Chromosomal alterations involving the RUNXI or CBFB genes are specifically and recurrently associated with human acute myeloid leukemia (AML). One such chromosomal alteration, a pericentric inversion of chromosome 16, is present in the majority of cases of the AML subtype M4Eo. This inversion joins CBFB with the smooth muscle myosin gene MYH11 creating the fusion CBFB-MYH11. Knock-in studies in the mouse have demonstrated that expression of the protein product of the Cbfb-MYH11fusion, Cbfβ-SMMHC, predisposes mice to AML and that chemical mutagenesis both accelerates and increases the penetrance of the disease (Castilla et al., 1999). However, the mechanism of transformation and the associated collaborating genetic events remain to be resolved. As detailed in Chapter 2, we used retroviral insertional mutagenesis (RIM) to identify mutations in Cbfb-MYH11 chimeric mice that contribute to AML. The genetic screen identified 54 independent candidate cooperating genes including 6 common insertion sites: Plag1, Plagl2, Runx2, H2T23, Pstpip2, and Dok1. Focusing on the 2 members of the Plag family of transcription factors, Chapter 3 presents experiments demonstrating that Plag1 and Plagl2 independently cooperate with Cbfβ-SMMHC in vivo to efficiently trigger leukemia with short latency in the mouse. In addition, Plag1 and PLAGL2 increased proliferation and in vitro cell renewal in Cbfβ-SMMHC hematopoietic progenitors. Furthemore, PLAG1 and PLAGL2 expression was increased in 20% of human AML samples suggesting that PLAG1 and PLAGL2 may also contribute to human AML. Interestingly, PLAGL2was preferentially increased in samples with chromosome 16 inversion, t(8;21), and t(15;17). To define the mechanism by which PLAGL2 contributes to leukemogenesis, Chapter 4 presents studies assessing the role of the Mp1 signaling cascade as a Plagl2 downstream pathway in leukemia development. Using microarray analysis we discovered that PLAGL2 induces the expression of Mp1 transcript in primary bone marrow cells that express Cbfβ-SMMHC and that this induction is maintained in leukemogenesis. We have also performed luciferase assays to confirm that the Mp1 proximal promoter can be directly bound and activated by PLAGL2. Furthermore, we demonstrate increased Mp1 expression leads to hypersensitivity to the Mp1 ligand thrombopoietin (TPO) in PLAGL2/Cbfβ-SMMHC leukemic cells. To test the functional relevance in leukemia formation, we performed a bone-marrow transplantation assay and demonstrate that overexpression of Mp1 is indeed sufficient to cooperate with Cbfβ-SMMHC in leukemia induction. This data reveals that PLAGL2 cooperates with Cbfβ-SMMHC at least in part by inducing the expression of the cytokine receptor Mp1. Thus, we have identified the Mp1 signal transduction pathway as a novel target for therapeutic intervention in AML.

INTRODUCÃO: A cirurgia de Fontan ou conexão cavo-pulmonar total é o último procedimento de uma estratégia estadiada, empregada no tratamento de cardiopatias congênitas complexas com ventrículo único anatômico ou funcional (também chamada de cardiopatia congênita com coração univentricular). Pacientes com um único ventrículo, situação incompatível com a vida, passaram a sobreviver até a idade adulta após a realização dessa cirurgia. A conexão direta da cavas com as artérias pulmonares, exclui do sistema circulatório o ventrículo subpulmonar, criando nesses indivíduos uma circulação do tipo univentricular. A falta do ventrículo subpulmonar e a subsequente ausência de fluxo sanguíneo pulsátil no pulmão, reduz o volume de enchimento do ventrículo único e consequentemente, o débito cardíaco. Mecanismos adaptativos do sistema periférico são desencadeados para garantir uma adequada redistribuição do fluxo sanguíneo para órgãos vitais, porém não são suficientes para garantir uma capacidade funcional adequada nesses indivíduos. Acreditamos que, uma análise detalhada dessas alterações e seus respectivos mecanismos adaptativos, possam contribuir na avaliação e entendimento dessa complexa fisiologia. A interferência nos componentes relacionados a baixa capacidade física, poderiam em teoria, modificar a história natural da doença nesses indivíduos, submetidos a cirurgia de Fontan. OBJETIVOS: 1. Comparar variáveis do sistema cardiovascular, pulmonar e músculo esquelético de pacientes com idade entre 12 e 30 anos, submetidos a cirurgia de Fontan com 5 anos ou mais de evolução pós-operatória com indivíduos saudáveis. 2. Correlacionar variáveis do sistema cardiovascular, pulmonar e músculo esquelético com a capacidade funcional no grupo de pacientes com cirurgia de Fontan. 3. Identificar as variáveis preditoras de baixa capacidade funcional nesta população. MÉTODOS: Estudo transversal, tipo caso-controle. Foram incluídos 30 pacientes no Grupo Fontan (GF) e 27 indivíduos saudáveis, que compuseram o grupo controle (GC). Os indivíduos foram submetidos à ressonância magnética cardiovascular, ecodopplercardiografia, teste de esforço cardiopulmonar, prova de função pulmonar completa, microneurografia direta no nervo fibular, pletismografia de oclusão venosa, dosagem plasmática de catecolaminas e peptídeo natriurético cerebral, teste de caminhada de seis minutos, espectroscopia de fósforo por ressonância magnética do quadríceps femoral, imagem musculoesquelética por ressonância magnética da musculatura da coxa e avaliação da qualidade de vida pelo questionário SF-36 (Short-Form Health Survey). Foram realizadas análises comparativas entre os grupos nos diferentes sistemas e posteriormente, testes para identificar os preditores de baixa capacidade funcional no GF. RESULTADOS: O consumo máximo de oxigênio (VO2 pico) no GF foi menor comparada ao GC, em valores absolutos, relativo ao peso corpóreo e em porcentagem do predito para sexo e idade. [1,65 (±0,54) vs 2,81 (±0,77) L/min p < 0,001]; [29,3 (±6,0) vs 41,5 (±9,2) mL/kg/min p < 0,001] e [70 (±14) vs 100% do predito (±20) p < 0,001] respectivamente. Os volumes e capacidades pulmonares foram significativamente menores no GF comparados ao GC, e demonstraram correlação positiva com o VO2 absoluto [capacidade vital forçada (CVF) r=0,836 p < 0,001; capacidade pulmonar total (CPT) r=0,730 p < 0,001 e capacidade de difusão do monóxido de carbono (DLCO) r=0,539 p=0,002], mas foram preditores de baixa capacidade funcional, a CVF (Constante= -0,306; B=0,393; IC=0,272-0,513 e p<0,001) e a DLCO (Constante= - 0,306; B=0,042; IC= 0,018-0,067; e p=0,002). O diâmetro da artéria pulmonar esquerda, também foi identificado como preditor de baixo VO2 (Constante=0,274; B=0,111; IC=0,061-0,161 e p < 0,001). E pela análise de periférica, a área seccional transversa da musculatura da coxa foi significativamente menor no GF, demonstrando ser mais um preditor de baixa capacidade funcional nesta população (Constante: 0,380; B=0,024; IC=0,018-0,030; p < 0,001). A atividade nervosa simpática muscular foi maior no GF [30 (±4) vs 22 (±3) disparos/min p < 0,001] e o fluxo sanguíneo muscular menor [1,59 (±0,3) vs 2,17 (±0,5) mL/min/100mL p < 0,001] comparados ao GC, porém estas variáveis não foram preditoras de baixa capacidade funcional. Em relação ao metabolismo oxidativo muscular, o GF apresentou menor amplitude de pico da fosfocreatina comparado ao GC [0,43 (0,41-0,45) vs 0,45 (0,42-0,50) p=0,023]. Porém essas alterações não se correlacionaram com o VO2 pico. CONCLUSÕES: Concluímos que a função pulmonar, o controle neurovascular e a capacidade funcional de pacientes com ventrículo único funcional, clinicamente estáveis, encontram-se comprometidas quando comparadas com indivíduos saudáveis. O menor diâmetro da artéria pulmonar esquerda, a capacidade vital forçada diminuída, a capacidade de difusão do monóxido de carbono comprometida e a área seccional transversa da musculatura da coxa reduzida foram os preditores de baixa capacidade funcional na população estudada / BACKGROUND: The Fontan operation or total cavo-pulmonary connection is the last procedure of a staged strategy, performed to treat complex congenital heart diseases in patients with a functional or anatomic single ventricle, also known as a univentricular heart. After the inception of the Fontan procedure, patients are surviving to adulthood due this remarkable technique. This operation, creates a direct connection of the superior and inferior vena cava with the pulmonary arteries, excluding the subpulmonary ventricle of the circulatory system an univentricular circulation. The lack of a subpulmonary ventricle and subsequent absence of pulsatile blood flow in the lungs, reduce the filling volume of the single ventricle and consequently the cardiac output. Although some adaptive mechanisms at the peripheral system are triggered to ensure an adequade blood flow to vital organs, they are not enough for an adequate functional capacity in these patients. A detailed analysis and evaluation of these changes and their respective mechanisms may contribute to understand this complex physiology. Hopefully, it might be possible to modify and improve the long term outcomes of these individuals. The aims of the study were: 1. To compare the variables of the cardiovascular, pulmonary and musculoskeletal systems in clinically stable Fontan patients with a control group. 2. To correlate the variables of the cardiovascular, pulmonary and skeletal muscle with the functional capacity in Fontan patients. 3. To identify predictors of low functional capacity in this population. METHODS: A prospective cross-sectional study of 30 FP of (20 +/- 6 years) and 10 (8-15) years of follow-up and 27 healthy controls (HC) (22 +/- 5years) was performed. They underwent cardiovascular magnetic resonance, echocardiography, cardiopulmonary exercise test, complete lung function, catecholamine and B-type natriuretic peptide (BNP) plasmatic levels, microneurography, venous occlusion plethismography, six-minute walk test, phosphorus magnetic resonance spectroscopy (31P MRS) and magnetic resonance imaging (MRI) of skeletal muscle and quality of life (QoL) using the Short Form Health Survey (SF36). Comparative analyzes of the different systems of two groups were done as well as tests to identify the predictors of low functional capacity in Fontan groups (FG). RESULTS: The maximal oxygen consumption (VO2) in the FG was lower compared to control group (CG), in absolute values, relative to body weight and percentage predicted for gender and age [1,65 (±0,54) vs 2,81 (±0,77) L/min p < 0,001]; [29,3 (±6,0) vs 41,5 (±9,2) mL/kg/min p < 0,001] e [70 (±14) vs 100% of the predicted value (±20) p < 0,001] respectively. Pulmonary volumes and capacities were also significantly lower in FG compared to CG, and demonstrated a positive correlation with absolute peak VO2 [Forced vital capacity (FVC) r=0,836 p < 0,001; total lung capacity (TLC) r=0,730 p < 0,001 and carbon monoxide diffusion capacity (DLCO) r=0,539 p=0,002]. But, the FVC and DLCO were predictors of reduced VO2 (Constant= -0,306; B=0,393; CI=0,272-0,513 e p < 0,001) and (Constant= -0,306; B=0,042; CI=0,018-0,067; e p=0,002) respectively. The diameter of the left pulmonary artery was also other of the predictors of low functional capacity (Constant=0,274; B=0,111; CI=0,061-0,161 e p < 0,001). Analyzing the volume of the thigh muscles, a significant difference between the groups was found, and the transversal sectional area of this muscle was a predictor of low functional capacity (Constant: 0,380; B=0,024; CI=0,018-0,030; p < 0,001). Muscle sympathetic nerve activity was higher in FG [30 (±4) vs 22 (±3) burst/min p < 0,001] and forearm blood flow was lower [1,59 (±0,3) vs 2,17 (±0,5) mL/min/100mL p < 0,001] compared with CG, however, these variables were not predictive of low functional capacity. Muscle oxidative metabolism showed difference in the intracellular pH value and the peak amplitude of phosphocreatine between the groups. The peak amplitude of phosphocreatine was lower in FG compared CG [0,43 (0,41-0,45) vs 0,45 (0,42-0,50) p=0,023] however, there were no correlation with the functional capacity. CONCLUSIONS: Pulmonary function, neurovascular control and functional capacity of clinically stable Fontan patients were impaired when compared to healthy subjects. Lower diameter of the left pulmonary artery, decreased forced vital capacity, impaired carbon monoxide diffusion capacity, and reduced transverse sectional area of the thigh musculature were the predictors of low functional capacity in the study population

Submitted in partial compliance with the requirements for the Master’s Degree in Technology: Chiropractic, Department of Chiropractic and Somatology, Faculty of Health Sciences, Durban University of Technology, Durban, South Africa. 2017. / Background The growing popularity and competitive nature of the beauty industry has resulted in beauty therapists performing an extensive range of treatments to meet the demand, often at the jeopardy of their own well-being. Treatments include massage, waxing, pedicures and facials. These techniques involve vigorous and repetitive movements that may lead to overuse and strain on the musculoskeletal system, resulting in injury. Work-related activities together with factors such as socio-demographic, psychosocial, environmental and anthropometric can influence the prevalence of work-related musculoskeletal disorders (WRMDs). There has been little investigation into the WRMDs of beauty therapists both locally and internationally. Aim The aim of this study was to determine the prevalence, selected risk factors and impact of self-reported work related musculoskeletal disorders (WRMDs) in beauty therapists working within the hotel spa industry in the eThekwini municipality. Method A cross sectional epidemiological design was used to survey 254 beauty therapists working in the hotel spa industry in the eThekwini municipality of Durban. A pre-validated questionnaire was hand delivered to beauty therapists working at all hotel spas that agreed to participate. Informed consent was obtained. The survey contained questions related to musculoskeletal pain, socio-demographic, psycho-social, lifestyle and occupational factors. Results A response rate of 70% was obtained (n=178). The majority of the respondents were female (94.3%; n=165), the mean age of the respondents was 27.74 (SD±4.83) years. The respondents had been working as beauty therapists for an average of 5.91 (SD±4.19; n=176) years and performed on average 27.89 (SD±13.33; n=170) treatments per week, working 47.38 (SD±13.36; n=175) hours per week. Those working overtime worked on average 5.43 (SD±2.94; n=59) hours of overtime per week. There was a high rate of reported WRMDs with the lifetime, current and 12 month period prevalence being 86% (n=153), 85% (n=151) and 83% (n=148), respectively. The area most affected were the low back followed by the neck, the hand and wrist. Ninety percent of the respondents indicated that their pain was mild to moderate in nature, with half expressing an inability to cope with the pain, and 78% reporting that it interfered with their ability to work. Mental exhaustion (p=0.032), suffering from a concomitant co-morbidity (p=0.031), years worked as a beauty therapist (p˂0.001) and treating clients after hours (p=0.007) were significantly associated with the presence of musculoskeletal pain. Performing manual massage (p=0.043) and reporting feelings of exhaustion whilst performing specialised massage techniques (p=0.014) and applying makeup/eyelashes (p=0.022) were associated with the presence of WRMDs. Those who reported having co-worker (p=0.006) and managerial support (p˂0.001) were less likely to experience musculoskeletal pain. In contrast, feeling frustrated by work (p=0.007), being under pressure (p=0.005) and over worked (p=0.009) increased the chances of experiencing WRMDs. Working with the back in an awkward position (p˂0.001), standing for prolonged periods (p˂0.001), lifting heavy loads (p=0.019) and working in a hot and humid environment (p<0.001) increased the risk of WRMDs. Conclusion WRMDs have a significant impact on beauty therapists’ ability to work. Effective strategies to prevent and manage WRMDs in hotel spa beauty therapists is required, along with governmental regulation. / M

The focus of this investigation was an analysis of the work demands being placed on South African automotive industry workers as there is a recognised problem with regard to the prevalence of musculoskeletal disorders (MSDs). Preliminary work was conducted to highlight the dominant risks and areas which elicited higher numbers of MSDs within the chosen automotive plant. An area of concern was highlighted through medical record analysis and the use of risk assessment tools, thereby prioritising the need for ergonomic intervention. In particular, the effects of varying restricted and overhead work heights on the biomechanical, physiological and psychophysical responses of an individual were investigated. Twenty-eight subjects were required to complete sixteen conditions. The conditions consisted of the adoption of restricted and upright overhead static postures, with half requiring the holding of four kilograms of weight in the hands and the remaining eight conditions having no weight. Testing was carried out using an electromyography unit, ergospirometer and a perceptual Body Discomfort Map and Scale. This involved a habituation and testing session. The results of the testing revealed the biomechanical and physiological responses were dependant on the change in height. Body discomfort was also shown to be variable over the changing height conditions. This indicates that there is a significant effect of height on an individual’s responses during overhead work. The extreme restricted (-200mm and -100mm) and upright (+300mm and +400mm) overhead conditions within this study were limiting, as they elicited the highest muscle activation, physiological responses and body discomfort ratings. Positions that are preferable to adopt, which were identified from the results in this study, indicate conditions closer to head height (0mm and +100mm) were favourable. The results therefore illustrate how awkward working postures during work are likely to elicit higher demands from an individual, which could lead to an increased risk for the development of a musculoskeletal disorder. The added factor of weight elicited significant results over all variables, excluding a respiratory The focus of this investigation was an analysis of the work demands being placed on South African automotive industry workers as there is a recognised problem with regard to the prevalence of musculoskeletal disorders (MSDs). Preliminary work was conducted to highlight the dominant risks and areas which elicited higher numbers of MSDs within the chosen automotive plant. An area of concern was highlighted through medical record analysis and the use of risk assessment tools, thereby prioritising the need for ergonomic intervention. In particular, the effects of varying restricted and overhead work heights on the biomechanical, physiological and psychophysical responses of an individual were investigated. Twenty-eight subjects were required to complete sixteen conditions. The conditions consisted of the adoption of restricted and upright overhead static postures, with half requiring the holding of four kilograms of weight in the hands and the remaining eight conditions having no weight. Testing was carried out using an electromyography unit, ergospirometer and a perceptual Body Discomfort Map and Scale. This involved a habituation and testing session. The results of the testing revealed the biomechanical and physiological responses were dependant on the change in height. Body discomfort was also shown to be variable over the changing height conditions. This indicates that there is a significant effect of height on an individual’s responses during overhead work. The extreme restricted (-200mm and -100mm) and upright (+300mm and +400mm) overhead conditions within this study were limiting, as they elicited the highest muscle activation, physiological responses and body discomfort ratings. Positions that are preferable to adopt, which were identified from the results in this study, indicate conditions closer to head height (0mm and +100mm) were favourable. The results therefore illustrate how awkward working postures during work are likely to elicit higher demands from an individual, which could lead to an increased risk for the development of a musculoskeletal disorder. The added factor of weight elicited significant results over all variables, excluding a respiratory individual.

INTRODUÇÃO: O aumento da cavidade ou da espessura do ventrículo esquerdo (VE) e da alteração morfológica do quadríceps se dão de forma diferente dependendo do tipo de atividade esportiva realizada (aeróbica ou anaeróbica). O presente estudo se propôs em verificar o grau de desenvolvimento da hipertrofia cardíaca e da musculatura periférica (quadríceps) em atletas fundistas e rasistas. MÉTODOS: Participaram deste estudo 16 (dezesseis) atletas de alta performance, voluntários, divididos em dois grupos: 8 (oito) atletas fundistas (corredores de 10000 metros) e 8 (oito) atletas rasistas (corredores de100 metros livre). Cada atleta foi submetido a uma coleta de sangue em jejum para as análises laboratoriais de rotina. Foram submetidos a exame de ressonância magnética cardíaca e do quadríceps femoral e, por fim, realizaram teste de esforço com a utilização da ergoespirometria. Para a análise estatística foram utilizados teste de Shapiro-Wilk para normalidade, quando não confirmado a normalidade, foi realizado o teste não paramétrico de Mann-Whitney, teste de Levene para homocedasticidade, teste t student para análise descritiva e comparativa entre os grupos e o teste de correlação de Spearman. RESULTADOS: Os fundistas e rasistas, respectivamente, apresentaram semelhanças nas comparações entre as variáveis antropométricas e clinicas, apresentando diferenças significativas nas variáveis sanguíneas direcionadas à troponina (2,2±1,0 e 3,0±2,4 ng/mL) com p = 0,042, creatina quinase (CKMB) com valores de 0,007 ng/mL e 0,016 ng/mL com p=0,001 e creatina fosfoquinase (CPK) com valores de 166,3±84,1 U/L e 465,1±221,6 U/L com p = 0,017, respectivamente. Em relação a variável relacionada à capacidade aeróbica dos corredores, não se observou diferenças entre corredores fundistas e rasistas. No que tange aos resultados apresentados na ressonância magnética, verificou-se diferença estatística comparativa no volume do quadríceps entre fundistas (3263,7 cm3) e rasistas (4946,4 cm3) sendo p < 0,01. Foi apresentado correlação significante no tamanho da área do quadríceps no grupo fundista (161,8±19,6 cm2) com a variável do diâmetro diastólico final do ventrículo esquerdo (DDF do VE) (5,3±0,5 cm) e no diâmetro sistólico final (DSF do VE) (3,5±0,4 cm), com p=0,05 em ambos. Ainda no grupo fundista, observou-se uma correlação positiva do DDF do VE com o volume do quadríceps (3263,7±212,2 cm3), com p = 0,04. No grupo rasista a significância estatística (p = 0,04) foi na aorta ascendente (2,7±0,5 cm) em correlação com a área do quadríceps (198,8±21,1 cm2) e no DSF do VE (3,5±0,5 cm) com o volume do quadríceps (4946,4±470,4 cm3). CONCLUSÃO: Com esses resultados pode-se afirmar que o quadríceps e o miocárdio adaptam-se estrutural e funcionalmente ao treinamento físico, sendo essas adaptações distintas de acordo com o tipo de modalidade esportiva realizada / INTRODUCTION: the increase of the cavity or the thickness of the left ventricle (LV) and the morphological changes of the quadriceps happen differently depending on the type of sports activity performed (aerobic or anaerobic). The proposal of the present study is to verifythe degree of development of cardiac hypertrophy and peripheral muscle (quadriceps) in short and long distance running athletes. METHOD: 16 (sixteen) high performance volunteer athletes participated in this study and were divided into two groups: 8 (eight) long distance athletes (10000 meter runners) and 8 (eight) short distance athletes (100 m runners /freestyle). Each athlete was subjected to a fasting blood collection for the routine laboratory analysis. They underwent a magnetic resonance imaging examination of the heart and quadriceps femoris and finally performed a stress test using ergo-spirometry.As statistical analysis we used Shapiro-Wilk test for normality and the non-parametric Mann-Whitney when normality was not confirmed; Levene test for homoscedasticity, t student test for descriptive and comparative analysis between the groups and the Spearman correlation test. RESULTS: The long and short distance athletes, respectively, presented similarities in the comparisons between the anthropometric and clinic variables, showing significant differences in blood variables directed to troponin (2.2 ± 1.0 and 3.0 ± 2.4 ng/mL) with p = 0.042, creatine kinase (CKMB) with values of 0.007 ng/mL and 0.016 ng/mL with p = 0.001 and creatine phosphokinase (CPK) with values of 166.3 ± U/L and 84.1 ± 465.1 221.6 U/L with p = 0.017, respectively. Regarding the variable related to the aerobic capacity of the runners, no differences were observed between long and short distance running athletes. With respect to the results presented in the MRI, there was comparative statistical difference in the quadriceps volume between long distance athletes (3263.7 cm3) and short distance athletes (4946.4 cm3) being p < 0,01. There was significant correlation in the size of the quadriceps area in long-distance group (161.8 ± 19.6 cm2) with the variable of final diastolic diameter of left ventricle (FDD of LV) (5.3 ± 0.5 cm) and in the final systolic diameter (FSD of LV) (3.5 ± 0.4 cm), with p = 0.05 in both. Also in the long distance group, it was noticed a positive correlation of FDD of LV with the volume of the quadriceps (3263.7 ± 212.2 cm3), with p = 0.04. In the short distance group, the statistical significance (p = 0.04) was in the ascending aorta (2.7 ± 0.5 cm) in correlation to the quadriceps area (198.8 ± 21.1 cm2) and in the FSD of LV (3.5 ± 0.5 cm) with the volume of the quadriceps (4946.4 ± 470.4 cm3). CONCLUSION: Based on this results we can state that both quadriceps and myocardium adapt structurally and functionally to the physical training, being these distinct adaptations according to the type of sport

Osteoclast-mediated focal articular bone erosion is a hallmark of rheumatoid arthritis, a disease of inflammation-induced bone loss. Inflammation in the bone microenvironment enhances osteoclast differentiation leading to bone erosion. Simultaneously, inflammation also inhibits osteoblast-mediated bone formation, further contributing to the net loss of bone. Previous studies have shown a paucity of mature osteoblasts at eroded bone surfaces correlating with suppression of bone formation and upregulation of antagonists of the Wnt pathway, a signaling cascade essential for osteoblast lineage commitment. Despite these observations, the exact pathogenesis of impaired bone formation in the setting of inflammation is not clearly understood. This dissertation aims to delineate the mechanisms by which inflammation suppresses osteoblast differentiation and activity in inflammatory arthritis. Specifically, this research elucidates how inflammation-induced alterations in the Wnt and bone morphogenetic protein (BMP) osteogenic signaling pathways contribute to bone loss and formation at distinct inflammatory microenvironments within the bone. Secondly, the means by which cellular mediators, including lymphocytes and macrophages, facilitate bone erosion and formation was addressed. Taken together, the research in this dissertation underscores the relationship between inflammation-induced bone loss and alterations in osteogenic signaling. Using an innovative murine inflammatory arthritis model, this study definitively demonstrates that resolving inflammation promotes osteoblast-mediated bone formation. Repair of erosions correlates with upregulation of synovial expression of Wnt10b, a Wnt agonist, and downregulation of sFRP1 and sFRP2, Wnt antagonists. This work also directly evaluates the contribution of sFRP1 to inflammation-induced bone destruction. Furthermore, this research demonstrates that expression of BMP3, a negative regulator of BMP signaling, is upregulated in osteoblasts by IL-17, a pro-inflammatory cytokine. BMP3-expressing osteoblasts are also observed at erosion sites in murine arthritis. Lastly, evaluation of the mediators of inflammation-induced periosteal bone formation implicates BMP2 as a means by which inflammation may positively regulate osteoblast function. This dissertation further elucidates the role of T cells and macrophages in the erosion and formation processes, respectively. In the absence of lymphocytes, bone erosion occurred normally, demonstrating that RANKL-expressing lymphocytes are not absolutely required for the bone erosion. Preliminary studies also suggest that M2 macrophages are potential mediators of bone formation via the expression of BMP2. In conclusion, this dissertation explores the ability of inflammation to act as a rheostat, which controls the fate of bone by modulating not only osteoclast differentiation, but also osteogenic signaling pathways and cellular mediators in the bone microenvironment. The soluble mediators and cell types identified in this research highlight novel mechanisms by which inflammation may regulate osteoblast activity within the bone microenvironment. Collectively, these data imply that strict control of inflammation may be necessary in order to create an anabolic environment that preserves bone architecture in diseases of inflammation-induced bone loss.

Indiana University-Purdue University Indianapolis (IUPUI) / Articular cartilage is subjected to dynamic mechanical loading during normal daily activities. This complex mechanical loading, including cell deformation and interstitial fluid flow, affects chondrocyte mechano-chemical signaling and subsequent cartilage homeostasis and remodeling. Focal adhesion kinase (FAK) and Src are known to be main mechanotransduction proteins, but little is known about the effect of mechanical loading on FAK and Src under its varying magnitudes and types. In this study, we addressed two questions using C28/I2 chondrocytes subjected to the different types and magnitudes of mechanical loading: Does a magnitude of the mechanical loading affect activities of FAK and Src? Does a type of the mechanical loading also affect their activities? Using fluorescence resonance energy transfer (FRET)-based FAK and Src biosensor in live C28/I2 chondrocytes, we monitored the effects of interstitial fluid flow and combined effects of cell deformation/interstitial fluid flow on FAK and Src activities. The results revealed that both FAK and Src activities in C28/I2 chondrocytes were dependent on the different magnitudes of the applied fluid flow. On the other hand, the type of mechanical loading differently affected FAK and Src activities. Although FAK and Src displayed similar activities in response to interstitial fluid flow only, simultaneous application of cell deformation and interstitial fluid flow induced differential FAK and Src activities possibly due to the additive effects of cell deformation and interstitial fluid flow on Src, but not on FAK. Collectively, the data suggest that the intensities and types of mechanical loading are critical in regulating FAK and Src activities in chondrocytes.

Indiana University-Purdue University Indianapolis (IUPUI) / As our population ages, treating bone and joint ailments is becoming increasingly important. Both osteoporosis, a bone disease characterized by a decreased density of mineral in bone, and osteoarthritis, a joint disease characterized by the degeneration of cartilage on the ends of bones, are major causes of decreased movement ability and increased pain. To combat these diseases, many treatments are offered, including drugs and exercise, and much biomedical research is being conducted. However, how can we get the most out of the research we perform and the treatment we do have? One approach is through computational analysis and mathematical modeling. In this thesis, quantitative methods of analysis are applied in different ways to two systems: osteoporosis and osteoarthritis. A mouse model simulating osteoporosis is treated with salubrinal and knee loading. The bone and cell data is used to formulate a system of differential equations to model the response of bone to each treatment. Using Particle Swarm Optimization, optimal treatment regimens are found, including a consideration of budgetary constraints. Additionally, an in vitro model of osteoarthritis in chondrocytes receives RNA silencing of Lrp5. Microarray analysis of gene expression is used to further elucidate the mode of regulation of ADAMTS5, an aggrecanase associated with cartilage degradation, by Lrp5, including the development of a mathematical model. The math model of osteoporosis reveals a quick response to salubrinal and a delayed but substantial response to knee loading. Consideration of cost effectiveness showed that as budgetary constraints increased, treatment did not start until later. The quantitative analysis of ADAMTS5 regulation suggested the involvement of IL1B and p38 MAPK. This research demonstrates the application of quantitative methods to further the usefulness of biomedical and biomolecular research into treatment and signaling pathways. Further work using these techniques can help uncover a bigger picture of osteoarthritis's mode of action and ideal treatment regimens for osteoporosis.

Indiana University-Purdue University Indianapolis (IUPUI) / Bone is a dynamic organ that responds to its external environment. Cell signaling cascades are initiated within bone cells when changes in mechanical loading occur. To describe these molecular signaling networks that sense a mechanical signal and convert it into a transcriptional response, we proposed the mechanosome model. “GO” and “STOP” mechansomes contain an adhesion-associated protein and a nucleocytoplasmic shuttling transcription factor. “GO” mechanosomes functions to promote the anabolic response of bone to mechanical loading, while “STOP” mechanosomes function to suppress the anabolic response of bone to mechanical loading. While much work has been done to describe the molecular mechanisms that enhance the anabolic response of bone to loading, less is known about the signaling mechanisms that suppress bone’s response to loading. We studied two adhesion-associated proteins, Src and Pyk2, which may function as “STOP” mechanosomes. Src kinase is involved in a number of signaling pathways that respond to changes in external loads on bone. An inhibition of Src causes an increase in the expression of the anabolic bone gene osteocalcin. Additionally, mechanical stimulation of osteoblasts and osteocytes by fluid shear stress further enhanced expression of osteocalcin when Src activity was inhibited. Importantly, fluid shear stress stimulated an increase in nuclear Src activation and activity. The mechanism by which Src participates in attenuating anabolic gene transcription remains unknown. The studies described here suggest Src and Pyk2 increase their association in response to fluid shear stress. Pyk2, a protein-tyrosine kinase, exhibits nucleocytoplasmic shuttling, increased association with methyl-CpG-binding protein 2 (MBD2), and suppression of osteopontin expression in response to fluid shear stress. MBD2, known to be involved in DNA methylation and interpretation of DNA methylation patterns, may aid in fluid shear stress-induced suppression of anabolic bone genes. We conclude that both Src and Pyk2 play a role in regulating bone mass, possibly through a complex with MBD2, and function to limit the anabolic response of bone cells to fluid shear stress through the suppression of anabolic bone gene expression. Taken together, these data support the hypothesis that “STOP” mechanosomes exist and their activity is simulated in response to fluid shear stress.

Объектом исследования явилась жизнестойкость подростков с нарушениями опорно-двигательного аппарата как психологический феномен. Предметом исследования стало психологическое содержание жизнестойкости подростков с НОДА. Магистерская диссертация состоит из введения, двух глав, заключения, списка литературы (38 источника) и приложения, включающего в себя бланки применявшихся методик. Объем магистерской диссертации 75 страниц, на которых размещены 14 таблиц. Во введении раскрывается актуальность проблемы исследования, разработанность проблематики, ставятся цель и задачи исследования, определяются объект и предмет исследования, формулируются гипотезы, указываются методы и эмпирическая база. Первая глава включает в себя обзор литературы по теме исследования, описание теоретических подходов к изучению жизнестойкости. Представлены разделы, посвященные факторам жизнестойкости личности, роли жизнестойкости в преодолении возникающих трудностей и психолого-педагогическим особенностям подростков с НОДА. Выводы по первой главе представляют собой итоги по изучению теоретического материала. Вторая глава посвящена эмпирической части исследования. В ней представлено описание организации и методов проведенного исследования и результатов, полученных по всем использованным методикам: Тесту жизнестойкости С. Мадди (в адаптации Д. А. Леонтьева, Е. И. Рассказовой), Личностному опроснику Г. Айзенка (EPI), методике Дембо-Рубинштейн (в модификации А. М. Прихожан), копинг-тесту Р. Лазаруса и С. Фолкман (в адаптации Л. И. Вассермана). Также в главе представлены сравнительный, корреляционный и факторный анализ результатов исследования. Выводы по главе 2 включают в себя основные результаты эмпирического исследования. В заключении в обобщенном виде изложены результаты теоретической и эмпирической частей работы, а также выводы по выдвинутым гипотезам. / The subject matter of the study is hardiness in adolescents with disorders of the musculoskeletal system as a psychological phenomenon. The object matter of the study is psychological content of hardiness of adolescents with disorders of the musculoskeletal system. The master's thesis consists of an introduction, two chapters, conclusions, a list of references (38 sources) and an annex including the blank forms of the applied methods. The volume of the master`s thesis is 75 pages, which includes 14 tables. The introduction reveals the relevance of the chosen research problem, the level of development of the problematics; the objective and the tasks of the study are set, the subject matter and the scope of the research are determined, hypotheses are formulated, the methods and the empirical base are defined. The first chapter includes a review of literature on the research topic, a description of theoretical approaches to the study of hardiness. The sections devoted to the factors of the hardiness of the individual, the role of hardiness in overcoming the emerging difficulties and the psychological and pedagogical characteristics of adolescents with disorders of the musculoskeletal system, are presented. Conclusions on the first chapter are the results of the study of theoretical material. The second chapter is devoted to the empirical part of the study. It includes a description of the organization and methods of the study, as well as the results obtained due to the applied methods: a Hardiness Test by S. Maddi (adapted by D. Leontiev, E. Rasskazova), a G. Eysenck Personality Questionnaire (EPI), a Method of Self-Esteem Measurement by Dembo-Rubinstein (in the modification of A.Prikhozhan), a Ways of Coping Questionnaire by R. Lazarus and S. Folkman (adapted by L. Wasserman). Also, the chapter presents a comparative, correlation and factor analysis of the results of the study. The findings of Chapter 2 are the main results of the empirical study. In the conclusion, brief results of the theoretical and empirical parts of the work are presented, as well as conclusions on the hypotheses.