The study was approved by the Ethics Committee of the Second Affiliated Hospital of Shantou University Medical College (Shantou, China).
1. Preparations
<ol>
	<li>Apply strict exclusion criteria to select patients and obtain signed informed consent forms12 (Table 1).</li>
	<li>Apply the following admission criteria:
	<ol>
		<li>Select patients who voluntarily received the two-step PTCSL treatment.</li>
		<li>Select those patients who were confirmed to have hepatic and extrahepatic bile duct stones by adjuvant examination.</li>
		<li>Select patients who had undergone previous complicated abdominal surgery, such as partial gastrectomy combined with biliary-intestinal anastomosis or Billroth II reconstruction, or who had anatomical abnormalities and were unable to undergo an endoscopic retrograde cholangiao pancreatography (ERCP) examination.</li>
		<li>Select patients with liver function preoperatively assessed as Child-Pugh A or B grades.</li>
		<li>Select patients with normal function of the heart, lung, and other organs.</li>
	</ol>
	</li>
	<li>Ensure the availability of the laparoscopic imaging system, images of the pneumoperitoneum, Wolf nephroscope, adjustable pressure pump, and hydroelectric lithotripter.</li>
</ol>
2. Procedure
<ol>
	<li>Percutaneous transhepatic cholangial drainage (PTCD)
	<ol>
		<li>Insert an indwelling 8F biliary drainage tube 1 week before the operation to drain and reduce the swelling of bile duct.</li>
		<li>Wait for the sinus tract to become stable at 1 week after PTCD. 
		&#8203;NOTE: The puncture methodology must be chosen according to the particular area of bile conduit calculi when operating. For example, the left hepatic duct approach is performed under the xiphoid process and abdominal wall of the lower edge of the right costal arch.</li>
	</ol>
	</li>
</ol>
3. Percutaneous transhepatic choledochoscopic lithotomy (PTCSL)
<ol>
	<li>Before administering general anesthesia, instruct the patient to lie flat on the operating table. Following general anesthesia, disinfect the surgical site 3x with iodophor.</li>
	<li>Inject 20 mL of saline into the bile duct along the drainage tube; pull the PTCD tube out after a guide wire is inserted into it.</li>
	<li>For placement of the fascia dilator and sheath, expand the sinus using a 16F expander and join a 16F belt dilator so that the 16F sheath can reach the bile duct where the stone might have been placed.</li>
	<li>After locating the stones, use clamping forceps to both break and remove the stones. For hard stones, use a hydraulic lithotripter to break the stone. Wash out small stones and powdered stones using an adjustable pressure pump. At the end of the procedure, leave a drainage tube inside the patient for 1 week (Figure 1).</li>
</ol>

<ol>
	<li>Shu, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Robotic-assisted+laparoscopic+surgery+for+complex+hepatolithiasis:+a+propensity+score+matching+analysis.">Robotic-assisted laparoscopic surgery for complex hepatolithiasis: a propensity score matching analysis.</a> Surgical Endoscopy. 33 (8), 2539-2547 (2019).</li><li>Korkes, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+treatment+of+complex+biliary+stone+disease+using+endourological+technique+and+literature+review.">Percutaneous treatment of complex biliary stone disease using endourological technique and literature review.</a> Einstein. 13 (4), 611-615 (2015).</li><li>Gamal, E. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+video+choledochoscopic+treatment+of+retained+biliary+stones+via+dilated+t-tube+tract.">Percutaneous video choledochoscopic treatment of retained biliary stones via dilated t-tube tract.</a> Surgical Endoscopy. 15 (5), 473-476 (2001).</li><li>Khalid, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+endoscopic+biliary+exploration+in+complex+biliary+stone+disease:+Case+series+study.">Percutaneous endoscopic biliary exploration in complex biliary stone disease: Case series study.</a> International Journal of Surgery. 24, 73-78 (2020).</li><li>Kim, J. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+transhepatic+cholangioscopic+treatment+of+patients+with+benign+bilio-enteric+anastomotic+strictures.">Percutaneous transhepatic cholangioscopic treatment of patients with benign bilio-enteric anastomotic strictures.</a> Gastrointestinal Endoscopy. 58 (5), 733-738 (2003).</li><li>Hwang, J. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Risk+factors+for+recurrent+cholangitis+after+initial+hepatolithiasis+treatment.">Risk factors for recurrent cholangitis after initial hepatolithiasis treatment.</a> Journal of Clinical Gastroenterology. 38 (4), 364-367 (2004).</li><li>Bonnel, D. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Common+bile+duct+and+intrahepatic+stones:+results+of+transhepatic+electrohydraulic+lithotripsy+in+50+patients.">Common bile duct and intrahepatic stones: results of transhepatic electrohydraulic lithotripsy in 50 patients.</a> Radiology. 180 (2), 345-348 (1991).</li><li>Kondo, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+clinicopathologic+study+of+primary+cholesterol+hepatolithiasis.">A clinicopathologic study of primary cholesterol hepatolithiasis.</a> Hepato-gastroenterology. 42 (5), 478-486 (1995).</li><li>Williams, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Updated+guideline+on+the+management+of+common+bile+duct+stones+(CBDS).">Updated guideline on the management of common bile duct stones (CBDS).</a> Gut. 66 (5), 765-782 (2017).</li><li>Makuuchi, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasonically+guided+cholangiography+and+bile+drainage.">Ultrasonically guided cholangiography and bile drainage.</a> Ultrasound in Medicine &amp; Biology. 10 (5), 617-623 (1984).</li><li>Jeng, K. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+transhepatic+cholangioscopy+in+the+treatment+of+complicated+intrahepatic+biliary+strictures+and+hepatolithiasis+with+internal+metallic+stent.">Percutaneous transhepatic cholangioscopy in the treatment of complicated intrahepatic biliary strictures and hepatolithiasis with internal metallic stent.</a> Surgical Laparoscopy. 10 (5), 278-283 (2000).</li><li>Zhuo, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+transhepatic+choledochoscopic+lithotomy+(PTCSL)+is+effective+for+the+treatment+of+intrahepatic+and+extrahepatic+choledocholithiasis.">Percutaneous transhepatic choledochoscopic lithotomy (PTCSL) is effective for the treatment of intrahepatic and extrahepatic choledocholithiasis.</a> Surgical Laparoscopy, Endoscopy &amp; Percutaneous Techniques. 31 (3), 326-330 (2020).</li><li>Hazey, J. W., McCreary, M., Guy, G., Melvin, W. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficacy+of+percutaneous+treatment+of+biliary+tract+calculi+using+the+holmium:+YAG+laser.">Efficacy of percutaneous treatment of biliary tract calculi using the holmium: YAG laser.</a> Surgical Endoscopy. 21 (7), 1180-1183 (2007).</li><li>Ozturk, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+surgical+technique+applied+with+urological+instruments+in+bilobar+multiple+hepatolithiasis:+Ultra-mini+percutaneous+hepatolithotomy.">New surgical technique applied with urological instruments in bilobar multiple hepatolithiasis: Ultra-mini percutaneous hepatolithotomy.</a> Turkish Journal of Urology. 43 (3), 371-377 (2017).</li><li>Ozcan, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+transhepatic+removal+of+bile+duct+stones:+results+of+261+patients.">Percutaneous transhepatic removal of bile duct stones: results of 261 patients.</a> Cardiovascular and Interventional Radiology. 35 (3), 621-627 (2012).</li><li>Kint, J. 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Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+trans-hepatic+cholangioscopic+lithotomy+for+hepatolithiasis:+long-term+results.">Percutaneous trans-hepatic cholangioscopic lithotomy for hepatolithiasis: long-term results.</a> Chang Gung Medical Journal. 17 (1), 149 (1994).</li><li>Cheon, Y. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+long-term+results+and+recurrent+factors+after+operative+and+nonoperative+treatment+for+hepatolithiasis.">Evaluation of long-term results and recurrent factors after operative and nonoperative treatment for hepatolithiasis.</a> Surgery. 146 (5), 843-853 (2009).</li><li>Tsutsumi, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparative+evaluation+of+treatment+methods+for+bile+duct+stones+after+hepaticojejunostomy+between+percutaneous+transhepatic+cholangioscopy+and+peroral,+short+double-balloon+enteroscopy.">A comparative evaluation of treatment methods for bile duct stones after hepaticojejunostomy between percutaneous transhepatic cholangioscopy and peroral, short double-balloon enteroscopy.</a> Therapeutic Advances in Gastroenterology. 10 (1), 54-67 (2017).</li></ol>

The authors have no conflicts of interest to disclose.

Intrahepatic and extrahepatic choledocholithiasis contribute to biliary obstruction, and the associated cholangitis could lead to gram-negative endotoxemia with clinical manifestations of septic shock or multiorgan dysfunction. Thus, biliary stones represent a complex disease that can be challenging for surgeons, especially in cases of patients who have had previous complex abdominal surgeries or patients with abnormal anatomical structure. PTCSL is being widely used around the world, but there remains a need for innovat...

Intrahepatic and extrahepatic choledocholithiasis is a complex medical problem faced by surgeons and is very common in East Asian countries1. At present, surgical intervention is required for most choledocholithiasis, while the main treatments for bile duct stones include open surgery, percutaneous transhepatic choledochoscopic lithotomy (PTCSL), laparoscopic lithotomy2, percutaneous endoscopic biliary exploration3,4, and endoscopic retrograde cholangiopancreatography. However, patients with bilioenteric anastomosis or Billroth II reconstruction, with an abnormal anatomical structure such as a duodenal periampullary diverticulum, which makes the biliary duct difficult to access through an endoscopic approach5,6,7,8, or who have had previous complex abdominal surgeries, including partial gastrectomy, could not undergo endoscopic therapy or laparoscopic lithotomy. Endoscopic therapy may also not be suitable for hepatolithiasis with large and impacted stones. Therefore, PTCSL, in general, may be a better choice9.
The traditional PTCS approach follows such a procedure to expand the sinus gradually and perform percutaneous transhepatic biliary drainage10,11, which requires a relatively long hospitalization cycle, usually between 2 and 3 weeks, leading to the potential for biliary fistula, biliary tract infection, bleeding, and peritonitis, with multiple expanding sheath sizes. The procedure is composed of two parts. In the first step, the target bile duct is punctured, and the guide wire is inserted while observing the bile outflow. Then, a dilator is used to slowly expand the channel along the guide wire. After the channel is successfully established, the dilator is pulled out, a drain is placed into the target bile duct, and the guide wire is pulled out.
The percutaneous liver puncture and drainage (PTCD) is complete if the bile drainage is unobstructed and the drain is fixed without blood outflow. The sinus tract becomes stable 1 week after PTCD, when it can then be expanded. The guide wire is placed into the drain tube and expanded through the sinus tract along the guide wire with a 10F dilator, and finally placed into a 10F drain. The sinus tract is expanded once a week after surgery, with a 2F size increase each time, so that the sinus tract can be expanded to 16F at 4 weeks after PTCD. In the second step, after expansion of the sinus tract to 16F, the surgery to remove the stone is performed using a biliary fiber lens. Therefore, the traditional PTCSL described above requires a treatment cycle and contributes to high economic pressure for the patients. Moreover, low stone extraction efficiency and repeated and multiple surgical procedures for stone removal are also inevitable disadvantages of traditional PTCS, which limits the use of this technology.
From January 2013, we began to apply PTCS for the minimally invasive treatment of patients with complex intrahepatic and extrahepatic choledocholithiasis in our hospital but modified some key steps of traditional PTCS. In the first step, the surgery was performed at 1 week after PTCD, just when the sinus tract became stable, and the sinus was expanded to 16F directly, using a 16F expander which greatly reduced the preoperative preparation time. In the second step, different from the traditional technology, a Wolf nephroscope was used instead of a traditional choledochoscope to perform the operation, which we refer to as the &#34;two-step PTCSL method.&#34; To determine whether the PTCSL procedure was safe, minimally invasive, and efficient, this study retrospectively reviewed 81 patients with intrahepatic and extrahepatic choledocholithiasis treated by the two-step PTCSL method from January 2013 to January 2020.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.9% Normal saline solution</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>16-F series fascia dilator</td>
			<td></td>
			<td>type PLVW, Cook Medical</td>
			<td></td>
		</tr>
		<tr>
			<td>adjustable pressure pump</td>
			<td></td>
			<td>type APL; Guangzhou Jielun Medical Equipment Co. , Ltd, Guangdong Province, China</td>
			<td></td>
		</tr>
		<tr>
			<td>biliary balloon dilator</td>
			<td></td>
			<td>ATB advance; Cook Medical</td>
			<td></td>
		</tr>
		<tr>
			<td>blade</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>camera system</td>
			<td></td>
			<td>1088i HD Camera Control Unit, PAL 220 V; Stryker Corporation</td>
			<td></td>
		</tr>
		<tr>
			<td>Cavity mirror protective sleeve</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>clamp</td>
			<td></td>
			<td>5Fr, Richard Wolf GmbH, Germany</td>
			<td></td>
		</tr>
		<tr>
			<td>color ultrasound machine</td>
			<td></td>
			<td>DC-N2S, Mindray</td>
			<td></td>
		</tr>
		<tr>
			<td>Cook net basket</td>
			<td></td>
			<td>NGE-017115-MB, COOK Medical Inc. , Bloomington, USA</td>
			<td></td>
		</tr>
		<tr>
			<td>drainage pack</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>expander</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>guide wire</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>hydroelectric lithotripter</td>
			<td></td>
			<td>Aymed Medical, <img alt="Equation 1" src="/files/ftp_upload/63206/63206eq01.jpg" style="margin: auto;" />stanbul, Turkey</td>
			<td></td>
		</tr>
		<tr>
			<td>&#160;Iodophor</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>&#160;Kidney basin</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>light source</td>
			<td></td>
			<td>Voice-control compatible X8000 Xenon Light Source of 300 W;Stryker Corporation, MI</td>
			<td></td>
		</tr>
		<tr>
			<td>Medical syringe</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>puncture needle</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>suction apparatus</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>surgical gauze</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>trocar</td>
			<td></td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Wolf nephroscope</td>
			<td></td>
			<td>12-degree Ultra-Wide-Angle Ureteroreno-scope; Richard Wolf GmbH, Germany</td>
			<td></td>
		</tr>
		<tr>
			<td>zebra guide wire</td>
			<td></td>
			<td>type HAW; Cook Medical, Bloomington, IN</td>
			<td></td>
		</tr>
	</tbody>
</table>

a two-step method for percutaneous transhepatic choledochoscopic lithotomy

Intrahepatic and extrahepatic choledocholithiasis is a challenge in the field of biliary surgery. We present our experience using a two-step percutaneous transhepatic choledochoscopic lithotomy (PTCSL) procedure to treat challenging biliary stones. We retrospectively reviewed 81 patients with intrahepatic and extrahepatic choledocholithiasis treated using this two-step PTCSL from January 2013 to January 2020, including 40 males and 41 females, with an average age of 66 years. In contrast to traditional percutaneous transhepatic cholangioscopy (PTCS), a channel was established directly through a 16F Amplatz sheath, and the stone in the channel was removed with the aid of a nephroscope. The clinical efficacy and complications of all patients were analyzed. Eighty-one patients (81/81, 100%) had their biliary stones successfully removed; 62/81 patients (76.5%) had biliary stones completely removed after the first operation; 17/81 patients (21%) underwent a second operation; 2/81 patients (2.5%) needed a third operation to completely remove the stones. The incidence of severe bleeding during the operation was 0%, and there were no deaths. The use of the two-step PTCSL method is safe and efficacious, and contributes to a better prognosis of intrahepatic and extrahepatic choledocholithiasis.

All patients had their biliary tract stones cleared successfully, with 58 patients (58/81, 72%) having bile duct stones completely removed after the first operation, 18 patients (18/81, 22%) requiring a second operation, and five patients (5/81, 6%) undergoing a third operation to completely remove the stones. Three patients had recurrent intrahepatic cholelithiasis in the 5th and 6th postoperative years. There were 56 patients (56/81, 68.7%) who underwent percutaneous liver surgery, as well as 25 p...

Watch this Scientific Journal Video about A Two-Step Method for Percutaneous Transhepatic Choledochoscopic Lithotomy at JoVE.com

A Two-Step Method for Percutaneous Transhepatic Choledochoscopic Lithotomy

This "two-step method" significantly improved the success rate of percutaneous transhepatic choledochoscopy and achieved a better prognosis of intrahepatic and extrahepatic choledocholithiasis.

Intrahepatic and extrahepatic choledocholithiasis is a challenge in the field of biliary surgery. We present our experience using a two-step percutaneous ...

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2.2K Views.  Shantou University Medical College. This "two-step method" significantly improved the success rate of percutaneous transhepatic choledochoscopy and achieved a better prognosis of intrahepatic and extrahepatic choledocholithiasis.

Video: A Two-Step Method for Percutaneous Transhepatic Choledochoscopic Lithotomy

A Method for Murine Islet Isolation and Subcapsular Kidney Transplantation

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize their blood glucose levels, islet transplantation has been proposed to be a potential treatment for type 1 diabetes 1,2. More recently, advances in human islet transplantation have further strengthened this view 1,3. However, two major limitations prevent islet transplantation from being a widespread clinical reality: (a) the requirement for large numbers of islets per patient, which severely reduces the number of potential recipients, and (b) the need for heavy immunosuppression, which significantly affects the pediatric population of patients due to their vulnerability to long-term immunosuppression. Strategies that can overcome these limitations have the potential to enhance the therapeutic utility of islet transplantation.
Islet transplantation under the mouse kidney capsule is a widely accepted model to investigate various strategies to improve islet transplantation. This experiment requires the isolation of high quality islets and implantation of islets to the diabetic recipients. Both procedures require surgical steps that can be better demonstrated by video than by text. Here, we document the detailed steps for these procedures by both video and written protocol. We also briefly discuss different transplantation models: syngeneic, allogeneic, syngeneic autoimmune, and allogeneic autoimmune.

Transplantation of isolated islets has been proposed to be a potential treatment for type 1 diabetes. Here we describe a method to isolate islets from mouse pancreata and transplant them to the subcapsular space of the kidney.

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize ...

A Modified Method for Heterotopic Mouse Heart Transplantion

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents available. A difficulty is presented due to the small size of the animal and the considerable technical challenges of the microsurgery involved in heart transplantation. In particular, a high rate of technical failure early after transplantation may result from recipient death and post-operative complications such as hind limb paralysis or a non-beating heart. Here, the complete technique for heterotopic mouse heart transplantation is demonstrated, involving harvesting the donor heart and its subsequent implantation into a recipient mouse. The donor heart is harvested immediately following in situ perfusion with cold heparinized saline and transection of the ascending aorta and pulmonary artery. The recipient operation involves preparation of the abdominal aorta and inferior vena cava (IVC), followed by end-to-side anastomosis of the donor aorta with the recipient aorta using a single running 10-0 microsuture and a similar anastomosis of the donor pulmonary artery with the recipient IVC. Following the operation the animal is injected with 0.6 ml normal saline subcutaneously and allowed to recover on a 37&#160;&#176;C heating pad. The results from 227 mouse heart transplants are summarized with a success rate at 48 hr of 86.8%. Of the 13.2% failures within 48 hr, 5 (2.2%) experienced hind limb paralysis, 10 (4.4%) had a non-beating heart due to graft ischemic injury and/or thrombosis, while 15 (6.6%) died within 48 hr.

A technique is demonstrated for the microsurgical procedure for heterotopic transplantation of hearts in mice, including simplified methods for donor harvesting and recipient vessel anastomosis.

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents ...

Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining samples of human Vastus lateralis, isolating mitochondria from minimal amounts of skeletal muscle tissue, and plate based respirometric profiling using an extracellular flux (XF) analyzer. Comparison of respirometric profiles obtained using 1.0, 2.5 and 5.0 &#956;g of mitochondria indicate that 1.0 &#956;g is sufficient to measure respiration and that 5.0 &#956;g provides most consistent results based on comparison of standard errors. Western blot analysis of isolated mitochondria for mitochondrial marker COX IV and non-mitochondrial tissue marker GAPDH indicate that there is limited non-mitochondrial contamination using this protocol. The ability to study mitochondrial respirometry in as little as 20 mg of muscle tissue allows users to utilize individual biopsies for multiple study endpoints in clinical research projects.

Methods for biopsy of Vastus lateralis, preparation of purified mitochondria, and respirometric profiling are described. The use of small muscle volume makes this technique suitable for clinical research applications.

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining ...

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.

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

Percutaneous Contrast Echocardiography-guided Intramyocardial Injection and Cell Delivery in a Large Preclinical Model

Cell and gene therapy are exciting and promising strategies for the purpose of cardiac regeneration in the setting of heart failure with reduced ejection fraction (HFrEF). Before they can be considered for use, and implemented in humans, extensive preclinical studies are required in large animal models to evaluate the safety, efficacy, and fate of the injectate (e.g., stem cells) once delivered into the myocardium. Small rodent models offer advantages (e.g., cost effectiveness, amenability for genetic manipulation); however, given inherent limitations of these models, the findings in these rarely translate into the clinic. Conversely, large animal models such as rabbits, have advantages (e.g., similar cardiac electrophysiology compared to humans and other large animals), whilst retaining a good cost-effective balance. Here, we demonstrate how to perform a percutaneous contrast echocardiography-guided intramyocardial injection (IMI) technique, which is minimally invasive, safe, well tolerated, and very effective in the targeted delivery of injectates, including cells, into several locations within the myocardium of a rabbit model. For the implementation of this technique, we also have taken advantage of a widely available clinical echocardiography system. After putting in practice the protocol described here, a researcher with basic ultrasound knowledge will become competent in the performance of this versatile and minimally invasive technique for routine use in experiments, aimed at hypothesis testing of the capabilities of cardiac regenerative therapeutics in the rabbit model. Once competency is achieved, the whole procedure can be performed within 25 min after anaesthetizing the rabbit.

Novel therapeutic strategies in cardiac regenerative medicine require extensive and detailed studies in large preclinical animal models before they can be considered for use in humans. Here, we demonstrate a percutaneous contrast echocardiography-guided intramyocardial injection technique in rabbits, which is valuable for hypothesis testing the efficacy of such novel therapies.

Cell and gene therapy are exciting and promising strategies for the purpose of cardiac regeneration in the setting of heart failure with reduced ejection ...

Three-Dimensional Printing Guide Template Assisted Percutaneous Vertebroplasty (PVP)

Percutaneous vertebroplasty (PVP) is considered an effective treatment for the back pain caused by osteoporotic vertebral compression fracture. The accuracy of PVP mainly depends on the surgeons&#39; experience and multiple fluoroscopes during a traditional procedure. Puncture related complications were reported all over the world. To make the surgical procedure more precise and decrease the rate of puncture-related complications, our team applied a three-dimensional printing guide template to PVP to modify the traditional procedure. This protocol introduces how to model target vertebrae DICOM imaging data into three-dimensions in the software, how to simulate operation in this 3-D model, and how to use all of the surgical data to reconstruct a patient specific template for application. Using this template, surgeons can identify suitable puncture points accurately to improve the accuracy of the operation. The whole protocol includes: 1) diagnosis of the osteoporotic vertebral compression fracture; 2) acquisition of CT imaging of the target vertebra; 3) simulation of the operation in the software; 4) design and fabrication of the 3-D printing guide template; and 5) application of the template into an operation procedure.

Three-Dimensional Printing Guide Template Assisted Percutaneous Vertebroplasty PVP

Herein, we present a three-dimensional printing guide template for percutaneous vertebraplasty. A patient with a T11 vertebral compression fracture was selected as a case study.

Percutaneous vertebroplasty (PVP) is considered an effective treatment for the back pain caused by osteoporotic vertebral compression fracture. The ...

A Revised Method for Inducing Secondary Lymphedema in the Hindlimb of Mice

Animal models are of paramount importance in the research of lymphedema in order to understand the pathophysiology of the disease but also to explore potential treatment options. This mouse model allows researchers to induce significant lymphedema lasting at least 8 weeks. Lymphedema is induced using a combination of fractioned radiotherapy and surgical ablation of lymphatics. This model requires that the mice get a dose of 10 Gray (Gy) radiation before and after surgery. The surgical part of the model involves ligation of three lymph vessels and extraction of two lymph nodes from the mouse hindlimb. Having access to microsurgical tools and a microscope is essential, due to the small anatomical structures of mice. The advantage of this model is that it results in statistically significant lymphedema, which provides a good basis for evaluating different treatment options. It is also a great and easily available option for microsurgical training. The limitation of this model is that the procedure can be time consuming, especially if not practiced in advance. The model results in objectively quantifiable lymphedema in mice, without causing severe morbidity and has been tested in three separate projects.

This animal model enables researchers to induce statistically significant secondary lymphedema in the hindlimb of mice, lasting at least 8 weeks. The model can be used to study the pathophysiology of lymphedema and to investigate novel treatment options.

Animal models are of paramount importance in the research of lymphedema in order to understand the pathophysiology of the disease but also to explore ...

A Simplified Stepwise Approach to Echo Guidance during Percutaneous Mitral Valve Repair

Percutaneous transcatheter edge-to-edge reconstruction of the mitral valve is a safe and well-established therapy for severe symptomatic mitral regurgitation in patients with high surgical risk. Echocardiographic guidance in addition to fluoroscopy is the gold standard and should be performed using a standardized technique.
This article lays out our reproducible step by step echocardiographic guide including views, measurements as well as highlighting possible difficulties that may arise during the procedure.
This article provides detailed and chronological echocardiographic views for each step of the procedure, especially preferences between 2D and 3D imaging. If needed, pulse wave, continuous wave and color doppler measurements are described. Furthermore, as there are no official recommendations for the quantification of mitral regurgitation during the percutaneous edge-to-edge-repair procedure, advice is also included for echocardiographic quantification after grasping the mitral leaflets and after device deployment. In addition, the article deals with important echocardiographic views to prevent and deal with possible complications during the procedure.
Echocardiographic guidance during transcatheter mitral valve repair is mandatory. A structured approach improves the collaboration between interventionist and imager and is indispensable for a safe and effective procedure.

This protocol presents in detail how to perform real-time echocardiographic guidance during transcatheter mitral valve repair. The fundamental views and the necessary measurements are described for each stage of the procedure.

Percutaneous transcatheter edge-to-edge reconstruction of the mitral valve is a safe and well-established therapy for severe symptomatic mitral ...

Use of a Percutaneous Ventricular Assist Device/Left Atrium to Femoral Artery Bypass System for Cardiogenic Shock

The left atrial to femoral artery bypass (LAFAB) system is a mechanical circulatory support (MCS) device used in cardiogenic shock (CS) that bypasses the left ventricle by draining blood from the left atrium (LA) and returning it to the systemic arterial circulation via the femoral artery. It can provide flows ranging from 2.5-5 L/min depending on the size of the cannula. Here, we discuss the mechanism of action of LAFAB, available clinical data, indications for its use in cardiogenic shock, steps of implantation, post-procedural care, and complications associated with the use of this device and their management.
We also provide a brief video of the procedural component of device therapy, including the pre-placement preparation, percutaneous placement of the device via transseptal puncture under echocardiographic guidance and the post-operative management of device parameters.

The following article describes the stepwise procedure for placement of a device (e.g., Tandemheart) in cardiogenic shock (CS) that is a percutaneous left ventricular assist device (pLVAD) and a left atrial to femoral artery bypass (LAFAB) system that bypasses and supports the left ventricle (LV) in CS.

The left atrial to femoral artery bypass (LAFAB) system is a mechanical circulatory support (MCS) device used in cardiogenic shock (CS) that bypasses the ...

Myocardial Infarction by Percutaneous Embolization Coil Deployment in a Swine Model

Myocardial infarction (MI) is the leading cause of mortality worldwide. Despite the use of evidence-based treatments, including coronary revascularization and cardiovascular drugs, a significant proportion of patients develop pathological left-ventricular remodeling and progressive heart failure following MI. Therefore, new therapeutic options, such as cellular and gene therapies, among others, have been developed to repair and regenerate injured myocardium. In this context, animal models of MI are crucial in exploring the safety and efficacy of these experimental therapies before clinical translation. Large animal models such as swine are preferred over smaller ones due to the high similarity of swine and human hearts in terms of coronary artery anatomy, cardiac kinetics, and the post-MI healing process. Here, we aimed to describe an MI model in pig by permanent coil deployment. Briefly, it comprises a percutaneous selective coronary artery cannulation through retrograde femoral access. Following coronary angiography, the coil is deployed at the target branch under fluoroscopic guidance. Finally, complete occlusion is confirmed by repeated coronary angiography. This approach is feasible, highly reproducible, and emulates the pathogenesis of human non-revascularized MI, avoiding the traditional open-chest surgery and the subsequent postoperative inflammation. Depending on the time of follow-up, the technique is suitable for acute, sub-acute, or chronic MI models.

Myocardial infarction (MI) animal models that emulate the natural process of the disease in humans are crucial to understanding pathophysiological mechanisms and testing the safety and efficacy of new emergent therapies. Here, we describe an MI swine model created by deploying a percutaneous embolization coil.

Myocardial infarction (MI) is the leading cause of mortality worldwide. Despite the use of evidence-based treatments, including coronary revascularization ...