Simple Single-Pass Technique for Ultrathin Descemet Stripping Automated Endothelial Keratoplasty: A Pilot Study.

PURPOSE: To assess intraoperative and postoperative graft thickness (GT) after donor deturgescence for ultrathin Descemet stripping automated endothelial keratoplasty and to evaluate visual outcomes, endothelial cell density, and patient satisfaction at 1 year. METHODS: Prospective interventional case series of patients with Fuchs endothelial dystrophy, Fuchs endothelial dystrophy and cataract, and pseudophakic bullous keratopathy (n = 12 grafts). The donor cornea was allowed to thin out by simple evaporation on an artificial anterior chamber, to the required precut thickness, before a single microkeratome pass. GT after microkeratome cut, at 1 week, 1, 3, 6, and 12, months was measured. Early Treatment Diabetic Retinopathy Study (ETDRS) best-corrected visual acuity, Pelli-Robson contrast sensitivity, endothelial cell density, and score on the visual function questionnaire (VFQ-25) were assessed. RESULTS: Mean intraoperative postmicrokeratome cut GT was 78.9 ± 33.3 µm. Mean GT at 1 week, 1, 3, 6, and 12 months was 70.7, 70.9, 62.8, 66.5, and 58.9 µm, respectively. Mean initial donor corneal thickness was 647 ± 67 µm, and mean precut thickness was 526 ± 4.5 µm (mean thinning time: 17 min). Best-corrected visual acuity at 1 week, 1, 3, 6, and 12 months was 68.8, 76.9, 76.3, 76.9, and 78.6 letters with 9-letter gain at 12 months (P = 0.02). Mean endothelial cell loss at 3, 6, and 12 months was 36.8% ± 6.75%, 37.2% ± 8%, and 37.9% ± 9.75% loss, respectively. At 1 year, 83.3% of patients achieved ≥20/40 (6/12) and 66.7% of patients achieved ≥20/32 (6/9.5). VFQ-25 testing showed an improvement in the visual function. CONCLUSIONS: This pilot study demonstrates a simple graft deturgescence technique that reproducibly creates ultrathin grafts without donor wastage.

Discovery of Heteroaromatic Sulfones As a New Class of Biologically Compatible Thiol-Selective Reagents.

The selective reaction of chemical reagents with reduced protein thiols is critical to biological research. This reaction is utilized to prevent cross-linking of cysteine-containing peptides in common proteomics workflows and is applied widely in discovery and targeted redox investigations of the mechanisms underlying physiological and pathological processes. However, known and commonly used thiol blocking reagents like iodoacetamide, N-ethylmaleimide, and others were found to cross-react with oxidized protein sulfenic acids (-SOH) introducing significant errors in studies employing these reagents. We have investigated and are reporting here a new heteroaromatic alkylsulfone, 4-(5-methanesulfonyl-[1,2,3,4]tetrazol-1-yl)-phenol (MSTP), as a selective and highly reactive -SH blocking reagent compatible with biological applications.

Strained cycloalkynes as new protein sulfenic acid traps.

Protein sulfenic acids are formed by the reaction of biologically relevant reactive oxygen species with protein thiols. Sulfenic acid formation modulates the function of enzymes and transcription factors either directly or through the subsequent formation of protein disulfide bonds. Identifying the site, timing, and conditions of protein sulfenic acid formation remains crucial to understanding cellular redox regulation. Current methods for trapping and analyzing sulfenic acids involve the use of dimedone and other nucleophilic 1,3-dicarbonyl probes that form covalent adducts with cysteine-derived protein sulfenic acids. As a mechanistic alternative, the present study describes highly strained bicyclo[6.1.0]nonyne (BCN) derivatives as concerted traps of sulfenic acids. These strained cycloalkynes react efficiently with sulfenic acids in proteins and small molecules yielding stable alkenyl sulfoxide products at rates more than 100× greater than 1,3-dicarbonyl reagents enabling kinetic competition with physiological sulfur chemistry. Similar to the 1,3-dicarbonyl reagents, the BCN compounds distinguish the sulfenic acid oxoform from the thiol, disulfide, sulfinic acid, and S-nitrosated forms of cysteine while displaying an acceptable cell toxicity profile. The enhanced rates demonstrated by these strained alkynes identify them as new bioorthogonal probes that should facilitate the discovery of previously unknown sulfenic acid sites and their parent proteins.

Nd:YAG laser treatment for epithelial ingrowth after laser refractive surgery.

Epithelial ingrowth under refractive surgery flaps occurs in primary treatment and in further flap lift. Nd:YAG treatment was first described in 2008. In the only series published to date, 100% of patients treated had topographic and symptomatic improvement. The authors pictorially illustrate two cases of epithelial ingrowth following laser in situ keratomileusis achieving complete symptomatic relief and marked microscopic improvement following Nd:YAG laser treatment.

Regioselective semihydrogenation of dienes.

A one-pot, three-step strategy for the regioselective semihydrogenation of dienes is described. This procedure uses 9-BBN-H as a temporary protective group for alkenes. Yields range from 55% to 95%, and the reaction is tolerant of a variety of common functional groups. Additionally, the final elimination step of the sequence can be replaced with a peroxide-mediated alkylborane oxidation, generating regioselectively semihydrogenated product alcohols.

Corneal graft surgery combined with subconjunctival bevacizumab (avastin).

PURPOSE: To describe the use of subconjunctival bevacizumab (Avastin) as an adjunctive treatment in a vascularized cornea at the time of lamellar keratoplasty to reduce the risk of graft rejection. MATERIALS AND METHODS: After a significant ocular high-velocity thermal injury, a patient developed extensive corneal scarring and a traumatic cataract. To improve vision, a corneal graft was indicated, but the presence of extensive neovascularization increased the risk of early graft rejection. RESULTS: Bevacizumab was injected subconjunctivally before surgery to reduce the corneal vessel load. There was a dramatic response in terms of vessel regression, but this was short lived and the vessels quickly regrew. The subconjunctival bevacizumab injection was repeated at the time of combined cataract extraction and lamellar corneal graft surgery, and the feeder vessels were cauterized at the limbus. After 6 months, the graft remained clear of vessels. CONCLUSIONS: This case is of interest because the eye was treated twice with subconjunctival bevacizumab with good short-term results in both instances but different longer-term outcomes in terms of vessel regrowth. This case suggests that the antiangiogenic effects of bevacizumab may be effectively harnessed at the time of a definitive surgical procedure, which reduces the stimulus for vessel regrowth.

Neurovascular anatomy of the embryonic quail hindlimb.

Blood vessel and nerve development in the vertebrate embryo possess certain similarities in pattern and molecular guidance cues. To study the specific influence of shared guidance molecules on nervous and vascular development, an understanding of the normal neurovascular anatomy must be in place. The present study documents the pattern of nervous and vascular development in the Japanese quail hindlimb using immunohistochemistry and fluorescently labeled intravital injection combined with confocal and epifluorescent microscopy. The developmental patterns of major nerves and blood vessels of embryonic hindlimbs between stages E2.75 (HH18) and E6.0 (HH29) are described. By E2.75, the dorsal aortae have begun to fuse into a single vessel at the level of the hindlimb, and have completely fused by E3 (HH20). The posterior cardinal vein is formed at the level of the hindlimb by E3, as is the main artery of the early hindlimb, the ischiadic artery, as an offshoot of the dorsal aorta. Our data suggest that eight spinal segments, versus seven as reported by others (Tanaka and Landmesser,1986a; Tyrrell et al.,1990), contribute to innervation of the quail hindlimb. Lumbosacral neurites reach the plexus region by E3.5 (HH21 & 22), pause for approximately 24 hr, and then enter the hindlimb along with the ischiadic and crural arteries through shared foramina in the pelvic anlage. The degree of anterior-posterior spatial congruency between major nerves and blood vessels of the quail hindlimb was found to be highest medial to the pelvic girdle precursor, versus in the hindlimb proper.

Expression of vascular endothelial growth factor isoforms in the Japanese quail embryo.

Vascular endothelial growth factor (VEGF) i s required for vascular development. In the quail, four VEGF isoforms formed by alternative splicing, are translated into proteins with 122, 146, 166, and 190 amino acids. VEGF isoforms differ biochemically, with variable affinities for heparan sulfate proteoglycans, the extracellular matrix and VEGF receptors. There are few data on the functional significance of VEGF isoforms. RT-PCR was used to examine isoform expression during quail vascular development. Our results suggest that all quail isoforms are expressed during establishment of the vascular pattern in whole embryos and extraembryonic tissues at apparently equal levels. No isoform-specific expression patterns were detected in isolated endoderm or between embryo halves.

Vascular endothelial growth factor: a regulator of vascular morphogenesis in the Japanese quail embryo.

Experiments in mouse embryos indicate that a critical level of VEGF is required for normal vascular development, as mice lacking a single VEGF allele die at midgestation. Thus VEGF concentration may be a determinant of the size and location of major blood vessels during formation of the primary capillary plexus. Ectopic VEGF delivery was used to examine the effect of VEGF concentration on early vascular patterning in the quail embryo. VEGF was delivered by implanting VEGF-soaked heparin chromatography beads at three rostral-caudal locations in embryos with six somite pairs, which allowed us to study the effect of VEGF on different cellular activities. Ectopic VEGF resulted in significant changes in the vascular pattern at three rostral-caudal levels. Quantitation demonstrated an increased vascularity in the area of the implanted VEGF bead compared to the vascular pattern of embryos with control beads. Areas lateral to the dorsal aortae that are normally avascular became vascularized, and there was an apparent fusion between the dorsal aorta and lateral capillary plexus.