Name: Intraportal Transplantation of Pancreatic Islets in Mouse Model (Video) | JoVE Journal
Uploaded: 2018-05-05T15:00:00.000+00:00
Duration: 9 min 1 s
Description: Learn intraportal islet transplantation for type 1 diabetes research using mouse models. This method demonstrates microsurgery techniques to achieve normoglycemia and study islet engraftment in vivo.

ai_agent_IP

ai_agent

maintenance

instantTrialDurationMinutes

isBiopharma

enableNewMyJoveSectionDesign

educationFeatures

seoMetaApi

ai_copilot_institution_blacklist

ai_chat_institution_blacklist

freeAccessWindow

ssoRoleCapture

scheduledMaintenanceBanner

joveLabsFeatures

disable_autoplay_for_bots

extended_trial_duration

5 minutes

We would like to acknowledge Gundula Hertl for her support.

All procedures in this protocol have been approved by German Animal welfare law and guidelines. Consider using 10- to 12-week-old NMRI nude mice and C57Bl/6 mice (donor: old female; recipient: male) throughout the experiments. Use NMRI nude mice for islet isolation. Keep the mice under defined conditions according to local animal facility regulations.
1. Induction of Diabetes by Streptozotocin
<ol>
	<li>On day -4, measure the body weights of 10 to 12 week-old NMRI nude mice and C57Bl/6 mice using a weighing machine.</li>
	<li>According to the measured body weight, prepare a 180 mg/kg dose of streptozotocin in 0.1 mL of saline per mouse. 
	NOTE: Streptozotocin should be freshly prepared before use and covered with aluminum foil, as it is light-sensitive.</li>
	<li>Inject the syngeneic and allogeneic mice each with a single dose of 0.1 mL of streptozotocin (180 mg/kg) intraperitoneally using a 26-G needle.</li>
	<li>On day -3,-2 and -1 collect 2-4 &#181;L of blood by puncturing the tail veins of non-fasted mice.</li>
	<li>Use 2-4 &#181;L of the blood sample per test strip to measure the blood glucose in NMRI nude mice and C57Bl/6 mice 
	NOTE: At this point, 80% of the mice should become diabetic (20 mmol/L).
	<ol>
		<li>Use mice for islet transplantation when their blood glucose levels reach 20 mmol/L.</li>
	</ol>
	</li>
</ol>
2. Isolation of Pancreatic Islets from Mice
<ol>
	<li>Anesthetize the mice with ketamine (100 mg/kg body weight) and xylazine (20 mg/kg body weight) by intraperitoneal injection with a 26-G needle. 
	NOTE: The volumes of ketamine and xylazine were determined according to the body weights of the animals (0.10 mL/10 g).</li>
	<li>Keep the mice in supine position and disinfect with povidone-iodine solution. Confirm proper euthanasia by checking the pedal reflex. 
	NOTE: In case of C57Bl/6 shave the abdominal region.</li>
	<li>Hold the skin with fine forceps and make a midline incision with scissors to expose the abdominal cavity. Place the intestine situs to the left and outside of the abdominal cavity on a sterile gauze compress.</li>
	<li>Adjust the stomach in another direction to locate the pancreas and other associated tissues.</li>
	<li>Reach the aorta and make a small cut to drain the blood. Use sterile gauze for blood absorption.</li>
	<li>Carefully remove the intestine. Separate the pancreas from the stomach and spleen.</li>
	<li>Excise the pancreas and place it in a fresh Petri dish. Remove all the unwanted materials, such as fat, using forceps.</li>
	<li>Fill a 5 mL syringe with 4 mL of collagenase and inject into the pancreas with a 26-G needle (stock solution: 30 mg of collagenase in 12 mL of Hank&#39;s solution).</li>
	<li>Mince the pancreas with scissors to increase the surface area for an improved digestion and transfer to a 12 mL red cap tube. 
	NOTE: Hank&#39;s balanced salt solution: HBSS 10x (100 mL), Penicillin-Streptomycin (10 mL), Gentamycin (1 mL), Ciprofloxacin (10 mL), HEPES (35 mL), distilled H2O (900 mL)</li>
	<li>Perform a digestion for 10 min at 37 &#176;C in a shaking water bath. Vortex the sample thrice at a regular interval for 15 s.</li>
	<li>Mix the digested pancreas vigorously by hand for 3 min and place it on ice for 10 s.</li>
	<li>Add 6 mL of cold Hank&#39;s solution to stop the digestion reaction and centrifuge at 560 x g for 3 min at room temperature.</li>
	<li>Remove the supernatant and dissolve the pellet in 10 mL of Parker/fetal calf serum (P/FCS) medium at room temperature. 
	NOTE: P/FCS medium: TCM-199 10x (100 mL), FCS (50 mL), Penicillin-Streptomycin (10 mL), HEPES (20 mL), distilled H2O (900 mL)</li>
	<li>Identify the islets based on their morphology (spheroid, golden-brown color) under a light microscope. Avoid islets with dark centers. Check for purity and viability by performing dithizone staining under a light microscope. 
	NOTE: Dithizone binds to zinc ion and provides red-colored stain to the islets.</li>
	<li>Use a 1-mL pipette to count and handpick islets under a light microscope and transfer them to a fresh Petri dish (60 mm x 15 mm) containing 4 mL of P/FCS medium. 
	NOTE: The volume of P/FCS medium in the Petri dish should not exceed 4 mL in order to ensure oxygen availability.</li>
	<li>Keep the isolated islets in 4 mL of P/FCS medium overnight at 37 &#176;C in an incubator.</li>
</ol>
3. Preparation of Animals for Islet Transplantation
<ol>
	<li>Remove the isolated islets from the 37 &#176;C incubator and place them under a flow bench. Center the islets with a circular swinging motion of the Petri dish.</li>
	<li>Fill a 1-mL syringe (blue 23-G needle) with 0.1 mL of P/FCS medium. Then, add 350 islets in 0.3 mL of Hank&#39;s solution. Keep the syringe in a vertical position to allow the islets to settle.</li>
	<li>Decrease the volume to 0.05 mL and replace the blue 23-G needle with a brown 26-G needle. To the same syringe, add 0.15 mL of Ficoll (density gradient medium) to the top to reach a final volume of 0.2 mL in the syringe. Avoid air bubbles.</li>
	<li>Keep the syringe in a vertical position and rotate it slightly to avoid the clumping of islets. The islets will settle at the cone of the syringe within 2-3 min.</li>
	<li>Measure the body weight and blood glucose levels of the recipient mice; the blood glucose levels should be around 20 mmol/L.</li>
	<li>Anesthetize the mice with ketamine (100 mg/kg body weight) and xylazine (20 mg/kg body weight) by intraperitoneal injection with 26-G needle.</li>
	<li>Place the mice in supine position on a warm thermal plate (37 &#176;C). Check the depth of anesthesia by pinching the toe.</li>
	<li>Clean the abdominal region with povidone-iodine solution (skin disinfectant). To prevent the eyes from drying, keep them moist with eye ointment. 
	NOTE: In case of C57Bl/6 shave the abdominal region.</li>
</ol>
4. Islet Transplantation via the Intraportal Route
<ol>
	<li>On day 0, start with a laparotomy by holding the skin with forceps and making a 2-3 cm-long midline incision with a scissor. Place a sterile wet gauze compress around the wound edges to keep it moist.</li>
	<li>Place the intestine situs to the left and outside the abdominal cavity on the sterile gauze compress. 
	NOTE: Keep the intestine moist throughout the procedure with pre-warmed 0.9% NaCl solution or sterile gauze dipped in normal saline.</li>
	<li>Move the duodenum and locate the pancreas. Locate the portal vein situated at the ventral site by pushing the pancreas upward. Use a finger and thumb to expose the portal vein and, once exposed, puncture it near to the liver.</li>
	<li>Within 1 min, slowly and steadily inject the islets (0.2 mL from step 3.4) into the portal vein under the inspection of magnifying glass to avoid any leakage of islets. 
	NOTE: After injecting, always check for remaining islets in the syringe. Take 1 mL of P/FCS media in a syringe, flush into a fresh Petri dish and count under a microscope.</li>
	<li>Place a sterile gauge at the puncture site and apply pressure with the index finger (penetration depth: 0.5-1 cm) for 6 min to stop the bleeding. 
	NOTE: The volume injected should not exceed 0.2 mL. Excessive volume and pressure can damage the islets due to shear stress.</li>
	<li>Carefully place the intestine situs, peritoneum, abdominal muscle layer, and fascia back into position.</li>
	<li>Using forceps, lift the skin, close the muscle incision with absorbable silk suture, and upper skin layer with 2-3 clips.</li>
	<li>Clean the surface with skin disinfectant, apply a wound healer, and place the mice in individual cages. Wound healer provides a water-resistant acrylate layer and protect from maceration and secondary infection. 
	NOTE: Observe the mice until they have regained sufficient consciousness, provide tramadol (0.2 mg per mouse) along with fluids. In this protocol, mice recovered from anesthesia quickly and tramadol administration reduced the pain. After 7 days, the wound should heal.</li>
	<li>Feed the mice with regular food and water. Food and water consumption and urine excretion will increase after the streptozotocin injection.</li>
	<li>Measure the blood glucose levels every day for the first week and then once a week in syngeneic mice. In allogeneic mice, measure the blood glucose levels thrice a week for up to 15 days.</li>
</ol>

<ol><li>McCall, M., Shapiro, A. M. <a target="_blank" href="https://www.ncbi.nlm.nih.gov/pubmed/22762022">Update on islet transplantation.</a> Cold Spring Harbor Perspectives in Medicine. 2 (7), a007823(2012).</li>
<li>Shapiro, A. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Islet+transplantation+in+seven+patients+with+type+1+diabetes+mellitus+using+a+glucocorticoid-free+immunosuppressive+regimen%5BTitle%5D)+AND+%22New+England+Journal+of+Medicine%22%5BJournal%5D)">Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen.</a> New England Journal of Medicine. 343 (4), 230-238 (2000).</li>
<li>Cantarelli, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Bone+marrow+as+an+alternative+site+for+islet+transplantation%5BTitle%5D)+AND+%22Blood%22%5BJournal%5D)">Bone marrow as an alternative site for islet transplantation.</a> Blood. 114 (20), 4566-4574 (2009).</li>
<li>Echeverri, G. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Endoscopic+gastric+submucosal+transplantation+of+islets+%28ENDO-STI%29%3A+technique+and+initial+results+in+diabetic+pigs%5BTitle%5D)+AND+%22American+Journal+of+Transplantation%22%5BJournal%5D)">Endoscopic gastric submucosal transplantation of islets (ENDO-STI): technique and initial results in diabetic pigs.</a> American Journal of Transplantation. 9 (11), 2485-2496 (2009).</li>
<li>Korsgren, O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optimising+islet+engraftment+is+critical+for+successful+clinical+islet+transplantation%5BTitle%5D)+AND+%22Diabetologia%22%5BJournal%5D)">Optimising islet engraftment is critical for successful clinical islet transplantation.</a> Diabetologia. 51 (2), 227-232 (2008).</li>
<li>Merani, S., Toso, C., Emamaullee, J., Shapiro, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optimal+implantation+site+for+pancreatic+islet+transplantation%5BTitle%5D)+AND+%22British+Journal+of+Surgery%22%5BJournal%5D)">Optimal implantation site for pancreatic islet transplantation.</a> British Journal of Surgery. 95 (12), 1449-1461 (2008).</li>
<li>Lingwal, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Inhibition+of+gelatinase+B+%28matrix+metalloprotease-9%29+activity+reduces+cellular+inflammation+and+restores+function+of+transplanted+pancreatic+islets%5BTitle%5D)+AND+%22Diabetes%22%5BJournal%5D)">Inhibition of gelatinase B (matrix metalloprotease-9) activity reduces cellular inflammation and restores function of transplanted pancreatic islets.</a> Diabetes. 61 (8), 2045-2053 (2012).</li>
<li>Chen, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Improved+intraportal+islet+transplantation+outcome+by+systemic+IKK-beta+inhibition%3A+NF-kappaB+activity+in+pancreatic+islets+depends+on+oxygen+availability%5BTitle%5D)+AND+%22American+Journal+of+Transplantation%22%5BJournal%5D)">Improved intraportal islet transplantation outcome by systemic IKK-beta inhibition: NF-kappaB activity in pancreatic islets depends on oxygen availability.</a> American Journal of Transplantation. 11 (2), 215-224 (2011).</li>
<li>Wang, Y., Wang, T., Zhang, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Three+aspects+are+critical+for+carrying+out+intraportal+islet+transplants+successfully+in+a+diabetes+mouse+model%5BTitle%5D)+AND+%22Experimental+and+Clinical+Transplantation%22%5BJournal%5D)">Three aspects are critical for carrying out intraportal islet transplants successfully in a diabetes mouse model.</a> Experimental and Clinical Transplantation. 10 (3), 302-304 (2012).</li>
<li>Neuman, J. C., Truchan, N. A., Joseph, J. W., Kimple, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+method+for+mouse+pancreatic+islet+isolation+and+intracellular+cAMP+determination%5BTitle%5D)+AND+%22Journal+of+Visualized+Experiments%22%5BJournal%5D)">A method for mouse pancreatic islet isolation and intracellular cAMP determination.</a> Journal of Visualized Experiments. (88), e50374(2014).</li>
<li>Zmuda, E. J., Powell, C. A., Hai, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+method+for+murine+islet+isolation+and+subcapsular+kidney+transplantation%5BTitle%5D)+AND+%22Journal+of+Visualized+Experiments%22%5BJournal%5D)">A method for murine islet isolation and subcapsular kidney transplantation.</a> Journal of Visualized Experiments. (50), (2011).</li>
<li>Rother, K. I., Harlan, D. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Challenges+facing+islet+transplantation+for+the+treatment+of+type+1+diabetes+mellitus%5BTitle%5D)+AND+%22Journal+of+Clinical+Investigation%22%5BJournal%5D)">Challenges facing islet transplantation for the treatment of type 1 diabetes mellitus.</a> Journal of Clinical Investigation. 114 (7), 877-883 (2004).</li>
<li>Su, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Small+islets+are+essential+for+successful+intraportal+transplantation+in+a+diabetes+mouse+model%5BTitle%5D)+AND+%22Scandinavian+Journal+of+Immunology%22%5BJournal%5D)">Small islets are essential for successful intraportal transplantation in a diabetes mouse model.</a> Scandinavian Journal of Immunology. 72 (6), 504-510 (2010).</li>
<li>Rekittke, N. E., Ang, M., Rawat, D., Khatri, R., Linn, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Regenerative+Therapy+of+Type+1+Diabetes+Mellitus%3A+From+Pancreatic+Islet+Transplantation+to+Mesenchymal+Stem+Cells%5BTitle%5D)+AND+%22Stem+Cells+International%22%5BJournal%5D)">Regenerative Therapy of Type 1 Diabetes Mellitus: From Pancreatic Islet Transplantation to Mesenchymal Stem Cells.</a> Stem Cells International. 2016, 22(2016).</li>
</ol>

All authors declare that they have no conflict of interests.

In this study, the intraportal route of islet transplantation is explored. This protocol is highly efficient in relieving diabetic stress and provides an in-depth opportunity to explore islet transplantation studies. This protocol confirms that about 350 murine islets are capable of reversing the hyperglycemic state. Moreover, none of the mice died during the procedure, and glycemic control was achieved in all recipients. Body weight and blood glucose levels were regularly recorded to reflect the competence, standard, an...

Islet transplantation is a promising approach to treat type 1 diabetes mellitus1. The first attempt to transfer a sheep pancreatic fragment into a diabetic patient was performed by Watson Williams in 1893. However, a major breakthrough in clinical islet transplantation was achieved with the Edmonton protocol, and thereafter, a series of national programs were developed2. In the past, several sites, such as the bone marrow, omental pouch, intramuscular region, gastric mucosal surface, spleen, and kidney capsule, were explored in preclinical models, but the portal venous system is considered as one of the suitable and effective site for clinical programs3,4,5,6.
Exogenous administration of insulin can be substituted by islet infusion into the portal vein. This is considered a preferred site because the oxygen supply is comparable to that in the native pancreas due to the location downstream of the confluence of the portal artery and vein. Moreover, there is a large surface area, so that the three-dimensional structure of the islets may be preserved, and vascularization could be facilitated5. In a recipient diabetic mouse, 2,000 human or porcine islet equivalents or 350 mouse islets are adequate to reverse the hyperglycemic state7,8. Euglycemic levels were reported for 15 days in the mouse recipient model of xenogeneic islets and in the allogeneic mouse-to-mouse model, and for more than 120 days in the syngeneic mouse-to-mouse transplantation.
Factors pertinent to the efficacy of islet transplantation via portal vein are appropriate anesthesia, method of puncture, and hemostasis9. Anesthesia can be induced by inhalation (5% isoflurane) or injection (ketamine, xylazine, or pentobarbital), and the substances are often combined. For control of the depth and time of anesthesia, attention should be paid to the state of the mouse, such as the color of the mucous membrane, eye lid, corneal reflex, respiratory rate, and body temperature. It is important to ensure that the animal is not struggling and that it survives the operation. The temperature can be maintained by different heating devices, such as heating pads, red light bulbs, etc. A warm thermal plate has been used to maintain a temperature of 25-30 &#176;C between the mouse and the operation table, which prevents the occurrence of hypothermia. The dosages of the anesthetics are important, as all of them are metabolized by the liver, and hepatic function may be transiently disordered by the infusion of the islet suspension. The ideal puncture point for the portal vein is the position between the first and second tributary vein.
The number of islets and the intended injection volume also influence the outcome of the transplant, as an excessive volume may increase the shear stress. A 0.2-mL volume is considered appropriate for islet transplantation into the portal vein. The puncturing wound (26-gauge needle) induces bleeding from the portal vein, which needs to be stopped in a timely and effective manner. Sterile gauze or a finger can be applied with minimal pressure at the site of the puncture for about 6 min to stop the bleeding. Taken together, portal islet injection is effective and provides regulation over blood glucose levels in chemically induced diabetic mouse models.

<table><tbody><tr><td>C57Bl/6 mice&amp;nbsp;</td><td>Charles River (Sulzfeld, Germany)</td><td></td><td>(10-12 weeks)</td></tr><tr><td>BALB/c mice</td><td>Charles River (Sulzfeld, Germany)</td><td></td><td>(10-12 weeks)</td></tr><tr><td>NMRI nude mice</td><td>Charles River (Sulzfeld, Germany)</td><td></td><td>(10-12 weeks)</td></tr><tr><td>Ketamine</td><td>Bela-pharm</td><td></td><td></td></tr><tr><td>Xylazine</td><td>CEVA TIERGESUNDHEIT GmbH</td><td></td><td></td></tr><tr><td>Syringe, 23G and 26G needle</td><td>BD</td><td></td><td></td></tr><tr><td>Petri Dish</td><td>Falcon</td><td>303800</td><td></td></tr><tr><td>NaCl</td><td>Carl Roth</td><td>351007</td><td></td></tr><tr><td>Sterile Gauze</td><td>Fuhrmann</td><td>7647-14-5</td><td></td></tr><tr><td>Shaking Water bath</td><td>Kotterman</td><td>1501</td><td></td></tr><tr><td>Scissors</td><td>Braun</td><td>1501</td><td></td></tr><tr><td>Glucose Meter&amp;nbsp;</td><td>One Touch</td><td></td><td></td></tr><tr><td>Warm Thermal Plate</td><td>Thermo Fisher</td><td></td><td></td></tr><tr><td>Braunol (povidone-iodide solution)</td><td>B.Braun Melsungen AG</td><td>3864154</td><td></td></tr><tr><td>Liposic Edo - Sprinkled Ointment (eye ointment)</td><td>Dr. G Mann Chem.-Pharm.&amp;nbsp; GmbH</td><td></td><td></td></tr><tr><td>Incubator</td><td>Fa. Roth</td><td></td><td></td></tr><tr><td>Flow Bench</td><td>Herdus</td><td></td><td></td></tr><tr><td>Ficoll</td><td>Sigma-Aldrich</td><td>10771</td><td></td></tr><tr><td>5-0 Silk Suture Vicryl</td><td>Braun</td><td></td><td>(7.5 *1.75mm)</td></tr><tr><td>Michel Clamps (clips)</td><td>Aesculap&amp;nbsp;</td><td>BN507R</td><td></td></tr><tr><td>P/FCS Solution</td><td>Gibco</td><td>21180-021</td><td></td></tr><tr><td>TCM-199 (10x)</td><td>Gibco</td><td>21180-021</td><td></td></tr><tr><td>FCS</td><td>Biowest</td><td>S1810-500</td><td></td></tr><tr><td>Penicillin-streptomycin</td><td>Gibco</td><td>15140-122</td><td></td></tr><tr><td>HEPES (1 M)</td><td>Biochrom</td><td>L 1613</td><td></td></tr><tr><td>Gentamycin</td><td>Ratiopharm</td><td></td><td></td></tr><tr><td>Ciprofloxacin</td><td>Fresenius Kabi</td><td></td><td></td></tr><tr><td>Hank&#039;s Solution</td><td>Gibco</td><td>14060-040</td><td></td></tr><tr><td>Collagenase</td><td>Roche</td><td>11213865001</td><td></td></tr><tr><td>Streptozotocin</td><td>Calbiochem</td><td>572204</td><td></td></tr><tr><td>OPSITE-spray (wound healer)</td><td>Smith and nephew</td><td>66004978</td><td></td></tr><tr><td>Reugular food</td><td>Altromin</td><td></td><td></td></tr><tr><td>Head light LED 16042 (magnifying glass)</td><td>Eschenbach</td><td>16042</td><td></td></tr><tr><td>Rat anti-mouse PECAM-1(CD31) monoclonal primary antibody</td><td>Millipore, Chemicon</td><td>CBL1337</td><td>Dilution (1:100)</td></tr><tr><td>Donkey anti-rat secondary antibody</td><td>Dianova</td><td>712095153</td><td>Dilution (1:400)</td></tr><tr><td>Polyclonal guinea pig anti-insulin primay antibody</td><td>Dako</td><td>A0564</td><td>Dilution (1:500)</td></tr><tr><td>Donkey anti-guinea pig secondary antibody</td><td>Dianova</td><td>706-259-148</td><td>Dilution (1:400)</td></tr></tbody></table>

intraportal transplantation of pancreatic islets in mouse model

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.

The competency of transplanted islets was studied in chemically induced diabetic mice. The islets were transplanted via the portal venous system in diabetic mice model. The donor-recipient ratio was 2:1. In the syngeneic mouse model, two donor mice were utilized to obtain 350 islets. The islets were then transplanted into diabetic mice. A reduction in blood glucose level was observed on day 1 after transplantation, suggesting a role of transplanted pancreatic islets in reducing the blood ...

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 ...

intraportal-transplantation-of-pancreatic-islets-in-mouse-model

Research

JoVE Journal

Medicine

Our protocol was developed to cleanly and easily deliver islets or cells under the kidney capsule of mice.  Cells are concentrated into pellets in the final tubing used for transplanting the cells under the kidney capsule. The ease of this technique reduces stress to the cells and the mouse.

Transplantation of Pancreatic Islets Into the Kidney Capsule of Diabetic Mice

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.

A Method for Murine Islet Isolation and Subcapsular Kidney Transplantation

This video describes the protocol for grafting human pancreatic islets into the anterior chamber of the mouse’s eye to monitor revascularization of human islet grafts in vivo. It also helps to understand the contribution of recipient versus donor cells in promoting the encapsulation and vascularization of the graft.

Grafting Pancreatic Islets: A Method to Implant Pancreatic Islets into the Anterior Chamber of the Mouse Eye for In Vivo Imaging

This protocol demonstrates murine islet isolation and seeding onto a decellularized scaffold. Scaffold-supported islets were transplanted into the epididymal fat pad of streptozotocin (STZ)-induced diabetic mice. Islets survived at the transplantation site and reversed the hyperglycemic condition.

Scaffold-supported Transplantation of Islets in the Epididymal Fat Pad of Diabetic Mice

Among abdominal solid organ transplantation, pancreatic grafts are prone to develop severe ischemia reperfusion injury-associated graft damage, leading eventually to early graft loss. This protocol describes a model of murine pancreas transplantation using a non-suture cuff technique, ideally suited for analyzing these early, deleterious damages.

Mouse Model for Pancreas Transplantation Using a Modified Cuff Technique

This islet isolation protocol described a novel route of collagenase injection to digest the exocrine tissue and a simplified gradient procedure to purify the islets from mice. It involves enzymatic digestion, gradient separation/purification, and islet hand-picking. Successful isolation can yield 250–350 high quality and fully functional islets per mouse.

A Simple High Efficiency Protocol for Pancreatic Islet Isolation from Mice

A method for the omental transplantation of islets in a mouse is introduced. The isolated islets are mixed with hydrogel and the mixture is placed into the omental pouch of a diabetic mouse. Then, the blood glucose is monitored, and immuno-histochemical analysis is performed.

A Method for Islet Transplantation to the Omentum in Mouse

In this protocol, a method of murine islet isolation and transplantation into the inguinal subcutaneous white adipose tissue is described. Isolated syngeneic murine islets are transplanted into a murine recipient using a basement membrane hydrogel. The blood glucose level of the recipients is monitored, and histology analysis of the islet grafts is performed.

Inguinal Subcutaneous White Adipose Tissue (ISWAT) Transplantation Model of Murine Islets

Here we present refined surgical procedures on successfully performing intraportal islet transplantation, a clinically relevant but technically challenging surgical procedure, in mice.

Minimizing Post-Infusion Portal Vein Bleeding during Intrahepatic Islet Transplantation in Mice

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

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

We provide a minimally invasive protocol for the intramuscular transplantation of human stem cell-derived pancreatic stem cell islets into immunodeficient mice for in vivo implantation and downstream analysis.

Intramuscular Transplantation of Human Pluripotent Stem Cell-derived Pancreatic Endocrine Cells in Mice

Learn intraportal islet transplantation for type 1 diabetes research using mouse models. This method demonstrates microsurgery techniques to achieve normoglycemia and study islet engraftment in vivo.

Intraportal Transplantation of Pancreatic Islets in Mouse Model

Explore More Videos