Wound healing in rabbit corneas after flapless refractive lenticule extraction with a 345 nm ultraviolet femtosecond laser.

PURPOSE: To characterize corneal wound healing in a rabbit model after flapless refractive lenticule extraction with a 345 nm ultraviolet femtosecond laser. SETTING: Departments of Ophthalmology and Anatomy II, University of Erlangen-Nürnberg and Wavelight GmbH, Erlangen, Germany. DESIGN: Experimental study. METHODS: Flapless refractive lenticule extraction was performed in 1 eye each of 20 New Zealand white rabbits (-5.0 diopters). Groups of 4 animals were euthanized after 48 hours, 1 week, 2 weeks, 4 weeks, and 3 months, respectively. Corneal samples were prepared for histology and fluorescence microscopy. To assess corneal cell death, proliferation, and myofibroblastic transdifferentiation, terminal uridine deoxynucleotidyl nick end-labeling (TUNEL) assay as well as immunostaining for Ki67 and α-smooth muscle actin (αSMA) were performed on sagittal cryosections. RESULTS: Histology revealed a zone of keratocyte depletion with a thickness of approximately 50 µm around the extraction site. At 48 hours, pronounced TUNEL staining of keratocytes was detected around the interface (159.9 cells/mm ±â€¯18.4 [SD]), which steadily decreased to 74.9 ±â€¯19.8 cells/mm at 1 week and 5.7 ±â€¯4.8 cells/mm at 2 weeks. Ki67 staining of keratocytes was evident at 48 hours (10.0 ±â€¯3.8 cells/mm), which then decreased at 1 week (5.2 ±â€¯1.7 cells/mm) and 2 weeks (0.4 ±â€¯0.5 cells/mm). From 4 weeks onward, no TUNEL or Ki67 staining was detected. The corneal stroma was αSMA-negative at all timepoints. CONCLUSION: Application of the 345 nm laser showed no signs of problematic repair processes in the cornea, which supports the initiation of the clinical phase.

Evaluation of a 345 nm Femtosecond Laser for Corneal Surgery with Respect to Intraocular Radiation Hazard.

PURPOSE: We report our findings from a preclinical safety study designed to assess potential side effects of corneal ultraviolet femtosecond laser treatment on lens and retina. METHODS: Refractive lenticules (-5 dpt) with a diameter of 6 mm were created in the right cornea of eight Dutch Belted rabbits. Radiant exposure was 0.5 J/cm² in two animals and 18 J/cm² in six animals. The presence of lens opacities was assessed prior to and up to six months following laser application using Scheimpflug images (Pentacam, Oculus) and backscatter analysis (Opacity Lensmeter 702, Interzeag). Ganzfeld flash and flicker electroretinogram (ERG) recordings were obtained from both eyes prior to and up to six weeks following laser application. At the study endpoint, retinas were examined by light microscopy. RESULTS: Independent of energy dose applied, no cataract formation could be observed clinically or with either of the two objective methods used. No changes in ERG recordings over time and no difference between treated and untreated eye were detected. Histologically, retinal morphology was preserved and retinal pigment epithelium as well as photoreceptor inner and outer segments appeared undamaged. Quantitative digital image analysis did not reveal cell loss in inner or outer nuclear layers. CONCLUSIONS: Our analysis confirms theoretical considerations suggesting that ultraviolet femtosecond laser treatment of the cornea is safe for intraocular tissues. Transmitted light including stray light induces no photochemical effects in lens or retina at energy levels much higher than required for the clinical purpose. These conclusions cannot be applied to eyes with pre-existing retinal damage, as these may be more vulnerable to light.

Corneal tissue interactions of a new 345 nm ultraviolet femtosecond laser.

PURPOSE: To assess the suitability of a new 345 nm ultraviolet (UV) femtosecond laser for refractive surgery. SETTING: Department of Ophthalmology, University of Erlangen-Nürnberg, Erlangen, Germany. DESIGN: Experimental study. METHODS: Twenty-five porcine corneas were used for stromal flap or lamellar bed creation (stromal depth, 150 µm) and 15 rabbit corneas for lamellar bed creation near the endothelium. Ultraviolet femtosecond laser cutting-line morphology, gas formation, and keratocyte death rate were evaluated using light and electron microscopy and compared with a standard infrared (IR) femtosecond laser. Endothelial cell survival was examined after application of a laser cut near the endothelium. RESULTS: Flaps created by the UV laser were lifted easily. Gas formation was reduced 4.2-fold compared with the IR laser (P = .001). The keratocyte death rate near the interface was almost doubled; however, the death zone was confined to a region within 38 µm ± 10 (SD) along the cutting line. Histologically and ultrastructurally, a distinct and continuous cutting line was not found after UV femtosecond laser application if flap lifting was omitted and standard energy parameters were used. Instead, a regular pattern of vertical striations, presumably representing self-focusing induced regions of optical tissue breakdown, were identified. Lamellar bed creation with standard energy parameters 50 µm from the endothelium rendered the endothelial cells intact and viable. CONCLUSION: The new 345 nm femtosecond laser is a candidate for pending in vivo trials and future high-precision flap creation, intrastromal lenticule extraction, and ultrathin Descemet-stripping endothelial keratoplasty. FINANCIAL DISCLOSURES: Mr. Klenke and Ms. Skerl were paid employees of Wavelight GmbH when the study was performed. Dr. Seiler is a scientific consultant to Wavelight GmbH. No other author has a financial or proprietary interest in any material or method mentioned.

Novel collagen membranes for the reconstruction of the corneal surface.

No standardized biomaterial exists for the surgical treatment of persistent corneal erosions and ulcerations. We analyzed the suitability and biocompatibility of defined noncross-linked and UV/riboflavin cross-linked equine type I collagen membranes for the reconstruction of the corneal surface. Isolated human oral mucosa epithelial cells, a cell type in clinical use for the treatment of ocular surface diseases, were subcultivated on both types of membranes and examined concerning cell adhesion, proliferation, and differentiation. Biocompatibility was evaluated following superficial and intrastromal corneal transplantation in New Zealand white rabbits. In cell cultures all collagen membranes supported adhesion of oral mucosa epithelial cells leading to the formation of multilayered epithelial cell sheets. After intrastromal corneal implantation clinical signs of degradation were seen in all variants of collagen membranes, which was fastest in noncross-linked variants. The histological and ultrastructural level invasion of keratocytes and production of new collagen fibers inside the collagen membranes could be detected in noncross-linked variants. After superficial corneal implantation covering of the membranes by corneal epithelium over time was visible. Ultrastructural analysis showed a slower rate of degradation and less invading keratocytes in cross-linked variants compared with noncross-linked collagen membranes. Cross-linked and noncross-linked variants of the collagen membrane proofed to be suitable to serve as a carrier for epithelial stem cells in vitro and showed a high biocompatibility in vivo. These results indicate that the tested collagen membranes might be suitable for the reconstruction of the corneal surface in patients with nonhealing ulcerations. Whether membranes with faster or slower degradation properties are preferable for the treatment of persistent corneal ulcerations might depend on the underlying corneal pathology and the degree of concomitant inflammation.

Haploinsufficiency of ARID1B, a member of the SWI/SNF-a chromatin-remodeling complex, is a frequent cause of intellectual disability.

Intellectual disability (ID) is a clinically and genetically heterogeneous common condition that remains etiologically unresolved in the majority of cases. Although several hundred diseased genes have been identified in X-linked, autosomal-recessive, or syndromic types of ID, the establishment of an etiological basis remains a difficult task in unspecific, sporadic cases. Just recently, de novo mutations in SYNGAP1, STXBP1, MEF2C, and GRIN2B were reported as relatively common causes of ID in such individuals. On the basis of a patient with severe ID and a 2.5 Mb microdeletion including ARID1B in chromosomal region 6q25, we performed mutational analysis in 887 unselected patients with unexplained ID. In this cohort, we found eight (0.9%) additional de novo nonsense or frameshift mutations predicted to cause haploinsufficiency. Our findings indicate that haploinsufficiency of ARID1B, a member of the SWI/SNF-A chromatin-remodeling complex, is a common cause of ID, and they add to the growing evidence that chromatin-remodeling defects are an important contributor to neurodevelopmental disorders.