Carole Burillon

Cultured autologous oral mucosal epithelial cell sheet (CAOMECS) transplantation for the treatment of corneal limbal epithelial stem cell deficiency.

PURPOSE Total bilateral corneal limbal epithelial stem cell deficiency (LSCD) cannot be treated with the surgical transplantation of autologous limbus or cultured autologous limbal epithelium. Transplantation of allogenic limbal epithelium is possible but requires immunosuppressive treatments. Cultured autologous oral mucosal epithelial cell sheet (CAOMECS) is a transparent, resistant, viable, and rapidly bioadhesive cell sheet, cultured with the UpCell-Insert technology (CellSeed, Inc., Tokyo, Japan), which allows for grafting onto the patients corneal stroma without suturing. It has therefore been proposed as an alternative treatment for LSCD. METHODS The objectives were to assess the safety and efficacy of CAOMECS, using a prospective Gehans design. Safety was measured in terms of ocular adverse events during the study period, and efficacy was measured using a composite criterion based on epithelial defect, punctate epithelial keratopathy, conjunctival epithelium on the cornea, number of vascular pediculi, and vessel activity. RESULTS CAOMECS was found to be safe and effective. In total, 26 eyes of 25 patients received a graft. Two patients experienced serious adverse events classified as not product related. Twenty-five patients were included in the efficacy analysis, as one patient was lost to follow-up. The treatment was found to be effective in 16 of 25 patients at 360 days after grafting. Of the 23 patients who completed follow-up at 360 days, 22 had no ulcers, and 19 showed a decrease in the severity of the punctate epithelial keratopathy. CONCLUSIONS CAOMECS is a well-tolerated and safe tissue-engineered product. These results suggest its efficacy for reconstructing the ocular surface in patients with total bilateral corneal LSCD.

Malignant glaucoma induced by a phakic posterior chamber intraocular lens for myopia

A 23-year-old woman with -14.00 diopters of myopia requested emmetropia for professional reasons. An ICM 130 V2 myopic phakic intraocular lens (IOL) (Staar Surgical AG) was implanted in the posterior chamber. Three days later, the patient developed malignant glaucoma. Pupillary block glaucoma and choroidal hemorrhage or effusion were ruled out. As maximum medical treatment failed, rapid secondary surgery was performed with sclerotomy, aspiration in the midvitreous cavity, and removal of the IOL. The follow-up was 43 months.

Use of magnetically oriented orthogonal collagen scaffolds for hemi-corneal reconstruction and regeneration.

We recently showed that the highly organized architecture of the corneal stroma could be reproduced using scaffolds consisting of orthogonally aligned multilayers of collagen fibrils prepared using a high magnetic field. Here we show that such scaffolds permit the reconstruction in vitro of human hemi-corneas (stroma + epithelium), using primary human keratocytes and limbal stem cell derived human keratinocytes. On the surface of these hemi-corneas, a well-differentiated epithelium was formed, as determined both histologically and ultrastructurally and by the expression of characteristic markers. Within the stroma, the keratocytes aligned with the directions of the fibrils in the scaffold and synthesized a new extracellular matrix with typical collagen markers and small, uniform diameter fibrils. Finally, in vivo experiments using a rabbit model showed that these orthogonally oriented multi-layer scaffolds could be used to repair the anterior region of the stroma, leading to re-epithelialization and recovery of both transparency and ultrastructural organization.

Reconstruction of a full-thickness collagen-based human oral mucosal equivalent

Tissue engineered human oral mucosa has the potential to be applied to the closure of surgical wounds after tissue deficits due to facial trauma, malignant lesion surgery or preposthetic procedure. It can also be used to elucidate the biology and pathology of oral mucosa and as a model alternative to animals for safety testing of oral care products. Using the technology previously developed in our laboratory for the production of a skin equivalent, we were able to reconstruct a nonkeratinized full-thickness human oral mucosal equivalent closely mimicking human native oral mucosa. The successive coculture of human lamina propria fibroblasts and human oral epithelial cells isolated from the nonkeratinized region of oral cavity in a porous collagen-glycosaminoglycan (GAG)-chitosan scaffold gave rise to a lamina propria equivalent (LPE) and then to an oral mucosa equivalent (OME). The results of the histology, immunohistology and transmission electron microscopy of this OME demonstrated the presence of a nonkeratinized pluristratified and differentiated epithelium as in native nonkeratinized human oral mucosa expressing both K13 and K3/76. This epithelium was firmly anchored to the LPE by a continuous and ultrastructurally well-organized basement membrane. In the LPE, fibroblasts synthesized new extracellular matrix where the average collagen fibre diameter was 28.4 nm, close to that of native oral mucosa. The proliferative capacity of the basal cells was demonstrated by the expression of Ki67.

Endophthalmitis After Intravitreal Injections: Incidence, Presentation, Management, and Visual Outcome

PURPOSE To report the incidence and characteristics of endophthalmitis after intravitreal injections of anti-vascular endothelial growth factor agents or corticosteroids and to describe the clinical and bacteriologic characteristics, management, and outcome of these eyes with acute endophthalmitis in France. DESIGN Retrospective, nationwide multicenter case series. METHODS From January 2, 2008 to June 30, 2013, a total of 316,576 intravitreal injections from 25 French ophthalmic centers were included. For each center, the number of intravitreal injections was determined using billing codes and the injection protocol was recorded. A registry and hospital records were reviewed to identify patients treated for endophthalmitis after injection during the same time period. The main outcome measures were the incidence of clinical endophthalmitis and visual acuity of endophthalmitis cases. RESULTS During the study period, 65 cases of presumed endophthalmitis were found, giving an overall incidence of 0.021% (2.1 in 10,000 injections) (95% confidence interval [CI], 0.016%-0.026%). The median number of days from injection to presentation was 4 [1-26] days. The most common symptom was vision loss. Bacterial identification was achieved in 43.4%. The most frequent pathogens were gram-positive bacteria (91.3%), including coagulase-negative Staphylococcus in 78.3%. Neither the interval between injection and presentation for endophthalmitis nor the clinical signs differentiated culture-positive from culture-negative cases. In multivariate analysis, the use of a disposable conjunctival mould assist device and the use of prophylaxis with an antibiotic or antiseptic were significantly associated with an increased incidence of endophthalmitis (P = .001). The majority of patients had worse visual acuity after 3 months of follow-up when compared with acuity before endophthalmitis. CONCLUSIONS The incidence of presumed endophthalmitis after intravitreal injections of anti-vascular endothelial growth factors or corticosteroids was low and the prognosis poor. Prevention and management remain challenging. It remains to be determined whether the findings of this study are relevant for other countries using different techniques for intravitreal injections.

Value of two mortality assessment techniques for organ cultured corneal endothelium: trypan blue versus TUNEL technique

Background/aim: It is known that trypan blue staining is not a good predictor of loss of corneal endothelial cells (ECs) during organ culture. As it is primarily an indicator of membrane integrity, it would also not be expected to identify ECs undergoing apoptosis. The aim of this study was to determine the ability of the in situ TdT dUTP mediated nick end labelling (TUNEL) technique to detect cell death in the corneal endothelium caused by apoptosis during organ culture, compared with conventional vital staining with trypan blue. Methods: 31 human corneas were organ cultured at 31°C for 3–35 days. Staurosporine was used to induce apoptosis in five control corneas. The endothelium was assessed by trypan blue and by the in situ TUNEL technique. The percentages of trypan and TUNEL positive ECs were compared. Their links with sex, donor age, time from donor death and organ culture, initial and final EC density and cell loss were studied. Results: TUNEL stained ECs were observed in all corneas. TUNEL positive ECs were mostly located either in corneal folds or at the periphery of corneal folds showing central shedding. The mean percentage of cell death at the end of storage, assessed by the trypan blue technique, was 1.47% (SD 2.63, range 0.03–12); assessed by the TUNEL technique it was 12.7% (SD 16.4 range 0.6–65.5). There was a significant correlation between the two techniques (r = 0.7, p<0.001). The percentage of TUNEL stained ECs was correlated negatively with EC density at the end of storage (r = −0.47, p <0.005) and positively with percentage EC loss during storage (r = 0.46, p < 0.05). Conclusion: This study demonstrates that organ cultured corneas systematically carry non-viable ECs that are implicated in cell death by apoptosis and go undetected when trypan blue staining is used. Because the in situ TUNEL assay detects earlier events in the cell death process than does trypan blue, it should be used to quantify endothelial viability, especially for experiments with new storage media.

Prospective study of corneal collagen cross-linking efficacy and tolerance in the treatment of keratoconus and corneal ectasia: 3-year results.

Purpose: To assess the efficacy and tolerance of corneal collagen cross-linking with corneal epithelium debridement in the stabilizing treatment of primary or secondary corneal ectasia. Methods: Prospective, comparative, single-center study of patients presenting with progressive primary or secondary corneal ectasia. The control group, comprising the fellow eye of patients with bilateral involvement, was followed up for 6 months and then treated. The parameters examined were the biomicroscopic examination, visual acuity [best spectacle–corrected visual acuity (BSCVA) and uncorrected visual acuity (UCVA)], keratometry of the central 3 mm, intraocular pressure, central pachymetry, endothelial density, and macular profile. Results: Fifty-five eyes of 39 patients were treated; the mean follow-up period was 20.8 ± 6.8 months (range, 12–36 months). The control group comprised 16 eyes. UCVA and BSCVA were significantly improved between 3 and 12 months, reaching their minimum at 6 months, varying from 0.12 UCVA to 0.07 BSCVA (P < 0.05). These values and the keratometry values did not vary significantly after 36 months of follow-up. In contrast, analysis of the control group revealed significant keratometric deterioration of +1.2 diopters at 6 months (P < 0.05), with no further significant variation after treatment. Analysis of the subgroups of patients with post–laser in situ keratomileusis ectasia confirmed these results. At the end of the study, intraocular pressure, pachymetry, and endothelial density were not significantly modified, and no macular profile modification was observed. Conclusion: This study shows that corneal collagen cross-linking can stabilize progressive corneal ectasia, both primary and secondary, with no induced iatrogenic effects.

In-Vitro Study of Bacterial Adherence to Different Types of Intraocular Lenses

ABSTRACT The aim of this study was to determine the adherence of Staphylococcus epidermidis to intraocular lenses made of five different biomaterials: polymethylmethacrylate (PMMA), heparinized PMMA, silicone, hydrophilic acrylic, and hydrogel. The extent of bacterial binding was measured by counting. The results were compared using a one-factor variance analysis. Adherence was weakest on hydrogel and strongest on the silicone polymer. Bacterial adherence to the implant surface must therefore depend on the hydrophobicity or hydrophilicity of the biomaterial.

Migration en chambre antérieure de l’implant intravitréen de dexaméthasone Ozurdex® chez le pseudophake : à propos de trois cas ☆

INTRODUCTION Intravitreal implantation of Ozurdex(®) (Allergan Inc., Irvine, CA, USA) is being used widely for the treatment of macular edema secondary to retinal vein occlusion and in the setting of non-infectious posterior uveitis. We describe a complication little reported in the literature until now: migration of the dexamethasone implant into the anterior chamber. PATIENTS AND METHODS We report three cases of migration in two pseudophakic patients with iris claw lenses (on the anterior and posterior aspects of the iris) and in one pseudophakic patient with a posterior chamber IOL and zonular rupture. DISCUSSION The risk of anterior chamber migration of the Ozurdex(®) implant is increased in cases of prior vitrectomy (three cases), prone positioning and dilation of the pupil (mydriasis). Clinical tolerability of the implant in the anterior chamber is poor in all cases, with diffuse corneal edema. Endothelial cell loss occurs, as demonstrated by specular microscopy performed in two of our patients. Removal or repositioning of the Ozurdex(®) implant into the posterior segment must be performed without delay because of the risk of endothelial toxicity. CONCLUSION Patients without perfect zonular/posterior capsular integrity present a high risk of anterior chamber migration of the Ozurdex(®) implant. In such cases, anti-VEGF therapies should be discussed as an alternative.

Adherence and kinetics of biofilm formation of Staphylococcus epidermidis to different types of intraocular lenses under dynamic flow conditions

PURPOSE: To compare the adherence and biofilm formation of Staphylococcus epidermidis under in vitro flow conditions on intraocular lenses (IOLs) made of 4 biomaterials: poly(methyl methacrylate) (PMMA), silicone, hydrophilic acrylic, and hydrophobic acrylic. SETTING: Department of Ophthalmology, Croix‐Rousse University Hospital and University Research Laboratory, Lyon, France. METHODS: Intraocular lenses were placed in a bioreactor designed to replicate intraocular conditions. The model consisted of Tygon tubing connected to a vial. Three septa allowed the entry and elimination of the artificial aqueous humor and inoculation of the bacterial suspension.The first of 2 pumps moved the aqueous humor along the circuit; the second pump regulated the flow at which the nutritive environment was regenerated. At various times (12, 16, 24, 40, 48, 60, and 72 hours), IOLs were taken from this environment and the bound bacteria were removed and counted. The distribution of bacterial adhesion on the IOLs was modeled using polynomial Poisson regression. To test the effect of the IOL biomaterial on bacterial adhesion, likelihood ratio tests were performed. RESULTS: The model provided the kinetics of S epidermidis biofilm growth on IOLs. The biofilm growth on each of the 4 biomaterials occurred in 3 phases: latent, dynamic or accelerated growth, and linear growth. The extent of bacterial binding to IOLs increased from hydrophilic acrylic polymer to PMMA, hydrophobic acrylic, and silicone. The differences were statistically significant. CONCLUSION: Bacterial adhesion to and biofilm development on the IOL surface depended on the characteristics of the biomaterial.