Objetivo: A proposta deste trabalho é avaliar o tempo de fibroplasia em tela de polipropileno na correção de hérnias incisionais da parede abdominal, em ratos, por meio da quantidade de colágeno, correlacionando-o com a resposta inflamatória local. Métodos: Trinta e seis ratos machos da linhagem Wistar foram submetidos à ressecção longitudinal de um segmento músculo-aponeurótico e peritoneal (3x2 cm) da parede abdominal, seguida por reforço com tela de polipropileno, em forma de ponte sobre a aponeurose. Os animais foram distribuídos em seis grupos, de acordo com o tempo de fibroplasia a ser estudado (um, dois, três, sete, 21 e 30 dias de pósoperatório). Após os prazos estabelecidos para estudo da fibroplasia, os animais foram submetidos à eutanásia, e a área de fixação da tela de polipropileno foi avaliada histologicamente quanto à reação inflamatória e à percentagem de colágeno pela técnica videomorfométrica assistida por computador. Resultados: O colágeno total foi identificado junto à tela no 3º dia pós-implante, apresentou aumento progressivo na sua proporção em todos os dias subseqüentes até o 21º dia, quando atingiu sua proporção máxima (p<0,001). A partir do dia 3, o colágeno III sofreu um aumento progressivo até o dia 21, quando atingiu sua proporção máxima (p<0,001), e no 30º dia apresentou uma redução significativa (p<0,001). O colágeno tipo I surgiu entre o 7º e o 21º dia, apresentou sua máxima proporção no 21º dia e manteve-se inalterado até o final do período de observação. A relação colágeno tipo I/tipo III aumentou progressivamente até o 30º dia de observação (p<0,001). Os neutrófilos foram identificados no 1º dia pós-implante, mantendo-se junto à tela até o 21º dia. Os macrófagos, gigantócitos e linfócitos foram identificados no 2º dia. Trinta dias após a implantação da tela, desapareceram os neutrófilos e mantiveram-se estáveis as proporções de macrófagos, gigantócitos e linfócitos (p<0,001). Conclusões: Os resultados do presente estudo evidenciaram colágeno total no 3º dia pós-implante, aumentando progressivamente até o 21º dia. O colágeno tipo III foi observado no 3º dia, aumentou até o 21º dia, quando reduziu significativamente. O colágeno tipo I surgiu entre o 7º e o 21º dia, e sua máxima proporção ocorreu no 21º dia, atingindo um platô. A relação do colágeno tipo I/tipo III aumentou progressivamente até o 30º dia, indicando maior proporção de colágeno tipo I ao final do período. O prolongamento da resposta inflamatória da cicatrização e a persistência do processo inflamatório crônico junto à tela não interferiram no tempo da fibroplasia. / Objective: This study assessed the amount of collagen and correlated it with local inflammatory responses to evaluate the length of time required for fibroplasia when polypropylene meshes are used to repair incisional abdominal wall hernias in rats. Methods: Thirty-six male Wistar rats underwent longitudinal resection of a peritoneal and musculoaponeurotic tissue segment (3x2 cm) of the abdominal wall followed by defect reconstruction with polypropylene mesh bridging over aponeurosis. The animals were divided into six groups according to the time points for the analysis of fibroplasia: one, two, three, seven, 21 and 30 days post-implantation. Animals were sacrificed at each time point, and the site where the polypropylene mesh was implanted was evaluated histologically to assess inflammatory response and percentage of collagen using computer-assisted video morphometry. Results: Total collagen was found at the mesh site on the third post-implantation day, and increased progressively on all subsequent days up to the 21st day, when it reached its highest percentage (p<0.001). Type III collagen increased progressively from the 3rd to the 21st day, when it reached its greatest percentage (p<0.001); on the 30th day, it decreased significantly (p>0.001). Type I collagen was first found between the 7th and 21st days, reached its greatest percentage on the 21st day and then remained stable until the 30th day. The type I to type III collagen ratio increased significantly and progressively up to the 30th day (p<0.001). Neutrophils were found at the mesh site from the 1st to the 21st post-implantation day. Macrophages, giant cells and lymphocytes were seen on the 2nd day. Thirty days after mesh implantation, neutrophils disappeared, but the percentages of macrophages, giant cells and lymphocytes remained stable (p<0.001). Conclusion: This study showed that total collagen was fist seen on the 3rd day post-implantation, increased progressively up to the 21st day. Type III collagen was first seen on the 3rd day, increased up to the 21st day, and then decreased significantly. Type I collagen was first seen between the 7th and 21st days and reached its greatest percentage on the 21st day, after which it remained stable. The type I and type III collagen ratio progressively increased up to the 30th day, indicating a greater percentage of type I collagen at the last observational time point. The prolonged healing inflammatory response and the persistence of chronic inflammation surrounding to the mesh did not affect the length of time required for fibroplasia.

Objetivo: A proposta deste trabalho é avaliar o tempo de fibroplasia em tela de polipropileno na correção de hérnias incisionais da parede abdominal, em ratos, por meio da quantidade de colágeno, correlacionando-o com a resposta inflamatória local. Métodos: Trinta e seis ratos machos da linhagem Wistar foram submetidos à ressecção longitudinal de um segmento músculo-aponeurótico e peritoneal (3x2 cm) da parede abdominal, seguida por reforço com tela de polipropileno, em forma de ponte sobre a aponeurose. Os animais foram distribuídos em seis grupos, de acordo com o tempo de fibroplasia a ser estudado (um, dois, três, sete, 21 e 30 dias de pósoperatório). Após os prazos estabelecidos para estudo da fibroplasia, os animais foram submetidos à eutanásia, e a área de fixação da tela de polipropileno foi avaliada histologicamente quanto à reação inflamatória e à percentagem de colágeno pela técnica videomorfométrica assistida por computador. Resultados: O colágeno total foi identificado junto à tela no 3º dia pós-implante, apresentou aumento progressivo na sua proporção em todos os dias subseqüentes até o 21º dia, quando atingiu sua proporção máxima (p<0,001). A partir do dia 3, o colágeno III sofreu um aumento progressivo até o dia 21, quando atingiu sua proporção máxima (p<0,001), e no 30º dia apresentou uma redução significativa (p<0,001). O colágeno tipo I surgiu entre o 7º e o 21º dia, apresentou sua máxima proporção no 21º dia e manteve-se inalterado até o final do período de observação. A relação colágeno tipo I/tipo III aumentou progressivamente até o 30º dia de observação (p<0,001). Os neutrófilos foram identificados no 1º dia pós-implante, mantendo-se junto à tela até o 21º dia. Os macrófagos, gigantócitos e linfócitos foram identificados no 2º dia. Trinta dias após a implantação da tela, desapareceram os neutrófilos e mantiveram-se estáveis as proporções de macrófagos, gigantócitos e linfócitos (p<0,001). Conclusões: Os resultados do presente estudo evidenciaram colágeno total no 3º dia pós-implante, aumentando progressivamente até o 21º dia. O colágeno tipo III foi observado no 3º dia, aumentou até o 21º dia, quando reduziu significativamente. O colágeno tipo I surgiu entre o 7º e o 21º dia, e sua máxima proporção ocorreu no 21º dia, atingindo um platô. A relação do colágeno tipo I/tipo III aumentou progressivamente até o 30º dia, indicando maior proporção de colágeno tipo I ao final do período. O prolongamento da resposta inflamatória da cicatrização e a persistência do processo inflamatório crônico junto à tela não interferiram no tempo da fibroplasia. / Objective: This study assessed the amount of collagen and correlated it with local inflammatory responses to evaluate the length of time required for fibroplasia when polypropylene meshes are used to repair incisional abdominal wall hernias in rats. Methods: Thirty-six male Wistar rats underwent longitudinal resection of a peritoneal and musculoaponeurotic tissue segment (3x2 cm) of the abdominal wall followed by defect reconstruction with polypropylene mesh bridging over aponeurosis. The animals were divided into six groups according to the time points for the analysis of fibroplasia: one, two, three, seven, 21 and 30 days post-implantation. Animals were sacrificed at each time point, and the site where the polypropylene mesh was implanted was evaluated histologically to assess inflammatory response and percentage of collagen using computer-assisted video morphometry. Results: Total collagen was found at the mesh site on the third post-implantation day, and increased progressively on all subsequent days up to the 21st day, when it reached its highest percentage (p<0.001). Type III collagen increased progressively from the 3rd to the 21st day, when it reached its greatest percentage (p<0.001); on the 30th day, it decreased significantly (p>0.001). Type I collagen was first found between the 7th and 21st days, reached its greatest percentage on the 21st day and then remained stable until the 30th day. The type I to type III collagen ratio increased significantly and progressively up to the 30th day (p<0.001). Neutrophils were found at the mesh site from the 1st to the 21st post-implantation day. Macrophages, giant cells and lymphocytes were seen on the 2nd day. Thirty days after mesh implantation, neutrophils disappeared, but the percentages of macrophages, giant cells and lymphocytes remained stable (p<0.001). Conclusion: This study showed that total collagen was fist seen on the 3rd day post-implantation, increased progressively up to the 21st day. Type III collagen was first seen on the 3rd day, increased up to the 21st day, and then decreased significantly. Type I collagen was first seen between the 7th and 21st days and reached its greatest percentage on the 21st day, after which it remained stable. The type I and type III collagen ratio progressively increased up to the 30th day, indicating a greater percentage of type I collagen at the last observational time point. The prolonged healing inflammatory response and the persistence of chronic inflammation surrounding to the mesh did not affect the length of time required for fibroplasia.

Objetivo: A proposta deste trabalho é avaliar o tempo de fibroplasia em tela de polipropileno na correção de hérnias incisionais da parede abdominal, em ratos, por meio da quantidade de colágeno, correlacionando-o com a resposta inflamatória local. Métodos: Trinta e seis ratos machos da linhagem Wistar foram submetidos à ressecção longitudinal de um segmento músculo-aponeurótico e peritoneal (3x2 cm) da parede abdominal, seguida por reforço com tela de polipropileno, em forma de ponte sobre a aponeurose. Os animais foram distribuídos em seis grupos, de acordo com o tempo de fibroplasia a ser estudado (um, dois, três, sete, 21 e 30 dias de pósoperatório). Após os prazos estabelecidos para estudo da fibroplasia, os animais foram submetidos à eutanásia, e a área de fixação da tela de polipropileno foi avaliada histologicamente quanto à reação inflamatória e à percentagem de colágeno pela técnica videomorfométrica assistida por computador. Resultados: O colágeno total foi identificado junto à tela no 3º dia pós-implante, apresentou aumento progressivo na sua proporção em todos os dias subseqüentes até o 21º dia, quando atingiu sua proporção máxima (p<0,001). A partir do dia 3, o colágeno III sofreu um aumento progressivo até o dia 21, quando atingiu sua proporção máxima (p<0,001), e no 30º dia apresentou uma redução significativa (p<0,001). O colágeno tipo I surgiu entre o 7º e o 21º dia, apresentou sua máxima proporção no 21º dia e manteve-se inalterado até o final do período de observação. A relação colágeno tipo I/tipo III aumentou progressivamente até o 30º dia de observação (p<0,001). Os neutrófilos foram identificados no 1º dia pós-implante, mantendo-se junto à tela até o 21º dia. Os macrófagos, gigantócitos e linfócitos foram identificados no 2º dia. Trinta dias após a implantação da tela, desapareceram os neutrófilos e mantiveram-se estáveis as proporções de macrófagos, gigantócitos e linfócitos (p<0,001). Conclusões: Os resultados do presente estudo evidenciaram colágeno total no 3º dia pós-implante, aumentando progressivamente até o 21º dia. O colágeno tipo III foi observado no 3º dia, aumentou até o 21º dia, quando reduziu significativamente. O colágeno tipo I surgiu entre o 7º e o 21º dia, e sua máxima proporção ocorreu no 21º dia, atingindo um platô. A relação do colágeno tipo I/tipo III aumentou progressivamente até o 30º dia, indicando maior proporção de colágeno tipo I ao final do período. O prolongamento da resposta inflamatória da cicatrização e a persistência do processo inflamatório crônico junto à tela não interferiram no tempo da fibroplasia. / Objective: This study assessed the amount of collagen and correlated it with local inflammatory responses to evaluate the length of time required for fibroplasia when polypropylene meshes are used to repair incisional abdominal wall hernias in rats. Methods: Thirty-six male Wistar rats underwent longitudinal resection of a peritoneal and musculoaponeurotic tissue segment (3x2 cm) of the abdominal wall followed by defect reconstruction with polypropylene mesh bridging over aponeurosis. The animals were divided into six groups according to the time points for the analysis of fibroplasia: one, two, three, seven, 21 and 30 days post-implantation. Animals were sacrificed at each time point, and the site where the polypropylene mesh was implanted was evaluated histologically to assess inflammatory response and percentage of collagen using computer-assisted video morphometry. Results: Total collagen was found at the mesh site on the third post-implantation day, and increased progressively on all subsequent days up to the 21st day, when it reached its highest percentage (p<0.001). Type III collagen increased progressively from the 3rd to the 21st day, when it reached its greatest percentage (p<0.001); on the 30th day, it decreased significantly (p>0.001). Type I collagen was first found between the 7th and 21st days, reached its greatest percentage on the 21st day and then remained stable until the 30th day. The type I to type III collagen ratio increased significantly and progressively up to the 30th day (p<0.001). Neutrophils were found at the mesh site from the 1st to the 21st post-implantation day. Macrophages, giant cells and lymphocytes were seen on the 2nd day. Thirty days after mesh implantation, neutrophils disappeared, but the percentages of macrophages, giant cells and lymphocytes remained stable (p<0.001). Conclusion: This study showed that total collagen was fist seen on the 3rd day post-implantation, increased progressively up to the 21st day. Type III collagen was first seen on the 3rd day, increased up to the 21st day, and then decreased significantly. Type I collagen was first seen between the 7th and 21st days and reached its greatest percentage on the 21st day, after which it remained stable. The type I and type III collagen ratio progressively increased up to the 30th day, indicating a greater percentage of type I collagen at the last observational time point. The prolonged healing inflammatory response and the persistence of chronic inflammation surrounding to the mesh did not affect the length of time required for fibroplasia.

In recent years, concern about a new epidemic of retinopathy of prematurity (ROP) has focused attention on the increasing incidence of the disease and the factors responsible for its most severe consequences. Two studies designed to address these issues were done using data from three sources: the B.C. Health Surveillance Registry (Registry), Physicians's Notices of Livebirth (PNOB), and the Vancouver General Hospital (VGH). In the first study, Registry and PNOB records were used to determine crude annual birth weight-specific incidence rates for ROP in infants liveborn in the Province of British Columbia (B.C.) in the period 1952-1983. These rates showed that, in B.C., the original epidemic of the disease ended in 1954. Linear regression lines fitted for each of four birth weight categories showed that, in the 29 year period after 1954, there was a significant increase in the incidence of ROP-induced blindness in infants weighing less than 1000 grams at birth. To refine this observation, the data were sub-divided: the 29 year period, to two smaller periods, 1955-1964 and 1965-1983; the less than 1000 gram birth weight category to two sub-categories, 500-749 and 750-999 grams. Since the inter-period incidence should have been similar if the birth weight-specific incidence had not changed since the end of the original epidemic, the crude weight-specific rates for ROP-induced blindness in the early period were used to calculate the expected number of cases in the later period. When weight-standardized incidence ratios (SIR's) and 95% confidence limits were calculated, the results showed that, in the 750-999 gram sub-category, the SIR was significantly increased. Infants born in the period 1965-1983 were 3.07 times more likely to be ROP: blind than their equal weight counterparts in the earlier period. In infants weighing 500-749 and 1000 grams or more, there was no evidence to suggest an increase in incidence after 1954. The second study was done to determine the cofactors that differentiate infants who are blinded by ROP from those who are not. Infants were included if (i) they were born in B.C. between 1955 and 1983, (ii) they were known to the Registry as being ROP: blind (cases) or not blind (controls), and (iii) they were born in or admitted to the VGH within 28 days of birth. When the data from all three data sources were dichotomized and analyzed using univariate techniques, two variables, respiratory distress syndrome (RDS) and neonatal weight loss, showed a significantly protective effect. The effect of RDS disappeared when the data were stratified by birth interval indicating that the observed association was confounded by time. When the variables were reanalyzed in continuous form, none were significantly associated with visual outcome. However, since the power of the cofactor study was extremely low, none of the variables that were included can be eliminated as potential cofactors for the induction of blindness in infants with ROP. / Medicine, Faculty of / Population and Public Health (SPPH), School of / Graduate

La Retinopatía de la Prematuridad (ROP) es una retinopatía vasoproliferativa que afecta a los recién nacidos inmaduros cuya etiología es aún desconocida. Algunos estudios han sugerido la asociación entre la exposición precoz a la luz y ésta patología. Se hizo un estudio para demostrar si la oclusión ocular se relaciona con la incidencia de retinopatía en los recién nacidos prematuros en el Instituto Materno Perinatal. El estudio analítico cuasi-experimental, comprende a todos los prematuros de muy bajo peso al nacer (< 1301 grs.) y menor de 32 semanas, nacidos entre el 1º de Enero del 2005 al 31 de Diciembre del 2007. Se consideró 201 pacientes que cumplieron con los criterios de inclusión, éstos fueron divididos en dos grupos, uno a los que se les ocluyeron ambos ojos con antifaz de tela negra (estudio) y otro sin oclusión expuesto a la luz natural del ambiente (control) .Ambos grupos fueron homogéneos en edad, sexo peso y cuidados de enfermeria. Se comparó la incidencia de ROP en ambos grupos, presentando algún grado de ROP 25 de 97 pacientes con ojos ocluidos (25.8 %) y 34 de 104 pacientes con ojos no ocluidos (32.7 %). Se determinó que no hubo relación entre la incidencia de ROP de los niños expuestos o no a la luz ambiental, RR 1.27 IC 95% (0.82-1.96), y se encontró beneficio moderado para los grados severos (grado 2) de ésta enfermedad, en los casos ocluidos RR 0.55 IC 95 % (0.29 - 1.04). Se concluye que la oclusión no disminuye la incidencia para desarrollar retinopatía en nacidos prematuros de muy bajo peso., pero si tiene efecto protector para casos severos de ésta enfermedad. -- PALABRAS CLAVES: Retinopatía del prematuro, oclusión ocular, luz ambiental. / -- Retinopathy of Prematurity (ROP) is a vasoproliferative retinopathy that affects immature infants whose etiology remains unknown. Some studies have suggested an association between early exposure to light and this pathology. A study was done to demonstrate if the ocular occlusion is related to the incidence of retinopathy in premature infants in the Maternal Perinatal Institute. The analytical quasi-experimental study, includes all the premature babies of very low weight (<1301 grams.) and less than 32 weeks, born between January 01, 2005 to December 31, 2007. It was considered to be 201 patients who expired with the criteria of incorporation, they were divided in two groups, one of them whom became occlusion both eyes with mask of black fabric (study) and other one without occlusion exposed to the natural light of the environment (control). Ambos groups, were homogeneous in age, sex, weigh and taken care of nursery. The ROP's incident was compared in both groups presenting some ROP's degree 25 of 97 patients with eyes occluded (25.8 %) and 34 of 104 patients with eyes not occluded (32.7 %). The study determined that it was no relation between ROP's incident of the exposed children or not to the environmental light RR 1.27 IC 95% (0.82-1.96), and there were benefit moderated for the severe degree (degree 2) in the occluded patients RR 0.55 IC 95 % (0.29 - 1.04). It concludes that the occlusion does not diminish the incident to develop retinopathies in born premature babies of very low weight but it has protective effect in severe cases of this one disease. -- KEY WORDS: Retinopathy of prematurity, ocular occlusion, ambient light. / Tesis

Indiana University-Purdue University Indianapolis (IUPUI) / Prevention of pathological angiogenesis is a key strategy for treatment of common blinding ocular diseases such as retinopathy of prematurity, proliferative diabetic retinopathy, and wet age-related macular degeneration. The current treatment strategies are associated with partial vision loss and are ineffective in a significant patient population. Hence novel drugs as well as new ways to target ocular angiogenesis are needed for treating these diseases. I pursued a natural antiangiogenic compound, cremastranone, to develop novel drug leads and to find new targets. The objective of my doctoral thesis project was to elucidate cremastranone’s molecular mechanism of action and optimize its structureactivity relationship (SAR). In order to achieve this goal, with the help of chemistry collaborators cremastranone was synthesized for the first time. I showed that cremastranone has 50-fold more potency against endothelial cells as compared to nonendothelial cells, and also tested a novel active isomer, SH-11052. By SAR studies I identified a potent molecule, SH-11037, that has 10-fold more selectivity against retinal endothelial cells as compared to macrovascular endothelial cells. I then elucidated cremastranone’s molecular mechanism using a chemical proteomic approach. I identified ferrochelatase (FECH) as a specific interacting protein partner of cremastranone using photoaffinity chromatography. Hence, I hypothesized that cremastranone exerts its antiangiogenic activities through modulation of the functions of FECH. Cremastranone inhibited the enzymatic activity FECH in endothelial cells. Therefore, I investigated the role of FECH in ocular angiogenesis. Partial loss of FECH, using a siRNA-based knock down approach, decreased retinal angiogenesis both in vitro and in vivo in mouse models. Knock down of FECH decreased the expression levels of key proangiogenic proteins HIF-1α, eNOS, and VEGFR2. This work suggests that ferrochelatase plays an important, previously undocumented role in angiogenesis and that targeting of this enzyme by cremastranone might be exploited to inhibit pathological angiogenesis in ocular diseases.