Atherosclerosis is an inflammatory disease, occurring preferentially in arterial regions with disturbed flow. We have shown that disturbed flow induces inflammation in endothelial cells (ECs) by producing bone morphogenic protein-4 (BMP4). Moreover, chronic BMP4 infusion induces endothelial dysfunction and systemic hypertension in mice. Here, we examined which BMP receptors (BMPR) mediate BMP4 action in ECs. Western blot, immunostaining and RT-PCR studies using human and bovine ECs, mouse aortas and human coronary arteries (HCA) showed that BMPRI (ALK2 and 6) and BMP-RII were expressed in ECs. As a functional test, ECs were treated with a BMPRII siRNA to knockdown expression. BMPRII knockdown blocked a well-known BMP4 response - smad1/5/8 phosphorylation, as expected. Unexpectedly, BMPRII knockdown itself significantly stimulated ICAM-1 and VCAM-1 expression and monocyte adhesion in a BMP4-independent manner. Inflammatory responses caused by BMPRII knockdown were blocked by inhibitors of NADPH oxidase and NFκ B. From these results, we hypothesized that BMP-RII knockdown in ECs would cause inflammation, which is a critical event in atherosclerosis initiation and progression. Genetic mutations of BMPRII have been linked to primary pulmonary hypertension. However, it is not known whether BMP-RII is regulated by atherosclerotic conditions and plays a role in non-pulmonary vessels causing inflammation and atherosclerosis. We examined BMPRII levels in HCA by immunostaining. While non-diseased arteries showed intense staining of BMPRII, the expression decreased as lesions became more advanced. BMPRII was virtually undetectable in the most advanced lesions. These findings suggested a potential link between pro-atherosclerotic conditions and BMP-RII levels. We tested this hypothesis by treating ECs with pro-inflammatory cytokines found in atheromas: TNFα decreased BMPRII by 2-fold. In contrast, statins increased BMPRII by 4-fold. In summary, we demonstrate for the first time that BMPRII can be down- or up-regulated by pro- or anti-atherogenic conditions, respectively, and it is dramatically decreased in HCA with advanced plaques. Moreover, BMPRII knockdown in ECs induces inflammation, a critical atherogenic step. We propose that focal inflammation initiated by disturbed flow, together with circulating pro-atherogenic risk factors, may lead to a vicious cycle of BMPRII down-regulation causing secondary inflammation and atheroma progression.

[Truncated abstract] Bone is a living tissue and is maintained by the coordinate action of osteoblasts and osteoclasts. The intercellular communication between these two cells is the quintessential mechanism in bone remodelling. Unfortunately, the importance of this interaction is often neglected and its significance is only realised when disruption of this “cross-talk” results in debilitating bone diseases. Additionally, the number of known proteins that are involved in this “cross-talk”, especially those that are osteoclast-derived, and act specifically on osteoblasts, is limited. This discrepancy leads to the question: Can osteoclasts directly control the growth and function of osteoblastic cells by expressing specific proteins that bind directly to osteoblasts? If so, is it possible to use these proteins to control and, possibly, treat bone disease? The objective of this thesis is to identify and characterise osteoclast-derived factors that can modulate bone homeostasis, as well as contribute to the intercellular communication between osteoblasts and osteoclasts ... Collectively, the data in this thesis culminates in one important conclusion: the identification of a novel paracrine secretory factor that has the potential to directly induce the formation of bone. These findings represent the first ever characterisation of a protein that allows the osteoclasts to directly control the growth and function of osteoblasts. Due to the potential function of ODOF to induce bone formation, this protein may be used therapeutically to treat bone disease.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Diss. (sammanfattning) Uppsala : Univ., 2001. / Härtill 5 uppsatser.

Thesis (Ph. D.)--University of Hong Kong, 2010. / Includes bibliographical references (leaves 202-229). Also available in print.

Thesis (Ph. D.)--Cornell University, May, 2009. / Vita. Includes bibliographical references (leaves 152-178).

Thesis (Ph. D.)-- University of Washington, 2005. / Includes bibliographical references (leaves 107-127).

Thesis (M.Sc.)--York University, 2009. Graduate Programme in Biology. / Typescript. Includes bibliographical references (leaves 53-57). Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:MR51603

Made available in DSpace on 2014-06-11T19:31:07Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-07-29Bitstream added on 2014-06-13T21:02:11Z : No. of bitstreams: 1 guskuma_mh_dr_araca.pdf: 465143 bytes, checksum: 0281f4cc114abd1f60018aa084e64214 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A proposta deste estudo foi avaliar a expressão de proteínas que participam da fase de osteoindução (VEGF, BMP-2 e CBFA1) durante o processo de regeneração óssea de defeitos criados em calvária de ratos e preenchidos com enxerto autógeno em bloco. MATERIAIS E MÉTODOS: Para o presente estudo foram utilizados 10 ratos adultos machos (Rattus norvegicus albinus, Wistar) que receberam dois defeitos ósseos de 5 mm cada, em calvária. Os defeitos ósseos constituiram dois grupos experimentais (n=10): Grupo controle (CONT) (defeitos preenchidos com o próprio coágulo); Grupo enxerto (ENX) (defeitos preenchidos com osso autógeno removido do defeito contralateral). Os animais foram submetidos a eutanásia nos períodos de 7 e 30 dias pós-operatórios. RESULTADOS: A análise quantitativa demonstrou formação óssea significativamente maior no Grupo ENX, no entanto, a presença das proteínas estudadas foi significativamente maior no Grupo CONT em ambos os períodos de observação. CONCLUSÃO: O enxerto ósseo autógeno cortical em bloco não expressou de forma significativa as proteínas osteoindutoras estudadas durante o processo de reparo / AIMS: The proposal of this study was to evaluate the expression of proteins that act in osteoinduction (VEGF, BMP-2, CBFA1) phase during the bone defects regeneration created in rat calvaria and filled with autogenous bone graft in block. METHODS: For the present study, 10 adult male rats (Rattus norvegicus albinus, Wistar) had two 5mm- bone defects in calvaria. Bone defects constituted two experimental groups: CONTROL Group (defects filled with blood clot); GRAFT Group (defects filled with bone graft). Animals were sacrificed at 7 and 30 days post operative. RESULTS: Quantitative analysis showed significantly higher bone formation in Graft Group, however, the presence of studied proteins was significantly higher in Control Group in both observation periods. CONCLUSION: Autogenous cortical bone graft in block did not express the studied osteoinductive proteins during bone repair

Orientador: Roberta Okamoto / Banca: Wilson Roberto Poi / Banca: Idelmo Rangel Garcia Júnior / Banca: Thallita Pereira Queiroz / Banca: Paulo Tambasco de Oliveira / Resumo: A neoformação óssea em defeitos críticos em calvária de ratos depende fortemente das propriedades osteocondutoras dos enxertos e biomateriais. Ainda é controverso se os biomateriais podem substituir os enxertos de osso autógeno e se a suplementação dos biomateriais com Proteínas Ósseas Morfogenéticas (BMPs) é necessária para melhorar a formação óssea. Examinamos defeitos críticos em calvária de ratos (5 mm de diâmetro) tratados com β-tricálcio fosfato (TCP; Cerasorb ® M), gel de ácido polilático e poliglicólico (PLA/PGA; Fisiograft®) e cimento de fosfato de cálcio (CPC; Norian® CRS®), isoladamente ou na presença de 5μg de BMP-2 após 45 dias. Defeitos tratados com enxerto de osso autógeno particulado e defeitos não tratados serviram como controle. A formação óssea foi avaliada com base na análise de μCT, análise histomorfométrica e análise de fluorescência. Nós relatamos que o TCP apoia a formação óssea de forma mais eficiente do que o enxerto de osso autógeno particulado. A formação óssea na presença de TCP sozinho atingiu um nível máximo de neoformação óssea, enquanto que a suplementação de BMP-2 falhou em melhorar a neoformação óssea. Em contrapartida, não houve diferença significativa na formação óssea quando o PLA / PGA e o CPC foram comparados ao enxerto autógeno. Além disso, a presença de BMP-2 não alterou substancialmente as propriedades osteocondutoras de PLA/PGA ou de CPC. Conclui-se que as propriedades osteocondutoras do TCP são superiores aos dos enxertos autógenos e que o TCP não exige suplementação de BMP-2. Nossos resultados também mostram que a diminuição da capacidade osteocondutora do PLA/PGA e do CPC não podem ser superadas pela suplementação de BMP-2 em defeitos de calvária de ratos / Abstract: Bone formation in critical-sized calvaria defects is strongly dependent on the osteoconductive properties of grafts. It remains a matter of controversy whether biomaterials can replace autografts and whether the supplementation of biomaterials with Bone Morphogenetic Proteins (BMPs) is necessary to enhance bone formation. We examined rat calvaria critical-sized defects (5mm diameter) treated with β-tricalcium phosphate (TCP; Cerasorb® M), polylactic and polyglycolic acid gel (PLA/PGA; Fisiograft®) and calcium phosphate cement (CPC; Norian® CRS®), either alone or in the presence of 5μg of BMP-2 after 45 days. Autografts and untreated defects served as controls. Bone formation was evaluated based on μCT analysis, histomorphometric analysis and fluorescence analysis. We report that TCP supported bone formation more efficiently than did autografts. Bone formation in the presence of TCP alone reached a maximal level, as BMP-2 supplementation failed to enhance bone formation. By contrast, no significant difference in bone formation was observed when PLA/PGA and CPC were compared to autografts. Moreover, the presence of BMP-2 did not substantially change the osteoconductive properties of PLA/PGA or CPC. We conclude that the osteoconductive properties of TCP are superior to those of autografts and that TCP does not require BMP-2 supplementation. Our findings also show that the decreased osteoconductive properties of PLA/PGA and CPC cannot be overcome by BMP-2 supplementation in rat calvaria defects / Doutor