Production, Characterization and Potential Uses of a 3D Tissue-engineered Human Esophageal Mucosal Model

The incidence of both esophageal adenocarcinoma and its precursor, Barrett&#8217;s Metaplasia, are rising rapidly in the western world. Furthermore ...

University of Sheffield

&#x9274;&#x4E8E;&#x9178;&#x6838;&#x7CD6;&#x6838;&#x9178; &#xFF08;Rna&#xFF09; &#x7684;&#x57FA;&#x672C;&#x7EC6;&#x80DE;&#x89D2;&#x8272;&#x662F;&#x91CD;&#x8981;&#x7684;&#x662F;&#x8981;&#x7406;&#x89E3;&#x5F62;&#x6210;&#x8FD9;&#x4E9B;&#x5206;&#x5B50;&#x7684;&#x4E09;&#x7EF4;&#x7ED3;&#x6784;&#x7684;&#x7A33;&#x5B9A;&#x3002;&#x8FD9;&#x9879;&#x7814;&#x7A76;&#x7684;&#x76EE;&#x7684;&#x662F;&#x8C03;&#x67E5;&#x57FA;&#x4E8E;&#x539F;&#x5B50;&#x529B;&#x663E;&#x5FAE;&#x955C;&#x7684;&#x5355;&#x5206;&#x5B50;&#x529B;&#x8C31;&#x6D4B;&#x91CF;&#x901A;&#x8FC7;&#x7B80;&#x5355;&#x7684; RNA &#x7ED3;&#x6784;&#x89E3;&#x79BB;&#x80FD;&#x6E90;&#x683C;&#x5C40;&#x3002;&#x8FD9;&#x79CD;&#x65B9;&#x6CD5;&#x63D0;&#x4F9B;&#x8BE6;&#x7EC6;&#x4FE1;&#x606F;&#x7684;&#x4F4D;&#x7F6E;&#x548C;&#x76F8;&#x5BF9;&#x9AD8;&#x5EA6;&#x4E0A;&#x7684;&#x5206;&#x79BB;&#xFF0C;&#x4ECE;&#x800C;&#x5F62;&#x6210;&#x914D;&#x5408;&#x7269;&#x540E;&#x76F8;&#x5BF9;&#x52A8;&#x529B;&#x5B66;&#x7A33;&#x5B9A;&#x6027;&#x4FE1;&#x606F;&#x7684;&#x80FD;&#x91CF;&#x969C;&#x788D;&#x3002;&#x6211;&#x4EEC;&#x7684;&#x7814;&#x7A76;&#x7ED3;&#x679C;&#x8868;&#x660E;&#x7B80;&#x5355; dodecamer RNA &#x87BA;&#x65CB;&#x7ECF;&#x5386;&#x7C7B;&#x4F3C;&#x4E8E;&#x4EE5;&#x524D;&#x89C2;&#x5BDF;&#x5230;&#x7684; DNA &#x5BE1;&#x6838;&#x82F7;&#x9178;&#x88AB;&#x8FEB;&#x7684;&#x5206;&#x79BB;&#x8FC7;&#x7A0B;&#x3002;&#x7EB3;&#x5165;&#x544A;&#x8BEB;&#x80C0;&#x6BCD;&#x9898;&#x662F;&#x53D1;&#x73B0;&#x5411;&#x7ED1;&#x5B9A;&#x72B6;&#x6001;&#xFF0C;&#x4ECB;&#x7ECD;&#x66F4;&#x63A5;&#x8FD1;&#x66F4;&#x591A;&#x7684;&#x80FD;&#x91CF;&#x5C4F;&#x969C;&#x548C;&#x8FD8;&#x7834;&#x574F;&#x8FD9;&#x5957;&#x590D;&#x5F0F;&#x516C;&#x5BD3;&#x3002;&#x5728;&#x9541;&#x79BB;&#x5B50;&#x6CA1;&#x6709;&#x5305;&#x542B;&#x6B64;&#x544A;&#x8BEB;&#x80C0;&#x53CC;&#x5DE5;&#x7834;&#x574F;&#x7A33;&#x5B9A;&#xFF0C;&#x5F62;&#x6210;&#x4E00;&#x4E2A;&#x5355;&#x4E00;&#x7684;&#x3001; &#x77ED;&#x7684;&#x53CC;&#x5DE5;&#x3002;&#x8FD9;&#x4E9B;&#x7ED3;&#x679C;&#x663E;&#x793A;&#x80C0;&#x6BCD;&#x9898;&#x5F71;&#x54CD; RNA &#x7684;&#x5F3A;&#x8FEB;&#x5206;&#x79BB;&#x5E76;&#x4EA7;&#x751F;&#x654F;&#x611F;&#x7684;&#x9541;&#x79BB;&#x5B50;&#x5B58;&#x5728;&#x7684;&#x80FD;&#x6E90;&#x683C;&#x5C40;&#x3002;&#x6709;&#x8DA3;&#x7684;&#x662F;&#xFF0C;&#x83B7;&#x5F97;&#x7684;&#x8D44;&#x6599;&#x6BD4;&#x8F83;&#x597D;&#x4E0E;&#x4EE5;&#x524D;&#x6240;&#x62A5;&#x544A;&#x7684;&#x5408;&#x594F;&#x6D4B;&#x91CF;&#xFF0C;&#x8BF4;&#x660E;&#x8FD9;&#x79CD;&#x65B9;&#x6CD5;&#x7684;&#x6F5C;&#x529B;&#xFF0C;&#x4EE5;&#x6539;&#x5584;&#x6211;&#x4EEC;&#x7684; RNA &#x7A33;&#x5B9A;&#x4E0E;&#x89E3;&#x79BB;&#x52A8;&#x529B;&#x5B66;&#x7684;&#x7406;&#x89E3;&#x3002;

&#x5355;&#x5206;&#x5B50;&#x7684; RNA &#x5206;&#x89E3;&#x7684;&#x8C03;&#x67E5;&#x3002;

Given the essential cellular roles for ribonucleic acids (RNAs) it is important to understand the stability of three-dimensional structures formed by these molecules. This study aims to investigate the dissociation energy landscape for simple RNA structures via atomic-force-microscopy-based single-molecule force-spectroscopy measurements. This approach provides details on the locations and relative heights of the energy barriers to dissociation, and thus information upon the relative kinetic stabilities of the formed complexes. Our results indicate that a simple dodecamer RNA helix undergoes a forced dissociation process similar to that previously observed for DNA oligonucleotides. Incorporating a UCU bulge motif is found to introduce an additional energy barrier closer to the bound state, and also to destabilize the duplex. In the absence of magnesium ions a duplex containing this UCU bulge is destabilized and a single, shorter duplex is formed. These results reveal that a bulge motif impacts upon the forced dissociation of RNA and produces an energy landscape sensitive to the presence of magnesium ions. Interestingly, the obtained data compare well with previously reported ensemble measurements, illustrating the potential of this approach to improve our understanding of RNA stability and dissociation kinetics.

Single-molecule investigations of RNA dissociation.

&Eacute;tant donn&eacute; les r&ocirc;les cellulaires essentielles d&#39;acides ribonucl&eacute;iques (ARN), il est important de comprendre la stabilit&eacute; des structures tridimensionnelles form&eacute;es par ces mol&eacute;cules. Cette &eacute;tude vise &agrave; &eacute;tudier le paysage d&#39;&eacute;nergie de dissociation de simples structures d&#39;ARN par des mesures de force-spectroscopie de mol&eacute;cules simples atomic force microscope-dot&eacute;s. Cette approche fournit des d&eacute;tails sur l&#39;emplacement et la hauteur relative des barri&egrave;res d&#39;&eacute;nergie de dissociation et donc des informations sur les stabilit&eacute;s cin&eacute;tiques des complexes form&eacute;s. Nos r&eacute;sultats indiquent qu&#39;une h&eacute;lice simple dod&eacute;cam&egrave;re RNA subit un processus de dissociation forc&eacute;e similaire &agrave; celui observ&eacute; ant&eacute;rieurement pour oligonucl&eacute;otides d&#39;ADN. Incorporant un motif de renflement UCU se trouve &agrave; mettre en place une barri&egrave;re d&#39;&eacute;nergie suppl&eacute;mentaire plus pr&egrave;s &agrave; l&#39;&eacute;tat li&eacute; et aussi &agrave; d&eacute;stabiliser le duplex. En l&#39;absence d&#39;ions magn&eacute;sium un duplex contenant ce renflement UCU est d&eacute;stabilis&eacute; et un duplex unique, plus court est form&eacute;. Ces r&eacute;sultats r&eacute;v&egrave;lent qu&#39;un motif de renflement impacts sur la dissociation forc&eacute;e de l&#39;ARN et produit un paysage d&#39;&eacute;nergie sensible &agrave; la pr&eacute;sence d&#39;ions magn&eacute;sium. Fait int&eacute;ressant, les donn&eacute;es obtenues se comparent bien avec les mesures rapport&eacute;es ant&eacute;rieurement ensemble, illustrant le potentiel de cette approche pour am&eacute;liorer notre compr&eacute;hension de la cin&eacute;tique de stabilit&eacute; et de la dissociation de RNA.

Enqu&ecirc;tes de simple-mol&eacute;cule de dissociation de la RNA.

Einzelmolek&#xFC;l-Untersuchungen von RNA-Dissoziation.

Dato i ruoli cellulari essenziali per acidi ribonucleico (RNA) &egrave; importante capire la stabilit&agrave; di strutture tridimensionali formata da queste molecole. Questo studio si propone di indagare il paesaggio di energia di dissociazione per strutture semplici RNA tramite misurazioni di forza-spettroscopia atomica-forza-microscopia-base singola molecola. Questo approccio fornisce dettagli sulle posizioni e relative altezze delle barriere di energia di dissociazione e quindi informazioni sul relativo stabilit&agrave; cinetico dei complessi formati. I nostri risultati indicano che un semplice dodecamer RNA elica subisce un processo di dissociazione forzata simile a quello osservato in precedenza per oligonucleotidi a DNA. Incorporando un motivo di rigonfiamento UCU &egrave; trovato per introdurre una barriera di energia supplementare vicino alla stato legato e anche di destabilizzare il duplex. In assenza di ioni di magnesio &egrave; destabilizzato un duplex contenenti questo rigonfiamento UCU e un singolo, pi&ugrave; breve duplex &egrave; formato. Questi risultati rivelano che un motivo di rigonfiamento impatti sulla dissociazione forzata del RNA e produce un paesaggio energia sensibile alla presenza di ioni magnesio. &Egrave; interessante notare che, i dati ottenuti confrontare bene con le misure precedentemente riportato ensemble, illustrando le potenzialit&agrave; di questo approccio per migliorare la nostra comprensione della cinetica di stabilit&agrave; e la dissociazione di RNA.

Indagini di singola molecola di dissociazione di RNA.

&#x5FC5;&#x9808;&#x306E;&#x30BB;&#x30EB;&#x30E9;&#x30FC; &#x30ED;&#x30FC;&#x30EB; &#x30EA;&#x30DC;&#x6838;&#x9178; &#xFF08;Rna&#xFF09; &#x3092;&#x4E0E;&#x3048;&#x3089;&#x308C;&#x305F;&#x3053;&#x308C;&#x3089;&#x306E;&#x5206;&#x5B50;&#x306E;&#x4E09;&#x6B21;&#x5143;&#x69CB;&#x9020;&#x306E;&#x5B89;&#x5B9A;&#x6027;&#x3092;&#x7406;&#x89E3;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x91CD;&#x8981;&#x3067;&#x3059;&#x3002;&#x539F;&#x5B50;&#x9593;&#x529B;&#x9855;&#x5FAE;&#x93E1;-&#x7CFB;&#x5358;&#x4E00;&#x5206;&#x5B50;&#x529B;&#x5206;&#x5149;&#x6E2C;&#x5B9A;&#x306B;&#x3088;&#x308B;&#x5358;&#x7D14;&#x306A; RNA &#x69CB;&#x9020;&#x306E;&#x89E3;&#x96E2;&#x30A8;&#x30CD;&#x30EB;&#x30AE;&#x30FC;&#x5730;&#x5F62;&#x3092;&#x8ABF;&#x67FB;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x3053;&#x306E;&#x7814;&#x7A76;&#x3092;&#x76EE;&#x6307;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x3053;&#x306E;&#x30A2;&#x30D7;&#x30ED;&#x30FC;&#x30C1;&#x306E;&#x5834;&#x6240;&#x3068;&#x76F8;&#x5BFE;&#x7684;&#x306A;&#x9AD8;&#x3055;&#x89E3;&#x96E2;&#x3068;&#x3053;&#x306E;&#x3088;&#x3046;&#x306B;&#x5F62;&#x6210;&#x3055;&#x308C;&#x305F;&#x8907;&#x5408;&#x4F53;&#x306E;&#x76F8;&#x5BFE;&#x901F;&#x5EA6;&#x8AD6;&#x7684;&#x5B89;&#x5B9A;&#x6027;&#x306E;&#x8A55;&#x4FA1;&#x6642;&#x306B;&#x60C5;&#x5831;&#x306E;&#x30A8;&#x30CD;&#x30EB;&#x30AE;&#x30FC;&#x969C;&#x58C1;&#x3092;&#x8A73;&#x3057;&#x304F;&#x8AAC;&#x660E;&#x3057;&#x307E;&#x3059;&#x3002;&#x5358;&#x7D14;&#x306A;&#x7A2E;&#x985E; RNA &#x306E;&#x3089;&#x305B;&#x3093;&#x306B;&#x4EE5;&#x524D; DNA &#x306E;&#x30AA;&#x30EA;&#x30B4;&#x30CC;&#x30AF;&#x30EC;&#x30AA;&#x30C1;&#x30C9;&#x306E;&#x89B3;&#x5BDF;&#x306B;&#x4F3C;&#x3066;&#x5F37;&#x5236;&#x89E3;&#x96E2;&#x904E;&#x7A0B;&#x3092;&#x7D4C;&#x308B;&#x304C;&#x793A;&#x5506;&#x3055;&#x308C;&#x307E;&#x3057;&#x305F;&#x3002;UCU &#x30D0;&#x30EB;&#x30B8; &#x30E2;&#x30C1;&#x30FC;&#x30D5;&#x3092;&#x53D6;&#x308A;&#x5165;&#x308C;&#x3066;&#x30D0;&#x30A4;&#x30F3;&#x30C9;&#x72B6;&#x614B;&#x306B;&#x8FD1;&#x3044;&#x8FFD;&#x52A0;&#x30A8;&#x30CD;&#x30EB;&#x30AE;&#x30FC;&#x969C;&#x58C1;&#x3092;&#x7D39;&#x4ECB;&#x3057;&#x3001;&#x4E0D;&#x5B89;&#x5B9A;&#x306B;&#x4E8C;&#x91CD;&#x304C;&#x898B;&#x3064;&#x304B;&#x3063;&#x305F;&#x3002;&#x30DE;&#x30B0;&#x30CD;&#x30B7;&#x30A6;&#x30E0; &#x30A4;&#x30AA;&#x30F3;&#x306E;&#x4E0D;&#x5728;&#x3067;&#x306F;&#x3053;&#x306E; UCU &#x306E;&#x3075;&#x304F;&#x3089;&#x307F;&#x3092;&#x542B;&#x3080;&#x4E8C;&#x91CD;&#x304C;&#x4E0D;&#x5B89;&#x5B9A;&#x306B;&#x306A;&#x308B;&#x3057;&#x3001;&#x77ED;&#x3044;&#x5358;&#x4E00;&#x4E8C;&#x91CD;&#x304C;&#x5F62;&#x6210;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;&#x3053;&#x308C;&#x3089;&#x306E;&#x7D50;&#x679C;&#x3067;&#x306F;&#x3001;&#x81A8;&#x3089;&#x307F;&#x30E2;&#x30C1;&#x30FC;&#x30D5;&#x304C; RNA &#x306E;&#x5F37;&#x5236;&#x89E3;&#x96E2;&#x6642;&#x306B;&#x5F71;&#x97FF;&#x3092;&#x4E0E;&#x3048;&#x308B;&#x3057;&#x3001;&#x30DE;&#x30B0;&#x30CD;&#x30B7;&#x30A6;&#x30E0; &#x30A4;&#x30AA;&#x30F3;&#x306E;&#x5B58;&#x5728;&#x3059;&#x308B;&#x6A5F;&#x5BC6;&#x6027;&#x306E;&#x9AD8;&#x3044;&#x30A8;&#x30CD;&#x30EB;&#x30AE;&#x30FC;&#x306E;&#x98A8;&#x666F;&#x3092;&#x4F5C;&#x308A;&#x51FA;&#x3059;&#x3053;&#x3068;&#x3092;&#x660E;&#x3089;&#x304B;&#x306B;&#x3057;&#x307E;&#x3059;&#x3002;&#x8208;&#x5473;&#x6DF1;&#x3044;&#x3053;&#x3068;&#x306B;&#x3001;&#x5F97;&#x3089;&#x308C;&#x305F;&#x30C7;&#x30FC;&#x30BF;&#x306F; RNA &#x306E;&#x5B89;&#x5B9A;&#x6027;&#x3068;&#x89E3;&#x96E2;&#x306E;&#x53CD;&#x5FDC;&#x901F;&#x5EA6;&#x306E;&#x6211;&#x3005; &#x306E;&#x7406;&#x89E3;&#x3092;&#x6539;&#x5584;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x3053;&#x306E;&#x30A2;&#x30D7;&#x30ED;&#x30FC;&#x30C1;&#x306E;&#x53EF;&#x80FD;&#x6027;&#x3092;&#x793A;&#x3059;&#x4EE5;&#x524D;&#x306B;&#x5831;&#x544A;&#x3057;&#x305F;&#x30A2;&#x30F3;&#x30B5;&#x30F3;&#x30D6;&#x30EB;&#x6E2C;&#x5B9A;&#x3068;&#x3088;&#x304F;&#x6BD4;&#x8F03;&#x3057;&#x307E;&#x3059;&#x3002;

RNA &#x89E3;&#x96E2;&#x306E;&#x4E00;&#x5206;&#x5B50;&#x8ABF;&#x67FB;&#x3002;

Ribonucleic &#xC0B0; (RNAs)&#xC5D0; &#xB300; &#xD55C; &#xD544;&#xC218;&#xC801;&#xC778; &#xC138;&#xD3EC; &#xC5ED;&#xD560;&#xC744; &#xAC10;&#xC548;&#xD560; &#xB54C;&#xC774; &#xBD84;&#xC790;&#xC5D0; &#xC758;&#xD574; &#xD615;&#xC131; &#xB41C; 3 &#xCC28;&#xC6D0; &#xAD6C;&#xC870;&#xBB3C;&#xC758; &#xC548;&#xC815;&#xC131;&#xC744; &#xC774;&#xD574; &#xD558;&#xB294; &#xAC83;&#xC774; &#xC911;&#xC694;&#xC785;&#xB2C8;&#xB2E4;. &#xC774; &#xC5F0;&#xAD6C; &#xC870;&#xC0AC; &#xBD84;&#xB9AC; &#xC5D0;&#xB108;&#xC9C0; &#xD504;&#xB9AC; &#xC6D0;&#xC790; &#xD798; &#xD604;&#xBBF8;&#xACBD;-&#xAE30;&#xBC18; &#xB2E8;&#xC77C; &#xBD84;&#xC790; &#xD798; &#xBD84;&#xAD11;&#xD559; &#xCE21;&#xC815;&#xC744; &#xD1B5;&#xD574; &#xAC04;&#xB2E8;&#xD55C; RNA &#xAD6C;&#xC870;&#xC5D0; &#xB300; &#xD55C; &#xBAA9;&#xD45C;&#xB85C; &#xD55C;&#xB2E4;. &#xC774; &#xBC29;&#xBC95;&#xC740; &#xC704;&#xCE58;&#xC640; &#xBD84;&#xB9AC;, &#xADF8;&#xB9AC;&#xACE0; &#xC774;&#xB807;&#xAC8C; &#xD615;&#xC131;&#xB41C; &#xB41C; &#xB2E8;&#xC9C0;&#xC758; &#xC0C1;&#xB300; &#xC6B4;&#xB3D9; &#xC548;&#xC815;&#xC131;&#xC5D0; &#xB530;&#xB77C; &#xC815;&#xBCF4; &#xC5D0;&#xB108;&#xC9C0; &#xC7A5;&#xBCBD;&#xC758; &#xC0C1;&#xB300; &#xB192;&#xC774;&#xC5D0; &#xC790;&#xC138;&#xD788;&#xB97C; &#xC124;&#xBA85;&#xD569;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC;&#xC758; &#xACB0;&#xACFC; &#xB098;&#xD0C0;&#xB0C5;&#xB2C8;&#xB2E4; &#xAC04;&#xB2E8;&#xD55C; dodecamer RNA &#xB098;&#xC120; DNA oligonucleotides&#xC5D0; &#xB300; &#xD55C; &#xC774;&#xC804;&#xC5D0; &#xAD00;&#xCC30; &#xB41C; &#xBE44;&#xC2B7;&#xD55C; &#xAC15;&#xC81C; &#xBD84;&#xB9AC; &#xACFC;&#xC815;&#xC744; &#xACAA; &#xC2B5;. UCU &#xC19F;&#xC544;&#xC624;&#xB978; &#xBAA8;&#xD2F0;&#xBE0C;&#xB97C; &#xD1B5;&#xD569; &#xD558;&#xB294; &#xAC83;&#xC740; &#xBC14;&#xC778;&#xB529;&#xB41C; &#xC0C1;&#xD0DC;&#xC5D0; &#xAC00;&#xAE4C;&#xC774; &#xCD94;&#xAC00; &#xC5D0;&#xB108;&#xC9C0; &#xBC29; &#xBCBD;&#xC744; &#xC18C;&#xAC1C; &#xD558; &#xACE0; &#xB610;&#xD55C; &#xC774;&#xC911; &#xBD88;&#xC548;&#xC815; &#xD558; &#xAC8C; &#xBC1C;&#xACAC; &#xB41C;&#xB2E4;. &#xB9C8;&#xADF8;&#xB124;&#xC298; &#xC774;&#xC628; &#xACB0;&#xD54D;&#xC5D0;&#xC11C; UCU &#xC19F;&#xC544;&#xC624;&#xB978;&#xC774; &#xD3EC;&#xD568; &#xB41C; &#xC774;&#xC911; &#xBD88;&#xC548;&#xC815; &#xD558; &#xACE0; &#xB2E8;&#xC77C;, &#xC9E7;&#xC740; &#xC774;&#xC911; &#xD615;&#xC131; &#xB429;&#xB2C8;&#xB2E4;. &#xC774; &#xACB0;&#xACFC; &#xACF5;&#xAC1C; &#xC19F;&#xC544;&#xC624;&#xB978; &#xBAA8;&#xD2F0;&#xBE0C; RNA&#xC758; &#xAC15;&#xC81C; &#xBD84;&#xB9AC; &#xC2DC;&#xC5D0; &#xC601;&#xD5A5;&#xC744; &#xC90D;&#xB2C8;&#xB2E4; &#xBC0F; &#xB9C8;&#xADF8;&#xB124;&#xC298; &#xC774;&#xC628;&#xC758; &#xC874;&#xC7AC;&#xC5D0; &#xBBFC;&#xAC10;&#xD55C; &#xC5D0;&#xB108;&#xC9C0; &#xD504;&#xB9AC;&#xB97C; &#xC0DD;&#xC131;. &#xD765;&#xBBF8;&#xB86D;&#xAC8C;&#xB3C4;, &#xD68D;&#xB4DD;&#xB41C; &#xB370;&#xC774;&#xD130; &#xC774;&#xC804;&#xC5D0; &#xBCF4;&#xACE0; &#xB41C; &#xC559;&#xC0C1;&#xBE14; &#xCE21;&#xC815;, RNA &#xBD84;&#xB9AC; &#xC548;&#xC815;&#xC131;&#xACFC; &#xC18D;&#xB3C4; &#xB300; &#xD55C; &#xC6B0;&#xB9AC;&#xC758; &#xC774;&#xD574;&#xB97C; &#xD5A5;&#xC0C1; &#xC2DC;&#xD0A4;&#xAE30; &#xC704;&#xD574;&#xC774; &#xC811;&#xADFC;&#xC758; &#xC7A0;&#xC7AC;&#xB825;&#xC744; &#xBCF4;&#xC5EC;&#xC8FC;&#xB294;&#xC640; &#xC798; &#xBE44;&#xAD50;.

&#xB2E8;&#xC77C; &#xBD84;&#xC790; RNA &#xBD84;&#xB9AC;&#xC758; &#xC870;&#xC0AC;&#xC785;&#xB2C8;&#xB2E4;.

Teniendo en cuenta las funciones celulares esenciales para &aacute;cidos ribonucleicos (ARN) es importante entender la estabilidad de estructuras tridimensionales formadas por estas mol&eacute;culas. Este estudio pretende investigar el panorama de la energ&iacute;a de disociaci&oacute;n para simples estructuras de ARN mediante mediciones at&oacute;mica-fuerza-microscopia basadas en una sola mol&eacute;cula-espectroscopia de fuerza. Este enfoque proporciona detalles sobre los lugares y la altura relativa de las barreras de energ&iacute;a de disociaci&oacute;n y as&iacute; obtener informaci&oacute;n sobre las estabilidades cin&eacute;ticas relativas de los complejos formados. Nuestros resultados indican que una h&eacute;lice simple dodecamer RNA pasa por un proceso de disociaci&oacute;n forzado similar a la observada previamente de oligonucle&oacute;tidos de ADN. Incorporando un motivo de bombeo de la UCU se encuentra a introducir una barrera de energ&iacute;a adicional m&aacute;s cerca al estado consolidado y tambi&eacute;n para desestabilizar el d&uacute;plex. En ausencia de iones de magnesio un d&uacute;plex que contiene este abultamiento de la UCU se desestabiliz&oacute; y se forma un d&uacute;plex &uacute;nico, m&aacute;s corto. Estos resultados revelan que un motivo de bombeo impacta sobre la disociaci&oacute;n forzada de ARN y produce un paisaje energ&iacute;a sensible a la presencia de iones de magnesio. Curiosamente, los datos obtenidos comparan bien con medidas conjunto previamente divulgados, que ilustran el potencial de este enfoque para mejorar nuestra comprensi&oacute;n de la cin&eacute;tica de estabilidad y disociaci&oacute;n de RNA.

Investigaciones de una sola mol&eacute;cula de la disociaci&oacute;n de RNA.

A novel monoclonal antibody was raised by immunization of mice with colorectal tumor cell lines. The fusion was screened by immunohistochemistry for binding to primary colorectal tumors. Subsequent analysis on primary disaggregated colorectal tumors show that the antibody recognizes a cell surface antigen expressed by the majority of colorectal tumors. Antigen characterization has shown that the antibody recognizes a sialyltetraosylceramide but does not bind to GM1, GD1a, GT1b, or sialyl Lewis(X) antigens. Binding to a frozen panel of tumor and normal tissue sections revealed that the antigen was also strongly expressed on esophageal, gastric, and endometrial tumors. Its normal tissue distribution was largely restricted to moderate staining of large intestine. Surprisingly, SC104 antibody directly induces tumor cell death without the need for immune effector cells or complement. This may be related in part to its homophilic binding properties that allow cross-linking of antibody and receptors on the cell surface. Caspase activation can be detected following SC104 treatment of colorectal cells, and cotreatment with caspase inhibitors has been shown to inhibit cell death. This suggests that SC104 induces death by a classic apoptotic pathway. Furthermore, SC104 antibody shows additive killing with complement and 5-fluorouracil/leucovorin in vivo, suggesting a new therapeutic approach for this class of antibodies.

A new anticancer glycolipid monoclonal antibody, SC104, which directly induces tumor cell apoptosis.

&Ouml;sophageale Exposition gegen&uuml;ber Duodenogastro-&ouml;sophageale Refluxate f&uuml;hrt um zu reflux-&Ouml;sophagitis und ist beteiligt an der Entwicklung der Barrett-Metaplasie (BM). NF-&kappa;B Signalisierung in Epithelzellen ist die Aktivierung von Transkriptionsfaktoren, die vermutet, dass zentrale BM Entwicklung, w&auml;hrend NF-&kappa;B Aktivierung in Fibroblasten eine entscheidende Rolle bei der Umgestaltung der Matrix spielt zugeordnet. Unser Ziel war es, die Auswirkungen einer S&auml;ure auf NF-&kappa;B Aktivierung im prim&auml;ren menschlichen &ouml;sophageale Fibroblasten (HOFs) und Prim&auml;r- und verewigt &ouml;sophageale Squames zu studieren und epitheliale und Stromale Wechselwirkungen in der Antwort dieser Zellen zur S&auml;ure zu untersuchen. Prim&auml;re HOFs und &ouml;sophageale Epithelzellen von Prim&auml;r- und verewigt wurden ausgesetzt S&auml;ure (pH 7 - pH 4 &le; 120 min) in Einzel- oder gepulste Behandlungen. Konditionierten Medium aus Epithelzellen, die folgenden sauren Exposition auch Fibroblasten angewendet wurde. Zellviabilit&auml;t wurde von MTT-ESTA bestimmt. NF-&kappa;B Aktivierung wurde durch zellul&auml;re Lokalisierung von NF-&kappa;B/p65 visualisiert durch Immunfluoreszenz bestimmt. Konditioniert Medium aus &ouml;sophageale Epithelzellen, Belichtung, NF-&kappa;B in Fibroblasten, aktiviert, mit einigen inter-patient Variabilit&auml;t pH-Wert 5 pulsierender ausgesetzt, aber diese Bedingungen die NF-&kappa;B in den Epithelzellen selbst nicht direkt aktiviert haben. Bedeutende NF-&kappa;B Aktivierung galt in den Epithelzellen sondern nur mit mehr S&auml;ure und Belichtung Zeiten (pH 4, 60-120 min). Unsere Ergebnisse zeigen, dass die sauren Exposition indirekte Aktivierung stromal Zellen von Epithel-Stroma-Interaktionen f&uuml;hren kann. Dies kann dazu beitragen, die Pathogenese der &ouml;sophageale Krankheiten kann, und die inter-patient Variabilit&auml;t irgendwie zu erkl&auml;ren, warum einige Patienten mit Reflux-&Ouml;sophagitis entwickeln BM und andere nicht.

NF-&kappa;B wird in &ouml;sophageale Fibroblasten als Reaktion auf eine parakrine-Signal generiert durch S&auml;ure-gemachten prim&auml;re &ouml;sophagealen Plattenepithel Zellen aktiviert.

Oesophage exposition &agrave; duodenogastro-&oelig;sophageal &oelig;sophagienne permet une &oelig;sophagite de reflux et est impliqu&eacute;e dans le d&eacute;veloppement de la m&eacute;taplasie de Barrett (BM). Dans les cellules &eacute;pith&eacute;liales de NF-&kappa;B est associ&eacute;e &agrave; l&#39;activation de facteurs de transcription, cens&eacute; &ecirc;tre au centre de d&eacute;veloppement de la BM, tandis que l&#39;activation de NF-&kappa;B dans les fibroblastes joue un r&ocirc;le essentiel dans le remodelage de la matrice. Notre objectif &eacute;tait d&#39;&eacute;tudier les effets de l&#39;exposition &agrave; l&#39;acide sur l&#39;activation de NF-&kappa;B dans les fibroblastes humains oesophagiennes primaires (HOFs) et squames oesophagien primaire et immortalis&eacute;es et d&#39;enqu&ecirc;ter sur toutes les interactions &eacute;pith&eacute;liales/stroma dans la r&eacute;ponse de ces cellules &agrave; l&#39;acide. HOFs primaires et primaires et immortalis&eacute;s les cellules &eacute;pith&eacute;liales &oelig;sophagiennes ont &eacute;t&eacute; expos&eacute;s &agrave; l&#39;acide (pH 7 - pH 4 &le; 120 min) dans les traitements simples ou puls&eacute;s. Milieu conditionn&eacute; par les cellules &eacute;pith&eacute;liales apr&egrave;s exposition acide a &eacute;galement &eacute;t&eacute; appliqu&eacute;e &agrave; des fibroblastes. La viabilit&eacute; cellulaire a &eacute;t&eacute; d&eacute;termin&eacute;e par MTT-ESTA. Activation de NF-&kappa;B a &eacute;t&eacute; d&eacute;termin&eacute;e par la localisation cellulaire de NF-&kappa;B/p65 visualis&eacute;e par immunofluorescence. Conditionn&eacute; milieu &agrave; partir de cellules &eacute;pith&eacute;liales &oelig;sophagiennes, soumis &agrave; pH 5 pulsatile exposition, activ&eacute; NF-&kappa;B dans les fibroblastes, avec une certaine variabilit&eacute; inter-patient, mais ces conditions n&#39;activent pas directement de NF-&kappa;B dans les cellules &eacute;pith&eacute;liales eux-m&ecirc;mes. Activation de NF-&kappa;B significative a &eacute;t&eacute; observ&eacute;e dans les cellules &eacute;pith&eacute;liales, mais seulement avec grandes fois d&#39;acidit&eacute; et de l&#39;exposition (pH 4, 60-120 mn). Nos r&eacute;sultats montrent qu&#39;exposition &agrave; l&#39;acide peut causer une activation indirecte des cellules stromales par des interactions &eacute;pith&eacute;liales stromales. Cela peut contribuer &agrave; la pathogen&egrave;se des maladies &oelig;sophagiennes, et la variabilit&eacute; inter-patient peut aller de quelque fa&ccedil;on d&#39;expliquer pourquoi certains patients atteints de &oelig;sophagite de reflux d&eacute;veloppent BM et d&#39;autres pas.

NF-&kappa;B est activ&eacute; dans les fibroblastes &oelig;sophagiennes en r&eacute;ponse &agrave; un signal de paracrine g&eacute;n&eacute;r&eacute; par les cellules squameuses &oelig;sophagiennes primaires expos&eacute;s.

Duodenogastro &#x98DF;&#x7BA1; refluxate &#x98DF;&#x7BA1;&#x63A5;&#x89E6;&#x6F5C;&#x5728;&#x987E;&#x5BA2;&#xFF0C;&#x4EE5;&#x53CD;&#x6D41;&#x98DF;&#x9053;&#x708E;&#x548C;&#x88AB;&#x5377;&#x5165;&#x4E86;&#x5DF4;&#x96F7;&#x7279;&#x7684;&#x4E0A;&#x76AE;&#x5316;&#x751F; (BM) &#x7684;&#x53D1;&#x5C55;&#x3002;&#x76F8;&#x5173;&#x4E0E;&#x76F8;&#x4FE1;&#x4E3A; BM &#x53D1;&#x5C55;&#x7684;&#x6838;&#x5FC3;&#xFF0C;&#x867D;&#x7136; NF-&kappa; B &#x6D3B;&#x5316;&#x7684;&#x6210;&#x7EA4;&#x7EF4;&#x7EC6;&#x80DE;&#x57FA;&#x8D28;&#x91CD;&#x5851;&#x4E2D;&#x7684;&#x91CD;&#x8981;&#x4F5C;&#x7528;&#x7684;&#x8F6C;&#x5F55;&#x56E0;&#x5B50;&#x7684;&#x6FC0;&#x6D3B; NF-&kappa; B &#x4FE1;&#x53F7;&#x5728;&#x4E0A;&#x76AE;&#x7EC6;&#x80DE;&#x3002;&#x6211;&#x4EEC;&#x7684;&#x76EE;&#x6807;&#x662F;&#x7814;&#x7A76;&#x5BF9; NF-&kappa; B &#x6D3B;&#x5316;&#x6210;&#x4E3B;&#x98DF;&#x7BA1;&#x7EA4;&#x7EF4;&#x7EC6;&#x80DE; (HOFs) &#x5728;&#x9178;&#x66B4;&#x9732;&#x7684;&#x5F71;&#x54CD;&#x548C;&#x5C0F;&#x5B66;&#x548C;&#x6C38;&#x751F;&#x5316;&#x98DF;&#x7BA1; squames &#x5E76;&#x8C03;&#x67E5;&#x8FD9;&#x4E9B;&#x7EC6;&#x80DE;&#x5BF9;&#x9178;&#x5728;&#x54CD;&#x5E94;&#x4E2D;&#x7684;&#x4EFB;&#x4F55;&#x4E0A;&#x76AE;/&#x57FA;&#x8D28;&#x4EA4;&#x4E92;&#x3002;&#x4E3B; HOFs &#x548C;&#x5C0F;&#x5B66;&#x548C;&#x6C38;&#x751F;&#x5316;&#x98DF;&#x7BA1;&#x4E0A;&#x76AE;&#x7EC6;&#x80DE;&#x63A5;&#x89E6;&#x9178; (ph &#x503C; 7-pH 4 &le; 120 &#x5206;&#x949F;&#xFF09; &#x5355;&#x4E2A;&#x6216;&#x8109;&#x51B2;&#x6CBB;&#x7597;&#x3002;&#x4ECE;&#x4EE5;&#x4E0B;&#x7684;&#x9178;&#x66B4;&#x9732;&#x4E5F;&#x9002;&#x7528;&#x4E8E;&#x6210;&#x7EA4;&#x7EF4;&#x7EC6;&#x80DE;&#x7684;&#x4E0A;&#x76AE;&#x7EC6;&#x80DE;&#x7684;&#x57F9;&#x517B;&#x6DB2;&#x3002;&#x7EC6;&#x80DE;&#x6D3B;&#x6027;&#x7684; MTT ESTA &#x786E;&#x5B9A;&#x3002;NF-&kappa; B &#x6D3B;&#x5316;&#x7684; NF-&kappa; B/p65 &#x53EF;&#x89C6;&#x5316;&#x514D;&#x75AB;&#x8367;&#x5149;&#x7EC6;&#x80DE;&#x5B9A;&#x4F4D;&#x7684;&#x786E;&#x5B9A;&#x3002;&#x4ECE;&#x98DF;&#x7BA1;&#x4E0A;&#x76AE;&#x7EC6;&#x80DE;&#x7684;&#x57F9;&#x517B;&#x6DB2;&#xFF0C;&#x906D;&#x53D7; pH 5 &#x640F;&#x52A8;&#x6027;&#x66B4;&#x9732;&#xFF0C;&#x6FC0;&#x6D3B; NF-&kappa; B &#x7684;&#x6210;&#x7EA4;&#x7EF4;&#x7EC6;&#x80DE;&#xFF0C;&#x4E0E;&#x4E00;&#x4E9B;&#x5185;&#x7684;&#x53D8;&#x5316;&#xFF0C;&#x4F46;&#x8FD9;&#x4E9B;&#x6761;&#x4EF6;&#x4E0D;&#x4F1A;&#x76F4;&#x63A5;&#x6FC0;&#x6D3B; NF-&kappa; B &#x4E0A;&#x76AE;&#x7EC6;&#x80DE;&#x672C;&#x8EAB;&#x3002;&#x91CD;&#x5927;&#x7684; NF-&kappa; B &#x6FC0;&#x6D3B;&#x6709;&#x4EBA;&#x770B;&#x89C1;&#x4E0A;&#x76AE;&#x7EC6;&#x80DE;&#xFF0C;&#x4F46;&#x53EA;&#x4E0E;&#x66F4;&#x5927;&#x7684;&#x9178;&#x5EA6;&#x548C;&#x66B4;&#x9732;&#x65F6;&#x95F4; &#xFF08;ph &#x503C; 4&#x3001; 60-120 &#x5206;&#x949F;&#xFF09;&#x3002;&#x6211;&#x4EEC;&#x7684;&#x7814;&#x7A76;&#x7ED3;&#x679C;&#x663E;&#x793A;&#x9178;&#x66B4;&#x9732;&#x5BFC;&#x81F4;&#x95F4;&#x63A5;&#x6FC0;&#x6D3B;&#x57FA;&#x8D28;&#x7EC6;&#x80DE;&#x4E0A;&#x76AE;&#x95F4;&#x8D28;&#x7684;&#x4EA4;&#x4E92;&#x3002;&#x8FD9;&#x53EF;&#x80FD;&#x4F1A;&#x5BFC;&#x81F4;&#x98DF;&#x7BA1;&#x75BE;&#x75C5;&#x7684;&#x53D1;&#x75C5;&#x673A;&#x5236;&#x548C;&#x5185;&#x7684;&#x53D8;&#x5F02;&#x53EF;&#x80FD;&#x5728;&#x67D0;&#x79CD;&#x7A0B;&#x5EA6;&#x4E0A;&#x89E3;&#x91CA;&#x4E3A;&#x4EC0;&#x4E48;&#x4E00;&#x4E9B;&#x60A3;&#x8005;&#x98DF;&#x9053;&#x53D1;&#x5C55; BM&#xFF0C;&#x6709;&#x4E9B;&#x5219;&#x6CA1;&#x6709;&#x3002;

&#x6FC0;&#x6D3B; NF-&kappa; B &#x5219;&#x5728;&#x98DF;&#x7BA1;&#x6210;&#x7EA4;&#x7EF4;&#x7EC6;&#x80DE;&#x4E2D;&#x4EA7;&#x751F;&#x7684;&#x9178;&#x66B4;&#x9732;&#x539F;&#x53D1;&#x6027;&#x98DF;&#x7BA1;&#x9CDE;&#x72B6;&#x7EC6;&#x80DE;&#x65C1;&#x5206;&#x6CCC;&#x4FE1;&#x53F7;&#x7684;&#x54CD;&#x5E94;&#x3002;

Oesophageal exposure to duodenogastro-oesophageal refluxate leads to reflux oesophagitis and is implicated in the development of Barrett's metaplasia (BM). NF-ÎºB signalling in epithelial cells is associated with the activation of transcription factors believed to be central to BM development, whilst NF-ÎºB activation in fibroblasts plays a critical role in matrix remodelling. Our aim was to study the effects of acid exposure on NF-ÎºB activation in primary human oesophageal fibroblasts (HOFs) and primary and immortalized oesophageal squames and to investigate any epithelial/stromal interactions in the response of these cells to acid. Primary HOFs and primary and immortalized oesophageal epithelial cells were exposed to acid (pH 7 - pH 4 â‰¤ 120 min) in single or pulsed treatments. Conditioned medium from epithelial cells following acid exposure was also applied to fibroblasts. Cell viability was determined by MTT-ESTA. NF-ÎºB activation was determined by cellular localization of NF-ÎºB/p65 visualized by immunofluorescence. Conditioned medium from oesophageal epithelial cells, subjected to pH 5 pulsatile exposure, activated NF-ÎºB in fibroblasts, with some inter-patient variability, but these conditions did not directly activate NF-ÎºB in the epithelial cells themselves. Significant NF-ÎºB activation was seen in the epithelial cells but only with greater acidity and exposure times (pH 4, 60-120 min). Our findings show that acid exposure can cause indirect activation of stromal cells by epithelial-stromal interactions. This may contribute to the pathogenesis of oesophageal diseases, and the inter-patient variability may go some way to explain why some patients with reflux oesophagitis develop BM and others do not.

NF-ÎºB is activated in oesophageal fibroblasts in response to a paracrine signal generated by acid-exposed primary oesophageal squamous cells.

Esposizione esofagea al refluxate duodenogastro-esofagea conduce al reflusso esofagite ed &egrave; implicato nello sviluppo di metaplasia di Barrett (BM). Segnalazione delle cellule epiteliali NF-&kappa;B &egrave; associato con l&#39;attivazione di fattori di trascrizione ritiene che sia fondamentale per lo sviluppo di BM, mentre l&#39;attivazione di NF-&kappa;B in fibroblasti gioca un ruolo fondamentale nel rimodellamento della matrice. Il nostro obiettivo era di studiare gli effetti dell&#39;esposizione acida su attivazione di NF-&kappa;B in fibroblasti umani esofagei primari (HOFs) e squames esofageo primario e immortalata e per indagare su eventuali interazioni epitelio/stromale nella risposta di queste cellule all&#39;acido. HOFs primaria e primarie e immortalate nelle cellule epiteliali esofagei sono stati esposti ad acido (pH 7 - pH 4 &le; 120 min) nel singoli o pulsate trattamenti. Mezzo condizionato dalle cellule epiteliali seguente esposizione acida &egrave; stato applicato anche ai fibroblasti. Vitalit&agrave; cellulare &egrave; stata determinata da MTT-ESTA. Attivazione di NF-&kappa;B &egrave; stato determinato dalla localizzazione cellulare di NF-&kappa;B/p65 visualizzati mediante immunofluorescenza. Condizionata medio da cellule epiteliali esofagei, sottoposti a pH 5 pulsatile esposizione, attivato NF-&kappa;B in fibroblasti, con qualche variabilit&agrave; inter-patient, ma queste condizioni non ha fatto direttamente attivare NF-&kappa;B nelle cellule epiteliali stessi. Significativa attivazione di NF-&kappa;B &egrave; stato visto nelle cellule epiteliali ma solo con una maggiore acidit&agrave; ed esposizione volte (pH 4, 60-120 min). I nostri risultati mostrano che l&#39;esposizione acida pu&ograve; causare indiretti attivazione di cellule stromali da interazioni epiteliali-stromale. Ci&ograve; pu&ograve; contribuire alla patogenesi delle malattie dell&#39;esofago, e la variabilit&agrave; inter-patient pu&ograve; andare qualche modo per spiegare perch&eacute; alcuni pazienti con esofagite da reflusso sviluppano BM e gli altri non lo fanno.

NF-&kappa;B &egrave; attivato in esofagei fibroblasti in risposta ad un segnale paracrina generato da acido-esposti primarie esofagea cellule squamose.

Duodenogastro &#x98DF;&#x9053; refluxate &#x98DF;&#x9053;&#x66DD;&#x9732;&#x306F;&#x98DF;&#x9053;&#x708E;&#x9006;&#x6D41;&#x306B;&#x3064;&#x306A;&#x304C;&#x308B;&#x3057;&#x3001;&#x30D0;&#x30EC;&#x30C3;&#x30C8;&#x4E0A;&#x76AE;&#x5316;&#x751F; (BM) &#x306E;&#x958B;&#x767A;&#x306B;&#x95A2;&#x4E0E;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;NF-&kappa; &#x4E0A;&#x76AE;&#x7D30;&#x80DE;&#x306E;&#x30B7;&#x30B0;&#x30CA;&#x30EB;&#x4F1D;&#x9054;&#x306F;&#x7DDA;&#x7DAD;&#x82BD;&#x7D30;&#x80DE;&#x306B;&#x304A;&#x3051;&#x308B; NF-&kappa; &#x6D3B;&#x6027;&#x5316;&#x884C;&#x5217;&#x3092;&#x6539;&#x9020;&#x3067;&#x91CD;&#x8981;&#x306A;&#x5F79;&#x5272;&#x3092;&#x679C;&#x305F;&#x3057;&#x3066;&#x3044;&#x308B;&#x3057;&#x306A;&#x304C;&#x3089; BM &#x306E;&#x958B;&#x767A;&#x306B;&#x4E2D;&#x592E;&#x3067;&#x3042;&#x308B;&#x3068;&#x8003;&#x3048;&#x3089;&#x308C;&#x3066;&#x8EE2;&#x5199;&#x56E0;&#x5B50;&#x306E;&#x6D3B;&#x6027;&#x5316;&#x306B;&#x95A2;&#x9023;&#x4ED8;&#x3051;&#x3089;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x79C1;&#x305F;&#x3061;&#x306E;&#x76EE;&#x7684;&#x306F;&#x3001;&#x30D7;&#x30E9;&#x30A4;&#x30DE;&#x30EA;&#x3072;&#x3068;&#x98DF;&#x9053;&#x7D30;&#x80DE; (HOFs) &#x306B;&#x304A;&#x3051;&#x308B; NF-&kappa; &#x6D3B;&#x6027;&#x5316;&#x306B;&#x53CA;&#x307C;&#x3059;&#x9178;&#x66DD;&#x9732;&#x3068;&#x98DF;&#x9053;&#x76AE;&#x6027;&#x6241;&#x5E73;&#x7D30;&#x80DE;&#x306E;&#x4E00;&#x6B21;&#x4E0D;&#x6B7B;&#x5316;&#x7814;&#x7A76;&#x3057;&#x3001;&#x4EFB;&#x610F;&#x9178;&#x306B;&#x3053;&#x308C;&#x3089;&#x306E;&#x7D30;&#x80DE;&#x306E;&#x5FDC;&#x7B54;&#x3067;&#x4E0A;&#x76AE;&#x9593;&#x8CEA;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x3092;&#x8ABF;&#x67FB;&#x3059;&#x308B;&#x3060;&#x3063;&#x305F;&#x3002;&#x4E3B;&#x306A; HOFs &#x3068;&#x30D7;&#x30E9;&#x30A4;&#x30DE;&#x30EA;&#x304A;&#x3088;&#x3073;&#x4E0D;&#x6B7B;&#x5316;&#x98DF;&#x9053;&#x4E0A;&#x76AE;&#x7D30;&#x80DE;&#x306F;&#x9178;&#x306B;&#x3055;&#x3089;&#x3055;&#x308C;&#x305F; (pH 7 - pH 4 &le; 120 min) &#x5358;&#x4E00;&#x307E;&#x305F;&#x306F;&#x30D1;&#x30EB;&#x30B9;&#x6CBB;&#x7642;&#x306E;&#x3002;&#x6B21;&#x306E;&#x9178;&#x66B4;&#x9732;&#x306F;&#x7DDA;&#x7DAD;&#x82BD;&#x7D30;&#x80DE;&#x306B;&#x3082;&#x9069;&#x7528;&#x3055;&#x308C;&#x305F;&#x4E0A;&#x76AE;&#x7D30;&#x80DE;&#x99B4;&#x5316;&#x57F9;&#x5730;&#x3002;&#x7D30;&#x80DE;&#x751F;&#x5B58;&#x7387;&#x306F; MTT-ESTA &#x306B;&#x3088;&#x3063;&#x3066;&#x6C7A;&#x5B9A;&#x3055;&#x308C;&#x307E;&#x3057;&#x305F;&#x3002;NF-&kappa; &#x6D3B;&#x6027;&#x5316;&#x306F;&#x3001;NF-&kappa;B/p65 &#x87A2;&#x5149;&#x6297;&#x4F53;&#x6CD5;&#x306B;&#x3088;&#x308B;&#x53EF;&#x8996;&#x5316;&#x306E;&#x7D30;&#x80DE;&#x5C40;&#x5728;&#x306B;&#x3088;&#x3063;&#x3066;&#x6C7A;&#x5B9A;&#x3055;&#x308C;&#x307E;&#x3057;&#x305F;&#x3002;&#x98DF;&#x9053;&#x4E0A;&#x76AE;&#x7D30;&#x80DE;&#x304B;&#x3089;&#x306E;&#x5A92;&#x4F53;&#x3092;&#x5B8C;&#x5099;&#x3001;&#x9732;&#x51FA;&#x3001;&#x3044;&#x304F;&#x3064;&#x304B; inter-patient &#x306E;&#x5909;&#x52D5;&#x3068;&#x7DDA;&#x7DAD;&#x82BD;&#x7D30;&#x80DE;&#x306B;&#x304A;&#x3051;&#x308B; NF-&kappa; &#x30A2;&#x30AF;&#x30C6;&#x30A3;&#x30D6; pH 5 &#x62CD;&#x52D5;&#x3092;&#x53D7;&#x3051;&#x308B;&#x304C;&#x3001;&#x3053;&#x308C;&#x3089;&#x306E;&#x6761;&#x4EF6;&#x304C;&#x76F4;&#x63A5; NF-&kappa; &#x4E0A;&#x76AE;&#x7D30;&#x80DE;&#x81EA;&#x4F53;&#x3092;&#x30A2;&#x30AF;&#x30C6;&#x30A3;&#x30D6;&#x5316;&#x3057;&#x306A;&#x304B;&#x3063;&#x305F;&#x3002;&#x91CD;&#x8981;&#x306A; NF-&kappa; &#x6D3B;&#x6027;&#x5316;&#x4E0A;&#x76AE;&#x7D30;&#x80DE;&#x3067;&#x3059;&#x304C;&#x5927;&#x304D;&#x3044;&#x9178;&#x5473;&#x3068;&#x9732;&#x51FA;&#x6642;&#x9593;&#x3067;&#x306E;&#x307F;&#x898B;&#x3089;&#x308C;&#x305F; &#xFF08;pH 4&#x3001;60-120 &#x5206;&#x3060;)&#x3002;&#x6211;&#x3005; &#x306E;&#x8ABF;&#x67FB;&#x7D50;&#x679C;&#x306F;&#x9178;&#x66DD;&#x9732;&#x304C;&#x4E0A;&#x76AE;&#x30FB;&#x9593;&#x8CEA;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x306B;&#x3088;&#x308B;&#x9593;&#x8CEA;&#x7D30;&#x80DE;&#x306E;&#x9593;&#x63A5;&#x306E;&#x6D3B;&#x6027;&#x5316;&#x3092;&#x5F15;&#x304D;&#x8D77;&#x3053;&#x3059;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x3053;&#x308C;&#x306F;&#x98DF;&#x9053;&#x75BE;&#x60A3;&#x306E;&#x767A;&#x75C7;&#x306B;&#x8CA2;&#x732E;&#x3059;&#x308B;&#x304B;&#x3082;&#x3057;&#x308C;&#x306A;&#x3044;&#x3057;&#x3001;inter-patient &#x306E;&#x53EF;&#x5909;&#x6027;&#x306A;&#x305C; BM &#x8853;&#x306F;&#x9006;&#x6D41;&#x30E9;&#x30C3;&#x30C8;&#x30E2;&#x30C7;&#x30EB;&#x3092;&#x958B;&#x767A;&#x3057;&#x3001;&#x4ED6;&#x306F;&#x3042;&#x308A;&#x307E;&#x305B;&#x3093;&#x8AAC;&#x660E;&#x3059;&#x308B;&#x3044;&#x304F;&#x3064;&#x304B;&#x306E;&#x65B9;&#x6CD5;&#x3092;&#x884C;&#x304F;&#x3053;&#x3068;&#x304C;&#x3042;&#x308A;&#x307E;&#x3059;&#x3002;

NF-&kappa; &#x9178;&#x3055;&#x3089;&#x3055;&#x30D7;&#x30E9;&#x30A4;&#x30DE;&#x30EA;&#x98DF;&#x9053;&#x6241;&#x5E73;&#x4E0A;&#x76AE;&#x764C;&#x7D30;&#x80DE;&#x306B;&#x3088;&#x3063;&#x3066;&#x751F;&#x6210;&#x3055;&#x308C;&#x308B;&#x508D;&#x5206;&#x6CCC;&#x30B7;&#x30B0;&#x30CA;&#x30EB;&#x306B;&#x5FDC;&#x7B54;&#x3059;&#x308B;&#x98DF;&#x9053;&#x7DDA;&#x7DAD;&#x82BD;&#x7D30;&#x80DE;&#x3067;&#x30A2;&#x30AF;&#x30C6;&#x30A3;&#x30D6;&#x306B;&#x306A;&#x308A;&#x307E;&#x3059;&#x3002;

Duodenogastro &#xC2DD;&#xB3C4; refluxate &#xC2DD;&#xB3C4; &#xB178;&#xCD9C; &#xB9AC;&#xB4DC; &#xC2DD;&#xB3C4; &#xC5ED;&#xB958; &#xC131; &#xC2DD;&#xB3C4; &#xC5FC;&#xACFC; &#xBC14; &#xB81B;&#xC758; metaplasia (BM) &#xAC1C;&#xBC1C;&#xC5D0; &#xC5F0;&#xB8E8;. NF &kappa;B &#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xC5D0; &#xC2E0;&#xD638; &#xC5F0;&#xACB0; &#xC12C;&#xC720; &#xC544; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; NF &kappa;B &#xD65C;&#xC131;&#xD654; &#xB9E4;&#xD2B8;&#xB9AD;&#xC2A4; &#xAC1C;&#xC7A5;&#xC5D0; &#xC911;&#xC694; &#xD55C; &#xC5ED;&#xD560;&#xC744; &#xC7AC;&#xC0DD; &#xD558;&#xB294; &#xB3D9;&#xC548; &#xC911;&#xC559; BM &#xAC1C;&#xBC1C; &#xB420; &#xAC83;&#xC73C;&#xB85C; &#xCD94;&#xC815; &#xD558;&#xB294; &#xC804;&#xC0AC; &#xC778;&#xC790;&#xB97C; &#xD65C;&#xC131;&#xD654;&#xAC00; &#xB429;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC;&#xC758; &#xBAA9;&#xD45C;&#xB294; &#xAE30;&#xBCF8; &#xC778;&#xAC04;&#xC758; &#xC2DD;&#xB3C4; &#xAC8C;&#xC7AC; (HOFs)&#xC5D0; &#xC788;&#xB294; NF-&kappa;B &#xD65C;&#xC131;&#xD654;&#xC5D0; &#xC0B0; &#xB178;&#xCD9C; &#xD6A8;&#xACFC; &#xBD88;&#xBA78; &#xD558; &#xAC8C; &#xC2DD;&#xB3C4; squames &#xACF5;&#xBD80; &#xD558; &#xACE0; &#xADF8;&#xB9AC;&#xACE0; &#xC0B0;&#xC774; &#xC138;&#xD3EC;&#xC758; &#xC751;&#xB2F5;&#xC5D0; &#xC5B4;&#xB5A4; &#xC0C1;&#xD53C; stromal &#xC0C1;&#xD638; &#xC791;&#xC6A9;&#xC744; &#xC870;&#xC0AC;. &#xAE30;&#xBCF8; HOFs &#xBC0F; &#xC2DD;&#xB3C4; &#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xB97C; &#xBD88;&#xBA78; &#xD558; &#xAC8C; &#xD558; &#xACE0; &#xC8FC; &#xC0B0;&#xC5D0; &#xB178;&#xCD9C; &#xB41C; (pH 7-pH 4 &le; 120 &#xBD84;) &#xB2E8;&#xC77C; &#xB610;&#xB294; &#xD384;&#xC2A4; &#xCE58;&#xB8CC;&#xC5D0;. &#xB2E4;&#xC74C; &#xC0B0; &#xB178;&#xCD9C; &#xB610;&#xD55C; &#xC12C;&#xC720; &#xC544; &#xC138;&#xD3EC;&#xC5D0; &#xC801;&#xC6A9; &#xB41C; &#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; &#xBC14;&#xB978;&#xB41C; &#xB9E4;&#xCCB4;. &#xC138;&#xD3EC; &#xC0DD;&#xC874; &#xB2A5;&#xB825; MTT ESTA&#xC5D0; &#xC758;&#xD574; &#xACB0;&#xC815; &#xB418;&#xC5C8;&#xB2E4;. NF &kappa;B &#xD65C;&#xC131;&#xD654; NF-&kappa;B/p65 &#xBA74;&#xC5ED; &#xD615;&#xAD11; &#xAC80;&#xC0AC;&#xC5D0;&#xC11C; &#xC2DC;&#xAC01;&#xC758; &#xC140;&#xB8F0;&#xB7EC; &#xC9C0;&#xC5ED;&#xD654;&#xC5D0; &#xC758;&#xD574; &#xACB0;&#xC815; &#xB418;&#xC5C8;&#xB2E4;. &#xC2DD;&#xB3C4; &#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; &#xB9E4;&#xCCB4;&#xB97C; &#xC870;&#xC808;, &#xB178;&#xCD9C;, &#xC77C;&#xBD80; inter-patient &#xB2E4;&#xC591;&#xC131;&#xC73C;&#xB85C; &#xAC8C;&#xC7AC;, NF-&kappa;B &#xD65C;&#xC131;&#xD654; pH 5 &#xD0C0;&#xC545;&#xAE30;&#xB97C; &#xBCF5;&#xC885; &#xD558;&#xC9C0;&#xB9CC; &#xC774;&#xB7EC;&#xD55C; &#xC870;&#xAC74;&#xC744; &#xC9C1;&#xC811; &#xC2A4;&#xC2A4;&#xB85C; &#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; NF-&Kappa;b&#xC744; &#xD65C;&#xC131;&#xD654; &#xD558;&#xC9C0; &#xC54A;&#xC740;. &#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xC5D0;&#xB9CC; &#xD070; &#xC0B0;&#xB3C4; &#xBC0F; &#xB178;&#xCD9C; &#xC2DC;&#xAC04; &#xB9CC;&#xC73C;&#xB85C; &#xC0C1;&#xB2F9;&#xD55C; NF &kappa;B &#xD65C;&#xC131;&#xD654; &#xBCF4;&#xC600;&#xB2E4; (pH 4, 60-120 &#xBD84;). &#xC6B0;&#xB9AC;&#xC758; &#xC5F0;&#xAD6C; &#xACB0;&#xACFC; &#xBCF4;&#xC5EC; &#xC0B0; &#xB178;&#xCD9C; &#xC0C1;&#xD53C; stromal &#xC0C1;&#xD638; &#xC791;&#xC6A9;&#xC5D0; &#xC758;&#xD574; stromal &#xC138;&#xD3EC;&#xC758; &#xAC04;&#xC811;&#xC801;&#xC778; &#xD65C;&#xC131;&#xD654;&#xB97C; &#xC77C;&#xC73C;&#xD0AC; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. &#xC2DD;&#xB3C4; &#xC9C8;&#xD658;&#xC758; pathogenesis&#xC758; &#xC6D0;&#xC778;&#xC774; &#xB420; &#xC218;&#xC774; &#xACE0; inter-patient &#xB2E4;&#xC591;&#xC131; &#xC65C; &#xC5ED;&#xB958; &#xC131; &#xC2DD;&#xB3C4; &#xC5FC; &#xC2DD;&#xB3C4; &#xC77C;&#xBD80; &#xD658;&#xC790; BM &#xAC1C;&#xBC1C; &#xD558; &#xACE0; &#xB2E4;&#xB978; &#xC0AC;&#xB78C;&#xC740; &#xC124;&#xBA85; &#xD558;&#xB294; &#xBA87; &#xAC00;&#xC9C0; &#xBC29;&#xBC95;&#xC774; &#xAC08; &#xC218;&#xB3C4; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;.

NF &kappa;B &#xC2DD;&#xB3C4; &#xAC8C;&#xC7AC; paracrine &#xC2E0;&#xD638; &#xC0B0; &#xB178;&#xCD9C; &#xAE30;&#xBCF8; &#xC2DD;&#xB3C4; &#xD3B8;&#xD3C9; &#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xC5D0; &#xC758;&#xD574; &#xC0DD;&#xC131; &#xB41C;&#xC5D0; &#xB300; &#xD55C; &#xC751;&#xB2F5;&#xC5D0;&#xC11C; &#xD65C;&#xC131;&#xD654; &#xB429;&#xB2C8;&#xB2E4;.

Exposici&oacute;n esof&aacute;gica al refluxate duodenogastro-esof&aacute;gica conduce a esofagitis de reflujo y est&aacute; implicada en el desarrollo de la metaplasia de Barrett (BM). NF-&kappa;B se&ntilde;alizaci&oacute;n en las c&eacute;lulas epiteliales se asocia con la activaci&oacute;n de factores de transcripci&oacute;n que cree que es fundamental para el desarrollo del BM, mientras que la activaci&oacute;n de NF-&kappa;B en fibroblastos desempe&ntilde;a un papel cr&iacute;tico en la remodelaci&oacute;n de la matriz. Nuestro objetivo fue estudiar los efectos de la exposici&oacute;n de &aacute;cido en la activaci&oacute;n de NF-&kappa;B en fibroblastos esof&aacute;gicos humanos primarios (HOFs) y escamas esof&aacute;gica primaria e inmortalizado y cualquier interacci&oacute;n epitelial/stromal en la respuesta de estas c&eacute;lulas al &aacute;cido. HOFs primarios y las c&eacute;lulas epiteliales esof&aacute;gicas primarias e inmortalizadas fueron expuestas al &aacute;cido (pH 7 - pH 4 &le; 120 min) en tratamientos individuales o pulsados. Medio condicionado de c&eacute;lulas epiteliales siguiente exposici&oacute;n &aacute;cida tambi&eacute;n se aplic&oacute; a los fibroblastos. Viabilidad celular se determin&oacute; por MTT-ESTA. Activaci&oacute;n de NF-&kappa;B determin&oacute; la localizaci&oacute;n celular de NF-&kappa;B/p65 visualizado por inmunofluorescencia. Acondicionado en medio de las c&eacute;lulas epiteliales esof&aacute;gicas, sometidos a pH 5 puls&aacute;til exposici&oacute;n, activan NF-&kappa;B en fibroblastos, con cierta variabilidad entre, pero estas condiciones no activar directamente NF-&kappa;B en las propias c&eacute;lulas epiteliales. Activaci&oacute;n de NF-&kappa;B significativa se observ&oacute; en las c&eacute;lulas epiteliales, pero s&oacute;lo con una mayor acidez y exposici&oacute;n veces (pH 4, 60-120 min). Nuestros resultados demuestran que la exposici&oacute;n de &aacute;cido puede causar activaci&oacute;n indirecta de las c&eacute;lulas estromales por interacciones epiteliales-stromal. Esto puede contribuir a la patogenesia de enfermedades esof&aacute;gicas, y la variabilidad entre puede ir de alguna manera para explicar por qu&eacute; algunos pacientes con esofagitis de reflujo desarrollan BM y otros no.

NF-&kappa;B interviene en fibroblastos esof&aacute;gicos en respuesta a una se&ntilde;al paracrina generada por c&eacute;lulas escamosas esof&aacute;gicas primarias expuestos al &aacute;cido.

The first transition-metal complex-based two-photon absorbing luminescence lifetime probes for cellular DNA are presented. This allows cell imaging of DNA free from endogenous fluorophores and potentially facilitates deep tissue imaging. In this initial study, ruthenium(II) luminophores are used as phosphorescent lifetime imaging microscopy (PLIM) probes for nuclear DNA in both live and fixed cells. The DNA-bound probes display characteristic emission lifetimes of more than 160 ns, while shorter-lived cytoplasmic emission is also observed. These timescales are orders of magnitude longer than conventional FLIM, leading to previously unattainable levels of sensitivity, and autofluorescence-free imaging.

Dinuclear ruthenium(II) complexes as two-photon, time-resolved emission microscopy probes for cellular DNA.

The biological and mechanical function of connective tissues is largely determined by controlled cellular alignment and therefore it seems appropriate that tissue-engineered constructs should be architecturally similar to the in vivo tissue targeted for repair or replacement. Collagen organisation dictates the tensile properties of most tissues and so monitoring the deposition of cell-secreted collagen as the construct develops is essential for understanding tissue formation. In this study, electrospun fibres with a random or high degree of orientation, mimicking two types of tissue architecture found in the body, were used to culture human fibroblasts for controlling cell alignment. The minimally-invasive technique of second harmonic generation was used with the aim of monitoring and profiling the deposition and organisation of collagen at different construct depths over time while construct mechanical properties were also determined over the culture period. It was seen that scaffold fibre organisation affected cell migration and orientation up to 21 days which in turn had an effect on collagen organisation. Collagen in random fibrous constructs was deposited in alternating configurations at different depths however a high degree of organisation was observed throughout aligned fibrous constructs orientated in the scaffold fibre direction. Three-dimensional second harmonic generation images showed that deposited collagen was more uniformly distributed in random constructs but aligned constructs were more organised and had higher intensities. The tensile properties of all constructs increased with increasing collagen deposition and were ultimately dictated by collagen organisation. This study highlights the importance of scaffold architecture for controlling the development of well-organised tissue engineered constructs and the usefulness of second harmonic generation imaging for monitoring collagen maturation in a minimally invasive manner.

Monitoring fibrous scaffold guidance of three-dimensional collagen organisation using minimally-invasive second harmonic generation.

Oesophageal exposure to duodenogastroesophageal refluxate is implicated in the development of Barrett's metaplasia (BM), with increased risk of progression to oesophageal adenocarcinoma. The literature proposes that reflux exposure activates NF-κB, driving the aberrant expression of intestine-specific caudal-related homeobox (CDX) genes. However, early events in the pathogenesis of BM from normal epithelium are poorly understood. To investigate this, our study subjected a 3D model of the normal human oesophageal mucosa to repeated, pulsatile exposure to specific bile components and examined changes in gene expression. Initial 2D experiments with a range of bile salts observed that taurochenodeoxycholate (TCDC) impacted upon NF-κB activation without causing cell death. Informed by this, the 3D oesophageal model was repeatedly exposed to TCDC in the presence and absence of acid, and the epithelial cells underwent gene expression profiling. We identified ~300 differentially expressed genes following each treatment, with a large and significant overlap between treatments. Enrichment analysis (Broad GSEA, DAVID and Metacore™; GeneGo Inc) identified multiple gene sets related to cell signalling, inflammation, proliferation, differentiation and cell adhesion. Specifically NF-κB activation, Wnt signalling, cell adhesion and targets for the transcription factors PTF1A and HNF4α were highlighted. Our data suggest that HNF4α isoform switching may be an early event in Barrett's pathogenesis. CDX1/2 targets were, however, not enriched, suggesting that although CDX1/2 activation reportedly plays a role in BM development, it may not be an initial event. Our findings highlight new areas for investigation in the earliest stages of BM pathogenesis of oesophageal diseases and new potential therapeutic targets.

Pulsatile exposure to simulated reflux leads to changes in gene expression in a 3D model of oesophageal mucosa.

Within drinking water distribution systems (DWDS), microorganisms form multi-species biofilms on internal pipe surfaces. A matrix of extracellular polymeric substances (EPS) is produced by the attached community and provides structure and stability for the biofilm. If the EPS adhesive strength deteriorates or is overcome by external shear forces, biofilm is mobilised into the water potentially leading to degradation of water quality. However, little is known about the EPS within DWDS biofilms or how this is influenced by community composition or environmental parameters, because of the complications in obtaining biofilm samples and the difficulties in analysing EPS. Additionally, although biofilms may contain various microbial groups, research commonly focuses solely upon bacteria. This research applies an EPS analysis method based upon fluorescent confocal laser scanning microscopy (CLSM) in combination with digital image analysis (DIA), to concurrently characterize cells and EPS (carbohydrates and proteins) within drinking water biofilms from a full-scale DWDS experimental pipe loop facility with representative hydraulic conditions. Application of the EPS analysis method, alongside DNA fingerprinting of bacterial, archaeal and fungal communities, was demonstrated for biofilms sampled from different positions around the pipeline, after 28 days growth within the DWDS experimental facility. The volume of EPS was 4.9 times greater than that of the cells within biofilms, with carbohydrates present as the dominant component. Additionally, the greatest proportion of EPS was located above that of the cells. Fungi and archaea were established as important components of the biofilm community, although bacteria were more diverse. Moreover, biofilms from different positions were similar with respect to community structure and the quantity, composition and three-dimensional distribution of cells and EPS, indicating that active colonisation of the pipe wall is an important driver in material accumulation within the DWDS.

Characterisation of the physical composition and microbial community structure of biofilms within a model full-scale drinking water distribution system.

Nicola H. Green

Department of Materials Science and Engineering

University of Sheffield

Production, Characterization and Potential Uses of a 3D Tissue-engineered Human Esophageal Mucosal Model

Nicola H. Green

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Materials Science and Engineering

University of Sheffield

Production, Characterization and Potential Uses of a 3D Tissue-engineered Human Esophageal Mucosal Model

Department of Materials Science and Engineering