EVIDENCE FOR MECHANICAL STRAINS INFLUENCE IN OSTEOPHYTE DEVELOPMENT

Purpose: Osteophytes are osteo-cartilaginous metaplastic tissue outgrowths of bone capped by cartilage usually found in degenerative and inflammatory joint disease. The presence and degree of maturity of osteophytes, along with joint space narrowing, are the main radiographic criteria for diagnosis and grading osteoarthritis (OA). Although osteophytes are known for being anatomic signs of advanced OA, they can occur in non-symptomatic joints, in joints with no other observable alterations, and in early stage OA. It remains unclear if they develop from molecular, physiological and/or mechanical stimuli. We hypothesized that mechanical strains play a role in osteophyte development. The overall objective of this thesis was to find evidence that osteophytes are influenced by mechanical strains. Methods: The first project was to develop a mechanically-induced osteophyte animal model. One single impact load that was reported to induce moderate joint damage was applied to the periosteum of the rat knee. Animals were sacrificed at four time points to characterize the evolution of damaged tissue and the joint by histology. A second study using human mature hip osteophytes was conducted to evaluate if mature osteophyte presented histological signs of proliferating and developmental processes. The histological characterization of mature osteophyte was used to compare findings of the mechanically-induced osteophyte in the animal model to validate the use of this rodent model in studying some aspect of osteophyte development of human. Lastly, a detailed three-dimensional (3D) radiological morphometric analysis was performed on microscopic computed tomography (µCT) scanned femoral heads collected from total hip arthroplasty patients presenting mature hip osteophytes. Quantitative morphometric measures of osteophytes internal structure was compared to three regions of the femoral head of known quality of organisation and mechanical constraint. Results and Conclusion: Osteophyte can be mechanically induced by a single load impact to the joint periosteum, indicating that a moderate trauma to the periosteal layer of the joint may play a role in osteophyte development. Mature osteophytes have proliferation, developing and remodelling zones and have trabecular structures. Mechanically-induced osteophytes and mature osteophytes presented similar histological composition. Mature osteophytes have organized internal structure. These results provide evidence that mechanical strain can influence osteophyte development.

Steroid induced neuroprotection of damaged photoreceptor cells

Retinitis Pigmentosa (RP) is the name given to a group of hereditary diseases causing progressive and degenerative blindness. RP affects over 1 in 4000 individuals, making it the most prevalent inherited retinal disease worldwide, yet currently there is no cure. In 2011, our group released a paper detailing the protective effects of the synthetic progestin ‘Norgestrel’. A common component of the female oral contraceptive pill, Norgestrel was shown to protect against retinal cell death in two distinct mouse models of retinal degeneration: in the Balb/c light damage model and the Pde6brd10 (rd10) model. Little was known of the molecular workings of this compound however and thus this study aimed to elucidate the protective manner in which Norgestrel worked. To this aim, the 661W cone photoreceptor-like cell line and ex vivo retinal explanting was utilised. We found that Norgestrel induces a increase in neuroprotective basic fibroblast growth factor (bFGF) with subsequent downstream actions on the inhibition of glycogen synthase kinase 3β. Progesterone receptor expression was subsequently characterised in the C57 and rd10 retinas and in the 661W cell line. Norgestrel caused nuclear trafficking of progesterone receptor membrane complex one (PGRMC1) in 661W cells and thus Norgestrel was hypothesised to work primarily through the actions of PGRMC1. This trafficking was shown to be responsible for the critical upregulation of bFGF and PGRMC1- Norgestrel binding was proven to cause a neuroprotective bFGF-mediated increase in intracellular calcium. The protective properties of Norgestrel were further studied in the rd10 mouse model of retinitis pigmentosa. Using non-invasive diet supplementation (80mg/kg), we showed that Norgestrel gave significant retinal protection out to postnatal day 40 (P40). Overactive microglia have previously been shown to potentiate photoreceptor cell loss in the degenerating rd10 retina and thus we focussed on Norgestrel-mediated changes in photoreceptor-microglial crosstalk. Norgestrel acted to dampen pro-inflammatory microglial cell reactivity, decreasing chemokine (MCP1, MCP3, MIP-1α, MIP-1β) and subsequent damaging cytokine (TNFα, Il-1β) production. Critically, Norgestrel up-regulated photoreceptor-microglial, fractalkine-CX3CR1 signalling 1000-fold in the P20 rd10 mouse. Known to prevent microglial activation, we hypothesise that Norgestrel acts as a vital anti-inflammatory in the diseased retina, driving fractalkine-CX3CR1 signalling to delay retinal degeneration. This study stands to highlight some of the neuroprotective mechanisms utilised by Norgestrel in the prevention of photoreceptor cell death. We identify for the first time, not only a pro-survival pathway activated directly in photoreceptor cells, but also a Norgestreldriven mediation of an otherwise damaging microglial cell response. All taken, these results form the beginning of a case to bring Norgestrel to clinical trials, as a potential therapeutic for the treatment of RP.

Progesterone attenuates microglial-driven retinal degeneration and stimulates protective Fractalkine-CX3CR1 signaling

Retinitis pigmentosa (RP) is a degenerative disease leading to photoreceptor cell loss. Mouse models of RP, such as the rd10 mouse (B6.CXBl-Pde6brd10/J), have enhanced our understanding of the disease, allowing for development of potential therapeutics. In 2011, our group first demonstrated that the synthetic progesterone analogue ‘Norgestrel’ is neuroprotective in two mouse models of retinal degeneration, including the rd10 mouse. We have since elucidated several mechanisms by which Norgestrel protects stressed photoreceptors, such as upregulating growth factors. This study consequently aimed to further characterize Norgestrel’s neuroprotective effects. Specifically, we sought to investigate the role that microglia might play; for microglial-derived inflammation has been shown to potentiate neurodegeneration. Dams of post-natal day (P) 10 rd10 pups were given a Norgestrel-supplemented diet (80mg/kg). Upon weaning, pups remained on Norgestrel. Tissue was harvested from P15-P50 rd10 mice on control or Norgestrel-supplemented diet. Norgestrel-diet administration provided significant retinal protection out to P40 in rd10 mice. Alterations in microglial activity coincided with significant protection, implicating microglial changes in Norgestrel-induced neuroprotection. Utilizing primary cultures of retinal microglia and 661W photoreceptor-like cells, we show that rd10 microglia drive neuronal cell death. We reveal a novel role of Norgestrel, acting directly on microglia to reduce pro-inflammatory activation and prevent neuronal cell death. Norgestrel effectively suppresses cytokine, chemokine and danger-associated molecular pattern molecule (DAMP) expression in the rd10 retina. Remarkably, Norgestrel upregulates fractalkine-CX3CR1 signaling 1 000-fold at the RNA level, in the rd10 mouse. Fractalkine-CX3CR1 signaling has been shown to protect neurons by regulating retinal microglial activation and migration. Ultimately, these results present Norgestrel as a promising treatment for RP, with dual actions as a neuroprotective and anti-inflammatory agent in the retina.

Culturas celulares

As culturas celulares apareceram no século XX e têm vindo a evoluir como método de estudo e análise de patologias e de novos medicamentos para as combater. Com o passar dos anos surgiram diferentes métodos de cultivo de células diversificando assim as culturas celulares em mono-camada, suspensas e imobilizadas. Um dos propósitos das culturas celulares é a manipulação das células de modo a permitir a visualização das alterações celulares como o envelhecimento da célula (senescência), a apoptose, a necrose e a autofagia da célula. Este método tem provado ser útil tanto no diagnostico e tratamento de várias patologias como em doenças oncológicas, neuro-degenerativas ou do sistema imunitário.

Microglia as therapeutic targets in retinal degeneration: role of translocator protein (18 κDa) (TSPO) and minocycline.

Microglia are the resident immune cells of the central nervous system (CNS) and play an important role in innate immune defense as well as tissue homeostasis. Chronic microglial reactivity, microgliosis, is a general hallmark of inflammatory and degenerative diseases that affect the CNS, including the retina. There is increasing evidence that chronic microgliosis is more than just a bystander effect, but rather actively contributes to progression of degeneration through processes such as toxic nitric oxide (NO) production and even phagocytosis of stressed but viable photoreceptors. Therefore immunmodulation of microglia presents a possible therapeutic strategy for retinal degenerations. Notably, the expression of the mitochondrial translocator protein 18 (κDa) (TSPO) is highly elevated in reactive microglia as seen in several neuroinflammatory diseases such as Alzheimer’s disease, Parkinson’s disease and multiple sclerosis. Therefore it is used as a gliosis biomarker in the brain. Moreover TSPO ligands show potent effects in resolving neuroinflammatory brain disorders. However, TSPO expression in the eye had not been investigated before. Further, it was unknown whether TSPO ligands’ potent immunomodulatory effects could be used to treat retinal degenerations. To fill this gap, the study aimed to analyze whether TSPO is also a potential biomarker for degenerative processes in the retina. Moreover the thesis attempted to determine whether a specific TSPO ligand, XBD173, might modulate microglial reactivity and is a potent therapeutic, to treat retinal degenerative diseases. The findings revealed that TSPO is strongly upregulated in microglial cells of retinoschisin-deficient (RS1-/y) mice, a model of inherited retinal degeneration and in a murine light damage model. A co-localization of TSPO and microglia was furthermore detectable in human retinal sections, indicating a potential role for TSPO as a biomarker for retinal degenerations. In vitro assays showed that the TSPO ligand XBD173 effectively inhibited features of microglial activation such as morphological transformation into reactive phagocytes and enhanced expression of pro-inflammatory cytokines. XBD173 also reduced microglial migration and proliferation and reduced their neurotoxic potential on photoreceptor cells. In two independent mouse models of light-induced retinal degeneration, the treatment with XBD173 reduced accumulation of amoeboid, reactive microglia in the outer retina and attenuated degenerative processes, indicated by a nearly preserved photoreceptor layer. A further question addressed in this thesis was whether minocycline, an antibiotic with additional anti-inflammatory properties is able to reduce microglial neurotoxicity and to protect the retina from degeneration. Minocycline administration dampened microglial pro-inflammatory gene expression, NO production and neurotoxicity on photoreceptors. Interestingly, in addition to its immunomodulatory effect, minocycline also increased the viability of photoreceptors in a direct manner. In the light damage model, minocycline administration counter-acted microglial activation and blocked retinal degeneration. Taken together these results identified TSPO as a biomarker for microglial reactivity and as therapeutic target in the retina. Targeting TSPO with XBD173 was able to reverse microglial reactivity and could prevent degenerative processes in the retina. In addition, the study showed that the antibiotic minocycline effectively counter-regulates microgliosis and light-induced retinal degeneration. Considering that microgliosis is a major contributing factor for retinal degenerative disorders, this thesis supports the concept of a microglia-directed therapy to treat retinal degeneration.

Evaluación de desenlace: trasplante meniscal versus segunda meniscectomía

Introducción: El incremento de pacientes sintomáticos de rodilla y la osteoartrosis en jóvenes con limitadas posibilidades terapéuticas después de una meniscectomía, genera la búsqueda de alternativas terapéuticas. A pesar que es poco utilizado en Colombia, el trasplante meniscal es una propuesta para el manejo sintomático. Según cifras norteamericanas, se practican entre 700.000 a 1.500.000 artroscopias de rodilla anualmente, el 50% termina en meniscectomía y de este un 40% persisten sintomáticos. Métodos: Estudio de cohorte retrospectivo, con el objetivo de evaluar dolor (Escala Visual Análoga-EVA) y funcionalidad (Escala de Tegner y Lysholm) en los pacientes a quienes se les realizó trasplante meniscal o meniscectomía por segunda vez, entre los años 2007 a 2015. Resultados: A partir de los 6 meses la EVA mostró una tendencia a la mejoría en el grupo de trasplante meniscal, pasando de Moderado a Leve (p: <0.000). La Escala de Tegner y Lysholm cambió de Pobre a Bueno en el grupo de segunda meniscectomía (p= 0.008) y de Bueno a Excelente en el grupo trasplantado (p=0.225). La calificación promedio de la EVA en el grupo de trasplante presentó mejoría (p=<0.000), a diferencia del grupo de segunda meniscectomía (p=0.591). La escala de Tegner y Lysholm, mostró significancia estadística con tendencia a la mejoría en el grupo de segunda meniscectomía. Discusión: Los resultados muestran que con trasplante meniscal hay mejoría del dolor y la funcionalidad versus un segunda meniscectomía. Para fortalecer la evidencia de este tratamiento son necesarios estudios prospectivos complementarios.

Factores asociados a transfusión sanguínea en el manejo quirúrgico del canal lumbar estrecho en la Fundación Santa Fe de Bogotá 2003 - 2013

Introduccion: El canal lumbar estrecho es un motivo de consulta frecuente en el servicio de columna de la Fundación Santa Fe de Bogotá. Derivado del tratamiento quirurgico se pueden generar múltiples complicaciones, entre las que se encuentra la transfusión sanguínea. Objetivo: Identificar los factores sociodemográficos, antecedentes personales y factores quirúrgicos asociados a transfusión sanguínea en cirugía canal lumbar estrecho en la Fundación Santa Fe de Bogotá 2003- 2013. Materiales y métodos: Se aplicó en diseño de estudio observacional analítico transversal. Se incluyeron 367 pacientes sometidos a cirugía de canal lumbar estrecho a quienes se les analizaron variables de antecedentes personales, características sociodemograficas y factores quirúrgicos. Resultados: La mediana de la edad fue de 57 años y la mayoría de pacientes fueron mujeres (55,6%). La mediana del Índice de Masa Corporal (IMC) fue de 24,9 clasificado como normal. Entre los antecedentes patológicos, la hipertensión arterial fue el más común (37,3%). La mayoría de pacientes (59,1%) presentaron clasificación ASA de II. El tipo de cirugía más prevalente fue el de descompresión (55,6%). En el 79,8% de los pacientes se intervinieron 2 niveles. Se realizó transfusión de glóbulos rojos en 26 pacientes correspondiente a 7,1% del total. En la mayoría de procedimientos quirúrgicos (42,5%) el sangrado fue clasificado como moderado (50-500 ml). En el modelo explicativo transfusión sanguínea en cirugía de canal lumbar estrecho se incluyen: antecedente de cardiopatía (OR 4,68, P 0,034, IC 1,12 – 19,44), Sangrado intraoperatorio >500ml (OR 6,74, p 0,001, 2,09 – 21,74) y >2 niveles intervenidos (OR 3,97, p 0,023, IC 1,20 – 13,09). Conclusión: Como factores asociados a la transfusión sanguínea en el manejo quirúrgico del canal lumbar estrecho a partir de la experiencia de 10 años en la Fundación Santa Fe de Bogotá se encontraron: enfermedad cardiaca, sangrado intraoperatorio mayor de 500ml y más de dos niveles intervenidos.

Methodology of Care Humanitude in Promoting Self-care in Dependent People: An Integrative Review

Abstract. The aging of the population has led to an increase in the prevalence of chronic degenerative diseases and dependence. We need to implement humanized health care that will improve the quality of life and well-being of these people and help maintain their autonomy and self-care. Objective: To identify the implications of the caring in Humanitude in promoting self-care in the dependent person. Methods: Integrative review of the literature of the period between 2007 and 2015, using the databases Medline, EBSCO and Google Scholar. In using the PI[C]OD methodology and criteria for inclusion and exclusion, we obtained 54 items where 7 were selected for analysis. Results: There are several health benefits in the promotion of self-care, by ap-plying the Humanitude caring philosophy, mainly regarding the relationship be-tween the nurse and the patient. Conclusions: It is essential to develop further studies focused on the implications of caring in Humanitude in self-care in the dependent person.

Clínica e cirurgia de equinos

O presente relatório de estágio, elaborado como etapa final do curso de Mestrado Integrado de Medicina Veterinária na Universidade de Évora, refere-se ao estágio curricular realizado na Clínica Equicare – Serviços Médico-Veterinários Lda., em Coimbra, Portugal, no âmbito da clínica e cirurgia de equinos. Este encontra-se dividido em três partes, onde, numa primeira fase se realiza uma descrição da casuística observada durante os seis meses de estágio, inseridas nas áreas de controlo e clínica reprodutiva, medicina preventiva e identificação equina, clínica médica e clínica cirúrgica. De seguida, realiza-se uma monografia sobre a doença degenerativa articular da extremidade distal de equinos e, por fim, uma apresentação de quatro casos clínicos onde se detetava a presença da mesma; EQUINE CLINICS AND SURGERY ABSTRACT: The present report was carried out as the final step of the Master of Science degree in Veterinary Medicine at the University of Évora and refers to the curricular externship held at the Equicare Clinic in Coimbra, Portugal, performed in Equine Clinics and Surgery. The report is divided in three parts, where initially takes place a casuistic analysis of the followed activities and clinical cases during the six month period of the externship, set in the areas of reproductive clinics, preventive medicine and equine identification, internal medicine and surgery. Afterwords a literature review about Degenerative Joint Disease of the distal limb in horses is performed and, finally, four clinical cases, about this same disease, are presented.

Antioxidant activity and cholinesterase inhibition studies if four flavoring herb from Alentejo.

In Alzheimer’s disease, the most common form of dementia, the loss of cholinergic neurons leads to the progressive reduction of acetylcholine in the brain, resulting cognitive impairment. Inhibition of the hydrolysis of acetylcholine by blocking acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) has been considered as a potential target in the treatment of Alzheimer’s disease. Essential oils and extracts of aromatic plants may have an important role in the oxidative stress protection. Traditionally, in Alentejo (Portugal), aromatic herbs Calamintha nepeta, Foeniculum vulgare, Mentha spicata and Thymus mastichina are often used by local population as condiments in food preparations. In this study, essential oils (EOs) and aqueous extracts (decoction waters) of these flavouring herbs were selected in order to evaluate its antioxidant potential and ability to inhibit AChE and BChE activities. Results suggest the potential use of EOs and extracts as nutraceutical or pharmaceutical preparations in the prevention of the oxidative stress and degenerative diseases.

Self-care in older people with functional deficit: needs of long-term care.

The group of 65-year-olds is becoming more numerous and with greater needs for health care. So, is necessary the reflection about new models of provision, organization, and allocation of health resources. According to the United Nations Organization, 2015, in 2050 elderly people will reach two million people (20% of the world’s population), what mean that the number of people over 60 years old will exceed a population of young people under 15 years. Parallel to aging, less healthy lifestyles have contributed to the prevalence of chronic diseases, especially cerebrovascular diseases. Hypertension and diabetes mellitus are risk factors and increase predisposition to other diseases. With aging, there is an increased risk for developing chronic, oncological and degenerative diseases, which account for more than 50% of the burden of diseases, with profound implications on independency, use of health care and services.

Antioxidant activity of jacaranda decurrens Cham.

Many diseases and related degenerative processes including heart, câncer and Parkinson diseases are associated with rcactive oxygen species (ROS). In an effort Io preveni diseases, search for com-pounds with antioxidant activity hás been a major interestof dif-ferent research groups in natural products research. The objective of this research was to investigatethe antioxidant activity of EtOH leaf extract ofjacarando decurrrens and its fractions using the 2,2-diphenyl-2-picrylhydrazyl hydrate (DPPH) assay [lJ, that uses s pé ciro me t ric method to determine radical scavenging activity, Rutinattheconcçntrationof l.Omg-L was used as the standard. Our results ha vê shown lhat crude extract and fractions had anti-oxidant activities mainly if tested at concentrations of 5.0 to 10.0mg-L. such activities, however. were lesser or equal to the standard (Table 1). The triterpenes, ursolic and oleanohc acids, were detected in the crude extract, Jd-1 and Jd-2, Possibly, these triterpenes are the active constituents responsible for the antioxi-dant activity |2j. At lower concentrations (0,6mg-L than the standard. Jd-3 fraction was the most active. The presenceof flavo-noids and glycosilated compounds were detected in jcl-3 fraction (2], This is the first attempt to demonstrate antioxidant activities found i n Já cara n (ia decurrens leaf extracts. Acknowledgements. Re-search funded by FAPESP. The State of São Paulo Research Founda-tion. Brazil, Referentes: 11) Koleva I. et ai. (2002). Phytochem Anal. 13: 8-17, (2! Oh CJ, et ai. (2007)- Free Radie Rés. 41(6): 638-44.