Corneal Buttons Research Articles

Use of different stains for microscopic evaluation for the diagnosis of Pythium keratitis

PurposeTo evaluate the efficacy of various staining techniques for detection of Pythium in keratitis cases. MethodsData of nineteen consecutive culture-positive cases of Pythium keratitis were retrospectively analysed. Corneal scrapings and corneal buttons (in the cases which underwent therapeutic penetrating keratoplasty [TPK]) were sent for microbiological and histopathological examination. The direct smears were stained with Potassium hydroxide and calcofluor white (KOH ​+ ​CFW), Gram and Iodine–Potassium Iodide–Sulphuric Acid (IKI–H2SO4) stains. The corneal buttons were stained with Gomori's Methanamine Silver (GMS), Periodic Acid-Schiff (PAS) and Iodine–Potassium Iodide–Sulphuric Acid (IKI–H2SO4) stains. The positivity of various stains in detecting Pythium was studied. ResultsGram and KOH ​+ ​CFW staining from smear was done in 16 out of 19 (84.2%) cases. KOH ​+ ​CFW and Gram stains were suggestive of Pythium in 10 (62.5%) and 7 (43.8%) cases, respectively. IKI–H2SO4 staining in scraping samples was positive for Pythium in all the 4 (100%) cases in which it was performed. Half corneal buttons were positive for Pythium with IKI–H2SO4 stain as well as GMS stain in all the 18 cases that underwent TPK (100%). PAS stain showed weak to faint pink staining of Pythium filaments in 7 out of 18 cases (38.9%). ConclusionIKI–H2SO4 stain followed by KOH ​+ ​CFW stain detects Pythium filaments most accurately in corneal scraping samples from keratitis patients, although the differences were not statistically significant. The positivity of the stains depends on astute observation by an experienced ocular microbiologist and pathologist.

Preservation of donor corneal epithelium in McCarey-Kaufman medium.

To evaluate the role of McCarey-Kaufman (MK) medium in maintaining the integrity of donor corneal epithelium. Nineteen corneal buttons were harvested and stored in MK media at 2°C-8°C for four days. Serial photographs were done every day till the 3rd day, and images were then analyzed with ImageJ software (LOCI, University of Wisconsin, USA). The area of exposure and epithelial defect (ED) was calculated every day for each corneal button. The average age of the donors was 56.5 ± 22.7 years and mean time from death to preservation of the corneal buttons was 7.7 ± 3.1 hours. The average corneal area was 145.6 ± 18.8 mm2. The total mean area of exposure was 3.6 ± 4.8, 7.2 ± 9.2, and 9.0 ± 11.9 mm2, and ED was 1.7 ± 4.6, 2.8 ± 5.3, and 3.3 ± 5.9 mm2 on days 1, 2, and 3, respectively. The percentage of increase in the area of exposure and ED in MK media was 3.71% and 1.1% from day 1 to day 3, respectively. Six out of 19 corneal buttons (31.57%) were utilized for keratoplasties, of which two were utilized in house and four were distributed outside. Of the two utilized corneas, none had epithelial defect on postoperative day 1. Rest 13 corneas were either used for training and research purposes, stored in glycerol media, or discarded. Since the percentage change in area of exposure/ED is not much at the end of day 3, corneas stored in MK media can be safely used even after three days of storage. Hence, MK medium serves as an excellent medium in maintaining the integrity of donor corneal epithelium.

Extending the utility of anterior corneal buttons through refrigeration and glycerol cryopreservation: utility rate and outcome analysis

Background/aimsTo evaluate the utility rate, indication, outcome, and cost of refrigeration and glycerol cryopreservation for storing anterior corneal buttons during endothelial keratoplasty for subsequent use in tectonic lamellar patch grafting.MethodAnterior...

A retrospective study to analyse epidemiological profile of community acquired infectious keratitis in a tertiary care hospital located in Northern Mumbai

Background: Epidemiology of infectious keratitis varies with geographical location. The aim of the study was to review the etiology of infectious keratitis in community that will help in management strategy.Methods: A retrospective study was conducted at tertiary care hospital from November 2017 to December 2020. Corneal scrapings and corneal button were processed as per microbiological standards. Bacterial identification and antimicrobial sensitivity testing was done by automated Vitek-2 compact system. Significant fungal growth was confirmed by its colony morphology, pigment production and lacto phenol cotton blue (LPCB) mount examination.Results: Of total 130 patients, culture positivity was 63 (48%), of these 59 (53%) were from corneal scrapping and 4 (21%) from corneal button. The most common bacterial isolates were Streptococcus pneumoniae 12 (38%), followed by Pseudomonas aeruginosa 6 (19%) and Staphylococcus aureus 4 (12%). The most common fungi isolated was Fusarium spp 14 (47%) followed by Aspergillus spp 9 (30%). One specimen showed Acanthamoba spp in wet mount preparation. Major associated factors were injury with vegetative matter 30 (48%) followed by contact lens use 12 (19%) and diabetes mellitus 11 (17%). Antibiotic susceptibility tests showed that all gram-positive cocci were susceptible to vancomycin (100%) and fluoroquinolones (100%) followed by third generation cephalosporins (80%). Whereas all gram-negative bacilli were susceptible to aminoglycosides (100%) followed by fluoroquinolones (90%). Conclusions: Epidemiology of infectious keratitis vary with geographical location. Understanding local microbiological profile assist in empirical therapy when diagnostic facilities are not readily available.

Effect of corneal collagen crosslinking on viscoelastic shear properties of the cornea

The cornea is responsible for most of the refractive power in the eye and acts as a protective layer for internal contents of the eye. The cornea requires mechanical strength for maintaining its precise shape and for withstanding external and internal forces. Corneal collagen crosslinking (CXL) is a treatment option to improve corneal mechanical properties. The primary objective of this study was to characterize CXL effects on viscoelastic shear properties of the porcine cornea as a function of compressive strain. For this purpose, corneal buttons were prepared and divided into three groups: control group (n = 5), pseudo-crosslinked group (n = 5), and crosslinked group (n = 5). A rheometer was used to perform dynamics torsional shear experiments on corneal disks at different levels of compressive strain (0%–40%). Specifically, strain sweep experiments and frequency sweep tests were done in order to determine the range of linear viscoelasticity and frequency dependent shear properties, respectively. It was found that the shear properties of all samples were dependent on the shear strain magnitude, loading frequency, and compressive strain. With increasing the applied shear strain, all samples showed a nonlinear viscoelastic response. Furthermore, the shear modulus of samples increased with increasing the frequency of the applied shear strain and/or increasing the compressive strain. Finally, the CXL treatment significantly increased the shear storage and loss moduli when the compressive strain was varied from 0% to 30% (p < 0.05); larger shear moduli were observed at compressive 40% strain but the difference was not significant (P = 0.12).

Clinical application of a shape-preserving rapid corneal donor dehydrater.

To describe the design and clinical application of a corneal donor dehydrator which can quickly dehydrate corneas and keep its original shape. The corneal donor material is placed on stainless steel beads with different diameters in the dehydrating box to make the cornea the same shape as the steel ball. Then, the cornea is placed inside the dehydrater for rapid dehydrating using the internal cleaning and ventilation system. Totally 83 eyes underwent deep anterior lamellar keratoplasty (DALK) using corneal donor tissue preserved with corneal dehydrater, and 60 patients (60 eyes) received DALK by the same surgeon using corneal donor tissue preserved with glycerol were included in the control group. The best corrected visual acuity (BCVA), the thickness and transparency of the corneal buttons were recorded. After the completion of dehydrating, all the donor corneas maintained a normal shape without any shrinkage or distortion, and the average intraoperative rehydration time was 43.3±12.1s during operation. The mean BCVA of the dehydrater group was 0.30±0.18 at 1wk and 0.32±0.16 at 1mo, which were statistically better than that of the control group (P<0.001). The score of corneal buttons transparency were lower than that of the control group with statistical difference (P<0.001). The thickness of corneal buttons at 1wk and at 1mo in the dehydrater group was significantly better than that of the control group respectively (P<0.001). One week after operation, no corneal button turbidity or edema was observed in both groups. The dehydrater can quickly dehydrate the corneal material in a clean and airtight environment and maintain the original shape of the corneal donor during the dehydrating process. This dehydrater is recommended for long-term high-quality preservation in areas where corneal materials cannot be used within a reasonable time period.

Elucidating the clinical, microbiological and molecular diagnostic aspects of Macrophomina phaseolina keratitis.

This study reports the clinico-microbiological features of Macrophomina phaseolina keratitis. Clinically diagnosed as microbial keratitis, six patients underwent microbiological evaluation. Fungal culture isolates from cornea were subjected to DNA sequencing of the ITS region, phylogenetic analysis and reconfirmation by polymerase chain reaction (PCR). Minimum inhibitory concentrations (MICs) of six antifungal drugs were determined by microbroth dilution method against the six isolates. All patients were treated with antifungals. Failed medical therapy necessitated therapeutic penetrating keratoplasty (TPK). Corneal buttons were processed for histopathology. In all patients, the corneal scraping showed septate hyaline fungal filaments. The BLAST analysis for ITS sequences of all six fungal isolates suggested M. phaseolina, however, when limited to sequences from type material, they matched M. pseudophaseolina. Phylogenetic analysis could not differentiate between these two species and clustered in a single clade. PCR assay of specific gene sequence [MpCal (calmodulin)] reconfirmed all isolates as M. phaseolina. The MICs of voriconazole and posaconazole were lowest (0.03 to 2 and 0.1 to 2µg/mL respectively) and all isolates were susceptible to natamycin. Except for case 1, which healed with a scar on treatment, all other cases worsened, despite medical treatment, necessitating TPK. Histopathology of 3 out of 4 buttons showed the presence of fungal filaments. While direct microscopic examination of corneal scrapings is helpful in diagnosis, identification of M. phaseolina in culture is challenging. Although MICs of commonly used antifungals are low response to medical therapy is not encouraging; patients may require TPK for resolution of infection in M. phaseolina keratitis.

Global and Comparative Proteome Signatures in the Lens Capsule, Trabecular Meshwork, and Iris of Patients With Pseudoexfoliation Glaucoma.

Pseudoexfoliation (PXF) is characterized by the accumulation of the exfoliative material in the eye and high rates of blindness if left untreated. Pseudoexfoliation glaucoma (PXG) is generally diagnosed too late due to its asymptomatic nature, necessitating the development of new effective screening tools for the early diagnosis of the disease. Thus, the increasing prevalence of this disease due to an aging population has demanded the identification of suitable biomarkers for the early detection of the disease or detection of the onset of glaucoma in the eyes with PXF. We applied a proteomics strategy based on a high-throughput screening method for the determination of proteins involving PXF and PXG pathogenesis. The lens capsule (LC), iris, and trabecular meshwork (TM) samples with PXF and PXG were taken by surgical trabeculectomy, and control samples were taken from the donor corneal buttons obtained from the institutional eye bank to characterize the proteome profile. Peptides from the LC were analyzed using liquid chromatography with tandem mass spectrometry (LC-MS/MS). The protein of interest and cytokine/chemokine profiles were verified using immunohistochemistry and the bio-plex kit assay, respectively. There were a total of 1433 proteins identified in the human LC, of which 27 proteins were overexpressed and eight proteins were underexpressed in PXG compared with PXF. Overexpressed proteins such as fibromodulin, decorin, lysyl oxidase homolog 1, collagen alpha-1(I) chain, collagen alpha-3(VI) chain, and biglycan were the major components of the extracellular matrix (ECM) proteins involved in cell-matrix interactions or ECM proteoglycans and the assembly and cross-linking of collagen fibrils. The ECM composition and homeostasis are altered in glaucoma. Thus, quantitative proteomics is a method to discover molecular markers in the eye. Monitoring these events can help evaluate disease progression in future studies.

Clinical Outcomes of Rose Bengal Mediated Photodynamic Antimicrobial Therapy on Fungal Keratitis with Their Microbiological and Pathological Correlation

Purpose To report the clinical outcome of Photodynamic Antimicrobial Therapy (PDAT) with Rose Bengal (RB) used as an early adjuvant therapy in patients with fungal keratitis and their microbiological and pathological correlation. Methods Patients with microbiologically confirmed fungal keratitis underwent PDAT-RB along with topical natamycin 5% drops hourly and oral ketoconazole 200 mg twice a day. This was performed by applying rose bengal (0.1%) to the de-epithelialized cornea for 30 minutes, followed by irradiation with a 6 mW/cm2 custom-made green LED source for 15 minutes (5.4 J/cm2). The corresponding fungal isolates were tested in vitro using PDAT-RB and corneal buttons were evaluated for correlation. Results Following informed consent, seven patients (male-5, female-2, mean age 47.7 years) with fungal keratitis were recruited. There were 3 cases each of Fusarium and Aspergillus flavus and 1 case of Acremonium sp. The average vertical and horizontal diameters of the corneal infiltrate were 4.12 ± 0.55 and 3.99 ± 1.19 mm, respectively. The average depth of corneal involvement was 283 ± 75.27µ as measured by anterior segment OCT. The clinical resolution was achieved in the cases with Fusarium keratitis with an average time of 39 days. Three cases of A. flavus and a single patient with Acremonium keratitis worsened and needed therapeutic keratoplasty (TPK) for resolution. Post-TPK, the corneal tissues grew A. flavus in one out of three cases and Acremonium sp. in one case. In vitro PDAT-RB experiment was performed on the corresponding fungal isolates grown from the corneal scraping. PDAT-RB produced clear inhibition of Fusarium and Acremonium sp. with no effect on the growth of A. flavus. Histopathologically, 2 out of 4 (50%) corneal buttons showed fungal filaments. Conclusions While the in vitro and in vivo results of PDAT-RB matched for Fusarium sp. and Aspergillus flavus keratitis being favourable in the former and non-favourable in the latter, these results were discrepant in Acremonium sp.

Effect of phacoemulsification fluid flow on the corneal endothelium: experimental study in rabbit eyes.

To quantify the damage to the corneal endothelium from the flow of a balanced salt solution during phacoemulsification. John A. Moran Eye Center, University of Utah, Salt Lake City, Utah. Experimental study. 12 New Zealand white rabbits received bilateral surgery. 6 eyes served as controls with no irrigation (incision only). In 18 eyes, the Intrepid Balanced tip of the Centurion Ozil (Alcon Laboratories, Inc.) handpiece was inserted into the anterior chamber for continuous irrigation at 50 mL/min with either 250 or 500 mL of balanced salt solution. After killing humanely and enucleation, the corneas were removed, stained with trypan blue-alizarin red, and photographed in a standardized manner (×400 photographs from 5 specific areas and 1 overview photograph from each corneal button). The ImageJ program was used to evaluate cell damage and loss in the photographs obtained from each cornea. Analysis of the ×400 photographs showed no statistically significant differences between control, 250 mL, and 500 mL groups in the percentage of intact, damaged, or lost cells (P = .896, .851, and .972 respectively). For the overview photographs, the differences in areas of intact and damaged cells among the groups were statistically significant, likely because of the peripheral areas of damage related to touches between the phacoemulsification tip and the endothelium during irrigation. The volume of balanced salt solution flow alone did not seem to be a major contributing source of endothelial cell damage and loss, providing further insight into mechanisms of corneal endothelium damage during phacoemulsification.

A hospital-based observational comparative study of efficacy of intracameral voriconazole and oral ketoconazole in deep keratomycosis

Background: Fungal keratitis, one of the most common causes of ocular mycosis, is the second most common cause of blindness in the world, after cataracts. The aim of the study was to compare the efficacy of conventional topical, systemic medications and intracameral voriconazole injection in visual and structural outcomes in keratomycosis. Material and methods: We conducted a hospital-based observational study of 45 patients of 45 eyes with smear-positive fungal keratitis. Patients were categorized into three groups: Group I received systemic topical with oral ketoconazole 200 mg, Group II — topical medications with intracameral voriconazole 50 μgm/0.1 mL, Group III — topical medications with both oral ketoconazole 200 mg and intracameral voriconazole 50 μgm/0.1 mL. Results: The common fungal organism is identified as Fusarium. The mean final visual acuity (VA) was 1.25 ± 0.32, 1.47 ± 1.05, and 1.22 ± 0.37 logMAR in Group I, group II, and Group III, respectively. The mean improvement in VA was 0.33 ± 0.07, 0.01 ± 0.71, and –0.19 ± 0.02 logMAR without significant change (p = 0.9). There was a significant difference in VA between the final postoperative follow-up period and baseline in Group I cases (p = 0.0019). Whereas no difference in VA between the final postoperative follow-up period and baseline in either Group II (p = 0.0671) or Group III (p = 0.1505) cases. The difference in time between the disappearance of hypopyon and the mean time to infection healing was not statistically significant (p = 0.1). Three cases in each group were perforated, and keratoplasty was performed. These perforated cases did not show culture positive. Histopathology identified the isolated organisms as Aspergillus species (n = 3) and Fusarium species (n = 2) in the corneal buttons. Conclusion: The differences in VA between the three methods were not statistically significant, indicating no treatment method superior to others (inter-group). However, in Group I, there was a significant difference in VA between the final postoperative follow-up period and baseline (p = 0.0019). There was no difference in VA between these time intervals in either Group II (p = 0.0671) or Group III (p = 0.1505). Within-group or intra-group analysis reveals that the Group I method is more effective for VA. The success rate of the method depended cumulatively on the duration of intracameral voriconazole in the anterior chamber, non-drainage of hypopyon, and individual clinical response.

Effect of longitudinal and torsional ultrasound on corneal endothelial cells: experimental study in rabbit eyes.

To compare corneal endothelial damage from longitudinal and torsional ultrasound during phacoemulsification. John A. Moran Eye Center, University of Utah, Salt Lake City, Utah, USA. Experimental study. 9 New Zealand white rabbits underwent bilateral surgery. After incision, the Intrepid Balanced Tip of the Centurion Ozil handpiece was inserted into the anterior chamber, and the following settings were used: 50 mL/min flow, 70 mm Hg intraocular pressure, 600 mm Hg vacuum, and 60% longitudinal (1 eye) or torsional (contralateral eye) ultrasound for 30 seconds. Cumulative dissipated energy (CDE) was noted. After euthanasia and enucleation, the corneas were removed, stained with trypan blue/alizarin red, and photographed (400× photographs from 5 specific areas and 1 overview photograph from each corneal button). The ImageJ program was used to evaluate cell damage and loss in the photographs obtained from each cornea. Cavitation bubbles around the phacoemulsification tip were generally observed in the longitudinal group. CDE was 17.4 ± 0.58 and 6.93 ± 0.15 in the longitudinal and torsional groups, respectively (P = .003). The percentage of intact cells was statistically higher in the torsional group (94.22% ± 5.99% vs 56.85% ± 22.29%), and the percentage of lost cells was statistically higher in the longitudinal group (42.10% ± 22.02% vs 4.97% ± 6.23%) (P = .003). This was observed in the analysis of the 400× photographs and the overview photographs. This study suggests that torsional ultrasound was associated with significantly less corneal endothelial cell damage than classical longitudinal tip motion, providing further insight on mechanisms of corneal endothelial damage during phacoemulsification.

Transmembrane Mucin 1 Blocks Fluorescein Ingress to Corneal Epithelium.

PurposeTo determine the role of transmembrane mucins in blocking fluorescein ingress to the corneal epithelium and its deficiency in contributing to corneal fluorescein punctate staining.MethodsA dry eye model was established by extirpating lacrimal and Harderian glands in rabbits to correlate the expression of mucins with fluorescein-stained areas on the corneal button using immunofluorescence. Expression of transmembrane mucins was promoted in human corneal epithelial cells (HCECs) by culturing with the mucin-promoting medium (MPM) or diquafosol treatment. Conversely, the expression of mucins was downregulated by knockdown with short hairpin RNA. The role of mucin1 extracellular domain in fluorescein ingress was further verified by overexpression of N-terminally truncated mucin1 in HCECs.ResultsIn the rabbit dry eye model, the expression level of mucin1 was significantly decreased in superficial corneal epithelial cells where fluorescein punctate staining was observed. Upregulation of mucin1 and mucin16 in HCECs promoted by MPM or by diquafosol treatment impeded intracellular fluorescein ingress. Downregulation of mucin1 and mucin16 enhanced fluorescence ingress in HCECs after fluorescein staining. Overexpression of truncated mucin1 did not alter the fluorescein intensity of fluorescein-stained HCECs, supporting the notion that the ability of mucin1 to block fluorescein ingress was primarily mediated by its extracellular domain. Minimal inherent expression of mucin16 in the rabbit cornea limited the validation of its role in blocking fluorescein ingress in vivo.ConclusionTransmembrane mucin1 blocks fluorescein ingress in the corneal epithelium, explaining how fluorescein staining is positive when the level of transmembrane mucins is disturbed in dry eyes.

Labial mucosal epithelium grafting in an ex vivo human donor cornea model

The purpose of the study was to establish a simple ex vivo corneal re-epithelization model and study the labial mucosal epithelium grafting as a potential approach for ocular surface reconstruction. Four human donor corneal buttons were overstored in a corneal cold storage solution at 4 °C for 32–52 days. Four labial oral mucosa strips were dissected from four patients during fornix reconstruction after they signed informed consent. The substantia propria was trimmed off, and the resulting graft was sutured near the corneal limbus with running sutures (thus forming the tissue construct). Constructs were cultured under the standard conditions with the anterior corneal side outwards. After 3 weeks of culture, constructs were removed, washed, and fixed. Sections were stained with hematoxylin and eosin (HE), anti-keratins 4, 13, 19, and p63. Nuclei were counterstained with Hoechst. After the cultivation, all constructs were integral with the attached graft and non-loosened sutures. The native cells were absent in all donor corneas. Histological evaluation demonstrated that the labial mucosal grafts were attached to the Bowman's membrane (BM), and its cellular outgrowths were found to be transit from the graft to the BM over the anterior surface in all constructs. Cells expressed mucosal epithelial keratins 4, 13, and 19, and several were p63-positive in nuclei. In the study, a simple ex vivo corneal re-epithelization model was successfully established. The model was potent in studying the labial mucosal epithelium grafting as an option for autologous ocular surface reconstruction in patients with bilateral limbal stem cell deficiency.

Epithelial lip mucosa transplant re‐populates corneal anterior surface. An ex vivo study

AbstractPurposeTo establish a new ex vivo model and study the labial mucosal epithelium grafting as a potential approach for cornea re‐epithelization in patients with bilateral limbal stem cell deficiency.MethodsFour human donor corneal buttons were collected from the Eye tissue bank in corneal cold storage solution and left at 4°C for 32–52 days. Labial oral mucosa strips were dissected from four patients during fornix reconstruction after they signed an informed consent. The labial mucosa epithelial graft was prepared by dissecting substantia propria. The resulted graft was sutured near the limbus of the corneal button with running sutures (construct). Each construct was cultured under the standard conditions with the concave side upwards. After 21 days of culture, constructs were removed, washed and fixed. First half of the constructs were stained with hematoxylin and eosin (HE). 5 um crosscut cryo‐sections of the second half were stained with anti‐ keratin 13, 19, and p63, and paired secondary fluorescent antibodies. Nuclei were counterstained with Hoechst. Sections were accessed and photographed.ResultsAfter the culture, all constructs were integral with the attached epithelium graft and non‐loosened sutures. The graft became slightly flattened and brighter. HE staining demonstrated absence of the native cells in donor corneas. The labial mucosa grafts were located on the surface of the Bowman's membrane (BM) at the cornea periphery. Epithelial outgrowths were found to be transit from the labial mucosa graft to the BM. Cellular outgrows of the constructs had positive staining for keratin 13 and 19. Few cells were p63‐positive in nuclei.ConclusionsIn the study, a new ex vivo model composed of processed human donor cornea and live human donor labial mucosal epithelium graft was successfully established. The study confirmed the consistency of the new approach for corneal re‐epithelization by transplantation of an oral mucosal epithelial graft.

Volcano explosion in the eye: Expulsive choroidal haemorrhage.

Background :This video shows the dreaded complication of expulsive suprachoroidal haemorrhage in a penetrating keratoplasty . It teaches the various measures that help in preventing and managing the challenge peri-operatively.Purpose:This video is intended to sensitize the corneal surgeons about this rare complication using real-time surgical video and prepare them to face this challenge by demonstrating the preventive and mitigating strategies.Synopsis:A patient who was suitably taken up for an autokeratoplasty had expulsive choroidal haemorrhage in the non-seeing eye. This complication was managed using glycerin-preserved donor corneal button. Further, the video discusses various pre-operative and intra-operative optimizations to avoid such a complication and mitigate the damage caused by it, should it ever happen during a surgery.Highlights:Expulsive choroidal haemorrhage can be a traumatizing event and one must be mentally prepared to manage it. Pre-operative high-risk factors should always be managed and intra-operative maneuvers performed to prevent the complication, as shown in the video.Video link: https://youtu.be/s_ImBugPELw

Long-Term Observation of Deep Anterior Lamellar Keratoplasty in Patients with Post-LASIK Granular Corneal Dystrophy Type 2: Two Case Reports

IntroductionLaser-assisted in situ keratomileusis (LASIK) exacerbates granular corneal dystrophy type 2. Post-LASIK granular corneal dystrophy type 2 is treated by several surgical techniques. To the best of our knowledge, no report has addressed the results of deep anterior lamellar keratoplasty in affected patients. Here, we report our experience regarding deep anterior lamellar keratoplasty treatment of patients with post-LASIK granular corneal dystrophy type 2.MethodsWe describe two Japanese women who underwent deep anterior lamellar keratoplasty to treat corneal opacities that worsened after LASIK.ResultsOne patient had a family history of corneal dystrophies. During the initial visit to our clinic, numerous fine opacities were found at the LASIK flap interface in both patients. The clinical findings were compatible with post-LASIK granular corneal dystrophy type 2. Both patients underwent deep anterior lamellar keratoplasty by one of the authors (T.C.). In both procedures, the surgeon used a visco-dissection technique and successfully removed the whole corneal stroma. Histopathological examination of the excised corneal button from each eye revealed amyloid and hyaline deposits at the LASIK flap interface. Neither patient experienced recurrent corneal opacity during the follow-up visit at 8 years (patient 1) and 6 years (patient 2).ConclusionDeep anterior lamellar keratoplasty can be used for the treatment of post-LASIK granular corneal dystrophy type 2. Removal of the entire host stroma may be important for the prevention of recurrent corneal opacity.

Ultrastructural study of collagen fibrils, proteoglycans and lamellae of the cornea treated with iontophoresis - UVA cross-linking and hypotonic riboflavin solution.

To investigate the effects of iontophoresis–ultraviolet A (UVA) cross-linking (CXL) with hypotonic riboflavin solution on the ultrastructural changes in the lamellae, collagen fibrils (CFs), and proteoglycans (PGs) in the central and peripheral stroma of the human corneal buttons. The iontophoresis method was used for the trans-epithelial application of hypotonic riboflavin in ex vivo corneal culture for 5 min. The corneas were irradiated using three methods: Group 1 (G1), a UVA irradiance of 3 mW/cm2 for 30 min; Group 2 (G2), a UVA irradiance of 10 mW/cm2 for 9 min; Group 3 (G3), without UVA irradiation. Three untreated corneas were used as controls (G0). After the CXL procedure, the corneas were processed for electron microscopy. The CF diameter and PGs in each sample were analyzed using the iTEM program. The keratocyte organelles and stromal architecture in the peripheral cornea were better preserved than those in the central cornea. In G1 and G2, the mean CF diameter in the peripheral cornea was significantly higher than that in the central cornea. In G3, the CF diameter in the central cornea was significantly larger than that in the peripheral cornea. Furthermore, differences in PG area size were observed between the central and peripheral corneas in all groups. Riboflavin + UVA application at 3 mW/cm2 for 30 min and 10 mW/cm2 for 9 min was a suitable method of CXL; however, 3 mW/cm2 for 30 min improved the organization and size of the collagen fibrils. CXL treatment applied at the periphery was more effective than that applied at the center.

Efficacy and outcome of simple limbal epithelial transplantation for limbal stem cell deficiency verified by epithelial phenotypes integrated with clinical evaluation

PurposeTo evaluate the efficacy and outcome of simple limbal epithelial transplantation (SLET) for limbal stem cell deficiency (LSCD) using epithelial phenotype detection integrated with clinical manifestation. MethodsThis prospective multicenter study included patients with LSCD who underwent autologous SLET (autoSLET) and living-related allogenic SLET (Lr-alloSLET). All patients were assessed by slit-lamp biomicroscopy, in vivo confocal microscopy (IVCM), and impression cytology with immunofluorescence staining (ICIF) before and after surgery. The criteria for success were the presence of a clinically non-conjunctivalized cornea and corneal epithelium detected by IVCM or ICIF. Otherwise, the case would be considered a failure. Visual improvement and risk factors for SLET failure were analyzed. ResultsA total of 28 eyes of 26 patients (11 autoSLET and 17 Lr-alloSLET) were included. The median age was 53 years (range, 35–63), and the follow-up time was 29.5 months (range, 17.5–39.8). The overall survival rate was 89.3% at 2 years and 75.6% at 3 years with no difference between autoSLET and Lr-alloSLET (p = 0.24). Seven eyes subsequently underwent penetrating keratoplasty. Immunohistochemistry analysis showed that all corneal buttons had corneal epithelium and limbal stem cell markers. Visual improvement was achieved in both SLET groups (p < 0.001). Failed SLET developed between 5 and 32 months postoperatively. However, absolute risk factors for SLET failure were unidentified. ConclusionThe efficacy of autoSLET and Lr-alloSLET for LSCD was excellent. Limbal explants can regenerate and restore the corneal surface while maintaining the characteristics of limbal stem cells as shown by epithelial phenotype detection and immunohistochemistry integrated with clinical evaluation.

Atypical Corneal Phenotype in Patients With Trachoma and Secondary Amyloidosis.

Purpose:To report clinical presentation, in vivo confocal microscopic features, and corneal phenotype in patients with trachomatous keratopathy (TK) and secondary amyloidosis.Methods:Histopathological records of all patients undergoing keratoplasty at the Dr. Rajendra Prasad Centre for Ophthalmic Sciences over a 3-year period were scanned retrospectively for a diagnosis of TK and amyloidosis. Demographic profile and details of preoperative comprehensive ophthalmic assessment were extracted. The histopathology was freshly reviewed.Results:Fifteen patients (29 eyes) with TK and atypical corneal involvement due to amyloid deposition were identified. Herbert's pits and upper palpebral conjunctival scarring were present in all cases. Central or total diffuse corneal scarring was present involving the anterior stroma in 5 (31%) and the full thickness of the cornea in 11 (69%) of the eyes. Eight (73%) of 11 patients with deep stromal amyloid deposits revealed bilateral, discrete, blue-white opacities at the level of deep stroma and Descemet membrane (DM). Endothelial cells were atrophic and flattened with gutta formation. Confoscans revealed hyperreflective, needle-shaped crystalline deposits of extracellular amyloid at various depths of the corneal stroma up to DM. All host corneal buttons demonstrated Congo red–positive amyloid deposits on histopathological examination.Conclusions:We describe a distinct form of TK unlike the usual presentation of dense, leucomatous, vascularized corneal scarring in trachoma. We believe that amyloid deposits in DM and the corneal endothelium have not previously been reported in patients with trachoma.