Gli autori desiderano ringraziare John Marston esperto per la zootecnia e il Dott. Marco A. Passini di assistenza tecnica nelle fasi iniziali di questo progetto. Questo lavoro è stato finanziato dalla sovvenzione da parte del National Institutes of Health a CLL (R01NS41584; R01HD054413).

1. Iniezione intracerebroventricolare <ol><li> Il primo passo è la preparazione delle soluzioni madre l&#39;iniezione; queste soluzioni sono vettore virale, DNA plasmidico, droghe, e deve essere iniettato in condizioni sterili. </li><li> Mescolare un titolo desiderato di vettore virale (5-7 totale mL) con il 0,05% w / v in PBS tripano blu per la visualizzazione del sito di iniezione. </li><li> Soluzione di DNA plasmidico (5 in totale l mL) contiene D-(+)-glucosio al 20% (w / v) (1 mL), trypan blu (0,05%) PBS (1 mL), il plasmide (~ 5 ~ g / L ~ ) (2 ~ L), e 2,5 kDa lineare PEI omopolimero (150 mm) (1 ~ L). </li><li> Immobilizzare il PND 2 neonati tramite crio-anestesia per 1-2 minuti. </li><li> L&#39;ago utilizzato per questa iniezione è un micro-litro micropipetta tarata vetro sterilizzati che è collegato ad una siringa da 3 ml 3 ml Hamilton attraverso un lungo tubo. </li><li> Rompere la punta dell&#39;ago (con spatola) per regolare la penetrazione per 2 mm nel cranio. Posizionare l&#39;ago in diagonale nella provetta contenente la soluzione di iniezione. Caricare la soluzione iniezione l&#39;ago con attenzione tirando lo stantuffo della siringa. </li><li> Dopo l&#39;assorbimento della soluzione, dissociare la siringa dal tubo; tirare lo stantuffo fino ulteriormente e quindi collegare l&#39;ago di nuovo. </li><li> Tenere l&#39;ago nella mano destra tra il pollice e le dita indice e posizionare la siringa tra il medio e l&#39;anulare con lo stantuffo toccando il palmo della mano destra. </li><li> Afferrare il topo immobilizzato con fermezza dalla pelle dietro la testa con la mano sinistra e posto su una fibra ottica la luce per illuminare le strutture anatomiche rilevanti che possono essere utilizzati come guida. </li><li> Inserire le 2 mm di profondità dell&#39;ago, perpendicolare alla superficie del cranio, in una località a circa 0,25 mm lateralmente alla sutura sagittale e 0,50-0,75 millimetri anteriore del neonatali sutura coronarica (Figura 1A). Quindi, spingere lo stantuffo con il palmo della mano destra molto lentamente e con attenzione. Monitor per le navi rottura o gonfiore del viso. </li><li> Rimuovere l&#39;ago 15 secondi dopo l&#39;interruzione del movimento di stantuffo per impedire il riflusso. </li><li> Per recuperare, mantenere i topi per 5-10 minuti in un contenitore riscaldato fino a quando il movimento e la reattività generale viene ripristinato. </li></ol> 2. L&#39;iniezione endovenosa attraverso la vena temporale / viso <ol><li> La soluzione da iniettare deve essere preparata e integrata con filtrata colorante alimentare verde alla diluizione 1:100 e iniettato in condizioni sterili. </li><li> Allegare un piccolo plastico luer alla fine di una siringa di vetro 100 microlitri Hamilton. Per il luer, collegare un manometro 33 ago ipodermico 0,25 pollici. Assicurarsi che tutte le parti siano saldamente collegati. </li><li> Caricare la siringa con il volume da iniettare, assicurando che non ci sia aria nella siringa, come l&#39;iniezione di aria nel vaso è letale. È spesso utile pipetta il volume da iniettare fino un pezzo di parafilm. Questo permette la visualizzazione chiara di assorbimento della soluzione nella siringa. </li><li> Un Transilluminatore Sight Wee è usato per visualizzare facilmente la vena temporale superficiale, o vena facciale, nel neonato (Figura 2A). Prima dell&#39;iniezione, fissare il neonato al transilluminatore con nastro chirurgico. Garza deve essere collocato tra il neonato e il nastro per evitare che il nastro di danneggiare la pelle dell&#39;animale. Fissare l&#39;animale al transilluminatore dalla sua parte, assicurando che il nastro non impedisce la respirazione. Collo del neonato deve essere delicatamente girato in modo che la vena facciale è facilmente visibile e il naso dovrebbe essere registrato per stabilizzare la testa. </li><li> Utilizzando una lente d&#39;ingrandimento 2.25X archetto per consentire la visualizzazione più semplice, inserire lentamente l&#39;ago nella vena. La vena è molto superficiale modo che l&#39;ago deve rimanere visibili sotto la pelle. </li><li> Infondere lentamente il volume di soluzione nella vena. La tinta verde permette una facile visualizzazione di iniezione. </li><li> Attendere 15 secondi prima di rimuovere l&#39;ago in quanto vi è un ritardo tra completamente lo stantuffo della siringa e l&#39;espulsione della soluzione rimanenti a causa del foro ago molto sottile. </li><li> Dopo aver rimosso l&#39;ago, usare una garza per applicare pressione sul sito di iniezione fino a quando il sanguinamento cessa. </li><li> Monitorare il neonato per i segni di sofferenza. Dopo una iniezione appropriate, nessun disagio palese devono essere osservati. Il neonato deve essere somministrata circa 5 minuti per riprendersi prima di essere restituita alla gabbia. </li><li> Sciacquare l&#39;ago / siringa unità con PBS tra iniezioni di sangue in l&#39;ago si intasano rapidamente a causa di foro di piccole dimensioni. </li></ol> 3. Rappresentante dei risultati: ICV iniezione: Il cervello può essere raccolte in diversi punti in tempo post-iniezione e visualizzati per dimostrare la tecnica di iniezione di successo. Se il colorante non viene aggiunto alla soluzione da iniettare, sarà difficile per verificare l&#39;accuratezza del aproiezione. Iniezione appropriate di uno dei ventricoli permetterà la distribuzione del trypan blu sul lato del cervello iniettato circa 10-15 minuti dopo l&#39;iniezione (Figura 1B). Una distribuzione uniforme di blu tripano in entrambi gli emisferi destro e sinistro cerebrale e bulbi olfattivi saranno visibili circa 60 minuti dopo l&#39;iniezione (figura 1C). Ciò è dovuto alla connettività dei ventricoli cerebrali che permetterà la diffusione del colorante blu per il ventricolo adiacenti. Iniezioni accurato può anche essere visualizzati dalla dispersione del colorante blu nel rostrale canale centrale spinale circa entro 12 ore dopo l&#39;iniezione (Figura 1D). Iniezioni inesatte si distingue per la mancanza di colore blu in emisferi cerebrali. In questo caso, la profondità dell&#39;ago non era sufficiente e la soluzione di iniezione è probabilmente distribuito sotto la pelle. Una possibilità alternativa è che l&#39;ago penetra troppo in profondità nel cervello, passando così al di là del ventricolo e la soluzione è svuotato sotto i ventricoli cerebrali. IV iniezione: accurata ed efficiente iniezione vena temporale / facciali possono essere visualizzati immediatamente dopo l&#39;iniezione. Dato che la soluzione viene iniettata, il colorante verde permette la visualizzazione della soluzione che entrano e che fluisce attraverso la vena facciale (Figura 2A). Inoltre, il colore della pelle alterata del mouse sarà osservato dopo l&#39;iniezione. Colore normale rosa del neonato è convertito al verde, a causa della presenza di colorante alimentare verde nella soluzione da iniettare (Figura 2B). L&#39;entità del cambiamento di colore verde osservato nel neonato dipende dalla concentrazione di colorante usato. Per la semplice visualizzazione di iniezione e durante un periodo di pratica, una diluizione 1:50 è raccomandato. Tuttavia, il tono della pelle drammaticamente alterato provoca spesso il cucciolo di essere rifiutato dalla madre e, quindi, la rimozione del colorante o di una concentrazione significativamente ridotto dovrebbero essere presi in considerazione durante le procedure sperimentali. Un&#39;iniezione imprecisi comporterà l&#39;accumulo di colorante verde sotto la pelle al sito di iniezione. Iniezione incompleta, che si verifica quando l&#39;apertura dell&#39;ago non è completamente inserito all&#39;interno della vena, si vede quando qualche soluzione fluisce nella vena e alcuni si accumula sotto la pelle. Se questa è osservato, l&#39;ago deve essere inserito ulteriormente nel vaso. <img src="/files/ftp_upload/2968/2968fig1.jpg" alt="Figura 1"/> Figura 1. Dimostrazione del successo iniezioni ICV in PDN2 topi. A) Schema di iniezione utilizzati in iniezioni ICV nei ventricoli. B) Cervello da ICV iniettato Pup con tripano blu in PBS sul ventricolo sinistro, foto scattata 15 minuti dopo l&#39;iniezione. C ) Cervello da ICV iniettato Pup con tripano blu in PBS a ventricolo sinistro, foto scattata 60 minuti dopo l&#39;iniezione. D) viene mostrato rostrate centrale canale spinale in ICV iniettato cucciolo con tripano blu in PBS sul ventricolo sinistro, foto scattata 12 ore dopo l&#39;iniezione. <img src="/files/ftp_upload/2968/2968fig2.jpg" alt="Figura 2"/> Figura 2. Dimostrazione del successo iniezioni IV PDN2 topi. A) Schema di vena facciale utilizzata per iniezioni IV. B) Pup IV iniettato con colorante verde in PBS rispetto ai non-iniettati cucciolata, Foto scattata immediatamente dopo l&#39;iniezione.

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D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rescue+of+the+spinal+muscular+atrophy+phenotype+in+a+mouse+model+by+early+postnatal+delivery+of+SMN.">Rescue of the spinal muscular atrophy phenotype in a mouse model by early postnatal delivery of SMN.</a> Nat. Biotechnol. 28, 271-274 (2010).</li><li>Foust, K. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intravascular+AAV9+preferentially+targets+neonatal+neurons+and+adult+astrocytes.">Intravascular AAV9 preferentially targets neonatal neurons and adult astrocytes.</a> Nat. Biotechnol. 27, 59-65 (2009).</li><li>Passini, M. A., Watson, D. J., Wolfe, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gene+delivery+to+the+mouse+brain+with+adeno-associated+virus.">Gene delivery to the mouse brain with adeno-associated virus.</a> Methods Mol. Biol. 246, 225-236 (2004).</li><li>Coady, T. H., Lorson, C. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trans-splicing-mediated+improvement+in+a+severe+mouse+model+of+spinal+muscular+atrophy.">Trans-splicing-mediated improvement in a severe mouse model of spinal muscular atrophy.</a> J. Neurosci. 30, 126-130 (2010).</li><li>Baughan, T. D., Dickson, A., Osman, E. Y., Lorson, C. 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Nessun conflitto di interessi dichiarati.

La ricerca che utilizza modelli murini di malattie spesso richiede la somministrazione di farmaci o altre sostanze per neonati. In questo video, dimostriamo il passo per passo le procedure che coinvolgono due tipi di strategie di iniezione che possono essere utilizzati per indirizzare il sistema nervoso centrale: 1) iniezione diretta nel sistema nervoso centrale che utilizza intracerebroventricolare (ICV) di iniezione, oppure 2) intese a promuovere l&#39;iniezione IV / temporali vena facciale. I tempi di queste iniezion...

<table border="1"><tbody><tr><th> Nome del reagente </th><th> Azienda </th><th> Numero di catalogo </th><th> Commenti (opzionale) </th></tr><tr><td> Green Food Dye </td><td> McCormick </td><td> n / a </td><td> Devono essere filtrati </td></tr><tr><td> Hamilton siringa di vetro (100 mL) </td><td> Sigma Aldrick </td><td> 20702 </td><td></td></tr><tr><td> LuerMxF filo di nylon Style Bianco </td><td> Parti piccole, Inc. </td><td> VPLF-LC78-1-25 </td><td></td></tr><tr><td> Belle calibro aghi ipodermici </td><td> Bottone automatico </td><td> 7111 </td><td> Dimensioni: 33 (SWG) x ¼ &quot;(6,35 mm) </td></tr><tr><td> Vista Transilluminatore Wee </td><td> Respironics </td><td> 1017920 </td><td></td></tr><tr><td> 2.25X fascia Magnifier </td><td> MagEyes </td><td> Modello n. 5 </td><td> Seleziona ingrandimento per soddisfare le preferenze individuali </td></tr></tbody></table>

delivery of therapeutic agents through intracerebroventricular (icv) and intravenous (iv) injection in mice

Nonostante il ruolo protettivo che la barriera sanguigna del cervello gioca nella protezione del cervello, che limita l&#39;accesso al sistema nervoso centrale (SNC), che il più delle volte si traduce in mancanza di terapie potenziali previsti per le malattie neurodegenerative 1,2. Malattie neurodegenerative come l&#39;atrofia muscolare spinale (SMA), in cui i neuroni motori inferiori sono interessati, possono trarre grandi vantaggi di introdurre gli agenti terapeutici nel sistema nervoso centrale. Lo scopo di questo video è quello di dimostrare due paradigmi diversi di iniezione per fornire materiali terapeutici in topo neonato subito dopo la nascita. Uno di questi metodi è l&#39;iniezione direttamente nel cervello ventricoli laterali (intracerebroventricolare) che si traduce in consegna dei materiali nel sistema nervoso centrale attraverso il fluido cerebrospinale 3,4. Il secondo metodo è una iniezione vena temporale (per via endovenosa) che possono introdurre terapie diverse nel sistema circolatorio, con conseguente consegna sistemiche tra cui il sistema nervoso centrale 5. Trasduzione diffusa del sistema nervoso centrale è realizzabile se un vettore virale appropriato e sierotipo virale viene utilizzato. Visualizzazione e l&#39;utilizzo della vena temporale per l&#39;iniezione è possibile fino al giorno dopo la nascita 6. Tuttavia, se il materiale consegnato è destinato a raggiungere il sistema nervoso centrale, queste iniezioni dovrebbero avvenire mentre la barriera emato-encefalica è più permeabile per il suo status immaturi, preferibilmente prima del giorno post-natale 2. La pienamente sviluppato barriera emato-encefalica limita fortemente l&#39;efficacia della consegna per via endovenosa. Entrambi i sistemi di consegna sono semplici ed efficaci, una volta l&#39;attitudine chirurgica è raggiunto. Essi non richiedono dispositivi chirurgici ampio e può essere eseguita da una sola persona. Tuttavia, queste tecniche non sono senza sfide. Le piccole dimensioni dei post-natale giorno 2 cuccioli e le aree successive bersaglio di piccole dimensioni può rendere le iniezioni di difficile esecuzione e inizialmente difficili da replicare.

Watch this Scientific Journal Video about Delivery of Therapeutic Agents Through Intracerebroventricular ICV and Intravenous IV Injection in Mice at JoVE.com

Delivery of Therapeutic Agents Through Intracerebroventricular ICV and Intravenous IV Injection in Mice

Questo articolo viene illustrato due metodi molto diversi di iniezione: 1) nel cervello (intracerebroventricolare) e 2) sistemica (endovenosa) per introdurre gli agenti terapeutici nel sistema nervoso centrale di topi neonati.

Consegna di agenti terapeutici attraverso intracerebroventricolare (ICV) e per via endovenosa (ev) nei topi

Despite the protective role that blood brain barrier plays in shielding the brain, it limits the access to the central nervous system (CNS) which most ...

delivery-therapeutic-agents-through-intracerebroventricular-icv

Research

JoVE Journal

Medicine

Delivery of Therapeutic Agents Through Intracerebroventricular (ICV) and Intravenous (IV) Injection in Mice

58.7K Views.  University of Missouri. Questo articolo viene illustrato due metodi molto diversi di iniezione: 1) nel cervello (intracerebroventricolare) e 2) sistemica (endovenosa) per introdurre gli agenti terapeutici nel sistema nervoso centrale di topi neonati.

Video: Delivery of Therapeutic Agents Through Intracerebroventricular ICV and Intravenous IV Injection in Mice

Orthotopic Xenografting of Human Luciferase-Tagged Malignant Peripheral Nerve Sheath Tumor Cells for in vivo Testing of Candidate Therapeutic Agents

Although in vitro screens are essential for the initial identification of candidate therapeutic agents, a rigorous assessment of the drug's ability to inhibit tumor growth must be performed in a suitable animal model. The type of animal model that is best for this purpose is a topic of intense discussion. Some evidence indicates that preclinical trials examining drug effects on tumors arising in transgenic mice are more predictive of clinical outcome1and so candidate therapeutic agents are often tested in these models. Unfortunately, transgenic models are not available for many tumor types. Further, transgenic models often have other limitations such as concerns as to how well the mouse tumor models its human counterpart, incomplete penetrance of the tumor phenotype and an inability to predict when tumors will develop.
Consequently, many investigators use xenograft models (human tumor cells grafted into immunodeficient mice) for preclinical trials if appropriate transgenic tumor models are not available. Even if transgenic models are available, they are often partnered with xenograft models as the latter facilitate rapid determination of therapeutic ranges. Further, this partnership allows a comparison of the effectiveness of the agent in transgenic tumors and genuine human tumor cells. Historically, xenografting has often been performed by injecting tumor cells subcutaneously (ectopic xenografts). This technique is rapid and reproducible, relatively inexpensive and allows continuous quantitation of tumor growth during the therapeutic period2. However, the subcutaneous space is not the normal microenvironment for most neoplasms and so results obtained with ectopic xenografting can be misleading due to factors such as an absence of organ-specific expression of host tissue and tumor genes. It has thus been strongly recommended that ectopic grafting studies be replaced or complemented by studies in which human tumor cells are grafted into their tissue of origin (orthotopic xenografting)2. Unfortunately, implementation of this recommendation is often thwarted by the fact that orthotopic xenografting methodologies have not yet been developed for many tumor types.
Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive sarcomas that occur sporadically or in association with neurofibromatosis type 13and most commonly arise in the sciatic nerve4. Here we describe a technically straightforward method in which firefly luciferase-tagged human MPNST cells are orthopically xenografted into the sciatic nerve of immunodeficient mice. Our approach to assessing the success of the grafting procedure in individual animals and subsequent non-biased randomization into study groups is also discussed.

Orthotopic Xenografting of Human Luciferase-Tagged Malignant Peripheral Nerve Sheath Tumor Cells for in vivo Testing of Candidate Therapeutic Agents

A method for reliably grafting luciferase-tagged human malignant peripheral nerve sheath tumor cells into the sciatic nerve of immunodeficient mice is described. The use of bioluminescence imaging to demonstrate proper establishment of tumor grafts and criteria for random segregation of animals into study groups are also discussed.

Xenotrapianto ortotopico di Human-luciferasi con tag cellule maligne del tumore della guaina dei nervi periferici per In vivo Test di agenti terapeutici candidati

Although in vitro screens are essential for the initial identification of candidate therapeutic agents, a rigorous assessment of the drug's ability to ...

Ricerca

Medicina

Slow-release Drug Delivery through Elvax 40W to the Rat Retina: Implications for the Treatment of Chronic Conditions

Diseases of the retina are difficult to treat as the retina lies deep within the eye. Invasive methods of drug delivery are often needed to treat these diseases. Chronic retinal diseases such as retinal oedema or neovascularization usually require multiple intraocular injections to effectively treat the condition. However, the risks associated with these injections increase with repeated delivery of the drug. Therefore, alternative delivery methods need to be established in order to minimize the risks of reinjection. Several other investigations have developed methods to deliver drugs over extended time, through materials capable of releasing chemicals slowly into the eye. In this investigation, we outline the use of Elvax 40W, a copolymer resin, to act as a vehicle for drug delivery to the adult rat retina. The resin is made and loaded with the drug. The drug-resin complex is then implanted into the vitreous cavity, where it will slowly release the drug over time. This method was tested using 2-amino-4-phosphonobutyrate (APB), a glutamate analogue that blocks the light response of the retina. It was demonstrated that the APB was slowly released from the resin, and was able to block the retinal response by 7 days after implantation. This indicates that slow-release drug delivery using this copolymer resin is effective for treating the retina, and could be used therapeutically with further testing.

This paper details how Elvax 40W can be used as a slow-release method for drug delivery to the adult rat retina. The protocol for preparing, loading, and delivering the drug-resin complex to the eye is described.

Lento rilascio Drug Delivery attraverso Elvax 40W al Rat Retina: Implicazioni per il trattamento di condizioni croniche

Diseases of the retina are difficult to treat as the retina lies deep within the eye. Invasive methods of drug delivery are often needed to treat these ...

Ultrasound-guided Transthoracic Intramyocardial Injection in Mice

Murine models of cardiovascular disease are important for investigating pathophysiological mechanisms and exploring potential regenerative therapies. Experiments involving myocardial injection are currently performed by direct surgical access through a thoracotomy. While convenient when performed at the time of another experimental manipulation such as coronary artery ligation, the need for an invasive procedure for intramyocardial delivery limits potential experimental designs. With ever improving ultrasound resolution and advanced noninvasive imaging modalities, it is now feasible to routinely perform ultrasound-guided, percutaneous intramyocardial injection. This modality efficiently and reliably delivers agents to a targeted region of myocardium. Advantages of this technique include the avoidance of surgical morbidity, the facility to target regions of myocardium selectively under ultrasound guidance, and the opportunity to deliver injectate to the myocardium at multiple, predetermined time intervals. With practiced technique, complications from intramyocardial injection are rare, and mice quickly return to normal activity on recovery from anesthetic. Following the steps outlined in this protocol, the operator with basic echocardiography experience can quickly become competent in this versatile, minimally invasive technique.

Echocardiography-guided percutaneous intramyocardial injection represents an efficient, reliable, and targetable modality for the delivery of gene transfer agents or cells into the murine heart. Following the steps outlined in this protocol, the operator can quickly become competent in this versatile, minimally invasive technique.

Ultrasound-guided transtoracica intramiocardico iniezione nei topi

Murine models of cardiovascular disease are important for investigating pathophysiological mechanisms and exploring potential regenerative therapies. ...

Testing the Efficacy of Pharmacological Agents in a Pericardial Target Delivery Model in the Swine

To date, many pharmacological agents used to treat or prevent arrhythmias in open-heart cases create undesired systemic side effects. For example, antiarrhythmic drugs administered intravenously can produce drops in systemic pressure in the already compromised cardiac patient. While performing open-heart procedures, surgeons will often either create a small port or form a pericardial cradle to create suitable fields for operation. This access yields opportunities for target pharmacological delivery (antiarrhythmic or ischemic preconditioning agents) directly to the myocardial tissue without undesired side effects.
We have developed a swine model for testing pharmacological agents for target delivery within the pericardial fluid. While fully anesthetized, each animal was instrumented with a Swan-Ganz catheter as well as left and right ventricle pressure catheters, and pacing leads were placed in the right atrial appendage and the right ventricle. A medial sternotomy was then performed and a pericardial access cradle was created; a plunge pacing lead was placed in the left atrial appendage and a bipolar pacing lead was placed in the left ventricle. Utilizing a programmer and a cardiac mapping system, the refractory period of the atrioventricular node (AVN), atria and ventricles was determined. In addition, atrial fibrillation (AF) induction was produced utilizing a Grass stimulator and time in AF was observed. These measurements were performed prior to treatment, as well as 30 min&#160;and 60 min&#160;after pericardial treatment. Additional time points were added for selected studies. The heart was then cardiopleged and reanimated in a four chamber working mode. Pressure measurements and function were recorded for 1 hr after reanimation. This treatment strategy model allowed us to observe the effects of pharmacological agents that may decrease the incidence of cardiac arrhythmias and/or ischemic damage, during and after open-heart surgery.

We have developed a swine model for the target delivery of pharmacological agents within the pericardial space/fluid. Using this approach, the relative benefits of administered agents on induced atrial fibrillation, relative refractory periods and/or ischemic protection can be investigated.

Testare l&#39;efficacia di agenti farmacologici in un pericardica target Delivery Model nel suina

To date, many pharmacological agents used to treat or prevent arrhythmias in open-heart cases create undesired systemic side effects. For example, ...

Disposable Dosators for Pulmonary Insufflation of Therapeutic Agents to Small Animals

Development of new therapeutic products requires efficacy testing in an animal model. The pulmonary route of administration can be utilized to deliver drugs locally and systemically. Evaluation of dry powder aerosols necessitates an efficient dispersion mechanism to maintain high concentrations in an exposure chamber or for direct endotracheal administration. While solutions exist to expose animals by passive inhalation to dry powder aerosols, most require masses of powder in large excess of the dose delivered. This precludes conducting early feasibility studies as insufficient drug is available at the research or early development stage to support the dose delivery requirements for conventional aerosol delivery systems. When designing an aerosol drug product, aerodynamic performance can relate directly to delivery efficiency and efficacy. Dispersion of powder into an aerosol requires energy input sufficient to overcome interparticulate forces, and particle engineering approaches can substantially improve aerosol performance. We have developed a dispersion system (dosator) which can aerosolize engineered dry powder aerosols efficiently for the purpose of direct pulmonary insufflation, dispersion into an exposure system or generation for analytical purposes.

During development of drugs for pulmonary delivery, it is necessary to evaluate pharmacokinetics and efficacy in an animal model. We present a method to build a disposable aerosol dispersion system from of-the-shelf components that can be used to administer intrapulmonary dry powder aerosol to rodents.

Monouso Dosators per insufflazione polmonare di agenti terapeutici ai piccoli animali

Development of new therapeutic products requires efficacy testing in an animal model. The pulmonary route of administration can be utilized to deliver ...

Intranasal Delivery of Therapeutic Stem Cells to Glioblastoma in a Mouse Model

The intrinsic tropism towards brain malignancies renders stem cells as promising carriers of therapeutic agents against malignant tumors. The delivery of therapeutic stem cells via the intranasal route is a recently discovered alternative strategy, with strong potential for clinical translation, due to its non-invasive nature compared to intracranial implantation or delivery via systemic routes. The lack of blood brain barrier further strengthens the therapeutic potential of stem cells undergoing intranasal brain entry. This article summarizes the essential techniques utilized in our studies and outlines the basic principles of intranasal strategy for stem cell delivery using a mouse model of intracranial glioma xenografts. We demonstrate the optimized procedures that generate consistent and reproducible results with specific predetermined experimental parameters and offer guidelines for streamlined work flow that ensure efficient execution and reliable experimental outcome. The article is designed to serve as a baseline for further experimental customization based on hypothesis, stem cell types, or tumor specifics.

Stem cells are promising therapeutic carriers to treat brain tumors due to their intrinsic tumor tropism. Non-invasive intranasal stem cell delivery bypasses the blood brain barrier and demonstrates strong potential for clinical translation. This article summarizes the basic principles of intranasal stem cell delivery in a mouse model of glioma.

Intranasale consegna delle cellule staminali terapeutiche a Glioblastoma in un modello di mouse

The intrinsic tropism towards brain malignancies renders stem cells as promising carriers of therapeutic agents against malignant tumors. The delivery of ...

A Rat Carotid Artery Pressure-Controlled Segmental Balloon Injury with Periadventitial Therapeutic Application

Cardiovascular disease remains the leading cause of death and disability worldwide, in part due to atherosclerosis. Atherosclerotic plaque narrows the luminal surface area in arteries thereby reducing adequate blood flow to organs and distal tissues. Clinically, revascularization procedures such as balloon angioplasty with or without stent placement aim to restore blood flow. Although these procedures reestablish blood flow by reducing plaque burden, they damage the vessel wall, which initiates the arterial healing response. The prolonged healing response causes arterial restenosis, or re-narrowing, ultimately limiting the long-term success of these revascularization procedures. Therefore, preclinical animal models are integral for analyzing the pathophysiological mechanisms driving restenosis, and provide the opportunity to test novel therapeutic strategies. Murine models are cheaper and easier to operate on than large animal models. Balloon or wire injury are the two commonly accepted injury modalities used in murine models. Balloon injury models in particular mimic the clinical angioplasty procedure and cause adequate damage to the artery for the development of restenosis. Herein we describe the surgical details for performing and histologically analyzing the modified, pressure-controlled rat carotid artery balloon injury model. Additionally, this protocol highlights how local periadventitial application of therapeutics can be used to inhibit neointimal hyperplasia. Lastly, we present light sheet fluorescence microscopy as a novel approach for imaging and visualizing the arterial injury in three-dimensions.

The rat carotid artery balloon injury mimics the clinical angioplasty procedure performed to restore blood flow in atherosclerotic vessels. This model induces the arterial injury response by distending the arterial wall, and denuding the intimal layer of endothelial cells, ultimately causing remodeling and an intimal hyperplastic response.

Una lesione segmentale del palloncino segmentale controllato dalla pressione carotide del ratto con applicazione terapeutica periadventitiale

Cardiovascular disease remains the leading cause of death and disability worldwide, in part due to atherosclerosis. Atherosclerotic plaque narrows the ...

Intratracheal Administration of Dry Powder Formulation in Mice

In the development of inhalable dry powder formulations, it is essential to evaluate their biological activities in preclinical animal models. This paper introduces a noninvasive method of intratracheal delivery of dry powder formulation in mice. A dry powder loading device that consists of a 200 &#181;L gel loading pipette tip connected to an 1 mL syringe via a three-way stopcock is presented. A small amount of dry powder (1-2 mg) is loaded into the pipette tip and dispersed by 0.6 mL of air in the syringe. Because pipette tips are disposable and inexpensive, different dry powder formulations can be loaded into different tips in advance. Various formulations can be evaluated in the same animal experiment without device cleaning and dose refilling, thereby saving time and eliminating the risk of cross-contamination from residual powder. The extent of powder dispersion can be inspected by the amount of powder remaining in the pipette tip. A protocol of intubation in mouse with a custom-made light source and a guiding cannula is included. Proper intubation is one of the key factors that influences the intratracheal delivery of dry powder formulation to the deep lung region of the mouse.

Dry powder formulations for inhalation have great potential in treating respiratory diseases. Before entering human studies, it is necessary to evaluate the efficacy of the dry powder formulation in preclinical studies. A simple and noninvasive method of the administration of dry powder in mice through the intratracheal route is presented.

Somministrazione intratracheale della formulazione di polvere secca nei topi

In the development of inhalable dry powder formulations, it is essential to evaluate their biological activities in preclinical animal models. This paper ...

Decotto Liujunzi: un trattamento efficace per la prevenzione dell'UC-CRC

Establishment of Coloproctitis Cancer Model in Mice and Evaluation of Therapeutic Effect of Chinese Medicine

Colorectal cancer (CRC) is a common malignancy of the digestive system and has become the third most common malignancy worldwide and the second leading cause of malignancy-related death. Ulcerative coloproctitis (UC) is a precancerous lesion, and UC-associated CRC (UC-CRC) is the most common subtype of CRC. Therefore, a reasonable UC-CRC model is the cornerstone and guarantee of new drug development. Traditional Chinese medicine (TCM) has been widely used in the treatment of UC-CRC due to its good efficacy. As a classic tonic prescription of TCM, Liujunzi decoction (LJZD) has been widely used in the treatment of UC-CRC. In this study, a UC-CRC model was established by combining azomethane and dextran sulfate sodium, and the LJZD was administered. The data confirmed that LJZD can effectively inhibit cancer transition in UC-CRC by using mouse body weight, colorectal length, pathological and inflammatory factors, colorectal barrier function, and cancer markers. This protocol provides a system for evaluating the efficacy of TCM in the prevention and treatment of UC-CRC.

Autore in primo piano: Il decotto di Liujunzi come trattamento tradizionale cinese per il cancro della coloproctite

This protocol provides a mouse model of ulcerative coloproctitis-associated colorectal cancer induced by azomethane combined with dextran sulfate sodium. The model was used to evaluate the efficacy of traditional Chinese medicine compounds in the prevention and treatment of colorectal cancer.

Creazione di un modello di cancro della coloproctite nei topi e valutazione dell'effetto terapeutico della medicina cinese

Colorectal cancer (CRC) is a common malignancy of the digestive system and has become the third most common malignancy worldwide and the second leading ...