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Summary

Abstract

Introduction

Protocol

Representative Results

Discussion

Acknowledgements

Materials

References

Medicine

Fat-Covered Islet Transplantation Using Epididymal White Adipose Tissue

Published: May 25th, 2021

DOI:

10.3791/62096

1Department of Regenerative Medicine and Transplantation, Faculty of Medicine, Fukuoka University, 2Center for Regenerative Medicine, Fukuoka University Hospital

This fat-covered islet transplantation method is suitable for the detection of engrafted islets in the intraperitoneal cavity. Notably, it does not require the use of biobinding agents or suturing.

Islet transplantation is a cellular replacement therapy for severe diabetes mellitus. The intraperitoneal cavity is typically the transplant site for this procedure. However, intraperitoneal islet transplantation has some limitations, including poor transplant efficacy, difficult graft detection ability, and a lack of graftectomy capability for post-transplant analysis. In this paper, "fat-covered islet transplantation", an intraperitoneal islet transplantation method that utilizes epididymal white adipose tissue, is used to assess the therapeutic effects of bioengineered islets. The simplicity of the method lies in the seeding of islets onto epididymal white adipose tissue and using the tissue to cover the islets. While this method can be categorized as an intraperitoneal islet transplantation technique, it shares characteristics with intra-adipose tissue islet transplantation. The fat-covered islet transplantation method demonstrates more robust therapeutic effects than intra-adipose tissue islet transplantation, however, including the improvement of blood glucose and plasma insulin levels and the potential for graft removal. We recommend the adoption of this method for assessing the mechanisms of islet engraftment into white adipose tissue and the therapeutic effects of bioengineered islets.

Islet transplantation is a cellular replacement therapy for patients with severe diabetes mellitus. Recent reports have shown that rates of insulin-independence at three years after transplantation improve up to 44%1 and that approximately 80% of recipients who receive more than 600,000 total islet equivalents achieve insulin independence2. Furthermore, in the most recent Collaborative Islet Transplant Registry report, it was revealed that fasting blood glucose levels were maintained at 60-140 mg/dL for over a period of 5 years in over 70% of patients who underwent islet transplant alone. The study also determined that a....

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The following procedure is performed in three steps. The first step includes the induction of diabetes in the recipient mice and the isolation of donor islets. The second step involves the preparation of islets before transplantation. In the third step, islet transplantation onto epididymal adipose tissue and covering of the islets using the adipose tissue is performed. After that, the therapeutic effects were assessed. The handling of the mice and the experimental procedures performed in this study comply with the '.......

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To compare the transplant efficacy of fat-covered islet transplantation to that after intraperitoneal islet transplantation, the same number of islets was implanted onto the peritoneum at the left paracolic space of control recipient diabetic animals. The blood glucose levels of mice with fat-covered islet transplantation were observed to gradually and significantly decrease compared to intraperitoneal islet transplanted mice (p = 0.0023; Figure 3A). One month after transplantation, the bloo.......

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The fat-covered islet transplantation method incorporates techniques from two different transplant techniques: intraperitoneal islet transplantation and intra-adipose tissue islet transplantation. As the surface membrane of epididymal white adipose tissue is considered to be the white adipose tissue that is covered by the peritoneum and that is attached to the epididymis, the fat-covered islet transplantation method can be anatomically categorized as a type of intraperitoneal islet transplantation. The technique by which.......

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This study was funded by a Grant-in-Aid for Scientific Research (C) (19K09839, NS) from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

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Name Company Catalog Number Comments
4-0 Nylon Alfresa ER2004NA45-KF2 Closing abdomen
Alexa 488-conjugated donkey anti-guinea pig Jackson Immunoresearch 706-546-148 Secondary antibody for insulin antibody
Alexa 647-conjugated donkey anti-rabbit Jackson Immunoresearch 711-606-152 Secondary antibody for von Willebrand factor antibody
DMEM, low glucose, pyruvate ThermoFisher Scientific 11885084 Culturing islets, transplanting islets
Eosin Fujifilm Wako Chemicals 051-06515 Using for staining tissue by eosin
Eppendorf Safe-Lock Tubes, 1.5 mL Eppendorf 30120086 Collecting islets 
Falcon 15 mL Conical Centrifuge Tubes Corning 352095 Collecting islets
Falcon 40 µm Cell Strainer Falcon 352340 Using for separating islets from other pancreatic tissue
Falcon 50 mL Conical Centrifuge Tubes Corning 352070 Discarding excessive medium/buffer
Guinea pig anti-insulin Agilent Technologies Japan, Ltd. (Dako) IR002 Primary antibody for murine insulin
Hematoxylin Muto Pure Chemicals Co., Ltd. 30002 Using for staining tissue by hematoxylin
Isodine solution 10% Shionogi&Co., Ltd. no catalog number Using for disinfection
Isoflurane Fujifilm Wako Chemicals 095-06573 Using for anesthesia
Labcon 1000 µL ZapSilk Low Retention Pipette Tips Labcon 1177-965-008 Using for separating islets from other pancreatic tissue
Labcon 200 µL ZapSilk Low Retention Pipette Tips Labcon 1179-965-008 Using for seeding islets onto epididymal white adipose tissue
Mintsensor Sanwa Kagaku Kenkyusho Co. Ltd., 8AEB02E Using for monitoring blood glucose
Pipetteman P-1000 Gilson F123602 Using for separating islets from other pancreatic tissue
Pipetteman P-200 Gilson F123601 Using for seeding islets onto epididymal white adipose tissue
Rabbit anti-vWF Abcam ab6994 Primary antibody for murine von Willebrand factor

  1. Barton, F. B., et al. Improvement in outcomes of clinical islet transplantation: 1999-2010. Diabetes Care. 35 (7), 1436-1445 (2012).
  2. Balamurugan, A. N., et al. Islet product characteristics and factors related to successful human islet transplantation from the Collaborative Islet Transplant Registry (CITR) 1999-2010. American Journal of Transplantation. 14 (11), 2595-2606 (2014).
  3. Collaborative Islet Transplant Registry. . Collaborative Islet Transplant Registry. Annual Report. , (2017).
  4. Rajab, A., et al. Total Pancreatectomy and Islet Autotransplantation Following Treated Hepatitis C Infection. Cell Transplantation. 27 (10), 1569-1573 (2018).
  5. Mellgren, A., Schnell Landstrom, A. H., Petersson, B., Andersson, A. The renal subcapsular site offers better growth conditions for transplanted mouse pancreatic islet cells than the liver or spleen. Diabetologia. 29 (9), 670-672 (1986).
  6. Hiller, W. F., Klempnauer, J., Luck, R., Steiniger, B. Progressive deterioration of endocrine function after intraportal but not kidney subcapsular rat islet transplantation. Diabetes. 40 (1), 134-140 (1991).
  7. Yasunami, Y., Lacy, P. E., Finke, E. H. A new site for islet transplantation--a peritoneal-omental pouch. Transplantation. 36 (2), 181-182 (1983).
  8. Kin, T., Korbutt, G. S., Rajotte, R. V. Survival and metabolic function of syngeneic rat islet grafts transplanted in the omental pouch. American Journal of Transplantation. 3 (3), 281-285 (2003).
  9. Kasoju, N., et al. Bioengineering a pre-vascularized pouch for subsequent islet transplantation using VEGF-loaded polylactide capsules. Biomaterials Science. 8 (2), 631-647 (2020).
  10. Sakata, N., Yoshimatsu, G., Kodama, S. White Adipose Tissue as a Site for Islet Transplantation. Transplantology. 1 (2), 55-70 (2020).
  11. Osama Gaber, A., Chamsuddin, A., Fraga, D., Fisher, J., Lo, A. Insulin independence achieved using the transmesenteric approach to the portal vein for islet transplantation. Transplantation. 77 (2), 309-311 (2004).
  12. Fujita, M., et al. Technique of endoscopic biopsy of islet allografts transplanted into the gastric submucosal space in pigs. Cell Transplantation. 22 (12), 2335-2344 (2013).
  13. Sakata, N., et al. Strategy for clinical setting in intramuscular and subcutaneous islet transplantation. Diabetes/Metabolism Research and Reviews. 30 (1), 1-10 (2014).
  14. Cantarelli, E., et al. Transplant Site Influences the Immune Response After Islet Transplantation: Bone Marrow Versus Liver. Transplantation. 101 (5), 1046-1055 (2017).
  15. White, S. A., et al. The risks of total pancreatectomy and splenic islet autotransplantation. Cell Transplantation. 9 (1), 19-24 (2000).
  16. Itoh, T., Nishinakamura, H., Kumano, K., Takahashi, H., Kodama, S. The Spleen Is an Ideal Site for Inducing Transplanted Islet Graft Expansion in Mice. PLoS One. 12 (1), 0170899 (2017).
  17. Sakata, N., Yoshimatsu, G., Kodama, S. The Spleen as an Optimal Site for Islet Transplantation and a Source of Mesenchymal Stem Cells. International Journal of Molecular Sciences. 19 (5), (2018).
  18. Sakata, N., et al. Effect of rat-to-mouse bioartificial pancreas xenotransplantation on diabetic renal damage and survival. Pancreas. 32 (3), 249-257 (2006).
  19. Nagaya, M., et al. Effectiveness of bioengineered islet cell sheets for the treatment of diabetes mellitus. Journal of Surgical Research. 227, 119-129 (2018).
  20. Weaver, J. D., et al. Vasculogenic hydrogel enhances islet survival, engraftment, and function in leading extrahepatic sites. Science Advances. 3 (6), 1700184 (2017).
  21. Dufour, J. M., et al. Development of an ectopic site for islet transplantation, using biodegradable scaffolds. Tissue Engineering. 11 (9-10), 1323-1331 (2005).
  22. Chen, X., et al. The epididymal fat pad as a transplant site for minimal islet mass. Transplantation. 84 (1), 122-125 (2007).
  23. Sakata, N., et al. Mechanism of Transplanted Islet Engraftment in Visceral White Adipose Tissue. Transplantation. 104 (12), 2516-2527 (2020).
  24. Navarro-Requena, C., et al. PEG hydrogel containing calcium-releasing particles and mesenchymal stromal cells promote vessel maturation. Acta Biomaterialia. 67, 53-65 (2018).
  25. Phelps, E. A., Headen, D. M., Taylor, W. R., Thule, P. M., Garcia, A. J. Vasculogenic bio-synthetic hydrogel for enhancement of pancreatic islet engraftment and function in type 1 diabetes. Biomaterials. 34 (19), 4602-4611 (2013).
  26. Manzoli, V., et al. Immunoisolation of murine islet allografts in vascularized sites through conformal coating with polyethylene glycol. American Journal of Transplantation. 18 (3), 590-603 (2018).
  27. Gotoh, M., Maki, T., Kiyoizumi, T., Satomi, S., Monaco, A. P. An improved method for isolation of mouse pancreatic islets. Transplantation. 40 (4), 437-438 (1985).
  28. Brandhorst, D., Brandhorst, H., Hering, B. J., Bretzel, R. G. Long-term survival, morphology and in vitro function of isolated pig islets under different culture conditions. Transplantation. 67 (12), 1533-1541 (1999).
  29. Noguchi, H., et al. Low-temperature preservation of isolated islets is superior to conventional islet culture before islet transplantation. Transplantation. 89 (1), 47-54 (2010).
  30. Itoh, T., et al. Low temperature condition prevents hypoxia-induced islet cell damage and HMGB1 release in a mouse model. Cell Transplantation. 21 (7), 1361-1370 (2012).
  31. Komatsu, H., et al. Optimizing Temperature and Oxygen Supports Long-term Culture of Human Islets. Transplantation. 103 (2), 299-306 (2019).
  32. Unger, R. H. Lipid overload and overflow: metabolic trauma and the metabolic syndrome. Trends in Endocrinology, Metabolism. 14 (9), 398-403 (2003).
  33. Mao, D., et al. A macroporous heparin-releasing silk fibroin scaffold improves islet transplantation outcome by promoting islet revascularisation and survival. Acta Biomaterialia. 59, 210-220 (2017).
  34. Wang, K., Wang, X., Han, C. S., Chen, L. Y., Luo, Y. Scaffold-supported Transplantation of Islets in the Epididymal Fat Pad of Diabetic Mice. Journal of Visualized Experiments. (125), e54995 (2017).
  35. Wang, X., Wang, K., Zhang, W., Qiang, M., Luo, Y. A bilaminated decellularized scaffold for islet transplantation: Structure, properties and functions in diabetic mice. Biomaterials. 138, 80-90 (2017).
  36. Rios, P. D., Zhang, X., Luo, X., Shea, L. D. Mold-casted non-degradable, islet macro-encapsulating hydrogel devices for restoration of normoglycemia in diabetic mice. Biotechnology and Bioengineering. 113 (11), 2485-2495 (2016).

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