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Our aim was to investigate the involvement of posterior pole structures in eyes affected by keratoconus (KC). Optical coherence tomography (OCT) and OCT angiography (OCTA) were used to determine the status of lamina cribrosa (LC), peripapillary retinal nerve fiber layer (RNFL), macular and peripapillary microvasculature.Observational, cross-sectional, case-control analysis.Single-center investigation involving patients with KC and healthy control subjects. Enrolled subjects underwent anterior segment OCT combined with Placido-disk topography, macular and optic nerve head swept-source OCT and swept-source OCTA scans, and 3D wide glaucoma module for peripapillary RNFL thickness measurement. The LC curvature index was used to express the degree of LC posterior bowing. We calculated the vessel density and vessel length density at the macular superficial capillary plexus, deep capillary plexus, choriocapillaris, and nerve radial peripapillary capillary plexus.Overall, 32 eyes with KC and an equal number of age- and axial length-matched control eyes were included in the analysis. Almost all (97%) of eyes with KC were classified as early stage. KC displayed a reduction in peripapillary RNFL thickness (104.8 ± 11.9 µm vs 110.7 ± 10.5 µm; P = .039) and nerve radial peripapillary capillary plexus vessel density (46.31% ± 3% vs 43.82% ± 4%; P = .006) when compared with control subjects; these differences were more evident in the temporal sector and were associated with a higher LC curvature index (9.9% ± 2.6% vs 8.48% ± 1.7%; P = .012). Mean macular superficial capillary plexus vessel density was 3 percentage points lower in eyes with KC than in healthy controls (P.001).Early-stage KC may be characterized by a posterior bowing of the LC along with a subtle peripapillary RNFL thinning and vascular impairment. These findings support the hypothesis that KC may be a corneal manifestation of a more generalized "eye collagen disease."

SummaryMost ocular diseases are associated with pain. While pain has been generally considered a mere (deleterious) additional symptom, it is now emerging that it is a key modulator of innate/adaptive immunity. Because the cornea receives the highest nerve density of the entire body, it is an ideal site to demonstrate interactions between pain and the immune response. Indeed, most neuropeptides involved in pain generation are also potent regulators of innate and adaptive leukocyte physiology. On the other hand, most inflammatory cells can modulate the generation of ocular pain through release of specific mediators (cytokines, chemokines, growth factors, and lipid mediators). This review will discuss the reciprocal role(s) of ocular surface (and specifically: corneal) pain on the immune response of the eye. Finally, we will discuss the clinical implications of such reciprocal interactions in the context of highly prevalent corneal diseases.

A large retrospective study evaluated the safety of a post-operative therapy protocol after epithelium-off corneal collagen cross-linking (CXL). In total, 1703 eyes of the 1190 patients with progressive keratoconus were enrolled in a retrospective cohort study in a tertiary care university hospital. CXL was performed using a standardized technique (Dresden protocol: 0.1% riboflavin solution containing dextran 20% for 30 min during the soaking phase followed by 30-min ultraviolet A irradiation (3 mW/cm2)). Postoperatively, a bandage contact lens was applied, and therapy included a topical fluoroquinolone antibiotic until the epithelium healed, followed by topical fluorometholone treatment for three weeks. Post-operative complications were recorded and analyzed. No cases of infectious keratitis occurred, whereas peripheral sterile infiltrates were observed in 1.17% of cases. Trace haze was typically present but did not have an impact on visual acuity. In fifteen cases (0.88%), visually significant anterior stromal opacity developed. Mild signs of dry eye were observed in 22 eyes (1.29%). The present study demonstrates that a post-operative treatment protocol including fluoroquinolone antibiotics and a BCL in the first phase until complete epithelial healing, followed by a three-week period of topical steroid treatment is safe and not associated with the development of microbial keratitis.

Background/Objectives:Large retrospective study evaluating the effectiveness and safety of corneal collagen cross-linking (CXL)Subjects/Methods: 886 eyes with progressive keratoconus were enrolled in a retrospective cohort study in a tertiary care university hospital. CXL was performed using a standardised technique (Dresden protocol: 0.1% riboflavin solution containing dextran 20% for 30 minutes during the soaking phase followed by 30 minute ultraviolet A irradiation (3mW/cm2)). Visual outcomes, maximum keratometry (Kmax), demarcation line measurements, and complications were recorded.Results:Uncorrected visual acuity (UCVA) improved from 0.49 ± 0.38 LogMAR to 0.47 ± 0.39 LogMAR (p=0.03, n=610) three years after the procedure, while best spectacle corrected visual acuity (BSCVA) improved from 0.15 ± 0.14 LogMAR to 0.14 ± 0.15 LogMAR (p=0.007, n=610). A significant reduction of Kmax from 56.28 ± 6.10 to 54.98 ± 6.19 (p

Purpose In recent decades, the medical and surgical treatment of limbal stem cell deficiency (LSCD) has evolved significantly through the incorporation of innovative pharmacological strategies, surgical techniques, bioengineering, and cell therapy. With such a wide variety of options, there is a need to establish a global consensus on the preferred approaches for the medical and surgical treatment of LSCD. Methods An international LSCD Working Group was established by the Cornea Society in 2012 and divided into subcommittees. Four face-to-face meetings, frequent email discussions, and teleconferences were conducted since then to reach agreement on a strategic plan and methods after a comprehensive literature search. A writing group drafted the current study. Results A consensus in the medical and surgical management of LSCD was reached by the Working Group. Optimization of the ocular surface by eyelid and conjunctival reconstruction, antiinflammatory therapy, dry eye and meibomian gland dysfunction treatment, minimization of ocular surface toxicity from medications, topical medications that promote epithelialization, and use of a scleral lens is considered essential before surgical treatment of LSCD. Depending on the laterality, cause, and stage of LSCD, surgical strategies including conjunctival epitheliectomy, amniotic membrane transplantation, transplantation of limbal stem cells using different techniques and sources (allogeneic vs. autologous vs. ex vivo-cultivated), transplantation of oral mucosal epithelium, and keratoprosthesis can be performed as treatment. A stepwise flowchart for use in treatment decision-making was established. Conclusions This global consensus provides an up-to-date and comprehensive framework for the management of LSCD.

The aim of this work was to assess corneal endothelial morphology in a well-established acute graft-versus-host disease (GVHD) murine model and to quantify the expression of neurokinin-1 receptor (NK1R) in the corneal endothelium during ocular GVHD (oGVHD). Pre-conditioning was performed in BALB/c using myeloablative total body irradiation. Subsequently, allogeneic bone marrow transplantation was performed without (BM) or with mature T cells (BM + T). Corneal transparency was monitored with in vivo biomicroscopy. After sacrifice, corneal thickness and endothelial cell number were measured, and the expression of NK1R was investigated in the corneal endothelium through immunofluorescence and quantified by immunohistochemistry. Mice presenting oGVHD showed a significant reduction in endothelial cell number compared to control animals (p 0.0001). NK1R expression was significantly increased in oGVHD mice endothelium (p 0.05). Corneal transparency and thickness were unchanged in all groups. Our results suggest that oGVHD affects the corneal endothelium, inducing a reduction of the cell number, and that this is associated with increased expression of the pro-inflammatory marker NK1R.

Dry eye disease (DED) is a highly prevalent, chronic and progressive condition that affects 5–33% of the world’s adult population [...]

To evaluate the impact of vitamin D (Vit D) supplementation on systemic biomarkers of collagen degradation, inflammation, oxidative stress, and copper metabolism in adolescent patients with keratoconus (KC).This was a prospective observational pilot study. Twenty patients (age range, 16-19 years) presenting KC and Vit D insufficiency (30 ng/mL) were included. Vit D supplementation was prescribed by their general practitioner as per the standard of care. Patients were followed up for 12 months. At each visit, best spectacle-corrected visual acuity (BSCVA), maximal keratometry (Kmax), and thinnest corneal thickness (TCT) were evaluated. The primary outcome of the study was the proportion of patients with Kmax progression of less than 1 D throughout the 12-month follow-up time. Blood samples were collected at different time points to evaluate Vit D levels and systemic markers of collagen degradation, inflammation, oxidative stress, and copper metabolism by ELISA or RT-PCR.Lower Vit D levels in the plasma were correlated with higher levels of systemic biomarkers of collagen degradation. Vit D supplementation increased the cell availability of copper. Moreover, stabilization of KC progression was found in 60% of patients (72% of eyes) after 12 months with Vit D supplementation. BSCVA, Kmax, and TCT rates remained stable during the observation period.Our findings support that Vit D administration could affect ocular and systemic biomarkers in KC and illuminate a possible mechanism that can be used to develop new treatment alternatives.Although KC therapy currently relies exclusively on surgical procedures, Vit D supplementation may offer a non-invasive and inexpensive alternative with minimal associated side effects.

Purpose To describe four cases of herpetic keratitis after cultivated limbal stem-cell transplantation (CLET). Methods Four patients affected by limbal stem cell deficiency (LSCD) caused by alkali burns received CLET. During regular follow-up, epithelial defects were detected and corneal scraping was performed to identify active viral keratitis. Results Herpetic epithelial keratitis was successfully diagnosed by corneal scraping and polymerase chain reaction of HSV-1 DNA in all four patients affected by epithelial defects. The defects resolved after appropriate therapy with topical and/or systemic antiviral drugs. Conclusion Epithelial defects due to herpetic keratitis after cultivated limbal stem-cell transplantation may mimic chronic stem-cell deficiency. A low-threshold for corneal scraping can yield rapid identification of herpetic viral DNA allowing for appropriate treatment.

AimsTo test the hypothesis that Acanthamoeba keratitis (AK) outcomes differ for different topical antiamoebic therapies (AAT) and to provide the detailed patient outcome data.MethodsA retrospective cohort study of 227 patients developing AK between 25 July 1991 and 10 August 2012. Inclusion criteria required a complete record of AAT treatment for both the primary outcome of a medical cure rate at 12 months and the secondary outcome of Snellen visual acuity ≤6/24 and/or surgical intervention. Analysis used multivariable regression to control for differences in baseline disease characteristics for both primary and secondary outcomes with unadjusted analyses for other outcomes. Subjects were categorised for analysis both by the AAT used at baseline and also by mutually exclusive AAT (patients exposed to all the drugs in each group, and no others, for some period). AAT categories were PHMB monotherapy, PHMB+diamidine, PHMB+chlorhexidine+diamidine, diamidine monotherapy and other AAT.ResultsAnalysis by baseline AAT showed no notable difference between treatments for both a medical cure at 12 months in 60.79% (138/227) or for a poor outcome in 49.34% (112/227). When AATs were analysed by mutually exclusive groups, PHMB monotherapy provided the best outcomes. These findings are subject to bias requiring careful interpretation. Overall cure rates for the 214 subjects with resolved outcomes were 94.27% (214/227), median time to cure 5 months (IQR 3.25–9.00 months) and range 1–26.24 months.ConclusionPHMB 0.02% monotherapy for the initial treatment of AK is as effective as biguanide+diamidine combination therapy. Chlorhexidine monotherapy was too infrequent for comparison. The outcome data are the most detailed available.

Purpose:To test the safety and efficacy of high frequency electrotherapy (ET) on the clinical signs and symptoms of patients affected by dry eye and meibomian gland dysfunction (MGD).Methods:Twenty-five patients affected by MGD were enrolled. Quantum Molecular Resonance ET was administered by means of the Rexon-Eye device 4 times, once per week for 4 weeks. Patients were reexamined 1 month after the last treatment. The primary endpoint was reduction in corneal fluorescein staining. Additional endpoints were tear break-up time, Ocular Surface Disease Index score, meibomian gland secretion score, and the number of expressible meibomian glands. Safety endpoints were Logarithm of the Minimum Angle of Resolution (LogMar) best spectacle-corrected visual acuity and intraocular pressure.Results:Corneal fluorescein staining improved by 62.5% (P < 0.0001), tear breakup time increased by 30.9% (P < 0.0001), and the Ocular Surface Disease Index score improved by 37% (P < 0.001). The meibum quality and the number of expressible meibomian glands also increased (35.7% and 12%, P < 0.001 and P < 0.0001, respectively). Schirmer test scores increased after treatment by 16.5% (P = 0.01). No adverse events were observed.Conclusions:Quantum Molecular Resonance ET appears to be safe and significantly reduces symptoms and signs associated with MGD. It may have a relevant role in the treatment of evaporative dry eye disease.

Severe ocular surface diseases can lead to limbal stem cell deficiency (LSCD), which is accompanied by defective healing. We aimed to evaluate the role of the substance P (SP)/neurokinin-1 receptor (NK1R) pathway in corneal epithelium wound healing in a pre-clinical model of LSCD. SP ablation or NK1R blockade significantly increased epithelial wound healing (p 0.001) and corneal transparency (p 0.001), compared with wild type (WT). In addition, a reduced number of infiltrating goblet and conjunctival cells (p 0.05) and increased number of epithelial stem cells (p 0.01), which also expressed NK1R, was observed. The mammalian target of rapamycin (mTOR) pathway was significantly inhibited (p 0.05) and expression of γH2AX was significantly reduced (p 0.05) after SP ablation. These results suggest that excessive expression of SP is associated with LSCD and results in accelerated senescence and exhaustion of residual stem cells. Topical treatment with NK1R antagonist ameliorates clinical signs associated with LSCD and could be used as an adjuvant treatment in LSCD.

Purpose: In cases of corneal opacity with vascularization and peripheral thinning, traditional keratoplasty techniques have several risks and drawbacks. We report the results of a two-step surgical strategy consisting in performing a large diameter tectonic lamellar keratoplasty (TLK) to restore appropriate corneal thickness and an avascular recipient bed, followed by central optical PK within the lamellar graft at a later date. Methods: This single-institution study analyzes the results of 7 eyes of 7 patients who received PK after large diameter TLK. All patients were affected by deep post-infectious corneal opacity with persistent stromal vascularization and peripheral thinning. The main outcomes measured were graft survival, visual acuity, refraction, and endothelial cell density. Results: TLK was performed in all cases with 10/10.1-mm diameters. After a mean interval of 14months, central PK was performed with a median host-graft diameter of 7.75/8.25mm. Mean follow-up after PK was 52months. At last follow-up, 6/7 (86%) grafts were clear. Endothelial rejection occurred in 5/7 (71%) eyes, with one patient having multiple episodes and subsequent graft failure. At 2years, all patients had a visual acuity ≥ 20/40, with an average refractive astigmatism of 3.75 diopters. Conclusion: Optical PK within a previous TLK is a safe and efficient technique for treating deep corneal opacity associated with extensive vascularization and peripheral thinning. Extended follow-up period is necessary to assess endothelial cell loss and long-term efficacy of the procedure.[Figure not available: see fulltext.]

PURPOSE. The purpose of this study was to test the role of substance P (SP) and its receptor neurokinin 1 (NK1R) on ocular surface pain. METHODS. Eight-week-old C57BL6/N (wild type [WT]) and B6.Cg-Tac1tm1Bbm/J (TAC1-KO) male mice were used. 5 M NaCl was topically applied on the cornea, followed by topical fosaprepitant 2, 10, and 50 mg/mL; 4 mg/mL oxybuprocaine chloride, or 0.1% diclofenac. Th eye wiping test was used to quantify ocular surface pain. SP content was quantified in the tear fluid and trigeminal ganglia (TG), and TAC1 mRNA was assessed in the cornea. Corneas were immunostained for β3-tubulin and NK1R, or CD45, to quantify leukocyte infiltration. RESULTS. TAC1-KO mice displayed a significant reduction of ocular pain (P < 0.001). Similarly, a single dose of 10 or 50 mg/mL fosaprepitant applied topically to WT mice reduced ocular pain as compared to vehicle (P < 0.001). Fosaprepitant 2 mg/mL, instead, induced corneal analgesia only when it was administered for 10 days, 6 times/day (P < 0.05). Diclofenac or oxybuprocaine reduced corneal nociception when compared to vehicle or fosaprepitant (P < 0.05). Fosaprepitant or oxybuprocaine groups showed lower SP content in tear secretions and TG (P < 0.05), and reduction in TAC1 mRNA (P < 0.05), and leukocyte infiltration (P < 0.05) in the cornea. Colocalization of NK1R and β3-tubulin was detected in mouse corneas. CONCLUSIONS. Topical administration of the NK1R antagonist fosaprepitant effectively reduces ocular surface nociception by decreasing SP release in the tear fluid and TG, and corneal leukocyte infiltration. Fosaprepitant repurposing shows promise for the treatment of ocular pain.

The management of fungal keratitis is complex since signs and symptoms are subtle and ocular inflammation is minimal in the preliminary stages of infection. Initial misdiagnosis of the condition and consequent management of inflammation with corticosteroids is a frequent occurrence. Topical steroid use is considered to be a principal factor for development of fungal keratitis. In this review, we assess the studies that have reported outcomes of fungal keratitis in patients receiving steroids prior to diagnosis. We also assess the possible rebound effect present when steroids are abruptly discontinued and the clinical characteristics of three patients in this particular clinical scenario. Previous reports and the three clinical descriptions presented suggest that in fungal keratitis, discontinuing topical steroids can induce worsening of clinical signs. In these cases, we recommend to slowly taper steroids and continue or commence appropriate antifungal therapy.

Purpose: To quantify the severity and location of corneal neovascularization (cNV) and its impact on the visual acuity and corneal sensitivity in a cohort of the patients referred to a specialist cornea clinic and also to describe the etiology of cNV in the cohort.Methods: We retrospectively evaluated the charts of 13,493 subjects referred to the San Raffaele Cornea Unit between January 2004 and December 2018 to search for cNV diagnosis. The corneal neovascularization severity was measured in the quadrants (range: 1–4) and location was defined as superficial, deep, or both. Best spectacle corrected visual acuity (BSCVA) was measured in logMar. We used the multiple regression analysis to identify the independent predictors of logMAR, after adjusting for age, gender, keratoconus, herpes keratitis, penetrating keratoplasty, trauma, and cataract surgery.Results: Corneal neovascularization was diagnosed in 10.4% of the patients analyzed. The most prevalent etiology of cNV in our population was non-infectious corneal dystrophies/degenerations followed by herpes simplex virus infection. cNV affected OD, OS, or both eyes in 35.6, 40.2, and 24.2 of cases, respectively. Mean BSCVA (SD) was 0.59 (0.76), 0.74 (0.94), and 1.24 (1.08) in cNV one, two, and three or four of the quadrant groups. Superficial, deep, or mixed cNV occurred in 1,029, 348, and 205 eyes. Severe cNV (three or four of the quadrants) was a significant predictor of low visual acuity (p < 0.001) and reduced corneal sensitivity (p < 0.05). cNV location and its severity were associated (p < 0.05). In addition, corneal anesthesia was associated with lower BSCVA (p < 0.001).Conclusion: Severe and deep cNV are associated with the reduced visual acuity and corneal sensitivity. Our data strongly support the relevance of appropriate follow-up as cNV is a major risk factor for graft rejection.

Purpose: to assess the effect of topical administration of the Neurokin-1 receptor (NK1R) antagonist Fosaprepitant in a pre-clinical model of ocular Graft-versus-Host disease (GVHD). Methods: BALB/c mice were pre-conditioned by myeloablative total body irradiation and subjected to allogeneic bone marrow transplantation and mature T cell infusion (BM + T). BM-transplanted mice (BM) were used as controls. Ocular GVHD was specifically assessed by quantifying corneal epithelial damage, tear secretion, blepharitis and phimosis, 3 times/week for 28 days post-transplantation. A group of BM + T mice received Fosaprepitant 10 mg/mL, 6 times/day, topically, from day 7–29 after transplantation. After sacrifice, the expression of NK1R, CD45, CD3, and CXCL10 was quantified in the cornea, conjunctiva, and lacrimal gland by immunohistochemistry. Results: BM + T mice developed corneal epithelial damage (day 0–29, p < 0.001), blepharitis (day 0–29, p < 0.001), and phimosis (day 0–29, p < 0.01), and experienced decreased tear secretion (day 21, p < 0.01) compared to controls. NK1R was found upregulated in corneal epithelium (p < 0.01) and lacrimal gland (p < 0.01) of BM + T mice. Fosaprepitant administration significantly reduced corneal epithelial damage (p < 0.05), CD45+ (p < 0.05) and CD3+ (p < 0.01) immune cell infiltration in the cornea and conjunctiva (p < 0.001 and p < 0.001, respectively). In addition, Fosaprepitant reduced the expression of CXCL10 in the cornea (p < 0.05) and in the lacrimal gland (p < 0.05). Conclusions: Our results suggest that NK1R represents a novel druggable pathway for the therapy of ocular GVHD.

The complexity of inverse finite element modelling methods used in ocular biomechanics research has significantly increased in recent years in order to produce material parameters that capture microscale tissue behaviour. This study presents a more accessible method for researchers to optimise sclera material parameters for use in finite element studies where macroscale sclera displacements are required. Five human donor sclerae aged between 36 and 72 years were subjected to cycles of internal pressure up to 61 mmHg using a custom-built inflation rig. Displacements were measured using a laser beam and two cameras through a digital image correlation algorithm. Specimen-specific finite element models incorporating regional thickness variation and sclera surface topography were divided into six circumferential regions. An inverse finite element procedure was used to optimise Ogden material parameters for each region. The maximum root mean squared (RMS) error between the numerical and experimental displacements within individual specimens was 17.5 µm. The optimised material parameters indicate a gradual reduction in material stiffness (as measured by the tangent modulus) from the equator to the posterior region at low-stress levels up to 0.005 MPa. The variation in stiffness between adjacent regions became gradually less apparent and statistically insignificant at higher stresses. The study demonstrated how inflation testing combined with inverse modelling could be used to effectively characterise regional material properties capable of reproducing global sclera displacements. The material properties were found to vary between specimens, and it is expected that age could be a contributing factor behind this variation.

Limbal stem-cell deficiency (LSCD) is a disease characterized by persistent or recurrent epithelial defects, chronic inflammation, and conjunctiva migration onto the cornea (conjunctivalization) with severe visual impairment. Lamellar and/or penetrating keratoplasty cannot be successful as donor corneal epithelium is replaced by that of the recipient within months. In the presence of LSCD graft re-epithelialisation will not take place, with subsequent recurrence of conjunctivalization and graft failure. Stem-cell transplantation to treat LSCD is a step in the reconstruction of the ocular surface, while lamellar or penetrating corneal grafting will finally restore corneal transparency, leading to the recovery of visual capacity. The source of stem cells is typically classified as autologous and allogeneic. Unilateral or partial bilateral LSCDs can be treated with autologous limbal stem cell transplantation, while total bilateral deficiency requires allogeneic LSCs, or other sources of autologous cells such as oral epithelial stem cells.

Purpose: To present a series of two patients affected by Tourette syndrome (TS) and progressive keratoconus. Case series: Two young male patients with keratoconus and TS were referred to our center. In both patients eye rubbing was present and in one patient, an ocular tic was present determining blepharospasm. Progression of keratoconus occurred in both cases and corneal collagen cross-linking (CXL) was performed. All treated eyes showed topographic stability with stable refraction and conserved visual acuity, with a follow-up period ranging from 1.5 to 2.5 years. Conclusion: Patients with keratoconus and TS should be observed frequently to document topographical and refractive changes, and in case of progressing disease, CXL should be performed in order to prevent further progression.

Neurotrophic Keratopathy (NK) refers to a condition where corneal epitheliopathy leading to frank epithelial defect with or without stromal ulceration (melting) is associated with reduced or absent corneal sensations. Sensory nerves serve nociceptor and trophic functions, which can be affected independently or simultaneously. Loss of trophic function and consequent epithelial breakdown exposes the stroma making it susceptible to enzymatic degradation. Nerve pathology can range from attrition to aberrant re-generation with corresponding symptoms from anaesthesia to hyperaesthesia/allodynia. Many systemic and ocular conditions, including surgery and preserved medications can lead to NK. NK can be mild (epithelium and tear film changes), moderate (non-healing epithelial defect) or severe (stromal melting and perforation). Moderate and severe NK can profoundly affect vision and adversely impact on the quality of life. Medical management with lubricating agents from artificial tears to serum/plasma drops, anti-inflammatory agents, antibiotics and anti-proteases all provide non-specific relief, which may be temporary. Contact lenses, punctal plugs, lid closure with botulinum toxin and surgical interventions like tarsorrhaphy, conjunctival flaps and amniotic membrane provide greater success but often at the cost of obscuring sight. Corneal surgery in a dry ocular surface with reduced sensation is at high risk of failure. The recent advent of biologicals such as biopolymers mimicking heparan sulfate; coenzyme Q10 and antisense oligonucleotide that suppress connexin 43 expression, all offer promise. Recombinant nerve growth factor (cenegermin), recently approved for human use targets the nerve pathology and has the potential of addressing the underlying deficit and becoming a specific therapy for NK.

Eye-drop recombinant human nerve growth factor (ed-rhNGF) has proved to recover the retina and optic nerve damage in animal models, including the unilateral optic nerve crush (ONC), and to improve visual acuity in humans. These data, associated with evidence that ed-rhNGF stimulates the brain derived neurotrophic factor (BDNF) in retina and cortex, suggests that NGF might exert retino-fugal effects by affecting BDNF and its receptor TrkB. To address these questions, their expression and relationship with the GABAergic and glutamatergic transmission markers, GAD65 and GAD67, vesicular inhibitory amino acid transporter (VGAT), and vesicular glutamate transporters 1 and 2 (VGLUT-1 and VGLUT-2) were investigated in adult ONC rats contralateral and ipsilateral visual cortex (VCx). Ed-rhNGF recovers the ONC-induced alteration of GABAergic and glutamatergic markers in contralateral VCx, induces an upregulation of TrkB, which is positively correlated with BDNF precursor (proBDNF) decrease in both VCx sides, and strongly enhances TrkB+ cell soma and neuronal endings surrounded by GAD65 immuno-reactive afferents. These findings contribute to enlarging the knowledge on the mechanism of actions and cellular targets of exogenously administrated NGF, and suggest that ed-rhNGF might act by potentiating the activity-dependent TrkB expression in GAD+ cells in VCx following retina damage and/or ONC.

Background/aimsTo evaluate the effectiveness of corneal collagen cross-linking (CXL) in paediatric patients.MethodsFifty-two eyes of 43 paediatric patients with progressive keratoconus were enrolled in a prospective cohort study. Corneal CXL was performed using a conventional technique with instillation of 0.1% riboflavin solution containing dextran 20% for 30 min during the soaking phase and during the 30 min ultraviolet A irradiation (3 mW/cm2). Visual outcomes, topographic keratometry, maximum keratometry (K-max), refractive astigmatism, demarcation line and endothelial cell density were measured postoperatively.ResultsA significant decrease of K-max from 59.30±7.08 to 57.07±6.46 (p2 (p=0.90).ConclusionCXL is an effective treatment for avoiding keratoconus progression in paediatric patients. The procedure is safe and successful in stabilising keratoconus in eyes with more advanced forms of the disease, characterised by topographic K-max values greater than 60 D.

Purpose To review the literature on the etiopathogenesis of keratoconus (KC). Methods A literature search was conducted using PUBMED and Google Scholar for keratoconus. The authors analyzed epidemiology studies, reviews, and case reports. Results Atopy and ocular surface inflammation are a common features of KC and should lead to a reconsideration of the traditional definition of KC as a “non inflammatory” condition. Co-morbidities suggest that KC may be the ocular manifestation of a systemic disease. Finally, KC shows higher prevalence in certain ethnicities, which calls into question the status of KC as a rare disease, at least in these communities. Conclusion We believe that future studies should test whether selected, high prevalence populations exhibit specific genetic background and/or ethno-specific environmental risk factors.

Retinal ganglion cell (RGC) degeneration occurs within 2 weeks following optic nerve crush (ONC) as a consequence of reduced retro-transport of growth factors including nerve growth factor (NGF). The hypothesis that intravitreal (ivt) and eye drop (ed) administration of recombinant human NGF (rhNGF) might counteract ONC in adult rats is explored in this study. We found that both ivt- and ed-rhNGF reduced RGC loss and stimulated axonal regrowth. Chiefly, survival and regenerative effects of rhNGF were associated with a reduction of cells co-expressing Nogo-A/p75NTR at crush site borders, which contribute to glia scar formation following nerve injury, and induce further degeneration. We also found that ocular application of rhNGF reduced p75NTR and proNGF and enhanced phosphorylation of TrkA and its intracellular signals at retina level. Nogo-R and Rock2 expression was also normalized by ed-rhNGF treatment in both ONC and contralateral retina. Our findings that ocular applied NGF reaches and exerts biological actions on posterior segment of the eye give a further insight into the neurotrophin diffusion/transport through eye structures and/or their trafficking in optic nerve. In addition, the use of a highly purified NGF form in injury condition in which proNGF/p75NTR binding is favored indicates that increased availability of mature NGF restores the balance between TrkA and p75NGF, thus resulting in RGC survival and axonal growth. In conclusion, ocular applied NGF is confirmed as a good experimental paradigm to study mechanisms of neurodegeneration and regeneration, disclose biomarkers, and time windows for efficacy treatment following cell or nerve injury.

Cell therapy for treatment of corneal lesions, with associated corneal (limbal) stem cell deficiency due to ocular burns, has recently gained regulatory approval from the European Medicines Agency (EMA), and it is the first choice to restore corneal surface integrity when corneal epithelial stem cells have been lost. LSCD can be due to a number of genetic, inflammatory, or traumatic disorders and frequently results in mono- or binocular blindness. In this chapter, we will review different limbal stem cell therapies for LSCD. We will do so by considering the epidemiology of LSCD and different types of cultivated limbal stem cell treatments. Alternative treatments will also be briefly reviewed. Potential advantages, and pitfalls, of autologous versus allogenic limbal stem cell transplantation will be considered.

http://hdl.handle.net/20.500.11768/96890

Purpose of Review: Aniridia is a complex ocular disorder that causes limbal stem cell deficiency determining progressive aniridic keratopathy and the loss of corneal transparency. Allogeneic limbal stem cell transplantation followed by penetrating keratoplasty can stabilize the ocular surface but requires prolonged systemic immunosuppression. The type-1 Boston Keratoprosthesis can be a valid alternative allowing faster visual recovery without the need for systemic immunosuppression. Recent Findings: Several studies have reported good visual outcomes and good retention rates for keratoprosthesis in aniridic patients. Nonetheless, concerns still exist regarding complications that might lead to irreversible loss of function, such as glaucoma progression or intraocular infection. Other complications include retroprosthetic membrane formation, hypotony, and device extrusion. The presence of macular pathology and prevalence of glaucoma are two factors that may affect final visual acuity outcomes in aniridic patients. Summary: Implantation of the type-1 Boston Keratoprosthesis for aniridic keratopathy has shown promising results. However, further studies are needed to provide more evidence regarding long-term survival and complications.

In 1997, the human corneal epithelium was reconstructed in vitro and transplanted on patients. Later, it became a routine treatment, before regulations considered advanced therapy medicinal products and drugs on the same lines. Manufacturing, before and after good manufacturing practice setting, was established in different facilities and the clinical application in several hospitals. Advanced therapy medicinal products, including stem cells, are unique products with different challenges than other drugs: some uncertainties, in addition to benefit, cannot be avoided. This review will focus on all recent developments in the stem cell-based corneal therapy.

Purpose To describe a case of unilateral limbal stem cell deficiency (LSCD) with previously failed autologous graft, resolved by ocular surface reconstruction using cultured autologous limbal stem cells from the contralateral eye. Case Report A 35-year-old patient presented to our clinic with LSCD due to a unilateral alkali burn. The patient had received a previous limbal graft from the contralateral eye that had failed to impede corneal conjunctivalization. We decided to repeat limbal stem cell transplantation using an ex vivo cultivation procedure to reduce the risk of tissue harvesting on the healthy fellow eye. A small limbal biopsy (1.5 × 1.5 mm) near the previously excised limbus was performed. Stem cells were then isolated and cultured on fibrin and a 3T3 feeder cell layer using a standard protocol. Four months later, the cultivated cells on fibrin were grafted after pannus removal. In the subsequent months, the ocular surface stabilized and inflammation decreased. Two years later, the patient underwent large tectonic lamellar keratoplasty for severe corneal thinning involving the entire cornea, and 6 months later central penetrating keratoplasty and extracapsular cataract extraction with intraocular lens implantation and pupilloplasty was performed. Following reconstruction, the patient showed improved best-corrected vision from count fingers to 20/200 due to amblyopia, and the ocular surface was stable with a transparent corneal graft. Conclusions Ex vivo limbal stem cell transplantation is a valid technique for treating LSCD and can be utilized for treating patients who have had previous failed limbal grafts.

Purpose Aging impairs corneal nerve density and sensitivity. Substance P (SP), a neuropeptide secreted by sensory nerves, regulates nerve morphology and nociception. Here, we investigate the relationship between aging, nerve morphology, and SP expression in mouse and human corneas. Methods SP levels in mouse corneas (wild type and substance P-knockout) and human corneas and tears were quantified with an ELISA assay. Corneal total nerve length (TNL) was measured with whole-mount β3-tubulin immunofluorescence in mouse and in vivo laser corneal confocal microscopy in humans. SP and β3-tubulin stained cross-sections were used to assess the colocalization of SP and nerves in human and mouse corneas. Ocular surface nociception was assessed with a wiping test in mice. Results SP colocalizes with sub-basal neurons in mice and humans. In WT mice, SP levels decrease with age (P = 0.0045, 8 vs. 52 weeks; P = 0.004, 26 vs. 52 weeks) as well as TNL (P = 0.018, 8 vs. 26 weeks; P = 0.0001, 8 vs. 52 weeks). Knockout mice show a greater TNL reduction (8 vs. 26 weeks, P = 0.0016) than WT mice. In the oldest WT and age-matched KO mice, nociception is impaired (P = 0.007 and P < 0.0001, respectively), and KO mice sensitivity is restored by topical SP treatment. In humans, SP levels are reduced in old subject corneas and correlate, in tears, with age (P = 0.0368); TNL also decreases in older patients (P = 0.0002). Conclusions Age-associated corneal nerve loss is paralleled by reduction of SP expression in mice and humans. SP promotes the maintenance of normal nerve morphology in the long term and modulates nociception in the cornea.

PURPOSE: To evaluate the safety and efficacy of topical recombinant human nerve growth factor (rhNGF) for treating moderate-to-severe neurotrophic keratitis (NK), a rare degenerative corneal disease resulting from loss of corneal innervation. DESIGN: Phase II multicenter, randomized, double-masked, vehicle-controlled trial. PARTICIPANTS: Patients with stage II (moderate) or stage III (severe) NK in one eye. METHODS: The REPARO Phase II study assessed safety and efficacy in 156 patients randomized 1:1:1 to rhNGF 10μg/ml, 20 μg/ml, or vehicle. Treatment was administered 6 drops per day for 8 weeks. Patients then entered a 48- or 56-week follow-up period. Safety was assessed in all patients who received study drug, while efficacy was by intention to treat. MAIN OUTCOME MEASURES: Corneal healing (defined as