The authors have no acknowledgements. Funding was received from the Foundation of Health and Family Planning Commission of Zhejiang Province (Grant 20146242) and the Foundation of Wenzhou Science &amp; Technology Bureau (Grant Y20140248).

All animal experiments were approved by the Animal Research and Care Committee at the Children&#39;s Hospital of Pittsburgh and the University of Pittsburgh Institutional Animal Care and Use Committee.
1. Preoperative Preparation
<ol>
	<li>Administer inhaled isoflurane (1 - 3% for maintenance and up to 5% for induction) via a nose cone to an appropriate anesthetic level. Monitor the adequacy of anesthesia by toe pinch with forceps. The lack of a response is considered adequate, and be sure not to overmedicate, as respiratory depression may occur.</li>
	<li>Place the animal on its back on the dissecting microscope stage, with the abdomen centered in view of the objective. Immobilize the animal with surgical tape. Apply ophthalmic ointment to the eyes of the animal to prevent dryness while under anesthesia.</li>
	<li>Apply a smooth, thick layer of hair removal cream and leave it in place for 3 min. Check a small area for hair removal. Gently wipe off the cream and hair with 70% ethanol-soaked gauze.</li>
	<li>Cleanse and disinfect the abdomen of the animal by wiping it with a 70% ethanol-soaked gauze followed by a betadine-soaked gauze. Maintain sterile conditions throughout the surgical procedure.</li>
</ol>
2. Proper Exposure
<ol>
	<li>Make a midline incision in the skin from the xyphoid process to the umbilicus using a scalpel. Lift the peritoneum with Adson forceps and make a small hole in the midline using scissors.</li>
	<li>Extend the peritoneal incision to the length of the skin incision, taking care to remain on the midline (linea alba). 
	NOTE: This will produce an upper midline abdominal laparotomy.</li>
	<li>Elevate the liver with blunt retractors to expose the common bile duct. Clamp the common bile duct with a curved bulldog vascular clamp. 
	NOTE: This maneuver prevents undesired infusion into the liver.</li>
</ol>
3. Identification of the Pancreatic Papilla and Duodenotomy
<ol>
	<li>Identify the duodenum and, with Arruga forceps, retract it caudally to display the papilla. 
	NOTE: The papilla is a white spot on the anterior surface of the second portion of the duodenum.</li>
	<li>Retract the duodenum caudally, allowing for the identification of the course of the pancreatic duct. With a 30&#189;-gauge needle, puncture the wall of the duodenum tangentially, directly opposite the papilla. 
	NOTE: The needle should follow the same angle as the duct as it enters the duodenum.</li>
</ol>
4. Infusion.
<ol>
	<li>Pass the catheter through the duodenotomy created in step 3.2 until the catheter is visible within the duct. Advance the catheter into the duct no more than 1 cm to prevent the bypass of minor ductal tributaries.</li>
	<li>Clamp the catheter in place with a second curved bulldog vascular clamp. Turn on the pump and infuse the selected volume. The volume infused may range from 25 - 250 &#181;L, with an ideal volume of approximately 150 &#181;L.</li>
	<li>Administer an intraperitoneal injection of approximately 1.5 mL of saline to offset losses. Cover the exposed bowel with a moist gauze to prevent desiccation.</li>
	<li>At the conclusion of the infusion, remove the bulldog clamp that was holding the catheter in place. Use the bulldog clamp to gently remove the catheter from the pancreatic duct. Release the clamp from the common bile duct.</li>
	<li>Close the peritoneum and the skin in one or two layers using a running suture. 
	NOTE: There is no need to close the duodenotomy, as it will seal on its own.</li>
	<li>As postoperative analgesia, administer a dose of ketofen at 5 mg/kg during the procedure and one on the following day.</li>
	<li>Return the mouse to its cage under a heat lamp until it recovers. Provide the mouse with food and water ad libitum.</li>
	<li>Do not leave the animal unattended until it has regained sufficient consciousness to maintain sternal recumbency. Do not return an animal that has undergone surgery to the company of other animals until it has fully recovered.</li>
</ol>
5. Sample Preparation
<ol>
	<li>Place the animal in a carbon dioxide chamber or perform cervical dislocation.</li>
	<li>Harvest the pancreas, with the spleen, liver, and duodenum as controls. Fix the tissues in paraformaldehyde overnight at 4 &#176;C. Cryoprotect the sample in 30% sucrose overnight, as previously described6.</li>
	<li>Snap-freeze the samples. Section the tissue at a thickness of 6 &#181;m using a microtome. Perform imaging on a microscope with fluorescence imaging, as previously described7.</li>
</ol>

<ol>
	<li>Gittes, G. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+biology+of+the+pancreas:+a+comprehensive+review.">Developmental biology of the pancreas: a comprehensive review.</a> Dev Biol. 326 (1), 4-35 (2009).</li><li>Gu, G., Dubauskaite, J., Melton, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+evidence+for+the+pancreatic+lineage:+NGN3++cells+are+islet+progenitors+and+are+distinct+from+duct+progenitors.">Direct evidence for the pancreatic lineage: NGN3+ cells are islet progenitors and are distinct from duct progenitors.</a> Development. 129 (10), 2447-2457 (2002).</li><li>Weinberg, N., Ouziel-Yahalom, L., Knoller, S., Efrat, S., Dor, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lineage+tracing+evidence+for+in+vitro+dedifferentiation+but+rare+proliferation+of+mouse+pancreatic+beta-cells.">Lineage tracing evidence for in vitro dedifferentiation but rare proliferation of mouse pancreatic beta-cells.</a> Diabetes. 56 (5), 1299-1304 (2007).</li><li>Raper, S. E., DeMatteo, R. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adenovirus-mediated+in+vivo+gene+transfer+and+expression+in+normal+rat+pancreas.">Adenovirus-mediated in vivo gene transfer and expression in normal rat pancreas.</a> Pancreas. 12 (4), 401-410 (1996).</li><li>Xiao, X., 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=Pancreatic+cell+tracing,+lineage+tagging+and+targeted+genetic+manipulations+in+multiple+cell+types+using+pancreatic+ductal+infusion+of+adeno-associated+viral+vectors+and/or+cell-tagging+dyes.">Pancreatic cell tracing, lineage tagging and targeted genetic manipulations in multiple cell types using pancreatic ductal infusion of adeno-associated viral vectors and/or cell-tagging dyes.</a> Nat Protoc. 9 (12), 2719-2724 (2014).</li><li>Song, Z., 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=EGFR+signaling+regulates+beta+cell+proliferation+in+adult+mice.">EGFR signaling regulates beta cell proliferation in adult mice.</a> J Biol Chem. 291 (43), (2016).</li><li>Xiao, X., 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=M2+macrophages+promote+beta-cell+proliferation+by+up-regulation+of+SMAD7.">M2 macrophages promote beta-cell proliferation by up-regulation of SMAD7.</a> Proc Natl Acad Sci U S A. 111 (13), E1211-E1220 (2014).</li><li>Laukkarinen, J. M., Van Acker, G. J., Weiss, E. R., Steer, M. L., Perides, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+mouse+model+of+acute+biliary+pancreatitis+induced+by+retrograde+pancreatic+duct+infusion+of+Na-taurocholate.">A mouse model of acute biliary pancreatitis induced by retrograde pancreatic duct infusion of Na-taurocholate.</a> Gut. 56 (11), 1590-1598 (2007).</li><li>Lichtenstein, 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=Acute+lung+injury+in+two+experimental+models+of+acute+pancreatitis:+infusion+of+saline+or+sodium+taurocholate+into+the+pancreatic+duct.">Acute lung injury in two experimental models of acute pancreatitis: infusion of saline or sodium taurocholate into the pancreatic duct.</a> Crit Care Med. 28 (5), 1497-1502 (2000).</li><li>Perides, G., van Acker, G. J., Laukkarinen, J. M., Steer, M. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+acute+biliary+pancreatitis+induced+by+retrograde+infusion+of+bile+acids+into+the+mouse+pancreatic+duct.">Experimental acute biliary pancreatitis induced by retrograde infusion of bile acids into the mouse pancreatic duct.</a> Nat Protoc. 5 (2), 335-341 (2010).</li><li>Guo, P., 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=Specific+transduction+and+labeling+of+pancreatic+ducts+by+targeted+recombinant+viral+infusion+into+mouse+pancreatic+ducts.">Specific transduction and labeling of pancreatic ducts by targeted recombinant viral infusion into mouse pancreatic ducts.</a> Lab Invest. 93 (11), 1241-1253 (2013).</li><li>Xiao, X., 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=Neurogenin3+activation+is+not+sufficient+to+direct+duct-to-beta+cell+transdifferentiation+in+the+adult+pancreas.">Neurogenin3 activation is not sufficient to direct duct-to-beta cell transdifferentiation in the adult pancreas.</a> J Biol Chem. 288 (35), 25297-25308 (2013).</li></ol>

The authors have nothing to disclose.

This work describes in detail the methodology behind pancreatic duct infusion, an effective mouse model for the delivery of genes and other molecules specifically and effectively to the pancreas. Ambiguity in the specifics of the procedure between various groups highlighted the need for a standardized protocol8,9,10.
There are several critical steps of the procedure, beginning with the selection of yo...

The pancreas is a bifunctional organ, with both endocrine and exocrine components. A number of pathologies can afflict the pancreas, including diabetes, pancreatitis, and pancreatic cancer1. All three of these diseases mark active areas of study, not only to develop immediate therapy, but also to better understand their pathophysiology.
A targeted therapeutic delivery method would be beneficial. We have developed a pancreatic duct infusion technique that is an effective and selective method for drug and viral delivery to pancreatic cells. In this model, the pancreatic duct is selectively cannulated, and the common bile duct is occluded, allowing for delivery exclusively to the pancreas.
This technique can allow for the lineage tracing of specific cell types to further elucidate the developmental pathways important to pancreatic development and pathology2,3. Different promoters in viral vectors allow for the specific targeting of cell lines and for the introduction of genes into these cell lines4,5. This work demonstrates the expression of a green fluorescent protein (GFP) within the pancreas after the pancreatic duct infusion of a viral vector expressing GFP versus a sham surgery.

<table><tbody><tr><td>Mice (25 - 30g, 8-12 weeks old)</td><td>Jackson Laboratory</td><td></td><td></td></tr><tr><td>Ketofen</td><td>Henry Schein Inc.</td><td>5487</td><td></td></tr><tr><td>Ethanol (70%)</td><td>Sigma-Aldrich</td><td>34923</td><td></td></tr><tr><td>Sterile Saline Solution</td><td>Baxter Healthcare Corp.</td><td>2B-13-00</td><td></td></tr><tr><td>Protective Equipment</td><td></td><td></td><td></td></tr><tr><td>Hair Removal Product</td><td></td><td></td><td>Nair or trimmer</td></tr><tr><td>Gauze Pads 2in x 2in</td><td>Fisher Healthcare</td><td>22-362-178</td><td></td></tr><tr><td>Needles (30.5G and 25G)</td><td>Becton Dicksinson and Co.</td><td>305106 and 305122</td><td></td></tr><tr><td>Syringe for Ketofen and Saline</td><td>Becton Dicksinson and Co.</td><td>309657</td><td></td></tr><tr><td>Syringe for Pump</td><td>Henke Sass Wolf</td><td>4010.200V0</td><td></td></tr><tr><td>Infusion Pump</td><td>Pump Systems Inc.</td><td>NE-1000</td><td></td></tr><tr><td>Infusion Catheter</td><td>World Precision Instruments</td><td>CMF31G</td><td></td></tr><tr><td>Curved Bulldog Clamps (2)</td><td>Roboz</td><td>RS-7439</td><td></td></tr><tr><td>Arruga Forceps</td><td>Roboz</td><td>RS-5163</td><td></td></tr><tr><td>Adson Forcep</td><td>Roboz</td><td>RS-5234</td><td></td></tr><tr><td>Needle Holder</td><td>Roboz</td><td>RS-7882</td><td></td></tr><tr><td>Scissor</td><td>Roboz</td><td>RS-5982</td><td></td></tr><tr><td>Cotton Tip Applicators</td><td>Physician Sales and Services</td><td>22-9988</td><td></td></tr><tr><td>Dissecting Microscope</td><td></td><td></td><td></td></tr><tr><td>Suture</td><td>Owens and Minor</td><td>SXMD1B402</td><td></td></tr></tbody></table>

pancreatic duct infusion: an effective and selective method of drug and viral delivery

The pancreas is a bifunctional organ with both endocrine and exocrine components. A number of pathologies can afflict the pancreas, including diabetes, pancreatitis, and pancreatic cancer. All three of these diseases mark active areas of study, not only to develop immediate therapy, but also to better understand their pathophysiology. There are few tools to further these areas of study. Pancreatic duct infusion is an important technique that can allow for lineage tracing, gene introduction, and cell line-specific targeting. The technique requires the intricate dissection of the second portion of the duodenum and ampulla, followed by the occlusion of the bile duct and the cannulation of the pancreatic duct. Although the technique is technically challenging at first, the applications are myriad. Ambiguity in the specifics of the procedure between groups highlighted the need for a standard protocol. This work describes the expression of a green fluorescent protein (GFP) within the pancreas after the pancreatic duct infusion of a viral vector expressing GFP versus a sham surgery. The infusion and therefore expression is specific to the pancreas, without expression present in any other tissue type.

With practice and careful surgical technique, the survival rate of these mice should be greater than 95%. One week after infusion, the desired effect should be evident in the pancreas. Mice at 10 to 12 weeks of age were utilized for pancreatic duct infusions. Here we use an adeno-associated virus serotype 8 (AAV8) with a CMV promoter to express green fluorescent protein (GFP), as compared to a sham surgery. Mice pancreases were harvested 7 days after the infusions. Sections of pancreas, s...

Watch this Scientific Journal Video about Pancreatic Duct Infusion: An Effective and Selective Method of Drug and Viral Delivery at JoVE.com

Pancreatic Duct Infusion: An Effective and Selective Method of Drug and Viral Delivery

Pancreatic duct infusion is an important technique that can allow for lineage tracing, gene introduction, and cell line-specific targeting. A pancreatic duct infusion technique for drug and viral delivery to pancreatic cells is presented here.

The pancreas is a bifunctional organ with both endocrine and exocrine components. A number of pathologies can afflict the pancreas, including diabetes, ...

pancreatic-duct-infusion-an-effective-selective-method-drug-viral

Research

JoVE Journal

Medicine

4.6K Views.  Children's Hospital of Pittsburgh of UPMC. Het algemene doel van deze pancreaskanaalinfusieprocedure is om afstammingstracering, genintroductie en cellijnspecifieke targeting in de alvleesklier mogelijk te maken. Deze methode kan helpen bij het beantwoorden van belangrijke vragen over ziekten die de alvleesklier aantasten, waaronder chronische pancreatitis, diabetes en alvleesklierkanker. Plaats om te beginnen een verdoofde muis onder een ontleedmicroscoop, zodat zijn buik in het midden van het gezichtsveld wordt geplaatst.Immo...

Video: Pancreatic Duct Infusion: An Effective and Selective Method of Drug and Viral Delivery

Ultrasound-Guided Orthotopic Implantation of Murine Pancreatic Ductal Adenocarcinoma

The recent success of immune checkpoint blockade in melanoma and lung adenocarcinoma has galvanized the field of immuno-oncology as well as revealed the limitations of current treatments, as the majority of patients do not respond to immunotherapy. Development of accurate preclinical models to quickly identify novel and effective therapeutic combinations are critical to address this unmet clinical need. Pancreatic ductal adenocarcinoma (PDA) is a canonical example of an immune checkpoint blockade resistant tumor with only 2% of patients responding to immunotherapy. The genetically engineered KrasG12D+/-;Trp53R172H+/-;Pdx-1 Cre (KPC) mouse model of PDA recapitulates human disease and is a valuable tool for assessing therapies for immunotherapy resistant in the preclinical setting, but time to tumor onset is highly variable. Surgical orthotopic tumor implantation models of PDA maintain the immunobiological hallmarks of the KPC tissue-specific tumor microenvironment (TME) but require a time-intensive procedure and introduce aberrant inflammation. Here, we use an ultrasound-guided orthotopic tumor implantation model (UG-OTIM) to non-invasively inject KPC-derived PDA cell lines directly into the mouse pancreas. UG-OTIM tumors grow in the endogenous tissue site, faithfully recapitulate histological features of the PDA TME, and reach enrollment-sized tumors for preclinical studies by four weeks after injection with minimal seeding on the peritoneal wall. The UG-OTIM system described here is a rapid and reproducible tumor model that may allow for high throughput analysis of novel therapeutic combinations in the murine PDA TME.

We describe a protocol for the ultrasound guided implantation of murine-derived pancreatic ductal adenocarcinoma cell lines directly into the native tumor site. This approach resulted in pancreatic tumors detectable by ultrasound scanning within 2-4 weeks of injection, and significantly reduced the proportion tumor cell seeding on the peritoneal wall as compared to surgical orthotopic implantation.

The recent success of immune checkpoint blockade in melanoma and lung adenocarcinoma has galvanized the field of immuno-oncology as well as revealed the ...

Cancer Research

Reprogramming PDAC and Pancreatic Cells into Induced Pluripotent Stem Cells

Reprogramming Pancreatic Ductal Adenocarcinoma to Pluripotency

The generation of induced pluripotent stem cells (iPSCs) using transcription factors has been achieved from almost any differentiated cell type and has proved highly valuable for research and clinical applications. Interestingly, iPSC reprogramming of cancer cells, such as pancreatic ductal adenocarcinoma (PDAC), has been shown to revert the invasive PDAC phenotype and override the cancer epigenome. The differentiation of PDAC-derived iPSCs can recapitulate PDAC progression from its early pancreatic intraepithelial neoplasia (PanIN) precursor, revealing the molecular and cellular changes that occur early during PDAC progression. Therefore, PDAC-derived iPSCs can be used to model the earliest stages of PDAC for the discovery of early-detection diagnostic markers. This is particularly important for PDAC patients, who are typically diagnosed at the late metastatic stages due to a lack of reliable biomarkers for the earlier PanIN stages. However, reprogramming cancer cell lines, including PDAC, into pluripotency remains challenging, labor-intensive, and highly variable between different lines. Here, we describe a more consistent protocol for generating iPSCs from various human PDAC cell lines using bicistronic lentiviral vectors. The resulting iPSC lines are stable, showing no dependence on the exogenous expression of reprogramming factors or inducible drugs. Overall, this protocol facilitates the generation of a wide range of PDAC-derived iPSCs, which is essential for discovering early biomarkers that are more specific and representative of PDAC cases.

Author Spotlight: Reprogramming Cancer Cells to iPSCs to Study Disease Progression and Treatment Targets

The present protocol describes the reprogramming of Pancreatic Ductal Adenocarcinoma (PDAC) and normal pancreatic ductal epithelial cells into induced pluripotent stem cells (iPSCs). We provide an optimized and detailed, step-by-step procedure, from preparing lentivirus to establishing stable iPSC lines.

The generation of induced pluripotent stem cells (iPSCs) using transcription factors has been achieved from almost any differentiated cell type and has ...

Intraluminal Drug Delivery to the Mouse Arteriovenous Fistula Endothelium

Delivery of therapeutic agents to enhance arteriovenous fistula (AVF) maturation can be administered either via intraluminal or external routes. The simple murine AVF model was combined with intraluminal administration of drug solution to the venous endothelium at the same time as fistula creation. Technical aspects of this model are discussed. Under general anesthesia, an abdominal incision is made and the aorta and inferior vena cava (IVC) are exposed. The infra-renal aorta and IVC are dissected for clamping. After proximal and distal clamping, the puncture site is exposed and a 25 G needle is used to puncture both walls of the aorta and into the IVC. Immediately after the puncture, a reporter gene-expressing viral vector was infused in the IVC via the same needle, followed by 15 min of incubation. The intraluminal administration method enabled more robust viral gene delivery to the venous endothelium compared to administration by the external route. This novel method of delivery will facilitate studies that explore the role of the endothelium in AVF maturation and enable intraluminal drug delivery at the time of surgical operation.

After puncturing the aorta through the inferior vena cava (IVC) to create an aorto-caval fistula in the mouse, solution containing a drug is infused into the IVC via the same needle, followed by incubation. This method enables more robust drug delivery to the venous endothelium compared to the external route.

Delivery of therapeutic agents to enhance arteriovenous fistula (AVF) maturation can be administered either via intraluminal or external routes. The ...

Intraportal Transplantation of Pancreatic Islets in Mouse Model

Pancreatic islet transplantation to reduce hyperglycemia is highly successful in rodents with chemically-induced diabetes. The most common transplantation site in experimental islet transplantation is the kidney capsule. However, as less is known about the interaction of pancreatic islets with blood constituents, it also makes sense to utilize the portal vein approach in experimental islet transplantation.
This protocol demonstrates an intraportal islet transplantation technique in NMRI nude mice. Streptozotocin (180 mg/kg) is injected intraperitoneally to induce hyperglycemia in recipient mice. They are considered as diabetic at a non-fasting blood glucose level greater than 20 mmol/L. One day prior to transplantation, mouse pancreatic islets are isolated from the donor pancreas by collagenase digestion; a minimum of 350 islets are utilized per diabetic recipient. Depending upon the islet isolation yield, two or more donor mice are utilized per recipient. After overnight culture at 37 &#176;C, islets are administered into the recipient liver via the portal vein. After surgery, the mice are protected in red Makrolon houses and observed until are awake. This protocol maintains glycemic control for 120 days in syngeneic mice and 15 days in allogeneic mice.

Pancreatic islet transplantation is a way to achieve normoglycemia in type 1 diabetes. This article focuses on the transplantation technique through the intraportal route to maintain normal glucose levels in diabetic mice.

Pancreatic islet transplantation to reduce hyperglycemia is highly successful in rodents with chemically-induced diabetes. The most common transplantation ...

Retrograde Parotid Gland Infusion through Stensen's Duct in a Non-Human Primate for Vectored Gene Delivery

Salivary glands are an attractive tissue target for gene therapy with promising results already leading to human trials. They are inherently capable of secreting proteins into the bloodstream and are easily accessible, making them potentially superior tissue sites for replacement hormone production or vaccination by gene transfer. Suggested methods for gene delivery include transcutaneous injection and retrograde infusion through salivary ducts. We demonstrate how to perform Retrograde Salivary Gland Infusion (RSGI) in non-human primates. We describe the important anatomic landmarks including identification of the parotid papilla, an atraumatic method of cannulating and sealing Stensen&#39;s Duct utilizing basic dental tools, polyethylene tubing, and cyanoacrylate, and the appropriate rate of infusion. While this is the least traumatic method of delivery, the method is still limited by the volume able to be delivered (&#60;0.5 mL) and the potential for trauma to the duct and gland. We demonstrate using fluoroscopy that an infusate can be fully delivered into the gland, and further demonstrate by immunohistochemistry the transduction of a typical vector and expression of the delivered gene.

Salivary glands have been proposed as a tissue target site for gene therapy, especially in the area of vaccination by gene transfer. We demonstrate gene delivery in a non-human primate model utilizing retrograde parotid infusion.

Salivary glands are an attractive tissue target for gene therapy with promising results already leading to human trials. They are inherently capable of ...

Immunology and Infection

Stabilized Longitudinal In Vivo Cellular-Level Visualization of the Pancreas in a Murine Model with a Pancreatic Intravital Imaging Window

Direct in vivo cellular-resolution imaging of the pancreas in a live small animal model has been technically challenging. A recent intravital imaging study, with an abdominal imaging window, enabled visualization of the cellular dynamics in abdominal organs in vivo. However, due to the soft sheet-like architecture of the mouse pancreas that can be easily influenced by physiologic movement (e.g., peristalsis and respiration), it was difficult to perform stabilized longitudinal in vivo imaging over several weeks at the cellular level to identify, track, and quantify islets or cancer cells in the mouse pancreas. Herein, we describe a method for implanting a novel supporting base, an integrated pancreatic intravital imaging window, that can spatially separate the pancreas from the bowel for longitudinal time-lapse intravital imaging of the pancreas microstructure. Longitudinal in vivo imaging with the imaging window enables stable visualization, allowing for the tracking of islets over a period of 3 weeks and high-resolution three-dimensional imaging of the microstructure, as evidenced here in an orthotopic pancreatic cancer model. With our method, further intravital imaging studies can elucidate the pathophysiology of various diseases involving the pancreas at the cellular level.

Stabilized Longitudinal In Vivo Cellular-Level Visualization of the Pancreas in a Murine Model with a Pancreatic Intravital Imaging Window

In vivo high-resolution imaging of the pancreas was facilitated with the pancreatic intravital imaging window.

Direct in vivo cellular-resolution imaging of the pancreas in a live small animal model has been technically challenging. A recent intravital imaging ...

Ultrasound-Guided Implantation: A Technique to Implant Pancreatic Ductal Adenocarcinoma Cells into an Adult Mouse

Source: Hay, C. A. et al.&#160;&#160;<a href="https://www.jove.com/v/60497/" target="_blank">Ultrasound-Guided Orthotopic Implantation of Murine Pancreatic Ductal Adenocarcinoma</a>. J. Vis. Exp. (2019).
This video describes the protocol for ultrasound-guided orthotopic implantation of murine-derived pancreatic ductal adenocarcinoma cells directly into the pancreas of the mouse. This causes less inflammation at the implantation site compared to surgical procedures.

Source: Hay, C. A. et al.&#160;&#160;Ultrasound-Guided Orthotopic Implantation of Murine Pancreatic Ductal Adenocarcinoma. J. Vis. Exp. (2019).
This video ...

JoVE Encyclopedia of Experiments

Pancreatic Cancer

Assays & techniques

Targeting Liver Tumors with Oncolytic Viruses via the Hepatic Artery

Source: Altomonte, J. et al., <a href="https://app.jove.com/v/53757">Transarterial Administration of Oncolytic Viruses for Locoregional Therapy of Orthotopic HCC in Rats.</a>&#160;J. Vis. Exp. (2016)
The video demonstrates locoregional delivery of oncolytic viruses through hepatic arterial injection as a therapeutic approach for hepatocellular carcinoma. These viruses selectively replicate in tumor cells, inducing cell lysis and releasing tumor cell-associated antigens. This process triggers immunogenic cell death, inducing an anti-tumor response.

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...

Immunology

Immunotherapy

Model Development and Disease Study

Intracardiac Delivery of Viral Vectors: A Technique to Inject Viral Vectors Intracardially to Study Pancreatic Metastasis in Murine Model

Source: Zhang, G., et al. <a href="https://www.jove.com/v/55890/" target="_blank">Identification and Characterization of Metastatic Factors by Gene Transfer into the Novel RIP-Tag; RIP-tva Murine Model</a>. J. Vis. Exp. (2017)
This video describes the protocol for intracardiac injection of retroviruses into the transgenic mouse to ensure gene transfer into proliferating pancreatic &#946; premalignant lesions. This method can also help in assessing the functions of candidate genes in tumor metastasis.

Source: Zhang, G., et al. Identification and Characterization of Metastatic Factors by Gene Transfer into the Novel RIP-Tag; RIP-tva Murine Model. J. Vis. ...

Acute Biliary Pancreatitis Model: A Method to Generate Acute Biliary Pancreatitis Mouse Model via Pancreatic Duct Infusion

Source: Serra, M. B. et al. <a href="https://www.jove.com/video/61547" target="_blank">Sodium Taurocholate Induced Severe Acute Pancreatitis in C57BL/6 Mice</a>. J. Vis. Exp. (2021)
This video describes the procedure for the generation of a mouse model with acute biliary pancreatitis via pancreatic infusion of sodium taurocholate. This model helps study pathophysiological changes induced by sodium taurocholate in the pancreas.

Source: Serra, M. B. et al. Sodium Taurocholate Induced Severe Acute Pancreatitis in C57BL/6 Mice. J. Vis. Exp. (2021)
This video describes the procedure ...

Pancreatic Duct Infusion: An Effective and Selective Method of Drug and Viral Delivery

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