Ocular penetration of caspofungin in a rabbit uveitis model

Background: Little is known about the ocular penetration of echinocandin antifungals. We studied the ocular distribution of systemically administered caspofungin in a rabbit uveitis model. Methods: Caspofungin (1mg/kg per day) was given intravenously to rabbits as a single dose or as repeated daily doses on 7days starting 24h after induction of unilateral uveitis by intravitreal endotoxin injection. Caspofungin concentrations were determined by high-performance liquid chromatography in the cornea, aqueous humor, vitreous humor, and serum 4, 8, 16, and 24h after administration of a single dose and 24h after the last of seven doses. Results: The mean caspofungin concentration in the aqueous of the inflamed eye 4 and 8h after single-dose administration was 1.30 ± 0.39μg/ml and 1.12 ± 0.34μg/ml, respectively. Drug concentrations decreased to 0.24 ± 0.09μg/ml at 16h and 0.26 ± 0.14μg/ml at 24h. In the vitreous of inflamed eyes drug levels were undetectable at all time points. No drug was found in the aqueous of inflamed eyes 24h after the last of seven repeated doses, and the vitreous only contained trace amounts. In the corneas of inflamed eyes concentrations reached 1.64 ± 0.48μg/g at 4h, peaked at 2.16 ± 1.14μg/g at 8h, and declined to 1.87 ± 0.52μg/g and 1.49 ± 0.48μg/g at 16and 24h, respectively. After repeated dosing, corneal concentrations of caspofungin were 0.8 and 1.0μg/g and below the limit of detection in two of four animals. In non-inflamed eyes no drug was detectable in the aqueous and vitreous humor, and the corneas at any time point. Conclusions: In our model, caspofungin reached therapeutically relevant levels in the aqueous and cornea but not in the vitreous humor of inflamed eyes. Intraocular drug deposition was critically dependent on a disrupted blood-eye barrier. These findings suggest a limited role for caspofungin in the treatment of fungal endophthalmiti

Tocotrienol inhibits proliferation of human Tenon's fibroblasts in vitro: a comparative study with vitamin E forms and mitomycin C

Purpose: To evaluate the potential of the vitamin E compound α-tocotrienol as antifibrotic agent in vitro. Methods: Using human Tenon's capsule fibroblast cultures, the antiproliferative and cytotoxic effects of the different vitamin E forms α-tocopherol, α-tocopheryl acetate, α-tocopheryl succinate and α-tocotrienol were compared with those of mitomycin C. To mimic subconjunctival and regular oral application in vivo, exposure time of serum-stimulated and serum-restimulated fibroblasts (SF and RF, respectively) to vitamin E forms was set at 6 days. Cultures were only exposed for 5min to mitomycin C due to its known acute toxicity and to mimic the short-time intraoperative administration. Proliferation (expressed as % of control) was determined by DNA content quantification on days 2, 4 and 6, whereas cytotoxicity was assessed by cell morphology and glucose 6-phosphate dehydrogenase (G6PD) release after 24h. Results: α-Tocopherol and α-tocopheryl acetate stimulated growth of SF, but not RF. Reduction of fibroblast content by α-tocopheryl succinate was accompanied by increased G6PD release and necrosis. Contrary to α-tocopheryl succinate, 50μM or repeatedly 20μM of α-tocotrienol significantly inhibited proliferation without causing cellular toxicity (maximal effect: 46.8%). RF were more sensitive to this effect than SF. Mitomycin C 100-400μg/ml showed a stronger antiproliferative effect than α-tocotrienol (maximal effect: 13.8%). Morphologic characteristics of apoptosis were more commonly found under treatment with mitomycin C. Conclusions: Of the vitamin E forms tested, only α-tocotrienol significantly inhibited growth at non-toxic concentrations. In this in vitro study, antiproliferative effects of mitomycin C were stronger than those of α-tocotrieno

Antifibrotic effects of tocotrienols on human Tenon's fibroblasts

Purpose: To compare the antifibrotic effect of vitamin E isoforms α-, γ-, and δ-tocotrienol on human Tenon's fibroblasts (hTf) to the antimetabolite mitomycin C. Methods: Antifibrotic effects of α- (40, 60, 80, 100, and 120μM), γ- (10, 20, 30, and 40μM) and δ-tocotrienol (10, 20, 30, and 40μM) on hTf cultures were evaluated by performing proliferation, migration and collagen synthesis assays. Whereas for vitamin E the exposure time was set to 7days to mimic subconjunctival application, cultures were exposed only 5 min to mitomycin C 100μg/ml to mimic intraoperative administration. Cell morphology (phase contrast microscopy) as an assessment for cytotoxicity and cell density by measuring DNA content in a fluorometric assay to determine proliferation inhibition was performed on day 0, 4, and 7. Migration ability and collagen synthesis of fibroblasts were measured. Results: All tested tocotrienol isoforms were able to significantly inhibit hTf proliferation in a dose-dependent manner (maximal inhibitory effect without relevant morphological changes at day 4 for α-tocotrienol 80μM with 36.7% and at day 7 for α-tocotrienol 80μM with 42.6% compared to control). Degenerative cell changes were observed in cultures with concentrations above 80μM for α- and above 30μM for γ- and δ-tocotrienol. The highest collagen synthesis inhibition has been found with 80 µM α-tocotrienol (62.4%) and no significant inhibition for mitomycin C (2.5%). Migration ability was significantly reduced in cultures exposed to 80 µM α- and 30 µM γ-tocotrienol (inhibition of 82.2% and 79.5%, respectively, compared to control) and also after mitomycin C treatment (60.0%). Complete growth inhibition without significant degenerative cell changes could only be achieved with mitomycin C. Conclusion: In vitro, all tested tocotrienol isoforms were able to inhibit proliferation, migration and collagen synthesis of human Tenon's fibroblasts and therefore may have the potential as an anti-scarring agent in filtrating glaucoma surger

Impact of cerebral hypoperfusion-reperfusion on optic nerve integrity and visual function in the DBA/2J mouse model of glaucoma.

OBJECTIVE One of the most important risk factors for developing a glaucomatous optic neuropathy is elevated intraocular pressure. Moreover, mechanisms such as altered perfusion have been postulated to injure the optical path. In a mouse model, we compare first negative effects of cerebral perfusion/reperfusion on the optic nerve structure versus alterations by elevated intraocular pressure. Second, we compare the alterations by isolated hypoperfusion-reperfusion and isolated intraocular pressure to the combination of both. METHODS AND ANALYSIS Mice were divided in four groups: (1) controls; (2) perfusion altered mice that underwent transient bi-common carotid artery occlusion (BCCAO) for 40 min; (3) glaucoma group (DBA/2J mice); (4) combined glaucoma and altered perfusion (DBA/2J mice with transient BCCAO). Optic nerve sections were stereologically examined 10-12 weeks after intervention. RESULTS All experimental groups showed a decreased total axon number per optic nerve compared with controls. In DBA/2J and combined DBA/2J & BCCAO mice the significant decrease was roughly 50%, while BCCAO leaded to a 23% reduction of axon number, however reaching significance only in the direct t-test. The difference in axon number between BCCAO and both DBA/2J mice was almost 30%, lacking statistical significance due to a remarkably high variation in both DBA/2J groups. CONCLUSION Elevated intraocular pressure in the DBA/2J mouse model of glaucoma leads to a much more pronounced optic nerve atrophy compared with transient forebrain hypoperfusion and reperfusion by BCCAO. A supposed worsening effect of an altered perfusion added to the pressure-related damage could not be detected

Antifibrotic effects of tocotrienols on human Tenon's fibroblasts

Antifibrotic effects of α- (40, 60, 80, 100, and 120 μM), γ- (10, 20, 30, and 40 μM) and δ-tocotrienol (10, 20, 30, and 40 μM) on hTf cultures were evaluated by performing proliferation, migration and collagen synthesis assays. Whereas for vitamin E the exposure time was set to 7 days to mimic subconjunctival application, cultures were exposed only 5 min to mitomycin C 100 μg/ml to mimic intraoperative administration. Cell morphology (phase contrast microscopy) as an assessment for cytotoxicity and cell density by measuring DNA content in a fluorometric assay to determine proliferation inhibition was performed on day 0, 4, and 7. Migration ability and collagen synthesis of fibroblasts were measured. Results All tested tocotrienol isoforms were able to significantly inhibit hTf proliferation in a dose-dependent manner (maximal inhibitory effect without relevant morphological changes at day 4 for α-tocotrienol 80 μM with 36.7% and at day 7 for α-tocotrienol 80 μM with 42.6% compared to control). Degenerative cell changes were observed in cultures with concentrations above 80 μM for α- and above 30 μM for γ- and δ-tocotrienol. The highest collagen synthesis inhibition has been found with 80 µM α-tocotrienol (62.4%) and no significant inhibition for mitomycin C (2.5%). Migration ability was significantly reduced in cultures exposed to 80 µM α- and 30 µM γ-tocotrienol (inhibition of 82.2% and 79.5%, respectively, compared to control) and also after mitomycin C treatment (60.0%). Complete growth inhibition without significant degenerative cell changes could only be achieved with mitomycin C. Conclusion In vitro, all tested tocotrienol isoforms were able to inhibit proliferation, migration and collagen synthesis of human Tenon’s fibroblasts and therefore may have the potential as an anti-scarring agent in filtrating glaucoma surge

Lens epithelial cell apoptosis and intracellular Ca(2+) increase in the presence of xanthurenic acid

BACKGROUND: Xanthurenic acid is an endogenous product of tryptophan degradation by indoleamine 2,3-dioxygenase (IDO). We have previously reported that IDO is present in mammalian lenses, and xanthurenic acid is accumulated in the lenses with aging. Here, we studied the involvement of xanthurenic acid in the human lens epithelial cell physiology. METHODS: Human lens epithelial cells primary cultures were used. Control cells, and cells in the presence of xanthurenic acid grow in the dark. Western blot analysis and immunofluorescence studies were performed. RESULTS: In the presence of xanthurenic acid human lens epithelial cells undergo apoptosis-like cell death. In the control cells gelsolin stained the perinuclear region, whereas in the presence of 10 μM xanthurenic acid gelsolin is translocated to the cytoskeleton, but does not lead to cytoskeleton breakdown. In the same condition caspase-3 activation, and DNA fragmentation was observed. At low (5 to 10 μM) of xanthurenic acid concentration, the elongation of the cytoskeleton was associated with migration of mitochondria and cytochrome c release. At higher concentrations xanthurenic acid (20 μM and 40 μM) damaged mitochondria were observed in the perinuclear region, and nuclear DNA cleavage was observed. We observed an induction of calpain Lp 82 and an increase of free Ca(2+) in the cells in a xanthurenic acid concentration-dependent manner. CONCLUSIONS: The results show that xanthurenic acid accumulation in human lens epithelial cells disturbs the normal cell physiology and leads to a cascade of pathological events. Xanthurenic acid induces calpain Lp82 and caspases in the cells growing in the dark and can be involved in senile cataract development

Efficacy of Amniotic Membrane Transplantation for the Treatment of Corneal Ulcers.

PURPOSE To evaluate the outcome of amniotic membrane transplantation (AMTX) as a treatment for corneal ulcers. METHODS Patients treated with AMTX for refractory corneal ulcers between 2012 and 2017 were evaluated in a retrospective analysis. Primary outcome measure was complete reepithelialization. RESULTS A total of 149 patients were included (mean age 68 ± 18 years). The mean duration between ulcer onset and AMTX was 42 ± 46 days. The longest time between ulcer diagnosis and AMTX was found in bacterial ulcers and the shortest time to AMTX in eyes with trauma/chemical burns (mean 65 ± 15 days and 14 ± 4 days, respectively). In 70% of the patients, a single AMTX procedure was sufficient to achieve epithelial closure (21% <1 month, 40% within 1 -3 months, and 9% within 3-6 months). Treatment failure was observed in 30% of all patients, and most of them underwent further interventions. Highest closure rates were found in bacterial ulcers, herpetic ulcers, and neurotrophic ulcers (80%, 85%, and 93%, respectively), whereas the lowest reepithelialization rates were found in ulcers after corneal surgery and ulcers associated with rheumatic disease (52% and 57%, respectively). CONCLUSIONS AMTX is a valuable treatment option to achieve corneal epithelial wound healing in cases refractory to conventional treatment. Success rates differ depending on the etiology of ulcer

Keratoconus Progression After Corneal Cross-Linking in Eyes With Preoperative Maximum Keratometry Values of 58 Diopters and Steeper.

PURPOSE To evaluate the effectiveness of cross-linking (CXL) in treating keratoconus eyes with Kmax values ≥58.0 D. METHODS Retrospective analysis of outcomes of standard Dresden epi-off CXL in progressive keratoconus with preoperative Kmax ≥58.0 Diopters (D). Inclusion criteria were Kmax ≥58.0 D and minimum follow-up of 1 year. Corneal topography and tomography were performed preoperatively and at 1 and 2 years. Sixty-one eyes of 56 patients with mean age of 24.9 ± 8.6 years (mean ± SD, range 12-57 years) had 1-year follow-up. Fifty of these eyes had 2-year follow-up. The definition of progression was an increase in Kmax of ≥1.0 D over 1 year. RESULTS Mean Kmax was 63.9 ± 6.1 D (mean ± SD, range 58.2-87.0 D) preoperatively (n = 61) and 62.9 ± 5.9 D (range 54.6-82.5 D) after 1 year. This represented a significant decrease in steepness (P = 0.0029). Mean pachymetry decreased significantly from 433.7 ± 44.8 μm preoperatively to 423.0 ± 41.8 μm (P = 0.001) at 1 year. Progression occurred in 14 of the 61 eyes (23%) at 1 year, and 5 (8.2%) steepened more than 2.0 D. In the group with 2-year follow-up, mean Kmax was 63.0 ± 5.0 D (range 58.2-87 D) before CXL and decreased to 61.5 ± 4.8 D (range 53.6-78.3 D) at 2 years (P = 0.001). Nine of the 50 eyes (18%) showed an increase of Kmax of ≥ 1 D. CONCLUSIONS The incidence of progression (23% at 1 and 18% at 2 years, respectively) is considerably higher than in previously reported results of CXL in eyes with mean Kmax ≥58.0 D. To the best of our knowledge, this study represents the largest number of such steep corneas analyzed with respect to long-term progression after CXL