Research reported in this publication was supported by the National Institute of General Medical Sciences of the National Institutes of Health under award number 5U54GM104942-04 (BAB).
The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

All aspects of this protocol fall within our institutions ethical guidelines of the human research ethics committee
1. Pre-operative planning
<ol>
	<li>Evaluate the patient pre-operatively. 
	NOTE: Patients present generally with vague abdominal complains and will be diagnosed primarily by imaging studies. This patient is a 65 year old Caucasian female presented with vague abdominal pain and underwent several CT imaging studies, eventually resulting in a diagnosis of pancreatic mass involving the body of the pancreas and abutting the common hepatic and splenic arteries (Figure 1).</li>
	<li>Preoperative Imaging and Biopsy
	<ol>
		<li>Proceed initially with cross-sectional imaging to diagnose the mass and identify anatomical relationships, arterial/venous involvement, and any aberrant arterial and venous anatomy. Once identified, proceed with biopsy for tissue diagnosis, if the mass is accessible. This patient&#39;s lesion was biopsied during EUS and confirmed as pancreatic ductal adenocarcinoma.</li>
		<li>Carefully note and consider any aberrant arterial anatomy or portosplenic confluence aberrancies and involvement of additional structure outside of the celiac axis prior to pursuing surgical resection.</li>
	</ol>
	</li>
	<li>Consider preoperative therapy. 
	NOTE: Many different preoperative treatment protocols are available for treatment of locally advanced pancreatic cancer. Patient tolerance and standards of practice at various institutions can guide therapy. In the case of our patient, she was initially started on FOLFIRINOX, but after significant intolerance requiring hospitalization, she was ultimately transitioned to gemcitabine/nab-paclitaxel and completed 6 months of treatment.</li>
	<li>Repeat imaging and serum studies.
	<ol>
		<li>Consider follow up imaging to evaluate treatment response prior to moving forward with resection. Serum studies, such as CA 19-9, will additionally help evaluate treatment response. In our patient, imaging demonstrated a promising treatment response as well as a 94% reduction in her serum CA 19-9 levels. However, she continued to have persistent soft tissue infiltration of her celiac axis on repeat imaging (Figure 2). As a result, she was scheduled for a robotic DP-CAR.</li>
	</ol>
	</li>
</ol>
2. Initial Operative Steps: Diagnostic Laparoscopy and Robot Docking
<ol>
	<li>Begin with a diagnostic laparoscopy to ensure no evidence of metastatic disease.</li>
	<li>Once no metastatic disease is confirmed, place the remainder of the ports and dock the robot from the right side of the operating table. 
	NOTE: Several variations of port placement have been successfully utilized. However, our approach utilizes 4 robotic ports across the upper abdomen, two assistant ports and a liver retractor. A skilled bedside assistant is critical to successful completion of the procedure, utilizing the two assistant ports for operating the bipolar vessel sealing device, suction, and endovascular stapler.</li>
</ol>
3. Robot-Assisted Dissection
<ol>
	<li>Open lesser sac and mobilize stomach along greater curve.
	<ol>
		<li>Following robot docking, proceed with opening the lesser sac with electrocautery and bipolar vessel sealer.</li>
		<li>Completely mobilize the greater curve of the stomach with care to avoid sacrificing the gastroepiploic vessels (Figure 3).</li>
		<li>Cauterize the short gastric arteries with vessel sealing device to the level of the diaphragm. Then retract the stomach and liver edge cephalad to allow for dissection of the porta hepatis.</li>
	</ol>
	</li>
	<li>Identify hepatic arterial anatomy and assess for adequate retrograde flow into proper hepatic artery and liver through the gastroduodenal artery.
	<ol>
		<li>After cephalad retraction of the stomach and liver, identify the porta hepatis. Dissect and isolate the hepatic artery node (Station 8a) for removal and permanent pathological evaluation.</li>
		<li>At this time, carefully dissect and identify the gastroduodenal artery (GDA). Evaluate for adequate retrograde flow through the GDA into the hepatic circulation with intra-operative Doppler ultrasound.</li>
		<li>Identify pulsatile flow in the liver and GDA before and after clamping of the common hepatic artery (Figure 4).</li>
	</ol>
	</li>
	<li>Dissect the inferior border of the pancreas and create a retropancreatic tunnel.
	<ol>
		<li>After identification of adequate retrograde flow through the GDA, dissect along the inferior border of the pancreas. Continue dissection with electrocautery and bipolar vessel sealer to identify the superior mesenteric vein.</li>
		<li>Create a retropancreatic tunnel over the vein with a sufficient margin from the tumor (Figure 5).</li>
	</ol>
	</li>
	<li>Divide the pancreas and continue the retropancreatic dissection to divide the splenic vein and coronary vein.
	<ol>
		<li>Divide the pancreas with an endovascular stapler. Mobilize the inferior and posterior border of the pancreas laterally with electrocautery and bipolar assisted dissection.</li>
		<li>Divide the splenic vein and coronary vein to assist in visualization of arterial anatomy and retraction. Additionally, the inferior mesenteric vein may be ligated if it inserts into the splenic vein.</li>
	</ol>
	</li>
	<li>Divide the hepatic artery and expose the superior mesenteric artery.
	<ol>
		<li>Turn attention back to the hepatic artery and continue dissection to fully delineate the anatomy if required. Divide the hepatic artery proximal the gastroduodenal artery (Figure 6).</li>
		<li>Once the hepatic artery is divided, retract the specimen laterally and continue to mobilize the pancreas to expose the superior mesenteric artery (SMA).</li>
	</ol>
	</li>
	<li>Identify and trace the superior mesenteric artery to its root on the aorta.
	<ol>
		<li>Once the SMA is identified, dissect cephalad along the superior border to trace it back to the origin on the aorta (Figure 7). Continue the dissection cephalad with electrocautery and bipolar cautery through dense connective tissue.</li>
	</ol>
	</li>
	<li>Dissect cephalad from the root of the SMA through dense connective tissue, nerve bundles, and lymphatic tissue until muscle fibers from the diaphgramatic crura are identified.
	<ol>
		<li>During this en bloc mobilization of the tissue overlying the superior mesenteric artery and celiac axis as well as the final lateral dissection, sample lymph node stations 14, 16 and 18.</li>
	</ol>
	</li>
	<li>Ligate the left gastric artery near its origin with an endovascular stapler to maximize collateral blood flow to the stomach.</li>
	<li>Expose and divide the celiac axis.
	<ol>
		<li>Continue retracting the sample to the patient&#39;s left and dissect through the crural muscle fibers until the origin of the celiac axis is visualized (Figure 8). It is of utmost importance to maintain sufficient lateral retraction on the specimen to rotate the celiac axis to the left of the patient. This will facilitate ligation of the celiac trunk by providing a favorable angle for the endovascular stapling device.</li>
	</ol>
	</li>
	<li>Continue retroperitoneal dissection laterally to fully mobilize pancreas and spleen and remove specimen.
	<ol>
		<li>Continue the retroperitoneal dissection laterally with a combination of hook and bipolar cautery. Remove the specimen through the left lower quadrant incision, which can be extended if necessary. A detailed view of the final resection bed vascular anatomy is available in the supplementary figures (Figure 9).</li>
		<li>Complete a final intra-operative ultrasound of the proper hepatic artery continues to demonstrate pulsatile, triphasic flow in the artery and parenchyma of the liver. Evaluate the stomach for external signs of ischemia.</li>
		<li>Once a final inspection is completed, undock the robot, and close the fascia and skin. The procedure is complete.</li>
	</ol>
	</li>
</ol>

<ol>
	<li>American Cancer Society. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=American+Cancer+Society+(2016)+Cancer+Facts+&amp;+Figures.">American Cancer Society (2016) Cancer Facts &amp; Figures.</a> American Cancer Society. , (2016).</li><li>Suker, 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=FOLFIRINOX+for+locally+advanced+pancreatic+cancer:+a+systematic+review+and+patient-level+meta-analysis.">FOLFIRINOX for locally advanced pancreatic cancer: a systematic review and patient-level meta-analysis.</a> Lancet Oncology. 17 (6), 801-810 (2016).</li><li>Faris, J. 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=FOLFIRINOX+in+locally+advanced+pancreatic+cancer:+the+Massachusetts+General+Hospital+Cancer+Center+experience.">FOLFIRINOX in locally advanced pancreatic cancer: the Massachusetts General Hospital Cancer Center experience.</a> Oncologist. 18 (5), 543-548 (2013).</li><li>Ueno, 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=Randomized+phase+III+study+of+gemcitabine+plus+S-1,+S-1+alone,+or+gemcitabine+alone+in+patients+with+locally+advanced+and+metastatic+pancreatic+cancer+in+Japan+and+Taiwan:+GEST+study.">Randomized phase III study of gemcitabine plus S-1, S-1 alone, or gemcitabine alone in patients with locally advanced and metastatic pancreatic cancer in Japan and Taiwan: GEST study.</a> Journal of Clinical Oncology. 31 (13), 1640-1648 (2013).</li><li>Hirono, 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=Treatment+Strategy+for+Borderline+Resectable+Pancreatic+Cancer+With+Radiographic+Artery+Involvement.">Treatment Strategy for Borderline Resectable Pancreatic Cancer With Radiographic Artery Involvement.</a> Pancreas. 45 (10), 1438-1446 (2016).</li><li>Schmocker, R. 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=An+Aggressive+Approach+to+Locally+Confined+Pancreatic+Cancer:+Defining+Surgical+and+Oncologic+Outcomes+Unique+to+Pancreatectomy+with+Celiac+Axis+Resection+(DP-CAR).">An Aggressive Approach to Locally Confined Pancreatic Cancer: Defining Surgical and Oncologic Outcomes Unique to Pancreatectomy with Celiac Axis Resection (DP-CAR).</a> Annals of Surgical Oncology. , (2020).</li><li>Klompmaker, 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=E-AHPBA+DP-CAR+study+group.+Outcomes+and+Risk+Score+for+Distal+Pancreatectomy+with+Celiac+Axis+Resection+(DP-CAR),+An+International+Multicenter+Analysis.">E-AHPBA DP-CAR study group. Outcomes and Risk Score for Distal Pancreatectomy with Celiac Axis Resection (DP-CAR), An International Multicenter Analysis.</a> Annals of Surgical Oncology. 26 (3), 772-781 (2019).</li><li>Ocuin, L. 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=Robotic+and+open+distal+pancreatectomy+with+celiac+axis+resection+for+locally+advanced+pancreatic+body+tumors:+a+single+institutional+assessment+of+perioperative+outcomes+and+survival.">Robotic and open distal pancreatectomy with celiac axis resection for locally advanced pancreatic body tumors: a single institutional assessment of perioperative outcomes and survival.</a> HPB. 18 (10), 835-842 (2016).</li><li>Caba Molina, D., Lambreton, F., Arrangoiz Majul, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trends+in+Robotic+Pancreaticoduodenectomy+and+Distal+Pancreatectomy.">Trends in Robotic Pancreaticoduodenectomy and Distal Pancreatectomy.</a> Journal of Laparoendoscopic &amp; Advanced Surgical Techniques A. 29 (2), 147-151 (2019).</li><li>Kornaropoulos, 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=Total+robotic+pancreaticoduodenectomy:+a+systematic+review+of+the+literature.">Total robotic pancreaticoduodenectomy: a systematic review of the literature.</a> Surgical Endoscopy. 31 (11), 4382-4392 (2017).</li><li>Peng, L., Lin, S., Li, Y., Xiao, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Systematic+review+and+meta-analysis+of+robotic+versus+open+pancreaticoduodenectomy.">Systematic review and meta-analysis of robotic versus open pancreaticoduodenectomy.</a> Surgical Endoscopy. 31 (8), 3085-3097 (2017).</li><li>Zureikat, A. 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=A+Multi-institutional+Comparison+of+Perioperative+Outcomes+of+Robotic+and+Open+Pancreaticoduodenectomy.">A Multi-institutional Comparison of Perioperative Outcomes of Robotic and Open Pancreaticoduodenectomy.</a> Annals of Surgery. 264 (4), 640-649 (2016).</li><li>Zureikat, A. 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=500+Minimally+Invasive+Robotic+Pancreatoduodenectomies:+One+Decade+of+Optimizing+Performance.">500 Minimally Invasive Robotic Pancreatoduodenectomies: One Decade of Optimizing Performance.</a> Annals of Surgery. 273 (5), 966-972 (2021).</li><li>Cai, 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+Pancreaticoduodenectomy+Is+Associated+with+Decreased+Clinically+Relevant+Pancreatic+Fistulas:+a+Propensity-Matched+Analysis.">Robotic Pancreaticoduodenectomy Is Associated with Decreased Clinically Relevant Pancreatic Fistulas: a Propensity-Matched Analysis.</a> Journal of Gastrointestinal Surgery. 24 (5), 1111-1118 (2020).</li><li>Strasberg, S. M., Linehan, D. C., Hawkins, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Radical+antegrade+modular+pancreatosplenectomy+procedure+for+adenocarcinoma+of+the+body+and+tail+of+the+pancreas:+ability+to+obtain+negative+tangential+margins.">Radical antegrade modular pancreatosplenectomy procedure for adenocarcinoma of the body and tail of the pancreas: ability to obtain negative tangential margins.</a> Journal of the American College of Surgeons. 204 (2), 244-249 (2007).</li><li>Chun, Y. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+Radical+Antegrade+Modular+Pancreatosplenectomy+(RAMPS)+and+Pancreatic+Cancer.">Role of Radical Antegrade Modular Pancreatosplenectomy (RAMPS) and Pancreatic Cancer.</a> Annals of Surgical Oncology. 25 (1), 46-50 (2018).</li></ol>

No financial conflicts of interest to disclose on the part of any of the authors involved.

With proper pre-operative planning, patient selection, and surgeon experience, it is clinically feasible and safe to approach locally advanced pancreatic tumors of the body/tail of the pancreas with celiac involvement with robot assisted distal pancreatectomy, splenectomy, and celiac axis resection. Proper patient selection requires comprehensive pre-operative planning with cross-sectional imaging to identify the tumor and its anatomical relationship to surrounding vascular structures. At this time, it is also imperative...

Pancreatic cancers involving the body and tail of the pancreas are traditionally surgically managed with a distal pancreatectomy and splenectomy. Approximately 30% of pancreatic cancers present in a locally advanced stage with involvement of structures beyond the pancreas1. A subset of these patients present&#160;with involvement of the celiac axis or proximal hepatic artery without involvement of the aorta. In this circumstance, an aggressive pre-operative strategy involving neo-adjuvant chemotherapy of FOLFIRINOX2,3 or Gemcitabine-Abraxane4 with potential neoadjuvant radiation prior to surgical resection with a modified version of the original Appleby procedure is considered5. The procedure involves resecting the celiac axis at its origin and relying on collateral flow to hepatic artery proper through the GDA. While this aggressive approach for locally advanced pancreatic cancer is performed only in highly selected patients, there is suggestion of potential oncologic benefit in retrospective series6,7,8.
The robotic surgical platform offers numerous technical advantages compared with open and laparoscopic techniques, including enhanced three-dimensional visualization, instrument wrist articulation and the ability for the operating surgeon to control multiple instruments and the camera. Additionally, limited retrospective case series of patients undergoing robotic pancreatic surgery have suggested decreased intra-operative blood loss, decreased peri-operative pain, lower pancreatic fistula rates and improved recovery when compared with open pancreatic resections9,10,11,12,13,14. These technical and clinical benefits along with increased robotic training have led to an expansion of the robotic approach in pancreatic surgery, demonstrating the versatility of the platform to perform a variety of pancreatic resections and procedures, including pancreaticoduodenectomy and distal pancreatectomy with and without splenic preservation. Herein, we will&#160;provide the pre-surgical and surgical evaluation and decision making that is involved in proper selection of patients as well as detail the patient characteristics, pre-operative management, and a detailed review of the surgical technique of the DP-CAR performed with the robotic platform on a singular patient in our practice.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Da Vinci Robotic Platform XI</td>
			<td>Intuitive Surgical</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Lightworks Video Editer</td>
			<td>Lightworks</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Studio 3 Video logging platform</td>
			<td>Stryker</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

robot assisted distal pancreatectomy with celiac axis resection (dp-car) for pancreatic cancer: surgical planning and technique

Malignant pancreatic tumors involving the celiac artery can be resected with a distal pancreatectomy, splenectomy and celiac axis resection (DP-CAR), relying on collateral flow to the liver through the gastroduodenal artery (GDA). In the current manuscript, the technical conduct of robotic DP-CAR is outlined. The greater curve of the stomach is mobilized with care to avoid sacrificing the gastroepiploic vessels. The stomach and liver are retracted cephalad to facilitate dissection of the porta hepatis. The hepatic artery (HA) is dissected and encircled with a vessel loop. The gastroduodenal artery (GDA) is carefully preserved. The common HA is clamped and triphasic flow in the proper HA via the GDA is confirmed using intra-operative ultrasound. A retropancreatic tunnel is made over the superior mesenteric vein (SMV). The pancreas is divided with an endovascular stapler at the neck. The inferior mesenteric vein (IMV) and splenic vein are ligated. The HA is stapled proximal to the GDA. The entire specimen is retracted laterally with further dissection cephalad to expose the superior mesenteric artery (SMA). The SMA is then traced back to the aorta. The dissection continues cephalad along the aorta with the bipolar energy device used to divide the crural fibers and celiac nerve plexus. The specimen is mobilized from the patient's right to left until the origin of the celiac axis is identified and oriented towards the left. The trunk is circumferentially dissected and stapled. Additional dissection with hook cautery and the bipolar energy device fully mobilizes the pancreatic tail and spleen. The specimen is removed from the left lower quadrant extraction site and one drain is left in the resection bed. A final intra-operative ultrasound of the proper HA confirms pulsatile, triphasic flow in the artery and liver parenchyma. The stomach is inspected for evidence of ischemia. Robotic DP-CAR is safe, feasible and when used in conjunction with multi-modality therapy, offers potential for long-term survival in selected patients.

The duration of the procedure was 228 minutes with a blood loss of 50 mL. Post-treatment final pathology revealed a moderately differentiated (G2) ypT1c ductal adenocarcinoma. No nodal involvement was noted (0/21 total nodes). The circumferential resection margin was negative. The patient&#39;s post-operative course was uncomplicated. Her drain amylase levels post-operatively were in the normal range and the drain was removed on post-operative day 3. She was discharged home on post-operative day 4 tolerating a regular di...

Watch this Scientific Journal Video about Robot Assisted Distal Pancreatectomy with Celiac Axis Resection DP-CAR for Pancreatic Cancer: Surgical Planning and Technique at JoVE.com

Robot Assisted Distal Pancreatectomy with Celiac Axis Resection DP-CAR for Pancreatic Cancer: Surgical Planning and Technique

We present our operative approach to robot assisted distal pancreatectomy, splenectomy, and celiac axis resection (DP-CAR), demonstrating that the procedure is safe and feasible with proper planning, patient selection, and surgeon experience.

Robot Assisted Distal Pancreatectomy with Celiac Axis Resection (DP-CAR) for Pancreatic Cancer: Surgical Planning and Technique

Malignant pancreatic tumors involving the celiac artery can be resected with a distal pancreatectomy, splenectomy and celiac axis resection (DP-CAR), ...

robot-assisted-distal-pancreatectomy-with-celiac-axis-resection-dp

Research

JoVE Journal

Cancer Research

6.1K Views.  West Virginia University. We present our operative approach to robot assisted distal pancreatectomy, splenectomy, and celiac axis resection (DP-CAR), demonstrating that the procedure is safe and feasible with proper planning, patient selection, and surgeon experience.

Video: Robot Assisted Distal Pancreatectomy with Celiac Axis Resection DP-CAR for Pancreatic Cancer: Surgical Planning and Technique

Laparoscopic Pancreatoduodenectomy With Modified Blumgart Pancreaticojejunostomy

Minimally invasive pancreatic resections are technically demanding but rapidly increasing in popularity. In contrast to laparoscopic distal pancreatectomy, laparoscopic pancreatoduodenectomy (LPD) has not yet obtained wide acceptance, probably due to technical challenges, especially regarding the pancreatic anastomosis.
The study describes and demonstrates all steps of LPD, including the modified Blumgart pancreaticojejunostomy. Indications for LPD are all pancreatic and peri-ampullary tumors without vascular involvement. Relative contra-indications are body mass index &#62;35 kg/m2, chronic pancreatitis, mid-cholangiocarcinomas and large duodenal cancers.
The patient is in French position, 6 trocars are placed, and dissection is performed using an (articulating) sealing device. A modified Blumgart end-to-side pancreaticojejunostomy is performed with 4 large needles (3/0) barbed trans-pancreatic sutures and 4 to 6 duct-to-mucosa sutures using 5/0 absorbable multifilament combined with a 12 cm, 6 or 8 Fr internal stent using 3D laparoscopy. Two surgical drains are placed alongside the pancreaticojejunostomy.
The described technique for LPD including a modified Blumgart pancreatico-jejunostomy is well standardized, and its merits are currently studied in the randomized controlled multicenter trial. This complex operation should be performed at high-volume centers where surgeons have extensive experience in both open pancreatic surgery and advanced laparoscopic gastro-intestinal surgery.

Laparoscopic pancreatoduodenctomy (LPD) may offer advantages over open pancreatoduodenectomy, including early postoperative mobilization, less delayed gastric emptying and a shorter hospital stay. However, LPD is technically challenging and not well-standardized, especially regarding the pancreatic anastomosis. We describe a standardized technique for the pancreatic anastomosis during LPD: modified Blumgart pancreaticojejunostomy.

Minimally invasive pancreatic resections are technically demanding but rapidly increasing in popularity. In contrast to laparoscopic distal ...

The Application of 1% Methylene Blue Dye As a Single Technique in Breast Cancer Sentinel Node Biopsy

In this study, we injected 1% methylene blue dye (MBD) into the subareolar or peritumoral space of the breast. In the case of breast conserving surgery (BCS), a separate incision in the lower axilla hairline was made to find the sentinel nodes (SNs). In mastectomy, the SNs were identified through the same mastectomy incision. The SNs were described as blue nodes or nodes with lymphatic blue channels. An anatomical landmark in the axilla was used to facilitate SNs identification. The SNs metastases were evaluated by intraoperative frozen section analysis and histopathology examination as it is a gold standard. Here, we described the MBD as the lone technique in breast cancer sentinel node biopsy (SNB) which could be useful when radioisotope tracer or patent or isosulfan blue dye (PBD) cannot be provided.

In Indonesia, sentinel node biopsy (SNB) is not routinely performed for breast cancer surgery because of the limitation to provide radioisotope tracer and isosulfan or patent blue dye (PBD). To overcome this obstacle, we applied 1% methylene blue dye (MBD) as a single agent to map the sentinel nodes (SNs).

In this study, we injected 1% methylene blue dye (MBD) into the subareolar or peritumoral space of the breast. In the case of breast conserving surgery ...

Isolation and Characterization of Patient-derived Pancreatic Ductal Adenocarcinoma Organoid Models

Pancreatic ductal adenocarcinoma (PDAC) is amongst the most lethal malignancies. Recently, next-generation organoid culture methods enabling the 3-dimensional (3D) modeling of this disease have been described. Patient-derived organoid (PDO) models can be isolated from both surgical specimens as well as small biopsies and form rapidly in culture. Importantly, organoid models preserve the pathogenic genetic alterations detected in the patient&#39;s tumor and are predictive of the patient&#39;s treatment response, thus enabling translational studies. Here, we provide comprehensive protocols for adapting tissue culture workflow to study 3D, matrix embedded, organoid models. We detail methods and considerations for isolating and propagating primary PDAC organoids. Furthermore, we describe how bespoke organoid media is prepared and quality controlled in the laboratory. Finally, we describe assays for downstream characterization of the organoid models such as isolation of nucleic acids (DNA and RNA), and drug testing. Importantly we provide critical considerations for implementing organoid methodology in a research laboratory.

Patient-derived organoid cultures of pancreatic ductal adenocarcinoma are a rapidly established 3-dimensional model that represent epithelial tumor cell compartments with high fidelity, enabling translational research into this lethal malignancy. Here, we provide detailed methods to establish and propagate organoids as well as to perform relevant biological assays using these models.

Pancreatic ductal adenocarcinoma (PDAC) is amongst the most lethal malignancies. Recently, next-generation organoid culture methods enabling the ...

Generation of Orthotopic Pancreatic Tumors and Ex vivo Characterization of Tumor-Infiltrating T Cell Cytotoxicity

In vivo models of pancreatic cancer provide invaluable tools for studying disease dynamics, immune infiltration and new therapeutic strategies. The orthotopic murine model can be performed on large cohorts of immunocompetent mice simultaneously, is relatively inexpensive and preserves the cognate tissue microenvironment. The quantification of T cell infiltration and cytotoxic activity within orthotopic tumors provides a useful indicator of an antitumoral response.
This protocol describes the methodology for surgical generation of orthotopic pancreatic tumors by injection of a low number of syngeneic tumor cells resuspended in 5 &#181;L basement membrane directly into the pancreas. Mice bearing orthotopic tumors take approximately 30 days to reach endpoint, at which point tumors can be harvested and processed for characterization of tumor-infiltrating T cell activity. Rapid enzymatic digestion using collagenase and DNase allows a single-cell suspension to be extracted from tumors. The viability and cell surface markers of immune cells extracted from the tumor are preserved; therefore, it is appropriate for multiple downstream applications, including flow-assisted cell sorting of immune cells for culture or RNA extraction, flow cytometry analysis of immune cell populations. Here, we describe the ex vivo stimulation of T cell populations for intracellular cytokine quantification (IFN&#947; and TNF&#945;) and degranulation activity (CD107a) as a measure of overall cytotoxicity. Whole-tumor digests were stimulated with phorbol myristate acetate and ionomycin for 5 h, in the presence of anti-CD107a antibody in order to upregulate cytokine production and degranulation. The addition of brefeldin A and monensin for the final 4 h was performed to block extracellular transport and maximize cytokine detection. Extra- and intra-cellular staining of cells was then performed for flow cytometry analysis, where the proportion of IFN&#947;+, TNF&#945;+ and CD107a+ CD4+ and CD8+ T cells was quantified.
This method provides a starting base to perform comprehensive analysis of the tumor microenvironment.

Generation of Orthotopic Pancreatic Tumors and Ex vivo Characterization of Tumor-Infiltrating T Cell Cytotoxicity

This protocol describes the surgical generation of orthotopic pancreatic tumors and the rapid digestion of freshly isolated murine pancreatic tumors. Following digestion, viable immune cell populations can be used for further downstream analysis, including ex vivo stimulation of T cells for intracellular cytokine detection by flow cytometry.

In vivo models of pancreatic cancer provide invaluable tools for studying disease dynamics, immune infiltration and new therapeutic strategies. The ...

Modeling Primary Bone Tumors and Bone Metastasis with Solid Tumor Graft Implantation into Bone

Primary bone tumors or bone metastasis from solid tumors result in painful osteolytic, osteoblastic, or mixed osteolytic/osteoblastic lesions. These lesions compromise bone structure, increase the risk of pathologic fracture, and leave patients with limited treatment options. Primary bone tumors metastasize to distant organs, with some types capable of spreading to other skeletal sites. However, recent evidence suggests that with many solid tumors, cancer cells that have spread to bone may be the primary source of cells that ultimately metastasize to other organ systems. Most syngeneic or xenograft mouse models of primary bone tumors involve intra-osseous (orthotopic) injection of tumor cell suspensions. Some animal models of skeletal metastasis from solid tumors also depend on direct bone injection, while others attempt to recapitulate additional steps of the bone metastatic cascade by injecting cells intravascularly or into the organ of the primary tumor. However, none of these models develop bone metastasis reliably or with an incidence of 100%. In addition, direct intra-osseous injection of tumor cells has been shown to be associated with potential tumor embolization of the lung. These embolic tumor cells engraft but do not recapitulate the metastatic cascade. We reported a mouse model of osteosarcoma in which fresh or cryopreserved tumor fragments (consisting of tumor cells plus stroma) are implanted directly into the proximal tibia using a minimally invasive surgical technique. These animals developed reproducible engraftment, growth, and, over time, osteolysis and lung metastasis. This technique has the versatility to be used to model solid tumor bone metastasis and can readily employ grafts consisting of one or multiple cell types, genetically-modified cells, patient-derived xenografts, and/or labeled cells that can be tracked by optical or advanced imaging. Here, we demonstrate this technique, modeling primary bone tumors and bone metastasis using solid tumor graft implantation into bone.

Bone metastasis models do not develop metastasis uniformly or with a 100% incidence. Direct intra-osseous tumor cell injection can result in embolization of the lung. We present our technique modeling primary bone tumors and bone metastasis using solid tumor graft implantation into bone, leading to reproducible engraftment and growth.

Primary bone tumors or bone metastasis from solid tumors result in painful osteolytic, osteoblastic, or mixed osteolytic/osteoblastic lesions. These ...

An Orthotopic Resectional Mouse Model of Pancreatic Cancer

There is a lack of satisfactory animal models to study adjuvant and/or neoadjuvant therapy in patients being considered for surgery of pancreatic cancer (PC). To address this deficiency, we describe a mouse model involving orthotopic implantation of PC followed by distal pancreatectomy and splenectomy. The model has been demonstrated to be safe and suitably flexible for the study of various therapeutic approaches in adjuvant and neo adjuvant settings.
In this model, a pancreatic tumor is first generated by implanting a mixture of human pancreatic cancer cells (luciferase-tagged AsPC-1) and human cancer associated pancreatic stellate cells into the distal pancreas of Balb/c athymic nude mice. After three weeks, the cancer is resected by re-laparotomy, distal pancreatectomy and splenectomy. In this model, bioluminescence imaging can be used to follow the progress of cancer development and effects of resection/treatments. Following resection, adjuvant therapy can be given. Alternatively, neoadjuvant treatment can be given prior to resection.
Representative data from 45 mice are presented. All mice underwent successful distal pancreatectomy/splenectomy with no issues of hemostasis. A macroscopic proximal pancreatic margin greater than 5 mm was achieved in 43 (96%) mice. The technical success rate of pancreatic resection was 100%, with 0% early mortality and morbidity. None of the animals died during the week after resection.
In summary, we describe a robust and reproducible technique for a surgical resection model of pancreatic cancer in mice which mimics the clinical scenario. The model may be useful for the testing of both adjuvant and neoadjuvant treatments.

In the clinical context, patients with localized pancreatic cancer will undergo pancreatectomy followed by adjuvant treatment. This protocol reported here aims to establish a safe and effective method of modelling this clinical scenario in nude mice, through orthotopic implantation of pancreatic cancer followed by distal pancreatectomy and splenectomy.

There is a lack of satisfactory animal models to study adjuvant and/or neoadjuvant therapy in patients being considered for surgery of pancreatic cancer ...

Robotic Pancreatoduodenectomy for Pancreatic Head Cancer: a Case Report of a Standardized Technique

Robotic pancreatoduodenectomy (RPD) for pancreatic cancer is a challenging procedure. Aberrant vasculature may increase the technical difficulty. Several studies have described the safety of RPD in case of a replaced or aberrant right hepatic artery, but detailed video descriptions of the approach are lacking. This case report describes a step-&#173;by-&#173;step technical video in case of a replaced right hepatic artery. A 58-year-old woman presented with an incidental finding of a 1.7 cm pancreatic head mass. RPD was performed using the da Vinci Xi system and involves a robotic-assisted pancreatico- and hepatico-jejunostomy and open gastro-jejunostomy at the specimen extraction site. The operation time was 410 min with 220 mL of blood loss. The patient had an uncomplicated postoperative course and was discharged after 5 days. Pathology revealed a pancreatic head cancer. RPD is a feasible and safe procedure in case of a replaced hepatic artery when performed in selected patients in high-volume centers by experienced surgeons.

Robotic pancreatoduodenctomy (RPD) has been highly standardized in recent years and may be used in selected patients with pancreatic head cancer, including those with a replaced right hepatic artery. This case report describes a standardized and reproducible technique for RPD, which includes the approach of the Dutch LAELAPS-3 training program to an aberrant vasculature.

Robotic pancreatoduodenectomy (RPD) for pancreatic cancer is a challenging procedure. Aberrant vasculature may increase the technical difficulty. Several ...

Syngeneic Mouse Orthotopic Allografts to Model Pancreatic Cancer

Pancreatic ductal adenocarcinoma (PDAC) is a very complex disease characterized by a heterogeneous tumor microenvironment made up of a diverse stroma, immune cells, vessels, nerves, and extracellular matrix components. Over the years, different mouse models of PDAC have been developed to address the challenges posed by its progression, metastatic potential, and phenotypic heterogeneity. Immunocompetent mouse orthotopic allografts of PDAC have shown good promise owing to their fast and reproducible tumor progression in comparison to genetically engineered mouse models. Moreover, combined with their ability to mimic the biological features observed in autochthonous PDAC, cell line-based orthotopic allograft mouse models enable large-scale in vivo experiments. Thus, these models are widely used in preclinical studies for rapid genotype-phenotype and drug-response analyses. The aim of this protocol is to provide a reproducible and robust approach to successfully inject primary mouse PDAC cell cultures into the pancreas of syngeneic recipient mice. In addition to the technical details, important information is given that must be considered before performing these experiments.

Syngeneic mouse orthotopic allografts of pancreatic ductal adenocarcinoma (PDAC) recapitulate the biology, phenotypes, and therapeutic responses of disease subtypes. Owing to their fast, reproducible tumor progression, they are widely used in preclinical studies. Here, we show common practices to generate these models, injecting syngeneic murine PDAC cultures into the pancreas.

Pancreatic ductal adenocarcinoma (PDAC) is a very complex disease characterized by a heterogeneous tumor microenvironment made up of a diverse stroma, ...

Surgical Technique for Superior Cervical Ganglionectomy in a Murine Model

Growing evidence suggests that the sympathetic nervous system plays an important role in cancer progression. Adrenergic innervation regulates salivary gland secretion, circadian rhythm, macular degeneration, immune function, and cardiac physiology. Murine surgical sympathectomy is a method for studying the effects of adrenergic innervation by allowing for complete, unilateral adrenergic ablation while avoiding the need for repeated pharmacologic intervention and the associated side effects. However, surgical sympathectomy in mice is technically challenging because of the small size of the superior cervical ganglion. This study describes a surgical technique for reliably identifying and resecting the superior cervical ganglion to ablate the sympathetic nervous system. The successful identification and removal of the ganglion are validated by imaging the fluorescent sympathetic ganglia using a transgenic mouse, identifying post-resection Horner's syndrome, staining for adrenergic markers in the resected ganglia, and observing diminished adrenergic immunofluorescence in the target organs following sympathectomy. This model enables future studies of cancer progression as well as other physiological processes regulated by the sympathetic nervous system.

The present protocol describes a mouse model of the ablation of adrenergic innervation by identifying and resecting the superior cervical ganglion.

Growing evidence suggests that the sympathetic nervous system plays an important role in cancer progression. Adrenergic innervation regulates salivary ...

Glioblastoma Relapse Post-Resection Model for Therapeutic Hydrogel Investigations

Tumor recurrence is an important factor indicative of a poor prognosis in glioblastoma (GBM). Many studies are attempting to identify effective therapeutic strategies to prevent the recurrence of GBM after surgery. Bioresponsive therapeutic hydrogels capable of sustaining locally released drugs are frequently used for the local treatment of GBM after surgery. However, research is limited due to the lack of a suitable GBM relapse post-resection model. Here, a GBM relapse post-resection model was developed and applied in therapeutic hydrogel investigations. This model was constructed based on the orthotopic intracranial GBM model, which is widely used in studies on GBM. Subtotal resection was performed on the orthotopic intracranial GBM model mouse to mimic the clinical treatment. The residual tumor was used to indicate the size of the tumor growth. This model is easy to build, can better mimic the situation of GBM surgical resection, and can be applied in various studies on the local treatment of GBM relapse post-resection. As a result, the GBM relapse post-resection model provides a unique GBM recurrence model for effective local treatment studies of relapse post-resection.

The present protocol establishes a glioblastoma (GBM) relapse post-resection model using microscopy to investigate the therapeutic effect of an injectable, bioresponsive hydrogel in vivo.