope. A unique PENL structures was identified at the posterior peripheral cornea at a radial distance of approximately 70-638 µm, from the endpoint of Descemet's membrane. The PENL thinned out gradually and disappeared in the center. The contained an electron-dense sheath with periodic structures (narrow-spacing fibers), wide-spacing fibers, and numerous microfibrils. Typical elastic fibers were present in the sclera and limbus but were not observed in the PENL. This study revealed the existence of a new acellular PENL, containing unique fibrillar structures that were unseen in the corneal stroma. From the evidence describe in this paper we therefore suggest that PENL is a distinct morphological structure present at the corneal periphery.

Fibrosis Is a Basement Membrane-Related Disease in the Cornea: Injury and Defective Regeneration of Basement Membranes May Underlie Fibrosis in Other Organs.

Every organ develops fibrosis that compromises functions in response to infections, injuries, or diseases. The cornea is a relatively simple, avascular organ that offers an exceptional model to better understand the pathophysiology of the fibrosis response. Injury and defective regeneration of the epithelial basement membrane (EBM) or the endothelial Descemet's basement membrane (DBM) triggers the development of myofibroblasts from resident corneal fibroblasts and bone marrow-derived blood borne fibrocytes due to the increased entry of TGF beta-1/-2 into the stroma from the epithelium and tears or residual corneal endothelium and aqueous humor. The myofibroblasts, and disordered extracellular matrix these cells produce, persist until the source of injury is removed, the EBM and/or DBM are regenerated, or replaced surgically, resulting in decreased stromal TGF beta requisite for myofibroblast survival. A similar BM injury-related pathophysiology can underly the development of fibrosis in other organs such as skin and lung. The normal liver does not contain traditional BMs but develops sinusoidal endothelial BMs in many fibrotic diseases and models. However, normal hepatic stellate cells produce collagen type IV and perlecan that can modulate TGF beta localization and cognate receptor binding in the space of Dissé. BM-related fibrosis is deserving of more investigation in all organs.

The EP2 agonist, omidenepag, alters the physical stiffness of 3D spheroids prepared from human corneal stroma fibroblasts differently depending on the osmotic pressure.

The objective of the current study was to examine the drug-induced effects of the EP2 agonist, omidenapag (OMD), on human corneal stroma, two- and three-dimensional (2D and 3D) cultures of human corneal stroma fibroblasts (HCSFs). The drug-induced effects on 2D monolayers and 3D spheroids were characterized by examining the ultrastructures by scanning electron microscope (SEM), transendothelial electrical resistance (TEER) measurements, and fluorescein isothiocyanate (FITC)-dextran permeability. The physical properties of 3D spheroids with respect to size and stiffness were also examined. In addition, the gene expressions of extracellular matrix (ECM) molecules, including collagen (COL) 1, 4, and 6, and fibronectin (FN), a tissue inhibitor of metalloproteinase (TIMP) 1-4, matrix metalloproteinase (MMP) 2, 9, and 14, aquaporin1 (AQP1), and several endoplasmic reticulum (ER) stress-related factors were evaluated. In the 2D HCSFs, OMD induced (1) a significant increase in ECM deposits, as evidenced by SEM, the mRNA expression of COL4 and FN, and (2) a decrease in TEER values and a concentration-dependent increase in FITC-dextran permeability. In the case of 3D spheroids, OMD had no effect on size but a substantial increase in stiffness was observed. Furthermore, such OMD-induced effects on stiffness were dramatically modulated by the osmotic pressure of the system. In contrast to the above 2D cultures, among the ECM molecules and the modulators of 3D spheroids, namely, TIMPS and MMPs, the down-regulation of COL1, TIMP1 and 2 and the up-regulation of MMP9 were observed. Interestingly, such diversity in terms of OMD-induced gene expressions between 2D and 3D cultures was also recognized in AQP1 (2D; no significant change, 3D; significant up-regulation) and ER stress-related genes. The findings presented herein suggest that the EP2 agonist, OMD, alters the physical stiffness of 3D spheroids obtained from human corneal stroma fibroblasts and this alteration is dependent on the osmotic pressures. 2D and 3D cell cultures may be useful for evaluating the drug induced effects of OMD toward human corneal stroma.

Manual DALK in Keratoconus: An Ex Vivo Light and Transmission Electron Microscopy Analysis 2 Years After Surgery.

PURPOSE: The aim of this study was to evaluate the microscopic structure of a human cornea 2 years after manual deep anterior lamellar keratoplasty (DALK) for keratoconus with a recipient residual stromal bed thickness of 100 µm, using light and transmission electron microscopy. METHODS: A human cornea treated with manual DALK for keratoconus 2 years before was removed during penetrating keratoplasty because of stromal opacity of unknown origin, involving about half of the sample. The transparent half of the specimen was processed for light and transmission electron microscopy. RESULTS: Light microscopy examination performed with different staining techniques (hematoxylin and eosin, Picrosirius red, and Masson trichrome) revealed a homogeneous stroma. No interface was detected. Electron microscopy confirmed these findings. CONCLUSIONS: This study confirmed the available clinical and confocal studies that show progressive stromal remodeling after manual DALK. Two years after surgery, no posterior stromal interface was detected.

Analysis of Cryopreservation Protocols and Their Harmful Effects on the Endothelial Integrity of Human Corneas.

Corneal cryopreservation can partially solve the worldwide concern regarding donor cornea shortage for keratoplasties. In this study, human corneas were cryopreserved using two standard cryopreservation protocols that are employed in the Tissue Bank of the Teresa Herrera Hospital (Spain) to store corneas for tectonic keratoplasties (TK protocol) and aortic valves (AV protocol), and two vitrification protocols, VS55 and DP6. Endothelial viability and general corneal state were evaluated to determine the protocol that provides the best results. The potential corneal cryopreservation protocol was studied in detail taking into consideration some cryopreservation-related variables and the endothelial integrity and stroma arrangement of the resulting cryopreserved corneas. TK corneas showed mostly viable endothelial cells, while the others showed few (AV) or none (DP6 and VS55). The corneal structure was well maintained in TK and AV corneas. TK corneas showed endothelial acellular areas surrounded by injured cells and a normal-like stromal fiber arrangement. Cryoprotectant solutions of the TK protocol presented an increasing osmolality and a physiological pH value. Cooling temperature rate of TK protocol was of 1 °C/min to -40 °C and 3 °C/min to -120 °C, and almost all of dimethyl sulfoxide left the tissue after washing. Future studies should be done changing cryopreservation-related variables of the TK protocol to store corneas of optical grade.

Corneal in vivo confocal microscopy to detect belantamab mafodotin-induced ocular toxicity early and adjust the dose accordingly: a case report.

BACKGROUND: New targeted antibody-drug conjugates (ADCs) against multiple myeloma are known to induce adverse effects that may lead to treatment discontinuation. Preclinical studies reported early severe ocular damage related to the use of belantamab mafodotin (belamaf), including ocular surface inflammation, severe dry eye, and a specific toxicity to the cornea, namely microcystic keratopathy. While belamaf-induced ocular changes have not been prospectively studied, a better understanding of mechanisms involved as well as kinetics may aid in anticipating dose adjustment rather than stopping the treatment once clinical ocular damage is too severe. CASE PRESENTATION: A 61-year-old woman scheduled for belamaf as a fifth-line treatment against multiple myeloma was prospectively included. Clinical examinations were performed before and every 3 weeks afterward, together with in vivo confocal microscopy (IVCM) of the cornea. Visual acuity, symptoms, slit-lamp examination, and ultrastructural changes of the cornea were recorded according to the received dose of belamaf. More precisely, kinetics, shape, density, and location of the toxic corneal lesions have been followed and analyzed using IVCM. Also, specific lesions at the sub-basal nerve plexus layer were detected and characterized for the first time. This advanced approach allowed a better understanding of the belamaf-induced toxicity, further balancing the dose to maintain good vision and eye health while continuing the treatment. CONCLUSIONS: Systematic ultrastructural analysis and follow-up of the corneal state during ADCs treatment for multiple myeloma may open new avenues in the therapeutic approach. Early preclinical detection of ocular damage may accurately contribute to finding the correct dose for each patient and not stopping the treatment due to severe ocular adverse effects.

Collagen XIV Is an Intrinsic Regulator of Corneal Stromal Structure and Function.

Collagen XIV is poorly characterized in the body, and the current knowledge of its function in the cornea is limited. The aim of the current study was to elucidate the role(s) of collagen XIV in regulating corneal stromal structure and function. Analysis of collagen XIV expression, temporal and spatial, was performed at different postnatal days (Ps) in wild-type C57BL/6 mouse corneal stromas and after injury. Conventional collagen XIV null mice were used to inquire the roles that collagen XIV plays in fibrillogenesis, fibril packing, and tissue mechanics. Fibril assembly and packing as well as stromal organization were evaluated using transmission electron microscopy and second harmonic generation microscopy. Atomic force microscopy was used to assess stromal stiffness. Col14a1 mRNA expression was present at P4 to P10 and decreased at P30. No immunoreactivity was noted at P150. Abnormal collagen fibril assembly with a shift toward larger-diameter fibrils and increased interfibrillar spacing in the absence of collagen XIV was found. Second harmonic generation microscopy showed impaired fibrillogenesis in the collagen XIV null stroma. Mechanical testing suggested that collagen XIV confers stiffness to stromal tissue. Expression of collagen XIV is up-regulated following injury. This study indicates that collagen XIV plays a regulatory role in corneal development and in the function of the adult cornea. The expression of collagen XIV is recapitulated during wound healing.

Mutation-induced dimerization of transforming growth factor-ß-induced protein may drive protein aggregation in granular corneal dystrophy.

Protein aggregation in the outermost layers of the cornea, which can lead to cloudy vision and in severe cases blindness, is linked to mutations in the extracellular matrix protein transforming growth factor-ß-induced protein (TGFBIp). Among the most frequent pathogenic mutations are R124H and R555W, both associated with granular corneal dystrophy (GCD) characterized by the early-onset formation of amorphous aggregates. The molecular mechanisms of protein aggregation in GCD are largely unknown. In this study, we determined the crystal structures of R124H, R555W, and the lattice corneal dystrophy-associated A546T. Although there were no changes in the monomeric TGFBIp structure of any mutant that would explain their propensity to aggregate, R124H and R555W demonstrated a new dimer interface in the crystal packing, which is not present in wildtype TGFBIp or A546T. This interface, as seen in both the R124H and R555W structures, involves residue 124 of the first TGFBIp molecule and 555 in the second. The interface is not permitted by the Arg124 and Arg555 residues of wildtype TGFBIp and may play a central role in the aggregation exhibited by R124H and R555W in vivo. Using cross-linking mass spectrometry and in-line size exclusion chromatography-small-angle X-ray scattering, we characterized a dimer formed by wildtype and mutant TGFBIps in solution. Dimerization in solution also involves interactions between the N- and C-terminal domains of two TGFBIp molecules but was not identical to the crystal packing dimerization. TGFBIp-targeted interventions that disrupt the R124H/R555W crystal packing dimer interface might offer new therapeutic opportunities to treat patients with GCD.

Displacement between anterior chamber width obtained by swept-source anterior segment optical coherence tomography and white-to-white distance.

PURPOSE: To determine the relationship between the external limbal location, represented by white-to-white (WTW) distance, and the actual angle location, represented by spur-to-spur (STS) and angle-to-angle (ATA) distances. METHODS: 166 eyes from 166 participants were imaged using CASIA2 anterior chamber optical coherence tomography (AS-OCT) and LenStar LS 900 optical biometer. The horizontal ATA and STS were measured using the swept-source Fourier-domain AS-OCT (CASIA2). The horizontal WTW was automatically measured using LenStar. The displacement lengths (DL) between WTW-STS and WTW-ATA were calculated. Bland-Altman plots and intraclass correlation were performed. RESULTS: The study showed that WTW has a positive correlation with STS (ICC = 0.82, p<0.001) and ATA (ICC = 0.82, p<0.001). The Bland-Altman analysis demonstrated that the mean difference of WTW-STS is 0.10 mm (95% CI 0.06 to 0.14 mm) with limits of agreement of -0.42 to 0.63 mm between WTW and STS, and the mean difference of WTW-ATA is 0.10 mm (95% CI 0.06 to 0.15 mm) with limits of agreement of -0.48 to 0.64 mm between WTW and ATA. Linear regression with adjustment showed that a WTW value greater than 12.07 mm is associated with a greater DL (WTW-STS DL ß 0.18, p = 0.003; WTW-ATA DL ß 0.14, p = 0.03). CONCLUSIONS: Greater WTW was significantly associated with higher displacement of WTW from the two distances representing anterior chamber width. External limbal location may not accurately represent the actual angle location in eyes with larger WTW.

Functional morphology of the cornea of the Little Penguin Eudyptula minor (Aves).

The cornea is a specialized component of the vertebrate eye that provides protection, refractive power, transparency for optical imaging and mechanical support. However, the corneas of birds have received little attention with no comprehensive study of their functional morphology. Using light microscopy and both scanning and transmission electron microscopy, the first description of the ultrastructure of all of the main components of the cornea in two different-sized individuals of the Little Penguin Eudyptula minor is presented. Two types of microprojections protrude from the surface of the cornea with a predominance of microridges and microvilli found in central (flattened) and peripheral regions, respectively. Epithelial cell density is higher in peripheral cornea, especially in the larger (older) individual, while there is a reduction of epithelial cell density with age. The cornea comprises a thick epithelium uniquely attached to the basement membrane with numerous incursions rather than anchoring fibres and anchoring plaques as is found in other vertebrate corneas. Posterior to Bowman's layer, the orthogonally-arranged collagen fibril lamellae in the stroma form extensive branches and anastomoses. Desçemet's membrane is well-developed with an anterior or foetal portion with long banding. However, the thickness of Desçemet's membrane is larger in the older individual with the inclusion of an additional irregular pale-staining posterior portion. Polygonal endothelial cells extend across the cornea as a monolayer with often tortuous cell junctions. Endothelial cell density increases towards the periphery, but decreases with age. Primary cilia are observed protruding through the central region of some endothelial cells into the anterior segment but subsurface structures resembling cilia suggest that these features may be more common. The ultrastructure of the corneal components reveals a range of functional adaptations that reflect the amphibious lifestyle of this seabird.

Comparison of porcine corneal decellularization methods and importance of preserving corneal limbus through decellularization.

BACKGROUND: The aim of this study is to compare the three previously applied, conventional porcine corneal decellularization methods and to demonstrate the importance of preserving the corneal limbus through decellularization. METHODS: Fresh, wild-type (with or without) limbus porcine corneas were decellularized using three different methods, including (i) sodium dodecyl sulfate (SDS), (ii) hypertonic saline (HS), and (iii) N2 gas (NG). Post-treatment evaluation was carried out using histological, residual nuclear material, and ultrastructural analyses. Glycerol was used to help reduce the adverse effects of decellularization. The corneas were preserved for two weeks in cornea storage medium. RESULTS: All three decellularization methods reduced the number of keratocytes at different rates in the stromal tissue. However, all methods, except SDS, resulted in the retention of large numbers of cells and cell fragments. The SDS method (0.1% SDS, 48h) resulted in almost 100% decellularization in corneas without limbus. Low decellularization capacity of the NG method (<50%) could make it unfavorable. Although HS method had a more balanced damage-decellularization ratio, its decellularization capacity was lower than SDS method. Preservation of the corneoscleral limbus could partially prevent structural damage and edema, but it would reduce the decellularization capacity. CONCLUSION: Our results suggest that SDS is a very powerful decellularization method, but it damages the cornea irreversibly. Preserving the corneoscleral limbus reduces the efficiency of decellularization, but also reduces the damage.

Safety profiles of terahertz scanning in ophthalmology.

Terahertz (THz) technology has emerged recently as a potential novel imaging modality in biomedical fields, including ophthalmology. However, the ocular biological responses after THz electromagnetic exposure have not been investigated. We conducted a rabbit study to evaluate the safety profiles of THz scanning on eyes, at a tissue, cellular, structural and functional level. Eight animals (16 eyes) were analysed after excessive THz exposure (control, 1 h, 4 h, and 1 week after continuous 4-h exposure; THz frequency = 0.3 THz with continuous pulse generated at 40 µW). We found that at all the time points, the corneas and lens remained clear with no corneal haze or lens opacity formation clinically and histopathologically. No thermal effect, assessed by thermographer, was observed. The rod and cone cell-mediated electroretinography responses were not significantly altered, and the corneal keratocytes activity as well as endothelial viability, assessed by in-vivo confocal microscopy, was not affected. Post-exposed corneas, lens and retinas exhibited no significant changes in the mRNA expression of heat shock protein (HSP)90AB1), DNA damage inducible transcript 3 (DDIT3), and early growth response (EGR)1. These tissues were also negative for the inflammatory (CD11b), fibrotic (fibronectin and α-smooth muscle actin), stress (HSP-47) and apoptotic (TUNEL assay) responses on the immunohistochemical analyses. The optical transmittance of corneas did not change significantly, and the inter-fibrillar distances of the corneal stroma evaluated with transmission electron microscopy were not significantly altered after THz exposure. These results provide the basis for future research work on the development of THz imaging system for its application in ophthalmology.

Animal model with structural similarity to human corneal collagen fibrillar arrangement.

Rabbit and porcine corneas have been used in scientific research due to their structural similarity to the human cornea. Currently, there are no studies that have compared corneal collagen fibrillar diameter, interfibrillar distance and interlamellar distance between human and animal models. Ten pairs of porcine, rabbit, and human corneas were used. These were analysed using light and Transmission Electron microscopy. The collagen fibrillar diameter, interfibrillar distance and interlamellar distance were statistically compared between porcine, rabbit and human corneas. The human, porcine and rabbit; mean collagen fibrillar diameters were: 24.52 ± 2.09 nm; 32.87 ± 0.87 nm; and 33.67 ± 1.97 nm. The mean interfibrillar distances were: 46.10 ± 2.44 nm; 53.33 ± 2.24 nm; and 52.87 ± 2.73 nm, respectively. The collagen fibrillar diameter and interfibrillar distance of porcine and rabbit corneas were significantly different (p < 0.001) to the human corneal values but not form each other. The interlamellar distance of human, porcine and rabbit corneas was: 2190 ± 820 nm; 6460 ± 1180 nm; and 4410 ± 1330 nm, respectively. All the comparisons were statistically different, in porcine versus rabbit at the p < 0.01 level and both porcine and rabbit versus human at the p < 0.001 level. Histologically, all five layers (epithelium, Bowman's layer, stroma, Descemet membrane and endothelium) of the cornea were visible in all the three species. While neither animal model was structurally identical to the human cornea, they are both relatively close to being used as models to study the biomechanical effects of external insults/treatments to be extrapolated to the human cornea.

A comparison of the ultrastructure of the cornea of the pre- and post-metamorphic axolotl (Ambystoma mexicanum, Amphibia).

The corneal ultrastructure of the pre- and post-metamorphic stages of the neotenic axolotl Ambystoma mexicanum is examined using light microscopy and both scanning and transmission electron microscopy to reveal whether there are any morphological changes associated with a switch in lifestyle. Although the complement of corneal layers remains the same, there are significant quantitative changes in corneal, epithelial and stromal thickness, epithelial and endothelial cell size and density, and the thickness of Bowman's layer and Desçemet's membrane. Microholes in the epithelium and vertical sutures within the stroma are predominant features in the pre-metamorphic stage but are rarely seen in the post-metamorphic stage. There are also significant quantitative centro-peripheral differences in the thickness of the whole cornea, primarily due to differences in the thickness of the stroma in both metamorphic stages. These changes may reflect the physiological demands on the cornea as it switches from a purely aquatic to an amphibious lifestyle, which includes venturing onto land.