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.

Pazienti hanno fornito consenso informato scritto e lo studio è stato approvato dalla Regione Ethical Review Board presso l&#39;Università di Göteborg. 1. Anestesia <ol><li> Impostare tutto il monitoraggio (ECG, pressione sanguigna bracciale) 17. </li><li> Inserire un catetere epidurale in basso a metà livello toracico 17. </li><li> Indurre l&#39;anestesia con propofol 2 mg / kg, 150 mg di fentanyl, e Rocuronium 0,6 mg / kg attraverso un catetere venoso periferico precedentemente inserito. </li><li> Preoxygenate e intubare il paziente. Mantenere l&#39;anestesia con sevoflurano, l&#39;ossigeno e l&#39;aria 17. </li><li> Inserire un doppio lume catetere venoso centrale e un catetere arterioso. Monitorare CVP e pressione arteriosa 17. </li></ol> 2. Isolamento del Fegato e Incannulazione delle navi <ol><li> Somministrare 100 IE / kg di eparina e dare profilassi antibiotica (ad esempio, 160 mg di trimetoprim e 800 mg di Sulfamethoxazole). </li><li> Utilizzando la tecnica Seldinger percutanea, posizionare un catetere P. 17 nella vena femorale e un padre del catetere 15 nella vena giugulare esterna 18. Verificare le posizioni dei cateteri dalla fluoroscopia. </li><li> Eseguire una laparotomia da un&#39;incisione a forma di L dal processo xifoideo lungo la linea mediana e poi esteso a un diritto un&#39;incisione sottocostale 19. </li><li> Verificare che non vi siano metastasi extra epatica nell&#39;addome per una completa ispezione e palpazione. <ol><li> Se vengono rilevati lesioni sospette, verificare malignità da sezioni congelate. </li><li> Terminare la procedura se vi è evidenza di manifestazione extraepatica (adenopatia periaortal, metastasi addominali). Linfoadenopatia nel epatico non è considerata una controindicazione. </li></ol></li><li> Mobilitare il fegato dividendo il legamento falciforme, il legamento triangolare di sinistra e destra e il legamento coronario 19. </li><li> Sezionare il Cavalvenosa partire dalle vene renali fino al diaframma con legatura delle vene tributarie retroperitoneali. </li><li> Identificare il diritto vena surrenalica e mettere un cappio nave intorno ad esso. </li><li> Identificare il diritto della vena gonadica. Sezionarlo per 5-10 cm. Collocare una sutura a borsa di intorno all&#39;ingresso della vena cava. <ol><li> Posizionare una legatura distale sulla vena gonadica. Utilizzando forbici, aprire la vena gonadica 3-5 cm dalla vena cava. </li><li> Inserire un 18 Fr catetere filo-rinforzato attraverso il diritto della vena gonadica e posizionare la punta retrohepatically sopra le vene renali. </li><li> Inserire una legatura intorno alla vena gonadica per fissare la posizione del catetere. </li><li> Verifica ritorno del sangue attraverso il catetere. Eparinizzare il catetere 18. </li></ol></li><li> Sezionare il legamento epatoduodenale e visualizzare l&#39;arteria epatica comune, l&#39;arteria gastroduodenale e l&#39;arteria epatica propria. <ol><li> Luogo nave loop intorno arte epatico comunery e l&#39;arteria gastrodoudenal. </li><li> Sezionare la vena porta e mettere un cappio nave intorno ad esso. </li><li> Isolare il resto del legamento contenente il dotto biliare comune e comprendono tutto questo in un ciclo vaso. </li></ol></li><li> Sollevare l&#39;arteria gastroduodenale con il ciclo nave. Dopo l&#39;interruzione del flusso arterioso, fare un&#39;incisione con le forbici. <ol><li> Inserire un 8-12 Fr catetere con la sua punta nell&#39;arteria epatica propria. Fissare la posizione con una legatura intorno all&#39;arteria gastroduodenale. </li><li> Verificare ritorno del sangue nel catetere e poi eparinizzare esso 18. </li></ol></li><li> Prime il sistema di perfusione con una unità di eritrociti confezionati, 100 ml di albumina (50 g / L), 100 ml di Tribonat (bicarbonato di sodio), 2500 unità di eparina (5.000 IE / ml) e 250 ml di cristalloidi. </li><li> Collegare arterioso e cateteri vena cava al sistema di perfusione costituiti da una macchina cuore-polmone dotato di una pompa a rulli e un riscaldatore together con venose sistemi reservoir, ossigenatori e tubi di tipo usa e getta. </li><li> Collegare i cateteri femorali giugulari ed esterni alla pompa bypass venovenosa costituito da una pompa centrifuga, con un riscaldatore e iniziare bypass. </li><li> Completare l&#39;isolamento del fegato stringendo un laccio emostatico Rumel intorno alla vena cava infrahepatically sopra le vene renali e bloccare la vena cava suprahepatically utilizzando un morsetto vascolare Klintmalm. <ol><li> Stringere un laccio emostatico Rumel intorno al dotto biliare comune, la vena porta e la vena surrenalica destra e infine bloccare l&#39;arteria epatica con un morsetto vascolare. </li></ol></li></ol> 3. La perfusione <ol><li> Avviare la perfusione e controllare manualmente il livello ematico nel serbatoio venoso. Regolare il flusso fino a raggiungere un livello di stato stazionario, di solito una portata tra 500-1,200 ml / min. </li><li> Mettere sonde termistore nei giusti e fegato lasciato lobi e uno direttamente inil catetere arterioso. </li><li> Riscaldare il perfusato mira a una temperatura obiettivo di fegato di 40 o C. Mantenere questa temperatura per tutta la perfusione. </li><li> Posizionare una sonda scintillazione sopra la testa della pompa bypass venovenosa. </li><li> Calibrare il sistema di monitoraggio di dispersione, iniettando una piccola dose di 10 MBq 99m Tc-albumina (1 ml) nella circolazione sistemica e registrare l&#39;attività basale. <ol><li> Iniettare un dieci volte più alta dose, 100 MBq 99m Tc-albumina (1 ml), nell&#39;unità di perfusione epatica. </li><li> Registra qualsiasi fuoriuscita dal sistema di perfusione alla circolazione sistemica come un aumento dell&#39;attività misurata dalla sonda scintillazione. </li><li> Registra qualsiasi perdita dalla circolazione sistemica al sistema di perfusione come un aumento del volume nel serbatoio venoso. </li></ol></li><li> Quando viene stabilita stato stazionario della portata, nessuna perdita è presente e la temperatura del fegato ha raggiunto 40 ° C, iniettare melphalan alla dose di1 mg / kg di peso corporeo nel sistema di perfusione diviso in due dosi somministrate 30 min a parte. </li></ol> 4. Terminare la perfusione e Rimuovi Cateteri <ol><li> Dopo 60 min di perfusione, svuotare il perfusato dal sistema e irrigare il fegato con 1.000 ml di crystalloid. </li><li> Aggiungere una unità di eritrociti confezionati all&#39;unità perfusione, infondere nel fegato e quindi fermare la perfusione. </li><li> Rilasciare il morsetto Klintmalm e il laccio emostatico Rumel dalla vena cava. </li><li> Rilasciare il laccio emostatico Rumel dal legamento epatoduodenale. </li><li> Rimuovere la legatura dalla arteria gastroduodenale. Rimuovere il catetere e legare il vaso. </li><li> Rilasciare il morsetto dalla arteria epatica. </li><li> Rimuovere il catetere dalla vena gonadica e legare il vaso. </li><li> Fermare la pompa di bypass veno-venosa e scollegare i tubi. </li><li> Chiudere l&#39;addome con una sutura fasciale ininterrotta e poi una sutura cutanea intracutaneous. </li><li> Rimuovere la catheters nelle vene giugulari femorali ed esterni e applicare pressione locale per 5 min. </li></ol> 5. Cura postoperatoria <ol><li> Estubare il paziente e il trasferimento di unità di terapia post-operatoria per 24 ore. Mantenere il controllo della pressione venosa centrale e pressione arteriosa. </li><li> Trasferire il paziente in un reparto di chirurgia per un tipico di 3-7 giorni di cure post-operatorie. </li><li> Continuare con basso peso molecolare eparina (LMWH) alla dose di 5000 IE / giorno per un totale di 4 settimane. </li></ol>

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The authors have nothing to disclose.

I pazienti con melanoma metastatico uveale hanno poche opzioni terapeutiche. Chemioterapia 6, immunoterapia 23 e terapia mirata 24 non sono stati in grado di mostrare una migliore OS. Per i pazienti con poche metastasi epatiche, resezione epatica è un&#39;opzione 12. Altri trattamenti includono la chemioterapia intra-arteriosa epatica (HIA) 9, chemioembolizzazione transarteriosa (TACE) 25 e la terapia selettiva interna di radiazioni (SIRT) 26....

Il melanoma uveale è il tumore maligno intraoculare primario più comune negli adulti, con l&#39;essere la più alta incidenza nelle popolazioni caucasiche 1. Trattamento locale si compone di enucleazione o placca locale radioterapia 2,3. Indipendentemente dal trattamento locale, metastasi in definitiva in circa il 35-50% dei pazienti 4,5. Il fegato è il sito più comune di metastasi e circa il 50% dei pazienti presentano isolate metastasi epatiche. La sopravvivenza globale mediana (OS) per i pazienti con metastasi epatiche è di 6-12 mesi 4. Diversi diverse strategie di trattamento sistemici e regionali sono state esplorate, ma i tassi di sopravvivenza non sono migliorati 2. Il trattamento con la chemioterapia sistemica, sia singola sostanza (dacarbazina o temozolomide 6) o trattamenti di combinazione (regime BOLD 7, gemcitabina con Treosulfan 8), mostra tassi di risposta inferiori poi10%. Epatica chemioterapia intra-arteriosa (HIA) ha mostrato un tasso leggermente superiore complessiva risposta rispetto alla chemioterapia sistemica, ma nessun aumento della sopravvivenza globale 9. Chemioembolizzazione transarteriosa (TACE) con cisplatino e carboplatino ha mostrato una risposta parziale nel 57% dei pazienti con una sopravvivenza mediana di 9 mesi 10. Radioterapia interna selettiva (SIRT) ha mostrato una risposta parziale nel 62% dei pazienti con una sopravvivenza mediana di 7 mesi 11. In uno studio su resezione epatica, inclusi 255 pazienti, è stata riportata una sopravvivenza complessiva di 14 mesi; in 76 dei pazienti in cui una resezione microscopicamente completo è stato possibile, la sopravvivenza era 27 mesi 12. Perfusione epatica isolata (IHP) è una opzione di trattamento regionale, originariamente sviluppato da Ryan e Ausman. La logica alla base della tecnica è quello di isolare chirurgicamente e profumato fegato con una dose elevata di un agente chemioterapico, evitando sistemicoxicity 13. Inoltre, ipertermia concomitante media un maggiore assorbimento dell&#39;agente chemioterapico da variazioni del flusso sanguigno tumorale e la permeabilità delle membrane cellulari 14. Completo isolamento vascolare è confermata con una tecnica di tracciante radioattivo con Tc-99 etichettata albumina sierica umana 15. Una retrospettiva studio di fase II ha mostrato un potenziale beneficio di sopravvivenza di 14 mesi (26 vs 12 mesi) per i pazienti sottoposti IHP rispetto ai sopravvissuti più lunghe con metastasi epatiche melanoma uveale in Svezia 16. Ecco a voi la tecnica di IHP con una breve rassegna dei risultati attuali e prospettive future.

<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

Isolato perfusione epatica (IHP) è una procedura in cui il fegato è chirurgicamente isolato e perfuso con un&#39;alta concentrazione di melfalan agente chemioterapico. In breve, la procedura inizia con la messa a punto di un veno-venosa bypass percutanea dalla vena femorale alla vena giugulare esterna. Via una laparotomia, cateteri vengono poi inseriti in arteria epatica propria e la vena cava. La vena porta e la vena cava, sia sopra- e infrahepatically, vengono poi bloccato. Il arteriosa e cateteri venosi sono collegati ad una macchina cuore-polmone e fegato vengono perfuse con melfalan (1 mg / kg di peso corporeo) per 60 min. In questo modo è possibile defluire in localmente fegato con una dose elevata di un agente chemioterapico, senza perdite nella circolazione sistemica. In studi precedenti, inclusi i pazienti con metastasi epatiche isolate di melanoma uveale, un tasso di risposta complessiva del 33-100% e una sopravvivenza media tra i 9 ei 13 mesi,sono stati riportati. L&#39;obiettivo di questo protocollo è quello di fornire una chiara descrizione di come eseguire la procedura e discutere IHP come opzione di trattamento per le metastasi epatiche di melanoma uveale.

Una prima relazione comprendeva 22 pazienti trattati con melfalan con o senza TNF-alfa. I risultati hanno mostrato un tasso di risposta globale del 62%, tra cui due pazienti con CR (10%), e una sopravvivenza globale mediana di 11 mesi 20. Uno studio successivo ha riferito sui risultati di 29 pazienti consecutivi trattati con melfalan solo. Il risultato è stato molto simile, con una risposta globale del 62% di cui 10% di risposta completa e una sopravvivenza media di 12 mesi 21. <p class="jove_...

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.

Isolata epatica perfusione come Trattamento di metastasi epatiche di melanoma uveale

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 Views.  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.

Modello Murine dei tumori metastatici del fegato nell'impostazione della lesione alla riperfusione dell'ischemia

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

Ricerca

Ricerca sul cancro

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.

Generazione di una piattaforma di Melanoma Uvenoma Uvenoft umano del fegato in topi immunodeficienti

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.

Iniezione di vena porta di organoidi del cancro del colon-retto per studiare le metastasi epatiche Stroma

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.

Tecnica di Subnormothermic<em&gt; Ex Vivo</em&gt; Fegato perfusione per la Conservazione, valutazione e riparazione dei marginali fegato innesti

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.

Impianto e valutazione del melanoma nella coroide murina mediante tomografia a coerenza ottica

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.

Percutanea epatica Perfusione (PHP) con melfalan come trattamento per non resecabili metastasi confinata al fegato

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.

Visualizzazione dei Vascolare e parenchimale rigenerazione dopo il 70% epatectomia parziale in topi normali

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 piccolo modello animale di aspersione epatica normotermica 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.

Isolamento di epatociti rigeneranti dopo epatectomia parziale nei topi

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 ...