Aim: Recent interest in cardiogenesis has focused on the epicardium, the outer epithelial layer that envelops the heart. Epicardial-derived cells (EPDCs) contribute vascular smooth muscle to developing coronary vessels and provide critical signalling cues to facilitate myocardial functionality. However, the precise molecular mechanisms that underpin epicardial biology remain unclear. Ablation of Myh10 in the EHC mouse results in embryonic lethal cardiac malformations, including epicardial and coronary defects. We sought to establish the role of Myh10 in epicardial cell function to further dissect the coronary vessel developmental pathway, a deeper understanding of which may inform the design of therapeutics to regenerate and repair the injured heart. Methods: Utilising multiple cell and developmental biology techniques, we generated a pathological evaluation of the EHC phenotype. EPDC migration was investigated in vivo with Wt1 immunohistochemistry, and in vitro by performing scratch wound assays on epicardial cell cultures. Congruently, we examined the ability of epicardial cells to undergo EMT in vivo by employing Snail and Phosphohistone-H3 immunohistochemistry. Results: Our studies reveal that EHC epicardial cells have a reduced capacity to invade the ventricular myocardium. Furthermore, we discovered increased proliferation and reduced Snail expression specifically within the EHC epicardium, consistent with EMT dysregulation. Interestingly, epicardial cell function did not appear to be disrupted in vitro. Conclusion: These results demonstrate a novel role for Myh10 in both EPDC migration and the promotion of epicardial EMT. Our finding that migration is unaffected in vitro suggests that the unique properties of the in vivo epicardial microenvironment dictate a requirement for Myh10 in order to elicit correct epicardial function. Together, this research enhances our understanding of the dysfunctional processes that contribute to abnormal cardiogenesis; these insights may aid our ability to determine the molecular regulators of coronary vessel development, and create therapeutics to regenerate vessel growth and repair injured cardiac tissue in cardiovascular disease.

The metabolism of 5'-AMP by 5'-nucleotidase, adenylate deaminase and adenylate kinase was examined in heart homogenates of rat, rabbit, dog, pigeon and turtle. The study was conducted in consideration of the possibility that adenosine, a catabolic product of 5'-AMP, may control vasotone for the autoregulation of coronary blood flow. The relative activities of homogenates of hearts from various species to form adenosine by the action of 5'-nucleotidase generally supported such a role for this nucleoside. Those species anticipated to have the largest potential requirements for coronary vasodilation, i.e. those whose oxygen consumption is known to increase significantly during physical exertion, had the highest levels of cardiac 5'-nucleotidase. An exception to this was the pigeon which had no detectable cardiac 5'-nucleotidase; the order of levels of this enzyme in hearts of the other species tested was: rat > dog > rabbit > turtle. The turtle ventricle, by virtue of its high content of adenylate deaminase and low content of 5'-nucleotidase appeared to catabolize 5'-AMP largely by deamination to IMP. Homogenates of pigeon ventricle contained the greatest activity of adenylate kinase, indicating that the heart of this species is equipped for preservation of ATP by resynthesis from ADP. Enzyme histochemistry revealed that most 5'-nucleotidase of mammalian hearts was localized in the endothelial cells of capillaries. Therefore, if adenosine is involved in regulation of coronary perfusion, its source may be capillary endothelial cells rather than cardiac muscle cells. 5'-Nucleotidase was partially purified from an acetone powder of rat heart. It was active over a broad range of pH with an optimum at pH 8.5. The enzyme was stimulated up to 5-fold by Mg(++) [formula omitted]; Mn(++) and Ni(++) also stimulated activity. The K for 5'-AMP was 2.1 x 10(-5)M in the absence of 16Mg and 2.3 x 10 M in the presence of 16 mM MgCl(2). Certain of its properties indicated that the production of adenosine might be favoured under conditions in which coronary vasodilation would be required and vice-versa. For example, the enzyme was inhibited by ATP, whose levels are greatest in well oxygenated hearts in which energy charge is high. Not all properties of 5'-nucleotidase were consistent with enhanced adenosine formation at reduced energy charge. Both ADP and orthophosphate, the levels of which increase when energy charge decreases, inhibited the enzyme; in fact ADP was a more powerful inhibitor than ATP. In addition, the enzyme was not specific for 5'-AMP but hydrolyzed a variety of nucleoside 5'-monophosphates; and the hydrolysis of 5'-AMP was competitively inhibited by UMP. In the absence of Mg(++) , inhibition by ADP was of the mixed (competitive- non-competitive) type. In the presence of 16 mM MgCl(2), inhibition was non-competitive. On the basis of these data and Dixon plots of inhibition as a function of ADP concentration, it is suggested that two conformations of the enzyme are possible; one which is competitively inhibited by ADP. The simple non-competitive inhibition by ADP, observed in the presence of 16 mM MgCl(2), is attributed to Mg(++) -induced preference for the latter conformation. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

Coronary heart diseases are a significant cause of death among both men and women in the industrialized world; thus finding ways to detect factors which cause coronary heart diseases is worthy challenge for researchers. Ultrasound elastography system (50MHz) for measuring mechanical properties of arteries was developed as part of this thesis. Ability to discriminate between various tissue types was demonstrated using fresh and modified porcine coronary arteries, which closely models the plaque in human atherosclerosis. Elastographic measurements agreed well with uniaxial mechanical testing over a range of compression moduli.

Thesis (Ph. D.)--University of Washington, 1997. / Vita. Includes bibliographical references (leaves [133]-140).

No description available.

Thesis (Ph. D.)--University of Missouri--Columbia, 1999. / "December 1999." Typescript. Vita. Includes bibliographical references (leaves [174]-186). Also available on the Internet.

FUNDAMENTOS: A reestenose após intervenção coronária percutânea é maior nos pacientes com vasos de pequeno calibre em comparação aos vasos grandes. Os stents com sirolimus demonstraram importante redução da reestenose em vasos maiores que 3,0mm. O desempenho destes stents nos vasos pequenos deve ser investigado. MATERAIS E MÉTODOS: O propósito deste estudo foi avaliar a redução do volume de hiperplasia intimal após angioplastia com stents com sirolimus (Cypher®) comparados com os stents não recobertos de estrutura metálica fina (Pixel®), em pacientes com vasos pequenos. Oitenta pacientes com doença arterial coronária foram prospectivamente incluídos em duas séries consecutivas de tratamento, sendo a primeira empregando stents com sirolimus (50) e a segunda stents não recobertos de estrutura metálica fina (30). Os resultados foram: menor porcentual de obstrução da prótese através da análise volumétrica do ultrasom intracoronário [5,0% (EP=0,77) versus 39,0% (EP=4,72), p<0,001], menor perda tardia intra-stent [0,25mm (EP=0,03) versus 1,11mm (EP=0,13), p<0,001] e no segmento do vaso-alvo [0,30mm (EP=0,04) versus 0,83mm (EP=0,11), p<0,001], e também menor reestenose intra-stent (0% versus 33,3%, p<0,001) e no segmento do vaso (4% versus 36,7%, p<0,001) com os stents com sirolimus. A sobrevivência livre de eventos aos oito meses de evolução foi de 96% com os stents com sirolimus versus 86,7% com os stents não recobertos (p=0,190). CONCLUSÃO: Os pacientes com vasos de pequeno calibre após o implante de stents com sirolimus evoluem com menor hiperplasia intimal (menor porcentual de obstrução intra-stent e menor perda tardia) do que quando são utilizados stents não recobertos de estrutura metálica fina. Isto resulta em redução significativa da reestenose angiográfica aos oito meses de evolução / BACKGROUND: Patients with small vessels treated with percutaneous coronary interventions are at high risk of restenosis. Sirolimus-eluting stents has proved safety and effectiveness in reducing restenosis in large vessels. The outcomes after sirolimus-eluting stents in small vessels have not been adequately investigated. METHODS: We conducted a prospective study in 80 patients with small vessels treated with percutaneous intervention with sirolimus-eluting stents (Cypher(TM)) compared to thin-strut-thickness stents (Pixel(TM)). The primary end point was the reduction in intimal hyperplasia volume after coronary stenting accessed by intravascular ultrasound. OUTCOMES: the use of sirolimus-eluting stents compared with the use of the thin-strut-thickness stents reduced in-stent obstruction as a percent of volume [5,0% (EP=0,77) versus 39,0% (EP=4,72), p<0,001], in-stent lateloss [0,25mm (EP=0,03) versus 1,11mm (EP=0,13), p<0,001], in-segment late-loss [0,30mm (EP=0,04) versus 0,83mm (EP=0,11), p<0,001], and instent and in-segment restenosis [0% versus 33,3%, (p<0,001); 4% versus 36,7%, p<0,001), respectively]. The event-free-survival at 8 months was 96% for sirolimus-eluting stents and 86,7% for the thin-strut-thickness stents (p=0,190). CONCLUSIONS: In this study, the use of sirolimus eluting stents in patients with small vessels reduce intimal hyperplasia, in-stent and insegment late-loss, and in-stent and in-segment restenosis in comparison to thin-strut-thickness stents

Epithelial to mesenchymal transition (EMT) is an essential process during embryogenesis for the development of organ systems, including the heart and its vasculature. The development of both coronary vessels and heart valves depends on EMT. In this dissertation, we first present data demonstrating that increasedoligosaccharide hyaluronan (o-HA) levels after EMT induction within atrioventricular (AV) valves leads to a decrease in EMT due to the induction of VEGF expression. Regulated EMT inhibition prevents the formation of hyperplastic valves. Next, we show that the proepicardium, which provides the precursor cells required for epicardial and coronary vessel development, migrates to the developing heart via direct contact of multicellular proepicardial villi to the developing myocardium. This shifts the paradigm from a migration consisting of floating cysts to one of direct contact and differential adhesion forces to form the initial epicardium. A subset of epicardial cells undergoes EMT, migrates into the developing heart, and differentiates into cardiac fibroblast, vascular endothelial, and smooth muscle cells. In order to more effectively study epicardial EMT in vitro, we developed several new methods for the in vitro study of coronary vessel development. We developed an improved protocol for isolating embryonic myocyte cells, for use in co-cultures with epicardial cells. This co-culture system allows investigation into the effects of myocyte derived soluble factors uponepicardial EMT and mesenchymal cell differentiation. We also present a protocol for isolating epicardial clonal colonies from an epicardial cell line derived from the ImmortoMouse. These clones provided direct evidence that the epicardium is a heterogeneous population of cells. These unique clones allow for to study into specific epicardial cell lineages and phenotypes. Finally, we provide data defining the expression of Wnts within the developing heart and the role may play during epicardial EMT. We conclude that canonical Wnts are both necessary and sufficient to inhibit epicardial EMT. These results provide the first direct evidence for a role of Wnt proteins during coronary vessel development. Collectively our results provide significant advancements in our understanding of EMT regulation during cardiac development.

Thesis (Ph.D.)--University of Adelaide, Dept. of Medicine, 2002. / Amendments inserted at back. "May 2002" Includes bibliographical references (leaves 237-290).

Made available in DSpace on 2015-07-22T20:50:20Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-05-30. Added 1 bitstream(s) on 2015-08-11T03:26:18Z : No. of bitstreams: 1 Publico-13256.pdf: 2482735 bytes, checksum: eaf78b153b33731b3ff4250c42b843ae (MD5) / Introdução: não há consenso na literatura a respeito do impacto que angioplastia com implante de stents teria sobre a função endotelial vasomotriz. Inúmeros trabalhos têm mostrado piora da função endotelial após o implante de stents farmacológicos quando comparados aos stents convencionais. Objetivos: o principal objetivo deste trabalho foi avaliar o impacto dos stents convencionais e do Sirolimus, administrado por via oral, sobre a função endotelial vasomotriz. Material e métodos: foram selecionados pacientes em três grupos: grupo 1 - stent convencional com alta dose de Sirolimus (15 mg de ataque seguidos de 6 mg diariamente por 4 semanas); grupo 2 - stents convencionais com baixa dose de Sirolimus (6 mg de dose de ataque seguidos de 2 mg diariamente por 4 semanas) e grupo 3 - stent convencional sem Sirolimus. Modificações na vasomotricidade foram avaliadas com uso de infusão de acetilcolina intracoronariana em doses crescentes. A função vasomotriz independente do endotélio foi avaliada com uso de nitroglicerina. O segmento alvo de análise foram os 15 mm distais à borda do stent. Este segmento também foi avaliado com ultrassonografia intracoronariana. Resultados: os três grupos apresentavam características clínicas e angiográficas semelhantes. A porcentagem de variação do diâmetro foi semelhante em todos os três grupos nos dois momentos da avaliação (4 horas após implante do stent e aos 8 meses de acompanhamento) (p=0,469). Quatro horas após implante dos stents o segmento alvo apresentou disfunção endotelial leve a moderada caracterizada por vasoconstrição em média de -3% que se manteve aos oito meses e foi semelhante entre os grupos. Não houve correlação do volume de placa pela ultrassonografia com o grau de disfunção endotelial nos dois momentos da avaliação. A função vasomotriz independente do endotélio encontrava-se preservada nos dois momentos da avaliação nos três grupos. Conclusão: a disfunção endotelial foi semelhante entre os grupos nos dois momentos da avaliação e não sofreu alteração com implante dos stents e nem com o uso do Sirolimus oral por 4 semanas. Não houve correlação do volume de placa com disfunção endotelial. / Rationale: There is no consensus regarding the impact of stenting on long-term endothelial function. There have been reports of increased endothelial dysfunction with Sirolimus-eluting stents as compared to bare metal stenting (SC). Objectives: This study aims to assess the impact of SC and the effect of oral Sirolimus on endothelial function. Methods: Forty-five patients were randomized into three groups: SC + high-dose oral Sirolimus (initial dose of 15 mg, followed by 6 mg/day during four weeks); SC + lowdose Sirolimus (6 mg followed by 2 mg daily during four weeks); and SC without Sirolimus. Changes in vasoconstriction or vasodilation in a 15 mm segment starting at the distal stent end in response to acetylcholine and nitroglycerin were assessed by quantitative angiography. Results: The groups had similar angiographic characteristics. The percent variation in diameter in response to acetylcholine was similar in all groups at the two time points (p = 0.469). Four hours after stenting, the target segment presented an endothelial dysfunction that was maintained after eight months in all groups. In all groups, endothelium-independent vasomotion in response to nitroglycerin was similar at four hours and eight months, with increased target segment diameter after nitroglycerin infusion (p = 0.001). Conclusions: The endothelial dysfunction was similarly present at the 15 mm segment distal to the treated segment, at 4 hours and 8 months after stenting. Sirolimus administered orally during 4 weeks to prevent restenosis did not affect the status of endothelium-dependent and independent vasomotion. / TEDE / BV UNIFESP: Teses e dissertações