Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

A prevascularized subcutaneous device-less site for islet and cellular transplantation

Nature Biotechnology volume 33, pages 518–523 (2015)Cite this article

Subjects

Abstract

Transplantation of donor-derived islets into the liver is a successful cellular replacement therapy for individuals with diabetes. However, the hepatic vasculature is not an optimal transplant site for several reasons, including graft attrition and the inability to retrieve or image the islets. Here we describe islet transplantation into a prevascularized, subcutaneous site created by temporary placement of a medically approved vascular access catheter. In mice with streptozotocin (STZ)-induced diabetes, transplantation of ∼500 syngeneic islets into the resulting 'device-less' space reversed diabetes in 91% of mice and maintained normoglycemia for >100 days. The approach was also effective in mice with pre-existing diabetes, in another mouse strain that mounts a more vigorous inflammatory response, and across an allogeneic barrier. These results demonstrate that transient priming of a subcutaneous site supports diabetes-reversing islet transplantation in mouse models without the need for a permanent cell-encapsulation device.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2: Histological analysis of islets transplanted long-term into the DL space.
Figure 3: The proinflammatory response elicited by angiocatheters composed of nylon (blue) or silicone (red) when implanted subcutaneously for 24 h, 1 week and 2 weeks.
Figure 4: Long-term function of syngeneic islet grafts transplanted into the DL space.
Figure 5: IPGTTs of syngeneic mouse islets transplanted under the KC or into the DL site, 100 days after transplant.
Figure 6: Long-term function of human islets transplanted into the DL space.

Similar content being viewed by others

References

  1. Shapiro, A.M. et al. Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen. N. Engl. J. Med. 343, 230–238 (2000).

    Article  CAS  Google Scholar 

  2. Ryan, E.A. et al. Five-year follow-up after clinical islet transplantation. Diabetes 54, 2060–2069 (2005).

    Article  CAS  Google Scholar 

  3. Shapiro, A.M. & Ricordi, C. in Textbook of Organ Transplantation, vol. 1. (eds. Kirk, A.D. et al.) 1–1,904 (Wiley-Blackwell, 2014).

  4. Harlan, D.M., Kenyon, N.S., Korsgren, O. & Roep, B.O. Current advances and travails in islet transplantation. Diabetes 58, 2175–2184 (2009).

    Article  CAS  Google Scholar 

  5. Plesner, A. & Verchere, C.B. Advances and challenges in islet transplantation: islet procurement rates and lessons learned from suboptimal islet transplantation. J. Transplant. 2011, 979527 (2011).

    Article  Google Scholar 

  6. Ricordi, C. & Strom, T.B. Clinical islet transplantation: advances and immunological challenges. Nat. Rev. Immunol. 4, 259–268 (2004).

    Article  CAS  Google Scholar 

  7. Shapiro, A.M. in Islet transplantation and beta cell replacement therapy. (eds. Shapiro, A.M. & Shaw, J.A.) (Informa Healthcare, New York, London, 2007).

  8. Brissova, M. & Powers, A.C. Revascularization of transplanted islets: can it be improved? Diabetes 57, 2269–2271 (2008).

    Article  CAS  Google Scholar 

  9. Pepper, A.R., Gala-Lopez, B., Ziff, O. & Shapiro, A.M. Revascularization of transplanted pancreatic islets and role of the transplantation site. Clin. Dev. Immunol. 2013, 352315 (2013).

    Article  Google Scholar 

  10. Merani, S., Toso, C., Emamaullee, J. & Shapiro, A.M. Optimal implantation site for pancreatic islet transplantation. Br. J. Surg. 95, 1449–1461 (2008).

    Article  CAS  Google Scholar 

  11. Vériter, S., Gianello, P. & Dufrane, D. Bioengineered sites for islet cell transplantation. Curr. Diab. Rep. 13, 745–755 (2013).

    Article  Google Scholar 

  12. Nishimura, R. et al. Assessment for revascularization of transplanted pancreatic islets at subcutaneous site in mice with a highly sensitive imaging system. Transplant. Proc. 43, 3239–3240 (2011).

    Article  CAS  Google Scholar 

  13. Saito, T. et al. Reversal of diabetes by the creation of neo-islet tissues into a subcutaneous site using islet cell sheets. Transplantation 92, 1231–1236 (2011).

    Article  CAS  Google Scholar 

  14. Sakata, N. et al. Strategy for clinical setting in intramuscular and subcutaneous islet transplantation. Diabetes Metab. Res. Rev. 30, 1–10 (2014).

    Article  Google Scholar 

  15. Simeonovic, C.J., Dhall, D.P., Wilson, J.D. & Lafferty, K.J. A comparative study of transplant sites for endocrine tissue transplantation in the pig. Aust. J. Exp. Biol. Med. Sci. 64, 37–41 (1986).

    Article  Google Scholar 

  16. Rajab, A. Islet transplantation: alternative sites. Curr. Diab. Rep. 10, 332–337 (2010).

    Article  Google Scholar 

  17. Anderson, J.M., Rodriguez, A. & Chang, D.T. Foreign body reaction to biomaterials. Semin. Immunol. 20, 86–100 (2008).

    Article  CAS  Google Scholar 

  18. Anderson, J.M. Biological response to materials. Annu. Rev. Mater. Res. 31, 81–110 (2001).

    Article  CAS  Google Scholar 

  19. Ward, W.K. A review of the foreign-body response to subcutaneously-implanted devices: the role of macrophages and cytokines in biofouling and fibrosis. J. Diabetes Sci. Technol. 2, 768–777 (2008).

    Article  Google Scholar 

  20. Fujiwara, N. & Kobayashi, K. Macrophages in inflammation. Curr. Drug Targets Inflamm. Allergy 4, 281–286 (2005).

    Article  CAS  Google Scholar 

  21. van Amerongen, M.J., Molema, G., Plantinga, J., Moorlag, H. & van Luyn, M.J. Neovascularization and vascular markers in a foreign body reaction to subcutaneously implanted degradable biomaterial in mice. Angiogenesis 5, 173–180 (2002).

    Article  CAS  Google Scholar 

  22. Grainger, D.W. All charged up about implanted biomaterials. Nat. Biotechnol. 31, 507–509 (2013).

    Article  CAS  Google Scholar 

  23. Sharkawy, A.A., Klitzman, B., Truskey, G.A. & Reichert, W.M. Engineering the tissue which encapsulates subcutaneous implants. I. Diffusion properties. J. Biomed. Mater. Res. 37, 401–412 (1997).

    Article  CAS  Google Scholar 

  24. Sharkawy, A.A., Klitzman, B., Truskey, G.A. & Reichert, W.M. Engineering the tissue which encapsulates subcutaneous implants. III. Effective tissue response times. J. Biomed. Mater. Res. 40, 598–605 (1998).

    Article  CAS  Google Scholar 

  25. Sharkawy, A.A., Klitzman, B., Truskey, G.A. & Reichert, W.M. Engineering the tissue which encapsulates subcutaneous implants. II. Plasma-tissue exchange properties. J. Biomed. Mater. Res. 40, 586–597 (1998).

    Article  CAS  Google Scholar 

  26. Pileggi, A. et al. Reversal of diabetes by pancreatic islet transplantation into a subcutaneous, neovascularized device. Transplantation 81, 1318–1324 (2006).

    Article  Google Scholar 

  27. Zhang, L. et al. Zwitterionic hydrogels implanted in mice resist the foreign-body reaction. Nat. Biotechnol. 31, 553–556 (2013).

    Article  CAS  Google Scholar 

  28. Olsson, R., Olerud, J., Pettersson, U. & Carlsson, P.O. Increased numbers of low-oxygenated pancreatic islets after intraportal islet transplantation. Diabetes 60, 2350–2353 (2011).

    Article  CAS  Google Scholar 

  29. Hellerström, C., Andersson, A., Korsgren, O., Jansson, L. & Sandler, S. Aspects of pancreatic islet transplantation in diabetes mellitus. Baillieres Clin. Gastroenterol. 3, 851–863 (1989).

    Article  Google Scholar 

  30. Korsgren, O. et al. Optimising islet engraftment is critical for successful clinical islet transplantation. Diabetologia 51, 227–232 (2008).

    Article  CAS  Google Scholar 

  31. Pileggi, A., Ricordi, C., Alessiani, M. & Inverardi, L. Factors influencing Islet of Langerhans graft function and monitoring. Clin. Chim. Acta 310, 3–16 (2001).

    Article  CAS  Google Scholar 

  32. Nyqvist, D., Kohler, M., Wahlstedt, H. & Berggren, P.O. Donor islet endothelial cells participate in formation of functional vessels within pancreatic islet grafts. Diabetes 54, 2287–2293 (2005).

    Article  CAS  Google Scholar 

  33. Zhang, H.F., Maslov, K., Stoica, G. & Wang, L.V. Functional photoacoustic microscopy for high-resolution and noninvasive in vivo imaging. Nat. Biotechnol. 24, 848–851 (2006).

    Article  CAS  Google Scholar 

  34. Costa, P.Z. & Soares, R. Neovascularization in diabetes and its complications. Unraveling the angiogenic paradox. Life Sci. 92, 1037–1045 (2013).

    Article  CAS  Google Scholar 

  35. Tamarat, R. et al. Impairment in ischemia-induced neovascularization in diabetes: bone marrow mononuclear cell dysfunction and therapeutic potential of placenta growth factor treatment. Am. J. Pathol. 164, 457–466 (2004).

    Article  CAS  Google Scholar 

  36. Brekken, R.A. et al. Selective inhibition of vascular endothelial growth factor (VEGF) receptor 2 (KDR/Flk-1) activity by a monoclonal anti-VEGF antibody blocks tumor growth in mice. Cancer Res. 60, 5117–5124 (2000).

    CAS  PubMed  Google Scholar 

  37. Kin, T. et al. Risk factors for islet loss during culture prior to transplantation. Transpl. Int. 21, 1029–1035 (2008).

    PubMed  Google Scholar 

  38. Ricordi, C., Lacy, P.E. & Scharp, D.W. Automated islet isolation from human pancreas. Diabetes 38 (suppl. 1), 140–142 (1989).

    Article  Google Scholar 

Download references

Acknowledgements

The authors thank H. Tanaka for Figure 1. We thank the Clinical Islet Laboratory and Alberta Health Services for providing human islet research preparations. This work was supported in part by the Diabetes Research Institute Foundation of Canada, Collaborative Research & Innovation Opportunities (CRIO)–Alberta Innovates Healthcare Solutions, and from University Hospital Foundation/C.F. “Curly” and Gladys B. MacLachlan Fund. All authors are members of the Alberta Diabetes Institute, and of the Canadian National Transplant Research Program. A.M.J.S. is supported through a Canada Research Chair in Transplantation Surgery and Regenerative Medicine, and through a Senior Clinical Scholarship from Alberta Innovates Healthcare Solutions.

Author information

Authors and Affiliations

  1. Clinical Islet Transplant Program, Alberta Diabetes Institute, University of Alberta, Edmonton, Alberta, Canada

    Andrew R Pepper, Boris Gala-Lopez, Rena Pawlick, Shaheed Merani, Tatsuya Kin & A M James Shapiro

  2. Department of Surgery, University of Alberta, Edmonton, Alberta, Canada

    Tatsuya Kin & A M James Shapiro

Authors
  1. Andrew R Pepper
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Boris Gala-Lopez
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Rena Pawlick
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Shaheed Merani
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Tatsuya Kin
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. A M James Shapiro
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

A.R.P. and A.M.J.S. initiated and designed the experiments. A.R.P., B.G.-L., S.M. and R.P. performed the experiments. T.K. isolated and provided human islets. A.R.P. and A.M.J.S. analyzed the data. All authors provided input for the manuscript writing and discussion.

Corresponding author

Correspondence to A M James Shapiro.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Overview of the subcutaneous, ‘device-less’ transplant approach

To create the ‘device-less’ transplant site, a 5-French textured nylon radiopaque angiographic catheter (Torcon NB® Advantage Beacon® tip Cook Medical, Indiana, USA) is: (a) Implanted beneath the skin; (b,c,d) Left for a period of 3-6 weeks; (e) Removed; (f) Subsequent to the implant period, the angiocatheter is removed (e) creating a vascularized void where the islet transplant is infused; (f) Islets are then infused via PE50 tubing (Instech Laboratories, Boston USA); (g) Incision site closed with a single surgical clip; and (h) The islet graft exhibited no visible profile post-transplant up to 100 days post-transplant.

Supplementary Figure 2 Rate of diabetes reversal, defined as percent euglycemic, in mouse recipients of syngeneic BALB/c islet grafts.

Glycemic control, measured by twice weekly non-fasting blood glucose levels, was monitored for 60 days post-islet transplant in chemically induced (STZ) diabetic mice. Reversal of diabetes was defined as a maintained non-fasting blood glucose level of <11.1 mM. Recipients received 500 BALC/c islets. Islet transplant groups: Kidney Capsule (KC – green, n=20), subcutaneous alone (SubQ – red, n=10), ‘device-less’ silicone (Silicone – purple, n=17) and ‘device-less’ nylon (Nylon – blue, n=21). Data points represent blood glucose mean ± s.e.m. Islets transplanted were from 10 separate isolations (n=20 pancreata per isolation).

Supplementary Figure 3 Intraperitoneal glucose tolerance tests in syngeneic BALB/c islet recipients.

Intraperitoneal glucose tolerance tests (3g/kg 50% dextrose intraperitoneal) in syngeneic BALB/c islet recipients under the Kidney Capsule (KC) or subcutaneous ‘device-less’ (DL) site, at 60 days post-transplant. (a) Blood glucose post-dextrose bolus (b) area under the curve (AUC) analysis did not differ between the KC (green, n=14) and nylon-DL (blue, n=15) recipients (p NS, one-way Anova-Newman-keuls post-hoc). Nylon-DL profiles were significantly improved compared to silicone-DL (pink, n=13), (**p<0.01 one-way Anova-Newman-keuls post-hoc). Islets transplanted beneath the skin without prevascularization, (SubQ – red, n=5), demonstrated diabetic profiles (***p<0.001 one-way Anova-Newman-keuls post-hoc test, compared with Nylon-DL). Naïve were normal, non-diabetic control BALB/c mice (black, n=16), and showed most optimal glycemic profiles (**p<0.01 and ***p<0.001 compared with Nylon-DL and KC respectively, one-way Anova-Newman-keuls post-hoc). Blood glucose was measured at 0, 15, 30, 60, 90 and 120 minutes. Data points represent blood glucose mean ± s.e.m. Islets transplanted were from 10 separate isolations (n=20 pancreata per isolation).

Supplementary Figure 4 Immunohistochemistry of representative syngeneic (BALB/c) islet grafts transplanted beneath the skin without prevascularization.

Immunohistochemistry of representative syngeneic (BALB/c) islet grafts transplanted beneath the skin without prevascularization, at 40 days post-transplant. Mason trichrome staining of cross-section of a subcutaneous islet graft (a) at 3x and (b) 10x magnification. Without prevascularization, islet necrosis and inflammatory destructive response ensued, resulting in graft loss.

Supplementary Figure 5 Vascular density of islet grafts post-transplantation.

(a) Islets transplanted into the unmodified subcutaneous space (red: SubQ Tx, n=8) had markedly less graft neovascularization compared to islets transplanted into the prevascularized DL site (blue: DL Tx, n=18) (p<0.01, unpaired t-test). Vascular density was quantified by measuring percentage of islet grafts staining positive for the vascular wall marker, von Willebrand (vWF) (green), using ImageJ software (ImageJ, National Institutes of Health, Bethesda MD). Representative images of vWF positive staining within (b) subcutaneous and (c) DL islet grafts. Scale bar represents 100μm. Values represent mean percentage of graft staining positive for vWF ± s.e.m.

Supplementary Figure 6 Time to normoglycemia in C57BL/6 vs. BALB/c mouse strains.

Time to normoglycemia in C57BL/6 (n=12) vs. BALB/c (n=21) mouse strains, using the device-less (DL) subcutaneous approach. Glycemic control was measured three times per week with non-fasting glucose levels, and reversal of diabetes defined as glucose <11.1mM. C57BL/6 mice reversed diabetes at a more rapid rate than BALB/c mice (11.3 ± 3.1 vs. 35.5 ± 6.1 days, p<0.05, unpaired t-test). Data points represent mean days post-transplant ± s.e.m. C57BL/6 transplants were conducted from 5 separate isolation isolations. BALB/c transplants were conducted from 10 separate islet isolations (n=20 pancreata per isolation).

Supplementary Figure 7 Impact of pre-existing diabetes before placement of the device-less (DL) catheter upon subsequent islet engraftment.

Mice were rendered diabetic 7 days ahead of DL catheter placement, and remained diabetic for a further 4 weeks before transplantation of 500 syngeneic islets. Glycemic control, measured by three times per week, was monitored for 50 days post-islet transplant in chemically induced streptozotocin (STZ) diabetic mice. Reversal of diabetes was defined as glucose <11.1 mM. No significant difference was found between pre-existing diabetic state (n=9) vs. post DL catheter placement diabetic state (n=25), upon subsequent islet engraftment (p NS, log-rank, Kaplan-Meier). Islets were transplanted from 15 separate islet isolations (n=20 pancreata per isolation).

Supplementary Figure 8 Impact of an allogeneic barrier upon diabetes reversal using the device-less (DL) subcutaneous site.

500 BALB/c islets were transplanted within the DL space of streptozotocin-diabetic C57BL/6 mice, in the presence or absence of immunosuppression (I.S.). Control mice (n=9) initially reversed diabetes, but then rapidly rejected allogeneic islet grafts. By contrast, with tacrolimus-based immunosuppression (0.5mg/kg/day for 28 days, n=3, subcutaneously via Alzet® mini-osmotic pump, (Alzet Cupertino, CA), rejection was delayed, and a proportion of grafts continued to function > 60 days. Hyperglycemia occurred promptly upon graft explantation. Kidney subcapsular allogeneic grafts (data not shown) rejected in a similar time-course. Insert depicts a representative islet allograft showing robust neovascularization, of similar response to that found in syngeneic grafts. Dashed lines indicate margins of prevascularized tract.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–8 and Supplementary Tables 1–3 (PDF 834 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Pepper, A., Gala-Lopez, B., Pawlick, R. et al. A prevascularized subcutaneous device-less site for islet and cellular transplantation. Nat Biotechnol 33, 518–523 (2015). https://doi.org/10.1038/nbt.3211

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt.3211

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research