Influence of Immobilization Strategies on the Antibacterial Properties of Antimicrobial Peptide-Chitosan Coatings.

It is key to fight bacterial adhesion to prevent biofilm establishment on biomaterials. Surface immobilization of antimicrobial peptides (AMP) is a promising strategy to avoid bacterial colonization. This work aimed to investigate whether the direct surface immobilization of Dhvar5, an AMP with head-to-tail amphipathicity, would improve the antimicrobial activity of chitosan ultrathin coatings. The peptide was grafted by copper-catalyzed azide-alkyne cycloaddition (CuAAC) chemistry by either its C- or N- terminus to assess the influence of peptide orientation on surface properties and antimicrobial activity. These features were compared with those of coatings fabricated using previously described Dhvar5-chitosan conjugates (immobilized in bulk). The peptide was chemoselectively immobilized onto the coating by both termini. Moreover, the covalent immobilization of Dhvar5 by either terminus enhanced the antimicrobial effect of the chitosan coating by decreasing colonization by both Gram-positive (Staphylococcus aureus, Staphylococcus epidermidis) and Gram-negative (Escherichia coli, Pseudomonas aeruginosa) bacteria. Relevantly, the antimicrobial performance of the surface on Gram-positive bacteria depended on how Dhvar5-chitosan coatings were produced. An antiadhesive effect was observed when the peptide was grafted onto prefabricated chitosan coatings (film), and a bactericidal effect was exhibited when coatings were prepared from Dhvar5-chitosan conjugates (bulk). This antiadhesive effect was not due to changes in surface wettability or protein adsorption but rather depended on variations in peptide concentration, exposure, and surface roughness. Results reported in this study show that the antibacterial potency and effect of immobilized AMP vary greatly with the immobilization procedure. Overall, independently of the fabrication protocol and mechanism of action, Dhvar5-chitosan coatings are a promising strategy for the development of antimicrobial medical devices, either as an antiadhesive or contact-killing surface.

How Charge, Size and Protein Corona Modulate the Specific Activity of Nanostructured Lipid Carriers (NLC) against Helicobacter pylori.

The major risk factor associated with the development of gastric cancer is chronic infection with Helicobacter pylori. The available treatments, based on a cocktail of antibiotics, fail in up to 40% of patients and disrupt their gut microbiota. The potential of blank nanostructured lipid carriers (NLC) for H. pylori eradication was previously demonstrated by us. However, the effect of NLC charge, size and protein corona on H. pylori-specific bactericidal activity herein studied was unknown at that time. All developed NLC formulations proved bactericidal against H. pylori. Although cationic NLC had 10-fold higher bactericidal activity than anionic NLC, they lacked specificity, since Lactobacillus acidophilus was also affected. Anionic NLC achieved complete clearance in both H. pylori morphologies (rod- and coccoid-shape) by inducing alterations in bacteria membranes and the cytoplasm, as visualized by transmission electron microscopy (TEM). The presence of an NLC protein corona, composed of 93% albumin, was confirmed by mass spectrometry. This protein corona delayed the bactericidal activity of anionic NLC against H. pylori and hindered NLC activity against Escherichia coli. Overall, these results sustain the use of NLC as a promising antibiotic-free strategy targeting H. pylori.

Graphene-based materials: the key for the successful application of pHEMA as a blood-contacting device.

Thrombosis and infection are the leading causes of blood-contacting device (BCD) failure, mainly due to the poor performance of existing biomaterials. Poly(2-hydroxyethyl methacrylate) (pHEMA) has excellent hemocompatibility but the weak mechanical properties impair its use as a bulk material for BCD. As such, pHEMA has been explored as a coating, despite the instability and difficulty of attachment to the underlying polymer compromise its success. This work describes the hydrogel composites made of pHEMA and graphene-based materials (GBM) that meet the biological and mechanical requirements for a stand-alone BCD. Five GBM differing in thickness, oxidation degree, and lateral size were incorporated in pHEMA, revealing that only oxidized-GBM can reinforce pHEMA. pHEMA/oxidized-GBM composites are cytocompatible and prevent the adhesion of endothelial cells, blood platelets, and bacteria (S. aureus), thus maintaining pHEMA's anti-adhesive properties. As a proof of concept, the thrombogenicity of the tubular prototypes of the best formulation (pHEMA/Graphene oxide (GO)) was evaluated in vivo, using a porcine arteriovenous-shunt model. pHEMA/GO conduits withstand the blood pressure and exhibit negligible adhesion of blood components, revealing better hemocompatibility than ePTFE, a commercial material for vascular access. Our findings reveal pHEMA/GO, a synthetic and off-the-shelf hydrogel, as a preeminent material for the design of blood-contacting devices that prevent thrombosis and bacterial adhesion.

Long-term results of topical mitomycin C 0.02% for primary and recurrent conjunctival-corneal intraepithelial neoplasia.

PURPOSE: To evaluate the efficacy, recurrence rate, and long-term complications of topical mitomycin C (MMC) 0.02% for conjunctival-corneal intraepithelial neoplasia (CCIN). METHODS: A prospective, nonrandomized, noncontrolled study was conducted of patients with primary or recurrent CCIN treated with topical MMC 0.02%, four times per day, for 28 consecutive days. The main outcome measures were complete resolution of the neoplasia by slit-lamp examination and cytology 1 month after treatment, tumor recurrence, and long-term complications. RESULTS: Between June 1999 and September 2005, 23 patients were included. Eighteen had primary CCIN (group 1) and 5 had recurrent CCIN (group 2). The mean follow-up was 46 months in group 1 and 54 months in group 2. All patients were treated with MMC 0.02% for 28 consecutive days. Complete resolution of the lesion was achieved in all patients after 1 month of treatment. Recurrence occurred in 1 patient (4.3%) after 24 months of treatment. Four patients developed corneal erosion (17.4%), 2 of them with primary CCIN and 2 with recurrent CCIN. Corneal erosion occurred 4 to 24 months after treatment and was treated successfully. The probability for corneal erosions by the log-rank test was equal for both groups (p = 0.1705). CONCLUSIONS: The use of topical MMC 0.02% for 28 consecutive days to treat primary or recurrent CCIN was effective and showed a low recurrence rate. Corneal erosion occurred in 17.4% of cases and can occur as late as 24 months after treatment.

Subretinal bevacizumab detection after intravitreous injection in rabbits.

PURPOSE: To evaluate subretinal detection of bevacizumab 2 hours after intravitreous injection of 1.25 mg in rabbit eyes. METHODS: Anterior chamber paracentesis using a 30-gauge needle was performed in nine female Dutch-belted rabbits by removal of 0.05 mL of aqueous humor. Transscleral retinal detachment was performed with a modified 25-gauge infusion cannula connected to a bottle of physiologic saline solution (PSS). The animals were divided into experimental group 1, with intravitreous injection of 0.05 mL of (1.25 mg) with a 30-gauge needle (n = 6) and the control group 2, with intravitreous injection of 0.05 mL of PSS with a 30-gauge needle (n = 3). Two hours after the intravitreous bevacizumab or PSS injection, subretinal fluid was aspirated and immunoassayed to measure the level of bevacizumab. The rabbits were killed by intravenous pentobarbital injection. The eyes were enucleated and fixed in 10% formaldehyde. The pars plana site at which the transscleral cannula was introduced was analyzed by light microscopy, to exclude iatrogenic retinal tears. Eyes with accidental retinal tears were excluded. RESULTS: Subretinal bevacizumab molecules were detected in the six eyes that received an intravitreous bevacizumab injection. No subretinal bevacizumab was detected in the control eyes. Light microscopy showed no evidence of retinal tears or holes in any rabbits used for the bevacizumab detection and control group. CONCLUSIONS: Bevacizumab molecules were detected in the subretinal space after intravitreous injection of 1.25 mg of bevacizumab, possibly as the result of diffusion through the retina in a rabbit model.

Fundus autofluorescence of choroidal nevus and melanoma.

BACKGROUND: To describe autofluorescence patterns of choroidal melanocytic lesions using the Heidelberg Retinal Angiograph 2 system (HRA2). METHODS: 20 patients with choroidal melanocytic lesions in the ocular fundus underwent ophthalmologic examination, fundus photography, autofluorescence and optical coherence tomography (OCT). Pathologic examination was performed on one enucleated eye with a large choroidal melanoma. RESULTS: 15 patients had choroidal nevi and 5 had malignant choroidal melanoma (1 small, 1 medium and 3 large tumours). Choroidal nevi did not show any characteristic autofluorescence pattern, although secondary retinal pigment epithelium (RPE) changes, such as drusen and pigment epithelium detachment, appeared faintly hyperautofluorescent in 2 patients. Only the small malignant choroidal melanomas had prominent orange pigmentation, although all melanomas had an intense confluent hyperautofluorescent signal over the lesions. Pathology of one large malignant melanoma revealed lipofuscin underlying RPE. CONCLUSION: Most nevi did not have characteristic hyperautofluorescent features, but choroidal melanomas seemed to have a pattern of confluent hyperautofluorescence. Therefore, autofluorescence may be a useful non-invasive tool to assess lipofuscin in pigmented choroidal lesions, which may contribute to the diagnosis of malignancy. This hypothesis, however, remains to be confirmed in large prospective studies.

[Diagnosis of Acanthamoeba corneal infection by impression cytology: case report]. / Citologia de impressão no diagnóstico de infecção corneana por Acanthamoeba: relato de caso.

To describe three cases of corneal infection due to Acanthamoeba sp in which was possible to detect Acanthamoeba sp cysts by the corneal impression cytology technique. Three patients referred to the External Eye Disease Laboratory in 2004 with superficial corneal alterations were submitted to corneal specimen collection by impression cytology filter paper to investigate the presence of Acanthamoeba sp cysts. Two impression cytology samples were obtained from each patient and were stained by PAS, hematoxylin and Papanicolaou. Routine microbiological investigation and culture were also performed using corneal scraping. Positive culture and impression cytology for Acanthamoeba sp was observed in all patients while smears with Giemsa stain were positive in two. Impression cytology Acanthamoeba sp cysts were observed among sheets of corneal epithelial cells and as isolated cells. Cysts were also found in the superficial epithelium in one of these patients after treatment while corneal scraping did not reveal any cyst. Histopathology revealed cysts in the epithelium and stroma in a transplanted cornea in one of these patients. The first description of impression cytology as a diagnostic method for Acanthamoeba keratitis occurred recently. In this study corneal impression cytology detected Acanthamoeba sp cysts successfully in these patients with only superficial involvement. Impression cytology as a non invasive technique can be used to facilitate early recognition of Acanthamoeba infection playing a useful role in the follow-up of the disease.

[Morphological assessment of different amniotic membrane epithelial denuding techniques]. / Avaliação morfológica de diferentes técnicas de desepitelização da membrana amniótica humana.

PURPOSE: To evaluate the morphological features of the amniotic membrane denuded by different techniques. METHODS: Human amniotic membrane was collected at the time of delivery, fixed in increasing concentrations of glycerol (0-50% in DMEM) and preserved at -80 degrees C until the time of use. The study consisted of 4 groups: intact epithelium (control) and denuded by trypsin (2 mg/mL at 1:250), dispase (1.2 U/mL in Mg2+ and Ca2+ free Hank's balanced salt solution) or ethylenediaminetetraacetic acid (EDTA), 0.02%. Specimens were submitted to electron (scanning and transmission) microscopy analysis. RESULTS: Scanning electron microscopy disclosed intact epithelium in the control group and its absence in the amniotic membranes denuded by trypsin and dispase. In those denuded by ethylenediaminetetraacetic acid there were areas with and without epithelium. When assessed by transmission electron microscopy, the epithelium was intact and firmly adhered to the basement membrane by hemidesmossomes in controls and in parts of ethylenediaminetetraacetic acid group. There were only collagen fibers in the dispase- and trypsin-treated groups. CONCLUSIONS: Trypsin and dispase treatment of the amniotic membrane may cause complete denuding of the epithelium and basement membrane whereas ethylenediaminetetraacetic acid may leave some intact epithelium-areas and partially destroy the basement membrane in others.

Aptamer-based fiber sensor for thrombin detection.

The detection of thrombin based on aptamer binding is studied using two different optical fiber-based configurations: long period gratings coated with a thin layer of titanium dioxide and surface plasmon resonance devices in optical fibers coated with a multilayer of gold and titanium dioxide. These structures are functionalized and the performance to detect thrombin in the range 10 to 100 nM is compared in transmission mode. The sensitivity to the surrounding refractive index (RI) of the plasmonic device is higher than 3100 nm RIU−1 in the RI range 1.335 to 1.355, a factor of 20 greater than the sensitivity of the coated grating. The detection of 10 nM of thrombin was accomplished with a wavelength shift of 3.5 nm and a resolution of 0.54 nM.

Ocular surface findings in impression cytology after interferon a2b or mitomycin C in rabbits / Achados em citologia de impressão da superfície ocular após uso de interferon alfa-2b ou mitomicina C: estudo em coelho

ABSTRACT Objective: To describe ocular surface findings in impression cytology obtained from healthy rabbit conjunctiva treated with interferon alpha-2b eyedrop, and compare them to findings after use of mitomycin C 0.02%. Methods: An experimental study using a rabbit model was performed between September 2013 and October 2014 at the Faculdade de Medicina de Marília, Universidade Federal de São Paulo, Clínica de Olhos Moacir Cunha. Thirty New Zealand white rabbits were divided into 6 groups and received interferon alpha-2b or mitomycin C 0.02%. Impression cytology (IC) was performed prior to topical applications and at15, 30 and 60 days of use. The following variables were analyzed in impression cytology: goblet cells, cellularity, cell-to-cell adhesion, nucleus/cytoplasm ratio, nuclear chromatin, inflammatory cells keratinization, and cytomegaly. Results: The major findings in impression cytology after us of interferon alpha-2b included loss of goblet cells (50.8%), reduced cell-to-cell adhesion (26.2%), abnormal nucleus/cytoplasm ratio (20%) and reduced cellularity (15.4%). After use of mitomycin C 0.02%, the most common changes included loss of goblet cells (46.2%), abnormal nucleus/cytoplasm ratio (25.6%), less cell-to-cell adhesion (23.1%), and reduced cellularity (20.5%). There were no significant differences in any variable when comparing impression cytology after interferon alpha-2b and after mitomycin C 0.02%. Goblet cell loss was more pronounced at days 30 and 60, as compared to impression cytology at day 15 for both drugs. Conclusion: The loss of goblet cells, reduced cell-to-cell adhesion and cellularity, along with abnormal nucleus/cytoplasm ratio were the most common findings in impression cytology after use of interferon alpha-2b. These findings are similar to those described for use of mitomycin C 0.02%. ..

RESUMO Objetivo: Descrever os achados em citologia de impressão de conjuntiva sadia de coelho submetida ao uso de colírio de interferon alfa-2b e compará-los ao que foi encontrado após uso da mitomicina C 0,02%. Métodos: Estudo experimental realizado em modelo animal no período entre setembro de 2013 e outubro de 2014 nas dependências da Faculdade de Medicina de Marília, da Universidade Federal de São Paulo e da Clínica de Olhos Moacir Cunha. Trinta coelhos albinos da raça Nova Zelândia foram divididos em seis grupos e receberam interferon alfa-2b ou mitomicina C. A citologia de impressão foi realizada antes do início dos colírios e após 15, 30, 60 dias de seu uso. As seguintes variáveis foram analisadas na citologia de impressão: células caliciformes, celularidade, adesão intercelular, razão núcleo/citoplasma, cromatina, células inflamatórias, queratinização e citomegalia. Resultados: Os principais achados na citologia de impressão após o uso do interferon alfa-2b foram a redução de células caliciformes (50,8%), a diminuição da adesão intercelular (26,2%), a alteração da razão N/C (20%) e a redução da celularidade (15,4%). Após o uso da mitomicina C 0,02%, foram mais frequentes a redução das células caliciformes (46,2%), a alteração da razão N/C (25,6%), a adesão intercelular (23,1%) e a redução da celularidade (20,5%). Não houve diferença estatisticamente significante para nenhuma das variáveis estudas quando se compararam as citologias de impressão após interferon alfa-2b com as citologias de impressão após mitomicina C 0,02%. Independentemente da substância utilizada, as citologias colhidas 30 e 60 dias após início das drogas apresentaram maior redução de células caliciformes quando comparadas com as citologias de impressão colhidas após 15 dias. Conclusão: A redução das células caliciformes, a diminuição da adesão intercelular, a alteração da razão N/C e a diminuição da celularidade foram as alterações mais frequentes na citologia de impressão colhida após o uso de interferon alfa-2b. Os achados em citologias de impressão após o uso de interferon alfa-2b são semelhantes àqueles encontrados após o uso da mitomicina C 0,02%.

Histopathological features and P-glycoprotein expression in retinoblastoma.

PURPOSE: To investigate the expression of P-glycoprotein (P-gp) in retinoblastoma specimens enucleated as a primary treatment or after conservative treatment and to correlate this expression with histopathological tumor features. METHODS: Retrospective analysis was performed on retinoblastoma specimens obtained consecutively between 1993 and 2003 by enucleation either as primary treatment (group I) or after the failure of conservative treatment (group II). Sections from the formalin-fixed, paraffin-embedded specimens were stained with hematoxylin and eosin. Group I tumor differentiation was classified according to the percentage of Flexner-Wintersteiner rosettes. Group II tumors, categorized as viable-appearing, regressed with a well-differentiated component (WDC), and regressed. Other features, such as choroidal and optic nerve invasion, were evaluated. P-gp expression was graded semiquantitatively as negative, low, or high. Variables were statistically analyzed by chi(2) and Student's t-tests. RESULTS: Histopathological assessment of group I revealed 65% moderately differentiated tumors, 30% well differentiated, and 5% poorly differentiated. Fifteen percent had optic nerve tumor invasion only, 20% choroidal invasion only, and 55% both choroidal and optic nerve invasion. Group II had 62.5% well-differentiated, regressed tumors; 25% had regressed tumors replaced by glial scarring; and 12.5% had tumors containing viable, poorly differentiated cells. Approximately 18% had choroidal tumor invasion only, 6.3% optic nerve tumor invasion only, and 6.3% simultaneous optic nerve and choroidal invasion. P-gp expression was observed in 60% of group I and 66.6% of group II. All P-gp-positive cases in group II had a high expression. P-gp was also expressed by 81.2% of well-differentiated tumors. CONCLUSIONS: P-gp was expressed more frequently by well-differentiated retinoblastomas, especially those treated by chemotherapy before enucleation. This finding could be related to treatment failure.

Protein adsorption on 18-alkyl chains immobilized on hydroxyl-terminated self-assembled monolayers.

Surfaces of devices that contact blood accumulate adsorbed and denatured proteins perhaps triggering activation of the coagulation system. A renewable layer of albumin would biologically "passivate" the surface and prevent thrombus formation. Based on the approach of selectively binding albumin to fatty acids, different percentages of a compound with 18 carbons (C18) were immobilized on OH-terminated self-assembled monolayers (SAMs). Fourier transform infrared reflection absorption spectroscopy (IRAS), ellipsometry, contact angle (and surface free energy) and X-ray photoelectron spectroscopy (XPS) measurements were used to characterize these surfaces and proved that there is an efficient immobilization of C18. There is an increase of the thickness and hydrophobicity of SAMs with an increasing percentage of C18. Adsorption of human serum albumin (HSA) was evaluated using radiolabelled (125)I-HSA and IRAS. This study showed a gradual increase of HSA adsorption with the increase of surface hydrophobicity. Regarding competitive binding and exchangeability of albumin towards fibrinogen, it was proved, by radiolabelling, that SAMs prepared from solutions with 2.5% C18 presented considerable adsorption in a selective and reversible way.

The genotype of Toxoplasma gondii strains causing ocular toxoplasmosis in humans in Brazil.

PURPOSE: To determine Toxoplasma gondii genotype in scars of retinochoroiditis in humans. DESIGN: Immunopathology analysis. METHODS: DNA was isolated from retinochoroidal specimens taken from the borders of the scars from eye bank eyes as well as enucleated eyes from uveitis patients with retinochoroidal toxoplasmic scars. The SAG2 region was amplified by polymerase chain reaction and analyzed by restriction fragment length polymorphism. RESULTS: Of the 92 eyes studied, 9 had toxoplasmic scars. All of them, as well as the 2 enucleated eyes, disclosed the presence of type I parasites by the gel electrophoresis analysis of the restriction fragment length polymorphism from SAG2. CONCLUSION: These results suggest that type I strain seems to be responsible for the ocular infections in the population studied.

Eyelid swelling as the only manifestation of ocular sarcoidosis.

PURPOSE: To describe a case of eyelid sarcoidosis without systemic manifestations with a three-year follow-up. METHODS: A 73-year-old woman presented complaining of a one-year history of swelling and hyperemia in the right upper eyelid. To confirm the diagnosis, we performed an incisional biopsy of the eyelid. RESULTS: Histopathological examination showed a non-caseating granulomatous process suggestive of sarcoidosis. After three years, the patient was asymptomatic and physical and laboratory examination showed no signs of systemic sarcoidosis. CONCLUSION: The histopathological exam was fundamental for the diagnosis.

Extranodal B-cell lymphoma of the uvea: a case report.

CASE REPORT: Ocular involvement by non-Hodgkin's lymphoma is a rare condition that can result from a primary intraocular lymphoma of the retina or an intraocular manifestation of systemic lymphoma. Uveal involvement is seldom the initial manifestation of extranodal lymphoma. We describe an 80-year-old patient with a blind and painful left eye and a history of recurrent uveitis. After ultrasound evaluation, the eye was enucleated and histopathologic examination revealed a malignant B-cell lymphoma of the uveal tract. The patient has been followed for 8 years after surgery, but she has had no further systemic manifestations of lymphoma and has not required subsequent treatment. COMMENTS: Primary extranodal lymphoma can be easily mistaken for recurrent uveitis or primary intraocular lymphoma of the retina and central nervous system; it is a differential diagnosis to be considered in cases of recurrent uveitis-like symptoms evolving to blind painful eye.

[Teleophthalmology as an auxiliary approach for the diagnosis of infectious and inflammatory ocular diseases: evaluation of an asynchronous method of consultation]. / Teleoftalmologia como auxílio diagnóstico nas doenças infecciosas e inflamatórias oculares.

PURPOSE: The purpose of this study was to evaluate the efficacy of teleophthalmology as an auxiliary approach for diagnosis of infectious and inflammatory ocular diseases and to study the main problems associated with the conclusion reached by consultation. METHODS: Two hundred and thirty three 35 mm slides (retinography or biomicroscopy photos) were digitalized and stored in a Compact Disc to be later analyzed in a computer monitor by two consultants, specialists in uveitis. The consultants filled out a form and a questionnaire, indicating the diagnosis and probable treatment for the analyzed cases also the main difficulties found to reach a conclusion of the consultation. The Kappa statistic, an assessment of the change-corrected agreement (reproducibility) among consultants, was calculated. Kappa values of > 0.7 indicated excellent agreement; values between 0.4 and 0.7 indicated fair to good agreement and values of < 0.4 indicated poor agreement. RESULTS: Agreement between the consultants and the final diagnosis was of 73.5%. Treatment suggestion or management was not possible in 8% of the cases for consultant A and in 10.4% for consultant B. The higher agreement rate among consultants, relating to the reasons that rendered diagnosis difficult, was found in relation to "lack of clinical data" and "low quality of images". CONCLUSION: Teleophthalmology, through the asynchronous method, was an efficient means for diagnosis of patients with infectious and inflammatory ocular diseases. The main problems affecting the teleophthalmology consultation were lack of detailed clinical data and low quality of the images.

[Histopathologic findings in retinoblastoma]. / Achados histopatológicos em retinoblastoma.

OBJECTIVE: To study histopathological findings of enucleated eyes with retinoblastoma. METHODS: Twenty-eight cases of retinoblastoma treated by enucleation at the Federal University of São Paulo from December 2000 to October 2002 were histopathologically reviewed. Clinical data included age, gender, race, unilateral or bilateral involvement and previous treatment. The histopathological review evaluated the presence of iris and/or angle neovascularization, tumor differentiation and optic nerve and choroidal invasion according to Khelfaoui's classification. RESULTS: Of 27 patients, 13 (48.5%) were boys and 14 (59.3%) were girls, 16 were white, 6 were black and 5 were asiatic, age ranging from 2 to 96 months (mean, 22.7 months). 13 cases were bilateral and 14 cases were unilateral. All tumors were histologically characterized by a proliferation of small cells with high nuclear-to-cytoplasmic ratios and 20 (71.4%) of them were well differentiated. Choroidal involvement was observed in 18 (64.2%) cases (degree II, III) and optic nerve invasion in 8 (28.5%) cases (degree III, IV, V). CONCLUSION: Neovascularization, necrosis and calcification were the most commonly observed feature. The invasion into the optic nerve and choroid, which are the two most important predictors of patient outcome were found in 28.5% and 64.2% of the cases, respectively.

Use of impression cytology for the detection of unsuspected ocular surface squamous neoplasia cells in pterygia.

Purpose: To evaluate the agreement between the methodologies of impression cytology (IC) and histopathology regarding epithelial lesions clinically diagnosed as pterygium and also regarding the detection of unsuspected and associated ocular surface squamous neoplasia (OSSN). Methods: Thirty-two Brazilian patients were included and IC was performed on all pterygia before excision. Histopathogical examination was considered the gold standard and was performed by two experienced ocular pathologists in which consensus existed regarding pterygia diagnosis. IC accuracy was assessed by sensitivity and specificity with a 95% confidence interval. Results: From the 32 primary lesions studied, histopathological examination confirmed the diagnosis of pterygium without atypical cells in 19 cases (60%) and showed unsuspected and associated OSSN cells in 13 cases (40%). IC demonstrated one false-negative and one false-positive result for atypia. Statistical analysis showed an estimated sensitivity of 92%, specificity of 94%, positive predictive value of 92%, and negative predictive value of 94%. Conclusion: IC demonstrated high agreement with histopathological analysis in the detection of atypical epithelial cells in unsuspected OSSN in Brazilian pterygia patients.

Accessibility of sports facilities for persons with reduced mobility and assessment of their motivation for practice.

This study aims to create awareness, both within the scientific community and among providers of sports facilities, for individuals with impaired or reduced mobility, promoting the development of technical solutions that allow greater autonomy and social integration of people with disabilities. The purpose of this work is, on the one hand, to evaluate the accessibility of sports facilities for people with reduced mobility and, on the other hand, to investigate why this user group has such low rates of participation in sporting activities. Firstly, using the Portuguese norms and legislation transcribed from European Community directives, a check list was created comprising all the items that sports facilities should abide by in order to provide accessibility and safety to people with reduced mobility. Another questionnaire was designed aimed at this user group, with questions pertaining to their desire and ability to use sports facilities. This questionnaire was distributed in Portugal, in the greater metropolitan area of Porto, to users of Rehabilitation Centres and Physiotherapy Clinics. The results obtained from the check-list showed the compliances and non-compliances of the respective sports facilities, proving that many barriers preventing the participation of people with reduced mobility still exist. Twenty-four people with permanent impairment of the lower extremities (paraplegia) answered the questionnaire pertaining to the desire and ability to perform physical activity. Two individuals (8%) had sporting activities available to them in their area of residence and only five (21%) performed any physical activity. The main reason given for not taking part in any activity was the lack of adapted sports facilities. All the participants felt that taking part in sports is beneficial. The benefits stated were: general well-being and development of the psychomotor component (e.g. coordination, balance, body posture), of physical condition (e.g. strength, resistance, flexibility) and social integration and quality of life. Besides wellbeing, they also mentioned social interaction, a way of occupying their time and an escape from the routine as the main motives for which they would like to take up physical exercise.

Bioactivity of immobilized EGF on self-assembled monolayers: optimization of the immobilization process.

Last trends in Biomaterials focus the mimic of cellular environments capable to control cellular responses. Epidermal growth factor (EGF) is a pleiotropic cytokine known to regulate cell proliferation, differentiation, and death. This study aims to optimize the immobilization of EGF on 11-mercapto-1-undecyl-tetra(ethylene)glycol (EG4)-self-assembled monolayers (SAMs) and to establish a new model surface to study EGF-mediated signaling. Gold substrates were modified with a monolayer of EG4 and N,N'-carbonyldiimidazole (CDI) was used to activate hydroxyl terminated groups of EG4-SAMs. EGF was then immobilized on activated EG4-SAMs at pH 7.4, 4 degrees C, and 100 rpm. Different immobilization reaction times were tested as well as different CDI concentrations to optimize the reaction conditions and obtain a range of immobilized EGF concentrations on the surfaces. Surface characterization of EGF-SAMs was performed using radiolabeling, water contact angle measurements, X-ray photoelectron spectroscopy, and ELISA. Phosphorylation of EGFR on BT-20 breast cancer cell line by EGF-SAMs was tested by immunostaining. EGF was successfully immobilized on EG4-SAMs, at 4 degrees C and pH 7.4 in a range of concentrations from 3.6 +/- 0.8 to 17.6 +/- 1.5 ng/cm(2). The concentration of EGF increases with immobilization time and with the CDI concentration reaching the maximum for surfaces activated with 30 mg/mL of CDI after 48 h. The bioactivity of EGF-SAMs was confirmed by immunostaining of phospho-EGFR of BT-20 cells. This study described EGF immobilization on EG4-SAMs at different concentrations, which could be important surface models to study specific protein interactions at the molecular level evolving EGF-family of proteins.