Intravital Microscopy of Leukocyte-endothelial and Platelet-leukocyte Interactions in Mesenterial Veins in Mice

Intravital microscopy is a method that can be used to investigate different processes in different regions and vessels in living animals. In this ...

Heart Center, University of Freiburg

CD40L &#x5728;&#x52A8;&#x8109;&#x7CA5;&#x6837;&#x786C;&#x5316;&#x7B49;&#x6162;&#x6027;&#x708E;&#x75C7;&#x75BE;&#x75C5;&#x4E2D;&#x5360;&#x91CD;&#x8981;&#x5730;&#x4F4D;&#x3002;&#x7136;&#x800C;&#xFF0C;&#x7531;&#x4E8E; CD40L &#x5148;&#x5929;&#x8C03;&#x8282;&#x514D;&#x75AB;&#x529F;&#x80FD;&#x548C;&#x4E0E; CD40 &#x53D7;&#x4F53;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x7684;&#x6B62;&#x8840;&#x548C;&#x8840;&#x5C0F;&#x677F; GPIIb/IIIa &#x6574;&#x5408;&#x7D20;&#xFF0C;&#x5176;&#x5168;&#x7403;&#x6291;&#x5236;&#x59A5;&#x534F;&#x4E3B;&#x673A;&#x9632;&#x5FA1;&#x548C;&#x751F;&#x6210;&#x8840;&#x6813;&#x6813;&#x585E;&#x5E76;&#x53D1;&#x75C7;&#x7684;&#x4E34;&#x5E8A;&#x8BD5;&#x9A8C;&#x3002;&#x6211;&#x4EEC;&#x6700;&#x8FD1;&#x62A5;&#x9053; CD40L &#x4ECB;&#x5BFC;&#x52A8;&#x8109;&#x7CA5;&#x6837;&#x786C;&#x5316;&#x5F62;&#x6210;&#x72EC;&#x7ACB;&#x4E8E; CD40 &#x548C;&#x62DF;&#x8BAE;&#x7684; Mac-1 &#x4F5C;&#x4E3A;&#x5907;&#x7528;&#x7684;&#x53D7;&#x4F53;&#x3002;

CD40L &#x5230; Mac 1-&#x57DF;&#x7ED1;&#x5B9A;&#x6D89;&#x53CA; EQLKKSKTL &#x6BCD;&#x9898;&#x548C;&#x4ECB;&#x5BFC;&#x767D;&#x7EC6;&#x80DE;&#x5F81;&#x8058;&#x548C;&#x52A8;&#x8109;&#x7CA5;&#x6837;&#x786C;&#x5316; &mdash; &mdash; &#x4F46;&#x4E0D;&#x5F71;&#x54CD;&#x514D;&#x75AB;&#x548C;&#x5C0F;&#x9F20;&#x8840;&#x6813;&#x5F62;&#x6210;&#x7684;&#x3002;

CD40L figures prominently in chronic inflammatory diseases such as atherosclerosis. However, since CD40L potently regulates immune function and hemostasis by interaction with CD40 receptor and the platelet integrin GPIIb/IIIa, its global inhibition compromises host defense and generated thromboembolic complications in clinical trials. We recently reported that CD40L mediates atherogenesis independently of CD40 and proposed Mac-1 as an alternate receptor.

Binding of CD40L to Mac-1's I-domain involves the EQLKKSKTL motif and mediates leukocyte recruitment and atherosclerosis--but does not affect immunity and thrombosis in mice.

CD40L occupe une place importante dans les maladies inflammatoires chroniques telles que l&#39;ath&eacute;roscl&eacute;rose. Toutefois, puisque le CD40L a puissamment r&eacute;gule la fonction immunitaire et h&eacute;mostase par interaction avec le r&eacute;cepteur CD40 et l&#39;int&eacute;grine plaquettaire GPIIb/IIIa, ses compromis global inhibition h&eacute;bergent d&eacute;fense et g&eacute;n&eacute;r&eacute;es des complications thrombo-emboliques lors d&#39;essais cliniques. Nous avons r&eacute;cemment rapport&eacute; que CD40L m&eacute;die ath&eacute;rogen&egrave;se ind&eacute;pendamment de CD40 et Mac-1 propos&eacute; comme un autre r&eacute;cepteur.

Liaison du CD40L &agrave; je-domaine de Mac-1 implique le motif EQLKKSKTL et intervient dans le recrutement des leucocytes et l&#39;ath&eacute;roscl&eacute;rose--mais n&#39;affecte pas l&#39;immunit&eacute; et thrombose chez la souris.

CD40L Zahlen prominent in chronisch entz&uuml;ndlichen Erkrankungen wie Atherosklerose. Jedoch da CD40L stark immune Funktion und Blutstillung durch Interaktion mit CD40-Rezeptor und der Thrombozyten-Integrin GPIIb/IIIa reguliert, hosten die globalen Hemmung Kompromissen Verteidigung und generierte thromboembolische Komplikationen in klinischen Studien. Vor kurzem berichteten wir, dass CD40L Atherogenesis unabh&auml;ngig von CD40 und vorgeschlagenen Mac-1 als eine Alternative-Rezeptor vermittelt.

Bindung von CD40L Mac-1-Dom&auml;ne umfasst das EQLKKSKTL-Motiv und vermittelt Leukozyten Rekrutierung und Atherosklerose-- aber hat keinen Einfluss auf die Immunit&auml;t und Thrombosen in den M&auml;usen.

CD40L cifre visibile nelle malattie infiammatorie croniche come l&#39;aterosclerosi. Tuttavia, poich&eacute; CD40L potentemente regola la funzione immunitaria e l&#39;emostasi di interazione con il recettore CD40 e l&#39;integrina piastrinica GPIIb/IIIa, suoi compromessi inibizione globale ospitano difesa e complicanze tromboemboliche generate in studi clinici. Abbiamo recentemente segnalato che CD40L da intermediario aterogenesi indipendentemente da CD40 e Mac-1 proposta come un&#39;alternativo del recettore.

Associazione di CD40L al dominio di Mac-1 coinvolge il motivo EQLKKSKTL e da intermediario reclutamento dei leucociti e aterosclerosi - ma non pregiudica l&#39;immunit&agrave; e trombosi nei topi.

CD40L &#x306F;&#x30A2;&#x30C6;&#x30ED;&#x30FC;&#x30E0;&#x6027;&#x52D5;&#x8108;&#x786C;&#x5316;&#x306A;&#x3069;&#x306E;&#x6162;&#x6027;&#x708E;&#x75C7;&#x6027;&#x75BE;&#x60A3;&#x3067;&#x9855;&#x8457;&#x306B;&#x73FE;&#x308C;&#x307E;&#x3059;&#x3002;&#x305F;&#x3060;&#x3057;&#x3001;&#x5F37;&#x529B;&#x3001;CD40L &#x306F;&#x514D;&#x75AB;&#x6A5F;&#x80FD;&#x3068; CD40 &#x53D7;&#x5BB9;&#x4F53;&#x3068;&#x306E;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x306B;&#x3088;&#x308B;&#x6B62;&#x8840;&#x3068;&#x8840;&#x5C0F;&#x677F;&#x30A4;&#x30F3;&#x30C6;&#x30B0;&#x30EA;&#x30F3; &#x30B7;&#x30ED;&#x30B9;&#x30BF;&#x30BE;&#x30FC;&#x30EB;&#x3092;&#x8ABF;&#x7BC0;&#x3059;&#x308B;&#x306E;&#x3067;&#x3001;&#x305D;&#x306E;&#x30B0;&#x30ED;&#x30FC;&#x30D0;&#x30EB;&#x6291;&#x5236;&#x59A5;&#x5354;&#x306F;&#x9632;&#x885B;&#x3068;&#x81E8;&#x5E8A;&#x8A66;&#x9A13;&#x3067;&#x751F;&#x6210;&#x3055;&#x308C;&#x305F;&#x8840;&#x6813;&#x585E;&#x6813;&#x5408;&#x4F75;&#x75C7;&#x3092;&#x30DB;&#x30B9;&#x30C8;&#x3057;&#x307E;&#x3059;&#x3002;&#x6211;&#x3005; &#x306F;&#x6700;&#x8FD1;&#x3001;CD40L &#x7CA5;&#x72B6;&#x52D5;&#x8108;&#x786C;&#x5316; CD40 &#x3057;&#x3001;&#x4EE3;&#x66FF;&#x306E;&#x53D7;&#x5BB9;&#x4F53;&#x3068;&#x3057;&#x3066;&#x63D0;&#x6848; Mac 1 &#x3068;&#x306F;&#x7121;&#x95A2;&#x4FC2;&#x3092;&#x4EF2;&#x4ECB;&#x3059;&#x308B;&#x3053;&#x3068;&#x3092;&#x5831;&#x544A;&#x3057;&#x307E;&#x3057;&#x305F;&#x3002;

CD40L &#x306E; Mac 1 &#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x3078;&#x306E;&#x30D0;&#x30A4;&#x30F3;&#x30C9;&#x306F; EQLKKSKTL &#x30E2;&#x30C1;&#x30FC;&#x30D5;&#x3092;&#x542B;&#x3080;&#x3068;&#x767D;&#x8840;&#x7403;&#x52D5;&#x54E1;&#x3068;&#x30A2;&#x30C6;&#x30ED;&#x30FC;&#x30E0;&#x6027;&#x52D5;&#x8108;&#x786C;&#x5316;--&#x4EF2;&#x4ECB;&#x3057;&#x307E;&#x3059;&#x304C;&#x3001;&#x514D;&#x75AB;&#x3068;&#x8840;&#x6813;&#x75C7;&#x306E;&#x30DE;&#x30A6;&#x30B9;&#x306B;&#x306F;&#x5F71;&#x97FF;&#x3057;&#x307E;&#x305B;&#x3093;&#x3002;

&#xB3D9;&#xB9E5; &#xACBD;&#xD654; &#xC99D;&#xACFC; &#xAC19;&#xC740; &#xB9CC;&#xC131; &#xC5FC;&#xC99D; &#xC131; &#xC9C8;&#xD658; &#xB208;&#xC5D0; &#xB744;&#xAC8C; CD40L &#xC778;&#xBB3C;. &#xADF8;&#xB7EC;&#xB098; CD40L potently &#xBA74;&#xC5ED; &#xAE30;&#xB2A5; &#xBC0F; CD40 &#xC218;&#xC6A9; &#xCCB4;&#xC640;&#xC758; &#xC0C1;&#xD638; &#xC791;&#xC6A9;&#xC5D0; &#xC758;&#xD574; &#xCDA9;&#xBD84;&#xD788; &#xBC0F; &#xD608;&#xC18C;&#xD310; integrin GPIIb/IIIa &#xC870;&#xC808;, &#xC774;&#xD6C4; &#xAE00;&#xB85C;&#xBC8C; &#xC800;&#xD574; &#xD0C0;&#xD611;&#xC758; &#xD638;&#xC2A4;&#xD2B8; &#xBC29;&#xC5B4; &#xBC0F; &#xC784;&#xC0C1; &#xC2E4;&#xD5D8;&#xC5D0;&#xC11C; &#xC0DD;&#xC131; &#xB41C; thromboembolic &#xD569;&#xBCD1;&#xC99D;. &#xC6B0;&#xB9AC;&#xB294; &#xCD5C;&#xADFC; CD40L cd40&#xC640; &#xB2E4;&#xB978; &#xC218;&#xC6A9; &#xCCB4;&#xB85C; &#xC11C; &#xC81C;&#xC548; &#xB41C; Mac 1 &#xBCC4;&#xB3C4;&#xB85C; atherogenesis&#xC744; &#xC911;&#xC7AC; &#xD588;&#xB2E4;.

&#xB3C4;&#xBA54;&#xC778; &#xB9E5; 1 cd40l&#xC758; &#xBC14;&#xC778;&#xB529; EQLKKSKTL &#xBAA8;&#xD2F0;&#xBE0C;&#xB97C; &#xD3EC;&#xD568; &#xD55C;&#xB2E4; &#xBC0F; &#xBC31;&#xD608;&#xAD6C; &#xC2E0;&#xADDC; &#xBAA8;&#xC9D1; &#xBC0F; &#xC544; &#xD14C; &#xB86C;-&#xC911;&#xC7AC; &#xD558;&#xC9C0;&#xB9CC; &#xBA74;&#xC5ED;&#xACFC; &#xC950;&#xC758; &#xD608;&#xC804; &#xC99D;&#xC5D0; &#xC601;&#xD5A5;&#xC744; &#xBBF8;&#xCE58;&#xC9C0; &#xC54A;&#xC2B5;&#xB2C8;&#xB2E4;.

Figuras de CD40L en enfermedades inflamatorias cr&oacute;nicas como la aterosclerosis. Sin embargo, puesto que CD40L potentemente regula la funci&oacute;n inmune y hemostasia por interacci&oacute;n con receptor CD40 y la plaqueta integrina GPIIb/IIIa, sus concesiones de inhibici&oacute;n global sede defensa y generadas complicaciones tromboemb&oacute;licas en ensayos cl&iacute;nicos. Nos inform&oacute; recientemente que el CD40L media aterog&eacute;nesis independientemente de CD40 y Mac-1 propuesto como un receptor suplente.

Uni&oacute;n de CD40L a dominios de Mac-1 consiste en el adorno EQLKKSKTL y media el reclutamiento leucocitario y aterosclerosis--pero no afecta a inmunidad y trombosis en ratones.

The majority of peripheral serotonin is stored in platelets, which secrete it on activation. Serotonin releases Weibel-Palade bodies (WPBs) and we asked whether absence of platelet serotonin affects neutrophil recruitment in inflammatory responses. Tryptophan hydroxylase (Tph)1–deficient mice, lacking non-neuronal serotonin, showed mild leukocytosis compared with wild-type (WT), primarily driven by an elevated neutrophil count. Despite this, 50% fewer leukocytes rolled on unstimulated mesenteric venous endothelium of Tph1(-/-) mice. The velocity of rolling leukocytes was higher in Tph1(-/-) mice, indicating fewer selectin-mediated interactions with endothelium. Stimulation of endothelium with histamine, a secretagogue of WPBs, or injection of serotonin normalized the rolling in Tph1(-/-) mice. Diminished rolling in Tph1(-/-) mice resulted in reduced firm adhesion of leukocytes after lipopolysaccharide treatment. Blocking platelet serotonin uptake with fluoxetine in WT mice reduced serum serotonin by > 80% and similarly reduced leukocyte rolling and adhesion. Four hours after inflammatory stimulation, neutrophil extravasation into lung, peritoneum, and skin wounds was reduced in Tph1(-/-) mice, whereas in vitro neutrophil chemotaxis was independent of serotonin. Survival of lipopolysaccharide-induced endotoxic shock was improved in Tph1(-/-) mice. In conclusion, platelet serotonin promotes the recruitment of neutrophils in acute inflammation, supporting an important role for platelet serotonin in innate immunity.

Platelet serotonin promotes the recruitment of neutrophils to sites of acute inflammation in mice.

We tested the feasibility of local thrombolytic therapy via a novel hollow flexible and perforated wire in a mouse model of deep vein thrombosis. Inferior vena cava (IVC) thrombosis was induced by vessel wall exposure to ferric chloride after laparotomy in anesthetized C57Bl/6 mice. Thrombus formation was visualized by intravital microscopy of rhodamine-labeled platelets and leukocytes. A nitinol hypotube coronary wire with perforated tip was inserted via a 0.8 × 40 mm canula into the IVC lumen distal to the site of ferric chloride exposure. Either tissue plasminogen activator (tPA, alteplase) or saline (control) was administered via the platinum wire distal to the thrombus, avoiding mechanical fragmentation. Thrombus size was assessed by immunohistochemistry (platelet CD41 staining). Intravital microscopy of the IVC demonstrated platelet-containing thrombus growth starting 1 min after ferric chloride exposure. Alteplase administration resulted in significant thrombus size reduction within 10-20 min observed by intravital microscopy and confirmed by histological assessment of IVC cross-sections. Saline-treated mice (n = 4) demonstrated near total IVC occlusion with thrombotic material (84 ± 8% of cross-sectional area in serial sections), whereas alteplase-treated mice showed a dose-dependent decrease of thrombotic area [56 ± 5% with 1.5, 39 ± 4 % with 15 and 21 ± 6% with 150 mg/kg, respectively (n = 4)]. We demonstrate that a flexible hollow and perforated wire enables the successful application of thrombolytic therapy to IVC thrombi in mice without vessel wall perforation. Flexible wire-based thrombolytic therapy appears to be a safe and reliable method for thrombus dissolution even in fragile small veins and may become a promising strategy for targeted therapy of small vessel thrombosis.

A novel hollow and perforated flexible wire allows the safe and effective local application of thrombolytic therapy in a mouse model of deep vein thrombosis.

Activated platelets release serotonin at sites of inflammation where it acts as inflammatory mediator and enhances recruitment of neutrophils. Chronic treatment with selective serotonin reuptake inhibitors (SSRI) depletes the serotonin storage pool in platelets, leading to reduced leukocyte recruitment in murine experiments. Here, we examined the direct and acute effects of SSRI on leukocyte recruitment in murine peritonitis.

Nadine Herr

Department of Cardiology and Angiology I

Heart Center, University of Freiburg

Intravital Microscopy of Leukocyte-endothelial and Platelet-leukocyte Interactions in Mesenterial Veins in Mice

Nadine Herr

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

Heart Center, University of Freiburg

Intravital Microscopy of Leukocyte-endothelial and Platelet-leukocyte Interactions in Mesenterial Veins in Mice

Department of Cardiology and Angiology I