Ricardo Luiz Smith

Human immature dental pulp stem cells share key characteristic features with limbal stem cells.

Objectives:  Limbal stem cells (LSC) are self‐renewing, highly proliferative cells in vitro, which express a set of specific markers and in vivo have the capacity to reconstruct the entire corneal epithelium in cases of ocular surface injury. Currently, LSC transplantation is a commonly used procedure in patients with either uni‐ or bilateral total limbal stem cells deficiency (TLSCD). Although LSC transplantation holds great promise for patients, several problems need to be overcome. In order to find an alternative source of cells that can partially substitute LSC in cornea epithelium reconstruction, we aimed at investigating whether human immature dental pulp stem cells (hIDPSC) would present similar key characteristics as LSC and whether they could be used for corneal surface reconstruction in a rabbit TLSCD model.

Effects of indocyanine green injection on the retinal surface and into the subretinal space in rabbits.

Purpose To evaluate the effects of indocyanine green (ICG) injection on the retinal surface and into the subretinal space of rabbit eyes. Methods Twenty-two Dutch-belted rabbits underwent two-port vitrectomy followed by injection of ICG (5 mg/mL) on the retinal surface and into the subretinal space. Balanced salt solution (BSS) was also injected subretinally. The locations where ICG was delivered (both epiretinal and subretinal) were exposed to light from an endoilluminator for 7 minutes. The animals were examined at 1, 7, and 14 days after surgery. The eyes were studied by fluorescein angiography as well as light and electron microscopy. Results No damage was observed after epiretinal ICG injection, but subretinal ICG injection resulted in damage to the outer nuclear layer, photoreceptor inner and outer segments, and retinal pigment epithelium. This damage was more severe with longer follow-up. Control experiments without ICG, in which balanced salt solution was injected into the subretinal space or light was delivered on the epiretinal surface, demonstrated only damage to the photoreceptor outer segments. Conclusion Subretinal delivery of ICG (5 mg/mL) in rabbits induces retinal pigment epithelium, photoreceptor inner and outer segment, and outer nuclear layer damage. These mechanisms of damage may explain the retinal pigment epithelium changes that are sometimes seen after ICG-assisted internal limiting membrane peeling in humans.

Choriocapillaris photodynamic therapy using indocyanine green

PURPOSE To evaluate the potential of photodynamic therapy using indocyanine green for occlusion of choroidal neovascularization, the authors studied efficiency and collateral damage of photodynamic therapy-induced photothrombosis in the rabbit choriocapillary layer. METHODS Fundus photography, fluorescein angiography, and light and transmission electron microscopy were used to study the efficiency of photodynamic therapy-induced photothrombosis using indocyanine green as the photosensitizer, and to assess the resultant collateral damage. The delivery system consisted of a modified infrared diode laser tuned to 810 nm, near the maximum absorption peak of indocyanine green. RESULTS Choriocapillary occlusion was achieved at indocyanine green doses of 10 and 20 mg/kg and a radiant as low as 6.3 J/cm(2). When photodynamic therapy was performed with indocyanine green doses of 10 mg/kg, damage to the neural retina was minimal. Only inner photoreceptor segments showed degeneration, probably secondary to choroidal ischemia. Bruch membrane remained intact. Retinal pigment epithelium was invariably damaged, as seen with other photosensitizers. Temporary occlusion of large choroidal vessels occurred at both dye doses. CONCLUSIONS In this experimental study, photodynamic therapy using indocyanine green and 810-nm light irradiation produced endothelium-bound intraluminal photothrombosis, with preservation of the retinal architecture and minimal loss of visual cells. Membrane targetability, hydrophilic and fluorescent properties, and activation at 805 nm suggest indocyanine green as a potential photosensitizer for choroidal neovascularization. These combined considerations point toward further study of photodynamic therapy using indocyanine green for the treatment of choroidal vascular disease.

Craniofacial Asymmetry in Development: An Anatomical Study

The purpose of this study was to evaluate the occurrence of craniofacial asymmetries in four areas of human skulls of various age groups to test the hypothesis that there is craniofacial symmetry before the chewing habit is established. The data were obtained from 95 skulls of fetuses, infants, children, and adults, from the collection of Federal University of São Paulo. The following measurements were taken on each skull with a digital caliper: from the infraorbital foramen to the anterior nasal spine (IOF); from the greater palatine foramen to the posterior nasal spine (GPF); from the spinous foramen to the basion (SF); and from the spinous foramen to the zygomatic arch (ZA). On different occasions, each measurement was taken three times on both sides of the skull in random order. The mean of the right-side measurements were subtracted from the mean of the left-side measurements, and the differences were transformed into percentages. Comparisons were made by analysis of variance. The presence of cranial asymmetry was statistically significant throughout the whole sample. The minimum value found was 2.8% and the maximum 6.5%. All age groups presented the same degree of asymmetry of distances IOF, GPF, and SF. The group of infants presented a higher degree of asymmetry on distance ZA, followed by the groups of fetuses, children, and adults. This study confirmed statistically significant craniofacial asymmetry in fetuses and infants (before dentition). Therefore, the hypothesis that craniofacial asymmetry only appears after establishment of the chewing habit was not supported.

Sex Determination Using Mastoid Process Measurements in Brazilian Skulls

Las caracteristicas del proceso mastoides son de gran utilidad para el diagnostico del sexo. De Paiva & Segre (2003) describieron que era posible el diagnostico del sexo, en base a la determinacion del area del triangulo formado entre los puntos porion, mastoidale y asterion. El proposito de este estudio fue determinar la existencia de dimorfismo sexual en las dimensiones y el area del triangulo mastoideo, mediante analisis estadistico y de funcion discriminante. Se utilizaron 81 craneos de individuos Brasilenos, pertenecientes a la coleccion de la UNIFESP, de sexo y edad conocidos, 50 hombres y 31 mujeres, de entre 40 y 70 anos (media 51.58 anos, DS 7.319). La mayoria de las dimensiones lineales y las areas calculadas fueron mayores en hombres que en mujeres. Solo la distancia porion-mastoidale (Po-Ma), el area del triangulo mastoideo derecho y el area total resultaron mayores y significativas en hombres con p<0.01. El analisis de la funcion discriminante mostro que el conjunto de dimensiones lineales analizadas presenta un bajo poder discriminante (Lambda de Wilks= 0.960, Correlacion Canonica= 0.199), solo Po-Ma permitio clasificar en grupos de hombres y de mujeres con una exactitud general del 64.2%, con una alta sensibilidad para clasificar hombres (93%) y una muy baja sensibilidad para mujeres (17.7%). Estos resultados indican que la determinacion del sexo en base a las dimensiones del triangulo mastoideo, tiende a subestimar a las mujeres, siendo de baja utilidad en la practica.

Evaluating Accuracy and Precisión in Morphologic Traits for Sexual Dimorphism in Malnutrition Human Skull: a Comparative Study

Uno de los propositos mas importantes del protocolo antropologico de estudios de osamentas humanas, es la determinacion del sexo. Algunos craneos pueden ser clasificados erroneamente debido a condiciones sistemicas presentadas durante la vida de individuos que hayan afectado a las fuerzas musculares que actuan sobre el hueso, como es el caso de la desnutricion severa. El proposito de este estudio fue determinar la exactitud y confiabilidad, sobre los parametros morfologicos determinantes de dimorfismo sexual, clasicamente estudiados en el analisis visual del craneo, para los dos grupos que componen la muestra, uno con desnutricion acompanando el momento del deceso y otro con causa de muerte sin ese factor acompanante. Se utilizaron 163 craneos humanos de sexo, edad y causa de muerte conocido, pertenecientes a la coleccion de la UNIFESP. Se realizaron la inspeccion visual del craneo y el diagnostico de sexo en base a: tamano y arquitectura general del craneo, aspecto del hueso cigomatico, tamano y rugosidades del proceso mastoides, rugosidades del hueso occipital y aspecto general de la mandibula. Los craneos fueron clasificados en dos grupos, de acuerdo a si presentaron desnutricion severa descrita en su causa de muerte (n=60) o presentaban otra causa de muerte (n=103). Los resultados de la inspeccion se compararon con la del registro para determinar la exactitud en el diagnostico y la sensibilidad del metodo para hombres y mujeres, en los dos grupos analizados. En el grupo con desnutricion, el mejor indicador resulto el aspecto del hueso cigomatico, con un 86,6% de exactitud. En el otro grupo, la mayor exactitud se encontro en el tamano del proceso mastoides (87,3%). En ambos grupos, los mayores valores de sensibilidad fueron para hombres (83,3-95,3%). La sensibilidad para mujeres presento menores valores en el grupo sin desnutricion (69,2-76,9%) que en el grupo con desnutricion (72,4-89,65%), estas diferencias resultaron estadisticamente significativas (p<0,05).

Sexual dimorphism in the foramen magnum dimensions

El proposito de este estudio fue evaluar la presencia de dimorfismo sexual en las dimensiones del foramen magno. Se analizaron 211 craneos humanos de la coleccion de la Universidade Federal de Sao Paulo, con registro de sexo y edad, se determinaron los diametros anteroposterior y transverso maximo del foramen magno y se establecieron las diferencias por sexo (p<0,05), se determinaron las funciones lineales discriminantes de Fischer y se calculo la utilidad para la clasificacion de estas variables. Todas las dimensiones resultaron mayores y significativas en craneos de hombres, las dimensiones del foramen magno presentaron un bajo poder discriminante y clasificaron correctamente el 66,5% de los craneos. Nuestros resultados muestran que este indicador cuantitativo es de limitada utilidad practica y debe ser complementado con los indicadores cualitativos de dimorfismo sexual en el hueso occipital para mejorar la exactitud en el diagnostico del sexo.

Ocular toxoplasmosis: the role of retinal pigment epithelium migration in infection

AbstractOur aim was to study the migration of retinal pigmented epithelium (RPE) into the retinal layer during infection of C57BL/6 mice with Toxoplasma gondii. Eyes from infected and non-infected animals were analyzed on the 60th day of infection by light and transmission electron microscopy. Non-infected eyes showed a normal morphology. In contrast, we observed free parasites in the retinal vasculature, the presence of mononuclear inflammatory infiltrate (MNII) and parasites in the vasculature of choroids in infected eyes. No inflammatory infiltrate was observed; RPE cells were identified near the MNII in nuclear and plexiforme layers. RPE cells were also found on the ganglion cell layer and in the outer segments of the photoreceptor. The morphology showed that RPE cells caused a discontinuity in the nuclear and plexiforme layers. Clusters of parasites were found surrounded by RPE cells and MNII in the inner plexiforme layers. Ultrastructural analysis showed that RPE cells migrated through the epithelium into the inner retinal layers. We did not observe Toxoplasma cysts in many eyes in which pathological changes were detected. Only 8.3% of the animals had Toxoplasma cysts in the inner nuclear layer in the absence of inflammatory cells. The migration of RPE cells can be triggered by a disruption of the RPE monolayer or injury to the neural retina, as in the case of toxoplasmosis.

Performance Evaluation as a Diagnostic Test for Traditional Methods for Forensic Identification of Sex

En el estudio fue evaluada la calidad, como una prueba diagnostica de los principales indicadores morfologicos de dimorfismo sexual a traves de la antropometria, herramientas bioestadisticas y epidemiologia clinica. Fueron utilizados 284 craneos de individuos adultos brasilenos, de los cuales 187 eran varones y 97 mujeres. Se realizo un estudio de corte transversal de evaluacion de prueba diagnostica, mediante un enfoque cualitativo en base al examen visual, de 16 indicadores tradicionales morfologicos de dimorfismo sexual, determinandose para cada uno de ellos los niveles de exactitud y de sensibilidad, valores predictivos, coeficientes de probabilidad y Odds ratios. Todos los indicadores estudiados presentaron altos niveles de exactitud (84,75-72,89%). Los mejores indicadores se encontraron en los rasgos cuya formacion esta relacionada con la insercion y la accion de los grandes grupos musculares. En 14 de los 16 indicadores, el error intraobservador fue <10%. Los mejores indicadores morfologicos de dimorfismo sexual fueron: proceso mastoides, hueso cigomatico, mandibula y rugosidades del hueso occipital. Los autores concluyeron que los indicadores morfologicos de dimorfismo presentan un adecuado rendimiento como pruebas de diagnostico; sin embargo, los valores de exactitud y sensibilidad deben ser complementados con indicadores mas solidos que sean independientes de la distribucion de la muestra, e integren a los errores diagnosticos, tales como los coeficientes de probabilidad, odds ratios y valores predictivos positivos.

Gap Junctions between Mast Cells and Fibroblasts in the Developing Avian Eye

Mast cells are present in the eye of chick embryos from the 14th day onward, displaying metachromatic granules, mainly in the iris anterior surface and pectinate ligament. Ultrastructurally these cells show electron-dense granules and a few thin and short cytoplasmic projections in close contact with fibroblasts. Sometimes these contacts are extensive, with long fibroblast projections partially involving the mast cells. Gap junctions between mast cells and fibroblasts are observed only in the eyes of 16- and 20-day-old embryos. These intercellular specializations are represented by a close apposition of cytoplasmic membranes with an extension up to 300 nm. Gap junctions between mast cells and fibroblasts were not observed previously in vivo or in vitro, although in vitro studies have shown that a number of functionally critical interactions may occur between these cells. Our morphological findings suggest that, in vivo, fibroblasts interact with mast cells and may influence their maturation.