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The challenge of treating corneal scarring through keratoplasties lies in the limited availability of donor tissue. Various studies have shown the therapeutic use of cultivated corneal stromal stem cells (CSSCs) to mitigate tissue inflammation and suppress fibrosis and scar tissue formation in preclinical corneal wound models. To develop CSSC therapy for clinical trials on patients with corneal scarring, it is necessary to generate clinical-grade CSSCs in compliant to Good Manufacturing Practice (GMP) regulations. This chapter elucidates human CSSC isolation, culture, and cryopreservation under GMP-compliant conditions. It underscores quality assessment encompassing morphological traits, expression of stemness markers, anti-inflammatory activity, and keratocyte differentiation potency., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Corneal scarring, a significant cause of global blindness, results from various insults, including trauma, infections, and genetic disorders. The conventional treatment to replace scarred corneal tissues includes partial or full-thickness corneal transplantation using healthy donor corneas. However, only 1 in 70 individuals with treatable corneal scarring can undergo surgery, due to the limited supply of transplantable donor tissue. Our research focuses on cell-based strategies, specifically ex vivo-expanded corneal stromal stem cells (CSSCs), to address corneal scarring. Preclinical studies have demonstrated the efficacy of CSSC treatment in reducing corneal inflammation and fibrosis, inhibiting scar formation, and regenerating native stromal tissue. Mechanisms include CSSC differentiation into stromal keratocytes and the expression of regenerative cytokines. Here, we present a good manufacturing practice (GMP)-compliant protocol to isolate and expand human CSSCs. This method paves the way to produce clinical-grade CSSCs for transplantation and clinical trials. Key features • This protocol utilizes surgical skills to dissect human corneal tissues for CSSC isolation. • The yield and features of CSSCs rely on donor tissue quality (freshness) and have donor-to-donor variability. • Up to 0.5 billion CSSCs can be generated from a single cornea specimen, and cells at passage 3 are suitable for treatment uses., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (©Copyright : © 2024 The Authors; This is an open access article under the CC BY license.)

Intrastromal cell therapy utilizing quiescent corneal stromal keratocytes (qCSKs) from human donor corneas emerges as a promising treatment for corneal opacities, aiming to overcome limitations of traditional surgeries by reducing procedural complexity and donor dependency. This investigation demonstrates the therapeutic efficacy of qCSKs in a male rat model of corneal stromal opacity, underscoring the significance of cell-delivery quality and keratocyte differentiation in mediating corneal opacity resolution and visual function recovery. Quiescent CSKs-treated rats display improvements in escape latency and efficiency compared to wounded, non-treated rats in a Morris water maze, demonstrating improved visual acuity, while stromal fibroblasts-treated rats do not. Advanced imaging, including multiphoton microscopy, small-angle X-ray scattering, and transmission electron microscopy, revealed that qCSK therapy replicates the native cornea's collagen fibril morphometry, matrix order, and ultrastructural architecture. These findings, supported by the expression of keratan sulfate proteoglycans, validate qCSKs as a potential therapeutic solution for corneal opacities., (© 2024. The Author(s).)

Corneal nerves originate from the ophthalmic branch of the trigeminal nerve, which enters the cornea at the limbus radially from all directions toward the central cornea. The cell bodies of the sensory neurons of trigeminal nerve are located in the trigeminal ganglion (TG), while the axons are extended into the three divisions, including ophthalmic branch that supplies corneal nerves. Study of primary neuronal cultures established from the TG fibers can therefore provide a knowledge basis for corneal nerve biology and potentially be developed as an in vitro platform for drug testing. However, setting up primary neuron cultures from animal TG has been dubious with inconsistency among laboratories due to a lack of efficient isolation protocol, resulting in low yield and heterogenous cultures. In this study, we used a combined enzymatic digestion with collagenase and TrypLE to dissociate mouse TG while preserving nerve cell viability. A subsequent discontinuous Percoll density gradient followed by mitotic inhibitor treatment effectively diminished the contamination of non-neuronal cells. Using this method, we reproducibly generated high yield and homogenous primary TG neuron cultures. Similar efficiency of nerve cell isolation and culture was further obtained for TG tissue cryopreserved for short (1 week) and long duration (3 months), compared to freshly isolated tissues. In conclusion, this optimized protocol shows a promising potential to standardize TG nerve culture and generate a high-quality corneal nerve model for drug testing and neurotoxicity studies.

The human corneal stroma contains corneal stromal keratocytes (CSKs) that synthesize and deposit collagens and keratan sulfate proteoglycans into the stromal matrix to maintain the corneal structural integrity and transparency. In adult corneas, CSKs are quiescent and arrested in the G0 phase of the cell cycle. Following injury, some CSKs undergo apoptosis, whereas the surviving cells are activated to become stromal fibroblasts (SFs) and myofibroblasts (MyoFBs), as a natural mechanism of wound healing. The SFs and MyoFBs secrete abnormal extracellular matrix proteins, leading to corneal fibrosis and scar formation (corneal opacification). The issue is compounded by the fact that CSK transformation into SFs or MyoFBs is irreversible in vivo, which leads to chronic opacification. In this scenario, corneal transplantation is the only recourse. The application of cell therapy by replenishing CSKs, propagated in vitro, in the injured corneas has been demonstrated to be efficacious in resolving early-onset corneal opacification. However, expanding CSKs is challenging and has been the limiting factor for the application in corneal tissue engineering and cell therapy. The supplementation of serum in the culture medium promotes cell division but inevitably converts the CSKs into SFs. Similar to the in vivo conditions, the transformation is irreversible, even when the SF culture is switched to a serum-free medium. In the current article, we present a detailed protocol on the isolation and propagation of bona fide human CSKs and the morphological and genotypic differences from SFs.

With the expected rise in patients undergoing refractive lenticule extraction worldwide, the number of discarded corneal stromal lenticules will increase. Therefore, establishing a lenticule bank to collect, catalog, process, cryopreserve, and distribute the lenticules (for future therapeutic needs) could be advantageous. In this study, we validated the safety of lenticule banking that involved the collection of human lenticules from our eye clinic, transportation of the lenticules to a Singapore Ministry of Health-licensed lenticule bank, processing, and cryopreservation of the lenticules, which, after 3 months or, a longer term, 12 months, were retrieved and transported to our laboratory for implantation in rabbit corneas. The lenticule collection was approved by the SingHealth Centralised Institutional Review Board (CIRB). Both short-term and long-term cryopreserved lenticules, although not as transparent as fresh lenticules due to an altered collagen fibrillar packing, did not show any sign of rejection and cytotoxicity, and did not induce haze or neovascularization for 16 weeks even when antibiotic and steroidal administration were withdrawn after 8 weeks. The lenticular transparency progressively improved and was mostly clear after 4 weeks, the same period when we observed the stabilization of corneal hydration. We showed that the equalization of the collagen fibrillar packing of the lenticules with that of the host corneal stroma contributed to the lenticular haze clearance. Most importantly, no active wound healing and inflammatory reactions were seen after 16 weeks. Our study suggests that long-term lenticule banking is a feasible approach for the storage of stromal lenticules after refractive surgery. Impact statement Since 2011, close to 3 million refractive lenticule extraction procedures have been performed. The majority of the extracted lenticules are discarded. The lenticules could have been cryopreserved and retrieved at a later date for therapeutic or refractive applications. Therefore, establishing a lenticule bank to collect, catalog, process, cryopreserve, and distribute the lenticules could be advantageous. In this study, we simulated a lenticule banking service in a validated health authority-licensed facility and showed that long-term cryopreservation of the lenticules in the facility was safe and feasible in vivo .

Purpose: Cornea epithelial-stromal scarring is related to the differentiation of fibroblasts into opaque myofibroblasts. Our study aims to assess the effectiveness of Lycium barbarum polysaccharide (LBP) solution as a pre-treatment in minimizing corneal scarring., Methods: Human corneal fibroblasts were cultured in a three-dimensional collagen type I-based hydrogel in an eye-on-a-chip model. Fibroblasts were pre-treated with 2 mg/mL LBP for 24 h, followed by another 24-h incubation with 10 ng/mL transforming growth factor-beta 1 (TGF-β1) to induce relevant physiological events after stromal injury. Intracellular pro-fibrotic proteins, extracellular matrix proteins, and pro-inflammatory cytokines that involved in fibrosis, were assessed using immunocytochemistry and enzyme-linked immunosorbent assays., Results: Compared to the positive control TGF-β1 group, LBP pre-treated cells had a significantly lower expression of alpha-smooth muscle actin, marker of myofibroblasts, vimentin (p < 0.05), and also extracellular matrix proteins both collagen type II and type III (p < 0.05) that can be found in scar tissues. Moreover, LBP pre-treated cells had a significantly lower secretion of pro-inflammatory cytokines interleukin-6 and interleukin-8 (p < 0.05). The cell-laden hydrogel contraction and stiffness showed no significant difference between LBP pre-treatment and control groups. Fibroblasts pretreated with LBP as well had reduced angiogenic factors expression and suppression of undesired proliferation (p < 0.05)., Conclusion: Our results showed that LBP reduced both pro-fibrotic proteins and pro-inflammatory cytokines on corneal injury in vitro. We suggest that LBP, as a natural Traditional Chinese Medicine, may potentially be a novel topical pre-treatment option prior to corneal refractive surgeries with an improved prognosis., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The study of corneal stromal keratocytes is motivated by its strong association with corneal health and visual function. They play a dominant role in the maintenance of corneal homeostasis and transparency through the production of collagens, proteoglycans and corneal crystallins. Trauma-induced apoptosis of keratocytes and replacement by fibroblasts and myofibroblasts disrupt the stromal matrix organization, resulting in corneal haze formation and vision loss. It is, therefore, important to understand the biology and behaviours of keratocytes and the associated stromal cell types (like fibroblasts, myofibroblasts, stromal stem cells) in wound healing, corneal pathologies (including keratoconus, keratitis, endothelial disorders) as well as different ophthalmic situations (such as collagen crosslinking/photodynamic treatment, keratoplasty and refractive surgery, and topical medications). The recent development of ex vivo propagation of keratocytes and stromal stem cells, and their translational applications, either via stromal injection or incorporated in bioscaffold, have been shown to restore the corneal transparency and regenerate native stromal tissue in animal models of corneal haze and other disorders., Competing Interests: Declaration of competing interest All authors indicated no potential conflicts of interest., (Published by Elsevier Ltd.)

Purpose: To investigate the postoperative inflammatory and wound-healing responses after laser scleral microporation for presbyopia., Methods: Thirty porcine eyes were used for the optimization of laser intensities first. Six monkeys (12 eyes) received scleral microporation with an erbium yttrium aluminum garnet (Er:YAG) laser, and half of the eyes received concurrent subconjunctival collagen gel to modulate wound-healing response. The intraocular pressure (IOP) and the laser ablation depth were evaluated. The animals were euthanized at 1, 6, and 9 months postoperatively. The limbal areas and scleras were harvested for histologic analysis and immunofluorescence of markers for inflammation (CD11b and CD45), wound healing (CD90, tenascin-C, fibronectin, and HSP47), wound contraction (α-smooth muscle actin [α-SMA]), vascular response (CD31), nerve injury (GAP43), and limbal stem cells (P63 and telomerase)., Results: In the nonhuman primate study, there was a significant reduction in IOP after the procedure. Overall, the ablation depth was 76.6% to 81.2% at 1 month and slightly decreased to 71.5% to 72.7% at 9 months. Coagulative necrosis around the micropores, as well as expression of CD11b, CD45, tenascin, fibronectin, HSP47, and GAP43, was distinct at 1 month but subsided with time. Collagen gel treatment significantly suppressed the upregulation of CD11b, CD45, fibronectin, and tenascin-C. The expression of CD90, α-SMA, and CD31 was minimal in all eyes., Conclusions: The study demonstrated the course of inflammatory and wound-healing responses following laser scleral microporation. The tissue responses were small and self-limited, resolved with time, and were suppressed by concurrent collagen treatment. It provides a useful understanding of this new procedure., Translational Relevance: The results would be helpful in the laser parameter modification to improve the long-term treatment stability., Competing Interests: Disclosure: Y.-C. Liu, None; B. Hall, None; N.C. Lwin, None; E.P.W. Teo, None; G.H.F. Yam, None; A. Hipsley, VisioLite Gen II (P); J.S. Mehta, None, (Copyright 2020 The Authors.)

Stromal keratophakia was first performed by José Ignacio Barraquer in the 1960s. The refractive lamellar keratoplasty technique was intensely pursued in the 1980s as a method to alter corneal refractive power. However, because sculpting of the donor stromal lenticule and lamellar keratectomy of the recipient's cornea were performed with a mechanical microkeratome, the quality of the cut was inconsistent. Consequently, the refractive outcomes of the lenticule implantation were poor. In addition, epithelial ingrowth, interface scarring, and induced astigmatism were common due to the manual resection. With the advancements of femtosecond laser, we are now able to optically sculpt a refractive lenticule and create an intrastromal pocket for implantation, with greater accuracy and precision compared to manual incisions. The lenticule can be decellularized, cryopreserved, and implanted on a later date to correct hyperopia and presbyopia, as well as to treat corneal ectasia and perforations. In this article, we will review the history of stromal keratophakia and the shortcomings of the previous attempts that led to its abandonment. We will then discuss the reinvigoration of stromal keratophakia with the emergence of advanced femtosecond laser technologies, including the basic science and clinical applications of femtosecond laser-assisted stromal keratophakia, methods to decellularize, cryopreserve and transport the refractive lenticule, lenticule banking, and regulatory framework that oversees the distribution and clinical translation of stromal lenticule implantation., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Recent literature has ascribed that the paracrine action of stem cells is mediated by exosomes. Exosomes are nano-sized extracellular vesicles (30 to 100 nm) of endocytic origin that play important roles in intercellular communication. They have the ability to deliver various therapeutic effects, e.g., skin regeneration or cardiac function recovery, when applied topically or injected systemically. However, injection of exosomes has been shown to result in rapid clearance from blood circulation and accumulation of the exosomes in the liver, spleen, lung, and gastrointestinal tract can be found as early as 2 h after injection. Topical administration of exosomes on the skin or ocular surface would suffer the same fate due to rapid fluid turnover (sweat or tears). Biodegradable or highly porous hydrogels have been utilized to load exosomes and to deliver a sustained therapeutic effect. They can also prevent the exosomes from being cleared prematurely and allow the delivery of a more localized and concentrated exosome dosage by placing the hydrogel directly at or in the proximity of the target site. In this mini-review, we elaborate on the challenges of conventional exosome administration and highlight the solution to the shortcomings in the form of exosome-incorporated hydrogels. Different techniques to encapsulate exosomes and examples of hydrogels that have been used to create sustained delivery systems of exosomes are also discussed., (Copyright © 2019 Riau, Ong, Yam and Mehta.)

Background: Human corneal stromal keratocytes propagated in culture media supplemented with human amnion extract (AME) can correct early corneal haze in an animal model. Clinical application of cultivated keratocytes is limited by infectious disease screening before amnion products can be used in humans. It remains unclear if AME from cryopreserved versus fresh human amnion can support human keratocyte propagation, and which components of the extract promote keratocyte growth., Methods: Three placentas were collected for the preparation of fresh and cryopreserved amnion tissues followed by homogenization and protein extraction. AME protein profiles were studied using isobaric tagging for relative and absolute quantitation (iTRAQ) proteomics. Enriched gene ontology (GO) terms and functional classes were identified. Primary human keratocytes from 4 donor corneas were cultured in media supplemented with fresh AME (F-AME) or cryopreserved AME (C-AME). Cell viability, proliferation and keratocyte marker expression were examined by confocal immunofluorescence and flow cytometry., Results: AME proteomics revealed 1385 proteins with similar expression levels (between 0.5- and 2-fold) between F- and C-AME, while 286 proteins were reduced (less than 0.5-fold) in C-AME. Enriched GO term and biological pathway analysis showed that those proteins with comparable expression between F-AME and C-AME were involved in cell metabolism, epithelial-mesenchymal transition, focal adhesion, cell-extracellular matrix interaction, cell stress regulation and complement cascades. Human corneal stromal keratocytes cultured with F-AME or C-AME showed similar morphology and viability, while cell proliferation was mildly suppressed with C-AME ( P  > 0.05). Expression of aldehyde dehydrogenase 3A1 (ALDH3A1) and CD34 was similar in both cultures., Conclusion: AME from cryopreserved amnion had limited influence on keratocyte culture. It is feasible to use protein extract from cryopreserved amnion to propagate human keratocytes for potential translational applications., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s). 2019.)

: Bacterial biofilm on medical devices is difficult to eradicate. Many have capitalized the anti-infective capability of silver ions (Ag + ) by incorporating nano-silver (nAg) in a biodegradable coating, which is then laid on polymeric medical devices. However, such coating can be subjected to premature dissolution, particularly in harsh diseased tissue microenvironment, leading to rapid nAg clearance. It stands to reason that impregnating nAg directly onto the device, at the surface, is a more ideal solution. We tested this concept for a corneal prosthesis by immobilizing nAg and nano-hydroxyapatite (nHAp) on poly(methyl methacrylate), and tested its biocompatibility with human stromal cells and antimicrobial performance against biofilm-forming pathogens, Pseudomonas aeruginosa and Staphylococcus aureus . Three different dual-functionalized substrates-high Ag (referred to as 75:25 HAp:Ag); intermediate Ag (95:5 HAp:Ag); and low Ag (99:1 HAp:Ag) were studied. The 75:25 HAp:Ag was effective in inhibiting biofilm formation, but was cytotoxic. The 95:5 HAp:Ag showed the best selectivity among the three substrates; it prevented biofilm formation of both pathogens and had excellent biocompatibility. The coating was also effective in eliminating non-adherent bacteria in the culture media. However, a 28-day incubation in artificial tear fluid revealed a ~40% reduction in Ag + release, compared to freshly-coated substrates. The reduction affected the inhibition of S. aureus growth, but not the P. aeruginosa . Our findings suggest that Ag + released from surface-immobilized nAg diminishes over time and becomes less effective in suppressing biofilm formation of Gram-positive bacteria, such as S. aureus . This advocates the coating, more as a protection against perioperative and early postoperative infections, and less as a long-term preventive solution., Competing Interests: The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript.

Purpose: To compare the aqueous oxidative stress, prostaglandin E 2 (PGE 2 ), and cytokine levels after low-energy femtosecond laser-assisted cataract surgery versus conventional phacoemulsification, and to evaluate the effect of a nonsteroidal antiinflammatory drug (NSAID) on the aqueous profiles., Setting: Singapore National Eye Center, Singapore., Design: Randomized controlled trial., Methods: Thirty-five patients were randomized to receive conventional phacoemulsification in one eye and femtosecond laser-assisted cataract surgery in the fellow eye. Another matched prospective cohort of 35 patients was included to receive femtosecond laser-assisted cataract surgery with a preoperative NSAID. Aqueous humor was collected after the laser was used or at the beginning of surgery (conventional phacoemulsification), and at the end of phacoemulsification. The levels of aqueous malondialdehyde (MDA), PGE 2 , and cytokines, chemokines, and growth factors were analyzed. The postoperative aqueous flare levels were evaluated., Results: Compared with the conventional group, the femtosecond-assisted group had a significantly higher PGE 2 (P = .01) and interleukin-1 receptor antagonist levels (P = 0.04). The preoperative NSAID significantly reduced the PGE 2 surge (P = .002) and the mean reduction in pupil diameter (P = .02). The MDA concentrations before phacoemulsification were 0.24 μmol/L ± 0.18 (SD), 0.51 ± 0.41 μmol/L and 0.59 ± 0.52 μmol/L for the conventional, femtosecond-assisted, and femtosecond-assisted NSAID groups, respectively (P = .42). After phacoemulsification, the PGE 2 and MDA levels increased in all groups. The MDA induction was significantly correlated with the phacoemulsification time (P = .002). The postoperative flare was insignificantly higher in the femtosecond-assisted group than the conventional group., Conclusions: Compared with conventional phacoemulsification, the femtosecond laser-assisted cataract surgery performed with the Femto LDV Z8 laser platform induced a significantly higher PGE 2 level. The MDA and postoperative aqueous flare level were insignificantly higher. The preoperative NSAID reduced the PGE 2 surge and occurrence of intraoperative miosis. The oxidative stress induced during phacoemulsification was strongly correlated with phacoemulsification time., (Copyright © 2018 ASCRS and ESCRS. Published by Elsevier Inc. All rights reserved.)

This pilot study uses a micro-optical coherence tomography (micro-OCT) system with ~1 μm axial resolution specifically to image the cornea and corneal scars in vivo. We used an established murine corneal scar model by irregular phototherapeutic keratectomy in ten C57BL/6 mice, with serial imaging using the micro-OCT and compared to anterior segment (AS-OCT) (RTvue, Optovue, Fremont, CA) before and after scar induction. Main outcome was agreement between the AS-OCT and micro-OCT using Bland-Altman plots (95% limits of agreement, LoA).We analysed 10 control eyes and 10 eyes with corneal scars and found that there was good agreement between AS-OCT and micro-OCT (P > 0.05) LOA: lower limit -14 µm (95% CI: -19 to -8.8 µm) upper limit 23 µm (95% CI: 18 to 28.5 µm) in terms of central corneal thickness. There was also good agreement between AS-OCT and micro-OCT in terms of corneal scar measurements (P > 0.5; correlation coefficient >0.99) LOA lower limit -2.1 µm (95% CI: -2.8 to -1.5 µm); upper limit 1.8 µm (95% CI: 1.1 to 2.4 µm). Our pilot study suggests that this novel in vivo micro-OCT imaging technique was able to measure central corneal thickness and scar thickness in agreement with current AS-OCT techniques.

Corneal inlays are a relatively new treatment option for presbyopia. Using biological inlays, derived from lenticules extracted from small incision lenticule extraction, may offer advantages over commercialized synthetic inlays in the aspect of biocompatibility. We conducted a non-human primate study to evaluate the safety, predictability, efficacy and tissue response after autogeneic, decellularized xenogeneic and xenogeneic lenticule implantation. The lenticule implantation effectively resulted in central corneal steepening (simulated keratometric values increased by 1.8-2.3 diopters), central hyper-prolate changes (asphericity Q values changed by -0.26 to -0.36), corneal anterior surface elevation (7.7-9.3 μm) and reasonable effective zone (1.5-1.8 times of the lenticule physical diameter), with no differences among the three groups. Slit lamp microscopy, transmission electron microscopy, confocal microscopy, histology and immunohistochemistry analyses confirmed the biocompatibility of the autogeneic and decellularized lenticules, whereas one eye in the xenogeneic group developed corneal stromal rejection during the study period. Our results showed that lenticule implantation has the potential for the management of presbyopia, and provide the basis for future clinical studies. The decellularization process may increase the potential utilization of lenticules without changing the efficacy.

Purpose: To explore the optimal lenticule storage conditions that maintain lenticule integrity and clarity., Methods: A total of 99 lenticules obtained from myopic patients undergoing small incision lenticule extraction (SMILE) were divided into four combinations for short-term storage conditions: PBS, Dulbecco's Modified Eagle's Medium (DMEM), Optisol GS, or anhydrous glycerol. Two thirds of the lenticules were further stored for 4 weeks under eight different conditions. Clarity evaluation with transmittance measurements, cell-death assays with terminal deoxynucleotidyl transferase-mediated nick end labeling assay (TUNEL), collagen fibril spacing and necrotic response assessed with transmission electron microscopy (TEM), and immunohistochemistry analysis for human leukocyte antigens (HLAs) and CD45 for immunogenicity, and matrix metalloproteinase (MMP)-2 for keratocyte response, were undertaken at baseline, 48 h (short term), and 4 weeks (long term)., Results: The TUNEL and immunogenicity results were comparable among the groups. The mean percentage of TUNEL-positive cells across all groups was 24.3% ± 11.8% and 62.9% ± 20.7% at the 48 h and 4 week time points, respectively. HLA-ABC+, HLA-DR+, and CD45+ cells were extremely rare, and MMP-2 expression ranged from non-detectable to minimal, under all conditions at all time points. Transmittance at 4 weeks was significantly different among groups with the greatest maintenance of clarity seen in the lenticules stored initially in DMEM at 4 °C for 48 h followed by cryopreservation in serum-free medium or glycerol at 4 °C followed by storage at room temperature. At TEM analysis at 4 weeks, the lenticules cryopreserved in liquid nitrogen, regardless of storage solutions, had significantly narrower inter-fibrillar distance than controls, while glycerol-preserved lenticules, at either room temperature or -80 °C, maintained the inter-fibrillar distance., Conclusions: Clarity, structural integrity, and low immunogenicity under various conditions, at 4 °C or room temperature for short-term storage, offer encouragement for lenticule storage. It can be undertaken without access to s specialized and potentially expensive laboratory setup at least within the first 48 h before transportation to larger facilities for long-term storage.

Purpose: To determine the effects of the Ziemer LDV Z8 liquid interface femtosecond laser platform during capsulotomy under different energy settings in the presence of corneal edema., Methods: Cadaveric porcine eyes (n = 36) employed at less than 6 and greater than 24 post enucleation hours to simulate clear/edematous corneas, underwent capsulotomy with the Ziemer LDV Z8 femtosecond laser (5-mm diameter, energy 90%, 130%, or 150%). Lens capsules were removed for evaluation by scanning electron microscopy and rupture strengths determined by the single column universal testing system. Following ethical approval, 23 patients had lens capsules removed during routine cataract surgery following manual or Z8 capsulotomy and subjected to TUNEL assay., Results: There was no difference in edge morphology or rupture strength (120, 113, and 118 mN at increasing energy, P = 0.42) in the clear cornea. Only 50% of capsulotomies succeeded at 90% energy in an edematous cornea, improving with increased energy (75% completion at 130%, 100% at 150%). Rupture strength in edematous corneas was not significantly different at 112, 133, and 114 mN for 90%, 130%, and 150%, respectively (P = 0.3). In human samples, increased TUNEL-positive cells were seen at 130% energy, but not at 150% (0.0 manual vs. 0.2 [90%] vs. 2.1 [130%] vs. 0.6 [150%], P < 0.05)., Conclusions: Because of the low energy delivered by a femtosecond nanojoule platform, even incremental increases in energy appeared to have minimal effect on lens capsule morphology and strength and negligible influence on cell death. Furthermore, increasing energy appeared to enhance consistency and the ability to complete a capsulotomy in an edematous cornea.

The protein composition of the nuclear matrix is both tissue and cell type specific, and it undergoes changes with differentiation and transformation. In the present study, nuclear matrix proteins of EGFR-antisense transfected glioblastoma cell lines, U87 and U343, were compared with untransfected cell lines using two dimensional-gel electrophoresis. After EGFR-antisense transfection, the protein compositions of the nuclear matrices in both cell lines were different. Several nuclear proteins were only found in EGFR-antisense transfected cell lines. There was no difference in NuMA expression in the transfected and untransfected cell lines. These results suggest that EGFR-antisense reduced tumorigenicity on human glioblastoma cells by changing nuclear matrix protein compositions.