The Vienna General Hospital and the Medical University of Vienna kindly provides the infrastructure for the Vienna Hepatic Hemodynamic Laboratory. Previous members of the Vienna Hepatic Hemodynamic Laboratory and coworkers should be acknoweledged for their valuable input that helped to continuously improve the methodology of HVPG measurement and transjugular liver biopsy at our institution. In addition, we thank the nurses of the Division of Gastroenterology and Hepatology who are an essential part of the Vienna Hemodynamic Laboratory and continuously provide patients with excellent care.

The protocol described here complies with the guidelines of the human research ethics committee of the Medical University of Vienna.
1. Preparations
<ol>
	<li>Use a specialized room dedicated/approved for procedures using x-ray equipped with a digital x-ray fluoroscopy&#160;system (Figure 1A), a monitor system (Figure 1B), a pressure recorder and transducer (Figure 1C) that are connected to a printer or recording software, and an ultrasound device (Figure 1D). Also, ensure there is sufficient space for a sterile working area (Figure 1E) next to the patient bench.</li>
	<li>Evaluate patients with suspected advanced chronic liver disease (ACLD) for CSPH by measurement of HVPG. In particular, patients with one of the following features might undergo HVPG measurement: thrombocytopenia &#60;150 G/L, portosystemic collaterals on cross-sectional imaging, gastroesophageal varices prior to initiation of non-selective betablocker therapy, ascites, and hepatic tumors that are scheduled for resection.</li>
	<li>Use the following contraindications for the procedure: (i), absence of vascular access due to jugular or caval vein thrombosis, (ii) clinically evident bleeding disorders (e.g. disseminated intravascular coagulation), (iii) contralateral pneumothorax, and (iv) significant cardiac arrhythmias.</li>
	<li>Ensure that the patient is fastened and gave written informed consent after being informed about the purpose and potential risks of the procedure.</li>
	<li>Explain that the procedure must be performed without general anesthesia. However, low doses of midazolam (up to 0.02 mg/kg bodyweight) might be offered to anxious patients.</li>
	<li>Monitor the patient&#8217;s vital signs by non-invasive arterial blood pressure measurement, pulse oximetry, and electrocardiography using a standard monitor system.</li>
	<li>Flush the pressure transducer set using sterile saline solution in a pressure bag.</li>
	<li>Calibrate the pressure transducer (if not precalibrated, calibrate against an external pressure reference before starting measurements, e.g. by using a water column where 13.6 cm H2O equals 10 mm Hg).</li>
	<li>Place the transducer at the level of the right atrium by aiming at the patient&#8217;s midaxillary line.</li>
	<li>Prepare the pressure recorder/recording software to be ready for recording pressure tracings.</li>
	<li>Ensure that all necessary sterile equipment is ready (see Table of Materials).</li>
	<li>Ask the patient to take a supine position on the patient bench.</li>
	<li>Have the operator wash and disinfect her/his hands and forearms.</li>
	<li>Put on surgical cap and face mask, sterile gloves and a sterile coat.</li>
	<li>Use a sterile table cover and prepare a sterile working area for the equipment needed for the central venous access (Figure 2A).</li>
</ol>
2. Central venous access under sterile conditions
<ol>
	<li>Instruct the patient to&#160;turn his/her head slightly to the left side, then disinfect the area of the right anterior and lateral neck with a disinfectant solution.</li>
	<li>Place sterile towels that cover the skin around the carotid triangle.</li>
	<li>Use a ultrasound probe (covered by a sterile US probe cover sheath) to identify the right internal jugular vein and the appropriate puncture site.</li>
	<li>Infiltrate the area of subsequent vascular access with a 21 G needle and apply a local anaesthetic (lidocaine 2%, 5 mL). Then, wait 1&#8211;2 min for its full effect.</li>
	<li>Prepare all materials needed for central venous access (see Table of Materials).</li>
	<li>Using the equipment of the 7.5 F catheter introducer sheath set, place the needle in the right internal jugular vein using ultrasound guidance and advance the guidewire through the needle using the Seldinger technique (Figure 3A). 
	NOTE: If a transjugular biopsy should be performed after HVPG measurement, a larger diameter 10 F liver biopsy introducer sheath set (with extra 18 G needle and guide wire) has to be used.</li>
	<li>Make a 2-3 mm incision along the guidewire using a blade 11 scalpel to ease introduction of the vascular access sheath loaded with the dilatator.</li>
	<li>Insert the vascular access percutaneous sheath&#160;loaded with the dilatator into the internal jugular vein over the guidewire.</li>
	<li>Remove the guidewire and the dilatator of the vascular access sheath.</li>
	<li>Make sure that the vascular access sheath remains in&#160;a stable intravascular position and orient the injection port of the vascular access sheath towards the lateral side (Figure 3B). 
	NOTE: Suturing or taping is not needed.</li>
</ol>
3. Placement of the balloon catheter in a hepatic vein
<ol>
	<li>Prepare all materials needed for placement of the balloon catheter into a&#160;hepatic vein (Figure 2B).</li>
	<li>Flush the balloon catheter with contrast media (dye) and check the integrity of the balloon by repetitive insufflation/deflation with the catheter tip immersed in saline solution. No air bubbles should occur. 
	NOTE: Depending on the visibility of the catheter itself on x-ray, you may also use just sterile saline or diluted dye to flush the vascular lumen of the catheter.</li>
	<li>Moisten the balloon catheter on the outside with sterile saline solution before inserting it into the vascular access sheath.</li>
	<li>Advance the balloon catheter under fluoroscopic guidance into the inferior cava vein (IVC). Aligning the tip of the balloon catheter towards the back of the patient and slight rotations of the catheter usually allows to advance from the right atrium to the IVC. Instructing the patient to inhale deeply might help in some cases (Figure 3B).</li>
	<li>Advance the balloon catheter from the IVC into the hepatic vein. Try to get access to the hepatic veins by repeatedly moving with the tip of the balloon catheter facing to the right towards the suspected area of the junction of the hepatic veins and the IVC (Figure 3C).</li>
	<li>Ensure that the catheter is advanced into a stable position that allows the repeated measurement of the free hepatic venous pressure (FHVP) at a 2-4 cm distance from its opening into the IVC and adequate space for the inflated balloon in the lumen of the hepatic vein in order to record the wedged hepatic venous pressure (WHVP).</li>
	<li>Check for an adequate occlusion of the vein (wedge position) by inflating the balloon (about 2 mL of air into the balloon lumen) and contrast agent&#160;injection (about 5 mL into the vascular lumen) until the hepatic vein distal to the inflated balloon is visualized (Figure 3D).</li>
	<li>Observe the stasis of the contrast media and exclude washout of the contrast media due to insufficient occlusion of the venous lumen by the balloon or due to the presence of vein-vein communications. If significant contrast media washout is observed, try to reposition the balloon catheter.</li>
	<li>Deflate the balloon and flush the lumen of the catheter with saline.</li>
</ol>
4. Hemodynamic readings for assessing the HVPG
<ol>
	<li>Connect the vascular lumen of the balloon catheter to the pressure transducer using an infusion line.</li>
	<li>Start recording the FHVP with the tip of the balloon 2-4 cm from the opening of the hepatic vein to the IVC. The waveform of the curve must be stable without variations over time. 
	NOTE: Stable values are usually obtained after 15 s.</li>
	<li>Inflate the balloon and continue recording the WHVP until the measurement becomes a stable horizontal line with no variations over time. 
	NOTE: Stable tracings of the wedged pressure are usually obtained after &#62;40 s.</li>
	<li>Repeat the measurement of FHVG (&#62;15 s) and WHVP (&#62;40 s) at least three times to obtain triplicate high-quality readings (Figure 3E). 
	NOTE: If significant discrepancies of &#8805;2 mm Hg are observed between the single FHVP/WHVP readings, additional measurements should be obtained. Note the reasons for potential artefacts, such as coughing, moving, or talking at the respective timepoints of hemodynamic recording.</li>
	<li>Record pressure in the IVC at the level of the ostium of the hepatic vein as well as the right atrial pressure (RAP).</li>
	<li>Stop recordings.</li>
	<li>Remove the balloon catheter.</li>
	<li>Calculate HVPG (FHVP subtracted from WHVP) as the mean of 3 measurements. 
	NOTE: If significant differences &#8805;2 mm Hg are noted between the sequentially obtained HVPG readings, additional measurements should be obtained.</li>
	<li>Proceed to transjugular biopsy (step 5.1) or remove the catheter introducer sheath from the internal jugular vein. 
	NOTE: Inserting a stiff guidewire may help to reach the same intrahepatic position for subsequent transjugular liver biopsy.</li>
	<li>Apply pressure on top of the vascular insertion site of the internal jugular vein for at least 5 min using a sterile gauze.</li>
</ol>
5. Preparation for transjugular liver biopsy
NOTE: Two different biopsy methodologies may be used to obtain a transjugular liver biopsy: aspiration (step 6) or core biopsy (step 7). First decide which system to use and then select the appropriate biopsy needle introducer sheath before proceeding with steps 5.1 to 5.3). Alternatively, the needle of the core TJBLX set can also be inserted into the aspiration set (NOTE: use appropriate diameters), which results in a more flexible core biopsy system that may be easier to introduce into a hepatic vein (NOTE: this approach is not described in detail here).
<ol>
	<li>Prepare the respective transjugular liver biopsy (TJLBX) set (aspiration TJLBX: Figure 4A-B; core TJLBX: Figure 4C; see Table of Materials).</li>
	<li>Flush the biopsy needle introducer sheath with sterile saline or, in case of transjugular aspiration liver biopsy, with contrast agent&#160;for better visualization.</li>
	<li>Advance the biopsy needle introducer sheath into a hepatic vein using the same technique as described in 3.4.</li>
	<li>Depending on the biopsy set used, proceed either to step 6.1 for aspiration TJBX&#160;or to step 7.1 for core TJBX.</li>
</ol>
6. Transjugular aspiration liver biopsy
<ol>
	<li>Use a 10 mL Luer-lock syringe to flush the aspiration TJLBX needle with sterile saline, however, leave about 3 mL in the syringe to facilitate aspiration.</li>
	<li>Advance the biopsy needle gently through the biopsy needle introducer sheath until the tip of the needle reaches the end of the introducer sheath. 
	NOTE: Avoid any force or fast movements while advancing the needle. Asking the patient to take a deep breath will decrease the angle between the IVC and the hepatic veins for easier advancement of the biopsy needle.</li>
	<li>Ask the patient to hold his breath.</li>
	<li>Apply suction using the 10 mL Luer-lock syringe and advance the needle into the liver parenchyma.</li>
	<li>Retract the needle while still applying suction (negative pressure).</li>
	<li>Advice the patient to continue breathing normally.</li>
	<li>Remove the needle (but not the needle introducer sheath) and harvest the liver sample. 
	NOTE: The liver sample is usually retained in the syringe, not in the needle (Figure 4D).</li>
	<li>Repeat steps 6.1 to 6.6 for additional needle passages until sufficient liver specimens are obtained.</li>
	<li>Inject 5-10 mL of contrast media over the catheter introducer sheath to rule out perforation of the liver capsule.</li>
	<li>Remove the biopsy needle introducer sheath.</li>
	<li>Remove the 10 F liver biopsy set introducer sheath and apply pressure on top of the vascular insertion site of the internal jugular vein for at least 5 min using sterile gauzes.</li>
</ol>
7. TRANSJUGULAR CORE LIVER BIOPSY
<ol>
	<li>Load the core TJLBX needle by pulling the grip until the shooting mechanism is locked. Advance the core biopsy needle gently through the biopsy needle introducer sheath until the tip of the needle approaches the end of the introducer sheath. 
	NOTE: Avoid any force or fast movements while advancing the needle.</li>
	<li>Ask the patient to hold his/her breath.</li>
	<li>Advance the needle into the liver parenchyma.</li>
	<li>Perform the core biopsy by pulling the trigger of the shooting mechanism.</li>
	<li>Advice the patient to continue breathing normally.</li>
	<li>Remove the needle (but not the needle introducer sheath) and harvest the liver sample. A small needle might help to remove the liver sample (Figure 4D).</li>
	<li>Repeat steps 7.1 to 7.6 for additional needle passages until sufficient liver specimens are obtained.</li>
	<li>Inject 5-10 mL of contrast media over the side port of the catheter introducer sheath to rule out perforation of the liver capsule.</li>
	<li>Remove the biopsy needle introducer sheath.</li>
	<li>Remove the 10 F liver biopsy set introducer sheath and apply pressure on top of the vascular insertion site of the internal jugular vein for about 5 min using sterile gauzes.</li>
</ol>

Thomas Reiberger has received grant and material support by Cook Medical as well as grant support and honoraria for lectures from W.L. Gore &#38; Associates. Philipp Schwabl, Markus Peck-Radosavljevic and Michael Trauner have nothing to disclose in regard to this manuscript. Mattias Mandorfer has received honoraria for lectures from W.L. Gore &#38; Associates.

While HVPG measurements require considerable resources and trained personal with interventional skills and expertise in the reading of pressure tracings, it improves prognostication and might guide treatment decisions, and thus, facilitates personalized medicine. In addition, the opportunity to safely obtain liver biopsy specimens via the transjugular route in the same session is another argument in favor of implementing hepatic hemodynamic laboratories at tertiary care centers. Indeed, guidelines support the use of HVPG...

Patients with cirrhosis are at risk for developing complications mostly related to portal hypertension (PHT), such as ascites or bleeding from gastric or esophageal varices1,2,3. The risk of hepatic decompensation is related to the degree of PHT2. Measurement of the hepatic venous pressure gradient (HVPG) is the gold standard to estimate portal venous pressure in patient with cirrhosis, i.e. assessing the severity of sinusoidal&#160;portal hypertension4. An HVPG of &#8805;6 mm Hg to 9 mm Hg indicates elevated portal pressure (&#39;subclinical portal hypertension&#39;), while an HVPG &#8805;10 mm Hg defines CSPH. This protocol provides a detailed description of the equipment and the procedure and also highlights potential pitfalls and offers advice for troubleshooting.
Clinically, measurement of HVPG is indicated (i) to establish the diagnosis of sinusoidal&#160;portal hypertension, (ii) to identify patients at risk for hepatic decompensation by diagnosing CSPH (HVPG &#8805;10 mm Hg), (iii) to guide pharmacological therapy in primary or secondary prophylaxis of variceal bleeding, and (iv) to assess the risk of hepatic failure after partial hepatectomy2,4. HVPG is used as an established surrogate marker for improvement and/or worsening of liver fibrosis/function, since a decrease in HVPG translates into a clinically meaningful benefit5, whereas higher HVPG values are associated with an increased variceal bleeding risk6. Based on observations on changes in HVPG in patients under non-selective beta-blocker (NSBB) or etiological therapies, a decrease in HVPG of 10% is considered to be clinically relevant7,8.
To date, there are no alternative, non-invasive parameters reflecting the degree of portal pressure with similar accuracy as HVPG. Even if HVPG is actually an &#8216;indirect&#8217; way to measure portal pressure, it strongly correlates and thus accurately reflects &#8216;directly&#8217; measured portal pressure in patients with cirrhosis9. Importantly, HVPG measurements should be performed using a balloon catheter to maximize the assessed amount of liver parenchyma10,11,12. Although HVPG measurements are invasive, resource-intensive, and require interventional skills and expertise in interpreting the reliability and plausibility of pressure readings, this method is the current gold standard for diagnosing and monitoring portal hypertension in patients with cirrhosis13,14,15.
Simple laboratory values, such as platelet count, may help to estimate the likelihood for CSPH. However, platelet count, or non-invasive scores that include platelet count, have limited predictive value16. Imaging modalities showing splenomegaly17 or portosystemic collaterals18 in patients with cirrhosis suggest the presence of CSPH, but are not helpful for quantifying the actual degree of portal hypertension. Novel non-invasive imaging tools, such as elastography of the liver19 and/or of the spleen20 are useful for ruling-in or ruling-out the presence of CSPH. Still, none of the available methods is able to directly measure dynamic changes in portal pressure21.
The prognostic value of HVPG has been underlined by several landmark studies, showing that a HVPG &#8805;10 mm Hg (i.e. CSPH) is predictive for the formation of varices8 (and for the development of complications related to portal hypertension22, while a (pharmacologically-induced) decrease of HVPG modulates the respective risk of variceal growth23 and decompensation7. HVPG-response is the only established surrogate for the effectiveness of NSBBs in preventing (recurrent) variceal bleeding. If HVPG decreases to a value of &#8804;12 mm Hg or is reduced by &#8805;10-20% during NSBB treatment, patients are protected from variceal bleeding and survival is increased24,25. Similarly, HVPG-response also decreases the incidence of ascites and related complications in patients with compensated cirrhosis5,26. Several studies have provided evidence supporting the use of HVPG-guided therapy27,28,29,30,31,32. Thus, in centers with sufficient experience, HVPG-response may&#160;guide treatment decisions, facilitating personalized medicine for patients with portal hypertension.
Moreover, measuring of HVPG might serve as a surrogate endpoint for proof-of-concept studies assessing the effectiveness of novel treatments for cirrhosis and/or portal hypertension being translated from bench to bedside, such as sorafenib33,34, simvastatin35,36, taurine37, or emricasan38. Ultimately, measurements of HVPG can also provide important prognostic information about the risk for development of HCC39 and for liver failure post hepatic resection40.
The infrastructure to measure HVPG should be readily available at secondary and tertiary care centers. Since the technique of HVPG measurement requires specialized training and equipment, it seems rational for academic and transplant centers to establish a hepatic hemodynamic laboratory, facilitating state-of-the-art diagnosis and management of portal hypertension. Large volume centers perform several hundred HVPG measurements per year. Based on our experience, sufficient expertise to perform accurate HVPG measurements is usually obtained after 50-100 supervised HVPG measurements.

<table><tbody><tr><td>10 mL Luer-Lock syringe</td><td>Braun</td><td>REF 4617100V, LOT 17G03C8</td><td>Luer-Lock Syringe for connection with the aspiration biopsy set</td></tr><tr><td>10 mL Syringe 2x</td><td>Braun</td><td>REF 4606108V</td><td>Snonpyrogenic, nontoxic 10 mL syringe</td></tr><tr><td>10 F liver biopsy&amp;nbsp; introducer sheath set</td><td>Cook Medical</td><td>REF RCFW-10.0-38, REF G07600</td><td>Percutaneous Sheath Introducer Set (TJBX), 10F Port, 13cm, Check-Flo Performer Introducer</td></tr><tr><td>18 G needle for biopsy introducer sheat</td><td>Arrow International</td><td>REF AN-04318</td><td>Introducer Needle for TJBX Set, 18 G, 6.35 cm</td></tr><tr><td>21 G needle</td><td>Henke Sass Wolf</td><td>REF 0086, Fine-Ject 21Gx2&quot;</td><td>Sterile injeciton needle, 21 Gx2&quot;, 0.8 x 50 mm, for local anesthesia</td></tr><tr><td>3-way channels</td><td>Becton Dickinson</td><td>BD Connecta Luer-Lok, REF 39402</td><td>Three way channel with Luer-Lok connection system</td></tr><tr><td>7.5 F catheter introducer sheath set</td><td>Arrow International</td><td>REF SI_09875-E</td><td>Percutaneous Sheath Introducer Set, 7.5 F Port</td></tr><tr><td>Aspiration TJLBX set</td><td>Cook Medical</td><td>REF RMT-16-51.0-TJL, REF G20521</td><td>TJ Liver Access and Biopsy Needle Set (Aspiration Set), 9 F-45 cm, 16 G-50.5 cm</td></tr><tr><td>Balloon catheter</td><td>Gerhard Pejcl Medizintechnik Austria</td><td>REF 500765B</td><td>Ferlitsch HVPG Catheter, 7F-65cm, Balloon:2.5mL, Pressure 50-90 kPa, GW: 0.032&quot;</td></tr><tr><td>Blade 11 scalpel</td><td>Medi-Safe Surgicals</td><td>MS Safety Scalpel REF /Batch 18012424</td><td>11 blade safety scalpels, retractable, single use, 10 scalpels per package</td></tr><tr><td>Blunt tip fill needle</td><td>Becton Dickinson</td><td>REF 303129</td><td>Sterile blunt tip fill needle, single use</td></tr><tr><td>Contrast media (dye)</td><td>Dr. Franz K&amp;ouml;hler Chemie GmbH, Bensheim, GER</td><td>ZNR 1-24112</td><td>Peritrast 300 mg Iod/mL, 50 mL, contrastmedia</td></tr><tr><td>Core TJLBX set</td><td>Cook Medical</td><td>REF RMT-14XT-50.5-LABS-100, REF G08285</td><td>TJ Needle Introducer and Bx-Needle 7 F-53.5 cm, 14 G-53.5 cm/20 mm, 18 G-60 cm</td></tr><tr><td>Digital x-ray fluoroscope system</td><td>Siemens</td><td>Model No 07721710</td><td>ARCADIS Varic, mobile x-ray fluoroscope system</td></tr><tr><td>Disinfectant solution</td><td>Gebro Pharma</td><td>1-20413</td><td>Isozid-H</td></tr><tr><td>Face mask</td><td>MSP Medizintechnik GmbH</td><td>REF HSO36984</td><td>Surgical face mask from double fleece, with tie, 50 pieces</td></tr><tr><td>Guide wire for biopsy introducer sheat</td><td>Arrow International</td><td>REF AW-14732</td><td>Marked Spring-Wire Guide, TJBX Set, 0.032&quot;, 0.81 mm, 68 cm</td></tr><tr><td>Infusion line</td><td>Rosstec Medical Products b Cardea GmbH &amp;amp; Co</td><td>REF 220010, 100m</td><td>Infusion line, Luer-Lok for connection of balloon catheter and pressure transducer</td></tr><tr><td>Lidocaine 2%</td><td>Gebro Pharma</td><td>Xylanaest, ZNR 17.792, 20mg/1mL (2%)</td><td>Sterile vials Xylanest including 2% Lidocain hydrochloride for local anesthesia</td></tr><tr><td>Midazolame</td><td>Roche</td><td>Dormicum, ZNr 1-18809, Midazolam 5mg/5mL</td><td>Sterile vials Dormicum including Midazolam for sedation</td></tr><tr><td>Monitor system</td><td>Datex-Ohmeda by GE</td><td>Type F-CMREC1</td><td>Patient monitoring system</td></tr><tr><td>Patient bench</td><td>Silerlen-MAQET</td><td>Model No 7474.00A</td><td>Mobile patient bench for x-ray fluoroscopy</td></tr><tr><td>Pressure bag</td><td>Ethox Corp</td><td>REF 4005</td><td>Pressure Infuser Bag 500 mL</td></tr><tr><td>Pressure recorder</td><td>Edwards Lifesciences</td><td>Ref T001631A, Lot 61202039</td><td>TruWave 3 cc/150 cm</td></tr><tr><td>Pressure transducer</td><td>Edwards Lifesciences TM</td><td>REF T001631A</td><td>Pressure Monitoring Set (1x), 3 cc/150 cm, TruWave TM</td></tr><tr><td>Recording software&amp;nbsp;</td><td>Datex Ohmeda by GE</td><td>Software S/5 is property of Instrumentarium Corp of Datex-Ohmeda TM</td><td>Datex-Ohmeda S/5TM Collect - Software to record pressure tracings of the patient monitor system</td></tr><tr><td>Sterile coat</td><td>Lohmann &amp;amp; Rauscher International GmbH &amp;amp; Co</td><td>REF 19351</td><td>Surgical Gown, Different sizes, e.g. L-130 cm</td></tr><tr><td>Sterile gauze</td><td>Hartmann</td><td>REF 401798, 10x10cm gauze</td><td>10 x 10 cm sterile gauze, 10 pieces per package</td></tr><tr><td>Sterile gloves</td><td>Meditrade</td><td>REF 9021</td><td>Gentle Skin sterile gloves, different sizes, e.g. 7.5</td></tr><tr><td>Sterile saline solution</td><td>Fresenius Kabi</td><td>NaCl 0.9%, B009827 REV 03</td><td>Physiologic Saline Solution 0.9% NaCl, 309 mosmol/L, pH-Wert: 4.5-7.0</td></tr><tr><td>Sterile saline vessel</td><td>KLS Martin</td><td>REF K8A,&amp;nbsp; 18/10 Jonas</td><td>Sterilizable Metal Vessel for sterile saline</td></tr><tr><td>Sterile table cover</td><td>Hartmann</td><td>REF 2502208</td><td>Table Cover, Foliodrape 150 x 100 cm</td></tr><tr><td>Sterile towel</td><td>BARRIER by M&amp;ouml;lnlycke Health Care</td><td>REF 706900</td><td>Adhesive OP-Towel, 100 x 100 cm</td></tr><tr><td>Sterile US probe cover sheath</td><td>Websinger</td><td>REF 07014</td><td>Sterile ultrasound probe cover, 20 x 60 cm, inluding two sterie adhesive tapes</td></tr><tr><td>Stiff guidewire</td><td>Cook Medical</td><td>REF TSMG-35-180-4-LES, G46729</td><td>Lunderquist Extra Stiff Wire Guide, 0.035&quot;-.89 mm, 180 cm, 4 cm flexible tip</td></tr><tr><td>Surgical cap</td><td>BARRIER by M&amp;ouml;lnlycke Health Care</td><td>REF 620500, PCS 100, Colour Green</td><td>Surgical Cap</td></tr><tr><td>Ultrasound device</td><td>FUJIFILM SonoSite</td><td>Model M-Turbo, REF P17000-17</td><td>Mobile ultrasound system</td></tr></tbody></table>

measurement of the hepatic venous pressure gradient and transjugular liver biopsy

Here we provide a detailed protocol describing the clinical procedure of hepatic venous pressure gradient (HVPG) measurement in patients with advanced chronic liver disease followed by an instruction for transjugular biopsy. Under local anesthesia and ultrasound guidance, a catheter introducer sheath is placed in the right internal jugular vein. Using&#160;fluoroscopic guidance, a balloon catheter is advanced into the inferior vena cava (IVC) and inserted into a large hepatic vein. Correct and sufficient wedge position of the catheter is ensured by injecting contrast media while the balloon is blocking the outflow of the cannulated hepatic vein. After calibrating the external pressure transducer, continuous pressure recordings are obtained with triplicate recordings of the wedged hepatic venous pressure (WHVP) and free hepatic venous pressure (FHVP). The difference between FHVP and WHVP is referred to as HVPG, with values &#8805;10 mm Hg indicating clinically significant portal hypertension (CSPH). Before removing the catheter, pressure readings obtained in the IVC at the same level, as well as the right atrial pressure are recorded.
Finally, a transjugular liver biopsy can be obtained via the same vascular route. Different systems are available; however, core biopsy needles are preferred over aspiration needles, especially for cirrhotic livers. Again, under fluoroscopic guidance a biopsy needle introducer sheath is advanced into an hepatic vein. Next, the transjugular biopsy needle is gently advanced through the introducer sheath: (i) in case of aspiration biopsy, the needle is advanced into the liver parenchyma under aspiration and then removed quickly, or (ii) in case of a core biopsy, the cutting-mechanism is triggered inside the parenchyma. Several separate passages can be safely performed to obtain sufficient liver specimens via transjugular biopsy. In experienced hands, the combination of these procedures takes about 30-45 min.

In compensated patients with well-preserved liver function (i.e. without any history of hepatic decompensation, such as ascites or variceal bleeding) the measured HVPG values might be normal or in the range of subclinical portal hypertension (HVPG 6&#8211;9 mm Hg). However, compensated patients might develop CSPH (HVPG &#8805;10 mm Hg) which indicates an increased risk for developing varices or hepatic decompensation. In turn, patients with esophageal or gastric varices, HVPG is usually in the range of CSPH, if not patie...

Watch this Scientific Journal Video about Measurement of the Hepatic Venous Pressure Gradient and Transjugular Liver Biopsy at JoVE.com

Measurement of the Hepatic Venous Pressure Gradient and Transjugular Liver Biopsy

Here, we present a protocol for measurement of hepatic venous pressure gradient (HVPG),the gold standard to diagnose clinically significant portal hypertension. Moreover, we describe how to perform a transjugular liver biopsy within the same session.

Here we provide a detailed protocol describing the clinical procedure of hepatic venous pressure gradient (HVPG) measurement in patients with advanced ...

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19.7K Views.  Medical University of Vienna. Hier presenteren we een protocol voor het meten van lever veneuze drukgradiënt (HVPG), de gouden standaard om klinisch significante portale hypertensie te diagnosticeren. Bovendien beschrijven we hoe we een transjugulaire leverbiopsie binnen dezelfde sessie kunnen uitvoeren.

Video: Measurement of the Hepatic Venous Pressure Gradient and Transjugular Liver Biopsy

Monitoring of Systemic and Hepatic Hemodynamic Parameters in Mice

The use of mouse models in experimental research is of enormous importance for the study of hepatic physiology and pathophysiological disturbances. However, due to the small size of the mouse, technical details of the intraoperative monitoring procedure suitable for the mouse were rarely described. Previously we have reported a monitoring procedure to obtain hemodynamic parameters for rats. Now, we adapted the procedure to acquire systemic and hepatic hemodynamic parameters in mice, a species ten-fold smaller than rats. This film demonstrates the instrumentation of the animals as well as the data acquisition process needed to assess systemic and hepatic hemodynamics in mice. Vital parameters, including body temperature, respiratory&#160;rate&#160;and heart&#160;rate were recorded throughout the whole procedure. Systemic hemodynamic parameters consist of carotid artery pressure (CAP) and central venous pressure (CVP). Hepatic perfusion parameters include portal vein pressure (PVP), portal flow rate as well as the flow rate of the common hepatic artery (table 1). Instrumentation and data acquisition to record the normal values was completed within 1.5 h. Systemic and hepatic hemodynamic parameters remained within normal ranges during this procedure.
This procedure is challenging but feasible. We have already applied this procedure to assess hepatic hemodynamics in normal mice as well as during 70% partial hepatectomy and in liver lobe clamping experiments. Mean PVP after resection (n= 20), was 11.41&#177;2.94 cmH2O which was significantly higher (P&#60;0.05) than before resection (6.87&#177;2.39 cmH2O). The results of liver lobe clamping experiment indicated that this monitoring procedure is sensitive and suitable for detecting small changes in portal pressure and portal flow rate. In conclusion, this procedure is reliable in the hands of an experienced micro-surgeon but should be limited to experiments where mice are absolutely needed.

This film demonstrates how to acquire systemic and hepatic hemodynamics in mice. The whole monitoring includes acquisition of vital parameters, systemic blood pressure, central venous pressure, common hepatic artery flow rate, and portal vein pressure as well as the portal flow rate in mice.

The use of mouse models in experimental research is of enormous importance for the study of hepatic physiology and pathophysiological disturbances. ...

Technique of Porcine Liver Procurement and Orthotopic Transplantation using an Active Porto-Caval Shunt

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation waiting list die without receiving an organ transplant or are delisted due to disease progression. Even after a successful transplantation, rejection and side effects of immunosuppression remain major concerns for graft survival and patient morbidity. 
Experimental animal research has been essential to the success of liver transplantation and still plays a pivotal role in the development of clinical transplantation practice. In particular, the porcine orthotopic liver transplantation model (OLTx) is optimal for clinically oriented research for its close resemblance to human size, anatomy, and physiology. 
Decompression of intestinal congestion during the anhepatic phase of porcine OLTx is important to guarantee reliable animal survival. The use of an active porto-caval-jugular shunt achieves excellent intestinal decompression. The system can be used for short-term as well as long-term survival experiments. The following protocol contains all technical information for a stable and reproducible liver transplantation model in pigs including post-operative animal care.

Experimental animal research plays a pivotal role in the development of clinical transplantation practice. The porcine orthotopic liver transplantation model (OLTx) closely resembles human conditions and is frequently used in clinically oriented research. The following protocol contains all information for a reliable porcine OLTx model using an active porto-caval-jugular shunt.

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation ...

Ex Vivo Hepatic Perfusion Through the Portal Vein in Mouse

Metabolic diseases such as diabetes, pre-diabetes, non-alcoholic fatty liver disease (NAFLD), and nonalcoholic steatohepatitis (NASH) are becoming increasingly common. Ex vivo liver perfusions allow for a comprehensive analysis of liver metabolism using nuclear magnetic resonance (NMR), in nutritional conditions that can be rigorously controlled. As in silico simulations remain a primarily theoretical means of assessing hormone actions and the effects of pharmaceutical intervention, the perfused liver remains one of the most valuable test beds for understanding hepatic metabolism. As these studies guide basic insights into hepatic physiology, results must be accurate and reproducible. The greatest factor in the reproducibility of ex vivo hepatic perfusion is the quality of surgery. Therefore, we have introduced an organized and streamlined method to perform ex vivo mouse liver perfusions in the context of in situ NMR experiments. We also describe a unique application and discuss common issues encountered in these studies. The overall purpose is to provide an uncomplicated guide to a technique we have refined over several years that we deem the golden standard for obtaining reproducible results in hepatic resections and perfusions in the context of in situ NMR experiments. The distance to the center of the field for the magnet as well as the inaccessibility of the tissue to intervention during the NMR experiment makes our methods novel.

Ex Vivo Hepatic Perfusion Through the Portal Vein in Mouse

The protocol describes a straightforward method of resectioning an intact mouse liver for metabolic studies through portal vein perfusion.

Metabolic diseases such as diabetes, pre-diabetes, non-alcoholic fatty liver disease (NAFLD), and nonalcoholic steatohepatitis (NASH) are becoming ...

Biochemistry

Contrast-Enhanced Subharmonic Aided Pressure Estimation (SHAPE) Using Ultrasound Imaging with a Focus on Identifying Portal Hypertension

Noninvasive, accurate measurement of pressures within the human body has long been an important but elusive clinical goal. Contrast agents for ultrasound imaging are gas-filled, encapsulated microbubbles (diameter &#60; 10 &#956;m) that traverse the entire vasculature and enhance signals by up to 30 dB. These microbubbles also produce nonlinear oscillations at frequencies ranging from the subharmonic (half of the transmit frequency) to higher harmonics. The subharmonic amplitude has an inverse linear relationship with the ambient hydrostatic pressure. Here an ultrasound system capable of performing real-time, subharmonic aided pressure estimation (SHAPE) is presented. During ultrasound contrast agent infusion, an algorithm for optimizing acoustic outputs is activated. Following this calibration, subharmonic microbubble signals (i.e., SHAPE) have the highest sensitivity to pressure changes and can be used to noninvasively quantify pressure. The utility of the SHAPE procedure for identifying portal hypertension in the liver is the emphasis here, but the technique has applicability across many clinical scenarios.

Contrast-Enhanced Subharmonic Aided Pressure Estimation SHAPE Using Ultrasound Imaging with a Focus on Identifying Portal Hypertension

A protocol for noninvasively estimating ambient pressures utilizing subharmonic ultrasound imaging of infused contrast microbubbles (following appropriate calibration) is described with examples from human patients with chronic liver disease.

Noninvasive, accurate measurement of pressures within the human body has long been an important but elusive clinical goal. Contrast agents for ultrasound ...

Bioengineering

Rat Model of the Associating Liver Partition and Portal Vein Ligation for Staged Hepatectomy (ALPPS) Procedure

Recent clinical data support an aggressive surgical approach to both primary and metastatic liver tumors. For some indications, like colorectal liver metastases, the amount of liver tissue left behind after liver resection has become the main limiting factor of resectability of large or multiple liver tumors. A minimal amount of functional tissue is required to avoid the severe complication of post-hepatectomy liver failure, which has high morbidity and mortality. Inducing liver growth of the prospective remnant prior to resection has become more established in liver surgery, either in the form of portal vein embolization by interventional radiologists or in the form of portal vein ligation several weeks prior to resection. Recently, it was shown that liver regeneration is more extensive and rapid, when the parenchymal transection is added to portal vein ligation in a first stage and then, after only one week of waiting, resection performed in a second stage (Associating Liver Partition and Portal vein ligation for Staged hepatectomy = ALPPS). ALPPS has rapidly become popular across the world, but has been criticized for its high perioperative mortality. The mechanism of accelerated and extensive growth induced by this procedure has not been well understood. Animal models have been developed to explore both the physiological and molecular mechanisms of accelerated liver regeneration in ALPPS. This protocol presents a rat model that allows mechanistic exploration of accelerated regeneration.

Rat Model of the Associating Liver Partition and Portal Vein Ligation for Staged Hepatectomy ALPPS Procedure

Inducing rapid liver hypertrophy using Associating Liver Partition and Portal vein ligation for a Staged hepatectomy (ALPPS) has been proposed for resection of borderline resectable liver tumors. This model may elucidate mechanisms involved in rapid hypertrophy and allows testing of drugs that promote or block the acceleration of regeneration.

Recent clinical data support an aggressive surgical approach to both primary and metastatic liver tumors. For some indications, like colorectal liver ...

Invasive Hemodynamic Characterization of the Portal-hypertensive Syndrome in Cirrhotic Rats

This is a detailed protocol describing invasive hemodynamic measurements in cirrhotic rats for the characterization of portal hypertensive syndrome. Portal hypertension (PHT) due to cirrhosis is responsible for the most severe complications in patients with liver disease. The full picture of the portal hypertensive syndrome is characterized by increased portal pressure (PP) due to the increased intrahepatic vascular resistance (IHVR), hyperdynamic circulation, and increased splanchnic blood flow. Progressive splanchnic arterial vasodilation and increased cardiac output with elevated heart rate (HR) but low arterial pressure characterizes the portal hypertensive syndrome.
Novel therapies are currently being developed that aim to decrease PP by either targeting IHVR or increased splanchnic blood flow &#8212; but side effects on systemic hemodynamics may occur. Thus, a detailed characterization of portal venous, splanchnic, and systemic hemodynamic parameters, including measurement of PP, portal venous blood flow (PVBF), mesenteric arterial blood flow, mean arterial pressure (MAP), and HR is needed for preclinical evaluation of the efficacy of novel treatments for PHT. Our video article provides the reader with a structured protocol for performing invasive hemodynamic measurements in cirrhotic rats. In particular, we describe the catheterization of the femoral artery and the portal vein via an ileocolic vein and the measurement of portal venous and splanchnic blood flow via perivascular Doppler-ultrasound flow probes. Representative results of different rat models of PHT are shown.

Here we describe a detailed protocol for invasive measurements of hemodynamic parameters including portal pressure, splanchnic blood flow, and systemic hemodynamics in order to characterize the portal hypertensive syndrome in rats.

This is a detailed protocol describing invasive hemodynamic measurements in cirrhotic rats for the characterization of portal hypertensive syndrome. ...

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

Plasma coagulation as a form of electrocautery is used in liver surgery for decades to seal the large liver cut surface after major hepatectomy to prevent hemorrhages at a later stage. The exact effects of plasma coagulation on liver tissue are only poorly examined. In our porcine model, the coagulation effects can be examined close to the clinical application. A combined laser Doppler flowmeter and spectrophotometer documents microcirculation changes during coagulation at 8 mm tissue depth noninvasively, providing quantifiable information about hemostasis beyond the subjective clinical impression. The temperature at coagulation site is assessed with an infrared thermometer prior and post coagulation and with a thermographic camera during coagulation, a measurement of the gas beam temperature is not possible due to the upper threshold of the devices. The depth of coagulation is measured microscopically on hematoxylin/eosin stained sections after calibration with an object micrometer and gives an exact information about the power setting-coagulation depth-relation. The sealing effect is examined on the bile ducts as it is not possible for a plasma coagulator to seal larger blood vessels. Burst pressure experiments are carried out on explanted organs to rule out blood pressure related effects.

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

Here we present a protocol to experimentally assess plasma coagulation in liver tissue in vivo. In a porcine model, microcirculation is examined by laser Doppler, coagulation depth is measured histologically, temperature at coagulation site by infrared thermometer and thermographic camera, and duct sealing effect is documented by burst pressure experiments.

Plasma coagulation as a form of electrocautery is used in liver surgery for decades to seal the large liver cut surface after major hepatectomy to prevent ...

3D Angiography for Venous Compliance Assessment in Sheep

Quantifying Inferior Vena Cava Compliance and Distensibility in an In Vivo Ovine Model Using 3D Angiography

Synthetic vascular grafts overcome some challenges of allografts, autografts, and xenografts but are often more rigid and less compliant than the native vessel into which they are implanted. Compliance matching with the native vessel is emerging as a key property for graft success. The current gold standard for assessing vessel compliance involves the vessel's excision and ex vivo biaxial mechanical testing. We developed an in vivo method to assess venous compliance and distensibility that better reflects natural physiology and takes into consideration the impact of a pressure change caused by flowing blood and by any morphologic changes present. 
This method is designed as a survival procedure, facilitating longitudinal studies while potentially reducing the need for animal use. Our method involves injecting a 20 mL/kg saline bolus into the venous vasculature, followed by the acquisition of pre and post bolus 3D angiograms to observe alterations induced by the bolus, concurrently with intravascular pressure measurements in target regions. We are then able to measure the circumference and the cross-sectional area of the vessel pre and post bolus. 
With these data and the intravascular pressure, we are able to calculate the compliance and distensibility with specific equations. This method was used to compare the inferior vena cava's compliance and distensibility in native unoperated sheep to the conduit of sheep implanted with a long-term expanded polytetrafluorethylene (PTFE) graft. The native vessel was found to be more compliant and distensible than the PTFE graft at all measured locations. We conclude that this method safely provides in vivo measurements of vein compliance and distensibility.

Author Spotlight: Improved Imaging for Neovascular Development in Congenital Heart Disease Research

This protocol allows for the in vivo quantification of venous compliance and distensibility using catheterization and 3D angiography as a survival procedure allowing for a variety of potential applications.

Synthetic vascular grafts overcome some challenges of allografts, autografts, and xenografts but are often more rigid and less compliant than the native ...

Novel In Vivo Micro-Computed Tomography Imaging Techniques for Assessing the Progression of Non-Alcoholic Fatty Liver Disease

Non-alcoholic fatty liver disease (NAFLD) is a growing global health issue, and the impact of NAFLD is compounded by the current lack of effective treatments. Considerable limiting factors hindering the timely and accurate diagnosis (including grading) and monitoring of NAFLD, as well as the development of potential therapies, are the current inadequacies in the characterization of the hepatic microenvironment structure and the scoring of the disease stage in a spatiotemporal and non-invasive manner. Using a diet-induced NAFLD mouse model, we investigated the use of in vivo micro-computed tomography (CT) imaging techniques as a non-invasive method to assess the progression stages of NAFLD, focusing predominantly on the hepatic vascular network due to its significant involvement in NAFLD-related hepatic dysregulation. This imaging methodology allows for longitudinal analysis of liver steatosis and functional tissue uptake, as well as the evaluation of the relative blood volume, portal vein diameter, and density of the vascular network. Understanding the adaptations of the hepatic vascular network during NAFLD progression and correlating this with other ways of characterizing the disease progression (steatosis, inflammation, fibrosis) using the proposed method can pave the way toward the establishment of new, more efficient, and reproducible approaches for NAFLD research in mice. This protocol is also expected to upgrade the value of preclinical animal models for investigating&#160;the development of novel therapies against disease progression.

Novel In Vivo Micro-Computed Tomography Imaging Techniques for Assessing the Progression of Non-Alcoholic Fatty Liver Disease

Using a diet-induced non-alcoholic fatty liver disease (NAFLD) mouse model, we describe the use of novel in vivo micro-computed tomography imaging techniques as a non-invasive method to assess the progression stages of NAFLD, focusing predominantly on the hepatic vascular network due to its significant involvement in NAFLD-related hepatic dysregulation.

Non-alcoholic fatty liver disease (NAFLD) is a growing global health issue, and the impact of NAFLD is compounded by the current lack of effective ...

Biology

Laparoscopic S7 Hepatectomy with Positive Fluorescence Staining

Laparoscopic liver resection for tumors located in the S7 segment of the liver typically adopts a traditional surgical approach. The main goal of this procedure is to accurately dissect the liver pedicle of the S7 segment. Dissecting the hepatic pedicle of the S7 segment along the hepatic hilum requires a relatively long path within the liver, which increases the risk of losing orientation and potentially injuring the adjacent hepatic pedicle of the S5 and S6 segments, thereby compromising the liver resection plane. A positive staining method was used to directly puncture the corresponding portal vein under ultrasound guidance (commonly for S7 and S8 segments) to specifically color the target liver segment, thereby avoiding extensive resection of the liver parenchyma, and reducing damage to the surrounding healthy liver tissue. However, the positive staining method requires a specific foundation in intraoperative procedures, which can be challenging for surgeons and has a certain learning curve. Currently, technologies such as three-dimensional reconstruction territory analysis, intraoperative ultrasound, and indocyanine green fluorescence imaging are popular and commonly used in laparoscopic liver resection. In this protocol, under laparoscopic ultrasound guidance, the tumor basin was punctured through the visceral and diaphragmatic surfaces of the liver to stain segment S7. Laparoscopic resection of segment S7 within the portal territory anatomic liver was successfully performed, further confirming the feasibility and advantages of positive fluorescence staining in laparoscopic liver resection at this stage.

The protocol demonstrates that, under laparoscopic ultrasound guidance, the S7 segment of the liver can be successfully stained by puncturing the visceral and diaphragmatic branches of the tumor, facilitating an anatomical hepatectomy of the S7 segment.

Laparoscopic liver resection for tumors located in the S7 segment of the liver typically adopts a traditional surgical approach. The main goal of this ...

Measurement of the Hepatic Venous Pressure Gradient and Transjugular Liver Biopsy

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