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<p>Primary hyperparathyroidism (pHPT) due to parathyroid tumours with hypersecretion of parathyroid hormone and hypercalcaemia is a common disease with incompletely understood etiology affecting more than 1 % of the population, primarily postmenopausal women. In secondary hyperparathyroidism (sHPT), parathyroid tumours develop in response to calcium and vitamin D deficiency generally in patients with uraemia. HPT is usually treated by surgical removal of enlarged parathyroid glands.</p><p>The aim of this thesis was to examine the Wnt/β-catenin signalling pathway in parathyroid tumours.</p><p>Aberrantly accumulated β-catenin was found in all analysed pHPT and sHPT tumours, with a stabilising homozygous mutation (Ser37Ala) in 7.3% of the pHPT tumours. Truncation of the APC protein was not found. MYC, a β-catenin target gene was overexpressed in a substantial fraction of pHPT and sHPT parathyroid tumours. </p><p>A parathyroid tumour cell line (sHPT-1) was established from a hyperplastic gland removed at operation of a patient with sHPT. The cells produced parathyroid hormone and grew with a doubling time of approximately 72 hours. Stabilised nonphosphorylated transcriptionally active β-catenin was expressed. Efficient transfection of siRNA against β-catenin decreased expression of cyclin D1 and MYC, and inhibited cell growth with ensuring cell death. </p><p>The Wnt coreceptor LRP5 was found expressed with an internal deletion of 142 amino acids (LRP5Δ) in 86% and 100% of pHPT and sHPT tumours, respectively. Stabilising mutation of β-catenin and expression of LRP5Δ was mutually exclusive. Expression of LRP5Δ was required to maintain the nonphosphorylated transcriptionally active ß-catenin level, MYC expression, parathyroid cell growth in vitro, and tumour growth in transplanted SCID mice. Wnt3 ligand and LRP5Δ strongly activated transcription, and LRP5Δ was insensitive to inhibition by DKK1.</p><p>Aberrant accumulation of β-catenin by stabilising mutation or expression of LRP5Δ appears as a common pathogenic pathway for hyperparathyroid disease. LRP5Δ in particular presents a potential target for therapeutic intervention.</p>

Primary hyperparathyroidism (pHPT) due to parathyroid tumours with hypersecretion of parathyroid hormone and hypercalcaemia is a common disease with incompletely understood etiology affecting more than 1 % of the population, primarily postmenopausal women. In secondary hyperparathyroidism (sHPT), parathyroid tumours develop in response to calcium and vitamin D deficiency generally in patients with uraemia. HPT is usually treated by surgical removal of enlarged parathyroid glands. The aim of this thesis was to examine the Wnt/β-catenin signalling pathway in parathyroid tumours. Aberrantly accumulated β-catenin was found in all analysed pHPT and sHPT tumours, with a stabilising homozygous mutation (Ser37Ala) in 7.3% of the pHPT tumours. Truncation of the APC protein was not found. MYC, a β-catenin target gene was overexpressed in a substantial fraction of pHPT and sHPT parathyroid tumours. A parathyroid tumour cell line (sHPT-1) was established from a hyperplastic gland removed at operation of a patient with sHPT. The cells produced parathyroid hormone and grew with a doubling time of approximately 72 hours. Stabilised nonphosphorylated transcriptionally active β-catenin was expressed. Efficient transfection of siRNA against β-catenin decreased expression of cyclin D1 and MYC, and inhibited cell growth with ensuring cell death. The Wnt coreceptor LRP5 was found expressed with an internal deletion of 142 amino acids (LRP5Δ) in 86% and 100% of pHPT and sHPT tumours, respectively. Stabilising mutation of β-catenin and expression of LRP5Δ was mutually exclusive. Expression of LRP5Δ was required to maintain the nonphosphorylated transcriptionally active ß-catenin level, MYC expression, parathyroid cell growth in vitro, and tumour growth in transplanted SCID mice. Wnt3 ligand and LRP5Δ strongly activated transcription, and LRP5Δ was insensitive to inhibition by DKK1. Aberrant accumulation of β-catenin by stabilising mutation or expression of LRP5Δ appears as a common pathogenic pathway for hyperparathyroid disease. LRP5Δ in particular presents a potential target for therapeutic intervention.

Made available in DSpace on 2015-07-22T20:49:32Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-11-24 / INTRODUÇÃO: Osteoporose Pseudoglioma (OPPG) é uma doença rara, caracterizada por osteoporose grave de início juvenil, associada a múltiplas fraturas e anormalidades oculares. É causada por uma mutação inativadora no gene da “low–density lipoprotein receptor-related protein 5” (LRP5), que está envolvido na formação óssea. O papel inicialmente proposto para o LRP5 seria sua participação na formação de um complexo receptor, juntamente com as proteínas Frizzled e Wnt na membrana dos osteoblastos, que, quando ativados, promoveriam a estabilização da beta-catenina no citoplasma destas células e induziram a formação óssea. A teriparatida, por sua vez, é um medicamento anabólico que, portanto, atua sobre a formação óssea. Seu mecanismo de ação, entretanto, ainda não está completamente esclarecido. Em camundongos com mutação inativadora do LRP5 -/- o tratamento com teriparatida induziu a um incremento da BMD, semelhante ao obtido em camundongos (LRP5 +/+), sugerindo que o efeito anabólico do PTH não utilize a via de sinalização do LRP5. Esta constatação nos incentivou a tratar com teriparatida um de nossos pacientes portador de OPPG. Ao longo deste tratamento, entretanto, uma nova teoria sobre o papel do LRP5 surgiu, associando-o à produção de serotonina intestinal. OBJETIVOS: Avaliar eficácia terapêutica da teriparatida em um paciente adulto jovem com OPPG sobre DMO e marcadores de remodelação óssea, além de dosar serotonina sérica neste paciente, em seu irmão também afetado e em controles saudáveis. PACIENTE E MÉTODOS: Paciente masculino, 19 anos com amaurose congênita e fraturas não traumáticas desde a infância. Após 3 anos de infusões periódicas de pamidronato com resposta insatisfatória, foi iniciado a teriparatida 20 mcg/dia por via subcutânea uma vez ao dia, durante 24 meses. Amostras sanguíneas e urinárias foram coletadas em jejum pela manhã nos tempos basal e a cada 3 meses até o final do tratamento. Foram determinadas a relação do cálcio/creatinina em amostra isolada de urina e as dosagens séricas de Telopeptideo Carboxiterminal do colágeno Tipo I (CTX), Peptídeo Aminoterminal do Procolágeno Tipo I (P1NP), cálcio total e ionizado, fósforo, ácido úrico, hemograma, função hepática e renal. Densitometria óssea foi realizada por Dual energy X-ray absorptiometry (DXA) a cada 12 meses. Serotonina sérica foi dosado por ELISA (kit Fitzgerald Cat# 55R- RE59121) em um único momento, logo após o término do tratamento. RESULTADOS: Os parâmetros sanguíneos e urinários estavam normais no basal e durante o tratamento. Com relação aos marcadores bioquímicos do metabolismo ósseo, notamos aumento inicial nos valores do CTX, atingindo pico no terceiro mês. O P1NP começou a aumentar no sexto mês, com pico tardio no nono mês. Apesar deste padrão não usual de resposta dos marcadores, houve aumento de 9,7% na densidade mineral óssea (DMO) em coluna lombar comparado ao basal e de 10,2% em fêmur total. As concentrações de serotonina foram duas vezes e meia maior no paciente, comparado aos valores obtidos em indivíduos normais. CONCLUSÃO: Em um paciente com OPPG previamente tratado com pamidronato, teriparatida mostrou-se segura e capaz de induzir aumento substancial da DMO de coluna lombar e fêmur proximal. Os valores elevados de serotonina plasmática concordam com os relatos descritos na literatura em outros pacientes com OPPG, mas seu papel no controle da formação óssea ainda precisa ser melhor estabelecido. / INTRODUCTION: Osteoporosis Pseudoglioma (OPPG) is characterized by severe juvenile-onset osteoporosis and ocular abnormalities. It is caused by one of several inactivating mutations in LRP5, a gene importantly involved in bone formation. OBJECTIVE: The objective of this study was to evaluate the efficacy of teriparatide in a young man with OPPG. PATIENT AND METHOD: The subject of this case report is a 19-year-old man with congenital blindness and low trauma fractures due to OPPG. A 2-year course of teriparatide, 20 mcg/day, was initiated after a 6-year course of intravenous pamidronate infusions, the latter 3 years of which had minimal effects on bone mineral density (BMD). Measurements in serum were made of C-terminal telopeptide of type I collagen (CTX), N-terminal propeptide of type I collagen (P1NP), total and ionized calcium, phosphate, uric acid, complete blood count, renal and liver function tests. Urinary calcium/creatinine ratio was determined. BMD was measured by DXA yearly. RESULTS: BMD increased by 9.7% in lumbar BMD and 10.2% in right femur hip. CTX rose early, peaking in month 3, followed by an increase in P1NP, peaking in month 9. Both indices returned to baseline by month 24. The increase in CTX followed by P1NP is an unusual time course when teriparatide is used to treat osteoporosis but may be typical of low bone turnover states. There were no adverse events. CONCLUSION: In a patient with OPPG, teriparatide markedly increased BMD in the lumbar spine and femur hip. / TEDE / BV UNIFESP: Teses e dissertações

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.