XHTW è un WM Keck Foundation Scholar Illustri giovani nella ricerca medica, ed è anche supportato da NIH / NHLBI concede R01-R01-HL089598 e R01HL091947 e Distrofia Muscolare associazione concedere # 69238. RJvO è il destinatario della Fellowship dell&#39;American Physiological Society 2008-2010 post-dottorato in Physiological Genomics. Questo lavoro è supportato anche in parte dall&#39;Alleanza Fondation Leducq per CaMKII segnalazione di Cuore.

Parte 1: Preparazione del campo operatorio <ol><li> Il campo operativo è disinfettata con il 75% di alcool isopropilico. </li><li> Assicurarsi che il pad di riscaldamento è acceso e alla giusta temperatura. Un sistema consigliato è una pompa Gaymar acqua circolante collegato ad un pad terapia che viene mantenuta a 37 ° C ± 1 ° C. E &#39;importante mantenere la temperatura corporea normale durante l&#39;intervento chirurgico per evitare una rapida diminuzione della frequenza cardiaca. </li><li> Strumenti chirurgici vengono sterilizzati in uno sterilizzatore a caldo perla prima dell&#39;intervento chirurgico. Per questa procedura, avrete bisogno dei seguenti strumenti chirurgici: forbici, pinzette ricurve corso X 2, fine 45 ° angolo X 2 pinze, forbici primavera angolato, retrattore del torace, e porta aghi. </li><li> Applicatori di cotone dovrebbe essere a portata di mano in caso di sanguinamento. </li></ol> Parte 2: Preparazione e intubazione di topi <ol><li> I topi vengono anestetizzati in una camera di induzione con isofluorano al 2% miscelato con 0,5 -1,0 l / min 100% O 2. </li><li> Tosatrici vengono utilizzate per accorciare il pelo dalla scollatura a livello del torace a metà. </li><li> Il mouse è posto in posizione supina in cima a una piastra elettrica al fine di mantenere la temperatura corporea. </li><li> Un elastico viene posizionato sopra i denti anteriori l&#39;animale s di estendere il collo. Utilizzando pinze curve in una mano, la lingua è delicatamente manipolato a lato. Con l&#39;altra mano, l&#39;intubazione endotracheale viene eseguita utilizzando tubi PE 90. Il tubo endotracheale viene poi collegato ad un volume-ciclismo pedalato roditori ventilatore Harvard a 125-150 respiri / minuto e un volume corrente di 0.1-0,3 ml. Durante la procedura chirurgica, l&#39;anestesia viene mantenuta al 1,5-2% isoflurano con 0,5-1,0 L / min 100% O 2. Corretto livello di anestesia è verificata applicando pressione sul letto ungueale del mouse (convergenza pizzico reflex). </li><li> Il campo chirurgico viene disinfettato con una soluzione di Betadine seguito dal 70% di alcol. Questa procedura viene ripetuta tre volte. </li><li> Per prevenire la contaminazione del campo operatorio durante l&#39;operazione, un telino sterile è posta sopra il mouse lasciando solo il campo operatorio esposti. </li><li> Un insieme di guanti sterili viene utilizzato per ogni individuo del mouse. </li></ol> Parte 3: legatura dell&#39;aorta trasversale <ol><li> Toracotomia parziale a seconda costa viene eseguito al microscopio chirurgico e lo sterno ritratta con un divaricatore petto. </li><li> Belle punta a 45 ° pinza angolata sono usati per separare delicatamente il tessuto del timo e il grasso dall&#39;arco aortico. </li><li> A seguito di identificazione dell&#39;aorta trasversale, un piccolo pezzo di una sutura 6,0 seta è posto tra le arterie innominate e carotide sinistra (Figura 1). </li><li> Due nodi sono sciolti legato intorno al aorta trasversali e un piccolo pezzo di un ago calibro 27 ½ ottuso è posto parallelamente alla aorta trasversale. Il primo nodo è legato rapidamente contro l&#39;ago, seguito dal secondo e l&#39;ago prontamente rimossi al fine di ottenere una costrizione di 0,4 millimetri di diametro. Nei topi di controllo sham, l&#39;intera procedura è identica tranne che per la legatura dell&#39;aorta. </li><li> Il retrattore petto viene rimosso e il deflusso del ventilatore pizzicato off per 2 secondi per ri-gonfiare i polmoni. </li><li> La gabbia toracica è chiuso con un 6,0 sutura prolene con un modello sutura interrotta. </li><li> La pelle è chiuso con un 6,0 sutura prolene con un modello di sutura continua. </li></ol> Parte 4: recupero post-operatorio <ol><li> Per analgesia post-operatoria, il mouse viene iniettato con buprenorfina (0,1 mg / kg) per via intraperitoneale. Se ci sono segni di disidratazione dopo l&#39;intervento chirurgico, soluzione salina sterile viene dato via intraperitoneale. </li><li> L&#39;anestesia è gradualmente abbassata in posizione off e il tubo endotracheale rimosso quando segni di respirazione spontanea si verificano. </li><li> Il mouse viene spostato nella posizione prona e ha permesso di recuperare su una piastra elettrica. </li></ol> Parte 5: Conferma di legatura successo della Aorta trasversale <ol><li> Una settimana dopo la TAC, il mouse viene nuovamente anestetizzati per determinare il grado di sovraccarico di pressione indotta dalla legatura dell&#39;aorta trasversale. Come descritto sopra, l&#39;anestesia viene mantenuta al 1,5-2% isoflurano con 0,5-1,0 L / min 100% O 2 e la temperatura corporea a 37 ° C ± 1 ° C. </li><li> A 20 MHz sonda doppler è posto sul lato destro e sinistro del collo in un angolo di 45 ° per rilevare velocità di flusso. Un computer basato su processore di segnale Doppler (Indo Instruments, Houston, TX) è stato utilizzato per visualizzare e memorizzare i segnali Doppler 5. </li><li> A seconda del grado di sovraccarico di pressione necessaria per il protocollo sperimentale, i topi solo con una carotide destra (RC) / carotide sinistra (LC) il rapporto di flusso entro un certo intervallo sono inclusi per ulteriori analisi. Ad esempio, un moderato grado di sovraccarico di pressione porta ad un rapporto di 5-8, mentre una costrizione stretto con conseguente sovraccarico di pressione gravi conduce toa rapporto di 8-10 (Figura 5.3A). Un animale finto (ma non gestite legatura), tuttavia, dovrebbe avere un rapporto di ~ 1 (Figura 5.3b). </li></ol> 6: Risultati Rappresentante La sopravvivenza tipico chirurgico dopo TAC nei topi wild-type è di circa 80-90%. Successo legatura chirurgica dell&#39;aorta trasversale porterà ad un rapporto di velocità di flusso Doppler tra destra e sinistra dell&#39;arteria carotidea (RC / LC) di 5-10 (Figura 1). Rispetto ai topi sham-operati (Figura 2A), i topi con sovraccarico di pressione sono tenuti a sviluppare l&#39;ipertrofia cardiaca entro 1-2 settimane, e la dilatazione cardiaca dopo 6-8 settimane, a seconda della tenuta della costrizione (Figura 2B). La mortalità durante il primo mese dopo la TAC è generalmente basso (&lt;20%) nei topi wild-type, anche se topi geneticamente modificati possono presentare tassi di sopravvivenza diversi. 6,7 <img src="/files/ftp_upload/1729/1729fig1tmb.jpg" alt="Figura 1" border="0" /> Figura 1. La conferma della legatura Aorta trasversale utilizzando misurazione del flusso Doppler. Rappresentante segnali di velocità Doppler del diritto (RC) e carotide sinistra (LC) arterie 6 giorni dopo la TAC. Queste registrazioni dimostrano una legatura di successo con un RC / LC rapporto tra il flusso di ~ 6.8. Si prega di <a href="https://www.jove.com/files/ftp_upload/1729/1729fig1.jpg">cliccare qui</a> per vedere una versione più grande della figura 1. <img src="/files/ftp_upload/1729/1729fig2.jpg" alt="Figura 2" /> Figura 2. Immagini rappresentative di tutto cuore Dopo TAC. A. Sham azionati cuore selvaggio del mouse tipo. B. Cuore selvaggio del mouse tipo 16 settimane post-TAC. Ogni linea = 1 mm.

<ol>
	<li>Rockman, H. A., Ross, R. S., Harris, A. N., Knowlton, K. U., Steinhelper, M. E., Field, L. J., Ross, J., Chein, K. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Segregation+of+atrial-specific+and+inducible+expression+of+an+atrial+natriuretic+factor+transgene+in+an+in+vivo+murine+model+of+cardiac+hypertrophy.">Segregation of atrial-specific and inducible expression of an atrial natriuretic factor transgene in an in vivo murine model of cardiac hypertrophy.</a> Proc. Natl. Acad. Sci. USA. 88, 8277-8281 (1991).</li><li>Heineke, J., Molkentin, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+of+cardiac+hypertrophy+by+intracellular+signaling+pathways.">Regulation of cardiac hypertrophy by intracellular signaling pathways.</a> Nat. Rev. Mol. Cell. Biol. 7, 589-600 (2006).</li><li>Hartley, C. J., Reddy, A. K., Madala, S., Michael, L. H., Entman, M. L., Taffett, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doppler+estimation+of+reduced+coronary+flow+reserve+in+mice+with+pressure+overload+cardiac+hypertrophy.">Doppler estimation of reduced coronary flow reserve in mice with pressure overload cardiac hypertrophy.</a> Ultrasound Med. Biol. 34, 892-901 (2008).</li><li>Reddy, A. K., Taffett, G. E., Li, Y. -. H., Lim, S. -. W., Pham, T. T., Pocius, J. S., Entman, M. L., Michael, L. H., Hartley, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doppler+signal+processing+for+use+in+mice:+applications.">Doppler signal processing for use in mice: applications.</a> IEEE Trans. Biomed. Eng. 52, 1771-1783 (2005).</li><li>Li, Y. -. H., Reddy, A. K., Ochoa, L. N., Pham, T. T., Hartley, C. J., Micheal, L. H., Entman, M. L., Taffett, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+age+on+peripheral+vascular+response+to+transverse+aortic+banding+in+mice.">Effect of age on peripheral vascular response to transverse aortic banding in mice.</a> J. Gerontol. 58A, 895-899 (2003).</li><li>Bourajjaj, M., Armand, A. S., Martins, d. a. C. o. s. t. a., Weijts, P. A., Nagel, B. V. a. n. d. e. r., Heeneman, R., Wehrens, S., Windt, X. H. D. e., J, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NFATc2+is+a+necessary+mediator+of+calcineurin-dependent+cardiac+hypertrophy+and+heart+failure.">NFATc2 is a necessary mediator of calcineurin-dependent cardiac hypertrophy and heart failure.</a> J. Biol. Chem. 28, 22295-22303 (2008).</li><li>Oort, R. J. v. a. n., Respress, J. L., Li, N., Reynolds, C., Almeida, A. C. D. e., Skapura, D. G., Windt, L. J. D. e., Wehrens, X. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Accelerated+development+of+pressure+overload+induced+cardiac+hypertrophy+and+dysfunction+in+an+RyR2-R176Q+knockin+mouse+model.">Accelerated development of pressure overload induced cardiac hypertrophy and dysfunction in an RyR2-R176Q knockin mouse model.</a> Hypertension. 55, (2010).</li><li>Tarnavski, O., McMullen, J. R., Schinke, M., Nie, Q., Kong, S., Izumo, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+cardiac+surgery:+comprehensive+techniques+for+the+generation+of+mouse+models+of+human+diseases+and+their+application+for+genomic+studies.">Mouse cardiac surgery: comprehensive techniques for the generation of mouse models of human diseases and their application for genomic studies.</a> Physiol Genomics. 16, 349-360 (2004).</li></ol>

TAC, che imita la stenosi aortica umana, è un metodo comune per indurre l&#39;ipertrofia cardiaca e insufficienza cardiaca nei topi. Siti alternativi per costrizione aortica includono l&#39;aorta ascendente e addominale. Restringimento dell&#39;aorta ascendente fornisce un sovraccarico estrema e più rapido del ventricolo sinistro (LV). Al contrario, la costrizione dell&#39;aorta addominale lascia intatta una porzione più ampia della circolazione come mezzo di eventuale risarcimento. 8 Perciò, TAC è spess...

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		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Gaymar T/pump</td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>Model TP-500</td>
<td>Circulating water pump</td>
</tr>
<tr>
<td>Temp. therapy pads</td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>60-3414</td>
<td>&nbsp;</td>
<tr>
<td>Hot bead sterilizer</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>18000-45</td>
<td>&nbsp;</td>
<tr>
<td>Blunt scissors</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5980</td>
<td>&nbsp;</td>
<tr>
<td>Angled spring scissors</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5668</td>
<td>&nbsp;</td>
<tr>
<td>Chest retractor</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>17002-02</td>
<td>&nbsp;</td>
<tr>
<td>Course curved forceps</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5138</td>
<td>&nbsp;</td>
<tr>
<td>Dumont fine 45&deg; angled forceps</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>11253-25</td>
<td>&nbsp;</td>
<tr>
<td>Needle holder</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>12565-14</td>
<td>&nbsp;</td>
<tr>
<td>Electric shaver</td>
<td>&nbsp;</td>
<td>General Electric</td>
<td>&nbsp;</td>
<td>For shaving mouse fur</td>
</tr>
<tr>
<td>PE 90 tubing</td>
<td>&nbsp;</td>
<td>BD</td>
<td>427420</td>
<td>For intubation</td>
</tr>
<tr>
<td>MiniVent (ventilator)</td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>Type 845</td>
<td>&nbsp;</td>
<tr>
<td>Betadine</td>
<td>&nbsp;</td>
<td>Fisher</td>
<td>19-027132</td>
<td>May be purchased at medical supply store</td>
</tr>
<tr>
<td>Sterile gloves</td>
<td>&nbsp;</td>
<td>Mckesson</td>
<td>20-1565</td>
<td>&nbsp;</td>
<tr>
<td>Stereo microscope</td>
<td>&nbsp;</td>
<td>Unico</td>
<td>ZM186</td>
<td>&nbsp;</td>
<tr>
<td>6-0 silk suture</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>18020-60</td>
<td>&nbsp;</td>
<tr>
<td>27 &frac12; gauge needle</td>
<td>&nbsp;</td>
<td>BD</td>
<td>305109</td>
<td>&nbsp;</td>
<tr>
<td>6-0 prolene suture</td>
<td>&nbsp;</td>
<td>Mckesson</td>
<td>3286</td>
<td>&nbsp;</td>
<tr>
<td>Buprenorphine</td>
<td>&nbsp;</td>
<td>Institutional Animal Facility</td>
<td>&nbsp;</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

transverse aortic constriction in mice

Restringimento aortico trasversale (TAC) nel topo è un modello comunemente usato sperimentale per un sovraccarico di pressione indotta ipertrofia cardiaca e scompenso cardiaco.<sup&gt; 1</sup&gt; TAC porta inizialmente a ipertrofia compensato del cuore, che spesso è associato ad un miglioramento temporaneo della contrattilità cardiaca. Nel corso del tempo, però, la risposta al cronico sovraccarico emodinamico diventa disadattamento, con conseguente dilatazione cardiaca e scompenso cardiaco.<sup&gt; 2</sup&gt; Il modello murino TAC è stato convalidato da Rockman<em&gt; Et al</em&gt;.<sup&gt; 1</sup&gt;, E da allora è stato ampiamente utilizzato come uno strumento prezioso per imitare umana malattie cardiovascolari e chiarire fondamentali processi di segnalazione coinvolte nella risposta ipertrofica cardiaca ed insufficienza cardiaca sviluppo. Rispetto ad altri modelli sperimentali di insufficienza cardiaca, come ad esempio l&#39;occlusione completa della discendente anteriore sinistra (LAD) delle arterie coronarie, TAC offre un modello più riproducibile di ipertrofia cardiaca e un corso di tempo più graduale per lo sviluppo di insufficienza cardiaca. Qui, descriviamo un passo-passo per eseguire procedura chirurgica TAC nei topi. Per determinare il livello di sovraccarico di pressione prodotta dalla legatura aortica, una sonda ad alta frequenza Doppler viene utilizzato per misurare il rapporto tra velocità di flusso sanguigno nelle arterie carotide destra e sinistra.<sup&gt; 3, 4</sup&gt; Con i tassi di sopravvivenza chirurgica del 80-90%, banding aortico trasversale è una tecnica efficace per indurre l&#39;ipertrofia ventricolare sinistra ed insufficienza cardiaca nei topi.

Watch this Scientific Journal Video about Transverse Aortic Constriction in Mice at JoVE.com

Transverse Aortic Constriction in Mice

Restringimento aortico trasversale (TAC) nel topo è un modello comunemente usato sperimentale per studiare meccanismi che sottendono l&#39;ipertrofia cardiaca e scompenso cardiaco sviluppo. Qui si descrivono le modalità per costrizione dell&#39;aorta per creare un grado riproducibile di ipertrofia cardiaca nei topi.

Aortica costrizione trasversale nei topi

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model for pressure overload-induced cardiac hypertrophy and heart ...

Research

JoVE Journal

Biology

66.7K Views.  Baylor College of Medicine (BCM). Restringimento aortico trasversale (TAC) nel topo è un modello comunemente usato sperimentale per studiare meccanismi che sottendono l'ipertrofia cardiaca e scompenso cardiaco sviluppo. Qui si descrivono le modalità per costrizione dell'aorta per creare un grado riproducibile di ipertrofia cardiaca nei topi.

Video: Transverse Aortic Constriction in Mice

Small Bowel Transplantation In Mice

Since 1990, the development of tacrolimus-based immunosuppression and improved surgical techniques, the increased array of potent immunosuppressive medications, infection prophylaxis, and suitable patient selection helped improve actuarial graft and patient survival rates for all types of intestine transplantation. Patients with irreversible intestinal failure and complications of parenteral nutrition should now be routinely considered for small intestine transplantation. However, Survival rates for small intestinal transplantation have been slow to improve compares increasingly favorably with renal, liver, heart and lung. The small bowel transplantation is still unsatisfactory compared with other organs. Further progress may depend on better understanding of immunology and physiology of the graft and can be greatly facilitated by animal models. A wider use of mouse small bowel transplantation model is needed in the study of immunology and physiology of the transplantation gut as well as efficient methods in diagnosing early rejection. However, this model is limited to use because the techniques involved is an extremely technically challenging. We have developed a modified technique. When making anastomosis of portal vein and inferior vena cava, two stay sutures are made at the proximal apex and distal apex of the recipient s inferior vena cava with the donor s portal vein. The left wall of the inferior vena cava and donor s portal vein is closed with continuing sutures in the inside of the inferior vena cava after, after one knot with the proximal apex stay suture the right wall of the inferior vena cava and the donor s portal vein are closed with continuing sutures outside the inferior vena cave with 10-0 sutures. This method is easier to perform because anastomosis is made just on the one side of the inferior vena cava and 10-0 sutures is the right size to avoid bleeding and thrombosis.  In this article, we provide details of the technique to supplement the video.

The mouse small bowel transplantation model has been recognized as an important tool to study mechanismes of immune rejection and screen new immunosuppressive drugs. However, this model is limited to use because the techniques involved is an extremely technically challenge. Now we introduce the modified technique.

Trapianto di intestino piccolo Nei topi

Since 1990, the development of tacrolimus-based immunosuppression and improved surgical techniques, the increased array of potent immunosuppressive ...

Ricerca

Biologia

Aortic Ring Assay

Angiogenesis, the sprouting of blood vessels from preexisting vasculature is associated with both natural and pathological processes. Various angiogenesis assays involve the study of individual endothelial cells in culture conditions (1). The aortic ring assay is an angiogenesis model that is based on organ culture. In this assay, angiogenic vessels grow from a segment of the aorta (modified from (2)). Briefly, mouse thoracic aorta is excised, the fat layer and adventitia are removed, and rings approximately 1 mm in length are prepared. Individual rings are then embedded in a small solid dome of basement matrix extract (BME), cast inside individual wells of a 48-well plate. Angiogenic factors and inhibitors of angiogenesis can be directly added to the rings, and a mixed co-culture of aortic rings and other cell types can be employed for the study of paracrine angiogenic effects. Sprouting is observed by inspection under a stereomicroscope over a period of 6-12 days. Due to the large variation caused by the irregularities in the aortic segments, experimentation in 6-plicates is strongly advised. Neovessel outgrowth is monitored throughout the experiment and imaged using phase microscopy, and supernatants are collected for measurement of relevant angiogenic and anti-angiogenic factors, cell death markers and nitrite.

Angiogenesis, the sprouting of blood vessels from pre-existing vasculature, is associated with both natural and pathological processes. Here we demonstrate an aortic ring assay that allows angiogenic potentiators and inhibitors to be directly added to aortic rings in culture. Sprouting and neovessel outgrowth can be determined by inspecting the aortic rings over a period of 6-12 days.

Aortica Anello Assay

Angiogenesis, the sprouting of blood vessels from preexisting vasculature is associated with both natural and pathological processes. Various angiogenesis ...

Ambulatory ECG Recording in Mice

Telemetric ECG recording in mice is essential to understanding the mechanisms behind arrhythmias, conduction disorders, and sudden cardiac death. Although the surface ECG is utilized for short-term measurements of waveform intervals, it is not practical for long-term studies of heart rate variability or the capture of rare episodes of arrhythmias. Implantable ECG telemeters offer the advantages of simple surgical implantation, long-term recording of electrograms in ambulatory mice, and scalability with simultaneous recordings of multiple animals. Here, we present a step-by-step guide to the implantation of telemeters for ambulatory ECG recording in mice. Careful adherence to aseptic technique is required for favorable survival results with the possibility of implantation and recording from weeks to months. Thus, implantable ECG telemetry is a valuable tool for detection of critical information on cardiac electrophysiology in ambulatory animal models such as the mouse. 

Telemetric ECG has emerged as an essential tool in evaluating animal models for cardiac arrhythmias and sudden cardiac death. Here, we present a stepwise guide to telemetric ECG recordings for application in long-term ambulatory ECG monitoring in mice.

La registrazione ECG ambulatoriale nei topi

Telemetric ECG recording in mice is essential to understanding the mechanisms behind arrhythmias, conduction disorders, and sudden cardiac death. Although ...

Intravenous Injections in Neonatal Mice

Intravenous injection is a clinically applicable manner to deliver therapeutics. For adult rodents and larger animals, intravenous injections are technically feasible and routine. However, some mouse models can have early onset of disease with a rapid progression that makes administration of potential therapies difficult. The temporal (or facial) vein is just anterior to the ear bud in mice and is clearly visible for the first two days after birth on either side of the head using a dissecting microscope. During this window, the temporal vein can be injected with volumes up to 50 &#956;l. The injection is safe and well tolerated by both the pups and the dams. A typical injection procedure is completed within 1-2 min, after which the pup is returned to the home cage. By the third postnatal day the vein is difficult to visualize and the injection procedure becomes technically unreliable. This technique has been used for delivery of adeno-associated virus (AAV) vectors, which in turn can provide almost body-wide, stable transgene expression for the life of the animal depending on the viral serotype chosen.

Animal models of pediatric disease can experience early onset and aggressive disease progression. Clinically relevant therapy delivery to young mouse models can be technically difficult. This protocol describes a non-invasive intravenous injection method for newborn mice within the first two postnatal days of life.

Le iniezioni per via endovenosa nel topo neonatale

Intravenous injection is a clinically applicable manner to deliver therapeutics. For adult rodents and larger animals, intravenous injections are ...

Isolation and Primary Culture of Mouse Aortic Endothelial Cells

The vascular endothelium is essential to normal vascular homeostasis. Its dysfunction participates in various cardiovascular disorders. The mouse is an important model for cardiovascular disease research. This study demonstrates a simple method to isolate and culture endothelial cells from the mouse aorta without any special equipment. To isolate endothelial cells, the thoracic aorta is quickly removed from the mouse body, and the attached adipose tissue and connective tissue are removed from the aorta. The aorta is cut into 1 mm rings. Each aortic ring is opened and seeded onto a growth factor reduced matrix with the endothelium facing down. The segments are cultured in endothelial cell growth medium for about 4 days. The endothelial sprouting starts as early as day 2. The segments are then removed and the cells are cultured continually until they reach confluence. The endothelial cells are harvested using neutral proteinase and cultured in endothelial cell growth medium for another two passages before being used for experiments. Immunofluorescence staining indicated that after the second passage the majority of cells were double positive for Dil-ac-LDL uptake, Lectin binding, and CD31 staining, the typical characteristics of endothelial cells. It is suggested that cells at the second to third passages are suitable for in vitro and in vivo experiments to study the endothelial biology. Our protocol provides an effective means of identifying specific cellular and molecular mechanisms in endothelial cell physiopathology.

The vascular endothelial cells play a significant role in many important cardiovascular disorders. This article describes a simple method to isolate and expand endothelial cells from the mouse aorta without using any special equipment. Our protocol provides an effective means of identifying mechanisms in endothelial cell physiopathology.

Isolamento e colture primarie di cellule di topo aortica endoteliali

The vascular endothelium is essential to normal vascular homeostasis. Its dysfunction participates in various cardiovascular disorders. The mouse is an ...

Induction of Right Ventricular Failure by Pulmonary Artery Constriction and Evaluation of Right Ventricular Function in Mice

The mechanism of right ventricular failure (RVF) requires clarification due to the uniqueness, high morbidity, high mortality, and refractory nature of RVF. Previous rat models imitating RVF progression have been described. Compared with rats, mice are more accessible, economical, and widely used in animal experiments. We developed a pulmonary artery constriction (PAC) approach which is comprised of banding the pulmonary trunk in mice to induce right ventricular (RV) hypertrophy. A special surgical latch needle was designed that allows for easier separation of the aorta and the pulmonary trunk. In our experiments, the use of this fabricated latch needle reduced the risk of arteriorrhexis and improved the surgical success rate to 90%. We used different padding needle diameters to precisely create quantitative constriction, which can induce different degrees of RV hypertrophy. We quantified the degree of constriction by evaluating the blood flow velocity of the PA, which was measured by noninvasive transthoracic echocardiography. RV function was precisely evaluated by right heart catheterization at 8 weeks after surgery. The surgical instruments made inhouse were composed of common materials using a simple process that is easy to master. Therefore, the PAC approach described here is easy to imitate using instruments made in the lab and can be widely used in other labs. This study presents a modified PAC approach that has a higher success rate than other models and an 8-week postsurgery survival rate of 97.8%. This PAC approach provides a useful technique for studying the mechanism of RVF and will enable an increased understanding of RVF.

Here, we provide a useful approach for studying the mechanism of right ventricular failure. A more convenient and efficient approach to pulmonary artery constriction is established using surgical instruments made inhouse. In addition, methods to evaluate the quality of this approach by echocardiography and catheterization are provided.

Induzione del fallimento ventricolare destro da parte della costrizione dell'arteria polmonare e valutazione della funzione ventricolare destra nei topi

The mechanism of right ventricular failure (RVF) requires clarification due to the uniqueness, high morbidity, high mortality, and refractory nature of ...

Inducing Apical Periodontitis in Mice

The mechanisms involved in local induced inflammation can be studied using several available animal models. One of these is the induction of apical periodontitis (AP). Apical periodontitis is a common pathology of an inflammatory nature in the periodontal tissues surrounding the tooth root. In order to better understand the nature and mechanism of this pathology it is advantageous to perform the procedure in mice. The induction of this odontogenic inflammation is achieved by drilling into the mouse tooth until the dental pulp is exposed. Next, the tooth pulp remains exposed to be contaminated by the natural oral flora over time, causing apical periodontitis. After this time period, the animal is sacrificed, and the tooth and the jaw bone can be analyzed in various ways. Typical analyses include micro-CT imaging (to evaluate bone resorption), histological staining, immunohistochemistry, and RNA expression. This protocol is useful for research in the field of oral biology to better understand this inflammatory process in an in vivo experimental setting with uniform conditions. The procedure requires a careful handling of the mice and the isolated jaw, and a visual demonstration of the technique is useful. All technical aspects of the procedures leading to induced apical periodontitis and its characterization in a mouse model are demonstrated.

Here, we present a protocol to locally induce apical periodontitis in mice. We show how to drill a hole in the mouse&#39;s tooth and expose its pulp, in order to cause local inflammation. Analysis methods to investigate the nature of this inflammation, such as micro-CT and histology, are also demonstrated.

Indurre la parodontite apicale nei topi

The mechanisms involved in local induced inflammation can be studied using several available animal models. One of these is the induction of apical ...

Transverse Fracture of the Mouse Femur with Stabilizing Pin

Fracture repair is an essential function of the skeleton that cannot be reliably modeled in vitro. A mouse injury model is an efficient approach to test whether a gene, gene product or drug influences bone repair because murine bones recapitulate the stages observed during human fracture healing. When a mouse or human breaks a bone, an inflammatory response is initiated, and the periosteum, a stem cell niche surrounding the bone itself, is activated and expands. Cells residing in the periosteum then differentiate to form a vascularized soft callus. The transition from the soft callus to a hard callus occurs as the recruited skeletal progenitor cells differentiate into mineralizing cells, and the bridging of the fractured ends results in the bone union. The mineralized callus then undergoes remodeling to restore the original shape and structure of the healed bone. Fracture healing has been studied in mice using various injury models. Still, the best way to recapitulate this entire biological process is to break through the cross-section of a long bone that encompasses both cortices. This protocol describes how a stabilized, transverse femur fracture can be safely performed to assess healing in adult mice. A surgical protocol including detailed harvesting and imaging techniques to characterize the different stages of fracture healing is also provided.

This protocol describes a method to perform fractures on adult mice and monitor the healing process.

Frattura trasversale del femore del topo con perno stabilizzante

Fracture repair is an essential function of the skeleton that cannot be reliably modeled in vitro. A mouse injury model is an efficient approach to test ...

A Modified Technique for Transverse Aortic Constriction in Mice

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of pressure overload in mouse models. The main challenge of this procedure is to clearly visualize the transverse aortic arch and precisely band the target vessel. Classical approaches perform a partial thoracotomy to expose the transverse aortic arch. However, it is an open-chest model that causes a rather large surgical trauma and requires a ventilator during the surgery. To prevent unnecessary trauma and simplify the operating procedure, the aortic arch is approached via the proximal proportion of the sternum, reaching and binding the target vessel using a small self-made retractor that contains a snare. This procedure can be conducted without entering the pleura cavity and does not need a ventilator or microsurgical operation, which leaves the mice with physiological breathing patterns, simplifies the procedure, and significantly reduces operation time. Due to the less invasive approach and less operation time, mice can undergo fewer stress reactions and recover rapidly.

The present protocol describes a modified and simplified technique with a minimally invasive transverse aortic constriction (TAC) procedure using a self-made retractor. This procedure can be conducted without a ventilator or microscope and introduces pressure overload, eventually leading to cardiac hypertrophy or heart failure.

Una tecnica modificata per la costrizione aortica trasversale nei topi

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of ...

A Calcium Phosphate-Induced Mouse Abdominal Aortic Aneurysm Model

An abdominal aortic aneurysm (AAA) is a life-threatening cardiovascular disease that occurs worldwide and is characterized by irreversible dilation of the abdominal aorta. Currently, several chemically induced murine AAA models are used, each simulating a different aspect of the pathogenesis of AAA. The calcium phosphate-induced AAA model is a rapid and cost-effective model compared to the angiotensin II- and elastase-induced AAA models. The application of CaPO4 crystals to the mouse aorta results in elastic fiber degradation, loss of smooth muscle cells, inflammation, and calcium deposition associated with aortic dilation. This article introduces a standard protocol for the CaPO4-induced AAA model. The protocol includes material preparation, the surgical application of the CaPO4&#160;to the adventitia of the infrarenal abdominal aorta, the harvesting of aortas to visualize aortic aneurysms, and histological analyses in mice.

This protocol describes a calcium phosphate-induced abdominal aortic aneurysm (AAA) mouse model to study the pathological features and molecular mechanisms of AAAs.

Un modello di aneurisma dell'aorta addominale del topo indotto da fosfato di calcio

An abdominal aortic aneurysm (AAA) is a life-threatening cardiovascular disease that occurs worldwide and is characterized by irreversible dilation of the ...