Vascular Gene Transfer from Metallic Stent Surfaces Using Adenoviral Vectors Tethered through Hydrolysable Cross-linkers

In-stent restenosis presents a major complication of stent-based revascularization procedures widely used to re-establish blood flow through critically ...

The Children's Hospital of Philadelphia, University of Pennsylvania

The cGATA-6 gene is flanked by an enhancer that selectively marks the atrioventricular conduction system (AVCS) in transgenic mice. This enhancer reads anterior/posterior and medial/lateral positional information very early in the cardiogenic program and remains active in progressively more restricted regions of primary myocardium leading up to the emergence of a histologically distinct AVCS. We undertook to parse this enhancer to resolve how the respective AVCS-specific transcription program is regulated at the molecular level. We determined that this AVCS enhancer includes a 102 bp module that is sufficient to restrict expression to primary nonchamber myocardium. This offers a novel tool to analyze the early molecular delineation of primary and chamber myocardium, which subsequently give rise to components of the central and peripheral conduction system, respectively. Furthermore, we show that this 102 bp module in turn contains a nested 47 bp core module that has the potential to direct expression specifically to the AVCS domain of primary myocardium, albeit with low efficiency. Accordingly, we show that a GATA site and a GC-rich site in the 102 bp region bolster the activity of the nested 47 bp AVCS core region even within the context of the parental 1,478 bp enhancer. These are the first functional elements to be reported for a cardiac conduction system-specific control region.

GATA-6 gene enhancer contains nested regulatory modules for primary myocardium and the embedded nascent atrioventricular conduction system.

&#x80CC;&#x666F;&#x3002;&#x901A;&#x8FC7;&#x805A;&#x96C6;&#x548C; (&#x6216;) &#x89C6;&#x9EC4;&#x9178; (RA) &#x6CBB;&#x7597;&#xFF0C;&#x53EF;&#x4EE5;&#x533A;&#x5206;&#x9F20;&#x6807; (ES) &#x7684;&#x80DA;&#x80CE;&#x5E72;&#x7EC6;&#x80DE;&#x4F53;&#x5916;&#x3002;&#x4E3B;&#x8981;&#x5206;&#x5316;&#x7EC6;&#x80DE;&#x7684;&#x5B9E;&#x9A8C;&#x662F;&#x80DA;&#x5916;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x5C42;&#x3002;Dab2&#x3001; &#x5C42;&#x7C98;&#x8FDE;&#x86CB;&#x767D;&#x3001; GATA4&#x3001; GATA5&#x3001; GATA6 &#x8868;&#x793A;&#x5728;&#x80DA;&#x80CE;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x5C42;&#x548C;&#x5176;&#x6CBF;&#x88AD;&#x627F;&#x8BFA;&#x4E2D;&#x53D1;&#x6325;&#x5173;&#x952E;&#x4F5C;&#x7528;&#x3002;&#x7ED3;&#x679C;&#x3002;&#x6211;&#x4EEC;&#x53D1;&#x73B0;&#x5728;&#x6CA1;&#x6709; GATA4 &#x6216; GATA5&#xFF0C;RA &#x8BF1;&#x5BFC;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x5C42;&#x5206;&#x5316;&#x7684;&#x80DA;&#x80CE;&#x5E72;&#x7EC6;&#x80DE;&#x51CF;&#x5C11;&#x3002;GATA4 (-/-) ES &#x7EC6;&#x80DE;&#x8868;&#x8FBE;&#x9AD8;&#x6C34;&#x5E73;&#x7684;&#x5185;&#x810F;&#x5185;&#x80DA;&#x5C42;&#x6CBF;&#x88AD;&#x7684; GATA5&#x3001; GATA6&#x3001; &#x548C;&#x809D;&#x7EC6;&#x80DE;&#x6838;&#x56E0;&#x5B50; 4 alpha &#x6807;&#x8BB0;&#x3002;GATA5 (-/-) ES &#x7EC6;&#x80DE;&#x8868;&#x8FBE;&#x7532;&#x80CE;&#x6807;&#x8BB0;&#x7684;&#x65E9;&#x671F;&#x809D;&#x53D1;&#x5C55;&#x7684;&#x66F4;&#x9AD8;&#x6C34;&#x5E73;&#x3002;GATA6 (-/-) ES &#x7EC6;&#x80DE;&#x8868;&#x8FBE;&#x9AD8;&#x6C34;&#x5E73;&#x7684; GATA5 &#x548C;&#x4E2D;&#x80DA;&#x5C42;&#x4EE5;&#x53CA;&#x5FC3;&#x808C;&#x7EC6;&#x80DE;&#x6807;&#x8BB0;&#x7684;&#x80F6;&#x539F;&#x4E09; &alpha;-1 &#x548C; tropomyosin1 alpha&#x3002;&#x56E0;&#x6B64;&#xFF0C;&#x5220;&#x9664; GATA6 &#x6392;&#x9664;&#x5185;&#x80DA;&#x5C42;&#x5206;&#x5316;&#xFF0C;&#x4F46;&#x63A8;&#x52A8;&#x4E2D;&#x80DA;&#x5C42;&#x5B97;&#x65CF;&#x3002;&#x7ED3;&#x8BBA;&#x3002;GATA4&#x3001; GATA5 &#x548C; GATA6 &#x4F20;&#x8FBE;&#x4E00;&#x79CD;&#x72EC;&#x7279;&#x7684;&#x57FA;&#x56E0;&#x8868;&#x8FBE;&#x6A21;&#x5F0F;&#x548C; ES &#x7EC6;&#x80DE;&#x5206;&#x5316;&#x7684;&#x5F71;&#x54CD;&#x3002;&#x6211;&#x4EEC;&#x8868;&#x660E;&#x907F;&#x514D;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x5C42;&#x5206;&#x5316;&#x5E76;&#x91C7;&#x53D6;&#x72EC;&#x7279;&#x7684;&#x5B97;&#x65CF;&#x4F53;&#x5916;&#x901A;&#x8FC7;&#x8C03;&#x8282;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#x56E0;&#x7D20;&#x53EF;&#x4EE5;&#x6307;&#x793A;&#x80DA;&#x80CE;&#x5E72;&#x7EC6;&#x80DE;&#x3002;&#x8FD9;&#x4E00;&#x53D1;&#x73B0;&#x63D0;&#x4F9B;&#x4E86;&#x6F5C;&#x5728;&#x7684;&#x529E;&#x6CD5;&#xFF0C;&#x4EA7;&#x751F;&#x7406;&#x60F3;&#x7684;&#x5355;&#x5143;&#x683C;&#x7C7B;&#x578B;&#x4ECE;&#x80DA;&#x80CE;&#x5E72;&#x7EC6;&#x80DE;&#xFF0C;&#x7528;&#x4E8E;&#x518D;&#x751F;&#x7EC6;&#x80DE;&#x7597;&#x6CD5;&#x3002;

&#x901A;&#x8FC7;&#x8C03;&#x8282;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#x8F6C;&#x5F55;&#x56E0;&#x5B50;&#x5728;&#x5C0F;&#x9F20;&#x80DA;&#x80CE;&#x5E72;&#x7EC6;&#x80DE;&#x7684;&#x5206;&#x5316;&#x6F5C;&#x80FD;&#x7684;&#x53D8;&#x5316;&#x3002;

Background. Mouse embryonic stem (ES) cells can be differentiated in vitro by aggregation and/or retinoic acid (RA) treatment. The principal differentiation lineage in vitro is extraembryonic primitive endoderm. Dab2, Laminin, GATA4, GATA5, and GATA6 are expressed in embryonic primitive endoderm and play critical roles in its lineage commitment. Results. We found that in the absence of GATA4 or GATA5, RA-induced primitive endoderm differentiation of ES cells was reduced. GATA4 (-/-) ES cells express higher level of GATA5, GATA6, and hepatocyte nuclear factor 4 alpha marker of visceral endoderm lineage. GATA5 (-/-) ES cells express higher level of alpha fetoprotein marker of early liver development. GATA6 (-/-) ES cells express higher level of GATA5 as well as mesoderm and cardiomyocyte markers which are collagen III alpha-1 and tropomyosin1 alpha. Thus, deletion of GATA6 precluded endoderm differentiation but promoted mesoderm lineages. Conclusions. GATA4, GATA5, and GATA6 each convey a unique gene expression pattern and influences ES cell differentiation. We showed that ES cells can be directed to avoid differentiating into primitive endoderm and to adopt unique lineages in vitro by modulating GATA factors. The finding offers a potential approach to produce desirable cell types from ES cells, useful for regenerative cell therapy.

Alteration of Differentiation Potentials by Modulating GATA Transcription Factors in Murine Embryonic Stem Cells.

Contexte. Souris souches embryonnaires) peuvent &#xEA;tre diff&#xE9;renci&#xE9;s les cellules in vitro par l&#x26;#39;agr&#xE9;gation et / ou de l&#x26;#39;acide r&#xE9;tino&#xEF;que (RA) de traitement. La lign&#xE9;e de diff&#xE9;renciation in vitro est principale extra-embryonnaire endoderme primitif. DAB2, la laminine, GATA4, GATA5, et GATA6 sont exprim&#xE9;s en endoderme primitif embryonnaire et jouent un r&#xF4;le critique dans la lign&#xE9;e de son engagement. R&#xE9;sultats. Nous avons constat&#xE9; que, en l&#x26;#39;absence de GATA4 ou GATA5, RA-diff&#xE9;renciation induite par endoderme primitif des cellules souches embryonnaires a &#xE9;t&#xE9; r&#xE9;duit. GATA4 (- / -) des cellules ES exprimer plus haut niveau de GATA5, GATA6, et des h&#xE9;patocytes marqueur nucl&#xE9;aire facteur alpha 4 de la lign&#xE9;e endoderme visc&#xE9;ral. GATA5 (- / -) des cellules ES exprimer niveau plus &#xE9;lev&#xE9; de marqueur alpha foetoprot&#xE9;ine de d&#xE9;veloppement du foie d&#xE9;but. GATA6 (- / -) des cellules ES exprimer niveau plus &#xE9;lev&#xE9; de GATA5 ainsi que m&#xE9;soderme et des marqueurs de cardiomyocytes, qui sont du collag&#xE8;ne III alpha-1 et alpha tropomyosin1. Ainsi, la suppression de la diff&#xE9;renciation GATA6 endoderme &#xE9;cart&#xE9;e, mais les lign&#xE9;es m&#xE9;soderme promus. Conclusions. GATA4, GATA5, et GATA6 chaque v&#xE9;hiculer un mod&#xE8;le g&#xE9;n&#xE9;tique unique d&#x26;#39;expression et influence la diff&#xE9;renciation des cellules ES. Nous avons montr&#xE9; que les cellules ES peuvent &#xEA;tre adress&#xE9;es &#xE0; &#xE9;viter de se diff&#xE9;rencier en endoderme primitif et d&#x26;#39;adopter des lign&#xE9;es uniques in vitro par des facteurs GATA modulation. La conclusion propose une approche possible de produire les types de cellules souhaitables &#xE0; partir de cellules ES, utiles pour la th&#xE9;rapie cellulaire r&#xE9;g&#xE9;n&#xE9;ratrice.

Alt&#xE9;ration des potentiels de diff&#xE9;renciation en modulant les facteurs de transcription GATA dans cellules souches embryonnaires murines

Hintergrund. M&auml;useder embryonalen Stammzellen (ES) k&ouml;nnen in-vitro durch Aggregation bzw. Retinoic S&auml;ure (RA) Behandlung unterschieden werden. Die wichtigsten Differenzierung-&Uuml;bertragungslinie in-vitro ist extraembryonic primitive Endodermis. Dab2, Laminin, GATA4, GATA5 und GATA6 in embryonalen primitiven Endodermis ausgedr&uuml;ckt und spielen wichtige Rollen in ihrem Herkunfts-Engagement. Ergebnisse. Wir fanden, dass in Ermangelung GATA4 oder GATA5, primitive Entoderm induziert durch RA Differenzierung von ES-Zellen reduziert wurde. GATA4 (- / -) ES Zellen express h&ouml;heres Ma&szlig; an GATA5, GATA6 und Hepatozyten nuklearer Faktor 4 alpha Marker der viszeralen Endodermis-Linie. GATA5 (- / -) ES Zellen express h&ouml;heren alpha-Fetoprotein-Marker f&uuml;r die fr&uuml;he Entwicklung der Leber. GATA6 (- / -) ES Zellen express h&ouml;heres Ma&szlig; an GATA5 und Mesoderm sowie Cardiomyocyte Markern Kollagen III Alpha-1 und tropomyosin1 Alpha sind. So l&ouml;schen von GATA6 ausgeschlossen Endodermis Differenzierung aber Mesoderm &Uuml;bertragungslinien gef&ouml;rdert. Schlussfolgerungen. GATA4, GATA5 und GATA6 vermitteln ein einzigartiges gen Ausdrucksmuster und ES Zelldifferenzierung beeinflusst. Wir haben gezeigt, dass ES-Zellen zu vermeiden, Differenzierung in primitiven Endodermis und in-vitro-einzigartige &Uuml;bertragungslinien erlassen durch Modulation GATA Faktoren gelenkt werden k&ouml;nnen. Die Feststellung bietet einen m&ouml;glichen Ansatz um w&uuml;nschenswert Zelltypen von ES-Zellen, n&uuml;tzlich f&uuml;r regenerative Zelltherapie zu produzieren.

&Auml;nderung der Differenzierung Potenziale durch Modulation GATA Transkriptionsfaktoren in murinen embryonalen Stammzellen.

Sullo sfondo. Le cellule staminali embrionali (ES) del mouse possono essere differenziate in vitro di aggregazione e/o retinoico trattamento acido (RA). Il lignaggio principale differenziazione in vitro &egrave; extraembryonic endoderma primitivo. Dab2, laminina, GATA4, GATA5 e GATA6 sono espressi in embrionale endoderma primitivo e svolgono un ruolo critico nel suo impegno di lignaggio. Risultati. Abbiamo trovato che, in assenza di GATA4 o GATA5, &egrave; stata ridotta differenziazione endoderma primitivo RA-indotta delle cellule di ES. GATA4 (- / -) ES cellule esprimono il pi&ugrave; alto livello di GATA5, GATA6 e del hepatocyte nuclear factor 4 alfa marcatore del lignaggio endoderma viscerale di. GATA5 (- / -) cellule ES express livello superiore del marcatore di alfa fetoproteina di sviluppo precoce di fegato. GATA6 (- / -) cellule ES esprimono il livello pi&ugrave; elevato di marcatori GATA5 cos&igrave; come mesoderma e cardiomyocyte che sono collagene III alfa alfa-1 e tropomyosin1. Cos&igrave;, eliminazione di GATA6 precluso differenziazione endoderma ma promosso lignaggi di mesoderma. Conclusioni. GATA4, GATA5 e GATA6 trasmettere un modello di espressione del gene unico e influenza la differenziazione delle cellule di ES. Abbiamo dimostrato che le cellule ES possono essere diretto per evitare differenziando in endoderma primitivo e adottare uniche lignaggi in vitro modulando i fattori di GATA. La scoperta offre un potenziale approccio per produrre i tipi di cella desiderabile da cellule ES, utile per la terapia rigenerativa cellulare.

Alterazione dei potenziali di differenziazione modulando i fattori di trascrizione GATA in cellule staminali embrionali Murine.

&#x80CC;&#x666F;&#x3002;&#x30DE;&#x30A6;&#x30B9;&#x80DA;&#x6027;&#x5E79;&#xFF08;ES&#xFF09;&#x7D30;&#x80DE;&#x306F;&#x3001;&#x51DD;&#x96C6;&#x304A;&#x3088;&#x3073;/&#x307E;&#x305F;&#x306F;&#x30EC;&#x30C1;&#x30CE;&#x30A4;&#x30F3;&#x9178;&#xFF08;RA&#xFF09;&#x6CBB;&#x7642;&#x306B;&#x3088;&#x3063;&#x3066;in vitro&#x3067;&#x5206;&#x5316;&#x3055;&#x305B;&#x308B;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x308B;&#x3002; in vitro&#x3067;&#x306E;&#x4E3B;&#x8981;&#x306A;&#x5206;&#x5316;&#x7CFB;&#x7D71;&#x306F;&#x80DA;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x8449;&#x3067;&#x3059;&#x3002; Dab2&#x3001;&#x30E9;&#x30DF;&#x30CB;&#x30F3;&#x3001;GATA4&#x3001;GATA5&#x3001;&#x3068;GATA6&#x306F;&#x3001;&#x80DA;&#x306E;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x8449;&#x306B;&#x767A;&#x73FE;&#x3057;&#x3001;&#x305D;&#x306E;&#x7CFB;&#x7D71;&#x306E;&#x30B3;&#x30DF;&#x30C3;&#x30C8;&#x30E1;&#x30F3;&#x30C8;&#x3067;&#x91CD;&#x8981;&#x306A;&#x5F79;&#x5272;&#x3092;&#x679C;&#x305F;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x7D50;&#x679C;&#x3002;&#x6211;&#x3005;&#x306F;&#x3001;GATA4&#x307E;&#x305F;&#x306F;GATA5&#x304C;&#x5B58;&#x5728;&#x3057;&#x306A;&#x3044;&#x5834;&#x5408;&#x306B;&#x3001;ES&#x7D30;&#x80DE;&#x306E;RA-&#x8A98;&#x5C0E;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x8449;&#x3078;&#x306E;&#x5206;&#x5316;&#x304C;&#x6E1B;&#x5C11;&#x3057;&#x305F;&#x3053;&#x3068;&#x304C;&#x308F;&#x304B;&#x3063;&#x305F;&#x3002; GATA4&#xFF08; -  /  - &#xFF09;ES&#x7D30;&#x80DE;&#x306F;GATA5&#x306E;&#x30EC;&#x30D9;&#x30EB;&#x304C;&#x9AD8;&#x3044;&#x307B;&#x3069;&#x3001;GATA6&#x3001;&#x5185;&#x81D3;&#x5185;&#x80DA;&#x8449;&#x7CFB;&#x7D71;&#x306E;&#x809D;&#x7D30;&#x80DE;&#x6838;&#x56E0;&#x5B50;4&#x30A2;&#x30EB;&#x30D5;&#x30A1;&#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x3092;&#x767A;&#x73FE;&#x3059;&#x308B;&#x3002; GATA5&#xFF08; -  /  - &#xFF09;ES&#x7D30;&#x80DE;&#x306F;&#x3001;&#x521D;&#x671F;&#x306E;&#x809D;&#x81D3;&#x958B;&#x767A;&#x306E;&#x30A2;&#x30EB;&#x30D5;&#x30A1;&#x30D5;&#x30A7;&#x30C8;&#x30D7;&#x30ED;&#x30C6;&#x30A4;&#x30F3;&#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x306E;&#x9AD8;&#x3044;&#x30EC;&#x30D9;&#x30EB;&#x3092;&#x8868;&#x73FE;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002; GATA6&#xFF08; -  /  - &#xFF09;ES&#x7D30;&#x80DE;&#x306F;&#x3001;&#x3088;&#x308A;&#x9AD8;&#x3044;GATA5&#x306E;&#x30EC;&#x30D9;&#x30EB;&#x3060;&#x3051;&#x3067;&#x306A;&#x304F;&#x3001;&#x4E2D;&#x80DA;&#x8449;&#x3001;&#x30B3;&#x30E9;&#x30FC;&#x30B2;&#x30F3;III&#x3001;&#x3B1;-1&#x3001;tropomyosin1&#x30A2;&#x30EB;&#x30D5;&#x30A1;&#x3067;&#x3042;&#x308A;&#x5FC3;&#x7B4B;&#x7D30;&#x80DE;&#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x3092;&#x767A;&#x73FE;&#x3059;&#x308B;&#x3002;&#x3057;&#x305F;&#x304C;&#x3063;&#x3066;&#x3001;GATA6&#x6392;&#x9664;&#x3055;&#x308C;&#x5185;&#x80DA;&#x8449;&#x3078;&#x306E;&#x5206;&#x5316;&#x304C;&#x4FC3;&#x9032;&#x3055;&#x80DA;&#x8449;&#x7CFB;&#x7D71;&#x306E;&#x524A;&#x9664;&#x3002;&#x7D50;&#x8AD6;&#x3002; GATA4&#x3001;GATA5&#x3001;&#x3068;GATA6&#x305D;&#x308C;&#x305E;&#x308C;&#x304C;&#x30E6;&#x30CB;&#x30FC;&#x30AF;&#x306A;&#x907A;&#x4F1D;&#x5B50;&#x767A;&#x73FE;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3092;&#x4F1D;&#x3048;&#x308B;&#x3068;&#x3001;ES&#x7D30;&#x80DE;&#x306E;&#x5206;&#x5316;&#x306B;&#x5F71;&#x97FF;&#x3092;&#x4E0E;&#x3048;&#x307E;&#x3059;&#x3002;&#x6211;&#x3005;&#x306F;&#x3001;ES&#x7D30;&#x80DE;&#x306F;&#x539F;&#x59CB;&#x5185;&#x80DA;&#x8449;&#x3078;&#x306E;&#x5206;&#x5316;&#x3092;&#x907F;&#x3051;&#x308B;&#x305F;&#x3081;&#x306B;&#x3001;&#x5909;&#x8ABF;GATA&#x56E0;&#x5B50;&#x306B;&#x3088;&#x3063;&#x3066;in vitro&#x3067;&#x306E;&#x30E6;&#x30CB;&#x30FC;&#x30AF;&#x306A;&#x7CFB;&#x7D71;&#x3092;&#x63A1;&#x7528;&#x3059;&#x308B;&#x3053;&#x3068;&#x3092;&#x6307;&#x793A;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x305F;&#x3002;&#x767A;&#x898B;&#x306F;&#x3001;&#x518D;&#x751F;&#x7D30;&#x80DE;&#x6CBB;&#x7642;&#x306E;&#x305F;&#x3081;&#x306E;&#x6709;&#x7528;&#x306A;ES&#x7D30;&#x80DE;&#x304B;&#x3089;&#x3001;&#x671B;&#x307E;&#x3057;&#x3044;&#x306E;&#x7D30;&#x80DE;&#x578B;&#x3092;&#x751F;&#x6210;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x6F5C;&#x5728;&#x7684;&#x306A;&#x30A2;&#x30D7;&#x30ED;&#x30FC;&#x30C1;&#x3092;&#x63D0;&#x4F9B;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;

&#x30DE;&#x30A6;&#x30B9;&#x80DA;&#x6027;&#x5E79;&#x7D30;&#x80DE;&#x306B;&#x304A;&#x3051;&#x308B;&#x5909;&#x8ABF;GATA&#x8EE2;&#x5199;&#x56E0;&#x5B50;&#x306B;&#x3088;&#x308B;&#x5206;&#x5316;&#x30DD;&#x30C6;&#x30F3;&#x30B7;&#x30E3;&#x30EB;&#x306E;&#x5909;&#x5316;

&#xBC30;&#xACBD;&#xC785;&#xB2C8;&#xB2E4;. &#xB9C8;&#xC6B0;&#xC2A4; &#xBC30;&#xC544; &#xC904;&#xAE30; (ES&#xC758;) &#xC138;&#xD3EC;&#xB294; &#xC9D1;&#xACC4; &#xB610;&#xB294; retinoic &#xC0B0; (RA) &#xCE58;&#xB8CC;&#xC5D0; &#xC758;&#xD574; &#xCCB4; &#xC678; &#xBD84;&#xD654; &#xB420; &#xC218; &#xC788;&#xB2E4;. &#xCCB4; &#xC678; &#xC8FC; &#xCC28;&#xBCC4;&#xD654; &#xD608;&#xD1B5;&#xC740; extraembryonic &#xAE30;&#xBCF8; endoderm. Dab2, Laminin, GATA4, GATA5, &#xBC0F; GATA6 &#xBC30;&#xC544; &#xC6D0;&#xC2DC;&#xC801;&#xC778; endoderm&#xC5D0;&#xC11C; &#xD45C;&#xD604; &#xD558; &#xACE0; &#xD608;&#xD1B5; &#xB178;&#xB825;&#xC5D0; &#xC911;&#xC694; &#xD55C; &#xC5ED;&#xD560;&#xC744; &#xC7AC;&#xC0DD;&#xD560; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. &#xACB0;&#xACFC;&#xC785;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC;&#xB294; GATA4 &#xB610;&#xB294; gata5&#xC758; &#xBD80;&#xC7AC;, &#xC6D0;&#xC2DC;&#xC801;&#xC778; endoderm RA &#xC720;&#xB3C4; &#xBD84;&#xD654; ES &#xC138;&#xD3EC;&#xC758; &#xAC10;&#xC18C; &#xD588;&#xB2E4; &#xBC1C;&#xACAC;. GATA4 (-/-) ES &#xC138;&#xD3EC; &#xD575; &#xC694;&#xC18C; 4 GATA5, GATA6, &#xBC0F; hepatocyte &#xB0B4;&#xC7A5; endoderm &#xD608;&#xD1B5;&#xC758; &#xC54C;&#xD30C; &#xB9C8;&#xCEE4; &#xB192;&#xC740; &#xC218;&#xC900;&#xC758; &#xD45C;&#xD604;. GATA5 (-/-) ES &#xC138;&#xD3EC; &#xAC04; &#xAC1C;&#xBC1C; &#xCD08;&#xAE30;&#xC758; &#xC54C;&#xD30C; fetoprotein &#xB9C8;&#xCEE4; &#xB192;&#xC740; &#xC218;&#xC900;&#xC758; &#xD45C;&#xD604;. GATA6 (-/-) ES &#xC138;&#xD3EC;&#xB294; &#xCF5C;&#xB77C;&#xAC90; III &#xC54C;&#xD30C; 1&#xACFC; tropomyosin1 &#xC54C;&#xD30C; GATA5 mesoderm &#xBC0F; cardiomyocyte &#xB9C8;&#xCEE4; &#xBCF4;&#xB2E4; &#xB192;&#xC740; &#xC218;&#xC900;&#xC758; &#xD45C;&#xD604;. &#xB530;&#xB77C;&#xC11C;, gata6&#xC758; &#xC0AD;&#xC81C; endoderm &#xCC28;&#xBCC4;&#xD654;&#xB97C; &#xBC30;&#xC81C; &#xD558;&#xC9C0;&#xB9CC; mesoderm &#xACC4;&#xBCF4;&#xB97C; &#xC2B9;&#xC9C4;. &#xACB0;&#xB860;&#xC785;&#xB2C8;&#xB2E4;. GATA4, GATA5, &#xADF8;&#xB9AC;&#xACE0; GATA6 &#xAC01; &#xC804;&#xB2EC;&#xC5D0; &#xACE0;&#xC720;&#xC758; &#xC720;&#xC804;&#xC790; &#xD328;&#xD134; ES &#xC138;&#xD3EC; &#xBD84;&#xD654;&#xC5D0; &#xC601;&#xD5A5;&#xC744; &#xBBF8;&#xCE69;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC;&#xB294; &#xC6D0;&#xC2DC;&#xC801;&#xC778; endoderm&#xC73C;&#xB85C; &#xCC28;&#xBCC4;&#xD654; &#xBC29;&#xC9C0; GATA &#xC694;&#xC18C;&#xB97C; &#xBCC0;&#xC870; &#xD558; &#xC5EC; &#xB3C5;&#xD2B9;&#xD55C; &#xACC4;&#xBCF4; &#xCCB4; &#xC678;&#xB97C; &#xCC44;&#xD0DD; &#xD558; &#xACE0; ES &#xC138;&#xD3EC;&#xB97C; &#xC774;&#xB3D9; &#xD558;&#xC2E4; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4; &#xBCF4;&#xC600;&#xB2E4;. &#xCC3E;&#xB294; ES &#xC138;&#xD3EC;, &#xC7AC;&#xC0DD; &#xC138;&#xD3EC; &#xCE58;&#xB8CC;&#xC5D0; &#xB300; &#xD55C; &#xC720;&#xC6A9;&#xD55C;&#xC5D0;&#xC11C; &#xBC14;&#xB78C;&#xC9C1;&#xD55C; &#xC140; &#xC720;&#xD615;&#xC744; &#xC0DD;&#xC0B0; &#xC7A0;&#xC7AC;&#xC801;&#xC778; &#xC811;&#xADFC;&#xC744; &#xC81C;&#xACF5; &#xD569;&#xB2C8;&#xB2E4;.

GATA Murine &#xBC30;&#xC544; &#xC904;&#xAE30; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; &#xC804;&#xC0AC; &#xC778;&#xC790; &#xBCC0;&#xC870; &#xD558; &#xC5EC; &#xCC28;&#xBCC4;&#xD654; &#xC7A0;&#xC7AC;&#xB825;&#xC758; &#xBCC0;&#xACBD;.

Antecedentes. Madre embrionarias de rat&#xF3;n (ES) las c&#xE9;lulas pueden diferenciarse in vitro mediante la agregaci&#xF3;n y / o &#xE1;cido retinoico (RA) de tratamiento. El linaje de diferenciaci&#xF3;n in vitro es director de endodermo extraembrionario primitivo. Dab2, laminina, GATA4, GATA5 y GATA6 se expresan en el endodermo primitivo embrionario y desempe&#xF1;ar un papel fundamental en su compromiso de linaje. Resultados. Hemos encontrado que, en ausencia de GATA4 o GATA5 AR diferenciaci&#xF3;n inducida endodermo primitivo de c&#xE9;lulas ES se redujo. GATA4 (- / -) de las c&#xE9;lulas madre embrionarias expresan un mayor nivel de GATA5 GATA6, y el factor nuclear de hepatocitos 4 alfa marcador de linaje endodermo visceral. GATA5 (- / -) de las c&#xE9;lulas madre embrionarias expresar un nivel m&#xE1;s alto del marcador alfa-fetoprote&#xED;na en el desarrollo temprano del h&#xED;gado. GATA6 (- / -) de las c&#xE9;lulas madre embrionarias expresan un mayor nivel de GATA5, as&#xED; como mesodermo y los marcadores de cardiomiocitos, que son el col&#xE1;geno III, alfa-1 y alfa tropomyosin1. Por lo tanto, la supresi&#xF3;n de la diferenciaci&#xF3;n del endodermo GATA6 impidi&#xF3;, pero promovidas linajes mesod&#xE9;rmicos. Conclusiones. GATA4, GATA5 y GATA6 cada transmitir un patr&#xF3;n de expresi&#xF3;n g&#xE9;nica &#xFA;nica e influye en la diferenciaci&#xF3;n de c&#xE9;lulas ES. Se demostr&#xF3; que las c&#xE9;lulas ES pueden ser dirigidos a evitar diferenciarse en endodermo primitivo y adoptar linajes &#xFA;nicos in vitro por factores GATA modulaci&#xF3;n. El hallazgo ofrece un enfoque potencial para producir los tipos celulares deseados a partir de c&#xE9;lulas madre embrionarias, &#xFA;tiles para la terapia celular regenerativa.

La alteraci&#xF3;n de los potenciales de diferenciaci&#xF3;n mediante la modulaci&#xF3;n de los factores de transcripci&#xF3;n GATA en c&#xE9;lulas madre embrionarias murinas

The importance of the epicardium for myocardial and valvuloseptal development has been well established; perturbation of epicardial development results in cardiac abnormalities, including thinning of the ventricular myocardial wall and malformations of the atrioventricular valvuloseptal complex. To determine the spatiotemporal contribution of epicardially derived cells to the developing fibroblast population in the heart, we have used a mWt1/IRES/GFP-Cre mouse to trace the fate of EPDCs from embryonic day (ED)10 until birth. EPDCs begin to populate the compact ventricular myocardium around ED12. The migration of epicardially derived fibroblasts toward the interface between compact and trabecular myocardium is completed around ED14. Remarkably, epicardially derived fibroblasts do not migrate into the trabecular myocardium until after ED17. Migration of EPDCs into the atrioventricular cushion mesenchyme commences around ED12. As development progresses, the number of EPDCs increases significantly, specifically in the leaflets which derive from the lateral atrioventricular cushions. In these developing leaflets the epicardially derived fibroblasts eventually largely replace the endocardially derived cells. Importantly, the contribution of EPDCs to the leaflets derived from the major AV cushions is very limited. The differential contribution of EPDCs to the various leaflets of the atrioventricular valves provides a new paradigm in valve development and could lead to new insights into the pathogenesis of abnormalities that preferentially affect individual components of this region of the heart. The notion that there is a significant difference in the contribution of epicardially and endocardially derived cells to the individual leaflets of the atrioventricular valves has also important pragmatic consequences for the use of endocardial and epicardial cre-mouse models in studies of heart development.

Epicardially derived fibroblasts preferentially contribute to the parietal leaflets of the atrioventricular valves in the murine heart.

Magnetic guidance is a physical targeting strategy with the potential to improve the safety and efficacy of a variety of therapeutic agents--including small-molecule pharmaceuticals, proteins, gene vectors, and cells--by enabling their site-specific delivery. The application of magnetic targeting for in-stent restenosis can address the need for safer and more efficient treatment strategies. However, its translation to humans may not be possible without revising the traditional magnetic targeting scheme, which is limited by its inability to selectively guide therapeutic agents to deep localized targets. An alternative two-source strategy can be realized through the use of uniform, deep-penetrating magnetic fields in conjunction with vascular stents included as part of the magnetic setup and the platform for targeted delivery to injured arteries. Studies showing the feasibility of this novel targeting strategy in in-stent restenosis models and considerations in the design of carrier formulations for magnetically guided antirestenotic therapy are discussed in this review.

Magnetically targeted delivery of therapeutic agents to injured blood vessels for prevention of in-stent restenosis.

Gene therapeutic strategies have shown promise in treating vascular disease. However, their translation into clinical use requires pharmaceutical carriers enabling effective, site-specific delivery as well as providing sustained transgene expression in blood vessels. While replication-deficient adenovirus (Ad) offers several important advantages as a vector for vascular gene therapy, its clinical applicability is limited by rapid inactivation, suboptimal transduction efficiency in vascular cells, and serious systemic adverse effects. We hypothesized that novel zinc oleate-based magnetic nanoparticles (MNPs) loaded with Ad would enable effective arterial cell transduction by shifting vector processing to an alternative pathway, protect Ad from inactivation by neutralizing factors, and allow site-specific gene transfer to arteries treated with stent angioplasty using a 2-source magnetic guidance strategy. Ad-loaded MNPs effectively transduced cultured endothelial and smooth muscle cells under magnetic conditions compared to controls and retained capacity for gene transfer after exposure to neutralizing antibodies and lithium iodide, a lytic agent causing disruption of free Ad. Localized arterial gene expression significantly stronger than in control animal groups was demonstrated after magnetically guided MNP delivery in a rat stenting model 2 and 9 d post-treatment, confirming feasibility of using Ad-loaded MNPs to achieve site-specific transduction in stented blood vessels. In conclusion, Ad-loaded MNPs formed by controlled precipitation of zinc oleate represent a novel delivery system, well-suited for efficient, magnetically targeted vascular gene transfer.

Site-specific gene delivery to stented arteries using magnetically guided zinc oleate-based nanoparticles loaded with adenoviral vectors.

The use of arterial stents and other medical implants as a delivery platform for surface immobilized gene vectors allows for safe and efficient localized expression of therapeutic transgenes. In this study we investigate the use of hydrolyzable cross-linkers with distinct kinetics of hydrolysis for delivery of gene vectors from polyallylamine bisphosphonate-modified metal surfaces. Three cross-linkers with the estimated t1/2 of ester bonds hydrolysis of 5, 12 and 50 days demonstrated a cumulative 20%, 39% and 45% vector release, respectively, after 30 days exposure to physiological buffer at 37 °C. Transgene expression in endothelial and smooth muscles cells transduced with substrate immobilized adenovirus resulted in significantly different expression profiles for each individual cross-linker. Furthermore, immobilization of adenoviral vectors effectively extended their transduction effectiveness beyond the initial phase of release. Transgene expression driven by adenovirus-tethered stents in rat carotid arteries demonstrated that a faster rate of cross-linker hydrolysis resulted in higher expression levels at day 1, which declined by day 8 after stent implantation, while inversely, slower hydrolysis was associated with increased arterial expression at day 8 in comparison with day 1. In conclusion, adjustable release of transduction-competent adenoviral vectors from metallic surfaces can be achieved, both in vitro and in vivo, through surface immobilization of adenoviral vectors using hydrolyzable cross-linkers with structure-specific release kinetics.

Adenoviral vector tethering to metal surfaces via hydrolyzable cross-linkers for the modulation of vector release and transduction.

Gene therapy with viral vectors encoding for NOS enzymes has been recognized as a potential therapeutic approach for the prevention of restenosis. Optimal activity of iNOS is dependent on the intracellular availability of L-Arg and BH4 via prevention of NOS decoupling and subsequent ROS formation. Herein, we investigated the effects of separate and combined L-Arg and BH4 supplementation on the production of NO and ROS in cultured rat arterial smooth muscle and endothelial cells transduced with AdiNOS, and their impact on the antirestenotic effectiveness of AdiNOS delivery to balloon-injured rat carotid arteries.

Modulation of NO and ROS production by AdiNOS transduced vascular cells through supplementation with L-Arg and BH4: implications for gene therapy of restenosis.

Richard F. Adamo

Department of Pediatrics,
Division of Cardiology,
Department of Pediatrics, Division of Cardiology

The Children's Hospital of Philadelphia, University of Pennsylvania

Vascular Gene Transfer from Metallic Stent Surfaces Using Adenoviral Vectors Tethered through Hydrolysable Cross-linkers

Richard F. Adamo

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Pediatrics,Division of Cardiology,Department of Pediatrics, Division of Cardiology

The Children's Hospital of Philadelphia, University of Pennsylvania

Vascular Gene Transfer from Metallic Stent Surfaces Using Adenoviral Vectors Tethered through Hydrolysable Cross-linkers

Department of Pediatrics,
Division of Cardiology,
Department of Pediatrics, Division of Cardiology