Maria Borrelli

Keratin films for ocular surface reconstruction

Human amniotic membrane (AM) is frequently used as a substrate for ocular surface reconstruction. Its disadvantages (e.g., reduced transparency and biomechanical strength, heterogeneity depending on donor) create the need for standardized alternatives. Keratin from hair or wool has been proposed as an appropriate material for producing films or cell cultivation scaffolds. The current study was performed to develop transparent, stable and transferable films based on human hair keratin that support cellular adhesion and proliferation. The films were engineered by a multi-step procedure including keratin extraction, neutral and alkaline dialysis, drying and a curing process. Keratin films were investigated by SDS-PAGE, SEM and X-ray analyses. Furthermore, swelling and water absorption of the films were studied, as were tensile strength and light transmission (UV/VIS). Finally, the growth behavior of corneal epithelial cells on the keratin films and AM was estimated in proliferation studies. In addition, we assessed the seeding efficiency and cell detachment behavior during trypsinization. The film-forming process resulted in transparent films composed of nanoparticulate keratin structures. The film characteristics could be varied by changing the protein composition, adding softening agents or varying the curing temperature and duration. Based on these findings, an optimized protocol was developed. The films showed improved light transmission and biomechanical strength in comparison to AM. Furthermore, cell behavior on the films was similar to that found on AM. We conclude that keratin films may represent a new, promising alternative for ocular surface reconstruction.

Long-term follow-up after submandibular gland transplantation in severe dry eyes secondary to cicatrizing conjunctivitis

PURPOSE To evaluate the long-term results of autologous submandibular gland transplantation in eyes with cicatrizing conjunctivitis and to determine biomechanical and biochemical features of the resulting salivary tear film. DESIGN Prospective, observational case series. METHODS Fifteen eyes with cicatrizing conjunctivitis with a viable autologous submandibular gland transplantation were compared with 10 eyes with cicatrizing conjunctivitis and a failed submandibular gland transplantation or no submandibular gland transplantation. Best-corrected visual acuity, frequency of tear substitute instillation, severity of dry eye discomfort, lid margin erythema, conjunctival hyperemia, corneal epithelial edema, tear film break-up time, Schirmer test results, and corneal fluorescein and conjunctival Rose Bengal staining were evaluated. In a subgroup central corneal thickness and sensitivity, corneal epithelial barrier function, conjunctival and lid margin flora, and conjunctival impression cytologic analysis results were evaluated. In 3 patients, preoperative and postoperative tear samples were analyzed for viscosity, surface tension, and presence of mucins. RESULTS Submandibular gland autotransplantation resulted in long-term improvement of subjective, objective, and some ocular surface parameters. Salivary mucins were detectable in salivary tears after submandibular gland transplantation. The viscosity of salivary tears was more similar to normal saliva and the surface tension was intermediate between the 2 original secretions. CONCLUSIONS Submandibular gland autotransplantation provides long-term relief from pain and reduces the need for frequent installation of lubricants.

Evaluation of an automated thermodynamic treatment (LipiFlow®) system for meibomian gland dysfunction: a prospective, randomized, observer-masked trial.

PURPOSE To compare the effectiveness of a single LipiFlow(®) treatment with combined lid warming and massage in patients with meibomian gland dysfunction (MGD). METHODS In this prospective, randomized, crossover, observer-masked clinical trial, subjects were randomized to receive either a single 12-min LipiFlow-LipiFlow Thermal Pulsation (LTP) system treatment or to perform combined twice-daily lid warming and massage for 3 months. All subjects were examined before, and 1 and 3 months after initiation of treatments. Investigated parameters included subjective symptoms, lipid layer thickness, meibomian gland assessment, tear break-up time, tear osmolarity, corneal and conjunctival staining, Schirmer test values, and tear meniscus height. RESULTS A total of 31 subjects completed the 3-month follow-up. At 1 and 3 months, patients in the LipiFlow treatment group had a significant reduction in Ocular Surface Disease Index (OSDI) scores compared with those in the lid-margin hygiene group. Both treatments produced a significant improvement in expressible meibomian glands compared to the baseline parameters, but no significant difference was noted between the two groups. The other investigated objective parameters did not show a significant difference. CONCLUSION Results of our study show that a single LipiFlow treatment is as least as effective as a 3-month, twice-daily lid margin hygiene regimen for MGD. However, the present study was observer-masked only, and therefore a placebo effect may have confounded any improvements in subjective symptoms and other parameters in both groups.

Comparison of central corneal thickness measured with Orbscan and Pentacam.

PURPOSE To compare central corneal thickness measurements obtained with Orbscan II using the acoustic equivalent correction factor (CF) and Pentacam in eyes screened for refractive surgery. METHODS Central corneal thickness in 91 eyes of 51 volunteers with a spherical equivalent refraction ranging from -17.00 to +5.50 diopters (D) (mean: -3.75+/-4.19 D) was measured with both Orbscan II (CF) and Pentacam prior to refractive surgery. RESULTS Central comeal thickness measurements ranged from 438 to 610 microm with Orbscan II (CF) and from 472 to 634 microm with Pentacam. The differences between Pentacam and Orbscan II (CF) measurements ranged from -43 to +52 microm (mean: -13.98+/-14.79 microm); these differences were statistically significant (P<.0001). No correlation was found between the difference in central corneal thickness and refractive error. CONCLUSIONS These results suggest measurements obtained with the Orbscan II (CF) are thinner than those obtained with the Pentacam. Further studies are needed to determine which instrument is more accurate in measuring central corneal thickness.

Review of Alternative Carrier Materials for Ocular Surface Reconstruction

Abstract Severe ocular surface disorders can result in deficiency of limbal stem cells that is potentially associated with chronic inflammation, impaired vision and even blindness. Advanced stem cells deficiency requires reconstruction of the OS with autologous or allogeneic limbal stem cells. To address such deficiency, a limbal tissue biopsy is taken and limbal cells are expanded on a carrier, which then can be used for OS reconstruction. Human amniotic membrane – currently the most common carrier for transplantation of limbal epithelial stem cells – has the downsides of carrying the risk of disease transmission, limited transparency, variable and unstable quality and low mechanical strength. This article reviews the advantages and disadvantages of the established carrier materials for limbal stem cell transplantation, as well as discussing emerging alternatives, including carriers based on collagen, fibrin, siloxane hydrogel contact lenses, poly(ε-caprolactone), gelatin–chitosan, silk fibroin, human anterior lens capsule, keratin, poly(lactide-co-glycolide), polymethacrylate, hydroxyethylmethacrylate and poly(ethylene glycol) for their potential use in the treatment of limbal stem cell deficiency.

Reliability of the IOLMaster in Measuring Corneal Power Changes After Hyperopic Photorefractive Keratectomy

PURPOSE To test the accuracy of the IOLMaster (Carl Zeiss Meditec) in detecting corneal power changes after hyperopic photorefractive keratectomy (PRK). METHODS Forty-five consecutive eyes that underwent hyperopic PRK with the SCHWIND ESIRIS excimer laser, ranging from +0.75 to +7.00 diopters (D) (mean: +3.84±1.56 D), were analyzed. Data included pre- and postoperative (1, 3, and 6 months) fogging refraction and automated keratometry (K). Statistical analysis was performed to determine the correlation between the changes in refraction at the corneal plane and the changes in keratometry. RESULTS The mean difference between the changes in refraction and the measured corneal changes was +0.27±1.19 D (range: -1.91 to +4.28 D) (P=.18) at 1 month; +0.56±0.97 D (range: -1.00 to +2.96 D) (P=.006) at 3 months; and +0.67±0.80 D (range: -0.73 to +2.31 D) (P=.00002) at 6 months. Based on these results, we suggest using the regression formula found at 6-month follow-up (y=0.8074 x + 0.092) to better calculate the effective corneal power. Comparison between the data obtained with IOLMaster measurements and equivalent K readings from the Holladay report obtained with the Pentacam (Oculus Optikgeräte GmbH) shows good agreement (R(2)=0.9). CONCLUSIONS Automated keratometry provided by the IOLMaster underestimates the effective refractive changes after hyperopic PRK, and a correcting factor is needed to calculate the corneal power in these cases.

Six-month effects of a thermodynamic treatment for MGD and implications of meibomian gland atrophy.

Purpose: The aim of this study was to evaluate the 6-month effect of a single automated thermodynamic treatment (LipiFlow) and implications of meibomian gland atrophy on treatment efficacy 6 months after application. Methods: We analyzed the data of 26 subjects with meibomian gland dysfunction before and 6 months after treatment. Investigated parameters included subjective symptoms, lipid layer thickness, meibomian gland assessment, tear osmolarity, corneal and conjunctival staining, lid margin parallel conjunctival folds, Schirmer test values, bulbar redness, tear meniscus height, meibomian gland atrophy, and noninvasive tear break-up time. Results: Subjective symptoms (mean Ocular Surface Disease Index, 42 ± 19 to 33 ± 21; P = 0.004, mean Standard Patient Evaluation of Eye Dryness 16 ± 7 to 12 ± 7; P = 0.0001), lipid layer thickness (44.0 ± 15.6 to 51.3 ± 20.4; P = 0.014), number of expressible glands (2.9 ± 1.6 to 6.4 ± 4.6; P < 0.0001), lid margin parallel conjunctival folds (2.3 ± 1.0 to 2.0 ± 0.9; P = 0.04), and bulbar redness (1.4 ± 0.5 to 1.2 ± 0.5; P = 0.0001) were all improved 6 months after treatment. Symptomatic improvement was higher in patients with less severe meibomian gland atrophy compared with patients with more dropout at treatment. There was no change of meibomian gland atrophy 6 months after treatment. Conclusions: In summary, the results showed that a single thermodynamic treatment is effective in the treatment of meibomian gland dysfunction and that the effects last for at least 6 months. We suggest performing meibography in every patient before treatment for better prediction of therapeutic effects.

Effect of oxybuprocaine eye drops on corneal volume and thickness measurements.

Purpose. To investigate the effect of oxybuprocaine eye drops on corneal volume (CV) and corneal thickness measurements. Methods. Central corneal thickness (CCT), corneal thinnest point (CTP), and CV of 78 eyes of 78 healthy volunteers were measured with Pentacam, before and 5 min after the administration of oxybuprocaine eye drops. The fellow non-anesthetized eyes were used as control. Results. Before topical anesthesia, the mean CCT was 546.76 ± 35.3 &mgr;m, after anesthesia, it was 547.76 ± 36.56 &mgr;m (p = 0.86). In the fellow eyes, the first mean CCT was 548.82 ± 35.2 &mgr;m and the second was 547.55 ± 35.9 &mgr;m (p = 0.82). The mean CTP before anesthesia was 543.99 ± 35.23 &mgr;m, after it was 544.89 ± 36.3 &mgr;m (p = 0.88). In the fellow eyes, the first mean CTP was 544.15 ± 35.35 &mgr;m and the second was 542.81 ± 36 &mgr;m (p = 0.81). Before topical anesthesia, the mean CV was 60.55 ± 3.84 mm3, after it was 60.66 ± 3.97 mm3 (p = 0.86). In the fellow eyes, the first mean CV was 60.93 ± 3.87 mm3 and the second was 60.73 ± 4 mm3 (p = 0.75). Conclusions. Oxybuprocaine eye drops do not appear to induce a significant corneal swelling and do not affect the measurements when comparing CCT measured with optical or ultrasound devices.

Keratin films for ocular surface reconstruction: evaluation of biocompatibility in an in-vivo model.

Amniotic membrane (AM) transplantation is the clinical standard for ocular surface reconstruction, however recently keratin film (KF) has been proposed as an alternative material. Aim of the current study was to evaluate corneal biocompatibility of KF in a rabbit model. Forty-six New Zealand white rabbits underwent dissection of a corneal intrastromal pocket in which an AM or KF implant was inserted and observed for 10 days and for 4 weeks. Half of animals received topical steroids, while the other half were left without. At the end of the follow-up clinical and histology examinations were performed to evaluate transparency, inflammation and degradation. After 10 days the clinical and the histology results appeared to be comparable in KF implanted eyes treated with and without steroids. After 4 weeks, comparable clinical results were observed in all KF implanted eyes, while the inflammation score was lower in non-steroid compared to steroid treated eyes along with a higher degradation rate of the keratin films. In conclusion, keratin films from human hair show a good biocompatibility and transparency in vivo. The administration of topical steroids seems to slow down implant degradation which might be important for the modulation of tissue integration and matrix regeneration.

Long-term results of corneal collagen crosslinking for progressive keratoconus

PURPOSE To evaluate long-term keratoconus stability after corneal crosslinking (CXL) with riboflavin. METHODS In this prospective study, 57 eyes of 55 patients with progressive keratoconus, consecutively treated with ultraviolet A (UVA) - riboflavin CXL, were examined with the corneal topographer Pentacam, the biometer IOLMaster and the analyzer of corneal biomechanics Ocular Response Analyzer before and during a 24 months follow-up after CXL. RESULTS Twenty-four months after CXL, there was a significant improvement in best corrected visual acuity (BCVA) (P<0.01), a significant decrease in corneal thinnest point (CTP), keratometry readings at the keratoconus apex (K max), and corneal volume (CV) (P<0.01), and a significant increase in axial eye length (AL) (P=0.01). No significant changes in anterior chamber volume (ACV) and depth (ACD), (P=0.8), corneal hysteresis (CH) (P=0.16) and corneal resistance factor (CRF) (P=0.06) were found. However, in the subgroup of patients with decreased K max readings 24 months after treatment, both CH and CRF showed a significant reduction (P<0.01). CONCLUSION In the first month after the procedure, CXL induces a reduction in corneal volume. During the 24 months follow-up the cornea tends to recover its original volume with a persistence of the CXL efficacy.