Name: isolated hepatic perfusion as a treatment for liver metastases of uveal melanoma
Uploaded: 2015-01-25T14:00:00.000+00:00
Duration: 9 min 52 s
Description: Watch this Scientific Journal Video about Isolated Hepatic Perfusion as a Treatment for Liver Metastases of Uveal Melanoma at JoVE.com

Lennart Wiman (photographer) is acknowledged for excellent video recording.

Los pacientes por escrito el consentimiento informado y el estudio fue aprobado por el Consejo Regional de Revisión Ética de la Universidad de Gotemburgo. 1. Anestesia <ol><li> Configure todo el seguimiento (ECG, manguito de presión arterial) 17. </li><li> Inserte un catéter epidural en la parte baja a mediados nivel torácico 17. </li><li> Inducir la anestesia con propofol 2 mg / kg, 150 g de fentanilo y rocuronio 0,6 mg / kg a través de un catéter venoso periférico insertado previamente. </li><li> Preoxigenar y intubar al paciente. Mantener la anestesia con sevoflurano, oxígeno y aire 17. </li><li> Inserte un catéter venoso central de doble luz y un catéter arterial. Monitorear CVP y arterial presión 17. </li></ol> 2. Aislamiento del Hígado y la canulación de los buques <ol><li> Administrar 100 IE / kg de heparina y dar profilaxis antibiótica (por ejemplo, 160 mg de trimetoprima y 800 mg de sulfametoxazol). </li><li> Utilizando la técnica de Seldinger percutánea, coloque un catéter 17 Fr en la vena femoral y un catéter 15 Fr en la vena yugular externa 18. Verifique la posición de los catéteres por fluoroscopia. </li><li> Realice una laparotomía por una incisión en forma de L de la apófisis xifoides hasta la línea media y luego se extendió a una incisión subcostal derecha 19. </li><li> Verifique que no existan metástasis extrahepáticas en el abdomen por una inspección completa y la palpación. <ol><li> Si se encuentra alguna lesión sospechosa, verificar malignidad en la sección congelada. </li><li> Terminar el procedimiento si hay evidencia de manifestación extrahepática (adenopatía periaortal, metástasis abdominales). Adenopatías en la porta del hígado no se considera una contraindicación. </li></ol></li><li> Movilizar el hígado dividiendo el ligamento falciforme, el ligamento triangular izquierdo y derecho y el ligamento coronario 19. </li><li> Diseccionar la vena cavavenosa a partir de las venas renales hasta el diafragma con la ligadura de venas tributarias retroperitoneales. </li><li> Identificar la vena suprarrenal derecha y colocar un asa vascular alrededor. </li><li> Identificar la vena gonadal derecha. Diseccionarlo para 5-10 cm. Coloque una sutura en bolsa de tabaco alrededor de la entrada de la vena cava. <ol><li> Coloque una ligadura distal de la vena gonadal. Con la tijera, abrir la vena gonadal 3-5 cm de la vena cava. </li><li> Inserte un 18 P. catéter reforzado con alambre a través de la vena gonadal derecha y coloque la punta retrohepatically encima de las venas renales. </li><li> Colocar una ligadura alrededor de la vena gonadal para asegurar la posición del catéter. </li><li> Verificar el retorno de la sangre a través del catéter. Heparinizar el catéter 18. </li></ol></li><li> Diseccionar el ligamento hepatoduodenal y visualizar la arteria hepática común, la arteria gastroduodenal y la arteria hepática propia. <ol><li> Lugar buque bucles alrededor del arte hepático comúnRY y la arteria gastrodoudenal. </li><li> Diseccionar la vena porta y colocar un asa vascular alrededor. </li><li> Aislar el resto del ligamento que contiene el conducto biliar común y incluir todo esto en un bucle buque. </li></ol></li><li> Levante la arteria gastroduodenal con el bucle buque. Después de interrumpir el flujo arterial, hacer una incisión con tijeras. <ol><li> Inserte un 8-12 Fr catéter con la punta dentro de la arteria hepática propia. Fije la posición con una ligadura alrededor de la arteria gastroduodenal. </li><li> Verificar retorno de la sangre en el catéter y luego se heparinizar 18. </li></ol></li><li> Primer sistema de perfusión con una unidad de hematíes, 100 ml de albúmina (50 g / L), 100 ml de Tribonat (bicarbonato de sodio), 2500 unidades de heparina (5000 IE / ml) y 250 ml de cristaloides. </li><li> Conectar el arterial y los catéteres de vena cava en el sistema de perfusión que consisten de una máquina corazón-pulmón equipado con una bomba de rodillos y un calentador tunto con venosas sistemas de depósito, oxigenadores y tubos de tipo desechable. </li><li> Conectar los catéteres yugulares femorales y externos a la bomba de derivación venovenosa que consta de una bomba centrífuga, junto con una unidad de calefacción y empezar bypass. </li><li> Completar el aislamiento del hígado apretando un torniquete de Rumel alrededor de la vena cava infrahepatically encima de las venas renales y la sujeción de la vena cava suprahepatically mediante una abrazadera vascular Klintmalm. <ol><li> Apriete un torniquete de Rumel alrededor del conducto biliar común, la vena porta y la vena suprarrenal derecha y finalmente fijar la arteria hepática usando una pinza vascular. </li></ol></li></ol> 3. La perfusión <ol><li> Iniciar la perfusión y controlar manualmente el nivel de sangre en el depósito venoso. Ajustar la velocidad de flujo hasta que se alcanza un nivel de estado estacionario, por lo general una velocidad de flujo entre 500-1,200 ml / min. </li><li> Coloque las sondas de termistor en los lóbulos derecho e izquierdo del hígado y uno directamente enel catéter arterial. </li><li> Calentar el líquido de perfusión destinado a una temperatura de hígado objetivo de 40 o C. Mantener esta temperatura durante toda la perfusión. </li><li> Colocar una sonda de centelleo sobre la cabeza de la bomba de bypass veno-venosa. </li><li> Calibrar el sistema de monitoreo de fugas, mediante la inyección de una pequeña dosis de 10 MBq 99m Tc-albúmina (1 ml) a la circulación sistémica y registrar la actividad de línea de base. <ol><li> Inyectar una dosis diez veces superior, 100 MBq 99m Tc-albúmina (1 ml), en la unidad de perfusión hepática. </li><li> Registrar cualquier fuga del sistema de perfusión a la circulación sistémica como un aumento en la actividad medida por la sonda de centelleo. </li><li> Registre cualquier fuga de la circulación sistémica al sistema de perfusión como un aumento en el volumen en el depósito venoso. </li></ol></li><li> Cuando se establece en estado estacionario de la velocidad de flujo, no hay fugas está presente y la temperatura de hígado ha alcanzado 40 ° C, se inyectan melfalán en una dosis de1 mg / kg de peso corporal en el sistema de perfusión dividido en dos dosis administradas 30 min aparte. </li></ol> 4. Poner fin a la perfusión y quitar catéteres <ol><li> Después de 60 minutos de perfusión, vaciar el líquido de perfusión del sistema y regar el hígado con 1000 ml de cristaloides. </li><li> Añadir una unidad de hematíes a la unidad de la perfusión, infundir en el hígado y luego detener la perfusión. </li><li> Suelte la abrazadera Klintmalm y el torniquete de Rumel de la vena cava. </li><li> Suelte el torniquete de Rumel del ligamento hepatoduodenal. </li><li> Retire la ligadura de la arteria gastroduodenal. Retire el catéter y ligar el vaso. </li><li> Soltar la abrazadera de la arteria hepática. </li><li> Retirar el catéter de la vena gonadal y ligar el vaso. </li><li> Parar la bomba de bypass veno-venoso y desconectar los tubos. </li><li> Cierre el abdomen con una sutura fascial ininterrumpida y luego una sutura intradérmica piel. </li><li> Quite la catheters en las venas femoral y yugular externas y aplican presión local durante 5 min. </li></ol> 5. Cuidado posoperatorio <ol><li> Retirar la intubación del paciente y transferir a la unidad de cuidados postoperatorios durante 24 horas. Mantener el seguimiento de la presión venosa central y la presión sanguínea arterial. </li><li> Traslado al paciente a un departamento de cirugía para un típico de 3-7 días de cuidados postoperatorios. </li><li> Continuar con heparina de bajo peso molecular (HBPM) a una dosis de 5000 IE / día para un total de 4 semanas. </li></ol>

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The authors have nothing to disclose.

Los pacientes con melanoma uveal metastásico tienen pocas opciones terapéuticas. Quimioterapia 6, inmunoterapia 23 y terapia dirigida 24 no han sido capaces de demostrar una mejoría de SG. Para los pacientes con pocas metástasis hepáticas, la resección hepática es una opción 12. Otros tratamientos incluyen quimioterapia hepática intraarterial (EIS) 9, la quimioembolización transarterial (TACE) 25 y la terapia selectiva interna radiación (SIRT) ...

El melanoma uveal es el tumor maligno intraocular primario más común en adultos, con una incidencia más alta de ser en poblaciones caucásicas 1. El tratamiento local consiste en la enucleación o placa local de radioterapia 2,3. Independientemente de tratamiento local, la enfermedad metastásica, en última instancia desarrollar en aproximadamente 35-50% de los pacientes 4,5. El hígado es el sitio más común de metástasis y aproximadamente el 50% de los pacientes presentan metástasis hepáticas aisladas. La mediana de supervivencia global (SG) para los pacientes con metástasis hepáticas es de 6-12 meses 4. Varios diferentes estrategias sistémicas y regionales de tratamiento se han explorado, pero las tasas de supervivencia no han mejorado 2. El tratamiento con quimioterapia sistémica, tanto sola sustancia (dacarbazina o temozolomida 6) o la combinación de tratamientos (régimen BOLD 7, gemcitabina con treosulfano 8), muestra tasas de respuesta de menos de10%. Quimioterapia intraarterial hepática (EIS) ha mostrado una tasa ligeramente superior de respuesta global en comparación con la quimioterapia sistémica, pero no un incremento en la supervivencia general 9. La quimioembolización transarterial (TACE) con cisplatino y carboplatino mostraron una respuesta parcial en el 57% de los pacientes con una supervivencia media de 9 meses 10. Radioterapia interna selectiva (SIRT) ha mostrado una respuesta parcial en el 62% de los pacientes con una supervivencia media de 7 meses 11. En un estudio sobre la resección hepática, incluyendo 255 pacientes, se informó de una supervivencia global de 14 meses; en 76 de los pacientes en los que era posible una resección microscópicamente completa, la supervivencia fue de 27 meses 12. Perfusión hepática aislada (PHI) es una opción de tratamiento regional, originalmente desarrollado por Ryan y Ausman. El fundamento de la técnica consiste en aislar quirúrgicamente y perfundir el hígado con una alta dosis de un agente quimioterapéutico, evitando sistémica axicidad 13. Además, la hipertermia concurrente media un aumento de la captación del agente quimioterapéutico por los cambios en el flujo sanguíneo del tumor y la permeabilidad de la membrana celular 14. Aislamiento vascular completa se confirma con una técnica trazador radioactivo utilizando Tc-99 etiquetados albúmina sérica humana 15. Una retrospectiva estudio de fase II ha demostrado un potencial beneficio de supervivencia de 14 meses (26 frente a 12 meses) para los pacientes que se sometieron a PHI en comparación con las más longevas con metástasis hepáticas melanoma uveal en Suecia 16. Aquí presentamos la técnica del PHI con una breve revisión de los resultados actuales y perspectivas futuras.

<table><tbody><tr><td>Wire re-inforced catethers</td><td>Medtronic Inc, Minnesota, USA</td><td></td><td>12-18Fr</td></tr><tr><td>St&amp;ouml;ckert S5</td><td>Sorin Group, Mirandola, Italy</td><td></td><td>Heart-lung machine</td></tr><tr><td>Rotaflow</td><td>Maquet, Jostra Medizintechnik AG, Hirrlingen, Germany</td><td></td><td>Centrifugal pump</td></tr><tr><td>HU 35</td><td>Maquet cardiopulmonary AG, Hirrlingen, Germany</td><td></td><td>Heater unit</td></tr><tr><td>Flexbumin 200 g/l</td><td>Baxter Medical AB, Kista, Sweden</td><td></td><td></td></tr><tr><td>Tribonat</td><td>Fresenius Kabi AB, Uppsala, Sweden</td><td></td><td></td></tr><tr><td>Heparin LEO, 5000 IE/ml</td><td>LEO Pharma AB, Sweden</td><td></td><td></td></tr><tr><td>Ringer Acetate</td><td>Baxter Medical AB, Kista, Sweden</td><td></td><td></td></tr><tr><td>Vasculosis</td><td>Cis-Bio International, Gif-Sur-Yvette, France</td><td></td><td>99mTc-albumin</td></tr><tr><td>MedicView</td><td>SystemData AB, Gothenburg, Sweden</td><td></td><td>Leakage monitoring system</td></tr><tr><td>Alkeran</td><td>Aspen Europe GmbH, Bad Oldesloe, Germany</td><td></td><td>melphalan</td></tr></tbody></table>

isolated hepatic perfusion as a treatment for liver metastases of uveal melanoma

La perfusión hepática aislada (PHI) es un procedimiento donde el hígado es aislado y perfundido con una alta concentración de la melfalán agente quimioterapéutico quirúrgicamente. Brevemente, el procedimiento se inicia con la configuración de una veno-venosa de bypass percutáneo de la vena femoral hasta la vena yugular externa. A través de una laparotomía, se insertan catéteres en la arteria hepática adecuada y la vena cava. La vena porta y la vena cava, tanto supra y infrahepatically, se fijan entonces. El catéteres arteriales y venosos están conectados a una máquina de corazón-pulmón y el hígado se perfunde con melfalán (1 mg / kg de peso corporal) durante 60 min. De esta manera es posible localmente perfundir el hígado con una alta dosis de un agente quimioterapéutico, sin fugas a la circulación sistémica. En estudios anteriores, que incluyeron pacientes con metástasis hepáticas aisladas de melanoma uveal, una tasa de respuesta global del 33-100% y una mediana de supervivencia entre 9 y 13 meses,han sido reportados. El objetivo de este protocolo es para dar una descripción clara de cómo llevar a cabo el procedimiento y debatir PHI como una opción de tratamiento para las metástasis hepáticas de melanoma uveal.

Un informe inicial incluyó 22 pacientes tratados con melfalán con o sin TNF-alfa. Los resultados mostraron una tasa de respuesta global del 62%, incluyendo dos pacientes con CR (10%), y una mediana de supervivencia global de 11 meses 20. Un estudio posterior informó sobre los resultados de 29 pacientes consecutivos tratados usando melfalán solo. El resultado fue muy similar con una respuesta global 62% incluyendo 10% de respuesta completa y una supervivencia media de 12 meses 21. <p class="...

Watch this Scientific Journal Video about Isolated Hepatic Perfusion as a Treatment for Liver Metastases of Uveal Melanoma at JoVE.com

Isolated Hepatic Perfusion as a Treatment for Liver Metastases of Uveal Melanoma

Here, we present a protocol how to perform an isolated liver perfusion (IHP) with melphalan and we also discuss IHP as a treatment option for liver metastases of uveal melanoma.

Aislado hepática de perfusión como un tratamiento para las metástasis hepáticas de melanoma uveal

Isolated hepatic perfusion (IHP) is a procedure where the liver is surgically isolated and perfused with a high concentration of the chemotherapeutic ...

isolated-hepatic-perfusion-as-treatment-for-liver-metastases-uveal

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

13.3K vistas.  Institute of Clinical Sciences. Here, we present a protocol how to perform an isolated liver perfusion (IHP) with melphalan and we also discuss IHP as a treatment option for liver metastases of uveal melanoma.

Video: Isolated Hepatic Perfusion as a Treatment for Liver Metastases of Uveal Melanoma

Murine Model of Metastatic Liver Tumors in the Setting of Ischemia Reperfusion Injury

Liver ischemia and reperfusion (I/R) injury, a common clinical challenge, remains an inevitable pathophysiological process that has been shown to induce multiple tissue and organ damage. Despite recent advances and therapeutic approaches, the overall morbidity has remained unsatisfactory especially in patients with underlying parenchymal abnormalities. In the context of aggressive cancer growth and metastasis, surgical I/R is suspected to be the promoter regulating tumor recurrence. This article aims to describe a clinically relevant murine model of liver I/R and colorectal liver metastasis. In doing so, we aim to assist other investigators in establishing and perfecting this model for their routine research practice to better understand the effects of liver I/R on promoting liver metastases.

We describe in detail a clinically relevant colorectal cancer liver metastases (CRLM) tumor model and the influence of liver ischemia reperfusion (I/R) in tumor growth and metastasis. This model can help to better understand the mechanisms underlying surgery-induced promotion of liver metastatic growth.

Modelo murino de tumores hepáticos metastásicos en el escenario de la lesión por reperfusión de isquemia

Liver ischemia and reperfusion (I/R) injury, a common clinical challenge, remains an inevitable pathophysiological process that has been shown to induce ...

Investigación

Investigación sobre el cáncer

Generation of a Liver Orthotopic Human Uveal Melanoma Xenograft Platform in Immunodeficient Mice

In recent decades, subcutaneously implanted patient-derived xenograft tumors or cultured human cell lines have been increasingly recognized as more representative models to study human cancers in immunodeficient mice than traditional established human cell lines in vitro. Recently, orthotopically implanted patient-derived tumor xenograft (PDX) models in mice have been developed to better replicate features of patient tumors. A liver orthotopic xenograft mouse model is expected to be a useful cancer research platform, providing insights into tumor biology and drug therapy. However, liver orthotopic tumor implantation is generally complicated. Here we describe our protocols for the orthotopic implantation of patient-derived liver-metastatic uveal melanoma tumors. We cultured human liver metastatic uveal melanoma cell lines into immunodeficient mice. The protocols can result in consistently high technical success rates using either a surgical orthotopic implantation technique with chunks of patient-derived uveal melanoma tumor or a needle injection technique with cultured human cell line. We also describe protocols for CT scanning to detect interior liver tumors and for re-implantation techniques using cryopreserved tumors to achieve re-engraftment. Together, these protocols provide a better platform for liver orthotopic tumor mouse models of liver metastatic uveal melanoma in translational research.

Orthotopic human liver metastatic uveal melanoma xenograft mouse models were created using surgical orthotopic implantation techniques with patient-derived tumor chunk and needle injection techniques with cultured human uveal melanoma cell lines.

Generación de una plataforma de xenoinjerto de melanoma uveal humano ortotópico hepático en ratones inmunodeficientes

In recent decades, subcutaneously implanted patient-derived xenograft tumors or cultured human cell lines have been increasingly recognized as more ...

Portal Vein Injection of Colorectal Cancer Organoids to Study the Liver Metastasis Stroma

Hepatic metastasis of colorectal cancer (CRC) is a leading cause of cancer-related death. Cancer-associated fibroblasts (CAFs), a major component of the tumor microenvironment, play a crucial role in metastatic CRC progression and predict poor patient prognosis. However, there is a lack of satisfactory mouse models to study the crosstalk between metastatic cancer cells and CAFs. Here, we present a method to investigate how liver metastasis progression is regulated by the metastatic niche and possibly could be restrained by stroma-directed therapy. Portal vein injection of CRC organoids generated a desmoplastic reaction, which faithfully recapitulated the fibroblast-rich histology of human CRC liver metastases. This model was tissue-specific with a higher tumor burden in the liver when compared to an intra-splenic injection model, simplifying mouse survival analyses. By injecting luciferase-expressing tumor organoids, tumor growth kinetics could be monitored by in vivo imaging. Moreover, this preclinical model provides a useful platform to assess the efficacy of therapeutics targeting the tumor mesenchyme. We describe methods to examine whether adeno-associated virus-mediated delivery of a tumor-inhibiting stromal gene to hepatocytes could remodel the tumor microenvironment and improve mouse survival. This approach enables the development and assessment of novel therapeutic strategies to inhibit hepatic metastasis of CRC.

Portal vein injection of colorectal cancer (CRC) organoids generates stroma-rich liver metastasis. This mouse model of CRC hepatic metastasis represents a useful tool to study tumor-stroma interactions and develop novel stroma-directed therapeutics such as adeno-associated virus-mediated gene therapies.

Inyección en la vena porta de organoides de cáncer colorrectal para estudiar la metástasis hepática del estroma

Hepatic metastasis of colorectal cancer (CRC) is a leading cause of cancer-related death. Cancer-associated fibroblasts (CAFs), a major component of the ...

Technique of Subnormothermic Ex Vivo Liver Perfusion for the Storage, Assessment, and Repair of Marginal Liver Grafts

The success of liver transplantation has resulted in a dramatic organ shortage. In most transplant regions 20-30% of patients on the waiting list for liver transplantation die without receiving an organ transplant or are delisted for disease progression. One strategy to increase the donor pool is the utilization of marginal grafts, such as fatty livers, grafts from older donors, or donation after cardiac death (DCD). The current preservation technique of cold static storage is only poorly tolerated by marginal livers resulting in significant organ damage. In addition, cold static organ storage does not allow graft assessment or repair prior to transplantation.
These shortcomings of cold static preservation have triggered an interest in warm perfused organ preservation to reduce cold ischemic injury, assess liver grafts during preservation, and explore the opportunity to repair marginal livers prior to transplantation. The optimal pressure and flow conditions, perfusion temperature, composition of the perfusion solution and the need for an oxygen carrier has been controversial in the past.
In spite of promising results in several animal studies, the complexity and the costs have prevented a broader clinical application so far. Recently, with enhanced technology and a better understanding of liver physiology during ex vivo perfusion the outcome of warm liver perfusion has improved and consistently good results can be achieved.
This paper will provide information about liver retrieval, storage techniques, and isolated liver perfusion in pigs. We will illustrate a) the requirements to ensure sufficient oxygen supply to the organ, b) technical considerations about the perfusion machine and the perfusion solution, and c) biochemical aspects of isolated organs.

Technique of Subnormothermic Ex Vivo Liver Perfusion for the Storage, Assessment, and Repair of Marginal Liver Grafts

Marginal grafts, such as fatty livers, grafts from older donors, or livers retrieved after cardiac death (DCD) tolerate conventional, cold static storage only poorly. We developed a novel model of subnormothermic ex vivo liver perfusion for preservation, assessment, and repair of marginal liver grafts prior to transplantation.

Técnica de Subnormothermic<em&gt; Ex Vivo</em&gt; Perfusión hepática para el Almacenamiento, Evaluación y Reparación de marginales injertos hepáticos

The success of liver transplantation has resulted in a dramatic organ shortage. In most transplant regions 20-30% of patients on the waiting list for ...

Medicina

Implantation and Evaluation of Melanoma in the Murine Choroid via Optical Coherence Tomography

Establishing experimental choroidal melanoma models is challenging in terms of the ability to induce tumors at the correct localization. In addition, difficulties in observing posterior choroidal melanoma in vivo limit tumor location and growth evaluation in real-time. The approach described here optimizes techniques for establishing choroidal melanoma in mice via a multi-step sub-choroidal B16LS9 cell injection procedure. To enable precision in injecting into the small dimensions of the mouse uvea, the complete procedure is performed under a microscope. First, a conjunctival peritomy is formed in the dorsal-temporal area of the eye. Then, a tract into the sub-choroidal space is created by inserting a needle through the exposed sclera. This is followed by the insertion of a blunt needle into the tract and the injection of melanoma cells into the choroid. Immediately after injection, noninvasive optical coherence tomography (OCT) imaging is utilized to determine tumor location and progress. Retinal detachment is evaluated as a predictor of tumor site and size. The presented method enables the reproducible induction of choroid-localized melanoma in mice and the live imaging of tumor growth evaluation. As such, it provides a valuable tool for studying intraocular tumors.

Implantation and Evaluation of Melanoma in the Murine Choroid via Optical Coherence Tomography

The present protocol describes the implantation and evaluation of melanoma in the murine choroid utilizing optical coherence tomography.

Implantación y evaluación del melanoma en la coroides murina mediante tomografía de coherencia óptica

Establishing experimental choroidal melanoma models is challenging in terms of the ability to induce tumors at the correct localization. In addition, ...

Percutaneous Hepatic Perfusion (PHP) with Melphalan as a Treatment for Unresectable Metastases Confined to the Liver

Unresectable liver metastases of colorectal cancer can be treated with systemic chemotherapy, aiming to limit the disease, extend survival or turn unresectable metastases into resectable ones. Some patients however, suffer from side effects or progression under systemic treatment. For patients with metastasized uveal melanoma there are no standard systemic therapy options. For patients without extrahepatic disease, isolated liver perfusion (IHP) may enable local disease control with limited systemic side effects. Previously, this was performed during open surgery with satisfying results, but morbidity and mortality related to the open procedure, prohibited a widespread application. Therefore, percutaneous hepatic perfusion (PHP) with simultaneous chemofiltration was developed. Besides decreasing morbidity and mortality, this procedure can be repeated, hopefully leading to a higher response rate and improved survival (by local control of disease). During PHP, catheters are placed in the proper hepatic artery, to infuse the chemotherapeutic agent, and in the inferior caval vein to aspirate the chemosaturated blood returning through the hepatic veins. The caval vein catheter is a double balloon catheter that prohibits leakage into the systemic circulation. The blood returning from the hepatic veins is aspirated through the catheter fenestrations and then perfused through an extra-corporeal filtration system. After filtration, the blood is returned to the patient by a third catheter in the right internal jugular vein. During PHP a high dose of melphalan is infused into the liver, which is toxic and would lead to life threatening complications when administered systemically. Because of the significant hemodynamic instability resulting from the combination of caval vein occlusion and chemofiltration, hemodynamic monitoring and hemodynamic support is of paramount importance during this complex procedure.

Percutaneous Hepatic Perfusion PHP with Melphalan as a Treatment for Unresectable Metastases Confined to the Liver

In this manuscript, we describe percutaneous isolated hepatic perfusion with simultaneous chemofiltration as treatment for unresectable liver metastases. This procedure is performed under general anaesthesia in the angiosuite by an experienced team, consisting of an interventional radiologist, a clinical perfusionist and anaesthesiologist.

Percutánea hepática perfusión (PHP) con melfalán como un tratamiento para la metástasis no resecables limitado al hígado

Unresectable liver metastases of colorectal cancer can be treated with systemic chemotherapy, aiming to limit the disease, extend survival or turn ...

Visualization of Vascular and Parenchymal Regeneration after 70% Partial Hepatectomy in Normal Mice

A modified silicone injection procedure was used for visualization of the hepatic vascular tree. This procedure consisted of in-vivo injection of the silicone compound, via a 26 G catheter, into the portal or hepatic vein. After silicone injection, organs were explanted and prepared for ex-vivo micro-CT (&#181;CT) scanning. The silicone injection procedure is technically challenging. Achieving a successful outcome requires extensive microsurgical experience from the surgeon. One of the challenges of this procedure involves determining the adequate perfusion rate for the silicone compound. The perfusion rate for the silicone compound needs to be defined based on the hemodynamic of the vascular system of interest. Inappropriate perfusion rate can lead to an incomplete perfusion, artificial dilation and rupturing of vascular trees.
The 3D reconstruction of the vascular system was based on CT scans and was achieved using preclinical software such as HepaVision. The quality of the reconstructed vascular tree was directly related to the quality of silicone perfusion. Subsequently computed vascular parameters indicative of vascular growth, such as total vascular volume, were calculated based on the vascular reconstructions. Contrasting the vascular tree with silicone allowed for subsequent histological work-up of the specimen after &#181;CT scanning. The specimen can be subjected to serial sectioning, histological analysis and whole slide scanning, and thereafter to 3D reconstruction of the vascular trees based on histological images. This is the prerequisite for the detection of molecular events and their distribution with respect to the vascular tree. This modified silicone injection procedure can also be used to visualize and reconstruct the vascular systems of other organs. This technique has the potential to be extensively applied to studies concerning vascular anatomy and growth in various animal and disease models.

Tools used for visualizing vascular regeneration require methods for contrasting the vascular trees. This film demonstrated a delicate injection technique used to achieve optimal contrasting of the vascular trees and illustrate the potential benefits resulting from a detailed analysis of the resulting specimen using &#181;CT and histological serial sections.

La visualización del parénquima vascular y la regeneración después del 70% hepatectomía parcial en ratones normales

A modified silicone injection procedure was used for visualization of the hepatic vascular tree. This procedure consisted of in-vivo injection of the ...

A Small Animal Model of Ex Vivo Normothermic Liver Perfusion

There is a significant shortage of liver allografts available for transplantation, and in response the donor criteria have been expanded. As a result, normothermic ex vivo liver perfusion (NEVLP) has been introduced as a method to evaluate and modify organ function. NEVLP has many advantages in comparison to hypothermic and subnormothermic perfusion including reduced preservation injury, restoration of normal organ function under physiologic conditions, assessment of organ performance, and as a platform for organ repair, remodeling, and modification. Both murine and porcine NEVLP models have been described. We demonstrate a rat model of NEVLP and use this model to show one of its important applications &#8212; the use of a therapeutic molecule added to liver perfusate. Catalase is an endogenous reactive oxygen species (ROS) scavenger and has been demonstrated to decrease ischemia-reperfusion in the eye, brain, and lung. Pegylation has been shown to target catalase to the endothelium. Here, we added pegylated-catalase (PEG-CAT) to the base perfusate and demonstrated its ability to mitigate liver preservation injury. An advantage of our rodent NEVLP model is that it is inexpensive in comparison to larger animal models. A limitation of this study is that it does not currently include post-perfusion liver transplantation. Therefore, prediction of the function of the organ post-transplantation cannot be made with certainty. However, the rat liver transplant model is well established and certainly could be used in conjunction with this model. In conclusion, we have demonstrated an inexpensive, simple, easily replicable NEVLP model using rats. Applications of this model can include testing novel perfusates and perfusate additives, testing software designed for organ evaluation, and experiments designed to repair organs.

A Small Animal Model of Ex Vivo Normothermic Liver Perfusion

There is a significant liver donor shortage, and criteria for liver donors have been expanded. Normothermic ex vivo liver perfusion (NEVLP) has been developed to evaluate and modify organ function. This study demonstrates a rat model of NEVLP and tests the ability of pegylated-catalase, to mitigate liver preservation injury.

Un pequeño modelo de la perfusión hepática normotérmica Ex Vivo

There is a significant shortage of liver allografts available for transplantation, and in response the donor criteria have been expanded. As a result, ...

Isolation of Regenerating Hepatocytes after Partial Hepatectomy in Mice

Partial hepatectomy has been widely used to investigate liver regeneration in mice, but the isolation of high yields of viable hepatocytes for downstream single-cell applications is challenging. A marked accumulation of lipids within regenerating hepatocytes is observed during the first 2 days of normal liver regeneration in mice. This so-called transient regeneration-associated steatosis (TRAS) is temporary but partially overlaps the major proliferative phase. Density-gradient purification is the backbone of most existing protocols for the isolation of primary hepatocytes. As gradient purification relies on the density and size of cells, it separates non-steatotic from steatotic hepatocyte populations. Therefore, fatty hepatocytes often are lost, yielding non-representative hepatocyte fractions. 
The presented protocol describes an easy and reliable method for the in vivo isolation of regenerating hepatocytes regardless of their lipid content. Hepatocytes from male C57BL/6 mice are isolated 24-48 h after hepatectomy by a classic two-step collagenase perfusion approach. A standard peristaltic pump drives the warmed solutions via the catheterized inferior vena cava into the remnant, using a retrograde perfusion technique with outflow through the portal vein. Hepatocytes are dissociated by collagenase for their release from the Glisson's capsule. After washing and careful centrifugation, the hepatocytes can be used for any downstream analyses. In conclusion, this paper describes a straightforward and reproducible technique for the isolation of a representative population of regenerating hepatocytes after partial hepatectomy in mice. The method may also aid the study of fatty liver disease.

Lipid-laden hepatocytes are inherent to liver regeneration but are usually lost upon density-gradient centrifugation. Here, we present an optimized cell isolation protocol that retains steatotic hepatocytes, yielding representative populations of regenerating hepatocytes after partial hepatectomy in mice.

Aislamiento de hepatocitos regeneradores después de hepatectomía parcial en ratones

Partial hepatectomy has been widely used to investigate liver regeneration in mice, but the isolation of high yields of viable hepatocytes for downstream ...

Development and Validation of a Porcine Ex Situ Isolated Liver Perfusion Model 

Porcine Normothermic Isolated Liver Perfusion

Porcine models of liver ex situ normothermic machine perfusion (NMP) are increasingly being used in transplant research. Contrary to rodents, porcine livers are anatomically and physiologically close to humans, with similar organ size and bile composition. NMP preserves the liver graft at near-to-physiological conditions by recirculating a warm, oxygenated, and nutrient-enriched red blood cell-based perfusate through the liver vasculature. NMP can be used to study ischemia-reperfusion injury, preserve a liver ex situ before transplantation, assess the liver's function prior to implantation, and provide a platform for organ repair and regeneration. Alternatively, NMP with a whole blood-based perfusate can be used to mimic transplantation. Nevertheless, this model is labor-intensive, technically challenging, and carries a high financial cost.
In this porcine NMP model, we use warm ischemic damaged livers (corresponding to donation after circulatory death). First, general anesthesia with mechanical ventilation is initiated, followed by the induction of warm ischemia by clamping the thoracic aorta for 60 min. Cannulas inserted in the abdominal aorta and portal vein allow flush-out of the liver with cold preservation solution. The flushed-out blood is washed with a cell saver to obtain concentrated red blood cells. Following hepatectomy, cannulas are inserted in the portal vein, hepatic artery, and infra-hepatic vena cava and connected to a closed perfusion circuit primed with a plasma expander and red blood cells. A hollow fiber oxygenator is included in the circuit and coupled to a heat exchanger to maintain a pO2 of 70-100 mmHg at 38 &#176;C. NMP is achieved by a continuous flow directly through the artery and via a venous reservoir through the portal vein. Flows, pressures, and blood gas values are continuously monitored. To evaluate the liver injury, perfusate and tissue are sampled at predefined time points; bile is collected via a cannula in the common bile duct.

The porcine model of liver normothermic machine perfusion (NMP), described here, can be successfully used to study NMP as a preservation strategy, a tool for viability assessment, and a platform for organ repair. It holds a high translational value, however it is technically challenging and labor-intensive.

Porcine models of liver ex situ normothermic machine perfusion (NMP) are increasingly being used in transplant research. Contrary to rodents, porcine ...