A Framework for Human Corneal Endothelial Cell Culture and Preliminary Wound Model Experiments with a New Cell Tracking Approach.

Cell injection therapy is emerging as an alternative to treat corneal endothelial dysfunction (CED) and to avoid corneal scarring due to bullous keratopathy. However, establishing a standardized culture procedure that provides appropriate cell yield while retaining functional features remains a challenge. Here, we describe a detailed framework obtained from in vitro culture of human corneal endothelial cells (HCECs) and comparative in vivo experimental models for CED treatment with a new cell tracking approach. Two digestion methods were compared regarding HCEC morphology and adhesion. The effect of Y-27632 (ROCKi) supplementation on final cell yield was also assessed. Cell adhesion efficacy with two cell delivery systems (superparamagnetic embedding and cell suspension) was evaluated in an ex vivo human cornea model and in an in vivo rabbit CED model. The injection of supplemented culture medium or balanced salt solution (BSS) was used for the positive and negative controls, respectively. HCEC isolation with collagenase resulted in better morphology and adhesion of cultured HCEC when compared to EDTA. Y-27632 supplementation resulted in a 2.6-fold increase in final cell yield compared to the control. Ex vivo and in vivo adhesion with both cell delivery systems was confirmed by cell tracker fluorescence detection. Corneal edema and opacity improved in both animal groups treated with cultured HCEC. The corneas in the control groups remained opaque. Both HCEC delivery systems seemed comparable as treatments for CED and for the prevention of corneal scarring.

Analysis of different conditioned media secreted by limbal progenitor cells in the modulation of corneal healing.

Ex vivo cultivation and transplantation of limbal epithelial cells has been reported as an alternative source for ocular surface reconstruction. However, until now, the functional improvement of these patients is limited due to the low survival rate of the transplanted cells. Consequently, the clinical benefits of this therapeutic strategy are only temporary and can assign them to paracrine effects associated with the transplanted cells. With this background in mind, we aimed to analyze the effect of different conditioned media containing growth factors secreted by limbal progenitor cells on corneal epithelial healing, both in vitro and in vivo. Limbal tissue was used to obtain different conditioned media (CM). For the in vitro assay, corneal epithelial cells were treated with CM and the epithelial migration was analyzed. Growth factors in the CM were identified with ELISA and multiplex. For the in vivo assay in rats, total limbal stem cell deficiency (LSCD) was induced with an abrasive injury to the ocular surface, and the animals were treated with different CM. Clinical and histological analyses were performed. In the in vitro assay, treatment with limbal fibroblast (LF CM) was more effective compared to the other CM, and analysis revealed high concentrations of keratinocyte growth factor (KGF) and hepatocyte growth factor (HGF). In the in vivo assay, animals treated with LF CM showed epithelial defect improvement, maintenance of thickness, and decreased opacity and neovascularization. This treatment also allowed better ocular surface tissue organization when compared to the other treatments. The in vitro and in vivo experiments showed better outcomes in the corneal wound healing for the LF CM treatment. The high concentrations of KGF and HGF, linked to epithelial cell migration and proliferation, may correlate to the best results found in this treatment.

Iatrogenic corneal diseases or conditions.

Any prescribed or self-administered therapy carries inherent risks of secondary adverse events. While the volume of treatments being administered through healthcare systems has been increasing, scientific advancements in our understanding of the mechanisms of pharmaceutical side effects and complications from procedures now allow us to reduce the risk of non-intentional damage to ocular health. This review summarizes the most common and leading causes of iatrogenic visual impairment, corneal diseases, and conditions that present in a general ophthalmologic practice, including a comprehensive analysis of their pathophysiology and recommendations for management and prophylaxis.Iatrogenic corneal diseases and conditions can arise from topical drugs, contact lens use, eye surgeries and procedures, systemic drugs, non-ophthalmological events, and cosmetic procedures. Topical and systemic drugs may disturb tear film homeostasis or result in ocular surface deposits. The use of ill-fitted contact lenses can trigger eye discomfort and poor hygiene conditions that can predispose to severe infections. Procedures to the eye may result in a variety of anatomical and functional complications that ophthalmologists should be aware of how to avoid or at least be prepared to manage if they occur. Even non-ophthalmological events such as non-invasive ventilation, radiation therapies and, immune-based conditions, or cosmetic procedures such as eyelash growth and fillers, can result in unwanted damage to the ocular surface.

IKZF1, a new susceptibility gene for cold medicine-related Stevens-Johnson syndrome/toxic epidermal necrolysis with severe mucosal involvement.

BACKGROUND: Stevens-Johnson syndrome (SJS) and its severe form, toxic epidermal necrolysis (TEN), are acute inflammatory vesiculobullous reactions of the skin and mucous membranes, including the ocular surface, oral cavity, and genitals. These reactions are very rare but are often associated with inciting drugs, infectious agents, or both. OBJECTIVE: We sought to identify susceptibility loci for cold medicine-related SJS/TEN (CM-SJS/TEN) with severe mucosal involvement (SMI). METHODS: A genome-wide association study was performed in 808 Japanese subjects (117 patients with CM-SJS/TEN with SMI and 691 healthy control subjects), and subsequent replication studies were performed in 204 other Japanese subjects (16 cases and 188 control subjects), 117 Korean subjects (27 cases and 90 control subjects), 76 Indian subjects (20 cases and 56 control subjects), and 174 Brazilian subjects (39 cases and 135 control subjects). RESULTS: In addition to the most significant susceptibility region, HLA-A, we identified IKZF1, which encodes Ikaros, as a novel susceptibility gene (meta-analysis, rs4917014 [G vs. T]; odds ratio, 0.5; P = 8.5 × 10(-11)). Furthermore, quantitative ratios of the IKZF1 alternative splicing isoforms Ik1 and Ik2 were significantly associated with rs4917014 genotypes. CONCLUSION: We identified IKZF1 as a susceptibility gene for CM-SJS/TEN with SMI not only in Japanese subjects but also in Korean and Indian subjects and showed that the Ik2/Ik1 ratio might be influenced by IKZF1 single nucleotide polymorphisms, which were significantly associated with susceptibility to CM-SJS/TEN with SMI.

Evaluation of Meibomian gland dysfunction using meibography in patients with xeroderma pigmentosum.

PURPOSE: To assess Meibomian gland dysfunction using meibography in patients with xeroderma pigmentosum and correlate with ocular surface changes. METHODS: This cross-sectional study evaluated patients with xeroderma pigmentosum. All patients underwent a comprehensive and standardized interview. The best-corrected visual acuity of each eye was determined. Detailed ophthalmic examination was conducted, including biomicroscopy examination of the ocular surface, Schirmer test type I, and meibography, and fundus examination was also performed when possible. Meibomian gland dysfunction was assessed by non-contact meibography using Oculus Keratograph® 5M (OCULUS Inc., Arlington, WA, USA). Saliva samples were collected using the Oragene DNA Self-collection kit (DNA Genotek Inc., Ottawa, Canada), and DNA was extracted as recommended by the manufacturer. Factors associated with abnormal meiboscores were assessed using generalized estimating equation models. RESULTS: A total of 42 participants were enrolled, and 27 patients underwent meibography. The meiboscore was abnormal in the upper eyelid in 8 (29.6%) patients and in the lower eyelid in 17 (62.9%). The likelihood of having abnormal meiboscores in the lower eyelid was 16.3 times greater than that in the upper eyelid. In the final multivariate model, age (p=0.001), mutation profile (p=0.006), and presence of ocular surface malignant tumor (OSMT) (p=0.014) remained significant for abnormal meiboscores. For a 1-year increase in age, the likelihood of abnormal meiboscores increased by 12%. Eyes with OSMT were 58.8 times more likely to have abnormal meiboscores than eyes without ocular surface malignant tumor. CONCLUSION: In the final model, age, xeroderma pigmentosum profile, previous cancer, and clinical alterations on the eyelid correlated with a meiboscore of ≥2. Meibomian gland dysfunction was common in patients with xeroderma pigmentosum, mainly in the lower eyelid. The severity of Meibomian gland dysfunction increases with age and is associated with severe eyelid changes.

Comparison of culture media for ex vivo cultivation of limbal epithelial progenitor cells.

PURPOSE: To compare the effectiveness of three culture media for growth, proliferation, differentiation, and viability of ex vivo cultured limbal epithelial progenitor cells. METHODS: Limbal epithelial progenitor cell cultures were established from ten human corneal rims and grew on plastic wells in three culture media: supplemental hormonal epithelial medium (SHEM), keratinocyte serum-free medium (KSFM), and Epilife. The performance of culturing limbal epithelial progenitor cells in each medium was evaluated according to the following parameters: growth area of epithelial migration; immunocytochemistry for adenosine 5'-triphosphate-binding cassette member 2 (ABCG2), p63, Ki67, cytokeratin 3 (CK3), and vimentin (VMT) and real-time reverse transcription polymerase chain reaction (RT-PCR) for CK3, ABCG2, and p63, and cell viability using Hoechst staining. RESULTS: Limbal epithelial progenitor cells cultivated in SHEM showed a tendency to faster migration, compared to KSFM and Epilife. Immunocytochemical analysis showed that proliferated cells in the SHEM had lower expression for markers related to progenitor epithelial cells (ABCG2) and putative progenitor cells (p63), and a higher percentage of positive cells for differentiated epithelium (CK3) when compared to KSFM and Epilife. In PCR analysis, ABCG2 expression was statistically higher for Epilife compared to SHEM. Expression of p63 was statistically higher for Epilife compared to SHEM and KSFM. However, CK3 expression was statistically lower for KSFM compared to SHEM. CONCLUSIONS: Based on our findings, we concluded that cells cultured in KSFM and Epilife media presented a higher percentage of limbal epithelial progenitor cells, compared to SHEM.

Measurement of the visual axis through two different methods: quantification and differences for measuring chord µ.

PURPOSE: To compare the differences between the apparent and actual chord µ. METHODS: In this prospective, comparative, non-randomized, and non-interventional study, imaging examinations using Pentacam and the HD Analyzer were performed in the same room under the same scotopic conditions. The inclusion criteria were patients aged 21-71 years, able to provide informed consent, myopia up to 4D, and anterior topographic astigmatism up to 1D. Patients using contact lenses, those with previous eye diseases or surgeries, corneal opacities, corneal tomographic changes, or suspected keratoconus were excluded. RESULTS: Altogether, 116 eyes of 58 patients were analyzed. The patients' mean age was 30.69 (±7.85) years. In the correlation analyses, Pearson's correlation coefficient of 0.647 indicates a moderate positive linear relationship between apparent and actual chord µ. The mean actual and apparent chord µ were 226.21 ± 128.53 and 278.66 ± 123.90 µm, respectively, with a mean difference of 52.45 µm (p=0.01). The analysis of mean pupillary diameter resulted in 5.76 mm using the HD Analyzer and 3.31 mm using the Pentacam. CONCLUSIONS: We found a correlation between the two measurement devices, and even though we found considerable differences, both can be used in daily practice. Given their differences, we should respect their peculiarities as well.

Severe ocular complications of SJS/TEN and associations among pre-onset, acute, and chronic factors: a report from the international ophthalmology collaborative group.

We formed an international research collaboration that included Japan, South Korea, Brazil, Thailand, Taiwan, the UK, and the US (682 patients from 13 hospitals between 2005 and 2020), to better evaluate the role of race, ethnicity, and other risk factors in the pathophysiology of Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). Ophthalmologists often see SJS/TEN patients with severe ocular complications (SOC; frequency 50% SJS/TEN patients) when the patients are referred to them in the chronic stage after the acute stage has passed. Global data were collected using a Clinical Report Form, capturing pre-onset factors, as well as acute and chronic ocular findings. Key conclusions of this retrospective observational cohort study were as follows: (1) Ingestion of cold medications [acetaminophen and non-steroidal anti-inflammatory drugs (NSAIDs)] was significantly and positively correlated with trichiasis, symblepharon, and/or conjunctivalization of the cornea in the chronic stage; (2) common cold symptoms prior to onset of SJS/TEN were significantly and positively correlated with acute conjunctivitis and ocular surface erosions in the acute stage and with trichiasis and symblepharon and/or conjunctivalization of the cornea in the chronic stage; (3) patients with SJS/TEN who presented with SOC tended to be female; (4) patients less than 30 years of age are more likely to develop SOC in the acute and chronic stages of SJS/TEN; (5) patients with acute severe conjunctivitis with ocular surface erosion and pseudomembrane formation in the acute stage are more likely to develop ocular sequelae in the chronic stage; and (6) onychopathy in the acute stage was positively correlated with ocular sequelae in the chronic stage. Our findings show that the ingestion of cold medications, common cold symptoms prior to the onset of SJS/TEN, and a young age might strongly contribute to developing the SOC of SJS/TEN.

Titanium Powder 3D-Printing Technology for a Novel Keratoprosthesis in Alkali-Burned Rabbits.

Purpose: To evaluate the surgical technique, clinical performance, and biocompatibility of a novel keratoprosthesis (KPro) named KPro of Brazil (KoBra) in an alkali-burned rabbit model. Methods: Two-piece three-dimensional-printed titanium powder and polymethyl methacrylate KPros were implanted into 14 alkali-burned corneas of 14 rabbits using an autologous full-thickness corneal graft as the KPro carrier. Rabbits were examined weekly for 12 months to evaluate retention and postoperative complications. Anterior segment optical coherence tomography (AS-OCT) and scanning electron microscopy (SEM) were performed at the end of the experiment to evaluate the relationship between the KoBra and the carrier graft. Results: All surgeries were performed without intraoperative complications, and the immediate postoperative period was uneventful. In 12 eyes (85.7%), the implanted KPros integrated into the operated eyes and maintained clear optics without extrusion or further complications over 12 months. Two eyes presented late postoperative complications that progressed to KPro extrusion: one had a presumed infectious keratitis, and the other had sterile stromal necrosis. AS-OCT demonstrated the correct relationship of the device and carrier graft in all remaining animals at the final follow-up. SEM findings indicate the integration of the porous structure of the back plate into the surrounding tissue. Conclusions: Clinical evaluations, AS-OCT, and SEM findings indicate good biointegr-ation of the implanted device into the corneal carrier graft. KoBra has the advantage of using recipients' own corneas as the prosthesis supporter, and its surgical procedure is relatively simple and safe. Translational Relevance: Titanium three-dimensional-printed technology used in an animal limbal stem-cell deficiency model holds great promise for the treatment of corneal blindness in humans.

Pterygium Surgery with Conjunctival Autograft Fixation Using Bipolar Electrocauterization.

PURPOSE: To report a case series of pterygium surgery with conjunctival autograft fixation using bipolar electrocautery. DESIGN: A noncomparative, retrospective, interventional case series of pterygium surgery with follow-up longer than 12 months to assess recurrence and other complication rates. PARTICIPANTS: Fifty-six eyes of 37 patients were treated between April 2011 and January 2018, either for primary (n = 53 cases) or recurrent (n = 3 cases) pterygia. INTERVENTION: After pterygium excision, free conjunctival grafts from the inferior bulbar conjunctiva of the same eye were harvested and fixated with the use of bipolar electrocautery. MAIN OUTCOME MEASURE: Recurrence of the pterygium and complications. RESULTS: The mean follow-up was 41 months (range 12 to 81 months). There were no intraoperative complications. Recurrence of the lesion was seen in three eyes (5.36%). There were no other postoperative complications such as graft detachment, or formation of dellen or granulomas. CONCLUSION: Conjunctival autograft fixation using bipolar electrocautery seems to be a fast, costly and safe procedure that can be applied in most cases of pterygium surgery.

Corneal densitometry in patients with keratoconus undergoing intrastromal Ferrara ring implantation.

OBJECTIVE: Evaluation of central corneal densitometry changes following Ferrara corneal ring segment implantation in patients with keratoconus, especially the correlation between corneal densitometry and keratometry. METHODS: Retrospective, non-comparative, interventional study based on the review of medical records of patients diagnosed with keratoconus who underwent Ferrara corneal ring segment implantation. Pre and post-operative corneal densitometry measurements obtained with Pentacam HR (Oculus, Wetzlar, Germany) were analyzed. The follow-up time was 3 months, and data comparison was made, using specific statistical analysis, with the data of 3 months postoperatively. RESULTS: The study sample consisted of 43 eyes of 36 patients. The mean corrected visual acuity improved from 0.82 LogMAR preoperatively (SD ± 0.33) to 0.19 LogMAR (SD ± 0.13) postoperatively. The mean spherical equivalent varied from -4.63 (SD ± 3.94) preoperatively to -2.16 (SD ± 2.63) postoperatively. Asphericity varied from -0.69 (SD ± 0.32) preoperatively to -0.27 (SD ± 0.31) postoperatively. The mean maximum K was 54.01D (SD ± 3.38) preoperatively and 51.50D (SD ± 2.90) postoperatively. The mean anterior densitometric value was 18.26 (SD ± 2.03) preoperatively and 17.66 (SD ± 1.84) postoperatively. CONCLUSION: Corneal densitometry is an interesting technology that should be studied in keratoconus patients. Our results suggest that the corneal densitometry in the cornea's anterior layer reduces after ICRS implantation and correlates with corneal keratometry. Further studies should be performed to increase the knowledge in this field.

Mass variability in drops of multidose eyedrops: a quality and reliability study.

PURPOSE: This quality and reliability study aimed to identify the mass variability of multidose eyedrops and to verify the existence of a reference pattern for the drop volume of eyedrops using standard lubricant eyedrops available on the Brazilian market. METHODS: Five brands of lubricant eyedrops were evaluated. An ideal standard 20 µL drop of eyedrops from each manufacturer was captured using an adjustable micropipette. The eyedrop bottles were randomly selected, and five measurements of the samples' masses were collected using calibrated precision scales. RESULTS: The mass of the 20 µL samples varied significantly (p<0.001) among the different manufacturers. However, among eyedrops of the same brand, mass variation was not statistically different. The global mean mass of all weighed drops was 18.24 mg, and non-uniformity was identified across all eyedrop brands. CONCLUSION: Significant variations in the drop masses of common lubricant eyedrops were identified using standard laboratory equipment. Heterogeneity in the drop volume of standard eyedrop medications suggest that potential dosage discrepancies exist, possibly altering treatment efficacy. A pre-established reference measure may lead to the production of more appropriately sized eyedrops for use in human eyes.

Prevalence and associated risk factors for dry eye disease among Brazilian undergraduate students.

PURPOSE: Dry eye is a common, complex, and multifactorial disease of the ocular surface and tear film that results in discomfort and visual disturbances. Prevalence rates vary and largely rely on studies involving older populations. This study sought to evaluate dry eye among a sample of young students in Brazil. METHODS: Cross-sectional survey included 2,140 students using 2 self-applicable questionnaires of dry eye symptoms: the Ocular Surface Disease Index (OSDI) and the Women's Health Study (WHS) questionnaire and a list of risk factors associated with dry eye. Participants with dry eye symptoms underwent a clinical evaluation. RESULTS: Participants were 23.4±5.2 years of age, 56.1% female and 43.9% male, 34.4% had an OSDI score greater than 22, and 23.5% had dry eye according to the WHS. Dry eye frequency differed consistently between the sexes: 42.6% women and 24.0% men based on the OSDI, and 27.1% women and 18.5% men based on the WHS. Univariate and multivariate analyses demonstrated that female sex, contact lens wear, the screen use for more than 6 hours per day, less than 6 hours of sleep a night, and certain medications were relevant related risk factors for dry eye. Despite symptoms, clinical evaluations demonstrated mild signs of dry eye. CONCLUSIONS: Dry eye symptoms were found to be a prevalent condition among Brazilian undergraduate students. Compared to the rates of dry eye among the general Brazilian population over 40 years of age, students present at higher dry eye symptoms rates and distinct odds for related risk factors were identified.

Clinical Aspects of Stevens-Johnson Syndrome and Toxic Epidermal Necrolysis With Severe Ocular Complications in Brazil.

Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) are acute and potentially fatal inflammatory vesiculobullous reactions that affect the skin and mucous membranes, and which are most often triggered by particular medications and infections. In Brazil, the drugs most frequently associated with TEN and SJS include cold medicine such as dipyrone and NSAIDs, followed by carbamazepine, phenobarbital, penicillin, and allopurinol. Genetic variations have been found to increase the risk of SJS/TEN in response to triggering factors such as medications. The most closely associated genes found in Brazilian cold-medicine-related SJS/TEN patients with severe ocular complications are HLA-A*66:01 in those of mixed African and European ancestry and HLA-B*44:03 and HLA-C*12:03 in those of solely European ancestry. Our classification system for grading ocular surface complication severity in SJS/TEN patients revealed the most severe complications to be limbal stem cell deficiency and dry eye. Changes to the conjunctival flora have also been observed in SJS/TEN patients. Our group identified bacterial colonization in 95% of the eyes (55.5% of which were gram-positive cocci, 25.5% of which were gram-negative bacilli, and 19% of which were gram-positive bacilli). Several new treatment options in the acute and chronic ocular management of the SJS/TEN patients have been described. This article highlights some Brazilian institutions' contributions to ocular surface care in both the acute phase (including the use of amniotic membrane transplantation) and the chronic phase (such as eyelid margin and fornix reconstruction, minor salivary gland transplantation, amniotic membrane and limbal transplantation, scleral contact lenses, anti-angiogenic eyedrops for corneal neovascularization, ex-vivo cultivated limbal epithelium transplantation, conjunctival-limbal autografting, oral mucosa transplantation, and keratoprosthesis).

Microbial Cross-contamination in Multidose Eyedrops: The Impact of Instillation Angle and Bottle Geometry.

Purpose: To evaluate the impact of instillation angle and nozzle tip geometry on cross-contamination risk of multidose ocular solution bottles. Methods: Pseudomonas aeruginosa solution was passed exclusively on the outside of the nozzle to simulate contamination on the exterior of topical agents. Three drops were administered from angles of 90° and 45° from bottles with either a round or sharp tip geometry, and the cultures were examined for growth. Two-hundred sixteen cultures from nine lubricant eyedrop brands currently existing in the Brazilian market were assessed for bacterial growth. Results: After seven days, bacterial contamination was detected in 53.7% of cultures when drops were administered at 90° and in 70.4% of cultures at 45°. Eyedrops collected from a rounded nozzle tip and an instillation angle of 90° transmitted bacteria in 69.4% of cases, whereas those administered from a sharp tip transmitted bacteria in only 22.2% of cases (P = 0.001). At an instillation angle of 45°, contamination was identified in 83.3% of bottles with a rounded tip geometry and in only eight of 18 bottles (44.4%) from those with a sharp nozzle geometry (P = 0.005). Conclusions: Adjusting the instillation angle of eyedrop solutions to 90°, as well as using a nozzle geometry that prevents flow of the solution to the side of the bottle, significantly reduced contamination rates. Translational Relevance: Standardizing drop bottles and adjusting delivery angle shows promise in reducing contamination rates and may critically impact the quality of care for patients requiring topical therapeutic agents.

Biological modulation of corneal epithelial wound healing.

The transparency and maintenance of corneal epithelial integrity are essential for its optical properties and, to preserve these characteristics, the epithelium undergoes continuous renewal. This renewal depends on the control of cell proliferation and differentiation mediated by mitogenic factors responsible for increasing mitoses and stimulating cellular migration. Cell-cell communication plays a pivotal role in epithelial healing process, and several cytokines and growth factors are involved in this process. Understanding the cross-talk and paracrine effects of these cytokines and growth factors released can help in the search for new therapeutic strategies to treat ocular surface diseases.

Evaluating the diameter of eyedropper tips using a computer vision system.

PURPOSE: This study aimed to determine the variation in diameters of outer and inner apertures of eyedropper tips using a computer vision system. Standardizing the size of eye drop nozzles is crucial to reduce the treatment cost of chronic eye diseases and to ensure a continued use of medication. An eyedropper volume of >20 µL (maximum storage of the conjunctival sac) causes medication wastage and increases treatment costs. METHODS: We measured the diameters of the outer and inner apertures of eyedropper tips and evaluated variations in diameters using a computerized visual inspection system. RESULTS: The computer visual inspection system identified anomalies in the apertures of eyedropper tips that resulted in diameter variations. CONCLUSIONS: The results of the present study show discrepancies in diameters of eyedropper tips, suggesting a variation in eyedropper size and medication wastage.

Amniotic membrane as a biological scaffold for dental pulp stem cell transplantation in ocular surface reconstruction.

PURPOSE: To evaluate the ability of human immature dental pulp stem cells, which are mesenchymal stem cells of neural crest origin, to differentiate into the corneal epithelium for purposes of corneal transplantation and tissue engineering when cultured on de-epithelized amniotic membranes. METHODS: We compared the immunophenotypes (ABCG2, K3/12, and vimentin) of cells grown on amniotic membranes or plastic surfaces under serum-free conditions or in culture media containing serum or serum replacement components. RESULTS: Immature dental pulp stem cells grown on amniotic membranes under basal conditions are able to maintain their undifferentiated state. Our data also suggest that the culture medium used in the present work can modulate the expression of immature dental pulp stem cell markers, thus inducing epithelial differentiation of these cells in vitro. CONCLUSIONS: Our results suggest that the amniotic membrane is a good choice for the growth and transplantation of mesenchymal stem cells, particularly immature dental pulp stem cells, in clinical ocular surface reconstruction.

Descemet Membrane Endothelial Keratoplasty Outcomes During the Initial Learning Curve of Cornea Fellows.

PURPOSE: To describe the outcomes of Descemet membrane endothelial keratoplasty (DMEK) performed by cornea fellows during their initial learning curve. METHODS: This prospective study included the first consecutive series of 53 DMEK surgeries performed by 14 corneal fellows at Sorocaba's Ophthalmological Hospital. Best spectacle-corrected visual acuity, endothelial cell density (ECD), and complications were evaluated. RESULTS: Of the eyes without ocular comorbidities and with complete visual acuity measurements, 97% of 33 eyes achieved a best spectacle-corrected visual acuity of ≥20/40 in 6 months. The mean (SD) preoperative donor ECD was 2453 (361) cells/mm, and at 6 months postoperatively, the in vivo mean ECD (SD) was found to be 1300 (587) cells/mm, an average loss of 47%. Preparation of the DMEK graft by the cornea fellows was successful in all cases. There were 9 eyes (17%) that experienced partial graft detachment requiring rebubbling. A total of 3 eyes (5.7%) failed to clear the cornea, all of which were managed with successful secondary endothelial keratoplasty. None of the eyes experienced pupillary block. CONCLUSIONS: Establishing DMEK in a fellowship program was successful with the supervision of an experienced DMEK surgeon. Even without the facility of a prestripped DMEK graft by an eye bank and with no backup tissue, no surgery was canceled because of graft preparation failure. Good visual outcomes were achieved with satisfactory ECD at 6-month follow-up, with an acceptable complication rate. Even the 3 eyes that failed to clear the cornea evolved well after secondary endothelial keratoplasty by the cornea fellows.

Measurement of the visual axis through two different methods: quantification and differences for measuring chord μ / Medição do eixo visual através de dois métodos diferentes: quantificação e diferenças para medição da corda μ

ABSTRACT Purpose: To compare the differences between the apparent and actual chord μ. Methods: In this prospective, comparative, non-randomized, and non-interventional study, imaging examinations using Pentacam and the HD Analyzer were performed in the same room under the same scotopic conditions. The inclusion criteria were patients aged 21-71 years, able to provide informed consent, myopia up to 4D, and anterior topographic astigmatism up to 1D. Patients using contact lenses, those with previous eye diseases or surgeries, corneal opacities, corneal tomographic changes, or suspected keratoconus were excluded. Results: Altogether, 116 eyes of 58 patients were analyzed. The patients' mean age was 30.69 (±7.85) years. In the correlation analyses, Pearson's correlation coefficient of 0.647 indicates a moderate positive linear relationship between apparent and actual chord μ. The mean actual and apparent chord μ were 226.21 ± 128.53 and 278.66 ± 123.90 μm, respectively, with a mean difference of 52.45 μm (p=0.01). The analysis of mean pupillary diameter resulted in 5.76 mm using the HD Analyzer and 3.31 mm using the Pentacam. Conclusions: We found a correlation between the two measurement devices, and even though we found considerable differences, both can be used in daily practice. Given their differences, we should respect their peculiarities as well.

RESUMO Objetivo: Comparar as diferenças entre a chord aparente μ e o chord real μ. Métodos: Estudo prospectivo, comparativo, não randomizado e não intervencionista. Os exames de imagem (Pentacam e HD Analyzer) foram realizados na mesma sala e nas mesmas condições escotópicas. Os critérios de inclusão foram idade de 21 a 71 anos; compreensão do termo de consentimento; miopia até 4D e astigmatismo topográfico anterior até 1D. Os critérios de exclusão foram usuários de lentes de contato; pacientes com doenças oculares prévias ou cirurgias; opacidades da córnea; a presença de alterações tomográficas da córnea ou suspeita de ceratocone. Resultados: Em nosso estudo foram analisados 116 olhos de 58 pacientes. A média de idade foi de 30,69 anos (± 7,85). Análises de correlação foram desenvolvidas e o coeficiente de correlação de Pearson (0,647) indica uma relação linear positiva moderada entre as variáveis. A média do chord μ real foi 226,21 ± 128,53 μm e a média do chord μ média foi 278,66 ± 123,90 μm, com diferença média de 52,45 μm (p=0,01). A análise do diâmetro pupilar médio apresentou: 5,76mm no HD Analyzer e 3,31mm no Pentacam. Conclusões: Entendemos a existência de uma diferença significativa entre os métodos e assim a medida de ambos os dispositivos com base em princípios diferentes devemos respeitar suas peculiaridades. Como encontramos correlação entre as duas medidas, acreditamos que ambas podem ser utilizadas na prática diária.