The global shortage of corneal endothelial graft tissue necessitates the exploration of alternative therapeutic strategies. Rho-associated protein kinase inhibitors (ROCKi), recognized for their regenerative potential in cardiology, oncology, and neurology, have shown promise in corneal endothelial regeneration. This study investigates the repurposing potential of additional ROCKi compounds. Through screening a self-assembled library of ROCKi on B4G12 corneal endothelial cells, we evaluated their dose-dependent effects on proliferation, migration, and toxicity using live-cell imaging. Nine ROCKi candidates significantly enhanced B4G12 proliferation compared to the basal growth rate. These candidates were further assessed for their potential to accelerate wound closure as another indicator for tissue regeneration capacity, with most demonstrating notable efficacy. To assess the potential impact of candidate ROCKi on key corneal endothelial cell markers related to cell proliferation, leaky tight junctions and ion efflux capacity, we analyzed the protein expression of cyclin E1, CDK2, p16, ZO-1 and Na+/K+-ATPase, respectively. Immunocytochemistry and western blot analysis confirmed the preservation of corneal endothelial markers post-treatment with ROCKi hits. However, notable cytoplasm enlargement and nuclear fragmentation were detected after the treatment with SR-3677 and Thiazovivin, indicating possible cellular stress. In compared parameters, Chroman-1 at a concentration of 10 nM outperformed other ROCKi, requiring significantly 1000-fold lower effective concentration than established ROCKi Y-27632 and Fasudil. Altogether, this study underscores the potential of repurposing ROCKi for treating corneal endothelial dysfunctions, offering a viable alternative to conventional grafting methods, and highlights Chroman-1 as a promising candidate structure for hit-to-lead development.
- Klíčová slova
- Chroman-1, Corneal endothelial regeneration, Drug repurposing, ROCK inhibitors, Small molecule screening,
- MeSH
- buněčné linie MeSH
- endoteliální buňky účinky léků MeSH
- inhibitory proteinkinas * farmakologie MeSH
- kinázy asociované s rho * antagonisté a inhibitory metabolismus MeSH
- lidé MeSH
- pohyb buněk účinky léků MeSH
- přehodnocení terapeutických indikací léčivého přípravku MeSH
- proliferace buněk * účinky léků MeSH
- regenerace * účinky léků MeSH
- rohovkový endotel * účinky léků MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- inhibitory proteinkinas * MeSH
- kinázy asociované s rho * MeSH
Fuchs endothelial corneal dystrophy (FECD) is an age-related cause of vision loss, and the most common repeat expansion-mediated disease in humans characterised to date. Up to 80% of European FECD cases have been attributed to expansion of a non-coding CTG repeat element (termed CTG18.1) located within the ubiquitously expressed transcription factor encoding gene, TCF4. The non-coding nature of the repeat and the transcriptomic complexity of TCF4 have made it extremely challenging to experimentally decipher the molecular mechanisms underlying this disease. Here we comprehensively describe CTG18.1 expansion-driven molecular components of disease within primary patient-derived corneal endothelial cells (CECs), generated from a large cohort of individuals with CTG18.1-expanded (Exp+) and CTG 18.1-independent (Exp-) FECD. We employ long-read, short-read, and spatial transcriptomic techniques to interrogate expansion-specific transcriptomic biomarkers. Interrogation of long-read sequencing and alternative splicing analysis of short-read transcriptomic data together reveals the global extent of altered splicing occurring within Exp+ FECD, and unique transcripts associated with CTG18.1-expansions. Similarly, differential gene expression analysis highlights the total transcriptomic consequences of Exp+ FECD within CECs. Furthermore, differential exon usage, pathway enrichment and spatial transcriptomics reveal TCF4 isoform ratio skewing solely in Exp+ FECD with potential downstream functional consequences. Lastly, exome data from 134 Exp- FECD cases identified rare (minor allele frequency <0.005) and potentially deleterious (CADD>15) TCF4 variants in 7/134 FECD Exp- cases, suggesting that TCF4 variants independent of CTG18.1 may increase FECD risk. In summary, our study supports the hypothesis that at least two distinct pathogenic mechanisms, RNA toxicity and TCF4 isoform-specific dysregulation, both underpin the pathophysiology of FECD. We anticipate these data will inform and guide the development of translational interventions for this common triplet-repeat mediated disease.
- MeSH
- alternativní sestřih genetika MeSH
- endoteliální buňky metabolismus MeSH
- expanze trinukleotidových repetic * genetika MeSH
- Fuchsova endoteliální dystrofie * genetika MeSH
- lidé MeSH
- rohovkový endotel metabolismus patologie MeSH
- transkripční faktor 4 * genetika metabolismus MeSH
- transkriptom genetika MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- TCF4 protein, human MeSH Prohlížeč
PURPOSE: To evaluate the safety and efficacy of using corneal stromal lenticules (CSLs) obtained during refractive surgery Refractive Lenticule Extraction (ReLEx) with the Small Incision Lenticule Extraction (SMILE) procedure for the treatment of corneal ulcers. METHODS: This retrospective study included 12 eyes of 12 patients, 7 men and 5 women with varying degrees of corneal ulcer. The mean age was 64 ± 18 (range 34 to 95 years). The monitoring included corrected distance visual acuity (CDVA), slit-lamp biomicroscopy examination, a Seidel test, stability of the graft and anterior segment optical coherence tomography (AS-OCT) inspection. Patients were closely monitored for possible postoperative complications for at least 6 months. RESULTS: In 7/12 (58%) eyes, the corneal ulcer was successfully sealed with CSL and amniotic membrane (AM) without the need for any additional surgical intervention. In 3 eyes, penetrating keratoplasty (PK) was needed in addition to CSL transplantation and in 2 eyes the scleral patch was used to fully seal after CSL transplantation. During the follow-up period no signs of rejection or infection were detected in any patient. CONCLUSION: The use of CSLs from ReLEx SMILE may be considered as an alternative method for the treatment of corneal ulcers before a more extensive and definitive solution - PK - is used. Our preliminary findings suggest that properly performed CSL transplantation using cryopreserved lenticules is a safe and effective method to temporarily cover the corneal partial-thickness defect or even perforation.
- Klíčová slova
- CSL transplantation, ReLEx SMILE, amniotic membrane, corneal stromal lenticule, corneal ulcer,
- MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- retrospektivní studie MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- stroma rohovky chirurgie MeSH
- transplantace rohovky * metody MeSH
- vřed rohovky * chirurgie MeSH
- vřed chirurgie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
AIM: To summarize the history and current trends in the use of scleral grafts in ophthalmology. MATERIALS AND METHODS: We conducted a review of the literature through the MEDLINE and Cochrane Library databases. The search terms were "sclera", "graft", and "surgery". The search resulted in 1596 articles, of which we evaluated 192 as relevant. The relevant articles were sorted chronologically and according to the method of using scleral grafts, which enabled the development of a review article. RESULTS: The sclera has been routinely used in ophthalmology since the 1950s in many different indications. Some of these indications have become practically obsolete over time (for example, use in the surgical management of retinal detachment), but a large number still find application today (especially use in glaucoma or oculoplastic surgery, or as a patch for a defect in the sclera or cornea). CONCLUSION: Even though allogeneic sclera is currently used less frequently in ophthalmology compared to other tissue banking products and the range of its indications has partially narrowed, it remains a useful material due to its availability and properties.
Pathogenic variants in the highly conserved OVOL2 promoter region cause posterior polymorphous corneal dystrophy (PPCD) 1 by inducing an ectopic expression of the endothelial OVOL2 mRNA. Here we produced an allelic series of Ovol2 promoter mutations in the mouse model including the heterozygous c.-307T>C variant (RefSeq NM_021220.4) causing PPCD1 in humans. Despite the high evolutionary conservation of the Ovol2 promoter, only some alterations of its sequence had phenotypic consequences in mice. Four independent sequence variants in the distal part of the Ovol2 promoter had no significant effect on endothelial Ovol2 mRNA level or caused any ocular phenotype. In contrast, the mutation c.-307T>C resulted in increased Ovol2 expression in the corneal endothelium. However, only a small fraction of adult mice c.-307T>C heterozygotes developed ocular phenotypes such as irido-corneal adhesions, and corneal opacity. Interestingly, phenotypic penetrance was increased at embryonic stages. Notably, c.-307T>C mutation is located next to the Ovol1/Ovol2 transcription factor binding site. Mice carrying an allele with a deletion encompassing the Ovol2 binding site c.-307_-320del showed significant Ovol2 gene upregulation in the cornea endothelium and exhibited phenotypes similar to the c.-307T>C mutation. In conclusion, although the mutations c.-307T>C and -307_-320del lead to a comparably strong increase in endothelial Ovol2 expression as seen in PPCD1 patients, endothelial dystrophy was not observed in the mouse model, implicating species-specific differences in endothelial cell biology. Nonetheless, the emergence of dominant ocular phenotypes associated with Ovol2 promoter variants in mice implies a potential role of this gene in eye development and disease.
- Klíčová slova
- Ovol2, PPCD1, cornea, corneal endothelium, pathogenic variant,
- MeSH
- dědičné dystrofie rohovky * genetika MeSH
- dospělí MeSH
- fenotyp MeSH
- lidé MeSH
- messenger RNA MeSH
- modely nemocí na zvířatech MeSH
- myši MeSH
- rohovkový endotel MeSH
- transkripční faktory genetika MeSH
- zvířata MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- messenger RNA MeSH
- Ovol2 protein, human MeSH Prohlížeč
- transkripční faktory MeSH
Refractive surgery with excimer laser has been a very common surgical procedure worldwide during the last decades. Currently, patients who underwent refractive surgery years ago are older, with a growing number of them now needing cataract surgery. To establish the power of the intraocular lens to be implanted in these patients, it is essential to define the true corneal power. However, since the refractive surgery modified the anterior, but not the posterior surface of the cornea, the determination of the corneal power in this group of patients is challenging. This article reviews the different sources of error in finding the true corneal power in these cases, and comments on several approaches, including the clinical history method as described originally by Holladay, and a modified version of it, as well as new alternatives based on corneal tomography, using devices that are able to measure the actual anterior and posterior corneal curvatures, which have emerged in recent years to address this issue.
- Klíčová slova
- corneal power, Refractive surgery, clinical history method, corneal tomography, refractive surgery,
- MeSH
- laserová modelace rohovky pod rohovkovou lamelou * MeSH
- lasery excimerové * terapeutické užití MeSH
- lidé MeSH
- refrakce oka MeSH
- rohovka chirurgie MeSH
- rohovková topografie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
AIM: To introduce the topic of pediatric keratoconus, highlighting the importance of routine corneal topography and tomography in children and adolescents from predisposed groups. To attempt to ensure the early detection of keratoconus and its subclinical form, enabling early treatment, which brings better expected postoperative results. Material and methods: Using the corneal tomograph Pentacam AXL we examined children and adolescents with astigmatism equal or greater than 2 diopters (in at least one eye) and patients with at least one risk factor such as eye rubbing in the case of allergic pathologies, positive family history of keratoconus or certain forms of retinal dystrophy. In total, we included 231 eyes (116 patients), of which 54 were girls and 62 were boys. RESULTS: The Belin-Ambrósio deviation index parameter was evaluated, in which we classified a total of 41 eyes as subclinical keratoconus and 12 eyes as clinical keratoconus. Next, the corneal maps were evaluated individually, in which we included a total of 15 eyes as subclinical keratoconus and 6 eyes as clinical keratoconus. In our group, compared to the control group, subclinical and clinical keratoconus occurred most often in the group of patients with astigmatism and in the group of so-called "eye rubbers". After individual evaluation, keratoconus occurred more frequently in boys than in girls in our cohort. CONCLUSION: Most patients with keratoconus are diagnosed when there is a deterioration of visual acuity and changes on the anterior surface of the cornea. Corneal topography and tomography allows us to monitor the initial changes on the posterior surface of the cornea, and helps us to detect the subclinical form of keratoconus and the possibility of its early treatment. Therefore, it is important to determine which groups are at risk and groups in which corneal topography and tomography should be performed routinely.
- Klíčová slova
- astigmatism, eye-rubbing, keratoconus, topography and tomography,
- MeSH
- astigmatismus * MeSH
- dítě MeSH
- keratokonus * diagnóza MeSH
- lidé MeSH
- mladiství MeSH
- oftalmologie * MeSH
- pachymetrie rohovky MeSH
- rohovka patologie MeSH
- rohovková topografie metody MeSH
- Check Tag
- dítě MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
Glaucoma is a leading cause of blindness worldwide, with elevated intraocular pressure being a major risk factor for its development and progression. First-line treatment for glaucoma relies on the administration of prostaglandin analogs, with latanoprost being the most widely used. However, before latanoprost reaches the cornea, it must pass through the tear film and tear film lipid layer (TFLL) on the ocular surface. Given the significant lipophilicity of latanoprost, we hypothesize that TFLL could, to a certain extent, act as a reservoir for latanoprost, releasing it on longer time scales, apart from the fraction being directly delivered to the cornea in a post-instillation mechanism. We investigated this possibility by studying latanoprost behavior in acellular in vitro TFLL models. Furthermore, we employed in silico molecular dynamics simulations to rationalize the experimental results and obtain molecular-level insight into the latanoprost-TFLL interactions. Our experiments demonstrated that latanoprost indeed accumulates in the TFLL models, and our simulations explain the basis of the accumulation mechanism. These results support the hypothesis that TFLL can serve as a reservoir for latanoprost, facilitating its prolonged release. This finding could have significant implications for optimizing glaucoma treatment, especially in the development of new drug delivery systems targeting the TFLL.
- Klíčová slova
- Glaucoma, Latanoprost, Ophthalmology, Tear film, Tear film lipid layer, Topical delivery,
- MeSH
- antihypertenziva terapeutické užití MeSH
- glaukom * farmakoterapie MeSH
- latanoprost terapeutické užití MeSH
- lidé MeSH
- nitrooční tlak MeSH
- počítačová simulace MeSH
- rohovka MeSH
- slzy MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- antihypertenziva MeSH
- latanoprost MeSH
Cannabidiol (CBD) is the non-psychoactive component of the plant Cannabis sativa (L.) that has great anti-inflammatory benefits and wound healing effects. However, its high lipophilicity, chemical instability, and extensive metabolism impair its bioavailability and clinical use. Here, we report on the preparation of a human cornea substitute in vitro and validate this substitute for the evaluation of drug penetration. CBD nanoemulsion was developed and evaluated for stability and biological activity. The physicochemical properties of CBD nanoemulsion were maintained during storage for 90 days under room conditions. In the scratch assay, nanoformulation showed significantly ameliorated wound closure rates compared to the control and pure CBD. Due to the lower cytotoxicity of nanoformulated CBD, a higher anti-inflammatory activity was demonstrated. Neither nanoemulsion nor pure CBD can penetrate the cornea after the four-hour apical treatment. For nanoemulsion, 94 % of the initial amount of CBD remained in the apical compartment while only 54 % of the original amount of pure CBD was detected in the apical medium, and 7 % in the cornea, the rest was most likely metabolized. In summary, the nanoemulsion developed in this study enhanced the stability and biological activity of CBD.
- Klíčová slova
- Anti-inflammatory, Bioavailability, Cannabidiol, Human corneal substitute, Nanoemulsion, Wound healing,
- MeSH
- antiflogistika farmakologie MeSH
- biologická dostupnost MeSH
- hojení ran MeSH
- kanabidiol * chemie MeSH
- lidé MeSH
- rohovka MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- antiflogistika MeSH
- kanabidiol * MeSH
PURPOSES: The primary aim of the study was to assess the response of intraocular pressure (IOP) to the acute ingestion of hydrogen-rich water (HRW) compared to hydrogen-free water (placebo) in healthy subjects. The effect of HRW intake on central corneal thickness (CCT) was also monitored. SUBJECTS AND METHODS: Twenty-four healthy volunteers (5 men, 19 women) aged between 20 and 33 were included in the study, in which one eye of each subject was measured. The study was prospective, randomized and double-blind, with crossover design. Each subject underwent two parts of the experiment, each part on a different day and in random order. In each part of experiment, a total volume of 1260 ml of HRW or placebo was administered over 15 minutes in three doses. IOP and CCT were measured before and during the course of 75 minutes from the start of the HRW or placebo intake. RESULTS: Administration of both HRW and the placebo caused a significant increase in IOP. The maximum IOP increase was 2.7 mmHg ±2.0 mmHg in minute 25 after the commencement of the experiment (HRW intake), and 1.4 mmHg ±2.0 mmHg in minute 35 (placebo intake). The values of IOP did not differ significantly between both parts, but there were significantly more clinically significant individual IOP increases after HRW intake (58%) compared to the placebo (25%). CCT did not change significantly during the experiment. CONCLUSION: The rapid intake of 1260 ml of both HRW and hydrogen-free water causes a statistically significant increase in IOP compared to the baseline in healthy individuals. In the case of HRW, the increase was also clinically significant in most of the subjects. Thus, the results indicate that acute intake of HRW may pose a higher risk than placebo intake in terms of IOP. However, in the case of risk groups such as subjects with glaucoma, ocular hypertension or suspected glaucoma, it is necessary to verify this conclusion by further studies.
- Klíčová slova
- corneal thickness, hydrogen-rich water, intraocular pressure, molecular hydrogen, water,
- MeSH
- dospělí MeSH
- dvojitá slepá metoda MeSH
- glaukom * MeSH
- klinické křížové studie MeSH
- lidé MeSH
- mladý dospělý MeSH
- nitrooční tlak MeSH
- oční hypertenze * MeSH
- pití MeSH
- prospektivní studie MeSH
- rohovka MeSH
- tonometrie oční MeSH
- voda MeSH
- zdraví dobrovolníci pro lékařské studie MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- randomizované kontrolované studie MeSH
- Názvy látek
- voda MeSH
