A Novel <em>Ex vivo</em> Culture Method for the Embryonic Mouse Heart

Developmental studies in the mouse are hampered  by the inaccessibility of the embryo during gestation. Thus, protocols to  isolate and culture individual ...

Isolation of Embryonic Ventricular Endothelial Cells

Cell culture has greatly enhanced our ability to assess individual populations of cells under myriad culture conditions. While immortalized cell lines ...

Protein Isolation from the Developing Embryonic Mouse Heart Valve Region

Western blot analysis is a commonly employed technique for detecting and quantifying protein levels. However, for small tissue samples, this analysis ...

University of North Carolina at Chapel Hill 

The Sonic hedgehog (Shh)-null mouse was initially described as a phenotypic mimic of Tetralogy of Fallot with pulmonary atresia (Washington Smoak, I., Byrd, N.A., Abu-Issa, R., Goddeeris, M.M., Anderson, R., Morris, J., Yamamura, K., Klingensmith, J., and Meyers, E.N. 2005. Sonic hedgehog is required for cardiac outflow tract and neural crest cell development. Dev. Biol. 283, 357-372.); however, subsequent reports describe only a single outflow tract, leaving the phenotype and its developmental mechanism unclear. We hypothesized that the phenotype that occurs in response to Shh knockdown is pulmonary atresia and is directly related to the abnormal development of the secondary heart field. We found that Shh was expressed by the pharyngeal endoderm adjacent to the secondary heart field and that its receptor Ptc2 was expressed in a gradient in the secondary heart field, with the most robust expression in the caudal secondary heart field, closest to the Shh expression. In vitro culture of secondary heart field with the hedgehog inhibitor cyclopamine significantly reduced proliferation. In ovo, cyclopamine treatment before the secondary heart field adds to the outflow tract reduced proliferation only in the caudal secondary heart field, which coincided with the region of high Ptc2 expression. After outflow tract septation should occur, embryos treated with cyclopamine exhibited pulmonary atresia, pulmonary stenosis, and persistent truncus arteriosus. In hearts with pulmonary atresia, cardiac neural crest-derived cells, which form the outflow tract septum, migrated into the outflow tract and formed a septum. However, this septum divided the outflow tract into two unequal sized vessels and effectively closed off the pulmonary outlet. These experiments show that Shh is necessary for secondary heart field proliferation, which is required for normal pulmonary trunk formation, and that embryos with pulmonary atresia have an outflow tract septum.

Sonic hedgehog maintains proliferation in secondary heart field progenitors and is required for normal arterial pole formation.

La Sonic hedgehog (Shh)-souris nulle a &eacute;t&eacute; initialement d&eacute;crite comme un imitateur ph&eacute;notypique de la t&eacute;tralogie de Fallot avec l&#39;atr&eacute;sie pulmonaire (Washington Smoak, I., Byrd, N.A., Abu-Issa, R., Goddeeris, M.M., Anderson, r., Morris, J., Yamamura, K., Klingensmith, J. et Meyers, E.N. 2005. Sonic hedgehog est n&eacute;cessaire pour le d&eacute;veloppement des cellules cardiaques de l&#39;&eacute;coulement des voies et de la cr&ecirc;te neurale. Biol d&eacute;v. 283, 357-372.) ; Cependant, les rapports suivants d&eacute;crivent seulement une &eacute;tendue unique sortie, laissant le ph&eacute;notype et son m&eacute;canisme de perfectionnement pas clair. Nous avons &eacute;mis l&#39;hypoth&egrave;se que le ph&eacute;notype qui survient en r&eacute;ponse &agrave; Shh knockdown est l&#39;atr&eacute;sie pulmonaire et est directement li&eacute; au d&eacute;veloppement anormal du champ cardiaque secondaire. Nous avons trouv&eacute; que Shh est exprim&eacute;e par l&#39;endoderme pharyngien adjacent au champ secondaire cardiaque et que son r&eacute;cepteur Ptc2 est exprim&eacute;e dans un d&eacute;grad&eacute; dans le domaine cardiaque secondaire, avec l&#39;expression plus robuste dans le domaine de la caudale cardiaque secondaire, plus proche de l&#39;expression de Shh. Culture in vitro du champ secondaire coeur avec le h&eacute;risson inhibiteur cyclopamine a r&eacute;duit significativement la prolif&eacute;ration. In ovo, cyclopamine traitement avant le champ cardiaque secondaire ajoute des voies d&#39;&eacute;coulement r&eacute;duit la prolif&eacute;ration seulement dans le domaine de la caudale cardiaque secondaire, qui a co&iuml;ncid&eacute; avec la r&eacute;gion de haute expression Ptc2. Apr&egrave;s que la sortie des voies septation survenait, embryons trait&eacute;s avec cyclopamine expos&eacute; l&#39;atr&eacute;sie pulmonaire, st&eacute;nose pulmonaire et persistants truncus arteriosus. Dans les coeurs avec l&#39;atr&eacute;sie pulmonaire, cardiaques cellules d&eacute;riv&eacute;es de cr&ecirc;te neurale, qui forment le septum de voies de sortie, a migr&eacute; dans les voies d&#39;&eacute;coulement et forment un septum. Toutefois, cette cloison divise les voies de sortie en deux navires de tailles in&eacute;gales et effectivement ferm&eacute; la sortie pulmonaire. Ces exp&eacute;riences d&eacute;montrent que Shh est n&eacute;cessaire pour la prolif&eacute;ration de champ cardiaque secondaire, qui est requise pour la formation du tronc pulmonaire normale, et que les embryons avec l&#39;atr&eacute;sie pulmonaire ont un septum de voies de sortie.

Sonic hedgehog maintient la prolif&eacute;ration en prog&eacute;niteurs champ secondaire coeur et est requis pour la formation de p&ocirc;le art&eacute;riel normal.

Il Sonic hedgehog (Shh)-mouse null &egrave; stato inizialmente descritto come un mimic fenotipica della tetralogia di Fallot con atresia polmonare (Washington Smoak, I., Byrd, N.A., Abu-Issa, R., Goddeeris, M.M., Anderson, r., Morris, J., Yamamura, K., Klingensmith, J. e Meyers, E.N. 2005. Sonic hedgehog &egrave; necessaria per lo sviluppo delle cellule del tratto e cresta neurale efflusso cardiaco. Dev Biol 283, 357-372.); Tuttavia, le relazioni successive descrivono solo un tratto di efflusso del singolo, lasciando poco chiaro il fenotipo e il meccanismo dello sviluppo. Abbiamo ipotizzato che il fenotipo che si verifica in risposta all&#39;atterramento di Shh &egrave; atresia polmonare ed &egrave; direttamente correlato allo sviluppo anormale del campo cuore secondario. Abbiamo trovato che Shh &egrave; stata espressa dall&#39;endoderma faringeo adiacente al campo di cuore secondario e che il recettore Ptc2 &egrave; stata espressa in una sfumatura nel campo secondario di cuore, con l&#39;espressione pi&ugrave; robusto nel campo caudale cuore secondario, pi&ugrave; vicino all&#39;espressione di Shh. Coltura in vitro del campo del cuore secondario con il riccio inibitore cyclopamine significativamente ridotta proliferazione. In ovo, cyclopamine trattamento prima che il campo del cuore secondaria aggiunge il tratto di efflusso ridotta proliferazione solo nel campo caudale cuore secondario, che coincide con la regione di alta espressione Ptc2. Dopo che dovrebbe verificarsi settazione del tratto di efflusso, embrioni trattati con cyclopamine esposto atresia polmonare, stenosi polmonare e persistente Tronco arterioso. Nei cuori con atresia polmonare, cardiache cellule derivate da cresta neurale, che formano il setto del tratto di efflusso, migrarono verso il tratto di efflusso e formarono un setto. Tuttavia, questo setto diviso il tratto di efflusso in due vasi di dimensioni ineguali ed efficacemente chiusi fuori uscita della polmonare. Questi esperimenti dimostrano che Shh &egrave; necessaria per la proliferazione di campo cuore secondario, che &egrave; necessario per la formazione del tronco polmonare normale, e che gli embrioni con atresia polmonare hanno un setto del tratto di efflusso.

Sonic hedgehog mantiene la proliferazione in progenitori campo cuore secondario ed &egrave; necessaria per la formazione del Polo arterioso normale.

Although de la Cruz and colleagues showed as early as 1977 that the outflow tract was added after the heart tube formed, the source of these secondarily added cells was not identified for nearly 25 years. In 2001, three pivotal publications described a secondary or anterior heart field that contributed to the developing outflow tract. This review details the history of the heart field, the discovery and continuing elucidation of the secondarily adding myocardial cells, and how the different populations identified in 2001 are related to the more recent lineage tracing studies that defined the first and second myocardial heart fields/lineages. Much recent work has focused on secondary heart field progenitors that give rise to the myocardium and smooth muscle at the definitive arterial pole. These progenitors are the last to be added to the arterial pole and are particularly susceptible to abnormal development, leading to conotruncal malformations in children. The major signaling pathways (Wnt, BMP, FGF8, Notch, and Shh) that control various aspects of secondary heart field progenitor behavior are discussed.

The role of secondary heart field in cardiac development.

Bien que de la Cruz et ses coll&egrave;gues ont montr&eacute; d&egrave;s 1977 que les voies de sortie a &eacute;t&eacute; ajout&eacute; apr&egrave;s que le tube de coeur form&eacute;, ne constituait pas la source de ces cellules secondairement ajout&eacute;s depuis pr&egrave;s de 25 ans. En 2001, trois publications pivotales d&eacute;crit un champ cardiaque secondaire ou ant&eacute;rieur qui a contribu&eacute; aux voies d&#39;&eacute;coulement en d&eacute;veloppement. Cette d&eacute;tails de revue l&#39;histoire du domaine du coeur, la d&eacute;couverte et l&#39;&eacute;lucidation continue des cellules myocardiques ajoutant secondairement, et comment les diff&eacute;rentes populations identifi&eacute;es en 2001 concernent les &eacute;tudes plus r&eacute;centes de tra&ccedil;age de lign&eacute;e qui d&eacute;finie les premi&egrave;re et deuxi&egrave;me myocardique c&oelig;ur champs/lign&eacute;es. Beaucoup de r&eacute;cent travaux ont port&eacute; sur prog&eacute;niteurs de champ secondaire coeur qui suscitent le myocarde et le muscle lisse au p&ocirc;le art&eacute;riel d&eacute;finitif. Ces cellules souches sont les derniers &agrave; &ecirc;tre ajout&eacute; au p&ocirc;le art&eacute;riel et sont particuli&egrave;rement sensibles &agrave; un d&eacute;veloppement anormal, menant &agrave; conotroncale chez les enfants. Les auteurs discutent les grandes voies de signalisation (Wnt, BMP, FGF8, Notch et Shh) qui contr&ocirc;lent les diff&eacute;rents aspects du comportement de prog&eacute;niteurs cardiaques secondaires champ.

Le r&ocirc;le du champ cardiaque secondaire dans le d&eacute;veloppement cardiaque.

Anche se de la Cruz e colleghi hanno mostrato fin dal 1977 che il tratto di efflusso &egrave; stato aggiunto dopo che il tubo di cuore formata, la fonte di queste cellule secondariamente aggiunte non &egrave; stata identificata per quasi 25 anni. Nel 2001, tre pubblicazioni pivotal descritto un campo secondario o anteriore il cuore che ha contribuito per il tratto di efflusso in via di sviluppo. Questa recensione dettagli la storia del campo del cuore, la scoperta e delucidazione continua delle cellule del miocardio secondariamente aggiunge, e come le popolazioni differenti identificate nel 2001 sono relativi ai pi&ugrave; recenti studi analisi lignaggio che definito i cuore di primo e secondo del miocardio campi/lignaggi. Lavoro molto recente ha concentrato sui progenitori campo cuore secondario che danno origine al miocardio e muscolo liscio al Polo arterioso definitivo. Questi progenitori sono l&#39;ultimo ad essere aggiunto al Polo arterioso e sono particolarmente sensibili allo sviluppo anormale, portando a troncoconali nei bambini. Vengono discussi i principali percorsi segnalazione (Wnt, BMP, FGF8, tacca e Shh) che controllano vari aspetti del comportamento del progenitore campo cuore secondario.

Il ruolo del campo del cuore secondaria nello sviluppo cardiaco.

A healthy vasculature is an essential component of development and is regulated by different signaling pathways. One of the most critical pathways involved is the vascular endothelial growth factor (VEGF) pathway. Components of this pathway serve as the first marker of primitive endothelial cells and are instrumental in inducing the initial differentiation of endothelial cells and later refining them into either arteries or veins. However, the regulation of VEGF signaling remains a mystery, with most studies focusing on the downstream components of this signaling cascade. New evidence shows that the protein cerebral cavernous malformation 3 (CCM3) is a key regulator of the VEGF pathway, bringing to light a previously unknown component of the VEGF signaling axis and opening the door to an exciting new era of vasculogenic research.

Gyrate: CCM3 dances with a different angiogenic partner.

Un syst&egrave;me vasculaire saine est une composante essentielle du d&eacute;veloppement et est r&eacute;gul&eacute;e par diff&eacute;rentes voies de signalisation. Une des plus importantes voies impliqu&eacute;es est la voie du facteur de croissance endoth&eacute;lial vasculaire (VEGF). Composants de cette voie de servent le premier marqueur des cellules endoth&eacute;liales primitifs et jouent un r&ocirc;le essentiel en induisant la diff&eacute;renciation initiale des cellules endoth&eacute;liales et leur affinage plus tard dans les art&egrave;res ou des veines. Toutefois, la r&eacute;glementation de la signalisation de VEGF reste un myst&egrave;re, avec la plupart des &eacute;tudes portant sur les composants en aval de cette cascade de signalisation. Nouveaux &eacute;l&eacute;ments de preuve montrent que la malformation caverneuse c&eacute;r&eacute;brale prot&eacute;ine 3 (CCM3) est un important r&eacute;gulateur de la voie VEGF, mettant en lumi&egrave;re un &eacute;l&eacute;ment inconnu du VEGF axe de signalisation et ouvrant la voie &agrave; une nouvelle &egrave;re passionnante de recherche vasculogenic.

Tournoyer : CCM3 danse avec un partenaire diff&eacute;rent angiog&eacute;nique.

Un sano sistema vascolare &egrave; un componente essenziale di sviluppo ed &egrave; regolata da diverse vie di segnalazione. Uno dei percorsi pi&ugrave; critici coinvolti &egrave; il percorso del fattore di crescita endoteliale vascolare (VEGF). Componenti di questo pathway serviscono come primo marcatore delle cellule endoteliali primitive e sono strumentali nell&#39;indurre la differenziazione iniziale delle cellule endoteliali e successivamente li raffinazione in vene o arterie. Tuttavia, il regolamento di VEGF segnalazione rimane un mistero, con la maggior parte dei studi concentrandosi sui componenti a valle di questa cascata di segnalazione. Nuove prove mostrano che la malformazione cavernosa cerebrale della proteina 3 (CCM3) &egrave; un regolatore chiave del percorso VEGF, riportando alla luce un componente precedentemente sconosciuti di VEGF segnalazione asse e aprendo le porte a una nuova era emozionante di vasculogenic ricerca.

Gyrate: CCM3 danze con un partner diverso angiogenic.

The endothelium is composed of specialized epithelial cells that line the vasculature, the lymph vessels, and the heart. These endothelial cells are characterized by their stratification and are connected via intercellular junctions that confer specific permeability. Although all endothelium acts as a barrier, considerable heterogeneity exists among different organs and even within vessels. During development, the endothelial cells are specified before they migrate to their final destination, and then they commit to an arterial or venous fate. From the venous endothelial cell population, a subset of cells is further specified as lymphatic endothelium. The endothelium can be highly permeable, as in the lymph vessels, or impenetrable, as in the blood-brain barrier. These differences arise during development and are orchestrated through a series of signaling pathways. This review details how endothelial cells arise and are directed to their specific fate, specifically targeting what differentiates endothelial populations.

Development of the endothelium: an emphasis on heterogeneity.

L&#39;endoth&eacute;lium est compos&eacute;e de cellules &eacute;pith&eacute;liales sp&eacute;cialis&eacute;es qui tapissent les vaisseaux, les vaisseaux lymphatiques et le c&oelig;ur. Ces cellules endoth&eacute;liales sont caract&eacute;ris&eacute;s par leur stratification et sont reli&eacute;s par des jonctions intercellulaires qui conf&egrave;rent une perm&eacute;abilit&eacute; sp&eacute;cifique. Bien que tous les actes de l&#39;endoth&eacute;lium comme une barri&egrave;re, une h&eacute;t&eacute;rog&eacute;n&eacute;it&eacute; consid&eacute;rable existe entre les diff&eacute;rents organes et au sein m&ecirc;me de navires. Au cours du d&eacute;veloppement, les cellules endoth&eacute;liales sont sp&eacute;cifi&eacute;es avant leur migration vers leur destination finale, et alors ils s&#39;engagent &agrave; un destin art&eacute;riel ou veineux. De la population de cellules endoth&eacute;liales veineux, un sous-ensemble de cellules est pr&eacute;cis&eacute; comme l&#39;endoth&eacute;lium lymphatique. L&#39;endoth&eacute;lium peut &ecirc;tre tr&egrave;s perm&eacute;ables, comme dans les vaisseaux lymphatiques ou imp&eacute;n&eacute;trables, comme dans la barri&egrave;re h&eacute;mato - enc&eacute;phalique. Ces diff&eacute;rences surviennent au cours du d&eacute;veloppement et sont orchestr&eacute;s par une s&eacute;rie de voies de signalisation. Cette revue d&eacute;taille comment endoth&eacute;lium surviennent et s&#39;adressent &agrave; leur sort sp&eacute;cifique, ciblant sp&eacute;cifiquement ce qui diff&eacute;rencie les populations endoth&eacute;liales.

D&eacute;veloppement de l&#39;endoth&eacute;lium : mettant l&#39;accent sur l&#39;h&eacute;t&eacute;rog&eacute;n&eacute;it&eacute;.

L&#39;endotelio &egrave; composto da cellule epiteliali specializzate che la linea del sistema vascolare, i vasi linfatici e del cuore. Queste cellule endoteliali sono caratterizzate dalla loro stratificazione e sono collegate tramite giunzioni intercellulari che conferiscono permeabilit&agrave; specifica. Anche se tutti endotelio agisce come una barriera, una notevole eterogeneit&agrave; esistente tra i diversi organi e anche all&#39;interno di vasi. Durante lo sviluppo, le cellule endoteliali sono specificate prima che migrano verso la loro destinazione finale, e quindi eseguire il commit di un destino arterioso o venoso. Dalla popolazione delle cellule endoteliali venose, un sottoinsieme delle cellule &egrave; ulteriormente specificato come endotelio linfatico. L&#39;endotelio pu&ograve; essere altamente permeabile, come i vasi linfatici, o impenetrabile, come la barriera emato - encefalica. Queste differenze sorgono durante lo sviluppo e sono orchestrate attraverso una serie di vie di segnalazione. Questa recensione particolari cellule endoteliali come nascono e sono diretti al loro destino specifico, mira specificamente che cosa differenzia popolazioni endoteliali.

Sviluppo dell&#39;endotelio: l&#39;accento sulla eterogeneit&agrave;.

Sonic hedgehog signaling in the secondary heart field has a clear role in cardiac arterial pole development. In the absence of hedgehog signaling, proliferation is reduced in secondary heart field progenitors, and embryos predominantly develop pulmonary atresia. While it is expected that proliferation in the secondary heart field would be increased with elevated hedgehog signaling, this idea has never been tested. We hypothesized that up-regulating hedgehog signaling would increase secondary heart field proliferation, which would lead to arterial pole defects. In culture, secondary heart field explants proliferated up to 6-fold more in response to the hedgehog signaling agonist SAG, while myocardial differentiation and migration were unaffected. Treatment of chick embryos with SAG at HH14, just before the peak in secondary heart field proliferation, resulted unexpectedly in stenosis of both the aortic and pulmonary outlets. We examined proliferation in the secondary heart field and found that SAG-treated embryos exhibited a much milder increase in proliferation than was indicated by the in vitro experiments. To determine the source of other signaling factors that could modulate increased hedgehog signaling, we co-cultured secondary heart field explants with isolated pharyngeal endoderm or outflow tract and found that outflow tract co-cultures prevented SAG-induced proliferation. BMP2 is made and secreted by the outflow tract myocardium. To determine whether BMP signaling could prevent SAG-induced proliferation, we treated explants with SAG and BMP2 and found that BMP2 inhibited SAG-induced proliferation. In vivo, SAG-treated embryos showed up-regulated BMP2 expression and signaling. Together, these results indicate that BMP signaling from the outflow tract modulates hedgehog-induced proliferation in the secondary heart field.

BMP signaling modulates hedgehog-induced secondary heart field proliferation.

Sonic hedgehog signalisation dans le domaine du c&oelig;ur secondaire a un r&ocirc;le pr&eacute;cis dans le d&eacute;veloppement du p&ocirc;le art&eacute;riel cardiaque. En l&#39;absence de signalisation de h&eacute;risson, prolif&eacute;ration est r&eacute;duite en prog&eacute;niteurs champ secondaire coeur et embryons d&eacute;veloppent principalement l&#39;atr&eacute;sie pulmonaire. Alors qu&#39;il est pr&eacute;vu que la prolif&eacute;ration dans le domaine du c&oelig;ur secondaire augmenterait avec la signalisation de h&eacute;risson &eacute;lev&eacute;e, cette id&eacute;e n&#39;a jamais &eacute;t&eacute; test&eacute;e. Nous avons suppos&eacute; que h&eacute;risson up-r&eacute;gulation de signalisation augmenterait la prolif&eacute;ration de champ cardiaque secondaire, qui conduirait &agrave; des d&eacute;fauts de p&ocirc;le art&eacute;riel. En culture, explants de champ secondaire coeur prolif&eacute;raient jusqu&#39;&agrave; 6 fois plus en r&eacute;ponse &agrave; l&#39;h&eacute;risson signalisation agoniste SAG, tandis que la migration et la diff&eacute;renciation du myocarde ont &eacute;t&eacute; affect&eacute;s. Traitement des embryons de poulet avec SAG &agrave; CM14, juste avant le sommet de la prolif&eacute;ration de c&oelig;ur secondaire de champ, a provoqu&eacute; inopin&eacute;ment st&eacute;nose des deux points de vente aortiques et pulmonaires. Nous avons examin&eacute;e la prolif&eacute;ration dans le domaine du c&oelig;ur secondaire et que les embryons trait&eacute;s SAG montraient une augmentation beaucoup plus douce en prolif&eacute;ration a &eacute;t&eacute; indiqu&eacute;e par les exp&eacute;riences in vitro. Pour d&eacute;terminer la source d&#39;autres facteurs de signalisation qui pourrait moduler h&eacute;risson accrue de signalisation, nous co-culture secondaire coeur explants de champ avec isolement pharyng&eacute;e tract endoderme ou &eacute;coulement et trouvent cette prolif&eacute;ration d&#39;induite par le SAG sortie voies co-cultures emp&ecirc;ch&eacute;s. BMP2 est faite et s&eacute;cr&eacute;t&eacute;e par le myocarde de voies de sortie. Pour d&eacute;terminer si BMP signaling pourrait emp&ecirc;cher la prolif&eacute;ration induite par le SAG, nous trait&eacute;s des explants avec SAG et BMP2 et trouv&eacute; que BMP2 inhibait la prolif&eacute;ration induite par le SAG. Les embryons in vivo, impr&eacute;gn&eacute;es de SAG montrent BMP2 expression r&eacute;gul&eacute;e et signalisation. Ensemble, ces r&eacute;sultats indiquent que le BMP de signalisation dans le tube de sortie module prolif&eacute;ration induite par le h&eacute;risson dans le domaine du c&oelig;ur secondaire.

BMP signaling module prolif&eacute;ration champ cardiaque secondaire induite par le h&eacute;risson.

Sonic hedgehog signaling in campo cuore secondario ha un chiaro ruolo nello sviluppo del Polo arterioso cardiaco. In assenza di hedgehog signaling, proliferazione &egrave; ridotto in progenitori campo cuore secondario ed embrioni sviluppano prevalentemente atresia polmonare. Mentre ci si aspetta che proliferazione nel campo secondario cuore sarebbe essere aumentata con elevata hedgehog signaling, questa idea non &egrave; mai stata testata. Abbiamo ipotizzato che up-regolazione hedgehog signaling aumenterebbe la proliferazione di campo cuore secondario, che porterebbe a difetti Polo arterioso. Nella cultura, cuore secondario campo espianti moltiplicate fino a 6-fold pi&ugrave; in response to the hedgehog signaling agonista SAG, mentre la migrazione e la differenziazione del miocardio sono state influenzate. Trattamento degli embrioni di pulcino con SAG a HH14, appena prima del picco nella proliferazione di campo del cuore secondaria, portato inaspettatamente stenosi delle uscite aortiche e polmonare. Abbiamo esaminato la proliferazione nel campo secondario cuore e ha trovato che embrioni trattati SAG esibito un aumento molto pi&ugrave; mite di proliferazione che &egrave; stato indicato dagli esperimenti in vitro. Per determinare l&#39;origine di altri fattori di segnalazione che potrebbe modulare aumentato hedgehog signaling, abbiamo co-cultured cuore secondario campo espianti con isolato tratto faringeo endoderma o deflusso e trovano quella proliferazione di SAG-indotta di efflusso del tratto co-cultures impedito. BMP2 &egrave; fatto e secreto del miocardio del tratto di efflusso. Per determinare la segnalazione BMP potrebbero impedire la proliferazione indotta da SAG, abbiamo trattato espianti con SAG e BMP2 e trovato che BMP2 inibito la proliferazione indotta da SAG. Embrioni in vivo, SAG-trattate hanno mostrato up-regolato BMP2 espressione e segnalazione. Insieme, questi risultati indicano che BMP segnalazione nel tratto di efflusso modula la proliferazione indotta da riccio nel campo cuore secondario.

Segnalazione BMP modula la proliferazione di campo indotta da riccio cuore secondario.

Prote&iacute;nas morfogen&eacute;ticas &oacute;seas (Bmps) son importantes mediadores de la inflamaci&oacute;n y aterosclerosis, aunque su mecanismo de acci&oacute;n no se entiende completamente. Para entender mejor la contribuci&oacute;n de la Bmp se&ntilde;alizaci&oacute;n v&iacute;a la inflamaci&oacute;n vascular, investigamos el papel de Bmper (Bmp c&eacute;lula endotelial derivado del precursor regulador), un modulador de Bmp extracelular, en un modelo in vivo inducido de la inflamaci&oacute;n y aterosclerosis.

Bmper inhibe la expresi&oacute;n endotelial de mol&eacute;culas de adhesi&oacute;n inflamatoria y protege contra la arterioesclerosis.

&#x9AA8;&#x5F62;&#x6001;&#x53D1;&#x751F;&#x86CB;&#x767D; &#xFF08;Bmps&#xFF09; &#x662F;&#x884C;&#x52A8;&#x7684;&#x708E;&#x75C7;&#x548C;&#x52A8;&#x8109;&#x7CA5;&#x6837;&#x786C;&#x5316;&#xFF0C;&#x91CD;&#x8981;&#x8C03;&#x89E3;&#x4EBA;&#xFF0C;&#x867D;&#x7136;&#x4ED6;&#x4EEC;&#x7684;&#x673A;&#x5236;&#x4ECD;&#x672A;&#x5B8C;&#x5168;&#x7406;&#x89E3;&#x3002;&#x4E3A;&#x4E86;&#x66F4;&#x597D;&#x5730;&#x7406;&#x89E3;&#x6240;&#x4F5C;&#x8D21;&#x732E;&#x7684; Bmp &#x4FE1;&#x53F7;&#x901A;&#x8DEF;&#x5728;&#x8840;&#x7BA1;&#x708E;&#x75C7;&#x53CD;&#x5E94;&#xFF0C;&#x6211;&#x4EEC;&#x8C03;&#x67E5;&#x7684; Bmper (Bmp &#x8840;&#x7BA1;&#x5185;&#x76AE;&#x7EC6;&#x80DE;&#x524D;&#x4F53;&#x6D3E;&#x751F;&#x8C03;&#x8282;&#x5668;) &#x4F5C;&#x7528;&#x7684;&#x80DE;&#x5916;&#x7684; Bmp &#x8C03;&#x5236;&#x5668;&#x3001; &#x708E;&#x75C7;&#x548C;&#x52A8;&#x8109;&#x7CA5;&#x6837;&#x786C;&#x5316;&#x7684;&#x8BF1;&#x5BFC;&#x4F53;&#x5185;&#x6A21;&#x578B;&#x4E2D;&#x3002;

Bmper &#x6291;&#x5236;&#x8840;&#x7BA1;&#x5185;&#x76AE;&#x708E;&#x75C7;&#x7C98;&#x9644;&#x5206;&#x5B50;&#x7684;&#x8868;&#x8FBE;&#x548C;&#x9632;&#x6B62;&#x52A8;&#x8109;&#x7CA5;&#x6837;&#x786C;&#x5316;&#x3002;

Bone morphogenetic proteins (Bmps) are important mediators of inflammation and atherosclerosis, though their mechanism of action is not fully understood. To better understand the contribution of the Bmp signaling pathway in vascular inflammation, we investigated the role of Bmper (Bmp endothelial cell precursor-derived regulator), an extracellular Bmp modulator, in an induced in vivo model of inflammation and atherosclerosis.

Bmper inhibits endothelial expression of inflammatory adhesion molecules and protects against atherosclerosis.

Prot&eacute;ines morphog&eacute;n&eacute;tiques osseuses (PGB) sont des m&eacute;diateurs importants de l&#39;inflammation et l&#39;ath&eacute;roscl&eacute;rose, bien que leur m&eacute;canisme d&#39;action n&#39;est pas enti&egrave;rement comprise. Pour mieux comprendre la contribution de la Bmp signalisation dans l&#39;inflammation vasculaire, nous avons &eacute;tudi&eacute; le r&ocirc;le de Bmper (Bmp endoth&eacute;lial pr&eacute;curseur d&eacute;riv&eacute; r&eacute;gulateur), un modulateur de Bmp extracellulaire, dans un mod&egrave;le in vivo induite par de l&#39;inflammation et l&#39;ath&eacute;roscl&eacute;rose.

BMPER inhibe l&#39;expression endoth&eacute;liale de mol&eacute;cules d&#39;adh&eacute;sion inflammatoires et prot&egrave;ge contre l&#39;ath&eacute;roscl&eacute;rose.

Bone morphogenetic Proteins (Bmps) sind wichtige Mediatoren der Entz&uuml;ndungen und Arteriosklerose, obwohl ihr Wirkmechanismus nicht vollst&auml;ndig verstanden wird. Zum besseren Verst&auml;ndnis den Beitrag von der Bmp Signalisierung Biosyntheseweges in vaskul&auml;ren Entz&uuml;ndung, wir untersucht die Rolle der Bmper (Bmp endothelial Zelle Vorl&auml;ufer abgeleitete Spannungsregler), eine extrazellul&auml;re Bmp-Modulator, in einem induzierten in-vivo-Modell von Entz&uuml;ndungen und Arteriosklerose.

BMPER hemmt endothelial Ausdruck der entz&uuml;ndlichen Adh&auml;sionsmolek&uuml;le und sch&uuml;tzt vor Arteriosklerose.

Proteine morfogenetiche ossee (Bmps) sono importanti mediatori dell&#39;infiammazione e aterosclerosi, anche se il loro meccanismo di azione completamente non &egrave; capito. Per capire meglio il contributo delle Bmp signaling pathway in infiammazione vascolare, abbiamo studiato il ruolo di Bmper (regolatore di derivati da precursori delle cellule endoteliali Bmp), un modulatore Bmp extracellulare, in un modello in vivo indotto della infiammazione e aterosclerosi.

Bmper inibisce l&#39;espressione endoteliale di molecole di adesione infiammatoria e protegge contro l&#39;aterosclerosi.

&#x305D;&#x306E;&#x4F5C;&#x7528;&#x30E1;&#x30AB;&#x30CB;&#x30BA;&#x30E0;&#x306F;&#x5B8C;&#x5168;&#x306B;&#x308F;&#x304B;&#x3089;&#x306A;&#x3044;&#x304C;&#x9AA8;&#x5F62;&#x6210;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA; (bmp &#x5F62;&#x5F0F;) &#x91CD;&#x8981;&#x306A;&#x30E1;&#x30C7;&#x30A3;&#x30A8;&#x30FC;&#x30BF;&#x30FC;&#x306E;&#x708E;&#x75C7;&#x3068;&#x52D5;&#x8108;&#x786C;&#x5316;&#x75C7;&#x3067;&#x3059;&#x3002;&#x6211;&#x3005; &#x306F; Bmper &#xFF08;Bmp &#x5185;&#x76AE;&#x7D30;&#x80DE;&#x524D;&#x99C6;&#x4F53;&#x7531;&#x6765;&#x30EC;&#x30AE;&#x30E5;&#x30EC;&#x30FC;&#x30BF;&#xFF09; &#x306E;&#x5F79;&#x5272;&#x3092;&#x8ABF;&#x3079;&#x305F;&#x30B7;&#x30B0;&#x30CA;&#x30EB;&#x4F1D;&#x9054;&#x7D4C;&#x8DEF;&#x8840;&#x7BA1;&#x708E;&#x75C7;&#x306B; Bmp &#x306E;&#x8CA2;&#x732E;&#x3092;&#x3088;&#x308A;&#x3088;&#x304F;&#x7406;&#x89E3;&#x3059;&#x308B;&#x8A98;&#x8D77; in vivo &#x30E2;&#x30C7;&#x30EB;&#x306B;&#x304A;&#x3051;&#x308B;&#x708E;&#x75C7;&#x3068;&#x52D5;&#x8108;&#x786C;&#x5316;&#x75C7;&#x306E;&#x7D30;&#x80DE;&#x5916;&#x306E; Bmp &#x5909;&#x8ABF;&#x3002;

Bmper &#x708E;&#x75C7;&#x6027;&#x63A5;&#x7740;&#x5206;&#x5B50;&#x306E;&#x5185;&#x76AE;&#x767A;&#x73FE;&#x3092;&#x963B;&#x5BB3;&#x3057;&#x3066;&#x52D5;&#x8108;&#x786C;&#x5316;&#x306B;&#x5BFE;&#x3057;&#x3066;&#x4FDD;&#x8B77;&#x3057;&#x307E;&#x3059;&#x3002;

&#xBF08; &#xC804;&#xCCB4;&#xC801; &#xB2E8;&#xBC31;&#xC9C8; (Bmps) &#xC5FC;&#xC99D;&#xACFC; &#xB3D9;&#xB9E5; &#xACBD;&#xD654; &#xC99D;&#xC758; &#xC911;&#xC694; &#xD55C; &#xC911;&#xC7AC;&#xC790;&#xB294; &#xADF8;&#xB4E4;&#xC758; &#xD589;&#xB3D9;&#xC758; &#xBA54;&#xCEE4;&#xB2C8;&#xC998;&#xC744; &#xC644;&#xC804;&#xD788; &#xC774;&#xD574; &#xB418;&#xC9C0; &#xC54A;&#xC2B5;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC; Bmper (Bmp &#xB0B4; &#xD53C; &#xC138;&#xD3EC; &#xC804;&#xC870; &#xD30C;&#xC0DD; &#xC870;&#xC815;&#xAE30;), &#xC5ED;&#xD560; &#xC870;&#xC0AC; &#xD608;&#xAD00; &#xC5FC;&#xC99D;&#xC758; &#xD1B5;&#xB85C;&#xAC00; &#xC2E0;&#xD638; Bmp&#xC758; &#xAE30;&#xC5EC;&#xB97C; &#xC774;&#xD574; &#xD558;&#xAE30; &#xC704;&#xD574; &#xC5FC;&#xC99D;&#xACFC; &#xB3D9;&#xB9E5; &#xACBD;&#xD654;&#xC758; &#xC720;&#xBC1C;&#xB41C; in vivo &#xBAA8;&#xB378;&#xC5D0;&#xC11C; &#xC138;&#xD3EC; &#xC678; Bmp &#xBCC0;&#xC870;&#xAE30;.

Bmper &#xC5FC;&#xC99D; &#xC131; &#xC811;&#xCC29; &#xBD84;&#xC790;&#xC758; &#xB0B4; &#xD53C; &#xD45C;&#xD604;&#xC744; &#xC5B5;&#xC81C; &#xD558; &#xACE0; &#xB3D9;&#xB9E5; &#xACBD;&#xD654; &#xC99D;&#xC5D0; &#xB300; &#xD55C; &#xBCF4;&#xD638;.

Among the extracellular modulators of Bmp (bone morphogenetic protein) signaling, Bmper (Bmp endothelial cell precursor-derived regulator) both enhances and inhibits Bmp signaling. Recently we found that Bmper modulates Bmp4 activity via a concentration-dependent, endocytic trap-and-sink mechanism.

LRP1-dependent endocytic mechanism governs the signaling output of the bmp system in endothelial cells and in angiogenesis.

Parmi les modulateurs extracellulaires de la signalisation de Bmp (prot&eacute;ine morphog&eacute;n&eacute;tique osseuse), Bmper (r&eacute;gulateur de d&eacute;riv&eacute;s de pr&eacute;curseurs de cellules endoth&eacute;liales Bmp) am&eacute;liore et inhibe la Bmp signaling. R&eacute;cemment, nous avons trouv&eacute; que le Bmper module Bmp4 activit&eacute; via un m&eacute;canisme de pi&egrave;ge et-r&eacute;cepteur d&#39;endocytose, concentration-d&eacute;pendant.

M&eacute;canisme endocytose LRP1 d&eacute;pendant r&eacute;git la sortie de signalisation du syst&egrave;me bmp dans les cellules endoth&eacute;liales et dans l&#39;angiogen&egrave;se.

Tra i modulatori extracellulari della segnalazione Bmp (bone morphogenetic protein), Bmper (regolatore Bmp derivati da precursori di cellule endoteliali) migliora e inibisce la segnalazione Bmp. Recentemente abbiamo scoperto che Bmper modula Bmp4 attivit&agrave; attraverso un meccanismo di trappola e lavello endocitico, concentrazione-dipendente.

LRP1-dipendente endocitico meccanismo governa l&#39;uscita di segnalazione del sistema bmp in cellule endoteliali e l&#39;angiogenesi.

BMPER, an orthologue of Drosophila melanogaster Crossveinless-2, is a secreted factor that regulates bone morphogenetic protein activity in endothelial cell precursors and during early cardiomyocyte differentiation. Although previously described in the heart, the role of BMPER in cardiac development and function remain unknown.

BMPER regulates cardiomyocyte size and vessel density in vivo.

BMPER, un orthologue de Drosophila melanogaster Crossveinless-2, est un facteur s&eacute;cr&eacute;t&eacute; qui r&eacute;gule l&#39;activit&eacute; de prot&eacute;ines morphog&eacute;n&eacute;tiques osseuses en pr&eacute;curseurs des cellules endoth&eacute;liales et au cours de la diff&eacute;renciation pr&eacute;coce de cardiomyocyte. Bien que d&eacute;crite pr&eacute;c&eacute;demment dans le coeur, le r&ocirc;le de BMPER dans la fonction et le d&eacute;veloppement cardiaque restent inconnues.

BMPER r&eacute;gule cardiomyocyte taille et navire densit&eacute; in vivo.

BMPER, un orthologue di Drosophila melanogaster Crossveinless-2, &egrave; un fattore secreta che regola l&#39;attivit&agrave; di proteine morfogenetiche ossee dei precursori delle cellule endoteliali e durante l&#39;inizio del cardiomyocyte differenziazione. Anche se precedentemente descritto nel cuore, il ruolo di BMPER in funzione e sviluppo cardiaco rimangono sconosciute.

BMPER regola cardiomyocyte dimensione vaso densit&agrave; e in vivo.

Angiogenesis plays a key role in cancer progression and correlates with disease aggressiveness and poor clinical outcomes. Affinity ligands discovered by screening phage display random peptide libraries can be engineered to molecularly target tumor blood vessels for noninvasive imaging and early detection of tumor aggressiveness. In this study, we tested the ability of a phage-display-selected peptide sequence recognizing specifically bone marrow- derived pro-angiogenic tumor-homing cells, the QFP-peptide, radiolabeled with 64Cu radioisotope to selectively image tumor vasculature in vivo by positron emission tomography (PET). To prepare the targeted PET tracer we modified QFP-phage with the DOTA chelator and radiolabeled the purified QFP-phage-DOTA intermediate with 64Cu to obtain QFP-targeted radioconjugate with high radiopharmaceutical yield and specific activity. We evaluated the new PET tracer in vivo in a subcutaneous (s.c.) Lewis lung carcinoma (LLC) mouse model and conducted tissue distribution, small animal PET/CT imaging study, autoradiography, histology, fluorescence imaging, and dosimetry assessments. The results from this study show that, in the context of the s.c. LLC immunocompetent mouse model, the QFP-tracer can target tumor blood vessels selectively. However, further optimization of the biodistribution and dosimetry profile of the tracer is necessary to ensure efficient radiopharmaceutical applications enabled by the biological specificity of the QFP-peptide.

Development of a new positron emission tomography tracer for targeting tumor angiogenesis: synthesis, small animal imaging, and radiation dosimetry.

Angiogen&egrave;se joue un r&ocirc;le cl&eacute; dans la progression du cancer et est en corr&eacute;lation avec l&#39;agressivit&eacute; de la maladie et de mauvais r&eacute;sultats cliniques. Ligands affinit&eacute; d&eacute;couvert par criblage al&eacute;atoire de phage display biblioth&egrave;ques peptide peuvent &ecirc;tre con&ccedil;ues pour cibler une tumeur des vaisseaux sanguins pour l&#39;imagerie non invasive et la d&eacute;tection pr&eacute;coce de l&#39;agressivit&eacute; de la tumeur. Dans cette &eacute;tude, nous avons test&eacute; la capacit&eacute; d&#39;une phage-affichage-s&eacute;lectionn&eacute; s&eacute;quence peptidique reconnaissant sp&eacute;cifiquement moelle osseuse-d&eacute;riv&eacute; pro-angiog&eacute;nique tumoral-homing cellules, QFP-peptide, radiomarqu&eacute;e &agrave; 64Cu radio-isotopes &agrave; s&eacute;lectivement image tumeur vascularisation in vivo par positons d&#39;&eacute;mission (TEP). Pour pr&eacute;parer le traceur de PET cibl&eacute;, nous avons modifi&eacute; les QFP-phage avec le ch&eacute;lateur DOTA et radioactif de l&#39;interm&eacute;diaire de QFP-phage-DOTA purifi&eacute; avec 64Cu obtenir cibl&eacute;es QFP radioconjugate avec radiopharmaceutique &agrave; haut rendement et l&#39;activit&eacute; sp&eacute;cifique. Nous avons &eacute;valu&eacute; le nouveau traceur de PET in vivo dans un sous-cutan&eacute;e (s.c.) carcinome pulmonaire de Lewis (LLC) souris distribution tissulaire mod&egrave;le et conduites, petite &eacute;tude d&#39;imagerie animale de la TEP/CT, autoradiographie, histologie, imagerie de fluorescence et des &eacute;valuations de dosim&eacute;trie. Les r&eacute;sultats de cette &eacute;tude d&eacute;montrent que, dans le contexte de la s.c. LLC immunocomp&eacute;tent souris mod&egrave;le, QFP-traceur peut cibler s&eacute;lectivement tumeur des vaisseaux sanguins. Cependant, optimisation du profil biodistribution et dosim&eacute;trie du traceur est n&eacute;cessaire pour assurer des applications efficaces de radiopharmaceutiques activ&eacute;es par la sp&eacute;cificit&eacute; biologique du BAA-peptide.

D&eacute;veloppement d&#39;un nouveau traceur de tomographie par &eacute;mission de positrons pour cibler l&#39;angiogen&egrave;se tumorale : synth&egrave;se, petite imagerie animale et dosim&eacute;trie des rayonnements.

L&#39;angiogenesi svolge un ruolo chiave nella progressione del cancro e correla con aggressivit&agrave; di malattia e scarsi risultati clinici. Ligandi affinit&agrave; scoperto da esposizione dei fagi casuale di screening biblioteche del peptide possono essere progettate per molecolarmente bersaglio vasi sanguigni tumorali per imaging non invasivo e individuazione precoce di aggressivit&agrave; del tumore. In questo studio, abbiamo testato la capacit&agrave; di una peptide phage-display-selezionato sequenza riconoscendo in particolare midollo osseo-derivato pro-angiogenica tumore-homing cellule, QFP-peptide, radiomarcata con radioisotopi 64Cu a selettivamente immagine tumore vascolare in vivo di tomografia ad emissione di positroni (PET). Per preparare il tracciante PET mirato abbiamo modificato QFP-fago con il chelante DOTA e radioattivi intermedi QFP-fago-DOTA purificato con 64Cu di ottenere radioconjugate QFP mirati con alto rendimento radiofarmaci e attivit&agrave; specifiche. Abbiamo valutato il nuovo tracciante PET in vivo in una sottocutanea (s.c.) carcinoma polmonare di Lewis (LLC) mouse distribuzione tissutale modello e condotti, piccolo animale studio di imaging PET/CT, autoradiografia, istologia, imaging in fluorescenza e valutazioni di dosimetria. I risultati di questo studio Visualizza che, nel contesto della s.c. LLC immunitarie mouse modello, QFP-tracciante pu&ograve; bersaglio vasi sanguigni tumorali selettivamente. Tuttavia, ulteriore ottimizzazione del profilo biodistribuzione e dosimetria del tracciante &egrave; necessario garantire efficienti applicazioni radiofarmaci abilitate dalla specificit&agrave; biologica del QFP-peptide.

Sviluppo di un nuovo tracciante di tomografia ad emissione di positroni per designare come bersaglio l&#39;angiogenesi tumorale: sintesi piccoli animali imaging e dosimetria di radiazioni.

The bone morphogenetic protein (BMP) family of proteins has a multitude of roles throughout the body. In embryonic development, BMPs promote endothelial specification and subsequent venous differentiation. The BMP pathway also plays important roles in the adult vascular endothelium, promoting angiogenesis and mediating shear and oxidative stress. The canonical BMP pathway functions through the Smad transcription factors; however, other intracellular signaling cascades can be activated, and receptor complexes beyond the traditional type I and type II receptors add additional layers of regulation. Dysregulated BMP signaling has been linked to vascular diseases including pulmonary hypertension and atherosclerosis. This review addresses recent advances in the roles of BMP signaling in the endothelium and how BMPs affect endothelial dysfunction and human disease.

Laura A. Dyer

McAllister Heart Institute

University of North Carolina at Chapel Hill

A Novel Ex vivo Culture Method for the Embryonic Mouse Heart

Isolation of Embryonic Ventricular Endothelial Cells

Protein Isolation from the Developing Embryonic Mouse Heart Valve Region

Laura A. Dyer

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}McAllister Heart Institute

University of North Carolina at Chapel Hill

A Novel Ex vivo Culture Method for the Embryonic Mouse Heart

Isolation of Embryonic Ventricular Endothelial Cells

Protein Isolation from the Developing Embryonic Mouse Heart Valve Region

McAllister Heart Institute