A novel gelatin hydrogel carrier sheet for corneal endothelial transplantation.

We examined the feasibility of using gelatin hydrogels as carrier sheets for the transplantation of cultivated corneal endothelial cells. The mechanical properties, transparency, and permeability of gelatin hydrogel sheets were compared with those of atelocollagen sheets. Immunohistochemistry (ZO-1, Na(+)/K(+)-ATPase, and N-cadherin), hematoxylin and eosin staining, and scanning electron microscopy were performed to assess the integrity of corneal endothelial cells that were cultured on gelatin hydrogel sheets. The gelatin hydrogel sheets displayed greater transparency, elastic modulus, and albumin permeability compared to those of atelocollagen sheets. The corneal endothelial cells on gelatin hydrogel sheets showed normal expression levels of ZO-1, Na(+)/K(+)-ATPase, and N-cadherin. Hematoxylin and eosin staining revealed the formation of a continuous monolayer of cells attached to the gelatin hydrogel sheet. Scanning electron microscopy observations showed that the corneal endothelial cells were arranged in a regular, mosaic, and polygonal pattern with normal cilia. These results indicate that the gelatin hydrogel sheet is a promising material to transport corneal endothelial cells during transplantation.

Tumor necrosis factor-alpha mediates photoreceptor death in a rodent model of retinal detachment.

PURPOSE: Photoreceptor degeneration is a major cause of visual loss in various retinal diseases, including retinal detachment (RD) and neovascular AMD, but the underlying mechanisms remain elusive. In this study, the role of TNFα in RD-induced photoreceptor degeneration was investigated. METHODS: RD was induced by subretinal injection of hyaluronic acid. Photoreceptor degeneration was assessed by counting the number of apoptotic cells with TdT-dUTP terminal nick-end labeling (TUNEL) 3 days after RD and measurement of the outer nuclear layer (ONL) thickness 7 days after RD. As the target of anti-inflammatory treatment, the expression of TNFα, with or without dexamethasone (DEX) was examined in rats by real-time PCR. To understand the role of TNFα in photoreceptor degeneration, RD was induced in mice deficient in TNFα or its receptors (TNFR1, TNFR2, and TNFR1 and -2), or in wild-type (WT) mice by using a functionally blocking antibody to TNFα. CD11b(+) cells in the outer plexiform layer (OPL) and subretinal space were counted by immunohistochemistry (IHC). RESULTS: Treatment with DEX (P = 0.001) significantly suppressed RD-induced photoreceptor degeneration and the expression of TNFα. RD-induced photoreceptor degeneration was significantly suppressed with specific blockade of TNFα (P = 0.032), in mice deficient for TNFα (P < 0.001), TNFR2 (P = 0.001), or TNFR1 and -2 (P < 0.001). However, lack of TNFR1 did not protect against RD-induced photoreceptor degeneration (P = 0.060). Müller cell activation was unchanged in WT and TNFα(-/-) mice. Recruitment of CD11b(+) monocytes was significantly lower in the TNFα(-/-) mice compared to WT mice (P = 0.002). CONCLUSIONS: TNFα plays a critical role in RD-induced photoreceptor degeneration. This pathway may become an important target in the prevention of RD-induced photoreceptor degeneration.

Suppression of phagocytic cells in retinal disorders using amphiphilic poly(γ-glutamic acid) nanoparticles containing dexamethasone.

To investigate the potential of nanoparticles (NPs) composed of poly(γ-glutamic acid) conjugated with l-phenylalanine (γ-PGA-Phe NPs) for the treatment of retinal diseases, γ-PGA-Phe NPs (200nm) were tested with macrophages and microglia in vitro or by intravitreal administration into normal or pathological rat eyes. The anti-inflammatory effects of the NPs containing dexamethasone (DEX-NPs) were examined using qRT-PCR in vitro by counting activated microglia and Fluorogold-labeled retinal ganglion cells in the retinas under excitotoxicity or by counting TUNEL (+) photoreceptors in the detached retinas. The NPs were taken up efficiently by cultured macrophages or microglia. At day 7, 60-80% of the diffuse signal remained in the cytoplasm of these cells. In normal rat eyes, the NPs did not accumulate in the retinas and no inflammatory cells were recruited. Conversely, under pathological conditions, the NPs were localized in activated CD11b-positive cells in the retina. DEX-NPs suppressed the expression of TNFα and MCP-1 in cultured macrophages or microglia, the activation of microglia, the loss of retinal ganglion cells (RGCs) in excitotoxic retinas, and the number of TUNEL (+) photoreceptors in detached retinas. These data suggest that γ-PGA-Phe NPs can be a powerful tool for suppressing inflammatory cells in pathological conditions in the retina.

Observation of posterior corneal vesicles with in vivo confocal microscopy and anterior segment OCT.

The histopathology of posterior corneal vesicles (PCV) has not yet been revealed. A 15-year-old girl, who was diagnosed by slit-lamp microscopy as PCV, was examined using specular microscopy, in vivo confocal microscopy, and anterior segment OCT (optical coherence tomography). Anterior segment OCT showed that the thickness of both corneas was within normal limits. At the same time, in vivo confocal microscopy revealed endothelial cells in the rounded dark areas, acellular hyporeflective layers on the Descemet's membrane, and hyperreflective linear lesions. These findings were not reported previously by slit-lamp and specular microscopy. The abnormal findings only existed at the Descemet's membrane and corneal endothelial layer. Previous reports dealing with posterior polymorphous dystrophy (PPMD) examined using in vivo confocal microscopy reported almost the same findings, suggesting that PCV and PPMD may be the same at the microstructural level.

Fluoroquinolone antibacterial eye drops: effects on normal human corneal epithelium, stroma, and endothelium.

BACKGROUND: In vitro studies have suggested the corneal cytotoxicity of third-generation fluoroquinolone levofloxacin (LVFX) and fourth-generation fluoroquinolone moxifloxacin hydrochloride (MFLX) among fluoroquinolone antibacterial eye drops. This study investigated the effects of these two eye drops on the human cornea in vivo. METHODS: We evaluated 30 healthy adults (19 men and 11 women, 38.3 ± 6.3 years old). Each subject received an LVFX ophthalmic solution 0.5% in one eye and an MFLX ophthalmic solution 0.5% in the other eye three times daily for 7 days. Functional and morphological corneal changes before and after instillation were evaluated through ophthalmic examinations including breakup time of tear film (BUT) as measured by fluorescein staining and DR-1, Schirmer I test, Heidelberg Retina Tomograph II Rostock Cornea Module (HRTII-RCM), specular microscope, and Pentacum examination. RESULTS: Both the LVFX and MFLX groups had no significant change in each examination before and after instillation. There was also no statistically significant difference in measurements after the 7-day instillation between the groups. CONCLUSION: Our study results suggest that as with LVFX, MFLX used in a normal clinical setting is unlikely to cause any obvious adverse effects on human normal cornea.