Sensory adaptation to blur improves visual acuity under defocused conditions. This phenomenon has been successfully demonstrated using subjective measures of acuity and is known as blur adaptation. This study investigates aspects of the mechanism of blur adaptation in human vision using subjective and objective methods. Parafoveal visual acuity measured under defocused conditions demonstrates that blur adaptation is not limited to the fovea. The presence of the adaptive mechanism in the parafovea also suggests that the neural compensation that takes places under defocused conditions acts across a spatial range and is not limited to specific frequency bands. An evaluation of the contrast sensitivity function under defocus provides further evidence. Electrophysiological methods measure the effect of blur adaptation at the retina and at the visual cortex to provide objective evidence for the presence of the blur adaptation mechanism. Finally enhanced-depth imaging optical coherence tomography examines whether a period of prolonged defocus triggers any short-term changes in choroidal thickness in a similar manner to that reported in animal emmetropisation. / Engineering and Physical Sciences Research Council

After over a decade of its discovery, the Electroretinogram (ERG) still remains the objective tool that is conventionally used in assessment of retinal function in health and disease. Although there is ongoing research in developing ERG recording techniques, interpretation and clinical applications, there is still a limited understanding on how each photoreceptor class contribute to the ERG waveform and their role and/or susceptibilities in various retinal diseases still remains unclear. Another limitation with currently used conventional testing protocols in a clinical setting is the requirement of an adaptation period which is time consuming. Furthermore, the ERG responses derived in this manner are recorded under different stimulus conditions, thus, making comparison of these signals difficult. To address these issues and develop a new testing method, we employed silent substitution paradigm in obtaining cone- and rod- isolating ERGs using sine- and square- wave temporal profiles. The ERGs achieved in this manner were shown to be photoreceptor-selective. Furthermore, these responses did not only provide the functional index of photoreceptors but their contributions to their successive postreceptoral pathways. We believe that the substitution stimuli used in this thesis could be a valuable tool in functional assessment of individual photoreceptor classes in normal and pathological conditions. Furthermore, we speculate that this method of cone/rod activity isolation could possibly be used in developing faster and efficient photoreceptor-selective testing protocols without the need of adaptation. / Bradford School of Optometry and Vision Sciences scholarship

Central to ocular health is the vitreous body, a complex, gelatinous tissue filling the space between the lens and retina. It is a natural polymeric hydrogel whose delicate architecture of collagen and hyaluronic acid loses its mechanical structure under the influence of degeneration or destruction, leaving the retina vulnerable to injury and disease. Since World War II, combat ocular trauma has increased six-fold while the population of aging veterans continues to grow in tandem. Compared to injuries in the civilian sector, injuries in theaters of combat operations are sustained in dirty, dusty, high-stress environments under hostile fire. These penetrating and perforating ocular injuries have predicable consequences, cascading into scarring on or under the retina (known as proliferative vitreoretinopathy or PVR). The growing population of aging veterans also faces a multitude of vitreous-related and vision-threatening pathologies. Current standards of care call for the removal and replacement of the vitreous, but contemporary substitutes are ill suited for long-term use. As such, there is critical need for development of a successful, long-term vitreous substitute. A biomimetic in situ forming hydrogel has been developed that utilizes a reversible disulfide cross-linker, enabling easy injection into the vitreous cavity. Recently, a copolymer has been introduced with this new formulation that possesses a unique comb-like structure whose characteristic bristles inhibit protein adsorption and cellular adherence, perfectly suited for the inhibition of PVR. The objective of this dissertation was to evaluate select formulations of this unique in situ forming hydrogel as potential vitreous substitutes. This was accomplished through rigorous in vivo rabbit modeled testing of long-term biocompatibility and bioperformance utilizing electroretinography, clinical examination, and histopathological assessment. We hypothesized that the in situ forming hydrogels would serve as a substantial improvement over the current gold standard, silicone oil, in terms of biocompatibility and the ability to inhibit PVR. / Bioengineering

Age-related maculopathy (ARM) is a central retinal disease with unclear pathogenesis. It is the major cause of permanent vision loss in adults over 50 years and is increasing in prevalence and incidence, faster than the aging population would suggest. Early in the disease process (early ARM) there is little or no vision loss and there are only slight retinal changes with abnormal deposits within Bruch's membrane. As the disease progresses (late ARM or age-related macular degeneration, AMD) vision loss may be quite severe due to atrophy (dry AMD) or the development of chorioretinal neovascularisation (CNV, wet AMD). It is hard to predict from conventional eye examinations and clinical vision tests which cases will progress to the severe, dry or wet forms of the disease. Moreover, most of the conventional clinical tests are based upon subjective vision measures. Objective tests which detect ARM earlier would be a useful aid to diagnosis and to monitoring progression. The multifocal electroretinogram (mfERG) is a relatively new clinical tool which enables the recording of electrical potentials from multiple, small areas of the central retina and thus assesses function from specific retinal locations. It is therefore useful in detecting focal retinal diseases such as hereditary or acquired maculopathies or in monitoring retinal laser or surgical treatment effects. There is cone and rod impairment in ARM and histopathological and psychophysical evidence for a preferential vulnerability of rods compared to cones. This research project investigated if an objective tool such as the mfERG could detect early ARM,its progression and the treatment effects of multiple photodynamic therapies (PDT) on retinal function in late ARM, prior to a battery of subjective vision measures. For comparison purposes a subjective assessment of central retinal function was performed using high and low contrast distance visual acuities (VA), near VA, low luminance VA (SKILL cards), contrast sensitivity (Pelli-Robson, P-R), saturated and desaturated Panel D-15 (sat Panel D-15, desat Panel D-15) and central visual fields (Humphrey 10-2, mean sensitivity, MS and mean defects, MD). As an objective assessment of central retinal function the cone- and rod-mediated multifocal electroretinograms were recorded. Subjective and objective tests of retinal function were compared in early ARM and an age-matched control group (chapter 3). Seventeen eyes of seventeen subjects with early ARM and twenty control subjects with normal vision were measured. For the cone-mediated mfERG responses conventional averaging methods were used and results were correlated with subjective vision tests. The conventional cone-mediated mfERG failed to distinguish between the early ARM and control subjects whereas subjective vision measures such as HC- and LC-VA, desat Panel D-15, MS, P-R were significantly reduced in the ARM group. However, there were significant correlations between the cone-mediated mfERG and the desat Panel D-15 results in the ARM group. This suggests that the mfERG measures similar retinal processes that detect colour vision deficiency under desaturated conditions. There was no significant correlation between cone-mediated mfERG measures and funduscopic changes. The conclusion from this study was that the subjective vision tests detected early ARM better than the objective cone-mediated mfERG. Thus the aim of detecting early ARM objectively was not met by the cone-mediated mfERG suggesting the need to develop other objective tests such as a rod-mediated mfERG. Whether the preferential rod vulnerability others have reported in early ARM could be detected by the rod-mediated mfERG was determined in the next study (chapter 4). A protocol for recording rod-mediated mfERG responses was developed by determining the optimal testing luminance to reduce the effect of stray light and elicit maximal rod-mediated responses. Sixteen of the seventeen ARM subjects and seventeen control subjects from the previous study were tested. For analysis, a customized computer template fitting method was developed in MATLAB (Mathworks, Natick, MA, USA). This method has been shown to be useful for low signal-to-noise ratio responses that characterize the rod-mediated mfERG. Significantly delayed rod-mediated mfERG responses were found whereas cone-mediated mfERG responses were within the normal range. This suggested that the effect of ARM on the rod system could be detected objectively with the rod-mediated mfERG before changes in the cone-mediated mfERG. Which of the tests best detected progression of vision loss was investigated in chapter 5. Visual function of 26 (13 ARM and 13 control subjects) of the original 37 subjects (17 ARM and 20 control subjects) had cone- and rod-mediated mfERG and the subjective vision measures repeated after one year. The main purpose was to determine which of the tests best detected progression of vision loss. The mfERG results were analysed by using both averaged and local responses and by using the computer template fitting procedure. On average no significant worsening of either objective or subjective function measures was evident after one year. These results reinforce the slow progression of the disease. With a longer follow-up period progression of ARM may translate into measurable changes in the mfERG and the other visual function tests. The effect of multiple photodynamic therapies (PDT) on cone- and rod-mediated function was assessed with the mfERG in the last study (chapter 6). The cumulative treatment effects of PDT in five subjects with late ARM were determined. Having demonstrated that the rod-mediated mfERG was applicable in early ARM, this study also aimed to investigate how useful it was in late ARM where there is substantially greater rod loss. Cone- and rod-mediated mfERGs, visual acuities, contrast sensitivities and central visual fields were investigated a week before treatment began and then one month after each PDT treatment. The subjects received three treatments each over an average period of five and a half months. In some subjects there were significant transient reductions in cone- and rod-mediated amplitudes possibly reflecting alterations in choroidal hypoperfusion dynamics one month after treatment. Further, b-wave component of the mfERG became increasingly misshapen after each PDT treatment suggesting an ischemic insult mainly targeting post-receptoral sites. However, objective and subjective function was stabilized after multiple PDT treatments in most of the subjects. This pilot study of five cases showed that there was no additional damage to cone- and rod-mediated outer retinal function after three PDT treatments. One of the novel findings of this research was that the rod-mediated function measured with the mfERG was impaired in early ARM. This finding supports histopathological and psychophysical evidence of rod vulnerability in early ARM. The results of these studies also suggest that early ARM affects different aspects of visual function which is reflected by different outcomes from objective and subjective vision tests. A model (chapter 7) based upon the results was developed proposing a hypoxic insult with a preferential alteration of post-receptoral sites in early ARM. The cone-mediated mfERG documented the retinal damage and possible treatment effects on outer retinal function of the multiple PDTs which did not further deteriorate. Thus, this technique might assist in the development of optimal treatment modalities for ARM, especially in retreatment regimes. Greater variability was found for the rod-mediated mfERG and its clinical use in PDT treatment regimes still needs to be investigated. In conclusion, this research has provided a better understanding of the disease process and treatment effects in ARM and might contribute to improvements in diagnosis and treatment of ARM.

Background: Optical coherence tomography (OCT), multifocal electroretinography (mfERG) and full-field electroretinography (ffERG) give important information about retinal structure and function. Purpose: To collect normative data of macular Cirrus Spectral domain (SD)-OCT assessments and of mfERG measurements of healthy children (papers I and II). To assess the macular thickness with Cirrus SD-OCT and the retinal function with ffERG in 6.5-year-old children born extremely preterm and in children born at term (papers III and IV). Methods: Study participants aged 5-15 years and living in Uppsala County were randomly chosen from the Swedish Birth Register (papers I and II). In papers III and IV, the study participants consisted of children born extremely preterm and children born at term – all were aged 6.5 years. In paper III, the children were living in Stockholm and Uppsala health care regions and, in paper IV, in Uppsala health care region only. Macular thickness was assessed with Cirrus SD-OCT and macular function with mfERG, using the Espion Multifocal system and DTL-electrodes. The retinal function was assessed with ffERG and DTL-electrodes, using the Espion Ganzfield system. Results: Altogether, 58 children participated in paper I and 49 children in paper II. In paper I, the repeatability and reproducibility of the OCT assessments were good. In paper II, the results of the mfERG measurements were in accordance with retinal cone density and there were no significant differences between the right and left eyes. In paper III, 134 preterm children and 145 children born at term constituted the study population. The central macular thickness was significantly thicker in the preterm group than in the control group. Within the preterm group, gestational age (GA), former retinopathy of prematurity (ROP) and male gender were all important risk factors for an increased macular thickness. In paper IV, 52 preterm children and 45 control children constituted the study population. Significantly lower amplitudes and prolonged implicit times of the combined rod and cone responses, as well as of the isolated cone responses, were found in the preterm group when compared with the control group. In paper IV, there was no association between GA, ROP or male gender and the ffERG assessments. Conclusion: Normative data of Cirrus SD-OCT and mfERG assessments were reported. The results of the assessments were reliable. Children aged 6.5 years, born extremely preterm, had a significantly thicker central macula and both rod and cone function were significantly reduced in comparison to children born at term. ROP had an influence on retinal structure but not retinal function in the present cohorts. Our results suggest that retinal development is abnormal in children born extremely preterm. Long-term follow-up studies are necessary in order to evaluate the functional ophthalmological outcome in this vulnerable population of children growing up today.

Uveítes são inflamações intraoculares geralmente crônicas e uma das principais causas de cegueira no mundo. Os corticosteroides são as drogas de primeira escolha para o tratamento das uveítes autoimunes, mas, muitas vezes, há necessidade do uso de outras drogas imunossupressoras, como o micofenolato de mofetila, uma pró-droga, que é rapidamente convertida em ácido micofenólico (MPA) e tem se mostrado eficaz no tratamento de uveíte autoimune, reduzindo e estabilizando a inflamação. Contudo, tanto os corticosteroides quanto os imunossupressores estão associados aos efeitos colaterais graves que podem levar à necessidade de diminuição da dose ou à suspensão da droga. Assim, como uma alternativa às vias tradicionais (sistêmica e tópica) e visando diminuir os efeitos colaterais, este trabalho tem como objetivos 1) verificar a viabilidade do implante intravítreo de liberação lenta de MPA em coelhos sadios, 2) reproduzir o modelo de uveíte autoimune experimental (UAE) em coelhos através da injeção intravítrea de M. tuberculosis e 3) avaliar os efeitos terapêuticos do implante biodegradável de MPA em modelos de uveíte autoimune experimental (UAE) através de análises longitudinais clínicas e eletrorretinográficas e, após 50 dias, de análises morfológicas. Nossos resultados mostraram a viabilidade do implante intravítreo de MPA e indicaram que o modelo de UAE desenvolveu uma uveíte anterior autolimitada, com pico de inflamação em 21 dias indicado pelo aumento de células na câmara anterior, flare e hiperemia conjuntival, assim como pela diminuição das amplitudes e aumento dos picos de latências. Com relação ao estudo do efeito terapêutico no modelo de UAE, os exames clínicos demonstraram que o implante de MPA foi capaz de diminuir o grau de uveíte durante o pico da inflamação, reduzindo a hiperemia conjuntival, as células presentes na câmara anterior e o flare. Porém, após 35 dias de tratamento, foi observado o descolamento da retina em 60% dos animais. Os resultados eletrorretinográficos também apontaram uma redução da inflamação através da recuperação das amplitudes das ondas-b escotópicas e dos picos de tempo implícito das respostas fotópicas tanto da onda-a quanto da onda-b nos animais tratados. Após 50 dias de tratamento, a imunohistoquímica revelou que o grupo tratado apresentava menos ativação das células da glia, indicando uma diminuição do processo inflamatório. A partir desses resultados, podemos concluir que o implante intravítreo de liberação lenta de MPA apresentou um efeito terapêutico no pico da inflamação, indicando uma diminuição do grau da doença e uma recuperação dos fotorreceptores, das células bipolares ON e das células de Müller, as quais foram alteradas com a inflamação. Nossos resultados também sugerem que o efeito terapêutico de MPA ocorre quando a concentração da droga se aproxima de 700 ng/mL no vítreo, pois a diminuição da inflamação ocular coincidiu com o aumento da taxa de liberação do MPA no vítreo / Chronic uveitis are generally intraocular inflammations and the major cause of blindness in the world. The corticosteroids are the drugs of first choice for the treatment of autoimmune uveitis, but is often need for use of other immunosupressive drugs. However, both immunossuppressants ans corticosteroids are associated with serious side effects that may lead to the need for dose reduction or dicontinuation of the drug. As an alternative to traditional drug delivery (systemic and topical) and to reduce the side effects, this study aims 1) to verify the viability of intravitreal MPA implant in the control rabbits, 2) to reproduce the experimental autoimmune uveitis model (EAU) induced with M. tuberculosis and 3) to evaluate the therapeutic effects of MPA biodegradable implant in EAU rabbits through ERG and clinical longitudinal analysis and morphological analyzes after 50 days. The results confirmed the viability and tolerance of the MPA intravitreal implant. The EAU model developed a self-limiting anterior uveitis, an inflammation peak at 21 days indicated by the increase in the anterior chamber cells, flare and conjunctival hyperemia. The ERG showed that this model present a decrease of the amplitudes and an increase of the implicit times. Our clinical results showed that the MPA implant decreased the uveitis degree during the peak of inflammation reducing conjunctival hyperemia, the presence of cells in the anterior chamber and the presence of flare. However, after 35 days of treatment, we observed retinal detachment in 60% of animals. Electroretinography results also showed a reduction of inflammation through the recovery of the scotopic b-waves amplitude and implicit time of the photopic responses of both a- and b-waves for the treated group. After 50 days of treatment, immunohistochemistry revealed that the activation of glial cells had become smaller without, indicating a reduction of the inflammatory process. With these results, we concluded that MPA intravitreal implant supressed the inflammation during the peak of inflammationin a rabbit model of uveitis. And besides reducing the clinical signs of disease, the drug recovered the photoreceptors, ON bipolar and Müller cells functions that have been altered by inflammation. Our results suggest thatthe therapeutic effect of MPA occurs when the drug concentration approaches 700 ng / mL in the vitreous

Uveítes são inflamações intra-oculares geralmente crônicas e constituem uma das principais causas de cegueira no mundo. Os corticosteroides são a droga de primeira escolha para o tratamento das uveítes não infecciosas, mas muitas vezes há necessidade do uso de outras drogas imunossupressoras. O micofenolato de mofetila (MMF) é um potente imunossupressor administrado por via oral que vem sendo utilizado com sucesso no tratamento das uveítes, mas cujos efeitos colaterais muitas vezes tornam necessária sua suspensão. O MMF é uma pró-droga, que é transformada no fígado em ácido micofenólico (MPA), o imunossupressor ativo. Para minimizar os efeitos colaterais do uso do MPA e permitir que o olho receba uma dose maior da droga, testamos os efeitos da injeção intravítrea do MPA em um modelo de uveíte crônica experimental (UCE) em olhos de coelhos. Os objetivos deste estudo foram: 1) reproduzir um modelo de UCE em coelhos através da injeção intravítrea de M. tuberculosis; 2) estabelecer uma dose segura de MPA a ser injetada no vítreo; e 3) analisar os efeitos morfológicos, clínicos e eletrofisiológicos da injeção intravítrea de MPA em coelhos utilizados como modelo de UCE. O modelo de UCE reproduzido apresentou uma inflamação autolimitada, possuindo um pico de inflamação no 17° dia após a indução da uveíte. As doses de MPA testadas (0,1 e 1mg) não foram toxicas para a retina do coelho. O modelo de UCE recebeu uma injeção intravítrea de 0,1mg de MPA e as análises clinicas demonstraram uma redução na inflamação. As análises realizadas com o eletrorretinograma (ERG) também apontaram uma melhora na inflamação através da recuperação da latência das ondas-a e b (fotópicas e escotópica) e recuperação da amplitude da onda-a (fotópica). As análises morfológicas com HE não apresentaram alterações na estrutura retinia, porem a imunohistoquimica para proteína GFAP evidenciou gliose das células de Müller, sinalizando um processo inflamatório. Concluímos que o modelo de UCE reproduziu uma uveíte anterior semelhante à uveíte causada em humanos e a dose de MPA utilizada apresentou efeitos terapêuticos durante o pico de inflamação, mostrando uma diminuição da inflamação e promovendo a recuperação de fotorreceptores e células bipolares-ON. Este resultado faz das injeções intravítreas de MPA um recurso promissor no tratamento de uveítes. Porém, novos experimentos são necessários para padronizar os resultados encontrados / Uveitis is an intraocular inflammation usually chronic and a major cause of blindness in the world. Corticosteroids are the first choice drug for treatment of non-infectious uveitis, but other immunosuppressive drugs are usually required. Mycophenolate mofetil is a powerful immunosuppressant orally administered that has been used successfully in the treatment of uveitis, but whose side effects often lead to the suspension of the drug intake. The Mycophenolate mofetil is a prodrug, which isconverted in the liver to mycophenolic acid (MPA), the active immunosuppressant. To minimize side effects of the use of mycophenolic acid and to enable a higher administration dose in the eye, we tested the effects of the intravitreal injection of mycophenolic acid in an experimental model of chronic uveitis (ECU) in rabbit eyes. The objectives of this study were: 1) to reproduce an ECU model in rabbits by intravitreal injection of M. tuberculosis; 2) to establish a safe dose of mycophenolic acid to be injected into the vitreous; and 3) to analyze the morphological, clinical and electrophysiological effects of the intravitreal injection of mycophenolic acid in rabbits used as a model of ECU. The ECU model reproduced showed a self-limiting inflammation, having a peak of inflammation at 17 days after uveitis induction. Both MPA doses we tested (0.1 and 1 mg) were not toxic to the retina. The ECU model received an intravitreal injection of 0.1 mg of MPA and clinical analysis demonstrated a reduction of the inflammation. The electroretinography (ERG) analysis also indicated an improvement in the inflammation process by restoring the latency of the a-wave and b-wave (photopic and scotopic) and by the recovery of the a-wave amplitude (photopic). The morphological analysis with HE showed no changes in the retinal structure, however the immunohistochemistry for GFAP protein showed gliosis of Müller cells, indicating an inflammatory process. We conclude that the ECU model reproduced an anterior uveitis similar to the human uveitis and the MPA dose we used showed therapeutic effects during the inflammation peak, reducing the inflammation and promoting the recovery of photoreceptors and ON bipolar cells. This makes intravitreal injections of MPA a promising resource in the treatment of uveitis. Future studies are necessary to standardize the present findings

A high frequency of retinitis pigmentosa (RP) is found in Northern Sweden. In an inventory of autosomal recessive RP patients in Västerbotten County, a great number of cases with a unique phenotype was noticed, denoted Bothnia Dystrophy (BD). The aim of the study was to describe the phenotype, to determine the chromosomal location, and to identify the gene. Patients typically show night blindness from early childhood. Symptoms of defect macular function with a decrease of visual acuity can appear in early adulthood. The retinal fundus shows irregular white spots in a central, and parafoveal pattern along the arcades. Centrally areolar maculopathy develops and round circular atrophies are observed in the periphery. The disease was shown to be associated with a missense mutation in the RLBP1 gene resulting in an amino acid substitution (R234W) in the cellular retinaldehyde-binding protein (CRALBP). The R234W mutation was found in a homozygous state in 61 patients affected with BD. Ten patients were heterozygous for the R234W mutation, and presented a similar phenotype. No additional mutations in the coding sequence or exon-intron junctions were found. CRALBP is localised in retinal pigment epithelium (RPE), and Müller cells of the retina. In the RPE, CRALBP functions as a carrier protein for endogenous retinoids. Dark adaptometry and electrophysiologic testing showed an initial loss of rod function followed by a progressive reduction of the cone responses in older ages. A compromised rod function, dysfunction of the Müller cells, and indications of a disturbed function of the inner retina were found. With prolonged dark adaptation, a gradual increase in retinal sensitivity to light and an improvement of the ERG components occurred. The findings indicate a prolonged synthesis of photopigments, retardation of the visual process in the retinal pigment epithelium and a loss of retinal cells probably starting at a relative early age in BD. To evaluate the subjective visual function in BD patients, a battery of objective tests of visual function and composite score of the 25-item NEI-Visual Function Questionnaire (VFQ-25) were analyzed. We found that weighted distance logMAR visual acuity (WVA), had the strongest association with subjective visual function, and that there was a considerable loss of subjective and objective visual function with increasing age in BD patients. The prevalence of BD is as high as 1:3600 in Västerbotten County. The possibility that recycling of retinoids localized in the RPE might be impaired in BD might give future therapeutic possibilities. Due to the large and clinically well-characterized set of patients with this disease, they constitute a suitable study group.

Uveítes são inflamações intra-oculares geralmente crônicas e constituem uma das principais causas de cegueira no mundo. Os corticosteroides são a droga de primeira escolha para o tratamento das uveítes não infecciosas, mas muitas vezes há necessidade do uso de outras drogas imunossupressoras. O micofenolato de mofetila (MMF) é um potente imunossupressor administrado por via oral que vem sendo utilizado com sucesso no tratamento das uveítes, mas cujos efeitos colaterais muitas vezes tornam necessária sua suspensão. O MMF é uma pró-droga, que é transformada no fígado em ácido micofenólico (MPA), o imunossupressor ativo. Para minimizar os efeitos colaterais do uso do MPA e permitir que o olho receba uma dose maior da droga, testamos os efeitos da injeção intravítrea do MPA em um modelo de uveíte crônica experimental (UCE) em olhos de coelhos. Os objetivos deste estudo foram: 1) reproduzir um modelo de UCE em coelhos através da injeção intravítrea de M. tuberculosis; 2) estabelecer uma dose segura de MPA a ser injetada no vítreo; e 3) analisar os efeitos morfológicos, clínicos e eletrofisiológicos da injeção intravítrea de MPA em coelhos utilizados como modelo de UCE. O modelo de UCE reproduzido apresentou uma inflamação autolimitada, possuindo um pico de inflamação no 17° dia após a indução da uveíte. As doses de MPA testadas (0,1 e 1mg) não foram toxicas para a retina do coelho. O modelo de UCE recebeu uma injeção intravítrea de 0,1mg de MPA e as análises clinicas demonstraram uma redução na inflamação. As análises realizadas com o eletrorretinograma (ERG) também apontaram uma melhora na inflamação através da recuperação da latência das ondas-a e b (fotópicas e escotópica) e recuperação da amplitude da onda-a (fotópica). As análises morfológicas com HE não apresentaram alterações na estrutura retinia, porem a imunohistoquimica para proteína GFAP evidenciou gliose das células de Müller, sinalizando um processo inflamatório. Concluímos que o modelo de UCE reproduziu uma uveíte anterior semelhante à uveíte causada em humanos e a dose de MPA utilizada apresentou efeitos terapêuticos durante o pico de inflamação, mostrando uma diminuição da inflamação e promovendo a recuperação de fotorreceptores e células bipolares-ON. Este resultado faz das injeções intravítreas de MPA um recurso promissor no tratamento de uveítes. Porém, novos experimentos são necessários para padronizar os resultados encontrados / Uveitis is an intraocular inflammation usually chronic and a major cause of blindness in the world. Corticosteroids are the first choice drug for treatment of non-infectious uveitis, but other immunosuppressive drugs are usually required. Mycophenolate mofetil is a powerful immunosuppressant orally administered that has been used successfully in the treatment of uveitis, but whose side effects often lead to the suspension of the drug intake. The Mycophenolate mofetil is a prodrug, which isconverted in the liver to mycophenolic acid (MPA), the active immunosuppressant. To minimize side effects of the use of mycophenolic acid and to enable a higher administration dose in the eye, we tested the effects of the intravitreal injection of mycophenolic acid in an experimental model of chronic uveitis (ECU) in rabbit eyes. The objectives of this study were: 1) to reproduce an ECU model in rabbits by intravitreal injection of M. tuberculosis; 2) to establish a safe dose of mycophenolic acid to be injected into the vitreous; and 3) to analyze the morphological, clinical and electrophysiological effects of the intravitreal injection of mycophenolic acid in rabbits used as a model of ECU. The ECU model reproduced showed a self-limiting inflammation, having a peak of inflammation at 17 days after uveitis induction. Both MPA doses we tested (0.1 and 1 mg) were not toxic to the retina. The ECU model received an intravitreal injection of 0.1 mg of MPA and clinical analysis demonstrated a reduction of the inflammation. The electroretinography (ERG) analysis also indicated an improvement in the inflammation process by restoring the latency of the a-wave and b-wave (photopic and scotopic) and by the recovery of the a-wave amplitude (photopic). The morphological analysis with HE showed no changes in the retinal structure, however the immunohistochemistry for GFAP protein showed gliosis of Müller cells, indicating an inflammatory process. We conclude that the ECU model reproduced an anterior uveitis similar to the human uveitis and the MPA dose we used showed therapeutic effects during the inflammation peak, reducing the inflammation and promoting the recovery of photoreceptors and ON bipolar cells. This makes intravitreal injections of MPA a promising resource in the treatment of uveitis. Future studies are necessary to standardize the present findings

Uveítes são inflamações intraoculares geralmente crônicas e uma das principais causas de cegueira no mundo. Os corticosteroides são as drogas de primeira escolha para o tratamento das uveítes autoimunes, mas, muitas vezes, há necessidade do uso de outras drogas imunossupressoras, como o micofenolato de mofetila, uma pró-droga, que é rapidamente convertida em ácido micofenólico (MPA) e tem se mostrado eficaz no tratamento de uveíte autoimune, reduzindo e estabilizando a inflamação. Contudo, tanto os corticosteroides quanto os imunossupressores estão associados aos efeitos colaterais graves que podem levar à necessidade de diminuição da dose ou à suspensão da droga. Assim, como uma alternativa às vias tradicionais (sistêmica e tópica) e visando diminuir os efeitos colaterais, este trabalho tem como objetivos 1) verificar a viabilidade do implante intravítreo de liberação lenta de MPA em coelhos sadios, 2) reproduzir o modelo de uveíte autoimune experimental (UAE) em coelhos através da injeção intravítrea de M. tuberculosis e 3) avaliar os efeitos terapêuticos do implante biodegradável de MPA em modelos de uveíte autoimune experimental (UAE) através de análises longitudinais clínicas e eletrorretinográficas e, após 50 dias, de análises morfológicas. Nossos resultados mostraram a viabilidade do implante intravítreo de MPA e indicaram que o modelo de UAE desenvolveu uma uveíte anterior autolimitada, com pico de inflamação em 21 dias indicado pelo aumento de células na câmara anterior, flare e hiperemia conjuntival, assim como pela diminuição das amplitudes e aumento dos picos de latências. Com relação ao estudo do efeito terapêutico no modelo de UAE, os exames clínicos demonstraram que o implante de MPA foi capaz de diminuir o grau de uveíte durante o pico da inflamação, reduzindo a hiperemia conjuntival, as células presentes na câmara anterior e o flare. Porém, após 35 dias de tratamento, foi observado o descolamento da retina em 60% dos animais. Os resultados eletrorretinográficos também apontaram uma redução da inflamação através da recuperação das amplitudes das ondas-b escotópicas e dos picos de tempo implícito das respostas fotópicas tanto da onda-a quanto da onda-b nos animais tratados. Após 50 dias de tratamento, a imunohistoquímica revelou que o grupo tratado apresentava menos ativação das células da glia, indicando uma diminuição do processo inflamatório. A partir desses resultados, podemos concluir que o implante intravítreo de liberação lenta de MPA apresentou um efeito terapêutico no pico da inflamação, indicando uma diminuição do grau da doença e uma recuperação dos fotorreceptores, das células bipolares ON e das células de Müller, as quais foram alteradas com a inflamação. Nossos resultados também sugerem que o efeito terapêutico de MPA ocorre quando a concentração da droga se aproxima de 700 ng/mL no vítreo, pois a diminuição da inflamação ocular coincidiu com o aumento da taxa de liberação do MPA no vítreo / Chronic uveitis are generally intraocular inflammations and the major cause of blindness in the world. The corticosteroids are the drugs of first choice for the treatment of autoimmune uveitis, but is often need for use of other immunosupressive drugs. However, both immunossuppressants ans corticosteroids are associated with serious side effects that may lead to the need for dose reduction or dicontinuation of the drug. As an alternative to traditional drug delivery (systemic and topical) and to reduce the side effects, this study aims 1) to verify the viability of intravitreal MPA implant in the control rabbits, 2) to reproduce the experimental autoimmune uveitis model (EAU) induced with M. tuberculosis and 3) to evaluate the therapeutic effects of MPA biodegradable implant in EAU rabbits through ERG and clinical longitudinal analysis and morphological analyzes after 50 days. The results confirmed the viability and tolerance of the MPA intravitreal implant. The EAU model developed a self-limiting anterior uveitis, an inflammation peak at 21 days indicated by the increase in the anterior chamber cells, flare and conjunctival hyperemia. The ERG showed that this model present a decrease of the amplitudes and an increase of the implicit times. Our clinical results showed that the MPA implant decreased the uveitis degree during the peak of inflammation reducing conjunctival hyperemia, the presence of cells in the anterior chamber and the presence of flare. However, after 35 days of treatment, we observed retinal detachment in 60% of animals. Electroretinography results also showed a reduction of inflammation through the recovery of the scotopic b-waves amplitude and implicit time of the photopic responses of both a- and b-waves for the treated group. After 50 days of treatment, immunohistochemistry revealed that the activation of glial cells had become smaller without, indicating a reduction of the inflammatory process. With these results, we concluded that MPA intravitreal implant supressed the inflammation during the peak of inflammationin a rabbit model of uveitis. And besides reducing the clinical signs of disease, the drug recovered the photoreceptors, ON bipolar and Müller cells functions that have been altered by inflammation. Our results suggest thatthe therapeutic effect of MPA occurs when the drug concentration approaches 700 ng / mL in the vitreous